Pub PHEUR-SE04 PDF

EUROPEAN
PHARMACOPOEIA
Free access to supportive pharmacopoeial texts
during the coronavirus (COVID-19) outbreak
April 2020
Published in accordance with the

Convention on the Elaboration of a European Pharmacopoeia
(European Treaty Series No. 50)
European Directorate Direction européenne

for the Quality de la qualité
of Medicines du médicament
& HealthCare & soins de santé
Council of Europe
Strasbourg
Free access to supportive pharmacopoeial texts during the coronavirus
(COVID-19) outbreak
The British Pharmacopoeia (BP) and European Pharmacopoeia (Ph. Eur. ) are committed
to supporting users during the coronavirus (COVID-19) outbreak and, as part of the wider
healthcare system response to this challenging period, the listed pharmacopoeial texts
(monographs, general chapters, appendices and supplementary chapters) are temporarily being
made freely available, at no cost, to all professionals involved in public health protection. This
is to support those involved in the development, manufacture or testing of these substances
and products worldwide. This list is being reviewed regularly and will be updated as required,
and ultimately withdrawn when appropriate. For example, when the outbreak is suitably under
control.
This in no way affects the existing legal status of the European or British Pharmacopoeias, nor
does it imply or confer any demonstrated effectiveness for the treatment of COVID-19 using
any of the substances or products in specific monographs. This is confirmed by the inclusion
of the following text at the bottom of pharmacopoeial texts reproduced in this document: “Not
official text. Please refer to the current legally effective version of the Pharmacopoeia to ensure
compliance.”
The British and European Pharmacopoeias have coordinated the release of these pharmacopoeial
texts to help maximise their distribution to potential users. These texts will be available on the
EDQM freepub website (https://register.edqm.eu/freepub) and on www.pharmacopoeia.com
as two separate PDFs; one which solely includes Ph. Eur. content and another which includes
the BP content incorporating Ph. Eur. content. Texts from the British Pharmacopoeia are from
the BP 2020 and texts from the European Pharmacopoeia are from the 10th Edition including
Supplement 10.1.
All rights reserved. Apart from any fair dealing for the purposes of research or private study,
this publication may not be reproduced or transmitted in any form or by any means without
the prior permission in writing of the publisher.
If you have any questions or comments please contact bpcom@mhra.gov.uk (for the British
Pharmacopoeia) and EDQM, via its Helpdesk (https://helpdesk.edqm.eu/servicedesk), (for
the European Pharmacopoeia).
The European Pharmacopoeia is published by the European Directorate for the Quality of Medicines &
HealthCare of the Council of Europe (EDQM).
© Council of Europe, 67075 Strasbourg Cedex, France - 2020

Title ID
1. General Notices 10000
2. Metyhods of analysis
2.2.1. Clarity and degree of opalescence of liquids 20201
2.2.2. Degree of coloration of liquids 20202
2.2.3. Potentiometric determination of pH 20203
2.2.5. Relative density 20205
2.2.7. Optical rotation 20207
2.2.13. Determination of water by distillation 20213
2.2.14. Melting point - capillary method 20214
2.2.15. Melting point - open capillary method 20215
2.2.16. Melting point - instantaneous method 20216
2.2.19. Amperometric titration 20219
2.2.20. Potentiometric titration 20220
2.2.21. Fluorimetry 20221
2.2.24. Absorption spectrophotometry, infrared 20224
2.2.25. Absorption spectrophotometry, ultraviolet and visible 20225
2.2.27. Thin-layer chromatography 20227
2.2.28. Gas chromatography 20228
2.2.29. Liquid chromatography 20229
2.2.32. Loss on drying 20232
2.2.43. Mass spectrometry 20243
2.2.46. Chromatographic separation techniques 20246
2.2.65. Voltametric titration 20265
2.4.14. Sulfated ash 20414
2.4.24. Identification and control of residual solvents 20424
2.5.8. Determination of primary aromatic amino-nitrogen 20508
2.5.37. Methyl, ethyl and isopropyl methanesulfonate in methanesulfonic acid 20537
2.5.38. Methyl, ethyl and isopropyl methanesulfonate in active substances 20538
2.5.39. Methanesulfonyl chloride in methanesulfonic acid 20539
2.6.1. Sterility 20601
2.6.14. Bacterial endotoxins 20614
2.9.1. Disintegration of tablets and capsules 20901
2.9.2. Disintegration of suppositories and pessaries 20902
2.9.5. Uniformity of mass of single-dose preparations 20905
2.9.6. Uniformity of content of single-dose preparations 20906
2.9.12. Sieve test 20912
2.9.17. Test for extractable volume of parenteral preparations 20917
Please refer to the current legally effective version of the Pharmacopoeia to access the official standards
2.9.18. Preparations for inhalation: aerodynamic assessment of fine particles 20918
2.9.27. Uniformity of mass of delivered doses from multidose containers 20927
2.9.35. Powder fineness 20935
2.9.40. Uniformity of dosage units 20940
2.9.44. Preparations for nebulisation: characterisation 20944
2.9.47. Demonstration of uniformity of dosage units using large sample sizes 20947
3. Materials for containers and containers

3.2.1. Glass containers for pharmaceutical use 30201
3.3.4. Sterile plastic containers for human blood and blood components 30304
3.3.5. Empty sterile containers of plasticised poly(vinyl chloride) for human 30305
blood and blood components
3.3.6. Sterile containers of plasticised poly(vinyl chloride) for human blood 30306
containing anticoagulant solution
4. Reagents
4.1.1. Reagents 40100
4.1. Reagents, standard solutions, buffer solutions 40101
4.1.2. Standard solutions for limit tests 40102
4.1.3. Buffer solutions 40103
4.2. Volumetric analysis 40200
4.2.1. Primary standards for volumetric solutions 40201
4.2.2. Volumetric solutions 40202
5. General texts
5.1.4. Microbiological quality of non-sterile pharmaceutical preparations 50104
and substances for pharmaceutical use
5.4. Residual solvents 50400
5.9. Polymorphism 50900
5.10. Control of impurities in substances for pharmaceutical use 51000
6. General monographs
Substances for Pharmaceutical Use 2034
Products of Fermentation 1468
Pharmaceutical Preparations 2619
7. Dosage form monographs
Capsules 16
Eye Preparations 1163
Liquid preparations for oral use 672
Parenteral Preparations 520
Tablets 478
Semi-solid preparations for cutaneous application 132
Individual monographs
Abacavir Sulfate 2589
Aciclovir 968
Atazanavir Sulfate 2898
Azithromycin 1649
Chloroquine Phosphate 544
Chloroquine Sulfate 545
Didanosine 2200
Disulfiram 603
Foscarnet sodium hexahydrate 1520
Ganciclovir 1752
Hydroxychloroquine Sulfate 2849
Idoxuridine 669
Indinavir Sulfate 2214
Lamivudine 2217
Lopinavir 2615
Nevirapine 2255
Nevirapine Hemihydrate 2479
Oseltamivir Phosphate 2422
Raltegravir Chewable Tablets 2939
Raltegravir Potassium 2887
Raltegravir Tablets 2938
Ribavirin 2109
Ritonavir 2136
Saquinavir Mesilate 2267
Stavudine 2130
Valaciclovir Hydrochloride 1768
Valaciclovir Hydrochloride Hydrate 2751
Zidovudine 1059
EUROPEAN PHARMACOPOEIA 1. General notices
07/2014:10000 (2) An enhanced approach to quality control could utilise

corrected 10.0 process analytical technology (PAT) and/or real-time release
testing (including parametric release) strategies as alternatives
to end-product testing alone. Real-time release testing
in circumstances deemed appropriate by the competent
authority is thus not precluded by the need to comply with the
Pharmacopoeia.
1. GENERAL NOTICES
(3) Reduction of animal testing : the European Pharmacopoeia
1.1. GENERAL STATEMENTS is dedicated to phasing out the use of animals for test purposes,
The General Notices apply to all monographs and other texts in accordance with the 3Rs (Replacement, Reduction,
of the European Pharmacopoeia. Refinement) set out in the European Convention for the
Protection of Vertebrate Animals used for Experimental and
The official texts of the European Pharmacopoeia are Other Scientific Purposes. In demonstrating compliance with
published in English and French. Translations in other the Pharmacopoeia as indicated above (1), manufacturers
languages may be prepared by the signatory States of the may consider establishing additional systems to monitor
European Pharmacopoeia Convention. In case of doubt consistency of production. With the agreement of the
or dispute, the English and French versions are alone competent authority, the choice of tests performed to assess
authoritative. compliance with the Pharmacopoeia when animal tests are
In the texts of the European Pharmacopoeia, the word prescribed is established in such a way that animal usage is
‘Pharmacopoeia’ without qualification means the European minimised as much as possible.
Pharmacopoeia. The official abbreviation Ph. Eur. may be Grade of materials. Certain materials that are the subject of
used to indicate the European Pharmacopoeia. a pharmacopoeial monograph may exist in different grades
The use of the title or the subtitle of a monograph implies suitable for different purposes. Unless otherwise indicated
that the article complies with the requirements of the relevant in the monograph, the requirements apply to all grades of
monograph. Such references to monographs in the texts of the material. In some monographs, particularly those on
the Pharmacopoeia are shown using the monograph title and excipients, a list of functionality-related characteristics that are
reference number in italics. relevant to the use of the substance may be appended to the
A preparation must comply throughout its period of validity ; monograph for information. Test methods for determination
a distinct period of validity and/or specifications for opened of one or more of these characteristics may be given, also for
or broached containers may be decided by the competent information.
authority. The subject of any other monograph must comply General monographs. Substances and preparations that are
throughout its period of use. The period of validity that is the subject of an individual monograph are also required
assigned to any given article and the time from which that to comply with relevant, applicable general monographs.
period is to be calculated are decided by the competent Cross-references to applicable general monographs are not
authority in light of experimental results of stability studies. normally given in individual monographs.
Unless otherwise indicated in the General Notices or in the General monographs apply to all substances and preparations
monographs, statements in monographs constitute mandatory within the scope of the Definition section of the general
requirements. General chapters become mandatory when monograph, except where a preamble limits the application,
referred to in a monograph, unless such reference is made in a for example to substances and preparations that are the subject
way that indicates that it is not the intention to make the text of a monograph of the Pharmacopoeia.
referred to mandatory but rather to cite it for information.
General monographs on dosage forms apply to all preparations
The active substances, excipients, pharmaceutical preparations of the type defined. The requirements are not necessarily
and other articles described in the monographs are intended comprehensive for a given specific preparation and
for human and veterinary use (unless explicitly restricted to requirements additional to those prescribed in the general
one of these uses). monograph may be imposed by the competent authority.
Quality systems. The quality standards represented by General monographs and individual monographs are
monographs are valid only where the articles in question are complementary. If the provisions of a general monograph do
produced within the framework of a suitable quality system. not apply to a particular product, this is expressly stated in the
The quality system must assure that the articles consistently individual monograph.
meet the requirements of the Pharmacopoeia.
Validation of pharmacopoeial methods. The test methods
Alternative methods. The tests and assays described given in monographs and general chapters have been validated
are the official methods upon which the standards of the in accordance with accepted scientific practice and current
Pharmacopoeia are based. With the agreement of the recommendations on analytical validation. Unless otherwise
competent authority, alternative methods of analysis may stated in the monograph or general chapter, validation of the
be used for control purposes, provided that the methods test methods by the analyst is not required.
used enable an unequivocal decision to be made as to
whether compliance with the standards of the monographs Implementation of pharmacopoeial methods. When
would be achieved if the official methods were used. In the implementing a pharmacopoeial method, the user must assess
event of doubt or dispute, the methods of analysis of the whether and to what extent the suitability of the method
Pharmacopoeia are alone authoritative. under the actual conditions of use needs to be demonstrated
according to relevant monographs, general chapters and
Demonstration of compliance with the Pharmacopoeia
quality systems.
(1) An article is not of Pharmacopoeia quality unless it
complies with all the requirements stated in the monograph. Conventional terms. The term ‘competent authority’
This does not imply that performance of all the tests in a means the national, supranational or international body or
monograph is necessarily a prerequisite for a manufacturer in organisation vested with the authority for making decisions
assessing compliance with the Pharmacopoeia before release concerning the issue in question. It may, for example, be a
of a product. The manufacturer may obtain assurance that national pharmacopoeia authority, a licensing authority or
a product is of Pharmacopoeia quality on the basis of its an official control laboratory.
design, together with its control strategy and data derived, for The expression ‘unless otherwise justified and authorised’
example, from validation studies of the manufacturing process. means that the requirements have to be met, unless the
General Notices (1) apply to all monographs and other texts 1

1. General notices EUROPEAN PHARMACOPOEIA
competent authority authorises a modification or an measured using a pipette, a volumetric flask or a burette, as
exemption where justified in a particular case. appropriate ; otherwise, a graduated measuring cylinder or a
Statements containing the word ‘should’ are informative or graduated pipette may be used. Volumes stated in microlitres
advisory. are measured using a micropipette or microsyringe.
In certain monographs or other texts, the terms ‘suitable’ and It is recognised, however, that in certain cases the precision
‘appropriate’ are used to describe a reagent, micro-organism, with which quantities are stated does not correspond to the
test method etc. ; if criteria for suitability are not described in number of significant figures stated in a specified numerical
the monograph, suitability is demonstrated to the satisfaction limit. The weighings and measurements are then carried out
of the competent authority. with a sufficiently improved accuracy.
Medicinal product. (a) Any substance or combination of Apparatus and procedures. Volumetric glassware complies
substances presented as having properties for treating or with Class A requirements of the appropriate International
preventing disease in human beings and/or animals ; or (b) Standard issued by the International Organisation for
any substance or combination of substances that may be used Standardisation.
in or administered to human beings and/or animals with a Unless otherwise prescribed, analytical procedures are carried
view either to restoring, correcting or modifying physiological out at a temperature between 15 °C and 25 °C.
functions by exerting a pharmacological, immunological or Unless otherwise prescribed, comparative tests are carried out
metabolic action, or to making a medical diagnosis. using identical tubes of colourless, transparent, neutral glass
Herbal medicinal product. Any medicinal product, exclusively with a flat base ; the volumes of liquid prescribed are for use
containing as active ingredients one or more herbal drugs or with tubes having an internal diameter of 16 mm, but tubes
one or more herbal drug preparations, or one or more such with a larger internal diameter may be used provided the
herbal drugs in combination with one or more such herbal volume of liquid used is adjusted (2.1.5). Equal volumes of
drug preparations. the liquids to be compared are examined down the vertical
Active substance. Any substance intended to be used in axis of the tubes against a white background, or if necessary
the manufacture of a medicinal product and that, when so against a black background. The examination is carried out in
used, becomes an active ingredient of the medicinal product. diffuse light.
Such substances are intended to furnish a pharmacological Any solvent required in a test or assay in which an indicator is
activity or other direct effect in the diagnosis, cure, mitigation, to be used is previously neutralised to the indicator, unless a
treatment or prevention of disease, or to affect the structure blank test is prescribed.
and function of the body. Water-bath. The term ‘water-bath’ means a bath of boiling
Excipient (auxiliary substance). Any constituent of a medicinal water unless water at another temperature is indicated.
product that is not an active substance. Adjuvants, stabilisers, Other methods of heating may be substituted provided the
antimicrobial preservatives, diluents, antioxidants, for temperature is near to but not higher than 100 °C or the
example, are excipients. indicated temperature.
Interchangeable methods. Certain general chapters contain Drying and ignition to constant mass. The terms ‘dried
a statement that the text in question is harmonised with to constant mass’ and ‘ignited to constant mass’ mean that
the corresponding text of the Japanese Pharmacopoeia 2 consecutive weighings do not differ by more than 0.5 mg,
and/or the United States Pharmacopeia and that these texts the 2nd weighing following an additional period of drying or
are interchangeable. This implies that if a substance or of ignition respectively appropriate to the nature and quantity
preparation is found to comply with a requirement using an of the residue.
interchangeable method from one of these pharmacopoeias Where drying is prescribed using one of the expressions ‘in a
it complies with the requirements of the European desiccator’ or ‘in vacuo’, it is carried out using the conditions
Pharmacopoeia. In the event of doubt or dispute, the text of described in chapter 2.2.32. Loss on drying.
the European Pharmacopoeia is alone authoritative. Reagents. The proper conduct of the analytical procedures
References to regulatory documents. Monographs and described in the Pharmacopoeia and the reliability of the
general chapters may contain references to documents results depend, in part, upon the quality of the reagents used.
issued by regulatory authorities for medicines, for example The reagents are described in general chapter 4. It is assumed
directives and notes for guidance of the European Union. that reagents of analytical grade are used ; for some reagents,
These references are provided for information for users for tests to determine suitability are included in the specifications.
the Pharmacopoeia. Inclusion of such a reference does not Solvents. Where the name of the solvent is not stated, the
modify the status of the documents referred to, which may be term ‘solution’ implies a solution in water.
mandatory or for guidance.
Where the use of water is specified or implied in the
1.2. OTHER PROVISIONS APPLYING TO GENERAL analytical procedures described in the Pharmacopoeia or
CHAPTERS AND MONOGRAPHS for the preparation of reagents, water complying with the
requirements of the monograph Purified water (0008) is
Quantities. In tests with numerical limits and assays, the used, except that for many purposes the requirements for
quantity stated to be taken for examination is approximate. bacterial endotoxins (Purified water in bulk) and microbial
The amount actually used, which may deviate by not more contamination (Purified water in containers) are not relevant.
than 10 per cent from that stated, is accurately weighed or The term ‘distilled water’ indicates purified water prepared
measured and the result is calculated from this exact quantity. by distillation.
In tests where the limit is not numerical, but usually depends
upon comparison with the behaviour of a reference substance The term ‘ethanol’ without qualification means anhydrous
in the same conditions, the stated quantity is taken for ethanol. The term ‘alcohol’ without qualification means
examination. Reagents are used in the prescribed amounts. ethanol (96 per cent). Other dilutions of ethanol are indicated
by the term ‘ethanol’ or ‘alcohol’ followed by a statement of the
Quantities are weighed or measured with an accuracy percentage by volume of ethanol (C2H6O) required.
commensurate with the indicated degree of precision. For
weighings, the precision corresponds to plus or minus 5 units Expression of content. In defining content, the expression
after the last figure stated (for example, 0.25 g is to be ‘per cent’ is used according to circumstances with one of
interpreted as 0.245 g to 0.255 g). For the measurement of 2 meanings :
volumes, if the figure after the decimal point is a zero or ends – per cent m/m (percentage, mass in mass) expresses the
in a zero (for example, 10.0 mL or 0.50 mL), the volume is number of grams of substance in 100 g of final product ;
2 See the information section on general monographs (cover pages)

– per cent V/V (percentage, volume in volume) expresses Limits of content. Where limits of content are prescribed,
the number of millilitres of substance in 100 mL of final they are those determined by the method described under
product. Assay.
The expression ‘parts per million’ (or ppm) refers to mass in Herbal drugs. In monographs on herbal drugs, the definition
mass, unless otherwise specified. indicates whether the subject of the monograph is, for
Temperature. Where an analytical procedure describes example, the whole drug or the drug in powdered form.
temperature without a figure, the general terms used have the Where a monograph applies to the drug in several states, for
following meaning : example both to the whole drug and the drug in powdered
form, the definition states this.
– in a deep-freeze : below − 15 °C ;
– in a refrigerator : 2 °C to 8 °C ;
– cold or cool : 8 °C to 15 °C ; PRODUCTION
Statements under the heading Production draw attention
– room temperature : 15 °C to 25 °C. to particular aspects of the manufacturing process but are
not necessarily comprehensive. They constitute mandatory
1.3. GENERAL CHAPTERS requirements for manufacturers, unless otherwise stated.
Containers. Materials used for containers are described They may relate, for example, to source materials ; to the
in general chapter 3.1. General names used for materials, manufacturing process itself and its validation and control ; to
particularly plastic materials, each cover a range of products in-process testing ; or to testing that is to be carried out by the
varying not only in the properties of the principal constituent manufacturer on the final article, either on selected batches
but also in the additives used. The test methods and limits or on each batch prior to release. These statements cannot
for materials depend on the formulation and are therefore necessarily be verified on a sample of the final article by an
applicable only for materials whose formulation is covered by independent analyst. The competent authority may establish
the preamble to the specification. The use of materials with that the instructions have been followed, for example, by
different formulations, and the test methods and limits applied examination of data received from the manufacturer, by
to them, are subject to agreement by the competent authority. inspection of manufacture or by testing appropriate samples.
The specifications for containers in general chapter 3.2 The absence of a Production section does not imply that
have been developed for general application to containers attention to features such as those referred to above is not
of the stated category, but in view of the wide variety of required.
containers available and possible new developments, the
publication of a specification does not exclude the use, in Choice of vaccine strain, Choice of vaccine composition.
justified circumstances, of containers that comply with The Production section of a monograph may define the
other specifications, subject to agreement by the competent characteristics of a vaccine strain or vaccine composition.
authority. Unless otherwise stated, test methods given for verification of
these characteristics are provided for information as examples
Reference may be made within the monographs of the of suitable methods. Subject to approval by the competent
Pharmacopoeia to the definitions and specifications for authority, other test methods may be used without validation
containers provided in chapter 3.2. Containers. The general against the method shown in the monograph.
monographs for pharmaceutical dosage forms may, under
the heading Definition/Production, require the use of certain
types of container ; certain other monographs may, under
the heading Storage, indicate the type of container that is POTENTIAL ADULTERATION
recommended for use. Due to the increasing number of fraudulent activities and
cases of adulteration, information may be made available to
1.4. MONOGRAPHS Ph. Eur. users to help detect adulterated materials (i.e. active
substances, excipients, intermediate products, bulk products
TITLES and finished products).
Monograph titles are in English and French in the respective To this purpose, a method for the detection of potential
versions and there is a Latin subtitle. adulterants and relevant limits, together with a reminder that
all stages of production and sourcing are subjected to a suitable
quality system, may be included in this section of monographs
RELATIVE ATOMIC AND MOLECULAR MASSES on substances for which an incident has occurred or that
The relative atomic mass (Ar) or the relative molecular present a risk of deliberate contamination. The frequency of
mass (Mr) is shown, as and where appropriate, at the beginning testing by manufacturers or by users (e.g. manufacturers of
of each monograph. The relative atomic and molecular masses intermediate products, bulk products and finished products,
and the molecular and graphic formulae do not constitute where relevant) depends on a risk assessment, taking into
analytical standards for the substances described. account the level of knowledge of the whole supply chain and
national requirements.
CHEMICAL ABSTRACTS SERVICE (CAS) REGISTRY This section constitutes requirements for the whole supply
NUMBER chain, from manufacturers to users (e.g. manufacturers of
CAS registry numbers are included for information in intermediate products, bulk products and finished products,
monographs, where applicable, to provide convenient access where relevant). The absence of this section does not imply
to useful information for users. CAS Registry Number® is a that attention to features such as those referred to above is
registered trademark of the American Chemical Society. not required.
DEFINITION
Statements under the heading Definition constitute an official CHARACTERS
definition of the substance, preparation or other article that is The statements under the heading Characters are not to be
the subject of the monograph. interpreted in a strict sense and are not requirements.

Solubility. In statements of solubility in the Characters Limits. The limits prescribed are based on data obtained
section, the terms used have the following significance, in normal analytical practice ; they take account of normal
referred to a temperature between 15 °C and 25 °C. analytical errors, of acceptable variations in manufacture and
compounding and of deterioration to an extent considered
Descriptive term Approximate volume of solvent in millilitres acceptable. No further tolerances are to be applied to the limits
per gram of solute prescribed to determine whether the article being examined
Very soluble less than 1 complies with the requirements of the monograph.
Freely soluble from 1 to 10 In determining compliance with a numerical limit, the
10 to 30
calculated result of a test or assay is first rounded to the
Soluble from
number of significant figures stated, unless otherwise
Sparingly soluble from 30 to 100 prescribed. The limits, regardless of whether the values are
100 to 1000
expressed as percentages or as absolute values, are considered
Slightly soluble from
significant to the last digit shown (for example 140 indicates 3
Very slightly soluble from 1000 to 10 000 significant figures). The last figure of the result is increased by
one when the part rejected is equal to or exceeds one half-unit,
Practically insoluble more than 10 000
whereas it is not modified when the part rejected is less than a
half-unit.
The term ‘partly soluble’ is used to describe a mixture where
only some of the components dissolve. The term ‘miscible’ is Indication of permitted limit of impurities. The acceptance
used to describe a liquid that is miscible in all proportions criteria for related substances are expressed in monographs
with the stated solvent. either in terms of comparison of peak areas (comparative tests)
or as numerical values. For comparative tests, the approximate
IDENTIFICATION content of impurity tolerated, or the sum of impurities, may
Scope. The tests given in the Identification section are not be indicated in brackets for information only. Acceptance
designed to give a full confirmation of the chemical structure or rejection is determined on the basis of compliance or
or composition of the product ; they are intended to give non-compliance with the stated test. If the use of a reference
confirmation, with an acceptable degree of assurance, that the substance for the named impurity is not prescribed, this
article conforms to the description on the label. content may be expressed as a nominal concentration of the
First and second identifications. Certain monographs substance used to prepare the reference solution specified in
have subdivisions entitled ‘First identification’ and ‘Second the monograph, unless otherwise described.
identification’. The test or tests that constitute the ‘First Herbal drugs. For herbal drugs, the sulfated ash, total ash,
identification’ may be used in all circumstances. The test or water-soluble matter, alcohol-soluble matter, water content,
tests that constitute the ‘Second identification’ may be used content of essential oil and content of active principle are
in pharmacies provided it can be demonstrated that the calculated with reference to the drug that has not been
substance or preparation is fully traceable to a batch certified specially dried, unless otherwise prescribed in the monograph.
to comply with all the other requirements of the monograph. Equivalents. Where an equivalent is given, for the purposes
Certain monographs give two or more sets of tests for the of the Pharmacopoeia only the figures shown are to be used in
purpose of the first identification, which are equivalent applying the requirements of the monograph.
and may be used independently. One or more of these sets Culture media. The culture media described in monographs
usually contain a cross-reference to a test prescribed in the and general chapters have been found to be satisfactory for
Tests section of the monograph. It may be used to simplify the intended purpose. However, the components of media,
the work of the analyst carrying out the identification and particularly those of biological origin, are of variable quality,
the prescribed tests. For example, one identification set and it may be necessary for optimal performance to modulate
cross-refers to a test for enantiomeric purity while the other the concentration of some ingredients, notably :
set gives a test for specific optical rotation : the intended
purpose of the two is the same, that is, verification that the – peptones and meat or yeast extracts, with respect to their
correct enantiomer is present. nutritive properties ;
Powdered herbal drugs. Monographs on herbal drugs may – buffering substances ;
contain schematic drawings of the powdered drug. These – bile salts, bile extract, deoxycholate, and colouring matter,
drawings complement the description given in the relevant depending on their selective properties ;
identification test. – antibiotics, with respect to their activity.
TESTS AND ASSAYS
STORAGE
Scope. The requirements are not framed to take account of all The information and recommendations given under the
possible impurities. It is not to be presumed, for example, that heading Storage do not constitute a pharmacopoeial
an impurity that is not detectable by means of the prescribed requirement but the competent authority may specify
tests is tolerated if common sense and good pharmaceutical particular storage conditions that must be met.
practice require that it be absent. See also below under
Impurities. The articles described in the Pharmacopoeia are stored
in such a way as to prevent contamination and, as far as
Calculation. Where the result of a test or assay is required possible, deterioration. Where special conditions of storage
to be calculated with reference to the dried or anhydrous are recommended, including the type of container (see section
substance or on some other specified basis, the determination 1.3. General chapters) and limits of temperature, they are
of loss on drying, water content or other property is carried stated in the monograph.
out by the method prescribed in the relevant test in the
monograph. The words ‘dried substance’ or ‘anhydrous The following expressions are used in monographs under
substance’ etc. appear in parentheses after the result. Storage with the meaning shown.
Where a quantitative determination of a residual solvent is In an airtight container means that the product is stored in an
carried out and a test for loss on drying is not carried out, airtight container (3.2). Care is to be taken when the container
the content of residual solvent is taken into account for the is opened in a damp atmosphere. A low moisture content
calculation of the assay content of the substance, the specific may be maintained, if necessary, by the use of a desiccant in
optical rotation and the specific absorbance. No further the container provided that direct contact with the product
indication is given in the specific monograph. is avoided.

Protected from light means that the product is stored either CRS Chemical reference substance
in a container made of a material that absorbs actinic light
sufficiently to protect the contents from change induced by
20
d 20 Relative density
such light, or in a container enclosed in an outer cover that λ Wavelength
provides such protection, or is stored in a place from which all
such light is excluded. HRS Herbal reference standard
LABELLING IU International Unit
In general, labelling of medicines is subject to supranational M Molarity
and national regulation and to international agreements. The
statements under the heading Labelling are not therefore Mr Relative molecular mass
comprehensive and, moreover, for the purposes of the mp Melting point
Pharmacopoeia only those statements that are necessary
to demonstrate compliance or non-compliance with the nD20 Refractive index
monograph are mandatory. Any other labelling statements are
included as recommendations. When the term ‘label’ is used Ph. Eur. U. European Pharmacopoeia Unit
in the Pharmacopoeia, the labelling statements may appear ppb Parts per billion (micrograms per kilogram)
on the container, the package, a leaflet accompanying the ppm
package, or a certificate of analysis accompanying the article, Parts per million (milligrams per kilogram)
as decided by the competent authority. R Substance or solution defined under
WARNINGS 4. Reagents
Materials described in monographs and reagents specified RF Retardation factor (see chapter 2.2.46)
for use in the Pharmacopoeia may be injurious to health Rst Used in chromatography to indicate the
unless adequate precautions are taken. The principles of ratio of the distance travelled by a substance
good quality control laboratory practice and the provisions to the distance travelled by a reference
of any appropriate regulations are to be observed at all substance
times. Attention is drawn to particular hazards in certain
monographs by means of a warning statement ; absence of such RV Substance used as a primary standard in
a statement is not to be taken to mean that no hazard exists. volumetric analysis (chapter 4.2.1)
IMPURITIES Abbreviations used in the monographs on
A list of all known and potential impurities that have been immunoglobulins, immunosera and vaccines
shown to be detected by the tests in a monograph may be CFU Colony-forming units
given. See also chapter 5.10. Control of impurities in substances
for pharmaceutical use. The impurities are designated by a LD50 The statistically determined quantity of a
letter or letters of the alphabet. Where a letter appears to substance that, when administered by the
be missing, the impurity designated by this letter has been specified route, may be expected to cause
deleted from the list during monograph development prior to the death of 50 per cent of the test animals
publication or during monograph revision. within a given period
FUNCTIONALITY-RELATED CHARACTERISTICS OF MLD Minimum lethal dose
EXCIPIENTS L+/10 dose The smallest quantity of a toxin that, in the
Monographs on excipients may have a section on conditions of the test, when mixed with
functionality-related characteristics. The characteristics, any 0.1 IU of antitoxin and administered by the
test methods for determination and any tolerances are not specified route, causes the death of the test
mandatory requirements ; they may nevertheless be relevant animals within a given period
for use of the excipient and are given for information (see also L+ dose The smallest quantity of a toxin that, in the
section 1.1. General statements). conditions of the test, when mixed with
REFERENCE STANDARDS 1 IU of antitoxin and administered by the
Certain monographs require the use of reference standards specified route, causes the death of the test
(chemical reference substances, herbal reference standards, animals within a given period
biological reference preparations, reference spectra). See lr/100 dose The smallest quantity of a toxin that, in
also chapter 5.12. Reference standards. The European the conditions of the test, when mixed
Pharmacopoeia Commission establishes the official with 0.01 IU of antitoxin and injected
reference standards, which are alone authoritative in case intracutaneously causes a characteristic
of arbitration. These reference standards are available from reaction at the site of injection within a
the European Directorate for the Quality of Medicines & given period
HealthCare (EDQM). Information on the available reference
standards and a batch validity statement can be obtained via Lp/10 dose The smallest quantity of toxin that, in the
the EDQM website. conditions of the test, when mixed with
0.1 IU of antitoxin and administered by the
specified route, causes paralysis in the test
1.5. ABBREVIATIONS AND SYMBOLS animals within a given period
A Absorbance Lo/10 dose The largest quantity of a toxin that, in the
conditions of the test, when mixed with
A11 cm
per cent Specific absorbance
0.1 IU of antitoxin and administered by the
Ar Relative atomic mass specified route, does not cause symptoms of
toxicity in the test animals within a given
[α]20 Specific optical rotation period
D
bp Boiling point Lf dose The quantity of toxin or toxoid that
flocculates in the shortest time with 1 IU of
BRP Biological reference preparation antitoxin

CCID50 The statistically determined quantity of NCPF National Collection of Pathogenic Fungi
virus that may be expected to infect 50 per London School of Hygiene and Tropical
cent of the cell cultures to which it is added Medicine
EID50 The statistically determined quantity of Keppel Street
virus that may be expected to infect 50 per
cent of the fertilised eggs into which it is London WC1E 7HT, Great Britain
inoculated NCTC National Collection of Type Cultures
ID50 The statistically determined quantity of Central Public Health Laboratory
a virus that may be expected to infect Colindale Avenue
50 per cent of the animals into which it is London NW9 5HT, Great Britain
inoculated
NCYC National Collection of Yeast Cultures
PD50 The statistically determined dose of a
vaccine that, in the conditions of the test, AFRC Food Research Institute
may be expected to protect 50 per cent of Colney Lane
the animals against a challenge dose of the Norwich NR4 7UA, Great Britain
micro-organisms or toxins against which it
is active NITE Biological Resource Center
ED50 The statistically determined dose of a Department of Biotechnology
vaccine that, in the conditions of the National Institute of Technology and
test, may be expected to induce specific Evaluation
antibodies in 50 per cent of the animals for 2-5-8 Kazusakamatari, Kisarazu-shi, Chiba,
the relevant vaccine antigens 292-0818
PFU Pock-forming units or plaque-forming units Japan
SPF Specified-pathogen-free S.S.I. Statens Serum Institut
80 Amager Boulevard, Copenhagen,
Collections of micro-organisms Denmark
ATCC American Type Culture Collection
10801 University Boulevard 1.6. UNITS OF THE INTERNATIONAL SYSTEM (SI) USED
Manassas, Virginia 20110-2209, USA IN THE PHARMACOPOEIA AND EQUIVALENCE WITH
OTHER UNITS
C.I.P. Collection de Bactéries de l’Institut Pasteur
B.P. 52, 25 rue du Docteur Roux INTERNATIONAL SYSTEM OF UNITS (SI)
The International System of Units comprises 2 main classes
75724 Paris Cedex 15, France of units, namely base units and derived units(1). The base units
IMI International Mycological Institute are the metre, the kilogram, the second, the ampere, the
Bakeham Lane kelvin, the mole and the candela.
Surrey TW20 9TY, Great Britain The derived units are formed as products of powers of the
I.P. Collection Nationale de Culture de base units according to the algebraic relationships linking the
Microorganismes (C.N.C.M.) corresponding quantities. Some of these derived units have
special names and symbols. The derived units used in the
Institut Pasteur Pharmacopoeia are shown in Table 1.6.-1.
25, rue du Docteur Roux
Some important and widely used units outside the
75724 Paris Cedex 15, France International System are shown in Table 1.6.-2.
NCIMB National Collection of Industrial and
Marine Bacteria Ltd The prefixes shown in Table 1.6.-3 are used to form the names
and symbols of the decimal multiples and submultiples of
23 St Machar Drive
SI units.
Aberdeen AB2 1RY, Great Britain
Table 1.6.-1. – Derived units used in the European Pharmacopoeia and equivalence with other units
Quantity Unit
Name Symbol Name Symbol Expression in Expression in Conversion of other units into SI units
SI base units other SI units
Wave number ν one per metre 1/m m− 1
Wavelength λ micrometre μm 10− 6m

nanometre nm 10− 9m
Area A, S square metre m2 m2
Volume V cubic metre m 3

m3 1 mL = 1 cm3 = 10− 6 m3
Frequency ν hertz Hz s − 1
Density ρ kilogram per kg/m 3

kg·m− 3 1 g/mL = 1 g/cm3 = 103 kg·m− 3
cubic metre
Velocity, speed v metre per m/s m·s− 1
second
(1) The definitions of the units used in the International System are given in the booklet ‘Le Système International d’Unités (SI)’, published by the Bureau International des Poids
et Mesures, Pavillon de Breteuil, F-92310 Sèvres.

Quantity Unit
Name Symbol Name Symbol Expression in Expression in Conversion of other units into SI units
SI base units other SI units
Force F newton N m·kg·s− 2 1 dyne = 1 g·cm·s− 2 = 10− 5 N
1 kp = 9.806 65 N
Pressure, stress p pascal Pa m− 1·kg·s− 2 N·m− 2 1 dyne/cm2 = 10− 1 Pa = 10− 1 N·m− 2
1 atm = 101 325 Pa = 101.325 kPa
1 bar = 105 Pa = 0.1 MPa
1 mm Hg = 133.322 387 Pa
1 Torr = 133.322 368 Pa
1 psi = 6.894 757 kPa
Dynamic η pascal second Pa·s m− 1·kg·s− 1 N·s·m− 2 1 P = 10− 1 Pa·s = 10− 1 N·s·m− 2
viscosity 1 cP = 1 mPa·s
Kinematic ν square metre m2/s m2·s− 1 Pa·s·m3·kg− 1 1 St = 1 cm2·s− 1 = 10− 4 m2·s− 1
viscosity per second N·m·s·kg− 1
Energy W joule J m2·kg·s− 2 N·m 1 erg = 1 cm2·g·s− 2 = 1 dyne·cm = 10− 7 J
1 cal = 4.1868 J
Power, P watt W m2·kg·s− 3 N·m·s− 1 1 erg/s = 1 dyne·cm·s− 1 =
radiant flux J·s− 1 10− 7 W = 10− 7 N·m·s− 1 = 10− 7 J·s− 1
Absorbed dose D gray Gy m2·s− 2 J·kg− 1 1 rad = 10− 2 Gy
(of radiant
energy)
Electric U volt V m2· kg·s− 3·A− 1 W·A− 1
potential
difference,
voltage
Electric R ohm Ω m2· kg·s− 3·A− 2 V·A− 1
resistance
Electric charge Q coulomb C A·s
Activity A becquerel Bq s− 1 1 Ci = 37·109 Bq = 37·109 s− 1

referred to a
radionuclide
Concentration c mole per cubic mol/m3 mol·m− 3 1 mol/L = 1 M = 1 mol/dm3 = 103 mol·m− 3
(of amount of metre
substance),
molar
concentration
Mass ρ kilogram per kg/m3 kg·m− 3 1 g/L = 1 g/dm3 = 1 kg·m− 3
concentration cubic metre
Catalytic Ζ katal kat mol·s-1
activity
NOTES
1. In the Pharmacopoeia, the Celsius temperature is used 4. In the Pharmacopoeia, conditions of centrifugation are
(symbol t). This is defined by the following equation : defined by reference to the acceleration due to gravity (g) :
t = T - T0 g = 9.806 65 m·s-2
where T0 = 273.15 K by definition. The Celsius or
centigrade temperature is expressed in degrees Celsius
(symbol °C). The unit ‘degree Celsius’ is equal to the unit
‘kelvin’.
5. Certain quantities without dimensions are used in the
Pharmacopoeia : relative density (2.2.5), absorbance
(2.2.25), specific absorbance (2.2.25) and refractive index
(2.2.6).
2. The practical expressions of concentrations used in the

Pharmacopoeia are defined in the General Notices.
6. The microkatal is defined as the enzymic activity that,
under defined conditions, produces the transformation
(e.g. hydrolysis) of 1 micromole of the substrate per second.
3. The radian is the plane angle between two radii of a circle

that cut off on the circumference an arc equal in length
to the radius.

Table 1.6.-2. – Non-SI units accepted for use with the SI units

Quantity Unit Value in SI units
Name Symbol
Time minute min 1 min = 60 s
hour h 1 h = 60 min = 3600 s
day d 1 d = 24 h = 86 400 s
Plane angle degree ° 1° = (π/180) rad
Volume litre L 1 L = 1 dm3 = 10− 3 m3
Mass tonne t 1 t = 103 kg
dalton Da 1 Da = 1.660539040(20) × 10-27 kg
Rotational revolution r/min 1 r/min = (1/60) s− 1
frequency per minute
Energy electronvolt eV 1eV=1.602176634 × 10-19J
Table 1.6.-3. – Decimal multiples and sub-multiples of SI units

Factor Prefix Symbol Factor Prefix Symbol
1018 exa E 10− 1 deci d
10 15
peta P 10 − 2
centi c
10 12
tera T 10 − 3
milli m
10 9
giga G 10 − 6
micro µ
106 mega M 10− 9 nano n
10 3
kilo k 10 − 12
pico p
10 2
hecto h 10 − 15
femto f
10 1
deca da 10 − 18
atto a

EUROPEAN PHARMACOPOEIA 2.2.1. Clarity and degree of opalescence of liquids
07/2017:20201 MEASUREMENTS IN RATIO MODE

The determination of opalescence of coloured liquids is done
using instruments with ratio mode, since colour provides a
negative interference, attenuating both incident and scattered
light and lowering the turbidity value. The effect is so great,
2.2.1. CLARITY AND DEGREE OF even for moderately coloured samples, that conventional
nephelometers cannot be used.
OPALESCENCE OF LIQUIDS In turbidimetry or nephelometry with ratio mode, the ratio
Opalescence is the effect of light being absorbed or scattered by of the transmission measurement to the 90° scattered light
submicroscopic particles or optical density inhomogeneities. measurement is determined. This procedure compensates
The absence of any particles or inhomogeneities in a solution for the light that is diminished by the colour of the sample.
results in a clear solution. Instruments with ratio mode use as light source a tungsten
A liquid is considered clear if its clarity is the same as that lamp with spectral sensitivity at about 550 nm operating
of water R or of the solvent used, or if its opalescence is at a filament colour temperature of 2700 K. Other suitable
not more pronounced than that of reference suspension I light sources may also be used. Silicon photodiodes and
(see Table 2.2.1.-1), when examined under the conditions photomultipliers are commonly used as detectors and record
described below. changes in light scattered or transmitted by the sample.
The light scattered at 90 ± 2.5° is measured by the primary
Requirements in monographs are expressed in terms of the detector. Other detectors measure back and forward scatter
visual method by comparing with the defined reference (reflected light) as well as transmitted light. The results are
suspensions (see Table 2.2.1.-1). However, instrumental obtained by calculating the ratio of the 90° scattered light
methods may also be used for determining compliance measured to the sum of the components of forward scattered
with monograph requirements once the suitability of the and transmitted light values.
instrument has been established as described below and
calibration with reference suspensions I-IV and with water R The instruments used are calibrated against standards of
or the solvent used has been performed. known turbidity and are capable of automatic measurement
of turbidity. The test results are obtained directly from
VISUAL METHOD the instrument and compared to the specifications in the
Using identical test-tubes of colourless, transparent, neutral individual monograph.
glass with a flat base and an internal diameter of 15-25 mm, Alternatively, the influence of the colour of the sample
compare the liquid to be examined with a reference suspension may also be eliminated by using an infrared light-emitting
freshly prepared as described below. Ensure that the depths of diode (IR LED) having an emission maximum at 860 nm
the layers in the 2 test-tubes are the same (about 40 mm). with a 60 nm spectral bandwidth as the light source of the
Compare the liquids in diffused daylight 5 min after instrument.
preparation of the reference suspension, viewing vertically INSTRUMENT REQUIREMENTS
against a black background. Instruments complying with the following characteristics and
System suitability. The diffusion of light must be such that verified using reference suspensions as described below may
reference suspension I can readily be distinguished from be used instead of visual examination for determination of
water R, and that reference suspension II can readily be compliance with monograph requirements.
distinguished from reference suspension I (see Table 2.2.1.-1). – Measuring unit : NTU (nephelometric turbidity units).
NTU is based on the turbidity of a primary standard
INSTRUMENTAL METHOD of formazin. FTU (formazin turbidity units) or FNU
The instrumental assessment of clarity and opalescence (formazin nephelometric units) are also used, and are
provides a more discriminatory test that does not depend on equivalent to NTU in regions of low turbidity (up to
the visual acuity of the analyst. Numerical results are more 40 NTU). These units are used in all 3 instrumental
useful for process control and quality monitoring, especially methods (nephelometry, turbidimetry and in ratio mode).
in stability studies. For example, previous numerical data on – Measuring range : 0.01-1100 NTU.
stability can be extrapolated to determine whether a given – Resolution : 0.01 NTU within the range 0-9.99 NTU ;
batch of a preparation will exceed shelf-life limits prior to the 0.1 NTU within the range 10.0-99.9 NTU ; and 1 NTU for
expiry date. the range > 100 NTU.
TURBIDIMETRY AND NEPHELOMETRY – Accuracy : ± (10 per cent of reading + 0.01 NTU) within
When a suspension is viewed at right angles to the direction the range 0-20 NTU ; ± 7.5 per cent within the range
of the incident light, the system appears opalescent due to the 20-1100 NTU.
scattering of light by the particles of the suspension (Tyndall – Repeatability : ± 0.05 NTU within the range
effect). A certain portion of the light beam entering a turbid 0-20 NTU ; ± 2 per cent of the reading within the
liquid is transmitted, another portion is absorbed and the range 20-1100 NTU.
remaining portion is scattered by the suspended particles.
Instruments with measuring range or resolution, accuracy and
The light-scattering effect of suspended particles can be
repeatability capabilities other than those mentioned above
measured either indirectly by observation of the transmitted
may be used provided they are sufficiently validated and are
light (turbidimetry) or directly by measuring the scattered
capable for the intended use.
light (nephelometry). Turbidimetry and nephelometry are
more reliable in low turbidity ranges, where there is a linear CONTROL OF INSTRUMENT PERFORMANCE
relationship between turbidity values and detector signals. – Calibration : performed with at least 4 reference suspensions
As the degree of turbidity increases, not all the particles of formazin covering the measuring range of interest.
are exposed to the incident light and the scattered or the Reference suspensions described in this chapter or suitable
transmitted radiation of other particles is hindered on its way reference standards calibrated against the primary reference
to the detector. suspensions may be used.
For quantitative measurements, the construction of calibration – Stray light : < 0.15 NTU within the range 0-10 NTU ;
curves is essential. Linearity must be based on at least 4 < 0.5 NTU within the range 10-1100 NTU. Stray light is
levels of concentrations. Reference suspensions must show a defined as that light that reaches the nephelometric detector
sufficiently stable degree of turbidity and must be produced without being a result of scatter from the sample. Stray light
under well-defined conditions. is always a positive interference and is a significant source

2.2.1. Clarity and degree of opalescence of liquids EUROPEAN PHARMACOPOEIA
of error in low-range turbidity measurements. Sources of the hydrazine sulfate solution. Mix and allow to stand for
of stray light include : imperfections in and scratches on 24 h. This suspension is stable for 2 months, provided it is
sample cells, internal reflections of the optical system, stored in a glass container free from surface defects. The
contamination of the optics or sample cell chamber with suspension must not adhere to the glass and must be mixed
dust, and electronic noise. Instrument design can also thoroughly before use.
affect stray light. The influence of stray light becomes Standard of opalescence. Dilute 15.0 mL of the primary
negligible in ratio mode measurements. opalescent suspension to 1000.0 mL with water R. This
The test methodology for the specific substance/product to be suspension is freshly prepared and may be stored for up to
analysed must also be verified to demonstrate its analytical 24 h.
capability. The instrument and methodology shall be
consistent with the attributes of the substance to be examined. Reference suspensions. Prepare the reference suspensions
Measurements of standards and samples should be carried out according to Table 2.2.1.-1. Mix and shake before use.
under the same temperature conditions, preferably between
20 °C and 25 °C. Table 2.2.1.-1
REFERENCE SUSPENSIONS I II III IV
Formazin has several desirable characteristics that make it an Standard of opalescence 5.0 mL 10.0 mL 30.0 mL 50.0 mL
excellent turbidity standard. It can be reproducibly prepared
from assayed raw materials. The physical characteristics Water R 95.0 mL 90.0 mL 70.0 mL 50.0 mL
make it a desirable light-scatter calibration standard. The
formazin polymer consists of chains of different lengths, Measurements of reference suspensions I-IV in ratio mode
which fold into random configurations. This results in a wide show a linear relationship between the concentrations and
variety of particle shapes and sizes, which allows the analysis measured NTU values (see Table 2.2.1.-2).
of different particle sizes and shapes that are found in real
samples. Stabilised formazin suspensions that can be used to
Table 2.2.1.-2
prepare stable, diluted turbidity standards are commercially
available and may be used after comparison with the standards Formazin suspensions Opalescent values (NTU)
prepared as described.
Reference suspension I 3
All steps of the preparation of reference suspensions as
described below are carried out at 25 ± 3 °C. Reference suspension II 6
Hydrazine sulfate solution. Dissolve 1.0 g of hydrazine Reference suspension III 18
sulfate R in water R and dilute to 100.0 mL with the same
solvent. Allow to stand for 4-6 h. Reference suspension IV 30
Primary opalescent suspension (formazin suspension). Standard of opalescence 60

In a 100 mL ground-glass-stoppered flask, dissolve 2.5 g of Primary opalescent suspension 4000
hexamethylenetetramine R in 25.0 mL of water R. Add 25.0 mL

EUROPEAN PHARMACOPOEIA 2.2.2. Degree of coloration of liquids
01/2008:20202 Blue primary solution. Dissolve 63 g of copper sulfate

pentahydrate R in about 900 mL of a mixture of 25 mL of
hydrochloric acid R and 975 mL of water R and dilute to
1000.0 mL with the same mixture. Titrate and adjust the
solution to contain 62.4 mg of CuSO4,5H2O per millilitre by
adding the same acidic mixture.
2.2.2. DEGREE OF COLORATION OF Titration. Place in a 250 mL conical flask fitted with a
LIQUIDS ground-glass stopper, 10.0 mL of the solution, 50 mL of
water R, 12 mL of dilute acetic acid R and 3 g of potassium
The examination of the degree of coloration of liquids in the iodide R. Titrate the liberated iodine with 0.1 M sodium
range brown-yellow-red is carried out by one of the 2 methods thiosulfate, using 0.5 mL of starch solution R, added towards
below, as prescribed in the monograph. the end of the titration, as indicator. The end-point is reached
A solution is colourless if it has the appearance of water R or when the solution shows a slight pale brown colour.
the solvent or is not more intensely coloured than reference 1 mL of 0.1 M sodium thiosulfate is equivalent to 24.97 mg
solution B9. of CuSO4,5H2O.
Standard solutions
METHOD I
Using the 3 primary solutions, prepare the 5 standard
Using identical tubes of colourless, transparent, neutral glass solutions as follows (Table 2.2.2.-1) :
of 12 mm external diameter, compare 2.0 mL of the liquid
to be examined with 2.0 mL of water R or of the solvent or Table 2.2.2.-1
of the reference solution (see Tables of reference solutions)
prescribed in the monograph. Compare the colours in diffused Volume in millilitres
daylight, viewing horizontally against a white background. Standard solution Yellow Red Blue Hydrochloric acid
solution solution solution (10 g/L HCl)
METHOD II B (brown) 3.0 3.0 2.4 1.6
Using identical tubes of colourless, transparent, neutral glass BY (brownish-yellow) 2.4 1.0 0.4 6.2
with a flat base and an internal diameter of 15 mm to 25 mm, Y (yellow) 2.4 0.6 0.0 7.0
compare the liquid to be examined with water R or the solvent
or the reference solution (see Tables of reference solutions) GY (greenish-yellow) 9.6 0.2 0.2 0.0
prescribed in the monograph, the depth of the layer being R (red) 1.0 2.0 0.0 7.0
40 mm. Compare the colours in diffused daylight, viewing
vertically against a white background. Reference solutions for Methods I and II
REAGENTS Using the 5 standard solutions, prepare the following reference

solutions.
Primary solutions
Table 2.2.2.-2. - Reference solutions B
Yellow solution. Dissolve 46 g of ferric chloride R in about
900 mL of a mixture of 25 mL of hydrochloric acid R and Volumes in millilitres
975 mL of water R and dilute to 1000.0 mL with the same
Reference Standard solution B Hydrochloric acid
mixture. Titrate and adjust the solution to contain 45.0 mg of solution (10 g/L HCl)
FeCl3,6H2O per millilitre by adding the same acidic mixture. B1 75.0 25.0
Protect the solution from light.
B2 50.0 50.0
Titration. Place in a 250 mL conical flask fitted with a
ground-glass stopper, 10.0 mL of the solution, 15 mL of B3 37.5 62.5
water R, 5 mL of hydrochloric acid R and 4 g of potassium B4 25.0 75.0
iodide R, close the flask, allow to stand in the dark for 15 min
and add 100 mL of water R. Titrate the liberated iodine with B5 12.5 87.5
0.1 M sodium thiosulfate, using 0.5 mL of starch solution R, B6 5.0 95.0
added towards the end of the titration, as indicator.
B7 2.5 97.5
1 mL of 0.1 M sodium thiosulfate is equivalent to 27.03 mg
of FeCl3,6H2O. B8 1.5 98.5
Red solution. Dissolve 60 g of cobalt chloride R in about B9 1.0 99.0

900 mL of a mixture of 25 mL of hydrochloric acid R and
975 mL of water R and dilute to 1000.0 mL with the same Table 2.2.2.-3. - Reference solutions BY
mixture. Titrate and adjust the solution to contain 59.5 mg of
CoCl2,6H2O per millilitre by adding the same acidic mixture. Volumes in millilitres
Titration. Place in a 250 mL conical flask fitted with a Reference Standard solution BY Hydrochloric acid
ground-glass stopper, 5.0 mL of the solution, 5 mL of dilute solution (10 g/L HCl)
hydrogen peroxide solution R and 10 mL of a 300 g/L solution BY1 100.0 0.0
of sodium hydroxide R. Boil gently for 10 min, allow to cool BY2 75.0 25.0
and add 60 mL of dilute sulfuric acid R and 2 g of potassium
iodide R. Close the flask and dissolve the precipitate by BY3 50.0 50.0
shaking gently. Titrate the liberated iodine with 0.1 M sodium BY4 25.0 75.0
thiosulfate, using 0.5 mL of starch solution R, added towards
the end of the titration, as indicator. The end-point is reached BY5 12.5 87.5
when the solution turns pink. BY6 5.0 95.0
1 mL of 0.1 M sodium thiosulfate is equivalent to 23.79 mg of BY7 2.5 97.5
CoCl2,6H2O.

2.2.2. Degree of coloration of liquids EUROPEAN PHARMACOPOEIA
Table 2.2.2.-4. - Reference solutions Y Table 2.2.2.-6. - Reference solutions R

Volumes in millilitres Volumes in millilitres
Reference Standard solution Y Hydrochloric acid Reference Standard solution R Hydrochloric acid
solution (10 g/L HCl) solution (10 g/L HCl)
Y1 100.0 0.0 R1 100.0 0.0
Y2 75.0 25.0 R2 75.0 25.0
Y3 50.0 50.0 R3 50.0 50.0
Y4 25.0 75.0 R4 37.5 62.5
Y5 12.5 87.5 R5 25.0 75.0
Y6 5.0 95.0 R6 12.5 87.5
Y7 2.5 97.5 R7 5.0 95.0
Table 2.2.2.-5. - Reference solutions GY Storage

Volumes in millilitres For Method I, the reference solutions may be stored in sealed
Reference Standard solution GY Hydrochloric acid tubes of colourless, transparent, neutral glass of 12 mm
solution (10 g/L HCl) external diameter, protected from light.
GY1 25.0 75.0 For Method II, prepare the reference solutions immediately
GY2 15.0 85.0
before use from the standard solutions.
GY3 8.5 91.5
GY4 5.0 95.0
GY5 3.0 97.0
GY6 1.5 98.5
GY7 0.75 99.25

EUROPEAN PHARMACOPOEIA 2.2.3. Potentiometric determination of pH
07/2016:20203 Management of electrodes. The electrodes are stored

appropriately and according to the manufacturer’s
recommendations (e.g. in an electrolyte solution or a suitable
storage solution). Before measurement, the electrodes are
visually checked. Refillable electrodes are checked for the
absence of air bubbles in the glass bulb and to ensure that
2.2.3. POTENTIOMETRIC the inner electrolyte solution level is satisfactory. The filling
DETERMINATION OF pH orifice has to remain open during the measurement. It is also
recommended that the diaphragm of the reference electrode is
checked. Before first use, or if the electrode has been stored
The pH of an aqueous solution is defined as the negative out of electrolyte solution, it is usually necessary to condition
logarithm of the activity of its hydrogen ions, expressed it, according to the recommendations of the manufacturer.
conventionally as the hydrogen ion concentration of If pH stabilisation is too slow (i.e. a long response time), or
the solution. For practical purposes, its definition is an a zero point shift, reduced slope or any other difficulties in
experimental one. The pH of a solution to be examined is calibration are observed, the electrode will probably need to
related to that of a reference solution (pHs) by the following be cleaned or replaced. The cleaning is performed depending
equation : on the type of sample and as prescribed in the manufacturer’s
manual. Regular cleaning is recommended.
E - Es
pH = pHs - Calibration and measurement conditions. Unless otherwise
k prescribed in the monograph, all measurements are carried out
in which E is the potential, expressed in volts, of the cell at the same temperature as that used for calibration (± 2.5 °C),
containing the solution to be examined and Es is the potential, usually between 20 °C and 25 °C. Table 2.2.3.-2 shows the
expressed in volts, of the cell containing the solution of known variation of pH with respect to temperature of a number of
pH (pHs), k is the change in potential per unit change in pH, reference buffer solutions used for calibration. Follow the
expressed in volts and calculated from the Nernst equation. manufacturer’s instructions for temperature correction.
The calibration consists of the determination of the slope
Table 2.2.3.-1. – Values of k at different temperatures (e.g. 95-105 per cent) and the offset of the measuring system.
Temperature (°C) k (V) Most commercial pH meters offer a “self test” or “start-up test”
where, for example, the slope and asymmetry potential are
15 0.0572 tested and compared to the manufacturer‘s specifications. The
20 0.0582 apparatus is calibrated using at least 2 buffer solutions chosen
so that the expected pH value of the solution to be examined
25 0.0592 lies between the pH values of the buffer solutions. The range
30 0.0601 must be at least 2 pH units. The pH of another buffer solution
of intermediate pH, read from the scale, must not differ by
35 0.0611 more than 0.05 pH units from the value corresponding to
that solution.
The potentiometric determination of pH is made by measuring Reference buffer solutions are preferably commercially
the potential difference between 2 appropriate electrodes available certified reference materials. Alternatively, buffer
immersed in the solution to be examined ; 1 of these electrodes solutions can be prepared in-house according to Table 2.2.3.-2.
is sensitive to hydrogen ions (usually a glass electrode) and These solutions must be traceable to primary standards.
the other is the reference electrode (e.g. a silver-silver chloride Calibration has to be performed on a regular basis, preferably
electrode). They are often combined as 1 compact electrode, each day of use or before each series of measurements.
together with a temperature probe.
Immerse the electrodes in the solution to be examined and
Apparatus. The measuring apparatus is usually a voltmeter take the reading in the same conditions as those applied for
with an input resistance at least 100 times that of the electrodes the reference buffer solutions.
used. It is normally graduated in pH units and has a sensitivity
such that discrimination of at least 0.05 pH unit or at least If suspensions, emulsions or solutions of non-aqueous or
0.003 V may be achieved. partially non-aqueous character are measured on a system
calibrated as described above, the pH reading can only be
Recent pH meters are microprocessor-controlled and are considered to be an approximation of the true value. Suitable
operated using the manufacturer’s firmware or software, electrodes have to be used for pH measurements of such
according to given instructions. mixtures.
Table 2.2.3.-2. – pH of reference buffer solutions at various temperatures
Temperature Potassium Potassium Potassium Potassium Potassium Potassium Disodium Sodium Calcium
(°C) tetraoxalate hydrogen dihydrogen hydrogen dihydrogen dihydrogen tetraborate carbonate hydroxide,
0.05 M tartrate citrate phthalate phosphate phosphate 0.01 M 0.025 M saturated
saturated at 0.05 M 0.05 M 0.025 M 0.0087 M + at 25°C
25 °C + + Sodium
Disodium Disodium bicarbonate
hydrogen hydrogen 0.025 M
phosphate phosphate
0.025 M 0.0303 M
C4H3KO8,2H2O C4H5KO6 C6H7KO7 C8H5KO4 KH2PO4+ KH2PO4+ Na2B4O7, Na2CO3+ Ca(OH)2
Na2HPO4 Na2HPO4 10H2O NaHCO3
15 1.67 3.80 4.00 6.90 7.45 9.28 10.12 12.81
20 1.68 3.79 4.00 6.88 7.43 9.23 10.06 12.63
25 1.68 3.56 3.78 4.01 6.87 7.41 9.18 10.01 12.45
30 1.68 3.55 3.77 4.02 6.85 7.40 9.14 9.97 12.29

2.2.3. Potentiometric determination of pH EUROPEAN PHARMACOPOEIA
Temperature Potassium Potassium Potassium Potassium Potassium Potassium Disodium Sodium Calcium
(°C) tetraoxalate hydrogen dihydrogen hydrogen dihydrogen dihydrogen tetraborate carbonate hydroxide,
0.05 M tartrate citrate phthalate phosphate phosphate 0.01 M 0.025 M saturated
saturated at 0.05 M 0.05 M 0.025 M 0.0087 M + at 25°C
25 °C + + Sodium
Disodium Disodium bicarbonate
hydrogen hydrogen 0.025 M
phosphate phosphate
0.025 M 0.0303 M
C4H3KO8,2H2O C4H5KO6 C6H7KO7 C8H5KO4 KH2PO4+ KH2PO4+ Na2B4O7, Na2CO3+ Ca(OH)2
Na2HPO4 Na2HPO4 10H2O NaHCO3
35 1.69 3.55 3.76 4.02 6.84 7.39 9.10 9.93 12.13
∆pH (1) + 0.001 − 0.0014 − 0.0022 + 0.0012 − 0.0028 − 0.0028 − 0.0082 − 0.0096 − 0.034
∆t
(1) pH variation per degree Celsius.
PREPARATION OF REFERENCE BUFFER SOLUTIONS Potassium dihydrogen phosphate 0.0087 M + Disodium

Potassium tetraoxalate 0.05 M. Dissolve 12.61 g of hydrogen phosphate 0.0303 M. Dissolve 1.18 g of KH2PO4
C4H3KO8,2H2O in carbon dioxide-free water R and dilute to and 4.30 g of Na2HPO4, both previously dried for 2 h at
1000.0 mL with the same solvent. 120 ± 2 °C, in carbon dioxide-free water R and dilute to
1000.0 mL with the same solvent.
Potassium hydrogen tartrate, saturated at 25 °C. Shake an Disodium tetraborate 0.01 M. Dissolve 3.80 g of
excess of C4H5KO6 vigorously with carbon dioxide-free water R Na B O ,10H O in carbon dioxide-free water R and dilute
2 4 7 2
at 25 °C. Filter or decant. Prepare immediately before use. to 1000.0 mL with the same solvent. Store protected from
atmospheric carbon dioxide.
Potassium dihydrogen citrate 0.05 M. Dissolve 11.41 g
of C6H7KO7 in carbon dioxide-free water R and dilute to Sodium carbonate 0.025 M + Sodium hydrogen carbonate
1000.0 mL with the same solvent. Prepare immediately before 0.025 M. Dissolve 2.64 g of Na2CO3 and 2.09 g of NaHCO3
use. in carbon dioxide-free water R and dilute to 1000.0 mL with
the same solvent. Store protected from atmospheric carbon
dioxide.
Potassium hydrogen phthalate 0.05 M. Dissolve 10.13 g of
C8H5KO4, previously dried for 1 h at 110 ± 2 °C, in carbon
dioxide-free water R and dilute to 1000.0 mL with the same Calcium hydroxide, saturated at 25 °C. Shake an excess of
solvent. calcium hydroxide R with carbon dioxide-free water R and
decant at 25 °C. Store protected from atmospheric carbon
dioxide.
Potassium dihydrogen phosphate 0.025 M + Disodium
hydrogen phosphate 0.025 M. Dissolve 3.39 g of KH2PO4 and
3.53 g of Na2HPO4, both previously dried for 2 h at 120 ± 2 °C, STORAGE OF BUFFER SOLUTIONS
in carbon dioxide-free water R and dilute to 1000.0 mL with Store buffer solutions in suitable chemically-resistant, airtight
the same solvent. containers, such as type I glass bottles or plastic containers
suitable for aqueous solutions.

EUROPEAN PHARMACOPOEIA 2.2.5. Relative density
1 c 1
01/2008:20205 f2 = = ´ 2
corrected 10.0 T2 m 4π
Hence :
æ M ρ ´ V ö÷
T 2 = çç + ÷ ´ 4π
2
çè c c ÷ø
2.2.5. RELATIVE DENSITY M = mass of the tube ;
The relative density of a substance is the ratio of the mass
d tt21 V = inner volume of the tube.
of a certain volume of a substance at temperature t1 to the Introduction of 2 constants A = c / (4π 2 ´ V ) and B = M / V ,
mass of an equal volume of water at temperature t2. leads to the classical equation for the oscillating transducer :
20
Unless otherwise indicated, the relative density d 20 is
ρ = A´T2-B
used. Relative density is also commonly expressed as d 420 . The constants A and B are determined by operating the
Density ρ20, defined as the mass of a unit volume of instrument with the U-tube filled with 2 different samples
the substance at 20 °C may also be used, expressed in of known density, for example, degassed water R and air.
kilograms per cubic metre or grams per cubic centimetre Control measurements are made daily using degassed water R.
(1 kg·m− 3 = 10− 3 g·cm− 3). These quantities are related by the The results displayed for the control measurement using
following equations where density is expressed in grams per degassed water R shall not deviate from the reference value
cubic centimetre : 20
(ρ20 = 0.998203 g·cm− 3, d 20 = 1.000000) by more than its
20 20
ρ20 = 0.998203 ´ d 20 or d 20 = 1.00180 ´ ρ20 specified error. For example, an instrument specified to
± 0.0001 g·cm− 3 shall display 0.9982 ± 0.0001 g·cm− 3 in
order to be suitable for further measurement. Otherwise a
ρ20 = 0.999972 ´ d 20 20
4 or d 4 = 1.00003 ´ ρ 20 re-adjustment is necessary. Calibration with certified reference
materials is carried out regularly. Measurements are made
d 420 = 0.998230 ´ d 20
20
using the same procedure as for calibration. The liquid to
Relative density or density is measured according to the be examined is equilibrated in a thermostat at 20 °C before
number of decimals prescribed in the monograph using introduction into the tube, if necessary, to avoid the formation
a density bottle (solids or liquids), a hydrostatic balance of bubbles and to reduce the time required for measurement.
(solids), a hydrometer (liquids) or a digital density meter Factors affecting accuracy include :
with an oscillating transducer (liquids and gases). When the – temperature uniformity throughout the tube ;
determination is made by weighing, the buoyancy of air is – non-linearity over a range of density ;
disregarded, which may introduce an error of 1 unit in the
3rd decimal place. When using a density meter, the buoyancy – parasitic resonant effects ;
of air has no influence. – viscosity, whereby solutions with a higher viscosity than
Oscillating transducer density meter. The apparatus consists of : the calibrant have a density that is apparently higher than
the true value.
– a U-shaped tube, usually of borosilicate glass, which The effects of non-linearity and viscosity may be avoided by
contains the liquid to be examined ; using calibrants that have density and viscosity close to those
– a magneto-electrical or piezo-electrical excitation system of the liquid to be examined (± 5 per cent for density, ± 50 per
that causes the tube to oscillate as a cantilever oscillator cent for viscosity). The density meter may have functions for
at a characteristic frequency depending on the density of automatic viscosity correction and for correction of errors
the liquid to be examined ; arising from temperature changes and non-linearity.
– a means of measuring the oscillation period (T), which may Precision is a function of the repeatability and stability of the
be converted by the apparatus to give a direct reading of oscillator frequency, which is dependent on the stability of the
density, or used to calculate density using the constants A volume, mass and spring constant of the cell.
and B described below. Density meters are able to achieve measurements with an
The resonant frequency (f) is a function of the spring error of the order of 1 × 10− 3 g·cm− 3 to 1 × 10− 5 g·cm− 3 and a
constant (c) and the mass (m) of the system : repeatability of 1 × 10− 4 g·cm− 3 to 1 × 10− 6 g·cm− 3.

EUROPEAN PHARMACOPOEIA 2.2.7. Optical rotation
07/2018:20207 – a temperature control system that indicates the temperature

with a readability of 0.1 °C ; it may be embedded in the
polarimeter (e.g. a Peltier system) or be an external unit
(e.g. a cycle-cryostat), and must be able to maintain
the temperature of the liquid to within ± 0.5 °C of that
prescribed.
2.2.7. OPTICAL ROTATION
EQUIPMENT PERFORMANCE
PRINCIPLE The accuracy of the scale is checked near the value to be
Optical rotation (also known as optical activity) is the measured or over an appropriate range, usually by means of
property displayed by chiral substances of rotating the plane certified quartz plates. Other certified reference materials may
of polarisation of linearly polarised light. also be suitable (e.g. sucrose solutions).
Optical rotation is considered to be positive (+) for Optical rotation measurements may be used to quantify the
dextrorotatory substances (i.e. those that rotate the plane amount of an enantiomer or the ratio of enantiomers present
of polarisation in a clockwise direction when viewed in the in a sample. For that purpose, the linearity must be checked,
direction facing the oncoming light beam) and negative (−) for example using sucrose solutions.
for laevorotatory substances (i.e. anticlockwise rotation).
PROCEDURE
The angle of optical rotation α of a liquid is the angle of Determine the zero of the polarimeter and the angle of rotation
rotation of the plane of polarisation, expressed in degrees (°), of the liquid at a wavelength of 589 nm and a temperature
at the wavelength of the D-line of sodium (λ = 589.3 nm) of 20 ± 0.5 °C, unless otherwise prescribed. The zero of the
measured at 20 °C through the liquid when using a path polarimeter is determined with the sample cell closed.
length of 1.00 dm.
For neat liquids, the zero is determined with an empty sample
The specific optical rotation [α]D20 of a substance in solution is cell.
calculated from the angle of optical rotation, as defined above, For solutions, the zero is determined with the sample cell
with reference to a path length of 1.00 dm and a concentration filled with the same solvent as that used for the solution to be
of the substance to be examined of 1 g/mL. The specific optical examined and measured at the same temperature. The sample
rotation of a substance in solution is always expressed with preparation procedure is prescribed in the monograph.
reference to a given solvent and concentration. Calculate the specific optical rotation at temperature t and
As some equipment may not use sodium lamps, the wavelength wavelength λ using the following formulae.
of measurement is given as 589 nm instead of 589.3 nm. For neat liquids, the density of the liquid is taken into account :
In certain cases specified in the monograph, the angle of α
[α]t =
optical rotation is measured at other temperatures, other λ l·ρt
wavelengths and/or in cells with a path length other than
1.00 dm. For solutions :
In the conventional system adopted by the Pharmacopoeia, the 1000α
[α]tλ =
specific optical rotation is expressed by its value without units ; l·c
the actual units, degree millilitres per decimetre gram α = angle of rotation measured at temperature t
[(°)·ml·dm− 1·g− 1] are understood.
and wavelength λ, in degrees ;
EQUIPMENT l = path length of the polarimeter sample cell, in
The polarimeter typically consists of : decimetres ;
ρt = density determined at the temperature of
– a light source, for example a sodium discharge lamp, a
measurement t, in grams per cubic centimetre ;
light-emitting diode (LED) or another light source capable
for the purposes of the Pharmacopoeia, density
of providing radiation at the desired wavelength (589 nm
is replaced by relative density (2.2.5);
unless otherwise prescribed in the monograph) ; if the light
source is polychromatic, a means of isolating the required c = concentration of the solution, in grams per
wavelength is necessary, e.g. an optical filter ; litre.
– a polariser and an analyser ;
– a sample cell with a path length of 1.00 dm, unless otherwise When the limits for optical rotation or specific optical rotation
specified in the monograph ; are expressed as the dried substance, the anhydrous substance
– a detection system to measure the angle of optical rotation, or the solvent-free substance, the result must be corrected for
which must be capable of giving readings to at least the loss on drying (2.2.32), water content (2.5.12 or 2.5.32) or
nearest 0.01°, unless otherwise specified in the monograph ; content of solvent as appropriate.

EUROPEAN PHARMACOPOEIA 2.2.13. Determination of water by distillation
01/2008:20213
2.2.13. DETERMINATION OF WATER

BY DISTILLATION
The apparatus (see Figure 2.2.13.-1) consists of a glass flask (A)
connected by a tube (D) to a cylindrical tube (B) fitted with
a graduated receiving tube (E) and reflux condenser (C). The
receiving tube (E) is graduated in 0.1 mL. The source of heat
is preferably an electric heater with rheostat control or an oil
bath. The upper portion of the flask and the connecting tube
may be insulated.
Method. Clean the receiving tube and the condenser of the
apparatus, thoroughly rinse with water, and dry.
Introduce 200 mL of toluene R and about 2 mL of water R into
the dry flask. Distil for 2 h, then allow to cool for about 30 min
and read the water volume to the nearest 0.05 mL. Place in the
flask a quantity of the substance, weighed with an accuracy
of 1 per cent, expected to give about 2 mL to 3 mL of water.
If the substance has a pasty consistency, weigh it in a boat of
metal foil. Add a few pieces of porous material and heat the
flask gently for 15 min. When the toluene begins to boil, distil
at the rate of about two drops per second until most of the
water has distilled over, then increase the rate of distillation to
about four drops per second. When the water has all distilled
over, rinse the inside of the condenser tube with toluene R.
Continue the distillation for 5 min, remove the heat, allow the
receiving tube to cool to room temperature and dislodge any
droplets of water which adhere to the walls of the receiving
tube. When the water and toluene have completely separated,
read the volume of water and calculate the content present in
the substance as millilitres per kilogram, using the formula :
1000(n2 - n1 )
m
m = the mass in grams of the substance to be examined,
n1 = the number of millilitres of water obtained in the
first distillation,
n2 = the total number of millilitres of water obtained in
the 2 distillations.
Figure 2.2.13.-1. – Apparatus for the determination of water

by distillation
Dimensions in millimetres

EUROPEAN PHARMACOPOEIA 2.2.14. Melting point - capillary method
04/2017:20214 Detection can be performed either visually or instrumentally.

In the case of instrumental detection, this is generally
performed by image recording and subsequent analysis or by
a photodetector that measures the transmitted or reflected
light from the sample.
Method. The substance is previously treated as described in
2.2.14. MELTING POINT - CAPILLARY the monograph. Coarse crystals are to be avoided as they
might lead to false results. If necessary, samples are crushed
METHOD into a fine powder. Unless otherwise prescribed, dry the finely
powdered substance in vacuo over anhydrous silica gel R for
The melting point determined by the capillary method is 24 h. Introduce a sufficient quantity into a capillary tube
the temperature at which the last solid particle of a compact to give a compact column as described by the instrument
column of a substance in a tube passes into the liquid phase manufacturer (e.g. 4-6 mm in height). Raise the temperature
(i.e. clear point). The melting point determined by this of the apparatus to about 5 °C below the presumed melting
method is specific to the methodology (e.g. heating rate) point. Allow the temperature to stabilise and then introduce
described in this chapter. Similarly, whenever the use of the capillary tube into the instrument. Finally, adjust the rate
certified reference materials is required, their certified values of heating to about 1 °C/min unless otherwise prescribed.
refer to the described analytical procedure. In the case of instrumental detection, follow the instrument
manufacturer’s requirements for the determination of the
When prescribed in the monograph, the same apparatus and
melting point. For visual detection, record the temperature at
method are used for the determination of other factors, such
which the last particle of the substance to be examined passes
as meniscus formation or melting range, that characterise the
into the liquid phase.
melting behaviour of a substance.
Samples can be measured in parallel if the instrument allows
Equipment. The equipment consists of a metal heating block multiple sample processing.
with 1 or more compartments for capillary tubes, or of a System suitability. Carry out a system suitability test before the
suitable glass vessel containing a liquid bath (e.g. water, measurements for example by choosing a suitable reference
liquid paraffin or silicone oil) and fitted with a suitable means material with a melting point close to that expected for the
of heating and stirring. The equipment is equipped with substance to be examined.
a temperature sensor or a suitable certified thermometer Qualification / Calibration of the equipment. The qualification
allowing readings at least to the nearest 0.1 °C. / calibration is carried out periodically according to the
Samples are introduced into the equipment in glass instrument manufacturer’s requirements, using at least
capillary tubes. The dimensions are chosen according to 2 certified reference materials. These are selected to cover
the manufacturer’s requirements, typically with an external the temperature range that is used on the equipment. Use
diameter of 1.3-1.5 mm and a wall thickness of 0.1-0.3 mm. In capillary tubes with the same dimensions as those used for
some equipment glass slides are used instead of capillary tubes. sample measurement.
Guidance on how to compare results obtained from certified
The equipment is capable of heating samples at a rate of reference materials with values from the certificates can be
1 °C/min or less. The accuracy of the equipment is at most found on the European Reference Materials (ERM) website
± 0.5 °C. (Application note 1).

EUROPEAN PHARMACOPOEIA 2.2.15. Melting point - open capillary method
01/2008:20215 Unless otherwise prescribed, substances with a waxy

consistency are carefully and completely melted on a
water-bath before introduction into the capillary tubes. Allow
the tubes to stand at 2-8 °C for 2 h.
2.2.15. MELTING POINT - OPEN Attach one of the tubes to a thermometer graduated in 0.5 °C
CAPILLARY METHOD so that the substance is close to the bulb of the thermometer.
Introduce the thermometer with the attached tube into a
For certain substances, the following method is used to beaker so that the distance between the bottom of the beaker
determine the melting point (also referred to as slip point and and the lower part of the bulb of the thermometer is 1 cm.
rising melting point when determined by this method). Fill the beaker with water to a depth of 5 cm. Increase the
Use glass capillary tubes open at both ends, about 80 mm temperature of the water gradually at a rate of 1 °C/min.
long, having an external diameter of 1.4 mm to 1.5 mm and
an internal diameter of 1.0 mm to 1.2 mm. The temperature at which the substance begins to rise in the
Introduce into each of 5 capillary tubes a sufficient amount of capillary tube is regarded as the melting point.
the substance, previously treated as described, to form in each
tube a column about 10 mm high and allow the tubes to stand Repeat the operation with the other 4 capillary tubes and
for the appropriate time and at the prescribed temperature. calculate the result as the mean of the 5 readings.

EUROPEAN PHARMACOPOEIA 2.2.16. Melting point - instantaneous method
07/2015:20216 maintained in the same position during the calibration of the

apparatus and the determination of the melting point of the
substance to be examined. The cylindrical cavity is parallel to
the upper polished surface of the block and about 3 mm from
it. The apparatus is calibrated using appropriate substances of
2.2.16. MELTING POINT - known melting point.
INSTANTANEOUS METHOD Method. Heat the block at a suitably rapid rate to a temperature
about 10 °C below the presumed melting temperature, then
The instantaneous melting point is calculated using the adjust the heating rate to about 1 °C/min. At regular intervals
following expression : drop a few particles of powdered and, where appropriate, dried
substance, prepared as for the capillary tube method, onto the
t1 + t2 block in the vicinity of the thermometer bulb, cleaning the
2 surface after each test. Record the temperature t1 at which the
in which t1 is the first temperature and t2 the second substance melts instantaneously for the first time in contact
temperature read under the conditions stated below. with the metal. Stop the heating. During cooling drop a few
Apparatus. The apparatus consists of a metal block resistant particles of the substance at regular intervals on the block,
to the substance to be examined, of good heat-conducting cleaning the surface after each test. Record the temperature t2
capacity, such as brass, with a carefully polished plane upper at which the substance ceases to melt instantaneously when it
surface. The block is uniformly heated throughout its mass by comes in contact with the metal.
means of a micro-adjustable gas heater or an electric heating Calibration of the apparatus. The apparatus may be calibrated
device with fine adjustment. The block has a cylindrical cavity, using melting point reference substances such as those of the
which is wide enough to accomodate a thermometer that is World Health Organization or other appropriate substances.

EUROPEAN PHARMACOPOEIA 2.2.19. Amperometric titration
01/2016:20219 platinum electrode, a rotating-disc electrode or a carbon

electrode) and a reference electrode (for example, a silver-silver
chloride electrode).
A three-electrode apparatus is sometimes used, consisting of
an indicator electrode, a reference electrode and a polarised
auxiliary electrode.
2.2.19. AMPEROMETRIC TITRATION Method. Set the potential of the indicator electrode as
prescribed and plot a graph of the initial current and the
In an amperometric titration, the end-point is determined values obtained during the titration as functions of the
by following the variation of the current measured between quantity of titrant added. Add the titrant in not fewer than
2 electrodes (either one indicator electrode and one reference 3 successive quantities equal to a total of about 80 per cent
electrode or 2 indicator electrodes) immersed in the solution of the theoretical volume corresponding to the presumed
to be examined and maintained at a constant potential equivalence point. The 3 values must fall on a straight line.
difference as a function of the quantity of titrant added. Continue adding the titrant beyond the presumed equivalence
point in not fewer than 3 successive quantities. The values
The potential of the measuring electrode is sufficient to ensure obtained must fall on a straight line. The point of intersection
a diffusion current for the electroactive substance. of the 2 lines represents the end-point of the titration.
Apparatus. The apparatus comprises an adjustable voltage For amperometric titrations with 2 indicator electrodes, the
source and a sensitive microammeter ; the detection system whole titration curve is recorded and used to determine the
generally consists of an indicator electrode (for example, a end-point.

EUROPEAN PHARMACOPOEIA 2.2.20. Potentiometric titration
01/2016:20220 Method. Prepare the sample solution as described. Add

the titrant in suitable aliquots paying particular attention
to the rate of addition and the volume increments near the
end-point. Continue the titration beyond this point to allow a
clear detection of the end-point.
2.2.20. POTENTIOMETRIC TITRATION
The end-point of the titration is reached when the maximum
In a potentiometric titration (volumetric titration with change in potential occurs in a plot of potential versus
potentiometric end-point determination) the end-point volume of titrant, and is expressed as the corresponding
is determined by recording the variation of the potential volume of titrant. Recording the first or second derivative
difference between 2 electrodes (either 1 indicator electrode curve can facilitate the determination of the end-point.
and 1 reference electrode, or a combined electrode) immersed In potentiometric titrations of weak acids or bases using
in the solution to be examined as a function of the volume non-aqueous solvents, if necessary, either carry out a blank
of titrant added. determination or pre-neutralise the solvent mixture. Where
it is impracticable to use potentiometric detection for this
Apparatus. The apparatus used comprises a millivoltmeter. purpose, the solvent mixture can be pre-neutralised by
Commercial autotitrator instruments may be used and are titration using a suitable indicator. Some examples are given
operated in accordance with the manufacturer’s instructions, below :
using electrodes recommended for the type of titration
described.
Titrant Indicator
The indicator electrode to be used depends on the substance
to be determined and may be a glass or metal electrode Perchloric acid Crystal violet solution R
(e.g. platinum, gold or silver). 3 g/L solution of thymol blue R
Tetrabutylammonium hydroxide
in methanol R
For acid-base titrations, a glass-silver-silver chloride electrode
Ethanolic sodium hydroxide Thymolphthalein solution R
combination is generally used.

EUROPEAN PHARMACOPOEIA 2.2.21. Fluorimetry
01/2008:20221 the second adjustment is made by altering the width of the

slits, a new zero setting must be made and the intensity of
the standard must be measured again. Finally introduce the
solution of unknown concentration and read the result on the
instrument. Calculate the concentration cx of the substance in
2.2.21. FLUORIMETRY the solution to be examined, using the formula :
Fluorimetry is a procedure which uses the measurement of the Ixcs

cx =
intensity of the fluorescent light emitted by the substance to Is
be examined in relation to that emitted by a given standard.
cx = concentration of the solution to be examined,
Method. Dissolve the substance to be examined in the solvent
or mixture of solvents prescribed in the monograph, transfer cs = concentration of the standard solution,
the solution to the cell or the tube of the fluorimeter and
Ix = intensity of the light emitted by the solution to be
illuminate it with an excitant light beam of the wavelength
prescribed in the monograph and as near as possible examined,
monochromatic. Is = intensity of the light emitted by the standard
Measure the intensity of the emitted light at an angle of 90° solution.
to the excitant beam, after passing it through a filter which If the intensity of the fluorescence is not strictly proportional
transmits predominantly light of the wavelength of the to the concentration, the measurement may be effected using
fluorescence. Other types of apparatus may be used provided a calibration curve.
that the results obtained are identical. In some cases, measurement can be made with reference
For quantitative determinations, first introduce into the to a fixed standard (for example a fluorescent glass or a
apparatus the solvent or mixture of solvents used to dissolve solution of another fluorescent substance). In such cases,
the substance to be examined and set the instrument to zero. the concentration of the substance to be examined must be
Introduce the standard solution and adjust the sensitivity determined using a previously drawn calibration curve under
of the instrument so that the reading is greater than 50. If the same conditions.

EUROPEAN PHARMACOPOEIA 2.2.24. Absorption spectrophotometry, infrared
04/2019:20224 – quantification of active substances in a sample matrix,

determination of water and solvent content ;
– quantification of impurities, e.g. in gases, inorganic
materials ;
– reaction monitoring, e.g. chemical synthesis.
2.2.24. ABSORPTION Physical analysis :
SPECTROPHOTOMETRY, INFRARED – determination of solid-state properties such as
polymorphism.
PRINCIPLE
Infrared absorption spectrophotometry (also known as LIMITATIONS
infrared (IR) spectroscopy) is based on the interaction of Notable limitations to the use of IR spectroscopy include the
infrared radiation with matter, which affects the vibrational following :
energy of molecules and induces intermolecular and
intramolecular vibrations at specific frequencies. This results – it may be necessary to use additional techniques to
in an infrared absorption spectrum with characteristic bands unambiguously identify a substance ;
that correspond to the functional groups of the molecule. – pure enantiomers of a substance cannot be discriminated ;
The infrared wavelength region can be further divided – it may not be a suitable method for trace analysis ;
into 3 subregions, namely near-infrared, mid-infrared and – sample preparation conditions (e.g. pressure, solvent) may
far-infrared. These subregions have wavelength ranges that are change the crystalline form of a substance that exhibits
generally accepted by convention to be 0.8-2.5 µm, 2.5-25 µm polymorphism ;
and 25-1000 µm respectively. However, in IR spectroscopy,
wavenumber is more commonly used than wavelength, and – for heterogeneous samples, the limited sampling volume
can be calculated using the following equation : may be problematic.
1 MEASUREMENT MODES
ν = ·104
λ IR measurements are based on passing radiation through or
where ν is the wavenumber in reciprocal centimetres into a sample and measuring the attenuation of the emerging
(cm-1) and λ is the wavelength in micrometres. Thus beam at various wavelengths. This corresponds to 2 main
12 500-4000 cm− 1 is near-infrared, 4000-400 cm− 1 is measurement modes, i.e. transmission and attenuated total
mid-infrared and 400-10 cm− 1 is far-infrared. reflection (ATR). However, other modes also exist for specific
applications (e.g. diffuse and specular reflection).
This chapter concerns only spectroscopy in the mid-infrared
region, i.e. 4000-400 cm− 1 (2.5-25 µm), which hereafter is TRANSMISSION MODE
referred to as infrared for simplicity. This region is where This mode is based on determination of the transmittance (T),
the fundamental molecular vibrations of functional groups namely the ability of the sample to transmit IR radiation at a
appear in the spectrum as absorption bands. The region below given wavelength (wavenumber). It is defined by the following
1500 cm− 1 is known as the ‘fingerprint region’, a very complex ratio :
and informative part of the spectrum which characterises the I
molecule being investigated. T=
I0
The mid-infrared region is flanked by the near-infrared
region, where overtones and combinations of fundamental I0 = intensity of incident radiation ;
vibrations, mainly C-H, N-H and O-H functional groups, are I = intensity of transmitted radiation.
detected (2.2.40) and the far-infrared region, where absorption
bands associated with crystal lattice modes, hydrogen bonds, The resulting spectrum is presented in terms of transmittance
angle deformation vibrations of heavy atoms and molecular (T) on the y-axis versus wavelength or wavenumber on the
rotations are observed. x-axis. It can also be presented in terms of absorbance (A)
APPLICATIONS on the y-axis, which is related to transmittance (T) by the
following equation :
As the absorption bands in IR spectra are characteristic of the
constituent functional groups of a compound, IR spectroscopy æ1 ö æI ö
is widely used to identify substances and provide information A = log10çç ÷÷ = log10çç 0 ÷÷÷ = a·b·c
çèT ÷ø çè I ÷ø
on their structure. It can also be used for quantitative
applications, which requires establishing a mathematical a molar absorption coefficient of the sample, in
relationship between the intensity of the radiation absorbed = square centimetres per mole (cm2·mol-1) ;
by the sample and the concentration of the investigated b = sample thickness, in centimetres ;
component in the sample. c = sample concentration, in moles per cubic
IR spectroscopy is widely used in the pharmaceutical field centimetre (mol·cm-3).
for chemical and physical analysis in the laboratory, and
has a wide variety of applications during the manufacturing ATTENUATED TOTAL REFLECTION MODE
process as outlined below. IR spectroscopy thereby enables the ATR mode is based on the phenomenon of total internal
application of Process Analytical Technology (PAT) as part of reflection. The sample, with a refractive index n2, is brought
an advanced control strategy. into close contact with a crystal (diamond, germanium, zinc
Chemical analysis : selenide or any other suitable material), having a refractive
index n1 which is greater than n2. A beam of IR light is then
– identification of active substances, excipients, dosage forms, passed through the crystal. When the angle α between the
manufacturing intermediates, chemicals and packaging incident beam and the sample-crystal interface exceeds a
materials ; critical value αc, theoretically all of the radiation is reflected
– quality assessment of active substances, excipients, dosage (total internal reflection). However, an evanescent wave is
forms, manufacturing intermediates and packaging produced which slightly penetrates the sample and part of the
materials, including batch-to-batch spectral comparison energy is absorbed. The total reflection is attenuated, which
and supplier change assessment ; makes it possible to generate an absorption spectrum. In

2.2.24. Absorption spectrophotometry, infrared EUROPEAN PHARMACOPOEIA
practice, multiple internal reflections are often used to amplify SPECTRAL RESOLUTION
the absorption intensity, although some accessories allow
absorption measurements with a single reflection.
The penetration depth dp is usually of the order of a few
micrometres and is given for a wavelength λ by the following
equation :
λ / n1
dp =
2π sin α - (n2 / n1 )2
2
where dp is the penetration depth, λ is the wavelength, α is the

angle of incidence and n1, n2 are the refractive indices of the
reflection element and the sample, respectively.
Due to the relationship between these parameters, the
absorption intensity in ATR is greater at higher wavelengths
(i.e. smaller wavenumbers) and slight band shifts occur
compared to the corresponding transmission spectrum.
It is therefore not advisable to compare ATR spectra with
transmission spectra when identifying compounds.
EQUIPMENT
The most commonly used IR spectrometers are
Fourier-transform (FT-IR) spectrometers which typically
consist of :
– a suitable polychromatic light source, e.g. a conducting
ceramic rod ; Figure 2.2.24.-1. – Typical IR absorbance spectrum of
– an interferometer ; polystyrene used to verify spectral resolution
– a sample presentation accessory, e.g. a sample holder ; Spectra recorded in transmission mode. The spectral
– a detector ; resolution is typically verified using a polystyrene film
– appropriate software for controlling the spectrometer, and approximately 35 µm thick.
for spectral evaluation and data processing. Acceptance criteria (see Figure 2.2.24.-1) : the difference
Other spectrometers based on alternative principles may between the absorbance values at the absorption minimum at
also be used if the requirements described under Control of 2870 cm-1 (A) and the absorption maximum at 2849.5 cm-1 (B)
equipment performance are fulfilled. is greater than 0.33 ; the difference between the absorbance
values at the absorption minimum at 1589 cm-1 (C) and the
IR spectrometers can also be used in association with a
absorption maximum at 1583 cm-1 (D) is greater than 0.08.
microscope for the study of a small part of the sample or for
chemical imaging. Spectra recorded in ATR mode. Appropriate assessment
IR spectroscopy can be coupled to other analytical techniques criteria for the control of spectral resolution according to the
such as thermal analysis or chromatography. specifications of each instrument need to be defined.
For measurement modes other than transmission or ATR,
CONTROL OF EQUIPMENT PERFORMANCE reference materials have to be defined by the user.
Accuracy of wavenumber scale and spectral resolution are
PROCEDURE
critical parameters and must be verified. The tests described
below can be used for the control of instrument performance SAMPLE PREPARATION AND PRESENTATION
and for qualification. They can also be used as system Sample preparation and presentation vary according to the
suitability tests. physical state of the sample and the measurement mode.
These parameters are checked using suitable reference Transmission mode is applied to transparent samples, such
materials which are selected and presented depending on the as neat liquids, solutions, gases or suitably prepared mulls
measurement mode (e.g. transmission or ATR). and alkali halide discs. For liquids and gases, cells with fixed
For quantitative analysis, appropriate assessment criteria for or variable pathlength and IR transparent windows can be
the control of absorption intensity must also be defined. used. For alkali-halide disks, specific sample holders are used.
Reflection mode, e.g. ATR, is appropriate for the measurement
WAVENUMBER SCALE of a wide range of samples in the solid and liquid state.
The wavenumber scale is typically verified using a polystyrene Some preparation modes (e.g. for discs and mulls in
film that exhibits IR absorption bands at the wavenumbers transmission mode or for solids in ATR mode) involve
shown in Table 2.2.24.-1. grinding and/or the application of pressure, which may induce
Table 2.2.24.-.1 - Band positions and associated acceptable unexpected crystal modifications.
tolerances of the polystyrene film used to verify wavenumber Transmission mode
accuracy Prepare the substance by one of the following methods
Band position (cm−1) depending on the sample state (solid, liquid or gas). If sample
Tolerance (cm−1) bands in a spectrum have a minimum transmission lower than
Transmission ATR
5 per cent, this spectrum is not used for further data analysis.
906.6 906.1 ± 1.0 Liquids. Examine liquids either in the form of a film between
1028.3 1027.7 ± 1.0
2 plates transparent to infrared radiation or in a cell of suitable
pathlength with windows that are transparent to infrared
1601.2 1601.0 ± 1.0 radiation.
3060.0 3059.7 ± 1.0 Liquids or solids in solution. Prepare a solution of the substance
to be examined in a suitable solvent. Choose a concentration
For measurement modes other than transmission or ATR, and a pathlength that give a satisfactory spectrum. Generally,
reference materials must be defined by the user. good results are obtained with concentrations of 10-100 g/L

EUROPEAN PHARMACOPOEIA 2.2.24. Absorption spectrophotometry, infrared
for a pathlength of 0.5-0.1 mm. The absorption due to the Solids. Ensure close and uniform contact between the
solvent is usually compensated by successively recording the substance to be examined and the whole crystal surface,
spectra of the solvent and the sample solution and subtracting either by applying pressure or by dissolving the substance
the solvent absorption bands from the spectrum of the sample in an appropriate solvent, then covering the crystal with the
solution. resulting solution and evaporating to dryness.
Solids dispersed in a solid (disc). Grind the substance to be METHODS
examined taking into consideration any possible changes Infrared spectroscopy is mostly used to identify substances,
(e.g. crystalline form) and mix with a suitable amount of but it may also be carried out for quantitative applications.
finely powdered and dried potassium bromide R or potassium Quantitative analysis (based on the Beer-Lambert law, which
chloride R, unless otherwise specified. A mixture of a few relates the absorbance of a sample to its concentration) will
milligrams (e.g. 1-2 mg) of the substance to be examined not be described in this chapter.
in a few hundred milligrams (e.g. 300-400 mg) of halide The measurement is performed on an appropriately prepared
is normally sufficient to give a disc of 10-15 mm diameter sample. The data is then processed and evaluated, either to
and a spectrum of suitable intensity. If the substance is a identify substances or quantify them (e.g. based on integration
hydrochloride salt, it is recommended to use potassium of IR-absorption bands).
chloride R. Carefully grind the mixture, spread it uniformly Spectral quality may be enhanced by mathematical
in a suitable die and apply a suitable pressure. A compacting pretreatments. In practice, these are limited to spectral
force of about 800 MPa is generally sufficient to prepare a disc. normalisation and subtraction of bands caused by
For substances that are unstable under normal atmospheric carbon-dioxide and water vapour. The same pretreatments are
conditions or are hygroscopic, the disc may be pressed under performed on both the sample and the reference spectra.
vacuum. Several factors may cause the formation of faulty
discs, such as insufficient or excessive grinding, humidity Identification
or impurities in the dispersion medium. For example, any Prepare the substance to be examined appropriately and record
water in either the sample or the potassium bromide will the spectra between 4000 and 650 cm-1, unless otherwise
cause clouding of the disc and produce a low transmission prescribed.
spectrum. A disc is rejected if visual examination shows a lack
of uniform transparency or when, in the absence of a specific Identification testing is performed by comparing the spectrum
absorption band, the transmittance is less than 60 per cent or of the substance to be examined with the spectrum obtained
the absorbance is more than 0.22 at about 2000 cm-1 (5 μm) from a Ph. Eur. chemical reference substance (CRS) or with a
and without compensation, unless otherwise prescribed. Ph. Eur. reference spectrum.
Solids dispersed in a liquid (mull). Triturate a small quantity of The spectrum of the current batch of the Ph. Eur. CRS may
the substance to be examined with the minimum quantity of be recorded for immediate use or stored, for example, in a
liquid paraffin R or other suitable liquid. A mixture of a few spectral library for future consultation. A stored spectrum
milligrams (e.g. 5-10 mg) of the substance to be examined may be used, provided traceability to the current batch of CRS
in 1 drop of liquid paraffin R is generally sufficient to make is ensured.
an adequate mull. Compress the mull between 2 plates In the case of substances that are not covered by individual
transparent to infrared radiation. A mull is rejected if a visual monographs, a suitable reference standard may be used.
examination shows lack of uniform transparency or where the In all cases, spectra must be recorded using the same
spectrum shows features such as : operating conditions and procedure, and especially the same
– low transmission at 4000 cm-1 ; measurement mode.
– a strongly sloping baseline between 4000 and about When comparison of the spectra recorded in the solid
2500 cm-1 ; state show differences (see below), treat the substance to be
examined and the reference substance in the same manner so
– a ratio of relative intensities of some absorption bands that that they recrystallise or are produced in the same crystalline
is less than expected. form, or proceed as prescribed in the monograph, then record
the spectra again. However, this procedure must only be
Molten solids. If prescribed in the monograph, make a film of done for substances where the monograph does not cover a
a molten mass and fix it on a suitable mount. particular form of a substance that exhibits polymorphism.
Evaporated solution. If prescribed in the monograph, dissolve Several comparison procedures may be used, and the analyst
the substance to be examined in a suitable solvent. Prepare a must document and justify the method used and the specific
film by evaporating the solvent on a suitable carrier and fix acceptance criteria that allow a conclusion for identification.
it on a suitable mount. The spectra can be compared either by overlaying the spectra
(in the whole spectral range or in the region of interest
Gases. Use a suitable cell transparent to infrared radiation. specified in the monograph) or by using mathematical
Evacuate the air from the cell and fill to the desired pressure calculations from the software. It is possible for example to
through a stopcock or needle valve using a suitable gas perform :
transfer line between the cell and the container of the gas
to be examined. If necessary, adjust the pressure in the cell – visual comparison based on band positions and relative
to atmospheric pressure using a gas transparent to infrared intensities unless otherwise specified - the transmission
radiation (e.g. nitrogen R or argon R), or purge with carbon minima (or absorption maxima) in the spectrum obtained
dioxide-free air. An appropriate measurement protocol must with the substance to be examined correspond in position
be followed to compensate for water, carbon dioxide or other and relative size to those of the reference ;
atmospheric gases. – calculation of the correlation coefficient between the 2
ATR mode spectra - this value is calculated by the software and the
identification threshold is defined by the user ;
ATR is suitable for liquid and solid samples, and requires no
preparation apart from simple treatments such as the grinding – evaluation by chemometric methods (e.g. Euclidean
of large crystals and coarse material. Proceed as follows distance, Mahalanobis distance, classification methods) ;
depending on the sample state (liquid or solid). these methods involve the set-up, assessment and validation
of the chemometric model by the analyst (see 5.21.
Liquids. Place the sample in contact with the crystal. Chemometric methods applied to analytical data).

2.2.24. Absorption spectrophotometry, infrared EUROPEAN PHARMACOPOEIA
Impurities in gases Fill the cell as prescribed under Gases. For detection and
For the analysis of impurities, use a cell transparent to infrared quantification of the impurities, proceed as prescribed in the
radiation and of suitable optical pathlength (e.g. 1-20 m). monograph.

EUROPEAN PHARMACOPOEIA 2.2.25. Absorption spectrophotometry, ultraviolet and visible
01/2020:20225 – a single-channel (e.g. photomultiplier, photodiode) or

multi-channel detector (e.g. photodiode array (PDA) or
charge-coupled device (CCD)) ;
– suitable computerised data processing and evaluation
systems.
2.2.25. ABSORPTION Control of cuvettes. For benchtop instruments, cuvettes or
SPECTROPHOTOMETRY, cells with a defined path length are used. These can be made
of different materials such as quartz or glass. The tolerance for
ULTRAVIOLET AND VISIBLE the path length of quartz and glass cuvettes is ± 0.5 per cent
PRINCIPLE (e.g. ± 0.005 cm for a 1 cm cuvette). Plastic cuvettes may also
be used but the tolerance interval is wider ; therefore, their use
Ultraviolet and visible (UV-Vis) spectroscopy (or
must be thoroughly justified and based on a risk assessment.
spectrophotometry) is based on the ability of atoms,
molecules and ions to absorb light at specific wavelengths The following method may be applied to check the cleanliness
in the ultraviolet (approximately 180-400 nm) and visible of optical cuvette windows and any significant differences in
(approximately 400-800 nm) range. This absorption is their thickness or parallelism : fill the cuvette with water R
associated with changes in electronic energy in the form of and measure its apparent absorbance against air at 240 nm
temporary transitions of electrons to an excited state at a for quartz cuvettes and 650 nm for glass cuvettes ; rotate
higher energy orbital. As each energy level of a molecule or the cuvette 180° in its holder and measure the apparent
molecular ion also has associated vibrational and rotational absorbance again at the same wavelength.
sub-levels, this results in many permitted transitions, which When using scanning instruments, it is recommended to scan
are generally impossible to separate, thereby producing over the optical region of interest.
absorption bands rather than sharp lines. These bands When using double-beam spectrophotometers, measures
are characteristic of the functional groups and bonds in a should be taken (e.g. matching the cuvettes) to ensure that
molecule. any difference between the absorbance of the cuvettes will not
UV-Vis spectroscopy measurements involve exposing a sample have a significant impact on the analysis to be performed.
to light and measuring the attenuation and/or scattering of Acceptance criteria :
the emerging (transmitted or reflected) light either at a single – the apparent absorbance is not greater than 0.093 for 1 cm
wavelength or over a specified wavelength range. quartz cuvettes (UV region) and 0.035 for 1 cm glass
APPLICATIONS cuvettes (visible region) ;
– the absorbance measured after rotation (180°) does not
UV-Vis spectroscopy is traditionally used for the quantitative
differ by more than 0.005 from the value previously
and qualitative analysis of liquid samples, but is also suitable
obtained.
for solid and gaseous analytes and has other applications such
as the determination of physical or chemical properties. MEASUREMENT
UV-Vis spectroscopy as described in this chapter can be Transmission mode. Transmission mode provides a measure
applied in various ways : of the transmittance (T), at a given UV-Vis wavelength, of
– when a monograph or general chapter refers to this chapter, a sample placed between the light source and the detector.
the requirements described in the relevant paragraphs of Transmittance is the ratio of the intensity of the transmitted
this chapter are mandatory ; light to the intensity of the incident light and is given by the
– when used as the detection method in chromatographic following equation :
systems as described in general chapter 2.2.46, the I
requirements listed in the relevant paragraphs of this T=
chapter are mandatory ; I0
– when used as a process analytical technology (PAT) tool I = intensity of transmitted radiation ;
for PAT applications similar to the applications described
in this chapter, the provisions herein apply ; for other I 0 = intensity of incident radiation.
PAT applications, the principles are the same, however A spectrum may be obtained by plotting the variation
the criteria are established bearing in mind the intended in transmittance (T) or absorbance (A) as a function of
purpose of the analysis, using a risk-based approach. wavelength.
EQUIPMENT The absorbance is defined as the logarithm to base 10 of the
Spectrophotometers used for carrying out measurements in reciprocal of the transmittance for monochromatic radiation.
the UV-Vis region typically consist of : It is a dimensionless quantity expressed in absorbance units
(AU), given by the following equation :
– a suitable light source (such as a deuterium lamp for the
UV region, a tungsten-halogen lamp for the visible region æ1 ö æI ö
or a xenon lamp to cover the entire UV-Vis range) ; UV-Vis A = log10çç ÷÷ = log10çç 0 ÷÷÷
çèT ÷ø çè I ÷ø
spectrophotometers often have 2 sources ;
According to the Beer-Lambert law, which applies to
– a monochromator such as a grating system ; clear diluted solutions, in the absence of interfering
– other optical components, such as lenses or mirrors, physico-chemical factors, the absorbance (A) is proportional
that relay light through the instrument and that may to the path length (l) of the radiation through the sample,
also be used to generate more than one beam of light, and to the concentration (c) of the substance in solution in
i.e. in double-beam spectrophotometers, as opposed to accordance with the following equation :
single-beam spectrophotometers ;
A = εcl
– a sample container, holder or sampling device ; examples
include conventional cuvettes, fibre-optic probes and ε = molar absorption coefficient, in litres per mole per
immersed transmission cells (e.g. high-purity quartz or centimetre ;
sapphire transparent to UV-Vis radiation) ; the choice c = molar concentration of the substance in solution,
depends on the intended application, paying particular in moles per litre ;
attention to its suitability for the type of sample to be
analysed ; l = absorption path length, in centimetres.

2.2.25. Absorption spectrophotometry, ultraviolet and visible EUROPEAN PHARMACOPOEIA
The specific absorbance ( A11 cm

per cent
) of the substance is Before an absorbance measurement is carried out, the zero
generally used in monographs and is related to absorbance position of the absorption (baseline correction) should be
(A) as follows : set or determined for the wavelengths of interest or over the
appropriate range of wavelengths.
1 per cent
A = A1 cm ´ cm ´ l For PAT applications, when measuring moving materials or
cm = mass concentration of the substance in solution, in samples, ensure that there is no fouling of the sensor (e.g. no
grams per 100 millilitres. contamination or build-up of material).
Unless otherwise prescribed in the monograph, measure the
absorbance using a path length of 1 cm at the prescribed
A11 cm
per cent
represents the specific absorbance of a dissolved wavelength. If a single value for the position of an absorption
substance and refers to the absorbance of a 1 g/100 mL (or maximum or minimum is given in a monograph, the user must
1 per cent m/V) solution in a 1 cm cuvette or cell and is determine the wavelength position. The value obtained may
measured at a defined wavelength. The relationship between differ by not more than ± 2 nm, unless otherwise prescribed.
1 per cent
A1 cm and ε is :
Quantitative measurements relying on absorption values
10 ε above 2.0 should be avoided.
A11 cm
per cent
=
Mr Background correction. Select a suitable spectroscopic blank
(e.g. air, blank solvent, solid material). Unless otherwise
Mr = relative molecular mass. prescribed, all measurements are carried out with reference to
the same solvent or the same mixture of solvents (blank).
Transmittance or absorbance measurements are generally used
for liquids (dispersions and solutions), but can also be used Measure the blank and the sample within a short time-frame
for solids (including tablets and capsules). For measurements either in parallel in double-beam spectrophotometers
of solids, a suitable sample accessory is used. Liquid samples or sequentially in single-beam spectrophotometers. The
are examined using a cell or cuvette with a suitable path absorbance values of both blank and sample must be in
length (typically 0.01-1 cm) and made of a material that is the working range of the equipment as specified by the
transparent to UV-Vis radiation, or by using a fibre-optic manufacturer.
probe of a suitable configuration immersed in the liquid. For benchtop instruments, the absorbance of the solvent
Diffuse reflection mode. Diffuse reflection mode provides measured against air and at the prescribed wavelength must
a measure of reflectance (R), which is given by the following not exceed 0.4 and is preferably less than 0.2.
equation : For chromatographic systems, the transmittance of the mobile
phase may be used as the blank.
I
R= In some PAT applications, it may be impossible to remove
I0 the probe for background data collection. Various options
are therefore to be considered, including the use of internal
I = intensity of light reflected and/or scattered from
references, measurement of a blank using a second detector,
the sample ; etc. Only spectra measured against a blank possessing the
I0 = intensity of light reflected and/or scattered from a same optical properties can be directly compared with one
blank or reference reflective surface. another.
Depending on the chemical composition and physical For reflectance measurements, common reflectance
characteristics of the sample, the UV-Vis radiation may be blank samples include ceramics, fluoropolymers such as
absorbed as it passes through the sample. In diffuse reflection polytetrafluoroethylene (PTFE) and powders such as barium
mode, it is the non-absorbed radiation which is partially sulfate (BaSO4) and magnesium oxide (MgO), but other
reflected and/or scattered back from the sample that is suitable materials may also be used.
measured by the detector. UV-Vis reflectance spectra are
typically obtained by calculating and plotting log10(1/R) as a MATHEMATICAL TREATMENT OF SPECTRAL DATA
function of the wavelength. In the case of single wavelength analysis used to determine the
This measurement mode is generally selected for solids. concentration of an unknown sample (e.g. as prescribed in
The sample is examined either in a suitable device (e.g. a monographs), mathematical treatment consists in determining
sample holder) or in direct contact with a probe. For process the regression of the photometric reading (absorbance) on the
monitoring, the material can be analysed through a polished concentration of the standard samples.
window interface (e.g. quartz or sapphire), or in-line using a In the case of full range spectra, data for both diffuse reflection
probe. Care must be taken to ensure that the measurement and transmission modes may have to be treated before a
conditions are as reproducible as possible from one sample classification or calibration model can be developed. The aim
to another. can be, for example, to reduce baseline drift or to correct for
Operation of the equipment. The factors below affect the scatter caused by particle size changes in solid samples. For
spectral response and must always be taken into account. example, first-, second- or higher-order derivative spectra
Choose a measurement mode that is appropriate for the can typically be used to improve resolution or sensitivity.
intended application and the sample type. This pretreatment may be a useful means of simplifying the
data and thereby reducing the variations that may cause
Define the measuring conditions taking into account interference in subsequently applied mathematical models.
the sample size and sample probe in such a way as to
obtain a satisfactory signal-to-noise ratio (e.g. beam size, A wide range of treatment methods, such as scaling,
measurement time and number of measurements). For smoothing, normalisation and derivatisation, can be applied
scanning spectrophotometers, also select the scan range, either singly or in combination. More information is available
scan rate and slit-width that provide the necessary optical in general chapter 5.21. Chemometric methods applied to
resolution for the intended application without losing the analytical data.
required signal-to-noise ratio or the linearity of the analytical
method. When using spectrophotometers with array sensors, CONTROL OF EQUIPMENT PERFORMANCE
there is no need to adjust the beam size, scan range, scan rate Spectrophotometer performance is controlled (automatically
or slit-width since the optical resolution is typically fixed and or manually) at regular intervals as defined in the quality
the full spectrum is always recorded. management system and dictated by the use of the equipment

EUROPEAN PHARMACOPOEIA 2.2.25. Absorption spectrophotometry, ultraviolet and visible
and the application. For example, equipment exposed to Wavelength accuracy, absorbance accuracy and linearity are
variations in temperature and humidity may need more controlled using either certified reference materials such as
frequent performance testing. solid filters or liquid filters in appropriate sealed cells, or
Requirements for control of equipment performance solutions prepared in the laboratory as described below.
for the various measurement modes are summarised in
Table 2.2.25.-1. Further such tests may be performed if
appropriate.
Table 2.2.25.-1. – Minimum tests to be carried out for the control of equipment performance
Purpose Wavelength Absorbance Photometric Resolution/spectral

Method Stray light
accuracy accuracy linearity bandwidth
Based on
measurement of the
absorbance at one
Quantitative or
or more identified X X X X If required in the monograph
limit test
wavelengths (e.g.
assay or impurities
test)
Based on wavelength
of absorption maxima X - - X -
and minima
Based on absorption
measurement and X X - X -
Identification test
wavelength of
absorption maxima
Based on comparison
of spectrum with that X X - - -
of reference substance
Control of wavelength accuracy. Control the wavelength Acceptance criteria

accuracy of an appropriate number of bands in the intended It is recommended to test at least 2 wavelengths that bracket
spectral range using one or more reference materials ; for the intended spectral range.
example, use solid or liquid filters (e.g. holmium perchlorate For benchtop instruments, the tolerance for wavelength
solution R) to verify the position of absorption bands, accuracy of UV-Vis spectroscopy in cuvettes is ± 1 nm at
or measure the emission from a light source to check wavelengths below 400 nm and ± 3 nm at wavelengths of
emission-line position. Table 2.2.25.-2 shows examples of 400 nm and above.
wavelengths used to check wavelength accuracy. When
certified reference materials are used, the reference wavelength For chromatographic systems, the tolerance for wavelength
is that stated on the corresponding certificate. accuracy is ± 2 nm for the whole UV-Vis range.
For PAT applications, a tolerance of ± 2 nm for the UV-Vis
Some instruments may have an automatic or inbuilt
range is recommended. However, wider tolerance intervals
wavelength accuracy control feature.
may be needed for some PAT applications, in which case the
Table 2.2.25.-2. – Examples of wavelengths requisite wavelength accuracy must be defined by the user
used for the control of wavelength accuracy depending on the intended purpose, and using a risk-based
*
Note : the wavelength varies with the resolution of approach.
the instrument The instrument parameters (especially the entrance optics
*
such as slit-width or optical fibre diameter) influence the
Material Wavelengths (nm) resolution and must be the same as those intended for the
201.1 ; 211.4 ; 222.6 ; 240.4 ; actual measurements.
Solutions Cerium in sulfuric acid 253.7 Control of absorbance accuracy. Control the absorbance
accuracy at an appropriate number of wavelengths in the
Didymium in perchloric
acid
511.8 ; 731.6 ; 794.2 intended spectral range, using suitable solid or liquid filters
to check that the absorbance measured at the test wavelength
Holmium in perchloric 241.1 ; 287.2 ; 361.3 ; 451.4 ; matches the certified absorbance of the filter or the absorbance
acid 485.2 ; 536.6 ; 640.5 value that is calculated from a certified specific absorbance.
Nicotinic acid for equipment qualification CRS may be used.
Solid filters Didymium glass 513.5
It is recommended to test absorbance accuracy at selected
Holmium glass 279.3 ; 360.9 ; 453.4 ; 637.5 wavelengths using one or more solid or liquid filters with
different absorbance levels ; as a minimum, values at
Lamps Deuterium 486.0 ; 656.1 approximately the 2 limits of the expected absorbance range
should be verified.
184.9 ; 253.7 ; 312.5 ; 365.0 ;
Mercury (low pressure) 404.7 ; 435.8 ; 546.1 ; 577.0 ; For chromatographic systems and PAT applications, the
579.1 testing of absolute absorbance accuracy may not be necessary,
providing that a standard curve is measured as required.
Neon 717.4 For measurements using nicotinic acid for equipment
Xenon 541.9 ; 688.2 ; 764.2
qualification CRS, the certified specific absorbance is given
in the corresponding leaflet.
For chromatographic systems, it is also possible to control The solution of nicotinic acid can be prepared as follows :
wavelength accuracy by measuring the absorbance of dissolve 57.0-63.0 mg of nicotinic acid for equipment
a 0.05 mg/mL solution of caffeine R in methanol R ; the qualification CRS in a 0.1 M hydrochloric acid solution
absorption maximum is obtained at 272 nm and the minimum prepared from hydrochloric acid R and dilute to 200.0 mL
at 244 nm. with the same acid solution ; dilute 2.0 mL of the solution

2.2.25. Absorption spectrophotometry, ultraviolet and visible EUROPEAN PHARMACOPOEIA
to 50.0 mL with the same acid solution to obtain a final The acceptance criterion depends on the filters or solutions
concentration of 12 mg/L. These volumes can be adjusted used, for example :
to obtain nicotinic acid solutions with other concentrations
(up to about 40 mg/L), for the purposes of testing different – the absorbance is not less than 3.0 when using a 10 g/L
absorbance levels. The absorbance is measured at 213 nm solution of sodium iodide R at 220 nm, a 10 g/L solution
and 261 nm. of potassium iodide R at 250 nm or a 50 g/L solution of
sodium nitrite R at 340 nm and 370 nm ;
Acceptance criteria
The difference between the measured absorbance and the – the absorbance is not less than 2.0 when using a 12 g/L
absorbance of the certified material is ± 0.010 or ± 1 per cent, solution of potassium chloride R at 198 nm.
whichever is greater, for each combination of wavelength and These values apply when using a 1 cm cell and water R as the
absorbance assessed (applies to absorbance values not greater compensation liquid.
than 2). Tolerances for higher absorbance values should be
defined on the basis of a risk assessment. Control of resolution. Where prescribed in a monograph,
measure the resolution of the equipment either using suitable
Control of photometric linearity. Control the photometric certified reference materials, or by recording the spectrum
linearity in the intended spectral range. In the ultraviolet of a 0.02 per cent V/V solution of toluene R in hexane R or
range, the filters used to control absorbance accuracy may heptane R, with respectively hexane R or heptane R as the
be used, as can solutions of nicotinic acid or caffeine. In compensation liquid.
the visible range, neutral glass filters may be used. Prior
to performing the test, ensure that the absorbance of the Acceptance criterion
standards is compatible with the intended linear range. For measurements taken with a solution prepared as described
Solutions with increasing concentrations (e.g. 5-40 mg/L) above, the minimum ratio of the absorbance at the maximum
of nicotinic acid for equipment qualification CRS in a 0.1 M (269 nm) to that at the minimum (266 nm) is stated in the
hydrochloric acid solution prepared from hydrochloric acid R monograph.
may be used. The absorbance is measured at 213 nm and
261 nm.
SYSTEM SUITABILITY
For chromatographic systems, it is also possible to check
photometric linearity using 0.5-50 mg/L solutions of caffeine R System suitability tests may be required prior to sample
in water for chromatography R. The absorbance is measured measurement to verify critical parameters that may have an
at 273 nm. impact on the result.
Acceptance criterion These tests may cover wavelength accuracy, absorbance
The coefficient of determination (R²) is not less than 0.999. accuracy, stray light and photometric linearity. System
Limit of stray light. Stray light is determined at an appropriate functionality tests, for example those performed as part of
wavelength using suitable solid or liquid filters or solutions equipment autotesting, may be considered part of the system
prepared in-house. The instrument parameters used for the suitability tests.
test, such as slit-width and type of light source (e.g. deuterium In the case of UV-Vis detection for chromatographic
or tungsten lamp), must be the same as those intended for the systems, additional system suitability tests are applicable
actual measurements. if prescribed in the monograph and/or in general chapter
Acceptance criterion 2.2.46. Chromatographic separation techniques.

EUROPEAN PHARMACOPOEIA 2.2.27. Thin-layer chromatography
01/2008:20227 used. For saturation of the chromatographic tank, replace the

corrected 9.0 lid and allow to stand at 20-25 °C for 1 h. Unless otherwise
indicated in the monograph, the chromatographic separation
is performed in a saturated tank. Apply the prescribed
volume of solutions as described above. When the solvent
has evaporated from the applied solutions, place the plate
2.2.27. THIN-LAYER in the chromatographic tank, ensuring that the plate is as
vertical as possible and that the spots or bands are above the
CHROMATOGRAPHY surface of the mobile phase. Close the chromatographic tank,
Thin-layer chromatography is a separation technique in maintain it at 20-25 °C and protect from sunlight. Remove the
which a stationary phase consisting of an appropriate material plate when the mobile phase has moved over the prescribed
is spread in a uniform thin layer on a support (plate) of distance, measured between the points of application and the
glass, metal or plastic. Solutions of analytes are deposited solvent front. Dry the plate and visualise the chromatograms
on the plate prior to development. The separation is based as prescribed.
on adsorption, partition, ion-exchange or on combinations For two-dimensional chromatography, dry the plates after the
of these mechanisms and is carried out by migration first development and carry out a second development in a
(development) of solutes (solutions of analytes) in a solvent direction perpendicular to that of the first development.
or a suitable mixture of solvents (mobile phase) through the Horizontal development. Apply the prescribed volume of the
thin-layer (stationary phase). solutions as described above. When the solvent has evaporated
from the applied solutions, introduce a sufficient quantity
APPARATUS
of the mobile phase into the trough of the chamber using a
Plates. The chromatography is carried out using pre-coated syringe or pipette, place the plate in the chamber after verifying
plates as described under Reagents (4.1.1). The particle size of that the latter is horizontal and connect the mobile phase
the silica gel is indicated after the name of the reagent in the direction device according to the manufacturer’s instructions.
tests where it is used. If prescribed, develop the plate starting simultaneously at both
Pre-treatment of the plates. It may be necessary to wash the ends. Close the chamber and maintain it at 20-25 °C. Remove
plates prior to separation. This can be done by migration of the plate when the mobile phase has moved over the distance
an appropriate solvent. The plates may also be impregnated prescribed in the monograph. Dry the plate and visualise the
by procedures such as development, immersion or spraying. chromatograms as prescribed.
At the time of use, the plates may be activated, if necessary, by For two-dimensional chromatography, dry the plates after the
heating in an oven at 120 °C for 20 min. first development and carry out a second development in a
Chromatographic tank with a flat bottom or twin trough, direction perpendicular to that of the first development.
of inert, transparent material, of a size suitable for the plates VISUAL EVALUATION
used and provided with a tightly fitting lid. For horizontal
development the tank is provided with a trough for the mobile Identification. The principal spot in the chromatogram
phase and it additionally contains a device for directing the obtained with the test solution is visually compared to the
mobile phase to the stationary phase. corresponding spot in the chromatogram obtained with the
reference solution by comparing the colour, the size and the
Micropipettes, microsyringes, calibrated disposable retardation factor (RF) of both spots.
capillaries or other application devices suitable for the proper
application of the solutions. The retardation factor (RF) is defined as the ratio of the
distance from the point of application to the centre of the spot
Fluorescence detection device to measure direct fluorescence and the distance travelled by the solvent front from the point
or the inhibition of fluorescence. of application.
Visualisation devices and reagents. Suitable devices are used Verification of the separating power for identification.
for derivatisation to transfer to the plate reagents by spraying, Normally the performance given by the suitability test
immersion or exposure to vapour and, where applicable, to described in Reagents (4.1.1) is sufficient. Only in special
facilitate heating for visualisation of separated components. cases an additional performance criterion is prescribed in the
Documentation. A device may be used to provide monograph.
documentation of the visualised chromatogram, for example a Related substances test. The secondary spot(s) in the
photograph or a computer file. chromatogram obtained with the test solution is (are)
visually compared to either the corresponding spot(s) in
METHOD the chromatogram obtained with the reference solution
Sample application. Apply the prescribed volume of the containing the impurity(ies) or the spot in the chromatogram
solutions at a suitable distance from the lower edge and obtained with the reference solution prepared from a dilution
from the sides of the plate and on a line parallel to the of the test solution.
lower edge ; allow an interval of at least 10 mm (5 mm on Verification of the separating power. The requirements for
high-performance plates) between the centres of circular the verification of the separating power are prescribed in the
spots and 5 mm (2 mm on high-performance plates) between monographs concerned.
the edges of bands. Apply the solutions in sufficiently small
Verification of the detecting power. The detecting power
portions to obtain circular spots 2-5 mm in diameter (1-2 mm
is satisfactory if a spot or band is clearly visible in the
on high-performance plates) or bands 10-20 mm (5-10 mm
chromatogram obtained with the most dilute reference
on high-performance plates) by 1-2 mm.
solution.
In a monograph, where both normal and high-performance
plates may be used, the working conditions for QUANTITATIVE MEASUREMENT
high-performance plates are given in the brackets [ ] after The requirements for resolution and separation are prescribed
those for normal plates. in the monographs concerned.
Vertical development. Line the walls of the chromatographic Substances separated by thin-layer chromatography and
tank with filter paper. Pour into the chromatographic tank responding to UV-Vis irradiation can be determined directly
a sufficient quantity of the mobile phase for the size of the on the plate, using appropriate instrumentation. While
tank to give after impregnation of the filter paper a layer of moving the plate or the measuring device, examine the plate
appropriate depth related to the dimension of the plate to be by measuring the reflectance of the incident light. Similarly,

2.2.27. Thin-layer chromatography EUROPEAN PHARMACOPOEIA
fluorescence may be measured using an appropriate optical Method. Prepare the solution of the substance to be examined
system. Substances containing radionuclides can be quantified (test solution) as prescribed in the monograph and, if
in 3 ways : either directly by moving the plate alongside a necessary, prepare the reference solutions of the substance to
suitable counter or vice versa (see Radiopharmaceutical be determined using the same solvent as in the test solution.
preparations (0125)), by cutting the plates into strips and Apply the same volume of each solution to the plate and
measuring the radioactivity on each individual strip using develop.
a suitable counter or by scraping off the stationary phase,
dissolving it in a suitable scintillation cocktail and measuring Substances responding to UV-Vis irradiation. Prepare and
the radioactivity using a liquid scintillation counter. apply not fewer than 3 reference solutions of the substance to
Apparatus. The apparatus for direct measurement on the be examined, the concentrations of which span the expected
plate consists of : value in the test solution (about 80 per cent, 100 per cent and
– a device for exact positioning and reproducible dispensing 120 per cent). Treat with the prescribed reagent, if necessary,
of the amount of substances onto the plate ; and record the reflectance, the transmittance or fluorescence
in the chromatograms obtained with the test and reference
– a mechanical device to move the plate or the measuring solutions. Use the measured results for the calculation of the
device along the x-axis or the y-axis ; amount of substance in the test solution.
– a recorder and a suitable integrator or a computer ;
– for substances responding to UV-Vis irradiation : a Substances containing radionuclides. Prepare and apply a
photometer with a source of light, an optical device able to test solution containing about 100 per cent of the expected
generate monochromatic light and a photo cell of adequate value. Determine the radioactivity as a function of the path
sensitivity are used for the measurement of reflectance length and report the radioactivity in each resulting peak as a
or transmittance ; if fluorescence is measured, a suitable percentage of the total amount of radioactivity.
filter is required to prevent light used for excitation from
reaching the detector while permitting emitted light or a Criteria for assessing the suitability of the system are described
specific portion thereof to pass ; in the chapter on Chromatographic separation techniques
– for substances containing radionuclides : a suitable counter (2.2.46). The extent to which adjustments of parameters of the
for radioactivity. The linearity range of the counting device chromatographic system can be made to satisfy the criteria of
is to be verified. system suitability are also given in this chapter.

EUROPEAN PHARMACOPOEIA 2.2.28. Gas chromatography
01/2019:20228 The carrier gas flow rate is usually expressed in millilitres per
minute at atmospheric pressure and at the stated temperature.
Flow rate is measured at the detector outlet, either with a
calibrated mechanical device or with a bubble tube, while the
column is at operating temperature.
2.2.28. GAS CHROMATOGRAPHY The linear velocity of the carrier gas through a column is
inversely proportional to the square of the internal diameter
PRINCIPLE of the column for a given flow volume.
Gas chromatography (GC) is a chromatographic separation Helium, nitrogen and hydrogen are commonly used carrier
technique based on the difference in the distribution of species gases.
between 2 non-miscible phases in which the mobile phase
is a carrier gas moving through or passing the stationary DETECTORS
phase contained in a column. It is applicable to substances or Flame-ionisation detectors are usually employed but other
their derivatives which are volatilised under the temperatures detectors such as electron-capture, nitrogen-phosphorus,
employed. mass spectrometric, thermal conductivity or infrared
GC is mainly based on mechanisms of adsorption or mass spectrophotometric detectors may also be used.
distribution. PROCEDURE
EQUIPMENT Equilibrate the column, the injector and the detector at the
The equipment typically consists of : temperatures and the gas flow rates/pressures specified in the
– an injector ; monograph until a stable baseline is achieved. Prepare the test
solution(s) and the reference solution(s) as prescribed. The
– a chromatographic column contained in an oven ; solutions injected must be free from solid particles.
– one or more detector(s) ; Criteria for assessing the suitability of the system are described
– a data acquisition system. in general chapter 2.2.46 Chromatographic separation
The carrier gas flows through the column and then through techniques. The extent to which adjustments of parameters of
the detector at a controlled rate or pressure. the chromatographic system can be made to satisfy the criteria
The chromatography is carried out either at a constant of system suitability are also given in this general chapter.
temperature or according to a given temperature programme.
INJECTORS Static head-space gas chromatography
Injection may be carried out either into a vaporisation
Static head-space gas chromatography is a technique
chamber which may be equipped with a stream splitter, or
particularly suitable for separating and determining volatile
directly at the head of the column using a syringe or an
compounds present in solid or liquid samples. The method
injection valve.
is based on the analysis of the vapour phase in equilibrium
Injections of vapour phase may be effected by static or dynamic with the solid or liquid phase.
head-space injection systems.
Dynamic head-space (purge and trap) injection systems EQUIPMENT
include a sparging device by which volatile substances in The equipment consists of a gas chromatograph provided
solution are swept into an absorbent column maintained at a with a sample-introduction device that may be connected to
low temperature. Retained substances are then desorbed into a module that automatically controls the pressure and the
the mobile phase by rapid heating of the absorbent column. temperature. If necessary, a device for eliminating solvents
Static head-space injection systems include a thermostatically can be added.
controlled sample heating chamber in which closed vials The sample to be analysed is introduced into a container fitted
containing solid or liquid samples are placed for a fixed period with a suitable stopper and a valve-system which permits
of time to allow equilibration of the volatile components of the passage of the carrier gas. The container is placed in
the sample between the non-gaseous phase and the vapour a thermostatically controlled chamber at a temperature set
phase. After equilibration, a predetermined amount of the according to the substance to be examined.
head-space of the vial is flushed into the gas chromatograph.
The sample is held at this temperature long enough to allow
STATIONARY PHASES equilibration between the solid or liquid phase and the vapour
Stationary phases are contained in columns which may be : phase.
– a capillary column whose stationary phase may be The carrier gas is introduced into the container and, after
a solid coating the inner surface of the column (e.g. the prescribed time, a suitable valve is opened so that the gas
macrogol 20 000), or a liquid deposited on the inner surface expands towards the chromatographic column taking the
(e.g. dimethylpolysiloxane) ; in the latter case it may be volatilised compounds with it.
chemically bonded to the inner surface ;
Instead of using a chromatograph specifically equipped for
– a column packed with the stationary phase which may be a the introduction of samples, it is also possible to use airtight
solid phase (e.g. alumina, silica) or an inert solid support syringes and a conventional chromatograph. Equilibration is
(usually a porous polymer) impregnated or coated with a then carried out in a separate chamber and the vapour phase
liquid. is carried onto the column, while necessary precautions are
Capillary columns, made of fused silica, are 0.1 mm to taken to avoid any changes in the equilibrium.
0.53 mm in internal diameter (Ø) and at least 5 m in length.
The stationary phase is a film 0.1 µm to 5.0 µm thick. PROCEDURE
Packed columns, made of glass or metal, are usually 1 m to Using the reference preparations, determine suitable
3 m in length with an internal diameter (Ø) of 2 mm to 4 mm. instrument settings to produce an adequate response.
MOBILE PHASES DIRECT CALIBRATION
Retention time and peak efficiency depend on the carrier gas Introduce into separate, identical containers the preparation
flow rate ; retention time is directly proportional to column to be examined and each of the reference preparations, as
length and resolution is proportional to the square root of prescribed in the monograph, avoiding contact between the
the column length. sampling device and the samples.

2.2.28. Gas chromatography EUROPEAN PHARMACOPOEIA
Close the containers hermetically and place in the Calculate the linear equation of the graph using a least-squares
thermostatically controlled chamber set to the temperature fit, and derive from it the concentration of the substance to be
and pressure prescribed in the monograph ; after equilibration, determined in the preparation to be examined.
carry out the chromatography under the prescribed conditions. Alternatively, plot on a graph the mean of readings against the
STANDARD ADDITIONS added quantity of the substance to be determined. Extrapolate
Add to a set of identical suitable containers equal volumes the line joining the points on the graph until it meets the
of the preparation to be examined. Add to all but one of concentration axis. The distance between this point and the
the containers, suitable quantities of a reference preparation intersection of the axes represents the concentration of the
containing a known concentration of the substance to substance to be determined in the preparation to be examined.
be determined so as to produce a series of preparations
containing steadily increasing concentrations of the substance.
Close the containers hermetically and place in the
thermostatically controlled chamber set to the temperature
and pressure prescribed in the monograph ; after equilibration,
carry out the chromatography under the prescribed conditions.

EUROPEAN PHARMACOPOEIA 2.2.29. Liquid chromatography
01/2019:20229 – a variety of chemically modified supports prepared from

corrected 10.0 polymers, silica or porous graphite, used in normal-phase
and reversed-phase LC (non-polar stationary phase
and polar mobile phase), where the separation is based
principally on partition of the molecules ;
– resins or polymers with acidic or basic groups, used in
2.2.29. LIQUID CHROMATOGRAPHY ion-exchange chromatography, where separation is based
on competition between the ions to be separated and those
PRINCIPLE in the mobile phase ;
Liquid chromatography (LC) is a method of chromatographic – porous silica or polymers, used in size-exclusion
separation based on the difference in the distribution of chromatography (2.2.30), where separation is based
species between 2 non-miscible phases, in which the mobile on differences between the volumes of the molecules,
phase is a liquid which percolates through a stationary phase corresponding to steric exclusion ;
contained in a column. – specially modified stationary phases, e.g. cellulose or
LC is mainly based on mechanisms of adsorption, mass amylose derivatives, proteins or peptides, cyclodextrins etc.,
distribution, ion exchange, size exclusion or stereochemical for the separation of enantiomers (chiral chromatography).
interaction. Most separations are based on reversed-phase LC utilising
Unless otherwise specified, all the information below is valid chemically modified silica as the stationary phase. The surface
for both standard LC and LC using reduced particle-size of the support, i.e. the silanol groups of silica, is reacted
columns (e.g. sub-2 µm). with various silane reagents to produce covalently bound
The latter requires instrumentation that is able to withstand silyl derivatives covering a varying number of active sites on
higher pressures (typically up to 100 MPa, i.e. about 15 000 the surface of the support. The nature of the bonded phase
psi), generates lower extra-column band broadening, provides is an important parameter for determining the separation
improved gradient mixing and allows a higher sampling rate properties of the chromatographic system.
in the detection system. Unless otherwise stated by the manufacturer, silica-based
reversed-phase columns are considered to be stable in mobile
EQUIPMENT phases having an apparent pH in the range 2.0 to 8.0. Columns
The equipment typically consists of : containing porous graphite or particles of polymeric materials
such as styrene-divinylbenzene copolymer are stable over a
– a pumping system ; wider pH range.
– an injector ; Analysis using normal-phase LC with unmodified silica or
– a chromatographic column (a column temperature polar chemically modified silica (e.g. cyanopropyl or diol)
controller may be used) ; as the stationary phase, with a non-polar mobile phase is
– 1 or more detector(s) ; applicable in certain cases.
– a data acquisition system. For analytical separations, the particle size of the most
commonly used stationary phases varies between 2 and
The mobile phase is supplied from 1 or more reservoirs and is
10 µm. The particles may be spherical or irregular, and of
pumped to the injector, then through the column, usually at a
varying porosity and specific surface area. These properties
constant rate, and then through the detector(s).
contribute to the chromatographic behaviour of a particular
PUMPING SYSTEMS stationary phase. In the case of reversed phases, the nature
LC pumping systems deliver the mobile phase at a controlled of the stationary phase, the extent of bonding, e.g. expressed
flow rate. Pressure fluctuations are to be minimised, for as the carbon loading, and whether the stationary phase
example by passing the pressurised solvent through a is end-capped (i.e. part of the residual silanol groups are
pulse-dampening device. Tubing and connections are capable silylated) are additional determining factors. Tailing of peaks,
of withstanding the pressures developed by the pumping particularly of basic substances, can occur when residual
system. LC pumps may be fitted with a facility for ‘bleeding’ silanol groups are present.
the system of entrapped air bubbles. In addition to porous particles, superficially porous or
Microprocessor-controlled pumping systems are capable monolithic materials may be used.
of accurately delivering a mobile phase of either constant Unless otherwise prescribed in the monograph, columns
(isocratic elution) or varying composition (gradient elution), made of stainless steel of varying length and internal diameter
according to a defined programme. In the case of gradient (Ø) are used for analytical chromatography. Columns with
elution, pumping systems which deliver solvent(s) from several internal diameters of less than 2 mm are often referred to as
reservoirs are available and solvent mixing can be achieved on microbore columns.
either the low or high-pressure side of the pump(s). The temperature of the mobile phase and the column must
INJECTORS be kept constant during the analysis. Most separations are
The sample solution is introduced into the flowing mobile performed at room temperature, but some require a different
phase at or near the head of the column using an injection temperature for optimal performance.
system which can operate at high pressure. Fixed-loop MOBILE PHASES
and variable volume devices operated manually or by an For normal-phase LC, low-polarity organic solvents are
autosampler are used. Partial filling of loops during manual generally employed. The residual water content of the solvents
injection may adversely affect injection volume precision. used in the mobile phase is to be strictly controlled to obtain
STATIONARY PHASES reproducible results.
There are many types of stationary phases employed in LC, In reversed-phase LC, aqueous mobile phases, usually with
including : organic solvents and/or modifiers, are employed.
– silica or alumina, commonly used in normal-phase LC The components of the mobile phase are usually filtered to
(polar stationary phase and non-polar mobile phase), remove particles greater than 0.45 µm in size (or greater
where the separation is based on differences in adsorption than 0.2 µm when the stationary phase is made of sub-2 µm
on the stationary phase and/or mass distribution between particles, and when special detectors, e.g. light scattering
the mobile phase and the stationary phase (partition detectors, are used). Multicomponent mobile phases are
chromatography) ; prepared by measuring the required volumes (unless masses

2.2.29. Liquid chromatography EUROPEAN PHARMACOPOEIA
are specified) of the individual components, followed by DETECTORS

mixing. Alternatively, the solvents may be delivered by Ultraviolet/visible (UV/Vis) spectrophotometers (including
individual pumps controlled by proportioning valves, diode array detectors) (2.2.25), are the most commonly
by which mixing is performed according to the desired employed detectors. Fluorescence spectrophotometers,
proportion. Solvents are normally degassed before pumping differential refractometers (RI), electrochemical detectors
by sparging with helium, sonication and/or using in-line (ECD), light scattering detectors, charged aerosol detectors
membrane/vacuum modules to avoid the creation of gas (CAD), mass spectrometers (MS) (2.2.43), radioactivity
bubbles in the detector cell. detectors, multi-angle light scattering (MALS) detectors or
Solvents for the preparation of the mobile phase are normally other detectors may be used.
free of stabilisers and, if an ultraviolet detector is employed,
are transparent at the wavelength of detection. Solvents and
other components employed are to be of appropriate quality. PROCEDURE
In particular, water for chromatography R is used for the
Equilibrate the column with the prescribed mobile phase
preparation of mobile phases when water, or an aqueous
and flow rate, at room temperature or at the temperature
solution, is 1 of the components. Any necessary adjustments
specified in the monograph, until a stable baseline is achieved.
of the pH are made to the aqueous component of the mobile
Prepare the solution(s) of the substance to be examined and
phase and not the mixture. If buffer solutions or saline
the reference solution(s) required. The solutions must be free
solutions are used, adequate rinsing of the system is carried
from solid particles.
out with a mixture of water and a small proportion of the
organic part of the mobile phase (5 per cent V/V) to prevent Criteria for assessing the suitability of the system are described
crystallisation of salts after completion of the analysis. in general chapter 2.2.46. Chromatographic separation
Mobile phases may contain other components, for example a techniques. The extent to which adjustments of parameters of
counter-ion for ion-pair chromatography or a chiral selector the chromatographic system can be made to satisfy the criteria
for chiral chromatography using an achiral stationary phase. of system suitability are also given in this chapter.

EUROPEAN PHARMACOPOEIA 2.2.32. Loss on drying
07/2019:20232 Equipment using other means of drying such as microwaves,

halogen lamps, infrared lamps or mixed technologies may be
used provided they are demonstrated to be fit for purpose.
PROCEDURE
It is recommended to perform the test in an environment that
2.2.32. LOSS ON DRYING has minimal impact on sample measurement (e.g. humidity).
Weigh an empty weighing bottle that has been previously
PRINCIPLE dried under the conditions prescribed for the substance to
be examined for at least 30 min, then weigh the weighing
Loss on drying is the loss of mass after drying under specified bottle filled with the prescribed quantity of substance to be
conditions, calculated as a percentage (m/m). examined. Dry to constant mass or for the prescribed time.
Drying to constant mass means that 2 consecutive weighings Where the drying temperature is indicated by a single value
do not differ by more than 0.5 mg, the 2nd weighing following rather than a range, drying is carried out at the prescribed
an additional period of at least 30 min of drying under the temperature ± 2 °C. Use one of the following procedures,
conditions prescribed for the substance to be examined. unless otherwise prescribed in the monograph.
– In a desiccator : the drying is carried out over about 100 g
EQUIPMENT of molecular sieve R at atmospheric pressure and at room
temperature.
The equipment typically consists of :
– In vacuo : the drying is carried out over about 100 g of
– weighing bottles that are made of suitable inert material molecular sieve R at a pressure not exceeding 2.5 kPa, at
and can easily be dried to constant mass ; their diameter room temperature or at the temperature prescribed in the
is large enough so that the layer of the substance to be monograph.
examined does not exceed about 5 mm ; – In an oven at a specified temperature : the drying is carried
– an analytical balance by which it is possible to determine a out at atmospheric pressure in an oven at the temperature
change in mass of 0.1 mg ; prescribed in the monograph.
– depending on the procedure to be applied, a desiccator, a After drying in an oven, allow the weighing bottle and the
vacuum cabinet, a vacuum oven or an ordinary laboratory sample to cool to room temperature in a desiccator and weigh
oven ; in any case, the temperature of ovens is adjustable to the weighing bottle containing the dried sample.
the specified temperature ± 2 °C ; vacuum ovens in which The mass of the sample is the difference between the mass
the pressure can at least be reduced to about 2 kPa are of the filled weighing bottle and the mass of the dried empty
suitable ; ovens are qualified according to established quality weighing bottle.
system procedures, for example by using a suitable certified The loss on drying is the difference in the mass of the sample
reference material (sodium aminosalicylate dihydrate for before and after drying, expressed as a percentage, m/m being
equipment qualification CRS may be used). implicit.

EUROPEAN PHARMACOPOEIA 2.2.43. Mass spectrometry
01/2008:20243 to be analysed are successively carried to the ion source

of the apparatus where they are ionised ; this technique is
rather poorly suited to very polar or heat-labile compounds.
Other types of coupling (electrospray, thermospray,
atmospheric-pressure chemical ionisation) are considered to
2.2.43. MASS SPECTROMETRY be ionisation techniques in their own right and are described
in the section on modes of ionisation.
Mass spectrometry is based on the direct measurement of
the ratio of the mass to the number of positive or negative Supercritical fluid chromatography/mass spectrometry.
elementary charges of ions (m/z) in the gas phase obtained The mobile phase, usually consisting of supercritical carbon
from the substance to be analysed. This ratio is expressed in dioxide enters the gas state after passing a heated restrictor
atomic mass units (1 a.m.u. = one twelfth the mass of 12C) or between the column and the ion source.
in daltons (1 Da = the mass of the hydrogen atom). Capillary electrophoresis/mass spectrometry. The eluent
The ions, produced in the ion source of the apparatus, are is introduced into the ion source, in some cases after adding
accelerated and then separated by the analyser before reaching another solvent so that flow rates of the order of a few
the detector. All of these operations take place in a chamber microlitres per minute can be attained. This technique is
where a pumping system maintains a vacuum of 10− 3 to limited by the small quantities of sample introduced and the
10 Pa.
− 6 need to use volatile buffers.
The resulting spectrum shows the relative abundance of the
various ionic species present as a function of m/z. The signal MODES OF IONISATION
corresponding to an ion will be represented by several peaks
corresponding to the statistical distribution of the various Electron impact. The sample, in the gas state, is ionised by
a beam of electrons whose energy (usually 70 eV) is greater
isotopes of that ion. This pattern is called the isotopic profile
and (at least for small molecules) the peak representing than the ionisation energy of the sample. In addition to the
the most abundant isotopes for each atom is called the molecular ion M+, fragments characteristic of the molecular
monoisotopic peak. structure are observed. This technique is limited mainly by the
need to vaporise the sample. This makes it unsuited to polar,
Information obtained in mass spectrometry is essentially heat-labile or high molecular mass compounds. Electron
qualitative (determination of the molecular mass, information impact is compatible with the coupling of gas chromatography
on the structure from the fragments observed) or quantitative to mass spectrometry and sometimes with the use of liquid
(using internal or external standards) with limits of detection chromatography.
ranging from the picomole to the femtomole.
Chemical ionisation. This type of ionisation involves a
INTRODUCTION OF THE SAMPLE reagent gas such as methane, ammonia, nitrogen oxide,
nitrogen dioxide or oxygen. The spectrum is characterised
The very first step of an analysis is the introduction of by ions of the (M + H)+ or (M − H)– types, or adduct ions
the sample into the apparatus without overly disturbing formed from the analyte and the gas used. Fewer fragments
the vacuum. In a common method, called direct liquid are produced than with electron impact. A variant of this
introduction, the sample is placed on the end of a cylindrical technique is used when the substance is heat-labile : the
rod (in a quartz crucible, on a filament or on a metal surface). sample, applied to a filament, is very rapidly vaporised by the
This rod is introduced into the spectrometer after passing Joule-Thomson effect (desorption chemical ionisation).
through a vacuum lock where a primary intermediate
vacuum is maintained between atmospheric pressure and the Fast-atom bombardment (FAB) or fast-ion bombardment
secondary vacuum of the apparatus. ionisation (liquid secondary-ion mass spectrometry
LSIMS). The sample, dissolved in a viscous matrix such as
Other introduction systems allow the components of a glycerol, is applied to a metal surface and ionised by a beam of
mixture to be analysed as they are separated by an appropriate neutral atoms such as argon or xenon or high-kinetic-energy
apparatus connected to the mass spectrometer. caesium ions. Ions of the (M + H)+ or (M − H)– types or adduct
Gas chromatography/mass spectrometry. The use of ions formed from the matrix or the sample are produced.
suitable columns (capillary or semi-capillary) allows the end This type of ionisation, well suited to polar and heat-labile
of the column to be introduced directly into the source of the compounds, allows molecular masses of up to 10 000 Da to
apparatus without using a separator. be obtained. The technique can be combined with liquid
chromatography by adding 1 per cent to 2 per cent of glycerol
Liquid chromatography/mass spectrometry. This to the mobile phase ; however, the flow rates must be very low
combination is particularly useful for the analysis of polar (a few microlitres per minute). These ionisation techniques
compounds, which are insufficiently volatile or too heat-labile also allow thin-layer chromatography plates to be analysed by
to be analysed by gas chromatography coupled with mass applying a thin layer of matrix to the surface of these plates.
spectrometry. This method is complicated by the difficulty of
obtaining ions in the gas phase from a liquid phase, which Field desorption and field ionisation. The sample is
requires very special interfaces such as : vaporised near a tungsten filament covered with microneedles
(field ionisation) or applied to this filament (field desorption).
– direct liquid introduction : the mobile phase is nebulised, A voltage of about 10 kV, applied between this filament and a
and the solvent is evaporated in front of the ion source of counter-electrode, ionises the sample. These two techniques
the apparatus, mainly produce molecular ions M+, and (M + H)+ ions and are
– particle-beam interface : the mobile phase, which may flow used for low polarity and/or heat-labile compounds.
at a rate of up to 0.6 mL/min, is nebulised in a desolvation Matrix-assisted laser desorption ionisation (MALDI). The
chamber such that only the analytes, in neutral form, reach sample, in a suitable matrix and deposited on a metal support,
the ion source of the apparatus ; this technique is used is ionised by a pulsed laser beam whose wavelength may range
for compounds of relatively low polarity with molecular from UV to IR (impulses lasting from a picosecond to a few
masses of less than 1000 Da, nanoseconds). This mode of ionisation plays an essential role
– moving-belt interface : the mobile phase, which may flow in the analysis of very high molecular mass compounds (more
at a rate of up to 1 mL/min, is applied to the surface of a than 100 000 Da) but is limited to time-of flight analysers (see
moving belt ; after the solvent evaporates, the components below).

2.2.43. Mass spectrometry EUROPEAN PHARMACOPOEIA
Electrospray. This mode of ionisation is carried out at in field-free regions between the ion source and the detector.
atmospheric pressure. The samples, in solution, are introduced Examination of these decompositions is very useful for the
into the source through a capillary tube, the end of which has determination of the structure as well as the characterisation
a potential of the order of 5 kV. A gas can be used to facilitate of a specific compound in a mixture and involves tandem mass
nebulisation. Desolvation of the resulting microdroplets spectrometry. There are many such techniques depending on
produces singly or multiply charged ions in the gas phase. the region where these decompositions occur :
The flow rates vary from a few microlitres per minute to – daughter-ion mode (determination of the decomposition
1 mL/min. This technique is suited to polar compounds and ions of a given parent ion) : B/E = constant, MIKES
to the investigation of biomolecules with molecular masses of (Mass-analysed Ion Kinetic Energy Spectroscopy),
up to 100 000 Da. It can be coupled to liquid chromatography – parent-ion mode (determination of all ions which by
or capillary electrophoresis. decomposition give an ion with a specific m/z ratio) :
Atmospheric-pressure chemical ionisation (APCI). B2/E = constant,
Ionisation is carried out at atmospheric pressure by the – neutral-loss mode (detection of all the ions that lose the
action of an electrode maintained at a potential of several same fragment) :
kilovolts and placed in the path of the mobile phase, which is B/E(1 − E/E0)1/2 = constant, where E0 is the basic voltage
nebulised both by thermal effects and by the use of a stream of the electric sector.
of nitrogen. The resulting ions carry a single charge and are of
the (M + H)+ type in the positive mode and of the (M − H)– Quadrupoles. The analyser consists of four parallel metal
type in the negative mode. The high flow rates that can be rods, which are cylindrical or hyperbolic in cross-section.
used with this mode of ionisation (up to 2 mL/min) make this They are arranged symmetrically with respect to the trajectory
an ideal technique for coupling to liquid chromatography. of the ions ; the pairs diagonally opposed about the axis of
symmetry of rods are connected electrically. The potentials to
Thermospray. The sample, in the mobile phase consisting the two pairs of rods are opposed. They are the resultant of a
of water and organic modifiers and containing a volatile constant component and an alternating component. The ions
electrolyte (generally ammonium acetate) is introduced in produced at the ion source are transmitted and separated by
nebulised form after having passed through a metal capillary varying the voltages applied to the rods so that the ratio of
tube at controlled temperature. Acceptable flow rates are of the continuous voltage to alternating voltage remains constant.
order of 1 mL/min to 2 mL/min. The ions of the electrolyte The quadrupoles usually have a mass range of 1 a.m.u. to
ionise the compounds to be analysed. This ionisation process 2000 a.m.u., but some may range up to 4000 a.m.u. Although
may be replaced or enhanced by an electrical discharge they have a lower resolving power than magnetic sector
of about 800 volts, notably when the solvents are entirely analysers, they nevertheless allow the monoisotopic profile of
organic. This technique is compatible with the use of liquid single charged ions to be obtained for the entire mass range. It
chromatography coupled with mass spectrometry. is possible to obtain spectra using three quadrupoles arranged
ANALYSERS in series, Q1, Q2, Q3 (Q2 serves as a collision cell and is not
really an analyser ; the most commonly used collision gas is
Differences in the performance of analysers depend mainly on argon).
two parameters :
The most common types of scans are the following :
– the range over which m/z ratios can be measured, ie, the – daughter-ion mode : Q1 selects an m/z ion whose fragments
mass range, obtained by collision in Q2 are analysed by Q3,
– their resolving power characterised by the ability to separate – parent-ion mode : Q3 filters only a specific m/z ratio, while
two ions of equal intensity with m/z ratios differing by ∆M, Q1 scans a given mass range. Only the ions decomposing to
and whose overlap is expressed as a given percentage of give the ion selected by Q3 are detected,
valley definition ; for example, a resolving power (M/∆M) of
– neutral loss mode : Q1 and Q3 scan a certain mass range
1000 with 10 per cent valley definition allows the separation
but at an offset corresponding to the loss of a fragment
of m/z ratios of 1000 and 1001 with the intensity returning
characteristic of a product or family of compounds.
to 10 per cent above baseline. However, the resolving power
may in some cases (time-of-flight analysers, quadrupoles, It is also possible to obtain spectra by combining quadrupole
ion-trap analysers) be defined as the ratio between the analysers with magnetic or electrostatic sector instruments ;
molecular mass and peak width at half height (50 per cent such instruments are called hybrid mass spectrometers.
valley definition). Ion-trap analyser. The principle is the same as for a
Magnetic and electrostatic analysers. The ions produced quadrupole, this time with the electric fields in three
in the ion source are accelerated by a voltage V, and focused dimensions. This type of analyser allows product-ion spectra
towards a magnetic analyser (magnetic field B) or an over several generations (MSn) to be obtained.
electrostatic analyser (electrostatic field E), depending on the Ion-cyclotron resonance analysers. Ions produced in a cell
configuration of the instrument. They follow a trajectory of and subjected to a uniform, intense magnetic field move in
radius r according to Laplace’s law : circular orbits at frequencies which can be directly correlated
to their m/z ratio by applying a Fourier transform algorithm.
m B 2r 2 This phenomenon is called ion-cyclotron resonance.
=
z 2V Analysers of this type consist of superconducting magnets and
Two types of scans can be used to collect and measure the are capable of very high resolving power (up to 1000 000 and
various ions produced by the ion source : a scan of B holding more) as well as MSn spectra. However, very low pressures are
V fixed or a scan of V with constant B. The magnetic analyser required (of the order of 10− 7 Pa).
is usually followed by an electric sector that acts as a kinetic Time-of-flight analysers. The ions produced at the ion
energy filter and allows the resolving power of the instrument source are accelerated at a voltage V of 10 kV to 20 kV. They
to be increased appreciably. The maximum resolving power pass through the analyser, consisting of a field-free tube, 25 cm
of such an instrument (double sector) ranges from 10 000 to to 1.5 m long, generally called a flight tube. The time (t) for
150 000 and in most cases allows the value of m/z ratios to an ion to travel to the detector is proportional to the square
be calculated accurately enough to determine the elemental root of the m/z ratio. Theoretically the mass range of such an
composition of the corresponding ions. For monocharged analyser is infinite. In practice, it is limited by the ionisation
ions, the mass range is from 2000 Da to 15 000 Da. Some ions or desorption method. Time-of-flight analysers are mainly
may decompose spontaneously (metastable transitions) or by used for high molecular mass compounds (up to several
colliding with a gas (collision-activated dissociation (CAD)) hundred thousand daltons). This technique is very sensitive (a

EUROPEAN PHARMACOPOEIA 2.2.43. Mass spectrometry
few picomoles of product are sufficient). The accuracy of the using suitable internal standards (for example, deuterated
measurements and the resolving power of such instruments standards). This type of analysis can be performed only on
may be improved considerably by using an electrostatic mirror apparatus fitted with three quadrupoles in series, ion-trap
(reflectron). analysers or cyclotron-resonance analysers.
SIGNAL ACQUISITION CALIBRATION
There are essentially three possible modes. Calibration allows the corresponding m/z value to be
Complete spectrum mode. The entire signal obtained over attributed to the detected signal. As a general rule, this is done
a chosen mass range is recorded. The spectrum represents using a reference substance. This calibration may be external
the relative intensity of the different ionic species present as (acquisition file separate from the analysis) or internal (the
a function of m/z. The results are essentially qualitative. The reference substance(s) are mixed with the substance to be
use of spectral reference libraries for more rapid identification examined and appear on the same acquisition file). The
is possible. number of ions or points required for reliable calibration
depends on the type of analyser and on the desired accuracy
Fragmentometric mode (Selected-ion monitoring). The of the measurement, for example, in the case of a magnetic
acquired signal is limited to one (single-ion monitoring (SIM)) analyser where the m/z ratio varies exponentially with the
or several (multiple-ion monitoring (MIM)) ions characteristic value of the magnetic field, there should be as many points
of the substance to be analysed. The limit of detection as possible.
can be considerably reduced in this mode. Quantitative or
semiquantitative tests can be carried out using external or SIGNAL DETECTION AND DATA PROCESSING
internal standards (for example, deuterated standards). Such Ions separated by an analyser are converted into electric
tests cannot be carried out with time-of-flight analysers. signals by a detection system such as a photomultiplier or
Fragmentometric double mass spectrometry mode an electron multiplier. These signals are amplified before
(multiple reaction monitoring (MRM)). The unimolecular being re-converted into digital signals for data processing,
or bimolecular decomposition of a chosen precursor ion allowing various functions such as calibration, reconstruction
characteristic of the substance to be analysed is followed of spectra, automatic quantification, archiving, creation or use
specifically. The selectivity and the highly specific nature of of libraries of mass spectra. The various physical parameters
this mode of acquisition provide excellent sensitivity levels required for the functioning of the apparatus as a whole are
and make it the most appropriate for quantitative studies controlled by computer.

EUROPEAN PHARMACOPOEIA 2.2.46. Chromatographic separation techniques
07/2016:20246 chromatograms are represented as a sequence of Gaussian

corrected 9.2 peaks on a baseline (Figure 2.2.46.-1).
Peak
The portion of a chromatogram recording the detector
response when a single component (or 2 or more unresolved
components) is eluted from the column.
2.2.46. CHROMATOGRAPHIC The peak may be defined by the peak area, or the peak
SEPARATION TECHNIQUES height (h) and the peak width at half-height (wh), or the
peak height (h) and the peak width between the points of
Chromatographic separation techniques are multi-stage
inflection (wi). In Gaussian peaks (Figure 2.2.46.-1) there is
separation methods in which the components of a sample
the following relationship :
are distributed between 2 phases, one of which is stationary,
while the other is mobile. The stationary phase may be a w h = 1.18wi
solid or a liquid supported on a solid or a gel. The stationary
phase may be packed in a column, spread as a layer, or Retention time (tR)
distributed as a film, etc. The mobile phase may be gaseous Time required for elution of a component (Figure 2.2.46.-1,
or liquid or supercritical fluid. The separation may be based baseline scale being in minutes).
on adsorption, mass distribution (partition), ion-exchange, Retention volume (VR)
etc., or may be based on differences in the physico-chemical Volume of the mobile phase required for elution of a
properties of the molecules such as size, mass, volume, etc. component. It may be calculated from the retention time and
This chapter contains definitions and calculations of common the flow rate (F) in millilitres per minute using the following
parameters and generally applicable requirements for system equation :
suitability. Principles of separation, apparatus and methods
are given in the following general methods : VR = tR ´ F
– paper chromatography (2.2.26); Hold-up time (tM)
– thin-layer chromatography (2.2.27) ; Time required for elution of an unretained component
– gas chromatography (2.2.28); (Figure 2.2.46.-1, baseline scale being in minutes). In
– liquid chromatography (2.2.29) ; size-exclusion chromatography, the symbol t0 (see below) is
used.
– size-exclusion chromatography (2.2.30) ;
– supercritical fluid chromatography (2.2.45). Hold-up volume (VM)
Volume of the mobile phase required for elution of an
DEFINITIONS unretained component. It may be calculated from the hold-up
The system suitability and acceptance criteria in monographs time and the flow rate (F) in millilitres per minute using the
have been set using parameters as defined below. With some following equation :
equipment, certain parameters, such as the signal-to-noise ratio
and resolution, can be calculated using software provided by VM = tM ´ F
the manufacturer. It is the responsibility of the user to ensure In size-exclusion chromatography, the symbol V0 (see below)
that the calculation methods used in the software are equivalent is used.
to the requirements of the European Pharmacopoeia and to Retention factor (k)
make any necessary corrections if this is not the case.
The retention factor (also known as mass distribution ratio
Chromatogram (Dm) or capacity factor (k′)) is defined as :
A graphical or other representation of detector response,
effluent concentration or other quantity used as a measure amount of component in stationary phase V
k= = KC s
of effluent concentration, versus time or volume. Idealised amount of component in mobile phase VM
Figure 2.2.46.-1.

2.2.46. Chromatographic separation techniques EUROPEAN PHARMACOPOEIA
Figure 2.2.46.-2.
KC = distribution constant (also known as equilibrium gel pores. It may be calculated from the total mobile phase
distribution coefficient) ; time and the flow rate (F) in millilitres per minute using the
VS = volume of the stationary phase ; following equation :
VM = volume of the mobile phase. Vt = tt ´ F
The retention factor of a component may be determined from Retention time of an unretained compound (t0)
the chromatogram using the following equation : In size-exclusion chromatography, retention time of a
tR - tM component whose molecules are larger than the largest gel
k= pores (Figure 2.2.46.-2).
tM
Retention volume of an unretained compound (V0)
Total mobile phase time (tt)
In size-exclusion chromatography, retention time of a In size-exclusion chromatography, retention volume of a
component whose molecules are smaller than the smallest component whose molecules are larger than the largest gel
gel pores (Figure 2.2.46.-2). pores. It may be calculated from the retention time of an
unretained compound and the flow rate (F) in millilitres per
Total mobile phase volume (Vt) minute using the following equation :
In size-exclusion chromatography, retention volume of a
component whose molecules are smaller than the smallest V0 = t0 ´ F

Distribution constant (K0) Time Mobile phase A Mobile phase B

In size-exclusion chromatography, the elution characteristics (min) (per cent V/V) (per cent V/V)
of a component in a particular column may be given by 0 - 20 100 → 0 0 → 100
the distribution constant (also referred to as distribution 20 - 30 0 100
coefficient), which is calculated using the following equation :
tR - t0 Flow rate : set to obtain sufficient back-pressure (e.g.
K0 = 2 mL/min).
tt - t0
Detection : spectrophotometer at 265 nm.
Retardation factor (RF) Determine the time (t0.5) in minutes when the absorbance has
The retardation factor (also known as retention factor (Rf)), increased by 50 per cent (Figure 2.2.46.-4).
used in planar chromatography, is the ratio of the distance
from the point of application to the centre of the spot and D = tD ´ F
the distance travelled by the solvent front from the point of
application (Figure 2.2.46.-3). tD = t0.5 − 0.5tG (in minutes) ;
b
tG = pre-defined gradient time (= 20 min) ;
RF =
a F = flow rate (in millilitres per minute).
b = migration distance of the component ;
a = migration distance of the solvent front.
b
Figure 2.2.46.-4
Symmetry factor (As)
C The symmetry factor of a peak (Figure 2.2.46.-5) is calculated
using the following equation :
w0.05
As =
A. mobile phase front B. spot C. line of application 2d
Figure 2.2.46.-3. w0.05 = width of the peak at one-twentieth of the peak
Plate number (N) height ;
The column performance (apparent efficiency) may be d = distance between the perpendicular dropped from
calculated from data obtained under either isothermal, the peak maximum and the leading edge of the
isocratic or isodense conditions, depending on the peak at one-twentieth of the peak height.
technique, as the plate number (also referred to as number of An As value of 1.0 signifies symmetry. When As > 1.0, the peak
theoretical plates), using the following equation, the values of is tailing. When As < 1.0, the peak is fronting.
tR and wh being expressed in the same units :
2
æt ö÷
N = 5.54çç R ÷÷
ççè w h÷
÷ø
tR = retention time of the peak corresponding to the

component ;
wh = width of the peak at half-height.
The plate number varies with the component as well as with
the column, the column temperature, the mobile phase and
the retention time.
Dwell volume (D)
The dwell volume (also known as gradient delay volume)
is the volume between the point at which the eluents meet
and the top of the column. It can be determined using the
following procedure.
Column : replace the chromatographic column by an Figure 2.2.46.-5
appropriate capillary tubing (e.g. 1 m × 0.12 mm). Resolution (Rs)
Mobile phase : The resolution between peaks of 2 components
– mobile phase A : water R ; (Figure 2.2.46.-1) may be calculated using the following
– mobile phase B : 0.1 per cent V/V solution of acetone R ; equation :

1.18(tR2 - tR1) The unadjusted relative retention (rG) is calculated using the
Rs =
w h1 + w h2 following equation :
tR2 > tR1 tRi
rG =
tR1, tR2 = retention times of the peaks ; tRst
wh1, wh2 = peak widths at half-height. Unless otherwise indicated, values for relative retention stated
in monographs correspond to unadjusted relative retention.
In quantitative planar chromatography, using densitometry,
the migration distances are used instead of retention times In planar chromatography, the retardation factors RFst and RFi
and the resolution between peaks of 2 components may be are used instead of tRst and tRi.
calculated using the following equation : Signal-to-noise ratio (S/N)
The short-term noise influences the precision of quantification.
1.18a(R F 2 - R F1) The signal-to-noise ratio is calculated using the following
Rs =
w h1 + w h2 equation :
RF1, RF2 = retardation factors of the peaks ; 2H
S/N =
wh1, wh2 = peak widths at half-height ; h
a = migration distance of the solvent front. H = height of the peak (Figure 2.2.46.-7) corresponding
to the component concerned, in the chromatogram
Peak-to-valley ratio (p/v) obtained with the prescribed reference solution,
The peak-to-valley ratio may be employed as a system measured from the maximum of the peak to the
suitability criterion in a test for related substances when extrapolated baseline of the signal observed over
baseline separation between 2 peaks is not achieved a distance equal to at least 5 times the width at
(Figure 2.2.46.-6). half-height ;
h = range of the noise in a chromatogram obtained
Hp after injection or application of a blank, observed
p/v=
Hv over a distance equal to at least 5 times the width
at half-height of the peak in the chromatogram
Hp = height above the extrapolated baseline of the minor obtained with the prescribed reference solution
peak ; and, if possible, situated equally around the place
Hv = height above the extrapolated baseline at the lowest where this peak would be found.
point of the curve separating the minor and major
peaks.
Figure 2.2.46.-7.
System repeatability
The repeatability of response is expressed as an estimated
percentage relative standard deviation (sr(%)) of a consecutive
series of measurements for not fewer than 3 injections or
applications of a reference solution, and is calculated using
the following equation :
Figure 2.2.46.-6 100

2
å(yi - y )
Relative retention (r) sr (%) =
y n-1
Relative retention is calculated as an estimate using the
following equation : yi = individual values expressed as peak area,
peak height, or ratio of areas by the internal
tRi - tM standardisation method ;
r=
tRst - tM y = mean of individual values ;
tRi = retention time of the peak of interest ; n = number of individual values.
tRst = retention time of the reference peak (usually SYSTEM SUITABILITY
the peak corresponding to the substance to be
examined) ; The various components of the equipment employed must
be qualified and be capable of achieving the performance
tM = hold-up time. required to conduct the test or assay.

The system suitability tests represent an integral part of the ADJUSTMENT OF CHROMATOGRAPHIC CONDITIONS
method and are used to ensure adequate performance of the
chromatographic system. Apparent efficiency, retention factor The extent to which the various parameters of a
(mass distribution ratio), resolution and symmetry factor chromatographic test may be adjusted to satisfy the system
are the parameters that are usually employed in assessing suitability criteria without fundamentally modifying the
the performance of the column. Factors that may affect the methods are listed below. Adjustment of conditions with
chromatographic behaviour include : gradient elutions is more critical than with isocratic elutions,
since it may lead to shifts in peaks to a different step of the
– the composition, ionic strength, temperature and apparent gradient, thus leading to the incorrect assignment of peaks,
pH of the mobile phase ; and to the masking of peaks or a shift such that elution
– flow rate, column dimensions, column temperature and occurs beyond the prescribed elution time. Changes other
pressure ; than those indicated require revalidation of the method. The
chromatographic conditions described have been validated
– stationary phase characteristics including type of during the elaboration of the monograph.
chromatographic support (particle-based or monolithic),
particle or macropore size, porosity, specific surface area ; The system suitability tests are included to verify that the
– reversed-phase and other surface-modification of the separation required for satisfactory performance of the test or
stationary phases, the extent of chemical modification (as assay is achieved. Nonetheless, since the stationary phases are
expressed by end-capping, carbon loading etc.). described in a general way and there is such a variety available
commercially, with differences in chromatographic behaviour,
The following requirements and any supplementary some adjustments of the chromatographic conditions may
requirements given in the individual monograph are to be be necessary to achieve the prescribed system suitability
fulfilled unless otherwise prescribed : requirements. With reversed-phase liquid chromatographic
– in a related substances test or assay, for a peak in the methods in particular, adjustment of the various parameters
chromatogram obtained with a reference solution used for will not always result in satisfactory chromatography. In that
quantification, the symmetry factor is 0.8 to 1.5, unless case, it may be necessary to replace the column with another
otherwise prescribed ; of the same type (e.g. octadecylsilyl silica gel), which exhibits
the desired chromatographic behaviour. The Knowledge
– in an assay of an active substance where the value is 100 per database on the EDQM website usually contains information
cent for a pure substance, the maximum permitted relative on the column(s) used during monograph elaboration.
standard deviation (sr(%)max) for the defined limits is
calculated for a series of injections of the reference solution For critical parameters the adjustments are defined clearly in
using the following equation : the monograph to ensure the system suitability.
Thin-layer chromatography and paper chromatography
KB n
sr (%) =
max t90%, n - 1 Composition of the mobile phase : the amount of the minor
solvent component may be adjusted by ± 30 per cent relative
K = constant (0.349), obtained from the expression or ± 2 per cent absolute, whichever is the larger ; for a minor
K=
0.6 t
´ 90 %6 ,5 in which
0.6
represents the component at 10 per cent of the mobile phase, a 30 per cent
2 2 relative adjustment allows a range of 7-13 per cent whereas
required percentage relative standard a 2 per cent absolute adjustment allows a range of 8-12 per
deviation after 6 injections for B = 1.0 ; cent, the relative value therefore being the larger ; for a minor
B = upper limit given in the definition of the component at 5 per cent of the mobile phase, a 30 per cent
individual monograph minus 100 per cent ; relative adjustment allows a range of 3.5-6.5 per cent whereas
n = number of replicate injections of the reference a 2 per cent absolute adjustment allows a range of 3-7 per
solution (3 ≤ n ≤ 6) ; cent, the absolute value being the larger in this case ; no other
component is altered by more than 10 per cent absolute.
t90%,n−1 = Student’s t at the 90 per cent probability level
(double sided) with n−1 degrees of freedom. pH of the aqueous component of the mobile phase : ± 0.2 pH,
unless otherwise prescribed, or ± 1.0 pH when non-ionisable
Unless otherwise prescribed, the maximum permitted substances are to be examined.
relative standard deviation does not exceed the appropriate
value given in Table 2.2.46.-1. This requirement does not Concentration of salts in the buffer component of a mobile
apply to tests for related substances. phase : ± 10 per cent.
Table 2.2.46.-1. – Repeatability requirements Application volume : 10-20 per cent of the prescribed volume
if using fine particle size plates (2-10 µm).
Number of individual injections
Liquid chromatography : isocratic elution
3 4 5 6
B (per cent) Maximum permitted relative standard deviation

Composition of the mobile phase : the amount of the minor
solvent component may be adjusted by ± 30 per cent relative
2.0 0.41 0.59 0.73 0.85 or ± 2 per cent absolute, whichever is the larger (see example
2.5 0.52 0.74 0.92 1.06
above) ; no other component is altered by more than 10 per
cent absolute.
3.0 0.62 0.89 1.10 1.27
pH of the aqueous component of the mobile phase : ± 0.2 pH,
– in a related substances test, the limit of quantification unless otherwise prescribed, or ± 1.0 pH when non-ionisable
(corresponding to a signal-to-noise ratio of 10) is equal to substances are to be examined.
or less than the disregard limit.
Concentration of salts in the buffer component of a mobile
Compliance with the system suitability criteria is required phase : ± 10 per cent.
throughout the chromatographic procedure. Depending on
various factors, such as the frequency of use of the procedure Flow rate : ± 50 per cent ; a larger adjustment is acceptable
and experience with the chromatographic system, the analyst when changing the column dimensions (see the formula
chooses an appropriate verification scheme to monitor this. below).

Column parameters D = dwell volume, in millilitres ;

Stationary phase : D0 = dwell volume used for development of the method,
– no change of the identity of the substituent of the in millilitres ;
stationary phase permitted (e.g. no replacement of C18 F = flow rate, in millilitres per minute.
by C8) ;
The isocratic step introduced for this purpose may be omitted
– particle size : maximum reduction of 50 per cent ; no if validation data for application of the method without this
increase permitted. step is available.
Column dimensions : pH of the aqueous component of the mobile phase : no
– length : ± 70 per cent ; adjustment permitted.
Concentration of salts in the buffer component of a mobile
– internal diameter : ± 25 per cent. phase : no adjustment permitted.
When column dimensions are changed, the flow rate may Flow rate : adjustment is acceptable when changing the column
be adjusted as necessary using the following equation : dimensions (see the formula below).
Column parameters
l d2 Stationary phase :
F2 = F1 2 22
l1d1 – no change of the identity of the substituent of the
stationary phase permitted (e.g. no replacement of C18
F1 = flow rate indicated in the monograph, in by C8) ;
millilitres per minute ;
– particle size : no adjustment permitted.
F2 = adjusted flow rate, in millilitres per minute ; Column dimensions :
l1 = length of the column indicated in the – length : ± 70 per cent ;
monograph, in millimetres ; – internal diameter : ± 25 per cent.
l2 = length of the column used, in millimetres ; When column dimensions are changed, the flow rate may
d1 = internal diameter of the column indicated in the be adjusted as necessary using the following equation :
monograph, in millimetres ; l d2
d2 = internal diameter of the column used, in F2 = F1 2 22
l1d1
millimetres.
Temperature : ± 10 °C, where the operating temperature is F1 = flow rate indicated in the monograph, in
specified, unless otherwise prescribed. millilitres per minute ;
F2 = adjusted flow rate, in millilitres per minute ;
Detector wavelength : no adjustment permitted.
Injection volume : may be decreased, provided detection and l1 = length of the column indicated in the
repeatability of the peak(s) to be determined are satisfactory ; monograph, in millimetres ;
no increase permitted. l2 = length of the column used, in millimetres ;
Liquid chromatography : gradient elution d1 = internal diameter of the column indicated in the
Adjustment of chromatographic conditions for gradient monograph, in millimetres ;
systems requires greater caution than for isocratic systems. d2 = internal diameter of the column used, in
Composition of the mobile phase/gradient elution : minor millimetres.
adjustments of the composition of the mobile phase and the Temperature : ± 5 °C, where the operating temperature is
gradient are acceptable provided that : specified, unless otherwise prescribed.
– the system suitability requirements are fulfilled ; Detector wavelength : no adjustment permitted.
Injection volume : may be decreased, provided detection and
– the principal peak(s) elute(s) within ± 15 per cent of the repeatability of the peak(s) to be determined are satisfactory ;
indicated retention time(s) ; no increase permitted.
– the final composition of the mobile phase is not weaker in Gas chromatography
elution power than the prescribed composition.
Column parameters
Where compliance with the system suitability requirements Stationary phase :
cannot be achieved, it is often preferable to consider the dwell
volume or to change the column. – particle size : maximum reduction of 50 per cent ; no
increase permitted (packed columns);
Dwell volume. The configuration of the equipment employed – film thickness : − 50 per cent to + 100 per cent (capillary
may significantly alter the resolution, retention time and columns).
relative retentions described. Should this occur, it may be due
Column dimensions :
to excessive dwell volume. Monographs preferably include an
isocratic step before the start of the gradient programme so – length : ± 70 per cent ;
that an adaptation can be made to the gradient time points – internal diameter : ± 50 per cent.
to take account of differences in dwell volume between the Flow rate : ± 50 per cent.
system used for method development and that actually used. Temperature : ± 10 per cent.
It is the user’s responsibility to adapt the length of the isocratic Injection volume and split volume : may be adjusted, provided
step to the analytical equipment used. If the dwell volume detection and repeatability are satisfactory.
used during the elaboration of the monograph is given in the
monograph, the time points (t min) stated in the gradient table Supercritical fluid chromatography
may be replaced by adapted time points (tc min), calculated Composition of the mobile phase : for packed columns, the
using the following equation : amount of the minor solvent component may be adjusted by
± 30 per cent relative or ± 2 per cent absolute, whichever is
(D - D0 ) the larger ; no adjustment is permitted for a capillary column
tc = t - system.
F

Detector wavelength : no adjustment permitted. solution and a reference solution. The internal standard
Column parameters is chosen such that it does not react with the substance
to be examined, is stable and does not contain impurities
Stationary phase : with the same retention time as that of the substance to
– particle size : maximum reduction of 50 per cent ; no be examined. The concentration of the substance to be
increase permitted (packed columns). examined is determined by comparing the ratio of the peak
Column dimensions : areas or peak heights due to the substance to be examined
– length : ± 70 per cent ; and the internal standard in the test solution with the ratio
of the peak areas or peak heights due to the substance to
– internal diameter : be examined and the internal standard in the reference
± 25 per cent (packed columns) ; solution.
± 50 per cent (capillary columns). – Normalisation procedure. The percentage content of a
Flow rate : ± 50 per cent. component of the substance to be examined is calculated
by determining the area of the corresponding peak as a
Temperature : ± 5 °C, where the operating temperature is percentage of the total area of all the peaks, excluding those
specified. due to solvents or reagents or arising from the mobile
Injection volume : may be decreased, provided detection and phase or the sample matrix, and those at or below the
repeatability are satisfactory ; no increase permitted. disregard limit.
QUANTIFICATION – Calibration procedure. The relationship between the
measured or evaluated signal (y) and the quantity
Peaks due to solvents and reagents or arising from the (concentration, mass, etc.) of substance (x) is determined
mobile phase or the sample matrix are disregarded during and the calibration function is calculated. The analytical
quantification. results are calculated from the measured signal or evaluated
– Detector sensitivity. The detector sensitivity is the signal signal of the analyte by means of the inverse function.
output per unit concentration or unit mass of a substance In tests for related substances for both the external standard
in the mobile phase entering the detector. The relative method, when a dilution of the test solution is used for
detector response factor, commonly referred to as response comparison, and the normalisation procedure, any correction
factor, expresses the sensitivity of a detector for a given factors indicated in the monograph are applied (i.e. when the
substance relative to a standard substance. The correction response factor is outside the range 0.8-1.2).
factor is the reciprocal of the response factor.
When the related substances test prescribes the total of
– External standard method. The concentration of the impurities or there is a quantitative determination of an
component(s) to be analysed is determined by comparing impurity, it is important to choose an appropriate threshold
the response(s) (peak(s)) obtained with the test solution setting and appropriate conditions for the integration of the
to the response(s) (peak(s)) obtained with a reference peak areas. In such tests the disregard limit, i.e. the limit at
solution. or below which a peak is disregarded, is generally 0.05 per
– Internal standard method. Equal amounts of a component cent. Integration of the peak area of any impurity that is not
that will be resolved from the substance to be examined completely separated from the principal peak is preferably
(the internal standard) are introduced into the test performed by valley-to-valley extrapolation (tangential skim).

EUROPEAN PHARMACOPOEIA 2.2.65. Voltametric titration
01/2013:20265 electrode, a rotating-disc electrode or a carbon electrode)

and a 2nd electrode (for example, a platinum electrode, a
rotating-disc electrode or a carbon electrode).
Method. Set the current to the indicator electrode as
prescribed in the monograph and plot a graph of the initial
voltage and the values obtained during the titration as
2.2.65. VOLTAMETRIC TITRATION functions of the quantity of titrant added. Add the titrant
in not fewer than 3 successive quantities equal to a total of
In voltametric titration the end-point of the titration is about 80 per cent of the theoretical volume corresponding
determined by following the variation of the voltage measured to the presumed equivalence point. The 3 values must fall
between 2 electrodes (either 1 indicator electrode and on a straight line. Continue adding the titrant beyond the
1 reference electrode or 2 indicator electrodes) immersed in presumed equivalence point in not fewer than 3 successive
the solution to be examined and maintained at a constant quantities. The values obtained must fall on another straight
current as a function of the quantity of titrant added. line. The point of intersection of the 2 lines represents the
end-point of the titration.
Apparatus. The apparatus comprises an adjustable current Using titration systems for voltametric titration with
source and a voltmeter; the detection system generally 2 indicator electrodes, the whole titration curve is recorded
consists of an indicator electrode (for example, a platinum and used to determine the end-point.

EUROPEAN PHARMACOPOEIA 2.4.14. Sulfated ash
04/2010:20414 the residue with a small amount of sulfuric acid R (usually

1 mL), heat gently until white fumes are no longer evolved
and ignite at 600 ± 50 °C until the residue is completely
incinerated. Ensure that flames are not produced at any
time during the procedure. Allow the crucible to cool in a
desiccator over silica gel or other suitable desiccant, weigh it
2.4.14. SULFATED ASH (1)
again and calculate the percentage of residue.
Ignite a suitable crucible (for example, silica, platinum, If the amount of the residue so obtained exceeds the prescribed
porcelain or quartz) at 600 ± 50 °C for 30 min, allow to cool limit, repeat the moistening with sulfuric acid R and ignition,
in a desiccator over silica gel or other suitable desiccant and as previously, for 30 min periods until 2 consecutive weighings
weigh. Place the prescribed amount of the substance to be do not differ by more than 0.5 mg or until the percentage of
examined in the crucible and weigh. Moisten the substance to residue complies with the prescribed limit.
be examined with a small amount of sulfuric acid R (usually The amount of substance used for the test (usually 1-2 g) is
1 mL) and heat gently at as low a temperature as practicable chosen so that at the prescribed limit the mass of the residue
until the sample is thoroughly charred. After cooling, moisten (usually about 1 mg) can be measured with sufficient accuracy.
(1) This chapter has undergone pharmacopoeial harmonisation. See chapter 5.8. Pharmacopoeial harmonisation.

EUROPEAN PHARMACOPOEIA 2.4.24. Identification and control of residual solvents
04/2020:20424
Solvent solution (a). To 1.0 mL of Class 1 residual solvent
solution CRS, add 9 mL of dimethyl sulfoxide R and dilute
to 100.0 mL with water R. Dilute 1.0 mL of this solution to
100 mL with water R. Dilute 1.0 mL of this solution to 10.0 mL
2.4.24. IDENTIFICATION AND with water R.
CONTROL OF RESIDUAL SOLVENTS
The reference solutions correspond to the following limits :
The test procedures described in this general method may – benzene : 2 ppm ;
be used :
– carbon tetrachloride : 4 ppm ;
i. for the identification of the majority of Class 1 and Class 2
residual solvents in an active substance, excipient or medicinal – 1,2-dichloroethane : 5 ppm ;
product when the residual solvents are unknown ; – 1,1-dichloroethene : 8 ppm ;
ii. as a limit test for Class 1 and Class 2 solvents when present – 1,1,1-trichloroethane : 10 ppm.
in an active substance, excipient or medicinal product ;
iii. for the quantification of Class 2 solvents when the limits are Solvent solution (b). Dissolve appropriate quantities of the
greater than 1000 ppm (0.1 per cent) or for the quantification Class 2 residual solvents in dimethyl sulfoxide R and dilute to
of Class 3 solvents when required. 100.0 mL with the same solvent. Dilute in water R to give a
concentration of 1/20 of the limits stated in Table 2 (see 5.4.
Residual solvents).
Class 1, Class 2 and Class 3 residual solvents are listed in
general chapter 5.4. Residual solvents.
Three diluents are described for sample preparation and Solvent solution (c). Dissolve 1.00 g of the solvent or solvents
the conditions to be applied for head-space injection of the present in the substance to be examined in dimethyl sulfoxide R
gaseous sample onto the chromatographic system. Two or water R, if appropriate, and dilute to 100.0 mL with water R.
chromatographic systems are prescribed but System A Dilute to give a concentration of 1/20 of the limit(s) stated in
is preferred whilst System B is employed normally for Table 1 or 2 (see 5.4. Residual solvents).
confirmation of identity. The choice of sample preparation
procedure depends on the solubility of the substance to be Blank solution. Prepare as described for solvent solution (c)
examined and in certain cases the residual solvents to be but without the addition of solvent(s) (used to verify the
controlled. absence of interfering peaks).
Test solution. Introduce 5.0 mL of the sample solution and
The following residual solvents are not readily detected by 1.0 mL of the blank solution into an injection vial.
the head-space injection conditions described : formamide,
2-ethoxyethanol, 2-methoxyethanol, ethylene glycol, Reference solution (a) (Class 1). Introduce 1.0 mL of solvent
N-methylpyrrolidone and sulfolane. Other appropriate solution (a) and 5.0 mL of the appropriate diluent into an
procedures should be employed for the control of these injection vial.
residual solvents. Reference solution (a1) (Class 1). Introduce 5.0 mL of the
When the test procedure is applied quantitatively to control sample solution and 1.0 mL of solvent solution (a) into an
residual solvents in a substance, then it must be validated. injection vial.
Reference solution (b) (Class 2). Introduce 1.0 mL of solvent
solution (b) and 5.0 mL of the appropriate diluent into an
PROCEDURE injection vial.
Examine by gas chromatography with static head-space
injection (2.2.28).
Reference solution (c). Introduce 5.0 mL of the sample solution
Sample preparation 1. This is intended for the control of and 1.0 mL of solvent solution (c) into an injection vial.
residual solvents in water-soluble substances.
Reference solution (d). Introduce 1.0 mL of the blank solution
Sample solution (1). Dissolve 0.200 g of the substance to be and 5.0 mL of the appropriate diluent into an injection vial.
examined in water R and dilute to 20.0 mL with the same
solvent.
Close the vials with a tight rubber membrane stopper coated
with polytetrafluoroethylene and secure with an aluminium
Sample preparation 2. This is intended for the control of crimp cap. Shake to obtain a homogeneous solution.
residual solvents in water-insoluble substances.
Sample solution (2). Dissolve 0.200 g of the substance to The following static head-space injection conditions may be
be examined in dimethylformamide R (DMF) and dilute to used :
Sample preparation
procedure
Sample preparation 3. This is intended for the control of Operating parameters 1 2 3
N,N-dimethylacetamide and/or N,N-dimethylformamide,
when it is known or suspected that one or both of these Equilibration temperature (°C) 80 105 80
substances are present in the substance to be examined. Equilibration time (min) 60 45 45
Sample solution (3). Dissolve 0.200 g of the substance to be 85 110 105
Transfer-line temperature (°C)
examined in 1,3-dimethyl-2-imidazolidinone R (DMI) and
dilute to 20.0 mL with the same solvent. Carrier gas : nitrogen for chromatography R or helium for chromatography R
at an appropriate pressure
In some cases none of the above sample preparation
procedures are appropriate, in which case the diluent to be Pressurisation time (s) 30 30 30
used for the preparation of the sample solution and the static Injection volume (mL) 1 1 1
head-space conditions to be employed must be demonstrated
to be suitable.

2.4.24. Identification and control of residual solvents EUROPEAN PHARMACOPOEIA
The chromatographic procedure may be carried out using : the residual solvent peaks in the chromatograms obtained
SYSTEM A with reference solution (a) or (b), then the substance to be
examined meets the requirements of the test. If any peak in
– a fused-silica capillary or wide-bore column 30 m long the chromatogram obtained with the test solution corresponds
and 0.32 mm or 0.53 mm in internal diameter coated with to any of the residual solvent peaks obtained with reference
cyanopropyl(3)phenyl(3)methyl(94)polysiloxane R (film solution (a) or (b) then System B is to be employed.
thickness 1.8 μm or 3 μm);
Inject 1 mL of the gaseous phase of reference solution (a)

– nitrogen for chromatography R or helium for
onto the column described in System B and record the
chromatography R as the carrier gas, split ratio 1:5 with a
chromatogram under such conditions that the signal-to-noise
linear velocity of about 35 cm/s ;
ratio for benzene can be measured. The signal-to-noise ratio
must be at least 5. A typical chromatogram is shown in
– a flame-ionisation detector (a mass spectrometer may also Figure 2.4.24.-3.
be used or an electron-capture detector for the chlorinated
residual solvents of Class 1) ; Inject 1 mL of the gaseous phase of reference solution (a1)
onto the column described in System B. The peaks due to the
maintaining the temperature of the column at 40 °C for Class 1 residual solvents are still detectable.
20 min, then raising the temperature at a rate of 10 °C
per min to 240 °C and maintaining it at 240 °C for 20 min Inject 1 mL of the gaseous phase of reference solution (b)
and maintaining the temperature of the injection port at onto the column described in System B and record the
140 °C and that of the detector at 250 °C ; or, where there is chromatogram under such conditions that the resolution
interference from the matrix, use : between acetonitrile and 1,1,2-trichloroethene can be
determined. The system is suitable if the chromatogram
SYSTEM B obtained resembles the chromatogram shown in
Figure 2.4.24.-4 and the resolution between acetonitrile and
– a fused-silica capillary or wide-bore column 30 m long 1,1,2-trichloroethene is at least 1.0.
and 0.32 mm or 0.53 mm in internal diameter coated with
macrogol 20 000 R (film thickness 0.25 µm) ;
Inject 1 mL of the gaseous phase of the test solution onto
– nitrogen for chromatography R or helium for the column described in System B. If in the chromatogram
chromatography R as the carrier gas, split ratio 1:5 with a obtained, there is no peak which corresponds to any of the
linear velocity of about 35 cm/s ; residual solvent peaks in the chromatogram obtained with
the reference solution (a) or (b), then the substance to be
examined meets the requirements of the test. If any peak in
– a flame-ionisation detector (a mass spectrophotometer the chromatogram obtained with the test solution corresponds
may also be used or an electron-capture detector for the to any of the residual solvent peaks obtained with reference
chlorinated residual solvents of Class 1) ; solution (a) or (b) and confirms the correspondence obtained
when using System A, then proceed as follows.
maintaining the temperature of the column at 50 °C for

20 min, then raising the temperature at a rate of 6 °C per min Inject 1 mL of the gaseous phase of reference solution (c) onto
to 165 °C and maintaining it at 165 °C for 20 min and the column described for System A or System B. If necessary,
maintaining the temperature of the injection port at 140 °C adjust the sensitivity of the system so that the height of the
and that of the detector at 250 °C. peak corresponding to the identified residual solvent(s) is at
least 50 per cent of the full scale of the recorder.
Inject 1 mL of the gaseous phase of reference solution (a)

onto the column described in System A and record the Inject 1 mL of the gaseous phase of reference solution (d) onto
chromatogram under such conditions that the signal-to-noise the column. No interfering peaks should be observed.
ratio for 1,1,1-trichloroethane can be measured. The
signal-to-noise ratio must be at least 5. A typical
Inject 1 mL of the gaseous phase of the test solution and
chromatogram is shown in Figure 2.4.24.-1.
1 mL of the gaseous phase of reference solution (c) on to the
column. Repeat these injections twice more.
Inject 1 mL of the gaseous phase of reference solution (a1)
onto the column described in System A. The peaks due to the The mean area of the peak of the residual solvent(s) in the
Class 1 residual solvents are still detectable. chromatograms obtained with the test solution is not greater
than half the mean area of the peak of the corresponding
Inject 1 mL of the gaseous phase of reference solution (b) residual solvent(s) in the chromatograms obtained with
onto the column described in System A and record the reference solution (c). The test is not valid unless the relative
chromatogram under such conditions that the resolution standard deviation of the differences in areas between the
between acetonitrile and dichloromethane can be determined. analyte peaks obtained from 3 replicate paired injections of
The system is suitable if the chromatogram obtained reference solution (c) and the test solution, is at most 15 per
resembles the chromatogram shown in Figure 2.4.24.-2 and cent.
the resolution between acetonitrile and dichloromethane is
at least 1.0. A flow diagram of the procedure is shown in Figure 2.4.24.-5.
When a residual solvent (Class 2 or Class 3) is present
Inject 1 mL of the gaseous phase of the test solution onto at a level of 0.1 per cent or greater then the content may
the column described in System A. If in the chromatogram be quantitatively determined by the method of standard
obtained, there is no peak which corresponds to one of additions.

1. 1,1-dichloroethene 2. 1,1,1-trichloroethane 3. carbon tetrachloride 4. benzene 5. 1,2-dichloroethane
Figure 2.4.24.-1. – Typical chromatogram of Class 1 solvents using the conditions described for System A and Procedure 1.
Flame-ionisation detector
1. methanol 5. 1,2- 9. cyclohexane 13. 1,4-dioxane 17. methylbutylketone 20. cumene

dichloroethene
2. acetonitrile 6. nitromethane 10. 1,2- 14. pyridine 18. chlorobenzene 21. tetralin
dimethoxyethane
3. dichloromethane 7. tetrahydrofuran 11. 1,1,2- 15. methylisobutylke- 19. xylene mix
trichloroethene tone a. ethylbenzene
b. p-xylene
c. m-xylene
d. o-xylene
4. hexane 8. chloroform 12. methylcyclohexane 16. toluene
Figure 2.4.24.-2. – Chromatogram of Class 2 solvents (solvent solution (b)) using the conditions described for System A and
Procedure 1. Flame-ionisation detector

2.4.24. Identification and control of residual solvents EUROPEAN PHARMACOPOEIA
1. 1,1-dichloroethene 2. 1,1,1-trichloroethane 3. carbon tetrachloride 4. benzene 5. 1,2-dichloroethane
Figure 2.4.24.-3. – Chromatogram of Class 1 residual solvents using the conditions described for System B and Procedure 1.
Flame-ionisation detector
1. methanol 5. 1,2-dichloroethene 9. cyclohexane 13. 1,4-dioxane 17. methylbutylketone 20. cumene
2. acetonitrile 6. nitromethane 10. 1,2-dimethoxyethane 14. pyridine 18. chlorobenzene 21. tetralin (tR = 27 min)
3. dichloromethane 7. tetrahydrofuran 11. 1,1,2-trichloroethene 15. methylisobutylke- 19. xylene mix
tone a. ethylbenzene
b. p-xylene
c. m-xylene
d. o-xylene
4. hexane 8. chloroform 12. methylcyclohexane 16. toluene
Figure 2.4.24.-4. – Typical chromatogram of Class 2 residual solvents (solvent solution (b)) using the conditions described
for System B and Procedure 1. Flame-ionisation detector

Figure 2.4.24.-5. – Diagram relating to the identification of residual solvents and the application of limit tests

EUROPEAN PHARMACOPOEIA 2.5.8. Determination of primary aromatic amino-nitrogen
01/2008:20508 Determine the end-point electrometrically or by the use of

the prescribed indicator.
2.5.8. DETERMINATION OF PRIMARY

AROMATIC AMINO-NITROGEN
Dissolve the prescribed quantity of the substance to be
examined in 50 mL of dilute hydrochloric acid R or in another
prescribed solvent and add 3 g of potassium bromide R. Cool
in ice-water and titrate by slowly adding 0.1 M sodium nitrite
with constant stirring.

EUROPEAN PHARMACOPOEIA 2.5.37. MMS, EMS and IMS in methanesulfonic acid
04/2011:20537 Detection : mass spectrometer as described below ; adjust the

corrected 10.0 detector settings so as to comply with the system suitability
criteria :
– quadrupole mass spectrometer equipped with an electron

impact ionisation mode (70 eV) ;
2.5.37. METHYL, ETHYL AND – mass spectrometer parameters for the fragmentometric
ISOPROPYL METHANESULFONATE IN mode (single-ion monitoring (SIM)) set as follows :
METHANESULFONIC ACID
Duration of
The following method has been validated for the methyl, ethyl Substance m/z
monitoring
and isopropyl esters of methanesulfonic acid at concentrations tR between 7.0 min
in the range of 0.5 ppm to 100 ppm. Butyl methanesulfonate (BMS) 56
and 9.0 min
If it is intended to be used to determine levels of tR between 2.0 min
Methyl methanesulfonate (MMS) 80
methanesulfonic acid esters outside this validated range, for and 3.5 min
example in early steps of the synthesis prior to their removal, tR between 4.0 min
Ethyl methanesulfonate (EMS) 79
the concentration of the test solution has to be adjusted and 4.7 min
accordingly. tR between 4.7 min
Isopropyl methanesulfonate (IMS) 123
Gas chromatography (2.2.28) coupled with mass spectrometry and 5.5 min
(2.2.43).
Internal standard solution. Dilute 7 µL of butyl Injection : 2 µL.
methanesulfonate CRS (BMS) to 10.0 mL with methylene
chloride R. Dilute 10 µL of the solution to 100.0 mL with Relative retention with reference to the internal
methylene chloride R. standard (BMS) (retention time = about 7.6 min) :
MMS = about 0.3 ; EMS = about 0.5 ; IMS = about 0.6.
Test solution. Add 0.74 g of the substance to be examined to
10.0 mL of water R and extract with 10.0 mL of the internal System suitability :
standard solution. Allow to separate and transfer the organic
layer to a vial containing anhydrous sodium sulfate R. Shake – resolution : minimum 3.0 between the peaks due to EMS
and filter. and IMS in the chromatogram obtained with reference
Reference solution (a). Dissolve 50 mg each of methyl solution (a) ;
methanesulfonate R (MMS), ethyl methanesulfonate R (EMS)
and isopropyl methanesulfonate R (IMS) in the internal – signal-to-noise ratio : minimum 10 for the peaks due to
standard solution and dilute to 50.0 mL with the same MMS, EMS and IMS in the chromatogram obtained with
solution. Dilute 74 µL of the solution to 10.0 mL with the reference solution (b).
internal standard solution. Dilute 100 µL of this solution to
10.0 mL with the internal standard solution. Calculate the content of MMS, EMS or IMS in parts per
Reference solution (b). Dilute 3.0 mL of reference solution (a) million using the following expression :
to 10.0 mL with the internal standard solution.
Column : A 2 ´ I1 ´ W1 ´ C ´ 0.148
– material : fused silica ; A1 ´ I 2 ´ W2
– size : l = 15 m, Ø = 0.25 mm ;
– stationary phase : methylpolysiloxane R (film thickness
A1 = area of the peak due to MMS, EMS or IMS
1 µm). in the chromatogram obtained with reference
solution (a) ;
Carrier gas : helium for chromatography R.
A2 = area of the peak due to MMS, EMS or IMS in the
Flow rate : 1 mL/min. chromatogram obtained with the test solution ;
Pulsed splitless : 250 kPa, 0.25 min.
C = percentage content of MMS, EMS or IMS ;
Temperature :
I1 = area of the peak due to the internal standard
Time Temperature
in the chromatogram obtained with reference
(min) (°C)
solution (a) ;
Column 0-1 55
I2 = area of the peak due to the internal standard in the
1-9 55 → 135 chromatogram obtained with the test solution ;
Injection port 240 W1 = mass of MMS, EMS or IMS used to prepare
reference solution (a), in milligrams ;
Detector : transfer line 280
W2 = mass of the substance to be examined in the test
source 230 solution, in milligrams ;
analyser 150 0.148 = dilution factor.

EUROPEAN PHARMACOPOEIA 2.5.38. MMS, EMS and IMS in active substances
04/2016:20538 The use of an inert inlet liner without glass wool significantly
corrected 10.0 reduces the effect of carry-over between the injections.
Flow rate : 0.5 mL/min.
Split ratio : 1:20.
Static head-space conditions that may be used :
2.5.38. METHYL, ETHYL AND – equilibration temperature : 60 °C ;
ISOPROPYL METHANESULFONATE IN –– equilibration time : 30 min ;
transfer-line temperature : 120 °C.
ACTIVE SUBSTANCES Temperature :
The following general method has been validated for the Time Temperature
determination of methyl, ethyl and isopropyl esters of (min) (°C)
methanesulfonic acid (in concentrations between 0.2 ppm and Column 0-1 40
5 ppm) in betahistine mesilate.
1 - 10 40 → 130
If it is intended to use the method for other active substances,
particularly those that contain different concentrations of the Injection port 220
methanesulfonic acid esters, the concentrations of the test Detector 280
transfer line
solution and reference solutions must be adjusted accordingly
and the method must be suitably validated. source 250
METHOD analyser 200
Head-space gas chromatography (2.2.28) coupled with mass At the end of analysis, the temperature of the column is raised
spectrometry (2.2.43). Prepare the test solution and reference to 240 °C and maintained at this temperature for 7 min.
solutions immediately before use.
Detection : mass spectrometer as described below ; adjust the
Solvent mixture : water R, acetonitrile R (20:80 V/V). The use detector settings so as to comply with the system suitability
of acetonitrile of appropriate purity is essential. criteria ; alternatively a suitable electron-capture detector may
Solution A. Dissolve with the aid of ultrasound 30 mg of be used :
anhydrous sodium thiosulfate R and 60.0 g of sodium iodide R – quadrupole mass spectrometer equipped with an electron
in water R and dilute to 50.0 mL with the same solvent. impact ionisation mode (70 eV) ;
Internal standard solution. Dilute 10 µL of butyl – mass spectrometer parameters for the fragmentometric
methanesulfonate CRS (BMS) to 10.0 mL with the solvent mode (single-ion monitoring (SIM)) set as follows :
mixture. Dilute 20 µL of the solution to 100.0 mL with the
solvent mixture. Substance
Quantitation ion Qualification ion
(m/z) (m/z)
Blank solution. Introduce 0.50 mL of solution A and 0.50 mL
of the internal standard solution into a headspace vial and seal Butyl iodide (BuI)* 184 127
the vial immediately with a polytetrafluoroethylene-coated Methyl iodide (MeI)* 142 127
silicon membrane and an aluminium cap.
Ethyl iodide (EtI)* 156 127
Test solution. Weigh 25.0 mg of the substance to be examined
into a 20 mL headspace vial. Add 0.50 mL of solution A and Isopropyl iodide (iPrI)* 170 127
0.50 mL of the internal standard solution and seal the vial
* formed from BMS, MMS, EMS and IMS in the derivatisation reaction.
immediately with a polytetrafluoroethylene-coated silicon
membrane and an aluminium cap. Injection : 1 mL of the gas phase of the test solution, reference
Following the derivatisation reaction, a precipitate may be solutions (b) and (c) and the blank solution.
observed, however this does not affect the validity of the Relative retention with reference to the internal standard
quantification. (BuI) (retention time = about 8.5 min) : MeI = about 0.51 ;
Reference solution (a). Dissolve 25.0 mg each of methyl EtI = about 0.63 ; iPrI = about 0.68.
methanesulfonate R (MMS), ethyl methanesulfonate R (EMS) System suitability :
and isopropyl methanesulfonate R (IMS) in toluene R and – resolution : minimum 1.5 between the peaks due to EtI
dilute to 5.0 mL with the same solvent. Dilute 50 µL of the and iPrI in the chromatogram obtained with reference
solution to 25.0 mL with the internal standard solution. solution (c);
Reference solution (b). Dilute 20 µL of reference solution (a) – signal-to-noise ratio : minimum 10 for the peak due to each
to 20.0 mL with the internal standard solution. Introduce alkyl iodide in the chromatogram obtained with reference
0.50 mL of this solution and 0.50 mL of solution A into a solution (b).
20 mL headspace vial and seal the vial immediately with a
polytetrafluoroethylene-coated silicon membrane and an Calculate the content in parts per million of each alkyl
aluminium cap. methanesulfonate using the following expression :
Reference solution (c). Dilute 500 μL of reference solution (a) A 2 ´ I1 ´ W1 ´ C ´ 0.05
to 20.0 mL with the internal standard solution. Introduce A1 ´ I 2 ´ W2
0.50 mL of this solution and 0.50 mL of solution A into a
20 mL headspace vial and seal the vial immediately with a A1 = area of the peak due to each alkyl iodide in
polytetrafluoroethylene-coated silicon membrane and an the chromatogram obtained with reference
aluminium cap. solution (c);
Column : A2 = area of the peak due to each alkyl iodide in the
– material : fused silica ; chromatogram obtained with the test solution ;
– size : l = 30 m, Ø = 0.25 mm ; C = percentage content of each ester ;
– stationary phase : polar-deactivated macrogol R (film I1 = area of the peak due to the internal standard
thickness 1 µm). in the chromatogram obtained with reference
Carrier gas : helium for chromatography R. solution (c);

2.5.38. MMS, EMS and IMS in active substances EUROPEAN PHARMACOPOEIA
I2 = area of the peak due to the internal standard in the W1 = mass of each ester used to prepare reference
chromatogram obtained with the test solution ; solution (a), in milligrams ;
W2 = mass of the substance to be examined in the test
solution, in milligrams ;
0.05 = dilution factor.

EUROPEAN PHARMACOPOEIA 2.5.39. Methanesulfonyl chloride in methanesulfonic acid
04/2013:20539 At the end of analysis the temperature of the column is raised

corrected 10.0 to 270 °C and maintained at this temperature for 8 min.
Detection : mass spectrometer as described below ; adjust the
detector settings so as to comply with the system suitability
criteria :
2.5.39. METHANESULFONYL – quadrupole mass spectrometer equipped with an electron
impact ionisation mode (70 eV) ;
CHLORIDE IN METHANESULFONIC
ACID – mass spectrometer parameters for the fragmentometric
mode (single-ion monitoring (SIM)) set as follows :
The following method has been validated for the determination
of methanesulfonyl chloride in methanesulfonic acid at Substance m/z Duration of monitoring
concentrations in the range of 0.05 ppm to 50 ppm. Methanesulfonyl 79 tR between 3.3 min and 6.0 min
Gas chromatography (2.2.28) coupled with mass spectrometry chloride
(2.2.43). Butyl methane- 56 tR between 6.0 min and 8.0 min
Internal standard solution. Dissolve 7 µL of butyl sulfonate (BMS)
methanesulfonate CRS (BMS) in methylene chloride R and
dilute to 10.0 mL with the same solvent. Dilute 5.0 mL of this Injection : 5 µL of the test solution, reference solutions (b)
solution to 50.0 mL with methylene chloride R. and (c), the internal standard solution and methylene
chloride R.
Test solution. To 5 mL of water R, add 7.4 g of the substance
to be examined and mix slowly. After cooling, add 5.0 mL Relative retention with reference to the internal standard
of methylene chloride R and 100 µL of the internal standard (BMS) (retention time = about 7.2 min) : methanesulfonyl
solution and shake. Allow to separate and transfer the chloride = about 0.68.
organic layer to a vial containing 1 g of anhydrous sodium System suitability :
sulfate R. Repeat the extraction twice with 5.0 mL of methylene
chloride R each time, combine the organic layers and filter. – in the chromatogram obtained with the internal standard
Reference solution (a). Dissolve 50.0 mg of methanesulfonyl solution, there is no peak with the same retention time as
chloride R in methylene chloride R and dilute to 10.0 mL with methanesulfonyl chloride ;
the same solvent. Dilute 1.0 mL of the solution to 10.0 mL – resolution : minimum 5.0 between the peaks due to
with methylene chloride R. Dilute 300 µL of this solution to methanesulfonyl chloride and BMS in the chromatogram
10.0 mL with methylene chloride R. obtained with reference solution (b) ;
Reference solution (b). Dilute 500 µL of reference solution (a)
and 100 µL of the internal standard solution to 15.0 mL with – signal-to-noise ratio : minimum 10 for the peak due to
methylene chloride R. methanesulfonyl chloride in the chromatogram obtained
with reference solution (c).
Reference solution (c). Dilute 25 µL of reference solution (a)
and 100 µL of the internal standard solution to 15.0 mL with Calculate the content of methanesulfonyl chloride in parts per
methylene chloride R. million using the following expression :
Column :
– material : fused silica ; A 2 ´ I1 ´ W1 ´ C ´ 1.5
– size : l = 15 m, Ø = 0.25 mm ; A1 ´ I 2 ´ W2
– stationary phase : methylpolysiloxane R (film thickness
1 µm).
A1 = area of the peak due to methanesulfonyl chloride
in the chromatogram obtained with reference
Carrier gas : helium for chromatography R. solution (b) ;
Flow rate : 1 mL/min. A2 = area of the peak due to methanesulfonyl chloride in
Pulsed splitless : 60 kPa, 0.1 min. the chromatogram obtained with the test solution ;
Temperature : C = percentage content of methanesulfonyl chloride ;
Time Temperature
(min) (°C)
I1 = area of the peak due to BMS in the chromatogram
obtained with reference solution (b) ;
Column 0-4 40
I2 = area of the peak due to BMS in the chromatogram
4-8 40 → 200 obtained with the test solution ;
Injection port 240 W1 = mass of methanesulfonyl chloride used to prepare
Detector : 280
reference solution (a), in milligrams ;
transfer line
source
W2 = mass of the sample in the test solution, in
230
milligrams ;
analyser 150 1.5 = dilution factor.

EUROPEAN PHARMACOPOEIA 2.6.1. Sterility
04/2011:20601 the containers in a water-bath or in free-flowing steam until

corrected 7.7 the pink colour disappears and cooling quickly, taking care to
prevent the introduction of non-sterile air into the container.
Do not use the medium for a longer storage period than has
been validated.
Fluid thioglycollate medium is to be incubated at 30-35 °C.
2.6.1. STERILITY(1) For products containing a mercurial preservative that
The test is applied to substances, preparations or articles cannot be tested by the membrane-filtration method, fluid
which, according to the Pharmacopoeia, are required to be thioglycollate medium incubated at 20-25 °C may be used
sterile. However, a satisfactory result only indicates that no instead of soya-bean casein digest medium provided that it
contaminating micro-organism has been found in the sample has been validated as described in growth promotion test.
examined in the conditions of the test. Where prescribed or justified and authorised, the following
alternative thioglycollate medium may be used. Prepare a
PRECAUTIONS AGAINST MICROBIAL mixture having the same composition as that of the fluid
CONTAMINATION thioglycollate medium, but omitting the agar and the resazurin
The test for sterility is carried out under aseptic conditions. sodium solution, sterilise as directed above. The pH after
In order to achieve such conditions, the test environment sterilisation is 7.1 ± 0.2. Heat in a water-bath prior to use and
has to be adapted to the way in which the sterility test is incubate at 30-35 °C under anaerobic conditions.
performed. The precautions taken to avoid contamination are Soya-bean casein digest medium
such that they do not affect any micro-organisms which are to Pancreatic digest of casein 17.0 g
be revealed in the test. The working conditions in which the
tests are performed are monitored regularly by appropriate Papaic digest of soya-bean meal 3.0 g
sampling of the working area and by carrying out appropriate Sodium chloride 5.0 g
controls.
Dipotassium hydrogen phosphate 2.5 g
CULTURE MEDIA AND INCUBATION TEMPERATURES
Glucose monohydrate/Glucose 2.5 g/2.3 g
Media for the test may be prepared as described below, or
equivalent commercial media may be used provided that they Water R 1000 mL
comply with the growth promotion test. pH after sterilisation 7.3 ± 0.2
The following culture media have been found to be suitable for
the test for sterility. Fluid thioglycollate medium is primarily Dissolve the solids in water R, warming slightly to effect
intended for the culture of anaerobic bacteria ; however, it will solution. Cool the solution to room temperature. Add 1 M
also detect aerobic bacteria. Soya-bean casein digest medium sodium hydroxide, if necessary, so that after sterilisation the
is suitable for the culture of both fungi and aerobic bacteria. solution will have a pH of 7.3 ± 0.2. Filter, if necessary, to
clarify, distribute into suitable vessels and sterilise using a
Fluid thioglycollate medium
validated process. Store at a temperature between 2 °C and
L-Cystine 0.5 g 25 °C in a sterile well-closed container, unless it is intended for
Agar 0.75 g immediate use. Do not use the medium for a longer storage
period than has been validated.
Sodium chloride 2.5 g
Soya-bean casein digest medium is to be incubated at 20-25 °C.
Glucose monohydrate/Glucose 5.5 g/5.0 g The media used comply with the following tests, carried
Yeast extract (water-soluble) 5.0 g out before or in parallel with the test on the product to be
examined.
Pancreatic digest of casein 15.0 g
Sterility. Incubate portions of the media for 14 days. No
Sodium thioglycollate or 0.5 g growth of micro-organisms occurs.
Thioglycollic acid 0.3 mL Growth promotion test of aerobes, anaerobes and fungi.
1.0 mL
Test each batch of ready-prepared medium and each batch of
Resazurin sodium solution (1 g/L of resazurin
sodium), freshly prepared medium prepared either from dehydrated medium or from
Water R 1000 mL
ingredients. Suitable strains of micro-organisms are indicated
in Table 2.6.1.-1.
pH after sterilisation 7.1 ± 0.2 Inoculate portions of fluid thioglycollate medium with a
small number (not more than 100 CFU) of the following
Mix the L-cystine, agar, sodium chloride, glucose,
micro-organisms, using a separate portion of medium for
water-soluble yeast extract and pancreatic digest of casein
each of the following species of micro-organism : Clostridium
with the water R and heat until solution is effected. Dissolve
sporogenes, Pseudomonas aeruginosa, Staphylococcus aureus.
the sodium thioglycollate or thioglycollic acid in the solution
Inoculate portions of soya-bean casein digest medium with
and, if necessary, add 1 M sodium hydroxide so that, after
a small number (not more than 100 CFU) of the following
sterilisation, the solution will have a pH of 7.1 ± 0.2. If
micro-organisms, using a separate portion of medium for
filtration is necessary, heat the solution again without boiling
each of the following species of micro-organism : Aspergillus
and filter while hot through moistened filter paper. Add the
brasiliensis, Bacillus subtilis, Candida albicans. Incubate for
resazurin sodium solution, mix and place the medium in
not more than 3 days in the case of bacteria and not more
suitable vessels which provide a ratio of surface to depth of
than 5 days in the case of fungi.
medium such that not more than the upper half of the medium
has undergone a colour change indicative of oxygen uptake at Seed lot culture maintenance techniques (seed-lot systems) are
the end of the incubation period. Sterilise using a validated used so that the viable micro-organisms used for inoculation
process. If the medium is stored, store at a temperature are not more than 5 passages removed from the original
between 2 °C and 25 °C in a sterile, airtight container. If master seed-lot.
more than the upper one-third of the medium has acquired The media are suitable if a clearly visible growth of the
a pink colour, the medium may be restored once by heating micro-organisms occurs.

2.6.1. Sterility EUROPEAN PHARMACOPOEIA
Table 2.6.1.-1. – Strains of the test micro-organisms suitable for use in the growth promotion test and the method suitability test
Aerobic bacteria
Staphylococcus aureus ATCC 6538, CIP 4.83, NCTC 10788, NCIMB 9518, NBRC 13276

Bacillus subtilis ATCC 6633, CIP 52.62, NCIMB 8054, NBRC 3134
Pseudomonas aeruginosa ATCC 9027, NCIMB 8626, CIP 82.118, NBRC 13275
Anaerobic bacterium
Clostridium sporogenes ATCC 19404, CIP 79.3, NCTC 532, ATCC 11437, NBRC 14293
Fungi
Candida albicans ATCC 10231, IP 48.72, NCPF 3179, NBRC 1594
Aspergillus brasiliensis ATCC 16404, IP 1431.83, IMI 149007, NBRC 9455
METHOD SUITABILITY TEST alcoholic solutions and cellulose acetate filters, for example,
Carry out a test as described below under Test for sterility offor strongly alcoholic solutions. Specially adapted filters may
the product to be examined using exactly the same methods be needed for certain products, e.g. for antibiotics.
except for the following modifications. The technique described below assumes that membranes
Membrane filtration. After transferring the contents of the about 50 mm in diameter will be used. If filters of a different
container or containers to be tested to the membrane add an diameter are used the volumes of the dilutions and the
inoculum of a small number of viable micro-organisms (not washings should be adjusted accordingly. The filtration
more than 100 CFU) to the final portion of sterile diluent apparatus and membrane are sterilised by appropriate means.
used to rinse the filter. The apparatus is designed so that the solution to be examined
can be introduced and filtered under aseptic conditions ; it
Direct inoculation. After transferring the content of the permits the aseptic removal of the membrane for transfer to
container or containers to be tested (for catgut and other the medium or it is suitable for carrying out the incubation
surgical sutures for veterinary use : strands) to the culture after adding the medium to the apparatus itself.
medium add an inoculum of a small number of viable
micro-organisms (not more than 100 CFU) to the medium. Aqueous solutions. If appropriate, transfer a small quantity
of a suitable, sterile diluent such as a 1 g/L neutral solution
In both cases use the same micro-organisms as those described of meat or casein peptone pH 7.1 ± 0.2 onto the membrane
above under Growth promotion test of aerobes, anaerobes and in the apparatus and filter. The diluent may contain suitable
fungi. Perform a growth promotion test as a positive control. neutralising substances and/or appropriate inactivating
Incubate all the containers containing medium for not more substances for example in the case of antibiotics.
than 5 days.
If clearly visible growth of micro-organisms is obtained after Transfer the contents of the container or containers to be
the incubation, visually comparable to that in the control tested to the membrane or membranes, if necessary after
vessel without product, either the product possesses no diluting to the volume used in the method suitability test
antimicrobial activity under the conditions of the test or suchwith the chosen sterile diluent but in any case using not less
than the quantities of the product to be examined prescribed
activity has been satisfactorily eliminated. The test for sterility
may then be carried out without further modification. in Table 2.6.1.-2. Filter immediately. If the product has
antimicrobial properties, wash the membrane not less than
If clearly visible growth is not obtained in the presence of the
3 times by filtering through it each time the volume of the
product to be tested, visually comparable to that in the control
chosen sterile diluent used in the method suitability test. Do
vessels without product, the product possesses antimicrobial not exceed a washing cycle of 5 times 100 mL per filter, even
activity that has not been satisfactorily eliminated under the if during the method suitability test it has been demonstrated
conditions of the test. Modify the conditions in order to that such a cycle does not fully eliminate the antimicrobial
eliminate the antimicrobial activity and repeat the method activity. Transfer the whole membrane to the culture medium
suitability test. or cut it aseptically into 2 equal parts and transfer one half
This method suitability test is performed : to each of 2 suitable media. Use the same volume of each
a) when the test for sterility has to be carried out on a new medium as in the method suitability test. Alternatively,
product ; transfer the medium onto the membrane in the apparatus.
b) whenever there is a change in the experimental conditions Incubate the media for not less than 14 days.
of the test. Soluble solids. Use for each medium not less than the quantity
The method suitability test may be performed simultaneously prescribed in Table 2.6.1.-2 of the product dissolved in
with the test for sterility of the product to be examined. a suitable solvent such as the solvent provided with the
preparation, water for injections, saline or a 1 g/L neutral
TEST FOR STERILITY OF THE PRODUCT TO BE solution of meat or casein peptone and proceed with the test
EXAMINED as described above for aqueous solutions using a membrane
The test may be carried out using the technique of membrane appropriate to the chosen solvent.
filtration or by direct inoculation of the culture media Oils and oily solutions. Use for each medium not less than
with the product to be examined. Appropriate negative the quantity of the product prescribed in Table 2.6.1.-2. Oils
controls are included. The technique of membrane filtration and oily solutions of sufficiently low viscosity may be filtered
is used whenever the nature of the product permits, that without dilution through a dry membrane. Viscous oils may
is, for filterable aqueous preparations, for alcoholic or oily be diluted as necessary with a suitable sterile diluent such as
preparations and for preparations miscible with or soluble in isopropyl myristate shown not to have antimicrobial activity
aqueous or oily solvents provided these solvents do not have in the conditions of the test. Allow the oil to penetrate the
an antimicrobial effect in the conditions of the test. membrane by its own weight then filter, applying the pressure
Membrane filtration. Use membrane filters having a nominal or suction gradually. Wash the membrane at least 3 times
pore size not greater than 0.45 µm whose effectiveness to by filtering through it each time about 100 mL of a suitable
retain micro-organisms has been established. Cellulose nitrate sterile solution such as 1 g/L neutral meat or casein peptone
filters, for example, are used for aqueous, oily and weakly containing a suitable emulsifying agent at a concentration

EUROPEAN PHARMACOPOEIA 2.6.1. Sterility
Table 2.6.1.-2. – Minimum quantity to be used for each medium

Minimum quantity to be used for each medium unless
Quantity per container
otherwise justified and authorised
Liquids
– less than 1 mL The whole contents of each container

– 1-40 mL Half the contents of each container but not less than 1 mL
– greater than 40 mL and not greater than 100 mL 20 mL
– greater than 100 mL 10 per cent of the contents of the container but not less than 20 mL
Antibiotic liquids 1 mL
Insoluble preparations, creams and ointments to be suspended or emulsified Use the contents of each container to provide not less than 200 mg
Solids
– less than 50 mg The whole contents of each container
– 50 mg or more but less than 300 mg Half the contents of each container but not less than 50 mg
– 300 mg to 5 g 150 mg
– greater than 5 g 500 mg
Catgut and other surgical sutures for veterinary use 3 sections of a strand (each 30 cm long)
shown to be appropriate in the method suitability test, for aseptic precautions and remove 3 sections of the strand for
example polysorbate 80 at a concentration of 10 g/L. Transfer each culture medium. Carry out the test on 3 sections, each
the membrane or membranes to the culture medium or media 30 cm long, cut off from the beginning, the centre and the
or vice versa as described above for aqueous solutions, and end of the strand. Use whole strands from freshly opened
incubate at the same temperatures and for the same times. cassette packs. Transfer each section of the strand to the
Ointments and creams. Use for each medium not less than selected medium. Use sufficient medium to cover adequately
the quantities of the product prescribed in Table 2.6.1.-2. the material to be tested (20 mL to 150 mL).
Ointments in a fatty base and emulsions of the water-in-oil
type may be diluted to 1 per cent in isopropyl myristate as OBSERVATION AND INTERPRETATION OF RESULTS
described above, by heating, if necessary, to not more than
40 °C. In exceptional cases it may be necessary to heat to not At intervals during the incubation period and at its conclusion,
more than 44 °C. Filter as rapidly as possible and proceed as examine the media for macroscopic evidence of microbial
described above for oils and oily solutions. growth. If the material being tested renders the medium
turbid so that the presence or absence of microbial growth
Direct inoculation of the culture medium. Transfer the cannot be readily determined by visual examination, 14 days
quantity of the preparation to be examined prescribed in after the beginning of incubation transfer portions (each not
Table 2.6.1.-2 directly into the culture medium so that the less than 1 mL) of the medium to fresh vessels of the same
volume of the product is not more than 10 per cent of the medium and then incubate the original and transfer vessels
volume of the medium, unless otherwise prescribed. for not less than 4 days.
If the product to be examined has antimicrobial activity, carry
out the test after neutralising this with a suitable neutralising If no evidence of microbial growth is found, the product to be
substance or by dilution in a sufficient quantity of culture examined complies with the test for sterility. If evidence of
medium. When it is necessary to use a large volume of the microbial growth is found the product to be examined does
product it may be preferable to use a concentrated culture not comply with the test for sterility, unless it can be clearly
medium prepared in such a way that it takes account of the demonstrated that the test was invalid for causes unrelated
subsequent dilution. Where appropriate, the concentrated to the product to be examined. The test may be considered
medium may be added directly to the product in its container. invalid only if one or more of the following conditions are
fulfilled :
Oily liquids. Use media to which have been added a suitable
emulsifying agent at a concentration shown to be appropriate a) the data of the microbiological monitoring of the sterility
in the method suitability test, for example polysorbate 80 at a testing facility show a fault ;
concentration of 10 g/L.
b) a review of the testing procedure used during the test in
Ointments and creams. Prepare by diluting to about 1 in 10 question reveals a fault ;
by emulsifying with the chosen emulsifying agent in a suitable
sterile diluent such as a 1 g/L neutral solution of meat or c) microbial growth is found in the negative controls ;
casein peptone. Transfer the diluted product to a medium not d) after determination of the identity of the micro-organisms
containing an emulsifying agent. isolated from the test, the growth of this species or these
Incubate the inoculated media for not less than 14 days. species may be ascribed unequivocally to faults with respect
Observe the cultures several times during the incubation to the material and/or the technique used in conducting the
period. Shake cultures containing oily products gently each sterility test procedure.
day. However when fluid thioglycollate medium is used for
the detection of anaerobic micro-organisms keep shaking If the test is declared to be invalid it is repeated with the same
or mixing to a minimum in order to maintain anaerobic number of units as in the original test.
conditions. If no evidence of microbial growth is found in the repeat
Catgut and other surgical sutures for veterinary use. Use for test the product examined complies with the test for sterility.
each medium not less than the quantities of the product If microbial growth is found in the repeat test the product
prescribed in Table 2.6.1.-2. Open the sealed package using examined does not comply with the test for sterility.

2.6.1. Sterility EUROPEAN PHARMACOPOEIA
Table 2.6.1.-3. – Minimum number of items to be tested

Minimum number of items to be tested for each medium, unless
Number of items in the batch*
otherwise justified and authorised**
Parenteral preparations
– Not more than 100 containers 10 per cent or 4 containers, whichever is the greater
– More than 100 but not more than 500 containers 10 containers
2 per cent or 20 containers (10 containers for large-volume parenterals)
– More than 500 containers
whichever is less
Ophthalmic and other non-injectable preparations
– Not more than 200 containers 5 per cent or 2 containers, whichever is the greater
– More than 200 containers 10 containers
– If the product is presented in the form of single-dose containers, apply
the scheme shown above for preparations for parenteral administration
2 per cent or 5 packages whichever is the greater, up to a maximum total of
Catgut and other surgical sutures for veterinary use
20 packages
Bulk solid products
– Up to 4 containers Each container
– More than 4 containers but not more than 50 containers 20 per cent or 4 containers, whichever is the greater
– More than 50 containers 2 per cent or 10 containers, whichever is the greater
* If the batch size is not known, use the maximum number of items prescribed.
**If the contents of one container are enough to inoculate the 2 media, this column gives the number of containers needed for both the media
together.
APPLICATION OF THE TEST TO PARENTERAL sterility are carried out on the same sample of the product to
PREPARATIONS, OPHTHALMIC AND OTHER be examined. When the volume or the quantity in a single
NON-INJECTABLE PREPARATIONS REQUIRED TO container is insufficient to carry out the tests, the contents of 2
COMPLY WITH THE TEST FOR STERILITY or more containers are used to inoculate the different media.
When using the technique of membrane filtration, use,
whenever possible, the whole contents of the container, but MINIMUM NUMBER OF ITEMS TO BE TESTED
not less than the quantities indicated in Table 2.6.1.-2, diluting
where necessary to about 100 mL with a suitable sterile The minimum number of items to be tested in relation to the
solution, such as 1 g/L neutral meat or casein peptone. size of the batch is given in Table 2.6.1.-3.
When using the technique of direct inoculation of media,
use the quantities shown in Table 2.6.1.-2, unless otherwise Guidelines on the test for sterility are given in general
justified and authorised. The tests for bacterial and fungal chapter 5.1.9.

EUROPEAN PHARMACOPOEIA 2.6.14. Bacterial endotoxins
01/2018:20614 3. PREPARATION OF THE STANDARD ENDOTOXIN

STOCK SOLUTION
The standard endotoxin stock solution is prepared from
an endotoxin reference standard that has been calibrated
against the International Standard, for example endotoxin
2.6.14. BACTERIAL ENDOTOXINS(1) standard BRP.
The test for bacterial endotoxins (BET) is used to detect Endotoxin is expressed in International Units (IU). The
or quantify endotoxins from gram-negative bacteria using equivalence in IU of the International Standard is stated by
amoebocyte lysate from the horseshoe crab (Limulus the World Health Organization.
polyphemus or Tachypleus tridentatus). There are 3 techniques NOTE : one International Unit (IU) of endotoxin is equal to
for this test : the gel-clot technique, which is based on one Endotoxin Unit (E.U.).
gel formation ; the turbidimetric technique, based on the
development of turbidity after cleavage of an endogenous Follow the specifications in the package leaflet and on the
substrate ; and the chromogenic technique, based on label for preparation and storage of the standard endotoxin
the development of colour after cleavage of a synthetic stock solution.
peptide-chromogen complex.
The following 6 methods are described in the present chapter : 4. PREPARATION OF THE STANDARD ENDOTOXIN
SOLUTIONS
Method A. Gel-clot method : limit test
After vigorously mixing the standard endotoxin stock solution,
Method B. Gel-clot method : quantitative test prepare appropriate serial dilutions of this solution using
Method C. Turbidimetric kinetic method water for BET.
Method D. Chromogenic kinetic method Use the solutions as soon as possible to avoid loss of activity
by adsorption.
Method E. Chromogenic end-point method
Method F. Turbidimetric end-point method 5. PREPARATION OF THE TEST SOLUTIONS
Proceed by any of the 6 methods for the test. In the event Prepare the test solutions by dissolving or diluting active
of doubt or dispute, the final decision is made based upon substances or medicinal products using water for BET.
method A unless otherwise indicated in the monograph. Some substances or preparations may be more appropriately
dissolved or diluted in other aqueous solutions. If necessary,
The test is carried out in a manner that avoids endotoxin
adjust the pH of the test solution (or dilution thereof) so that
contamination.
the pH of the mixture of the lysate and test solution falls within
1. APPARATUS the pH range specified by the lysate manufacturer, usually
6.0 to 8.0. The pH may be adjusted by the use of acid, base or
Depyrogenate all glassware and other heat-stable apparatus a suitable buffer, as recommended by the lysate manufacturer.
in a hot-air oven using a validated process. A commonly Acids and bases may be prepared from concentrates or solids
used minimum time and temperature is 30 min at 250 °C. If with water for BET in containers free of detectable endotoxin.
employing plastic apparatus, such as microtitre plates and Buffers must be validated to be free of detectable endotoxin
pipette tips for automatic pipetters, use apparatus shown to and interfering factors.
be free of detectable endotoxin and which does not interfere
in the test.
6. DETERMINATION OF THE MAXIMUM VALID
NOTE : in this chapter, the term ‘tube’ includes all types of DILUTION
receptacles, for example microtitre plate wells.
The Maximum Valid Dilution (MVD) is the maximum
2. REAGENTS, TEST SOLUTIONS allowable dilution of a sample at which the endotoxin limit
(1) Amoebocyte lysate can be determined. Determine the MVD using the following
formulae :
Amoebocyte lysate is a lyophilised product obtained from
amoebocyte lysate from the horseshoe crab (Limulus
endotoxin limit × concentration of test solution
polyphemus or Tachypleus tridentatus). This reagent refers MVD =
λ
only to a product manufactured in accordance with the
regulations of the competent authority. Endotoxin limit : the endotoxin limit for active substances
NOTE : amoebocyte lysate reacts with some β-glucans in administered parenterally, defined on the basis of dose, is
addition to endotoxins. Amoebocyte lysate preparations which equal to :
do not react with glucans are available ; they are prepared by
removing from amoebocyte lysate the G factor, which reacts K
with glucans, or by inhibiting the G factor reacting system M
of amoebocyte lysate. These preparations may be used for
endotoxin testing in the presence of glucans. K = threshold pyrogenic dose of endotoxin per
kilogram of body mass,
(2) Lysate solution
M = maximum recommended bolus dose of product
Dissolve amoebocyte lysate in water for BET or in a buffer, as per kilogram of body mass.
recommended by the lysate manufacturer, by gentle stirring.
Store the reconstituted lysate, refrigerated or frozen, as When the product is to be injected at frequent intervals
indicated by the manufacturer. or infused continuously, M is the maximum total dose
(3) Water for BET (water for bacterial endotoxins test) administered in a single hour period.
Water for injections R or water produced by other procedures The endotoxin limit for active substances administered pa-
that shows no reaction with the lysate employed at the renterally is specified in units such as IU/mL, IU/mg, IU/Unit
detection limit of the reagent. of biological activity, etc., in monographs.

2.6.14. Bacterial endotoxins EUROPEAN PHARMACOPOEIA
Concentration of test solution : concentration by calculting the mean of the logarithms of

– mg/mL if the endotoxin limit is specified by mass (IU/mg), the end-point concentrations of the 4 dilution series, take
the antilogarithm of this value, as indicated by the following
– Units/mL if the endotoxin limit is specified by unit of
expression :
biological activity (IU/Unit),
– mL/mL if the endotoxin limit is specified by volume Geometric mean end-point concentration = antilog
åe
(IU/mL). f
λ = the labelled lysate sensitivity in the gel-clot å e = sum of the log10 end-point concentrations of the
technique (IU/mL) or the lowest concentration dilution series used,
used in the standard curve of the turbidimetric or
chromogenic techniques. f = number of replicates.
The geometric mean end-point concentration is the measured
7. GEL-CLOT TECHNIQUE (METHODS A AND B) sensitivity of the lysate solution (IU/mL). If this is not less
The gel-clot technique allows detection or quantification than 0.5λ and not more than 2λ, the labelled sensitivity is
of endotoxins and is based on clotting of the lysate in the confirmed and is used in the tests performed with this lysate.
presence of endotoxins. The minimum concentration of (ii) Test for interfering factors
endotoxins required to cause the lysate to clot under standard
conditions is the labelled lysate sensitivity. To ensure both Prepare solutions A, B, C and D as shown in Table 2.6.14.-1,
the precision and validity of the test, confirm the labelled and use the test solutions at a dilution less than the MVD, not
lysate sensitivity and perform the test for interfering factors as containing any detectable endotoxins, operating as described
described under 1. Preparatory testing. under 1. Preparatory testing, (i) Confirmation of the labelled
lysate sensitivity.
1. PREPARATORY TESTING
The geometric mean end-point concentrations of solutions B
(i) Confirmation of the labelled lysate sensitivity and C are determined using the expression described in
Confirm in 4 replicates the labelled sensitivity λ, expressed 1. Preparatory testing, (i) Confirmation of the labelled lysate
in IU/mL, of the lysate solution prior to use in the test. sensitivity.
Confirmation of the lysate sensitivity is carried out when a
new lot of lysate is used or when there is any change in the test The test for interfering factors must be repeated when any
conditions which may affect the outcome of the test. changes are made to the experimental conditions that are
likely to influence the result of the test.
Prepare standard solutions of at least 4 concentrations
equivalent to 2λ, λ, 0.5λ and 0.25λ by diluting the standard The test is considered valid when all replicates of solutions A
endotoxin stock solution with water for BET. and D show no reaction and the result of solution C confirms
the labelled lysate sensitivity.
Mix a volume of the lysate solution with an equal volume
of 1 of the standard solutions (such as 0.1 mL aliquots) in If the sensitivity of the lysate determined with solution B is
each tube. When single test vials or ampoules containing not less than 0.5λ and not greater than 2λ, the test solution
lyophilised lysate are employed, add solutions of standards does not contain interfering factors under the experimental
directly to the vial or ampoule. Incubate the reaction mixture conditions used. Otherwise, the test solution interferes with
for a constant period according to the recommendations of the test.
the lysate manufacturer (usually at 37 ± 1 °C for 60 ± 2 min), If the preparation being examined interferes with the test at
avoiding vibration. Test the integrity of the gel : for tubes, take a dilution less than the MVD, repeat the test for interfering
each tube in turn directly from the incubator and invert it factors using a greater dilution, not exceeding the MVD. The
through approximately 180° in one smooth motion. If a firm use of a more sensitive lysate permits a greater dilution of the
gel has formed that remains in place upon inversion, record preparation being examined and this may contribute to the
the result as positive. A result is negative if an intact gel is not elimination of interference.
formed.
Interference may be overcome by suitable validated treatment,
The test is considered valid when the lowest concentration of such as filtration, neutralisation, dialysis or heat treatment.
the standard solutions shows a negative result in all replicate To establish that the treatment chosen effectively eliminates
tests. interference without loss of endotoxins, repeat the test for
The end-point is the lowest concentration in the series interfering factors using the preparation being examined to
of decreasing concentrations of standard endotoxin that which the standard endotoxin has been added and which has
clots the lysate. Determine the geometric mean end-point then been submitted to the chosen treatment.
Table 2.6.14.-1
Solution Endotoxin concentration/Solution to Diluent Dilution factor Endotoxin Number of replicates
which endotoxin is added concentration
A None/Test solution - - - 4
B 2λ/Test solution Test solution 1 2λ 4

2 1λ 4
4 0.5λ 4
8 0.25λ 4
C 2λ/Water for BET Water for BET 1 2λ 2
2 1λ 2
4 0.5λ 2
8 0.25λ 2
D None/Water for BET - - - 2
Solution A = solution of the preparation being examined that is free of detectable endotoxins.
Solution B = test for interference.
Solution C = control of the labelled lysate sensitivity.
Solution D = negative control (water for BET).

2. LIMIT TEST (METHOD A) with the test if a negative result is found for both replicates of
solution A. The preparation does not comply with the test if a
(i) Procedure
positive result is found for one or both replicates of solution A.
Prepare solutions A, B, C and D as shown in Table 2.6.14.-2, However, if the preparation does not comply with the test at a
and perform the test on these solutions following the procedure dilution less than the MVD, the test may be repeated using a
described under 1. Preparatory testing, (i) Confirmation of greater dilution, not exceeding the MVD.
the labelled lysate sensitivity.
3. QUANTITATIVE TEST (METHOD B)
(i) Procedure
The test quantifies bacterial endotoxins in the test solution by
Table 2.6.14.-2 titration to an end-point. Prepare solutions A, B, C and D as
shown in Table 2.6.14.-3, and test these solutions according
Solution Endotoxin concentration/Solution Number of replicates
to which endotoxin is added to the procedure described under 1. Preparatory testing, (i)
A None/Diluted test solution 2 Confirmation of the labelled lysate sensitivity.
B 2λ/Diluted test solution 2
(ii) Calculation and interpretation
C 2λ/Water for BET 2 The test is considered valid when the following 3 conditions
are met :
D None/Water for BET 2
(a) both replicates of solution D (negative control) are negative,
(b) both replicates of solution B (positive product control)
are positive,
Prepare solution A and solution B (positive product control)
using a dilution not greater than the MVD and treatments as (c) the geometric mean end-point concentration of solution C
described in 1. Preparatory testing, (ii) Test for interfering is in the range of 0.5λ to 2λ.
factors. Solutions B and C (positive controls) contain the
standard endotoxin at a concentration corresponding to twice To determine the endotoxin concentration of solution A,
calculate the end-point concentration for each replicate, by
the labelled lysate sensitivity. Solution D (negative control)
multiplying each end-point dilution factor by λ.
consists of water for BET.
(ii) Interpretation The endotoxin concentration in the test solution is the
end-point concentration of the replicates. If the test
The test is considered valid when both replicates of solution B is conducted with a diluted test solution, calculate the
and C are positive and those of solution D are negative. concentration of endotoxin in the original solution by
multiplying the result by the dilution factor.
When a negative result is found for both replicates of If none of the dilutions of the test solution is positive in a
solution A, the preparation being examined complies with valid test, report the endotoxin concentration as less than
the test. λ (or, if a diluted sample was tested, report as less than the
lowest dilution factor of the sample × λ). If all dilutions are
When a positive result is found for both replicates of positive, the endotoxin concentration is reported as equal to
solution A, the preparation being examined does not comply or greater than the largest dilution factor multiplied by λ (e.g.
with the test. in Table 2.6.14.-3, the initial dilution factor × 8 × λ).
When a positive result is found for one replicate of solution A The preparation being examined meets the requirements of
and a negative result is found for the other, repeat the test. the test if the endotoxin concentration in both replicates is
In the repeat test, the preparation being examined complies less than that specified in the monograph.
Table 2.6.14.-3
Solution Endotoxin concentration/Solution to which Diluent Dilution factor Endotoxin Number of replicates
endotoxin is added concentration
A None/Test solution Water for BET 1 - 2

2 - 2
4 - 2
8 - 2
B 2λ/Test solution 1 2λ 2
C 2λ/Water for BET Water for BET 1 2λ 2

2 1λ 2
4 0.5λ 2
8 0.25λ 2
D None/Water for BET - - - 2
Solution A = test solution at the dilution, not exceeding the MVD, with which the test for interfering factors was carried out. Subsequent dilution of
the test solution must not exceed the MVD. Use water for BET to make a dilution series of 4 tubes containing the test solution at concentrations of 1,
1/2, 1/4 and 1/8, relative to the dilution used in the test for interfering factors. Other dilutions up to the MVD may be used as appropriate.
Solution B = solution A containing standard endotoxin at a concentration of 2λ (positive product control).
Solution C = a dilution series of 4 tubes of water for BET containing the standard endotoxin at concentrations of 2λ, λ, 0.5λ and 0.25λ.
Solution D = water for BET (negative control).

2.6.14. Bacterial endotoxins EUROPEAN PHARMACOPOEIA
8. PHOTOMETRIC QUANTITATIVE TECHNIQUES Table 2.6.14.-4

(METHODS C, D, E AND F)
Solution Endotoxin Solution to which Number of
1. TURBIDIMETRIC TECHNIQUE (METHODS C AND F) concentration endotoxin is added replicates
This technique is a photometric test to measure the increase in A None Test solution Not less than 2
turbidity. Based on the test principle employed, this technique B Middle concentration Test solution Not less than 2
may be classified as being either the end-point-turbidimetric of the standard curve
test or the kinetic-turbidimetric test. C At least 3 concentra- Water for BET Each concentration
tions (lowest concen- not less than 2
The end-point-turbidimetric test (Method F) is based on the tration is designated λ)
quantitative relationship between the endotoxin concentration D None Water for BET Not less than 2
and the turbidity (absorbance or transmission) of the reaction
mixture at the end of an incubation period. Solution A = test solution, that may be diluted not to exceed the MVD.
Solution B = preparation to be examined at the same dilution as
The kinetic-turbidimetric test (Method C) is a method to solution A, containing added endotoxin at a concentration equal to or
measure either the time (onset time) needed for the reaction near the middle of the standard curve.
mixture to reach a predetermined absorbance or transmission, Solution C = standard endotoxin solution at the concentrations used in
or the rate of turbidity development. the validation of the method as described under 3. Preparatory testing,
(i) Assurance of criteria for the standard curve (positive controls).
The test is carried out at the incubation temperature Solution D = water for BET (negative control).
recommended by the lysate manufacturer (usually 37 ± 1 °C).
The test is considered valid when the following conditions
2. CHROMOGENIC TECHNIQUE (METHODS D AND E) are met :
This technique is used to measure the chromophore released – the absolute value of the correlation coefficient of the
from a suitable chromogenic peptide by the reaction of standard curve generated using solution C is greater than
endotoxins with the lysate. Depending on the test principle or equal to 0.980 ;
employed, this technique may be classified as being either the
end-point-chromogenic test or the kinetic-chromogenic test. – the result with solution D does not exceed the limit of the
blank value required in the description of the lysate reagent
The end-point-chromogenic test (Method E) is based on the employed, or it is less than the endotoxin detection limit of
quantitative relationship between the endotoxin concentration the lysate reagent employed.
and the quantity of chromophore released at the end of an
incubation period. Calculate the mean recovery of the added endotoxin by
subtracting the mean endotoxin concentration in the solution
The kinetic-chromogenic test (Method D) measures either the (if any) (solution A, Table 2.6.14.-4) from that in the solution
time (onset time) needed for the reaction mixture to reach a containing the added endotoxin (solution B, Table 2.6.14.-4).
predetermined absorbance, or the rate of colour development.
The test solution is considered free of interfering factors if
The test is carried out at the incubation temperature under the conditions of the test, the measured concentration
recommended by the lysate manufacturer (usually 37 ± 1 °C). of the endotoxin added to the test solution is within 50-200 per
cent of the known added endotoxin concentration, after
3. PREPARATORY TESTING subtraction of any endotoxin detected in the solution without
To assure the precision or validity of the turbidimetric and added endotoxin.
chromogenic techniques, preparatory tests are conducted to
show that the criteria for the standard curve are satisfied and When the endotoxin recovery is out of the specified range,
that the test solution does not interfere with the test. the test solution is considered to contain interfering factors.
Repeat the test using a greater dilution, not exceeding
Validation of the test method is required when any changes the MVD. Furthermore, interference of the test solution
are made to the experimental conditions that are likely to or diluted test solution not to exceed the MVD may be
influence the result of the test. eliminated by suitable validated treatment, such as filtration,
(i) Assurance of criteria for the standard curve neutralisation, dialysis or heat treatment. To establish that the
treatment chosen effectively eliminates interference without
The test must be carried out for each lot of lysate reagent. loss of endotoxins, repeat the test for interfering factors
using the preparation being examined to which the standard
Using the standard endotoxin solution, prepare at least endotoxin has been added and which has then been submitted
3 endotoxin concentrations within the range indicated by the to the chosen treatment.
lysate manufacturer to generate the standard curve. Perform
the test using at least 3 replicates of each standard endotoxin 4. TEST
solution as recommended by the lysate manufacturer (volume (i) Procedure
ratios, incubation time, temperature, pH, etc.).
Follow the procedure described in 3. Preparatory testing,
If the desired range is greater than 2 log10 in the kinetic (ii) Test for interfering factors.
methods, additional standards must be included to bracket
(ii) Calculation
each log10 increase in the range of the standard curve.
Calculate the endotoxin concentration of each replicate of
The absolute value of the correlation coefficient, | r |, must solution A using the standard curve generated by the positive
be greater than or equal to 0.980, for the range of endotoxin control solution C.
concentrations set up.
The test is considered valid when the following 3 requirements
(ii) Test for interfering factors are met :
Select an endotoxin concentration at or near the middle of (1) the results obtained with solution C comply with the
the endotoxin standard curve. requirements for validation defined under 3. Preparatory
testing, (i) Assurance of criteria for the standard curve,
Prepare solutions A, B, C and D as shown in Table 2.6.14.-4.
Perform the test on at least 2 replicates of these solutions as (2) the endotoxin recovery, calculated from the endotoxin
recommended by the lysate manufacturer (volume of test concentration found in solution B after subtracting the
solution and lysate solution, volume ratio of test solution to endotoxin concentration found in solution A, is within the
lysate solution, incubation time, etc.). range of 50-200 per cent,

(3) the result obtained with solution D (negative control) after correction for dilution and concentration, is less than the
does not exceed the limit of the blank value required in endotoxin limit for the product.
the description of the lysate employed, or it is less than the
endotoxin detection limit of the lysate reagent employed. Guidelines on the test for bacterial endotoxins are given in
general chapter 5.1.10.
(iii) Interpretation
The preparation being examined complies with the test if the
mean endotoxin concentration of the replicates of solution A,

EUROPEAN PHARMACOPOEIA 2.9.1. Disintegration of tablets and capsules
01/2020:20901 and to each other, as defined in Figure 2.9.1.-1. 4 identical

trapezoidal-shaped planes are cut into the wall of the cylinder,
nearly perpendicular to the ends of the cylinder. The
trapezoidal shape is symmetrical ; its parallel sides coincide
with the ends of the cylinder and are parallel to an imaginary
2.9.1. DISINTEGRATION OF TABLETS the line connecting the centres of 2 adjacent holes 6 mm from
cylindrical axis. The parallel side of the trapezoid on the
AND CAPSULES(1) bottom of the cylinder has a length of 1.6 ± 0.1 mm and its
bottom edges lie at a depth of 1.5 mm to 1.8 mm from the
This test is provided to determine whether tablets or capsules cylinder’s circumference. The parallel side of the trapezoid
disintegrate within the prescribed time when placed in a liquid on the top of the cylinder has a length of 9.4 ± 0.2 mm and
medium under the experimental conditions presented below. its centre lies at a depth of 2.6 ± 0.1 mm from the cylinder’s
For the purposes of this test, disintegration does not circumference. All surfaces of the disc are smooth.
imply complete dissolution of the unit or even of its active If the use of discs is specified, add a disc to each tube and
constituent. Complete disintegration is defined as that state in operate the apparatus as directed under Procedure. The discs
which any residue of the unit, except fragments of insoluble conform to the dimensions shown in Figure 2.9.1.-1.
coating or capsule shell, remaining on the screen of the test
apparatus or adhering to the lower surface of the discs, if used, The use of automatic detection employing modified discs
is a soft mass having no palpably firm core. is permitted where the use of discs is specified or allowed.
Such discs must comply with the requirements of density and
♦Use apparatus A for tablets and capsules that are not dimension given in this chapter.
greater than 18 mm long. For larger tablets or capsules use
apparatus B.♦ Procedure. Place 1 dosage unit in each of the 6 tubes of the
basket and, if prescribed, add a disc. Operate the apparatus
TEST A - TABLETS AND CAPSULES OF NORMAL SIZE using the specified medium, maintained at 37 ± 2 °C, as
Apparatus. The apparatus consists of a basket-rack assembly, the immersion fluid. At the end of the specified time, lift
a 1 L, low-form beaker, 149 ± 11 mm in height and having the basket from the fluid and observe the dosage units :
an inside diameter of 106 ± 9 mm for the immersion fluid, a all of the dosage units have disintegrated completely. If
thermostatic arrangement for heating the fluid between 35 °C 1 or 2 dosage units fail to disintegrate, repeat the test on
and 39 °C, and a device for raising and lowering the basket in 12 additional dosage units. The requirements of the test are
the immersion fluid at a constant frequency rate between 29 met if not less than 16 of the 18 dosage units tested have
and 32 cycles per minute, through a distance of 55 ± 2 mm. disintegrated.
The volume of the fluid in the vessel is such that at the highest ♦TEST B – LARGE TABLETS AND LARGE CAPSULES
point of the upward stroke the wire mesh remains at least
15 mm below the surface of the fluid, and descends to not less Apparatus. The main part of the apparatus (Figure 2.9.1.-2.)
than 25 mm from the bottom of the vessel on the downward is a rigid basket-rack assembly supporting 3 cylindrical
stroke. At no time should the top of the basket-rack assembly transparent tubes 77.5 ± 2.5 mm long, 33.0 mm ± 0.5 mm in
become submerged. The time required for the upward stroke internal diameter, and with a wall thickness of 2.5 ± 0.5 mm.
is equal to the time required for the downward stroke, and the Each tube is provided with a cylindrical disc 31.4 ± 0.13 mm
change in stroke direction is a smooth transition, rather than in diameter and 15.3 ± 0.15 mm thick, made of transparent
an abrupt reversal of motion. The basket-rack assembly moves plastic with a relative density of 1.18-1.20. Each disc is pierced
vertically along its axis. There is no appreciable horizontal by 7 holes, each 3.15 ± 0.1 mm in diameter, 1 in the centre
motion or movement of the axis from the vertical. and the other 6 spaced equally on a circle of radius 4.2 mm
Basket-rack assembly. The basket-rack assembly consists of from the centre of the disc. The tubes are held vertically
6 open-ended transparent tubes, each 77.5 ± 2.5 mm long by 2 separate and superimposed rigid plastic plates 97 mm
and having an inside diameter of 21.85 ± 1.15 mm and a wall in diameter and 9 mm thick, with 3 holes. The holes are
1.9 ± 0.9 mm thick ; the tubes are held in a vertical position equidistant from the centre of the plate and equally spaced.
by 2 plates, each 90 ± 2 mm in diameter and 6.75 ± 1.75 mm Attached to the under side of the lower plate is a piece of
in thickness, with 6 holes, each 24 ± 2 mm in diameter, woven gauze made from stainless steel wire 0.63 ± 0.03 mm
equidistant from the centre of the plate and equally spaced in diameter and having mesh apertures of 2.0 ± 0.2 mm.
from one another. Attached to the under surface of the lower The plates are held rigidly in position and 77.5 mm apart
plate is a woven stainless steel wire cloth, which has a plain by vertical metal rods at the periphery. A metal rod is also
square weave with 2.0 ± 0.2 mm mesh apertures and with a fixed to the centre of the upper plate to enable the assembly
wire diameter of 0.615 ± 0.045 mm. The parts of the apparatus to be attached to a mechanical device capable of raising and
are assembled and rigidly held by means of 3 bolts passing lowering it smoothly at a constant frequency of between
through the 2 plates. A suitable means is provided to suspend 29 and 32 cycles per minute, through a distance of 55 ± 2 mm.
the basket-rack assembly from the raising and lowering device The assembly is suspended in the specified liquid medium
using a point on its axis. in a suitable vessel, preferably a 1 L beaker. The volume of
The design of the basket-rack assembly may be varied the liquid is such that when the assembly is in the highest
somewhat provided the specifications for the glass tubes and position the wire mesh is at least 15 mm below the surface of
the screen mesh size are maintained. The basket-rack assembly the liquid, and when the assembly is in the lowest position the
conforms to the dimensions shown in Figure 2.9.1.-1. wire mesh is at least 25 mm above the bottom of the beaker
and the upper open ends of the tubes remain above the surface
Discs. The use of discs is permitted only where specified of the liquid. A suitable device maintains the temperature of
or allowed. Each tube is provided with a cylindrical disc the liquid at 35-39 °C.
9.5 ± 0.15 mm thick and 20.7 ± 0.15 mm in diameter. The
disc is made of a suitable, transparent plastic material having The design of the basket-rack assembly may be varied provided
a specific gravity of 1.18-1.20. 5 parallel 2 ± 0.1 mm holes the specifications for the tubes and wire mesh are maintained.
extend between the ends of the cylinder. One of the holes Method. Test 6 tablets or capsules either by using
is centered on the cylindrical axis. The other holes are 2 basket-rack assemblies in parallel or by repeating the
parallel to the cylindrical axis and centered 6 ± 0.2 mm procedure. In each of the 3 tubes, place 1 tablet or capsule
from the axis on imaginary lines perpendicular to the axis and, if prescribed, add a disc; suspend the assembly in the

2.9.1. Disintegration of tablets and capsules EUROPEAN PHARMACOPOEIA
beaker containing the specified liquid. Operate the apparatus for the prescribed period, withdraw the assembly and examine
the state of the tablets or capsules. To pass the test, all 6 of the
tablets or capsules must have disintegrated.♦
Figure 2.9.1.-1. – Disintegration apparatus A


EUROPEAN PHARMACOPOEIA 2.9.1. Disintegration of tablets and capsules
Figure 2.9.1.-2. – Disintegration apparatus B


EUROPEAN PHARMACOPOEIA 2.9.2. Disintegration of suppositories and pessaries
01/2008:20902
2.9.2. DISINTEGRATION OF
SUPPOSITORIES AND PESSARIES
The disintegration test determines whether the suppositories

or pessaries soften or disintegrate within the prescribed
time when placed in a liquid medium in the experimental
conditions described below.
Disintegration is considered to be achieved when :
a) dissolution is complete,
b) the components of the suppository or pessary have

separated : melted fatty substances collect on the surface of
the liquid, insoluble powders fall to the bottom and soluble
components dissolve, depending on the type of preparation,
the components may be distributed in one or more of these
ways,
c) there is softening of the sample that may be accompanied

by appreciable change of shape without complete separation
of the components, the softening is such that the suppository
or pessary no longer has a solid core offering resistance to
pressure of a glass rod,
d) rupture of the gelatin shell of rectal or vaginal capsules

occurs allowing release of the contents,
e) no residue remains on the perforated disc or if a residue

remains, it consists only of a soft or frothy mass having no
solid core offering resistance to pressure of a glass rod (vaginal
tablets).
Apparatus. The apparatus (Figure 2.9.2.-1) consists of a

sleeve of glass or suitable transparent plastic, of appropriate
thickness, to the interior of which is attached by means of three
hooks a metal device consisting of two perforated stainless
metal discs each containing 39 holes 4 mm in diameter ; the
diameter of the discs is similar to that of the interior of the Figure 2.9.2.-1. – Apparatus for disintegration of suppositories
sleeve ; the discs are about 30 mm apart. The test is carried out and pessaries
using three such apparatuses each containing a single sample. Dimensions in millimetres
Each apparatus is placed in a beaker with a capacity of at least
4 L filled with water maintained at 36-37 °C, unless otherwise METHOD OF OPERATION FOR VAGINAL TABLETS
prescribed. The apparatuses may also be placed together in a Use the apparatus described above, arranged so as to rest on
vessel with a capacity of at least 12 L. The beaker is fitted with a the hooks (see Figure 2.9.2.-2). Place it in a beaker of suitable
slow stirrer and a device that will hold the cylinders vertically diameter containing water maintained at 36-37 °C with the
not less than 90 mm below the surface of the water and allow level just below the upper perforated disc. Using a pipette,
them to be inverted without emerging from the water. adjust the level with water at 36-37 °C until a uniform film
covers the perforations of the disc. Use three vaginal tablets.
Method. Use three suppositories or pessaries. Place each one Place each one on the upper plate of an apparatus and cover
on the lower disc of a device, place the latter in the sleeve the latter with a glass plate to maintain appropriate conditions
and secure. Invert the apparatuses every 10 min. Examine of humidity. Examine the state of the samples after the period
the samples after the period prescribed in the monograph. To prescribed in the monograph. To pass the test all the samples
pass the test all the samples must have disintegrated. must have disintegrated.

2.9.2. Disintegration of suppositories and pessaries EUROPEAN PHARMACOPOEIA
A. glass plate D. water
B. vaginal tablet E. dish, beaker

C. water surface
Figure 2.9.2.-2.

EUROPEAN PHARMACOPOEIA 2.9.5. Uniformity of mass of single-dose preparations
01/2008:20905 Table 2.9.5.-1
Pharmaceutical Form Average Mass Percentage
deviation
Tablets (uncoated and 80 mg or less 10
film-coated)
More than 80 mg and 7.5
less than 250 mg
2.9.5. UNIFORMITY OF MASS OF 250 mg or more 5
SINGLE-DOSE PREPARATIONS Capsules, granules Less than 300 mg 10

(uncoated, single-dose)
and powders (single-dose) 300 mg or more 7.5
Powders for parenteral More than 40 mg 10

Weigh individually 20 units taken at random or, for single-dose administration*
preparations presented in individual containers, the contents (single-dose)
of 20 units, and determine the average mass. Not more than
Suppositories and pessaries All masses 5
2 of the individual masses deviate from the average mass by
more than the percentage deviation shown in Table 2.9.5.-1 Powders for eye-drops and Less than 300 mg 10
and none deviates by more than twice that percentage. powders for eye lotions 300 mg or more 7.5
(single-dose)
For capsules and powders for parenteral administration, * When the average mass is equal to or below 40 mg, the preparation
is not submitted to the test for uniformity of mass but to the test for
proceed as described below. uniformity of content of single-dose preparations (2.9.6).
POWDERS FOR PARENTERAL ADMINISTRATION

Remove any paper labels from a container and wash and dry
CAPSULES the outside. Open the container and without delay weigh the
container and its contents. Empty the container as completely
Weigh an intact capsule. Open the capsule without losing any as possible by gentle tapping, rinse it if necessary with water R
part of the shell and remove the contents as completely as and then with alcohol R and dry at 100-105 °C for 1 h, or, if the
possible. For soft shell capsules, wash the shell with a suitable nature of the container precludes heating at this temperature,
solvent and allow to stand until the odour of the solvent is no dry at a lower temperature to constant mass. Allow to cool
longer perceptible. Weigh the shell. The mass of the contents in a desiccator and weigh. The mass of the contents is the
is the difference between the weighings. Repeat the procedure difference between the weighings. Repeat the procedure with
with another 19 capsules. another 19 containers.

EUROPEAN PHARMACOPOEIA 2.9.6. Uniformity of content of single-dose preparations
01/2017:20906 dosage units taken at random. The preparation complies with

the test if not more than one of the individual contents of the
30 units is outside 85 per cent to 115 per cent of the average
content and none is outside the limits of 75 per cent to 125 per
cent of the average content.
2.9.6. UNIFORMITY OF CONTENT OF
SINGLE-DOSE PREPARATIONS TEST B
The preparation complies with the test if not more than one
The test for uniformity of content of single-dose preparations
individual content is outside the limits of 85 per cent to
is based on the assay of the individual contents of active
115 per cent of the average content and none is outside the
substance(s) of a number of single-dose units to determine
limits of 75 per cent to 125 per cent of the average content.
whether the individual contents are within limits set with
The preparation fails to comply with the test if more than
reference to the average content of the sample.
3 individual contents are outside the limits of 85 per cent
The test is not required for multivitamin and trace-element to 115 per cent of the average content or if one or more
preparations and in other justified and authorised individual contents are outside the limits of 75 per cent to
circumstances. 125 per cent of the average content.
Method. Using a suitable analytical method, determine the
individual contents of active substance(s) of 10 dosage units If 2 or 3 individual contents are outside the limits of 85 per
taken at random. cent to 115 per cent but within the limits of 75 per cent to
125 per cent, determine the individual contents of another
Apply the criteria of test A, test B or test C as specified in the 20 dosage units taken at random. The preparation complies
monograph for the dosage form in question. with the test if not more than 3 individual contents of the
TEST A 30 units are outside the limits of 85 per cent to 115 per cent
of the average content and none is outside the limits of 75 per
The preparation complies with the test if each individual
cent to 125 per cent of the average content.
content is between 85 per cent and 115 per cent of the average
content. The preparation fails to comply with the test if
more than one individual content is outside these limits or if TEST C
one individual content is outside the limits of 75 per cent to The preparation complies with the test if the average content
125 per cent of the average content. of the 10 dosage units is between 90 per cent and 110 per cent
If one individual content is outside the limits of 85 per of the content stated on the label and if the individual content
cent to 115 per cent but within the limits of 75 per cent to of each dosage unit is between 75 per cent and 125 per cent
125 per cent, determine the individual contents of another 20 of the average content.

EUROPEAN PHARMACOPOEIA 2.9.12. Sieve test
01/2008:20912 Coarse powder. Not less than 95 per cent by mass passes
through a number 1400 sieve and not more than 40 per cent
by mass passes through a number 355 sieve.
Moderately fine powder. Not less than 95 per cent by mass
passes through a number 355 sieve and not more than 40 per
cent by mass passes through a number 180 sieve.
2.9.12. SIEVE TEST Fine powder. Not less than 95 per cent by mass passes
through a number 180 sieve and not more than 40 per cent by
The degree of fineness of a powder may be expressed by mass passes through a number 125 sieve.
reference to sieves that comply with the specifications for Very fine powder. Not less than 95 per cent by mass passes
non-analytical sieves (2.1.4). through a number 125 sieve and not more than 40 per cent by
mass passes through a number 90 sieve.
Where the degree of fineness of powders is determined by
sieving, it is defined in relation to the sieve number(s) used If a single sieve number is given, not less than 97 per cent of
either by means of the following terms or, where such terms the powder passes through the sieve of that number, unless
cannot be used, by expressing the fineness of the powder as a otherwise prescribed.
percentage m/m passing the sieve(s) used. Assemble the sieves and operate in a suitable manner until
sifting is practically complete. Weigh the separated fractions
The following terms are used in the description of powders : of the powder.

EUROPEAN PHARMACOPOEIA 2.9.17. Test for extractable volume of parenteral preparations
04/2010:20917 The contents of containers holding 10 mL or more may be

determined by opening them and emptying the contents
directly into the graduated cylinder or tared beaker.
The volume is not less than the nominal volume in case of
containers examined individually, or, in case of containers
2.9.17. TEST FOR EXTRACTABLE with a nominal volume of 2 mL or less, is not less than the sum
of the nominal volumes of the containers taken collectively.
VOLUME OF PARENTERAL
MULTIDOSE CONTAINERS
PREPARATIONS(1)
For injections in multidose containers labelled to yield
Suspensions and emulsions are shaken before withdrawal of a specific number of doses of a stated volume, select one
the contents and before the determination of the density. Oily container and proceed as directed for single-dose containers
and viscous preparations may be warmed according to the using the same number of separate syringe assemblies as the
instructions on the label, if necessary, and thoroughly shaken number of doses specified.
immediately before removing the contents. The contents are The volume is such that each syringe delivers not less than
then cooled to 20-25 °C before measuring the volume. the stated dose.
SINGLE-DOSE CONTAINERS CARTRIDGES AND PREFILLED SYRINGES
Select 1 container if the nominal volume is 10 mL or more, Select 1 container if the nominal volume is 10 mL or more,
3 containers if the nominal volume is more than 3 mL and less 3 containers if the nominal volume is more than 3 mL and less
than 10 mL, or 5 containers if the nominal volume is 3 mL or than 10 mL, or 5 containers if the nominal volume is 3 mL
less. Take up individually the total contents of each container or less. If necessary, fit the containers with the accessories
selected into a dry syringe of a capacity not exceeding 3 times required for their use (needle, piston, syringe) and transfer the
the volume to be measured, and fitted with a 21-gauge needle entire contents of each container without emptying the needle
not less than 2.5 cm in length. Expel any air bubbles from the into a dry tared beaker by slowly and constantly depressing
syringe and needle, then discharge the contents of the syringe the piston. Determine the volume in millilitres calculated as
without emptying the needle into a standardised dry cylinder the mass in grams divided by the density.
(graduated to contain rather than to deliver the designated The volume measured for each of the containers is not less
volumes) of such size that the volume to be measured occupies than the nominal volume.
at least 40 per cent of its graduated volume. Alternatively, the
volume of the contents in millilitres may be calculated as the PARENTERAL INFUSIONS
mass in grams divided by the density. Select one container. Transfer the contents into a dry
For containers with a nominal volume of 2 mL or less, the measuring cylinder of such a capacity that the volume to
contents of a sufficient number of containers may be pooled to be determined occupies at least 40 per cent of the nominal
obtain the volume required for the measurement provided that volume of the cylinder. Measure the volume transferred.
a separate, dry syringe assembly is used for each container. The volume is not less than the nominal volume.

EUROPEAN PHARMACOPOEIA 2.9.18. Preparations for inhalation
01/2008:20918 depth of about 10 mm, lines up along the horizontal axis of

the throat and the open end of the actuator, which accepts the
pressurised container, is uppermost and in the same vertical
plane as the rest of the apparatus.
Introduce 7 mL and 30 mL of a suitable solvent into the upper
and lower impingement chambers, respectively.
2.9.18. PREPARATIONS FOR
INHALATION : AERODYNAMIC
ASSESSMENT OF FINE PARTICLES Connect all the component parts. Ensure that the assembly
is vertical and adequately supported and that the lower
This test is used to determine the fine particle characteristics jet-spacer peg of the lower jet assembly just touches the
of the aerosol clouds generated by preparations for inhalation. bottom of the lower impingement chamber. Connect a
suitable pump to the outlet of the apparatus. Adjust the air
Unless otherwise justified and authorised, one of the following flow through the apparatus, as measured at the inlet to the
apparatus and test procedures is used. throat, to 60 ± 5 L/min.
Stage mensuration is performed periodically together with Prime the metering valve by shaking for 5 s and discharging
confirmation of other dimensions critical to the effective once to waste ; after not less than 5 s, shake and discharge
operation of the impactor. again to waste. Repeat a further 3 times.
Re-entrainment (for apparatus D and E). To ensure efficient Shake for about 5 s, switch on the pump to the apparatus
particle capture, coat each plate with glycerol, silicone oil or and locate the mouthpiece end of the actuator in the adapter,
similar high viscosity liquid, typically deposited from a volatile discharge once immediately. Remove the assembled inhaler
solvent. Plate coating must be part of method validation and from the adapter, shake for not less than 5 s, relocate the
may be omitted where justified and authorised. mouthpiece end of the actuator in the adapter and discharge
again. Repeat the discharge sequence. The number of
Mass balance. The total mass of the active substance is not less
discharges should be minimised and typically would not be
than 75 per cent and not more than 125 per cent of the average
greater than 10. After the final discharge wait for not less than
delivered dose determined during testing for uniformity of
5 s and then switch off the pump. Dismantle the apparatus.
delivered dose. This is not a test of the inhaler but it serves to
ensure that the results are valid.
Wash the inner surface of the inlet tube to the lower

APPARATUS A - GLASS IMPINGER impingement chamber and its outer surface that projects into
the chamber with a suitable solvent, collecting the washings
The apparatus is shown in Figure 2.9.18.-1 (see also in the lower impingement chamber. Determine the content
Table 2.9.18.-1). of active substance in this solution. Calculate the amount of
Procedure for nebulisers active substance collected in the lower impingement chamber
per discharge and express the results as a percentage of the
Introduce 7 mL and 30 mL of a suitable solvent into the upper dose stated on the label.
Connect all the component parts. Ensure that the assembly is
vertical and adequately supported and that the jet spacer peg Procedure for powder inhalers
of the lower jet assembly just touches the bottom of the lower
Introduce 7 mL and 30 mL of a suitable solvent into the upper
impingement chamber. Connect a suitable pump fitted with
a filter (of suitable pore size) to the outlet of the apparatus.
Adjust the air flow through the apparatus, as measured at the
inlet to the throat, to 60 ± 5 L/min.
Connect all the component parts. Ensure that the assembly is
Introduce the liquid preparation for inhalation into the vertical and adequately supported and that the jet-spacer peg
reservoir of the nebuliser. Fit the mouthpiece and connect it of the lower jet assembly just touches the bottom of the lower
by means of an adapter to the device. impingement chamber. Without the inhaler in place, connect
Switch on the pump of the apparatus and after 10 s switch on a suitable pump to the outlet of the apparatus. Adjust the air
the nebuliser. flow through the apparatus, as measured at the inlet to the
throat, to 60 ± 5 L/min.
After 60 s, unless otherwise justified, switch off the nebuliser,
wait for about 5 s and then switch off the pump of the Prepare the inhaler for use and locate the mouthpiece in the
apparatus. Dismantle the apparatus and wash the inner apparatus by means of a suitable adapter. Switch on the pump
surface of the upper impingement chamber collecting the for 5 s. Switch off the pump and remove the inhaler. Repeat
washings in a volumetric flask. Wash the inner surface of the the discharge sequence. The number of discharges should
lower impingement chamber collecting the washings in a be minimised and typically would not be greater than 10.
second volumetric flask. Finally, wash the filter preceding the Dismantle the apparatus.
pump and its connections to the lower impingement chamber
and combine the washings with those obtained from the
lower impingement chamber. Determine the amount of active Wash the inner surface of the inlet tube to the lower
substance collected in each of the 2 flasks. Express the results impingement chamber and its outer surface that projects into
for each of the 2 parts of the apparatus as a percentage of the the chamber with a suitable solvent, collecting the washings
total amount of active substance. in the lower impingement chamber. Determine the content
Procedure for pressurised inhalers of active substance in this solution. Calculate the amount of
active substance collected in the lower impingement chamber
Place the actuator adapter in position at the end of the throat per discharge and express the results as a percentage of the
so that the mouthpiece end of the actuator, when inserted to a dose stated on the label.

2.9.18. Preparations for inhalation EUROPEAN PHARMACOPOEIA
Code Item Description Dimen-

sions*
E Coupling tube Medium-wall glass tubing :
– ground-glass cone 14/23
Bent section and upper vertical

section:
– external diameter 13
Lower vertical section:

– external diameter 8
F Screwthread, Plastic screw cap 28/13
side-arm Silicone rubber ring 28/11
adaptor PTFE washer 28/11
Glass screwthread :
– thread size 28
Side-arm outlet to vacuum pump:

– minimum bore diameter 5
G Lower jet Modified polypropylene filter see

assembly holder connected to lower vertical Figure
section of coupling tube by PTFE 2.9.18.-1
tubing.
Acetal circular disc with the centres
of four jets arranged on a projected
circle of diameter 5.3 mm with an
integral jet spacer peg: 10
– peg diameter 2
– peg protrusion 2
H Lower Conical flask 250 mL
Figure 2.9.18.-1. – Apparatus A : glass impinger
impingement – ground-glass inlet socket 24/29
Dimensions in millimetres (tolerances ± 1 mm unless otherwise chamber
prescribed) * Dimensions in millimetres, unless otherwise stated.
Fine particle dose and particle size

distribution
Table 2.9.18.-1. – Component specification for apparatus A APPARATUS C - MULTI-STAGE LIQUID IMPINGER
in Figure 2.9.18.-1 The multi-stage liquid impinger consists of impaction stages 1
(pre-separator), 2, 3 and 4 and an integral filter stage (stage 5),
Code Item Description Dimen- see Figures 2.9.18.-4/6. An impaction stage comprises an
sions* upper horizontal metal partition wall (B) through which
A Mouthpiece Moulded rubber adapter for
a metal inlet jet tube (A) with its impaction plate (D) is
adaptor actuator mouthpiece. protruding. A glass cylinder (E) with sampling port (F) forms
the vertical wall of the stage, and a lower horizontal metal
B Throat Modified round-bottomed flask : 50 mL partition wall (G) through which the tube (H) connects to the
– ground-glass inlet socket 29/32 next lower stage. The tube into stage 4 (U) ends in a multi-jet
arrangement. The impaction plate (D) is secured in a metal
– ground-glass outlet cone 24/29 frame (J) which is fastened by 2 wires (K) to a sleeve (L)
C Neck Modified glass adapter : secured on the jet tube. The horizontal face of the collection
plate is perpendicular to the axis of the jet tube and centrally
– ground-glass inlet socket 24/29 aligned. The upper surface of the impaction plate is slightly
– ground-glass outlet cone 24/29 raised above the edge of the metal frame. A recess around the
perimeter of the horizontal partition wall guides the position
Lower outlet section of of the glass cylinder. The glass cylinders are sealed against
precision-bore glass tubing: the horizontal partition walls with gaskets (M) and clamped
– bore diameter 14 together by 6 bolts (N). The sampling ports are sealed by
stoppers. The bottom-side of the lower partition wall of
Selected bore light-wall glass stage 4 has a concentrical protrusion fitted with a rubber
tubing :
O-ring (P) which seals against the edge of a filter placed in
– external diameter 17 the filter holder. The filter holder (R) is constructed as a
basin with a concentrical recess in which a perforated filter
D Upper Modified round-bottomed flask 100 mL
support (S) is flush-fitted. The filter holder is dimensioned for
impingement – ground-glass inlet socket 24/29 76 mm diameter filters. The assembly of impaction stages is
clamped onto the filter holder by 2 snap-locks (T). Connect
chamber – ground-glass outlet cone 24/29
an induction port (see Figure 2.9.18.-7) onto the stage 1 inlet

Figure 2.9.18.-4. – Apparatus C : multi-stage liquid impinger

jet tube of the impinger. A rubber O-ring on the jet tube stoppers. Tilt the apparatus to wet the stoppers, thereby
provides an airtight connection to the induction port. A neutralising electrostatic charge. Place a suitable filter capable
suitable mouthpiece adapter is used to provide an airtight seal of quantitatively collecting the active substance in stage 5
between the inhaler and the induction port. The front face and assemble the apparatus. Place a suitable mouthpiece
of the inhaler mouthpiece must be flush with the front face adapter in position at the end of the induction port so that
of the induction port. the mouthpiece end of the actuator, when inserted, lines up
Procedure for pressurised inhalers along the horizontal axis of the induction port and the inhaler
is positioned in the same orientation as intended for use.
Dispense 20 mL of a solvent, capable of dissolving the
Connect a suitable vacuum pump to the outlet of the apparatus
active substance into each of stages 1 to 4 and replace the

Figure 2.9.18.-5. – Apparatus C : details of jet tube and impaction plate. Inserts show end of multi-jet tube U leading to stage 4.
(Numbers and lowercase letters refer to Table 2.9.18.-3 and uppercase letters refer to Figure 2.9.18.-4).
and adjust the air flow through the apparatus, as measured Dismantle the filter stage of the apparatus. Carefully remove
at the inlet to the induction port, to 30 L/min (± 5 per cent). the filter and extract the active substance into an aliquot of the
Switch off the pump. solvent. Remove the induction port and mouthpiece adapter
from the apparatus and extract the active substance into an
aliquot of the solvent. If necessary, rinse the inside of the inlet
Unless otherwise prescribed in the patient instructions, shake jet tube to stage 1 with solvent, allowing the solvent to flow
the inhaler for 5 s and discharge 1 delivery to waste. Switch into the stage. Extract the active substance from the inner
on the pump to the apparatus, locate the mouthpiece end of walls and the collection plate of each of the 4 upper stages
the actuator in the adapter and discharge the inhaler into of the apparatus into the solution in the respective stage by
the apparatus, depressing the valve for a sufficient time to carefully tilting and rotating the apparatus, observing that no
ensure complete discharge. Wait for 5 s before removing the liquid transfer occurs between the stages.
assembled inhaler from the adapter. Repeat the procedure.
The number of discharges should be minimised and typically
would not be greater than 10. The number of discharges is Using a suitable method of analysis, determine the quantity of
sufficient to ensure an accurate and precise determination of active substance contained in each of the aliquots of solvent.
the fine particle dose. After the final discharge, wait for 5 s
and then switch off the pump. Calculate the fine particle dose (see Calculations).

Table 2.9.18.-2. – Component specification for apparatus C in Code* Item Description Dimen-
sions**
Figures 2.9.18.-4/6
T Snap-locks
Code* Item Description Dimen-
sions** U Multi-jet Jet tube (H) ending in multi-jet see inserts
A,H Jet tube Metal tube screwed onto partition see Figure tube arrangement. Figure
wall sealed by gasket (C), polished 2.9.18.-5 2.9.18.-5
inner surface * Refers to Figure 2.9.18.-4.
B,G Partition Circular metal plate ** Measures in millimetres with tolerances according to iso 2768-m
wall unless otherwise stated.
– diameter 120
– thickness see Figure

2.9.18.-5
C Gasket e.g. PTFE to fit jet
tube
D Impaction Porosity 0 sintered-glass disk
plate – diameter see Figure
2.9.18.-5
E Glass Plane polished cut glass tube
cylinder
– height, including gaskets 46
– outer diameter 100
– wall thickness 3.5
– sampling port (F) diameter 18
– stopper in sampling port ISO 24/25 Figure 2.9.18.-6. – Apparatus C : details of the filter stage
(stage 5). Numbers refer to dimensions (Ø = diameter).
J Metal frame L-profiled circular frame with slit
Uppercase letters refer to Table 2.9.18.-2.
– inner diameter to fit Dimensions in millimetres unless otherwise stated
impaction
plate Table 2.9.18.-3. – Dimensions(1) of jet tube with impaction plate
– height 4 of apparatus C
– thickness of horizontal section 0.5 Type Code(2) Stage 1 Stage 2 Stage 3 Stage 4 Filter
(stage 5)
– thickness of vertical section 2
Distance 1 9.5 5.5 4.0 6.0 n.a.
K Wire Steel wire interconnecting metal (-.0+.5) (-.0+.5) (-.0+.5) (-.0+.5)
frame and sleeve (2 for each Distance 2 26 31 33 30.5 0
frame)
– diameter 1 Distance 3 8 5 5 5 5
L Sleeve Metal sleeve secured on jet tube Distance 4 3 3 3 3 n.a.
by screw
Distance 5 0 3 3 3 3
– inner diameter to fit jet
tube Distance 6 (3)
20 25 25 25 25
– height 6
Distance 7 n.a. n.a. n.a. 8.5 n.a.
– thickness 5
Diameter c 25 14 8.0 21 14
M Gasket e.g. silicone to fit glass (± .1)
cylinder Diameter 50 30 20 30 n.a.
d
N Bolt Metal bolt with nut (6 pairs)
Diameter e 27.9 16.5 10.5 23.9 n.a.
– length 205
Diameter f 31.75 22 14 31 22
– diameter 4 (-.0+.5)
Diameter g 25.4 21 13 30 21
P O-ring Rubber O-ring
Diameter h n.a. n.a. n.a. 2.70 n.a.
– diameter × thickness 66.34 × 2.62
(± .5)
Q O-ring Rubber O-ring Diameter j n.a. n.a. n.a. 6.3 n.a.
– diameter × thickness 29.1 × 1.6 Diameter k n.a. n.a. n.a. 12.6 n.a.
R Filter holder Metal housing with stand and see Figure Radius (4) r 16 22 27 28.5 0
outlet 2.9.18.-6
Radius s 46 46 46 46 n.a.
S Filter Perforated sheet metal
support t n.a. 50 50 50 50
Radius
– diameter 65
Angle w 10° 53° 53° 53° 53°
– hole diameter 3
Angle u n.a. n.a. n.a. 45° n.a.
– distance between holes 4
(centre-points) Angle v n.a. n.a. n.a. 60° n.a.
(1) Measures in millimetres with tolerances according to ISO 2768-m

unless otherwise stated
(2) Refer to Figure 2.9.18.-5
(3) Including gasket
(4) Relative centreline of stage compartment
n.a. = not applicable

1. Material may be aluminium, stainless steel or other suitable material.

2. Machine from 38 mm bar stock.
3. Bore 19 mm hole through bar.
4. Cut tube to exact 45° as shown.
5. The inner bores and tapers should be smooth – surface roughness Ra approx. 0.4 µm.
6. Mill joining cads of stock to provide a liquid tight leak-free seal.
7. Set up a holding fixture for aligning the inner 19 mm bore and for drilling and tapping M4 × 0.7 threads. There must be virtually no mismatch of
the inner bores in the miter joint.
Figure 2.9.18.-7. – Induction port

Dimensions in millimetres unless otherwise stated
Procedure for powder inhalers P0 = atmospheric pressure,

Place a suitable low resistance filter capable of quantitatively ∆P = pressure drop over the meter.
collecting the active substance in stage 5 and assemble the Adjust the flow control valve to achieve steady flow through
apparatus. Connect the apparatus to a flow system according the system at the required rate, Qout (± 5 per cent). Switch off
to the scheme specified in Figure 2.9.18.-8 and Table 2.9.18.-4. the pump. Ensure that critical flow occurs in the flow control
Unless otherwise defined, conduct the test at the flow rate, valve by the following procedure.
Qout, used in the test for uniformity of delivered dose, drawing
4 L of air from the mouthpiece of the inhaler and through the With the inhaler in place and the test flow rate established,
apparatus. measure the absolute pressure on both sides of the control
valve (pressure reading points P2 and P3 in Figure 2.9.18.-8).
Connect a flowmeter to the induction port. Use a flowmeter A ratio P3/P2 of less than or equal to 0.5 indicates critical flow.
calibrated for the volumetric flow leaving the meter, or Switch to a more powerful pump and re-measure the test flow
calculate the volumetric flow leaving the meter (Qout) using rate if critical flow is not indicated.
the ideal gas law. For a meter calibrated for the entering Dispense 20 mL of a solvent, capable of dissolving the active
volumetric flow (Qin), use the following expression : substance into each of the 4 upper stages of the apparatus and
replace the stoppers. Tilt the apparatus to wet the stoppers,
thereby neutralising electrostatic charge. Place a suitable
Qin ´ P0 mouthpiece adapter in position at the end of the induction
Qout =
P0 - ∆P port.

APPARATUS D - ANDERSEN CASCADE IMPACTOR

The Andersen 1 ACFM non-viable cascade impactor
consists of 8 stages together with a final filter. Material of
construction may be aluminium, stainless steel or other
suitable material. The stages are clamped together and sealed
with O-rings. Critical dimensions applied by the manufacturer
of apparatus D are provided in Table 2.9.18.-5. In use, some
occlusion and wear of holes will occur. In-use mensuration
tolerances need to be justified. In the configuration used
for pressurised inhalers (Figure 2.9.18.-9) the entry cone
of the impactor is connected to an induction port (see
Figure 2.9.18.-7). A suitable mouthpiece adapter is used to
provide an airtight seal between the inhaler and the induction
port. The front face of the inhaler mouthpiece must be flush
with the front face of the induction port.
Figure 2.9.18.-8. – Experimental set-up for testing powder
inhalers
Table 2.9.18.-4. – Component specification for Figure 2.9.18.-8

Code Item Description
A Connector ID ≥ 8 mm, e.g., short metal coupling, with
low-diameter branch to P3.
B Vacuum tubing A length of suitable tubing having an
ID ≥ 8 mm and an internal volume of
25 ± 5 mL.
C 2-way solenoid A 2-way, 2-port solenoid valve having a
valve minimum airflow resistance orifice with
ID ≥ 8 mm and an opening time ≤ 100 ms.
(e.g. type 256-A08, Bürkert GmbH,
D-74653 Ingelfingen), or equivalent.
D Vacuum pump Pump must be capable of drawing the required
flow rate through the assembled apparatus
with the powder inhaler in the mouthpiece
adapter (e.g. product type 1023, 1423 or 2565,
Gast Manufacturing Inc., Benton Harbor, MI
49022), or equivalent. Connect the pump to the
2-way solenoid valve using short and/or wide
(ID ≥ 10 mm) vacuum tubing and connectors
to minimise pump capacity requirements.
E Timer Timer capable to drive the 2-way solenoid
valve for the required duration (e.g. type
G814, RS Components International, Corby,
NN17 9RS, UK), or equivalent.
P2 P3 Pressure Determine under steady-state flow condition
measurements with an absolute pressure transducer.
F Flow control Adjustable regulating valve with maximum
valve Cv ≥ 1, (e.g. type 8FV12LNSS, Parker Hannifin
plc., Barnstaple, EX31 1NP, UK), or equivalent.
Prepare the powder inhaler for use according to patient

instructions. With the pump running and the 2-way solenoid
valve closed, locate the mouthpiece of the inhaler in the
mouthpiece adapter. Discharge the powder into the apparatus
by opening the valve for the required time, T (± 5 per cent).
Repeat the procedure. The number of discharges should be
minimised and typically would not be greater than 10. The
number of discharges is sufficient to ensure an accurate and
precise determination of fine particle dose.
Dismantle the filter stage of the apparatus. Carefully remove
the filter and extract the active substance into an aliquot of the
solvent. Remove the induction port and mouthpiece adapter
from the apparatus and extract the active substance into an
aliquot of the solvent. If necessary, rinse the inside of the inlet
jet tube to stage 1 with solvent, allowing the solvent to flow
into the stage. Extract the active substance from the inner Figure 2.9.18.-9. – Apparatus D : Andersen cascade impactor
walls and the collection plate of each of the 4 upper stages used for pressurised inhalers
of the apparatus into the solution in the respective stage by
carefully tilting and rotating the apparatus, observing that no In the configuration for powder inhalers, a pre-separator
liquid transfer occurs between the stages. is placed above the top stage to collect large masses of
non-respirable powder. It is connected to the induction port
Using a suitable method of analysis, determine the amount of as shown in Figure 2.9.18.-10. To accommodate high flow
active substance contained in each of the aliquots of solvent. rates through the impactor, the outlet nipple, used to connect
the impactor to the vacuum system is enlarged to have an
Calculate the fine particle dose (see Calculations). internal diameter of greater than or equal to 8 mm.

Table 2.9.18.-5. – Critical dimensions for apparatus D the horizontal axis of the induction port and the inhaler unit
Description Number Dimension (mm) is positioned in the same orientation as the intended use.
Connect a suitable pump to the outlet of the apparatus and
Stage 0 nozzle diameter 96 2.55 ± 0.025 adjust the air flow through the apparatus, as measured at
Stage 1 nozzle diameter 96 1.89 ± 0.025 the inlet to the induction port, to 28.3 L/min (± 5 per cent).
Switch off the pump.
Stage 2 nozzle diameter 400 0.914 ± 0.0127
Stage 3 nozzle diameter 400 0.711 ± 0.0127

Unless otherwise prescribed in the patient instructions, shake
Stage 4 nozzle diameter 400 0.533 ± 0.0127 the inhaler for 5 s and discharge one delivery to waste. Switch
Stage 5 nozzle diameter 400 0.343 ± 0.0127 on the pump to the apparatus, locate the mouthpiece end of
the actuator in the adapter and discharge the inverted inhaler
Stage 6 nozzle diameter 400 0.254 ± 0.0127 into the apparatus, depressing the valve for a sufficient time to
Stage 7 nozzle diameter 201 0.254 ± 0.0127 ensure complete discharge. Wait for 5 s before removing the
assembled inhaler from the adapter. Repeat the procedure.
Procedure for pressurised inhalers The number of discharges should be minimised and typically
Assemble the Andersen impactor with a suitable filter in would not be greater than 10. The number of discharges is
place. Ensure that the system is airtight. In that respect, follow sufficient to ensure an accurate and precise determination of
the manufacturer’s instructions. Place a suitable mouthpiece the fine particle dose. After the final discharge, wait for 5 s
adapter in position at the end of the induction port so that the and then switch off the pump.
mouthpiece end of the actuator, when inserted, lines up along
Figure 2.9.18.-10. – Connection of the induction port to the preseparator of the Andersen cascade impactor
Dimensions in millimetres unless otherwise stated

Dismantle the apparatus. Carefully remove the filter and evenly spaced on a logarithmic scale. In this flow range, there
extract the active substance into an aliquot of the solvent. are always at least 5 stages with D50 values between 0.5 µm
Remove the induction port and mouthpiece adapter from the and 6.5 µm. The collection efficiency curves for each stage are
apparatus and extract the active substance into an aliquot of sharp and minimise overlap between stages.
the solvent. Extract the active substance from the inner walls Material of construction may be aluminium, stainless steel or
and the collection plate of each of the stages of the apparatus other suitable material.
into aliquots of solvent.
The impactor configuration has removable impaction cups
Using a suitable method of analysis, determine the quantity of with all the cups in one plane (Figures 2.9.18.-11/14). There
active substance contained in each of the aliquots of solvent. are 3 main sections to the impactor ; the bottom frame
Calculate the fine particle dose (see Calculations). that holds the impaction cups, the seal body that holds the
Procedure for powder inhalers jets and the lid that contains the interstage passageways
(Figures 2.9.18.-11/12). Multiple nozzles are used at all but the
The aerodynamic cut-off diameters of the individual stages of
first stage (Figure 2.9.18.-13). The flow passes through the
this apparatus are currently not well-established at flow rates
impactor in a saw-tooth pattern.
other than 28.3 L/min. Users must justify and validate the use
of the impactor in the chosen conditions, when flow rates Critical dimensions are provided in Table 2.9.18.-6.
different from 28.3 L/min are selected.
Table 2.9.18.-6. – Critical dimensions for apparatus E
Assemble the Andersen impactor with the pre-separator and a
suitable filter in place and ensure that the system is airtight. Description Dimension
Depending on the product characteristics, the pre-separator (mm)
may be omitted, where justified and authorised. Stages 6 Pre-separator (dimension a - see Figure 2.9.18.-15) 12.8 ± 0.05
and 7 may also be omitted at high flow rates, if justified. The Stage 1* Nozzle diameter 14.3 ± 0.05
pre-separator may be coated in the same way as the plates
or may contain 10 mL of a suitable solvent. Connect the Stage 2* Nozzle diameter 4.88 ± 0.04
apparatus to a flow system according to the scheme specified Stage 3* Nozzle diameter 2.185 ± 0.02
in Figure 2.9.18.-8 and Table 2.9.18.-4.
Stage 4* Nozzle diameter 1.207 ± 0.01
Unless otherwise defined, conduct the test at the flow rate,
Qout, used in the test for uniformity of delivered dose drawing Stage 5* Nozzle diameter 0.608 ± 0.01
4 L of air from the mouthpiece of the inhaler and through the
Stage 6* Nozzle diameter 0.323 ± 0.01
apparatus.
Connect a flowmeter to the induction port. Use a flowmeter Stage 7* Nozzle diameter 0.206 ± 0.01
calibrated for the volumetric flow leaving the meter, or MOC* approx. 0.070
calculate the volumetric flow leaving the meter (Qout) using
the ideal gas law. For a meter calibrated for the entering Cup depth (dimension b - see Figure 2.9.18.-14) 14.625 ± 0.10
volumetric flow (Qin), use the following expression : Collection cup surface roughness (Ra) 0.5 - 2 µm
Qin ´ P0 Stage 1 nozzle to seal body distance** - dimension c 0 ± 1.18
Qout =
P0 - ∆P Stage 2 nozzle to seal body distance** - dimension c 5.236 ± 0.736
P0 = atmospheric pressure, Stage 3 nozzle to seal body distance** - dimension c 8.445 ± 0.410
∆P = pressure drop over the meter. Stage 4 nozzle to seal body distance** - dimension c 11.379 ± 0.237
Adjust the flow control valve to achieve steady flow through Stage 5 nozzle to seal body distance** - dimension c 13.176 ± 0.341
the system at the required rate, Qout (± 5 per cent). Ensure that Stage 6 nozzle to seal body distance** - dimension c 13.999 ± 0.071
critical flow occurs in the flow control valve by the procedure
described for Apparatus C. Switch off the pump. Stage 7 nozzle to seal body distance** - dimension c 14.000 ± 0.071
Prepare the powder inhaler for use according to the patient MOC nozzle to seal body distance** - dimension c 14.429 to 14.571
instructions. With the pump running and the 2-way solenoid
* See Figure 2.9.18.-13
valve closed, locate the mouthpiece of the inhaler in the
** See Figure 2.9.18.-14
mouthpiece adapter. Discharge the powder into the apparatus
by opening the valve for the required time, T (± 5 per cent). In routine operation, the seal body and lid are held together as
Repeat the discharge sequence. The number of discharges a single assembly. The impaction cups are accessible when this
should be minimised and typically would not be greater assembly is opened at the end of an inhaler test. The cups are
than 10. The number of discharges is sufficient to ensure an held in a support tray, so that all cups can be removed from
accurate and precise determination of fine particle dose. the impactor simultaneously by lifting out the tray.
Dismantle the apparatus. Carefully remove the filter and An induction port with internal dimensions (relevant to
extract the active substance into an aliquot of the solvent. the airflow path) defined in Figure 2.9.18.-7 connects to the
Remove the pre-separator, induction port and mouthpiece impactor inlet. A pre-separator can be added when required,
adapter from the apparatus and extract the active substance typically with powder inhalers, and connects between the
into an aliquot of the solvent. Extract the active substance induction port and the impactor. A suitable mouthpiece
from the inner walls and the collection plate of each of the adapter is used to provide an airtight seal between the inhaler
stages of the apparatus into aliquots of solvent. and the induction port.
Using a suitable method of analysis, determine the quantity of Apparatus E contains a terminal Micro-Orifice
active substance contained in each of the aliquots of solvent. Collector (MOC) that for most formulations will eliminate
Calculate the fine particle dose (see Calculations). the need for a final filter as determined by method validation.
The MOC is an impactor plate with nominally 4032 holes,
APPARATUS E each approximately 70 μm in diameter. Most particles not
Apparatus E is a cascade impactor with 7 stages and a captured on stage 7 of the impactor will be captured on the cup
micro-orifice collector (MOC). Over the flow rate range of surface below the MOC. For impactors operated at 60 L/min,
30 L/min to 100 L/min the 50 per cent-efficiency cut-off the MOC is capable of collecting 80 per cent of 0.14 µm
diameters (D50 values) range between 0.24 μm to 11.7 µm, particles. For formulations with a significant fraction of

Figure 2.9.18.-11. – Apparatus E (shown with the pre-separator in place)

particles not captured by the MOC, there is an optional filter sufficient to ensure an accurate and precise determination of
holder that can replace the MOC or be placed downstream of the fine particle dose. After the final discharge, wait for 5 s
the MOC (a glass fibre filter is suitable). and then switch off the pump.
Procedure for pressurised inhalers Dismantle the apparatus and recover the active substance as
follows : remove the induction port and mouthpiece adapter
Place cups into the apertures in the cup tray. Insert the cup from the apparatus and recover the deposited active substance
tray into the bottom frame, and lower into place. Close the into an aliquot of solvent. Open the impactor by releasing
impactor lid with the seal body attached and operate the the handle and lifting the lid. Remove the cup tray, with the
handle to lock the impactor together so that the system is collection cups, and recover the active substance in each cup
airtight. into an aliquot of solvent.
Connect an induction port with internal dimensions defined Using a suitable method of analysis, determine the quantity of
in Figure 2.9.18.-7 to the impactor inlet. Place a suitable active substance contained in each of the aliquots of solvent.
mouthpiece adapter in position at the end of the induction Calculate the fine particle dose (see Calculations).
port so that the mouthpiece end of the actuator, when
inserted, lines up along the horizontal axis of the induction Procedure for powder inhalers
port. The front face of the inhaler mouthpiece must be flush Assemble the apparatus with the pre-separator
with the front face of the induction port. When attached to (Figure 2.9.18.-15). Depending on the product characteristics,
the mouthpiece adapter, the inhaler is positioned in the same the pre-separator may be omitted, where justified.
orientation as intended for use. Connect a suitable pump to Place cups into the apertures in the cup tray. Insert the cup
the outlet of the apparatus and adjust the air flow through the tray into the bottom frame, and lower into place. Close the
apparatus, as measured at the inlet to the induction port, to impactor lid with the seal body attached and operate the
30 L/min (± 5 per cent). Switch off the pump. handle to lock the impactor together so that the system is
Unless otherwise prescribed in the patient instructions, shake airtight.
the inhaler for 5 s and discharge 1 delivery to waste. Switch When used, the pre-separator should be assembled as follows :
on the pump to the apparatus. Prepare the inhaler for use assemble the pre-separator insert into the pre-separator base.
according to the patient instructions, locate the mouthpiece Fit the pre-separator base to the impactor inlet. Add 15 mL
end of the actuator in the adapter and discharge the inhaler of the solvent used for sample recovery to the central cup of
into the apparatus, depressing the valve for a sufficient time the pre-separator insert. Place the pre-separator body on top
to ensure a complete discharge. Wait for 5 s before removing of this assembly and close the 2 catches.
the assembled inhaler from the adapter. Repeat the procedure. Connect an induction port with internal dimensions defined
The number of discharges should be minimised, and typically in Figure 2.9.18.-7 to the impactor inlet or pre-separator
would not be greater than 10. The number of discharges is inlet. Place a suitable mouthpiece adapter in position at the

Figure 2.9.18.-12. – Apparatus E showing component parts
Figure 2.9.18.-13. – Apparatus E : nozzle configuration
end of the induction port so that the mouthpiece end of the Qin × P0
Qout =
inhaler, when inserted, lines up along the horizontal axis of P0 − ∆P
the induction port. The front face of the inhaler mouthpiece
must be flush with the front face of the induction port. When P0 = atmospheric pressure,
attached to the mouthpiece adapter, the inhaler is positioned ∆P = pressure drop over the meter.
in the same orientation as intended for use. Connect the
apparatus to a flow system according to the scheme specified Adjust the flow control valve to achieve steady flow through
in Figure 2.9.18.-8 and Table 2.9.18.-4. the system at the required rate, Qout (± 5 per cent). Ensure that
Unless otherwise prescribed, conduct the test at the flow rate, critical flow occurs in the flow control valve by the procedure
Qout, used in the test for uniformity of delivered dose drawing described for Apparatus C. Switch off the pump.
4 L of air from the mouthpiece of the inhaler and through the Prepare the powder inhaler for use according to the patient
apparatus. Connect a flowmeter to the induction port. Use instructions. With the pump running and the 2-way solenoid
a flowmeter calibrated for the volumetric flow leaving the valve closed, locate the mouthpiece of the inhaler in the
meter, or calculate the volumetric flow leaving the meter (Qout) mouthpiece adapter. Discharge the powder into the apparatus
using the ideal gas law. For a meter calibrated for the entering by opening the valve for the required time, T (± 5 per cent).
volumetric flow (Qin), use the following expression : Repeat the discharge sequence. The number of discharges

Figure 2.9.18.-14. – Apparatus E : configuration of interstage passageways
Figure 2.9.18.-15. – Apparatus E : pre-separator configuration

should be minimised and typically would not be greater CALCULATIONS
than 10. The number of discharges is sufficient to ensure an
accurate and precise determination of fine particle dose. From the analysis of the solutions, calculate the mass of active
substance deposited on each stage per discharge and the mass
Dismantle the apparatus and recover the active substance as of active substance per discharge deposited in the induction
follows : remove the induction port and mouthpiece adapter port, mouthpiece adapter and when used, the pre-separator.
from the pre-separator, when used, and recover the deposited
active substance into an aliquot of solvent. When used, Starting at the final collection site (filter or MOC), derive
remove the pre-separator from the impactor, being careful to a table of cumulative mass versus cut-off diameter of the
avoid spilling the cup liquid into the impactor. Recover the respective stage (see Tables 2.9.18.-7 for Apparatus C, 2.9.18.-8
active substance from the pre-separator. for Apparatus D, 2.9.18.-9 for Apparatus E). Calculate by
Open the impactor by releasing the handle and lifting the lid. interpolation the mass of the active substance less than 5 µm.
Remove the cup tray, with the collection cups, and recover the This is the Fine Particle Dose (FPD).
active substance in each cup into an aliquot of solvent. If necessary, and where appropriate (e.g., where there is a
Using a suitable method of analysis, determine the quantity of log-normal distribution), plot the cumulative fraction of active
active substance contained in each of the aliquots of solvent. substance versus cut-off diameter (see Tables 2.9.18.-7/9) on
Calculate the fine particle dose (see Calculations). log probability paper, and use this plot to determine values

Table 2.9.18.-7. – Calculations for Apparatus C. Use q = (60 / Q) , where Q is the test flow rate in litres per minute (Qout for
powder inhalers)
Cut-off diameter Mass of active substance deposited Cumulative mass of active substance Cumulative fraction of active substance
(μm) per discharge deposited per discharge (per cent)
d4 = 1.7 × q mass from stage 5, m5* c4 = m5 f4 = (c4/c) × 100
d3 = 3.1 × q mass from stage 4, m4 c3 = c4 + m4 f3 = (c3/c) × 100
d2 = 6.8 × q mass from stage 3, m3 c2 = c3 + m3 f2 = (c2/c) × 100
mass from stage 2, m2 c = c2 + m2 100
* Stage 5 is the filter stage
Table 2.9.18.-8. – Calculations for Apparatus D when used at a flow rate of 28.3 L/min
Cut-off diameter Mass of active substance deposited Cumulative mass of active substance Cumulative fraction of active
(μm) per discharge deposited per discharge substance (per cent)
d7 = 0.4 mass from stage 8, m8 c7 = m8 f7 = (c7/c) × 100
d6 = 0.7 mass from stage 7, m7 c6 = c7 + m7 f6 = (c6/c) × 100
Table 2.9.18.-9. – Calculations for Apparatus E. Use q = (60/Q)x, where Q is the test flow rate in litres per minute, and x
is listed in the table
Cut-off diameter x Mass of active substance Cumulative mass of active substance Cumulative fraction of active
(μm) deposited per discharge deposited per discharge substance (per cent)
d7 = 0.34 × q 0.67 mass from MOC or terminal c7 = m8 F7 = (c7/c) × 100
filter, m8
d6 = 0.55 × q 0.60 mass from stage 7, m7 c6 = c7 + m7 F6 = (c6/c) × 100
for the Mass Median Aerodynamic Diameter (MMAD)

and Geometric Standard Deviation (GSD) as appropriate.
Appropriate computational methods may also be used.

EUROPEAN PHARMACOPOEIA 2.9.27. Uniformity of doses from multidose containers
01/2008:20927 Weigh individually 20 doses taken at random from one or

more containers with the measuring device provided and
determine the individual and average masses. Not more than
2 of the individual masses deviate from the average mass by
more than 10 per cent and none deviates by more than 20 per
2.9.27. UNIFORMITY OF MASS cent.
OF DELIVERED DOSES FROM
MULTIDOSE CONTAINERS
The following test is intended for oral dosage forms such as
granules, powders for oral use and liquids for oral use, which
are supplied in multidose containers provided at manufacture
with a measuring device.

EUROPEAN PHARMACOPOEIA 2.9.35. Powder fineness
04/2019:20935 Qr(x) = cumulative distribution of particles with a dimension

less than or equal to x where the subscript r reflects the
distribution type.
r Distribution type
0 Number
2.9.35. POWDER FINENESS(1)
1 Length
Particle-size distribution is estimated by analytical
2 Area
sieving (2.9.38) or by application of other suitable methods
where appropriate. A simple descriptive classification of 3 Volume
powder fineness is provided in this chapter. For practical
reasons, sieves are commonly used to measure powder Therefore, by definition :
fineness. Sieving is most suitable where a majority of the Qr(x) = 0.90 when x = x90
particles are larger than about 75 μm, although it can be used Qr(x) = 0.50 when x = x50
for some powders having smaller particle sizes where the Qr(x) = 0.10 when x = x10
method can be validated. Light diffraction is also a widely used
technique for measuring the size of a wide range of particles. An alternative but less informative method of classifying
powder fineness is by use of the descriptive terms in
Where the cumulative distribution has been determined by Table 2.9.35.-1.
analytical sieving or by application of other methods, particle
size may be characterised in the following manner : Table 2.9.35.-1.
x90 = particle size corresponding to 90 per cent of the Classification of powders by fineness
cumulative undersize distribution ;
Cumulative
x50 = median particle size (i.e. 50 per cent of the particles Descriptive term x50 (μm) distribution by
are smaller and 50 per cent of the particles are volume basis, Q3(x)
larger) ;
Coarse > 355 Q3(355) < 0.50
x10 = particle size corresponding to 10 per cent of the
cumulative undersize distribution. Q3(180) < 0.50 and
Moderately fine 180 - 355
Q3(355) ≥ 0.50
It is recognised that the symbol d is also widely used to
designate these values. Therefore, the symbols d90, d50, d10 Q3(125) < 0.50 and
Fine 125 - 180
may be used. Q3(180) ≥ 0.50
The following parameters may be defined based on the
Very fine ≤ 125 Q3(125) ≥ 0.50
cumulative distribution.

EUROPEAN PHARMACOPOEIA 2.9.40. Uniformity of dosage units
04/2017:20940 The test for mass variation is applicable for the following
dosage forms :
(1) solutions enclosed in single-dose containers and in soft
capsules ;
2.9.40. UNIFORMITY OF (2) solids (including powders, granules and sterile solids) that
are packaged in single-dose containers and contain no added
DOSAGE UNITS(1) active or inactive substances ;
To ensure the consistency of dosage units, each unit in a batch (3) solids (including sterile solids) that are packaged in
should have an active substance content within a narrow single-dose containers, with or without added active or
range around the label claim. Dosage units are defined as inactive substances, that have been prepared from true
dosage forms containing a single dose or a part of a dose of solutions and freeze-dried in the final containers and are
an active substance in each dosage unit. ◊Unless otherwise labelled to indicate this method of preparation ;
stated,◊ the uniformity of dosage units specification is not (4) hard capsules, uncoated tablets, or film-coated tablets,
intended to apply to solutions, suspensions, emulsions or containing 25 mg or more of an active substance comprising
gels in single-dose containers intended for local action 25 per cent or more, by mass, of the dosage unit or, in the case
following cutaneous administration. ◊The test for content of hard capsules, the capsule contents, except that uniformity
uniformity is not required for multivitamin, single-vitamin of other active substances present in lesser proportions is
and trace-element preparations.◊ demonstrated by meeting content uniformity requirements.
The term ‘uniformity of dosage unit’ is defined as the degree
of uniformity in the amount of the active substance among The test for content uniformity is required for all dosage
dosage units. Therefore, the requirements of this chapter forms not meeting the above conditions for the mass
apply to each active substance being comprised in dosage units variation test. ♦Alternatively, products that do not meet the
containing 1 or more active substances, unless otherwise 25 mg/25 per cent threshold limit may be tested for uniformity
specified elsewere in this Pharmacopoeia. of dosage units by mass variation instead of the content
uniformity test on the following condition : the concentration
The uniformity of dosage units can be demonstrated by either Relative Standard Deviation (RSD) of the active substance in
of 2 methods : content uniformity or mass variation (see the final dosage units is not more than 2 per cent, based on
Table 2.9.40.-1). process validation data and development data, and if there has
The test for content uniformity of preparations presented in been regulatory approval of such a change. The concentration
dosage units is based on the assay of the individual contents of RSD is the RSD of the concentration per dosage unit (m/m
active substance(s) of a number of dosage units to determine or m/V), where concentration per dosage unit equals the assay
whether the individual contents are within the limits set. The result per dosage unit divided by the individual dosage unit
content uniformity method may be applied in all cases. mass. See the RSD formula in Table 2.9.40.-2.♦
Table 2.9.40.-1. – Application of Content Uniformity (CU) and Mass Variation (MV) test for dosage forms
Dosage forms Type Sub-Type Dose and ratio of active substance
≥ 25 mg and ≥ 25 per cent < 25 mg or < 25 per cent
Tablets uncoated MV CU
coated film-coated MV CU
others CU CU
Capsules hard MV CU
soft suspensions, emulsions, gels CU CU
solutions MV MV
Solids in single-dose MV MV
single component
containers
solution freeze-dried in final MV MV

multiple components
container
others CU CU
Solutions enclosed in MV MV
single-dose containers
Others : dosage forms CU CU

not addressed by the
other categories in this
table including but not
limited to suppositories,
transdermal patches and
semi-solid preparations
applied cutaneously and
intended for systemic
distribution of the active
substance

2.9.40. Uniformity of dosage units EUROPEAN PHARMACOPOEIA
Table 2.9.40.-2.
Variable Definition Conditions Value
Mean of individual contents (x1,

X x2,..., xn), expressed as a percentage
of the label claim
x1, x2,..., xn Individual contents of the

dosage units tested, expressed
as a percentage of the label claim
n Sample size (number of dosage units

in a sample)
k Acceptability constant If n = 10, then 2.4
If n = 30, then 2.0
s Sample standard deviation 1 /2

é n 2ù
ê å (x i - X ) ú
ê ú
êi=1 ú
ê n-1 ú
ê ú
ê ú
êë úû
RSD Relative standard deviation

100s
X
M (case 1) Reference value If 98.5 per cent ≤ X ≤ 101.5 per M = X

To be applied when T ≤ 101.5 cent, then (AV = ks)
If X < 98.5 per cent, then M = 98.5 per cent

(AV = 98.5 − X + ks)
If X > 101.5 per cent, then M = 101.5 per cent

(AV = X − 101.5 + ks)
M (case 2) Reference value If 98.5 per cent ≤ X ≤ T, then M = X

To be applied when T > 101.5 (AV = ks)
If X < 98.5 per cent, then M = 98.5 per cent

(AV = 98.5 − X + ks)
If X > T, then M = T per cent

(AV = X − T + ks)
Acceptance value (AV) General formula :

M - X + ks
Calculations are specified above
for the different cases.
L1 Maximum allowed acceptance value L1 = 15.0 unless otherwise specified
L2 Maximum allowed range for On the low side, no dosage unit L2 = 25.0 unless otherwise specified
deviation of each dosage unit tested result can be less than 0.75 M while
from the calculated value of M on the high side, no dosage unit
result can be greater than 1.25 M
(This is based on L2 value of 25.0)
T Target content per dosage unit at

time of manufacture, expressed
as a percentage of the label claim.
Unless otherwise stated, T is
equal to 100 per cent or T is the
manufacturer’s approved target
content per dosage unit
CONTENT UNIFORMITY Liquid or semi-solid dosage forms. Assay 10 units

Select not fewer than 30 units, and proceed as follows for the individually using an appropriate analytical method. Carry
dosage form designated. Where different procedures are used out the assay on the amount of well-mixed material that
for assay of the preparation and for the content uniformity is removed from an individual container in conditions of
test, it may be necessary to establish a correction factor to be normal use. Express the results as delivered dose. Calculate
applied to the results of the latter. the acceptance value (see Table 2.9.40.-2).
Solid dosage forms. Assay 10 units individually using an Calculation of Acceptance Value
appropriate analytical method. Calculate the acceptance value Calculate the Acceptance Value (AV) using the formula :
(see Table 2.9.40.-2).
M - X + ks
for which the terms are as defined in Table 2.9.40.-2.

EUROPEAN PHARMACOPOEIA 2.9.40. Uniformity of dosage units
MASS VARIATION Liquid ◊or semi-solid◊ dosage forms. Accurately weigh the
Carry out an assay for the active substance(s) on a amount of liquid or semi-solid that is removed from each of 10
representative sample of the batch using an appropriate individual containers in conditions of normal use. If necessary,
analytical method. This value is result A, expressed as compute the equivalent volume after determining the density.
percentage of label claim (see Calculation of Acceptance Calculate the active substance content in each container from
Value). Assume that the concentration (mass of active the mass of product removed from the individual containers
substance per mass of dosage unit) is uniform. Select not and the result of the assay. Calculate the acceptance value.
fewer than 30 dosage units, and proceed as follows for the Calculation of Acceptance Value. Calculate the acceptance
dosage form designated. value (AV) as shown in content uniformity, except that
the individual contents of the units are replaced with the
Uncoated or film-coated tablets. Accurately weigh 10 tablets individual estimated contents defined below.
individually. Calculate the active substance content, expressed
as percentage of label claim, of each tablet from the mass of x1, x2,..., xn = individual estimated contents of the
the individual tablets and the result of the assay. Calculate the dosage units tested ;
acceptance value. where
Hard capsules. Accurately weigh 10 capsules individually, A
taking care to preserve the identity of each capsule. Remove xi = wi ´
W
the contents of each capsule by suitable means. Accurately
weigh the emptied shells individually, and calculate for each w1, w2,..., wn = individual masses of the dosage units
capsule the net mass of its contents by subtracting the mass of tested ;
the shell from the respective gross mass. Calculate the active A = content of active substance (percentage of
substance content in each capsule from the mass of product label claim) obtained using an appropriate
removed from the individual capsules and the result of the analytical method (assay);
assay. Calculate the acceptance value. W = mean of individual masses (w1, w2,..., wn).
Soft capsules. Accurately weigh 10 intact capsules individually
to obtain their gross masses, taking care to preserve the CRITERIA
identity of each capsule. Then cut open the capsules by means Apply the following criteria, unless otherwise specified.
of a suitable clean, dry cutting instrument such as scissors or a Solid, semi-solid and liquid dosage forms. The requirements
sharp open blade, and remove the contents by washing with a for dosage uniformity are met if the acceptance value of
suitable solvent. Allow the occluded solvent to evaporate from the first 10 dosage units is less than or equal to L1 per
the shells at room temperature over a period of about 30 min, cent. If the acceptance value is greater than L1 per cent,
taking precautions to avoid uptake or loss of moisture. Weigh test the next 20 dosage units and calculate the acceptance
the individual shells, and calculate the net contents. Calculate value. The requirements are met if the final acceptance
the active substance content in each capsule from the mass of value of the 30 dosage units is less than or equal to L1 per
product removed from the individual capsules and the result cent and no individual content of the dosage unit is less
of the assay. Calculate the acceptance value. than (1 − L2 × 0.01)M or more than (1 + L2 × 0.01)M in
Solid dosage forms other than tablets and capsules. Proceed calculation of acceptance value under content uniformity or
as directed for hard capsules, treating each unit as described under mass variation. Unless otherwise specified, L1 is 15.0
therein. Calculate the acceptance value. and L2 is 25.0.

EUROPEAN PHARMACOPOEIA 2.9.44. Preparations for nebulisation : characterisation
01/2012:20944 Filter system. A suitably validated low-resistance filter,

capable of quantitatively collecting the aerosol and enabling
recovery of the active substance with an appropriate solvent,
is used for the test. The dead volume of the filter casing does
not exceed 10 per cent of the tidal volume used in the breath
2.9.44. PREPARATIONS simulation.
FOR NEBULISATION : METHOD
Attach the filter (contained in the filter holder) (A) to the
CHARACTERISATION breath simulator (B) according to Figure 2.9.44.-1. Fill the
Products used for nebulisation and intended for pulmonary nebuliser (C) with the volume of the medicinal product as
delivery are characterised using the following tests : specified in the patient instructions. Attach the mouthpiece
– Active substance delivery rate and total active substance of the nebuliser to the inhalation filter using a mouthpiece
delivered ; adapter if required, ensuring that connections are airtight.
Make sure the nebuliser is positioned in the same orientation
– Aerodynamic assessment of nebulised aerosols.
as intended for use ; this may require tilting the breathing
These tests standardise the approach given to the assessment simulator and filter holder. Set the breathing simulator to
of the dose that would be delivered to a patient but are generate the specified breathing pattern.
not intended to provide assessment of the nebuliser
device itself, which is described in the European standard
EN 13544-1:2007+A1:2009, Respiratory therapy equipment -
Part 1 : Nebulizing systems and their components.
The mass- rather than the number-weighted size distribution
is more appropriate to evaluate product performance. Indeed,
active substance mass as a function of aerodynamic diameter
is more indicative of therapeutic effect within the respiratory
tract. A. inhalation filter and filter holder B. breathing simulator C. nebuliser
ACTIVE SUBSTANCE DELIVERY RATE AND TOTAL Figure 2.9.44.-1. – Experimental set-up for breathing simulator
ACTIVE SUBSTANCE DELIVERED testing
These tests are performed to assess the rate of delivery to Start the breathing simulator then, at the beginning of an
the patient and the total active substance delivered to the inhalation cycle, start the nebuliser. Operate the nebuliser
patient, using standardised conditions of volumetric flow for a defined initial time period. The time chosen, usually
rate. It is essential that breath-enhanced and breath-actuated 60 ± 1 s, must allow sufficient active substance deposition
nebulisers be evaluated by a breathing simulator, as the output on the inhalation filter to allow quantitative analysis. If the
of these types of device is highly dependent on inhalation quantity of active substance deposited on the inhalation filter
flow rate. The methodology below describes the use of a in 60 s is insufficient for this analysis, the length of the time
standard breathing pattern defined for adults. Should a interval for aerosol collection can be increased. If the filter is
particular product for nebulisation only be indicated for soaked with the preparation, this time can be decreased. At
paediatric (i.e. neonate, infant or child) use, then paediatric the end of this initial period, stop the nebuliser.
breathing pattern(s) must be used. Breathing patterns are
used, rather than continuous flow rates, to provide a more Place a fresh filter and filter holder in position and continue
appropriate measure of the mass of active substance that until nebulisation ceases. Interrupt nebulisation and exchange
would be delivered to patients. filters if necessary, to avoid filter saturation.
Active substance delivery rate and total active substance RESULTS
delivered are appropriate characteristics because they allow Using a suitable method of analysis, determine the mass of
the mass delivered to be characterised in a standard way active substance collected on the filters and filter holders
regardless of the nebuliser used. Accordingly, the test during each time interval. Determine the active substance
methodology described below allows that the mass of active delivery rate by dividing the mass of active substance collected
substance delivered in the 1st period (typically 1 min) is on the first inhalation filter by the time interval used for
measured (consequently giving an assessment of active collection. Determine the total mass of active substance
substance delivery rate) as well as capturing the total mass delivered by summing the mass of active substance collected
of active substance delivered. on all inhalation filters and filter holders.
APPARATUS
AERODYNAMIC ASSESSMENT OF NEBULISED
Breathing simulator. A commercially available breathing AEROSOLS
simulator, which is able to generate the breathing profiles
specified in Table 2.9.44.-1, is used for the test. The breathing Nebulised products need to be size-characterised at flow
profile indicated for adults is used unless the medicinal product rates lower than the range that is normally used for powder
is specifically intended for use in paediatrics, where alternate inhalers and metered-dose inhalers. A flow rate of 15 L/min
patterns should be used, as indicated in Table 2.9.44.-1. is recommended in the European standard because this value
represents a good approximation to the mid-inhalation flow
Table 2.9.44.-1. – Breathing simulator specifications rate achievable by a tidally breathing healthy adult (500 mL
Item Specification tidal volume).
Adult Neonate Infant Child Although low-angle laser light scattering instruments
(laser diffractometers) can provide rapid size-distribution
Tidal 500 mL 25 mL 50 mL 155 mL measurements of nebuliser-generated aerosols, these
volume
techniques do not detect the active substance ; rather they
Frequency 15 40 30 25 measure the size distribution of the droplets irrespective of
cycles/min cycles/min cycles/min cycles/min
their content. This may not be a problem with homogeneous
Waveform sinusoidal sinusoidal sinusoidal sinusoidal solutions, but can result in significant error if the product
Inhala- 1:1 1:3 1:3 1:2 to be nebulised is a suspension, or if droplet evaporation is
tion/exha- significant as can be the case with certain nebuliser types.
lation ratio Cascade impactors enable the aerosol to be characterised

2.9.44. Preparations for nebulisation : characterisation EUROPEAN PHARMACOPOEIA
unambiguously in terms of the mass of active substance as a Re-entrainment. Droplet bounce and re-entrainment are
function of aerodynamic diameter. Laser diffraction may be less likely with nebuliser-produced droplets than with solid
used if validated against a cascade impaction method. particles from inhalers and for that reason coating would not
Apparatus E (see below under Apparatus), a cascade impactor, normally be required.
has been calibrated at 15 L/min specifically to meet the METHOD
recommendation of the European standard, and is therefore Pre-cool the assembled impactor and induction port in a
used for this test. Determining mass balance in the same refrigerator (set at about 5 °C) for not less than 90 min and
way as for powder inhalers and metered-dose inhalers is start the determination within about 5 min of removal of the
not straightforward, in that the dose is being captured as impactor from the refrigerator. Other methods that maintain
a continuous output, and hence is not included. As part the impactor at a constant temperature (for example, use of a
of method development, recovery experiments must be cooling cabinet) can also be employed when validated.
performed to validate the method.
Set up the nebuliser with a supply of driving gas (usually
It is also recognised that the control of evaporation of air or oxygen), or use a compressor, at the pressure and
droplets produced by nebulisers may be critical to avoid flow rate specified by the manufacturer of the nebuliser.
bias in the droplet size assessment process. Evaporation can Take precautions to ensure that the gas supply line does not
be minimised by cooling the impactor to a temperature of become detached from the nebuliser when under pressure.
about 5 °C, typically achieved by cooling the impactor in a Fill the nebuliser with the volume of the medicinal product as
refrigerator for about 90 min. Typically, at least after each specified in the patient instructions.
day of use, the apparatus must be fully cleaned, including the
inter-stage passageways, in view of the greater risk of corrosion Remove the impactor from the refrigerator. Attach the
caused by the condensation/accumulation of saline-containing induction port to the impactor, and connect the outlet of the
droplets on inter-stage metalwork associated with cooling impactor/external filter to a vacuum source that is capable of
the impactor. It is recommended to dry all surfaces of the drawing air through the system at 15 L/min as specified in
apparatus after each test, for example with compressed air. Figure 2.9.44.-2. Turn on the flow through the impactor.
Note : the micro-orifice collector (MOC) should not be dried Connect a flow meter, calibrated for the volumetric flow
with compressed air. leaving the meter, to the induction port. Adjust the flow
control valve located between the impactor and the vacuum
APPARATUS
source to achieve a steady flow through the system at 15 L/min
A detailed description of Apparatus E and the induction port (± 5 per cent). Remove the flow meter.
is contained in general chapter 2.9.18, and includes details
of critical dimensions and the qualification process for the Make sure the nebuliser is positioned in the same orientation
impactor (stage mensuration). as intended for use then attach the mouthpiece of the
nebuliser to the induction port, using a mouthpiece adapter if
A back-up filter in addition to the micro-orifice required, ensuring that connections are airtight. Switch on the
collector (MOC) must be used to ensure quantitative recovery flow/compressor for the nebuliser. Sample for a predetermined
of active substance from the nebulised aerosol at the specified time (T0). Once determined, this time (T0) must be defined
flow rate of 15 L/min. The filter is located below the MOC and used in the analytical method for a particular medicinal
(internal filter option) or a filter in holder, external to the product to ensure that mass fraction data can be compared.
impactor, is used to capture any fine droplets that pass beyond At the end of the sampling period, switch off the driving gas
the last size fractionating stage. flow/compressor to the nebuliser, remove the nebuliser from
A pre-separator is not used for testing nebuliser-generated the induction port and switch off the flow from the vacuum
aerosols. source to the impactor.
METHOD VALIDATION Dismantle the impactor and, using a suitable method of
Impactor stage overloading. During method development analysis, determine the mass of active substance collected
and validation, it is important to confirm that the volume in the induction port, on each stage and on the back-up
of liquid sampled from the nebuliser does not overload the filter/external filter as described for Apparatus E in general
impactor. Visual inspection of the collection surfaces on chapter 2.9.18. Add the mass of active substance collected in
stages collecting most of the droplets may reveal streaking the MOC to that deposited on the back-up filter/external filter
if overloading has occurred. This phenomenon is usually and treat as a single sample for the purpose of subsequent
also associated with an increase in mass of active substance calculations.
collected on the final stage and back-up filter. Reducing Calculate the mass fraction (Fm,comp) of the active substance
the sampling period (T0) is the most effective way to avoid deposited on each component of the impactor, commencing
overloading in any given system, balancing overloading with with the induction port and proceeding in order through the
analytical sensitivity. impactor, using the following expression:
A. nebuliser B. induction port C. impactor (apparatus E) D. flow control valve E. vacuum source
Figure 2.9.44.-2. – Apparatus E for measuring the size distribution of preparations for nebulisation

EUROPEAN PHARMACOPOEIA 2.9.44. Preparations for nebulisation : characterisation
mcomp If necessary, and where appropriate, determine values for

Fm,comp =
M the mass median aerodynamic diameter (MMAD) and the
mcomp = mass associated with the component under geometric standard deviation (GSD), as appropriate.
evaluation ;
M = total mass collected by the system.
Present Fm,comp in order of location within the measurement
equipment, beginning at the induction port and ending with
the back-up filter of the impactor (see Figure 2.9.44.-3). Where
appropriate, Fm,comp for adjacent stages of the impactor may be
combined in order to report the mass fraction collected on a
group of stages as a single value.
Determine the cumulative mass-weighted particle-size
distribution of the aerosol size-fractionated by the impactor in
accordance with the procedure given in general chapter 2.9.18.
Starting at the filter, derive a cumulative mass versus effective
cut-off diameter of the respective stages (see Table 2.9.44.-2
for the appropriate cut-off diameters at 15 L/min). Plot the
cumulative fraction of active substance versus cut-off diameter
in a suitable format, for example logarithmic or log-probability
format. Where appropriate, determine by interpolation the
fraction either below a given size or between an upper and
a lower size limit.
Table 2.9.44.-2. – Cut-off sizes for Apparatus E at 15 L/min
Stage Cut-off diameter (μm) Figure 2.9.44.-3. – Example of mass fraction of droplets
presented in terms of location within the sampling system
1 14.1
2 8.61
3 5.39
4 3.30
5 2.08
6 1.36
7 0.98

EUROPEAN PHARMACOPOEIA 2.9.47. Uniformity of dosage units using large sample sizes
04/2013:20947 ALTERNATIVE 1 (PARAMETRIC)

corrected 8.1 Select not fewer than 100 units according to a predefined
sampling plan.
The consistency of dosage units is evaluated by content
uniformity or mass variation as prescribed in Table 2.9.40.-1.
Calculate the acceptance value (AV) using the following
2.9.47. DEMONSTRATION OF expression :
UNIFORMITY OF DOSAGE UNITS M - X + ks
USING LARGE SAMPLE SIZES for which the terms are defined in Table 2.9.40.-2, but using the
The procedure is intended for, but not limited to, the evaluation sample size-dependent value for k defined in Table 2.9.47.-1.
of medicinal products that are manufactured using process CRITERIA
analytical technology (PAT) methodology. Apply the following criteria, unless otherwise specified.
Compliance with general chapter 2.9.40. Uniformity of The requirements for dosage form uniformity are met if :
dosage units can be demonstrated by the following procedure, 1. the acceptance value (AV) is less than or equal to L1 ; and
when large samples (sample size n ≥ 100) are evaluated.
2. in the calculation of acceptance value (AV) under content
Application of this chapter does not constitute a mandatory uniformity or under mass variation, the number of
requirement. It presents 2 alternative tests (Alternative 1 and individual dosage units outside (1 ± L2 × 0.01)M is less
Alternative 2). Fulfilling the requirements of either of the than or equal to c2 as defined for a given sample size n in
2 alternatives is considered as evidence that the medicinal Table 2.9.47.-1.
product tested complies with general chapter 2.9.40. The
2 alternatives are considered equivalent in their demonstration Unless otherwise specified, L1 is 15.0 and L2 is 25.0.
of compliance with general chapter 2.9.40. Table 2.9.47.-1. is to be interpreted as follows :
Table 2.9.47.-1. – Acceptability constant (k) and acceptable number of dosage units with a content outside (1 ± L2 × 0.01)M
(= c2) for a given sample size n
n (≥) k c2 n (≥) k c2 n (≥) k c2 n (≥) k c2 n (≥) k c2 n (≥) k c2
100 2.15 804 2.26 2480 2.29 23 4366 2.30 41 6252 2.31 59 8243 2.31 78
7
105 2.16 905 2.27 2585 2.29 24 4471 2.30 42 6357 2.31 60 8347 2.31 79
120 2.17 0 908 2.27 8 2690 2.29 25 4576 2.30 43 6462 2.31 61
8452 2.31 80
1013 2.27 9 2794 2.29 26 4680 2.30 44 6566 2.31 62
139 2.18
8557 2.31 81
1118 2.27 10 2899 2.29 27 4785 2.30 45 6671 2.31 63
161 2.19
8662 2.31 82
1223 2.27 3004 2.29 28 4890 2.30 46 6776 2.31 64
176 2.19
11 8767 2.31 83
1276 2.28 3109 2.29 4995 2.30 47 6881 2.31 65
189 2.20 29
1 8871 2.31 84
1328 2.28 12 3171 2.30 5099 2.30 48 6985 2.31 66
224 2.21
1432 2.28 13 3213 2.30 30 5204 2.30 49 7090 2.31 67 8976 2.31 85
270 2.22
1537 2.28 14 3318 2.30 31 5309 2.30 50 7195 2.31 68 9081 2.31 86
280 2.22
2 1642 2.28 15 3423 2.30 32 5414 2.30 51 7300 2.31 69 9186 2.31 87
328 2.23
1747 2.28 16 3528 2.30 33 5519 2.30 52 7404 2.31 70
9290 2.31 88
385 2.23
3 1851 2.28 3633 2.30 34 5623 2.30 53 7509 2.31 71
17 9395 2.31 89
407 2.24
1918 2.29 3737 2.30 35 5728 2.30 54 7614 2.31 72
9500 2.31 90
490 2.24
1956 2.29 18 3842 2.30 36 5833 2.30 55 7719 2.31 73
4
9605 2.31 91
516 2.25
2061 2.29 19 3947 2.30 37 5938 2.30 56 7824 2.31 74
594 2.25 9710 2.31 92

2166 2.29 20 4052 2.30 38 6042 2.30 7928 2.31 75
5 57
672 2.26 2270 2.29 21 4156 2.30 39 6136 2.31 8033 2.31 76 9814 2.31 93
699 2.26 6 2375 2.29 22 4261 2.30 40 6147 2.31 58 8138 2.31 77 9919 2.31 94

2.9.47. Uniformity of dosage units using large sample sizes EUROPEAN PHARMACOPOEIA
Table 2.9.47.-2. – Acceptable number of individual dosage units with a content outside (1 ± L1 × 0.01)T (= c1) and
(1 ± L2 × 0.01)T (= c2) respectively, for a given sample size n
n (≥) c1 c2 n (≥) c1 c2 n (≥) c1 c2 n (≥) c1 c2 n (≥) c1 c2 n (≥) c1 c2 n (≥) c1 c2
100 3 1432 35 2899 67 4366 98 5833 129 7300 160 8767 191
123 4 0 1476 36 13 2935 68 27 4377 99 41 5835 130 7304 161 8780 192 83
55 69
159 5 1521 37 2981 69 4424 100 5883 131 7351 162
8828 193
176 5 1537 37 3004 69 4471 101 5930 132 7399 163
8871 193
196 6 1566 38 14 3027 70 28 4518 102 42 5938 132 7404 163
1 8875 194
1611 39 3073 71 4565 103 5977 133 56 7447 164 70
234 7 84
1642 39 8923 195
273 8 3109 71 4576 103 6024 134 7494 165
1656 40 3120 72 6042 134 7509 165 8971 196
280 8 15 4612 104 43
29
1701 41 3166 73 6072 135 57 7542 166 71 8976 196
313 9 2 4658 105
1746 42 3212 74 6119 136 7589 167
353 10 4680 105 9019 197 85
1747 42 3213 74 6147 136 7614 167
385 10 4705 106 44 9066 198
1791 43 16
3259 75 30 6166 137 58 7637 168 72
394 11 4752 107 9081 198
3 1836 44
3305 76 6214 138 7684 169
434 12 4785 107 9114 199 86
1851 44
3318 76 6252 138 7719 169
476 13 4799 108 45
1882 45 17 9162 200
3351 77 31 6261 139 7732 170 73
490 13 4846 109 59
1927 46 9186 200
3398 78 6308 140 7779 171
517 14 4 4890 109
1956 46 9210 201 87
3423 78 6355 141 7824 171
559 15 4893 110
1972 47 18
3444 79 32 46 6357 141 7827 172 9257 202
594 15 2018 48 4940 111 74
3491 80 6403 142 60 7875 173 9290 202
601 16 2061 48 4987 112
5 3528 80 6450 143 7922 174
4995 112 9305 203 88
644 17 2063 49
19 3537 81 6462 143 7928 174
686 18 2109 50 33 5034 113 47 9353 204
3584 82 6498 144 61 7970 175 75
699 18 2154 51 5081 114 9395 204
3630 83 6545 145 8017 176
729 19 6 2166 51 5099 114 9401 205
3633 83 6566 145 8033 176 89
772 20 2200 52 20 5128 115 48 9449 206
3677 84 34 6592 146 62 8065 177 76
804 20 2246 53 5175 116
3723 85 6640 147 8113 178 9496 207
815 21 2270 53 5204 116
7 3737 85 6671 147 8138 178 9500 207
858 22 2291 54 21 5222 117 49
3770 86 35 6687 148 63 8160 179 77 9544 208 90
2337 55 5269 118
902 23
3817 87 6734 149 8208 180 9592 209
2375 55
908 23 5309 118
3842 87 6776 149 8243 180
2383 56 9605 209
945 24 8 22 5317 119
3863 88 36 50 6782 150 8256 181 78
2429 57 64 9640 210 91
989 25 5364 120
3910 89 6829 151 8303 182
2475 58
1013 25 5411 121 9688 211
3947 89 6877 152 8347 182
2480 58
1033 26 9 5414 121 9710 211
3956 90 6881 152 8351 183
2520 59 23
37 5458 122 51 79
1077 27 4003 91 6924 153 65 8399 184 9735 212 92
2566 60
1118 27 5505 123 9783 213
2585 60 4050 92 6972 154 8446 185
1121 28 5519 123
2612 61 24 4052 92 6985 154 8452 185 9814 213
10
1165 29 4097 93 38 5552 124 52 7019 155 66 8494 186 80
2658 62 9831 214 93
1209 30 4143 94 5599 125 7067 156 8542 187
2690 62 9879 215
1223 30 4156 94 5623 125 7090 156 8557 187
2704 63 25
9919 215
1253 31 11 4190 95 39 5647 126 53 7114 157 67 8589 188 81
2750 64
9927 216
1298 32 2794 64 4237 96 5694 127 7161 158 8637 189
5728 127 9975 217 94
1328 32 2796 65 4261 96 7195 158 8662 189
26
1342 33 12 2843 66 4284 97 40 5741 128 54 7209 159 68 8685 190 82 10023 218
1387 34 2889 67 4330 98 5788 129 7256 160 8732 191 10070 219

EUROPEAN PHARMACOPOEIA 2.9.47. Uniformity of dosage units using large sample sizes
– for a sample size of n = 400, enter the table at n ≥ 385 : CRITERIA

k = 2.23 and c2 = 3 ; Apply the following criteria, unless otherwise specified.
– for a sample size of n = 450, enter the table at n ≥ 407 : The requirements for dosage form uniformity are met if :
k = 2.24 and c2 = 3 ; 1. the number of individual dosage units outside
– for a sample size of n = 500, enter the table at n ≥ 490 : (1 ± L1 × 0.01)T is less than or equal to c1 ; and
k = 2.24 and c2 = 4. 2. the number of individual dosage units outside
ALTERNATIVE 2 (NON-PARAMETRIC) (1 ± L2 × 0.01)T is less than or equal to c2.
Select not fewer than 100 units according to a predefined c1 and c2 for a given sample size n are defined in Table 2.9.47.-2.
sampling plan. Unless otherwise specified, L1 is 15.0 and L2 is 25.0.
The consistency of dosage units is evaluated by content Table 2.9.47.-2 is to be interpreted as follows :
uniformity or mass variation as prescribed in Table 2.9.40.-1. – for a sample size of n = 400, enter the table at n ≥ 394 :
Assay individually or weigh the units and calculate individual c1 = 11 and c2 = 3 ;
contents as prescribed in general chapter 2.9.40. Count the
number of individual dosage units with a content outside – for a sample size of n = 450, enter the table at n ≥ 434 :
(1 ± L1 × 0.01)T and the number of individual dosage units c1 = 12 and c2 = 3 ;
with a content outside (1 ± L2 × 0.01)T. Evaluate if the values – for a sample size of n = 500, enter the table at n ≥ 490 :
are within the limits defined in Table 2.9.47.-2. c1 = 13 and c2 = 4.

EUROPEAN PHARMACOPOEIA 3.2.1. Glass containers for pharmaceutical use
01/2019:30201 or coloured glass is used for the other pharmaceutical

preparations. It is recommended that all glass containers
for liquid preparations and for powders for parenteral
administration permit the visual inspection of the contents.
The inner surface of glass containers may be specially treated
3.2.1. GLASS CONTAINERS FOR to improve hydrolytic resistance, to confer water-repellancy,
etc. The outer surface may also be treated, for example to
PHARMACEUTICAL USE reduce friction and to improve resistance to abrasion. The
Glass containers for pharmaceutical use are glass articles outer treatment is such that it does not contaminate the inner
intended to come into direct contact with pharmaceutical surface of the container.
preparations. Except for type I glass containers, glass containers for
Colourless glass is highly transparent in the visible spectrum. pharmaceutical preparations are not to be re-used. Containers
for human blood and blood components must not be re-used.
Coloured glass is obtained by the addition of small amounts
of metal oxides, chosen according to the desired spectral PRODUCTION
absorbance.
When glass containers for pharmaceutical use
Neutral glass is a borosilicate glass containing significant are manufactured under stressed conditions
amounts of boric oxide, aluminium oxide, alkali metal oxides (e.g. temperature-time profile) and/or are placed in
and/or alkaline earth oxides in the glass network. Due to its contact with particularly aggressive pharmaceutical
composition, neutral glass has a high hydrolytic resistance and preparations, they may undergo delamination, i.e the
a high thermal shock resistance. separation of the inner glass surface into thin layers called
Soda-lime-silica glass is a silica glass containing alkali metal lamellae or flakes. Glass delamination may be the result of a
oxides, mainly sodium oxide, and alkaline earth oxides, chemical attack that occurs according to well-known glass
mainly calcium oxide, in the glass network. Due to its corrosion mechanisms, such as dissolution by hydrolysis and
composition, soda-lime-silica glass has only a moderate ion exchange (leaching) as a function of the pH. The process of
hydrolytic resistance. interaction between the glass surface and the pharmaceutical
The hydrolytic stability of glass containers for pharmaceutical preparation requires incubation time, and flaking may only
use is expressed by the resistance to the release of soluble become visible a number of months after filling.
mineral substances into water under the prescribed conditions Several risk factors are known to increase the propensity
of contact between the inner surface of the container or glass of a glass to delaminate. The chemical composition of the
grains and water. The hydrolytic resistance is evaluated by pharmaceutical preparation, the presence of buffers like
titrating released alkali reacting ions. According to their citrate or phosphate, which are known to corrode glass, and
hydrolytic resistance, glass containers are classified as follows : the ionic strength of the liquid medium may all strongly
– type I glass containers : neutral glass, with a high hydrolytic favour delamination. The manufacturing process of the
resistance due to the chemical composition of the glass container, chemical treatments of the inner surface, and
itself ; terminal sterilisation and processing at the pharmaceutical
filling lines are other important risk factors to be considered.
– type II glass containers : usually of soda-lime-silica glass
It is recommended that the user of the container assesses the
with a high hydrolytic resistance resulting from suitable
compatibility of the glass container and the pharmaceutical
treatment of the inner surface ;
preparation on a case-by-case basis, considering for example
– type III glass containers : usually of soda-lime-silica glass the dosage form, properties of the formulation and glass
with only moderate hydrolytic resistance. quality.
The following italicised statements constitute general The propensity to delamination of glass containers from
recommendations concerning the type of glass container different sources can be assessed and ranked by exposing the
that may be used for different types of pharmaceutical container to accelerated degradation testing, carried out at
preparations. The manufacturer of a pharmaceutical product specified temperatures for a short time and using the solutions
is responsible for ensuring the suitability of the chosen associated with the actual pharmaceutical preparation as
container. extractants. The presence of particles in the extraction
Type I glass containers are suitable for most preparations solution, the occurrence of phase separation on the inner
whether or not for parenteral administration. surface, and the steep increase of silica concentration in the
Type II glass containers are suitable for most acidic and extraction solution are all indicators of a potential propensity
neutral, aqueous preparations whether or not for parenteral for delamination. Accelerated degradation testing can be used
administration. as a predictive tool to select the most appropriate container
for the intended preparation, but the full compatibility of the
Type III glass containers are in general suitable for non-aqueous active substance with the glass leachate can only be assessed
preparations for parenteral administration, for powders for by a stability test under normal conditions of use.
parenteral administration (except for freeze-dried preparations)
and for preparations not for parenteral administration. TESTS
Glass containers with a hydrolytic resistance higher than that Glass containers for pharmaceutical use comply with the
recommended above for a particular type of preparation may relevant test or tests for hydrolytic resistance. When glass
generally also be used. containers have non-glass components, the tests apply only to
The container chosen for a given preparation shall be such the glass part of the container.
that the glass material does not release substances in quantities
To define the quality of glass containers according to the
sufficient to affect the stability of the preparation or to
intended use, one or more of the following tests are necessary.
present a risk of toxicity. In justified cases, further detailed
information may be necessary to assess the impact on chronic Tests for hydrolytic resistance are carried out to define the type
use and for vulnerable patient groups. of glass (I, II or III) and to control its hydrolytic resistance.
Preparations for parenteral administration are normally In addition, containers for aqueous parenteral preparations
presented in colourless glass, but coloured glass may be are tested for arsenic release and coloured glass containers are
used for substances known to be light-sensitive. Colourless tested for spectral transmission.

3.2.1. Glass containers for pharmaceutical use EUROPEAN PHARMACOPOEIA
HYDROLYTIC RESISTANCE Ampoules. Place at least 6 dry ampoules on a flat, horizontal

surface and fill them with water R from a burette, until the
Table 3.2.1.-1. – Types of glass water reaches point A, where the body of the ampoule declines
Type of container Test to be performed to the shoulder (see Figure 3.2.1.-1). Read the capacities
Type I and type II glass containers Test A (surface test)
(expressed to 2 decimal places) and calculate the mean value.
(to distinguish from type III glass This volume, expressed to 1 decimal place, is the filling volume
containers) for the particular ampoule lot. The filling volume may also
Type I glass containers (to distinguish Test B (glass grains test) or test C be determined by weighing.
from type II and type III glass (etching test)
containers)
Type I and type II glass containers Tests A and B, or tests A and C
(if there are doubts whether the
high hydrolytic resistance is due to
the chemical composition or to the
surface treatment)
The test is carried out by titration of the extraction solutions
obtained under the conditions described for tests A, B and C.
Test C is performed if there are uncertainties whether the
container is type I or type II.
EQUIPMENT
– An autoclave or steam steriliser capable of withstanding
a pressure of 2.5 × 105 N/m2 (equivalent to
0.25 MPa = 2.5 bar) or more and capable of carrying
out the heating cycle described under Autoclaving
process. Preferably it is equipped with a constant-pressure
regulator or other suitable means in order to maintain the
temperature at 121 ± 1 °C. The autoclave vessel is equipped
with a heating device, a thermometer integrated in the
autoclave, a pressure gauge, a vent cock (for manually
operated autoclaves only) and a tray of sufficient capacity
to accommodate, above the water level, the number of
containers needed to carry out the test. The autoclave Figure 3.2.1.-1. – Filling volume of ampoules (up to point A)
has the possibility to connect a calibrated resistance
thermometer or a calibrated thermocouple from the Syringes and cartridges. Select 6 syringes or cartridges.
inner chamber to an external measuring device to allow a Close the small opening (mouth of cartridges and needle
temperature measurement independent from the autoclave and/or Luer cone of syringes) using an inert material (e.g. a
system. tip cap) or any other suitable means to prevent water leakage.
Determine the mean brimful volume in accordance with the
The autoclave vessel and all ancillary equipment must be
procedure described under Vials and bottles and multiply it
washed thoroughly with water R before use.
by 0.9. This volume, expressed to 1 decimal place, is the filling
– A calibrated resistance thermometer or calibrated volume for the particular container lot.
thermocouple connected to a suitable temperature
TEST A. HYDROLYTIC RESISTANCE OF THE INNER SURFACES OF
measuring device.
GLASS CONTAINERS (SURFACE TEST)
– Burettes with a suitable capacity. The determination is carried out on unused containers.
– One-mark volumetric flasks, with a capacity of 1000 mL. The volumes of the test liquid necessary are indicated in
– Pipettes and beakers. Table 3.2.1.-2.
– Conical flasks with capacities of 100 mL and 250 mL. Table 3.2.1.-2. – Volume of test liquid and number of titrations
– A water-bath. Filling volume (mL) Volume of test liquid Number of titrations
– Metal foil (e.g. aluminium, stainless steel). for 1 titration (mL)
Flasks and beakers must already have been used for the test Up to 3 25.0 1
or have been filled with water R and kept in an autoclave at
121 °C for at least 1 h before being used. Above 3 and up to 30 50.0 2
DETERMINATION OF THE FILLING VOLUME Above 30 and up to 100 100.0 2

The filling volume is the volume of water to be introduced 100.0 3
Above 100
into the container for the purpose of the test. For vials and
bottles the filling volume is 90 per cent of the brimful capacity. Cleaning. Remove any debris or dust. Shortly before the
For ampoules it is the volume up to the height of the shoulder. test, fill each container to the brim with water R and allow to
Vials and bottles. Select, at random, 6 containers from stand, filled with water, for 20 ± 5 min. Empty the containers,
the sample lot, or 3 if their capacity exceeds 100 mL, and carefully rinse twice with water R and once with water R1 and
remove any debris or dust. Weigh the empty containers with allow to drain.
an accuracy of 0.1 g. Place the containers on a horizontal Closed ampoules are not rinsed before testing. Closed
surface and fill them with water R until about the rim edge, ampoules may be warmed on a water-bath or in an oven at
avoiding overflow and introduction of air bubbles. Adjust the about 40 °C for approximately 2 min before opening to avoid
liquid levels to the brimful line. Weigh the filled containers to underpressure when opening.
obtain the mass of the water expressed to 2 decimal places
for containers having a nominal volume less than or equal to Filling. Fill the containers with water R1 up to the filling
30 mL, and expressed to 1 decimal place for containers having volume.
a nominal volume greater than 30 mL. Calculate the mean Loosely cap each container with an inert material, for example
value of the brimful capacity in millilitres and multiply it with inverted beakers of such a size that the bottoms of the
by 0.9. This volume, expressed to 1 decimal place, is the filling beakers fit snugly down on the rims of the sample. Ampoules
volume for the particular container lot. and vials capped with clean aluminium foil are further

examples. Place syringes and cartridges in a beaker and cover to the load used during the calibration stage. The use of the
the beaker with clean aluminium foil. calibrated thermocouple is no longer necessary provided the
Containers of a volume of 2 mL or less, in which the water is calibration is proved to be valid over a defined time span.
not sufficiently retained during the autoclaving process, may At the end of the cycle, remove the hot samples from the
be closed in a suitable way, e.g. with a stopper or plug of inert autoclave and cool them cautiously to room temperature
material, such as silicone, and fixed using a plunger or a stable within 30 min.
fixing or clamping device. NOTE : depending on the type or size of the autoclave the heat
Place the samples, gathered in groups in glass dishes or in transfer and thus the resulting thermal cycle in the containers
beakers or other suitable holders, on the rack in the autoclave may vary with the total load of the autoclave. It may therefore
containing water R at room temperature. Ensure that they are be necessary to adjust the autoclave load.
held above the level of the water in the autoclave. Method. Carry out the titration within 1 h of removal of the
Autoclaving process containers from the autoclave. Combine the liquids obtained
Reference thermal cycle from the containers and mix. Introduce the prescribed volume
(Table 3.2.1.-2) into a conical flask (test solution). Place the
The autoclave is run in such a way that the temperature same volume of water R1 into a 2nd similar flask as a blank.
in the containers to be tested follows a thermal cycle with Add to each flask 0.05 mL of methyl red solution R for each
the following characteristics : temperature raised from 25 mL of liquid. Titrate the blank with 0.01 M hydrochloric
room temperature to 100 °C within 20-30 min ; temperature acid. Titrate the test solution with the same acid until the
maintained at 100 ± 1 °C for 10 ± 1 min ; temperature in the colour of the resulting solution is the same as that obtained
containers raised from 100 °C to 121 °C within 20-22 min ; for the blank. Subtract the value found for the blank titration
temperature maintained at 121 ± 1 °C for 60 ± 1 min ; from that found for the test solution and express the results
temperature cooled to 100 °C within 40-44 min. in millilitres of 0.01 M hydrochloric acid per 100 mL. Express
Autoclave calibration titration values of less than 1.0 mL to 2 decimal places and
Before being used for the first time, the autoclave and the titration values of more than or equal to 1.0 mL to 1 decimal
temperature measuring system are calibrated to ensure that place.
the autoclave settings are suitable to guarantee that the Limits. The results, or the average of the results if more than
temperature inside the containers is 121 ± 1 °C. 1 titration is performed, is not greater than the values stated
NOTE : significant differences may be observed between the in Table 3.2.1.-3.
temperature measured in the autoclave chamber and inside
the containers. Table 3.2.1.-3. – Limit values in the test for surface hydrolytic
resistance
Take a set of containers of intermediate capacity (10 mL
Maximum volume of 0.01 M HCl per
for instance) and fill them with water R1. Select a sufficient 100 mL of test solution (mL)
number of containers to fill completely the tray within the Filling volume (mL) Types I and II Type III glass
autoclave chamber. Insert the end of the calibrated resistance glass containers containers
thermometer or calibrated thermocouple into one of the Up to 0.5 3.0 30.0
containers through a hole in the closure having approximately
the same diameter as the probe and connect it to the external Above 0.5 and up to 1 2.0 20.0
measuring device. If the container is too small to insert a Above 1 and up to 2 1.8 17.6
thermocouple, place the thermocouple in a similar container
of suitable size filled with water R1. Close the autoclave door Above 2 and up to 3 1.6 16.1
or lid securely and run the autoclave to achieve the target Above 3 and up to 5 1.3 13.2
thermal cycle in the containers. Where a manual autoclave is
run, leave the vent cock open. Heat the autoclave at a regular Above 5 and up to 10 1.0 10.2
rate so that steam issues vigorously from the vent cock after Above 10 and up to 20 0.80 8.1
20-30 min, and maintain a vigorous evolution of steam for
a further 10 min. Above 20 and up to 50 0.60 6.1
Close the vent cock, follow the temperature increase on the Above 50 and up to 100 0.50 4.8
calibrated thermocouple measuring device by comparison
with readings taken from the autoclave thermometer and Above 100 and up to 200 0.40 3.8
adjust the autoclave settings accordingly in order to match the Above 200 and up to 500 0.30 2.9
target thermal cycle. Keep the temperature ramp as smooth
as possible. Above 500 0.20 2.2
Using the calibrated thermocouple measuring device, ensure TEST B. HYDROLYTIC RESISTANCE OF GLASS GRAINS (GLASS
that deviations from the holding temperature of 121 ± 1 °C GRAINS TEST)
are within the tolerance. When cooling down, vent to prevent Check that the articles as received have been annealed to a
the formation of a vacuum. For safety reasons (boiling commercially acceptable quality.
retardation) do not open the autoclave before the water in the
containers has reached a temperature of 95 °C. Remove the The test may be performed on the canes used for the
hot samples from the autoclave and cool cautiously to room manufacture of tubing glass containers or on the containers.
temperature within 30 min. Equipment
Record the autoclave settings used to carry out the thermal – a mortar, pestle (see Figure 3.2.1.-2) and hammer made
cycle and use these settings for routine autoclave runs. of tempered, magnetic steel ;
At regular intervals verify the validation of the calibration. – as an alternative to the mortar, pestle and hammer, a ball
Establish a re-calibration plan based on quality assurance mill can be used ; the ball mill is made of agate, zirconia
criteria, recalibrate as appropriate and keep records. or stainless steel with a volume of 250 mL ; 2 balls with a
Routine autoclave runs diameter of 40 mm or 3 balls with a diameter of 30 mm
are suitable ;
Use the autoclave settings established during the calibration
stage and follow the same thermal cycle described above. – a set of 3 square-mesh sieves of stainless steel, mounted on
Container sets of different capacity can be tested during the frames of the same material and consisting of the following :
same run. Keep the glass load very similar in size and mass (a) sieve no. 710 ;

(b) sieve no. 425 ; sieve (a), the coarsest of the set. Repeat the operation until
(c) sieve no. 300 ; all fragments have been transferred to the sieve. Shake the
set of sieves for a short time by hand and remove the glass
– a mechanical sieve-shaker or a sieving machine may be that remains on sieves (a) and (b). Submit these portions to
used to sieve the grains ; further fracture, repeating the operation until about 10 g of
– a permanent magnet ; glass remains on sieve (a). Reject this portion and the portion
– metal foil (e.g. aluminium, stainless steel); that passes through sieve (c). Reassemble the set of sieves
and shake for 5 min. Transfer to a weighing bottle those glass
– a hot-air oven, capable of maintaining a temperature of grains that pass through sieve (b) and are retained on sieve (c).
140 ± 5 °C ;
– a balance, capable of weighing up to 500 g with an accuracy Where a ball mill is used, place in the ball mill beaker about
of 0.005 g ; 50 g of the pieces 10-30 mm across taken from 1 of the
samples, add the balls and crush thin-walled glass (wall
– a desiccator ; thickness up to 1.5 mm) for up to 2 min and thick-walled
– an ultrasonic bath. glass (wall thickness greater than 1.5 mm) for up to 5 min.
Transfer the grains to sieve (a), sieve for about 30 s and collect
the grains retained on sieve (c). Transfer the glass from
sieves (a) and (b) into the ball mill and crush and sieve again
as indicated above. Combine the grains retained on sieve (c).
Repeat the crushing and sieving procedure with the other
glass sample and thus 2 samples of grains, each of which shall
be in excess of 10 g, are obtained. Spread each sample on a
piece of clean glazed paper and remove any iron particles by
passing the magnet over them. Transfer each sample into a
beaker for cleaning. Add to the grains in each beaker 30 mL
of acetone R and scour the grains by suitable means, such as a
rubber- or plastic-coated glass rod. After scouring the grains,
allow to settle and decant as much acetone as possible. Add
another 30 mL of acetone R, swirl, allow to settle and decant
again, and add 30 mL of acetone R.
Fill the bath of the ultrasonic vessel with water at room
temperature, then place the beaker in the rack and immerse it
until the level of the acetone is at the level of the water ; apply
the ultrasound for 1 min. Swirl the beaker, allow to settle
and decant the acetone as completely as possible, add 30 mL
of acetone R and repeat the ultrasonic cleaning operation. If
a fine turbidity persists, repeat the ultrasonic cleaning and
acetone washing until the solution remains clear. Swirl and
decant the acetone then dry the grains, first by putting the
beaker on a warm plate to remove excess acetone and then by
heating at 140 °C for 20 min in the drying oven. Transfer the
dried grains from each beaker into separate weighing bottles,
insert the stoppers and cool in the desiccator. Weigh 10.00 g
of the cleaned and dried grains into 2 separate conical flasks.
Add 50 mL of water R1 into each by means of a pipette (test
solutions). Pipette 50 mL of water R1 into a 3rd conical flask
as a blank. Distribute the grains evenly over the flat bases
of the flasks by gentle shaking. Close the flasks with neutral
glass dishes or aluminium foil rinsed with water R, or with
inverted beakers so that the inner surface of the beakers fit
snugly down onto the top rims of the flasks. Place all 3 flasks
in the rack in the autoclave containing the water at room
temperature, and ensure that they are held above the level of
the water in the vessel. Carry out the autoclaving procedure in
a similar manner to that described under test A, but maintain
the temperature of 121 ± 1 °C only for 30 ± 1 min. Do not
open the autoclave until it has cooled to 95 °C. Remove the
hot samples from the autoclave and cool the flasks in running
tap water as soon as possible, avoiding thermal shock. To each
of the 3 flasks add 0.05 mL of methyl red solution R. Titrate
the blank solution immediately with 0.02 M hydrochloric
acid then titrate the test solutions with the same acid until
Figure 3.2.1.-2. – Mortar and pestle apparatus for glass grains the colour matches that obtained with the blank solution.
method (dimensions in millimetres) Subtract the titration volume for the blank solution from that
Method. Rinse the containers to be tested with water R and for the test solution.
dry in the oven. Wrap at least 3 of the glass articles in clean NOTE : where necessary to obtain a sharp end-point, the clear
paper and crush to produce 2 samples of about 100 g each, in solution is to be decanted into a separate 250 mL flask. Rinse
pieces not more than 30 mm across. the grains with 3 quantities, each of 15 mL, of water R1 by
Where a mortar, pestle and hammer are used, place in the swirling and add the washings to the main solution. Add
mortar 30-40 g of the pieces 10-30 mm across taken from 0.05 mL of methyl red solution R. Titrate and calculate as
1 of the samples, insert the pestle and strike it heavily, once described below. In this case also add 45 mL of water R1 and
only, with the hammer. Transfer the contents of the mortar to 0.05 mL of methyl red solution R to the blank solution.

Calculate the mean value of the results in millilitres of 0.02 M Reference solutions. Prepare the reference solutions using
hydrochloric acid per gram of the sample and if required its arsenic standard solution (1 ppm As) R. Add 10 mL of
equivalent in alkali extracted, calculated as micrograms of hydrochloric acid R and 5 mL of a 200 g/L solution of
sodium oxide per gram of glass grains. potassium iodide R. Heat on a water-bath at 80 °C for 20 min,
allow to cool and dilute to 100.0 mL with water R. The
1 mL of 0.02 M hydrochloric acid is equivalent to 620 µg of concentration range of the reference solutions is typically
sodium oxide. 0.005-0.015 ppm of As.
Repeat the test if the highest and lowest observed values differ Acid reservoir. Hydrochloric acid R.
by more than 20 per cent.
Reducing reservoir. Sodium tetrahydroborate reducing
Limits. Type I glass containers require not more than 0.1 mL solution R.
of 0.02 M hydrochloric acid per gram of glass, type II and
type III glass containers require not more than 0.85 mL of Use a hydride generation device to introduce the test
0.02 M hydrochloric acid per gram of glass. solution into the cuvette of the spectrometer. Establish and
standardise instrumental operating conditions according to
TEST C. TO DETERMINE WHETHER THE CONTAINERS HAVE BEEN the manufacturer’s instructions, optimise the uptake rate of
SURFACE-TREATED (ETCHING TEST) the peristaltic pump, then connect it to the acid reservoir, the
If there are uncertainties whether a container has been reducing reservoir and the test solution.
surface-treated, and/or to distinguish between type I and
type II glass containers, test C is used in addition to test A. Source : hollow-cathode lamp.
Alternatively, tests A and B may be used. Test C may be Wavelength : 193.7 nm.
carried out either on unused samples or on samples previously
used in test A. Atomisation device : air-acetylene flame.
Vials and bottles. The volumes of test liquid required are Limit : maximum 0.1 ppm of As.
shown in Table 3.2.1.-2. SPECTRAL TRANSMISSION FOR COLOURED GLASS
CONTAINERS
Rinse the containers twice with water R, fill to the brimful
point with a mixture of 1 volume of hydrofluoric acid R and Equipment. A UV-Vis spectrophotometer, equipped with a
9 volumes of hydrochloric acid R and allow to stand for 10 min. photodiode detector or equipped with a photomultiplier tube
Empty the containers and rinse carefully 5 times with water R. coupled with an integrating sphere.
Immediately before the test, rinse once again with water R. Preparation of the specimen. Break the glass container or
Submit the containers thus prepared to the same autoclaving cut it with a circular saw fitted with a wet abrasive wheel,
and determination procedure as described in Test A for surface such as a carborundum or a bonded-diamond wheel. Select
hydrolytic resistance. If the results are considerably higher sections representative of the wall thickness and trim them as
than those obtained from the original surfaces (by about a suitable for mounting in a spectrophotometer. If the specimen
factor of 5 to 10), the samples have been surface-treated. is too small to cover the opening in the specimen holder, mask
Ampoules, cartridges and syringes the uncovered portion with opaque paper or tape, provided
that the length of the specimen is greater than that of the slit.
NOTE : ampoules, cartridges and syringes made from glass Before placing in the holder, wash, dry and wipe the specimen
tubing are not normally subjected to internal surface treatment with a lens tissue. Mount the specimen with the aid of wax,
because their high chemical resistance is dependent upon the or by other convenient means, taking care to avoid leaving
chemical composition of the glass as a material. fingerprints or other marks.
Apply the test method as described above for vials and bottles. Method. Place the specimen in the spectrophotometer with
If the ampoules are not surface-treated, the new values are its cylindrical axis parallel to the slit and in such a way that the
slightly lower than those obtained in previous tests. light beam is perpendicular to the surface of the section and
Distinction between type I and type II glass containers that the losses due to reflection are at a minimum. Measure
the transmission of the specimen with reference to air in the
The results obtained in Test C are compared to those obtained spectral region of 290-450 nm, continuously or at intervals
in Test A. The interpretation of the result is shown in of 20 nm.
Table 3.2.1.-4. Limits. The observed spectral transmission for coloured
glass containers for preparations that are not for parenteral
Table 3.2.1.-4. – Distinction between type I and type II glass administration does not exceed 10 per cent at any wavelength
containers in the range of 290-450 nm, irrespective of the type and
Type I Type II the capacity of the glass container. The observed spectral
transmission in coloured glass containers for parenteral
The values are closely similar The values greatly exceed those found preparations does not exceed the limits given in Table 3.2.1.-5.
to those found in the test for in the test for surface hydrolytic
surface hydrolytic resistance for resistance and are similar to but not
type I glass containers. larger than those for type III glass Table 3.2.1.-5. – Limits of spectral transmission for coloured
containers. glass containers for parenteral preparations
ARSENIC Maximum percentage of spectral transmission
The test applies to glass containers for aqueous parenteral at any wavelength between 290 nm and 450 nm
preparations. Nominal volume (mL) Flame-sealed Containers with
containers closures
Hydride generation atomic absorption spectrometry (2.2.23,
Method I). Up to 1 50 25
Above 1 and up to 2 45 20
Test solution. Use the extraction solution obtained from
containers of types I and II, after autoclaving at 121 °C for 1 h Above 2 and up to 5 40 15
as described under Test A for surface hydrolytic resistance.
Above 5 and up to 10 35 13
Transfer 10.0 mL to a 100 mL volumetric flask. Add 10 mL
of hydrochloric acid R and 5 mL of a 200 g/L solution of Above 10 and up to 20 30 12
potassium iodide R. Heat on a water-bath at 80 °C for 20 min,
Above 20 25 10
allow to cool and dilute to 100.0 mL with water R.

Annex – test for surface hydrolytic resistance Use reference solutions containing 5 per cent V/V of the
– determination by flame spectrometry spectrochemical buffer solution.
METHOD
The surface hydrolytic resistance of glass of types I and II may Carry out preliminary measurements of the potassium oxide
be determined by analysis of the leaching solution by flame and calcium oxide concentrations on one of the extraction
spectrometry. A number of elements that, when present as solutions. If, for one container type, the concentration of
oxides in glass, contribute to the alkalinity of the solution, potassium oxide is less than 0.2 µg/mL and the concentration
are determined and used to express an alkali equivalent. of calcium oxide is less than 0.1 µg/mL, the remaining
The spectrometric method has the advantage of allowing extraction solutions of this container type need not be
the use of a much smaller sample of extract so that it can analysed for these ions. Aspirate the extraction solution
be applied to small individual containers. This enables an from each sample directly into the flame of the atomic
evaluation of the uniformity of the containers in a given batch absorption or atomic emission instrument and determine the
where this is critical. The results of this measurement are not approximate concentrations of sodium oxide (and potassium
equivalent to those of titrimetry and the 2 methods cannot be oxide and calcium oxide, if present) by reference to calibration
considered interchangeable. A correlation between the 2 is graphs produced from the reference solutions of suitable
dependent on the type of glass and the size and shape of the concentration.
container. The titrimetric method is the reference method of
the Pharmacopoeia ; the spectrometric method may be used in FINAL ANALYSIS
justified and authorised cases. If dilution is unnecessary, add to each container a volume
A method suitable for this type of analysis is shown below. of the spectrochemical buffer solution equivalent to 5 per
cent of the filling volume, mix well and determine sodium
The determination is carried out on unused containers. oxide, calcium oxide and potassium oxide, by reference to
The number of containers to be examined is indicated calibration graphs. For the determination of the calcium oxide
in Table 3.2.1.-6. concentration by flame spectrometry, a nitrous oxide/acetylene
Table 3.2.1.-6. – Number of containers to be examined for the flame is used.
spectrometric method If dilution is necessary, determine sodium oxide, calcium oxide
and potassium oxide, if present, following the procedures as
Filling volume (mL) Number of containers Additional containers
to be measured for preliminary
described above. The solutions shall contain 5 per cent V/V
separately measurements of the spectrochemical buffer solution. Concentration values
less than 1.0 µg/mL are expressed to 2 decimal places, values
Up to 2 20 2 greater than or equal to 1.0 µg/mL to 1 decimal place. Correct
Above 2 and up to 5 15 2 the result for the buffer addition and for any dilution.
Above 5 and up to 30 10 2 DETERMINATION
Determine the mean value of the concentration of individual
Above 30 and up to 100 5 1 oxides found in the samples tested, in micrograms of the
Above 100 3 1 oxide per millilitre of the extraction solution, and calculate
the sum of the individual oxides, expressed as micrograms of
Instructions on determination of the filling volume, cleaning sodium oxide per millilitre of the extraction solution, using
of the containers, filling and heating are given above under the following mass conversion factors :
Hydrolytic resistance and Test A.
– 1 µg of potassium oxide corresponds to 0.658 µg of sodium
SOLUTIONS oxide ;
Spectrochemical buffer solution. Dissolve 80 g of caesium – 1 μg of calcium oxide corresponds to 1.105 μg of sodium
chloride R in about 300 mL of water R1, add 10mL of oxide.
6 M hydrochloric acid R, dilute to 1.0 L with water R1 and mix.
Limits. The mean value is not greater than the value given
Stock solutions: in Table 3.2.1.-7.
– sodium oxide, c(Na2O) = 1 mg/mL ;
Table 3.2.1.-7. – Limit values in the test for surface hydrolytic
– potassium oxide, c(K2O) = 1 mg/mL ;
resistance by flame spectrometry, for type I and type II glass
– calcium oxide, c(CaO) = 1 mg/mL. containers
Commercially available stock solutions may also be used. Filling volume (mL) Limit values for the concentration
Standard solutions. Prepare standard solutions by diluting the of oxides, expressed as sodium
stock solutions with water R1 to obtain concentrations suitable oxide (μg/mL)
for establishing the reference solutions in an appropriate Up to 0.5 7.50
manner, e.g. with concentrations of 20 μg/mL of sodium Above 0.5 and up to 1 5.00
oxide, potassium oxide and calcium oxide, respectively.
Commercially available standard solutions may also be used. Above 1 and up to 2 4.50
Reference solutions. Prepare the reference solutions for Above 2 and up to 3 4.10
establishing the calibration graph (set of calibration solutions)
Above 3 and up to 5 3.20
by diluting suitable concentrated standard solutions with
water R1, so that the normal working ranges of the specific Above 5 and up to 10 2.50
elements are covered, taking into account the instrument used
for the measurement. Typical concentration ranges of the
reference solutions are : Above 20 and up to 50 1.50
– for determination by atomic emission spectrometry of Above 50 and up to 100 1.20
sodium oxide and potassium oxide : up to 10 µg/mL ;
– for determination by atomic absorption spectrometry of
sodium oxide and potassium oxide : up to 3 µg/mL ; Above 200 and up to 500 0.75
– for determination by atomic absorption spectrometry of Above 500 0.50
calcium oxide : up to 7 µg/mL.

EUROPEAN PHARMACOPOEIA 3.3.4. Sterile plastic containers for human blood
01/2020:30304 TESTS
Solution S1. Fill the container with 100 mL of a sterile,
pyrogen-free 9 g/L solution of sodium chloride R. Close the
container and heat it in an autoclave so that the contents are
maintained at 110 °C for 30 min.
3.3.4. STERILE PLASTIC CONTAINERS If the container to be examined contains an anticoagulant
FOR HUMAN BLOOD AND solution, first empty it, rinse the container with 250 mL of
water for injections R at 20 ± 1 °C and discard the rinsings.
BLOOD COMPONENTS
Solution S2. Introduce into the container a volume of water
Plastic containers for the collection, storage, processing and for injections R corresponding to the intended volume of
administration of blood and its components are manufactured anticoagulant solution. Close the container and heat it in an
from one or more polymers, if necessary with additives. autoclave so that the contents are maintained at 110 °C for
The composition and the conditions of manufacture of 30 min. After cooling, add sufficient water for injections R to
the containers are registered by the appropriate competent fill the container to its nominal capacity.
authorities in accordance with the relevant national legislation If the container to be examined contains an anticoagulant
and international agreements. solution, first empty it and rinse it as indicated above.
When the composition of the materials of the different parts of Resistance to centrifugation. Introduce into the container a
the containers corresponds to the appropriate specifications, volume of water R, acidified by the addition of 1 mL of dilute
their quality is controlled by the methods indicated in those hydrochloric acid R, sufficient to fill it to its nominal capacity.
specifications (see 3.1. Materials used for the manufacture of Envelop the container with absorbent paper impregnated with
containers and subsections and 3.3. Containers for human a 1 in 5 dilution of bromophenol blue solution R1 or other
blood and blood components, and materials used in their suitable indicator and then dried. Centrifuge at 5000 g for
manufacture ; transfusion sets and materials used in their 10 min. No leakage perceptible on the indicator paper and no
manufacture ; syringes and subsections). permanent distortion occur.
Materials other than those described in the Pharmacopoeia Resistance to stretch. Introduce into the container a
may be used provided that their composition is authorised by volume of water R, acidified by the addition of 1 mL of
the competent authority and that the containers manufactured dilute hydrochloric acid R, sufficient to fill it to its nominal
from them comply with the requirements prescribed in this capacity. Suspend the container by the suspending device at
general chapter. the opposite end from the blood-taking tube and apply along
the axis of this tube an immediate force of 20 N (2.05 kgf).
In normal conditions of use the materials do not release Maintain the traction for 5 s. Repeat the test with the force
monomers, or other substances, in amounts likely to be applied to each of the parts for filling and emptying. No break
harmful nor do they lead to any abnormal modifications of and no deterioration occur.
the blood.
Leakage. Place the container which has been submitted to
The containers may contain anticoagulant solutions, the stretch test between two plates covered with absorbent
depending on their intended use, and are supplied sterile. paper impregnated with a 1 in 5 dilution of bromophenol
Each container is fitted with attachments suitable for the blue solution R1 or other suitable indicator and then dried.
intended use. The container may be in the form of a single unit Progressively apply force to the plates to press the container
or the collecting container may be connected by one or more so that its internal pressure (i.e. the difference between the
tubes to one or more secondary containers to allow separation applied pressure and atmospheric pressure) reaches 67 kPa
of the blood components to be effected within a closed system. within 1 min. Maintain the pressure for 10 min. No signs of
leakage are detectable on the indicator paper or at any point of
The outlets are of a shape and size allowing for adequate attachment (seals, joints, etc.).
connection of the container with the blood-giving equipment. Vapour permeability. For a container containing an
The protective coverings on the blood-taking needle and on anticoagulant solution, fill with a volume of a 9 g/L solution of
the appendages must be such as to ensure the maintenance sodium chloride R equal to the volume of blood for which the
of sterility. They must be easily removable but must be container is intended.
tamper-evident.
The capacity of the containers is related to the nominal For an empty container, fill with the same mixture of
capacity prescribed by the national authorities and to the anticoagulant solution and sodium chloride solution. Close
appropriate volume of anticoagulant solution. The nominal the container, weigh it and store it at 5 ± 1 °C in an atmosphere
capacity is the volume of blood to be collected in the container. with a relative humidity of (50 ± 5) per cent for 21 days. At
The containers are of a shape such that when filled they may the end of this period the loss in mass is not greater than 1 per
be centrifuged. cent.
Emptying under pressure. Fill the container with a volume
The containers are fitted with a suitable device for suspending of water R at 5 ± 1 °C equal to the nominal capacity. Attach
or fixing which does not hinder the collection, storage, a transfusion set without an intravenous cannula to one of
processing or administration of the blood. the connectors. Compress the container so as to maintain
The containers are enclosed in sealed, protective envelopes. throughout the emptying an internal pressure (i.e the
difference between the applied pressure and atmospheric
pressure) of 40 kPa. The container empties in less than 2 min.
CHARACTERS
Speed of filling. Attach the container by means of the
The container is sufficiently transparent to allow adequate blood-taking tube fitted with the needle to a reservoir
visual examination of its contents before and after the taking containing a suitable solution having a viscosity equal to that
of the blood and is sufficiently flexible to offer minimum of blood, such as a 335 g/L solution of sucrose R at 37 °C.
resistance during filling and emptying under normal Maintain the internal pressure of the reservoir (i.e. the
conditions of use. The container contains not more than 5 mL difference between the applied pressure and atmospheric
of air. pressure) at 9.3 kPa with the base of the reservoir and the

3.3.4. Sterile plastic containers for human blood EUROPEAN PHARMACOPOEIA
upper part of the container at the same level. The volume of the tubes to be examined and the control tubes at exactly
liquid which flows into the container in 8 min is not less than 2500 g in the same horizontal centrifuge for 5 min. After
the nominal capacity of the container. centrifuging, measure the absorbances (2.2.25) of the liquids
Resistance to temperature variations. Place the container at 540 nm using the stock buffer solution as compensation
in a suitable chamber having an initial temperature of liquid. Calculate the haemolytic value as a percentage from
20-23 °C. Cool it rapidly in a deep-freeze to − 80 °C and the expression :
maintain it at this temperature for 24 h. Raise the temperature Aexp
to 50 °C and maintain for 12 h. Allow to cool to room ´ 100
temperature. The container complies with the tests for A100
resistance to centrifugation, resistance to stretch, leakage,
vapour permeability emptying under pressure and speed of A100 = absorbance of tube III ;
filling prescribed above. Aexp = absorbance of tube I or II or of the corresponding
Transparency. Fill the empty container with a volume equal control tubes.
to its nominal capacity of the primary opalescent suspension The solution in tube I gives a haemolytic value not greater
(2.2.1) diluted so as to have an absorbance (2.2.25) at 640 nm than 10 per cent and the haemolytic value of the solution in
of 0.37 to 0.43 (dilution factor about 1 in 16). The cloudiness tube II does not differ by more than 10 per cent from that of
of the suspension must be perceptible when viewed through the corresponding control tube.
the bag, as compared with a similar container filled with
water R. Sterility (2.6.1). The containers comply with the test for
sterility. Introduce aseptically into the container 100 mL of
Extractable matter. Tests are carried out by methods designed a sterile 9 g/L solution of sodium chloride and shake the
to simulate as far as possible the conditions of contact between container to ensure that the internal surfaces have been
the container and its contents which occur in conditions of entirely wetted. Filter the contents of the container through a
use. membrane filter and place the membrane in the appropriate
The conditions of contact and the tests to be carried out on culture medium, as prescribed in the test for sterility.
the eluates are prescribed, according to the nature of the
constituent materials, in the particular requirements for each Pyrogens (2.6.8). Solution S1 complies with the test for
type of container. pyrogens. Inject 10 mL of the solution per kilogram of the
rabbit’s mass.
Haemolytic effects in buffered systems
Stock buffer solution. Dissolve 90.0 g of sodium chloride R, PACKAGING
34.6 g of disodium hydrogen phosphate dodecahydrate R and The containers are packed in protective envelopes.
2.43 g of sodium dihydrogen phosphate R in water R and dilute On removal from its protective envelope the container shows
to 1000 mL with the same solvent. no leakage and no growth of micro-organisms. The protective
Buffer solution A0. To 30.0 mL of stock buffer solution add envelope is sufficiently robust to withstand normal handling.
10.0 mL of water R. The protective envelope is sealed in such a manner that it
Buffer solution B0. To 30.0 mL of stock buffer solution add cannot be opened and re-closed without leaving visible traces
20.0 mL of water R. that the seal has been broken.
Buffer solution C0. To 15.0 mL of stock buffer solution add
85.0 mL of water R. LABELLING
Introduce 1.4 mL of solution S2 into each of three centrifuge The labelling complies with the relevant national legislation
tubes. To tube I add 0.1 mL of buffer solution A0, to tube II and international agreements. The label states :
add 0.1 mL of buffer solution B0 and to tube III add 0.1 mL – the name and address of the manufacturer ;
of buffer solution C0. To each tube add 0.02 mL of fresh, – a batch number which enables the history of the container
heparinised human blood, mix well and warm on a water-bath and of the plastic material of which it is manufactured to
at 30 ± 1 °C for 40 min. Use blood collected less than be traced.
3 h previously or blood collected into an anticoagulant
citrate-phosphate-dextrose solution (CPD) less than 24 h A part of the label is reserved for :
previously. – the statement of the blood group, the reference number and
Prepare three solutions containing, respectively : all other information required by national legislation or
international agreements, and an empty space is provided
3.0 mL of buffer solution A0 and 12.0 mL of water R
for the insertion of supplementary labelling.
(solution A1),
4.0 mL of buffer solution B0 and 11.0 mL of water R The label of the protective envelope or the label on the
(solution B1), container, visible through the envelope, states :
4.75 mL of buffer solution B0 and 10.25 mL of water R – the expiry date ;
(solution C1). – that, once withdrawn from its protective envelope, the
To tubes I, II and III add, respectively, 1.5 mL of solution A1, container must be used within 10 days.
1.5 mL of solution B1 and 1.5 mL of solution C1. At the same The ink or other substance used to print the labels or the
time and in the same manner, prepare three other tubes, writing must not diffuse into the plastic material of the
replacing solution S2 by water R. Centrifuge simultaneously container and must remain legible up to the time of use.

EUROPEAN PHARMACOPOEIA 3.3.5. Sterile containers of plasticised PVC for human blood
01/2020:30305 The stock solutions may be stored at 4 °C for up to 2 weeks.

Stock solution (a). Dissolve 20.0 mg of plastic additive 01 CRS
in internal standard solution S4 and dilute to 20.0 mL with
internal standard solution S4.
Stock solution (b). Dissolve 20.0 mg of plastic additive 24 CRS
3.3.5. EMPTY STERILE CONTAINERS in internal standard solution S4 and dilute to 20.0 mL with
OF PLASTICISED POLY(VINYL internal standard solution S4.
CHLORIDE) FOR HUMAN BLOOD Stock solution (c). Dissolve 20.0 mg of plastic additive 25 CRS
AND BLOOD COMPONENTS internal standard solution S4.
Stock solution (d). Dissolve 20.0 mg of plastic additive 26 CRS
This general chapter is published for information.
DEFINITION internal standard solution S4.
Unless otherwise authorised as described in general chapter Stock solution (e). Dissolve 20.0 mg of plastic additive 27 CRS
3.3.4. Sterile plastic containers for human blood and blood in internal standard solution S4 and dilute to 20.0 mL with
components, the nature and composition of the material from internal standard solution S4.
which the containers are made comply with the requirements Reference solutions (a1)-(a5). Dilute stock solution (a) with
in general chapter 3.3.2. Materials based on plasticised internal standard solution S4 to obtain 5 reference solutions
poly(vinyl chloride) for containers for human blood and blood containing 10-40 µg/mL of plastic additive 01 CRS.
components. Reference solutions (b1)-(b5). Dilute stock solution (b) with
internal standard solution S4 to obtain 5 reference solutions
TESTS containing 10-40 µg/mL of plastic additive 24 CRS.
They comply with the tests prescribed in general chapter Reference solutions (c1)-(c5). Dilute stock solution (c) with
3.3.4. Sterile plastic containers for human blood and blood internal standard solution S4 to obtain 5 reference solutions
components and with the following tests. containing 10-40 µg/mL of plastic additive 25 CRS.
Reference solution. Heat water R in a borosilicate-glass flask Reference solutions (d1)-(d5). Dilute stock solution (d) with
in an autoclave at 110 °C for 30 min. internal standard solution S4 to obtain 5 reference solutions
Acidity or alkalinity. To a volume of solution S2 (see 3.3.4) containing 10-40 µg/mL of plastic additive 26 CRS.
corresponding to 4 per cent of the nominal value of the Reference solutions (e1)-(e5). Dilute stock solution (e) with
container add 0.1 mL of phenolphthalein solution R. The internal standard solution S4 to obtain 5 reference solutions
solution remains colourless. Add 0.4 mL of 0.01 M sodium containing 10-40 µg/mL of plastic additive 27 CRS.
hydroxide. The solution is pink. Add 0.8 mL of 0.01 M Column :
hydrochloric acid and 0.1 mL of methyl red solution R. The
solution is orange-red or red. – material : fused silica ;
Absorbance (2.2.25) : maximum 0.30, determined between – size : l = 30 m, Ø = 0.25 mm ;
wavelengths of 230 nm and 250 nm on solution S2 (see 3.3.4) ; – stationary phase : phenyl(5)methyl(95)polysiloxane R (film
maximum 0.10, determined between wavelengths of 251 nm thickness 0.25 µm).
and 360 nm on solution S2. Use the reference solution as the Carrier gas : helium for chromatography R.
compensation liquid.
Flow rate : 1 mL/min.
Reducing substances. Immediately after preparation of
Split ratio : 1:20.
solution S2 (see 3.3.4), transfer to a borosilicate-glass flask a
volume corresponding to 8 per cent of the nominal value of Temperature :
the container. At the same time, prepare a blank using an Time Temperature
equal volume of the freshly prepared reference solution in (min) (°C)
another borosilicate-glass flask. To each solution add 20.0 mL Column 0 - 3.3 100 → 200
of 0.002 M potassium permanganate and 1 mL of dilute sulfuric
acid R. Allow to stand protected from light for 15 min. To 3.3 - 20 200 → 250
each solution add 0.1 g of potassium iodide R. Allow to stand 20 - 22.5 250
protected from light for 5 min and titrate immediately with
0.01 M sodium thiosulfate, using 0.25 mL of starch solution R 22.5 - 23 250 → 270
as indicator. The difference between the 2 titrations is not 23 - 25 270
greater than 2.0 mL.
25 - 25.6 270 → 320
Plastic additives 01, 24, 25, 26 and 27. Gas chromatography
(2.2.28) coupled with mass spectrometry (2.2.43). 25.6 - 30.6 320
Internal standard solution S3 : 1 mg/mL solution of di-n-octyl Injection port 300
phthalate R in tetrahydrofuran for chromatography R.
Internal standard solution S4. 5 μg/mL solution of di-n-octyl Detection : mass spectrometer as described below ; adjust the
phthalate R in anhydrous ethanol R. detector settings so as to comply with the system suitability
criteria :
Test solution. Cut 0.2 g of the material to be examined into
pieces about 0.5 cm in length. Dissolve the pieces in 12.5 mL of – quadrupole mass spectrometer equipped with an electron
internal standard solution S3 using a polytetrafluoroethylene impact ionisation mode (70 eV) ;
magnetic stirring bar. Complete dissolution of the material to – ion source temperature : 230 °C ;
be examined is obtained after stirring for about 20-30 min. – acquisition system : performed on full-scan (m/z = 40-350)
The poly(vinyl chloride) is precipitated as a white powder by and on single-ion monitoring (SIM) modes ;
adding dropwise 37.5 mL of anhydrous ethanol R. Centrifuge,
then dilute 1.0 mL of the supernatant to 50.0 mL with – solvent delay : 2.5 min ;
anhydrous ethanol R. The final concentration of the internal – mass spectrometer parameters for the fragmentometric
standard in the test solution is 5 µg/mL. mode (SIM) set as follows :

3.3.5. Sterile containers of plasticised PVC for human blood EUROPEAN PHARMACOPOEIA
Substance Ion 1 Ion 2 Ion 3 – repeatability : maximum relative standard deviation of

[m/z] [m/z] [m/z] 1.0 per cent for the retention time of the peak due to the
Plastic additive 01 149 167 279 plastic additive, determined on 6 injections of a reference
155 127 299
solution of each plastic additive tested situated in the
Plastic additive 24
middle of the calibration range (e.g. 20 µg/mL) ; maximum
Plastic additive 25 71 213 315 relative standard deviation of 3.0 per cent for the ratio of
305 193 323
the area of the peak due to the plastic additive to that due
Plastic additive 26
to the internal standard, determined on 6 injections of a
Plastic additive 27 261 149 167 reference solution of each plastic additive tested situated in
149 279 167
the middle of the calibration range (e.g. 20 µg/mL).
DnOP (internal
standard) From the calibration curve obtained with the reference
solutions, calculate the percentage content of plastic additives
Injection : 1 µL.
in the material to be examined.
Relative retention with reference to di-n-octyl
Limits :
phthalate (retention time = about 22 min): plastic
additive 01 = about 0.80 ; plastic additive 24 = about 0.95-1.09 ; – plastic additive 01 : maximum 40 per cent ;
plastic additive 27 = about 1.02 ; plastic additive – plastic additive 24 : maximum 45 per cent ;
25 = about 1.14 ; plastic additive 26 = about 1.34. – plastic additive 25 : maximum 45 per cent ;
The specificity of the detection is checked by monitoring 3 – plastic additive 26 : maximum 45 per cent ;
different ions for each substance using a mass spectrometer
– plastic additive 27 : maximum 45 per cent.
in SIM mode. Ion ratios are determined from the peak areas
after the injection of a standard solution. The ratios in the Chlorides (2.4.4): maximum 0.4 ppm, determined with
table below are given for information. solution S2 (see 3.3.4).
Substance Ion 1 Ion 2 Ion 3 Ion ratio Ion ratio
Prepare the standard using a mixture of 1.2 mL of chloride
[m/z] [m/z] [m/z] 2/1 3/1 standard solution (5 ppm Cl) R and 13.8 mL of water R.
(%) (%) Residue on evaporation. Evaporate to dryness 100 mL
Plastic additive 01 149 167 279 50 30 of solution S2 (see 3.3.4) in a borosilicate-glass beaker of
Plastic additive 24 155 127 299 30 13
appropriate capacity, previously heated to 105 °C. Evaporate
to dryness in the same conditions 100 mL of the reference
Plastic additive 25 71 213 315 45 20 solution (blank test). Dry to constant mass at 100-105 °C. The
residue from solution S2 weighs a maximum of 3 mg, taking
into account the blank test.
STORAGE
DnOP (internal 149 279 167 / /
standard) See general chapter 3.3.4. Sterile plastic containers for human
blood and blood components.
System suitability :
– resolution : if plastic additive 27 is tested, minimum 1.5 LABELLING
between the peaks due to the internal standard and plastic See general chapter 3.3.4. Sterile plastic containers for human
additive 27 ; blood and blood components.

EUROPEAN PHARMACOPOEIA 3.3.6. Containers of plasticised PVC with anticoagulant solution
01/2020:30306 TESTS
They comply with the tests prescribed in general chapter
3.3.4. Sterile plastic containers for human blood and blood
components and with the following tests.
Volume of anticoagulant solution. Empty the container,
collecting the anticoagulant solution in a graduated cylinder.
3.3.6. STERILE CONTAINERS The volume does not differ by more than ± 10 per cent from
OF PLASTICISED POLY(VINYL the stated volume.
CHLORIDE) FOR HUMAN BLOOD Absorbance (2.2.25). Measure the absorbance of the
anticoagulant solution removed from the container between
CONTAINING ANTICOAGULANT 250 nm and 350 nm, using as the compensation liquid an
SOLUTION anticoagulant solution of the same composition that has not
been in contact with a plastic material. The absorbance at the
This general chapter is published for information. maximum at 280 nm is not greater than 0.5.
Plastic additives 01, 24, 25, 26 and 27. Gas chromatography
(2.2.28) coupled with mass spectrometry (2.2.43).
DEFINITION Carefully remove the anticoagulant solution by means of
the flexible transfer tube. Using a funnel fitted to the tube,
Sterile plastic containers containing an anticoagulant completely fill the container with water R, leave in contact
solution complying with the monograph Anticoagulant and for 1 min while squeezing the container gently, then empty
preservative solutions for human blood (0209) are used for completely. Repeat the rinsing. The container, emptied and
the collection, storage and administration of blood. Before rinsed in this manner, complies with the test for plastic
filling they comply with the description and characters given additives 01, 24, 25, 26 and 27 prescribed in general chapter
in general chapter 3.3.5. Empty sterile containers of plasticised 3.3.5. Empty sterile containers of plasticised poly(vinyl chloride)
poly(vinyl chloride) for human blood and blood components. for human blood and blood components.
Unless otherwise authorised as described in general chapter STORAGE
3.3.4. Sterile plastic containers for human blood and blood See general chapter 3.3.4. Sterile plastic containers for human
components, the nature and composition of the material from blood and blood components.
which the containers are made comply with the requirements
prescribed in general chapter 3.3.2. Materials based on LABELLING
plasticised poly(vinyl chloride) for containers for human blood See general chapter 3.3.4. Sterile plastic containers for human
and blood components. blood and blood components.

EUROPEAN PHARMACOPOEIA 4. Reagents
04/2013:40000 This information is given only to make it easier to obtain

such reagents and this does not suggest in any way that the
mentioned suppliers are especially recommended or certified by
the European Pharmacopoeia Commission or the Council of
Europe. It is therefore acceptable to use reagents from another
source provided that they comply with the standards of the
4. REAGENTS Pharmacopoeia.
Additional information for reagents that can only be fully
identified by a trademark or whose availability is limited may
be found in the Knowledge database on the EDQM website.

EUROPEAN PHARMACOPOEIA 4.1. Reagents, standard solutions, buffer solutions
07/2018:40100 reagent stocks. The description may also include a CAS

number (Chemical Abstract Service Registry Number)
recognisable by its typical format, for example 9002-93-1.
Some of the reagents included in the list are toxic and are to be
handled in conformity with good quality control laboratory
practice.
4.1. REAGENTS, STANDARD Reagents in aqueous solution are prepared using water R.
SOLUTIONS, BUFFER SOLUTIONS For liquid chromatography, water for chromatography R is
used for the preparation of mobile phases when water, or an
aqueous solution, is one of the components. Where a reagent
Where the name of a substance or a solution is followed by the
solution is described using an expression such as ‘hydrochloric
letter R (the whole in italics), this indicates a reagent included
acid (10 g/L HCl)’, the solution is prepared by an appropriate
in the following list. The specifications given for reagents do
dilution with water R of a more concentrated reagent solution
not necessarily guarantee their quality for use in medicines.
specified in this chapter. Reagent solutions used in the limit
Within the description of each reagent there is a 7-digit tests for barium, calcium and sulfates are prepared using
reference code in italics (for example, 1002501). This number, distilled water R. Where the name of the solvent is not stated,
which will remain unchanged for a given reagent during an aqueous solution is intended.
subsequent revisions of the list, is used for identification The reagents and reagent solutions are to be stored in
purposes by the Secretariat, and users of the Pharmacopoeia well-closed containers. The labelling should comply with the
may also find it useful, for example in the management of relevant national legislation and international agreements.

EUROPEAN PHARMACOPOEIA 4.1.1. Reagents
01/2020:40101 Dilute 30 g of glacial acetic acid R to 100 mL with water R.

Acetic acid, dilute. 1000402.
Content : 115 g/L to 125 g/L of C2H4O2 (Mr 60.1).
Dilute 12 g of glacial acetic acid R to 100 mL with water R.
4.1.1. REAGENTS Acetic acid, dilute R1. 1000403.
Acacia. 1000100. Content : 57.5 g/L to 62.5 g/L (Mr 60.1).
See Acacia (0307). Dilute 6 g of glacial acetic acid R to 100 mL with water R.
Acacia solution. 1000101. Acetic anhydride. C4H6O3. (Mr 102.1). 1000500. [108-24-7].
Dissolve 100 g of acacia R in 1000 mL of water R. Stir with Content : minimum 97.0 per cent m/m of C4H6O3.
a mechanical stirrer for 2 h. Centrifuge at about 2000 g for Clear, colourless liquid.
30 min to obtain a clear solution. bp : 136 °C to 142 °C.
Storage : in polyethylene containers of about 250 mL Assay. Dissolve 2.00 g in 50.0 mL of 1 M sodium hydroxide
capacity at a temperature of 0 °C to − 20 °C. in a ground-glass-stoppered flask and boil under a reflux
Acebutolol hydrochloride. 1148900. [34381-68-5]. condenser for 1 h. Titrate with 1 M hydrochloric acid, using
See Acebutolol hydrochloride (0871). 0.5 mL of phenolphthalein solution R as indicator. Calculate
the number of millilitres of 1 M sodium hydroxide required
Acetal. C6H14O2. (Mr 118.2). 1112300. [105-57-7]. for 1 g (n1). Dissolve 2.00 g in 20 mL of cyclohexane R in
Acetaldehyde diethyl acetal. 1,1-Diethoxyethane. a ground-glass-stoppered flask, cool in ice and add a cold
Clear, colourless, volatile liquid, miscible with water and with mixture of 10 mL of aniline R and 20 mL of cyclohexane R.
ethanol (96 per cent). Boil the mixture under a reflux condenser for 1 h, add 50.0 mL
: about 0.824. of 1 M sodium hydroxide and shake vigorously. Titrate with
1 M hydrochloric acid, using 0.5 mL of phenolphthalein
: about 1.382. solution R as indicator. Calculate the number of millilitres of
bp : about 103 °C. 1 M sodium hydroxide required for 1 g (n2). Calculate the
Acetaldehyde. C2H4O. (Mr 44.1). 1000200. [75-07-0]. percentage of C4H6O3 from the following expression :
Ethanal. 10.2(n1 - n2 )
Clear, colourless flammable liquid, miscible with water and
with ethanol (96 per cent). Acetic anhydride solution R1. 1000501.
: about 0.788. Dissolve 25.0 mL of acetic anhydride R in anhydrous
pyridine R and dilute to 100.0 mL with the same solvent.
: about 1.332.
Storage : protected from light and air.
bp : about 21 °C.
Acetic anhydride - sulfuric acid solution. 1000502.
Acetaldehyde ammonia trimer trihydrate.
C6H15N3,3H2O. (Mr 183.3). 1133500. [58052-80-5]. Carefully mix 5 mL of acetic anhydride R with 5 mL of
2,4,6-Trimethylhexahydro-1,3,5-triazine trihydrate. sulfuric acid R. Add dropwise and with cooling to 50 mL of
anhydrous ethanol R.
Content : minimum 95.0 per cent.
Colourless or white or pale yellow crystals or powder. Prepare immediately before use.
mp : 95 °C to 97 °C. Acetone. 1000600. [67-64-1].
Assay. Dissolve 0.900 g in water R and dilute to 50.0 mL See Acetone (0872).
with the same solvent. Titrate with 1 M hydrochloric acid,
determining the end-point potentiometrically (2.2.20). Acetonitrile. C2H3N. (Mr 41.05). 1000700. [75-05-8]. Methyl
cyanide. Ethanenitrile.
1 mL of 1 M hydrochloric acid is equivalent to 61.08 mg of
C6H15N3,3H2O. Clear, colourless liquid, miscible with water, with acetone and
with methanol.
Acetic acid, anhydrous. C2H4O2. (Mr 60.1). 1000300. : about 0.78.
[64-19-7]. : about 1.344.
Content : minimum 99.6 per cent m/m of C2H4O2. A 100 g/L solution is neutral to litmus paper.
Colourless liquid or white or almost white, shining, fern-like Distillation range (2.2.11). Not less than 95 per cent distils
crystals, miscible with or very soluble in water, in ethanol between 80 °C and 82 °C.
(96 per cent), in glycerol (85 per cent), and in most fatty and
essential oils. Acetonitrile used in spectrophotometry complies with the
following additional test.
: 1.052 to 1.053.
Absorbance (2.2.25) : maximum 0.01 from 255 nm to 420 nm,
bp : 117 °C to 119 °C. determined using water R as compensation liquid.
A 100 g/L solution is strongly acid (2.2.4).
A 5 g/L solution neutralised with dilute ammonia R2 gives Acetonitrile for chromatography. 1000701.
reaction (b) of acetates (2.3.1). See Acetonitrile R.
Freezing point (2.2.18) : minimum 15.8 °C. Acetonitrile used in chromatography complies with the
Water (2.5.12): maximum 0.4 per cent. If the water content following additional tests.
is more than 0.4 per cent it may be adjusted by adding the Absorbance (2.2.25) : maximum 0.01 at 240 nm and higher
calculated amount of acetic anhydride R. wavelengths, determined using water R as compensation
Storage : protected from light. liquid.
Content (2.2.28) : minimum 99.8 per cent.
Acetic acid, glacial. C2H4O2. (Mr 60.1). 1000400. [64-19-7].
See Acetic acid, glacial (0590). Acetonitrile R1. 1000702.
Complies with the requirements prescribed for
Acetic acid. 1000401. acetonitrile R and with the following additional
Content : 290 g/L to 310 g/L of C2H4O2 (Mr 60.1). requirements.

4.1.1. Reagents EUROPEAN PHARMACOPOEIA
Content : minimum 99.9 per cent. Content : minimum 98.0 per cent, calculated by the
Absorbance (2.2.25) : maximum 0.10, determined at 200 nm normalisation procedure.
using water R as the compensation liquid. N-Acetylglucosamine. C8H15NO6. (Mr 221.2). 1133600.
Acetoxyvalerenic acid. C17H24O4. (Mr 292.4). 1165800. [7512-17-6]. 2-(Acetylamino)-2-deoxy-D-glucopyranose.
[81397-67-3]. (2E)-3-[(1RS,4S,7R,7aR)-1-(Acetyloxy)- mp : about 202 °C.
3,7-dimethyl-2,4,5,6,7,7a-hexahydro-1H-inden-4-yl]-2-
methylprop-2-enoic acid. Acetyl-11-keto-β-boswellic acid. C32H48O5. (Mr 512.7).
1167700. [67416-61-9]. 3α-(Acetyloxy)-11-oxours-12-en-24-
Colourless or pale yellow viscous oil. oic acid. (4β)-3α-(Acetyloxy)-11-oxours-12-en-23-oic acid.
Absorbance (2.2.25). A solution in methanol R shows an White or almost white powder, insoluble in water, soluble in
absorption maximum at about 216 nm. acetone, in anhydrous ethanol and in methanol.
Acetylacetamide. C4H7NO2. (Mr 101.1). 1102600. mp : 271 °C to 274 °C.
[5977-14-0]. 3-Oxobutanamide. Acetyl-11-keto-β-boswellic acid used in liquid chromatography
mp : 53 °C to 56 °C. complies with the following additional test.
Assay. Liquid chromatography (2.2.29) as prescribed in the
Acetylacetone. C5H8O2. (Mr 100.1). 1000900. [123-54-6]. monograph on Indian frankincense (2310).
2,4-Pentanedione.
Content : minimum 90 per cent, calculated by the
Colourless or slightly yellow, easily flammable liquid, freely normalisation procedure.
soluble in water, miscible with acetone, with ethanol (96 per
cent) and with glacial acetic acid. N-Acetylneuraminic acid. C11H19NO9. (Mr 309.3). 1001100.
: 1.452 to 1.453. [131-48-6]. O-Sialic acid.
bp : 138 °C to 140 °C. White or almost white acicular crystals, soluble in water and
in methanol, slightly soluble in anhydrous ethanol, practically
Acetylacetone reagent R1. 1000901. insoluble in acetone.
To 100 mL of ammonium acetate solution R add 0.2 mL of : about − 36, determined on a 10 g/L solution.
acetylacetone R. mp : about 186 °C, with decomposition.
Acetylacetone reagent R2. 1000902. N-Acetyltryptophan. C13H14N2O3. (Mr 246.3). 1102800.
Dissolve 0.2 mL of acetylacetone R, 3 mL of glacial acetic [1218-34-4]. 2-Acetylamino-3-(indol-3-yl)propanoic acid.
acid R and 25 g of ammonium acetate R in water R and White or almost white powder or colourless crystals, slightly
dilute to 100 mL with the same solvent. soluble in water. It dissolves in dilute solutions of alkali
N-Acetyl-ε-caprolactam. C8H13NO2. (Mr 155.2). 1102700. hydroxides.
[1888-91-1]. N-Acetylhexane-6-lactam. mp : about 205 °C.
Colourless liquid, miscible with anhydrous ethanol. Assay. Liquid chromatography (2.2.29) as prescribed in the
monograph Tryptophan (1272).
: about 1.100.
Test solution. Dissolve 10.0 mg in a mixture of 10 volumes
: about 1.489.
of acetonitrile R and 90 volumes of water R and dilute to
bp : about 135 °C. 100.0 mL with the same mixture of solvents.
Acetyl chloride. C2H3ClO. (Mr 78.5). 1000800. [75-36-5]. Content : minimum 99.0 per cent, calculated by the
normalisation procedure.
Clear, colourless liquid, flammable, decomposes in contact
with water and with ethanol (96 per cent), miscible with Acetyltyrosine ethyl ester. C13H17NO4,H2O. (Mr 269.3).
ethylene chloride. 1001200. [36546-50-6]. N-Acetyl-L-tyrosine ethyl
: about 1.10. ester monohydrate. Ethyl (S)-2-acetamido-3-(4-
Distillation range (2.2.11). Not less than 95 per cent distils hydroxyphenyl)propionate monohydrate.
between 49 °C and 53 °C. White or almost white, crystalline powder suitable for the
assay of chymotrypsin.
Acetylcholine chloride. C7H16ClNO2. (Mr 181.7). 1001000. : + 21 to + 25, determined on a 10 g/L solution in ethanol
[60-31-1]. (96 per cent) R.
Crystalline powder, very soluble in cold water and in ethanol
: 60 to 68, determined at 278 nm in ethanol (96 per
(96 per cent). It decomposes in hot water and in alkalis.
cent) R.
Storage : at − 20 °C.
Acetyltyrosine ethyl ester, 0.2 M. 1001201.
Acetylene. C2H2. (Mr 26.04). 1199800. [74-86-2]. Ethyne.
Dissolve 0.54 g of acetyltyrosine ethyl ester R in ethanol
Content : minimum 99.0 per cent V/V. (96 per cent) R and dilute to 10.0 mL with the same solvent.
Acetyleugenol. C12H14O3. (Mr 206.2). 1100700. [93-28-7]. Acid blue 83. C45H44N3NaO7S2. (Mr 826). 1012200.
2-Methoxy-4-(2-propenyl)phenylacetate. [6104-59-2].
Yellow coloured, oily liquid, practically insoluble in water, Colour Index No. 42660.
freely soluble in ethanol (96 per cent). Brilliant blue. Coomassie brilliant blue R 250.
: about 1.521. Brown powder insoluble in cold water, slightly soluble in
bp : 281 °C to 282 °C. boiling water and in anhydrous ethanol, soluble in sulfuric
Acetyleugenol used in gas chromatography complies with the acid, glacial acetic acid and in dilute solutions of alkali
following additional test. hydroxides.
Assay. Gas chromatography (2.2.28) as prescribed in the Acid blue 90. C47H48N3NaO7S2. (Mr 854). 1001300.
monograph Clove oil (1091). [6104-58-1].
Test solution. The substance to be examined. Colour Index No. 42655.

Sodium [4-[[4-[(4-ethoxyphenyl)amino]phenyl][[4-(ethyl)(3- Actein. C37H56O11. (Mr 677). 1181500. [18642-44-9].

sulfonatobenzyl)amino]phenyl]methylene]cyclo-hexa-2,5- (23R,24R,25S,26S)-3β-(β-D-Xylopyranosyloxy)-
dien-1-ylidene](ethyl)-(3-sulfonatobenzyl)ammonium. 16β,23:23,26:24,25-triepoxy-26-hydroxy-9,19-cyclolanostan-
A dark brown powder, with a violet sheen and some particles 12β-yl acetate.
having a metallic lustre, soluble in water and in anhydrous
Acteoside. C29H36O15. (Mr 624.6). 1145100.
ethanol.
[61276-17-3]. 2-(3,4-Dihydroxyphenyl)ethyl
: greater than 500, determined at 577 nm in a 0.01 g/L 3-O-(6-deoxy-α-L-mannopyranosyl)-4-O-[(2E)-3-
solution in buffer solution pH 7.0 and calculated with (3,4-dihydroxyphenyl)prop-2-enoyl]-β-D-glucopyranoside.
reference to the dried substance. Verbascoside.
Loss on drying (2.2.32) : maximum 5.0 per cent, determined on Light yellowish powder, freely soluble in water and in
0.500 g by drying in an oven at 105 °C. methanol.
Acid blue 92. C26H16N3Na3O10S3. (Mr 696). 1001400. mp : about 140 °C, with decomposition.
[3861-73-2].
Adamantane. C10H16. (Mr 136.2). 1181600. [281-23-2].
Colour Index No. 13390. Tricyclo[3.3.1.13,7]decane.
Coomassie blue. Anazolene sodium. Trisodium 8-hydroxy-4′-
(phenylamino)azonaphthalene-3,5′,6-trisulfonate. mp : about 270 °C.
Dark blue crystals, soluble in water, in acetone and in ethylene Adenine. 1172800. [73-24-5].
glycol monoethylether, slightly soluble in ethanol (96 per See Adenine (0800).
cent).
Adenosine. C10H13N5O4. (Mr 267.2). 1001600. [58-61-7].
Acid blue 92 solution. 1001401.
6-Amino-9-β-D-ribofuranosyl-9H-purine.
Dissolve 0.5 g of acid blue 92 R in a mixture of 10 mL of
White or almost white, crystalline powder, slightly soluble in
glacial acetic acid R, 45 mL of ethanol (96 per cent) R and
water, practically insoluble in acetone and in ethanol (96 per
45 mL of water R.
cent). It dissolves in dilute solutions of acids.
Acid blue 93. C37H27N3Na2O9S3. (Mr 800). 1134200. mp : about 234 °C.
[28983-56-4].
Colour Index No. 42780. Adipic acid. C6H10O4. (Mr 146.1). 1095600. [124-04-9].
Methyl blue. Poirrier blue. Prisms, freely soluble in methanol, soluble in acetone,
Mixture of triphenylrosaniline di- and trisulfonate and of practically insoluble in light petroleum.
triphenylpararosaniline. mp : about 152 °C.
Dark blue powder. Adrenaline. C9H13NO3. (Mr 183.2). 1155000. [51-43-4].
Colour change : pH 9.4 to pH 14.0. (1R)-1-(3,4-Dihydroxyphenyl)-2-(methylamino)ethanol.
Acid blue 93 solution. 1134201. 4-[(1R)-1-hydroxy-2-(methylamino)ethyl]benzene-1,2-diol.
Dissolve 0.2 g of acid blue 93 R in water R and dilute to White or almost white powder, gradually becoming brown on
100 mL with the same solvent. exposure to light and air, very slightly soluble in water and
in ethanol (96 per cent), insoluble in acetone. It dissolves in
α1-Acid-glycoprotein silica gel for chiral separation. dilute solutions of mineral acids and alkali hydroxides.
1148700. mp : about 215 °C.
A very finely divided silica gel for chromatography consisting
of spherical particles coated with α1-acid glycoprotein. Adrenalone hydrochloride. C9H12ClNO3. (Mr 217.7).
1155100. [62-13-5]. 1-(3,4-Dihydroxyphenyl)-2-
Acrylamide. C3H5NO. (Mr 71.1). 1001500. [79-06-1]. (methylamino)ethanone hydrochloride. 3′,4′-Dihydroxy-2-
Propenamide. (methylamino)acetophenone hydrochloride.
Colourless or white flakes or a white or almost white, Pale yellow crystals, freely soluble in water, soluble in ethanol
crystalline powder, very soluble in water and in methanol, (96 per cent).
freely soluble in anhydrous ethanol. mp : about 244 °C.
mp : about 84 °C.
Aescin. 1001700. [6805-41-0].
30 per cent acrylamide/bisacrylamide (29:1) solution.
1001501. A mixture of related saponins obtained from the seeds of
Aesculus hippocastanum L.
Prepare a solution containing 290 g of acrylamide R and
10 g of methylenebisacrylamide R per litre of water R. Filter. Fine, almost white or slightly reddish or yellowish, amorphous
powder.
30 per cent acrylamide/bisacrylamide (36.5:1) solution. Chromatography. Thin-layer chromatography (2.2.27).
1001502.
Test solution. Dissolve 10 mg of aescin R in ethanol (70 per
Prepare a solution containing 292 g of acrylamide R and 8 g cent V/V) R and dilute to 10 mL with the same solvent.
of methylenebisacrylamide R per litre of water R. Filter.
Plate : TLC silica gel plate R.
Acrylic acid. C3H4O2. (Mr 72.1). 1133700. [79-10-7]. Mobile phase : the upper layer of a mixture of 10 volumes of
Prop-2-enoic acid. Vinylformic acid. glacial acetic acid R, 40 volumes of water R and 50 volumes
Content : minimum 99 per cent. of butanol R.
It is stabilised with 0.02 per cent of hydroquinone monomethyl Application : 20 μL of the test solution as bands of 20 mm by
ether. 3 mm.
Corrosive liquid, miscible with water and ethanol (96 per Development : over a path of 12 cm.
cent). It polymerises readily in the presence of oxygen. Drying : at 100-105 °C.
: about 1.05. Detection : spray with about 10 mL of anisaldehyde solution R
: about 1.421. for a plate 200 mm square and heat again at 100-105 °C.
bp : about 141 °C. Results : the chromatogram shows a principal band with an RF
mp : 12 °C to 15 °C. of about 0.4.

Aflatoxin B1. C17H12O6. (Mr 312.3). 1166000. White to light brownish-yellow powder.

[1162-65-8]. (6aR,9aS)-4-Methoxy-2,3,6a,9a-tetrahydro- Water (2.5.12) : maximum 3.0 per cent, determined on 0.800 g.
cyclopenta[c]furo[3′,2′:4,5]furo[2,3-h][1]benzopyran-1,11-
dione. Albumin, bovine R1. 1183500. [9048-46-8].
White or faint yellow crystals. Bovine serum albumin containing about 96 per cent of protein.
White or light brownish-yellow powder.
Agarose/cross-linked polyacrylamide. 1002200.
Agarose trapped within a cross-linked polyacrylamide Albumin, human. 1133800.
network ; it is used for the separation of globular proteins with Human serum albumin containing not less than 96 per cent
relative molecular masses of 2 × 104 to 35 × 104. of albumin.
Agarose-DEAE for ion-exchange chromatography. Albumin solution, human. 1002400. [9048-46-8].
1002100. [57407-08-6]. See Human albumin solution (0255).
Cross-linked agarose substituted with diethylaminoethyl
groups, presented as beads. Albumin solution, human R1. 1002401.
Dilute human albumin solution R with a 9 g/L solution of
Agarose for chromatography. 1001800. [9012-36-6]. sodium chloride R to a concentration of 1 g/L of protein.
Swollen beads 60-140 µm in diameter presented as a 4 per Adjust the pH to 3.5-4.5 with glacial acetic acid R.
cent suspension in water R.
Alcohol. 1002500. [64-17-5].
Used in size-exclusion chromatography for the separation of See Ethanol (96 per cent) R.
proteins with relative molecular masses of 6 × 104 to 20 × 106
and of polysaccharides with relative molecular masses of Alcohol (x per cent V/V). 1002502.
3 × 103 to 5 × 106. See Ethanol (x per cent V/V) R.
Agarose for chromatography, cross-linked. 1001900. Alcohol, aldehyde-free. 1002501.
[61970-08-9].
Mix 1200 mL of ethanol (96 per cent) R with 5 mL of a
Prepared from agarose by reaction with 2,3-dibromopropanol 400 g/L solution of silver nitrate R and 10 mL of a cooled
in strongly alkaline conditions. 500 g/L solution of potassium hydroxide R. Shake, allow to
It occurs as swollen beads 60-140 μm in diameter and is stand for a few days and filter. Distil the filtrate immediately
presented as a 4 per cent suspension in water R. before use.
Used in size-exclusion chromatography for the separation of Aldehyde dehydrogenase. 1103000.
proteins with relative molecular masses of 6 × 104 to 20 × 106
and of polysaccharides with relative molecular masses of Enzyme obtained from baker’s yeast which oxidises
3 × 103 to 5 × 106. acetaldehyde to acetic acid in the presence of
nicotinamide-adenine dinucleotide, potassium salts
Agarose for chromatography, cross-linked R1. 1001901. and thiols, at pH 8.0.
[65099-79-8].
Aldehyde dehydrogenase solution. 1103001.
Prepared for agarose by reaction with 2,3-dibromopropanol in
strongly alkaline conditions. Dissolve in water R a quantity of aldehyde dehydrogenase R,
equivalent to 70 units and dilute to 10 mL with the same
It occurs as swollen beads 60-140 μm in diameter and is solvent. This solution is stable for 8 h at 4 °C.
presented as a 4 per cent suspension in water R.
Used in size-exclusion chromatography for the separation of Aldrin. C12H8Cl6. (Mr 364.9). 1123100. [309-00-2].
proteins with relative molecular masses of 7 × 104 to 40 × 106 bp : about 145 °C.
and of polysaccharides with relative molecular masses of mp : about 104 °C.
1 × 105 to 2 × 107. A suitable certified reference solution (10 ng/µL in
Agarose for electrophoresis. 1002000. [9012-36-6]. cyclohexane) may be used.
A neutral, linear polysaccharide, the main component of Aleuritic acid. C16H32O5. (Mr 304.4). 1095700. [533-87-9].
which is derived from agar. (9RS,10SR)-9,10,16-Trihydroxyhexadecanoic acid.
White or almost white powder, practically insoluble in cold White or almost white powder, greasy to the touch, soluble
water, very slightly soluble in hot water. in methanol.
Agnuside. C22H26O11. (Mr 466.4). 1162000. mp : about 101 °C.
[11027-63-7]. (1RS,4aSR,5RS,7aRS)-5-Hydroxy- Alizarin S. C14H7NaO7S,H2O. (Mr 360.3). 1002600.
7-[[(4-hydroxybenzoyl)oxy]methyl]-1,4a,5,7a- [130-22-3].
tetrahydrocyclopenta[c]pyran-1-yl β-D-glucopyranoside. Schultz No. 1145.
White or almost white crystals. Colour Index No. 58005.
Air, hydrocarbon-free. 1188700. Sodium 1,2-dihydroxyanthraquinone-3-sulfonate
monohydrate. Sodium 3,4-dihydroxy-9,10-dioxo-9,10-
Complies with the requirements prescribed for the
dihydroanthracene-2-sulfonate monohydrate.
monograph Medicinal air (1238) with the following additional
requirement. Orange-yellow powder, freely soluble in water and in ethanol
(96 per cent).
Hydrocarbons : maximum 5 ppm V/V, calculated as CH4.
Alizarin S solution. 1002601.
Alanine. 1102900. [56-41-7].
A 1 g/L solution of alizarin S R.
See Alanine (0752).
Test for sensitivity. If alizarin S solution is used for the
β-Alanine. 1004500. [107-95-9]. standardisation of 0.05 M barium perchlorate, it shows
See 3-aminopropionic acid R. a colour change from yellow to orange-red when it is
tested according to the standardisation of 0.05 M barium
Albumin, bovine. 1002300. [9048-46-8]. perchlorate.
Bovine serum albumin containing about 96 per cent of protein. Colour change : pH 3.7 (yellow) to pH 5.2 (violet).

Aloe emodin. C15H10O5. (Mr 270.2). 1188800. [481-72-1]. Amido black 10B. C22H14N6Na2O9S2. (Mr 617). 1003100.
1,8-Dihydroxy-3-(hydroxymethyl)anthracene-9,10-dione. [1064-48-8].
1,8-Dihydroxy-3-(hydroxymethyl)anthraquinone. Schultz No. 299.
Colour Index No. 20470.
Alovudine. C10H13FN2O4. (Mr 244.2). 1185400. [25526-93-6]. Disodium 5-amino-4-hydroxy-6-[(4-nitrophenyl)azo]-3-
1-[(2R,4S,5R)-4-Fluoro-5-(hydroxymethyl)tetrahydrofuran- (phenylazo)naphthalene-2,7-disulfonate.
2-yl]-5-methylpyrimidine-2,4(1H,3H)-dione.
Fluorodeoxythymidine. 3′-Deoxy-3′-fluorothymidine. Dark-brown to black powder, sparingly soluble in water,
soluble in ethanol (96 per cent).
Content : minimum 95 per cent.
Colourless crystals. Amido black 10B solution. 1003101.
A 5 g/L solution of amido black 10B R in a mixture of
Aluminium. Al. (Ar 26.98). 1118200. [7429-90-5]. 10 volumes of acetic acid R and 90 volumes of methanol R.
White or almost white, malleable, flexible, bluish metal,
available as bars, sheets, powder, strips or wire. In moist air an Aminoazobenzene. C12H11N3. (Mr 197.2). 1003200.
oxide film forms which protects the metal from corrosion. [60-09-3].
Analytical grade. Colour Index No. 11000.
4-(Phenylazo)aniline.
Aluminium chloride. AlCl3,6H2O. (Mr 241.4). 1002700. Brownish-yellow needles with a bluish tinge, slightly soluble
[7784-13-6]. Aluminium chloride hexahydrate. in water, freely soluble in ethanol (96 per cent).
Content : minimum 98.0 per cent of AlCl3,6H2O. mp : about 128 °C.
White or slightly yellowish, crystalline powder, hygroscopic,
freely soluble in water and in ethanol (96 per cent). 2-Aminobenzoic acid. C7H7NO2. (Mr 137.1). 1003400.
Storage : in an airtight container. [118-92-3]. Anthranilic acid.
A white or pale-yellow, crystalline powder, sparingly soluble
Aluminium chloride reagent. 1002702. in cold water, freely soluble in hot water, in ethanol (96 per
Dissolve 2.0 g of aluminium chloride R in 100 mL of a 5 per cent) and in glycerol. Solutions in ethanol (96 per cent) or in
cent V/V solution of glacial acetic acid R in methanol R. ether and, particularly, in glycerol show a violet fluorescence.
Aluminium chloride solution. 1002701. mp : about 145 °C.
Dissolve 65.0 g of aluminium chloride R in water R and 3-Aminobenzoic acid. C7H7NO2. (Mr 137.1). 1147400.
dilute to 100 mL with the same solvent. Add 0.5 g of [99-05-8].
activated charcoal R, stir for 10 min, filter and add to the White or almost white crystals. An aqueous solution turns
filtrate, with continuous stirring, sufficient of a 10 g/L brown on standing in air.
solution of sodium hydroxide R (about 60 mL) to adjust
mp : about 174 °C.
the pH to about 1.5.
Storage : in an airtight container, protected from light.
Aluminium nitrate. Al(NO3)3,9H2O. (Mr 375.1). 1002800.
[7784-27-2]. Aluminium nitrate nonahydrate. 4-Aminobenzoic acid. C7H7NO2. (Mr 137.1). 1003300.
Crystals, deliquescent, very soluble in water and ethanol [150-13-0].
(96 per cent), very slightly soluble in acetone. White or almost white, crystalline powder, slightly soluble
Storage : in an airtight container. in water, freely soluble in ethanol (96 per cent), practically
insoluble in light petroleum.
Aluminium oxide, anhydrous. 1002900. [1344-28-1]. mp : about 187 °C.
Aluminium oxide, consisting of γ-Al2O3, dehydrated and Chromatography. Thin-layer chromatography (2.2.27) as
activated by heat treatment. prescribed in the monograph Procaine hydrochloride (0050) ;
Particle size : 75 µm to 150 μm. the chromatogram shows only one principal spot.
Storage : protected from light.
Aluminium oxide, basic. 1118300.
A basic grade of anhydrous aluminium oxide R suitable for 4-Aminobenzoic acid solution. 1003301.
column chromatography. Dissolve 1 g of 4-aminobenzoic acid R in a mixture of
pH (2.2.3). Shake 1 g with 10 mL of carbon dioxide-free 18 mL of anhydrous acetic acid R, 20 mL of water R and
water R for 5 min. The pH of the suspension is 9 to 10. 1 mL of phosphoric acid R. Immediately before use, mix
2 volumes of the solution with 3 volumes of acetone R.
Aluminium oxide for chromatography, deactivated.
1188900. N-(4-Aminobenzoyl)- L-glutamic acid. C12H14N2O5.
Aluminium oxide suitably deactivated for the separation and (Mr 266.3). 1141700. [4271-30-1]. ABGA. (2S)-2-[(4-
detection of traces of polar hydrocarbons, with porous layer Aminobenzoyl)amino]pentanedioic acid.
open tubular (PLOT) design. White or almost white, crystalline powder.
Aluminium oxide, neutral. 1118400. mp : about 175 °C, with decomposition.
See Aluminium oxide, hydrated (0311). 4-Aminobutanoic acid. C4H9NO2. (Mr 103.1). 1123200.
[56-12-2]. γ-Aminobutyric acid. GABA.
Aluminium potassium sulfate. 1003000. [7784-24-9].
Leaflets from methanol and ether, needles from water and
See Alum (0006). ethanol (96 per cent). Freely soluble in water, practically
Aluminium test strip. 1199900. insoluble or slightly soluble in other solvents.
Commercially available test strip for the determination of mp : about 202 °C (decreases on rapid heating).
aluminium in aqueous solvents at a level below 5 ppm. Aminobutanol. C4H11NO. (Mr 89.1). 1003500. [5856-63-3].
Americium-243 spiking solution. 1167500. 2-Aminobutanol.
Contains 50 Bq/L 243Am and a 134 mg/L solution of lanthanum Oily liquid, miscible with water, soluble in ethanol (96 per
chloride heptahydrate R in a 103 g/L solution of hydrochloric cent).
acid R. : about 0.94.

: about 1.453. Aminomethylalizarindiacetic acid reagent. 1003901.

bp : about 180 °C. Solution A. Dissolve 0.36 g of cerous nitrate R in water R
and dilute to 50 mL with the same solvent.
Aminochlorobenzophenone. C13H10ClNO. (Mr 231.7).
1003600. [719-59-5]. 2-Amino-5-chlorobenzophenone. Solution B. Suspend 0.7 g of aminomethylalizarindiacetic
acid R in 50 mL of water R. Dissolve with the aid of about
Yellow, crystalline powder, practically insoluble in water, freely
0.25 mL of concentrated ammonia R, add 0.25 mL of glacial
soluble in acetone, soluble in ethanol (96 per cent).
acetic acid R and dilute to 100 mL with water R.
mp : about 97 °C.
Solution C. Dissolve 6 g of sodium acetate R in 50 mL of
Content : minimum 95.0 per cent. water R, add 11.5 mL of glacial acetic acid R and dilute to
Storage : protected from light. 100 mL with water R.
4-Aminofolic acid. C19H20N8O5. (Mr 440.4). 1163700. To 33 mL of acetone R add 6.8 mL of solution C, 1.0 mL of
[54-62-6]. (2S)-2-[[4-[[(2,4-Diaminopteridin-6- solution B and 1.0 mL of solution A and dilute to 50 mL
yl)methyl]amino]benzoyl]amino]pentanedioic acid. with water R.
N-[4-[[(2,4-Diaminopteridin-6-yl)methyl]amino]benzoyl]-L- Test for sensitivity. To 1.0 mL of fluoride standard solution
glutamic acid. Aminopterine. (10 ppm F) R add 19.0 mL of water R and 5.0 mL of the
Yellowish powder. aminomethylalizarindiacetic acid reagent. After 20 min,
mp : about 230 °C. the solution assumes a blue colour.
Storage : use within 5 days.
6-Aminohexanoic acid. C6H13NO2. (Mr 131.2). 1103100.
[60-32-2]. Aminomethylalizarindiacetic acid solution. 1003902.
Colourless crystals, freely soluble in water, sparingly soluble Dissolve 0.192 g of aminomethylalizarindiacetic acid R
in methanol, practically insoluble in anhydrous ethanol. in 6 mL of freshly prepared 1 M sodium hydroxide. Add
mp : about 205 °C. 750 mL of water R, 25 mL of succinate buffer solution
pH 4.6 R and, dropwise, 0.5 M hydrochloric acid until the
Aminohippuric acid. C9H10N2O3. (Mr 194.2). 1003700. colour changes from violet-red to yellow (pH 4.5 to 5). Add
[61-78-9]. (4-Aminobenzamido)acetic acid. 100 mL of acetone R and dilute to 1000 mL with water R.
White or almost white powder, sparingly soluble in water,
soluble in ethanol (96 per cent). 4-Aminomethylbenzoic acid. C8H9NO2. (Mr 151.2).
mp : about 200 °C. 1167800. [56-91-7].
Aminohippuric acid reagent. 1003701. Aminonitrobenzophenone. C13H10N2O3. (Mr 242.2).

1004000. [1775-95-7]. 2-Amino-5-nitrobenzophenone.
Dissolve 3 g of phthalic acid R and 0.3 g of aminohippuric
acid R in ethanol (96 per cent) R and dilute to 100 mL with Yellow, crystalline powder, practically insoluble in water,
the same solvent. soluble in tetrahydrofuran, slightly soluble in methanol.
mp : about 160 °C.
Aminohydroxynaphthalenesulfonic acid. C10H9NO4S.
(Mr 239.3). 1112400. [116-63-2]. 4-Amino-3- : 690 to 720, determined at 233 nm using a 0.01 g/L
hydroxynaphthalene-1-sulfonic acid. solution in methanol R.
White or grey needles, turning pink on exposure to light, 6-Aminopenicillanic acid. C8H12N2O3S. (Mr 216.3). 1162100.
especially when moist, practically insoluble in water and in [551-16-6]. (2S,5R,6R)-6-Amino-3,3-dimethyl-7-oxo-4-thia-
ethanol (96 per cent), soluble in solutions of alkali hydroxides 1-azabicyclo[3.2.0]heptane-2-carboxylic acid.
and in hot solutions of sodium metabisulfite.
Appearance : white or almost white powder.
mp : about 205 °C, with decomposition.
Aminohydroxynaphthalenesulfonic acid solution.
1112401. Aminophenazone. C13H17N3O. (231.3). 1133900. [58-15-1].
4-(Dimethylamino)-1,5-dimethyl-2-phenyl-1,2-dihydro-3H-
Mix 5.0 g of anhydrous sodium sulfite R with pyrazol-3-one.
94.3 g of sodium hydrogensulfite R and 0.7 g of
aminohydroxynaphthalenesulfonic acid R. Dissolve 1.5 g of White or almost white, crystalline powder or colourless
the mixture in water R and dilute to 10.0 mL with the same crystals, soluble in water, freely soluble in ethanol (96 per
solvent. Prepare the solution daily. cent).
mp : about 108 °C.
cis-Aminoindanol. C9H11NO. (Mr 149.2). 1168300.
[126456-43-7]. (1S,2R)-1-Amino-2,3-dihydro-1H-inden-2-ol. 2-Aminophenol. C6H7NO. (Mr 109.1). 1147500. [95-55-6].
(−)-cis-1-Aminoindan-2-ol. Pale yellowish-brown crystals which rapidly become brown,
Content : minimum 98.0 per cent (sum of enantiomers, sparingly soluble in water, soluble in ethanol (96 per cent).
determined by gas chromatography). mp : about 172 °C.
: − 69 to − 59, determined on a 2 g/L solution in Storage : in an airtight container, protected from light.
chloroform R.
mp : 118 °C to 122 °C. 3-Aminophenol. C6H7NO. (Mr 109.1). 1147600. [591-27-5].
Aminomethylalizarindiacetic acid. C19H15NO8,2H2O. Pale yellowish-brown crystals, sparingly soluble in water.
(Mr 421.4). 1003900. [3952-78-1]. 2,2′-[(3,4-dihydroxy- mp : about 122 °C.
anthraquinon-3-yl)methylenenitrilo]diacetic acid dihydrate.
Alizarin complexone dihydrate. 4-Aminophenol. C6H7NO. (Mr 109.1). 1004300. [123-30-8].
Fine, pale brownish-yellow or orange-brown powder, Content : minimum 95 per cent.
practically insoluble in water, soluble in solutions of alkali White or slightly coloured, crystalline powder, becoming
hydroxides. coloured on exposure to air and light, sparingly soluble in
mp : about 185 °C. water, soluble in anhydrous ethanol.
Loss on drying (2.2.32) : maximum 10.0 per cent, determined mp : about 186 °C, with decomposition.
on 1.000 g. Storage : protected from light.

Aminopolyether. C18H36N2O6. (Mr 376.5). 1112500. Ammonia, dilute R2. 1004703.

[23978-09-8]. 4,7,13,16,21,24-hexaoxa-1,10- Content : 33 g/L to 35 g/L of NH3 (Mr 17.03).
diazabicyclo[8,8,8]hexacosane.
Dilute 14 g of concentrated ammonia R to 100 mL with
mp : 70 °C to 73 °C. water R.
3-Aminopropanol. C3H9NO. (Mr 75.1). 1004400. [156-87-6]. Ammonia, dilute R3. 1004704.
3-Aminopropan-1-ol. Propanolamine.
Content : 1.6 g/L to 1.8 g/L of NH3 (Mr 17.03).
Clear, colourless, viscous liquid.
Dilute 0.7 g of concentrated ammonia R to 100 mL with
: about 0.99. water R.
: about 1.461.
Ammonia, dilute R4. 1004706.
mp : about 11 °C.
Content : 8.4 g/L to 8.6 g/L of NH3 (Mr 17.03).
3-Aminopropionic acid. C3H7NO2. (Mr 89.1). 1004500. Dilute 3.5 g of concentrated ammonia R to 100 mL with
[107-95-9]. β-Alanine. water R.
Ammonia, lead-free. 1004705.
White or almost white, crystalline powder, freely soluble in
water, slightly soluble in ethanol (96 per cent), practically Complies with the requirements prescribed for dilute
insoluble in acetone. ammonia R1 with the following additional test : to 20 mL
of lead-free ammonia, add 1 mL of lead-free potassium
mp : about 200 °C, with decomposition. cyanide solution R, dilute to 50 mL with water R and add
0.10 mL of sodium sulfide solution R. The solution is not
Aminopyrazolone. C11H13N3O. (Mr 203.2). 1004600. more intensely coloured than a reference solution prepared
[83-07-8]. 4-Amino-2,3-dimethyl-1-phenylpyrazolin-5-one. without sodium sulfide.
Light-yellow needles or powder, sparingly soluble in water,
freely soluble in ethanol (96 per cent). Ammonia, concentrated R1. 1004800.
mp : about 108 °C. Content : minimum 30.0 per cent m/m of NH3 (Mr 17.03).
A clear, colourless liquid.
Aminopyrazolone solution. 1004601.
: less than 0.892.
A 1 g/L solution of aminopyrazolone R in buffer solution
pH 9.0 R. Assay. Weigh accurately a ground-glass-stoppered flask
containing 50.0 mL of 1 M hydrochloric acid. Introduce 2 mL
3-Aminosalicylic acid. C7H7NO3. (Mr 153.1). 1183600. of concentrated ammonia R1 and weigh again. Titrate the
[570-23-0]. 3-Amino-2-hydroxybenzoic acid. solution with 1 M sodium hydroxide, using 0.5 mL of methyl
red mixed solution R as indicator.
mp : about 240 °C.
1 mL of 1 M hydrochloric acid is equivalent to 17.03 mg of
Slightly soluble in water. NH3.
4-Aminosalicylic acid. C7H7NO3. (Mr 153.1). 1183700. Storage : protected from atmospheric carbon dioxide, at a
[65-49-6]. 4-Amino-2-hydroxybenzoic acid. temperature below 20 °C.
White or almost white, bulky powder, slightly soluble in water, Ammonium acetate. C2H7NO2. (Mr 77.1). 1004900.
soluble in ethanol (96 per cent), in dilute nitric acid and in [631-61-8].
sodium hydroxide. It darkens on exposure to air and light.
Colourless crystals, very deliquescent, very soluble in water
mp : 135 °C to 145 °C. and in ethanol (96 per cent).
Storage : at a temperature not exceeding 30 °C, in an airtight Storage : in an airtight container.
container, protected from light.
Ammonium acetate solution. 1004901.
Ammonia, concentrated. 1004700.
Dissolve 150 g of ammonium acetate R in water R. Add
See Concentrated ammonia solution (0877). 3 mL of glacial acetic acid R and dilute to 1000 mL with
water R.
Ammonia. 1004701.
Storage : use within 1 week.
Content : 170 g/L to 180 g/L of NH3 (Mr 17.03).
Dilute 67 g of concentrated ammonia R to 100 mL with Ammonium and cerium nitrate. (NH4)2Ce(NO3)6.
water R. (Mr 548.2). 1005000. [16774-21-3].
: 0.931 to 0.934. Orange-yellow, crystalline powder, or orange transparent
crystals, soluble in water.
When used in the test for iron, ammonia R complies with
the following additional requirement. Evaporate 5 mL of Ammonium and cerium sulfate. (NH4)4Ce(SO4)4,2H2O.
ammonia to dryness on a water-bath, add 10 mL of water R, (Mr 633). 1005100. [10378-47-9].
2 mL of a 200 g/L solution of citric acid monohydrate R
and 0.1 mL of thioglycollic acid R. Make alkaline by adding Orange-yellow, crystalline powder or crystals, slowly soluble
ammonia R and dilute to 20 mL with water R. No pink in water.
colour develops. (1R)-(–)-Ammonium 10-camphorsulfonate. C10H19NO4S.
Storage : protected from atmospheric carbon dioxide, at a (Mr 249.3). 1103200.
temperature below 20 °C. Content : minimum 97.0 per cent of (1R)-(–)-ammonium
10-camphorsulfonate.
Ammonia, dilute R1. 1004702.
Content : 100 g/L to 104 g/L of NH3 (Mr 17.03). : − 18 ± 2, determined on a 50 g/L solution.
Dilute 41 g of concentrated ammonia R to 100 mL with Ammonium carbamate. CH6N2O2. (Mr 78.1). 1168400.
water R. [1111-78-0]. Carbamic acid ammonium salt.

Ammonium carbonate. 1005200. [506-87-6]. A Storage : in amber flasks at 37 °C for 24 h.
mixture of varying proportions of ammonium hydrogen
carbonate (NH4HCO3, Mr 79.1) and ammonium carbamate Ammonium molybdate reagent R2. 1005708.
(NH2COONH4, Mr 78.1). Dissolve 50 g of ammonium molybdate R in 600 mL of
White or almost white translucent mass, slowly soluble in water R. To 250 mL of cold water R add 150 mL of sulfuric
about 4 parts of water. It is decomposed by boiling water. acid R and cool. Mix the 2 solutions together. Storage : use
Ammonium carbonate liberates not less than 30 per cent m/m within 1 day.
of NH3 (Mr 17.03).
Ammonium molybdate solution. 1005702.
Assay. Dissolve 2.00 g in 25 mL of water R. Slowly add 50.0 mL
of 1 M hydrochloric acid, titrate with 1 M sodium hydroxide, A 100 g/L solution of ammonium molybdate R.
using 0.1 mL of methyl orange solution R as indicator. Ammonium molybdate solution R2. 1005703.
1 mL of 1 M hydrochloric acid is equivalent to 17.03 mg of Dissolve 5.0 g of ammonium molybdate R with heating in
NH3. 30 mL of water R. Cool, adjust the pH to 7.0 with dilute
Storage : at a temperature below 20 °C. ammonia R2 and dilute to 50 mL with water R.
Ammonium carbonate solution. 1005201. Ammonium molybdate solution R3. 1005704.
A 158 g/L solution of ammonium carbonate R. Solution A. Dissolve 5 g of ammonium molybdate R in
Ammonium carbonate solution R1. 1005202. 20 mL of water R with heating.
Dissolve 20 g of ammonium carbonate R in 20 mL of dilute Solution B. Mix 150 mL of ethanol (96 per cent) R with
ammonia R1 and dilute to 100 mL with water R. 150 mL of water R. Add with cooling 100 mL of sulfuric
acid R.
Ammonium chloride. 1005300. [12125-02-9]. Immediately before use add 80 volumes of solution B to
See Ammonium chloride (0007). 20 volumes of solution A.
Ammonium chloride solution. 1005301. Ammonium molybdate solution R4. 1005705.
A 107 g/L solution of ammonium chloride R. Dissolve 1.0 g of ammonium molybdate R in water R
Ammonium citrate. C6H14N2O7. (Mr 226.2). 1103300. and dilute to 40 mL with the same solvent. Add 3 mL of
[3012-65-5]. Diammonium hydrogen citrate. hydrochloric acid R and 5 mL of perchloric acid R and dilute
to 100 mL with acetone R.
White or almost white, crystalline powder or colourless
crystals, freely soluble in water, slightly soluble in ethanol Storage : protected from light ; use within 1 month.
(96 per cent). Ammonium molybdate solution R5. 1005707.
pH (2.2.3) : about 4.3 for a 22.6 g/L solution. Dissolve 1.0 g of ammonium molybdate R in 40.0 mL of
Ammonium dihydrogen phosphate. (NH4)H2PO4. a 15 per cent V/V solution of sulfuric acid R. Prepare the
(Mr 115.0). 1005400. [7722-76-1]. Monobasic ammonium solution daily.
phosphate.
Ammonium molybdate solution R6. 1005709.
crystals, freely soluble in water. Slowly add 10 mL of sulfuric acid R to about 40 mL of
water R. Mix and allow to cool. Dilute to 100 mL with
pH (2.2.3) : about 4.2 for a 23 g/L solution. water R and mix. Add 2.5 g of ammonium molybdate R and
Ammonium formate. CH5NO2. (Mr 63.1). 1112600. 1 g of cerium sulfate R, and shake for 15 min to dissolve.
[540-69-2].
Ammonium nitrate. NH4NO3. (Mr 80.0). 1005800.
Deliquescent crystals or granules, very soluble in water, [6484-52-2].
mp : 119 °C to 121 °C. crystals, hygroscopic, very soluble in water, freely soluble in
Storage : in an airtight container. methanol, soluble in ethanol (96 per cent).
Ammonium hexafluorogermanate(IV). (NH4)2GeF6. Storage : in an airtight container.
(Mr 222.7). 1134000. [16962-47-3].
Ammonium nitrate R1. 1005801.
White or almost white crystals, freely soluble in water.
Complies with the requirements prescribed for ammonium
Ammonium hydrogen carbonate. NH4HCO3. (Mr 79.1). nitrate R with the following additional requirements.
1005500. [1066-33-7]. Acidity. The solution of the substance is slightly acid (2.2.4).
Content : minimum 99 per cent. Chlorides (2.4.4) : maximum 100 ppm, determined on
Ammonium molybdate. (NH4)6Mo7O24,4H2O. (Mr 1236). 0.50 g.
1005700. [12054-85-2]. Sulfates (2.4.13): maximum 150 ppm, determined on 1.0 g.
Colourless or slightly yellow or greenish crystals, soluble in Sulfated ash (2.4.14) : maximum 0.05 per cent, determined
water, practically insoluble in ethanol (96 per cent). on 1.0 g.
Ammonium molybdate reagent. 1005701. Ammonium oxalate. C2H8N2O4,H2O. (Mr 142.1). 1005900.
Mix, in the given order, 1 volume of a 25 g/L solution of [6009-70-7].
ammonium molybdate R, 1 volume of a 100 g/L solution of Colourless crystals, soluble in water.
ascorbic acid R and 1 volume of sulfuric acid R (294.5 g/L
H2SO4). Add 2 volumes of water R. Ammonium oxalate solution. 1005901.
Storage : use within 1 day. A 40 g/L solution of ammonium oxalate R.
Ammonium molybdate reagent R1. 1005706. Ammonium persulfate. (NH4)2S2O8. (Mr 228.2). 1006000.
Mix 10 mL of a 60 g/L solution of disodium arsenate R, [7727-54-0].
50 mL of ammonium molybdate solution R, 90 mL of dilute White or almost white, crystalline powder or granular crystals,
sulfuric acid R and dilute to 200 mL in water R. freely soluble in water.

Ammonium phosphate. (NH4)2HPO4. (Mr 132.1). 1006100. β-Amyrin. C30H50O. (Mr 426.7). 1141800. [559-70-6].
[7783-28-0]. Diammonium hydrogen phosphate. Olean-12-en-3β-ol.
White or almost white crystals or granules, hygroscopic, very White or almost white powder.
soluble in water, practically insoluble in ethanol (96 per cent). mp : 187 °C to 190 °C.
pH (2.2.3) : about 8 for a 200 g/L solution.
Andrographolide. C20H30O5. (Mr 350.4). 1198100.
Storage : in an airtight container. [5508-58-7]. (3E,4S)-3-[2-[(1R,4aS,5R,6R,8aS)-
Ammonium pyrrolidinedithiocarbamate. C5H12N2S2. 6-Hydroxy-5-(hydroxymethyl)-5,8a-dimethyl-2-
(Mr 164.3). 1006200. [5108-96-3]. Ammonium methylenedecahydronaphthalen-1-yl]ethylidene]-4-
1-pyrrolidinyl-dithioformate. hydroxydihydrofuran-2(3H)-one.
White or pale yellow, crystalline powder, sparingly soluble in Anethole. C10H12O. (Mr 148.2). 1006900. [4180-23-8].
water, very slightly soluble in ethanol (96 per cent). 1-Methoxy-4-(propen-1-yl)benzene.
Storage : in a bottle containing a piece of ammonium carbonate White or almost white, crystalline mass up to 20 °C to 21 °C,
in a muslin bag. liquid above 23 °C, practically insoluble in water, freely soluble
Ammonium reineckate. NH4[Cr(NCS)4(NH3)2],H2O. in anhydrous ethanol, soluble in ethyl acetate and in light
(Mr 354.4). 1006300. [13573-16-5]. Ammonium petroleum.
diamine-tetrakis(isothiocyanato)chromate(III) monohydrate. : about 1.56.
Red powder or crystals, sparingly soluble in cold water, soluble bp : about 230 °C.
in hot water and in ethanol (96 per cent). Anethole used in gas chromatography complies with the
Ammonium reineckate solution. 1006301.
Assay. Gas chromatography (2.2.28) as prescribed in the
A 10 g/L solution of ammonium reineckate R. Prepare monograph Anise oil (0804).
immediately before use.
Test solution. The substance to be examined.
Ammonium sulfamate. NH2SO3NH4. (Mr 114.1). 1006400. Content : minimum 99.0 per cent of trans-anethole (retention
[7773-06-0]. time : about 41 min), calculated by the normalisation
White or almost white, crystalline powder or colourless procedure.
crystals, hygroscopic, very soluble in water, slightly soluble in
ethanol (96 per cent). Anhydrous colloidal silica. 1202000. [7631-86-9].
mp : about 130 °C. See Anhydrous colloidal silica (0434).
Storage : in an airtight container. Aniline. C6H7N. (Mr 93.1). 1007100. [62-53-3].
Benzeneamine.
Ammonium sulfate. (NH4)2SO4. (Mr 132.1). 1006500.
[7783-20-2]. Colourless or slightly yellowish liquid, soluble in water,
miscible with ethanol (96 per cent).
Colourless crystals or white or almost white granules, very
soluble in water, practically insoluble in acetone and in ethanol : about 1.02.
(96 per cent). bp : 183 °C to 186 °C.
pH (2.2.3) : 4.5 to 6.0 for a 50 g/L solution in carbon Storage : protected from light.
dioxide-free water R.
Aniline hydrochloride. C6H8ClN. (Mr 129.6). 1147700.
Sulfated ash (2.4.14): maximum 0.1 per cent. [142-04-1]. Benzenamine hydrochloride.
Ammonium sulfide solution. 1123300. Crystals.
Saturate 120 mL of dilute ammonia R1 with hydrogen sulfide R It darkens on exposure to air and light.
and add 80 mL of dilute ammonia R1. Prepare immediately mp : about 198 °C.
before use. Storage : protected from light.
Ammonium thiocyanate. NH4SCN. (Mr 76.1). 1006700. Content : minimum 97.0 per cent.
[1762-95-4]. Anion-exchange resin. 1007200.
Colourless crystals, deliquescent, very soluble in water, soluble Resin in chlorinated form containing quaternary ammonium
in ethanol (96 per cent). groups [CH2N+(CH3)3] attached to a polymer lattice consisting
Storage : in an airtight container. of polystyrene cross-linked with 2 per cent of divinylbenzene.
Ammonium thiocyanate solution. 1006701. It is available as spherical beads.
A 76 g/L solution of ammonium thiocyanate R. Wash the resin with 1 M sodium hydroxide on a sintered-glass
filter (40) (2.1.2) until the washings are free from chloride,
Ammonium vanadate. NH4VO3. (Mr 117.0). 1006800. then wash with water R until the washings are neutral.
[7803-55-6]. Ammonium trioxovanadate(V). Suspend in freshly prepared ammonium-free water R and
White or slightly yellowish, crystalline powder, slightly soluble protect from atmospheric carbon dioxide.
in water, soluble in dilute ammonia R1. Anion-exchange resin R1. 1123400.
Ammonium vanadate solution. 1006801. Resin containing quaternary ammonium groups
Dissolve 1.2 g of ammonium vanadate R in 95 mL of [CH2N+(CH3)3] attached to a lattice consisting of methacrylate.
water R and dilute to 100 mL with sulfuric acid R. Anion-exchange resin R2. 1141900.
Amoxicillin trihydrate. 1103400. Conjugate of homogeneous 10 μm hydrophilic polyether
See Amoxicillin trihydrate (0260). particles, and a quaternary ammonium salt, providing a
matrix suitable for strong anion-exchange chromatography
α-Amylase. 1100800. 1,4-α-D-glucane-glucanohydrolase of proteins.
(EC 3.2.1.1).
White or light brown powder. Anion-exchange resin R3. 1180900.
Resin with quaternary ammonium groups attached to a
α-Amylase solution. 1100801. lattice of ethylvinylbenzene crosslinked with 55 per cent of
A solution of α-amylase R with an activity of 800 FAU/g. divinylbenzene.

Anion-exchange resin for chromatography, strongly basic. Anthracene. C14H10. (Mr 178.2). 1007400. [120-12-7].
1112700. White or almost white, crystalline powder, practically
Resin with quaternary amine groups attached to a lattice of insoluble in water, slightly soluble in chloroform.
latex cross linked with divinylbenzene. mp : about 218 °C.
Anion-exchange resin for chromatography, strongly
Anthrone. C14H10O. (Mr 194.2). 1007500. [90-44-8].
basic R1. 1187400.
9(10H)-Anthracenone.
Non-porous resin agglomerated with a 100 nm alkyl
quaternary ammonium functionalised latex. Pale yellow, crystalline powder.
mp : about 155 °C.
Anion-exchange resin for chromatography, strongly
basic R2. 1203000. Antimony potassium tartrate. C8H4K2O12Sb2,3H2O.
Non-porous resin agglomerated with a 43 nm (Mr 668). 1007600. [28300-74-5]. Dipotassium
quaternary amine functionalised latex, cross-linked di[tartrato(4-)O1,O2,O3,O4]bis[antimonate(III)] trihydrate.
with ethylvinylbenzene/divinylbenzene. White or almost white, granular powder or colourless,
transparent crystals, soluble in water and in glycerol, freely
Anion-exchange resin, strongly basic. 1026600. soluble in boiling water, practically insoluble in ethanol
Gel-type resin in hydroxide form containing quaternary (96 per cent). The aqueous solution is slightly acid.
ammonium groups [CH2N+(CH3)3, type 1] attached to a
polymer lattice consisting of polystyrene cross-linked with Antimony trichloride. SbCl3. (Mr 228.1). 1007700.
8 per cent of divinylbenzene. [10025-91-9].
Brown transparent beads. Colourless crystals or a transparent crystalline mass,
Particle size : 0.2 mm to 1.0 mm. hygroscopic, freely soluble in anhydrous ethanol. Antimony
trichloride is hydrolysed by water.
Moisture content : about 50 per cent.
Storage : in an airtight container, protected from moisture.
Total exchange capacity : minimum 1.2 meq/mL.
Anion-exchange resin, weak. 1146700. Antimony trichloride solution. 1007701.
Resin with diethylaminoethyl groups attached to a lattice Rapidly wash 30 g of antimony trichloride R with two
consisting of poly(methyl methacrylate). quantities, each of 15 mL, of ethanol-free chloroform R,
drain off the washings, and dissolve the washed crystals
Anisaldehyde. C8H8O2. (Mr 136.1). 1007300. [123-11-5]. immediately in 100 mL of ethanol-free chloroform R,
4-Methoxybenzaldehyde. warming slightly.
Oily liquid, very slightly soluble in water, miscible with Storage : over a few grams of anhydrous sodium sulfate R.
ethanol (96 per cent).
bp : about 248 °C. Antithrombin III. 1007800. [90170-80-2].
Anisaldehyde used in gas chromatography complies with the Antithrombin III is purified from human plasma by heparin
following additional test. agarose chromatography and should have a specific activity of
at least 6 IU/mg.
monograph Anise oil (0804). Antithrombin III solution R1. 1007801.
Test solution. The substance to be examined. Reconstitute antithrombin III R as directed by the manu-
Content : minimum 99.0 per cent, calculated by the facturer and dilute with tris(hydroxymethyl)aminomethane
normalisation procedure. sodium chloride buffer solution pH 7.4 R to 1 IU/mL.
Anisaldehyde solution. 1007301. Antithrombin III solution R2. 1007802.
Mix in the following order, 0.5 mL of anisaldehyde R, Reconstitute antithrombin III R as directed by the manu-
10 mL of glacial acetic acid R, 85 mL of methanol R and facturer and dilute with tris(hydroxymethyl)aminomethane
5 mL of sulfuric acid R. sodium chloride buffer solution pH 7.4 R to 0.5 IU/mL.
Anisaldehyde solution R1. 1007302. Antithrombin III solution R3. 1007803.
To 10 mL of anisaldehyde R add 90 mL of ethanol (96 per Reconstitute antithrombin III R as directed by the
cent) R, mix, add 10 mL of sulfuric acid R and mix again. manufacturer and dilute to 0.3 IU/mL with phosphate
Anise ketone. C10H12O2. (Mr 164.2). 1174700. [122-84-9]. buffer solution pH 6.5 R.
1-(4-Methoxyphenyl)propan-2-one. Antithrombin III solution R4. 1007804.
p-Anisidine. C7H9NO. (Mr 123.2). 1103500. [104-94-9]. Reconstitute antithrombin III R as directed by
4-Methoxyaniline. the manufacturer and dilute to 0.1 IU/mL with
White or almost white crystals, sparingly soluble in water, tris(hydroxymethyl)aminomethane-EDTA buffer solution
soluble in anhydrous ethanol. pH 8.4 R.
Content : minimum 97.0 per cent. Antithrombin III solution R5. 1007805.
Caution : skin irritant, sensitiser. Reconstitute antithrombin III R as directed by
Storage : protected from light, at 0 °C to 4 °C. the manufacturer and dilute to 0.125 IU/mL with
On storage, p-anisidine tends to darken as a result of oxidation. tris(hydroxymethyl)aminomethane-EDTA buffer solution
A discoloured reagent can be reduced and decolorised in the pH 8.4 R1.
following way : dissolve 20 g of p-anisidine R in 500 mL of
water R at 75 °C. Add 1 g of sodium sulfite heptahydrate R and Antithrombin III solution R6. 1007806.
10 g of activated charcoal R and stir for 5 min. Filter, cool the Reconstitute antithrombin III R as directed by
filtrate to about 0 °C and allow to stand at this temperature for the manufacturer and dilute to 1.0 IU/mL with
at least 4 h. Filter, wash the crystals with a small quantity of tris(hydroxymethyl)aminomethane-EDTA buffer solution
water R at about 0 °C and dry the crystals in vacuo (2.2.32). pH 8.4 R1.

Apigenin. C15H10O5. (Mr 270.2). 1095800. [520-36-5]. Aromadendrene. C15H24. (Mr 204.4 ). 1139100. [489-39-4].
4′,5,7-Trihydroxyflavone. (1R,2S,4R,8R,11R)-3,3,11-Trimethyl-7-methylenetricyclo-
Light yellowish powder, practically insoluble in water, [6.3.0.02,4]undecane.
sparingly soluble in ethanol (96 per cent). Clear, almost colourless liquid.
mp : about 310 °C, with decomposition. d 420 : about 0.911.
Chromatography. Thin-layer chromatography (2.2.27) as
prescribed in the monograph Roman chamomile flower (0380) : nD20 : about 1.497.
apply 10 μL of a 0.25 g/L solution in methanol R ; the [α]20 : about + 12.
D
chromatogram shows in the upper third a principal zone of
yellowish-green fluorescence. bp : about 263 °C.
Aromadendrene used in gas chromatography complies with
Apigenin 7-glucoside. C21H20O10. (Mr 432.4). 1095900. the following additional test.
[578-74-5]. Apigetrin. 7-(β-D-Glucopyranosyloxy)-5-
hydroxy-2-(4-hydroxyphenyl)-4H-1-benzopyran-4-one. Assay. Gas chromatography (2.2.28) as prescribed in the
monograph on Tea tree oil (1837).
Light yellowish powder, practically insoluble in water,
sparingly soluble in ethanol (96 per cent). Content : minimum 92 per cent, calculated by the
mp : 198 °C to 201 °C.
Chromatography. Thin-layer chromatography (2.2.27) as Arsenazo III. C22H18As2N4O14S2. (Mr 776). 1198200.
prescribed in the monograph Roman chamomile flower (0380) : [1668-00-4]. 3,6-Bis[(2-arsonophenyl)diazenyl]-4,5-
apply 10 μL of a 0.25 g/L solution in methanol R ; the dihydroxynaphthalene-2,7-disulfonic acid.
chromatogram shows in the middle third a principal zone of Brown powder.
yellowish fluorescence.
Apigenin-7-glucoside used in liquid chromatography complies Arsenious trioxide. As2O3. (Mr 197.8). 1008300. [1327-53-3].
with the following additional test. Arsenious anhydride. Diarsenic trioxide.
Assay. Liquid chromatography (2.2.29) as prescribed in the Crystalline powder or a white or almost white mass, slightly
monograph Matricaria flower (0404). soluble in water, soluble in boiling water.
Test solution. Dissolve 10.0 mg in methanol R and dilute to
Ascorbic acid. 1008400. [50-81-7].
Content : minimum 95.0 per cent, calculated by the See Ascorbic acid (0253).
normalisation procedure. Ascorbic acid solution. 1008401.
Aprotinin. 1007900. [9087-70-1]. Dissolve 50 mg in 0.5 mL of water R and dilute to 50 mL
See Aprotinin (0580). with dimethylformamide R.
Arabinose. C5H10O5. (Mr 150.1). 1008000. [87-72-9]. Asiaticoside. C48H78O19. (Mr 959). 1123500.

(3R,4S,5S)-Tetrahydro-2H-pyran-2,3,4,5-tetrol. [16830-15-2]. O-6-Deoxy-α-L-mannopyranosyl-
L-Arabinopyranose. L-(+)-Arabinose. (1→4)-O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl
White or almost white, crystalline powder, freely soluble in 2α,3β,23-trihydroxy-4α-urs-12-en-28-oate.
water. White or almost white powder, hygroscopic, soluble in
: + 103 to + 105, determined on a 50 g/L solution in methanol, slightly soluble in anhydrous ethanol, insoluble in
water R containing about 0.05 per cent of NH3. acetonitrile.
Arachidyl alcohol. C20H42O. (Mr 298.5). 1156300. [629-96-9].
1-Eicosanol. Water (2.5.12) : 6.0 per cent.
mp : about 65 °C. Asiaticoside used in liquid chromatography complies with the
Content : minimum 96 per cent of C20H42O.
Arbutin. C12H16O7. (Mr 272.3). 1008100. [497-76-7]. monograph Centella (1498).
Arbutoside. 4-Hydroxyphenyl-β-D-glucopyranoside. Content : minimum 97.0 per cent, calculated by the
Fine, white or almost white, shiny needles, freely soluble in normalisation procedure.
water, very soluble in hot water, soluble in ethanol (96 per Storage : protected from humidity.
cent).
Chromatography. Thin-layer chromatography (2.2.27) as Asparagine. C4H8N2O3. (Mr 132.12). 1200000. [70-47-3].
prescribed in the monograph Bearberry leaf (1054) ; the
chromatogram shows only one principal spot. Aspartic acid. 1134100. [56-84-8].
See Aspartic acid (0797).
Arginine. 1103600. [74-79-3].
See Arginine (0806). D-Aspartic acid. C4H7NO4. (Mr 133.1). 1200100. [1783-96-6].
Argon. Ar. (Ar 39.95). 1008200. [7440-37-1]. L-Aspartyl- L-phenylalanine. C13H16N2O5. (Mr 280.3).

Content : minimum 99.995 per cent V/V. 1008500. [13433-09-5]. (S)-3-Amino-N-[(S)-1-carboxy-2-
phenylethyl]-succinamic acid.
Carbon monoxide (2.5.25, Method I) : maximum 0.6 ppm V/V ;
after passage of 10 L of argon R at a flow rate of 4 L/h, not White or almost white powder.
more than 0.05 mL of 0.002 M sodium thiosulfate is required mp : about 210 °C, with decomposition.
for the titration.
Astragaloside IV. C41H68O14. (Mr 785). 1178200.
Argon R1. Ar. (Ar 39.95). 1176000. [7440-37-1]. [84687-43-4]. (20R,24S)-20,24-Epoxy-16β,25-dihydroxy-
Content : minimum 99.99990 per cent V/V. 3β-(β-D-xylopyranosyloxy)-9,19-cyclolanostan-6α-yl
β-D-glucopyranoside.
Argon for chromatography. Ar. (Ar 39.95). 1166200.
[7440-37-1]. Atropine sulfate. 1159000. [5908-99-6].
Content : minimum 99.95 per cent V/V. See Atropine sulfate (0068).

Aucubin. C15H22O9. (Mr 346.3 ). 1145200. [479-98-1]. Barium hydroxide. Ba(OH)2,8H2O. (Mr 315.5). 1009400.
[1S,4aR,5S,7aS)-5-Hydroxy-7-(hydroxymethyl)-1,4a,5,7a- [12230-71-6]. Barium dihydroxide.
tetrahydrocyclopenta[c]pyran-1-yl β-D-glucopyranoside. Colourless crystals, soluble in water.
Crystals, soluble in water, in ethanol (96 per cent) and in
Barium hydroxide solution. 1009401.
methanol, practically insoluble in light petroleum.
A 47.3 g/L solution of barium hydroxide R.
[α]D20 : about − 163.
Barium nitrate. Ba(NO3)2. (Mr 261.3). 1163800.
mp : about 181 °C. [10022-31-8].
Azomethine H. C17H12NNaO8S2. (Mr 445.4). 1008700. Crystals or crystalline powder, freely soluble in water, very
[5941-07-1]. Sodium hydrogeno-4-hydroxy-5-(2- slightly soluble in ethanol (96 per cent) and in acetone.
hydroxybenzylideneamino)-2,7-naphthalenedisulfonate. mp : about 590 °C.
Azomethine H solution. 1008701. Barium sulfate. 1009500. [7727-43-7].
Dissolve 0.45 g of azomethine H R and 1 g of ascorbic acid R See Barium sulfate (0010).
with gentle heating in water R and dilute to 100 mL with Benzalacetone. C10H10O. (Mr 146.2). 1168500. [122-57-6].
the same solvent. (3E)-4-phenylbut-3-en-2-one.
Baicalin. C21H18O11. (Mr 446.4). 1179200. [21967-41-9]. White or pale yellow mass.
5,6-Dihydroxy-4-oxo-2-phenyl-4H-1-benzopyran-7-yl-β-D- Content : minimum 98.0 per cent.
glucopyranosiduronic acid. bp : about 261 °C.
Barbaloin. C21H22O9,H2O. (Mr 436.4). 1008800. [1415-73-2]. mp : about 39 °C.
Aloin. 1,8-Dihydroxy-3-hydroxymethyl-10-β-D- Benzaldehyde. C7H6O. (Mr 106.1). 1009600. [100-52-7].
glucopyranosyl-10H-anthracen-9-one. Colourless or slightly yellow liquid, slightly soluble in water,
Yellow to dark-yellow, crystalline powder, or yellow needles, miscible with ethanol (96 per cent).
darkening on exposure to air and light, sparingly soluble in : about 1.05.
water and in ethanol (96 per cent), soluble in acetone, in : about 1.545.
ammonia and in solutions of alkali hydroxides.
Distillation range (2.2.11). Not less than 95 per cent distils
: about 192 at 269 nm, about 226 at 296.5 nm, about between 177 °C and 180 °C.
259 at 354 nm, determined on a solution in methanol R and
calculated with reference to the anhydrous substance. Storage : protected from light.
Chromatography. Thin-layer chromatography (2.2.27) as Benzene. C6H6. (Mr 78.1). 1009800. [71-43-2].
prescribed in the monograph Frangula bark (0025) ; the Clear, colourless, flammable liquid, practically insoluble in
chromatogram shows only one principal spot. water, miscible with ethanol (96 per cent).
Barbital. 1008900. [57-44-3]. bp : about 80 °C.
Where benzene is used to prepare a reference solution,
See Barbital (0170).
for safety reasons, the pure reagent may be replaced by
Barbital sodium. C8H11N2NaO3. (Mr 206.2). 1009000. a commercially available reference material containing a
[144-02-5]. Sodium derivative of 5,5-diethyl-1H,3H,5H- certified amount of benzene.
pyrimidine-2,4,6-trione. Benzene-1,2,4-triol. C6H6O3. (Mr 126.1). 1177500.
Content : minimum 98.0 per cent. [533-73-3]. Hydroxyhydroquinone. Hydroxyquinol.
A white or almost white, crystalline powder or colourless Freely soluble in water, in ethanol (96 per cent) and in ethyl
crystals, freely soluble in water, slightly soluble in ethanol acetate.
(96 per cent). mp : about 140 °C.
Barbituric acid. C4H4N2O3. (Mr 128.1). 1009100. [67-52-7]. Benzethonium chloride. C27H42ClNO2,H2O. (Mr 466.1).
1H,3H,5H-Pyrimidine-2,4,6-trione. 1009900. [121-54-0]. Benzyldimethyl[2-[2-[4-(1,1,3,3-
White or almost white powder, slightly soluble in water, freely tetramethylbutyl)phenoxy]ethoxy]ethyl]ammonium chloride
soluble in boiling water and in dilute acids. monohydrate.
mp : about 253 °C. Fine, white or almost white powder or colourless crystals,
soluble in water and in ethanol (96 per cent).
Barium acetate. C4H6BaO4. (Mr 255.4). 1162700. [543-80-6]. mp : about 163 °C.
Barium diacetate.
White or almost white powder, soluble in water.
20 Benzidine. C12H12N2. (Mr 184.2). 1145300. [92-87-5].
d 20 : 2.47. Biphenyl-4,4′-diamine.
Barium carbonate. BaCO3. (Mr 197.3). 1009200. [513-77-9]. Content : minimum 95 per cent.
White or almost white powder or friable masses, practically White or slightly yellowish or reddish powder, darkening on
insoluble in water. exposure to air and light.
mp : about 120 °C.
Barium chloride. BaCl2,2H2O. (Mr 244.3). 1009300. Storage : protected from light.
[10326-27-9]. Barium dichloride.
Colourless crystals, freely soluble in water, slightly soluble in Benzil. C14H10O2. (Mr 210.2). 1117800. [134-81-6].
ethanol (96 per cent). Diphenylethanedione.
Yellow, crystalline powder, practically insoluble in water,
Barium chloride solution R1. 1009301. soluble in ethanol (96 per cent), ethyl acetate and toluene.
A 61 g/L solution of barium chloride R. mp : 95 °C.
Barium chloride solution R2. 1009302. Benzocaine. C9H11NO2. (Mr 165.2). 1123600. [94-09-7].
A 36.5 g/L solution of barium chloride R. See Benzocaine (0011).

Benzohydrazide. C7H8N2O. (Mr 136.2). 1194400. [613-94-5]. Benzyl cyanide. C8H7N. (Mr 117.2). 1171100. [140-29-4].
Benzoyldiazane. Phenylacetonitrile.
Benzoic acid. 1010100. [65-85-0]. Content : minimum 95.0 per cent.
See Benzoic acid (0066). Clear, colourless or light yellow liquid.
: about 1.523.
Benzoin. C14H12O2. (Mr 212.3). 1010200. [579-44-2]. bp : about 233 °C.
2-Hydroxy-1,2-diphenylethanone.
Slightly yellowish crystals, very slightly soluble in water, freely Benzyl ether. C14H14O. (Mr 198.3). 1140900. [103-50-4].
soluble in acetone, soluble in hot ethanol (96 per cent). Dibenzyl ether.
mp : about 137 °C. Clear, colourless liquid, practically insoluble in water, miscible
with acetone and with anhydrous ethanol.
Benzophenone. C13H10O. (Mr 182.2). 1010300. [119-61-9]. : about 1.043.
Diphenylmethanone.
: about 1.562.
Prismatic crystals, practically insoluble in water, freely soluble
in ethanol (96 per cent). bp : about 296 °C, with decomposition.
mp : about 48 °C. Benzylpenicillin sodium. 1011000. [69-57-8].
1,4-Benzoquinone. C6H4O2. (Mr 108.1). 1118500. See Benzylpenicillin sodium (0114).
[106-51-4]. Cyclohexa-2,5-diene-1,4-dione. 2-Benzylpyridine. C12H11N. (Mr 169.2). 1112900. [101-82-6].
Content : minimum 98.0 per cent. Content : minimum 98.0 per cent.
Benzoylarginine ethyl ester hydrochloride. Yellow liquid.
C15H23ClN4O3. (Mr 342.8). 1010500. [2645-08-1]. mp : 13 °C to 16 °C.
N-Benzoyl-L-arginine ethyl ester hydrochloride. Ethyl
(S)-2-benzamido-5-guanidinovalerate hydrochloride. 4-Benzylpyridine. C12H11N. (Mr 169.2). 1181200.
[2116-65-6].
White or almost white, crystalline powder, very soluble in
water and in anhydrous ethanol. Content : minimum 98.0 per cent.
: − 15 to − 18, determined on a 10 g/L solution. Yellow liquid.
mp : about 129 °C. mp : 72 °C to 78 °C.
: 310 to 340, determined at 227 nm using a 0.01 g/L Benzyltrimethylammonium chloride. C10H16ClN.
solution. (Mr 185.7). 1155700. [56-93-9]. N,N,N-Trimethylphenylme-
thanaminium chloride. N,N,N-Trimethylbenzenemethana-
Benzoyl chloride. C7H5ClO. (Mr 140.6). 1010400. [98-88-4]. minium chloride.
Colourless, lachrymatory liquid, decomposed by water and by White or almost white powder, soluble in water.
: about 1.21.
bp : about 197 °C. Berberine chloride. C20H18ClNO4,2H2O. (Mr 407.8). 1153400.
[5956-60-5]. 9,10-Dimethoxy-5,6-dihydrobenzo[g]-1,3-
N-Benzoyl- L-prolyl- L-phenylalanyl- L-arginine benzodioxolo[5,6-a]quinolizinium chloride.
4-nitroanilide acetate. C35H42N8O8. (Mr 703). 1010600. Yellow crystals, slightly soluble in water, practically insoluble
3-Benzoylpropionic acid. C10H10O3. (Mr 178.2). 1171000. in ethanol (96 per cent).
[2051-95-8]. 4-Oxo-4-phenylbutanoic acid. mp : 204 °C to 206 °C.
mp : about 118 °C. Berberine chloride used in liquid chromatography complies
with the following additional test.
2-Benzoylpyridine. C12H9NO. (Mr 183.2). 1134300. Assay. Liquid chromatography (2.2.29) as prescribed in the
[91-02-1]. Phenyl(pyridin-2-yl)methanone. monograph Goldenseal rhizome (1831).
Colourless crystals, soluble in ethanol (96 per cent). Content : minimum 95 per cent, calculated by the
mp : about 43 °C. normalisation procedure.
Benzyl alcohol. 1010700. [100-51-6]. Bergapten. C12H8O4. (Mr 216.2). 1103700. [484-20-8].
See Benzyl alcohol (0256). 5-Methoxypsoralen.
Colourless crystals, practically insoluble in water, sparingly
Benzyl benzoate. 1010800. [120-51-4].
soluble in ethanol (96 per cent) and slightly soluble in glacial
See Benzyl benzoate (0705). acetic acid.
Chromatography. Thin-layer chromatography (2.2.27) as mp : about 188 °C.
prescribed in the monograph Peru balsam (0754) : apply 20 µL
of a 0.3 per cent V/V solution in ethyl acetate R ; after spraying Betulin. C30H50O2. (Mr 442.7). 1011100. [473-98-3].
and heating, the chromatogram shows a principal band with Lup-20(39)-ene-3β,28-diol.
an RF of about 0.8. White or almost white, crystalline powder.
Benzyl cinnamate. C16H14O2. (Mr 238.3). 1010900. mp : 248 °C to 251 °C.
[103-41-3]. Benzyl 3-phenylprop-2-enoate. Bibenzyl. C14H14. (Mr 182.3). 1011200. [103-29-7].
Colourless or yellowish crystals, practically insoluble in water, 1,2-Diphenylethane.
soluble in ethanol (96 per cent). White or almost white, crystalline powder, practically
mp : about 39 °C. insoluble in water, very soluble in methylene chloride, freely
Chromatography. Thin-layer chromatography (2.2.27) as soluble in acetone, soluble in ethanol (96 per cent).
prescribed in the monograph Peru balsam (0754) : apply mp : 50 °C to 53 °C.
20 µL of a 3 g/L solution in ethyl acetate R ; after spraying and
heating, the chromatogram shows a principal band with an RF Biphenyl. C12H10. (Mr 154.2). 1168600. [92-52-4].
of about 0.6. mp : 68 °C to 70 °C.

(−)-α-Bisabolol. C15H26O. (Mr 222.4). 1128800. [23089-26-1]. Biuret reagent. 1011601.

(2S)-6-Methyl-2-[(1S)-4-methylcyclohex-3-enyl]hept-5-en- Dissolve 1.5 g of copper sulfate pentahydrate R and 6.0 g of
2-ol. Levomenol. sodium potassium tartrate R in 500 mL of water R. Add 300 mL
Colourless, viscous liquid with a slight, characteristic odour, of a carbonate-free 100 g/L solution of sodium hydroxide R,
practically insoluble in water, freely soluble in ethanol (96 per dilute to 1000 mL with the same solution and mix.
cent), in methanol, in toluene, in fatty oils and in essential oils.
20 Blocking solution. 1122400.
d 20 : 0.925 to 0.935.
A 10 per cent V/V solution of acetic acid R.
nD20 : 1.492 to 1.500.
Blue dextran 2000. 1011700. [9049-32-5].
[α]D20 : − 54.5 to − 58.0, determined on a 50 g/L solution in Prepared from dextran having an average relative molecular
ethanol (96 per cent) R. mass of 2 × 106 by introduction of a polycyclic chromophore
(− )-α-Bisabolol used for gas chromatography complies with that colours the substance blue. The degree of substitution is
the following additional test. 0.017.
Assay. Gas chromatography (2.2.28) as prescribed in the It is freeze-dried and dissolves rapidly and completely in water
monograph Matricaria oil (1836). and aqueous saline solutions.
Test solution. A 4 g/L solution in cyclohexane R. Absorbance (2.2.25). A 1 g/L solution in a phosphate buffer
Content : minimum 95.0 per cent, calculated by the solution pH 7.0 R shows an absorption maximum at 280 nm.
normalisation procedure. Boldine. C19H21NO4. (Mr 327.3). 1118800. [476-70-0].
Bisbenzimide. C25H27Cl3N6O,5H2O. (Mr 624). 1103800. 1,10-Dimethoxy-6aα-aporphine-2,9-diol.
[23491-44-3]. 4-[5-[5-(4-Methylpiperazin-1-yl)benzimidazol- White or almost white crystalline powder, very slightly
2-yl]benzimidazol-2-yl]phenol trihydrochloride pentahydrate. soluble in water, soluble in ethanol (96 per cent) and in dilute
solutions of acids.
Bisbenzimide stock solution. 1103801. : about + 127, determined on a 1 g/L solution in
Dissolve 5 mg of bisbenzimide R in water R and dilute to anhydrous ethanol R.
100 mL with the same solvent. mp : about 163 °C.
Storage : in the dark.
Boric acid. 1011800. [10043-35-3].
Bisbenzimide working solution. 1103802. See Boric acid (0001).
Immediately before use, dilute 100 µL of bisbenzimide
stock solution R to 100 mL with phosphate buffered saline Boric acid solution, saturated, cold. 1011801.
pH 7.4 R. To 3 g of boric acid R add 50 mL of water R and shake for
10 min. Place the solution for 2 h in the refrigerator.
Bis(diphenylmethyl) ether. C26H22O. (Mr 350.5). 1203100.
[574-42-5]. Borneol. C10H18O. (Mr 154.3). 1011900. [507-70-0].
[Oxybis(methanetriyl)]tetrakisbenzene. 1,1′,1′′,1′′′-(Oxy- endo-1,7,7-Trimethylbicyclo[2.2.1]heptan-2-ol.
methylidyne)tetrakisbenzene. Colourless crystals, readily sublimes, practically insoluble
in water, freely soluble in ethanol (96 per cent) and in light
Bismuth nitrate pentahydrate. Bi(NO3)3,5H2O. (Mr 485.1). petroleum.
1165600. [10035-06-0].
mp : about 208 °C.
mp : about 30 °C.
Chromatography. Thin-layer chromatography (2.2.27), using
Bismuth subnitrate. 4BiNO3(OH)2,BiO(OH). (Mr 1462). silica gel G R as the coating substance. Apply to the plate
1011500. [1304-85-4]. 10 µL of a 1 g/L solution in toluene R. Develop over a path of
White or almost white powder, practically insoluble in water. 10 cm using chloroform R. Allow the plate to dry in air, spray
with anisaldehyde solution R, using 10 mL for a plate 200 mm
Bismuth subnitrate R1. 1011501. square, and heat at 100-105 °C for 10 min. The chromatogram
obtained shows only one principal spot.
Content : 71.5 per cent to 74.0 per cent of bismuth (Bi),
and 14.5 per cent to 16.5 per cent of nitrate, calculated as Bornyl acetate. C12H20O2. (Mr 196.3). 1012000. [5655-61-8].
nitrogen pentoxide (N2O5). endo-1,7,7-Trimethylbicyclo[2.2.1]hept-2-yl acetate.
Bismuth subnitrate solution. 1011502. Colourless crystals or a colourless liquid, very slightly soluble
in water, soluble in ethanol (96 per cent).
Dissolve 5 g of bismuth subnitrate R1 in a mixture of 8.4 mL
of nitric acid R and 50 mL of water R and dilute to 250 mL mp : about 28 °C.
with water R. Filter if necessary. Chromatography. Thin-layer chromatography (2.2.27), using
Acidity. To 10 mL add 0.05 mL of methyl orange solution R. silica gel G R as the coating substance. Apply to the plate
5.0 mL to 6.25 mL of 1 M sodium hydroxide is required to 10 µL of a 2 g/L solution in toluene R. Develop over a path of
change the colour of the indicator. 10 cm using chloroform R. Allow the plate to dry in air, spray
with anisaldehyde solution R, using 10 mL for a plate 200 mm
Bis-tris propane. C11H26N2O6. (Mr 282.3). 1185500. square, and heat at 100-105 °C for 10 min. The chromatogram
[64431-96-5]. 2,2′-(Propane-1,3-diyldiimino)bis[2- obtained shows only one principal spot.
(hydroxymethyl)-1,3-propanediol.
Boron trichloride. BCl3. (Mr 117.2). 1112000. [10294-34-5].
Colourless gas. Reacts violently with water. Available as
Biuret. C2H5N3O2. (Mr 103.1). 1011600. [108-19-0]. solutions in suitable solvents (2-chloroethanol, methylene
White or almost white crystals, hygroscopic, soluble in water, chloride, hexane, heptane, methanol).
sparingly soluble in ethanol (96 per cent). : about 1.420.
mp : 188 °C to 190 °C, with decomposition. bp : about 12.6 °C.
Storage : in an airtight container. Caution : toxic and corrosive.

Boron trichloride-methanol solution. 1112001. Bromocresol purple. C21H16Br2O5S. (Mr 540.2). 1012700.

A 12 per cent m/m solution of boron trichloride R in [115-40-2]. 3′,3″-Dibromo-o-cresolsulfonphthalein.
methanol R. 4,4′-(3H-2,1-Benzoxathiol-3-ylidene)bis(2-bromo-6-
methylphenol)-S,S-dioxide.
Storage : protected from light at − 20 °C, preferably in
sealed tubes. Pinkish powder, practically insoluble in water, soluble
in ethanol (96 per cent) and in dilute solutions of alkali
Boron trifluoride. BF3. (Mr 67.8). 1012100. [7637-07-2]. hydroxides.
Colourless gas. Bromocresol purple solution. 1012701.
Dissolve 50 mg of bromocresol purple R in 0.92 mL of 0.1 M
Boron trifluoride-methanol solution. 1012101.
sodium hydroxide and 20 mL of ethanol (96 per cent) R and
A 140 g/L solution of boron trifluoride R in methanol R. dilute to 100 mL with water R.
Brilliant blue. 1012200. [6104-59-2]. Test for sensitivity. To 0.2 mL of the bromocresol purple
solution add 100 mL of carbon dioxide-free water R and
See acid blue 83 R. 0.05 mL of 0.02 M sodium hydroxide. The solution is
bluish-violet. Not more than 0.2 mL of 0.02 M hydrochloric
Bromelains. 1012300. [37189-34-7]. acid is required to change the colour to yellow.
Concentrate of proteolytic enzymes derived from Ananas Colour change : pH 5.2 (yellow) to pH 6.8 (bluish-violet).
comosus Merr.
Dull-yellow powder. 5-Bromo-2′-deoxyuridine. C9H11BrN2O5. (Mr 307.1).
1012500. [59-14-3]. 5-Bromo-1-(2-deoxy-β-d-erythro-
Activity. 1 g liberates about 1.2 g of amino-nitrogen from a pentofuranosyl)-1H,3H-pyrimidine-2,4-dione.
solution of gelatin R in 20 min at 45 °C and pH 4.5.
mp : about 194 °C.
Bromelains solution. 1012301. Chromatography. Thin-layer chromatography (2.2.27) as
A 10 g/L solution of bromelains R in a mixture of 1 volume prescribed in the monograph Idoxuridine (0669) : apply 5 µL
of phosphate buffer solution pH 5.5 R and 9 volumes of a of a 0.25 g/L solution ; the chromatogram shows only one
9 g/L solution of sodium chloride R. principal spot.
Bromomethoxynaphthalene. C11H9BrO. (Mr 237.1).
Bromine. Br2. (Mr 159.8). 1012400. [7726-95-6].
1159100. [5111-65-9]. 2-Bromo-6-methoxynaphthalene.
Brownish-red fuming liquid, slightly soluble in water, soluble mp : about 109 °C.
in ethanol (96 per cent).
: about 3.1. Bromophenol blue. C19H10Br4O5S. (Mr 670). 1012800.
[115-39-9]. 3′,3″,5′,5″-Tetrabromophenolsulfonphthalein.
Bromine solution. 1012401. 4,4′-(3H-2,1-Benzoxathiol-3-ylidene)bis(2,6-dibromophenol)
Dissolve 30 g of bromine R and 30 g of potassium bromide R S,S-dioxide.
in water R and dilute to 100 mL with the same solvent. Light orange-yellow powder, very slightly soluble in water,
slightly soluble in ethanol (96 per cent), freely soluble in
Bromine water. 1012402. solutions of alkali hydroxides.
Shake 3 mL of bromine R with 100 mL of water R to Bromophenol blue solution. 1012801.
saturation.
Dissolve 0.1 g of bromophenol blue R in 1.5 mL of 0.1 M
Storage : over an excess of bromine R, protected from light. sodium hydroxide and 20 mL of ethanol (96 per cent) R and
dilute to 100 mL with water R.
Bromine water R1. 1012403.
Test for sensitivity. To 0.05 mL of the bromophenol blue
Shake 0.5 mL of bromine R with 100 mL of water R. solution add 20 mL of carbon dioxide-free water R and
Storage : protected from light ; use within 1 week. 0.05 mL of 0.1 M hydrochloric acid. The solution is yellow.
Not more than 0.1 mL of 0.1 M sodium hydroxide is
Bromocresol green. C21H14Br4O5S. (Mr 698). 1012600. required to change the colour to bluish-violet.
[76-60-8]. 3′,3″,5′,5″-Tetrabromo-m-cresol-sulfonphthalein. Colour change : pH 2.8 (yellow) to pH 4.4 (bluish-violet).
4,4′-(3H-2,1-Benzoxathiol-3-ylidene)bis(2,6-dibromo-3-
methylphenol)-S,S-dioxide. Bromophenol blue solution R1. 1012802.
Brownish-white powder, slightly soluble in water, soluble Dissolve 50 mg of bromophenol blue R with gentle heating
in ethanol (96 per cent) and in dilute solutions of alkali in 3.73 mL of 0.02 M sodium hydroxide and dilute to
hydroxides. 100 mL with water R.
Bromocresol green-methyl red solution. 1012602. Bromophenol blue solution R2. 1012803.
Dissolve 0.15 g of bromocresol green R and 0.1 g of methyl Dissolve with heating 0.2 g of bromophenol blue R in 3 mL
red R in 180 mL of anhydrous ethanol R and dilute to of 0.1 M sodium hydroxide and 10 mL of ethanol (96 per
200 mL with water R. cent) R. After solution is effected, allow to cool and dilute
to 100 mL with ethanol (96 per cent) R.
Bromocresol green solution. 1012601.
Bromophos. C8H8BrCl2O3PS. (Mr 366.0). 1123700.
Dissolve 50 mg of bromocresol green R in 0.72 mL of 0.1 M [2104-96-3].
sodium hydroxide and 20 mL of ethanol (96 per cent) R and
dilute to 100 mL with water R. A suitable certified reference solution (10 ng/µL in iso-octane)
may be used.
Test for sensitivity. To 0.2 mL of the bromocresol green
solution add 100 mL of carbon dioxide-free water R. Bromophos-ethyl. C10H12BrCl2O3PS. (Mr 394.0). 1123800.
The solution is blue. Not more than 0.2 mL of 0.02 M [4824-78-6].
hydrochloric acid is required to change the colour to green. A suitable certified reference solution (10 ng/µL in iso-octane)
Colour change : pH 3.6 (yellow) to pH 5.2 (blue). may be used.

Bromothymol blue. C27H28Br2O5S. (Mr 624). 1012900. : about 0.81.

[76-59-5]. 3′,3″-Dibromothymolsulfonphthalein. Distillation range (2.2.11). Not less than 95 per cent distils
4,4′-(3H-2,1-Benzoxathiol-3-ylidene)bis(2-bromo-6- between 99 °C and 100 °C.
isopropyl-3-methylphenol) S,S-dioxide. Assay. Gas chromatography (2.2.28) as prescribed in the
Reddish-pink or brownish powder, practically insoluble in monograph Isopropyl alcohol (0970).
water, soluble in ethanol (96 per cent) and in dilute solutions
of alkali hydroxides. Butyl acetate. C6H12O2. (Mr 116.2). 1013400. [123-86-4].
Clear, colourless liquid, flammable, slightly soluble in water,
Bromothymol blue solution R1. 1012901. miscible with ethanol (96 per cent).
Dissolve 50 mg of bromothymol blue R in a mixture of 4 mL : about 0.88.
of 0.02 M sodium hydroxide and 20 mL of ethanol (96 per : about 1.395.
cent) R and dilute to 100 mL with water R.
Test for sensitivity. To 0.3 mL of bromothymol blue between 123 °C and 126 °C.
solution R1 add 100 mL of carbon dioxide-free water R. The
solution is yellow. Not more than 0.1 mL of 0.02 M sodium Butyl acetate R1. 1013401.
hydroxide is required to change the colour to blue. Content : minimum 99.5 per cent, determined by gas
Colour change : pH 5.8 (yellow) to pH 7.4 (blue). chromatography.
Clear, colourless liquid, flammable, slightly soluble in
Bromothymol blue solution R2. 1012902.
water, miscible with ethanol (96 per cent).
A 10 g/L solution of bromothymol blue R in : about 0.883.
dimethylformamide R.
: about 1.395.
Bromothymol blue solution R3. 1012903. Butanol : maximum 0.2 per cent, determined by gas
Warm 0.1 g of bromothymol blue R with 3.2 mL of 0.05 M chromatography.
sodium hydroxide and 5 mL of ethanol (90 per cent V/V) R. n-Butyl formate : maximum 0.1 per cent, determined by
After solution is effected, dilute to 250 mL with ethanol gas chromatography.
(90 per cent V/V) R. n-Butyl propionate : maximum 0.1 per cent, determined by
Bromothymol blue solution R4. 1012904. gas chromatography.
Dissolve 100 mg of bromothymol blue R in a mixture of Water : maximum 0.1 per cent.
equal volumes of ethanol (96 per cent) R and water R and Butylamine. C4H11N. (Mr 73.1). 1013600. [109-73-9].
dilute to 100 mL with the same mixture of solvents. Filter Butan-1-amine.
if necessary. Distil and use within one month.
BRP indicator solution. 1013000. Colourless liquid, miscible with water, with ethanol (96 per
Dissolve 0.1 g of bromothymol blue R, 20 mg of methyl red R cent).
and 0.2 g of phenolphthalein R in ethanol (96 per cent) R and : about 1.401.
dilute to 100 mL with the same solvent. Filter. bp : about 78 °C.
Brucine. C23H26N2O4. (Mr 394.5). 1013100. [357-57-3]. tert-Butylamine. 1100900. [75-64-9].
2,3-Dimethoxystrychnidin-10-one. 2,3-Dimethoxystrychnine. See 1,1-dimethylethylamine R.
Colourless crystals, slightly soluble in water, freely soluble in
ethanol (96 per cent). 4-(Butylamino)benzoic acid. C11H15NO2. (Mr 193.2).
1206700. [4740-24-3].
mp : about 178 °C.
White or almost white powder.
Butanal. C4H8O. (Mr 72.1). 1134400. [123-72-8]. Content : 96.5 per cent to 103.5 per cent.
Butyraldehyde.
Butylated hydroxytoluene. 1013800. [128-37-0].
: 0.806.
See Butylhydroxytoluene R.
: 1.380.
bp : 75 °C. Butylboronic acid. C4H11BO2. (Mr 101.9). 1013700.
[4426-47-5].
i-Butane. C4H10. (Mr 58.12). 1189000. [75-28-5]. Isobutane. Content : minimum 98 per cent.
2-Methylpropane. mp : 90 °C to 92 °C.
Content : minimum 99.0 per cent V/V.
tert-Butylhydroperoxide. C4H10O2. (Mr 90.1). 1118000.
n-Butane. C4H10. (Mr 58.12). 1189100. [106-97-8]. Butane. [75-91-2]. 1,1-Dimethylethylhydroperoxide.
Content : minimum 99.0 per cent V/V. Flammable liquid, soluble in organic solvents.
: 0.898.
Butane-1,4-diol. HO(CH2)4OH. (Mr 90.12). 1174800.
[110-63-4]. : 1.401.
bp : 35 °C.
Butanol. C4H10O. (Mr 74.1). 1013200. [71-36-3]. Butan-1-ol.
Clear, colourless liquid, miscible with ethanol (96 per cent). Butyl 4-hydroxybenzoate. 1103900. [94-26-8].
See Butyl parahydroxybenzoate R.
: about 0.81.
bp : 116 °C to 119 °C. Butylhydroxytoluene. 1013800. [128-37-0].
See Butylhydroxytoluene (0581).
2-Butanol R1. C4H10O. (Mr 74.1). 1013301. [78-92-2].
Butan-2-ol. sec-Butyl alcohol. Butyl methacrylate. C8H14O2. (Mr 142.2). 1145400.
Content : minimum 99.0 per cent. [97-88-1]. Butyl 2-methylpropenoate.
Clear, colourless liquid, soluble in water, miscible with ethanol Clear, colourless solution.
(96 per cent). d 420 : about 0.894.

nD20 : about 1.424. Calcium chloride solution, 0.02 M. 1014603.

bp : about 163 °C. Dissolve 2.94 g of calcium chloride R in 900 mL of water R,
adjust to pH 6.0 to 6.2 and dilute to 1000.0 mL with water R.
tert-Butyl methyl ether. 1013900. [1634-04-4]. Storage : at 2 °C to 8 °C.
See 1,1-dimethylethyl methyl ether R.
Calcium chloride solution, 0.025 M. 1014604.
2-Butyloctanol. C12H26O. (Mr 186.3). 1206100. [3913-02-8]. Dissolve 0.368 g of calcium chloride R in water R and dilute
(2Ξ)-2-Butyloctan-1-ol. to 100.0 mL with the same solvent.
Butyl parahydroxybenzoate. 1103900. [94-26-8]. Calcium chloride R1. CaCl2,4H2O. (Mr 183.1). 1014700.
See Butyl parahydroxybenzoate (0881). Calcium chloride tetrahydrate.
Iron : maximum 0.05 ppm.
Butyric acid. C4H8O2. (Mr 88.1). 1014000. [107-92-6].
Butanoic acid. Calcium chloride, anhydrous. CaCl2. (Mr 111.0). 1014800.
Content : minimum 99.0 per cent. [10043-52-4].
Oily liquid, miscible with water and with ethanol (96 per cent). Content : minimum 98.0 per cent (dried substance).
: about 0.96. White or almost white granules, deliquescent, very soluble in
water, freely soluble in ethanol (96 per cent) and in methanol.
: about 1.398.
Loss on drying (2.2.32) : maximum 5.0 per cent, determined by
bp : about 163 °C. drying in an oven at 200 °C.
Butyrolactone. C4H6O2. (Mr 86.1). 1104000. [96-48-0]. Storage : in an airtight container, protected from moisture.
Dihydro-2(3H)-furanone. γ-Butyrolactone.
Calcium hydroxide. Ca(OH)2. (Mr 74.1). 1015000.
Oily liquid, miscible with water, soluble in methanol. [1305-62-0]. Calcium dihydroxide.
: about 1.435. White or almost white powder, almost completely soluble in
bp : about 204 °C. 600 parts of water.
Cadmium. Cd. (Ar 112.4). 1014100. [7440-43-9]. Calcium hydroxide solution. 1015001.
Silvery-white, lustrous metal, practically insoluble in water, A freshly prepared saturated solution.
freely soluble in nitric acid and in hot hydrochloric acid.
Calcium lactate pentahydrate. 1015100. [41372-22-9].
Cadmium nitrate tetrahydrate. Cd(NO3)2,4H2O. (Mr 308.5). See Calcium lactate pentahydrate (0468).
1174900. [10022-68-1].
Calcium phosphate monobasic monohydrate.
Hygroscopic orthorhombic crystals, very soluble in water, CaH4O8P2,H2O. (Mr 252.1). 1157200. [10031-30-8]. Calcium
soluble in acetone and in ethanol (96 per cent). tetrahydrogen bisphosphate monohydrate. Phosphoric acid
mp : about 59.5 °C. calcium salt (2:1) monohydrate.
Caesium chloride. CsCl. (Mr 168.4). 1014200. [7647-17-8]. White or almost white, crystalline powder, soluble in water.
White or almost white powder, very soluble in water, freely Calcium sulfate. CaSO4,1/2H2O. (Mr 145.1). 1015200.
soluble in methanol, practically insoluble in acetone. [10034-76-1]. Calcium sulfate hemihydrate.
White or almost white powder, soluble in about 1500 parts of
Caffeic acid. C9H8O4. (Mr 180.2). 1014300. [331-39-5]. water, practically insoluble in ethanol (96 per cent). When
(E)-3-(3,4-Dihydroxyphenyl)propenoic acid. mixed with half its mass of water it rapidly solidifies to a hard
White or almost white crystals or plates, freely soluble in hot and porous mass.
water and in ethanol (96 per cent), sparingly soluble in cold
water. Calcium sulfate solution. 1015201.
Absorbance (2.2.25). A freshly prepared solution at pH 7.6 Shake 5 g of calcium sulfate R with 100 mL of water R for
shows 2 absorption maxima at about 288 nm and about 1 h and filter.
313 nm. Calconecarboxylic acid. C21H14N2O7S. (Mr 438.4).
Caffeine. 1014400. [58-08-2]. 1015300. [3737-95-9]. 2-Hydroxy-1-(2-hydroxy-4-sulfo-1-
naphthylazo)naphthalene-3-carboxylic acid.
See Caffeine (0267).
Brownish-black powder, slightly soluble in water, very slightly
Calcium acetate. C4H6CaO4. (Mr 158.2). 1191600. [62-54-4]. soluble in acetone and in ethanol (96 per cent), sparingly
Calcium diacetate. See Calcium acetate (2128). soluble in dilute solutions of sodium hydroxide.
Calcium carbonate. 1014500. [471-34-1]. Calconecarboxylic acid triturate. 1015301.
See Calcium carbonate (0014). Mix 1 part of calconecarboxylic acid R with 99 parts of
sodium chloride R.
Calcium carbonate R1. 1014501. Test for sensitivity. Dissolve 50 mg of calconecarboxylic
Complies with the requirements prescribed for calcium acid triturate in a mixture of 2 mL of strong sodium
carbonate R with the following additional requirement. hydroxide solution R and 100 mL of water R. The solution
Chlorides (2.4.4) : maximum 50 ppm. is blue but becomes violet on addition of 1 mL of a 10 g/L
solution of magnesium sulfate R and 0.1 mL of a 1.5 g/L
Calcium chloride. 1014600. [10035-04-8]. solution of calcium chloride R and turns pure blue on
See Calcium chloride (0015). addition of 0.15 mL of 0.01 M sodium edetate.
Calcium chloride solution. 1014601. Camphene. C10H16. (Mr 136.2). 1139200. [79-92-5].
A 73.5 g/L solution of calcium chloride R. 2,2-Dimethyl-3-methylenebicyclo[2.2.1]heptane.
Camphene used in gas chromatography complies with the
Calcium chloride solution, 0.01 M. 1014602. following additional test.
Dissolve 0.147 g of calcium chloride R in water R and dilute Assay. Gas chromatography (2.2.28) as prescribed in the
to 100.0 mL with the same solvent. monograph Rosemary Oil (1846).

Content : minimum 90 per cent, calculated by the : about 1.428.

normalisation procedure. bp : about 239.7 °C.
Camphor. 1113000. [76-22-2]. mp : about 16.7 °C.
See Camphor, racemic (0655). Caprylic acid used in the assay of total fatty acids in Saw
palmetto fruit (1848) complies with the following additional
Camphor used in gas chromatography complies with the test.
Assay. Gas chromatography (2.2.28) as prescribed in the monograph Saw palmetto fruit (1848).
monograph Lavender oil (1338). Content : minimum 98 per cent, calculated by the
Test solution. A 10 g/L solution of the substance to be normalisation procedure.
examined in hexane R.
Capsaicin. C18H27NO3. (Mr 305.4). 1147900. [404-86-4].
Content : minimum 95.0 per cent, calculated by the
(E)-N-[(4-Hydroxy-3-methoxyphenyl)methyl]-8-methylnon-
6-enamide.
(1S)-(+)-10-Camphorsulfonic acid. C10H16O4S. (Mr 232.3). White or almost white, crystalline powder, practically
1104100. [3144-16-9]. (1S,4R)-(+)-2-Oxo-10-bornenesulfonic insoluble in water, freely soluble in anhydrous ethanol.
acid. [(1S)-7,7-Dimethyl-2-oxobicyclo[2.2.1]heptan-1- mp : about 65 °C.
yl]methanesulfonic acid. Reychler’s acid. Capsaicin used in the assay in Capsicum (1859) complies with
Prismatic crystals, hygroscopic, soluble in water. the following additional test.
Content : minimum 99.0 per cent of (1S)-(+)-10- Assay. Liquid chromatography (2.2.29) as prescribed in the
camphorsulfonic acid. monograph Capsicum (1859).
: + 20 ± 1, determined on a 43 g/L solution. Content : minimum 95.0 per cent, calculated by the
mp : about 194 °C, with decomposition. normalisation procedure.
∆A (2.2.41): 10.2 × 103 determined at 290.5 nm on a 1.0 g/L Carbazole. C12H9N. (Mr 167.2). 1015400. [86-74-8].
solution. Dibenzopyrrole.
Crystals, practically insoluble in water, freely soluble in
Capric acid. C10H20O2. (Mr 172.3). 1142000. [334-48-5]. acetone, slightly soluble in anhydrous ethanol.
Decanoic acid.
mp : about 245 °C.
Crystalline solid, very slightly soluble in water, soluble in
anhydrous ethanol. Carbomer. 1015500. [9007-20-9].
bp : about 270 °C. A cross-linked polymer of acrylic acid ; it contains a large
mp : about 31.4 °C. proportion (56 per cent to 68 per cent) of carboxylic acid
(CO2H) groups after drying at 80 °C for 1 h. Average relative
Capric acid used in the assay of total fatty acids in Saw palmetto molecular mass about 3 × 106.
fruit (1848) complies with the following additional test. pH (2.2.3) : about 3 for a 10 g/L suspension.
monograph Saw palmetto fruit (1848). Carbon dioxide. 1015600. [124-38-9].
Content : minimum 98 per cent, calculated by the See Carbon dioxide (0375).
normalisation procedure. Carbon dioxide R1. CO2. (Mr 44.01). 1015700. [124-38-9].
Capric alcohol. 1024700. Content : minimum 99.995 per cent V/V.
See Decanol R. Carbon monoxide : less than 5 ppm.
Oxygen : less than 25 ppm.
Caproic acid. C6H12O2. (Mr 116.2). 1142100. [142-62-1]. Nitric oxide : less than 1 ppm.
Hexanoic acid.
Oily liquid, sparingly soluble in water. Carbon dioxide R2. CO2. (Mr 44.01). 1134500. [124-38-9].
Content : minimum 99 per cent V/V.
d 420 : about 0.926.
Carbon disulfide. CS2. (Mr 76.1). 1015800. [75-15-0].
nD20 : about1.417.
Colourless or yellowish, flammable liquid, practically insoluble
bp : about 205 °C. in water, miscible with anhydrous ethanol.
Caproic acid used in the assay of total fatty acids in Saw : about 1.26.
palmetto fruit (1848) complies with the following additional
bp : 46 °C to 47 °C.
test.
Assay. Gas chromatography (2.2.28) as prescribed in the Carbon for chromatography, graphitised. 1015900.
monograph Saw palmetto fruit (1848). Carbon chains having a length greater than C9 .
Content : minimum 98 per cent, calculated by the Particle size : 400 µm to 850 µm.
normalisation procedure. Relative density : 0.72.
ε-Caprolactam. C6H11NO. (Mr 113.2). 1104200. [105-60-2]. Surface area : 10 m2/g.
Hexane-6-lactam. Do not use at a temperature higher than 400 °C.
Hygroscopic flakes, freely soluble in water, in anhydrous Carbon for chromatography, graphitised R1. 1153500.
ethanol and in methanol. Porous spherical carbon particles comprised of flat sheets of
mp : about 70 °C. hexagonally arranged carbon atoms.
Caprylic acid. C8H16O2. (Mr 144.2). 1142200. [124-07-2]. Particle size : 5 µm to 7 µm.
Octanoic acid. Pore volume : 0.7 cm3/g.
Slightly yellow, oily liquid. Carbon monoxide. CO. (Mr 28.01). 1016000. [630-08-0].
: about 0.910. Content : minimum 99.97 per cent V/V.

Carbon monoxide R1. CO. (Mr 28.01). 1134600. [630-08-0]. Carveol. C10H16O. (Mr 152.2). 1160400. [99-48-9]. p-Mentha-
Content : minimum 99 per cent V/V. 1(6),8-dien-2-ol. 2-Methyl-5-(1-methylethenyl)cyclohex-2-
enol.
Carbon tetrachloride. CCl4. (Mr 153.8). 1016100. [56-23-5]. The substance contains a variable content of trans- and
Tetrachloromethane. cis-carveol.
Clear, colourless liquid, practically insoluble in water, miscible Carveol used in gas chromatography complies with the following
with ethanol (96 per cent). additional test.
: 1.595 to 1.598. Assay. Gas chromatography (2.2.28) as prescribed in the
bp : 76 °C to 77 °C. test for chromatographic profile in the monograph Caraway
oil (1817).
Carbophenothion. C11H16ClO2PS3. (Mr 342.9). 1016200. Content : minimum 97 per cent, calculated by the
[786-19-6]. O,O-Diethyl S-[[(4-chlorophenyl)thio]methyl]- normalisation procedure.
phosphorodithioate.
Yellowish liquid, practically insoluble in water, miscible with Carvone. C10H14O. (Mr 150.2). 1016500. [2244-16-8].
organic solvents. (+)-p-Mentha-6,8-dien-2-one. (5S)-2-Methyl-5-(1-
methylethenyl)-cyclohex-2-enone.
: about 1.27.
Liquid, practically insoluble in water, miscible with ethanol
For the monograph Wool Fat (0134), a suitable certified
(96 per cent).
reference solution (10 ng/µL in iso-octane) may be used.
: about 0.965
Car-3-ene. C10H16. (Mr 136.2). 1124000. [498-15-7]. : about 1.500.
3,7,7-Trimethylbicyclo[4.1.0]hept-3-ene. 4,7,7-Trimethyl-3-
norcarene. : about + 61.
Liquid with a pungent odour, slightly soluble in water, soluble bp : about 230 °C.
in organic solvents. Carvone used in gas chromatography complies with the
: about 0.864. following additional test.
: 1.473 to 1.474. Assay. Gas chromatography (2.2.28) as prescribed in the
: + 15 to + 17. monograph Peppermint oil (0405) using the substance to be
examined as the test solution.
bp : 170 °C to 172 °C.
Car-3-ene used in gas chromatography complies with the normalisation procedure.
Assay. Gas chromatography (2.2.28) as prescribed in the Carvone R1. 1016501.
monograph Nutmeg oil (1552). Complies with the requirements prescribed for carvone R
Content : minimum 95.0 per cent, calculated by the with the following additional requirement.
normalisation procedure. Assay. Gas chromatography (2.2.28) as prescribed in the
test for chiral purity in the monograph Caraway oil (1817).
Carminic acid. C22H20O13. (Mr 492.4). 1156700. [1260-17-9].
7-α-D-Glucopyranosyl-3,5,6,8-tetrahydroxy-1-methyl-9,10- Content : minimum 98 per cent.
dioxo-9,10-dihydroanthracene-2-carboxylic acid.
(−)-Carvone. C10H14O. (Mr 150.2). 1160500.
Dark red powder, very slightly soluble in water, soluble in [6485-40-1]. (–)-p-Mentha-1(6),8-dien-2-one.
dimethyl sulfoxide, very slightly soluble in ethanol (96 per (5R)-2-Methyl-5-(1-methylethenyl)cyclohex-2-enone.
cent).
Liquid.
Carob bean gum. 1104500. : about 0.965.
The ground endosperm of the fruit kernels of Ceratonia : about 1.4988.
siliqua L. Taub.
[α]20 : about − 62.
White or almost white powder containing 70 per cent to D
80 per cent of a water-soluble gum consisting mainly of bp : about 230 °C.
galactomannoglycone.
Assay. Gas chromatography (2.2.28) as prescribed in the test
Carvacrol. C10H14O. (Mr 150.2). 1016400. [499-75-2]. for chiral purity in the monograph Caraway oil (1817).
5-Isopropyl-2-methylphenol. Content : minimum 99 per cent.
Brownish liquid, practically insoluble in water, very soluble in
ethanol (96 per cent). β-Caryophyllene. C15H24. (Mr 204.4). 1101000.
[87-44-5]. (E)-(1R,9S)-4,11,11-Trimethyl-8-methylene-
: about 0.975. bicyclo[7.2.0]undec-4-ene.
: about 1.523. Oily liquid, practically insoluble in water, miscible with
bp : about 237 °C. ethanol (96 per cent).
Carvacrol used in gas chromatography complies with the β-Caryophyllene used in gas chromatography complies with
following additional test. the following additional test.
Assay. Gas chromatography (2.2.28) as prescribed in the Assay. Gas chromatography (2.2.28) as prescribed in the
monograph Peppermint oil (0405). monograph Clove oil (1091).
Test solution. Dissolve 0.1 g in about 10 mL of acetone R. Test solution. The substance to be examined.
Content : minimum 95.0 per cent, calculated by the Content : minimum 90.0 per cent, calculated by the
normalisation procedure. normalisation procedure.

Caryophyllene oxide. C15H24O. (Mr 220.4). 1149000. [1139- Cation-exchange resin, weak. 1203200.
30-6]. (-)-β-Caryophyllene epoxide. (1R,4R,6R,10S)-4,12,12- Weak cation-exchange resin in protonated form with
Trimethyl-9-methylene-5-oxatricyclo[8.2.0.04,6]dodecane. carboxylate functional groups attached to a polymer lattice
Colourless, fine crystals with lumps. consisting of polystyrene cross-linked with divinylbenzene.
mp : 62 °C to 63 °C. Cellulose for chromatography. 1016800. [9004-34-6].
Caryophyllene oxide used in gas chromatography complies with Fine, white or almost white, homogeneous powder with an
the following additional test. average particle size less than 30 µm.
Assay. Gas chromatography (2.2.28) as prescribed in the Preparation of a thin layer. Suspend 15 g in 100 mL of water R
monograph Turpentine oil, Pinus pinaster type (1627). and homogenise in an electric mixer for 60 s. Coat carefully
Content : minimum 99.0 per cent, calculated by the cleaned plates with a layer 0.1 mm thick using a spreading
normalisation procedure. device. Allow to dry in air.
Casein. 1016600. [9000-71-9]. Cellulose for chromatography R1. 1016900.
Mixture of related phosphoproteins obtained from milk. Microcrystalline cellulose.
White or almost white, amorphous powder or granules, very Preparation of a thin layer. Suspend 25 g in 90 mL of water R
slightly soluble in water and in non-polar organic solvents. It and homogenise in an electric mixer for 60 s. Coat carefully
dissolves in concentrated hydrochloric acid giving a pale-violet cleaned plates with a layer 0.1 mm thick using a spreading
solution. It forms salts with acids and bases. Its isoelectric device. Allow to dry in air.
point is at about pH 4.7. Alkaline solutions are laevorotatory.
Cellulose for chromatography F254. 1017000.
Casticin. C19H18O8. (Mr 374.3). 1162200. [479-91-4]. Microcrystalline cellulose F254. A fine, white or almost white,
5-Hydroxy-2-(3-hydroxy-4-methoxyphenyl)-3,6,7- homogeneous powder with an average particle size less than
trimethoxy-4H-1-benzopyran-4-one. 30 µm, containing a fluorescent indicator having an optimal
Yellow crystals. intensity at 254 nm.
Preparation of a thin layer. Suspend 25 g in 100 mL of water R
Catalpol. C15H22O10. (Mr 362.3). 1142300. [2415-24-9]. and homogenise using an electric mixer for 60 s. Coat
(1aS,1bS,2S,5aR,6S,6aS)-6-Hydroxy-1a-(hydroxymethyl)- carefully cleaned plates with a layer 0.1 mm thick using a
1a,1b,2,5a,6,6a-hexahydrooxireno[4,5]cyclopenta[1,2- spreading device. Allow to dry in air.
c]pyran-2-yl β-D-glucopyranoside.
mp : 203 °C to 205 °C. Cerium sulfate. Ce(SO4)2,4H2O. (Mr 404.3). 1017300.
[10294-42-5]. Cerium(IV) sulfate tetrahydrate. Ceric sulfate.
Catechin. C15H14O6,xH2O. (Mr 290.3 for the anhydrous Yellow or orange-yellow, crystalline powder or crystals, very
substance). 1119000. [154-23-4]. (+)-(2R,3S)-2-(3,4- slightly soluble in water, slowly soluble in dilute acids.
Dihydroxyphenyl)-3,4-dihydro-2H-chromene-3,5,7-triol.
Catechol. Cianidanol. Cyanidol. Cerous nitrate. Ce(NO3)3,6H2O. (Mr 434.3). 1017400.
[10294-41-4]. Cerium trinitrate hexahydrate.
Cathine hydrochloride. C9H14ClNO. (Mr 187.7). 1206800. Colourless or pale yellow, crystalline powder, freely soluble in
[2153-98-2]. (1S,2S)-2-Amino-1-phenylpropan-1-ol water and in ethanol (96 per cent).
hydrochloride. Norpseudoephedrine hydrochloride.
White or almost white solid. Cetostearyl alcohol. 1017500. [67762-27-0].
Content : minimum 95.0 per cent. See Cetostearyl alcohol (0702).
Cation-exchange resin. 1016700. Cetrimide. 1017600. [8044-71-1].

A resin in protonated form with sulfonic acid groups attached See Cetrimide (0378).
to a polymer lattice consisting of polystyrene cross-linked with Cetyl alcohol. C16H34O. (Mr 242.4). 1160600. [36653-82-4].
8 per cent of divinylbenzene. It is available as spherical beads. Hexadecan-1-ol.
Cation-exchange resin R1. 1121900. Content : minimum 95.0 per cent.
A resin in protonated form with sulfonic acid groups attached mp : about 48 °C.
to a polymer lattice consisting of polystyrene cross-linked with
4 per cent of divinylbenzene. It is available as spherical beads. Cetylpyridinium chloride monohydrate. C21H38ClN,H2O.
(Mr 358.0). 1162800. [6004-24-6]. 1-Hexadecylpyridinium
Cation-exchange resin R2. 1195400. chloride monohydrate.
Resin containing strongly acidic propylenesulfonic acid White or almost white powder, freely soluble in water and in
groups. ethanol (96 per cent).
mp : 80 °C to 83 °C.
Cation-exchange resin (calcium form), strong. 1104600.
Resin in calcium form with sulfonic acid groups attached to Cetyltrimethylammonium bromide. C19H42BrN.
a polymer lattice consisting of polystyrene cross-linked with (Mr 364.5). 1017700. [57-09-0]. Cetrimonium bromide.
8 per cent of divinylbenzene. N-Hexadecyl-N,N,N-trimethylammonium bromide.
White or almost white, crystalline powder, soluble in water,
Cation-exchange resin (sodium form), strong. 1176100. freely soluble in ethanol (96 per cent).
Resin in sodium form with sulfonic acid groups attached to mp : about 240 °C.
a polymer lattice consisting of polystyrene cross-linked with
divinylbenzene. Chamazulene. C14H16. (Mr 184.3). 1148000. [529-05-5].
7-Ethyl-1,4-dimethylazulene.
Cation-exchange resin, strong. 1156800. Blue liquid, very slightly soluble in water, soluble in ethanol
Strong cation-exchange resin in protonated form with sulfonic (96 per cent), miscible with fatty oils, with essential oils and
acid groups attached to a polymer lattice consisting of with liquid paraffin, soluble with discolouration in phosphoric
polystyrene cross-linked with divinylbenzene. acid (85 per cent m/m) and sulfuric acid (50 per cent V/V).

Appearance of solution. 50 mg is soluble in 2.5 mL of hexane R. 2-Chlorobenzoic acid. C7H5ClO2. (Mr 156.6). 1139300.
The blue solution is clear in a thin-layer obtained by tilting [118-91-2].
the test-tube. Soluble in water, slightly soluble in anhydrous ethanol.
Chamazulene used for gas chromatography complies with the bp : about 285 °C.
following additional test. mp : about 140 °C.
monograph Matricaria oil (1836). Chlorobutanol. 1018400. [57-15-8].
Test solution : a 4 g/L solution in cyclohexane R. See Chlorobutanol (0382).
Content : minimum 95.0 per cent, calculated by the 2-Chloro-2-deoxy- D-glucose. C6H11ClO5. (Mr 198.6).
normalisation procedure. 1134700. [14685-79-1].
Charcoal, activated. 1017800. [64365-11-3]. White or almost white crystalline, very hygroscopic powder,
soluble in water and in dimethyl sulfoxide, practically
See Activated charcoal (0313). insoluble in ethanol (96 per cent).
Chloral hydrate. 1017900. [302-17-0]. 2-Chloroethanol. C2H5ClO. (Mr 80.5). 1097500. [107-07-3].
See Choral hydrate (0265). Colourless liquid, soluble in ethanol (96 per cent).
Chloral hydrate solution. 1017901. : about 1.197.
A solution of 80 g in 20 mL of water R. : about 1.442.
bp : about 130 °C.
Chloramine. 1018000. [7080-50-4].
mp : about − 89 °C.
See Tosylchloramide sodium (0381).
2-Chloroethanol solution. 1097501.
Chloramine solution. 1018001.
Dissolve 125 mg of 2-chloroethanol R in 2-propanol R and
A 20 g/L solution of chloramine R. Prepare immediately dilute to 50 mL with the same solvent. Dilute 5 mL of the
before use. solution to 50 mL with 2-propanol R.
Chloramine solution R1. 1018002. Chloroethylamine hydrochloride. C2H7Cl2N. (Mr 116.0).
A 0.1 g/L solution of chloramine R. Prepare immediately 1124300. [870-24-6]. 2-Chloroethanamine hydrochloride.
before use. mp : about 145 °C.
Chloramine solution R2. 1018003. (2-Chloroethyl)diethylamine hydrochloride. C6H15Cl2N.
A 0.2 g/L solution of chloramine R. Prepare immediately (Mr 172.1). 1018500. [869-24-9].
before use. White or almost white, crystalline powder, very soluble in
water and in methanol, freely soluble in methylene chloride,
Chlordane. C10H6Cl8. (Mr 409.8). 1124100. [12789-03-6]. practically insoluble in hexane.
bp : about 175 °C. mp : about 211 °C.
mp : about 106 °C.
A suitable certified reference solution of technical grade Chloroform. CHCl3. (Mr 119.4). 1018600. [67-66-3].
(10 ng/µL in iso-octane) may be used. Trichloromethane.
Clear, colourless liquid, slightly soluble in water, miscible with
Chlordiazepoxide. 1113200. [58-25-3]. ethanol (96 per cent).
See Chlordiazepoxide (0656). : 1.475 to 1.481.
Chlorfenvinphos. C12H14Cl3O4P. (Mr 359.6). 1124200. bp : about 60 °C.
[470-90-6]. Ethanol : 0.4 per cent m/m to 1.0 per cent m/m.
A suitable certified reference solution (10 ng/µL in Chloroform, acidified. 1018601.
cyclohexane) may be used. To 100 mL of chloroform R add 10 mL of hydrochloric
Chloroacetanilide. C8H8ClNO. (Mr 169.6). 1018100. acid R. Shake, allow to stand and separate the 2 layers.
[539-03-7]. 4′-Chloroacetanilide. Chloroform, ethanol-free. 1018602.
Content : minimum 95 per cent. Shake 200 mL of chloroform R with four quantities, each
Crystalline powder, practically insoluble in water, soluble in of 100 mL, of water R. Dry over 20 g of anhydrous sodium
ethanol (96 per cent). sulfate R for 24 h. Distil the filtrate over 10 g of anhydrous
mp : about 178 °C. sodium sulfate R. Discard the first 20 mL of distillate.
Prepare immediately before use.
Chloroacetic acid. C2H3ClO2. (Mr 94.5). 1018200. [79-11-8].
Chlorogenic acid. C16H18O9. (Mr 354.3). 1104700. [327-97-9].
Colourless or white or almost white crystals, deliquescent,
(1S,3R,4R,5R)-3-[(3,4-Dihydroxycinnamoyl)oxy]-1,4,5-
very soluble in water, soluble in ethanol (96 per cent).
trihydroxycyclohexanecarboxylic acid.
Storage : in an airtight container.
White or almost white, crystalline powder or needles, freely
Chloroaniline. C6H6ClN. (Mr 127.6). 1018300. [106-47-8]. soluble in boiling water, in acetone and in ethanol (96 per
4-Chloroaniline. cent).
Crystals, soluble in hot water, freely soluble in ethanol (96 per : about − 35.2.
cent). mp : about 208 °C.
mp : about 71 °C. Chromatography. Thin-layer chromatography (2.2.27) as
prescribed on Identification A in the monograph Belladonna
4-Chlorobenzenesulfonamide. C6H6ClNO2S. (Mr 191.6). leaf dry extract, standardised (1294) ; the chromatogram shows
1097400. [98-64-6]. only one principal zone.
White or almost white powder. Chlorogenic acid used in liquid chromatography complies with
mp : about 145 °C. the following additional test.

Assay. Liquid chromatography (2.2.29) as prescribed in the 5-Chlorosalicylic acid. C7H5ClO3. (Mr 172.6). 1019100.
monograph Artichoke Leaf (1866). [321-14-2].
Content : minimum 97.0 per cent. White or almost white, crystalline powder, soluble in
methanol.
3-Chloro-2-methylaniline. C7H8ClN. (Mr 141.6). 1139400. mp : about 173 °C.
[87-60-5]. 6-Chloro-2-toluidine.
Not miscible with water, slightly soluble in anhydrous ethanol. Chlorothiazide. C7H6ClN3O4S2. (Mr 295.7). 1112100. [58-
94-6]. 6-Chloro-2H-1,2,4-benzothiadiazine-7-sulfonamide
: about 1.171. 1,1-dioxide.
: about 1.587. Content : minimum 98.0 per cent.
bp : about 115 °C. White or almost white, crystalline powder, very slightly
mp : about 2 °C. soluble in water, sparingly soluble in acetone, slightly soluble
in ethanol (96 per cent). It dissolves in dilute solutions of
2-Chloro-N-(2,6-dimethylphenyl)acetamide. C10H12ClNO. alkali hydroxides.
(Mr 197.7). 1168700. [1131-01-7].
Chlorotrimethylsilane. C3H9ClSi. (Mr 108.6). 1019300.
2-Chloronicotinic acid. C6H4ClNO2. (Mr 157.6). 1157300. [75-77-4].
[2942-59-8]. 2-Chloropyridine-3-carboxylic acid. Clear, colourless liquid, fuming in air.
White or almost white powder. : about 0.86.
mp : about 177 °C. : about 1.388.
Content : minimum 95 per cent. bp : about 57 °C.
2-Chloro-4-nitroaniline. C6H5ClN2O2. (Mr 172.6). 1018800. Chlorpyriphos. C9H11Cl3NO3PS. (Mr 350.6). 1124400.

[121-87-9]. [2921-88-2].
Yellow, crystalline powder, freely soluble in methanol. bp : about 200 °C.
mp : 42 °C to 44 °C.
mp : about 107 °C.
A suitable certified reference solution (10 ng/µL in
Storage : protected from light. cyclohexane) may be used.
2-Chloro-5-nitrobenzoic acid. C7H4ClNO4. (Mr 201.6). Chlorpyriphos-methyl. C7H7Cl3NO3PS. (Mr 322.5). 1124500.
1183800. [2516-96-3]. [5598-13-0].
mp : 165 °C to 168 °C. mp : 45 °C to 47 °C.
Chlorophenol. C6H5ClO. (Mr 128.6). 1018900. [106-48-9]. A suitable certified reference solution (10 ng/µL in
4-Chlorophenol. cyclohexane) may be used.
Colourless or almost colourless crystals, slightly soluble in Chlortetracycline hydrochloride. 1145500.
water, very soluble in ethanol (96 per cent) and in solutions of See Chlortetracycline hydrochloride (0173).
alkali hydroxides.
(5α)-Cholestane. C27H48. (Mr 372.7). 1167900. [481-21-0].
mp : about 42 °C.
Slightly soluble in anhydrous ethanol.
2-[2-(4-Chlorophenyl)acetyl]benzoic acid. C15H11ClO3. mp : about 81 °C.
(Mr 274.7). 1194500. [53242-76-5].
Cholesterol. 1019400. [57-88-5].
Chloroplatinic acid. H2Cl6Pt,6H2O. (Mr 517.9). 1019000. See Cholesterol (0993).
[18497-13-7]. Hydrogen hexachloroplatinate(IV) hexahydrate.
Choline chloride. C5H14ClNO. (Mr 139.6). 1019500.
Content : minimum 37.0 per cent m/m of platinum (Ar 195.1). [67-48-1]. (2-Hydroxyethyl)trimethylammonium chloride.
Brownish-red crystals or a crystalline mass, very soluble in Deliquescent crystals, very soluble in water and in ethanol
water, soluble in ethanol (96 per cent). (96 per cent).
Assay. Ignite 0.200 g to constant mass at 900 ± 50 °C and Chromatography. Thin-layer chromatography (2.2.27) as
weigh the residue (platinum). prescribed in the monograph Suxamethonium chloride (0248) :
Storage : protected from light. apply 5 µL of a 0.2 g/L solution in methanol R ; the
chromatogram shows one principal spot.
3-Chloropropane-1,2-diol. C3H7ClO2. (Mr 110.5). 1097600. Storage : in an airtight container.
[96-24-2].
Colourless liquid, soluble in water and ethanol (96 per cent). Chondroitinase ABC. 1162900.
: about 1.322. Pectin lyase-like enzyme secreted by Flavobacterium
heparinum. Available in vials containing 5-10 units. It cleaves
: about 1.480. both glucuronate-containing disaccharides, e.g. chondroitin
bp : about 213 °C. sulfate, and iduronate-containing disaccharides, e.g. dermatan
sulfate.
5-Chloroquinolin-8-ol. C9H6ClNO. (Mr 179.6). 1156900.
[130-16-5]. 5-Chlorooxine. Chondroitinase AC. 1163000.
Sparingly soluble in cold dilute hydrochloric acid. Pectin lyase-like enzyme secreted by Flavobacterium
heparinum. Available in vials containing 5-10 units. It cleaves
mp : about 123 °C. only glucuronate-containing disaccharides, e.g. chondroitin
Content : minimum 95.0 per cent. sulfate.
4-Chlororesorcinol. C6H5ClO2. (Mr 144.6). Chromazurol S. C23H13Cl2Na3O9S. (Mr 605). 1019600.
1177700. [95-88-5]. 4-Chlorobenzene-1,3-diol. [1667-99-8].
1,3-Dihydroxy-4-chlorobenzene. Schultz No. 841.
mp : 106 °C to 108 °C. Colour Index No. 43825.

Trisodium 5-[(3-carboxylato-5-methyl-4-oxocyclohexa-2,5- Chromotropic acid, sodium salt solution. 1020301.

dien-1-ylidene)(2,6-dichloro-3-sulfonatophenyl)methyl]-2- Dissolve 0.60 g of chromotropic acid, sodium salt R in about
hydroxy-3-methylbenzoate. 80 mL of water R and dilute to 100 mL with the same
Brownish-black powder, soluble in water, slightly soluble in solvent. Use this solution within 24 h.
Chromotropic acid-sulfuric acid solution. 1020302.
Chromic potassium sulfate. CrK(SO4)2,12H2O. (Mr 499.4). Dissolve 5 mg of chromotropic acid, sodium salt R in 10 mL
1019800. [7788-99-0]. Chrome alum. of a mixture of 9 mL of sulfuric acid R and 4 mL of water R.
Large, violet-red or black crystals, freely soluble in water, Chrysanthemin. C21H21ClO11. (Mr 485.8). 1134800.
practically insoluble in ethanol (96 per cent). [7084-24-4]. Cyanidin 3-O-glucoside chloride.
Chromium(III) acetylacetonate. C15H21CrO6. (Mr 349.3). Kuromanin chloride. 2-(3,4-Dihydroxyphenyl)-3-(β-D-
1172900. [21679-31-2]. (OC-6-11)-Tris(2,4-pentanedionato- glucopyranosyl)oxy-5,7-dihydroxy-1-benzopyrylium chloride.
κO,κO′)chromium. Reddish-brown crystalline powder, soluble in water and in
Chromium(III) trichloride hexahydrate. Absorbance (2.2.25). A 0.01 g/L solution in a mixture
[Cr(H2O)4Cl2]Cl,2H2O. (Mr 266.5). 1104800. [10060-12-5]. of 1 volume of hydrochloric acid R and 999 volumes of
Dark green crystalline powder, hygroscopic. methanol R shows an absorption maximum at 528 nm.
Storage : protected from humidity and oxidising agents. α-Chymotrypsin for peptide mapping. 1142400.
Chromium trioxide. CrO3. (Mr 100.0). 1019900. [1333-82-0]. α-Chymotrypsin of high purity, treated to eliminate tryptic
Dark brownish-red needles or granules, deliquescent, very activity.
soluble in water. Cimifugin. C16H18O6. (Mr 306.3). 1181700. [37921-38-3].
Storage : in an airtight glass container. (2S)-7-(Hydroxymethyl)-2-(1-hydroxy-1-methylethyl)-4-
methoxy-2,3-dihydro-5H-furo[3,2-g][1]benzopyran-5-one.
Chromogenic substrate R1. 1020000.
Dissolve N-α-benzyloxycarbonyl-D-arginyl-L- Cinchonidine. C19H22N2O. (Mr 294.4). 1020400. [485-71-2].
glycyl-L-arginine-4-nitroanilide dihydrochloride (R)-(Quinol-4-yl)[(2S,4S,5R)-5-vinylquinuclidin-2-
in water R to give a 0.003 M solution. Dilute in yl]methanol.
tris(hydroxymethyl)aminomethane-EDTA buffer solution White or almost white, crystalline powder, very slightly
pH 8.4 R to 0.0005 M before use. soluble in water and in light petroleum, soluble in ethanol
(96 per cent).
Chromogenic substrate R2. 1020100. : − 105 to − 110, determined on a 50 g/L solution in
Dissolve D-phenylalanyl-L-pipecolyl-L-arginine-4-nitroanilide ethanol (96 per cent) R.
dihydrochloride in water R to give a 0.003 M solution. Dilute mp : about 208 °C, with decomposition.
before use in titrating in tris(hydroxymethyl)aminomethane- Storage : protected from light.
EDTA buffer solution pH 8.4 R to give a 0.0005 M solution.
Cinchonine. C19H22N2O. (Mr 294.4). 1020500. [118-10-5].
Chromogenic substrate R3. 1149100. (S)-(Quinol-4-yl)[(2R,4S,5R)-5-vinylquinuclidin-2-
Dissolve D-valyl-leucyl-lysyl-4-nitroanilide dihydrochloride in yl]methanol.
water R to give a 0.003 M solution. White or almost white, crystalline powder, very slightly
Chromogenic substrate R4. 1163100. soluble in water, sparingly soluble in ethanol (96 per cent) and
in methanol.
Dissolve D-phenylalanyl-L-pipecolyl-L-arginine-4-nitroanilide
: + 225 to + 230, determined on a 50 g/L solution in
dihydrochloride in water R to give a 0.008 M solution. Dilute
ethanol (96 per cent) R.
to 0.0025 M with phosphate buffer solution pH 8.5 R before use.
mp : about 263 °C.
Chromogenic substrate R5. 1163200. Storage : protected from light.
Dissolve N-benzoyl-L-isoleucyl-L-glutamyl-glycyl-L-arginine-
Cineole. C10H18O. (Mr 154.3). 1020600. [470-82-6].
4-nitroanilide hydrochloride in water R to give a 0.003 M
1,8-Cineole. Eucalyptol. 1,8-Epoxy-p-menthane.
solution.
Colourless liquid, practically insoluble in water, miscible with
Chromotrope II B. C16H9N3Na2O10S2. (Mr 513.4). 1020200. anhydrous ethanol.
[548-80-1]. : 0.922 to 0.927.
Schultz No. 67. : 1.456 to 1.459.
Colour Index No. 16575. Freezing point (2.2.18) : 0 °C to 1 °C.
Disodium 4,5-dihydroxy-3-(4-nitrophenylazo)naphthalene- Distillation range (2.2.11): 174 °C to 177 °C.
2,7-disulfonate.
Phenol. Shake 1 g with 20 mL of water R. Allow to separate
Reddish-brown powder, soluble in water giving a yellowish-red and add to 10 mL of the aqueous layer 0.1 mL of ferric chloride
colour, practically insoluble in ethanol (96 per cent). solution R1. No violet colour develops.
Chromotrope II B solution. 1020201. Turpentine oil. Dissolve 1 g in 5 mL of ethanol (90 per
cent V/V) R. Add dropwise freshly prepared bromine water R.
A 0.05 g/L solution of chromotrope II B R in sulfuric acid R. Not more than 0.5 mL is required to give a yellow colour
Chromotropic acid, sodium salt. C10H6Na2O8S2,2H2O. lasting for 30 min.
(Mr 400.3). 1020300. [5808-22-0]. Residue on evaporation : maximum 0.05 per cent.
Schultz No. 1136. To 10.0 mL add 25 mL of water R, evaporate on a water-bath
Disodium 4,5-dihydroxynaphthalene-2,7-disulfonate and dry the residue to constant mass at 100-105 °C.
dihydrate. Disodium 1,8-dihydroxynaphthalene-3,6- Cineole used in gas chromatography complies with the following
disulfonate dihydrate. additional test.
A yellowish-white powder, soluble in water, practically Assay. Gas chromatography (2.2.28) as prescribed in the
insoluble in ethanol (96 per cent). monograph Peppermint oil (0405).

Test solution. The substance to be examined. Citrated rabbit plasma. 1020900.

Content : minimum 98.0 per cent, calculated by the Collect blood by intracardiac puncture from a rabbit kept
normalisation procedure. fasting for 12 h, using a plastic syringe with a No. 1 needle
containing a suitable volume of 38 g/L solution of sodium
1,4-Cineole. C10H18O. (Mr 154.3). 1142500. [470-67-7]. citrate R so that the final volume ratio of citrate solution to
1-Methyl-4-(1-methylethyl)-7-oxabicyclo[2.2.1]heptane. blood is 1 : 9. Separate the plasma by centrifugation at 1500 g
1-Isopropyl-4-methyl-7-oxabicyclo[2.2.1]heptane. to 1800 g at 15 °C to 20 °C for 30 min.
Colourless liquid. Storage : at 0 °C to 6 °C ; use within 4 h of collection.
: about 0.900. Citric acid, anhydrous. 1021200. [77-92-9].
: about 1.445. See Citric acid (0455).
bp : about 173 °C.
Citric acid monohydrate. 1021000. [5949-29-1].
Cinnamamide. C9H9NO. (Mr 147.2). 1154800. [621-79-4]. See Citric acid monohydrate (0456).
(E)-3-Phenylprop-2-enamide. When used in the test for iron, it complies with the following
White or almost white powder. additional requirement.
mp : about 149 °C. Dissolve 0.5 g in 10 mL of water R, add 0.1 mL of thioglycollic
acid R, mix and make alkaline with ammonia R. Dilute to
trans-Cinnamic acid. C9H8O2. (Mr 148.2). 1159200. 20 mL with water R. No pink colour appears in the solution.
[140-10-3]. trans-3-Phenylacrylic acid. (2E)-3-Phenylprop-
2-enoic acid. Citronellal. C10H18O. (Mr 154.3). 1113300. [106-23-0].
3,7-Dimethyl-6-octenal.
Colourless crystals, very slightly soluble in water, freely soluble Very slightly soluble in water, soluble in ethanol (96 per cent).
: 0.848 to 0.856.
mp : 133 °C.
: about 1.446.
Cinnamic aldehyde. C9H8O. (Mr 132.2). 1020700. [104-55-2]. Citronellal used in gas chromatography complies with the
3-Phenylpropenal. following additional test.
Yellowish or greenish-yellow, oily liquid, slightly soluble in Assay. Gas chromatography (2.2.28) as prescribed in the
water, very soluble in ethanol (96 per cent). monograph Citronella oil (1609).
: about 1.620. Content : minimum 95.0 per cent, calculated by the
Storage : protected from light. normalisation procedure.
Citronellol. C10H20O. (Mr 156.3). 1134900. [106-22-9].
trans-Cinnamic aldehyde. C9H8O. (Mr 132.2). 1124600. 3,7-Dimethyloct-6-en-1-ol.
[14371-10-9]. (E)-3-Phenylprop-2-enal.
Clear, colourless liquid, practically insoluble in water, miscible
trans-Cinnamic aldehyde used in gas chromatography complies with ethanol (96 per cent).
with the following additional test.
: 0.857.
Assay. Gas chromatography (2.2.28) as prescribed in the : 1.456.
monograph Cassia oil (1496).
bp : 220 °C to 222 °C.
normalisation procedure. Citronellol used in gas chromatography complies with the
Cinnamyl acetate. C11H12O2. (Mr 176.2). 1124700. Assay. Gas chromatography (2.2.28) as prescribed in the
[103-54-8]. 3-Phenylprop-2-en-1-yl acetate. monograph Citronella oil (1609).
: about 1.542. Content : minimum 95.0 per cent, calculated by the
bp : about 262 °C. normalisation procedure.
Cinnamyl acetate used in gas chromatography complies with Storage : in an airtight container, protected from light.
the following additional test. Citronellyl acetate. C12H22O2. (Mr 198.3). 1135000.
Assay. Gas chromatography (2.2.28) as prescribed in the [150-84-5]. 3,7-Dimethyl-6-octen-1-yl acetate.
monograph Cassia oil (1496). : 0.890.
Content : minimum 99.0 per cent, calculated by the : 1.443.
normalisation procedure. bp : 229 °C.
Citral. C10H16O. (Mr 152.2). 1020800. [5392-40-5]. Mixture of Citronellyl acetate used in gas chromatography complies with
(2E)- and (2Z)-3,7-Dimethylocta-2,6-dienal. the following additional test.
Light yellow liquid, practically insoluble in water, miscible Assay. Gas chromatography (2.2.28) as prescribed in the
with ethanol (96 per cent) and with propylene glycol. monograph Citronella oil (1609).
Chromatography. Thin-layer chromatography (2.2.27), using
silica gel GF254 R as the coating substance : apply to the plate
10 µL of a 1 g/L solution in toluene R. Develop over a path of Storage : in an airtight container, protected from light.
15 cm using a mixture of 15 volumes of ethyl acetate R and Citropten. C11H10O4. (Mr 206.2). 1021300. [487-06-9].
85 volumes of toluene R. Allow the plate to dry in air and Limettin. 5,7-Dimethoxy-2H-1-benzopyran-2-one.
examine in ultraviolet light at 254 nm. The chromatogram Needle-shaped crystals, practically insoluble in water and
shows only one principal spot. in light petroleum, freely soluble in acetone and in ethanol
Citral used in gas chromatography complies with the following (96 per cent).
additional test. mp : about 145 °C.
Assay. Gas chromatography (2.2.28) as prescribed in the Chromatography. Thin-layer chromatography (2.2.27), using
monograph Citronella oil (1609). silica gel GF254 R as the coating substance : apply to the plate
Content of citral (neral + geranial): minimum 95.0 per cent, 10 µL of a 1 g/L solution in toluene R. Develop over a path of
calculated by the normalisation procedure. 15 cm using a mixture of 15 volumes of ethyl acetate R and

85 volumes of toluene R. Allow the plate to dry in air and Congo red paper. 1022002.
examine in ultraviolet light at 254 nm. The chromatogram Immerse strips of filter paper for a few minutes in congo
obtained shows only one principal spot. red solution R. Allow to dry.
Clobetasol propionate. C25H32ClFO5. (Mr 467.0). 1097700. Congo red solution. 1022001.
[25122-46-7]. 21-Chloro-9-fluoro-11β,17-dihydroxy-16β-
methylpregna-1,4-diene-3,20-dione 17-propionate. Dissolve 0.1 g of congo red R in a mixture of 20 mL of
ethanol (96 per cent) R and water R and dilute to 100 mL
White or almost white crystalline powder, insoluble in water,
with water R.
soluble in ethanol (96 per cent) and in acetone.
Test for sensitivity. To 0.2 mL of the congo red solution add
: about + 104 (in dioxan).
100 mL of carbon dioxide-free water R and 0.3 mL of 0.1 M
mp : about 196 °C. hydrochloric acid. The solution is blue. Not more than
Coagulation factor V solution. 1021400. 0.3 mL of 0.1 M sodium hydroxide is required to change
the colour to pink.
Coagulation factor V solution may be prepared by the
following method or by any other method which excludes Colour change : pH 3.0 (blue) to pH 5.0 (pink).
factor VIII. Coomassie blue. 1001400. [3861-73-2].
Prepare the factor V reagent from fresh oxalated bovine
See acid blue 92 R.
plasma, by fractionation at 4 °C with a saturated solution of
ammonium sulfate R prepared at 4 °C. Separate the fraction Coomassie blue solution. 1001401.
which precipitates between 38 per cent and 50 per cent
of saturation, which contains factor V without significant See acid blue 92 solution R.
contamination with factor VIII. Remove the ammonium Coomassie staining solution. 1012201.
sulfate by dialysis and dilute the solution with a 9 g/L solution
of sodium chloride R to give a solution containing between A 1.25 g/L solution of acid blue 83 R in a mixture consisting of
10 per cent and 20 per cent of the quantity of factor V present 1 volume of glacial acetic acid R, 4 volumes of methanol R and
in fresh human normal plasma. 5 volumes of water R. Filter.
Assay of factor V. Prepare two dilutions of the preparation Coomassie staining solution R1. 1173000.
of factor V in imidazole buffer solution pH 7.3 R containing
Dissolve 0.275 g of acid blue 83 R in 200 mL of methanol R.
1 volume of the preparation in 10 volumes and in 20 volumes
Stir until complete dissolution of the crystals (for about 2 h).
of the buffer solution respectively. Test each dilution
Add 750 mL of water R and 50 mL of glacial acetic acid R. Stir
as follows : mix 0.1 mL of plasma substrate deficient in
overnight (for at least 16 h) ; filter.
factor V R, 0.1 mL of the solution to be examined, 0.1 mL of
thromboplastin R and 0.1 mL of a 3.5 g/L solution of calcium Copper. Cu. (Ar 63.55). 1022100. [7440-50-8].
chloride R and measure the coagulation times, i.e. the interval
between the moment at which the calcium chloride solution is Cleaned foil, turnings, wire or powder of the pure metal of
added and the first indication of the formation of fibrin, which electrolytic grade.
may be observed visually or by means of a suitable apparatus. Copper acetate. C4H6CuO4,H2O. (Mr 199.7). 1022200.
In the same manner, determine the coagulation time (in [6046-93-1].
duplicate) of four dilutions of human normal plasma in Blue-green crystals or powder, freely soluble in boiling water,
imidazole buffer solution pH 7.3 R, containing respectively, soluble in water and in ethanol (96 per cent), slightly soluble
1 volume in 10 (equivalent to 100 per cent of factor V), in glycerol (85 per cent).
1 volume in 50 (20 per cent), 1 volume in 100 (10 per cent),
and 1 volume in 1000 (1 per cent). Using two-way logarithmic Copper edetate solution. 1022300.
paper plot the average coagulation times for each dilution of To 2 mL of a 20 g/L solution of copper acetate R add 2 mL of
human plasma against the equivalent percentage of factor V 0.1 M sodium edetate and dilute to 50 mL with water R.
and read the percentage of factor V for the two dilutions of the
factor V solution by interpolation. The mean of the two results Copper nitrate. Cu(NO3)2,3H2O. (Mr 241.6). 1022400.
gives the percentage of factor V in the solution to be examined. [10031-43-3]. Copper dinitrate trihydrate.
Storage : in the frozen state at a temperature not higher than Dark blue crystals, hygroscopic, very soluble in water giving a
− 20 °C. strongly acid reaction, freely soluble in ethanol (96 per cent)
Cobalt chloride. CoCl2,6H2O. (Mr 237.9). 1021600. and in dilute nitric acid.
[7791-13-1]. Storage : in an airtight container.
Red, crystalline powder or deep-red crystals, very soluble in
Copper sulfate, anhydrous. CuSO4. (Mr 159.6). 1199000.
water, soluble in ethanol (96 per cent).
[7758-98-7].
Cobalt nitrate. Co(NO3)2,6H2O. (Mr 291.0). 1021700. Greenish-grey powder, hygroscopic, freely soluble in water,
[10026-22-9]. slightly soluble in methanol and practically insoluble in
Small garnet-red crystals, very soluble in water. ethanol (96 per cent).
Codeine. 1021800. [6059-47-8]. Copper sulfate solution R1. 1199001.
See Codeine monohydrate (0076). To 600 mL of water R slowly add 80 mL of phosphoric
acid R. Dissolve with stirring 100 g of anhydrous copper
Codeine phosphate. 1021900. [52-28-8]. sulfate R and dilute to 1 L with water R.
See Codeine phosphate hemihydrate (0074).
Copper sulfate pentahydrate. CuSO4,5H2O. (Mr 249.7).
Congo red. C32H22N6Na2O6S2. (Mr 697). 1022000. [573-58-0]. 1022500. [7758-99-8].
Schultz No. 360. Blue powder or deep-blue crystals, slowly efflorescent, very
Colour Index No. 22120. soluble in water, slightly soluble in ethanol (96 per cent).
Disodium (biphenyl-4,4′-diyl-bis-2,2′-azo)bis(1-amino-
naphthalene-4-sulfonate). Copper sulfate solution. 1022501.
Brownish-red powder, soluble in water. A 125 g/L solution of copper sulfate pentahydrate R.

Copper tetrammine, ammoniacal solution of. 1022600. p-Cresol. C7H8O. (Mr 108.1). 1153100. [106-44-5].
Dissolve 34.5 g of copper sulfate pentahydrate R in 100 mL 4-Methylphenol.
of water R and, whilst stirring, add dropwise concentrated Colourless or white or almost white crystals or crystalline
ammonia R until the precipitate which forms dissolves mass.
completely. Keeping the temperature below 20 °C, add 20
d 20 : about 1.02.
dropwise with continuous shaking 30 mL of strong sodium
hydroxide solution R. Filter through a sintered-glass filter (40) bp : about 202 °C.
(2.1.2), wash with water R until the filtrate is clear and take up
m-Cresol purple. C21H18O5S. (Mr 382.44). 1121700.
the precipitate with 200 mL of concentrated ammonia R. Filter
[2303-01-7]. m-Cresolsulfonphthalein.
through a sintered-glass filter (2.1.2) and repeat the filtration
to reduce the residue to a minimum. Olive-green, crystalline powder, slightly soluble in water,
soluble in ethanol (96 per cent), in glacial acetic acid and in
Cortisone. C21H28O5. (Mr 360.4). 1175000. [53-06-5]. methanol.
m-Cresol purple solution. 1121701.
mp : 223-228 °C.
Dissolve 0.1 g of m-cresol purple R in 13 mL of 0.01 M
Cortisone acetate. 1097800. [50-04-4]. sodium hydroxide, dilute to 100 mL with water R and mix.
See Cortisone acetate (0321). Colour change: pH 1.2 (red) to pH 2.8 (yellow) ; pH 7.4
(yellow) to pH 9.0 (purple).
Corydaline. C22H27NO4. (Mr 369.4). 1204400. [518-69-4].
(13S,13aR)-5,8,13,13a-Tetrahydro-2,3,9,10-tetramethoxy-13- Cresol red. C21H18O5S. (Mr 382.4). 1022800. [1733-12-6].
methyl-6H-dibenzo[a,g]quinolizine. Cresolsulfonphthalein. 4,4′-(3H-2,1-Benzoxathiol-3-
ylidene)bis-(2-methylphenol) S,S-dioxide.
Costunolide. C15H20O2. (Mr 232.3). 1194600. [553-21-9].
(3aS,6E,10E,11aR)-6,10-Dimethyl-3-methylene-3a,4,5,8,9,11a- A reddish-brown crystalline powder, slightly soluble in water,
hexahydrocyclodeca[b]furan-2(3H)-one. soluble in ethanol (96 per cent) and in dilute solutions of
alkali hydroxides.
Coumaphos. C14H16ClO5PS. (Mr 362.8). 1124800. [56-72-4].
Cresol red solution. 1022801.
mp : 91 °C to 92 °C.
Dissolve 0.1 g of cresol red R in a mixture of 2.65 mL of
A suitable certified reference solution (10 ng/µL in iso-octane) 0.1 M sodium hydroxide and 20 mL of ethanol (96 per
may be used. cent) R and dilute to 100 mL with water R.
o-Coumaric acid. C9H8O3. (Mr 164.2). 1157400. [614-60-8]. Test for sensitivity. A mixture of 0.1 mL of the cresol
(E)-2-Hydroxycinnamic acid. (2E)-3-(2-Hydroxyphenyl)prop- red solution and 100 mL of carbon dioxide-free water R
2-enoic acid. to which 0.15 mL of 0.02 M sodium hydroxide has been
White or almost white powder. added is purple-red. Not more than 0.15 mL of 0.02 M
hydrochloric acid is required to change the colour to yellow.
mp : about 217 °C.
Colour change : pH 7.0 (yellow) to pH 8.6 (red).
Coumarin. C9H6O2. (Mr 146.1). 1124900. [91-64-5].
2H-Chromen-2-one. 2H-1-Benzopyran-2-one. Crown-ether silica gel for chiral separation. 1192400.
Colourless, crystalline powder or orthorhombic or rectangular A very finely divided silica gel for chromatography coated
crystals, very soluble in boiling water, soluble in ethanol with the following chiral crown ether :
(96 per cent). It dissolves in solutions of alkali hydroxides.
mp : 68 °C to 70 °C.
Coumarin used in gas chromatography complies with the
Cresol. C7H8O. (Mr 108.1). 1022700. [95-48-7]. o-Cresol.
2-Methylphenol. (Ra)-6,23-Diphenyl-8,9,11,12,14,15,17,18,20,21-
Crystals or a super-cooled liquid becoming dark on exposure decahydrodinaphtho[2,1-q:1′,2′-s][1,4,7,10,13,16]-
to light and air, miscible with anhydrous ethanol, soluble hexaoxacycloicosine.
in about 50 parts of water and soluble in solutions of alkali
hydroxides. Crystal violet. C25H30ClN3. (Mr 408.0). 1022900. [548-62-9].
: about 1.05. Schultz No. 78.
: 1.540 to 1.550. Colour Index No. 42555.
Hexamethyl-pararosanilinium chloride.
bp : about 190 °C.
Dark-green powder or crystals, soluble in water and in ethanol
Freezing point (2.2.18) : minimum 30.5 °C. (96 per cent).
Residue on evaporation : maximum 0.1 per cent m/m,
determined by evaporating on a water-bath and drying in an Crystal violet solution. 1022901.
oven at 100-105 °C. Dissolve 0.5 g of crystal violet R in anhydrous acetic acid R
Storage : protected from light, moisture and oxygen. and dilute to 100 mL with the same solvent.
Distil before use. Test for sensitivity. To 50 mL of anhydrous acetic acid R
add 0.1 mL of the crystal violet solution. On addition of
m-Cresol. 1177100. [108-39-4]. 0.1 mL of 0.1 M perchloric acid the bluish-purple solution
See metacresol (2077). turns bluish-green.

Cupric chloride. CuCl2,2H2O. (Mr 170.5). 1023000. Mix 1 part of solution A with 25 parts of solution B
[10125-13-0]. Cupric chloride dihydrate. immediately before use.
Greenish-blue powder or crystals, deliquescent in moist air,
efflorescent in dry air, freely soluble in water, in ethanol Curcumin. C21H20O6. (Mr 368.4). 1023500. [458-37-7]. 1,7-
(96 per cent) and in methanol, sparingly soluble in acetone. Bis(4-hydroxy-3-methoxyphenyl)hepta-1,6-diene-3,5-dione.
Storage : in an airtight container. Orange-brown, crystalline powder, practically insoluble in
water, soluble in glacial acetic acid.
Cupri-citric solution. 1023100. mp : about 183 °C.
Dissolve 25 g of copper sulfate pentahydrate R, 50 g of
citric acid monohydrate R and 144 g of anhydrous sodium Curcuminoids. 1183900.
carbonate R in water R and dilute to 1000 mL with the same A mixture of curcumin (C21H20O6 ; Mr 368.4),
solvent. demethoxycurcumin (C20H18O5 ; Mr 338.4) and
Cupri-citric solution R1. 1023200. bis-demethoxycurcumin (C19H16O4 ; Mr 308.3).
Dissolve 25 g of copper sulfate pentahydrate R, 50 g of Cyanoacetic acid. C3H3NO2. (Mr 85.1). 1097900. [372-09-8].
citric acid monohydrate R and 144 g of anhydrous sodium
White or yellowish-white, hygroscopic crystals, very soluble in
carbonate R in water R and dilute to 1000 mL with the same
water.
solvent.
Adjust the solution so that it complies with the following Storage : in an airtight container.
requirements. Cyanocobalamin. 1023600. [68-19-9].
a) To 25.0 mL add 3 g of potassium iodide R. Add 25 mL of a See Cyanocobalamin (0547).
25 per cent m/m solution of sulfuric acid R with precaution
and in small quantities. Titrate with 0.1 M sodium thiosulfate Cyanogen bromide solution. 1023700. [506-68-3].
using 0.5 mL of starch solution R, added towards the end of
Add dropwise, with cooling 0.1 M ammonium thiocyanate to
the titration, as indicator.
bromine water R until the yellow colour disappears. Prepare
24.5 mL to 25.5 mL of 0.1 M sodium thiosulfate is used in the immediately before use.
titration.
b) Dilute 10.0 mL to 100.0 mL with water R and mix. To Cyanoguanidine. C2H4N4. (Mr 84.1). 1023800. [461-58-5].
10.0 mL of the solution, add 25.0 mL of 0.1 M hydrochloric Dicyandiamide. 1-Cyanoguanidine.
acid and heat for 1 h on a water-bath. Cool, adjust with White or almost white, crystalline powder, sparingly soluble
water R to the initial volume and titrate with 0.1 M sodium in water and in ethanol (96 per cent), practically insoluble in
hydroxide, using 0.1 mL of phenolphthalein solution R1 as methylene chloride.
indicator. mp : about 210 °C.
5.7 mL to 6.3 mL of 0.1 M sodium hydroxide is used in the
titration. Cyanopropyl(3)phenyl(3)methyl(94)polysiloxane.
c) Dilute 10.0 mL to 100.0 mL with water R and mix. Titrate 1114800.
10.0 mL of the solution with 0.1 M hydrochloric acid, using Polysiloxane substituted with 3 per cent of cyanopropyl
0.1 mL of phenolphthalein solution R1 as indicator. groups, 3 per cent of phenyl groups and 94 per cent of methyl
6.0 mL to 7.5 mL of 0.1 M hydrochloric acid is used in the groups.
titration.
Cyanopropyl(7)phenyl(7)methyl(86)polysiloxane.
Cupriethylenediamine hydroxide solution. 3008700. 1109200.
[14552-35-3]. Polysiloxane substituted with 7 per cent of cyanopropyl
The molar ratio of ethylenediamine to copper is 2.00 ± 0.04. groups, 7 per cent of phenyl groups and 86 per cent of methyl
This solution is commercially available. groups.
Cupri-tartaric solution. 1023300. Cyanopropyl(25)phenyl(25)methyl(50)polysiloxane.
Solution A. Dissolve 34.6 g of copper sulfate pentahydrate R in 1066500.
water R and dilute to 500 mL with the same solvent. Polysiloxane substituted with 25 per cent of cyanopropyl
Solution B. Dissolve 173 g of sodium potassium tartrate R groups, 25 per cent of phenyl groups and 50 per cent of methyl
and 50 g of sodium hydroxide R in 400 mL of water R. Heat groups.
to boiling, allow to cool and dilute to 500 mL with carbon
dioxide-free water R. Cyanopropylpolysiloxane. 1066700.
Mix equal volumes of the 2 solutions immediately before use. Polysiloxane substituted with 100 per cent of cyanopropyl
groups.
Cupri-tartaric solution R2. 1023302.
Add 1 mL of a solution containing 5 g/L of copper sulfate Cyasterone. C29H44O8. (Mr 520.7). 1204500. [17086-76-9].
pentahydrate R and 10 g/L of potassium tartrate R to 50 mL of (2β,3β,5β,22R,24S,241R,25S)-241,26-Epoxy-2,3,14,20,22-
sodium carbonate solution R1. Prepare immediately before use. pentahydroxystigmast-7-ene-6,26-dione.
Cupri-tartaric solution R3. 1023303. α-Cyclodextrin. C36H60O30. (Mr 972). 1176200.
Prepare a solution containing 10 g/L of copper sulfate [10016-20-3]. Cyclohexakis-(1→4)-(α-D-glucopyranosyl).
pentahydrate R and 20 g/L of sodium tartrate R. To 1.0 mL Cyclomaltohexaose. Alfadex.
of the solution add 50 mL of sodium carbonate solution R2.
Prepare immediately before use. β-Cyclodextrin. 1184000. [7585-39-9].
See Betadex (1070).
Cupri-tartaric solution R4. 1023304.
Solution A. 150 g/L copper sulfate pentahydrate R. β-Cyclodextrin for chiral chromatography, modified.
Solution B. Dissolve 2.5 g of anhydrous sodium carbonate R, 1154600.
2.5 g of sodium potassium tartrate R, 2.0 g of sodium hydrogen 30 per cent of 2,3-di-O-ethyl-6-O-tert-butyldimethylsilyl-β-
carbonate R, and 20.0 g of anhydrous sodium sulfate R in cyclodextrin dissolved in polysiloxane substituted with 15 per
water R and dilute to 100 mL with the same solvent. cent of phenyl groups and 85 per cent of methyl groups.

β-Cyclodextrin for chiral chromatography, modified R1. Assay. Gas chromatography (2.2.28) as prescribed in the
1160700. monograph Peppermint oil (0405).
30 per cent of 2,3-di-O-acetyl-6-O-tert-butylsilyl-β- Test solution. The substance to be examined.
cyclodextrin dissolved in polysiloxane substituted with 15 per Content : minimum 96.0 per cent, calculated by the
cent of phenyl groups and 85 per cent of methyl groups. normalisation procedure.
Cyclohexane. C6H12. (Mr 84.2). 1023900. [110-82-7]. Cynarin. C25H24O12. (Mr 516.4). 1159300. [30964-13-7].
Clear, colourless, flammable liquid, practically insoluble in (1α,3α,4α,5β)-1,3-Bis[[3-(3,4-Dihydroxyphenyl)-1-oxo-2-
water, miscible with organic solvents. propenyl]oxy]-4,5-dihydroxycyclohexanecarboxylic acid.
: about 0.78. White or almost white amorphous mass, odourless.
bp : about 80.5 °C. Cypermethrin. C22H19Cl2NO3. (Mr 416.3). 1125100.
Cyclohexane used in spectrophotometry complies with the [52315-07-8].
following additional test. bp : 170 °C to 195 °C.
Absorbance (2.2.25) : maximum 0.35 at 220 nm, 0.16 at mp : 60 °C to 80 °C.
235 nm, 0.05 at 240 nm, 0.01 at 250 nm, determined using A suitable certified reference solution (10 ng/µL in
water R as compensation liquid. cyclohexane) may be used.
Cyclohexane R1. 1023901. L-Cysteine. C3H7NO2S. (Mr 121.1). 1024200. [52-90-4].
Complies with the requirements prescribed for Powder, freely soluble in water, in ethanol (96 per cent) and in
cyclohexane R with the following additional requirement. acetic acid, practically insoluble in acetone.
The fluorescence, measured at 460 nm, under illumination
Cysteine hydrochloride. 1024300. [7048-04-6].
with an excitant light beam at 365 nm, is not more intense
than that of a solution containing 0.002 ppm of quinine R See Cysteine hydrochloride monohydrate (0895).
in dilute sulfuric acid R1. L-Cystine. C6H12N2O4S2. (Mr 240.3). 1024400. [56-89-3].
Cyclohexylamine. C6H13N. (Mr 99.2). 1024000. [108-91-8]. White or almost white, crystalline powder, practically
Cyclohexanamine. insoluble in water and in ethanol (96 per cent). It dissolves in
Colourless liquid, soluble in water, miscible with usual organic dilute solutions of alkali hydroxides.
solvents. : − 218 to − 224, determined in 1 M hydrochloric acid.
: about 1.460. mp : 250 °C, with decomposition.
bp : 134 °C to 135 °C. Cytosine. C4H5N3O. (Mr 111.1). 1160800. [71-30-7].
Cyclohexylenedinitrilotetra-acetic acid. C14H22N2O8,H2O. Content : minimum 95.0 per cent.
(Mr 364.4). 1024100. trans-Cyclohexylene-1,2-dinitrilo- Daidzein. C15H10O4. (Mr 254.2). 1178400. [486-66-8].
N,N,N’,N’-tetra-acetic acid. 7-Hydroxy-3-(4-hydroxyphenyl)-4H-1-benzopyran-4-one.
White or almost white, crystalline powder.
Daidzin. C21H20O9. (Mr 416.4). 1178300. [552-66-9].
mp : about 204 °C. Daidzein-7-O-glucoside. 7-(β-D-Glucopyranosyloxy)-3-(4-
Cyclohexylmethanol. C7H14O. (Mr 114.2). 1135200. hydroxyphenyl)-4H-1-benzopyran-4-one.
[100-49-2]. Cyclohexylcarbinol. Dantron. C14H8O4. (Mr 240.2). 1024500. [117-10-2].
Liquid with a slight odour of camphor, soluble in ethanol 1,8-Dihydroxyanthraquinone. 1,8-Dihydroxyanthracene-9,10-
(96 per cent). dione.
: about 1.464. Crystalline orange powder, practically insoluble in water,
bp : about 185 °C. slightly soluble in ethanol (96 per cent), soluble in solutions of
alkali hydroxides.
3-Cyclohexylpropionic acid. C9H16O2. (Mr 156.2). 1119200. mp : about 195 °C.
[701-97-3].
Clear liquid. o,p′-DDD. C14H10Cl4. (Mr 320.0). 1125200. [53-19-0].
1-(2-Chlorophenyl)-1-(4-chlorophenyl)-2,2-dichloroethane.
: about 0.998.
: about 1.4648. cyclohexane) may be used.
bp : about 130 °C.
p,p′-DDD. C14H10Cl4. (Mr 320.0). 1125300. [72-54-8].
Cyhalothrin. C23H19ClF3NO3. (Mr 449.9). 1125000. 1,1-Bis(4-chlorophenyl)-2,2-dichloroethane.
[91465-08-6]. bp : about 193 °C.
bp : 187 °C to 190 °C. mp : about 109 °C.
mp : about 49 °C. A suitable certified reference solution (10 ng/µL in
A suitable certified reference solution (10 ng/µL in cyclohexane) may be used.
cyclohexane) may be used.
o,p′-DDE. C14H8Cl4. (Mr 318.0). 1125400. [3424-82-6].
p-Cymene. C10H14. (Mr 134.2). 1113400. [99-87-6]. 1-(2-Chlorophenyl)-1-(4-chlorophenyl)-2,2-dichloroethylene.
1-Isopropyl-4-methylbenzene. A suitable certified reference solution (10 ng/µL in
Colourless liquid, practically insoluble in water, soluble in cyclohexane) may be used.
ethanol (96 per cent). p,p′-DDE. C14H8Cl4. (Mr 318.0). 1125500. [72-55-9].
: about 0.858. 1,1-Bis(4-chlorophenyl)-2,2-dichloroethylene.
: about 1.4895. bp : 316 °C to 317 °C.
bp : 175 °C to 178 °C. mp : 88 °C to 89 °C.
p-Cymene used in gas chromatography complies with the A suitable certified reference solution (10 ng/µL in
following additional test. cyclohexane) may be used.

o,p′-DDT. C14H9Cl5. (Mr 354.5). 1125600. [789-02-6]. 4-Deoxypyridoxine hydrochloride. C8H12NO2Cl.

1-(2-Chlorophenyl)-1-(4-chlorophenyl)-2,2,2-trichloroethane. (Mr 189.6). 1175500. [148-51-6]. 5-(Hydroxymethyl)-2,4-
A suitable certified reference solution (10 ng/µL in dimethylpyridin-3-ol.
cyclohexane) may be used. Desmethylmisonidazole. C6H9N3O4. (Mr 187.2). 1185600.
p,p′-DDT. C14H9Cl5. (Mr 354.5). 1125700. [50-29-3]. [13551-92-3]. (2RS)-3-(2-Nitro-1H-imidazol-1-yl)propane-
1,1-Bis(4-chlorophenyl)-2,2,2-trichloroethane. 1,2-diol.
bp : about 260 °C. Content : minimum 95 per cent.
mp : 108 °C to 109 °C. Yellow powder.
A suitable certified reference solution (10 ng/µL in Destaining solution. 1012202.
cyclohexane) may be used. A mixture consisting of 1 volume of glacial acetic acid R,
Decanal. C10H20O. (Mr 156.3). 1149200. [112-31-2]. Decyl 4 volumes of methanol R and 5 volumes of water R.
aldehyde. Deuterated acetic acid. C22H4O2. (Mr 64.1). 1101100.
Oily, colourless liquid, practically insoluble in water. [1186-52-3]. Tetradeuteroacetic acid. Acetic-d3 acid-d.
Decanal used in gas chromatography complies with the Degree of deuteration : minimum 99.7 per cent.
following additional test. : about 1.12.
Assay. Gas chromatography (2.2.28) as prescribed in the : about 1.368.
monograph Sweet orange oil (1811).
bp : about 115 °C.
mp : about 16 °C.
Deuterated acetone. C32H6O. (Mr 64.1). 1024900. [666-52-4].
Decane. C10H22. (Mr 142.3). 1024600. [124-18-5]. Acetone-d6. (2H6)-Acetone.
Colourless liquid, practically insoluble in water. Degree of deuteration : minimum 99.5 per cent.
: about 1.411. Clear, colourless liquid, miscible with water, with
bp : about 174 °C. dimethylformamide, with anhydrous ethanol and with
Decanol. C10H22O. (Mr 158.3). 1024700. [112-30-1]. methanol.
Decan-1-ol. : about 0.87.
Viscous liquid, solidifying at about 6 °C, practically insoluble : about 1.357.
in water, soluble in ethanol (96 per cent). bp : about 55 °C.
: about 1.436. Water and deuterium oxide. Not more than 0.1 per cent.
bp : about 230 °C. Deuterated acetonitrile. C22H3N. (Mr 44.1). 1173100.
Dehydrocostus lactone. C15H18O2. (Mr 230.3). [2206-26-0].
1194700. [477-43-0]. (3aS,6aR,9aR,9bS)-3,6,9- Degree of deuteration : minimum 99.8 per cent.
Trismethylenedecahydroazuleno[4,5-b]furan-2(3H)-one. Clear, colourless liquid, miscible with water, with acetone and
with methanol.
Deltamethrin. C22H19Br2NO3. (Mr 505.2). 1125800.
[52918-63-5]. : about 0.78.
bp : about 300 °C. : about 1.344.
mp : about 98 °C. Deuterated chloroform. C2HCl3. (Mr 120.4). 1025000.
A suitable certified reference solution (10 ng/µL in [865-49-6]. (2H)-Chloroform. Chloroform-d.
cyclohexane) may be used. Degree of deuteration : minimum 99.7 per cent.
Demeclocycline hydrochloride. 1145600. Clear, colourless liquid, practically insoluble in water, miscible
with acetone and with ethanol (96 per cent). It may be
See Demeclocycline hydrochloride (0176). stabilised over silver foil.
Demethylflumazenil. C14H12FN3O3. (Mr 289.3). 1149300. : about 1.51.
[79089-72-8]. Ethyl 8-fluoro-6-oxo-5,6-dihydro-4H- : about 1.445.
imidazo[1,5-a][1,4]benzodiazepine-3-carboxylate. bp : about 60 °C.
Colourless needles, soluble in dimethyl sulfoxide and in hot Water and deuterium oxide : maximum 0.05 per cent.
methanol.
mp : about 288 °C. Deuterated dimethyl sulfoxide. C22H6OS. (Mr 84.2).
1025100. [2206-27-1]. (2H6)-Dimethyl sulfoxide. Dimethyl
14-Deoxy-11,12-didehydroandrographolide. C20H28O4. sulfoxide-d6.
(Mr 332.4). 1198300. [42895-58-9]. 3-[(1E)-2- Degree of deuteration : minimum 99.8 per cent.
[(1R,4aS,5R,6R,8aR)-6-Hydroxy-5-(hydroxymethyl)-
5,8a-dimethyl-2-methylenedecahydronaphthalen-1- Very hygroscopic liquid, practically colourless, viscous, soluble
yl]ethenyl]furan-2(5H)-one. in water, in acetone and in anhydrous ethanol.
: about 1.18.
2-Deoxy- D-ribose. C5H10O4. (Mr 134.1). 1163900. mp : about 20 °C.
[533-67-5]. Thyminose. 2-Deoxy-D-erythro-pentose.
Water and deuterium oxide : maximum 0.1 per cent.
2′-Deoxyuridine. C9H12N2O5. (Mr 228.2). 1024800. Storage : in an airtight container.
[951-78-0]. 1-(2-Deoxy-β-d-erythro-pentofuranosyl)-1H,3H-
pyrimidine-2,4-dione. Deuterated methanol. C2H4O. (Mr 36.1). 1025200.
mp : about 165 °C. [811-98-3]. (2H)-Methanol. Methanol-d.
Chromatography. Thin-layer chromatography (2.2.27) as Degree of deuteration : minimum 99.8 per cent.
prescribed in the monograph Idoxuridine (0669) : apply 5 µL Clear, colourless liquid miscible with water, with ethanol
of a 0.25 g/L solution ; the chromatogram shows only one (96 per cent) and with methylene chloride.
principal spot. : about 0.888.

: about 1.326. Diatomaceous earth. 1025900. [91053-39-3].

bp : 65.4 °C. White or almost white, fine granular powder, made up of
siliceous frustules of fossil diatoms or of debris of fossil
Deuterated sodium trimethylsilylpropionate. diatoms, practically insoluble in water and in ethanol (96 per
C6H92H4NaO2Si. (Mr 172.3). 1179100. [24493-21-8]. Sodium cent).
3-(trimethylsilyl)(2,2,3,3-2H4)propionate. TSP-d4.
The substance may be identified by microscopic examination
Degree of deuteration : minimum 98 per cent. with a magnification of × 500.
Diatomaceous earth for gas chromatography. 1026000.
Deuterium chloride. 2HCl. (Mr 37.47). 1178800. [7698-05-7]. White or almost white, fine granular powder, practically
Deuterated hydrochloric acid. insoluble in water and in ethanol (96 per cent), made up
Gas. of siliceous frustules of fossil diatoms or of debris of fossil
diatoms. The substance may be identified by microscopic
Degree of deuteration : minimum 99 per cent. examination with a magnification of × 500. The substance is
Caution : toxic. acid-washed, then water-washed until neutral.
Deuterium chloride solution. 1178801.
Dilute 1 mL of deuterium chloride R (38 per cent m/m)

with 5 mL of deuterium oxide R. Diatomaceous earth for gas chromatography, silanised.
1026300.
Deuterium oxide. 2H2O. (Mr 20.03). 1025300. [7789-20-0]. Diatomaceous earth for gas chromatography R silanised with
Deuterated water. dimethyldichlorosilane or other suitable silanising agents.
Degree of deuteration : minimum 99.7 per cent.
: about 1.11. Diazinon. C12H21N2O3PS. (Mr 304.3). 1125900. [333-41-5].

: about 1.328. bp : about 306 °C.
bp : about 101 °C. A suitable certified reference solution (10 ng/µL in iso-octane)
may be used.
Deuterium oxide R1. 2H2O. (Mr 20.03). 1025301.
[7789-20-0]. Deuterated water. Diazobenzenesulfonic acid solution R1. 1026500.
Degree of deuteration : minimum 99.95 per cent. Dissolve 0.9 g of sulfanilic acid R in a mixture of 30 mL of
dilute hydrochloric acid R and 70 mL of water R. To 3 mL of
Developer solution. 1122500. the solution add 3 mL of a 50 g/L solution of sodium nitrite R.
Dilute 2.5 mL of a 20 g/L solution of citric acid monohydrate R Cool in an ice-bath for 5 min, add 12 mL of the sodium nitrite
and 0.27 mL of formaldehyde R to 500.0 mL with water R. solution and cool again. Dilute to 100 mL with water R and
keep the reagent in an ice-bath. Prepare extemporaneously
Dextran for chromatography, cross-linked R2. 1025500. but allow to stand for 15 min before use.
Bead-form dextran with a fraction range suitable for the Dibromomethane. CH2Br2. (Mr 173.8). 1195500. [74-95-3].
separation of peptides and proteins with relative molecular
masses of 15 × 10 to 30 × 10 . When dry, the beads have a
2 3 Colourless liquid, slightly soluble in water.
diameter of 20-80 µm. bp : about 96 °C.
Dextran for chromatography, cross-linked R3. 1025600. Dibutylamine. C8H19N. (Mr 129.3). 1126000. [111-92-2].
Bead-form dextran with a fraction range suitable for the N-Butylbutan-1-amine.
separation of peptides and proteins with relative molecular Colourless liquid.
masses of 4 × 103 to 15 × 104. When dry, the beads have a : about 1.417.
diameter of 40-120 µm.
bp : about 159 °C.
Dextrose. 1025700. [50-99-7].
Dibutylammonium phosphate for ion-pairing. 1168800.
See glucose R.
A colourless solution of 10 per cent to 15 per cent V/V
3,3′-Diaminobenzidine tetrahydrochloride. of di-n-butylamine and 12 per cent to 17 per cent V/V of
C12H18Cl4N4, 2H2O. (Mr 396.1). 1098000. [7411-49-6]. phosphoric acid in water, suitable for ion-pairing in liquid
3,3′,4,4′-Biphenyl-tetramine. chromatography.
Almost white or slightly pink powder, soluble in water. Dibutyl ether. C8H18O. (Mr 130.2). 1026700. [142-96-1].
mp : about 280 °C, with decomposition. Colourless, flammable liquid, practically insoluble in water,
1,2-Diamino-4,5-methylenedioxybenzene dihydro- miscible with anhydrous ethanol.
chloride. C7H10Cl2N2O2. (Mr 225.1). 1202100. [81864-15-5]. : about 0.77.
2H-1,3-Benzodioxole-5,6-diamine dihydrochloride. : about 1.399.
Content : minimum 99 per cent (HPLC). Do not distil if the dibutyl ether does not comply with the test
for peroxides.
Diammonium 2,2′-azinobis(3-ethylbenzothiazoline-6-
sulfonate). C18H24N6O6S4. (Mr 548.7). 1153000. [30931-67-0]. Peroxides. Place 8 mL of potassium iodide and starch solution R
ABTS. Diammonium 2,2′-(diazanediylidene)bis[3-ethyl-2,3- in a 12 mL ground-glass-stoppered cylinder about 1.5 cm in
dihydrobenzothiazole-6-sulfonate]. diameter. Fill completely with the substance to be examined,
shake vigorously and allow to stand protected from light for
Chromogenic substrate suitable for use in ELISA procedures. 30 min. No colour is produced.
Green tablets, freely soluble in water. The name and concentration of any added stabiliser are stated
pH (2.2.3): 4.2 to 5.8 for a 0.1 g/L solution. on the label.

Dibutyl phthalate. C16H22O4. (Mr 278.3). 1026800. [84-74-2]. 2,6-Dichlorophenol. C6H4Cl2O. (Mr 163.0). 1177600.
Dibutyl benzene-1,2-dicarboxylate. [87-65-0].
Clear, colourless or faintly coloured, oily liquid, very slightly mp : 64 °C to 66 °C.
soluble in water, miscible with acetone and with ethanol
(96 per cent). Dichlorophenolindophenol, sodium salt.
C12H6Cl2NNaO2,2H2O. (Mr 326.1). 1027300. [620-45-1].
: 1.043 to 1.048. The sodium derivative of 2,6-dichloro-N-(4-hydroxy-
: 1.490 to 1.495. phenyl)-1,4-benzoquinone monoimine dihydrate.
Dicarboxidine hydrochloride. C20H26Cl2N2O6. (Mr 461.3). Dark-green powder, freely soluble in water and in anhydrous
1026900. [56455-90-4]. 4,4′-[(4,4′-Diaminobiphenyl-3,3′- ethanol. The aqueous solution is dark blue ; when acidified
diyl)dioxy]dibutanoic acid dihydrochloride. it becomes pink.
Dichlofenthion. C10H13Cl2O3PS. (Mr 315.2). 1126100. Dichlorophenolindophenol standard solution. 1027301.

[97-17-6]. Dissolve 50.0 mg of dichlorophenolindophenol, sodium
A suitable certified reference solution (10 ng/µL in salt R in 100.0 mL of water R and filter.
cyclohexane) may be used. Assay. Dissolve 20.0 mg of ascorbic acid R in 10 mL of a
freshly prepared 200 g/L solution of metaphosphoric acid R
Dichloroacetic acid. C2H2Cl2O2. (Mr 128.9). 1027000. and dilute to 250.0 mL with water R. Titrate 5.0 mL rapidly
[79-43-6]. with the dichloro-phenolindophenol standard solution,
Colourless liquid, miscible with water and ethanol (96 per added from a microburette graduated in 0.01 mL, until the
cent). pink colour persists for 10 s, the titration occupying not
: about 1.566. more than 2 min. Dilute the dichlorophenolindophenol
: about 1.466. solution with water R to make 1 mL of the solution
equivalent to 0.1 mg of ascorbic acid (C6H8O6).
bp : about 193 °C.
Storage : use within 3 days.
Dichloroacetic acid solution. 1027001. Standardise immediately before use.
Dilute 67 mL of dichloroacetic acid R to 300 mL with
5,7-Dichloroquinolin-8-ol. C9H5Cl2NO. (Mr 214.1).
water R and neutralise to blue litmus paper R using
1157000. [773-76-2]. 5,7-Dichlorooxine.
ammonia R. Cool, add 33 mL of dichloroacetic acid R and
dilute to 600 mL with water R. Yellow, crystalline powder, soluble in acetone, slightly soluble
3,5-Dichloroaniline. C6H5Cl2N. (Mr 162.0). 1177800. mp : about 179 °C.
[626-43-7]. 3,5-dichlorophenylamine.
mp : 46 °C to 52 °C.
Dichloroquinonechlorimide. C6H2Cl3NO. (Mr 210.4).
Dichlorobenzene. C6H4Cl2. (Mr 147.0). 1027100. [95-50-1]. 1027400. [101-38-2]. 2,6-Dichloro-N-chloro-1,4-
1,2-Dichlorobenzene. benzoquinone mono-imine.
Colourless, oily liquid, practically insoluble in water, soluble Pale yellow or greenish-yellow crystalline powder, practically
in anhydrous ethanol. insoluble in water, soluble in ethanol (96 per cent) and in
: about 1.31. dilute alkaline solutions.
2,4-Dichlorobenzoic acid. C7H4Cl2O2. (Mr 191.0). 1185700. Dichlorvos. C4H7Cl2O4P. (Mr 221). 1101200. [62-73-7].
[50-84-0]. 2,2-Dichlorovinyl dimethyl phosphate.
Faintly beige powder. Colourless or brownish-yellow liquid, soluble in water,
mp : about 160 °C. miscible with most organic solvents.
: about 1.452.
2,3-Dichloro-5,6-dicyanobenzoquinone. C8Cl2N2O2.
(Mr 227.0). 1153600. [84-58-2]. 4,5-Dichloro-3,6-dioxo- Dicyclohexyl. C12H22. (Mr 166.3). 1135300. [92-51-3].
cyclohexa-1,4-diene-1,2-dicarbonitrile. Bicyclohexyl.
Yellow or orange crystals, soluble in dioxan and in acetic acid, : about 0.864.
slightly soluble in methylene chloride. It decomposes in water. bp : about 227 °C.
mp : about 214 °C. mp : about 4 °C.
Storage : at a temperature of 2 °C to 8 °C.
Dicyclohexylamine. C12H23N. (Mr 181.3). 1027500.
(S)-3,5-Dichloro-2,6-dihydroxy-N-[(1-ethylpyrrolidin- [101-83-7]. N,N-Dicyclohexylamine.
2-yl)methyl]benzamide hydrobromide. C14H19BrCl2N2O3. Colourless liquid, sparingly soluble in water, miscible with the
(Mr 414.1). 1142600. [113310-88-6]. usual organic solvents.
White or almost white, crystalline powder. : about 1.484.
: + 11.4, determined on a 15.0 g/L solution in anhydrous bp : about 256 °C.
ethanol R. Freezing point (2.2.18) : 0 °C to 1 °C.
mp : about 212 °C.
Dicyclohexylurea. C13H24N2O. (Mr 224.4). 1027600.
Dichlorofluorescein. C20H10Cl2O5. (Mr 401.2). [2387-23-7]. 1,3-Dicyclohexylurea.
1027200. [76-54-0]. 2,7-Dichlorofluorescein. White or almost white, crystalline powder.
2-(2,7-Dichloro-6-hydroxy-3-oxo-3H-xanthen-9-yl)benzoic
acid. mp : about 232 °C.
Yellowish-brown or yellow-orange powder, slightly soluble Didocosahexaenoin. C47H68O5. (Mr 713.0). 1142700.
in water, freely soluble in ethanol (96 per cent) and in dilute [88315-12-2]. Diglyceride of docosahexaenoic acid (C22:6).
solutions of alkali hydroxides giving a solution showing a Glycerol didocosahexaenoate. (all-Z)-Docosahexaenoic acid,
yellowish-green fluorescence. diester with propane-1,2,3-triol.

Didodecyl 3,3′-thiodipropionate. C30H58O4S. (Mr 514.8). Diethylamine. C4H11N. (Mr 73.1). 1028000. [109-89-7].

1027700. [123-28-4]. Clear, colourless, flammable liquid, strongly alkaline, miscible
White or almost white, crystalline powder, practically with water and with ethanol (96 per cent).
insoluble in water, freely soluble in acetone and in light : about 0.71.
petroleum, slightly soluble in ethanol (96 per cent). bp : about 55 °C.
mp : about 39 °C.
Diethylamine R1. C4H11N. (Mr 73.1). 1028001. [109-89-7].
Dieldrin. C12H8Cl6O. (Mr 380.9). 1126200. [60-57-1]. N-Ethylethanamine.
bp : about 385 °C. Content : minimum 99.5 per cent.
mp : about 176 °C. Clear, colourless, flammable liquid, strongly alkaline,
miscible with water and with ethanol (96 per cent).
: about 0.71.
bp : about 55 °C.
Diethanolamine. C4H11NO2. (Mr 105.1). 1027800.
[111-42-2]. 2,2′-Iminobisethanol. Diethylaminoethyldextran. 1028200.
Anion-exchange resin presented as the hydrochloride.
Viscous, clear, slightly yellow liquid or deliquescent crystals
melting at about 28 °C, very soluble in water, in acetone and Powder forming gels with water.
in methanol. N,N-Diethylaniline. C10H15N. (Mr 149.2). 1028400.
: about 1.09. [91-66-7].
pH (2.2.3) : 10.0 to 11.5 for a 50 g/L solution. : about 0.938.
Diethanolamine used in the test for alkaline phosphatase bp : about 217 °C.
complies with the following additional test. mp : about − 38 °C.
Ethanolamine. Gas chromatography (2.2.28). Diethylene glycol. C4H10O3. (Mr 106.1). 1028300. [111-46-6].
Internal standard solution. Dissolve 1.00 g of 2,2′-Oxydiethanol.
3-aminopropanol R in acetone R and dilute to 10.0 mL with Content : minimum 99.5 per cent m/m.
the same solvent. Clear, colourless liquid, hygroscopic, miscible with water, with
Test solution (a). Dissolve 5.00 g of the substance to be acetone and with ethanol (96 per cent).
examined in acetone R and dilute to 10.0 mL with the same : about 1.118.
solvent. : about 1.447.
Test solution (b). Dissolve 5.00 g of the substance to be bp : 244 °C to 246 °C.
examined in acetone R, add 1.0 mL of the internal standard
solution and dilute to 10.0 mL with the same solvent.
Reference solutions. Dissolve 0.50 g of ethanolamine R in N,N-Diethylethane-1,2-diamine. C6H16N2. (Mr 116.2).
acetone R and dilute to 10.0 mL with the same solvent. To 1028500. [100-36-7]. N,N-Diethylethylenediamine.
0.5 mL, 1.0 mL and 2.0 mL of this solution, add 1.0 mL of Content : minimum 98.0 per cent.
the internal standard solution and dilute to 10.0 mL with Slightly oily liquid, colourless or slightly yellow, strong odour
acetone R. of ammonia, irritant to the skin, eyes and mucous membranes.
Column : : 0.827.
– size : l = 1 m, Ø = 4 mm ; bp : 145 °C to 147 °C.
– stationary phase : diphenylphenylene oxide polymer R Water (2.5.12) : maximum 1.0 per cent, determined on 0.500 g.
(180-250 μm).
Di(2-ethylhexyl) phthalate. C24H38O4. (Mr 390.5). 1028100.
Carrier gas : nitrogen for chromatography R. Di(2-ethylhexyl) benzene-1,2-dicarboxylate.
Flow rate : 40 mL/min. Colourless, oily liquid, practically insoluble in water, soluble
Temperature : in organic solvents.
: about 0.98.
Time Temperature
(min) (°C) : about 1.486.
Column 0→3 125 Viscosity (2.2.9) : about 80 mPa·s.
3 → 17.6 125 → 300 Diethylphenylenediamine sulfate. C10H18N2O4S. (Mr 262.3).
1028600. [6283-63-2]. N,N’-Diethyl-p-phenylenediamine
Injection port 250
sulfate. N,N’-Diethylbenzene-1,4-diamine sulfate.
Detector 280 White or slightly yellow powder, soluble in water.
Detection : flame-ionisation.
Injection : 1.0 µL.
Limit : Diethylphenylenediamine sulfate solution. 1028601.
To 250 mL of water R add 2 mL of sulfuric acid R and
– ethanolamine : maximum 1.0 per cent.
25 mL of 0.02 M sodium edetate. Dissolve in this solution
Diethoxytetrahydrofuran. C8H16O3. (Mr 160.2). 1027900. 1.1 g of diethylphenylenediamine sulfate R and dilute to
[3320-90-9]. 2,5-Diethoxytetrahydrofuran. A mixture of the 1000 mL with water R.
cis and trans isomers. Do not use if the solution is not colourless.
Clear, colourless or slightly yellowish liquid, practically Storage : protected from light and heat for 1 month.
insoluble in water, soluble in ethanol (96 per cent) and in most Diethyl sulfone. C4H10O2S. (Mr 122.2). 1203300.
other organic solvents. [597-35-3]. 1-(Ethylsulfonyl)ethane. 1-(Ethanesulfon-
: about 0.98. yl)ethane.
: about 1.418. Content : minimum 97 per cent.

Crystalline powder. 2,7-Dihydroxynaphthalene. C10H8O2. (Mr 160.2). 1029100.

mp : about 73 °C. [582-17-2]. Naphthalene-2,7-diol.
Needles, soluble in water and in ethanol (96 per cent).
Diflubenzuron. C14H9ClF2N2O2. (Mr 310.7). 1180000.
[35367-38-5]. 1-(4-Chlorophenyl)-3-(2,6-difluoro- mp : about 190 °C.
benzoyl)urea. 2,7-Dihydroxynaphthalene solution. 1029101.
Colourless or white or almost white crystals, practically Dissolve 10 mg of 2,7-dihydroxynaphthalene R in 100 mL
insoluble in water, freely soluble in dimethyl sulfoxide, slightly of sulfuric acid R and allow to stand until decolorised.
soluble in acetone. Storage : use within 2 days.
mp : 230 to 232 °C.
5,7-Diiodoquinolin-8-ol. C9H5I2NO. (Mr 397.0). 1157100.
Digitonin. C56H92O29. (Mr 1229). 1028700. [11024-24-1]. [83-73-8]. 5,7-Diiodooxine.
3β-[O-β-D-Glucopyranosyl-(1→3)-O-β-D-galactopyranosyl- Yellowish-brown powder, sparingly soluble in acetone and in
(1→2)-O-[β-D-xylopyranosyl-(1→3)]-O-β-D-galactopyranosyl- ethanol (96 per cent).
(1→4)-O-β-D-galactopyranosyloxy]-(25R)-5α-spirostan-
2α,15β-diol. Content : minimum 95.0 per cent.
Crystals, practically insoluble in water, sparingly soluble in Di-isobutyl ketone. C9H18O. (Mr 142.2). 1029200. [108-83-8].
anhydrous ethanol, slightly soluble in ethanol (96 per cent). Clear, colourless liquid, slightly soluble in water, miscible with
Digitoxin. 1028800. [71-63-6]. most organic solvents.
See Digitoxin (0078). : about 1.414
bp : about 168 °C.
Diglycine. C4H8N2O3. (Mr 132.1). 1191700. [556-50-3].
2-[(2-Aminoacetyl)amino]acetic acid. Glycylglycine. Di-isopropyl ether. C6H14O. (Mr 102.2). 1029300. [108-20-3].
Digoxin. 1203400. Clear, colourless liquid, very slightly soluble in water, miscible
with ethanol (96 per cent).
See Digoxin (0079).
: 0.723 to 0.728.
Dihydrocapsaicin. C18H29NO3. (Mr 307.4). 1148100. bp : 67 °C to 69 °C.
[19408-84-5]. N-[(4-Hydroxy-3-methoxyphenyl)methyl]-8- Do not distil if the di-isopropyl ether does not comply with the
methylnonanamide. test for peroxides.
White or almost white, crystalline powder, practically Peroxides. Place 8 mL of potassium iodide and starch solution R
insoluble in cold water, freely soluble in anhydrous ethanol. in a 12 mL ground-glass-stoppered cylinder about 1.5 cm in
10,11-Dihydrocarbamazepine. C15H14N2O. diameter. Fill completely with the substance to be examined,
(Mr 238.3). 1028900. [3564-73-6]. 10,11-Dihydro- shake vigorously and allow to stand protected from light for
5H-dibenzo[b,f]azepine-5-carboxamide. 30 min. No colour is produced.
mp : 205 °C to 210 °C. The name and concentration of any added stabiliser are stated
on the label.
Dihydrocarvone. C10H16O. (Mr 152.2). 1160900. Storage : protected from light.
[7764-50-3]. p-Menth-8-en-2-one. 2-Methyl-5-(1-
methylethenyl)cyclohexanone. N,N-Diisopropylethylamine. C8H19N. (Mr 129.2). 1204600.
Dihydrocarvone used in gas chromatography complies with the [7087-68-5]. N-Ethyl-N-(propan-2-yl)propan-2-amine.
following additional test. N-Ethyldiisopropylamine.
Assay. Gas chromatography (2.2.28) as prescribed in the Clear, colourless or light yellow liquid.
test for chromatographic profile in the monograph Caraway bp : 127 °C.
oil (1817).
N,N′-Diisopropylethylenediamine. C8H20N2. (Mr 144.3).
Content calculated by the normalisation procedure : 1140600. [4013-94-9]. N,N′-Bis(1-methylethyl)-1,2-
– major component (trans-dihydrocarvone) : minimum 70 per ethanediamine.
cent ; Colourless or yellowish, corrosive, flammable, hygroscopic
– sum of cis- and trans-dihydrocarvone : minimum 98 per liquid.
cent. : about 0.798.
2,5-Dihydroxybenzoic acid. C7H6O4. (Mr 154.1). 1148200. : about 1.429.
[490-79-9]. Gentisic acid. bp : about 170 °C.
Light yellow crystals.
4,4′-Dimethoxybenzophenone. C15H14O3. (Mr 242.3).
mp : about 200 °C. 1126300. [90-96-0]. Bis(4-methoxyphenyl)methanone.
5,7-Dihydroxy-4-methylcoumarin. C10H8O4. (Mr 192.2). White or almost white powder, practically insoluble in water
1149400. [2107-76-8]. 5,7-Dihydroxy-4-methyl-2H-1- and slightly soluble in ethanol (96 per cent).
benzopyran-2-one. mp : about 142 °C.
Light yellowish powder, practically insoluble in water,
sparingly soluble in ethanol (96 per cent). 3,4-Dimethoxy- L-phenylalanine. C11H15NO4.
(Mr 225.2). 1191800. [32161-30-1]. (2S)-2-Amino-3-(3,4-
mp : 295 °C to 303 °C. dimethoxyphenyl)propanoic acid.
Dihydroxynaphthalene. 1029000. [132-86-5]. Content : minimum 95 per cent.
See 1,3-dihydroxynaphthalene R. White or almost white powder.
1,3-Dihydroxynaphthalene. C10H8O2. (Mr 160.2). 1029000. Dimethoxypropane. C5H12O2. (Mr 104.1). 1105200.
[132-86-5]. Naphthalene-1,3-diol. [77-76-9]. 2,2-Dimethoxypropane.
Crystalline, generally brownish-violet powder, freely soluble Colourless liquid, decomposing on exposure to moist air or
in water and in ethanol (96 per cent). water.
mp : about 125 °C. : about 0.847.

: about 1.378. 4-Dimethylaminocinnamaldehyde. C11H13NO.

bp : about 83 °C. (Mr 175.2). 1029900. [6203-18-5]. 3-(4-Dimethylamino-
phenyl)prop-2-enal.
Dimethylacetamide. C4H9NO. (Mr 87.1). 1029700. Orange or orange-brown crystals or powder. Sensitive to light.
[127-19-5]. N,N-Dimethylacetamide.
mp : about 138 °C.
Colourless liquid, miscible with water and with many organic 4-Dimethylaminocinnamaldehyde solution. 1029901.
solvents. Dissolve 2 g of 4-dimethylaminocinnamaldehyde R in a
: about 0.94. mixture of 100 mL of hydrochloric acid R1 and 100 mL of
anhydrous ethanol R. Dilute the solution to four times its
: about 1.437. volume with anhydrous ethanol R immediately before use.
bp : about 165 °C.
Dimethylaminoethanol. C4H11NO. (Mr 89.1). 1195600.
Dimethylamine. C2H7N. (Mr 45.08). 1168900. [124-40-3]. [108-01-0]. 2-(Dimethylamino)ethan-1-ol.
N-Methylmethanamine. Colourless or slightly yellow liquid, miscible with water.
Colourless, flammable gas. bp : about 135 °C.
bp : about 7 °C.
2-(Dimethylamino)ethyl methacrylate. C8H15NO2.
mp : about − 92.2 °C. (Mr 157.2). 1147200. [2867-47-2]. 2-(Dimethylamino)ethyl
Dimethylamine solution. 1168901. 2-methylpropenoate.
A 400 g/L solution of dimethylamine R. d 420 : about 0.930.
Clear, colourless solution. bp : about 187 °C.
Density : about 0.89. Dimethylaminonaphthalenesulfonyl chloride.
bp : about 54 °C. C12H12ClNO2S. (Mr 269.8). 1030000. [605-65-2].
mp : about − 37 °C. 5-Dimethyl-amino-1-naphthalenesulfonyl chloride.
Yellow, crystalline powder, slightly soluble in water, soluble
Dimethylaminobenzaldehyde. C9H11NO. (Mr 149.2). in methanol.
1029800. [100-10-7]. 4-Dimethylaminobenzaldehyde. mp : about 70 °C.
White or yellowish-white crystals, soluble in ethanol (96 per
cent) and in dilute acids. 3-Dimethylaminophenol. C8H11NO. (Mr 137.2). 1156500.
mp : about 74 °C. [99-07-0]. 3-(Dimethylamino)phenol.
Grey powder, slightly soluble in water.
Dimethylaminobenzaldehyde solution R1. 1029801. mp : about 80 °C.
Dissolve 0.2 g of dimethylaminobenzaldehyde R in 20 mL
of ethanol (96 per cent) R and add 0.5 mL of hydrochloric 2-(Dimethylamino)thioacetamide hydrochloride.
acid R. Shake the solution with activated charcoal R and C4H11ClN2S. (Mr 154.7). 1181800. [27366-72-9].
filter. The colour of the reagent is less intense than that of
iodine solution R3. Prepare immediately before use. Dimethylaniline. 1030100. [121-69-7].
See N,N-Dimethylaniline R.
Dimethylaminobenzaldehyde solution R2. 1029802.
Dissolve 0.2 g of dimethylaminobenzaldehyde R, without N,N-Dimethylaniline. C8H11N. (Mr 121.2). 1030100.
heating, in a mixture of 4.5 mL of water R and 5.5 mL of [121-69-7].
hydrochloric acid R. Prepare immediately before use. Clear, oily liquid, almost colourless when freshly distilled,
darkening on storage to reddish-brown, practically insoluble
Dimethylaminobenzaldehyde solution R6. 1029803. in water, freely soluble in ethanol (96 per cent).
Dissolve 0.125 g of dimethylaminobenzaldehyde R in a : about 1.558.
cooled mixture of 35 mL of water R and 65 mL of sulfuric Distillation range (2.2.11). Not less than 95 per cent distils
acid R. Add 0.1 mL of a 50 g/L solution of ferric chloride R. between 192 °C and 194 °C.
Before use allow to stand for 24 h, protected from light.
Storage : when stored at room temperature, use within 2,3-Dimethylaniline. C8H11N. (Mr 121.2). 1105300.
1 week ; when stored in a refrigerator use within several [87-59-2]. 2,3-Xylidine.
months. Yellowish liquid, sparingly soluble in water, soluble in ethanol
(96 per cent).
Dimethylaminobenzaldehyde solution R7. 1029804.
: 0.993 to 0.995.
Dissolve 1.0 g of dimethylaminobenzaldehyde R in 50 mL
of hydrochloric acid R and add 50 mL of ethanol (96 per : about 1.569.
cent) R. bp : about 224 °C.
Storage : protected from light ; use within 4 weeks.
2,6-Dimethylaniline. C8H11N. (Mr 121.2). 1030200.
Dimethylaminobenzaldehyde solution R8. 1029805. [87-62-7]. 2,6-Xylidine.
Dissolve 0.25 g of dimethylaminobenzaldehyde R in a Colourless liquid, sparingly soluble in water, soluble in ethanol
mixture of 5 g of phosphoric acid R, 45 g of water R and 50 g (96 per cent).
of anhydrous acetic acid R. Prepare immediately before use. : about 0.98.
Dimethylaminobenzaldehyde solution R9. 1029806. 2,6-Dimethylaniline hydrochloride. C8H12ClN. (Mr 157.6).
Dissolve 1.0 g of dimethylaminobenzaldehyde R in 3.5 mL 1169000. [21436-98-6]. 2,6-Dimethylbenzenamide
of perchloric acid (600 g/L HCIO4) and slowly add 6.5 mL hydrochloride. 2,6-Xylidine hydrochloride.
of 2-propanol R. Prepare immediately before use. Content : minimum 98.0 per cent.

2,4-Dimethyl-6-tert-butylphenol. C12H18O. (Mr 178.3). Dimethylglyoxime. C4H8N2O2. (Mr 116.1). 1030400.

1126500. [1879-09-0]. [95-45-4]. 2,3-Butanedione dioxime.
Dimethyl carbonate. C3H6O3. (Mr 90.1). 1119300. crystals, practically insoluble in cold water, very slightly
[616-38-6]. Carbonic acid dimethyl ester. soluble in boiling water, soluble in ethanol (96 per cent).
Liquid, insoluble in water, miscible with ethanol (96 per cent). mp : about 240 °C, with decomposition.
: 1.065. Sulfated ash (2.4.14): maximum 0.05 per cent.
: 1.368.
1,3-Dimethyl-2-imidazolidinone. C5H10N2O. (Mr 114.2).
bp : about 90 °C. 1135400. [80-73-9]. N,N′-Dimethylethylene urea.
1,3-Dimethyl-2-imidazolidone.
Dimethyl-β-cyclodextrin. C56H98O35. (Mr 1331). 1169100.
[51166-71-3]. Heptakis(2,6-di-O-methyl)cyclomaltoheptaose. : 1.4720.
Cycloheptakis-(1→4)-(2,6-di-O-methyl-α-D-glucopyranosyl). bp : about 224 °C.
2A,2B,2C,2D,2E,2F,2G,6A,6B,6C,6D,6E,6F,6G-Tetradeca-O-methyl-β-
cyclodextrin. N,N-Dimethyloctylamine. C10H23N. (Mr 157.3). 1030500.
[7378-99-6]. Octyldimethylamine.
Colourless liquid.
Dimethyldecylamine. C12H27N. (Mr 185.4). 1113500. : about 0.765.
[1120-24-7]. N,N-dimethyldecylamine. : about 1.424.
Content : minimum 98.0 per cent m/m. bp : about 195 °C.
bp : about 234 °C.
2,5-Dimethylphenol. C8H10O. (Mr 122.2). 1162300.
1,1-Dimethylethylamine. C4H11N. (Mr 73.1). 1100900. [95-87-4]. p-Xylenol.
[75-64-9]. 2-Amino-2-methylpropane. tert-Butylamine. White or almost white crystals.
Liquid, miscible with ethanol (96 per cent). 2,6-Dimethylphenol. C8H10O. (Mr 122.2). 1030600.
: about 0.694. [576-26-1].
: about 1.378. Colourless needles, slightly soluble in water, very soluble in
bp : about 46 °C. ethanol (96 per cent).
bp : about 203 °C.
1,1-Dimethylethyl methyl ether. C5H12O. (Mr 88.1). mp : 46 °C to 48 °C.
1013900. [1634-04-4]. 2-Methoxy-2-methylpropane.
tert-Butyl methyl ether. 3,4-Dimethylphenol. C8H10O. (Mr 122.2). 1098100.
Colourless, clear, flammable liquid. [95-65-8].
: about 1.376. White or almost white crystals, slightly soluble in water, freely
Absorbance (2.2.25) : maximum 0.30 at 240 nm, 0.10 at
bp : about 226 °C.
255 nm, 0.01 at 280 nm, determined using water R as
compensation liquid. mp : 25 °C to 27 °C.
1,1-Dimethylethyl methyl ether R1. C5H12O. (Mr 88.1). N,N-Dimethyl- L-phenylalanine. C11H15NO2. (Mr 193.2).

1126400. [1634-04-4]. 2-Methoxy-2-methylpropane. 1164000. [17469-89-5]. (2S)-2-(Dimethylamino)-3-
tert-Butyl methyl ether. phenylpropanoic acid.
Content : minimum 99.5 per cent. mp : about 226 °C.
: about 0.741. Dimethylpiperazine. C6H14N2. (Mr 114.2). 1030700.
: about 1.369. [106-58-1]. 1,4-Dimethylpiperazine.
A colourless liquid, miscible with water and with ethanol
bp : about 55 °C.
(96 per cent).
Dimethylformamide. C3H7NO. (Mr 73.1). 1030300. : about 0.85.
[68-12-2]. : about 1.446.
Clear, colourless neutral liquid, miscible with water and with bp : about 131 °C.
Dimethylstearamide. C20H41NO. (Mr 311.6). 1030800.
: 0.949 to 0.952. N,N-Dimethylstearamide.
bp : about 153 °C. White or almost white solid mass, soluble in many organic
Water (2.5.12): maximum 0.1 per cent. solvents, including acetone.
mp : about 51 °C.
Dimethylformamide diethylacetal. C7H17NO2. (Mr 147.2).
1113600. [1188-33-6]. N,N-Dimethylformamide diethylacetal. Dimethylstearylamide. 1030800.
: about 1.40. See dimethylstearamide R.
bp : 128 °C to 130 °C. Dimethyl sulfone. C2H6O2S. (Mr 94.1). 1030900. [67-71-0].
N,N-Dimethylformamide dimethylacetal. C5H13NO2. White or almost white, crystalline powder, freely soluble in
(Mr 119.2). 1140700. [4637-24-5]. 1,1-Dimethoxytrimethyl- water, soluble in acetone and ethanol (96 per cent).
amine. mp : 108 °C to 110 °C.
Clear, colourless liquid. Dimethyl sulfoxide. 1029500. [67-68-5].
: about 0.896. See Dimethyl sulfoxide (0763).
: about 1.396. Dimethyl sulfoxide used in spectrophotometry complies with
bp : about 103 °C. the following additional test.

Absorbance (2.2.25) : maximum 1.00 at 262 nm, 0.46 at 270 nm, Dinitrophenylhydrazine-aceto-hydrochloric solution.

0.16 at 290 nm, 0.01 at 340 nm and higher wavelengths, 1031501.
determined using water R as compensation liquid. Dissolve 0.2 g of dinitrophenylhydrazine R in 20 mL of
Dimethyl sulfoxide R1. 1029501. methanol R and add 80 mL of a mixture of equal volumes of
acetic acid R and hydrochloric acid R1. Prepare immediately
Content : minimum 99.7 per cent, determined by gas before use.
chromatography.
Dinitrophenylhydrazine-hydrochloric solution.
Dimethyl sulfoxide R2. 1029502.
1031502.
Content : minimum 99.9 per cent, determined by gas
Dissolve by heating 0.50 g of dinitrophenylhydrazine R in
chromatography.
dilute hydrochloric acid R and dilute to 100 mL with the
Residue on evaporation : maximum 0.0005 per cent. same solvent. Allow to cool and filter. Prepare immediately
Water (2.5.32) : maximum 0.005 per cent. before use.
Dimeticone. 1105400. [9006-65-9]. Dinitrophenylhydrazine-sulfuric acid solution. 1031503.
See Dimeticone (0138). Dissolve 1.5 g of dinitrophenylhydrazine R in 50 mL of
Dimidium bromide. C20H18BrN3. (Mr 380.3). 1031100. a 20 per cent V/V solution of sulfuric acid R. Prepare
[518-67-2]. 3,8-Diamino-5-methyl-6-phenylphen- immediately before use.
anthridinium bromide. Dinonyl phthalate. C26H42O4. (Mr 418.6). 1031600.
Dark-red crystals, slightly soluble in water at 20 °C, sparingly
[28553-12-0].
soluble in water at 60 °C and in ethanol (96 per cent). Colourless to pale yellow, viscous liquid.
Dimidium bromide-sulfan blue mixed solution. : 0.97 to 0.98.
1031101. : 1.482 to 1.489.
Dissolve separately 0.5 g of dimidium bromide R and 0.25 g Acidity. Shake 5.0 g with 25 mL of water R for 1 min. Allow
of sulfan blue R in 30 mL of a hot mixture of 1 volume of to stand, filter the separated aqueous layer and add 0.1 mL of
anhydrous ethanol R and 9 volumes of water R, stir, mix the phenolphthalein solution R. Not more than 0.3 mL of 0.1 M
two solutions, and dilute to 250 mL with the same mixture sodium hydroxide is required to change the colour of the
of solvents. Mix 20 mL of this solution with 20 mL of a solution (0.05 per cent, calculated as phthalic acid).
14.0 per cent V/V solution of sulfuric acid R previously Water (2.5.12) : maximum 0.1 per cent.
diluted with about 250 mL of water R and dilute to 500 mL
with water R. Dioctadecyl disulfide. C36H74S2. (Mr 571.1). 1031700.
Storage : protected from light. [2500-88-1].
White or almost white powder, practically insoluble in water.
Dinitrobenzene. C6H4N2O4. (Mr 168.1). 1031200. [99-65-0].
1,3-Dinitrobenzene. mp : 53 °C to 58 °C.
Yellowish crystalline powder or crystals, practically insoluble 2,2′-Di(octadecyloxy)-5,5′-spirobi(1,3,2-dioxaphosphorin-
in water, slightly soluble in ethanol (96 per cent). ane). C41H82O6P2. (Mr 733). 1031800.
mp : about 90 °C. White or almost white, waxy solid, practically insoluble in
Dinitrobenzene solution. 1031201. water, soluble in hydrocarbons.
A 10 g/L solution of dinitrobenzene R in ethanol (96 per mp : 40 °C to 70 °C.
cent) R. Dioctadecyl 3,3′-thiodipropionate. C42H82O4S. (Mr 683).
Dinitrobenzoic acid. C7H4N2O6. (Mr 212.1). 1031300. 1031900. [693-36-7].
[99-34-3]. 3,5-Dinitrobenzoic acid. White or almost white, crystalline powder, practically
Almost colourless crystals, slightly soluble in water, very insoluble in water, freely soluble in methylene chloride,
soluble in ethanol (96 per cent). sparingly soluble in acetone, in ethanol (96 per cent) and in
light petroleum.
mp : about 206 °C.
mp : 58 °C to 67 °C.
Dinitrobenzoic acid solution. 1031301.
A 20 g/L solution of dinitrobenzoic acid R in ethanol (96 per Di-n-octyl phthalate. C24H38O4. (Mr 390.6). 1203500.
[117-84-0]. Dioctyl benzene-1,2-dicarboxylate.
cent) R.
Colourless viscous liquid, insoluble in water.
Dinitrobenzoyl chloride. C7H3ClN2O5. (Mr 230.6). 1031400. Density : about 0.98 g/mL (20 °C).
[99-33-2]. 3,5-Dinitrobenzoyl chloride.
Translucent, yellow or greenish-yellow powder or yellowish Dioxan. C4H8O2. (Mr 88.1). 1032000. [123-91-1].
crystals, soluble in acetone and in toluene. 1,4-Dioxane.
mp : about 68 °C. Clear, colourless liquid, miscible with water and with most
Suitability test. To 1 mL of anhydrous ethanol R and 0.1 g organic solvents.
of dinitrobenzoyl chloride R add 0.05 mL of dilute sulfuric : about 1.03.
acid R and boil under a reflux condenser for 30 min. After Freezing point (2.2.18) : minimum 11.0 °C.
evaporation on a water-bath add 5 mL of heptane R to the Water (2.5.12) : maximum 0.5 per cent.
residue and heat to boiling. Filter the hot solution. Wash the
crystals formed on cooling to room temperature with a small Do not distil if the dioxan does not comply with the test for
quantity of heptane R and dry in a desiccator. The crystals peroxides.
melt (2.2.14) at 94 °C to 95 °C. Peroxides. Place 8 mL of potassium iodide and starch solution R
in a 12 mL ground-glass-stoppered cylinder about 1.5 cm in
Dinitrophenylhydrazine. C6H6N4O4. (Mr 198.1). 1031500. diameter. Fill completely with the substance to be examined,
[119-26-6]. 2,4-Dinitrophenylhydrazine. shake vigorously and allow to stand in the dark for 30 min.
Reddish-orange crystals, very slightly soluble in water, slightly No colour is produced.
soluble in ethanol (96 per cent). Dioxan used for liquid scintillation is of a suitable analytical
mp : about 203 °C (instantaneous method). grade.

Dioxan solution. 1032002. Diphenylcarbazide solution. 1032501.

Dilute 50.0 mL of dioxan stock solution R to 100.0 mL with Dissolve 0.2 g of diphenylcarbazide R in 10 mL of glacial
water R. (0.5 mg/mL of dioxan). acetic acid R and dilute to 100 mL with anhydrous
ethanol R. Prepare immediately before use.
Dioxan solution R1. 1032003.
Dilute 10.0 mL of dioxan solution R to 50.0 mL with Diphenylcarbazone. C13H12N4O. (Mr 240.3). 1032600.
water R. (0.1 mg/mL of dioxan). [538-62-5]. 1,5-Diphenylcarbazone.
Orange-yellow, crystalline powder, practically insoluble in
Dioxan solution R2. 1032004. water, freely soluble in ethanol (96 per cent).
Dilute 2.0 mL of dioxan solution R to 50.0 mL with water R mp : about 157 °C, with decomposition.
(0.02 mg/mL of dioxan).
Diphenylcarbazone mercuric reagent. 1032601.
Dioxan stock solution. 1032001. Solution A. Dissolve 0.1 g of diphenylcarbazone R in
Dissolve 1.00 g of dioxan R in water R and dilute to anhydrous ethanol R and dilute to 50 mL with the same
100.0 mL with the same solvent. Dilute 5.0 mL of this solvent.
solution to 50.0 mL with water R (1.0 mg/mL). Solution B. Dissolve 1 g of mercuric chloride R in anhydrous
ethanol R and dilute to 50 mL with the same solvent.
Diphenylamine. C12H11N. (Mr 169.2). 1032100. [122-39-4].
Mix equal volumes of the two solutions.
White or almost white crystals, slightly soluble in water,
soluble in ethanol (96 per cent). 2,2-Diphenylglycine. C14H13NO2. (Mr 227.26). 1174300.
mp : about 55 °C. [3060-50-2]. Amino(diphenyl)acetic acid.
Storage : protected from light. 1,2-Diphenylhydrazine. C12H12N2. (Mr 184.3). 1140800.
[122-66-7]. Hydrazobenzene. 1,2-Diphenyldiazane.
Diphenylamine solution. 1032101.
Orange powder.
A 1 g/L solution of diphenylamine R in sulfuric acid R. mp : about 125 °C.
Diphenylmethanol. C13H12O. (Mr 184.2). 1145700. [91-01-0].
Diphenylamine solution R1. 1032102. Benzhydrol.
A 10 g/L solution of diphenylamine R in sulfuric acid R. White or almost white, crystalline powder.
The solution is colourless. mp : about 66 °C.
Diphenylamine solution R2. 1032103. Diphenyloxazole. C15H11NO. (Mr 221.3). 1032700. [92-71-7].
Dissolve 1 g of diphenylamine R in 100 mL of glacial acetic 2,5-Diphenyloxazole.
acid R and add 2.75 mL of sulfuric acid R. Use immediately. White or almost white powder, practically insoluble in water,
soluble in methanol, sparingly soluble in dioxan and in glacial
Diphenylanthracene. C26H18. (Mr 330.4). 1032200. acetic acid.
[1499-10-1]. 9,10-Diphenylanthracene.
mp : about 70 °C.
Yellowish or yellow, crystalline powder, practically insoluble
in water. : about 1260 determined at 305 nm in methanol R.
mp : about 248 °C. Diphenyloxazole used for liquid scintillation is of a suitable
analytical grade.
Diphenylbenzidine. C24H20N2. (Mr 336.4). 1032300. Diphenylphenylene oxide polymer. 1032800.
[531-91-9]. N,N’-Diphenylbenzidine. N,N’-Diphenylbiphenyl- 2,6-Diphenyl-p-phenylene oxide polymer.
4,4′-diamine.
White or almost white, porous beads. The size range of the
White or faintly grey, crystalline powder, practically insoluble beads is specified after the name of the reagent in the tests
in water, slightly soluble in acetone and in ethanol (96 per where it is used.
cent).
mp : about 248 °C. Diphosphorus pentoxide. P2O5. (Mr 141.9). 1032900.
[1314-56-3]. Phosphorus pentoxide. Phosphoric anhydride.
Nitrates. Dissolve 8 mg in a cooled mixture of 5 mL of water R
and 45 mL of nitrogen-free sulfuric acid R. The solution is White or almost white powder, amorphous, deliquescent. It is
colourless or very pale blue. hydrated by water with the evolution of heat.
Sulfated ash (2.4.14): maximum 0.1 per cent. Storage : in an airtight container.
Storage : protected from light. Dipotassium hydrogen phosphate. K2HPO4. (Mr 174.2).
1033000. [7758-11-4].
Diphenylboric acid aminoethyl ester. C14H16BNO.
(Mr 225.1). 1032400. [524-95-8]. White or almost white, crystalline powder, hygroscopic, very
soluble in water, slightly soluble in ethanol (96 per cent).
White or slightly yellow, crystalline powder, practically
insoluble in water, soluble in ethanol (96 per cent).
mp : about 193 °C. Dipotassium hydrogen phosphate trihydrate.
K2HPO4,3H2O. (Mr 228.2). 1157600. [16788-57-1].
Diphenylcarbazide. C13H14N4O. (Mr 242.3). 1032500. Colourless or white or almost white powder or crystals, freely
[140-22-7]. 1,5-Diphenylcarbonodihydrazide. soluble in water.
White or almost white, crystalline powder which gradually
becomes pink on exposure to air, very slightly soluble in water, Dipotassium sulfate. K2SO4. (Mr 174.3). 1033100.
soluble in acetone, in ethanol (96 per cent) and in glacial [7778-80-5].
acetic acid. Colourless crystals, soluble in water.
mp : about 170 °C. 2,2′-Dipyridylamine. C10H9N3. (Mr 171.2). 1157700.
Sulfated ash (2.4.14): maximum 0.1 per cent. [1202-34-2]. N-(Pyridin-2-yl)pyridin-2-amine.
Storage : protected from light. mp : about 95 °C.

Disodium arsenate. Na2HAsO4,7H2O. (Mr 312.0). 1102500. Dithiothreitol. C4H10O2S2. (Mr 154.2). 1098200.

[10048-95-0]. Disodium hydrogen arsenate heptahydrate. [27565-41-9]. threo-1,4-Dimercaptobutane-2,3-diol.
Dibasic sodium arsenate. Slightly hygroscopic needles, freely soluble in water, in acetone
Crystals, efflorescent in warm air, freely soluble in water, and in anhydrous ethanol.
soluble in glycerol, slightly soluble in ethanol (96 per cent). Storage : in an airtight container.
The aqueous solution is alcaline to litmus.
: about 1.87. Dithizone. C13H12N4S. (Mr 256.3). 1033900. [60-10-6].
1,5-Diphenylthiocarbazone.
mp : about 57 °C when rapidly heated.
A bluish-black, brownish-black or black powder, practically
Disodium bicinchoninate. C20H10N2Na2O4. insoluble in water, soluble in ethanol (96 per cent).
(Mr 388.3). 1126600. [979-88-4]. Disodium Storage : protected from light.
2,2′-biquinoline-4-4′-dicarboxylate.
Dithizone solution. 1033901.
Disodium hydrogen citrate. C6H6Na2O7,11/2H2O. (Mr 263.1). A 0.5 g/L solution of dithizone R in chloroform R. Prepare
1033200. [144-33-2]. Sodium acid citrate. Disodium hydrogen immediately before use.
2-hydroxypropane-1,2,3-tricarboxylate sesquihydrate.
White or almost white powder, soluble in less than 2 parts of Dithizone solution R2. 1033903.
water, practically insoluble in ethanol (96 per cent). Dissolve 40.0 mg of dithizone R in chloroform R and dilute
to 1000.0 mL with the same solvent. Dilute 30.0 mL of the
Disodium hydrogen phosphate, anhydrous. Na2HPO4. solution to 100.0 mL with chloroform R.
(Mr 142.0). 1033400. [7558-79-4]. Assay. Dissolve a quantity of mercuric chloride R equivalent
Disodium hydrogen phosphate dihydrate. 1033500. to 0.1354 g of HgCl2 in a mixture of equal volumes of dilute
[10028-24-7]. sulfuric acid R and water R and dilute to 100.0 mL with the
same mixture of solvents. Dilute 2.0 mL of this solution
See Disodium phosphate dihydrate (0602).
to 100.0 mL with a mixture of equal volumes of dilute
Disodium hydrogen phosphate dodecahydrate. 1033300. sulfuric acid R and water R. (This solution contains 20 ppm
[10039-32-4]. of Hg). Transfer 1.0 mL of the solution to a separating
See Disodium phosphate dodecahydrate (0118). funnel and add 50 mL of dilute sulfuric acid R, 140 mL of
water R and 10 mL of a 200 g/L solution of hydroxylamine
Disodium hydrogen phosphate solution. 1033301. hydrochloride R. Titrate with the dithizone solution ; after
A 90 g/L solution of disodium hydrogen phosphate each addition, shake the mixture twenty times and towards
dodecahydrate R. the end of the titration allow to separate and discard
the chloroform layer. Titrate until a bluish-green colour
Disodium hydrogen phosphate heptahydrate. is obtained. Calculate the equivalent in micrograms of
Na2HPO4,7H2O. (Mr 268.1). 1206900. [7782-85-6]. mercury per millilitre of the dithizone solution from the
expression 20/V, where V is the volume in millilitres of the
Disodium tetraborate. 1033600. [1303-96-4].
dithizone solution used in the titration.
See Borax (0013).
Dithizone R1. C13H12N4S. (Mr 256.3). 1105500. [60-10-6].
Borate solution. 1033601. 1,5-Diphenylthiocarbazone.
Dissolve 9.55 g of disodium tetraborate R in sulfuric acid R, Content : minimum 98.0 per cent.
heating on a water-bath, and dilute to 1 L with the same Bluish-black, brownish-black or black powder, practically
acid. insoluble in water, soluble in ethanol (96 per cent).
Ditalimphos. C12H14NO4PS. (Mr 299.3). 1126700. Storage : protected from light.
[5131-24-8]. O,O-Diethyl (1,3-dihydro-1,3-dioxo-2H-
isoindol-2-yl)phosphonothioate. Divanadium pentoxide. V2O5. (Mr 181.9). 1034000.
[1314-62-1]. Vanadic anhydride.
Very slightly soluble in water, in ethyl acetate and in anhydrous
ethanol. Content : minimum 98.5 per cent.
Yellow-brown or rust-brown powder, slightly soluble in water,
A suitable certified reference solution may be used.
soluble in strong mineral acids and in solutions of alkali
5,5′-Dithiobis(2-nitrobenzoic acid). C14H8N2O8S2. hydroxides with formation of salts.
(Mr 396.4). 1097300. [69-78-3]. 3-Carboxy-4- Appearance of solution. Heat 1 g for 30 min with 10 mL of
nitrophenyldisulfide. Ellman’s reagent. DTNB. sulfuric acid R. Allow to cool and dilute to 10 mL with the
Yellow powder sparingly soluble in ethanol (96 per cent). same acid. The solution is clear (2.2.1).
mp : about 242 °C. Sensitivity to hydrogen peroxide. Dilute 1.0 mL of the solution
prepared for the test for appearance of solution cautiously to
Dithioerythritol. C4H10O2S2. (Mr 154.3). 1187500. 50.0 mL with water R. To 0.5 mL of the solution add 0.1 mL
[6892-68-8]. of a solution of hydrogen peroxide (0.1 g/L of H2O2) prepared
(2R,3S)-1,4-Disulfanylbutane-2,3-diol. DTE. from dilute hydrogen peroxide solution R. The solution has a
mp : about 83 °C. distinct orange colour compared with a blank prepared from
0.5 mL of the solution to be examined and 0.1 mL of water R.
Dithiol. C7H8S2. (Mr 156.3). 1033800. [496-74-2]. After the addition of 0.4 mL of a solution of hydrogen peroxide
Toluene-3,4-dithiol. 4-Methylbenzene-1,2-dithiol. (0.1 g/L of H2O2) prepared from dilute hydrogen peroxide
White or almost white crystals, hygroscopic, soluble in solution R, the orange solution becomes orange-yellow.
methanol and in solutions of alkali hydroxides. Loss on ignition : maximum 1.0 per cent, determined on 1.00 g
mp : about 30 °C. at 700 ± 50 °C.
Storage : in an airtight container. Assay. Dissolve 0.200 g with heating in 20 mL of a 70 per
cent m/m solution of sulfuric acid R. Add 100 mL of water R
Dithiol reagent. 1033801. and 0.02 M potassium permanganate until a reddish colour is
To 1 g of dithiol R add 2 mL of thioglycollic acid R and dilute obtained. Decolorise the excess of potassium permanganate
to 250 mL with a 20 g/L solution of sodium hydroxide R. by the addition of a 30 g/L solution of sodium nitrite R. Add
Prepare immediately before use. 5 g of urea R and 80 mL of a 70 per cent m/m solution of

sulfuric acid R. Cool. Using 0.1 mL of ferroin R as indicator, Elementary standard solution for atomic spectrometry
titrate the solution immediately with 0.1 M ferrous sulfate (1.000 g/L). 5004000.
until a greenish-red colour is obtained. This solution is prepared, generally in acid conditions, from
1 mL of 0.1 M ferrous sulfate is equivalent to 9.095 mg of V2O5. the element or a salt of the element whose minimum content
is not less than 99.0 per cent. The quantity per litre of solution
Divanadium pentoxide solution in sulfuric acid. is greater than 0.995 g throughout the guaranteed period, as
1034001. long as the vial has not been opened. The starting material
Dissolve 0.2 g of divanadium pentoxide R in 4 mL of (element or salt) and the characteristics of the final solvent
sulfuric acid R and dilute to 100 mL with water R. (nature and acidity, etc.) are mentioned on the label.
Docosahexaenoic acid methyl ester. C23H34O2. (Mr 342.5). Emodin. C15H10O5. (Mr 270.2). 1034400. [518-82-1].
1142800. [301-01-9]. DHA methyl ester. Cervonic acid 1,3,8-Trihydroxy-6-methylanthraquinone.
methyl ester. (all-Z)-Docosa-4,7,10,13,16,19-hexaenoic acid Orange-red needles, practically insoluble in water, soluble in
methyl ester. ethanol (96 per cent) and in solutions of alkali hydroxides.
Content : minimum 90.0 per cent, determined by gas Chromatography. Thin-layer chromatography (2.2.27)
chromatography. as prescribed in the monograph Rhubarb (0291) ; the
chromatogram shows only one principal spot.
Docusate sodium. 1034100. [577-11-7].
See Docusate sodium (1418). Endoprotease LysC. 1173200.
Microbial extracellular proteolytic enzyme secreted by
Dodecyltrimethylammonium bromide. C15H34BrN. Achromobacter lyticus. A lyophilised powder, free of salts.
(Mr 308.4). 1135500. [1119-94-4]. N,N,N-Trimethyldodecan-
1-aminium bromide. α-Endosulfan. C9H6Cl6O3S. (Mr 406.9). 1126800. [959-98-8].
White or almost white crystals. bp : about 200 °C.
D-Dopa. C9H11NO4. (Mr 197.2). 1164100. [5796-17-8]. cyclohexane) may be used.
(2R)-2-Amino-3-(3,4-dihydroxyphenyl)propanoic acid.
3-Hydroxy-D-tyrosine. 3,4-Dihydroxy-D-phenylalanine. β-Endosulfan. C9H6Cl6O3S. (Mr 406.9). 1126900.
[33213-65-9].
: + 9.5 to + 11.5, determined on a 10 g/L solution in 1 M
hydrochloric acid. bp : about 390 °C.
Dotriacontane. C32H66. (Mr 450.9). 1034200. [544-85-4]. cyclohexane) may be used.
n-Dotriacontane.
Endrin. C12H8Cl6O. (Mr 380.9). 1127000. [72-20-8].
White or almost white plates, practically insoluble in water,
sparingly soluble in hexane. A suitable certified reference solution (10 ng/µL in
mp : about 69 °C.
Impurities. Not more than 0.1 per cent of impurities with (−)-Epicatechin. C15H14O6. (Mr 290.3). 1201300. [490-46-0].
the same tR value as α-tocopherol acetate, determined by the (2R,3R)-2-(3,4-Dihydroxyphenyl)-3,4-dihydro-2H-1-
gas chromatographic method prescribed in the monograph benzopyran-3,5,7-triol.
α-Tocopherol acetate (0439). (−)-Epigallocatechin-3-O-gallate. C22H18O11. (Mr 458.4).
Doxycycline. 1145800. 1201400. [989-51-5]. (2R,3R)-5,7-Dihydroxy-2-(3,4,5-
trihydroxyphenyl)-3,4-dihydro-2H-1-benzopyran-3-yl
See Doxycycline monohydrate (0820). 3,4,5-trihydroxybenzoate.
β-Ecdysterone. C27H44O7. (Mr 480.6). 1204700. [5289-74-7]. Epilactose. C12H22O11. (Mr 342.3). 1189200. [20869-27-6].
(2β,3β,5β,22R)-2,3,14,20,22,25-Hexahydroxycholest-7-en-6- 4-O-β-D-Galactopyranosyl-D-mannopyranose.
one.
Echinacoside. C35H46O20. (Mr 786.5). 1159400. [82854-37-3].
β-(3′,4′-Dihydroxyphenyl)-ethyl-O-α-L-rhamnopyranosyl Erucamide. C22H43NO. (Mr 337.6). 1034500. [112-84-5].
(1→3)-O-β-D-[β-D-glucopyranosyl(1→6)]-(4-O-caffeoyl)- (Z)-Docos-13-enoamide.
glucopyranoside. Yellowish or white powder or granules, practically insoluble
in water, very soluble in methylene chloride, soluble in
Pale yellow powder, odourless.
anhydrous ethanol.
Edotreotide. C65H92N14O18S2. (Mr 1422). 1182400. mp : about 70 °C.
[204318-14-9]. N-[[4,7,10-Tris(carboxymethyl)-1,4,7,10-
tetraazacyclododecan-1-yl]acetyl]-D-phenylalanyl-L-cysteinyl- Erythritol. 1113800. [149-32-6].
L-tyrosyl-D-tryptophyl-L-lysyl-L-threonyl-N-[(1R,2R)-2- See Erythritol (1803).
hydroxy-1-(hydroxymethyl)propyl]-L-cysteinamide cyclic
Esculetin. C9H6O4. (Mr 178.1). 1185800. [305-01-1].
(2→7)-disulfide. DOTATOC. DOTA-[Tyr3]-octreotide.
6,7-Dihydroxy-2H-1-benzopyran-2-one. Aesculetin.
Content : minimum 95.0 per cent. Esculin. C15H16O9,11/2H2O. (Mr 367.3). 1119400. [531-75-9].
6-(β-D-Glucopyranosyloxy)-7-hydroxy-2H-chromen-2-one.
Electrolyte reagent for the micro determination of water. White or almost white powder or colourless crystals, sparingly
1113700. soluble in water and in ethanol (96 per cent), freely soluble in
Commercially available anhydrous reagent or a combination hot water and in hot ethanol (96 per cent).
of anhydrous reagents for the coulometric titration of water, Chromatography (2.2.27). Thin-layer chromatography (2.2.27)
containing suitable organic bases, sulfur dioxide and iodide as prescribed in the monograph Eleutherococcus (1419) ; the
dissolved in a suitable solvent. chromatogram shows only one principal spot.

Estradiol. C18H24O2. (Mr 272.4). 1135600. [50-28-2].

Estra-1,3,5(10)-triene-3,17β-diol. β-Estradiol. a = percentage V/V content of methanol in the
Prisms stable in air, practically insoluble in water, freely reference solution,
soluble in ethanol (96 per cent), soluble in acetone and in b = area of the methanol peak in the chromatogram
dioxan, sparingly soluble in vegetable oils. obtained with the test solution,
mp : 173 °C to 179 °C. c = area of the methanol peak in the chromatogram
obtained with the reference solution.
17α-Estradiol. C18H24O2. (Mr 272.4). 1034600. [57-91-0].
Limit :
– methanol : maximum 0.005 per cent V/V.
crystals.
mp : 220 °C to 223 °C. Ethanol (96 per cent). 1002500. [64-17-5].
See Ethanol (96 per cent) (1317).
Estragole. C10H12O. (Mr 148.2). 1034700. [140-67-0].
1-Methoxy-4-prop-2-enylbenzene. Ethanol (x per cent V/V). 1002502.
Liquid, miscible with ethanol (96 per cent). Mix appropriate volumes of water R and ethanol (96 per
cent) R, allowing for the effects of warming and volume
: about 1.52. contraction inherent to the preparation of such a mixture,
bp : about 216 °C. to obtain a solution whose final content of ethanol
Estragole used in gas chromatography complies with the corresponds to the value of x.
following test. Ethanolamine. C2H7NO. (Mr 61.1). 1034900. [141-43-5].
Assay. Gas chromatography (2.2.28) as prescribed in the 2-Aminoethanol.
monograph Anise oil (0804). Clear, colourless, viscous, hygroscopic liquid, miscible with
Test solution. The substance to be examined. water and with methanol.
Content : minimum 98.0 per cent, calculated by the : about 1.014.
normalisation procedure. : about 1.454.
Ethane. C2H6. (Mr 30.07). 1189300. [74-84-0]. mp : about 11 °C.
Ether. C4H10O. (Mr 74.1). 1035000. [60-29-7].
Ethanol. 1034800. [64-17-5].
Clear, colourless, volatile and very mobile liquid, very
See Ethanol, anhydrous R. flammable, hygroscopic, soluble in water, miscible with
Ethanol, anhydrous. 1034800. [64-17-5].
: 0.713 to 0.715.
See Ethanol, anhydrous (1318).
bp : 34 °C to 35 °C.
Ethanol R1. 1034801. Do not distil if the ether does not comply with the test for
Complies with the requirements prescribed for the peroxides.
monograph Ethanol, anhydrous (1318) with the following Peroxides. Place 8 mL of potassium iodide and starch solution R
additional requirement. in a 12 mL ground-glass-stoppered cylinder about 1.5 cm in
diameter. Fill completely with the substance to be examined,
Methanol. Gas chromatography (2.2.28).
shake vigorously and allow to stand in the dark for 30 min.
Test solution. The substance to be examined. No colour is produced.
Reference solution. Dilute 0.50 mL of anhydrous methanol R The name and concentration of any added stabilisers are
to 100.0 mL with the substance to be examined. Dilute stated on the label.
1.0 mL of this solution to 100.0 mL with the substance to Storage : in an airtight container, protected from light, at a
be examined. temperature not exceeding 15 °C.
Column :
Ether, peroxide-free. 1035100.
– material : glass ;
See Anaesthetic ether (0367).
– size : l = 2 m, Ø = 2 mm ;
Ethion. C9H22O4P2S4. (Mr 384.5). 1127100. [563-12-2].
– stationary phase : ethylvinylbenzene-divinylbenzene
copolymer R (75-100 μm). mp : − 24 °C to − 25 °C.
Carrier gas : nitrogen for chromatography R. A suitable certified reference solution (10 ng/µL in
Temperature : Ethoxychrysoidine hydrochloride. C14H17ClN4O. (Mr 292.8).
1035200. [2313-87-3]. 4-[(4-Ethoxyphenyl)diazenyl]phenyl-
– column : 130 °C ; ene-1,3-diamine hydrochloride.
– injection port : 150 °C ; Reddish powder, soluble in ethanol (96 per cent).
– detector : 200 °C.
Ethoxychrysoidine solution. 1035201.
Detection : flame-ionisation. A 1 g/L solution of ethoxychrysoidine hydrochloride R in
Injection : 1 μL of the test solution and 1 μL of the reference ethanol (96 per cent) R.
solution, alternately, three times. Test for sensitivity. To a mixture of 5 mL of dilute
After each chromatography, heat the column to 230 °C hydrochloric acid R and 0.05 mL of the ethoxy-chrysoidine
for 8 min. Integrate the methanol peak. Calculate solution add 0.05 mL of 0.0167 M bromide-bromate. The
the percentage methanol content from the following colour changes from red to light yellow within 2 min.
expression :
Ethyl acetate. C4H8O2. (Mr 88.1). 1035300. [141-78-6].
a´b Clear, colourless liquid, soluble in water, miscible with ethanol
c-b (96 per cent).

: 0.901 to 0.904. : about 1.25.

bp : 76 °C to 78 °C. Distillation range (2.2.11). Not less than 95 per cent distils
between 82 °C and 84 °C.
Ethyl acetate, treated. 1035301.
Disperse 200 g of sulfamic acid R in ethyl acetate R and Ethylenediamine. C2H8N2. (Mr 60.1). 1036500. [107-15-3].
make up to 1000 mL with the same solvent. Stir the Ethane-1,2-diamine.
suspension obtained for three days and filter through a Clear, colourless, fuming liquid, strongly alkaline, miscible
filter paper. with water and with ethanol (96 per cent).
Storage : use within 1 month. bp : about 116 °C.
Ethyl acrylate. C5H8O2. (Mr 100.1). 1035400. [140-88-5].
Ethylene bis[3,3-di(3-tert-butyl-4-hydroxyphenyl)buty-
Ethyl prop-2-enoate.
rate]. 1035900. [32509-66-3].
Colourless liquid.
See ethylene bis[3,3-di(3-(1,1-dimethylethyl)-4-
: about 0.924. hydroxyphenyl)butyrate] R.
: about 1.406.
bp : about 99 °C. Ethylene bis[3,3-di(3-(1,1-dimethylethyl)-4-
hydroxyphenyl)butyrate]. C50H66O8. (Mr 795). 1035900.
mp : about − 71 °C. [32509-66-3]. Ethylene bis[3,3-di(3-tert-butyl-4-
4-[(Ethylamino)methyl]pyridine. C8H12N2. (Mr 136.2). hydroxyphenyl)butyrate].
1101300. [33403-97-3]. Crystalline powder, practically insoluble in water and in light
Pale yellow liquid. petroleum, very soluble in acetone and in methanol.
: about 0.98. mp : about 165 °C.
: about 1.516. (Ethylenedinitrilo)tetra-acetic acid. C10H16N2O8.
bp : about 98 °C. (Mr 292.2). 1105800. [60-00-4]. N,N’-1,2-Ethanediylbis[N-
(carboxymethyl)glycine]. Edetic acid.
Ethylbenzene. C8H10. (Mr 106.2). 1035800. [100-41-4].
Content : minimum 99.5 per cent m/m, determined by gas White or almost white crystalline powder, very slightly soluble
chromatography. in water.
Clear, colourless liquid, practically insoluble in water, soluble mp : about 250 °C, with decomposition.
in acetone, and in ethanol (96 per cent). Ethylene glycol. C2H6O2. (Mr 62.1). 1036100. [107-21-1].
: about 0.87. Ethane-1,2-diol.
: about 1.496. Content : minimum 99.0 per cent.
bp : about 135 °C. Colourless, slightly viscous liquid, hygroscopic, miscible with
Ethyl benzenesulfonate. C8H10O3S. (Mr 186.2). 1194800. water and with ethanol (96 per cent).
[515-46-8]. : 1.113 to 1.115.
Content : minimum 97.0 per cent. : about 1.432.
Colourless or slightly yellow liquid, slightly soluble in water, bp : about 198 °C.
Density : about 1.22 g/mL (25 °C).
Acidity. To 10 mL add 20 mL of water R and 1 mL of
Ethyl benzoate. C9H10O2. (Mr 150.2). 1135700. [93-89-0]. phenolphthalein solution R. Not more than 0.15 mL of 0.02 M
A clear, colourless, refractive liquid, practically insoluble sodium hydroxide is required to change the colour of the
in water, miscible with ethanol (96 per cent) and with light indicator to pink.
petroleum. Water (2.5.12) : maximum 0.2 per cent
: about 1.050.
Ethylene glycol monododecyl ether. C14H30O2. (Mr 230.4).
: about 1.506. 1191900. [4536-30-5]. 2-(Dodecyloxy)ethan-1-ol.
bp : 211 °C to 213 °C. Colourless or faintly green liquid.
Ethyl 5-bromovalerate. C7H13BrO2. (Mr 209.1). 1142900.
Ethylene glycol monoethyl ether. C4H10O2. (Mr 90.1).
[14660-52-7]. Ethyl 5-bromopentanoate.
1036200. [110-80-5]. 2-Ethoxyethanol.
Clear, colourless liquid.
: about 1.321.
Clear, colourless liquid, miscible with water, with acetone and
bp : 104 °C to 109 °C. with ethanol (96 per cent).
Ethyl clorazepate. C18H15ClN2O3. (Mr 342.8). 1204800. : about 0.93.
[5606-55-3]. Ethyl (3RS)-7-chloro-2-oxo-5-phenyl-2,3- : about 1.406.
dihydro-1H-1,4-benzodiazepine-3-carboxylate.
bp : about 135 °C.
Ethyl cyanoacetate. C5H7NO2. (Mr 113.1). 1035500.
[105-56-6]. Ethylene glycol monomethyl ether. C3H8O2. (Mr 76.1).
Colourless or pale yellow liquid, slightly soluble in water, 1036300. [109-86-4]. 2-Methoxyethanol.
miscible with ethanol (96 per cent). Content : minimum 99.0 per cent.
bp : 205 °C to 209 °C, with decomposition. Clear, colourless liquid, miscible with water, with acetone and
Ethylene chloride. C2H4Cl2. (Mr 99.0). 1036000. [107-06-2].
1,2-Dichloroethane. : about 0.97.
Clear, colourless liquid, soluble in about 120 parts of water : about 1.403.
and in 2 parts of ethanol (96 per cent). bp : about 125 °C.

Ethylene oxide. C2H4O. (Mr 44.05). 1036400. [75-21-8]. Assay. To 10 mL of a 500 g/L suspension of magnesium
Oxirane. chloride R in anhydrous ethanol R add 20.0 mL of 0.1 M
Colourless, flammable gas, very soluble in water and in alcoholic hydrochloric acid R in a flask. Stopper and
anhydrous ethanol. shake to obtain a saturated solution and allow to stand
overnight to equilibrate. Weigh 5.00 g of ethylene oxide
Liquefaction point : about 12 °C. stock solution (2.5 g/L) into the flask and allow to stand for
Ethylene oxide solution. 1036402. 30 min. Titrate with 0.1 M alcoholic potassium hydroxide R
determining the end-point potentiometrically (2.2.20).
Weigh a quantity of cool ethylene oxide stock solution R
equivalent to 2.5 mg of ethylene oxide into a cool flask and Carry out a blank titration, replacing the substance to be
dilute to 50.0 g with macrogol 200 R1. Mix well and dilute examined with the same quantity of macrogol 200 R1.
2.5 g of this solution to 25.0 mL with macrogol 200 R1 Ethylene oxide content in milligrams per gram is given by :
(5 μg of ethylene oxide per gram of solution). Prepare
immediately before use. (V0 - V1) ´ f ´ 4.404
The solution can be prepared using commercially available m
reagents instead of ethylene oxide stock solution R, making V 0, V 1 = volumes of 0.1 M alcoholic potassium
appropriate dilutions. hydroxide used respectively for the blank
Ethylene oxide solution R1. 1036403. titration and the assay,
Dilute 1.0 mL of cooled ethylene oxide stock solution R f = factor of the alcoholic potassium hydroxide
(check the exact volume by weighing) to 50.0 mL with solution,
macrogol 200 R1. Mix well and dilute 2.5 g of this solution m = mass of the sample taken, in grams.
to 25.0 mL with macrogol 200 R1. Calculate the exact
amount of ethylene oxide in parts per million from the Ethylene oxide stock solution R1. 1036406.
volume determined by weighing and taking the relative A 50 g/L solution of ethylene oxide R in methanol R.
density of macrogol 200 R1 as 1.127. Prepare immediately
before use. Either use a commercially available reagent or prepare the
solution corresponding to the aforementioned composition.
The solution can be prepared using commercially available
reagents instead of ethylene oxide stock solution R, making Ethylene oxide stock solution R2. 1036408.
appropriate dilutions. A 50 g/L solution of ethylene oxide R in methylene
Ethylene oxide solution R2. 1036404. chloride R.
Weigh 1.00 g of cold ethylene oxide stock solution R Either use a commercially available reagent or prepare the
(equivalent to 2.5 mg of ethylene oxide) into a cold flask solution corresponding to the aforementioned composition.
containing 40.0 g of cold macrogol 200 R1. Mix and Ethyl formate. C3H6O2. (Mr 74.1). 1035600. [109-94-4]. Ethyl
determine the exact mass and dilute to a calculated mass methanoate.
to obtain a solution containing 50 µg of ethylene oxide per
gram of solution. Weigh 10.00 g into a flask containing Clear, colourless, flammable liquid, freely soluble in water,
about 30 mL of water R, mix and dilute to 50.0 mL with miscible with ethanol (96 per cent).
water R (10 μg/mL of ethylene oxide). Prepare immediately : about 0.919.
before use. : about 1.36.
The solution can be prepared using commercially available bp : about 54 °C.
reagents instead of ethylene oxide stock solution R, making
appropriate dilutions. 2-Ethylhexane-1,3-diol. C8H18O2. (Mr 146.2). 1105900.
[94-96-2].
Ethylene oxide solution R3. 1036405. Slightly oily liquid, soluble in anhydrous ethanol, 2-propanol,
Dilute 10.0 mL of ethylene oxide solution R2 to 50.0 mL with propylene glycol and castor oil.
water R (2 µg/mL of ethylene oxide). Prepare immediately : about 0.942.
before use.
: about 1.451.
Ethylene oxide solution R4. 1036407. bp : about 244 °C.
Dilute 1.0 mL of ethylene oxide stock solution R1 to
100.0 mL with water R. Dilute 1.0 mL of this solution to 2-Ethylhexanoic acid. C8H16O2. (Mr 144.2). 1036600.
25.0 mL with water R. [149-57-5].
Colourless liquid.
Ethylene oxide stock solution. 1036401.
: about 0.91.
All operations carried out in the preparation of these : about 1.425.
solutions must be conducted in a fume cupboard. The
operator must protect both hands and face by wearing Related substances. Gas chromatography (2.2.28).
polyethylene protective gloves and an appropriate face mask. Injection : 1 µL of the test solution.
Store all solutions in an airtight container in a refrigerator Test solution : suspend 0.2 g of the 2-ethylhexanoic acid in
at 4 °C to 8 °C. Carry out all determinations three times. 5 mL of water R, add 3 mL of dilute hydrochloric acid R and
Into a dry, clean test-tube, cooled in a mixture of 1 part of 5 mL of hexane R, shake for 1 min, allow the layers to separate
sodium chloride R and 3 parts of crushed ice, introduce a and use the upper layer. Carry out the chromatographic
slow current of ethylene oxide R gas, allowing condensation procedure as prescribed in the test for 2-ethylhexanoic acid in
onto the inner wall of the test-tube. Using a glass the monograph on Amoxicillin sodium (0577).
syringe, previously cooled to − 10 °C, inject about 300 µL Limit : the sum of the area of any peaks, apart from the
(corresponding to about 0.25 g) of liquid ethylene oxide R principal peak and the peak due to the solvent, is not greater
into 50 mL of macrogol 200 R1. Determine the absorbed than 2.5 per cent of the area of the principal peak.
quantity of ethylene oxide by weighing before and after
absorption (Meo). Dilute to 100.0 mL with macrogol 200 R1. Ethyl 4-hydroxybenzoate. 1035700. [120-47-8].
Mix well before use. See Ethyl parahydroxybenzoate R.

N-Ethylmaleimide. C6H7NO2. (Mr 125.1). 1036700. In a container with a minimum capacity of 30 L in a chamber
[128-53-0]. 1-Ethyl-1H-pyrrole-2,5-dione. at 4 °C introduce 25 L of distilled water R at 4 °C and add about
Colourless crystals, sparingly soluble in water, freely soluble 500 g of solid carbon dioxide. Immediately add, while stirring,
in ethanol (96 per cent). the supernatant obtained from the plasma. A white precipitate
mp : 41 °C to 45 °C. is formed. Allow to settle at 4 °C for 10-15 h. Remove the clear
supernatant solution by siphoning. Collect the precipitate by
Storage : at a temperature of 2 °C to 8 °C. centrifuging at 4 °C. Suspend the precipitate by dispersing
Ethyl methanesulfonate. C3H8O3S. (Mr 124.2). 1179300. mechanically in 500 mL of distilled water R at 4 °C, shake
[62-50-0]. for 5 min and collect the precipitate by centrifuging at 4 °C.
Clear, colourless liquid. Disperse the precipitate mechanically in 60 mL of a solution
containing 9 g/L of sodium chloride R and 0.9 g/L sodium
Content : minimum 99.0 per cent. citrate R and adjust to pH 7.2-7.4 by adding a 10 g/L solution
Density : about 1.206 g/cm3 (20 °C). of sodium hydroxide R. Filter through a sintered glass filter
: about 1.418. (2.1.2); to facilitate the dissolution of the precipitate crush the
bp : about 213 °C. particles of the precipitate with a suitable instrument. Wash
the filter and the instrument with 40 mL of the chloride-citrate
Ethyl methyl ketone. 1054100. [78-93-3]. solution described above and dilute to 100 mL with the same
See methyl ethyl ketone R. solution. Freeze-dry the solution. The yields are generally 6 g
to 8 g of euglobulins per litre of bovine plasma.
2-Ethyl-2-methylsuccinic acid. C7H12O4. (Mr 160.2).
1036800. [631-31-2]. 2-Ethyl-2-methylbutanedioic acid. Test for suitability. For this test, prepare the solutions using
phosphate buffer solution pH 7.4 R containing 30 g/L of bovine
mp : 104 °C to 107 °C. albumin R.
Ethyl parahydroxybenzoate. 1035700. [120-47-8]. Into a test-tube 8 mm in diameter placed in a water-bath at
See Ethyl parahydroxybenzoate (0900). 37 °C introduce 0.2 mL of a solution of a reference preparation
of urokinase containing 100 IU/mL and 0.1 mL of a solution
2-Ethylpyridine. C7H9N. (Mr 107.2). 1133400. [100-71-0]. of human thrombin R containing 20 IU/mL. Add rapidly
Colourless or brownish liquid. 0.5 mL of a solution containing 10 mg of bovine euglobulins
: about 0.939. per millilitre. A firm clot forms in less than 10 s. Note the time
: about 1.496. that elapses between the addition of the solution of bovine
euglobulins and the lysis of the clot. The lysis time does not
bp : about 149 °C. exceed 15 min.
Ethyl toluenesulfonate. C9H12O3S. (Mr 200.3). 1191000. Storage : protected from moisture at 4 °C ; use within 1 year.
[80-40-0]. Ethyl 4-methylbenzenesulfonate. Ethyl tosilate.
Content : minimum 97.0 per cent. Euglobulins, human. 1037200.
Density : about 1.17 g/mL (25 °C). For the preparation, use fresh human blood collected into an
anticoagulant solution (for example sodium citrate solution)
bp : about 160 °C. or human blood for transfusion that has been collected in
mp : about 33 °C. plastic blood bags and which has just reached its expiry date.
Ethylvinylbenzene-divinylbenzene copolymer. 1036900. Discard any haemolysed blood. Centrifuge at 1500-1800 g
at 15 °C to obtain a supernatant plasma poor in platelets.
Porous, rigid, cross-linked polymer beads. Several grades are Iso-group plasmas may be mixed.
available with different sizes of bead. The size range of the
beads is specified after the name of the reagent in the tests To 1 L of the plasma add 75 g of barium sulfate R and shake
where it is used. for 30 min. Centrifuge at not less than 15 000 g at 15 °C and
draw off the clear supernatant. Add 10 mL of a solution
of aprotinin R containing 0.2 mg/mL and shake to ensure
Eugenol. C10H12O2. (Mr 164.2). 1037000. [97-53-0]. mixing. In a container with a minimum capacity of 30 L in
4-Allyl-2-methoxyphenol. a chamber at 4 °C introduce 25 L of distilled water R at 4 °C
Colourless or pale yellow, oily liquid, darkening on exposure and add about 500 g of solid carbon dioxide. Immediately
to air and light and becoming more viscous, practically add while stirring the supernatant obtained from the plasma.
insoluble in water, miscible with ethanol (96 per cent) and A white precipitate is formed. Allow to settle at 4 °C for
with fatty and essential oils. 10-15 h. Remove the clear supernatant solution by siphoning.
: about 1.07. Collect the precipitate by centrifuging at 4 °C. Suspend the
bp : about 250 °C. precipitate by dispersing mechanically in 500 mL of distilled
Eugenol used in gas chromatography complies with the water R at 4 °C, shake for 5 min and collect the precipitate by
following additional test. centrifuging at 4 °C. Disperse the precipitate mechanically in
60 mL of a solution containing 9 g/L of sodium chloride R and
Assay. Gas chromatography (2.2.28) as prescribed in the 0.9 g/L of sodium citrate R, and adjust the pH to 7.2-7.4 by
monograph Clove oil (1091). adding a 10 g/L solution of sodium hydroxide R. Filter through
Test solution. The substance to be examined. a sintered-glass filter (2.1.2) ; to facilitate the dissolution of the
Content : minimum 98.0 per cent, calculated by the precipitate crush the particles of the precipitate with a suitable
normalisation procedure. instrument. Wash the filter and the instrument with 40 mL
Storage : protected from light. of the chloride-citrate solution described above and dilute to
100 mL with the same solution. Freeze-dry the solution. The
Euglobulins, bovine. 1037100. yields are generally 6 g to 8 g of euglobulins per litre of human
Use fresh bovine blood collected into an anticoagulant plasma.
solution (for example, sodium citrate solution). Discard any Test for suitability. For this test, prepare the solutions using
haemolysed blood. Centrifuge at 1500-1800 g at 15-20 °C to phosphate buffer solution pH 7.2 R containing 30 g/L of
obtain a supernatant plasma poor in platelets. bovine albumin R. Into a test-tube 8 mm in diameter placed
To 1 L of bovine plasma add 75 g of barium sulfate R and in a water-bath at 37 °C introduce 0.1 mL of a solution of a
shake for 30 min. Centrifuge at not less than 1500-1800 g at reference preparation of streptokinase containing 10 IU of
15-20 °C and draw off the clear supernatant. Add 10 mL of a streptokinase activity per millilitre and 0.1 mL of a solution of
0.2 mg/mL solution of aprotinin R and shake to ensure mixing. human thrombin R containing 20 IU/mL. Add rapidly 1 mL

of a solution containing 10 mg of human euglobulins per Fast red B salt. C17H13N3O9S2. (Mr 467.4). 1037500.
millilitre. A firm clot forms in less than 10 s. Note the time [49735-71-9].
that elapses between the addition of the solution of human Schultz No. 155.
euglobulins and the lysis of the clot. The lysis time does not Colour Index No. 37125.
exceed 15 min. 2-Methoxy-4-nitrobenzenediazonium hydrogen
Storage : in an airtight container at 4 °C ; use within 1 year. naphthalene-1,5-disulfonate.
Evodiamine. C19H17N3O. (Mr 303.4). 1199400. Orange-yellow powder, soluble in water, slightly soluble in
[518-17-2]. (13bS)-14-Methyl-8,13,13b,14-tetrahydro- ethanol (96 per cent).
indolo[2′,3′:3,4]pyrido[2,1-b]quinazolin-5(7H)-one. Storage : in an airtight container, protected from light, at 2 °C
to 8 °C.
Extraction resin. 1204900.
Solid phase extraction resin containing 2,2′-oxybis(N,N- Fenchlorphos. C8H8Cl3O3PS. (Mr 321.5). 1127200.
dioctylacetamide) (N,N,N′,N′-tetra-n-octyldiglycolamide). [299-84-3].
Factor VII-deficient plasma. 1185900. mp : about 35 °C.
Plasma that is deficient in factor VII. A suitable certified reference solution (10 ng/µL in
Factor Xa, bovine, coagulation. 1037300. [9002-05-5].
Fenchone. C10H16O. (Mr 152.2). 1037600. [7787-20-4].
An enzyme which converts prothrombin to thrombin. The (1R)-1,3,3-Trimethylbicyclo[2.2.1]heptan-2-one.
semi-purified preparation is obtained from liquid bovine
plasma and it may be prepared by activation of the zymogen Oily liquid, miscible with ethanol (96 per cent), practically
factor X with a suitable activator such as Russell’s viper venom. insoluble in water.
Storage : freeze-dried preparation at − 20 °C and frozen : about 1.46.
solution at a temperature lower than − 20 °C. bp15mm : 192 °C to 194 °C.
Fenchone used in gas chromatography complies with the
Factor Xa solution, bovine. 1037301.
following test.
Reconstitute as directed by the manufacturer and dilute
with tris(hydroxymethyl)aminomethane sodium chloride Assay. Gas chromatography (2.2.28) as prescribed in the
buffer solution pH 7.4 R. monograph Bitter fennel (0824).
Any change in the absorbance of the solution, measured at Test solution. The substance to be examined.
405 nm (2.2.25) against tris(hydroxymethyl)aminomethane Content : minimum 98.0 per cent, calculated by the
sodium chloride buffer solution pH 7.4 R and from which normalisation procedure.
the blank absorbance has been substracted, is not more Fenvalerate. C25H22ClNO3. (Mr 419.9). 1127300.
than 0.20 per minute. [51630-58-1].
Factor Xa solution, bovine R1. 1037302. bp : about 300 °C.
Reconstitute as directed by the manufacturer and dilute to A suitable certified reference solution (10 ng/µL in
1.4 nkat/mL with tris(hydroxymethyl)aminomethane-EDTA cyclohexane) may be used.
buffer solution pH 8.4 R.
Ferric ammonium sulfate. FeNH4(SO4)2,12H2O. (Mr 482.2).
Factor Xa solution, bovine R2. 1037303. 1037700. [7783-83-7]. Ammonium iron disulfate
Reconstitute as directed by the manufacturer and dilute dodecahydrate.
with tris(hydroxymethyl)aminomethane-EDTA buffer Pale-violet crystals, efflorescent, very soluble in water,
solution pH 8.4 R1 to obtain a solution that gives an practically insoluble in ethanol (96 per cent).
absorbance between 0.65 and 1.25 at 405 nm when
determining the blank amidolytic activity according to Ferric ammonium sulfate solution R2. 1037702.
general chapter 2.7.5 using the end-point method. A 100 g/L solution of ferric ammonium sulfate R. If
necessary filter before use.
Fargesin. C21H22O6. (Mr 370.4). 1200200. [31008-19-2].
5-[(3SR,3aRS,6RS,6aRS)-6-(3,4-Dimethoxyphenyl)- Ferric ammonium sulfate solution R5. 1037704.
1,3,3a,4,6,6a-hexahydrofuro[3,4-c]furan-3-yl]-1,3- Shake 30.0 g of ferric ammonium sulfate R with 40 mL
benzodioxole. of nitric acid R and dilute to 100 mL with water R. If the
solution is turbid, centrifuge or filter it.
(E,E)-Farnesol. C15H26O. (Mr 222.4). 1161000. [106-28-5].
trans,trans-Farnesol. (2E,6E)-3,7,11-Trimethyldodeca-2,6,10- Storage : protected from light.
trien-1-ol. Ferric ammonium sulfate solution R6. 1037705.
Fast blue B salt. C14H12Cl2N4O2. (Mr 339.2). 1037400. Dissolve 20 g of ferric ammonium sulfate R in 75 mL of
[84633-94-3]. water R, add 10 mL of a 2.8 per cent V/V solution of sulfuric
Schultz No. 490. acid R and dilute to 100 mL with water R.
Colour Index No. 37235. Ferric chloride. FeCl3,6H2O. (Mr 270.3). 1037800.
3,3′-Dimethoxy(biphenyl)-4,4′-bisdiazonium dichloride. [10025-77-1]. Iron trichloride hexahydrate.
Dark green powder, soluble in water. It is stabilised by Yellowish-orange or brownish crystalline masses, deliquescent,
addition of zinc chloride. very soluble in water, soluble in ethanol (96 per cent). On
Storage : in an airtight container, at a temperature between exposure to light, ferric chloride and its solutions are partly
2 °C and 8 °C. reduced.
Fast blue B salt solution. 1037401. Storage : in an airtight container.
Dissolve 140 mg of fast blue B salt R in 10 mL of water R Ferric chloride solution R1. 1037801.
and mix with 50 mL of methylene chloride R and 140 mL of A 105 g/L solution of ferric chloride R.
methanol R.
Storage : protected from light at a temperature of 4 °C ; use Ferric chloride solution R2. 1037802.
within 1 week. A 13 g/L solution of ferric chloride R.

Ferric chloride solution R3. 1037803. Assay. Liquid chromatography (2.2.29) as prescribed in the
Dissolve 2.0 g of ferric chloride R in anhydrous ethanol R monograph Eleutherococcus (1419).
and dilute to 100.0 mL with the same solvent. Content : minimum 99 per cent, calculated by the
Ferric chloride-ferricyanide-arsenite reagent. 1037805.
Immediately before use mix 10 mL of a 27 g/L solution Fibrin blue. 1101400.
of ferric chloride R in dilute hydrochloric acid R, 7 mL of Mix 1.5 g of fibrin with 30 mL of a 5 g/L solution of indigo
potassium ferricyanide solution R, 3 mL of water R and carmine R in 1 per cent V/V dilute hydrochloric acid R.
10 mL of sodium arsenite solution R. Heat the mixture to 80 °C and maintain at this temperature
whilst stirring for about 30 min. Allow to cool. Filter.
Ferric chloride-sulfamic acid reagent. 1037804.
Wash extensively by resuspension in 1 per cent V/V dilute
A solution containing 10 g/L of ferric chloride R and 16 g/L hydrochloric acid R and mixing for about 30 min ; filter. Repeat
of sulfamic acid R. the washing operation three times. Dry at 50 °C. Grind.
Ferric nitrate. Fe(NO3)3,9H2O. (Mr 404). 1106100. Fibrin congo red. 1038400.
[7782-61-8].
Take 1.5 g of fibrin and leave overnight in 50 mL of a 20 g/L
Content : minimum 99.0 per cent m/m of Fe(NO3)3,9H2O. solution of congo red R in ethanol (90 per cent V/V) R. Filter,
Light-purple crystals or crystalline mass, very soluble in water. rinse the fibrin with water R and store under ether R.
Free acid : not more than 0.3 per cent (as HNO3).
Fibrinogen. 1038500. [9001-32-5].
Ferric sulfate. Fe2(SO4)3,xH2O. 1037900. [15244-10-7]. See Human fibrinogen, freeze-dried (0024).
Iron(III) trisulfate hydrated.
Yellowish-white powder, very hygroscopic, decomposes in air, Fixing solution. 1122600.
slightly soluble in water and in ethanol (96 per cent). To 250 mL of methanol R, add 0.27 mL of formaldehyde R and
Storage : in an airtight container, protected from light. dilute to 500.0 mL with water R.
Ferric sulfate solution. 1037901. Fixing solution for isoelectric focusing in polyacrylamide
Dissolve 50 g of ferric sulfate R in an excess of water R, gel. 1138700.
add 200 mL of sulfuric acid R and dilute to 1000 mL with A solution containing 35 g of sulfosalicylic acid R and 100 g of
water R. trichloroacetic acid R per litre of water R.
Ferric sulfate pentahydrate. Fe2(SO4)3,5H2O. (Mr 489.9). Flufenamic acid. C14H10F3NO2. (Mr 281.2). 1106200.
1153700. [142906-29-4]. [530-78-9]. 2-[[3-(Trifluoromethyl)phenyl]amino]benzoic
White or yellowish powder. acid.
Pale yellow, crystalline powder or needles, practically insoluble
Ferrocyphene. C26H16FeN6. (Mr 468.3). 1038000. in water, freely soluble in ethanol (96 per cent).
[14768-11-7]. Dicyanobis(1,10-phenanthroline)iron(II).
mp : 132 °C to 135 °C.
Violet-bronze, crystalline powder, practically insoluble in
water and in ethanol (96 per cent). Flumazenil. 1149600. [78755-81-4].
Storage : protected from light and moisture. See Flumazenil (1326).
Ferroin. 1038100. [14634-91-4]. Flunitrazepam. 1153800. [1622-62-4].
Dissolve 0.7 g of ferrous sulfate R and 1.76 g of phenanthroline See Flunitrazepam (0717).
hydrochloride R in 70 mL of water R and dilute to 100 mL
with the same solvent. Fluorene. C13H10. (Mr 166.2). 1127400. [86-73-7].
Test for sensitivity. To 50 mL of dilute sulfuric acid R add Diphenylenemethane.
0.1 mL of ferroin R. After the addition of 0.1 mL of 0.1 M White or almost white crystals, freely soluble in anhydrous
ammonium and cerium nitrate the colour changes from red acetic acid, soluble in hot ethanol (96 per cent).
to light blue. mp : 113 °C to 115 °C.
Ferrous ammonium sulfate. Fe(NH4)2(SO4)2,6H2O. (9-Fluorenyl)methyl chloroformate. C15H11ClO2.
(Mr 392.2). 1038200. [7783-85-9]. Diammonium iron (Mr 258.7). 1180100. [28920-43-6]. Fluoren-9-ylmethyl
disulfate hexahydrate. chloromethanoate.
Pale bluish-green crystals or granules, freely soluble in water, mp : about 63 °C.
practically insoluble in ethanol (96 per cent).
Storage : protected from light. Fluorescamine. C17H10O4. (Mr 278.3). 1135800. [38183-12-9].
4-Phenylspiro[furan-2(3H),1’(3’H)-isobenzofuran]-3,3’-
Ferrous sulfate. 1038300. [7782-63-0]. dione.
See Ferrous sulfate heptahydrate (0083). mp : 154 °C to 155 °C.
Ferrous sulfate solution R2. 1038301. Fluorescein. C20H12O5. (Mr 332.3). 1106300. [2321-07-5].
Dissolve 0.45 g of ferrous sulfate R in 50 mL of 0.1 M 3′,6′-Dihydroxyspiro[isobenzofurane-1(3H),9′-[9H]xanthen]-
hydrochloric acid and dilute to 100 mL with carbon 3-one.
dioxide-free water R. Prepare immediately before use. Orange-red powder, practically insoluble in water, soluble in
warm ethanol (96 per cent), soluble in alkaline solutions. In
Ferulic acid. C10H10O4. (Mr 194.2). 1149500.
solution, fluorescein displays a green fluorescence.
[1135-24-6]. 4-Hydroxy-3-methoxycinnamic acid.
3-(4-Hydroxy-3-methoxyphenyl)propenoic acid. mp : about 315 °C.
Faint yellow powder, freely soluble in methanol. Fluorescein-conjugated rabies antiserum. 1038700.
mp : 172.9 °C to 173.9 °C. Immunoglobulin fraction with a high rabies antibody titre,
Ferulic acid used in the assay of eleutherosides in prepared from the sera of suitable animals that have been
Eleutherococcus (1419) complies with the following additional immunised with inactivated rabies virus ; the immunoglobulin
test. is conjugated with fluorescein isothiocyanate.

Fluorocholine chloride. C5H13ClFNO. (Mr 157.6). Formaldehyde. 1039100. [50-00-0].

1195700. [459424-38-5]. N-(Fluoromethyl)-2-hydroxy-N,N- See Formaldehyde solution R.
dimethylethan-1-aminium chloride.
Colourless, hygroscopic crystals. Formaldehyde solution. 1039101.
mp : about 184 °C. See Formaldehyde solution (35 per cent) (0826).
Formaldehyde solution R1. 1039102.
2-Fluoro-2-deoxy- D-glucose. C6H11FO5. (Mr 182.2).
1113900. [86783-82-6]. Complies with the requirements prescribed in the monograph
Formaldehyde solution (35 per cent) (0826) with the following
White or almost white crystalline powder. modification.
mp : 174 °C to 176 °C. Content : 36.5 per cent m/m to 38.0 per cent m/m of
2-Fluoro-2-deoxy- D-mannose. C6H11FO5. (Mr 182.1). formaldehyde (CH2O ; Mr 30.03).
1172100. [38440-79-8]. Formamide. CH3NO. (Mr 45.0). 1039200. [75-12-7].
Colourless semi-solid. Clear, colourless, oily liquid, hygroscopic, miscible with water
Fluorodinitrobenzene. C6H3FN2O4. (Mr 186.1). 1038800. and with ethanol (96 per cent). It is hydrolysed by water.
[70-34-8]. 1-Fluoro-2,4-dinitrobenzene. : about 1.134.
Pale yellow liquid or crystals, soluble in propylene glycol. bp : about 210 °C.
mp : about 29 °C. Content : minimum 99.5 per cent.
Content : minimum 99.0 per cent, determined by gas Storage : in an airtight container.
chromatography. Formamide R1. 1039202.
1-Fluoro-2,4-dinitrophenyl-5- L-alaninamide. Complies with the requirements prescribed for
C9H9FN4O5. (Mr 272.2). 1194900. [95713-52-3]. formamide R with the following additional requirement.
Nα-(5-Fluoro-2,4-dinitrophenyl)-L-alaninamide. Marfey’s Water (2.5.12): maximum 0.1 per cent determined with an
reagent. FDAA. equal volume of anhydrous methanol R.
Yellow or orange powder.
Formamide, treated. 1039201.
mp : about 228 °C. Disperse 1.0 g of sulfamic acid R in 20.0 mL of formamide R
Enantiomeric purity : minimum 99.5 per cent. containing 5 per cent V/V of water R.
DL-6-Fluorodopa hydrochloride. C9H11ClFNO4. Formic acid, anhydrous. CH2O2. (Mr 46.03). 1039300.
(Mr 251.6). 1169200. (2RS)-2-Amino-3-(2-fluoro- [64-18-6].
4,5-dihydroxyphenyl)propanoic acid hydrochloride. Content : minimum 98.0 per cent m/m.
2-Fluoro-5-hydroxy-DL-tyrosine hydrochloride.
Colourless liquid, corrosive, miscible with water and with
White or almost white powder. ethanol (96 per cent).
Fluoroethyl(2-hydroxyethyl)dimethylammonium : about 1.22.
chloride. C6H15ClFNO. (Mr 171.6). 1195800. [479407-08-4]. Assay. Weigh accurately a conical flask containing 10 mL of
N-(2-Fluoroethyl)-2-hydroxy-N,N-dimethylethan-1-aminium water R, quickly add about 1 mL of the acid and weigh again.
chloride. Add 50 mL of water R and titrate with 1 M sodium hydroxide,
Slightly yellow powder. using 0.5 mL of phenolphthalein solution R as indicator.
1 mL of 1 M sodium hydroxide is equivalent to 46.03 mg of
Fluoroethyl- D-tyrosine hydrochloride. C11H15FNO3Cl. CH2O2.
(Mr 263.7). 1192000. (2R)-2-Amino-3-[4-(2-
fluoroethoxy)phenyl]propanoic acid hydrochloride. Fructose. 1106400. [57-48-7].
Content : minimum 95 per cent. See Fructose (0188).
Colourless or almost colourless crystals. Fuchsin, basic. 1039400. [632-99-5].
Fluoroethyl- L-tyrosine hydrochloride. C11H15FNO3Cl. A mixture of rosaniline hydrochloride (C20H20ClN3 ; Mr 337.9 ;
(Mr 263.7). 1192100. (2S)-2-Amino-3-[4-(2- Colour Index No. 42510 ; Schultz No. 780) and para-rosaniline
fluoroethoxy)phenyl]propanoic acid hydrochloride. hydrochloride (C19H18ClN3 ; Mr 323.8 ; Colour Index
Content : minimum 95 per cent. No. 42500 ; Schultz No. 779).
Colourless or almost colourless crystals. If necessary, purify in the following manner. Dissolve 1 g in
250 mL of dilute hydrochloric acid R. Allow to stand for 2 h at
6-Fluorolevodopa hydrochloride. C9H11ClFNO4. (Mr 251.6). room temperature, filter and neutralise with dilute sodium
1169300. [144334-59-8]. (2S)-2-Amino-3-(2-fluoro- hydroxide solution R and add 1 mL to 2 mL in excess. Filter
4,5-dihydroxyphenyl)propanoic acid hydrochloride. the precipitate through a sintered-glass filter (40) (2.1.2)
2-Fluoro-5-hydroxy-L-tyrosine hydrochloride. and wash with water R. Dissolve the precipitate in 70 mL of
Colourless or almost colourless solid, soluble in water. methanol R, previously heated to boiling, and add 300 mL of
water R at 80 °C. Allow to cool to room temperature, filter
Fluoromisonidazole. C6H8FN3O3. (Mr 189.1). 1186000. and dry the crystals in vacuo.
[13551-89-8]. (2RS)-1-Fluoro-3-(2-nitro-1H-imidazol-1- Crystals with a greenish-bronze sheen, soluble in water and in
yl)propan-2-ol. FMISO. ethanol (96 per cent).
Content : minimum 95 per cent. Storage : protected from light.
Yellow crystals.
Fuchsin solution, decolorised. 1039401.
1-Fluoro-2-nitro-4-(trifluoromethyl)benzene. C7H3F4NO2. Dissolve 0.1 g of basic fuchsin R in 60 mL of water R. Add
(Mr 209.1). 1038900. [367-86-2]. a solution containing 1 g of anhydrous sodium sulfite R
mp : about 197 °C. or 2 g of sodium sulfite heptahydrate R in 10 mL of
water R. Slowly and with continuous shaking add 2 mL of
Folic acid. 1039000. [75708-92-8]. hydrochloric acid R. Dilute to 100 mL with water R. Allow
See Folic acid hydrate (0067). to stand protected from light for at least 12 h, decolorise

with activated charcoal R and filter. If the solution becomes Gallium (68Ga) chloride solution. 68GaCl3. (Mr 174.3).
cloudy, filter before use. If on standing the solution becomes 1182500.
violet, decolorise again by adding activated charcoal R. Solution containing gallium-68 in the form of gallium chloride
Test for sensitivity. To 1.0 mL add 1.0 mL of water R and in dilute hydrochloric acid R.
0.1 mL of aldehyde-free alcohol R. Add 0.2 mL of a solution Content : 90 per cent to 110 per cent of the declared gallium-68
containing 0.1 g/L of formaldehyde (CH2O, Mr 30.03). A radioactivity at the date and time stated on the label.
pale-pink colour develops within 5 min.
Storage : protected from light. Gastric juice, artificial. 1039900.
Dissolve 2.0 g of sodium chloride R and 3.2 g of pepsin
Fuchsin solution, decolorised R1. 1039402. powder R in water R. Add 80 mL of 1 M hydrochloric acid and
To 1 g of basic fuchsin R add 100 mL of water R. Heat to dilute to 1000 mL with water R.
50 °C and allow to cool with occasional shaking. Allow to
stand for 48 h, shake and filter. To 4 mL of the filtrate add Gastrodin. C13H18O7. (Mr 286.3). 1203600. [62499-27-8].
6 mL of hydrochloric acid R, mix and dilute to 100 mL with 4-(Hydroxymethyl)phenyl α-D-glucopyranoside.
water R. Allow to stand for at least 1 h before use. (2R,3S,4S,5R,6S)-2-(Hydroxymethyl)-6-[4-(hydroxymethyl)-
phenoxy]oxane-3,4,5-triol.
Fucose. C6H12O5. (Mr 164.2). 1039500. [6696-41-9].
6-Deoxy-L-galactose. GC concentrical column. 1135100.
White or almost white powder, soluble in water and in ethanol A commercially available system consisting of 2 concentrically
(96 per cent). arranged tubes. The outer tube is packed with molecular
sieves and the inner tube is packed with a porous polymer
: about − 76, determined on a 90 g/L solution 24 h after mixture. The main application is the separation of gases.
dissolution.
mp : about 140 °C. Gelatin. 1040000. [9000-70-8].
See Gelatin (0330).
Fumaric acid. C4H4O4. (Mr 116.1). 1153200. [110-17-8].
(E)-Butenedioic acid. Gelatin, hydrolysed. 1040100.
White or almost white crystals, slightly soluble in water, Dissolve 50 g of gelatin R in 1000 mL of water R. Autoclave in
soluble in ethanol (96 per cent), slightly soluble in acetone. saturated steam at 121 °C for 90 min and freeze dry.
mp : about 300 °C. Geniposide. C17H24O10. (Mr 388.4). 1196800. [24512-63-8].
Furfural. C5H4O2. (Mr 96.1). 1039600. [98-01-1]. Methyl (1S,4aS,7aS)-1-(β-D-glucopyranosyloxy)-7-
2-Furaldehyde. 2-Furanecarbaldehyde. (hydroxymethyl)-1,4a,5,7a-tetrahydrocyclopenta[c]pyran-4-
Clear, colourless to brownish-yellow, oily liquid, miscible in carboxylate.
11 parts of water, miscible with ethanol (96 per cent). Geraniol. C10H18O. (Mr 154.2). 1135900. [106-24-1].
: 1.155 to 1.161. (E)-3,7-Dimethylocta-2,6-dien-1-ol.
Distillation range (2.2.11). Not less than 95 per cent distils Oily liquid, slight odour of rose, practically insoluble in water,
between 159 °C and 163 °C. miscible with ethanol (96 per cent).
Storage : in a dark place. Geraniol used in gas chromatography complies with the
Gadolinium chloride hexahydrate. GdCl3,6H2O. (Mr 371.7).
1198400. [13450-84-5]. Gadolinium trichloride hexahydrate. Assay. Gas chromatography (2.2.28) as prescribed in the
monograph Citronella oil (1609).
Gadolinium sulfate octahydrate. Gd2(SO4)3,8H2O. (Mr 747). normalisation procedure.
1195300. [13450-87-8]. Storage : in an airtight container, protected from light
Colourless, crystalline powder.
Geranyl acetate. C12H20O2. (Mr 196.3). 1106500. [105-87-3].
Galactose. C6H12O6. (Mr 180.2). 1039700. [59-23-4]. (E)-3,7-Dimethylocta-2,6-dien-1-yl acetate.
D-(+)-Galactose. Colourless or slightly yellow liquid, slight odour of rose and
White or almost white, crystalline powder, freely soluble in lavender.
water. Geranyl acetate used in gas chromatography complies with the
: + 79 to + 81, determined on a 100 g/L solution in following additional test.
water R containing about 0.05 per cent of NH3. Assay. Gas chromatography (2.2.28) as prescribed in the
1,6-Galactosylgalactose. C12H22O11. (Mr 342.3). 1195900. monograph Bitter-orange-flower oil (1175).
[5077-31-6]. 6-O-β-D-Galactopyranosyl-D-galactopyranose. Test solution. The substance to be examined.
White or almost white powder. Content : minimum 98.0 per cent, calculated by the
Galacturonic acid. C6H10O7. (Mr 194.1). 1196000.
[685-73-4]. D-(+)-galacturonic acid. (2S,3R,4S,5R)-2,3,4,5- Ginsenoside Rb1. C54H92O23,3H2O. (Mr 1163). 1127500.
Tetrahydroxy-6-oxo-hexanoic acid. [41753-43-9]. (20S)-3β-Di-D-glucopyranosyl-20-di-D-
: about + 53°, determined on a 100 g/L solution. glucopyranosylprotopanaxadiol. (20S)-3β-[(2-O-β-D-
Glucopyranosyl-β-D-glucopyranosyl)oxy]-20-[(6-O-β-D-
Gallic acid. C7H6O5,H2O. (Mr 188.1). 1039800. [5995-86-8]. glucopyranosyl-β-D-glucopyranosyl)oxy]-5α-dammar-
3,4,5-Trihydroxybenzoic acid monohydrate. 24-en-12β-ol. (20S)-3β-[(2-O-β-D-Glucopyranosyl-β-D-
Crystalline powder or long needles, colourless or slightly glucopyranosyl)oxy]-20-[(6-O-β-D-glucopyranosyl-β-D-
yellow, soluble in water, freely soluble in hot water, in ethanol glucopyranosyl)oxy]-4,4,8,14-tetramethyl-18-nor-5α-cholest-
(96 per cent) and in glycerol. 24-en-12β-ol.
It loses its water of crystallisation at 120 °C. A colourless solid, soluble in water, in anhydrous ethanol and
mp : about 260 °C, with decomposition. in methanol.
Chromatography. Thin-layer chromatography (2.2.27) as : + 11.3 determined on a 10 g/L solution in methanol R.
prescribed in the monograph Bearberry leaf (1054) ; the mp : about 199 °C.
chromatogram shows only one principal spot. Water (2.5.12) : maximum 6.8 per cent.

Assay. Liquid chromatography (2.2.29) as prescribed in the D-Glucuronic acid. C6H10O7. (Mr 194.1). 1119700.
monograph Ginseng (1523). [6556-12-3].
Test solution. Dissolve 3.0 mg, accurately weighed, of Content : minimum 96.0 per cent, calculated with reference to
ginsenoside Rb1 in 10 mL of methanol R. the substance dried in vacuo (2.2.32).
Content : minimum 95.0 per cent, calculated by the Soluble in water and in ethanol (96 per cent).
normalisation procedure. Shows mutarotation : : + 11.7 → + 36.3.
Ginsenoside Re. C48H82O18. (Mr 947.2). 1157800. Assay. Dissolve 0.150 g in 50 mL of anhydrous methanol R
[52286-59-6]. (3β,6α,12β)-20-(β-D-Glucopyranosyloxy)- while stirring under nitrogen. Titrate with 0.1 M
3,12-dihydroxydammar-24-en-6-yl 2-O-(6-deoxy-α-L- tetrabutylammonium hydroxide, protecting the solution from
mannopyranosyl)-β-D-glucopyranoside. atmospheric carbon dioxide throughout solubilisation and
titration. Determine the end-point potentiometrically (2.2.20).
Colourless solid, soluble in water, in ethanol (96 per cent) and
in methanol. 1 mL of 0.1 M tetrabutylammonium hydroxide is equivalent to
19.41 mg of C6H10O7.
Ginsenoside Rf. C42H72O14,2H2O. (Mr 837). 1127700.
[52286-58-5]. (20S)-6-O-[β-D-Glucopyranosyl-(1→2)-β-D- Glutamic acid. 1040400. [56-86-0].
glycopyranoside]-dammar-24-ene-3β,6α,12β,20-tetrol. See Glutamic acid (0750).
A colourless solid, soluble in water, in anhydrous ethanol and L-Glutamine. C5H10N2O3. (Mr 146.2). 1203700. [56-85-9].
in methanol. (S)-2,5-Diamino-5-oxopentanoic acid.
: + 12.8 determined on a 10 g/L solution in methanol R. White crystalline powder.
mp : about 198 °C. mp : about 185 °C, with decomposition.
Ginsenoside Rg1. C42H72O14,2H2O. (Mr 837). Glutamyl endopeptidase for peptide mapping. 1173300.
1127600. [22427-39-0]. (20S)-6β-D-Glucopyranosyl- [137010-42-5]. Endoproteinase Glu-C of high purity from
D-glucopyranosylprotopanaxatriol. (20S)-6α,20-Bis(β- Staphylococcus aureus strain V8 (EC 3.4.21.19).
D-glucopyranosyloxy)-5α-dammar-24-ene-3β,12β-diol.
(20S)-6α,20-Bis(β-D-glucopyranosyloxy)-4,4,8,14- L-γ-Glutamyl- L-cysteine. C8H14N2O5S. (Mr 250.3). 1157900.
tetramethyl-18-nor-5α-cholest-24-ene-3β,12β-diol. [636-58-8].
A colourless solid, soluble in water, in anhydrous ethanol and Glutaraldehyde. C5H8O2. (Mr 100.1). 1098300. [111-30-8].
in methanol. Oily liquid, soluble in water.
: + 31.2 determined on a 10 g/L solution in methanol R. : about 1.434.
mp : 188 °C to 191 °C. bp : about 188 °C.
Water (2.5.12) : maximum 4.8 per cent.
Glutaric acid. C5H8O4. (Mr 132.1). 1149700. [110-94-1].
Assay. Liquid chromatography (2.2.29) as prescribed in the Pentanedioic acid.
monograph Ginseng (1523).
Test solution. Dissolve 3.0 mg, accurately weighed, of
ginsenoside Rg1 in 10 mL of methanol R. L-Glutathione, oxidised. C20H32N6O12S2. (Mr 612.6).
Content : minimum 95.0 per cent, calculated by the 1158000. [27025-41-8]. Bis(L-γ-glutamyl-L-cysteinylglycine)
normalisation procedure. disulfide.
Ginsenoside Rg2. C42H72O13. (Mr 785). 1182600. Glycerol. 1040500. [56-81-5].

[52286-74-5]. 3β,12β,20-Trihydroxydammar-24-en-6α-yl See Glycerol (0496).
2-O-(6-deoxy-α-L-mannopyranosyl)-β-D-glucopyranoside.
Glycerol R1. 1040501.
Ginsenoside Ro. C48H76O19. (Mr 957). 1205000. [34367-04-9]. Complies with the requirements prescribed for the
(3β)-28-(β-D-Glucopyranosyloxy)-28-oxoolean-12-en-3-yl monograph Glycerol (0496) and free from diethylene glycol
2-O-β-D-glucopyranosyl-β-D-glucopyranosiduronic acid. when examined as prescribed in the test for impurity A and
related substances in that monograph.
Gitoxin. C41H64O14. (Mr 781). 1040200. [4562-36-1].
Glycoside of Digitalis purpurea L. 3β-(O-2,6-Dideoxy- Glycerol (85 per cent). 1040600.
β-d-ribo-hexopyranosyl-(1→4)-O-2,6-dideoxy-β-d- See Glycerol (85 per cent) (0497).
ribo-hexopyranosyl-(1→4)-2,6-dideoxy-β-d-ribo-
hexopyranosyloxy)-14,16β-dihydroxy-5β,14β-card-20(22)- Glycerol (85 per cent) R1. 1040601.
enolide. Complies with the requirements prescribed for the
A white or almost white, crystalline powder, practically monograph Glycerol 85 per cent (0497) and free from
insoluble in water and in most common organic solvents, diethylene glycol when examined as prescribed in the test
soluble in pyridine. for impurity A and related substances in that monograph.
: + 20 to + 24, determined on a 5 g/L solution in a Glycerol 1-decanoate. C13H26O4. (Mr 246.3). 1169400.
mixture of equal volumes of chloroform R and methanol R. [2277-23-8]. (2RS)-2,3-Dihydroxypropyl decanoate.
Chromatography. Thin-layer chromatography (2.2.27) α-Monocaprin. 1-Monodecanoyl-rac-glycerol.
as prescribed in the monograph Digitalis leaf (0117) ; the Content : about 99 per cent.
chromatogram shows only one principal spot.
Glycerol 1-octanoate. C11H22O4. (Mr 218.3). 1169500.
Glucosamine hydrochloride. C6H14ClNO5. (Mr 215.6). [502-54-5]. (2RS)-2,3-Dihydroxypropyl octanoate.
1040300. [66-84-2]. D-Glucosamine hydrochloride. α-Monocaprylin. 1-Monooctanoyl-rac-glycerol.
Crystals, soluble in water. Content : about 99 per cent.
: + 100, decreasing to + 47.5 after 30 min, determined Glycidol. C3H6O2. (Mr 74.1). 1127800. [556-52-5].
on a 100 g/L solution.
Slightly viscous liquid, miscible with water.
Glucose. 1025700. [50-99-7]. : about 1.115.
See Glucose (0177). : about 1.432.

Glycine. 1040700. [56-40-6]. Guaiacol. C7H8O2. (Mr 124.1). 1148300. [90-05-1].

See Glycine (0614). 2-Methoxyphenol. 1-Hydroxy-2-methoxybenzene.
Crystalline mass or colourless or yellowish liquid, hygroscopic,
Glycine anhydride. C4H6N2O2. (Mr 114.1). 1192200. slightly soluble in water, very soluble in methylene chloride,
[106-57-0]. Piperazine-2,5-dione (2,5-DKP). freely soluble in ethanol (96 per cent).
Glycolic acid. C2H4O3. (Mr 76.0). 1040800. [79-14-1]. bp : about 205 °C.
2-Hydroxyacetic acid. mp : about 28 °C.
Crystals, soluble in water, in acetone, in ethanol (96 per cent) Guaiacum resin. 1041400.
and in methanol.
Resin obtained from the heartwood of Guaiacum officinale L.
mp : about 80 °C. and Guaiacum sanctum L.
Glycyrrhetic acid. C30H46O4. (Mr 470.7). 1040900. Reddish-brown or greenish-brown, hard, glassy fragments ;
[471-53-4]. Glycyrrhetinic acid. 12,13-Didehydro-3β- fracture shiny.
hydroxy-11-oxo-olean-30-oic acid.
Guaiazulene. C15H18. (Mr 198.3). 1041500. [489-84-9].
A mixture of α- and β-glycyrrhetic acids in which the β-isomer 1,4-Dimethyl-7-isopropylazulene.
is predominant.
Dark-blue crystals or blue liquid, very slightly soluble in water,
White or yellowish-brown powder, practically insoluble in miscible with fatty and essential oils and with liquid paraffin,
water, soluble in anhydrous ethanol and in glacial acetic acid. sparingly soluble in ethanol (96 per cent), soluble in 500 g/L
: + 145 to + 155, determined on a 10.0 g/L solution in sulfuric acid and 80 per cent m/m phosphoric acid, giving a
anhydrous ethanol R. colourless solution.
Chromatography. Thin-layer chromatography (2.2.27) using mp : about 30 °C.
silica gel GF254 R as the coating substance ; prepare the slurry Storage : protected from light and air.
using a 0.25 per cent V/V solution of phosphoric acid R.
Apply to the plate 5 μL of a 5 g/L solution of the glycyrrhetic Guanidine hydrochloride. CH5N3HCl. (Mr 95.5). 1098500.
acid in a mixture of equal volumes of chloroform R and [50-01-1].
methanol R. Develop over a path of 10 cm using a mixture Crystalline powder, freely soluble in water and in ethanol
of 5 volumes of methanol R and 95 volumes of chloroform R. (96 per cent).
Examine the chromatogram in ultraviolet light at 254 nm. The
chromatogram shows a dark spot (RF about 0.3) corresponding Guanine. C5H5N5O. (Mr 151.1). 1041600. [73-40-5].
to β-glycyrrhetic acid and a smaller spot (RF about 0.5) 2-Amino-1,7-dihydro-6H-purin-6-one.
corresponding to α-glycyrrhetic acid. Spray with anisaldehyde Amorphous white or almost white powder, practically
solution R and heat at 100-105 °C for 10 min. Both spots are insoluble in water, slightly soluble in ethanol (96 per cent).
coloured bluish-violet. Between them a smaller bluish-violet It dissolves in ammonia and in dilute solutions of alkali
spot may be present. hydroxides.
18α-Glycyrrhetinic acid. C30H46O4. (Mr 470.7). 1127900. Haemoglobin. 1041700. [9008-02-0].
[1449-05-4]. (20β)-3β-Hydroxy-11-oxo-18α-olean-12-en-29- Nitrogen : 15 per cent to 16 per cent.
oic acid. Iron : 0.2 per cent to 0.3 per cent.
White or almost white powder, practically insoluble in water, Loss on drying (2.2.32) : maximum 2 per cent.
soluble in anhydrous ethanol, sparingly soluble in methylene
Sulfated ash (2.4.14) : maximum 1.5 per cent.
chloride.
Haemoglobin solution. 1041701.
Glyoxalhydroxyanil. C14H12N2O2. (Mr 240.3). 1041000.
[1149-16-2]. Glyoxal bis(2-hydroxyanil). Transfer 2 g of haemoglobin R to a 250 mL beaker and add
75 mL of dilute hydrochloric acid R2. Stir until solution is
White or almost white crystals, soluble in hot ethanol (96 per complete. Adjust the pH to 1.6 ± 0.1 using 1 M hydrochloric
cent). acid. Transfer to a 100 mL flask with the aid of dilute
mp : about 200 °C. hydrochloric acid R2. Add 25 mg of thiomersal R. Prepare
daily, store at 5 ± 3 °C and readjust to pH 1.6 before use.
Glyoxal solution. 1098400. [107-22-2].
Storage : at 2 °C to 8 °C.
Contains about 40 per cent ( m/m) glyoxal.
Assay. In a ground-glass stoppered flask place 1.000 g of Hamamelitannin. C20H20O14. (Mr 484.4). 1192700.
glyoxal solution, 20 mL of a 70 g/L solution of hydroxylamine [469-32-9]. (2R,3R,4R)-2-Formyl-2,3,4-trihydroxy-
hydrochloride R and 50 mL of water R. Allow to stand for pentane-1,5-diyl bis(3,4,5-trihydroxybenzoate).
30 min and add 1 mL of methyl red mixed solution R and 2-C-[(Galloyloxy)methyl]-D-ribose 5-gallate.
titrate with 1 M sodium hydroxide until the colour changes Harpagoside. C24H30O11. (Mr 494.5). 1098600.
from red to green. Carry out a blank titration.
White or almost white, crystalline powder, very hygroscopic,
1 mL of 1 M sodium hydroxide is equivalent to 29.02 mg of soluble in water and in ethanol (96 per cent).
glyoxal (C2H2O2).
mp : 117 °C to 121 °C.
Gonadotrophin, chorionic. 1041100. [9002-61-3]. Storage : in an airtight container.
See Chorionic gonadotrophin (0498).
Hederacoside C. C59H96O26. (Mr 1221). 1158100.
Gonadotrophin, serum. 1041200. [14216-03-6]. O-6-Deoxy-α-L-mannopyranosyl-(1→4)-
See Equine serum gonadotrophin for veterinary use (0719). O-β-D-glucopyranosyl-(1→6)-β-D-glucopyranosyl
(4R)-3β-[[2-O(-6-deoxy-α-L-mannopyranosyl)-α-L-
Gramine. C11H14N2. (Mr 174.2). 1189400. [87-52-5]. arabinopyranosyl]oxy]-23-hydroxyolean-12-en-28-oate.
1-(1H-Indol-3-yl)-N,N-dimethylmethanamine. Colourless crystals or white or almost white powder.
Flakes, practically insoluble in water, soluble in ethanol mp : about 220 °C.
(96 per cent), slightly soluble in acetone. Hederacoside C used in liquid chromatography complies with
mp : 132 °C to 134 °C. the following additional test.

Assay. Liquid chromatography (2.2.29) as prescribed in the : about 1.300.

monograph Ivy leaf (2148). bp : about 120 °C.
Test solution. Dissolve 5.0 mg of hederacoside C in 5.0 mL Content : minimum 99.5 per cent.
of methanol R.
Content : minimum 95 per cent, calculated by the Heptafluoro-N-methyl-N-(trimethylsilyl)butanamide.
normalisation procedure. C8H12F7NOSi. (Mr 299.3). 1139500. [53296-64-3].
2,2,3,3,4,4,4-Heptafluoro-N-methyl-N-(trimethylsilyl)-
Hederagenin. C30H48O4. (Mr 472.7). 1184100. butyramide.
[465-99-6]. Astrantiagenin E. Caulosapogenin. Clear, colourless liquid, flammable.
3β,23-Dihydroxy-4α-olean-12-en-28-oic acid.
: about 1.351.
α-Hederin. C41H66O12. (Mr 751.0). 1158200. [27013-91-8]. bp : about 148 °C.
(+)-(4R)-3β-[[2-O-(6-Deoxy-α-L-mannopyranosyl)-α-L-
arabinopyranosyl]oxy]-23-hydroxyolean-12-en-28-oic acid. Heptane. C7H16. (Mr 100.2). 1042000. [142-82-5].
White or almost white powder. Colourless, flammable liquid, practically insoluble in water,
mp : about 256 °C. miscible with anhydrous ethanol.
: 0.683 to 0.686.
Helium for chromatography. He. (Ar 4.003). 1041800. : 1.387 to 1.388.
[7440-59-7].
Content : minimum 99.995 per cent V/V of He. between 97 °C and 98 °C.
Heparin. 1041900. [9041-08-1]. Hesperidin. C28H34O15. (Mr 611). 1139000. [520-26-3].
See Heparin sodium (0333). (S)-7-[[6-O-(6-Deoxy-α-L-mannopyranosyl)-β-D-
glucopyranosyl]oxy]-5-hydroxy-2-(3-hydroxy-4-
Heparinase I. 1187600. [9025-39-2]. Heparin lyase methoxyphenyl)-2,3-dihydro-4H-1-benzopyran-4-one.
(EC 4.2.2.7).
Hygroscopic powder, slightly soluble in water and in methanol.
Enzyme from Flavobacterium heparinum that performs
eliminative cleavage of polysaccharides containing mp : 258 °C to 262 °C.
(1→4)-linked D-glucuronate or L-iduronate residues and Hexachlorobenzene. C6Cl6. (Mr 284.8). 1128200. [118-74-1].
(1→4)-α-linked 2-sulfoamino-2-deoxy-6-sulfo-D-glucose
residues to give oligosaccharides with terminal bp : about 332 °C.
4-deoxy-α-D-gluc-4-enuronosyl groups at their non-reducing mp : about 230 °C.
ends. A suitable certified reference solution (10 ng/µL in
Heparinase II. 1187700. [149371-12-0].
Enzyme from Flavobacterium heparinum that depolymerises α-Hexachlorocyclohexane. C6H6Cl6. (Mr 290.8). 1128300.
sulfated polysaccharide chains containing 1→4 linkages [319-84-6].
between hexosamines and uronic acid residues (both bp : about 288 °C.
iduronic and glucuronic acid residues). The reaction yields mp : about 158 °C.
oligosaccharide products (mainly disaccharides) containing
unsaturated uronic acids. A suitable certified reference solution (10 ng/µL in
Heparinase III. 1187800. [37290-86-1]. Heparin-sulfate lyase
(EC 4.2.2.8). β-Hexachlorocyclohexane. C6H6Cl6. (Mr 290.8). 1128400.
[319-85-7].
Enzyme from Flavobacterium heparinum that depolymerises
selectively sulfated polysaccharide chains containing 1→4 A suitable certified reference solution (10 ng/µL in
linkages between hexosamines and glucuronic acid residues cyclohexane) may be used.
to give oligosaccharide products (mainly disaccharides) δ-Hexachlorocyclohexane. C6H6Cl6. (Mr 290.8). 1128500.
containing unsaturated uronic acids. [319-86-8].
HEPES. C8H18N2O4S. (Mr 238.3). 1106800. [7365-45-9]. A suitable certified reference solution (10 ng/µL in
2-[4-(2-Hydroxyethyl)piperazin-1-yl]ethanesulfonic acid. cyclohexane) may be used.
White or almost white powder. Hexacosane. C26H54. (Mr 366.7). 1042200. [630-01-3].
mp : about 236 °C, with decomposition Colourless or white or almost white flakes.
Heptachlor. C10H5Cl7. (Mr 373.3). 1128000. [76-44-8]. mp : about 57 °C.
bp : about 135 °C. Hexadimethrine bromide. (C13H30Br2N2)n. 1042300.
mp : about 95 °C. [28728-55-4]. 1,5-Dimethyl-1,5-diazaundecamethylene
A suitable certified reference solution (10 ng/µL in polymethobromide. Poly(1,1,5,5-tetramethyl-1,5-azonia-
cyclohexane) may be used. undecamethylene dibromide).
White or almost white, amorphous powder, hygroscopic,
Heptachlor epoxide. C10H5Cl7O. (Mr 389.3). 1128100. soluble in water.
[1024-57-3].
bp : about 200 °C.
mp : about 160 °C. 2,2′,2″,6,6′,6″-Hexa(1,1-dimethylethyl)-4,4′,4″-[(2,4,6-
A suitable certified reference solution (10 ng/µL in trimethyl-1,3,5-benzenetriyl)trismethylene]triphenol.
cyclohexane) may be used. C54H78O3. (Mr 775). 1042100. 2,2′,2″,6,6′,6″-
Hexa-tert-butyl-4,4′,4″-[(2,4,6-trimethyl-1,3,5-
Heptafluorobutyric acid. C4HF7O2. (Mr 214.0). 1162400. benzenetriyl)trismethylene]triphenol.
[375-22-4]. HFBA. Crystalline powder, practically insoluble in water, soluble in
Clear, colourless liquid. Corrosive. acetone, slightly soluble in ethanol (96 per cent).
: about 1.645. mp : about 244 °C.

1,1,1,3,3,3-Hexafluoropropan-2-ol. C3H2F6O. (Mr 168.0). Holmium oxide. Ho2O3. (Mr 377.9). 1043100. [12055-62-8].

1136000. [920-66-1]. Diholmium trioxide.
Content : minimum 99.0 per cent, determined by gas Yellowish powder, practically insoluble in water.
chromatography.
Holmium perchlorate solution. 1043101.
Clear, colourless liquid, miscible with water and with
anhydrous ethanol. A 40 g/L solution of holmium oxide R in a solution of
perchloric acid R containing 141 g/L of HClO4.
: about 1.596.
bp : about 59 °C. DL-Homocysteine. C4H9NO2S. (Mr 135.2). 1136100.
[454-29-5]. (2RS)-2-Amino-4-sulfanylbutanoic acid.
Hexamethyldisilazane. C6H19NSi2. (Mr 161.4). 1042400. White or almost white, crystalline powder.
[999-97-3].
mp : about 232 °C.
: about 0.78. L-Homocysteine thiolactone hydrochloride.
C4H8ClNOS. (Mr 153.6). 1136200. [31828-68-9].
: about 1.408. (3S)-3-Aminodihydrothiophen-2(3H)-one hydrochloride.
bp : about 125 °C. White or almost white, crystalline powder.
Storage : in an airtight container. mp : about 202 °C.
Hexamethylenetetramine. C6H12N4. (Mr 140.2). 1042500. Homoorientin. C21H20O11. (Mr 448.4). 1189500.
[100-97-0]. Hexamine. 1,3,5,7-Tetraazatricyclo[3.3.1.13,7]- [4261-42-1]. 2-(3,4-Dihydroxyphenyl)-6-β-D-glucopyranosyl-
decane. 5,7-dihydroxy-4H-1-benzopyran-4-one. Isoorientin.
Colourless, crystalline powder, very soluble in water. Luteolin-6-C-glucoside.
Hexane. C6H14. (Mr 86.2). 1042600. [110-54-3]. Honokiol. C18H18O2. (Mr 266.3). 1182700. [35354-74-6].
Colourless, flammable liquid, practically insoluble in water, 3′,5-Di(prop-2-enyl)biphenyl-2,4′-diol. 3′,5-Diallyl-2,4′-
miscible with anhydrous ethanol. dihydroxybiphenyl. 3′,5-Di-2-propenyl-[1,1′-biphenyl]-2,4′-
diol.
: 0.659 to 0.663.
: 1.375 to 1.376. Human tissue factor solution. 1186100.
Distillation range (2.2.11). Not less than 95 per cent distils Solution containing human tissue factor, which may be
between 67 °C and 69 °C. produced by recombinant DNA technology, combined with
Hexane used in spectrophotometry complies with the following phospholipids and calcium buffers. Suitable stabilisers may
additional test. be added.
Absorbance (2.2.25) : maximum 0.01 from 260 nm to 420 nm, Hyaluronidase diluent. 1043300.
determined using water R as compensation liquid. Mix 100 mL of phosphate buffer solution pH 6.4 R with 100 mL
of water R. Dissolve 0.140 g of hydrolysed gelatin R in the
Hexylamine. C6H15N. (Mr 101.2). 1042700. [111-26-2].
solution at 37 °C.
Hexan-1-amine.
Storage : use within 2 h.
Colourless liquid, slightly soluble in water, soluble in ethanol
(96 per cent). Hydrastine hydrochloride. C21H22ClNO6. (Mr 419.9).
: about 0.766. 1154000. [5936-28-7]. (3S)-6,7-Dimethoxy-3-[(5R)-6-
: about 1.418. methyl-5,6,7,8-tetrahydro-1,3-dioxolo[4,5-g]isoquinolin-5-
yl]isobenzofuran-1(3H)-one hydrochloride.
bp : 127 °C to 131 °C.
White or almost white powder, hygroscopic, very soluble in
Hibifolin. C21H18O14. (Mr 494.4). 1207000. [55366-56-8]. water and in ethanol (96 per cent).
2-(3,4-Dihydroxyphenyl)-3,5,7-trihydroxy-4-oxo-4H-1- [α]17 : about + 127.
benzopyran-8-yl β-D-glucopyranosiduronic acid. Gossypetin D
8-O-glucuronide. Gossypetin 8-O-β-D-glucuropyranoside. mp : about 116 °C.
Storage : protected from light, at a temperature not exceeding Hydrastine hydrochloride used in liquid chromatography
8 °C, in a dry place. complies with the following additional test.
Histamine dihydrochloride. 1042800. [56-92-8]. monograph Goldenseal rhizome (1831).
See Histamine dihydrochloride (0143). Content : minimum 98 per cent, calculated by the
Histamine solution. 1042901.
A 9 g/L solution of sodium chloride R containing 0.1 µg Hydrazine. H4N2. (Mr 32.05). 1136300. [302-01-2]. Diazane.
per millilitre of histamine base (as the phosphate or Slightly oily liquid, colourless, with a strong odour of
dihydrochloride). ammonia, miscible with water. Dilute solutions in water are
commercially available.
Histidine. 1187900. [71-00-1]. (2S)-2-Amino-3-(1H-
imidazol-4-yl)propanoic acid. : about 1.470.
bp : about 113 °C.
Histidine monohydrochloride. C6H10ClN3O2,H2O. mp : about 1.5 °C.
(Mr 209.6). 1043000. [123333-71-1]. (RS)-2-Amino-3-
(imidazol-4-yl)propionic acid hydrochloride monohydrate. Caution : toxic and corrosive.
Crystalline powder or colourless crystals, soluble in water. Hydrazine sulfate. H6N2O4S. (Mr 130.1). 1043400.
mp : about 250 °C, with decomposition. [10034-93-2].
Chromatography. Thin-layer chromatography Colourless crystals, sparingly soluble in cold water, soluble
(2.2.27) as prescribed in the monograph Histamine in hot water (50 °C) and freely soluble in boiling water,
dihydrochloride (0143) ; the chromatogram shows only one practically insoluble in ethanol (96 per cent).
principal spot. Content : minimum 99 per cent.

Hydriodic acid. HI. (Mr 127.9). 1098900. [10034-85-2]. Pb : 0.001 ppm.

Prepare by distilling hydriodic acid over red phosphorus, Zn : 0.005 ppm.
passing carbon dioxide R or nitrogen R through the apparatus
during the distillation. Use the colourless or almost colourless, Hydrochloric acid, dilute R1. 1043504.
constant-boiling mixture (55 per cent to 58 per cent of HI) Contains 0.37 g/L of HCl.
distilling between 126 °C and 127 °C. Dilute 1.0 mL of dilute hydrochloric acid R to 200.0 mL
Place the acid in small, amber, glass-stoppered bottles with water R.
previously flushed with carbon dioxide R or nitrogen R, seal Hydrochloric acid, dilute R2. 1043505.
with paraffin.
Dilute 30 mL of 1 M hydrochloric acid to 1000 mL with
Storage : in a dark place. water R ; adjust to pH 1.6 ± 0.1.
Hydrobromic acid, 30 per cent. 1098700. [10035-10-6]. Hydrochloric acid, dilute R3. 1203800.
A 30 per cent solution of hydrobromic acid in glacial acetic Contains 3.7 g/L of HCl.
acid R.
Dilute 10.0 mL of dilute hydrochloric acid R to 200.0 mL
Degas with caution the contents before opening. with water R.
Hydrobromic acid, dilute. 1098701. Hydrochloric acid, ethanolic. 1043506.
Place 5.0 mL of 30 per cent hydrobromic acid R in amber Dilute 5.0 mL of 1 M hydrochloric acid to 500.0 mL with
vials equipped with polyethylene stoppers. Seal under ethanol (96 per cent) R.
argon R and store in the dark. Add 5.0 mL of glacial acetic
acid R immediately before use. Shake. Hydrochloric acid, heavy metal-free. 1043510.
Storage : in the dark. Complies with the requirements prescribed for hydrochloric
acid R with the following maximum contents of heavy
Hydrobromic acid, 47 per cent. 1118900. metals.
A 47 per cent m/m solution of hydrobromic acid. As : 0.005 ppm.
Hydrobromic acid, dilute R1. 1118901. Cd : 0.003 ppm.
Contains 7,9 g/L of HBr. Cu : 0.003 ppm.
Dissolve 16.81 g of 47 per cent hydrobromic acid R in Fe : 0.05 ppm.
water R and dilute to 1000 mL with the same solvent. Hg : 0.005 ppm.
Hydrochloric acid. 1043500. [7647-01-0]. Ni : 0.004 ppm.
See Concentrated hydrochloric acid (0002). Pb : 0.001 ppm.
Zn : 0.005 ppm.
0.1 M Hydrochloric acid, alcoholic. 3008800.
Dilute 9.0 mL of hydrochloric acid R to 1000.0 mL with Hydrochloric acid, lead-free. 1043508.
aldehyde-free alcohol R. Complies with the requirements prescribed for hydrochloric
acid R with the following additional requirement.
2 M Hydrochloric acid. 3001700.
Lead : maximum 20 ppb.
Dilute 206.0 g of hydrochloric acid R to 1000.0 mL with
water R. Atomic emission spectrometry (2.2.22, Method I).
Test solution. In a quartz crucible evaporate 200 g of the
3 M Hydrochloric acid. 3001600. acid to be examined almost to dryness. Take up the residue
Dilute 309.0 g of hydrochloric acid R to 1000.0 mL with in 5 mL of nitric acid prepared by sub-boiling distillation of
water R. nitric acid R and evaporate to dryness. Take up the residue
in 5 mL of nitric acid prepared by sub-boiling distillation
6 M Hydrochloric acid. 3001500. of nitric acid R.
Dilute 618.0 g of hydrochloric acid R to 1000.0 mL with Reference solutions. Prepare the reference solutions using
water R. lead standard solution (0.1 ppm Pb) R diluted with nitric
Hydrochloric acid R1. 1043501. acid prepared by sub-boiling distillation of nitric acid R.
Contains 250 g/L of HCl. Wavelength : 220.35 nm.
Dilute 70 g of hydrochloric acid R to 100 mL with water R. Hydrochloric acid, methanolic. 1043511.
Hydrochloric acid, brominated. 1043507. Dilute 4.0 mL of hydrochloric acid R to 1000.0 mL with
methanol R2.
To 1 mL of bromine solution R add 100 mL of hydrochloric
acid R. Hydrocortisone acetate. 1098800. [50-03-3].
Hydrochloric acid, dilute. 1043503. See Hydrocortisone acetate (0334).
Contains 73 g/L of HCl. Hydrofluoric acid. HF. (Mr 20.01). 1043600. [7664-39-3].
Dilute 20 g of hydrochloric acid R to 100 mL with water R. Content : minimum 40.0 per cent m/m.
Hydrochloric acid, dilute, heavy metal-free. 1043509. Clear, colourless liquid.
Complies with the requirements prescribed for dilute Loss on ignition : not more than 0.05 per cent m/m ; evaporate
hydrochloric acid R with the following maximum contents the hydrofluoric acid in a platinum crucible and gently ignite
of heavy metals. the residue to constant mass.
As : 0.005 ppm. Assay. Weigh accurately a glass-stoppered flask containing
50.0 mL of 1 M sodium hydroxide. Introduce 2 g of the
Cd : 0.003 ppm. hydrofluoric acid and weigh again. Titrate the solution with
Cu : 0.003 ppm. 0.5 M sulfuric acid, using 0.5 mL of phenolphthalein solution R
Fe : 0.05 ppm. as indicator.
Hg : 0.005 ppm. 1 mL of 1 M sodium hydroxide is equivalent to 20.01 mg of HF.
Ni : 0.004 ppm. Storage : in a polyethylene container.

Hydrogen for chromatography. H2. (Mr 2.016). 1043700. Hydroxylamine solution, alcoholic. 1044301.
[1333-74-0]. Dissolve 3.5 g of hydroxylamine hydrochloride R in 95 mL
Content : minimum 99.95 per cent V/V. of ethanol (60 per cent V/V) R, add 0.5 mL of a 2 g/L
solution of methyl orange R in ethanol (60 per cent V/V) R
Hydrogen peroxide solution, dilute. 1043800. [7722-84-1]. and sufficient 0.5 M potassium hydroxide in alcohol (60 per
See Hydrogen peroxide solution (3 per cent) (0395). cent V/V) to give a pure yellow colour. Dilute to 100 mL
Hydrogen peroxide solution, strong. 1043900. [7722-84-1]. with ethanol (60 per cent V/V) R.
See Hydrogen peroxide solution (30 per cent) (0396). Hydroxylamine solution, alkaline. 1044302.
Hydrogen sulfide. H2S. (Mr 34.08). 1044000. [7783-06-4]. Immediately before use, mix equal volumes of a 139 g/L
Gas, slightly soluble in water. solution of hydroxylamine hydrochloride R and a 150 g/L
solution of sodium hydroxide R.
Hydrogen sulfide solution. 1136400.
Hydroxylamine solution, alkaline R1. 1044303.
A recently prepared solution of hydrogen sulfide R in
water R. The saturated solution contains about 0.4 per cent Solution A. Dissolve 12.5 g of hydroxylamine
to 0.5 per cent of H2S at 20 °C. hydrochloride R in methanol R and dilute to 100 mL with
the same solvent.
Hydrogen sulfide R1. H2S. (Mr 34.08). 1106600. [7783-06-4]. Solution B. Dissolve 12.5 g of sodium hydroxide R in
Content : minimum 99.7 per cent V/V. methanol R and dilute to 100 mL with the same solvent.
Hydroquinone. C6H6O2. (Mr 110.1). 1044100. [123-31-9]. Mix equal volumes of solution A and solution B
Benzene-1,4-diol. immediately before use.
Fine, colourless or white or almost white needles, darkening Hydroxymethylfurfural. C6H6O3. (Mr 126.1). 1044400.
on exposure to air and light, soluble in water and in ethanol [67-47-0]. 5-Hydroxymethylfurfural.
(96 per cent).
Acicular crystals, freely soluble in water, in acetone and in
mp : about 173 °C. ethanol (96 per cent).
Storage : protected from light and air. mp : about 32 °C.
Hydroquinone solution. 1044101.
Hydroxynaphthol blue, sodium salt. C20H11N2Na3O11S3.
Dissolve 0.5 g of hydroquinone R in water R, add 20 µL of (Mr 620). 1044500. [63451-35-4]. Trisodium
sulfuric acid R and dilute to 50 mL with water R. 2,2′-dihydroxy-1,1′-azonaphthalene-3′,4,6′-trisulfonate.
4′-Hydroxyacetophenone. C8H8O2. (Mr 136.2). 1196900. 2-Hydroxypropylbetadex for chromatography. 1146000.
[99-93-4]. 1-(4-Hydroxyphenyl)ethan-1-one. Betacyclodextrin modified by the bonding of (R) or (RS)
2-Hydroxybenzimidazole. C7H6N2O. (Mr 134.1). 1169600. propylene oxide groups on the hydroxyl groups.
[615-16-7]. 1H-benzimidazol-2-ol.
Hydroxypropyl-β-cyclodextrin. 1128600. [94035-02-6].
4-Hydroxybenzohydrazide. C7H8N2O2. (Mr 152.2). 1145900. See Hydroxypropylbetadex (1804).
[5351-23-5]. p-Hydroxybenzohydrazide. pH (2.2.3) : 5.0 to 7.5 for a 20 g/L solution.
4-Hydroxybenzoic acid. C7H6O3. (Mr 138.1). 1106700. Hydroxyquinoline. C9H7NO. (Mr 145.2). 1044600.
[99-96-7]. [148-24-3]. 8-Hydroxyquinoline. Quinolin-8-ol.
Crystals, slightly soluble in water, very soluble in ethanol
White or slightly yellowish, crystalline powder, slightly soluble
(96 per cent), soluble in acetone.
in water, freely soluble in acetone, in ethanol (96 per cent)
mp : 214 °C to 215 °C. and in dilute mineral acids.
4-Hydroxycoumarin. C9H6O3. (Mr 162.2). 1169700. mp : about 75 °C.
[1076-38-6]. 4-Hydroxy-2H-1-benzopyran-2-one. Sulfated ash (2.4.14): maximum 0.05 per cent.
White or almost white powder, freely soluble in methanol.
12-Hydroxystearic acid. C18H36O3. (Mr 300.5). 1099000.
[106-14-9]. 12-Hydroxyoctadecanoic acid.
6-Hydroxydopa. C9H11NO5. (Mr 213.2). 1169800. White or almost white powder.
[21373-30-8]. (2RS)-2-Amino-3-(2,4,5-trihydroxyphenyl)- mp : 71 °C to 74 °C.
propanoic acid. 2,5-Dihydroxy-DL-tyrosine.
mp : about 257 °C. 5-Hydroxyuracil. C4H4N2O3. (Mr 128.1). 1044700.
[496-76-4]. Isobarbituric acid. Pyrimidine-2,4,5-triol.
4-Hydroxyisophthalic acid. C8H6O5. (Mr 182.1). 1106900.
[636-46-4]. 4-Hydroxybenzene-1,3-dicarboxylic acid.
Needles or platelets, very slightly soluble in water, freely mp : about 310 °C, with decomposition.
soluble in ethanol (96 per cent). Chromatography. Thin-layer chromatography (2.2.27)
mp : about 314 °C, with decomposition. as prescribed in the monograph Fluorouracil (0611) ; the
chromatogram shows a principal spot with an RF of about 0.3.
Hydroxylamine hydrochloride. NH4ClO. (Mr 69.5). Storage : in an airtight container.
1044300. [5470-11-1].
White or almost white, crystalline powder, very soluble in Hyoscine hydrobromide. 1044800. [6533-68-2].
water, soluble in ethanol (96 per cent). See Hyoscine hydrobromide (0106).
Hydroxylamine hydrochloride solution R2. 1044304. Hyoscyamine sulfate. 1044900. [620-61-1].
Dissolve 2.5 g of hydroxylamine hydrochloride R in 4.5 mL See Hyoscyamine sulfate (0501).
of hot water R and add 40 mL of ethanol (96 per cent) R
and 0.4 mL of bromophenol blue solution R2. Add 0.5 M Hypericin. C30H16O8. (Mr 504.4). 1149800. [548-04-9].
alcoholic potassium hydroxide until a greenish-yellow 1,3,4,6,8,13-Hexahydroxy-10,11-dimethylphenan-
colour is obtained. Dilute to 50.0 mL with ethanol (96 per thro[1,10,9,8-opqra]perylene-7,14-dione.
cent) R. Content : minimum 85 per cent.

Hyperoside. C21H20O12. (Mr 464.4). 1045000. The solution complies with the following test : add 10 mL
2-(3,4-Dihydroxyphenyl)-3-β-D-galactopyranosyloxy- to a solution of 1.0 mg of potassium nitrate R in 10 mL of
5,7-dihydroxychromen-4-one. water R, rapidly add 20 mL of nitrogen-free sulfuric acid R
Faint yellow needles, soluble in methanol. and heat to boiling. The blue colour is discharged within
1 min.
Absorbance (2.2.25). A solution in methanol R shows
2 absorption maxima at about 257 nm and at about 359 nm. Indigo carmine solution R1. 1045602.
Hypophosphorous reagent. 1045200. Dissolve 4 g of indigo carmine R in about 900 mL of water R
Dissolve with the aid of gentle heat, 10 g of sodium added in several portions. Add 2 mL of sulfuric acid R and
hypophosphite R in 20 mL of water R and dilute to 100 mL dilute to 1000 mL with water R.
with hydrochloric acid R. Allow to settle and decant or filter Assay. Place in a 100 mL conical flask with a wide
through glass wool. neck 10.0 mL of nitrate standard solution (100 ppm
NO3) R, 10 mL of water R, 0.05 mL of the indigo carmine
Hypoxanthine. C5H4N4O. (Mr 136.1). 1045300. [68-94-0]. solution R1, and then in a single addition, but with caution,
1H-Purin-6-one. 30 mL of sulfuric acid R. Titrate the solution immediately,
White or almost white, crystalline powder, very slightly soluble using the indigo carmine solution R1, until a stable blue
in water, sparingly soluble in boiling water, soluble in dilute colour is obtained.
acids and in dilute alkali hydroxide solutions, decomposes The number of millilitres used, n, is equivalent to 1 mg of
without melting at about 150 °C. NO3.
prescribed in the monograph Mercaptopurine (0096) ; the Indirubin. C16H10N2O2. (Mr 262.3). 1192900. [479-41-4].
chromatogram shows only one principal spot. 1,1′,2′,3-Tetrahydro-2,3′-bi(indolylidene)-2′,3-dione.
Ibuprofen. 1197000. [15687-27-1]. Indometacin. 1101500. [53-86-1].
See Ibuprofen (0721). See Indometacin (0092).
Imidazole. C3H4N2. (Mr 68.1). 1045400. [288-32-4]. Inosine. C10H12N4O5. (Mr 268.2). 1169900. [58-63-9].
White or almost white, crystalline powder, soluble in water 9-β-D-Ribofuranosylhypoxanthine. 9-β-D-Ribofuranosyl-1,9-
and in ethanol (96 per cent). dihydro-6H-purin-6-one.
mp : about 90 °C. mp : 222 °C to 226 °C.
Iminodiacetic acid. C4H7NO4. (Mr 133.1). 1192300. myo-Inositol. 1161100.
[142-73-4]. 2,2′-Iminodiacetic acid. See myo-Inositol (1805).
Iminodibenzyl. C14H13N. (Mr 195.3). 1045500. [494-19-9].
10,11-Dihydrodibenz[b,f]azepine. Iodine. 1045800. [7553-56-2].
Pale yellow, crystalline powder, practically insoluble in water, See Iodine (0031).
freely soluble in acetone. Iodine solution R1. 1045801.
mp : about 106 °C.
To 10.0 mL of 0.05 M iodine add 0.6 g of potassium iodide R
Imipramine hydrochloride. 1207100. [113-52-0]. and dilute to 100.0 mL with water R. Prepare immediately
before use.
See Imipramine hydrochloride (0029).
Iodine solution R2. 1045802.
Imperatorin. C16H14O4. (Mr 270.3). 1180200. [482-44-0].
9-[(3-Methylbut-2-enyl)oxy]-7H-furo[3,2-g][1]benzopyran- To 10.0 mL of 0.05 M iodine add 0.6 g of potassium iodide R
7-one. and dilute to 1000.0 mL with water R. Prepare immediately
before use.
2-Indanamine hydrochloride. C9H12ClN. (Mr 169.7).
1175800. [2338-18-3]. 2-Aminoindane hydrochloride. Iodine solution R3. 1045803.
2,3-Dihydro-1H-inden-2-amine hydrochloride. Dilute 2.0 mL of iodine solution R1 to 100.0 mL with
water R. Prepare immediately before use.
Indigo. C16H10N2O2. (Mr 262.3). 1192800. [482-89-3].
Indigotin. 1,1′,3,3′-Tetrahydro-2-2′-bi(indolylidene)-3,3′- Iodine solution R4. 1045806.
dione. Dissolve 14 g of iodine R in 100 mL of a 400 g/L solution of
Indigo carmine. C16H8N2Na2O8S2. (Mr 466.3). 1045600. potassium iodide R, add 1 mL of dilute hydrochloric acid R
[860-22-0]. and dilute to 1000 mL with water R.
Schultz No. 1309. Storage : protected from light.
Colour Index No. 73015. Iodine solution R5. 1045807.
3,3′-Dioxo-2,2′-bisindolylidene-5,5′-disulfonate disodium. E
132. Dissolve 12.7 g of iodine R and 20 g of potassium iodide R
in water R and dilute to 1000.0 mL with the same solvent
It usually contains sodium chloride. (0.05 M solution).
Blue or violet-blue powder or blue granules with a coppery
lustre, sparingly soluble in water, practically insoluble in Iodine solution, alcoholic. 1045804.
ethanol (96 per cent). It is precipitated from an aqueous A 10 g/L solution of iodine R in ethanol (96 per cent) R.
solution by sodium chloride.
Indigo carmine solution. 1045601.
Iodine solution, chloroformic. 1045805.
To a mixture of 10 mL of hydrochloric acid R and 990 mL
of 200 g/L nitrogen-free sulfuric acid R add 0.2 g of indigo A 5 g/L solution of iodine R in chloroform R.
carmine R. Storage : protected from light.

Iodine-123 and ruthenium-106 spiking solution. 1166700. 3-Iodobenzylammonium chloride. C7H9ClIN. (Mr 269.5).
Prepare immediately before use. Mix 3.5 mL of an 18.5 kBq/mL 1168000. [3718-88-5]. 1-(3-Iodophenyl)methanamine
solution of ruthenium-106 in the form of ruthenium hydrochloride. 1-(3-Iodophenyl)methanaminium chloride.
trichloride in a mixture of equal volumes of glacial acetic m-Iodobenzylamine hydrochloride.
acid R and water R with 200 µL of a 75 kBq/mL solution of White or almost white crystals.
iodine-123 in the form of sodium iodide in water R. mp : 188 °C to 190 °C.
Iodine bromide. IBr. (Mr 206.8). 1045900. [7789-33-5]. Iodoethane. C2H5I. (Mr 156.0). 1099100. [75-03-6].
Bluish-black or brownish-black crystals, freely soluble in Content : minimum 99 per cent.
water, in ethanol (96 per cent) and in glacial acetic acid. Colourless or slightly yellowish liquid, darkening on exposure
bp : about 116 °C. to air and light, miscible with ethanol (96 per cent) and most
mp : about 40 °C. organic solvents.
Storage : protected from light. : about 1.95.
: about 1.513.
Iodine bromide solution. 1045901.
bp : about 72 °C.
Dissolve 20 g of iodine bromide R in glacial acetic acid R
and dilute to 1000 mL with the same solvent. Storage : in an airtight container, protected from light.
Storage : protected from light. 2-Iodohippuric acid. C9H8INO3,2H2O. (Mr 341.1). 1046200.
[147-58-0]. 2-(2-Iodobenzamido)acetic acid.
Iodine chloride. ICl. (Mr 162.4). 1143000. [7790-99-0].
White or almost white, crystalline powder, sparingly soluble
Black crystals, soluble in water, in acetic acid and in ethanol in water.
(96 per cent).
mp : about 170 °C.
bp : about 97.4 °C.
Water (2.5.12) : 9 per cent to 13 per cent, determined on
Iodine chloride solution. 1143001. 1.000 g.
Dissolve 1.4 g of iodine chloride R in glacial acetic acid R Chromatography. Thin-layer chromatography (2.2.27), using
and dilute to 100 mL with the same acid. cellulose for chromatography F254 R as the coating substance :
Storage : protected from light. apply to the plate 20 μL of a solution of the 2-iodohippuric
acid, prepared by dissolving 40 mg in 4 mL of 0.1 M sodium
Iodine pentoxide, recrystallised. I2O5. (Mr 333.8). 1046000. hydroxide and diluting to 10 mL with water R. Develop over
[12029-98-0]. Di-iodine pentoxide. Iodic anhydride. a path of about 12 cm using as the mobile phase the upper
Content : minimum 99.5 per cent. layer obtained by shaking together 20 volumes of water R,
White or almost white, crystalline powder, or white or 40 volumes of glacial acetic acid R and 40 volumes of toluene R.
greyish-white granules, hygroscopic, very soluble in water Allow the plate to dry in air and examine in ultraviolet light at
forming HIO3. 254 nm. The chromatogram shows only one principal spot.
Stability on heating. Dissolve 2 g, previously heated for 1 h at Iodoplatinate reagent. 1046300.
200 °C, in 50 mL of water R. A colourless solution is obtained. To 3 mL of a 100 g/L solution of chloroplatinic acid R add
Assay. Dissolve 0.100 g in 50 mL of water R, add 3 g of 97 mL of water R and 100 mL of a 60 g/L solution of potassium
potassium iodide R and 10 mL of dilute hydrochloric acid R. iodide R.
Titrate the liberated iodine with 0.1 M sodium thiosulfate, Storage : protected from light.
using 1 mL of starch solution R as indicator.
1 mL of 0.1 M sodium thiosulfate is equivalent to 2.782 mg Iodoplatinate reagent R1. 1172200.
of I2O5. Mix 2.5 mL of a 50 g/L solution of chloroplatinic acid R,
Storage : in an airtight container, protected from light. 22.5 mL of a 100 g/L solution of potassium iodide R and 50 mL
of water R.
Iodoacetamide. C2H4INO. (Mr 185.0). 1186200. [144-48-9]. Storage : protected from light, at a temperature of 2-8 °C.
2-Iodoacetamide.
Slightly yellow, crystalline powder, soluble in water. Iodosulfurous reagent. 1046400.
mp : about 92 °C. The apparatus, which must be kept closed and dry
during the preparation, consists of a 3000 mL to 4000 mL
Iodoacetic acid. C2H3IO2. (Mr 185.9). 1107000. [64-69-7]. round-bottomed flask with three inlets for a stirrer and
Colourless or white or almost white crystals, soluble in water a thermometer and fitted with a drying tube. To 700 mL
and in ethanol (96 per cent). of anhydrous pyridine R and 700 mL of ethylene glycol
monomethyl ether R add, with constant stirring, 220 g of
mp : 82 °C to 83 °C. finely powdered iodine R, previously dried over diphosphorus
2-Iodobenzoic acid. C7H5IO2. (Mr 248.0). 1046100. pentoxide R. Continue stirring until the iodine has completely
[88-67-5]. dissolved (about 30 min). Cool to − 10 °C, and add quickly,
White or slightly yellow, crystalline powder, slightly soluble in still stirring, 190 g of sulfur dioxide R. Do not allow the
water, soluble in ethanol (96 per cent). temperature to exceed 30 °C. Cool.
mp : about 160 °C. Assay. Add about 20 mL of anhydrous methanol R to a titration
vessel and titrate to the end-point with the iodosulfurous
Chromatography. Thin-layer chromatography (2.2.27), using reagent (2.5.12). Introduce in an appropriate form a suitable
cellulose for chromatography f254 R as the coating substance : amount of water R, accurately weighed, and repeat the
apply to the plate 20 μL of a solution of the 2-iodobenzoic determination of water. Calculate the water equivalent in
acid, prepared by dissolving 40 mg in 4 mL of 0.1 M sodium milligrams per millilitre of iodosulfurous reagent.
hydroxide and diluting to 10 mL with water R. Develop over
a path of about 12 cm using as the mobile phase the upper The minimum water equivalent is 3.5 mg of water per
layer obtained by shaking together 20 volumes of water R, millilitre of reagent.
40 volumes of glacial acetic acid R and 40 volumes of toluene R. Work protected from humidity. Standardise immediately
Allow the plate to dry in air and examine in ultraviolet light at before use.
254 nm. The chromatogram shows only one principal spot. Storage : in a dry container.

5-Iodouracil. C4H3IN2O2. (Mr 238.0). 1046500. [696-07-1]. Isoamyl benzoate. C12H16O2. (Mr 192.3). 1164200. [94-46-2].
5-Iodo-1H,3H-pyrimidine-2,4-dione. Isopentyl benzoate. 3-Methylbutyl benzoate.
mp : about 276 °C, with decomposition. : about 1.494.
Chromatography. Thin-layer chromatography (2.2.27) as bp : about 261 °C.
prescribed in the monograph Idoxuridine (0669) : apply 5 µL Colourless or pale yellow liquid.
of a 0.25 g/L solution ; the chromatogram obtained shows only
one principal spot. Isoandrosterone. C19H30O2. (Mr 290.4). 1107100. [481-29-8].
Epiandrosterone. 3β-Hydroxy-5α-androstan-17-one.
Ion-exclusion resin for chromatography. 1131000. White or almost white powder, practically insoluble in water,
A resin with sulfonic acid groups attached to a polymer lattice soluble in organic solvents.
consisting of polystyrene cross-linked with divinylbenzene. : + 88, determined on 20 g/L solution in methanol R.
Ion-exchange resin, strongly acidic. 1085400. mp : 172 °C to 174 °C.
Resin in protonated form with sulfonic acid groups attached to ∆A (2.2.41) : 14.24 × 10 , determined at 304 nm on a 1.25 g/L
3
a lattice consisting of polystyrene cross-linked with 8 per cent solution.

of divinylbenzene. It is available as spherical beads ; unless N-Isobutyldodecatetraenamide. C16H25NO. (Mr 247.4).
otherwise prescribed, the particle size is 0.3 mm to 1.2 mm. 1159500. [866602-52-0]. (2E,4E,8Z,10EZ)-N-2-
Capacity. 4.5 mmol to 5 mmol per gram, with a water content (Methylpropyl)dodeca-2,4,8,10-tetraenamide.
of 50 per cent to 60 per cent. White or almost white or non-coloured crystals.
Preparation of a column. Unless otherwise prescribed, use a mp : about 70 °C.
tube with a fused-in sintered glass disc having a length of
400 mm, an internal diameter of 20 mm and a filling height N-Isobutyldodecatetraenamide solution. 1159501.
of about 200 mm. Introduce the resin, mixing it with water R A solution of N-isobutyldodecatetraenamide R, exactly
and pouring the slurry into the tube, ensuring that no air weighed, in methanol R at a concentration of about
bubbles are trapped between the particles. When in use, the 10 mg/mL.
liquid must not be allowed to fall below the surface of the
resin. If the resin is in its protonated form, wash with water R Isodrin. C12H8Cl6. (Mr 364.9). 1128700. [465-73-6].
until 50 mL requires not more than 0.05 mL of 0.1 M sodium 1,2,3,4,10,10-Hexachloro-1,4,4a,5,8,8a-hexahydro-endo,endo-
hydroxide for neutralisation, using 0.1 mL of methyl orange 1,4:5,8-dimethanonaphthalene.
solution R as indicator. Practically insoluble in water, soluble in common organic
If the resin is in its sodium form or if it requires regeneration, solvents such as acetone.
pass about 100 mL of a mixture of equal volumes of A suitable certified reference solution may be used.
hydrochloric acid R1 and water R slowly through the column
and then wash with water R as described above. Isoeugenol. C10H12O2. (Mr 164.2). 1206200. [97-54-1].
2-Methoxy-4-[(1Ξ)-prop-1-en-1-yl]phenol.
Irisflorentin. C20H18O8. (Mr 386.4). 1186300. [41743-73-1].
9-Methoxy-7-(3,4,5-trimethoxyphenyl)-8H-1,3-dioxolo- Isoleucine. 1185000. [73-32-5].
[4,5-g][1]benzopyran-8-one. See Isoleucine (0770).
Iron. Fe. (Ar 55.85). 1046600. [7439-89-6]. Isomalt. C12H24O11. (Mr 344.3). 1164300. [64519-82-0].
Grey powder or wire, soluble in dilute mineral acids. Mixture of 6-O-α-D-glucopyranosyl-D-glucitol and of
1-O-α-D-glucopyranosyl-D-mannitol.
Iron salicylate solution. 1046700. White or almost white powder or granules, freely soluble in
Dissolve 0.1 g of ferric ammonium sulfate R in a mixture of water.
2 mL of dilute sulfuric acid R and 48 mL of water R and dilute
to 100 mL with water R. Add 50 mL of a 11.5 g/L solution of Isomaltitol. C12H24O11. (Mr 344.3). 1161200. [534-73-6].
sodium salicylate R, 10 mL of dilute acetic acid R, 80 mL of 6-O-α-D-Glucopyranosyl-D-glucitol.
a 136 g/L solution of sodium acetate R and dilute to 500 mL White or almost white powder, freely soluble in water.
with water R. The solution should be recently prepared.
Isomenthol. C10H20O. (Mr 156.3). 1047000. [23283-97-8].
Storage : in an airtight container, protected from light. (+)-Isomenthol : (1S,2R,5R)-2-isopropyl-5-methylcyclo-
hexanol. (±)-Isomenthol : a mixture of equal parts of
Isatin. C8H5NO2. (Mr 147.1). 1046800. [91-56-5].
Indoline-2,3-dione. (1S,2R,5R)- and (1R,2S,5S)-2-isopropyl-5-methylcyclohexanol.
Colourless crystals, practically insoluble in water, very soluble
Small, yellowish-red crystals, slightly soluble in water, soluble
in hot water and in ethanol (96 per cent), soluble in solutions
of alkali hydroxides giving a violet colour becoming yellow : (+)-Isomenthol : about + 24, determined on a 100 g/L
on standing. solution in ethanol (96 per cent) R.
mp : about 200 °C, with partial sublimation. bp : (+)-Isomenthol : about 218 °C. (±)-Isomenthol : about
218 °C.
Sulfated ash (2.4.14): maximum 0.2 per cent.
mp : (+)-Isomenthol : about 80 °C. (±)-Isomenthol : about 53 °C.
Isatin reagent. 1046801.
(+)-Isomenthone. C10H18O. (Mr 154.2). 1047100.
Dissolve 6 mg of ferric sulfate R in 8 mL of water R and add (1R)-cis-p-Menthan-3-one. (1R)-cis-2-Isopropyl-5-
cautiously 50 mL of sulfuric acid R. Add 6 mg of isatin R methylcyclohexanone.
and stir until dissolved.
Contains variable amounts of menthone. A colourless liquid,
The reagent should be pale yellow, but not orange or red. very slightly soluble in water, soluble in ethanol (96 per cent).
Isoamyl alcohol. C5H12O. (Mr 88.1). 1046900. [123-51-3]. : about 0.904.
3-Methylbutan-1-ol. : about 1.453.
Colourless liquid, slightly soluble in water, miscible with : about + 93.2.
ethanol (96 per cent). Isomenthone used in gas chromatography complies with the
bp : about 130 °C. following additional test.

Assay. Gas chromatography (2.2.28) as prescribed in the Content : minimum 99 per cent, calculated by the
monograph Peppermint oil (0405). normalisation procedure.
Test solution. The substance to be examined. Isoquercitrin. C21H20O12. (Mr 464.4). 1201600. [482-35-9].
Content : minimum 80.0 per cent, calculated by the 2-(3,4-Dihydroxyphenyl)-3-(β-D-glucopyranosyloxy)-5,7-
normalisation procedure. dihydroxy-4H-1-benzopyran-4-one.
Isomethyleugenol. C11H14O2. (Mr 178.2). 1181900. [93-16-3]. Isoquercitroside. C21H20O12. (Mr 464.4). 1136500.
1,2-Dimethoxy-4-prop-1-enylbenzene. [21637-25-2]. 2-(3,4-Dihydroxyphenyl)-3-(β-D-
Isomethyleugenol used in gas chromatography complies with glucofuranosyloxy)-5,7-dihydroxy-4H-1-benzopyran-4-one.
the following additional test. Isorhamnetin-3-O-neohesperidoside. C28H32O16. (Mr 625).
Assay. Gas chromatography (2.2.28) as prescribed in the 1205100. [55033-90-4]. 3-[6-Deoxy-α-L-mannopyranosyl-
monograph Niaouli oil, cineole type (2468). (1→2)-β-D-glucopyranosyloxy]-5,7-dihydroxy-2-(4-hydroxy-
Content : minimum 97.0 per cent, calculated by the 3-methoxyphenyl)-4H-1-benzopyran-4-one.
normalisation procedure. Isorhynchophylline. C22H28N2O4. (Mr 384.5). 1197100.
Isonicotinamide. C6H6N2O. (Mr 122.1). 1193000. [6859-01-4]. Methyl (16E)-17-methoxy-2-oxo-16,17-
[1453-82-3]. 4-Pyridinecarboxamide. Pyridine-4- didehydro-20α-corynoxan-16-carboxylate. Methyl (16E)-16-
carboxamide. (methoxymethylidene)-2-oxo-20α-corynoxan-17-oate.
White or almost white, crystalline powder, soluble in water. Isosilibinin. C25H22O10. (Mr 482.4). 1149900. [72581-71-6].
3,5,7-Trihydroxy-2-[2-(4-hydroxy-3-methoxyphenyl)-3-
Isonicotinic acid. C6H5NO2. (Mr 123.1). 1202200. [55-22-1]. hydroxymethyl-2,3-dihydro-1,4-benzodioxin-6-yl]chroman-
Pyridine-4-carboxylic acid. 4-one.
Creamish-white powder, sparingly soluble in water. White to yellowish powder, practically insoluble in water,
mp : about 311 °C. soluble in acetone and in methanol.
Isopropylamine. C3H9N. (Mr 59.1). 1119800. [75-31-0]. Kaempferol. C15H10O6. (Mr 286.2). 1197200. [520-18-3].
Propan-2-amine. 3,5,7-Trihydroxy-2-(4-hydroxyphenyl)-4H-1-benzopyran-4-
one.
Colourless, highly volatile, flammable liquid.
: about 1.374. Kaolin, light. 1047400. [1332-58-7].
bp : 32 °C to 34 °C. A purified native hydrated aluminium silicate. It contains a
suitable dispersing agent.
Isopropyl iodide. C3H7I. (Mr 170.0). 1166600. [75-30-9]. Light, white or almost white powder free from gritty particles,
2-Iodopropane. unctuous to the touch, practically insoluble in water and in
Content : minimum 99 per cent. mineral acids.
Coarse particles : maximum 0.5 per cent.
Isopropyl methanesulfonate. C4H10O3S. (Mr 138.2). 1179400.
[926-06-7]. 1-methylethyl methanesulfonate. Place 5.0 g in a ground-glass-stoppered cylinder about
160 mm long and 35 mm in diameter and add 60 mL of a
Clear, colourless liquid. 10 g/L solution of sodium pyrophosphate R. Shake vigorously
Content : minimum 99.0 per cent. and allow to stand for 5 min. Using a pipette, remove 50 mL
Density : about 1.129 g/cm3 (20 °C). of the liquid from a point about 5 cm below the surface. To
: 1.418-1.421. the remaining liquid add 50 mL of water R, shake, allow to
stand for 5 min and remove 50 mL as before. Repeat the
bp : about 82 °C at 6 mm Hg. operations until a total of 400 mL has been removed. Transfer
Isopropyl myristate. 1047200. [110-27-0]. the remaining suspension to an evaporating dish. Evaporate
to dryness on a water-bath and dry the residue to constant
See Isopropyl myristate (0725). mass at 100-105 °C. The residue weighs not more than 25 mg.
4-Isopropylphenol. C9H12O. (Mr 136.2). 1047300. [99-89-8]. Fine particles. Disperse 5.0 g in 250 mL of water R by shaking
vigorously for 2 min. Immediately pour into a glass cylinder
50 mm in diameter and, using a pipette, transfer 20 mL to
bp : about 212 °C. a glass dish, evaporate to dryness on a water-bath and dry
mp : 59 °C to 61 °C. to constant mass at 100-105 °C. Allow the remainder of the
suspension to stand at 20 °C for 4 h and, using a pipette with
Isopropyl toluenesulfonate. C10H14O3S. (Mr 214.3). 1191100. its tip exactly 5 cm below the surface, withdraw a further
[2307-69-9]. 1-Methylethyl 4-methylbenzenesulfonate. 20 mL without disturbing the sediment, place in a glass dish,
Propan-2-yl 4-methylbenzenesulfonate. Isopropyl tosilate. evaporate to dryness on a water-bath and dry to constant mass
Content : minimum 97.0 per cent. at 100-105 °C. The mass of the second residue is not less than
Clear liquid. 70 per cent of that of the first residue.
mp : about 20 °C. 11-Keto-β-boswellic acid. C30H46O4. (Mr 470.7). 1167600.
[17019-92-0]. 3α-Hydroxy-11-oxours-12-en-24-oic acid.
Isopulegol. C10H18O. (Mr 154.2). 1139600. [89-79-2]. (4β)-3α-Hydroxy-11-oxours-12-en-23-oic acid.
(−)-Isopulegol. (1R,2S,5R)-2-Isopropenyl-5-methyl-
cyclohexanol. White or almost white powder, insoluble in water, soluble in
acetone, in anhydrous ethanol and in methanol.
: about 0.911. mp : 195 °C to 197 °C.
: about 1.472. 11-Keto-β-boswellic acid used in liquid chromatography
bp : about 91 °C. complies with the following additional test.
Isopulegol used in gas chromatography complies with the Assay. Liquid chromatography (2.2.29) as prescribed in the
following additional test. monograph Indian frankincense (2310).
Assay. Gas chromatography (2.2.28) as prescribed in the Content : minimum 90 per cent, calculated by the
monograph Mint oil, partly dementholised (1838). normalisation procedure.

Kieselguhr for chromatography. 1047500. Mix solutions A and B. Add solution C.

White or yellowish-white, light powder, practically insoluble
in water, in dilute acids and in organic solvents. Lactobionic acid. C12H22O12. (Mr 358.3). 1101600. [96-82-2].
Filtration rate. Use a chromatography column 0.25 m long and White or almost white, crystalline powder, freely soluble in
10 mm in internal diameter with a sintered-glass (100) plate water, practically insoluble in ethanol (96 per cent).
and two marks at 0.10 m and 0.20 m above the plate. Place mp : about 115 °C.
sufficient of the substance to be examined in the column to
reach the first mark and fill to the second mark with water R. β-Lactose. C12H22O11. (Mr 342.3). 1150100. [5965-66-2].
When the first drops begin to flow from the column, fill to β-D-Lactose.
the second mark again with water R and measure the time White or slightly yellowish powder.
required for the first 5 mL to flow from the column. The flow Content : minimum 99 per cent.
rate is not less than 1 mL/min. α-D-Lactose : not greater than 35 per cent.
Appearance of the eluate. The eluate obtained in the test for
Assay. Gas chromatography (2.2.28) : use the normalisation
filtration rate is colourless (2.2.2, Method I).
procedure.
Acidity or alkalinity. To 1.00 g add 10 mL of water R, shake
vigorously and allow to stand for 5 min. Filter the suspension Column :
on a filter previously washed with hot water R until the – size : l = 30 m, Ø = 0.25 mm ;
washings are neutral. To 2.0 mL of the filtrate add 0.05 mL – stationary phase : cyanopropyl(3)phenyl(3)methyl(94)polysi-
of methyl red solution R ; the solution is yellow. To 2.0 mL of loxane R (film thickness 1 µm).
the filtrate add 0.05 mL of phenolphthalein solution R1 ; the Carrier gas : helium for chromatography R.
solution is at most slightly pink.
Temperature :
Water-soluble substances. Place 10.0 g in a chromatography
column 0.25 m long and 10 mm in internal diameter and elute Time Temperature
with water R. Collect the first 20 mL of eluate, evaporate to (min) (°C)
dryness and dry the residue at 100 °C to 105 °C. The residue Column 0 - 32.5 20 → 280
weighs not more than 10 mg.
Injection port 250
Iron (2.4.9) : maximum 200 ppm.
Detector 250
To 0.50 g add 10 mL of a mixture of equal volumes of
hydrochloric acid R1 and water R, shake vigorously, allow to
Detection : flame ionisation.
stand for 5 min and filter. 1.0 mL of the filtrate complies with
the test for iron. Injection : an appropriate derivatised sample.
Loss on ignition : maximum 0.5 per cent. During heating to Lactose monohydrate. 1047900. [5989-81-1].
red heat (600 ± 50 °C) the substance does not become brown
or black. See Lactose monohydrate (0187).
Kieselguhr G. 1047600. α-Lactose monohydrate. C12H22O11,H2O. (Mr 360.3).

Consists of kieselguhr treated with hydrochloric acid and 1150000. [5989-81-1]. α-D-Lactose monohydrate.
calcined, to which is added about 15 per cent of calcium White or almost white powder.
sulfate hemihydrate. Content : minimum 97 per cent.
A fine greyish-white powder ; the grey colour becomes more β-D-Lactose : less than 3 per cent.
pronounced on triturating with water. The average particle Assay. Gas chromatography (2.2.28) : use the normalisation
size is 10-40 µm. procedure.
Calcium sulfate content. Determine by the method prescribed
Column :
for silica gel G R.
pH (2.2.3). Shake 1 g with 10 mL of carbon dioxide-free – size : l = 30 m, Ø = 0.25 mm ;
water R for 5 min. The pH of the suspension is 7 to 8. – stationary phase : methylpolysiloxane R (film thickness
Chromatographic separation. Thin-layer chromatography 1 μm).
(2.2.27). Prepare plates using a slurry of the kieselguhr G Carrier gas : helium for chromatography R.
with a 2.7 g/L solution of sodium acetate R. Apply 5 µL of a Temperature :
solution containing 0.1 g/L of lactose, sucrose, glucose and
fructose in pyridine R. Develop over a path of 14 cm using a Time Temperature
mixture of 12 volumes of water R, 23 volumes of 2-propanol R (min) (°C)
and 65 volumes of ethyl acetate R. The migration time of Column 0 - 12.5 230 → 280
the solvent is about 40 min. Dry, spray onto the plate about Injection port 250
10 mL of anisaldehyde solution R and heat for 5-10 min at
100-105 °C. The chromatogram shows four well-defined spots Detector 280
without tailing and well separated from each other.
Lactic acid. 1047800. [50-21-5]. Injection : an appropriate derivatised sample.
See Lactic acid (0458).
Lactulose. 1189600. [4618-18-2].
Lactic reagent. 1047801.
See Lactulose (1230).
Solution A. To 60 mL of lactic acid R add 45 mL of
previously filtered lactic acid R saturated without heating Lanatoside C. C49H76O20. (Mr 985). 1163300. [17575-22-3].
with Sudan red G R ; as lactic acid saturates slowly without 3β-[(β-D-Glucopyranosyl-(1→4)-3-O-acetyl-2,6-
heating, an excess of colorant is always necessary. dideoxy-β-D-ribo-hexopyranosyl-(1→4)-2,6-dideoxy-
Solution B. Prepare 10 mL of a saturated solution of β-D-ribo-hexopyranosyl-(1→4)-2,6-dideoxy-β-D-ribo-
aniline R. Filter. hexopyranosyl)oxy]-12β,14-dihydroxy-5β-card-20(22)-
Solution C. Dissolve 75 mg of potassium iodide R in water enolide.
and dilute to 70 mL with the same solvent. Add 10 mL of Long, flat prisms obtained after recrystallisation in ethanol
ethanol (96 per cent) R and 0.1 g of iodine R. Shake. (96 per cent), freely soluble in pyridine and in dioxan.

Lanthanum chloride heptahydrate. LaCl3,7H2O. (Mr 371.4). Lead acetate cotton. 1048101.

1167200. Immerse absorbent cotton in a mixture of 1 volume
White or almost white powder or colourless crystals, freely of dilute acetic acid R and 10 volumes of lead acetate
soluble in water. solution R. Drain off the excess of liquid, without squeezing
the cotton, by placing it on several layers of filter paper.
Lanthanum chloride solution. 1114001. Allow to dry in air.
To 58.65 g of lanthanum trioxide R slowly add 100 mL of Storage : in an airtight container.
hydrochloric acid R. Heat to boiling. Allow to cool and dilute
to 1000.0 mL with water R. Lead acetate paper. 1048102.
Lanthanum nitrate. La(NO3)3,6H2O. (Mr 433.0). 1048000. Immerse filter paper weighing about 80 g/m2 in a mixture
[10277-43-7]. Lanthanum trinitrate hexahydrate. of 1 volume of dilute acetic acid R and 10 volumes of lead
acetate solution R. After drying, cut the paper into strips
Colourless crystals, deliquescent, freely soluble in water. 15 mm by 40 mm.
Lead acetate solution. 1048103.
Lanthanum nitrate solution. 1048001. A 95 g/L solution of lead acetate R in carbon dioxide-free
A 50 g/L solution of lanthanum nitrate R. water R.
Lanthanum trioxide. La2O3. (Mr 325.8). 1114000. Lead dioxide. PbO2. (Mr 239.2). 1048200. [1309-60-0].
[1312-81-8]. Dark brown powder, evolving oxygen when heated, practically
An almost white, amorphous powder, practically insoluble in insoluble in water, soluble in hydrochloric acid with evolution
water R. It dissolves in dilute solutions of mineral acids and of chlorine, soluble in dilute nitric acid in the presence of
absorbs atmospheric carbon dioxide. hydrogen peroxide, oxalic acid or other reducing agents,
Calcium : maximum 5 ppm. soluble in hot, concentrated alkali hydroxide solutions.
Lauric acid. C12H24O2. (Mr 200.3). 1143100. [143-07-7]. Lead nitrate. Pb(NO3)2. (Mr 331.2). 1048300. [10099-74-8].
Dodecanoic acid. Lead dinitrate.
White or almost white, crystalline powder, practically White or almost white, crystalline powder or colourless
insoluble in water, freely soluble in ethanol (96 per cent). crystals, freely soluble in water.
mp : about 44 °C. Lead nitrate solution. 1048301.
Lauric acid used in the assay of total fatty acids in Saw palmetto A 33 g/L solution of lead nitrate R.
fruit (1848) complies with the following additional test.
Assay. Gas chromatography (2.2.28) as prescribed in the Lead subacetate solution. 1048400. [1335-32-6]. Basic lead
monograph Saw palmetto fruit (1848). acetate solution.
Content : minimum 98 per cent, calculated by the Content : 16.7 per cent m/m to 17.4 per cent m/m of Pb
normalisation procedure. (Ar 207.2) in a form corresponding approximately to the
formula C8H14O10Pb3.
Lauryl alcohol. C12H26O. (Mr 186.3). 1119900. [112-53-8]. Dissolve 40.0 g of lead acetate R in 90 mL of carbon
Dodecan-1-ol. dioxide-free water R. Adjust the pH to 7.5 with strong sodium
: about 0.820. hydroxide solution R. Centrifuge and use the clear colourless
mp : 24 °C to 27 °C. supernatant solution.
Content : minimum 98.0 per cent, determined by gas The solution remains clear when stored in a well-closed
chromatography. container.
Lavandulol. C10H18O. (Mr 154.2). 1114100. [498-16-8]. Leiocarposide. C27H34O16. (Mr 614.5). 1150200.

(R)-5-Methyl-2-(1-methylethenyl)-4-hexen-1-ol. [71953-77-0]. 2-(β-D-Glucopyranosyloxy)benzyl
Oily liquid with a characteristic odour. 3-(β-D-glucopyranosyloxy)-6-hydroxy-2-methoxybenzoate.
2-[[[3-(β-D-Glucopyranosyloxy)-6-hydroxy-2-
Lavandulol used in gas chromatography complies with the methoxybenzoyl]oxy]methyl]phenyl-β-D-glucopyranoside.
White or almost white powder, soluble in water, freely soluble
Assay. Gas chromatography (2.2.28) as prescribed in the in methanol, slightly soluble in ethanol (96 per cent).
monograph Lavender oil (1338).
mp : 190 °C to 193 °C.
Content : minimum 90.0 per cent, calculated by the Lemon oil. 1101700.
normalisation procedure. See Lemon oil (0620).
Lavandulyl acetate. C12H20O2. (Mr 196.3). 1114200. Leucine. 1048500. [61-90-5].
[25905-14-0]. 2-Isopropenyl-5-methylhex-4-en-1-yl acetate. See Leucine (0771).
Colourless liquid with a characteristic odour.
Levodopa. 1170000. [59-92-7].
Lavandulyl acetate used in gas chromatography complies with
the following additional test. See Levodopa (0038).
Assay. Gas chromatography (2.2.28) as prescribed in the (Z)-Ligustilide. C12H14O2. (Mr 190.2). 1180300. [81944-09-4].
monograph Lavender oil (1338). (3Z)-3-Butylidene-1,3,4,5-tetrahydroisobenzofuran-1-one.
Test solution. The substance to be examined. Limonene. C10H16. (Mr 136.2). 1048600. [5989-27-5].
Content : minimum 93.0 per cent, calculated by the D-Limonene. (+)-p-Mentha-1,8-diene. (R)-4-Isopropenyl-1-
normalisation procedure. methylcyclohex-1-ene.
Lead acetate. C4H6O4Pb,3H2O. (Mr 379.3). 1048100. Colourless liquid, practically insoluble in water, soluble in
[6080-56-4]. Lead di-acetate. ethanol (96 per cent).
Colourless crystals, efflorescent, freely soluble in water, soluble : about 0.84.
in ethanol (96 per cent). : 1.471 to 1.474.

: about + 124. nD20 : about 1.480.

bp : 175 °C to 177 °C. Linolenic acid used in the assay of total fatty acids in Saw
Limonene used in gas chromatography complies with the palmetto fruit (1848) complies with the following additional
following additional test. test.
Assay. Gas chromatography (2.2.28) as prescribed in the Assay. Gas chromatography (2.2.28) as prescribed in the
monograph Peppermint oil (0405). monograph Saw palmetto fruit (1848).
Test solution. The substance to be examined. Content : minimum 98 per cent, calculated by the
normalisation procedure. Linolenyl alcohol. C18H32O. (Mr 264.4). 1156200.
[24149-05-1]. (9Z,12Z,15Z)-Octadeca-9,12,15-trien-1-ol.
Linalol. C10H18O. (Mr 154.2). 1048700. [78-70-6]. α-Linolenyl alcohol.
(RS)-3,7-Dimethylocta-1,6-dien-3-ol.
Mixture of two stereoisomers (licareol and coriandrol).
Liquid, practically insoluble in water. Linoleyl alcohol. C18H34O. (Mr 266.5). 1155900. [506-43-4].
(9Z,12Z)-Octadeca-9,12-dien-1-ol.
: about 0.860.
Relative density : 0.830.
: about 1.462.
bp : about 200 °C.
Linalol used in gas chromatography complies with the following Linsidomine hydrochloride. C6H11ClN4O2. (Mr 206.6).
test. 1171200. [16142-27-1]. 3-(Morpholin-4-yl)sydnonimine
Assay. Gas chromatography (2.2.28) as prescribed in the hydrochloride. 3-(Morpholin-4-yl)-1,2,3-oxadiazol-3-ium-5-
monograph Anise oil (0804). aminide hydrochloride.
Content : minimum 98.0 per cent, calculated by the Liquid scintillation cocktail. 1167300.
normalisation procedure. Commercially available solution for the determination of
radioactivity by liquid scintillation counting. It contains
Linalyl acetate. C12H20O2. (Mr 196.3). 1107200. [115-95-7]. one or more fluorescent agents and mostly one or more
(RS)-1,5-Dimethyl-1-vinylhex-4-enyl acetate. emulsifying agents in a suitable organic solvent or mixture of
Colourless or slightly yellow liquid with a strong odour of organic solvents.
bergamot and lavender.
: 0.895 to 0.912. Liquid scintillation cocktail R1. 1176800.
: 1.448 to 1.451. To 1000 mL of dioxan R, add 0.3 g of methylphenyloxazolylben-
zene R, 7 g of diphenyloxazole R and 100 g of naphthalene R.
bp : about 215 °C.
Linalyl acetate used in gas chromatography complies with the Lithium. Li. (Ar 6.94). 1048800. [7439-93-2].
following additional test. A soft metal whose freshly cut surface is silvery-grey. It rapidly
Assay. Gas chromatography (2.2.28) as prescribed in the tarnishes in contact with air. It reacts violently with water,
monograph Bitter-orange-flower oil (1175). yielding hydrogen and giving a solution of lithium hydroxide ;
soluble in methanol, yielding hydrogen and a solution of
Test solution. The substance to be examined. lithium methoxide ; practically insoluble in light petroleum.
Content : minimum 95.0 per cent, calculated by the Storage : under light petroleum or liquid paraffin.
Lithium carbonate. Li2CO3. (Mr 73.9). 1048900. [554-13-2].
Lindane. C6H6Cl6. (Mr 290.8). 1128900. [58-89-9]. Dilithium carbonate.
γ-Hexachlorocyclohexane.
White or almost white, light powder, sparingly soluble
For the monograph Wool fat (0134), a suitable certified in water, very slightly soluble in ethanol (96 per cent). A
reference solution (10 ng/µL in cyclohexane) may be used. saturated solution at 20 °C contains about 13 g/L of Li2CO3.
Linoleic acid. C18H32O2. (Mr 280.5). 1143200. [60-33-3]. Lithium chloride. LiCl. (Mr 42.39). 1049000. [7447-41-8].
(9Z,12Z)-Octadeca-9,12-dienoic acid.
Crystalline powder or granules or cubic crystals, deliquescent,
Colourless, oily liquid. freely soluble in water, soluble in acetone and in ethanol
d 420 : about 0.903. (96 per cent). Aqueous solutions are neutral or slightly
alkaline.
nD20 : about 1.470.
Linoleic acid used in the assay of total fatty acids in Saw
palmetto fruit (1848) complies with the following additional Lithium hydroxide. LiOH,H2O. (Mr 41.96). 1049100.
test. [1310-66-3]. Lithium hydroxide monohydrate.
Assay. Gas chromatography (2.2.28) as prescribed in the White or almost white, granular powder, strongly alkaline, it
monograph Saw palmetto fruit (1848). rapidly absorbs water and carbon dioxide, soluble in water,
Content : minimum 98 per cent, calculated by the sparingly soluble in ethanol (96 per cent).
normalisation procedure. Storage : in an airtight container.
Linolenic acid. C18H30O2. (Mr 278.4). 1143300. [463-40-1]. Lithium metaborate, anhydrous. LiBO2. (Mr 49.75).
(9Z,12Z,15Z)-Octadeca-9,12,15-trienoic acid. α-Linolenic 1120000. [13453-69-5].
acid.
Lithium sulfate. Li2SO4,H2O. (Mr 128.0). 1049200.
Colourless liquid, practically insoluble in water, soluble in [10102-25-7]. Dilithium sulfate monohydrate.
organic solvents.
Colourless crystals, freely soluble in water, practically
d 420 : about 0.915. insoluble in ethanol (96 per cent).

Lithium trifluoromethanesulfonate. CF3LiO3S. (Mr 156.0). Lutetium chloride hexahydrate. LuCl3,6H2O. (Mr 389.4).

1173400. [33454-82-9]. 1199600. [15230-79-2].
Litmus. 1049300. [1393-92-6]. White to yellow, crystalline powder, freely soluble in water.
Schultz No. 1386. Lysyl endopeptidase. 1188000. [78642-25-8].
Indigo-blue fragments prepared from various species of Achromobacter endoproteinase I. Lysyl bond specific
Rocella, Lecanora or other lichens, soluble in water, practically proteinase (EC 3.4.21.50).
insoluble in ethanol (96 per cent). It belongs to the serine endopeptidase family. Initially isolated
Colour change : pH 5 (red) to pH 8 (blue). from Achromobacter lyticus. Enzymes with similar specificity
are produced by Lysobacter enzymogenes (endoproteinase
Litmus paper, blue. 1049301. Lys-C) and Pseudomonas aeruginosa (Ps-1). It cleaves
Boil 10 parts of coarsely powdered litmus R for 1 h with peptide bonds at the carboxy-terminal of both lysine residues
100 parts of ethanol (96 per cent) R. Decant the alcohol and S-aminoethylcysteine residues with a high degree of
and add to the residue a mixture of 45 parts of ethanol specificity. 1 amidase unit (U) is the amount of enzyme
(96 per cent) R and 55 parts of water R. After 2 days decant that will produce 1 micromole of p-nitroaniline from
the clear liquid. Impregnate strips of filter paper with the N-benzoyl-DL-lysine-p-nitroaniline per minute at 30 °C at
solution and allow to dry. pH 9.5.
Test for sensitivity. Immerse a strip measuring 10 mm by
Macrogol 23 lauryl ether. 1129000.
60 mm in a mixture of 10 mL of 0.02 M hydrochloric acid
and 90 mL of water R. On shaking the paper turns red See Macrogol lauryl ether (1124), the number of moles of
within 45 s. ethylene oxide reacted per mole of lauryl alcohol being 23
(nominal value).
Litmus paper, red. 1049302.
Macrogol 200. 1099200. [25322-68-3]. Polyethyleneglycol
To the blue litmus extract, add dilute hydrochloric acid R 200.
dropwise until the blue colour becomes red. Impregnate
strips of filter paper with the solution and allow to dry. Clear, colourless or almost colourless viscous liquid, very
soluble in acetone and in anhydrous ethanol, practically
Test for sensitivity. Immerse a strip measuring 10 mm by insoluble in fatty oils.
60 mm in a mixture of 10 mL of 0.02 M sodium hydroxide
and 90 mL of water R. On shaking the paper turns blue : about 1.127.
within 45 s. : about 1.450.
Loganin. C17H26O10. (Mr 390.4). 1136700. [18524-94-2]. Macrogol 200 R1. 1099201.
Methyl (1S,4aS,6S,7R,7aS)-1-(β-D-glucopyranosyloxy)- Introduce 500 mL of macrogol 200 R into a 1000 mL round
6-hydroxy-7-methyl-1,4a,5,6,7,7a-hexahydro- bottom flask. Using a rotation evaporator remove any
cyclopenta[c]pyran-4-carboxylate. volatile components applying for 6 h a temperature of 60 °C
mp : 220 °C to 221 °C. and a vacuum with a pressure of 1.5-2.5 kPa.
Longifolene. C15H24. (Mr 204.4). 1150300. [475-20-7]. Macrogol 300. 1067100. [25322-68-3]. Polyethyleneglycol
(1S,3aR,4S,8aS)-4,8,8-Trimethyl-9-methylenedecahydro-1,4- 300.
methanoazulene. See Macrogols (1444).
Oily, colourless liquid, practically insoluble in water, miscible
with ethanol (96 per cent). 400.
d 418 : 0.9319. See Macrogols (1444).
: 1.5050.
: + 42.7. 600.
bp : 254 °C to 256 °C. See Macrogols (1444).
Longifolene used in gas chromatography complies with the
following additional test. Macrogol 1000. 1067300. [25322-68-3]. Polyethyleneglycol
1000.
monograph Turpentine oil, Pinus pinaster type (1627). See Macrogols (1444).
Content : minimum 98.0 per cent, calculated by the Macrogol 1500. 1067400. [25322-68-3]. Polyethyleneglycol
normalisation procedure. 1500.
Low-vapour-pressure hydrocarbons (type L). 1049400. See Macrogols (1444).
Unctuous mass, soluble in benzene and in toluene. Macrogol 4000. 1198000. [25322-68-3]. Polyethyleneglycol
4000.
Lumiflavine. C13H12N4O2. (Mr 256.3). 1141000. [1088-56-8].
7,8,10-Trimethylbenzo[g]pteridine-2,4(3H,10H)-dione. See Macrogols (1444).
Yellow powder or orange crystals, very slightly soluble in Macrogol 6000. 1189800. [25322-68-3]. Polyethyleneglycol
water, freely soluble in methylene chloride. 6000.
Luteolin. C15H10O6. (Mr 286.2). 1198500. [491-70-3]. 2-(3,4- White or almost white solid with a waxy or paraffin-like
Dihydroxyphenyl)-5,7-dihydroxy-4H-1-benzopyran-4-one. appearance, very soluble in water and in methylene chloride,
practically insoluble in ethanol (96 per cent), in fatty oils and
Luteolin-7-glucoside. C21H20O11. (Mr 448.4). 1163400. in mineral oils.
[5373-11-5]. 2-(3,4-Dihydroxyphenyl)-7-(β-D-
glucopyranosyloxy)-5-hydroxy-4H-1-benzopyran-4-one. Macrogol 20 000. 1067600. Polyethyleneglycol 20 000.
Yellow powder. See Macrogols (1444).
Absorbance (2.2.25). A solution in methanol R shows Macrogol 20 000 2-nitroterephthalate. 1067601.
absorption maxima at 255 nm, 267 nm and 350 nm. Polyethyleneglycol 20 000 with embedded 2-nitroterephthalate
mp : about 247 °C. groups.

Macrogol, base-deactivated. 1170300. Maize oil. 1050400.

Base-deactivated polyethyleneglycol. See Maize oil, refined (1342).
Macrogol cetostearyl ether. 1196100. Makisterone A. C28H46O7. (Mr 494.7). 1207200. [20137-14-8].
See Macrogol cetostearyl ether (1123). (22R)-2β,3β,14,20,22,25-Hexahydroxy-5β-ergost-7-en-6-one.
Macrogol, polar-deactivated. 1179000. Malachite green. C23H25ClN2. (Mr 364.9). 1050500.
Polar-deactivated polyethyleneglycol. [123333-61-9].
Schultz No. 754.
Magnesium. Mg. (Ar 24.30). 1049500. [7439-95-4].
Silver-white ribbon, turnings or wire, or a grey powder. [4-[[4-(Dimethylamino)phenyl]phenylmethylene]cyclohexa-
Magnesium acetate. C4H6MgO4,4H2O. (Mr 214.5). 1049600. 2,5-dien-1-ylidene]dimethylammonium chloride.
[16674-78-5]. Magnesium diacetate tetrahydrate. Green crystals with a metallic lustre, very soluble in water
Colourless crystals, deliquescent, freely soluble in water and giving a bluish-green solution, soluble in ethanol (96 per cent)
in ethanol (96 per cent). and in methanol.
Storage : in an airtight container. Absorbance (2.2.25). A 0.01 g/L solution in ethanol (96 per
cent) R shows an absorption maximum at 617 nm.
Magnesium chloride. 1049700. [7791-18-6].
See Magnesium chloride hexahydrate (0402). Malachite green solution. 1050501.
A 5 g/L solution of malachite green R in anhydrous acetic
Magnesium nitrate. Mg(NO3)2,6H2O. (Mr 256.4). 1049800. acid R.
[13446-18-9]. Magnesium nitrate hexahydrate.
Colourless, clear crystals, deliquescent, very soluble in water, Malathion. C10H19O6PS2. (Mr 330.3). 1129200. [121-75-5].
freely soluble in ethanol (96 per cent). bp : about 156 °C.
Storage : in an airtight container. A suitable certified reference solution (10 ng/µL in iso-octane)
may be used.
Magnesium nitrate solution. 1049801.
Dissolve 17.3 g of magnesium nitrate R in 5 mL of water R Maleic acid. 1050600. [110-16-7].
warming gently and add 80 mL of ethanol (96 per cent) R. See Maleic acid (0365).
Cool and dilute to 100.0 mL with the same solvent.
Maleic anhydride. C4H2O3. (Mr 98.1). 1050700. [108-31-6].
Magnesium oxide. 1049900. [1309-48-4]. Butenedioic anhydride. 2,5-Furandione.
See Light magnesium oxide (0040). White or almost white crystals, soluble in water forming
Magnesium oxide R1. 1049901. maleic acid, very soluble in acetone and in ethyl acetate, freely
soluble in toluene, soluble in ethanol (96 per cent) with ester
Complies with the requirements prescribed for magnesium
formation, very slightly soluble in light petroleum.
oxide R with the following modifications.
mp : about 52 °C.
Arsenic (2.4.2, Method A) : maximum 2 ppm.
Dissolve 0.5 g in a mixture of 5 mL of water R and 5 mL of Any residue insoluble in toluene does not exceed 5 per cent
hydrochloric acid R1. (maleic acid).
Heavy metals (2.4.8) : maximum 10 ppm. Maleic anhydride solution. 1050701.
Dissolve 1.0 g in a mixture of 3 mL of water R and 7 mL Dissolve 5 g of maleic anhydride R in toluene R and dilute
of hydrochloric acid R1. Add 0.05 mL of phenolphthalein to 100 mL with the same solvent. Use within one month.
solution R and concentrated ammonia R until a pink colour If the solution becomes turbid, filter.
is obtained. Neutralise the excess of ammonia by the
addition of glacial acetic acid R. Add 0.5 mL in excess and Malic acid. 1200400. [6915-15-7].
dilute to 20 mL with water R. Filter, if necessary. 12 mL of See Malic acid (2080).
the solution complies with test A. Prepare the reference
solution using a mixture of 5 mL of lead standard solution Maltitol. 1136800. [585-88-6].
(1 ppm Pb) R and 5 mL of water R. See Maltitol (1235).
Iron (2.4.9) : maximum 50 ppm.
Maltol. C6H6O3. (Mr 126.1). 1202300. [118-71-8].
Dissolve 0.2 g in 6 mL of dilute hydrochloric acid R and 3-Hydroxy-2-methyl-4H-pyran-4-one.
White or almost white crystalline powder, soluble in hot water.
Magnesium oxide, heavy. 1050000. [1309-48-4]. mp : 161 °C to 162 °C.
See Heavy magnesium oxide (0041).
Maltose monohydrate. C12H22O11,H2O. (Mr 360.3). 1193100.
Magnesium silicate for pesticide residue analysis. 1129100. [6363-53-7]. 4-O-α-D-glucopyranosyl-D-glucopyranose
[1343-88-0]. monohydrate.
Magnesium silicate for chromatography (60-100 mesh).
Maltotriose. C18H32O16. (Mr 504.4). 1176300. [1109-28-0].
Magnesium sulfate. 1050200. [10034-99-8]. α-D-Glucopyranosyl-(1→4)-α-D-glucopyranosyl-(1→4)-D-
See Magnesium sulfate heptahydrate (0044). glucose.
White or almost white, crystalline powder, very soluble in
Magnolin. C23H28O7. (Mr 416.5). 1200300. [31008-18-1]. water.
(3S,3aR,6S,6aR)-3-(3,4-Dimethoxyphenyl)-6-(3,4,5- mp : about 134 °C.
trimethoxyphenyl)-1,3,3a,4,6,6a-hexahydrofuro[3,4-c]furan.
Mandelic acid. C8H8O3. (Mr 152.1). 1171300. [90-64-2].
Magnolol. C18H18O2. (Mr 266.3). 1182800. [528-43-8]. 2-Hydroxy-2-phenylacetic acid.
5,5′-Di(prop-2-enyl)biphenyl-2,2′-diol. 5,5′-Diallyl-2,2′-
dihydroxybiphenyl. 5,5′-Di-2-propenyl-[1,1′-biphenyl]-2,2′- White crystalline flakes, soluble in water.
diol. mp : 118 to 121 °C.

Manganese sulfate. MnSO4,H2O. (Mr 169.0). 1050900. Menthone. C10H18O. (Mr 154.2). 1051700. [14073-97-3].
[10034-96-5]. Manganese sulfate monohydrate. (2S,5R)-2-Isopropyl-5-methylcyclohexanone.
Pale-pink, crystalline powder or crystals, freely soluble in (–)-trans-p-Menthan-3-one.
water, practically insoluble in ethanol (96 per cent). Contains variable amounts of isomenthone.
Loss on ignition : 10.0 per cent to 12.0 per cent, determined on Colourless liquid, very slightly soluble in water, very soluble
1.000 g at 500 ± 50 °C. in ethanol (96 per cent).
: about 0.897.
Mannitol. 1051000. [69-65-8].
: about 1.450.
See Mannitol (0559).
Menthone used in gas chromatography complies with the
Mannose. C6H12O6. (Mr 180.2). 1051100. [3458-28-4]. following additional test.
D-(+)-Mannose. Assay. Gas chromatography (2.2.28) as prescribed in the
white or almost white, crystalline powder or small crystals, monograph Peppermint oil (0405).
very soluble in water, slightly soluble in anhydrous ethanol. Test solution. The substance to be examined.
: + 13.7 + 14.7, determined on a 200 g/L solution in Content : minimum 90.0 per cent, calculated by the
water R containing about 0.05 per cent of NH3. normalisation procedure.
Menthyl acetate. C12H22O2. (Mr 198.3). 1051800. [2623-23-6].
Marrubiin. C20H28O4. (Mr 332.4). 1158300. [465-92-9]. (1R,2S,5R)-5-Methyl-2-(propan-2-yl)cyclohexyl acetate.
(2aS,5aS,6R,7R,8aR,8bR)-6-[2-(Furan-3-yl)ethyl]-6-hydroxy- Colourless liquid, slightly soluble in water, miscible with
2a,5a,7-trimethyldecahydro-2H-naphtho[1,8-bc]furan-2-one. ethanol (96 per cent).
Colourless, microcrystalline powder. 20
d 20 : about 0.92.
Marrubiin used in liquid chromatography complies with the nD20 : about 1.447.
bp : about 228 °C.
monograph White horehound (1835). Menthyl acetate used in gas chromatography complies with the
normalisation procedure. Assay. Gas chromatography (2.2.28) as prescribed in the
monograph Peppermint oil (0405).
Meclozine dihydrochloride. 1051200. [1104-22-9]. Test solution. The substance to be examined.
See Meclozine dihydrochloride (0622). Content : minimum 97.0 per cent, calculated by the
Medronic acid. 1193200. [1984-15-2].
See Medronic acid for radiopharmaceutical preparations 2-Mercaptobenzimidazole. C7H6N2S. (Mr 150.2). 1170100.
(2350). [583-39-1]. 1H-benzimidazole-2-thiol.
mp : about 302 °C.
Melamine. C3H6N6. (Mr 126.1). 1051300. [108-78-1].
1,3,5-Triazine-2,4,6-triamine. 2-Mercaptoethanol. C2H6OS. (Mr 78.1). 1099300. [60-24-2].
A white or almost white, amorphous powder, very slightly Liquid, miscible with water.
soluble in water and in ethanol (96 per cent). : about 1.116.
Menadione. 1051400. [58-27-5]. bp : about 157 °C.
See Menadione (0507). Mercaptopurine. 1051900. [6112-76-1].
Menthofuran. C10H14O. (Mr 150.2). 1051500. See Mercaptopurine (0096).
[17957-94-7]. 3,9-Epoxy-p-mentha-3,8-diene. Mercuric acetate. C4H6HgO4. (Mr 318.7). 1052000.
3,6-Dimethyl-4,5,6,7-tetrahydro-benzofuran. [1600-27-7]. Mercury diacetate.
Slightly bluish liquid, very slightly soluble in water, soluble in White or almost white crystals, freely soluble in water, soluble
ethanol (96 per cent). in ethanol (96 per cent).
: about 0.965.
Mercuric acetate solution. 1052001.
: about 1.480.
Dissolve 3.19 g of mercuric acetate R in anhydrous acetic
: about + 93. acid R and dilute to 100 mL with the same acid. If
bp : 196 °C. necessary, neutralise the solution with 0.1 M perchloric acid
Menthofuran used in gas chromatography complies with the using 0.05 mL of crystal violet solution R as indicator.
following additional test. Mercuric chloride. 1052200. [7487-94-7].
Assay. Gas chromatography (2.2.28) as prescribed in the See Mercuric chloride (0120).
monograph Peppermint oil (0405).
Test solution. The substance to be examined. Mercuric chloride solution. 1052201.
Content : minimum 97.0 per cent, calculated by the A 54 g/L solution of mercuric chloride R.
normalisation procedure. Mercuric iodide. HgI2. (Mr 454.4). 1052300. [7774-29-0].
Menthol. 1051600. [2216-51-5]. Mercury di-iodide.
See Levomenthol (0619) and Racemic menthol (0623). Dense, scarlet, crystalline powder, slightly soluble in water,
sparingly soluble in acetone and in ethanol (96 per cent),
Menthol used in gas chromatography complies with the soluble in an excess of potassium iodide solution R.
related substances test included in the monograph Racemic Mercuric nitrate. Hg(NO3)2,H2O. (Mr 342.6). 1052400.
menthol (0623). [7783-34-8]. Mercury dinitrate monohydrate.
Content : minimum 98.0 per cent, calculated by the Colourless or slightly coloured crystals, hygroscopic, soluble
normalisation procedure. in water in the presence of a small quantity of nitric acid.

Storage : in an airtight container, protected from light. Methane. CH4. (Mr 16). 1166300. [74-82-8].
Mercuric oxide. HgO. (Mr 216.6). 1052500. [21908-53-2]. Content : minimum 99.0 per cent V/V.
Yellow mercuric oxide. Mercury oxide. Methane R1. CH4. (Mr 16). 1176400. [74-82-8].
A yellow to orange-yellow powder, practically insoluble in Content : minimum 99.995 per cent V/V.
water and in ethanol (96 per cent).
Storage : protected from light. Methanesulfonic acid. CH4O3S. (Mr 96.1). 1053100.
[75-75-2].
Mercuric sulfate solution. 1052600. [7783-35-9]. Clear, colourless liquid, solidifying at about 20 °C, miscible
Dissolve 1 g of mercuric oxide R in a mixture of 20 mL of with water, slightly soluble in toluene, practically insoluble
water R and 4 mL of sulfuric acid R. in hexane.
Mercuric thiocyanate. Hg(SCN)2. (Mr 316.7). 1052700. : about 1.48.
[592-85-8]. Mercury di(thiocyanate). : about 1.430.
White or almost white, crystalline powder, very slightly Methanesulfonyl chloride. CH3ClO2S. (Mr 114.6). 1181300.
soluble in water, slightly soluble in ethanol (96 per cent), [124-63-0].
soluble in solutions of sodium chloride. Clear, colourless or slightly yellow liquid.
Mercuric thiocyanate solution. 1052701. Content : minimum 99.0 per cent.
Dissolve 0.3 g of mercuric thiocyanate R in anhydrous Density : 1.48 g/cm3.
ethanol R and dilute to 100 mL with the same solvent. : about 1.452.
Storage : use within 1 week. bp : about 161 °C.
Mesityl oxide. C6H10O. (Mr 98.1). 1120100. [141-79-7]. Methanol. CH4O. (Mr 32.04). 1053200. [67-56-1].
4-Methylpent-3-en-2-one.
Clear, colourless, flammable liquid, miscible with water and
Colourless, oily liquid, soluble in 30 parts of water, miscible with ethanol (96 per cent).
with most organic solvents.
: 0.791 to 0.793.
: about 0.858.
bp : 64 °C to 65 °C.
bp : 129 °C to 130 °C.
Methanol R1. 1053201.
Metanil yellow. C18H14N3NaO3S. (Mr 375.4). 1052900.
Complies with the requirements prescribed for methanol R
[587-98-4].
with the following additional requirement.
Schultz No. 169.
Absorbance (2.2.25) : maximum 0.70 at 210 nm, 0.30 at
Colour Index No. 13065. 220 nm, 0.13 at 230 nm, 0.02 at 250 nm, 0.01 at 260 nm
Sodium 3-[4-(phenylamino)phenylazo]benzenesulfonate. and higher wavelengths, determined using water R as
A brownish-yellow powder, soluble in water and in ethanol compensation liquid.
(96 per cent).
Methanol R2. 1053202.
Metanil yellow solution. 1052901. Complies with the requirements prescribed for methanol R
A 1 g/L solution of metanil yellow R in methanol R. and the following additional requirements.
Test for sensitivity. To 50 mL of anhydrous acetic acid R Content : minimum 99.8 per cent.
add 0.1 mL of the metanil yellow solution. Add 0.05 mL of Absorbance (2.2.25) : maximum 0.17, determined at 225 nm
0.1 M perchloric acid ; the colour changes from pinkish-red using water R as the compensation liquid.
to violet.
Colour change : pH 1.2 (red) to pH 2.3 (orange-yellow). Methanol, hydrochloric. 1053203.
Dilute 1.0 mL of hydrochloric acid R1 to 100.0 mL with
Metaphosphoric acid. (HPO3)x. 1053000. [37267-86-0]. methanol R.
Glassy lumps or sticks containing a proportion of sodium
metaphosphate, hygroscopic, very soluble in water. Methanol, aldehyde-free. 1053300.
Nitrates. Boil 1.0 g with 10 mL of water R, cool, add 1 mL Dissolve 25 g of iodine R in 1 L of methanol R and pour the
of indigo carmine solution R, 10 mL of nitrogen-free sulfuric solution, with constant stirring, into 400 mL of 1 M sodium
acid R and heat to boiling. The blue colour is not entirely hydroxide. Add 150 mL of water R and allow to stand for
discharged. 16 h. Filter. Boil under a reflux condenser until the odour
Reducing substances : maximum 0.01 per cent, calculated as of iodoform disappears. Distil the solution by fractional
H3PO3. distillation.
Dissolve 35.0 g in 50 mL of water R. Add 5 mL of a 200 g/L Aldehydes and ketones: maximum 0.001 per cent.
solution of sulfuric acid R, 50 mg of potassium bromide R and Methanol, anhydrous. 1053400. [67-56-1].
5.0 mL of 0.02 M potassium bromate and heat on a water-bath Treat 1000 mL of methanol R with 5 g of magnesium R. If
for 30 min. Allow to cool and add 0.5 g of potassium iodide R. necessary initiate the reaction by adding 0.1 mL of mercuric
Titrate the liberated iodine with 0.1 M sodium thiosulfate, chloride solution R. When the evolution of gas has ceased,
using 1 mL of starch solution R as indicator. Carry out a blank distil the liquid and collect the distillate in a dry container
test. protected from moisture.
1 mL of 0.02 M potassium bromate is equivalent to 4.10 mg Water (2.5.12) : maximum 0.3 g/L.
of H3PO3.
Storage : in an airtight container. DL-Methionine. 1129400. [59-51-8].
See DL-Methionine (0624).
Methacrylic acid. C4H6O2. (Mr 86.1). 1101800. [79-41-4].
2-Methylprop-2-enoic acid. L-Methionine. 1053500. [63-68-3].
Colourless liquid. See Methionine (1027).
: about 1.431. L-Methionine sulfoxide. C5H11NO3S. (Mr 165.2). 1193300.
bp : about 160 °C. [3226-65-1]. (2S)-2-Amino-4-[(RS)-methylsulfinyl]butanoic
mp : about 16 °C. acid.

(RS)-Methotrexate. C20H22N8O5. 1120200. [60388-53-6]. Methyl 4-acetylbenzoate reagent. 1154101.

(RS)-2-[4-[[(2,4-diaminopteridin-6-yl)methyl]- Dissolve 0.25 g of methyl 4-acetylbenzoate R in a mixture of
methylamino]benzoylamino]pentanedioic acid. 5 mL of sulfuric acid R and 85 mL of cooled methanol R.
Methyl acrylate. C4H6O2. (Mr 86.1). 1199200. [96-33-3].
mp : about 195 °C. Methyl prop-2-enoate.
Methoxychlor. C16H15Cl3O2. (Mr 345.7). 1129300. [72-43-5]. Clear, colourless liquid.
1,1-(2,2,2-Trichloroethylidene)-bis(4-methoxybenzene). bp : about 80 °C.
Practically insoluble in water, freely soluble in most organic
solvents. Methylal. C3H8O2. (Mr 76.1). 1173500. [109-87-5].
Dimethoxymethane. Dioxapentane. Formaldehyde dimethyl
bp : about 346 °C. acetal. Methylene dimethyl ether.
mp : 78 °C to 86 °C. Clear, colourless, volatile, flammable liquid, soluble in water
A suitable certified reference solution (10 ng/µL in iso-octane) and miscible with ethanol (96 per cent).
may be used. : about 0.860.
trans-2-Methoxycinnamaldehyde. C10H10O2. (Mr 162.2). : about 1.354.
1129500. [60125-24-8]. bp : about 41 °C.
mp : 44 °C to 46 °C. Methylal used in gas chromatography complies with the
trans-2-Methoxycinnamaldehyde used in gas chromatography following additional test.
complies with the following additional test. Content : minimum 99.5 per cent, determined by gas
Assay. Gas chromatography (2.2.28) as prescribed in the chromatography.
Methylamine hydrochloride. CH6ClN. (Mr 67.5). 1198600.
Content : minimum 96.0 per cent, calculated by the [593-51-1]. Methanamine hydrochloride.
(1RS)-1-(6-Methoxynaphthalen-2-yl)ethanol. Content : minimum 98.0 per cent.
C13H14O2. (Mr 202.3). 1159600. [77301-42-9].
6-Methoxy-α-methyl-2-naphthalenemethanol. Methyl 4-aminobenzoate. C8H9NO2. (Mr 151.2). 1175600.
White or almost white powder. [619-45-4].
mp : about 113 °C. mp : 110 °C to 113 °C.
1-(6-Methoxynaphthalen-2-yl)ethanone. C13H12O2. 4-Methylaminophenol sulfate. C14H20N2O6S. (Mr 344.4).

(Mr 200.2). 1159700. [3900-45-6]. 6′-Methoxy-2′- 1053800. [55-55-0].
acetonaphthone. Colourless crystals, very soluble in water, slightly soluble in
White or almost white powder. ethanol (96 per cent).
6-Methoxy-2-naphthoic acid. C12H10O3. (Mr 202.2). 3-(Methylamino)-1-phenylpropan-1-ol. C10H15NO.

1184200. [2471-70-7]. 6-Methoxynaphthalene-2-carboxylic (Mr 165.2). 1186400. [42142-52-9].
acid. White or almost white powder.
White or almost white, crystalline powder. mp : 59 °C to 64 °C.
mp : 201 °C to 206 °C. Methyl anthranilate. C8H9NO2. (Mr 151.2). 1107300.
[134-20-3]. Methyl 2-aminobenzoate.
Methoxyphenylacetic acid. C9H10O3. (Mr 166.2). 1053600.
[7021-09-2]. (RS)-2-Methoxy-2-phenylacetic acid. Colourless crystals or a colourless or yellowish liquid, soluble
in water, freely soluble in ethanol (96 per cent).
White, crystalline powder or white or almost white crystals,
sparingly soluble in water, freely soluble in ethanol (96 per mp : 24 °C to 25 °C.
cent). Methyl anthranilate used in gas chromatography complies with
mp : about 70 °C. the following additional test.
Methoxyphenylacetic reagent. 1053601. monograph Bitter-orange-flower oil (1175).
Dissolve 2.7 g of methoxyphenylacetic acid R in 6 mL of Test solution. The substance to be examined.
tetramethylammonium hydroxide solution R and add 20 mL
of anhydrous ethanol R. Content : minimum 95.0 per cent, calculated by the
Storage : in a polyethylene container.
Methyl arachidate. C21H42O2. (Mr 326.6). 1053900.
3-Methoxy- L-tyrosine. C10H13NO4H2O. (Mr 229.2). 1164400. [1120-28-1]. Methyl eicosanoate.
[200630-46-2].
Off-white or yellow powder. chromatography (2.4.22).
Methyl acetate. C3H6O2. (Mr 74.1). 1053700. [79-20-9]. White or yellow, crystalline mass, soluble in ethanol (96 per
Clear, colourless liquid, soluble in water, miscible with ethanol cent) and in light petroleum.
(96 per cent). mp : about 46 °C.
: about 0.933. Methyl behenate. C23H46O2. (Mr 354.6). 1107500. [929-77-1].
: about 1.361. Methyl docosanoate.
bp : 56 °C to 58 °C. mp : 54 °C to 55 °C.
Methyl 4-acetylbenzoate. C10H10O3. (Mr 178.2). 1154100. Methyl benzenesulfonate. C7H8O3S. (Mr 172.2). 1159800.
[3609-53-8]. [80-18-2].
mp : about 94 °C. Content : minimum 98.0 per cent.

Clear, colourless liquid. Methyl 4-(butylamino)benzoate. C12H17NO2. (Mr 207.3).

bp : about 148 °C. 1207300. [71839-12-8].
White or almost white solid.
Methyl benzoate. C8H8O2. (Mr 136.2). 1164500. [93-58-3].
Benzoic acid, methyl ester. Content : minimum 99.9 per cent.
Colourless liquid. Methyl caprate. 1054000.
: 1.088.
See Methyl decanoate R.
bp : about 200 °C.
Methylbenzothiazolone hydrazone hydrochloride. Methyl caproate. C7H14O2. (Mr 130.2). 1120300. [106-70-7].
C8H10ClN3S,H2O. (Mr 233.7). 1055300. [38894-11-0]. Methyl hexanoate.
3-Methylbenzothiazol-2(3H)-one hydrazone hydrochloride : about 0.885.
monohydrate. : about 1.405.
Almost white or yellowish, crystalline powder. bp : 150 °C to 151 °C.
mp : about 270 °C.
Suitability for determination of aldehydes. To 2 mL of Methyl caprylate. C9H18O2. (Mr 158.2). 1120400. [111-11-5].
aldehyde-free methanol R add 60 µL of a 1 g/L solution of Methyl octanoate.
propionaldehyde R in aldehyde-free methanol R and 5 mL : about 0.876.
of a 4 g/L solution of methylbenzothiazolone hydrazone : about 1.417.
hydrochloride. Mix. Allow to stand for 30 min. Prepare a
blank omitting the propionaldehyde solution. Add 25.0 mL of bp : 193 °C to 194 °C.
a 2 g/L solution of ferric chloride R to the test solution and to
Methylcellulose 450. 1055500. [9004-67-5].
the blank, dilute to 100.0 mL with acetone R and mix. The
absorbance (2.2.25) of the test solution, measured at 660 nm See Methylcellulose (0345).
using the blank as compensation liquid, is not less than 0.62. Nominal viscosity : 450 mPa·s.
(R)-(+)-α-Methylbenzyl isocyanate. C9H9NO. (Mr 147.2). Methyl cinnamate. C10H10O2. (Mr 162.2). 1099400.
1171400. [33375-06-3]. (+)-(R)-α-Methylbenzyl isocyanate. [103-26-4].
(+)-[(1R)-1-Isocyanatoethyl]benzene. (+)-(1R)-1-Phenylethyl
isocyanate. Colourless crystals practically insoluble in water, soluble in
Colourless liquid. : about 1.56.
: about 1.045. bp : about 260 °C.
: about 1.513. mp : 34 °C to 36 °C.
bp : 55 °C to 56 °C at 2.5 mm Hg. Methylcyclohexane. C7H14. (Mr 98.2). 1189900. [108-87-2].
Enantiomeric purity : minimum 99.5.
Storage : at a temperature of 2 °C to 8 °C. Methyl decanoate. C11H22O2. (Mr 186.3). 1054000.
[110-42-9].
(S)-(−)-α-Methylbenzyl isocyanate. C9H9NO. (Mr 147.2).
1170200. [14649-03-7]. (−)-(S)-α-Methylbenzyl isocyanate.
(−)-[(1S)-1-Isocyanatoethyl]benzene. (−)-(1S)-1-Phenylethyl Clear, colourless or yellow liquid, soluble in light petroleum.
isocyanate. : 0.871 to 0.876.
Content : minimum 99.0 per cent. : 1.425 to 1.426.
Colourless liquid. Foreign substances. Gas chromatography (2.2.28), injecting
20
d 20 : about 1.045. equal volumes of each of the following :
nD20 : about 1.514. A 0.02 g/L solution of the substance to be examined in carbon
bp : 55 °C to 56 °C at 2.5 mm Hg. disulfide R (solution A), a 2 g/L solution of the substance to
be examined in carbon disulfide R (solution B), and carbon
Enantiomeric purity : minimum 99.5 per cent. disulfide R (solution C). Carry out the chromatographic
Storage : at a temperature of 2 °C to 8 °C. procedure under the conditions of the test for butylated
NOTE : do not use the reagent if it is coloured. hydroxytoluene prescribed in the monograph Wool fat (0134).
The total area of any peaks, apart from the solvent peak
2-Methylbutane. C5H12. (Mr 72.2). 1099500. [78-78-4]. and the principal peak, in the chromatogram obtained with
Isopentane. solution B is less than the area of the principal peak in the
Content : minimum 99.5 per cent of C5H12. chromatogram obtained with solution A.
Very flammable colourless liquid.
Methyldopa, racemic. C10H13NO4,1½H2O. (Mr 238.2).
: about 0.621.
1175100.
: about 1.354.
Mixture of equal volumes of (2S)- and (2R)-2-amino-3-(3,4-
bp : about 29 °C. dihydroxyphenyl)-2-methylpropanoic acids.
Water (2.5.12) : maximum 0.02 per cent.
Residue on evaporation : maximum 0.0003 per cent. 3-O-Methyldopamine hydrochloride. C9H14ClNO2.
Absorbance (2.2.25) : maximum 0.30 at 210 nm, 0.07 at (Mr 203.7). 1055600. [1477-68-5]. 4-(2-Aminoethyl)-2-
220 nm, 0.01 at 240 nm and higher wavelengths, determined methoxyphenol hydrochloride.
using water R as compensation liquid. mp : 213 °C to 215 °C.
2-Methylbut-2-ene. C5H10. (Mr 70.1). 1055400. [513-35-9]. 4-O-Methyldopamine hydrochloride. C9H14ClNO2.
Very flammable liquid, practically insoluble in water, miscible (Mr 203.7). 1055700. [645-33-0]. 5-(2-Aminoethyl)-2-
with ethanol (96 per cent). methoxyphenol hydrochloride.
bp : 37.5 °C to 38.5 °C. mp : 207 °C to 208 °C.

Methylenebisacrylamide. C7H10N2O2. (Mr 154.2). 1056000. 1-Methylimidazole. C4H6N2. (Mr 82.1). 1139700. [616-47-7].

[110-26-9]. N,N′-Methylenebispropenamide. 1-Methyl-1H-imidazole.
Fine, white or almost white powder, slightly soluble in water, Colourless or slightly yellowish liquid.
soluble in ethanol (96 per cent). : about 1.495.
mp : 300 °C, with decomposition. bp : 195 °C to 197 °C.
Methylene blue. C16H18ClN3S,xH2O. (Mr 319.9 for the Storage : in an airtight container, protected from light.
anhydrous substance). 1055800. [122965-43-9]. 1-Methylimidazole R1. 1139701.
Schultz No. 1038. Complies with the requirements prescribed for
Colour Index No. 52015. 1-methylimidazole R with the following additional
3,7-Dimethylaminophenothiazin-5-ium chloride. requirement.
It occurs in different hydrated forms and may contain up to Content : minimum 95.0 per cent.
22 per cent of water.
2-Methylimidazole. C4H6N2. (Mr 82.1). 1143400. [693-98-1].
Dark-green or bronze, crystalline powder, freely soluble in
water, soluble in ethanol (96 per cent). White or almost white, crystalline powder.
mp : about 145 °C.
Methylene blue solution. 1055801.
Methyl iodide. CH3I. (Mr 141.9). 1166400. [74-88-4].
Dissolve 3 mg of methylene blue R, 1.2 g of sulfuric acid R Iodomethane.
and 5.0 g of anhydrous sodium sulfate R in 100 mL of
water R. Content : minimum 99.0 per cent.
Methyl isobutyl ketone. C6H12O. (Mr 100.2). 1054300.
Methylene chloride. CH2Cl2. (Mr 84.9). 1055900. [75-09-2]. [108-10-1]. 4-Methyl-2-pentanone.
Dichloromethane.
Clear, colourless liquid, slightly soluble in water, miscible with
Colourless liquid, sparingly soluble in water, miscible with most organic solvents.
: about 0.80.
bp : 39 °C to 42 °C.
bp : about 115 °C.
Methylene chloride used in fluorimetry complies with the Distillation range (2.2.11). Distil 100 mL. The range of
following additional test. temperature of distillation from 1 mL to 95 mL of distillate
Fluorescence. Under irradiation at 365 nm, the fluorescence does not exceed 4.0 °C.
(2.2.21) measured at 460 nm in a 1 cm cell is not more intense Residue on evaporation : maximum 0.01 per cent, determined
than that of a solution containing 0.002 ppm of quinine R in by evaporating on a water-bath and drying at 100-105 °C.
0.5 M sulfuric acid measured in the same conditions.
Methyl isobutyl ketone R1. 1054301.
Methylene chloride, acidified. 1055901. Shake 50 mL of freshly distilled methyl isobutyl ketone R
To 100 mL of methylene chloride R add 10 mL of with 0.5 mL of hydrochloric acid R1 for 1 min. Allow the
hydrochloric acid R, shake, allow to stand and separate the phases to separate and discard the lower phase. Prepare
two layers. Use the lower layer. immediately before use.
Methyl eicosenoate. C21H40O2. (Mr 324.5). 1120500. Methyl isobutyl ketone R3. 1054302.
[2390-09-2]. Methyl (11Z)-eicos-11-enoate. Complies with the requirements for methyl isobutyl
Methyl erucate. C23H44O2. (Mr 352.6). 1146100. [1120-34-9]. ketone R and with the following limits.
Methyl (13Z)-docos-13-enoate. Cr : maximum 0.02 ppm.
: about 0.871. Cu : maximum 0.02 ppm.
: about 1.456. Pb : maximum 0.1 ppm.
Ni : maximum 0.02 ppm.
3-O-Methylestrone. C19H24O2. (Mr 284.4). 1137000. Sn : maximum 0.1 ppm.
[1624-62-0]. 3-Methoxy-1,3,5(10)-estratrien-17-one.
White to yellowish-white powder. Methyl isobutyl ketone, water-saturated. 1054303.
Shake methyl isobutyl ketone R with water R prior to use.
: about + 157.
mp : about 173 °C. Methyl laurate. C13H26O2. (Mr 214.4). 1054400. [111-82-0].
Methyl dodecanoate.
Methyl ethyl ketone. C4H8O. (Mr 72.1). 1054100. [78-93-3]. Content : minimum 98.0 per cent, determined by gas
Ethyl methyl ketone. 2-Butanone. chromatography (2.4.22).
Clear, colourless, flammable liquid, very soluble in water, Colourless or yellow liquid, soluble in ethanol (96 per cent)
miscible with ethanol (96 per cent). and in light petroleum.
: about 0.81. : about 0.87.
bp : 79 °C to 80 °C. : about 1.431.
Methyleugenol. C11H14O2. (Mr 178.2). 1182000. [93-15-2]. mp : about 5 °C.
1,2-Dimethoxy-4-prop-2-enylbenzene. Methyl lignocerate. C25H50O2. (Mr 382.7). 1120600.
Methyleugenol used in gas chromatography complies with [2442-49-1]. Methyl tetracosanoate.
the following additional test. Flakes.
Assay. Gas chromatography (2.2.28) as prescribed in the mp : about 58 °C.
monograph Niaouli oil, cineole type (2468).
Methyl linoleate. C19H34O2. (Mr 294.5). 1120700. [112-63-0].
Content : minimum 97.0 per cent, calculated by the Methyl (9Z,12Z)-octadeca-9,12-dienoate.
: about 0.888.
Methyl 4-hydroxybenzoate. 1055000. [99-76-3]. : about 1.466.
See Methyl parahydroxybenzoate R. bp : 207 °C to 208 °C.

Methyl linolenate. C19H32O2. (Mr 292.5). 1120800. : about 1.437.

[301-00-8]. Methyl (9Z,12Z,15Z)-octadeca-9,12,15-trienoate. mp : about 20 °C.
Methyl α-linolenate.
: about 0.901. Methyl nervonate. 1144800. [2733-88-2].
: about 1.471. See Tetracos-15-enoic acid methyl ester R.
bp : about 207 °C. Methyl oleate. C19H36O2. (Mr 296.4). 1054700. [112-62-9].
Methyl (9Z)-octadec-9-enoate.
Methyl γ-linolenate. C19H32O2. (Mr 292.5). 1158400.
[16326-32-2]. Methyl (6Z,9Z,12Z)-octadeca-6,9,12-trienoate. Content : minimum 98.0 per cent, determined by gas
Content : minimum 99.0 per cent, determined by gas chromatography (2.4.22).
chromatography. Colourless or slightly yellow liquid, soluble in ethanol (96 per
cent) and in light petroleum.
Methyl margarate. C18H36O2. (Mr 284.5). 1120900. : about 0.88.
[1731-92-6]. Methyl heptadecanoate.
: about 1.452.
mp : 32 °C to 34 °C. Methylophiopogonanone A. C19H18O6. (Mr 342.3). 1206500.
Methyl margarate used in the assay of total fatty acids in Saw [74805-92-8]. (3R)-3-[(1,3-Benzodioxol-5-yl)methyl]-2,3-
palmetto fruit (1848) complies with the following additional dihydro-5,7-dihydroxy-6,8-dimethyl-4H-1-benzopyran-4-
test. one.
Assay. Gas chromatography (2.2.28) as prescribed in the Methyl orange. C14H14N3NaO3S. (Mr 327.3). 1054800.
monograph Saw palmetto fruit (1848). [547-58-0].
Content : minimum 97 per cent, calculated by the Schultz No. 176.
normalisation procedure. Colour Index No. 13025.
Methyl methacrylate. C5H8O2. (Mr 100.1). 1054500. Sodium 4′-(dimethylamino)azobenzene-4-sulfonate.
[80-62-6]. Methyl 2-methylprop-2-enoate. Orange-yellow, crystalline powder, slightly soluble in water,
Colourless liquid. practically insoluble in ethanol (96 per cent).
: about 1.414. Methyl orange mixed solution. 1054801.
bp : about 100 °C. Dissolve 20 mg of methyl orange R and 0.1 g of bromocresol
mp : about − 48 °C. green R in 1 mL of 0.2 M sodium hydroxide and dilute to
It contains a suitable stabilising reagent. 100 mL with water R.
Colour change : pH 3.0 (orange) to pH 4.4 (olive-green).
Methyl methanesulfonate. C2H6O3S. (Mr 110.1). 1179500.
[66-27-3]. Methyl orange solution. 1054802.
Clear, colourless or slightly yellow liquid. Dissolve 0.1 g of methyl orange R in 80 mL of water R and
Content : minimum 99.0 per cent. dilute to 100 mL with ethanol (96 per cent) R.
Density : about 1.3 g/cm3 (25 °C). Test for sensitivity. A mixture of 0.1 mL of the methyl
: about 1.414. orange solution and 100 mL of carbon dioxide-free water R
bp : about 202 °C. is yellow. Not more than 0.1 mL of 1 M hydrochloric acid is
required to change the colour to red.
Methyl 2-methoxybenzoate. C9H10O3. (Mr 166.2). 1206300. Colour change : pH 3.0 (red) to pH 4.4 (yellow).
[606-45-1].
Colourless liquid. Methyl palmitate. C17H34O2. (Mr 270.5). 1054900. [112-39-0].
Methyl hexadecanoate.
Methyl 4-methoxybenzoate. C9H10O3. (Mr 166.2). 1206400. Content : minimum 98.0 per cent, determined by gas
[121-98-2]. chromatography (2.4.22).
White or almost white powder. White or yellow, crystalline mass, soluble in ethanol (96 per
Methyl N-methylanthranilate. C9H11NO2. (Mr 165.2). cent) and in light petroleum.
1164600. [85-91-6]. Methyl 2-(methylamino)benzoate. mp : about 30 °C.
Pale yellow liquid. Methyl palmitoleate. C17H32O2. (Mr 268.4). 1121000.
d 420 : about 1.128. [1120-25-8]. Methyl (9Z)-hexadec-9-enoate.
nD20 : about 1.579. : about 0.876.
bp : 255 °C to 258 °C. : about 1.451.
Methyl N-methylanthranilate used in gas chromatography Methyl parahydroxybenzoate. 1055000. [99-76-3].
complies with the following additional test. See Methyl parahydroxybenzoate (0409).
monograph Mandarin oil (2355). Methyl pelargonate. C10H20O2. (Mr 172.3). 1143500.
[1731-84-6]. Methyl nonanoate.
Content : minimum 97 per cent, calculated by the Clear, colourless liquid.
normalisation procedure. d 420 : about 0.873.
Methyl myristate. C15H30O2. (Mr 242.4). 1054600. [124-10-7]. nD20 : about 1.422.
Methyl tetradecanoate. bp : 91 °C to 92 °C.
Content : minimum 98.0 per cent, determined by gas Methyl pelargonate used in the assay of total fatty acids in Saw
chromatography (2.4.22). palmetto fruit (1848) complies with the following additional
Colourless or slightly yellow liquid, soluble in ethanol (96 per test.
cent) and in light petroleum. Assay. Gas chromatography (2.2.28) as prescribed in the
: about 0.87. monograph Saw palmetto fruit (1848).

Content : minimum 98 per cent, calculated by the 2-Methyl-2-propanol. C4H10O. (Mr 74.1). 1056500.
normalisation procedure. [75-65-0]. 1,1-Dimethyl ethyl alcohol. tert-Butyl alcohol.
2-Methylpentane. C6H14. (Mr 86.2). 1180400. [107-83-5]. Clear, colourless liquid or crystalline mass, soluble in water,
Isohexane.
Freezing point (2.2.18) : about 25 °C.
: about 0.653.
bp : about 60.0 °C. between 81 °C and 83 °C.
Colourless, flammable liquid, practically insoluble in water,
miscible with anhydrous ethanol. (15R)-15-Methylprostaglandin F2α. C21H36O5. (Mr 368.5).
1159900. [35864-81-4]. (5Z)-7-[(1R,2R,3R,5S)-3,5-
3-Methylpentan-2-one. C6H12O. (Mr 100.2). 1141100. Dihydroxy-2-[(1E)-(3R)-3-hydroxy-3-methyloct-1-
[565-61-7]. enyl]cyclopentyl]hept-5-enoic acid.
Colourless, flammable liquid. Available as a 10 g/L solution in methyl acetate R.
: about 0.815. Storage : at a temperature below − 15 °C.
: about 1.400.
5-Methylpyridin-2-amine. C6H8N2. (Mr 108.1). 1193500.
bp : about 118 °C [1603-41-4]. 6-Amino-3-picoline.
4-Methylpentan-2-ol. C6H14O. (Mr 102.2). 1114300. White or yellow crystals or crystalline powder.
[108-11-2]. mp : about 76 °C.
Clear, colourless, volatile liquid. 5-Methylpyridin-2(1H)-one. C6H7NO. (Mr 109.1). 1193600.
: about 0.802. [1003-68-5].
: about 1.411. White or almost white powder, soluble in anhydrous ethanol
bp : about 132 °C. and in methanol.
Methylphenyloxazolylbenzene. C26H20N2O2. (Mr 392.5). mp : about 181 °C.
1056200. [3073-87-8]. 1,4-Bis[2-(4-methyl-5-phenyl)- Storage : at a temperature of 2 °C to 8 °C.
oxazolyl]benzene. N-Methylpyrrolidine. C5H11N. (Mr 85.2). 1164700.
Fine, greenish-yellow powder with a blue fluorescence or [120-94-5].
small crystals, soluble in ethanol (96 per cent), sparingly Content : minimum 97.0 per cent.
soluble in xylene.
bp : about 80 °C.
mp : about 233 °C.
Methylphenyloxazolylbenzene used for liquid scintillation is N-Methylpyrrolidone. C5H9NO. (Mr 99.1). 1164800.
of a suitable analytical grade. [872-50-4]. 1-Methylpyrrolidin-2-one.
: about 1.028.
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine. C12H15N.
(Mr 173.3). 1137100. [28289-54-5]. MPTP. bp : about 202 °C.
White or almost white, crystalline powder, slightly soluble in mp : about − 24 °C.
water. Methyl red. C15H15N3O2. (Mr 269.3). 1055100. [493-52-7].
mp : about 41 °C. Schultz No. 250.
Methylpiperazine. C5H12N2. (Mr 100.2). 1056300. [109-01-3]. Colour Index No. 13020.
1-Methylpiperazine. 2-(4-Dimethylamino-phenylazo)benzoic acid.
Colourless liquid, miscible with water and with ethanol Dark-red powder or violet crystals, practically insoluble in
(96 per cent). water, soluble in ethanol (96 per cent).
: about 0.90. Methyl red mixed solution. 1055101.
: about 1.466. Dissolve 0.1 g of methyl red R and 50 mg of methylene
bp : about 138 °C. blue R in 100 mL of ethanol (96 per cent) R.
4-(4-Methylpiperidin-1-yl)pyridine. C11H16N2. (Mr 176.3). Colour change: pH 5.2 (red-violet) to pH 5.6 (green).
1114400. [80965-30-6]. Methyl red solution. 1055102.
Clear liquid. Dissolve 50 mg of methyl red R in a mixture of 1.86 mL
: about 1.565. of 0.1 M sodium hydroxide and 50 mL of ethanol (96 per
cent) R and dilute to 100 mL with water R.
Methylpolysiloxane. 1066800.
Test for sensitivity. To 0.1 mL of the methyl red solution
Polysiloxane substituted with 100 per cent of methyl groups. add 100 mL of carbon dioxide-free water R and 0.05 mL of
Methylprednisolone. C22H30O5. (Mr 374.5). 1193400. 0.02 M hydrochloric acid. The solution is red. Not more
[83-43-2]. 11β,17,21-Trihydroxy-6α-methylpregna-1,4-diene- than 0.1 mL of 0.02 M sodium hydroxide is required to
3,20-dione. change the colour to yellow.
White or almost white, crystalline powder. Colour change : pH 4.4 (red) to pH 6.0 (yellow).
2-Methylpropanol. C4H10O. (Mr 74.1). 1056400. [78-83-1]. Methyl salicylate. 1146200. [119-36-8].

Isobutyl alcohol. 2-Methylpropan-1-ol. See Methyl salicylate (0230)
Clear colourless liquid, soluble in water, miscible with ethanol Methyl stearate. C19H38O2. (Mr 298.5). 1055200. [112-61-8].
(96 per cent). Methyl octadecanoate.
: about 0.80. Content : minimum 98.0 per cent, determined by gas
: 1.397 to 1.399. chromatography (2.4.22).
bp : about 107 °C. White or yellow, crystalline mass, soluble in ethanol (96 per
Distillation range (2.2.11). Not less than 96 per cent distils cent) and in light petroleum.
between 107 °C and 109 °C. mp : about 38 °C.

Methylthymol blue. C37H40N2Na4O13S. (Mr 845). 1158500. Mordant black 11. C20H12N3NaO7S. (Mr 461.4). 1056800.
[1945-77-3]. Tetrasodium 2,2′,2″,2′″-[3H-2,1-benzoxathiol- [1787-61-7].
3-ylidenebis[[6-hydroxy-2-methyl-5-(1-methylethyl)-3,1- Schultz No. 241.
phenylene]methylenenitrilo]]tetraacetate S,S-dioxide. Colour Index No. 14645.
Produces a blue colour with calcium in alkaline solution. Sodium 2-hydroxy-1-[(1-hydroxynaphth-2-yl)azo]-6-
Methylthymol blue mixture. 1158501. nitronaph-thalene-4-sulfonate. Eriochrome black.
A mixture of 1 part of methylthymol blue R and 100 parts Brownish-black powder, soluble in water and in ethanol
of potassium nitrate R. (96 per cent).
Storage : in an airtight container, protected from light.
Methyl toluenesulfonate. C8H10O3S. (Mr 186.2). 1191200.
[80-48-8]. Methyl 4-methylbenzenesulfonate. Methyl tosilate. Mordant black 11 triturate. 1056801.
Content : minimum 97.0 per cent. Mix 1 g of mordant black 11 R with 99 g of sodium
Density : about 1.234 g/mL (25 °C). chloride R.
bp : about 292 °C. Test for sensitivity. Dissolve 50 mg in 100 mL of water R.
The solution is brownish-violet. On addition of 0.3 mL
mp : 25 °C to 28 °C. of dilute ammonia R1 the solution turns blue. On the
N-Methyl-m-toluidine. C8H11N. (Mr 121.2). 1175200. subsequent addition of 0.1 mL of a 10 g/L solution of
[696-44-6]. N,3-Dimethylaniline. N,3-Dimethylbenzenamine. magnesium sulfate R, it turns violet.
Methyl-m-tolylamine. Storage : in an airtight container, protected from light.
Content : minimum 97 per cent. Mordant black 11 triturate R1. 1056802.
Methyl tricosanoate. C24H48O2. (Mr 368.6). 1111500. Mix 1.0 g of mordant black 11 R, 0.4 g of methyl orange R
[2433-97-8]. Tricosanoic acid methyl ester. and 100 g of sodium chloride R.
Content : minimum 99.0 per cent. Morphine hydrochloride. 1056900.
White or almost white crystals, practically insoluble in water, See Morphine hydrochloride (0097).
soluble in hexane.
mp : 55 °C to 56 °C. Morpholine. C4H9NO. (Mr 87.1). 1057000. [110-91-8].
Tetrahydro-1,4-oxazine.
Methyl tridecanoate. C14H28O2. (Mr 228.4). 1121100. Colourless, hygroscopic liquid, flammable, soluble in water
[1731-88-0]. and in ethanol (96 per cent).
Colourless or slightly yellow liquid, soluble in ethanol (96 per : about 1.01.
cent) and in light petroleum.
: about 0.86. between 126 °C and 130 °C.
: about 1.441. Storage : in an airtight container.
mp : about 6 °C.
Morpholine for chromatography. 1057001.
Methyl 3,4,5-trimethoxybenzoate. C11H14O5. (Mr 226.23). Complies with the requirements prescribed for
1177200. [1916-07-0]. morpholine R with the following additional requirement.
N-Methyltrimethylsilyl-trifluoroacetamide. Content : minimum 99.5 per cent.
C6H12F3NOSi. (Mr 199.3). 1129600. [24589-78-4].
2,2,2-Trifluoro-N-methyl-N-(trimethylsilyl)acetamide. 2-[N-Morpholino]ethanesulfonic acid. C6H13NO4S.
(Mr 195.2). 1186500. [4432-31-9]. 2-(Morpholin-4-yl)sulfonic
: about 1.380. acid. MES.
bp : 130 °C to 132 °C. White or almost white, crystalline powder, soluble in water.
Minocycline hydrochloride. 1146300. mp : about 300 °C.
See Minocycline hydrochloride (1030). Murexide. C8H8N6O6,H2O. (Mr 302.2). 1137200.
Molecular sieve. 1056600. 5,5′-Nitrilobis(pyrimidine-2,4,6(1H,3H,5H)-trione)
monoammonium salt.
Molecular sieve composed of sodium aluminosilicate. It is
available as beads or powder with a pore size of 0.4 nm. Brownish-red crystalline powder, sparingly soluble in cold
water, soluble in hot water, practically insoluble in ethanol
When reused, it is recommended that the molecular sieve be
regenerated according to the manufacturer’s instructions. (96 per cent), soluble in solutions of potassium hydroxide or
sodium hydroxide giving a blue colour.
Molecular sieve for chromatography. 1129700.
Myosmine. C9H10N2. (Mr 146.2). 1121200. [532-12-7].
Molecular sieve composed of sodium aluminosilicate. The 3-(4,5-Dihydro-3H-pyrrol-2-yl)pyridine.
pore size is indicated after the name of the reagent in the tests
where it is used. If necessary, the particle size is also indicated. Colourless crystals.
mp : about 45 °C.
Molybdovanadic reagent. 1056700.
β-Myrcene. C10H16. (Mr 136.2). 1114500. [123-35-3].
In a 150 mL beaker, mix 4 g of finely powdered ammonium
molybdate R and 0.1 g of finely powdered ammonium 7-Methyl-3-methylenocta-1,6-diene.
vanadate R. Add 70 mL of water R and grind the particles Oily liquid with a pleasant odour, practically insoluble in
using a glass rod. A clear solution is obtained within a few water, miscible with ethanol (96 per cent), soluble in glacial
minutes. Add 20 mL of nitric acid R and dilute to 100 mL acetic acid. It dissolves in solutions of alkali hydroxides.
with water R. : about 0.794.
Monodocosahexaenoin. C25H38O4. (Mr 402.6). 1143600. : about 1.470.
[124516-13-8]. Monoglyceride of docosahexaenoic β-Myrcene used in gas chromatography complies with the
acid (C22:6). Glycerol monodocosahexaenoate. following additional test.
(all-Z)-Docosa-4,7,10,13,16,19-hexaenoic acid, monoester Assay. Gas chromatography (2.2.28) as prescribed in the
with propane-1,2,3-triol. monograph Peppermint oil (0405).

Test solution. The substance to be examined. Storage : protected from light ; use within 1 week.
Content : minimum 90.0 per cent, calculated by the Naphtharson solution R1. 1121402.
A 1 g/L solution in deionised distilled water R.
Myristic acid. C14H28O2. (Mr 228.4). 1143700. [544-63-8]. Test for sensitivity. To 50 mL of ethanol (96 per cent) R,
Tetradecanoic acid. add 20 mL of water R, 1 mL of dilute sulfuric acid R1 and
Colourless or white or almost white flakes. 1 mL of naphtharson solution R1. Titrate with 0.025 M
mp : about 58.5 °C. barium perchlorate ; the colour changes from orange-yellow
Myristic acid used in the assay of total fatty acids in Saw to orange-pink.
palmetto fruit (1848) complies with the following additional Storage : protected from light ; use within 1 week.
test. α-Naphthol. C10H8O. (Mr 144.2). 1057300. [90-15-3].
Assay. Gas chromatography (2.2.28) as prescribed in the 1-Naphthol.
monograph Saw palmetto fruit (1848).
White or almost white, crystalline powder or colourless or
Content : minimum 97 per cent, calculated by the white or almost white crystals, darkening on exposure to light,
normalisation procedure. slightly soluble in water, freely soluble in ethanol (96 per cent).
Myristicine. C11H12O3. (Mr 192.2). 1099600. [607-91-0]. mp : about 95 °C.
5-Allyl-1-methoxy-2,3-methylenedioxybenzene. Storage : protected from light.
4-Methoxy-6-(prop-2-enyl)-1,3-benzodioxole.
α-Naphthol solution. 1057301.
Oily colourless liquid, practically insoluble in water, slightly
soluble in anhydrous ethanol, miscible with toluene and with Dissolve 0.10 g of α-naphthol R in 3 mL of a 150 g/L
xylene. solution of sodium hydroxide R and dilute to 100 mL with
water R. Prepare immediately before use.
: about 1.144.
: about 1.540. β-Naphthol. C10H8O. (Mr 144.2). 1057400. [135-19-3].
bp : 276 °C to 277 °C. 2-Naphthol.
mp : about 173 °C. White or slightly pink plates or crystals, very slightly soluble
Chromatography. Thin-layer chromatography (2.2.27) in water, very soluble in ethanol (96 per cent).
as prescribed in the monograph Star anise (1153) ; the mp : about 122 °C.
chromatogram shows only one principal spot. Storage : protected from light.
Myristicine used in gas chromatography complies with the β-Naphthol solution. 1057401.
Dissolve 5 g of freshly recrystallised β-naphthol R in 40 mL
Assay. Gas chromatography (2.2.28) as prescribed in the of dilute sodium hydroxide solution R and dilute to 100 mL
monograph Nutmeg oil (1552). with water R. Prepare immediately before use.
normalisation procedure. β-Naphthol solution R1. 1057402.
Storage : protected from light. Dissolve 3.0 mg of β-naphthol R in 50 mL of sulfuric acid R
and dilute to 100.0 mL with the same acid. Use the recently
Myristyl alcohol. C14H30O. (Mr 214.4). 1121300. [112-72-1]. prepared solution.
Tetradecan-1-ol.
: about 0.823. Naphtholbenzein. C27H18O2. (Mr 374.4). 1057600.
[145-50-6]. α-Naphtholbenzein. 4-[(4-Hydroxynaphthalen-1-
mp : 38 °C to 40 °C. yl)(phenyl)methylidene] naphthalen-1(4H)-one.
Myrtillin. C21H21ClO12. (Mr 500.8). 1172300. [6906-38-3]. Brownish-red powder or shiny brownish-black crystals,
Delphinidin 3-O-glucoside chloride. practically insoluble in water, soluble in ethanol (96 per cent)
and in glacial acetic acid.
Naphthalene. C10H8. (Mr 128.2). 1057100. [91-20-3].
White or almost white crystals, practically insoluble in water, Naphtholbenzein solution. 1057601.
soluble in ethanol (96 per cent). A 2 g/L solution of naphtholbenzein R in anhydrous acetic
mp : about 80 °C. acid R.
Naphthalene used for liquid scintillation is of a suitable Test for sensitivity. To 50 mL of glacial acetic acid R add
analytical grade. 0.25 mL of the naphtholbenzein solution. The solution
is brownish-yellow. Not more than 0.05 mL of 0.1 M
2,3-Naphthalenediamine. C10H10N2. (Mr 158.2). perchloric acid is required to change the colour to green.
1199700. [771-97-1]. Naphthalene-2,3-diamine.
2,3-Diaminonaphthalene. Naphthol yellow. C10H5N2NaO5. (Mr 256.2). 1136600.
Brownish-yellow crystalline powder, slightly soluble in ethanol 2,4-Dinitro-1-naphthol, sodium salt.
(96 per cent), practically insoluble in acetone. Orange-yellow powder or crystals, freely soluble in water,
mp : 195 °C to 198 °C. slightly soluble in ethanol (96 per cent).
Naphtharson. C16H11AsN2Na2O10S2. (Mr 576.3). 1121400. Naphthol yellow S. C10H4N2Na2O8S. (Mr 358.2). 1143800.
[3688-92-4]. Thorin. Disodium 4-[(2-arsonophenyl)azo]-3- [846-70-8].
hydroxynaphthalene-2,7-disulfonate. Colour Index No. 10316.
Red powder, soluble in water. 8-Hydroxy-5,7-dinitro-2-naphthalenesulfonic acid disodium
salt. Disodium 5,7-dinitro-8-oxidonaphthalene-2-sulfonate.
Naphtharson solution. 1121401. Yellow or orange-yellow powder, freely soluble in water.
A 0.58 g/L solution of naphtharson R.
Test for sensitivity. To 50 mL of ethanol (96 per cent) R, 1-Naphthylacetic acid. C12H10O2. (Mr 186.2). 1148400.
add 20 mL of water R, 1 mL of dilute sulfuric acid R1 and [86-87-3]. (Naphthalen-1-yl)acetic acid.
1 mL of the naphtharson solution. Titrate with 0.025 M White or yellow crystalline powder, very slightly soluble in
barium perchlorate ; the colour changes from orange-yellow water, freely soluble in acetone.
to orange-pink. mp : about 135 °C.

Naphthylamine. C10H9N. (Mr 143.2). 1057700. [134-32-7]. Test solution. The substance to be examined.
1-Naphthylamine. Content : minimum 93.0 per cent, calculated by the
White or almost white, crystalline powder, turning pink on normalisation procedure.
exposure to light and air, slightly soluble in water, freely
soluble in ethanol (96 per cent). Nickel-aluminium alloy. 1058100.
mp : about 51 °C. Contains 48 per cent to 52 per cent of aluminium (Al ; Ar 26.98)
and 48 per cent to 52 per cent of nickel (Ni ; Ar 58.70).
Storage : protected from light. Before use, reduce to a fine powder (180) (2.9.12).
Naphthylethylenediamine dihydrochloride. It is practically insoluble in water and soluble in mineral acids.
C12H16Cl2N2. (Mr 259.2). 1057800. [1465-25-4].
N-(1-Naphthyl)ethylene-diamine dihydrochloride. Nickel-aluminium alloy (halogen-free). 1118100.
Contains 48 per cent to 52 per cent of aluminium (Al ; Ar 26.98)
It may contain methanol of crystallisation. and 48 per cent to 52 per cent of nickel (Ni ; Ar 58.71).
White or yellowish-white powder, soluble in water, slightly Fine, grey powder, practically insoluble in water, soluble in
soluble in ethanol (96 per cent). mineral acids with formation of salts.
Chlorides : maximum 10 ppm.
Naphthylethylenediamine dihydrochloride solution.
1057801. Dissolve 0.400 g in 40 mL of a mixture of 67 volumes
of sulfuric acid R and 33 volumes of dilute nitric acid R.
Dissolve 0.1 g of naphthylethylenediamine dihydrochloride R Evaporate the solution nearly to dryness, dissolve the residue
in water R and dilute to 100 mL with the same solvent. in water R and dilute to 20.0 mL with the same solvent. To
Prepare immediately before use. one half-aliquot of the solution, add 1.0 mL of 0.1 M silver
nitrate. Filter after 15 min and add 0.2 mL of sodium chloride
Naringin. C27H32O14. (Mr 580.5). 1137300. [10236-47-2]. solution (containing 10 µg of chlorides per millilitre) to the
7-[[2-O-(6-Deoxy-α-L-mannopyranosyl)-β-D- filtrate. After 5 min the solution is more opalescent than a
glucopyranosyl]oxy]-5-hydroxy-2-(4-hydroxyphenyl)- mixture of the second half-aliquot of the solution with 1.0 mL
2,3-dihydro-4H-chromen-4-one. of 0.1 M silver nitrate.
White or almost white crystalline powder, slightly soluble in
water, soluble in methanol and in dimethylformamide. Nickel chloride. NiCl2. (Mr 129.6). 1057900. [7718-54-9].
Nickel chloride, anhydrous.
mp : about 171 °C.
Yellow, crystalline powder, very soluble in water, soluble in
Absorbance (2.2.25). Naringin dissolved in a 5 g/L solution ethanol (96 per cent). It sublimes in the absence of air and
of dimethylformamide R in methanol R shows an absorption readily absorbs ammonia. The aqueous solution is acid.
maximum at 283 nm.
Nickel nitrate hexahydrate. Ni(NO3)2,6H2O. (Mr 290.8).
Neohesperidin. C28H34O15. (Mr 610.6). 1182200. 1175300. [13478-00-7].
[13241-33-3]. Hesperetin-7-neohesperidoside.
(2S)-7-[[2-O-(6-Deoxy-α-L-mannopyranosyl)-β-D- Nickel sulfate. NiSO4,7H2O. (Mr 280.9). 1058000.
glucopyranosyl]oxy]-5-hydroxy-2-(3-hydroxy-4- [10101-98-1]. Nickel sulfate heptahydrate.
methoxyphenyl)-2,3-dihydro-4H-1-benzopyran-4-one. Green, crystalline powder or crystals, freely soluble in water,
slightly soluble in ethanol (96 per cent).
trans-Nerolidol. C15H26O. (Mr 222.4). 1107900. [40716-66-3].
3,7,11-Trimethyldodeca-1,6,10-trien-3-ol. Nicotinamide-adenine dinucleotide. C21H27N7O14P2.
Slightly yellow liquid, slight odour of lily and lily of the valley, (Mr 663). 1108100. [-84-9]. NAD .
+
practically insoluble in water and in glycerol, miscible with White or almost white powder, very hygroscopic, freely
ethanol (96 per cent). soluble in water.
: about 0.876. Nicotinamide-adenine dinucleotide solution. 1108101.
: about 1.479. Dissolve 40 mg of nicotinamide-adenine dinucleotide R in
bp12 : 145 °C to 146 °C. water R and dilute to 10 mL with the same solvent. Prepare
trans-Nerolidol used in gas chromatography complies with the
following additional test. Nicotinic acid. 1158600. [59-67-6].
Assay. Gas chromatography (2.2.28) as prescribed in the See Nicotinic acid (0459).
monograph Bitter-orange-flower oil (1175).
Nicotinoyl hydrazide. C6H7N3O. (Mr 137.1). 1202400.
Test solution. The substance to be examined. [553-53-7]. Pyridine-3-carbohydrazide.
Content : minimum 90.0 per cent, calculated by the White or almost white powder or crystalline powder, soluble
normalisation procedure. in water.
mp : about 160 °C.
Neryl acetate. C12H20O2. (Mr 196.3). 1108000. [141-12-8].
(Z)-3,7-Dimethylocta-2,6-dienyl acetate. Nile blue A. C20H21N3O5S. (Mr 415.5). 1058200. [3625-57-8].
Colourless, oily liquid. Schultz No. 1029.
: about 0.907. Colour Index No. 51180.
: about 1.460. 5-Amino-9-(diethylamino)benzo[a]phenoxazinylium
hydrogen sulfate.
bp25 : 134 °C.
Green, crystalline powder with a bronze lustre, sparingly
Neryl acetate used in gas chromatography complies with the soluble in ethanol (96 per cent), in glacial acetic acid and in
following additional test. pyridine.
Assay. Gas chromatography (2.2.28) as prescribed in the Absorbance (2.2.25). A 0.005 g/L solution in ethanol (50 per
monograph Bitter-orange-flower oil (1175). cent V/V) R shows an absorption maximum at 640 nm.

Nile blue A solution. 1058201. Iron (2.4.9) : maximum 1 ppm.

A 10 g/L solution of Nile blue A R in anhydrous acetic Dissolve the residue from the determination of sulfated ash in
acid R. 1 mL of dilute hydrochloric acid R and dilute to 50 mL with
Test for sensitivity. To 50 mL of anhydrous acetic acid R water R. Dilute 5 mL of this solution to 10 mL with water R.
add 0.25 mL of the Nile blue A solution. The solution is Heavy metals (2.4.8) : maximum 2 ppm.
blue. On the addition of 0.1 mL of 0.1 M perchloric acid, Dilute 10 mL of the solution prepared for the limit test for
the colour changes to blue-green. iron to 20 mL with water R. 12 mL of the solution complies
Colour change : pH 9.0 (blue) to pH 13.0 (red). with test A. Prepare the reference solution using lead standard
solution (2 ppm Pb) R.
Ninhydrin. C9H4O3,H2O. (Mr 178.1). 1058300. [485-47-2].
1,2,3-Indanetrione monohydrate. Sulfated ash : maximum 0.001 per cent.
White or very pale yellow, crystalline powder, soluble in water Carefully evaporate 100 g to dryness. Moisten the residue with
and in ethanol (96 per cent). a few drops of sulfuric acid R and heat to dull red.
Storage : protected from light. Assay. To 1.50 g add about 50 mL of water R and titrate with
1 M sodium hydroxide, using 0.1 mL of methyl red solution R
Ninhydrin and stannous chloride reagent. 1058301. as indicator.
Dissolve 0.2 g of ninhydrin R in 4 mL of hot water R, add 1 mL of 1 M sodium hydroxide is equivalent to 63.0 mg of
5 mL of a 1.6 g/L solution of stannous chloride R, allow to HNO3.
stand for 30 min, then filter and store at a temperature of Storage : protected from light.
2 °C to 8 °C. Immediately before use dilute 2.5 mL of the
solution with 5 mL of water R and 45 mL of 2-propanol R. Nitric acid, cadmium- and lead-free. 1058401.
Complies with the requirements prescribed for nitric acid R
Ninhydrin solution. 1058303. and with the following additional test.
A 2 g/L solution of Ninhydrin R in a mixture of 5 volumes Test solution. To 100 g add 0.1 g of anhydrous sodium
of dilute acetic acid R and 95 volumes of butanol R. carbonate R and evaporate to dryness. Dissolve the residue
Ninhydrin solution R1. 1058304. in water R heating slightly, and dilute to 50.0 mL with the
same solvent.
Dissolve 1.0 g of ninhydrin R in 50 mL of ethanol (96 per
cent) R and add 10 mL of glacial acetic acid R. Cadmium : maximum 0.1 ppm.
Atomic absorption spectrometry (2.2.23, Method II).
Ninhydrin solution R2. 1058305.
Source : cadmium hollow-cathode lamp.
Dissolve 3 g of ninhydrin R in 100 mL of a 45.5 g/L solution
Wavelength : 228.8 nm.
of sodium metabisulfite R.
Atomisation device : air-acetylene or air-propane flame.
Ninhydrin solution R3. 1058306. Lead : maximum 0.1 ppm.
A 4 g/L solution in a mixture of 5 volumes of anhydrous Atomic absorption spectrometry (2.2.23, Method II).
acetic acid R and 95 volumes of butanol R. Source : lead hollow-cathode lamp.
Ninhydrin solution R4. 1058307. Wavelength : 283.3 nm or 217.0 nm.
A 3 g/L solution of ninhydrin R in a mixture of 5 volumes Atomisation device : air-acetylene flame.
of glacial acetic acid R and 95 volumes of 2-propanol R.
Nitric acid, dilute. 1058402.
Nitrazepam. 1143900. [146-22-5]. Contains about 125 g/L of HNO3 (Mr 63.0).
See Nitrazepam (0415). Dilute 20 g of nitric acid R to 100 mL with water R.
Nitric acid. HNO3. (Mr 63.0). 1058400. [7697-37-2]. Nitric acid, dilute R1. 1058407.
Content : 63.0 per cent m/m to 70.0 per cent m/m. Dilute 40 g of nitric acid R to 100 mL with water R.
Clear, colourless or almost colourless liquid, miscible with
water. Nitric acid, dilute R2. 1058409.
: 1.384 to 1.416. Dilute 30 g of nitric acid R to 100 mL with water R.
A 10 g/L solution is strongly acid and gives the reaction of Nitric acid, dilute, heavy metal-free. 1058410.
nitrates (2.3.1). Complies with the requirements prescribed for dilute nitric
Appearance. Nitric acid is clear (2.2.1) and not more intensely acid R with the following maximum contents of heavy
coloured than reference solution Y6 (2.2.2, Method II). metals.
Chlorides (2.4.4) : maximum 0.5 ppm. As : 0.005 ppm.
To 5 g add 10 mL of water R and 0.3 mL of silver nitrate Cd : 0.005 ppm.
solution R2 and allow to stand for 2 min protected from light. Cu : 0.001 ppm.
Any opalescence is not more intense than that of a standard Fe : 0.02 ppm.
prepared in the same manner using 13 mL of water R, 0.5 mL
Hg : 0.002 ppm.
of nitric acid R, 0.5 mL of chloride standard solution (5 ppm
Cl) R and 0.3 mL of silver nitrate solution R2. Ni : 0.005 ppm.
Sulfates (2.4.13) : maximum 2 ppm. Pb : 0.001 ppm.
Evaporate 10 g to dryness with 0.2 g of sodium carbonate R. Zn : 0.01 ppm.
Dissolve the residue in 15 mL of distilled water R. Prepare the Nitric acid, heavy metal-free. 1058404.
standard using a mixture of 2 mL of sulfate standard solution Complies with the requirements prescribed for nitric acid R
(10 ppm SO4) R and 13 mL of distilled water R. with the following maximum contents of heavy metals.
Arsenic (2.4.2, Method A) : maximum 0.02 ppm. As : 0.005 ppm.
Gently heat 50 g with 0.5 mL of sulfuric acid R until white Cd : 0.005 ppm.
fumes begin to evolve. To the residue add 1 mL of a 100 g/L
solution of hydroxylamine hydrochloride R and dilute to 2 mL Cu : 0.001 ppm.
with water R. Prepare the standard using 1.0 mL of arsenic Fe : 0.02 ppm.
standard solution (1 ppm As) R. Hg : 0.002 ppm.

Ni : 0.005 ppm. 4-Nitrobenzaldehyde. C7H5NO3. (Mr 151.1). 1198700.

Pb : 0.001 ppm. [555-16-8].
Zn : 0.01 ppm. Nitrobenzene. C6H5NO2. (Mr 123.1). 1058800. [98-95-3].
Colourless or very slightly yellow liquid, practically insoluble
Nitric acid, lead-free. 1058403. in water, miscible with ethanol (96 per cent).
Complies with the requirements prescribed for Nitric bp : about 211 °C.
acid R with the following additional test. Dinitrobenzene. To 0.1 mL add 5 mL of acetone R, 5 mL of
Lead : maximum 0.1 ppm. water R and 5 mL of strong sodium hydroxide solution R. Shake
and allow to stand. The upper layer is almost colourless.
Atomic absorption spectrometry (2.2.23, Method II).
4-Nitrobenzoic acid. C7H5NO4. (Mr 167.1). 1144000.
Test solution. To 100 g add 0.1 g of anhydrous sodium
[62-23-7].
carbonate R and evaporate to dryness. Dissolve the residue
in water R, heating slightly, and dilute to 50.0 mL with the Yellow crystals.
same solvent. mp : about 240 °C.
Source : lead hollow-cathode lamp. Nitrobenzoyl chloride. C7H4ClNO3. (Mr 185.6). 1058900.
Wavelength : 283.3 nm or 217.0 nm. [122-04-3]. 4-Nitrobenzoyl chloride.
Atomisation device : air-acetylene flame. Yellow crystals or a crystalline mass, decomposing in moist
air, completely soluble in sodium hydroxide solution giving a
Nitric acid, lead-free R1. 1058405. yellowish-orange colour.
mp : about 72 °C.
Nitric acid R containing not more than 1 μg/kg of lead.
Nitrobenzyl chloride. C7H6ClNO2. (Mr 171.6). 1059000.
Nitric acid, lead-free, dilute. 1058406. [100-14-1]. 4-Nitrobenzyl chloride.
Dilute 5 g of lead-free nitric acid R1 to 100 mL with Pale-yellow crystals, lachrymatory, practically insoluble in
deionised distilled water R. water, very soluble in ethanol (96 per cent).
Nitric acid, nickel-free. 1058408. 4-(4-Nitrobenzyl)pyridine. C12H10N2O2. (Mr 214.2).

1101900. [1083-48-3].
Complies with the requirements prescribed for nitric acid R Yellow powder.
with the following additional requirement.
mp : about 70 °C.
Nickel : maximum 0.005 ppm.
Nitroethane. C2H5NO2. (Mr 75.1). 1059200. [79-24-3].
Nitric acid, fuming. 1058500. [7697-37-2]. Clear, oily, colourless liquid.
Clear, slightly yellowish liquid, fuming on contact with air. bp : about 114 °C.
: about 1.5. Nitrofurantoin. 1099700. [67-20-9].
See Nitrofurantoin (0101).
Nitrilotriacetic acid. C6H9NO6. (Mr 191.1). 1137400.
[139-13-9]. Nitrogen. N2. (Mr 28.01). 1059300. [7727-37-9].
White or almost white crystalline powder, practically insoluble Nitrogen, washed and dried.
in water and in most organic solvents. Nitrogen gas mixture. 1136900.
mp : about 240 °C, with decomposition. Nitrogen R containing 1 per cent V/V of each of the
following gases : carbon dioxide R2, carbon monoxide R1
Nitroaniline. C6H6N2O2. (Mr 138.1). 1058600. [100-01-6]. and oxygen R1.
4-Nitroaniline.
Nitrogen, oxygen-free. 1059600.
Bright yellow, crystalline powder, very slightly soluble in water,
sparingly soluble in boiling water, soluble in ethanol (96 per Nitrogen R which has been freed from oxygen by passing it
cent), forms water-soluble salts with strong mineral acids. through alkaline pyrogallol solution R.
mp : about 147 °C. Nitrogen R1. N2. (Mr 28.01). 1059400. [7727-37-9].
Nitrobenzaldehyde. C7H5NO3. (Mr 151.1). 1058700. Carbon monoxide : less than 5 ppm.
[552-89-6]. 2-Nitrobenzaldehyde.
Oxygen : less than 5 ppm.
Yellow needles, slightly soluble in water, freely soluble in
ethanol (96 per cent), volatile in steam. Nitrogen dioxide. NO2. (Mr 46.01). 1179600. [10102-44-0].
mp : about 42 °C. Content : minimum 98.0 per cent V/V.
Nitrogen for chromatography. N2. (Mr 28.01). 1059500.
Nitrobenzaldehyde paper. 1058701. [7727-37-9].
Dissolve 0.2 g of nitrobenzaldehyde R in 10 mL of a 200 g/L Content : minimum 99.95 per cent V/V.
solution of sodium hydroxide R. Use the solution within
1 h. Immerse the lower half of a slow filter paper strip Nitrogen monoxide. NO. (Mr 30.01). 1108300.
10 cm long and 0.8-1 cm wide. Absorb the excess reagent Content : minimum 98.0 per cent V/V.
between two sheets of filter paper. Use within a few minutes
Nitromethane. CH3NO2. (Mr 61.0). 1059700. [75-52-5].
of preparation.
Clear, colourless, oily liquid, slightly soluble in water, miscible
Nitrobenzaldehyde solution. 1058702. with ethanol (96 per cent).
Add 0.12 g of powdered nitrobenzaldehyde R to 10 mL : 1.132 to 1.134.
of dilute sodium hydroxide solution R ; allow to stand for : 1.381 to 1.383.
10 min shaking frequently and filter. Prepare immediately Distillation range (2.2.11). Not less than 95 per cent distils
before use. between 100 °C and 103 °C.

Nitro-molybdovanadic reagent. 1060100. Nonylamine. C9H21N. (Mr 143.3). 1139800. [112-20-9].

Solution A. Dissolve 10 g of ammonium molybdate R in Nonan-1-amine. 1-Aminononane.
water R, add 1 mL of ammonia R and dilute to 100 mL with Corrosive, colourless, clear liquid.
water R. : about 0.788.
Solution B. Dissolve 2.5 g of ammonium vanadate R in hot : about 1.433.
water R, add 14 mL of nitric acid R and dilute to 500 mL with
water R. Nordazepam. C15H11ClN2O. (Mr 270.7). 1060200.
To 96 mL of nitric acid R add 100 mL of solution A and [1088-11-5]. 7-Chloro-2,3-dihydro-5-phenyl-1H-1,4-
100 mL of solution B and dilute to 500 mL with water R. benzodiazepin-2-one.
White or pale yellow, crystalline powder, practically insoluble
4-Nitrophenol. C6H5NO3. (Mr 139.1). 1146400. [100-02-7]. in water, slightly soluble in ethanol (96 per cent).
p-Nitrophenol. mp : about 216 °C.
Colourless or slightly yellow powder, sparingly soluble in DL-Norleucine. C6H13NO2. (Mr 131.2). 1060300. [616-06-8].
water and in methanol. (RS)-2-Aminohexanoic acid.
mp : about 114 °C. Shiny crystals, sparingly soluble in water and in ethanol
(96 per cent), soluble in acids.
3-Nitrosalicylic acid. C7H5NO5. (Mr 183.1). 1184300.
Noscapine hydrochloride. 1060500. [912-60-7].
[85-38-1]. 2-Hydroxy-3-nitrobenzoic acid.
See Noscapine hydrochloride (0515).
Yellowish crystals, slightly soluble in water, freely soluble in
ethanol (96 per cent). Ochratoxin A solution. 1175700.
mp : 142 °C to 147 °C. 50 μg/mL solution of (2S)-2-([[(3R)-5-chloro-8-hydroxy-
3-methyl-1-oxo-3,4-dihydro-1H-2-benzopyran-7-
N-Nitrosodiethanolamine. C4H10N2O3. (Mr 134.1). 1129800. yl]carbonyl]amino)-3-phenylpropanoic acid (ochratoxin A)
[1116-54-7]. 2,2′-(Nitrosoimino)diethanol. in a mixture of 1 volume of acetic acid R and 99 volumes of
Yellow liquid, miscible with anhydrous ethanol. benzene R.
: about 1.485.
Octadecyl [3-[3,5-bis(1,1-dimethylethyl)-4-
bp : about 125 °C. hydroxyphenyl]-propionate]. C35H62O3. (Mr 530.9).
N-Nitrosodiisopropanolamine. C6H14N2O3. (Mr 162.2). 1060600. [2082-79-3]. Octadecyl 3-(3,5-di-tert-butyl-4-
1176500. [53609-64-6]. 1,1′-(Nitrosoimino)bispropan-2-ol. hydroxyphenyl)propionate.
bp : 122-124 °C. White or slightly yellowish, crystalline powder, practically
insoluble in water, very soluble in acetone and in hexane,
Nitrosodipropylamine. C6H14N2O. (Mr 130.2). 1099900. slightly soluble in methanol.
[621-64-7]. Dipropylnitrosamine. mp : 49 °C to 55 °C.
Liquid, soluble in anhydrous ethanol and in strong acids.
Octanal. C8H16O. (Mr 128.2). 1150400. [124-13-0]. Octyl
: about 0.915. aldehyde.
bp : about 78 °C. Oily, colourless liquid. Practically insoluble in water.
Appropriate grade for chemiluminescence determination. Octanal used in gas chromatography complies with the
Nitrosodipropylamine solution. 1099901. following additional test.
Inject 78.62 g of anhydrous ethanol R through the septum Assay. Gas chromatography (2.2.28) as prescribed in the
of a vial containing nitrosodipropylamine R. Dilute 1/100 monograph Sweet orange oil (1811).
in anhydrous ethanol R and place 0.5 mL aliquots in Content : minimum 99 per cent, calculated by the
crimp-sealed vials. normalisation procedure.
Storage : in the dark at 5 °C. Octane. C8H18. (Mr 114.2). 1166500. [111-65-9]. n-Octane.
Nitrotetrazolium blue. C40H30Cl2N10O6. (Mr 818). 1060000. Content : minimum 99 per cent.
[298-83-9]. 3,3′-(3,3′-Dimethoxy-4,4′-diphenylene)di[2- Octanol. C8H18O. (Mr 130.2). 1060700. [111-87-5].
(4-nitrophenyl)-5-phenyl-2H-tetrazolium] dichloride. Octan-1-ol. Caprylic alcohol.
p-Nitro-tetrazolium blue.
Crystals, soluble in methanol, giving a clear, yellow solution. ethanol (96 per cent).
mp : about 189 °C, with decomposition. : about 0.828.
Nitrous oxide. N2O. (Mr 44.01). 1108500. bp : about 195 °C.
Content : minimum 99.99 per cent V/V. 3-Octanone. C8H16O. (Mr 128.2). 1114600. [106-68-3].
Nitrogen monoxide : less than 1 ppm. Octan-3-one. Ethylpentylketone.
Carbon monoxide : less than 1 ppm. Colourless liquid with a characteristic odour.
Nonivamide. C17H27NO3. (Mr 293.4). 1148500. [2444-46-4]. : about 0.822.
N-[(4-Hydroxy-3-methoxyphenyl)methyl]nonanamide. : about 1.415.
White or almost white, crystalline powder, practically bp : about 167 °C.
insoluble in cold water, freely soluble in anhydrous ethanol. 3-Octanone used in gas chromatography complies with the
Nonivamide used in the test for nonivamide in the monograph following additional test.
Capsicum (1859) complies with the following additional test. Assay. Gas chromatography (2.2.28) as prescribed in the
Assay. Liquid chromatography (2.2.29) as prescribed in the monograph Lavender oil (1338).
monograph Capsicum (1859). Test solution. The substance to be examined.
Content : minimum 98.0 per cent, calculated by the Content : minimum 98.0 per cent, calculated by the
normalisation procedure. normalisation procedure.

Octoxinol 10. C34H62O11 (average). (Mr 647). 1060800. Orcinol. C7H8O2,H2O. (Mr 142.2). 1108700. [6153-39-5].
[9002-93-1]. α-[4-(1,1,3,3-Tetramethylbutyl)phenyl]-ω- 5-Methylbenzene-1,3-diol monohydrate.
hydroxypoly-(oxyethylene). Crystalline powder, sensitive to light.
Clear, pale-yellow, viscous liquid, miscible with water, with bp : about 290 °C.
acetone and with ethanol (96 per cent), soluble in toluene. mp : 58 °C to 61 °C.
Organosilica polymer, amorphous, octadecylsilyl. 1144200.
Octreotide acetate. C49H66N10O10S2,xC2H4O2. 1182900. Synthetic, spherical hybrid particles, containing both
[79517-01-4]. (Acetate-free peptide : Mr 1019. inorganic (silica) and organic (organosiloxanes) components,
[83150-76-9]). D-Phenylalanyl-L-cysteinyl-L-phenylalanyl- chemically modified at the surface by trifunctionally bonded
D-tryptophyl-L-lysyl-L-threonyl-N-[(1R,2R)-2-hydroxy-1- octadecylsilyl groups.
(hydroxymethyl)propyl]-L-cysteinamide cyclic (2→7)-disulfide
acetate. It contains a variable quantity of acetic acid. Organosilica polymer, amorphous, polar-embedded
White or almost white powder, freely soluble in water and octadecylsilyl, end-capped. 1150600.
acetic acid. Synthetic, spherical hybrid particles containing both inorganic
Content : minimum 96.0 per cent. (silica) and organic (organosiloxanes) components, chemically
modified at the surface by the bonding of polar-embedded
Octylamine. C8H19N. (Mr 129.2). 1150500. [111-86-4]. octadecylsilyl groups. To minimise any interaction with basic
Octan-1-amine. compounds, they are carefully end-capped to cover most of
the remaining silanol groups.
Colourless liquid.
: about 0.782. Organosilica polymer, amorphous, propyl-2-phenylsilyl,
end-capped. 1178100.
bp : 175 °C to 179 °C.
Synthetic, spherical hybrid particles containing both inorganic
Oleamide. C18H35NO. (Mr 281.5). 1060900. (silica) and organic (organosiloxanes) components, chemically
(9Z)-Octadec-9-enoamide. modified at the surface by the bonding of propyl-2-phenylsilyl
Yellowish or white powder or granules, practically insoluble groups. To minimise any interaction with basic compounds,
in water, very soluble in methylene chloride, soluble in they are carefully end-capped to cover most of the remaining
anhydrous ethanol. silanol groups.
mp : about 80 °C. Organosilica polymer compatible with 100 per cent
aqueous mobile phases, octadecylsilyl, solid core,
Oleanolic acid. C30H48O3. (Mr 456.7). 1183000. [508-02-1]. end-capped. 1201700.
3β-Hydroxyolean-12-en-28-oic acid. Astrantiagenin C. Silica gel with spherical silica particles containing a solid
non-porous silica core surrounded by a thin outer organosilica
Oleic acid. C18H34O2. (Mr 282.5). 1144100. [112-80-1].
polymer coating with octadecylsilyl groups, suitable for use
(9Z)-Octadec-9-enoic acid.
with highly aqueous mobile phases including 100 per cent
Clear, colourless liquid, practically insoluble in water. aqueous phases. To minimise any interaction with basic
: about 0.891. compounds, it is carefully end-capped to cover most of the
: about 1.459. remaining silanol groups.
mp : 13 °C to 14 °C. Organosilica polymer for chromatography, amorphous,
Oleic acid used in the assay of total fatty acids in the monograph octadecylsilyl, end-capped. 1164900.
Saw palmetto fruit (1848) complies with the following Synthetic, spherical hybrid particles containing both inorganic
additional test. (silica) and organic (organosiloxanes) components, chemically
modified at the surface by the bonding of octadecylsilyl
Assay. Gas chromatography (2.2.28) as prescribed in the groups. To minimise any interaction with basic compounds,
monograph Saw palmetto fruit (1848). they are carefully end-capped to cover most of the remaining
Content : minimum 98 per cent, calculated by the silanol groups.
Organosilica polymer, multi-layered, octadecylsilyl,
Oleuropein. C25H32O13. (Mr 540.5). 1152900. [32619-42-4]. end-capped. 1202500.
2-(3,4-Dihydroxyphenyl)ethyl[(2S,3E,4S)-3-ethylidene-2-(β- Synthetic, spherical hybrid particles, multi-layered, containing
D-glucopyranosyloxy)-5-(methoxycarbonyl)-3,4-dihydro-2H- both inorganic (silica) and organic (organosiloxanes)
pyran-4-yl]acetate. components, chemically modified at the surface by the
Powder, soluble in methanol. bonding of octadecylsilyl groups. To minimise any interaction
Oleuropein used in Olive leaf (1878) complies with the following with basic compounds, they are carefully end-capped to cover
test. most of the remaining silanol groups.
Assay. Liquid chromatography (2.2.29) as prescribed in the Osthole. C15H16O3. (Mr 244.3). 1180500. [484-12-8].
monograph Olive leaf (1878). 7-Methoxy-8-(3-methylbut-2-enyl)-2H-1-benzopyran-2-one.
Content : minimum 80 per cent, calculated by the 7-Methoxy-8-isopentenylcoumarin.
normalisation procedure. Oxalic acid. C2H2O4,2H2O. (Mr 126.1). 1061400. [6153-56-6].
Ethanedioic acid dihydrate.
Oleyl alcohol. C18H36O. (Mr 268.5). 1156000. [143-28-2].
(9Z)-Octadec-9-en-1-ol. White or almost white crystals, soluble in water, freely soluble
bp : about 207 °C.
Oxalic acid and sulfuric acid solution. 1061401.
nD20 : 1.460.
A 50 g/L solution of oxalic acid R in a cooled mixture of
Content : minimum 85 per cent. equal volumes of sulfuric acid R and water R.
Olive oil. 1061000. [8001-25-0]. Oxazepam. 1144300. [604-75-1].
See Olive oil, virgin (0518). See Oxazepam (0778).

Ox brain, acetone-dried. 1061300. Palmitoleic acid. C16H30O2. (Mr 254.4). 1144400. [373-49-9].

Cut into small pieces a fresh ox brain previously freed (9Z)-Hexadec-9-enoic acid.
from vascular and connective tissue. Place in acetone R Clear, colourless liquid.
for preliminary dehydration. Complete the dehydration by bp : about 162 °C.
pounding in a mortar 30 g of this material with successive
quantities, each of 75 mL, of acetone R until a dry powder Palmitoleic acid used in the assay of total fatty acids in Saw
is obtained after filtration. Dry at 37 °C for 2 h or until the palmetto fruit (1848) complies with the following additional
odour of acetone is no longer present. test.
2,2′-Oxybis(N,N-dimethylethylamine). C8H20N2O. monograph Saw palmetto fruit (1848).
(Mr 160.3). 1141200. [3033-62-3]. Bis(2-dimethylaminoethyl)
ether. Content : minimum 98 per cent, calculated by the
Colourless, corrosive liquid.
: about 0.85. Palmityl alcohol. C16H34O. (Mr 242.4). 1156100.
: about 1.430. [36653-82-4]. Hexadecan-1-ol. Cetyl alcohol.
mp : about 48 °C.
Oxygen. O2. (Mr 32.00). 1108800.
Nitrogen and argon : less than 100 ppm. Pancreas powder. 1061700.
Carbon dioxide : less than 10 ppm. See Pancreas powder (0350).
Carbon monoxide : less than 5 ppm.
Papain. 1150700. [9001-73-4].
Oxygen R1. O2. (Mr 32.00). 1137600. A proteolytic enzyme obtained from the latex of the green
Content : minimum 99 per cent V/V. fruit and leaves of Carica papaya L.
Oxytetracycline hydrochloride. 1146500. Papaverine hydrochloride. 1061800. [61-25-6].
See Oxytetracycline hydrochloride (0198). See Papaverine hydrochloride (0102).
Paeoniflorin. C23H28O11. (Mr 480.5). 1197300. Paper chromatography performance test solutions.
[23180-57-6]. [(1R,2S,3R,5R,6R,8S)-3-(β-D- 1150800.
Glucopyranosyloxy)6-hydroxy-8-methyl-9,10-dioxate- Test solution (A). Sodium pertechnetate (99mTc) injection
tracyclo[4.3.1.02.5.03.8]decan-2-yl]methyl benzoate. (fission) (0124) or Sodium pertechnetate (99mTc) injection
Paeonol. C9H10O3. (Mr 166.2). 1197400. [552-41-0]. (non-fission) (0283).
1-(2-Hydroxy-4-methoxyphenyl)ethan-1-one. Test solution (B). In a closed vial mix 100 µL of a 5 g/L
2′-Hydroxy-4′-methoxyacetophenone. solution of stannous chloride R in 0.05 M hydrochloric acid
and 100 MBq to 200 MBq of Sodium pertechnetate (99mTc)
Palladium. Pd. (Ar 106.4). 1114700. [7440-05-3]. injection (fission) (0124) or Sodium pertechnetate (99mTc)
Grey white metal, soluble in hydrochloric acid. injection (non-fission) (0283) in a volume not exceeding 2 mL.
Palladium chloride. PdCl2. (Mr 177.3). 1061500. [7647-10-1]. Paper for chromatography. 1150900.
Red crystals. Pure cellulose grade thin paper with a smooth surface and
mp : 678 °C to 680 °C. a thickness of about 0.2 mm.
Palladium chloride solution. 1061501. Chromatographic separation. To 2 strips of paper for
Dissolve 1 g of palladium chloride R in 10 mL of warm chromatography R apply separately 2-5 μL of test solution (a)
and test solution (b) of paper chromatography performance
hydrochloric acid R. Dilute the solution to 250 mL with a
mixture of equal volumes of dilute hydrochloric acid R and test solutions R. Develop over a pathlength of 3/4 of the paper
water R. Dilute this solution immediately before use with height, using a mixture of equal volumes of methanol R
and water R. Allow to dry and determine the distribution
2 volumes of water R.
of radioactivity using a suitable detector. The paper is not
Palmatine. C21H22NO4+. (Mr 352.4). 1198800. [3486-67-7]. satisfactory, unless the chromatogram obtained with test
2,3,9,10-Tetramethoxy-5,6-dihydro-7λ5-isoquinolino- solution (a) shows a single radioactivity spot with an RF value
[3,2-a]isoquinolin-7-ylium. 7,8,13,13a-Tetradehydro-2,3,9,10- in the range 0.8-1.0 and the chromatogram obtained with test
tetramethoxyberbinium. solution (b) shows a single radioactivity spot at the application
point (RF value in the range 0.0-0.1).
Palmitic acid. C16H32O2. (Mr 256.4). 1061600. [57-10-3].
Hexadecanoic acid. Paracetamol. 1061900. [103-90-2].
White or almost white, crystalline scales, practically insoluble See Paracetamol (0049).
in water, freely soluble in hot ethanol (96 per cent).
mp : about 63 °C. Paracetamol, 4-aminophenol-free. 1061901.
Chromatography. Thin-layer chromatography (2.2.27) Recrystallise paracetamol R from water R and dry in vacuo
as prescribed in the monograph Chloramphenicol at 70 °C ; repeat the procedure until the product complies
palmitate (0473) ; the chromatogram shows only one principal with the following test : dissolve 5 g of the dried substance
spot. in a mixture of equal volumes of methanol R and water R
and dilute to 100 mL with the same mixture of solvents.
Palmitic acid used in the assay of total fatty acids in the Add 1 mL of a freshly prepared solution containing 10 g/L
monograph Saw palmetto fruit (1848) complies with the of sodium nitroprusside R and 10 g/L of anhydrous sodium
following additional test. carbonate R, mix and allow to stand for 30 min protected
Assay. Gas chromatography (2.2.28) as prescribed in the from light. No blue or green colour is produced.
monograph Saw palmetto fruit (1848).
Content : minimum 98 per cent, calculated by the Paraffin, liquid. 1062000. [8042-47-5].
normalisation procedure. See Liquid paraffin (0239).

Paraffin, white soft. 1062100. the hydrolysis of not less than 500 mg of benzylpenicillin to
A semi-liquid mixture of hydrocarbons obtained from benzylpenicilloic acid per hour) at 30 °C and pH 7, provided
petroleum and bleached, practically insoluble in water and that the concentration of benzylpenicillin does not fall below
in ethanol (96 per cent), soluble in light petroleum R1, the the level necessary for enzyme saturation.
solution sometimes showing a slight opalescence. The Michaelis constant for benzylpenicillin of the penicillinase
in penicillinase solution is approximately 12 μg/mL.
Paraldehyde. 1151000. [123-63-7].
Sterility (2.6.1). It complies with the test for sterility.
See Paraldehyde (0351).
Storage : at a temperature between 0 °C and 2 °C for 2 to
Pararosaniline hydrochloride. C19H18ClN3. (Mr 323.8). 3 days. When freeze-dried and kept in sealed ampoules, it
1062200. [569-61-9]. may be stored for several months.
Schultz No. 779. Pentaerythrityl tetrakis[3-(3,5-di(1,1-dimethylethyl)-
Colour Index No. 42500. 4-hydroxyphenyl)propionate]. C73H108O12. (Mr 1178).
4-[Bis(4-aminophenyl)methylene]cyclohexa-2,5-dieniminium 1062400. [6683-19-8]. Pentaerythrityl tetrakis[3-
chloride. (3,5-di-tert-butyl-4-hydroxyphenyl) propionate].
Bluish-red, crystalline powder, slightly soluble in water, 2,2′-Bis(hydroxymethyl)propane-1,3-diol tetrakis[3-[3,5-
soluble in anhydrous ethanol. Solutions in water and di(1,1-dimethylethyl)-4-hydroxyphenyl]]propionate.
anhydrous ethanol are deep-red ; solutions in sulfuric acid and White or slightly yellow, crystalline powder, practically
in hydrochloric acid are yellow. insoluble in water, very soluble in acetone, soluble in
mp : about 270 °C, with decomposition. methanol, slightly soluble in hexane.
Decolorised pararosaniline solution. 1062201. mp : 110 °C to 125 °C.
To 0.1 g of pararosaniline hydrochloride R in a α-form : 120 °C to 125 °C.
ground-glass-stoppered flask add 60 mL of water R and β-form : 110 °C to 115 °C.
a solution of 1.0 g of anhydrous sodium sulfite R or 2.0 g Pentafluoropropanoic acid. C3HF5O2. (Mr 164.0). 1151100.
of sodium sulfite heptahydrate R or 0.75 g of sodium [422-64-0].
metabisulfite R in 10 mL of water R. Slowly and with stirring
add 6 mL of dilute hydrochloric acid R, stopper the flask Clear, colourless liquid.
20
and continue stirring until dissolution is complete. Dilute d 20 : about 1.561.
to 100 mL with water R. Allow to stand for 12 h before use. 20
nD : about 1.284.
bp : about 97 °C.
Parthenolide. C15H20O3. (Mr 248.3). 1129900. [20554-84-1].
Pentafluoropropionic anhydride. C6F10O3. (Mr 310.0).
(4E)-(1aR,7aS,10aS,10bS)-1a,5-Dimethyl-8-methylene-
1177300. [356-42-3]. Pentafluoropropanoic anhydride.
2,3,6,7,7a,8,10a,10b-octahydro-oxireno[9,10]cyclodeca[1,2-
b]furan-9(1aH)-one. (E)-(5S,6S)-4,5-Epoxygermacra- Pentane. C5H12. (Mr 72.2). 1062500. [109-66-0].
1(10),11(13)-dieno-12(6)-lactone. Clear, colourless, flammable liquid, very slightly soluble in
White or almost white, crystalline powder, very slightly water, miscible with acetone and with anhydrous ethanol.
soluble in water, very soluble in methylene chloride, soluble : about 0.63.
in methanol.
: about 1.359.
: − 71.4, determined on a 2.2 g/L solution in methylene
chloride R. bp : about 36 °C.
mp : 115 °C to 116 °C. Pentane used in spectrophotometry complies with the following
additional test.
Absorbance (2.2.25). A 0.01 g/L solution in ethanol (96 per
cent) R shows an absorption maximum at 214 nm. Absorbance (2.2.25) : maximum 0.70 at 200 nm, 0.30 at
210 nm, 0.07 at 220 nm, 0.03 at 230 nm, 0.01 at 240 nm,
Assay. Liquid chromatography (2.2.29) as prescribed in the determined using water R as compensation liquid.
monograph Feverfew (1516), at the concentration of the
reference solution. 1,2-Pentanediol. C5H12O2. (Mr 104.2). 1155800. [5343-92-0].
Content : minimum 90 per cent, calculated by the (2RS)-Pentane-1,2-diol.
normalisation procedure. : about 0.971.
L-Penicillamine coated silica gel for chiral separations.
: about 1.439.
1200500. bp : about 201 °C.
A very finely divided silica gel for chromatography coated Pentanol. C5H12O. (Mr 88.1). 1062600. [71-41-0].
with L-penicillamine. Pentan-1-ol.
Penicillinase solution. 1062300. Colourless liquid, sparingly soluble in water, miscible with
Dissolve 10 g of casein hydrolysate, 2.72 g of potassium
dihydrogen phosphate R and 5.88 g of sodium citrate R in : about 1.410.
200 mL of water R, adjust to pH 7.2 with a 200 g/L solution bp : about 137 °C.
of sodium hydroxide R and dilute to 1000 mL with water R.
Dissolve 0.41 g of magnesium sulfate R in 5 mL of water R 3-Pentanone. C5H10O. (Mr 86.13). 1173600. [96-22-0].
and add 1 mL of a 1.6 g/L solution of ferrous ammonium Pentan-3-one. Diethyl ketone.
sulfate R and sufficient water R to produce 10 mL. Sterilise tert-Pentyl alcohol. C5H12O. (Mr 88.1). 1062700. [75-85-4].
both solutions by heating in an autoclave, cool, mix, distribute tert-Amyl alcohol. 2-Methyl-2-butanol.
in shallow layers in conical flasks and inoculate with Bacillus Volatile, flammable liquid, freely soluble in water, miscible
cereus (NCTC 9946). Allow the flasks to stand at 18 °C to with ethanol (96 per cent) and with glycerol.
37 °C until growth is apparent and then maintain at 35 °C
to 37 °C for 16 h, shaking constantly to ensure maximum : about 0.81.
aeration. Centrifuge and sterilise the supernatant by filtration Distillation range (2.2.11). Not less than 95 per cent distils
through a membrane filter. 1.0 mL of penicillinase solution between 100 °C and 104 °C.
contains not less than 0.4 microkatals (corresponding to Storage : protected from light.

Pentetic acid. C14H23N3O10. (Mr 393.3). 1183100. [67-43-6]. Petroleum, light R3. 1063103. Petroleum
[[(Carboxymethyl)imino]bis(ethylenenitrilo)]tetraacetic acid. ether 100-120 °C.
White or almost white powder, slightly soluble in water. Complies with the requirements prescribed for light
mp : 219 °C to 220 °C, with decomposition. petroleum R, with the following modifications.
20
Pepsin powder. 1062800. [9001-75-6]. d 20 : about 0.720.
See Pepsin powder (0682). Distillation range (2.2.11) : 100 °C to 120 °C.
Water (2.5.12): maximum 0.03 per cent.
Peptide N-glycosidase F. 1186600. [83534-39-8].
Peptide-N4-(N-acetyl-β-glucosaminyl)asparagine amidase Petroleum, light R4. 1063104. Petroleum ether 80-100 °C.
(EC 3.5.1.52). PNGase F. Complies with the requirements prescribed for light
petroleum R, with the following modifications.
Perchloric acid. HClO4. (Mr 100.5). 1062900. [7601-90-3].
: about 0.70.
Content : 70.0 per cent m/m to 73.0 per cent m/m.
Distillation range (2.2.11): 80 °C to 100 °C.
Clear, colourless liquid, miscible with water.
: about 1.7. pH indicator strip. 1178900.
Assay. To 2.50 g add 50 mL of water R and titrate with 1 M Paper strip, or plastic strip containing multiple segments
sodium hydroxide, using 0.1 mL of methyl red solution R as of different dye-impregnated papers, allowing visual
indicator. determination of pH in the prescribed range, by comparison
1 mL of 1 M sodium hydroxide is equivalent to 100.5 mg of with the corresponding master chart.
HClO4. α-Phellandrene. C10H16. (Mr 136.2). 1130400.
Perchloric acid solution. 1062901. [4221-98-1]. (R)-5-Isopropyl-2-methyl-cyclohexa-1,3-diene.
(–)-p-Mentha-1,5-diene.
Dilute 8.5 mL of perchloric acid R to 100 mL with water R.
: about 1.471.
Perfluoroheptanoic acid. C7HF13O2. (Mr 364.1). 1207400. bp : 171 °C to 174 °C.
[375-85-9]. Tridecafluoroheptanoic acid. α-Phellandrene used in gas chromatography complies with the
Periodic acetic acid solution. 1063000. following additional test.
Dissolve 0.446 g of sodium periodate R in 2.5 mL of a 25 per Assay. Gas chromatography (2.2.28) as prescribed in the
cent V/V solution of sulfuric acid R. Dilute to 100.0 mL with monograph Eucalyptus oil (0390).
glacial acetic acid R. Test solution. The substance to be examined.
Periodic acid. H 5IO6. (Mr 227.9). 1108900. [10450-60-9]. Content : 95.0 per cent, calculated by the normalisation
procedure.
Crystals, freely soluble in water and soluble in ethanol (96 per
cent). Phenanthrene. C14H10. (Mr 178.2). 1063200. [85-01-8].
mp : about 122 °C. White or almost white crystals, practically insoluble in water,
sparingly soluble in ethanol (96 per cent).
Permethrin. C21H20Cl2O3. (Mr 391.3). 1130000. [52645-53-1].
mp : about 100 °C.
mp : 34 °C to 35 °C.
A suitable certified reference solution (10 ng/µL in Phenanthroline hydrochloride. C12H9ClN2,H2O. (Mr 234.7).
cyclohexane) may be used. 1063300. [18851-33-7]. 1,10-Phenanthroline hydrochloride
monohydrate.
Peroxide test strips. 1147800. White or almost white, crystalline powder, freely soluble in
Use commercial test strips with a suitable scale in the range water, soluble in ethanol (96 per cent).
from 0 ppm to 25 ppm peroxide. mp : about 215 °C, with decomposition.
Perylene. C20H12. (Mr 252.3). 1130100. [198-55-0]. Phenazone. 1063400. [60-80-0].
Dibenz[de,kl]anthracene.
See Phenazone (0421).
Orange powder.
mp : about 279 °C. Phenol. 1063500. [108-95-2].
See Phenol (0631).
Petroleum, light. 1063100. [8032-32-4]. Petroleum ether
50-70 °C. Phenolphthalein. C20H14O4. (Mr 318.3). 1063700. [77-09-8].
Clear, colourless, flammable liquid without fluorescence, 3,3-Bis(4-hydroxyphenyl)-3H-isobenzofuran-1-one.
practically insoluble in water, miscible with ethanol (96 per White or yellowish-white powder, practically insoluble in
cent). water, soluble in ethanol (96 per cent).
: 0.661 to 0.664. Phenolphthalein paper. 1063704.
Distillation range (2.2.11) : 50 °C to 70 °C. Immerse strips of filter paper for a few minutes in
Petroleum, light R1. 1063101. Petroleum ether 40-60 °C. phenolphthalein solution R. Allow to dry.
Complies with the requirements prescribed for light Phenolphthalein solution. 1063702.
petroleum R, with the following modifications. Dissolve 0.1 g of phenolphthalein R in 80 mL of ethanol
: 0.630 to 0.656. (96 per cent) R and dilute to 100 mL with water R.
Distillation range (2.2.11) : 40 °C to 60 °C. It does not Test for sensitivity. To 0.1 mL of the phenolphthalein
become cloudy at 0 °C. solution add 100 mL of carbon dioxide-free water R. The
Petroleum, light R2. 1063102. Petroleum ether 30-40 °C. solution is colourless. Not more than 0.2 mL of 0.02 M
sodium hydroxide is required to change the colour to pink.
Complies with the requirements prescribed for light
petroleum R, with the following modifications. Colour change: pH 8.2 (colourless) to pH 10.0 (red).
: 0.620 to 0.630. Phenolphthalein solution R1. 1063703.
Distillation range (2.2.11) : 30 °C to 40 °C. It does not A 10 g/L solution of phenolphthalein R in ethanol (96 per
become cloudy at 0 °C. cent) R.

Phenol red. 1063600. [143-74-8]. D-Phenylglycine. C8H9NO2. (Mr 151.2). 1144500. [875-74-1].

Bright red or dark red, crystalline powder, very slightly soluble (2R)-2-Amino-2-phenylacetic acid.
in water, slightly soluble in ethanol (96 per cent). Content : minimum 99 per cent.
Phenol red solution. 1063601. White or almost white, crystalline powder.
Dissolve 0.1 g of phenol red R in a mixture of 2.82 mL Phenylhydrazine. C6H8N2. (Mr 108.1). 1190800. [100-63-0].
of 0.1 M sodium hydroxide and 20 mL of ethanol (96 per White or almost white, crystalline powder, becoming yellow
cent) R and dilute to 100 mL with water R. or dark red on exposure to air, melting at room temperature
Test for sensitivity. Add 0.1 mL of the phenol red solution giving an oily liquid, miscible with anhydrous ethanol,
to 100 mL of carbon dioxide-free water R. The solution is sparingly soluble in water.
yellow. Not more than 0.1 mL of 0.02 M sodium hydroxide bp : about 244 °C, with decomposition.
is required to change the colour to reddish-violet. mp : about 20 °C.
Colour change : pH 6.8 (yellow) to pH 8.4 (reddish-violet).
Phenylhydrazine hydrochloride. C6H9ClN2. (Mr 144.6).
Phenol red solution R2. 1063603. 1064500. [59-88-1].
Solution A. Dissolve 33 mg of phenol red R in 1.5 mL of White or almost white, crystalline powder, becoming brown
dilute sodium hydroxide solution R and dilute to 100 mL on exposure to air, soluble in water and in ethanol (96 per
with water R. cent).
Solution B. Dissolve 25 mg of ammonium sulfate R in mp : about 245 °C, with decomposition.
235 mL of water R ; add 105 mL of dilute sodium hydroxide Storage : protected from light.
solution R and 135 mL of dilute acetic acid R.
Add 25 mL of solution A to solution B. If necessary, adjust Phenylhydrazine hydrochloride solution. 1064501.
the pH of the mixture to 4.7. Dissolve 0.9 g of phenylhydrazine hydrochloride R in 50 mL
of water R. Decolorise with activated charcoal R and filter.
Phenol red solution R3. 1063604. To the filtrate add 30 mL of hydrochloric acid R and dilute
Solution A. Dissolve 33 mg of phenol red R in 1.5 mL of to 250 mL with water R.
dilute sodium hydroxide solution R and dilute to 50 mL
with water R. Phenylhydrazine-sulfuric acid solution. 1064502.
Solution B. Dissolve 50 mg of ammonium sulfate R in Dissolve 65 mg of phenylhydrazine hydrochloride R,
235 mL of water R ; add 105 mL of dilute sodium hydroxide previously recrystallised from ethanol (85 per cent V/V) R,
solution R and 135 mL of dilute acetic acid R. in a mixture of 80 volumes of water R and 170 volumes of
sulfuric acid R and dilute to 100 mL with the same mixture
Add 25 mL of solution A to solution B ; if necessary, adjust of solvents. Prepare immediately before use.
the pH of the mixture to 4.7.
Phenyl isothiocyanate. C7H5NS. (Mr 135.2). 1121500.
Phenoxyacetic acid. C8H8O3. (Mr 152.1). 1063800. [103-72-0].
[122-59-8]. 2-Phenoxyethanoic acid.
Liquid, insoluble in water, soluble in ethanol (96 per cent).
Almost white crystals, sparingly soluble in water, freely soluble
: about 1.13.
in ethanol (96 per cent), and in glacial acetic acid.
: about 1.65.
mp : about 98 °C.
bp : about 221 °C.
prescribed in the monograph Phenoxymethylpenicillin (0148) ; mp : about − 21 °C.
the chromatogram shows only one principal spot. Use a grade suitable for protein sequencing.
2-Phenoxyaniline. C12H11NO. (Mr 185.2). 1165500. Phenyl(5)methyl(95)polysiloxane. 1066900.
[2688-84-8]. 2-Phenoxybenzenamine. 2-Aminophenyl phenyl Polysiloxane substituted with 5 per cent of phenyl groups and
ether. 95 per cent of methyl groups.
Phenoxyethanol. C8H10O2. (Mr 138.2). 1064000. [122-99-6]. Phenyl(5)methyl(95)polysiloxane, base-deactivated.
2-Phenoxyethanol. 1176600.
Clear, colourless, oily liquid, slightly soluble in water, freely Base-deactivated polysiloxane substituted with 5 per cent of
soluble in ethanol (96 per cent). phenyl groups and 95 per cent of methyl groups.
: about 1.11. Phenyl(50)methyl(50)polysiloxane. 1067900.
: about 1.537. Polysiloxane substituted with 50 per cent of phenyl groups
Freezing point (2.2.18) : minimum 12 °C. and 50 per cent of methyl groups.
Phenylacetic acid. C8H8O2. (Mr 136.2). 1160000. [103-82-2]. 1-Phenylpiperazine. C10H14N2. (Mr 162.2). 1130500.
White or almost white powder, soluble in water. [92-54-6].
bp : about 265 °C. Slightly viscous, yellow liquid, not miscible with water.
mp : about 75 °C. : about 1.07.
: about 1.588.
Phenylalanine. 1064100. [63-91-2].
See Phenylalanine (0782). 1-Phenylpropan-2-ol. C9H12O. (Mr 136.2). 1205200.
[698-87-3]. (2RS)-1-Phenylpropan-2-ol.
p-Phenylenediamine dihydrochloride. C6H10Cl2N2. mp : 65 °C to 67 °C.
(Mr 181.1). 1064200. [615-28-1]. 1,4-Diaminobenzene
dihydrochloride. 1-Phenyl-1,2,3,4-tetrahydroisoquinoline. C15H15N.
Crystalline powder or white or slightly coloured crystals, (Mr 209.3). 1193700. [22990-19-8].
turning reddish on exposure to air, freely soluble in water, Phloroglucide. C12H10O5. (Mr 234.2). 1177400. [491-45-2].
slightly soluble in ethanol (96 per cent). 2,3′,4,5′,6-Biphenylpentol.
α-Phenylglycine. C8H9NO2. (Mr 151.2). 1064300. White or almost white powder, hygroscopic, light sensitive.
[2835-06-5]. (RS)-2-Amino-2-phenylacetic acid. Slowly discolours on exposure to light.

Phloroglucinol. C6H6O3,2H2O. (Mr 162.1). 1064600. Phthalaldehyde. C8H6O2. (Mr 134.1). 1065300. [643-79-8].

[6099-90-7]. Benzene-1,3,5-triol. Benzene-1,2-dicarboxaldehyde.
White or yellowish crystals, slightly soluble in water, soluble Yellow, crystalline powder.
in ethanol (96 per cent). mp : about 55 °C.
mp : about 223 °C (instantaneous method). Storage : protected from light and air.
Phloroglucinol solution. 1064601. Phthalaldehyde reagent. 1065301.
To 1 mL of a 100 g/L solution of phloroglucinol R in ethanol Dissolve 2.47 g of boric acid R in 75 mL of water R, adjust to
(96 per cent) R, add 9 mL of hydrochloric acid R. pH 10.4 using a 450 g/L solution of potassium hydroxide R
and dilute to 100 mL with water R. Dissolve 1.0 g of
phthalaldehyde R in 5 mL of methanol R, add 95 mL of the
Phosalone. C12H15ClNO4PS2. (Mr 367.8). 1130200. boric acid solution and 2 mL of thioglycollic acid R and
[2310-17-0]. adjust to pH 10.4 with a 450 g/L solution of potassium
hydroxide R.
mp : 45 °C to 48 °C
Storage : protected from light ; use within 3 days.
A suitable certified reference solution (10 ng/µL in iso-octane)
may be used. Phthalazine. C8H6N2. (Mr 130.1). 1065400. [253-52-1].
Pale yellow crystals, freely soluble in water, soluble in
Phosphomolybdic acid. 12MoO3,H3PO4,xH2O. 1064900. anhydrous ethanol, in ethyl acetate and in methanol.
[51429-74-4].
mp : 89 °C to 92 °C.
Orange-yellow, fine crystals, freely soluble in water, soluble in
ethanol (96 per cent). Phthalein purple. C32H32N2O12,xH2O. (Mr 637,
anhydrous substance). 1065500. [2411-89-4].
Phosphomolybdic acid solution. 1064901. Metalphthalein. 2,2′,2″,2″′-[o-Cresolphthalein-3′,3″-
Dissolve 4 g of phosphomolybdic acid R in water R and bis(methylenenitrilo)]tetra-acetic acid. (1,3-Dihydro-3-
dilute to 40 mL with the same solvent. Add cautiously and oxo-isobenzofuran-1-ylidene)bis[(6-hydroxy-5-methyl-3,1-
with cooling 60 mL of sulfuric acid R. Prepare immediately phenylene)bis(methyleneimino)diacetic acid].
before use. Yellowish-white or brownish powder, practically insoluble
in water, soluble in ethanol (96 per cent). The product may
Phosphomolybdotungstic reagent. 1065000. be found in commerce in the form of the sodium salt : a
Dissolve 100 g of sodium tungstate R and 25 g of sodium yellowish-white to pink powder, soluble in water, practically
molybdate R in 700 mL of water R. Add 100 mL of hydrochloric insoluble in ethanol (96 per cent).
acid R and 50 mL of phosphoric acid R. Heat the mixture Test for sensitivity. Dissolve 10 mg in 1 mL of concentrated
under a reflux condenser in a glass apparatus for 10 h. Add ammonia R and dilute to 100 mL with water R. To 5 mL of
150 g of lithium sulfate R, 50 mL of water R and a few drops the solution add 95 mL of water R, 4 mL of concentrated
of bromine R. Boil to remove the excess of bromine (15 min), ammonia R, 50 mL of ethanol (96 per cent) R and 0.1 mL
allow to cool, dilute to 1000 mL with water R and filter. The of 0.1 M barium chloride. The solution is blue-violet. Add
reagent should be yellow in colour. If it acquires a greenish 0.15 mL of 0.1 M sodium edetate. The solution becomes
tint, it is unsatisfactory for use but may be regenerated by colourless.
boiling with a few drops of bromine R. Care must be taken to
remove the excess of bromine by boiling. Phthalic acid. C8H6O4. (Mr 166.1). 1065600. [88-99-3].
Benzene-1,2-dicarboxylic acid.
White or almost white, crystalline powder, soluble in hot
Phosphomolybdotungstic reagent, dilute. 1065001. water and in ethanol (96 per cent).
To 1 volume of phosphomolybdotungstic reagent R add Phthalic anhydride. C8H4O3. (Mr 148.1). 1065700. [85-44-9].
2 volumes of water R. Isobenzofuran-1,3-dione.
Phosphoric acid. 1065100. [7664-38-2]. Content : minimum 99.0 per cent.
See Concentrated phosphoric acid (0004). White or almost white flakes.
mp : 130 °C to 132 °C.
Phosphoric acid, dilute. 1065101. Assay. Dissolve 2.000 g in 100 mL of water R and boil under a
See Dilute phosphoric acid (0005). reflux condenser for 30 min. Cool and titrate with 1 M sodium
hydroxide, using phenolphthalein solution R as indicator.
Phosphoric acid, dilute R1. 1065102. 1 mL of 1 M sodium hydroxide is equivalent to 74.05 mg of
Dilute 93 mL of dilute phosphoric acid R to 1000 mL with C 8H 4O 3.
water R.
Phthalic anhydride solution. 1065701.
Phosphorous acid. H3PO3. (Mr 82.0). 1130600. [13598-36-2]. Dissolve 42 g of phthalic anhydride R in 300 mL of
White or almost white, very hygroscopic and deliquescent anhydrous pyridine R. Allow to stand for 16 h.
crystalline mass ; slowly oxidised by oxygen (air) to H3PO4. Storage : protected from light ; use within 1 week.
Unstable, orthorhombic crystals, soluble in water, in ethanol Picein. C14H18O7. (Mr 298.3). 1130700. [530-14-3].
(96 per cent) and in a mixture of 3 volumes of ether and 1-[4-(β-D-Glucopyranosyloxy)phenyl]ethanone.
1 volume of ethanol (96 per cent). p-(Acetylphenyl)-β-D-glucopyranoside.
: 1.651. mp : 194 °C to 195 °C.
mp : about 73 °C.
Picric acid. C6H3N3O7. (Mr 229.1). 1065800. [88-89-1].
Phosphotungstic acid solution. 1065200. 2,4,6-Trinitrophenol.
Heat under a reflux condenser for 3 h, 10 g of sodium Yellow prisms or plates, soluble in water and in ethanol (96 per
tungstate R with 8 mL of phosphoric acid R and 75 mL of cent).
water R. Allow to cool and dilute to 100 mL with water R. Storage : moistened with water R.

Picric acid solution. 1065801. Piperitone. C10H16O. (Mr 152.2). 1151200. [89-81-6].

A 10 g/L solution of picric acid R. 6-Isopropyl-3-methyl-cyclohex-2-en-1-one.
Picric acid solution R1. 1065802. Pirimiphos-ethyl. C13H24N3O3PS. (Mr 333.4). 1130300.
Prepare 100 mL of a saturated solution of picric acid R and [23505-41-1].
add 0.25 mL of strong sodium hydroxide solution R. mp : 15 °C to 18 °C.
Picrotin. C15H18O7. (Mr 310.3). 1188100. [21416-53-5]. A suitable certified reference solution (10 ng/µL in
(1R,3R,5S,8S,9R,12S,13R,14S)-1-hydroxy-14-(2- cyclohexane) may be used.
hydroxypropan-2-yl)-13-methyl-4,7,10-trioxapenta- Plasma, platelet-poor. 1066100.
cyclo[6.4.1.1.9,12.03,5.05,13]tetradecane-6,11-dione.
Withdraw 45 mL of human blood into a 50 mL plastic syringe
White or colourless crystalline powder or crystals, soluble containing 5 mL of a sterile 38 g/L solution of sodium citrate R.
in boiling water and in ethanol (96 per cent), practically Without delay, centrifuge at 1500 g at 4 °C for 30 min. Remove
insoluble in methylene chloride. the upper two-thirds of the supernatant plasma using a plastic
mp : 248 °C to 250 °C. syringe and without delay centrifuge at 3500 g at 4 °C for
30 min. Remove the upper two-thirds of the liquid and freeze
Picrotoxinin. C15H16O6. (Mr 292.2). 1188200.
it rapidly in suitable amounts in plastic tubes at or below
[17617-45-7]. (1R,3R,5S,8S,9R,12S,13R,14R)-1-
− 40 °C. Use plastic or silicone-treated equipment.
hydroxy-13-methyl-14-(prop-1-en-2-yl)-4,7,10-trioxa-
pentacyclo[6.4.1.19,12.03,5.05,13]tetradecane-6,11-dione. Plasma substrate. 1066200.
White or colourless crystalline powder or crystals, soluble in Separate the plasma from human or bovine blood collected
methylene chloride, in ethanol (96 per cent) and in alkaline into one-ninth its volume of a 38 g/L solution of sodium
solutions. citrate R, or into two-sevenths its volume of a solution
mp : 207 to 210 °C. containing 20 g/L of disodium hydrogen citrate R and 25 g/L
of glucose R. With the former, prepare the substrate on the
α-Pinene. C10H16. (Mr 136.2). 1130800. [7785-70-8]. day of collection of the blood. With the latter, prepare within
(1R,5R)-2,6,6-Trimethylbicyclo[3.1.1]hept-2-ene. two days of collection of the blood.
Liquid not miscible with water. Storage : at − 20 °C.
: about 0.859.
: about 1.466. Plasma substrate R1. 1066201.
bp : 154 °C to 156 °C. Use water-repellent equipment (made from materials such
α-Pinene used in gas chromatography complies with the as suitable plastics or suitably silicone-treated glass) for
following additional test. taking and handling blood.
Assay. Gas chromatography (2.2.28) as prescribed in the Collect a suitable volume of blood from each of at least
monograph Bitter-orange-flower oil (1175). five sheep ; a 285 mL volume of blood collected into 15 mL
of anticoagulant solution is suitable but smaller volumes
Test solution. The substance to be examined. may be collected, taking the blood, either from a live
Content : minimum 99.0 per cent, calculated by the animal or at the time of slaughter, using a needle attached
normalisation procedure. to a suitable cannula which is long enough to reach the
β-Pinene. C10H16. (Mr 136.2). 1109000. [127-91-3]. bottom of the collecting vessel. Discarding the first few
6,6-Dimethyl-2-methylenebicyclo[3.1.1]heptane. millilitres and collecting only free-flowing blood, collect
the blood in a sufficient quantity of an anticoagulant
Colourless, oily liquid, odour reminiscent of turpentine, solution containing 8.7 g of sodium citrate R and 4 mg of
practically insoluble in water, miscible with ethanol (96 per aprotinin R per 100 mL of water R to give a final ratio of
cent). blood to anticoagulant solution of 19 to 1. During and
β-Pinene used in gas chromatography complies with the immediately after collection, swirl the flask gently to ensure
following additional test. mixing but do not allow frothing to occur. When collection
Assay. Gas chromatography (2.2.28) as prescribed in the is complete, close the flask and cool to 10-15 °C. When
monograph Bitter-orange-flower oil (1175). cold, pool the contents of all the flasks with the exception
Test solution. The substance to be examined. of any that show obvious haemolysis or clots and keep the
Content : minimum 95.0 per cent. pooled blood at 10-15 °C.
As soon as possible and within 4 h of collection, centrifuge
1,4-Piperazinediethanesulfonic acid. C8H18N2O6S2. the pooled blood at 1000-2000 g at 10-15 °C for 30 min.
(Mr 302.4). 1186700. [5625-37-6]. Piperazine-1,4- Separate the supernatant and centrifuge it at 5000 g for
bis(2-ethanesulfonic acid). 2,2′-(Piperazine-1,4- 30 min. (Faster centrifugation, for example 20 000 g for
diyl)bis(ethanesulfonic acid). Piperazine-N,N′-bis(2- 30 min, may be used if necessary to clarify the plasma,
ethanesulfonic acid). PIPES. but filtration procedures should not be used.) Separate
Content : minimum 99 per cent. the supernatant and, without delay, mix thoroughly and
White, crystalline powder. distribute the plasma substrate into small stoppered
containers in portions sufficient for a complete heparin
Piperazine hydrate. 1065900. [142-63-2]. assay (for example 10 mL to 30 mL). Without delay, rapidly
See Piperazine hydrate (0425). cool to a temperature below − 70 °C (for example by
immersing the containers into liquid nitrogen) and store at
Piperidine. C5H11N. (Mr 85.2). 1066000. [110-89-4].
a temperature below − 30 °C.
Hexahydropyridine.
Colourless to slightly yellow, alkaline liquid, miscible with The plasma is suitable for use as plasma substrate in the
water, with ethanol (96 per cent) and with light petroleum. assay for heparin if, under the conditions of the assay,
it gives a clotting time appropriate to the method of
bp : about 106 °C. detection used and if it provides reproducible, steep log
Piperine. C17H19NO3. (Mr 285.3). 1183200. [94-62-2]. dose-response curves.
(2E,4E)-1-(Piperidin-1-yl)-5-(1,3-benzodioxol-5-yl)penta- When required for use, thaw a portion of the plasma
2,4-dien-1-one. 1-Piperoyl-piperidine. 1-[(2E,4E)-5-(3,4- substrate in a water-bath at 37 °C, gently swirling until
Methylenedioxyphenyl)-1-oxo-2,4-pentadienyl]piperidine. thawing is complete ; once thawed it should be kept at

10-20 °C and used without delay. The thawed plasma Polymethacrylate gel, hydroxylated. 1151300.
substrate may be lightly centrifuged if necessary ; filtration Stationary phase for size-exclusion chromatography.
procedures should not be used. Gel based on hydroxylated methacrylic acid polymer.

Plasma substrate R2. 1066202.

Prepare from human blood containing less than 1 per Polyorganosiloxane for oxygen-containing compounds.
cent of the normal amount of factor IX. Collect the blood 1200600.
into one-ninth its volume of a 38 g/L solution of sodium

Combination of suitable polyorganosiloxanes with high

citrate R. affinity for oxygen-containing compounds.
Storage : in small amounts in plastic tubes at a temperature
of − 30 °C or lower. Polyoxyethylated castor oil. 1068200.
Light yellow liquid. It becomes clear above 26 °C.
Plasma substrate R3. 1066203.
Prepare from human blood containing less than 1 per Polysorbate 20. 1068300. [9005-64-5].
cent of the normal amount of factor XI. Collect the blood See Polysorbate 20 (0426).
into one-ninth its volume of a 38 g/L solution of sodium
Polysorbate 65. 1196200. [9005-71-4].
citrate R.
Storage : in small amounts in plastic tubes at a temperature Polysorbate 80. 1068400. [9005-65-6].
of − 30 °C or lower. See Polysorbate 80 (0428).
Plasma substrate deficient in factor V. 1066300. Polystyrene 900-1000. 1112200. [9003-53-6].
Use preferably a plasma which is congenitally deficient, or Organic standard used for calibration in gas chromatography.
prepare it as follows : separate the plasma from human blood Mw : about 950.
collected into one tenth of its volume of a 13.4 g/L solution Mw/Mn : 1.10.
of sodium oxalate R. Incubate at 37 °C for 24 h to 36 h. The
coagulation time determined by the method prescribed for Potassium acetate. 1175900. [127-08-2].
coagulation factor V solution R should be 70 s to 100 s. If the See Potassium acetate (1139).
coagulation time is less than 70 s, incubate again for 12 h to
24 h. Potassium bicarbonate. 1069900. [298-14-6].
Storage : in small quantities at a temperature of − 20 °C or See Potassium hydrogen carbonate R.
lower. Potassium bicarbonate solution, saturated methanolic.
Plasminogen, human. 1109100. [9001-91-6]. 1069901.
A substance present in blood that may be activated to plasmin, See potassium hydrogen carbonate solution, saturated
an enzyme that lyses fibrin in blood clots. methanolic R.
Plutonium-242 spiking solution. 1167400. Potassium bromate. KBrO3. (Mr 167.0). 1068700.

[7758-01-2].
Contains 50 Bq/L Pu and a 134 mg/L solution of lanthanum
242
chloride heptahydrate R in a 284 g/L solution of nitric acid R. White or almost white granular powder or crystals, soluble in
water, slightly soluble in ethanol (96 per cent).
Poloxamer 188. 1186800.
Potassium bromide. 1068800. [7758-02-3].
See Poloxamers (1464).
See Potassium bromide (0184).
Polyamine grafted poly(vinyl alcohol) copolymer. 1188300. Potassium bromide used for infrared absorption
Copolymer beads of poly(vinyl alcohol) to which polyamine spectrophotometry (2.2.24) also complies with the following
is covalently bonded. The size range of the beads is specified additional test.
after the name of the reagent in the tests where it is used. A disc 2 mm thick prepared from the substance previously
dried at 250 °C for 1 h, has a substantially flat baseline over

Polydatin. C20H22O8. (Mr 390.4). 1197500. [65914-17-2]. the range 4000 cm− 1 to 620 cm− 1. It exhibits no maxima
3-Hydroxy-5-[2-(4-hydroxyphenyl)eth-1-en-1-yl]phenyl with absorbance greater than 0.02 above the baseline, except
β-D-glucopyranoside. Resveratrol-3-β-mono-D-glucoside. maxima for water at 3440 cm− 1 and 1630 cm− 1.

Potassium carbonate. K2CO3. (Mr 138.2). 1068900.

Polyether hydroxylated gel for chromatography. 1067000. [584-08-7]. Dipotassium carbonate.
Gel with a small particle size having a hydrophilic surface White or almost white, granular powder, hygroscopic, very
with hydroxyl groups. It has an exclusion limit for dextran of soluble in water, practically insoluble in anhydrous ethanol.
relative molecular mass 2 × 105 to 2.5 × 106.
Polyethyleneglycol adipate. (C8H12O4)n. (Mr (172.2)n).
1067700. Potassium chlorate. KClO3. (Mr 122.6). 1069000.
[3811-04-9].
White or almost white, wax-like mass, practically insoluble in
water. A white or almost white powder, granules or crystals, soluble
in water.
mp : about 43 °C.

Potassium chloride. 1069100. [7447-40-7].

Polyethyleneglycol succinate. (C6H8O4)n. (Mr (144.1)n). See Potassium chloride (0185).
1067800. Potassium chloride used for infrared absorption
White or almost white, crystalline powder, practically spectrophotometry (2.2.24) also complies with the following
insoluble in water. additional test.
mp : about 102 °C. A disc 2 mm thick, prepared from the substance previously
dried at 250 °C for 1 h, has a substantially flat baseline over
Polymethacrylate gel. 1181100. the range 4000 cm− 1 to 620 cm− 1. It exhibits no maxima
A methacrylate-based size-exclusion stationary phase for with absorbance greater than 0.02 above the baseline, except
water-soluble samples. maxima for water at 3440 cm− 1 and 1630 cm− 1.

Potassium chloride, 0.1 M. 1069101. Potassium ferriperiodate solution. 1070801.

A solution of potassium chloride R containing the Dissolve 1 g of potassium periodate R in 5 mL of a freshly
equivalent of 7.46 g of KCl in 1000.0 mL. prepared 120 g/L solution of potassium hydroxide R. Add
20 mL of water R and 1.5 mL of ferric chloride solution R1.
Potassium chromate. K2CrO4. (Mr 194.2). 1069200. Dilute to 50 mL with a freshly prepared 120 g/L solution of
[7789-00-6]. Dipotassium chromate. potassium hydroxide R.
Yellow crystals, freely soluble in water.
Potassium ferrocyanide. K4[Fe(CN)6],3H2O. (Mr 422.4).
Potassium chromate solution. 1069201. 1069800. [14459-95-1]. Potassium hexacyanoferrate(II).
A 50 g/L solution of potassium chromate R. Transparent yellow crystals, freely soluble in water, practically
insoluble in ethanol (96 per cent).
Potassium citrate. 1069300. [6100-05-6].
Potassium ferrocyanide solution. 1069801.
See Potassium citrate (0400).
A 53 g/L solution of potassium ferrocyanide R.
Potassium cyanide. KCN. (Mr 65.1). 1069400. [151-50-8]. Potassium fluoride. KF. (Mr 58.1). 1137800. [7789-23-3].
White or almost white, crystalline powder or white or almost Colourless crystals or white or almost white crystalline
white mass or granules, freely soluble in water, slightly soluble powder, deliquescent, soluble in water, practically insoluble in
in ethanol (96 per cent). ethanol (96 per cent).
Potassium cyanide solution. 1069401. Potassium hydrogen carbonate. KHCO3. (Mr 100.1).
A 100 g/L solution of potassium cyanide R. 1069900. [298-14-6]. Potassium bicarbonate.
Transparent, colourless crystals, freely soluble in water,
Potassium cyanide solution, lead-free. 1069402. practically insoluble in ethanol (96 per cent).
Dissolve 10 g of potassium cyanide R in 90 mL of water R,
add 2 mL of strong hydrogen peroxide solution R diluted 1 Potassium hydrogen carbonate solution, saturated
to 5. Allow to stand for 24 h, dilute to 100 mL with water R methanolic. 1069901.
and filter. Dissolve 0.1 g of potassium hydrogen carbonate R in
The solution complies with the following test : take 10 mL of 0.4 mL of water R, heating on water-bath. Add 25 mL
the solution, add 10 mL of water R and 10 mL of hydrogen of methanol R and swirl, keeping the solution on the
sulfide solution R. No colour is evolved even after addition water-bath until dissolution is complete. Use a freshly
of 5 mL of dilute hydrochloric acid R. prepared solution.
Potassium hydrogen phthalate. C8H5KO4. (Mr 204.2).
Potassium dichromate. K2Cr2O7. (Mr 294.2). 1069500. 1070000. [877-24-7]. Potassium hydrogen benzene-1,2-
[7778-50-9]. Dipotassium dichromate. dicarboxylate.
Potassium dichromate used for the calibration of White or almost white crystals, soluble in water, slightly
spectrophotometers (2.2.25) contains not less than 99.9 per soluble in ethanol (96 per cent).
cent of K2Cr2O7, calculated with reference to the substance
dried at 130 °C. Potassium hydrogen phthalate, 0.2 M. 1070001.
Orange-red crystals, soluble in water, practically insoluble in A solution of potassium hydrogen phthalate R containing
ethanol (96 per cent). the equivalent of 40.84 g of C8H5KO4 in 1000.0 mL.
Assay. Dissolve 1.000 g in water R and dilute to 250.0 mL with Potassium hydrogen sulfate. KHSO4. (Mr 136.2). 1070100.
the same solvent. To 50.0 mL of this solution add a freshly [7646-93-7].
prepared solution of 4 g of potassium iodide R, 2 g of sodium Colourless, transparent, hygroscopic crystals, freely soluble in
hydrogen carbonate R and 6 mL of hydrochloric acid R in water giving a strongly acid solution.
100 mL of water R in a 500 mL flask. Stopper the flask and
allow to stand protected from light for 5 min. Titrate with Storage : in an airtight container.
0.1 M sodium thiosulfate, using 1 mL of iodide-free starch Potassium hydrogen tartrate. C4H5KO6. (Mr 188.2).
solution R as indicator. 1070200. [868-14-4]. Potassium hydrogen (2R,3R)-2,3-
1 mL of 0.1 M sodium thiosulfate is equivalent to 4.903 mg dihydroxybutane-1,4-dioate.
of K2Cr2O7. White or almost white, crystalline powder or colourless,
slightly opaque crystals, slightly soluble in water, soluble in
Potassium dichromate solution. 1069501. boiling water, practically insoluble in ethanol (96 per cent).
A 106 g/L solution of potassium dichromate R.
Potassium hydroxide. 1070300. [1310-58-3].
Potassium dichromate solution R1. 1069502. See Potassium hydroxide (0840).
A 5 g/L solution of potassium dichromate R. Potassium hydroxide, alcoholic, 2 M. 1070301.
Potassium dihydrogen phosphate. 1069600. [7778-77-0]. Dissolve 12 g of potassium hydroxide R in 10 mL of water R
See Potassium dihydrogen phosphate (0920). and dilute to 100 mL with ethanol (96 per cent) R.
Potassium hydroxide in alcohol (10 per cent V/V),
Potassium dihydrogen phosphate, 0.2 M. 1069601.
0.5 M. 1070302.
A solution of potassium dihydrogen phosphate R containing Dissolve 28 g of potassium hydroxide R in 100 mL of
the equivalent of 27.22 g of KH2PO4 in 1000.0 mL. ethanol (96 per cent) R and dilute to 1000 mL with water R.
Potassium ferricyanide. K3[Fe(CN)6]. (Mr 329.3). 1069700. Potassium hydroxide solution, alcoholic. 1070303.
[13746-66-2]. Potassium hexacyanoferrate(III).
Dissolve 3 g of potassium hydroxide R in 5 mL of water R
Red crystals, freely soluble in water. and dilute to 100 mL with aldehyde-free alcohol R. Decant
the clear solution. The solution should be almost colourless.
Potassium ferricyanide solution. 1069701.
Wash 5 g of potassium ferricyanide R with a little water R, Potassium hydroxide solution, alcoholic R1. 1070304.
dissolve and dilute to 100 mL with water R. Prepare Dissolve 6.6 g of potassium hydroxide R in 50 mL of water R
immediately before use. and dilute to 1000 mL with anhydrous ethanol R.

Potassium iodate. KIO3. (Mr 214.0). 1070400. [7758-05-6]. Potassium iodobismuthate solution R4. 1070605.
White or almost white, crystalline powder, soluble in water. Dissolve 1.7 g of bismuth subnitrate R in 20 mL of glacial acetic
acid R. Add 80 mL of distilled water R, 100 mL of a 400 g/L
Potassium iodide. 1070500. [7681-11-0]. solution of potassium iodide R, 200 mL of glacial acetic acid R
See Potassium iodide (0186). and dilute to 1000 mL with distilled water R. Mix 2 volumes
of this solution with 1 volume of a 200 g/L solution of barium
Potassium iodide and starch solution. 1070501. chloride R.
Dissolve 0.75 g of potassium iodide R in 100 mL of water R.
Potassium iodobismuthate solution R5. 1070606.
Heat to boiling and add whilst stirring a solution of 0.5 g
of soluble starch R in 35 mL of water R. Boil for 2 min and To 0.85 g of bismuth subnitrate R add 10 mL of glacial acetic
allow to cool. acid R and gently heat until completely dissolved. Add 40 mL
of water R and allow to cool. To 5 mL of this solution, add
Test for sensitivity. A mixture of 15 mL of the potassium 5 mL of a 400 g/L solution of potassium iodide R, 20 mL of
iodide and starch solution, 0.05 mL of glacial acetic acid R glacial acetic acid R and 70 mL of water R.
and 0.3 mL of iodine solution R2 is blue.
Potassium nitrate. KNO3. (Mr 101.1). 1070700. [7757-79-1].
Potassium iodide solution. 1070502. Colourless crystals, very soluble in water.
A 166 g/L solution of potassium iodide R.
Potassium periodate. KIO4. (Mr 230.0). 1070800.
Potassium iodide solution, iodinated. 1070503. [7790-21-8].
Dissolve 2 g of iodine R and 4 g of potassium iodide R in White or almost white, crystalline powder or colourless
10 mL of water R. When solution is complete dilute to crystals, soluble in water.
100 mL with water R.
Potassium permanganate. 1070900. [7722-64-7].
Potassium iodide solution, iodinated R1. 1070505. See Potassium permanganate (0121).
Dissolve 500 mg of iodine R and 1.5 g of potassium iodide R Potassium permanganate and phosphoric acid solution.
in water R and dilute to 25 mL with the same solvent. 1070901.
Potassium iodide solution, saturated. 1070504. Dissolve 3 g of potassium permanganate R in a mixture of
15 mL of phosphoric acid R and 70 mL of water R. Dilute to
A saturated solution of potassium iodide R in carbon
100 mL with water R.
dioxide-free water R. Make sure the solution remains
saturated as indicated by the presence of undissolved Potassium permanganate solution. 1070902.
crystals. A 30 g/L solution of potassium permanganate R.
Test by adding to 0.5 mL of the saturated potassium iodide
solution 30 mL of a mixture of 2 volumes of chloroform R Potassium perrhenate. KReO4. (Mr 289.3). 1071000.
and 3 volumes of glacial acetic acid R, as well as 0.1 mL [10466-65-6].
of starch solution R. Any blue colour formed should be White or almost white, crystalline powder, soluble in water,
discharged by the addition of 0.05 mL of 0.1 M sodium slightly soluble in ethanol (96 per cent), in methanol and in
thiosulfate. propylene glycol.
Storage : protected from light. Potassium persulfate. K2S2O8. (Mr 270.3). 1071100.
[7727-21-1]. Dipotassium peroxodisulfate.
Potassium iodobismuthate solution. 1070600.
Colourless crystals or white or almost white, crystalline
To 0.85 g of bismuth subnitrate R add 40 mL of water R, 10 mL powder, sparingly soluble in water, practically insoluble in
of glacial acetic acid R and 20 mL of a 400 g/L solution of ethanol (96 per cent). Aqueous solutions decompose at room
potassium iodide R. temperature and more rapidly on warming.
Potassium iodobismuthate solution, dilute. 1070603. Potassium plumbite solution. 1071200.
Dissolve 100 g of tartaric acid R in 500 mL of water R and Dissolve 1.7 g of lead acetate R, 3.4 g of potassium citrate R
add 50 mL of potassium iodobismuthate solution R1. and 50 g of potassium hydroxide R in water R and dilute to
Storage : protected from light. 100 mL with the same solvent.
Potassium iodobismuthate solution R1. 1070601. Potassium pyroantimonate. KSb(OH)6. (Mr 262.9). 1071300.

[12208-13-8]. Potassium hexahydroxoantimoniate.
Dissolve 100 g of tartaric acid R in 400 mL of water R and add
8.5 g of bismuth subnitrate R. Shake for 1 h, add 200 mL of a White or almost white, crystals or crystalline powder,
400 g/L solution of potassium iodide R and shake well. Allow sparingly soluble in water.
to stand for 24 h and filter. Potassium pyroantimonate solution. 1071301.
Storage : protected from light. Dissolve 2 g of potassium pyroantimonate R in 95 mL of hot
water R. Cool quickly and add a solution containing 2.5 g
Potassium iodobismuthate solution R2. 1070602. of potassium hydroxide R in 50 mL of water R and 1 mL of
Stock solution. Suspend 1.7 g of bismuth subnitrate R and 20 g dilute sodium hydroxide solution R. Allow to stand for 24 h,
of tartaric acid R in 40 mL of water R. To the suspension add filter and dilute to 150 mL with water R.
40 mL of a 400 g/L solution of potassium iodide R and stir for
1 h. Filter. The solution may be kept for several days in brown Potassium pyroantimonate solution R1. 1071302.
bottles. Dissolve 2.0 g of potassium pyroantimonate R in 100 mL
Spray solution. Mix immediately before use 5 mL of the stock of hot water R. Boil for about 5 min, cool quickly and add
solution with 15 mL of water R. 10 mL of a 150 g/L solution of potassium hydroxide R.
Allow to stand for 24 h and filter.
Potassium iodobismuthate solution R3. 1070604.

Dissolve 0.17 g of bismuth subnitrate R in a mixture of 2 mL Potassium 4-sulfobenzoate. C7H5KO5S. (Mr 240.3). 1190000.
of glacial acetic acid R and 18 mL of water R. Add 4 g of [5399-63-3]. 4-Sulfobenzoic acid potassium salt. Potassium
potassium iodide R, 1 g of iodine R and dilute to 100 mL with 4-carboxybenzenesulfonate.
dilute sulfuric acid R. White, crystalline powder.

Potassium tartrate. C4H4K2O6,1/2H2O. (Mr 235.3). 1071400. Absorbance (2.2.25) : maximum 0.60 at 210 nm, 0.26 at
[921-53-9]. Dipotassium (2R,3R)-2,3-dihydroxybutane-1,4- 220 nm, 0.13 at 230 nm, 0.02 at 250 nm, 0.01 at 260 nm,
dioate hemihydrate. determined using water R as compensation liquid.
White or almost white, granular powder or crystals, very
2-Propanol R2. 1184900. [67-63-0].
soluble in water, very slightly soluble in ethanol (96 per cent).
See Isopropyl alcohol (0970).
Potassium tetraiodomercurate solution. 1071500.
Dissolve 1.35 g of mercuric chloride R in 50 mL of water R. Add Propetamphos. C10H20NO4PS. (Mr 281.3). 1130900.
5 g of potassium iodide R and dilute to 100 mL with water R. [31218-83-4].
Potassium tetraiodomercurate solution, alkaline. 1071600. cyclohexane) may be used.
Dissolve 11 g of potassium iodide R and 15 g of mercuric
iodide R in water R and dilute to 100 mL with the same solvent. Propidium iodide. C27H34I2N4. (Mr 668.4). 1154200.
Immediately before use, mix 1 volume of this solution with an [25535-16-4]. 3,8-Diamino-5-[3(diethylmethylammonio)-
equal volume of a 250 g/L solution of sodium hydroxide R. propyl]-6-phenylphenanthridinium diiodide.
Dark red solid.
Potassium tetroxalate. C4H3KO8,2H2O. (Mr 254.2). 1071700.
[6100-20-5]. Propionaldehyde. C3H6O. (Mr 58.1). 1072300. [123-38-6].
White or almost white, crystalline powder, sparingly soluble Propanal.
in water, soluble in boiling water, slightly soluble in ethanol Liquid freely soluble in water, miscible with ethanol (96 per
(96 per cent). cent).
Potassium thiocyanate. KSCN. (Mr 97.2). 1071800. : about 0.81.
[333-20-0]. : about 1.365.
Colourless crystals, deliquescent, very soluble in water and in bp : about 49 °C.
ethanol (96 per cent). mp : about − 81 °C.
Propionic acid. C3H6O2. (Mr 74.1). 1072400. [79-09-4].
Potassium thiocyanate solution. 1071801.
Oily liquid, soluble in ethanol (96 per cent), miscible with
A 97 g/L solution of potassium thiocyanate R. water.
Povidone. 1068500. [9003-39-8]. : about 0.993.
See Povidone (0685). : about 1.387.
Procaine hydrochloride. 1109400. bp : about 141 °C.
See Procaine hydrochloride (0050). mp : about − 21 °C.
Proline. 1152200. [147-85-3]. Propionic anhydride. C6H10O3. (Mr 130.1). 1072500.
See Proline (0785). [123-62-6].
Clear, colourless liquid, soluble in ethanol (96 per cent).
Propane. C3H8. (Mr 44.10). 1190100. [74-98-6].
: about 1.01.
bp : about 167 °C.
Propane-1,3-diol. C3H8O2. (Mr 76.1). 1185100. [504-63-2].
1,3-Dihydroxypropane. Propionic anhydride reagent. 1072501.
Colourless, viscous liquid. Dissolve 1 g of toluenesulfonic acid R in 30 mL of glacial
bp : about 214 °C. acetic acid R, add 5 mL of propionic anhydride R and allow
to stand for at least 15 min before use.
Propanol. C3H8O. (Mr 60.1). 1072000. [71-23-8].
Propan-1-ol. Propyl acetate. C5H10O2. (Mr 102.1). 1072600. [109-60-4].
Clear colourless liquid, miscible with water and with ethanol : about 0.888.
(96 per cent). bp : about 102 °C.
: about 0.802 to 0.806. mp : about − 95 °C.
bp : about 97.2 °C.
Propyl parahydroxybenzoate. 1072700. [94-13-3].
between 96 °C and 99 °C. See Propyl parahydroxybenzoate (0431).
Propanol R1. 1184400. [71-23-8]. D-Prolyl- L-phenylalanyl- L-arginine
4-nitroanilide
See Propanol (2036). dihydrochloride. C26H36Cl2N8O5. (Mr 612). 1072800.
2-Propanol. C3H8O. (Mr 60.1). 1072100. [67-63-0]. Propylene glycol. 1072900. [57-55-6].

Propan-2-ol. Isopropyl alcohol. See Propylene glycol (0430).
Clear, colourless, flammable liquid, miscible with water and Propylene oxide. C3H6O. (Mr 58.1). 1121800. [75-56-9].
Colourless liquid, miscible with ethanol (96 per cent).
: about 0.785.
bp : 81 °C to 83 °C. Protamine sulfate. 1073000. [53597-25-4 (salmine)
9007-31-2 (clupeine)].
2-Propanol R1. 1072101.
See Protamine sulfate (0569).
Complies with the requirements prescribed for
2-propanol R with the following additional requirements. Protopine hydrochloride. C20H20ClNO5. (Mr 389.8).
: about 1.378. 1163500. [6164-47-2].
Water (2.5.12) : maximum 0.05 per cent, determined on 5-Methyl-4,6,7,14-tetrahydrobis[1,3]benzodioxolo[4,5-c:5′,6′-
10 g. g]azecin-13(5H)-one hydrochloride.

Pteroic acid. C14H12N6O3. (Mr 312.3). 1144600. Procedure. Transfer 4.0 mL of substrate to a test tube and add
[119-24-4]. 4-[[(2-Amino-4-oxo-1,4-dihydropteridin-6- 0.5 mL of buffer solution A, mix, and incubate at 30 °C. Add
yl)methyl]amino]benzoic acid. 0.5 mL of pullulanase diluent and mix thoroughly. After 30 s,
Crystals, soluble in solutions of alkali hydroxides. transfer 1.0 mL of this solution to a test tube labelled “pullulan
test solution 1”, add 2.0 mL of Somogyi reagent, and mix.
Puerarin. C21H20O9. (Mr 416.4). 1180600. [3681-99-0]. After 30.5 min, transfer 1.0 mL of the mixture of substrate and
7,4′-Dihydroxy-8-C-glucosyliso-haloprone. 8-β-D- pullulanase diluent to a second test tube labelled “pullulan test
Glucopyranosyl-7-hydroxy-3-(4-hydroxyphenyl)-4H-1- solution 2”, add 2.0 mL of Somogyi reagent, and mix. In a third
benzopyran-4-one. test tube labelled “standard blank”, mix 2.0 mL of Somogyi
reagent and 1.0 mL of water R. In a fourth test tube labelled
Pulegone. C10H16O. (Mr 152.2). 1073100. [89-82-7]. “glucose standard solution”, mix 2.0 mL of Somogyi reagent
(R)-2-Isopropylidene-5-methylcyclohexanone. and 1.0 mL of glucose standard solution, and add 1.0 mL
(+)-p-Menth-4-en-3-one. of water R. Incubate the fourth test tube in a water-bath for
Oily, colourless liquid, practically insoluble in water, miscible exactly 10 min. Remove the tube and allow it to cool under
with ethanol (96 per cent). running water. Add 2.0 mL of Nelson reagent, mix well, and
: about 0.936. allow the solution to stand for at least 15 min. Add 5.0 mL
: 1.485 to 1.489. of water R to each of the 4 test tubes and mix thoroughly.
Determine the absorbance at 520 nm of the standard blank
bp : 222 °C to 224 °C. (Ablank), the glucose standard solution (AStd), pullulan test
Pulegone used in gas chromatography complies with the solution 1 (A0) and pullulan test solution 2 (A30), using water R
following additional test. as the blank. One unit is defined as the enzymatic activity
Assay. Gas chromatography (2.2.28) as prescribed in the required to produce 1 μmol of maltotriose (measured as
monograph Peppermint oil (0405). glucose) from pullulan per minute. Calculate the pullulanase
activity, P, in units/mL, using the following expression :
Content : minimum 98.0 per cent, calculated by the [(A30 - A0 ) / (AStd - Ablank )] ´ 0.185 ´ D
normalisation procedure. MEASUREMENT OF PROTEIN CONTENT (MEASURED AS
Pullulanase. 1190200. [9075-68-7]. Pullulan-6- ALBUMINOID CONTENT) FOR THE CALCULATION OF SPECIFIC
glucanohydrolase obtained from Klebsiella pneumoniae. ACTIVITY
Content : minimum 30 units/mg of protein. Reagent A. Prepare a solution having known concentrations
of about 4 g/L of sodium hydroxide R and about 21 g/L of
One unit represents the enzymatic activity required to produce
anhydrous sodium carbonate R.
1.0 μmol of maltotriose from pullulan per minute at pH 5.0 at
30 °C. Reagent B. Transfer 0.5 g of copper sulfate pentahydrate R and
DETERMINATION OF PULLULANASE ACTIVITY
1.0 g of sodium citrate R to a volumetric flask, dissolve in and
dilute with water R to 100.0 mL, and mix.
Substrate. Dissolve 0.250 g of pullulan in 20.0 mL of water R,
adding pullulan to the water. Lowry solution. Mix 50 volumes of reagent A and 1 volume
of reagent B.
Buffer solution A. 21 g/L solution of citric acid monohydrate R
adjusted to pH 5.0 with a 27 g/L solution of disodium hydrogen Diluted Folin-Ciocalteu’s phenol reagent (for albuminoid
phosphate dodecahydrate R. quantification). Prepare a two-fold dilution of the
commercially available 2 N Folin-Ciocalteu’s phenol reagent
Buffer solution B. Prepare a 136 g/L solution of sodium or prepare a solution by making an appropriate dilution of
acetate R adjusted to pH 6.0 with dilute acetic acid R. Dilute phosphomolybdotungstic reagent R.
1 mL of this solution to 100 mL with water R.
Bovine albumin standard stock solution. Transfer 50.0 mg of
Somogyi reagent. To 28 g of anhydrous disodium hydrogen
bovine albumin R to a volumetric flask, dissolve in and dilute
phosphate R and 40 g of sodium potassium tartrate R add
with water R to 500.0 mL, and mix. It contains 100 μg/mL of
about 700 mL of water R. Add 100 mL of a 42 g/L solution
bovine albumin.
of sodium hydroxide R and mix. Add 80 mL of a 100 g/L
solution of copper sulfate pentahydrate R. Heat until complete Standard solutions. Using appropriate dilutions of bovine
dissolution. Add 180 g of anhydrous sodium sulfate R and albumin standard stock solution in water R, prepare 5 standard
adjust the volume to 1 L with water R. Allow to stand at room solutions having concentrations equally spaced between
temperature for 1 or 2 days to let insoluble matter precipitate. 5 μg/mL and 100 μg/mL of bovine albumin.
Filter the solution and keep the filtrate in a brown-glass bottle Test solution. Dilute pullulanase R with buffer solution B
with a ground-glass stopper. in order to obtain a solution having a concentration of
Nelson reagent. Dissolve 50 g of ammonium molybdate R 60-70 μg/mL of albuminoid. Water may be used as diluent.
in 900 mL of water R. Add 42 g of sulfuric acid R and mix. Record the dilution factor, Df.
Dissolve 6 g of disodium arsenate R in 50 mL of water R. Add Procedure. Introduce into separate tubes 0.3 mL of each
the latter solution to the 1st solution, and allow to stand in a of the standard solutions, the test solution and water R.
brown-glass bottle with a ground-glass stopper at 37 °C for Add 3.0 mL of Lowry solution to each tube and mix.
1 or 2 days. Incubate at room temperature for 10 min. Add 0.3 mL
Glucose standard solution. Dry glucose R at a pressure less of diluted Folin-Ciocalteu’s phenol reagent to each tube,
than 6 kPa at 60 °C for 5 h, and calculate the water content. mix immediately, and allow to stand at room temperature
Transfer 10.00 g of dried glucose to a volumetric flask, dissolve for 60 min. Determine the absorbances of the standard
with water R, dilute to 1.0 L with the same solvent, and mix. solutions and the test solution at the wavelength of maximum
Transfer 10.0 mL of this solution to a volumetric flask and absorbance, about 750 nm, using water R as the blank.
dilute to 1.0 L with water R. Each millilitre contains 100 µg Calculation. The relationship of absorbance to protein
of glucose. concentration is non-linear ; however, if the standard curve
Pullulanase diluent. Dilute pullulanase R with buffer concentration range is sufficiently small, it will approach
solution B to prepare a solution with an enzyme activity of linearity. Using linear regression method, plot the absorbances
about 0.2 units/mL. The measurement range is between 0.1 of the standard solutions versus the protein (bovine albumin)
and 0.4 units/mL. Record the dilution factor (D). This diluent concentrations, in µg/mL. Using the plot, determine the
is used as a diluted enzyme solution. concentration of protein (albuminoid content), Calbuminoid,

in µg/mL, in the test solution. Calculate the albuminoid Pyrocatechol. C6H6O2. (Mr 110.1). 1073600. [120-80-9].
concentration, in mg/mL, in pullulanase R using the following Benzene-1,2-diol.
expression : Colourless or slightly yellow crystals, soluble in water, in
C protein = (Calbuminoid ´ Df ) / 1000
acetone and in ethanol (96 per cent).
mp : about 102 °C.
Calculate the specific activity, in units/mg, of pullulanase
using the formula : Storage : protected from light.
P / C protein Pyrogallol. C6H6O3. (Mr 126.1). 1073700. [87-66-1].

Benzene-1,2,3-triol.
P = pullulanase activity in units/mL.
White or almost white crystals, becoming brownish on
Putrescine. C4H12N2. (Mr 88.15). 1137900. [110-60-1]. exposure to air and light, very soluble in water and in ethanol
1,4-Butanediamine. Tetramethylenediamine. (96 per cent), slightly soluble in carbon disulfide. On exposure
Colourless oily liquid, very soluble in water. Strong to air, aqueous solutions, and more rapidly alkaline solutions,
piperidine-like odour. become brown owing to the absorption of oxygen.
mp : about 23 °C. Storage : protected from light.
Pyrazine-2-carbonitrile. C5H3N3. (Mr 105.1). 1183300. Pyrogallol solution, alkaline. 1073701.

[19847-12-2]. 2-Cyanopyrazine. Dissolve 0.5 g of pyrogallol R in 2 mL of carbon dioxide-free
Clear, pale yellow liquid. water R. Dissolve 12 g of potassium hydroxide R in 8 mL
Content : minimum 99 per cent. of carbon dioxide-free water R. Mix the two solutions
Pyridin-2-amine. C5H6N2. (Mr 94.1). 1073400. [504-29-0].
2-Aminopyridine. Pyrrolidine. C4H9N. (Mr 71.1). 1165000. [123-75-1].
Large crystals soluble in water and in ethanol (96 per cent). Content : minimum 99 per cent.
bp : about 210 °C. bp : 87 °C to 88 °C.
mp : about 58 °C.
2-Pyrrolidone. C4H7NO. (Mr 85.1). 1138000. [616-45-5].
Pyridine. C5H5N. (Mr 79.1). 1073200. [110-86-1]. Pyrrolidin-2-one.
Clear, colourless liquid, hygroscopic, miscible with water and Liquid above 25 °C, miscible with water, with anhydrous
with ethanol (96 per cent). ethanol and with ethyl acetate.
bp : about 115 °C. : 1.116.
Storage : in an airtight container. Water (2.5.12): maximum 0.2 per cent determined on 2.00 g.
Pyridine, anhydrous. 1073300. Assay. Gas chromatography (2.2.28) : use the normalisation
procedure.
Dry pyridine R over anhydrous sodium carbonate R. Filter
and distil. Test solution. Dissolve 1.0 g in methanol R and dilute to
Water (2.5.12) : maximum 0.01 per cent m/m. 10.0 mL with the same solvent.
Column :
Pyridine-4-carbonitrile. C6H4N2. (Mr 104.1). 1190300.
– material : glass ;
[100-48-1]. 4-Cyanopyridine.
White or almost white crystalline powder. – size : l = 30 m ; Ø = 0.53 mm ;
bp : 194 °C to 196 °C. – stationary phase : macrogol 20 000 R (1.0 μm).
mp : 76 °C to 79 °C. Carrier gas : helium for chromatography R.
Flow rate : adjusted so that the retention time of 2-pyrrolidone
Pyridinium hydrobromide perbromide. C5H6Br3N. is about 10 min.
(Mr 319.8). 1166100. [39416-48-3]. Pyridinium
tribromide(1-). Split ratio : 1:20.
Red crystals. Temperature :
Time Temperature
Pyridylazonaphthol. C15H11N3O. (Mr 249.3). 1073500.
(min) (°C)
[85-85-8]. 1-(2-Pyridylazo)-2-naphthol.
Column 0-1 80
Brick-red powder, practically insoluble in water, soluble in
ethanol (96 per cent), in methanol and in hot dilute alkali 1 - 12 80 → 190
solutions. 12 - 32 190
mp : about 138 °C.
Injection port 200
Pyridylazonaphthol solution. 1073501.
A 1 g/L solution of pyridylazonaphthol R in anhydrous Detection : flame ionisation.
ethanol R. Injection : 1 µL of the test solution.
Test for sensitivity. To 50 mL of water R add 10 mL of Content : minimum 98.0 per cent.
acetate buffer solution pH 4.4 R, 0.10 mL of 0.02 M sodium
edetate and 0.25 mL of the pyridylazonaphthol solution. Pyruvic acid. C3H4O3. (Mr 88.1). 1109300. [127-17-3].
After addition of 0.15 mL of a 5 g/L solution of copper 2-Oxopropanoic acid.
sulfate pentahydrate R, the colour changes from light yellow Yellowish liquid, miscible with water and with anhydrous
to violet. ethanol.
4-(2-Pyridylazo)resorcinol monosodium salt. : about 1.267.
C11H8N3NaO2, H2O. (Mr 255.2). 1131500. [16593-81-0]. : about 1.413.
Orange crystalline powder. bp : about 165 °C.

Quercetin dihydrate. C15H10O7,2H2O. (Mr 338.2). 1138100. Quinine. C20H24N2O2. (Mr 324.4). 1074100. [130-95-0].
2-(3,4-Dihydroxyphenyl)-3,5,7-trihydroxy-4H-1-benzopyran- (R)-(6-Methoxyquinol-4-yl)[(2S,4S,5R)-5-vinylquinuclidin-2-
4-one. yl]methanol.
Yellow crystals or yellowish powder, practically insoluble in White or almost white, microcrystalline powder, very slightly
water, soluble in acetone and in methanol. soluble in water, slightly soluble in boiling water, very soluble
Water (2.5.12): maximum 12.0 per cent, determined on in anhydrous ethanol.
0.100 g. : about − 167, determined on a 10 g/L solution in
Assay. Liquid chromatography (2.2.29) as prescribed in the anhydrous ethanol R.
monograph Ginkgo leaf (1828). mp : about 175 °C.
Content :minimum 90 per cent (anhydrous substance) Storage : protected from light.
calculated by the normalisation procedure. Quinine hydrochloride. 1074200. [6119-47-7].
Storage : protected from light. See Quinine hydrochloride (0018).
Quercitrin. C21H20O11. (Mr 448.4). 1138200. Quinine sulfate. 1074300. [6119-70-6].
[522-12-3]. Quercetin 3-L-rhamnopyranoside. See Quinine sulfate (0019).
3-[(6-Deoxy-α-L-mannopyranosyl)oxy]-2-(3,4-
dihydroxyphenyl)-5,7-dihydroxy-4H-1-benzopyran-4-one. 3-Quinuclidinol. C7H13NO. (Mr 127.2). 1193800.
Quercitroside. [1619-34-7]. (3R)-1-Azabicyclo[2.2.2]octan-3-ol.
Yellow crystals, practically insoluble in cold water, soluble in Content : minimum 99 per cent.
ethanol (96 per cent). Light yellow powder.
mp : 176 °C to 179 °C. Rabbit erythrocyte suspension. 1074500.
Chromatography. Thin-layer chromatography (2.2.27) as Prepare a 1.6 per cent V/V suspension of rabbit erythrocytes
prescribed in the monograph Goldenrod (1892) : apply 20 µL as follows : defibrinate 15 mL of freshly drawn rabbit blood by
of the solution ; after spraying, the chromatogram shows a shaking with glass beads, centrifuge at 2000 g for 10 min and
yellowish-brown fluorescent zone with an RF of about 0.6. wash the erythrocytes with three quantities, each of 30 mL,
Storage : at a temperature of 2 °C to 8 °C. of a 9 g/L solution of sodium chloride R. Dilute 1.6 mL of
the suspension of erythrocytes to 100 mL with a mixture of
Quillaia saponins, purified. 1184500. 1 volume of phosphate buffer solution pH 7.2 R and 9 volumes
A mixture of related saponins obtained from the bark of of a 9 g/L solution of sodium chloride R.
Quillaja saponaria Molina s.l.
Raclopride tartrate. C19H26Cl2N2O9. (Mr 497.3). 1144700.
Chromatography. Thin-layer chromatography (2.2.27) as [98185-20-7]. Raclopride L-tartrate.
prescribed in the monograph Quillaia bark (1843) : apply 5 µL
of the solution ; after treating with a 10 per cent V/V solution White or almost white solid, sensitive to light, soluble in water.
of sulfuric acid R in methanol R, heat at 120 °C for 5 min and : + 0.3, determined on a 3 g/L solution.
examine in daylight ; the chromatogram shows 3 principal mp : about 141 °C.
zones in the upper part of the middle third.
Raffinose pentahydrate. C18H32O16,5H2O. (Mr 594.5).
Quinaldine red. C21H23IN2. (Mr 430.3). 1073800. [117-92-0]. 1201800. [17629-30-0]. β-D-Fructofuranosyl
2-[2-[4-(Dimethylamino)phenyl]ethenyl]-1-ethylquinolinium α-D-galactopyranosyl-(1→6)-α-D-glucopyranoside
iodide. pentahydrate.
Dark bluish-black powder, sparingly soluble in water, freely Content : minimum 98.0 per cent.
soluble in ethanol (96 per cent). Crystalline powder.
mp : about 80 °C.
Quinaldine red solution. 1073801.
Dissolve 0.1 g of quinaldine red R in methanol R and dilute Raltegravir potassium. C20H20FKN6O5. 1202600.
to 100 mL with the same solvent. [871038-72-1].
Colour change : pH 1.4 (colourless) to pH 3.2 (red). See Raltegravir potassium (2887).
Quinhydrone. C12H10O4. (Mr 218.2). 1073900. [106-34-3]. Rapeseed oil. 1074600.

Equimolecular compound of 1,4-benzoquinone and See Rapeseed oil, refined (1369).
hydroquinone. Reducing mixture. 1074700.
Dark green, lustrous crystals or a crystalline powder, slightly Grind the substances added in the following order to obtain a
soluble in water, sparingly soluble in hot water, soluble in homogeneous mixture : 20 mg of potassium bromide R, 0.5 g
ethanol (96 per cent) and in concentrated ammonia. of hydrazine sulfate R and 5 g of sodium chloride R.
mp : about 170 °C.
Reichstein’s substance S. C21H30O4. (Mr 346.5). 1175400.
Quinidine. C20H24N2O2. (Mr 324.4). 1074000. [56-54-2]. [152-58-9].
(S)-(6-Methoxyquinol-4-yl)[(2R,4S,5R)-5-vinylquinuclidin-2- Content : minimum 95.0 per cent.
yl]methanol. mp : about 208 °C.
White or almost white crystals, very slightly soluble in water,
sparingly soluble in ethanol (96 per cent), slightly soluble in Resin for hydrophobic interaction chromatography.
methanol. 1202700.
: about + 260, determined on a 10 g/L solution in Non-porous resin consisting of spherical polymethacrylate
anhydrous ethanol R. particles bonded with butyl groups.
pH limits of use : 2 to 12.
mp : about 172 °C.
Storage : protected from light. Resin for reversed-phase ion chromatography. 1131100.
A neutral, macroporous, high specific surface area with a
Quinidine sulfate. 1109500. [6591-63-5]. non-polar character resin consisting of polymer lattice of
See Quinidine sulfate (0017). polystyrene cross-linked with divinylbenzene.

Resin, weak cationic. 1096000. : about 1.472.

See weak cationic resin R. mp : about 285 °C, with decomposition.
Resorcinol. 1074800. [108-46-3]. Rosmarinic acid. C18H16O8. (Mr 360.3). 1138300.
See Resorcinol (0290). [20283-92-5].
mp : 170 °C to 174 °C.
Resorcinol reagent. 1074801.
Rutecarpine. C18H13N3O. (Mr 287.3). 1199500. [84-26-4].
To 80 mL of hydrochloric acid R1 add 10 mL of a 20 g/L 8,13-Dihydroindolo[2′,3′:3,4]pyrido[2,1-b]quinazolin-5(7H)-
solution of resorcinol R and 0.25 mL of a 25 g/L solution of one.
copper sulfate pentahydrate R and dilute to 100.0 mL with
water R. Prepare the solution at least 4 h before use. Ruthenium red. [(NH3)5RuORu(NH3)4ORu(NH3)5]Cl6,4H2O.
Storage : at 2 °C to 8 °C for 1 week. (Mr 858). 1075200. [11103-72-3].
Brownish-red powder, soluble in water.
Resveratrol. C14H12O3. (Mr 228.2). 1186900.
[501-36-0]. 3,4′,5-Stilbenetriol. 5-[(E)-2-(4-Hydroxy- Ruthenium red solution. 1075201.
phenyl)ethenyl]benzene-1,3-diol. A 0.8 g/L solution of ruthenium red R in lead acetate
solution R.
Rhamnose. C6H12O5,H2O. (Mr 182.2). 1074900. [6155-35-7].
(2R,3R,4R,5R,6S)-6-Methyltetrahydro-2H-pyran-2,3,4,5-tetrol Rutin. 1075300. [250249-75-3].
monohydrate. 6-Deoxy-α-L-mannopyranose monohydrate. See Rutoside trihydrate R.
α-L-Rhamnopyranose monohydrate. L-(+)-Rhamnose
monohydrate. Rutoside trihydrate. 1075300. [250249-75-3].
White or almost white, crystalline powder, freely soluble in See Rutoside trihydrate (1795).
water. Sabinene. C10H16. (Mr 136.2). 1109700. [3387-41-5]. Thuj-
: + 7.8 to + 8.3, determined on a 50 g/L solution in 4(10)-ene. 4-Methylene-1-isopropylbicyclo[3.1.0]hexane.
water R containing about 0.05 per cent of NH3. A colourless, oily liquid.
Rhaponticin. C21H24O9. (Mr 420.4). 1075000. [155-58-8]. Sabinene used in gas chromatography complies with the
3-Hydroxy-5-[2-(3-hydroxy-4-methoxyphenyl)ethenyl]phenyl following additional test.
β-D-glucopyranoside. Assay. Gas chromatography (2.2.28) as prescribed in the
Yellowish-grey, crystalline powder, soluble in ethanol (96 per monograph Bitter-orange-flower oil (1175).
cent) and in methanol. Test solution. The substance to be examined.
Chromatography. Thin-layer chromatography (2.2.27) Content : minimum 95.0 per cent, calculated by the
as prescribed in the monograph Rhubarb (0291) ; the normalisation procedure.
chromatogram shows only one principal spot. Saccharin sodium. 1131400. [128-44-9].
Rhein. C15H8O6. (Mr 284.2). 1197700. [478-43-3]. 4,5- See Saccharin sodium (0787).
Dihydroxy-9,10-dioxo-9,10-dihydroanthracene-2-carboxylic
acid. 1,8-Dihydroxy-3-carboxyanthraquinone. Safrole. C10H10O2. (Mr 162.2). 1131200. [94-59-7].
5-(Prop-2-enyl)-1,3-benzodioxole. 4-Allyl-1,2-
Rhodamine 6 G. C28H31ClN2O3. (Mr 479.0). 1153300. (methylenedioxy)benzene.
[989-38-8]. Colourless or slightly yellow, oily liquid, with the odour of
Colour Index No. 45160. sassafras, insoluble in water, very soluble in ethanol (96 per
9-[2-(Ethoxycarbonyl)phenyl]-3,6-bis(ethylamino)-2,7- cent), miscible with hexane.
dimethylxanthenylium chloride. : 1.095 to 1.096.
Brownish-red powder. : 1.537 to 1.538.
bp : 232 °C to 234 °C.
Rhodamine B. C28H31ClN2O3. (Mr 479.0). 1075100. [81-88-9].
Freezing point : about 11 °C.
Schultz No. 864.
Safrole used in gas chromatography complies with the following
Colour Index No. 45170. additional test.
[9-(2-Carboxyphen-yl)-6-(diethylamino)-3H-xanthen-3-
ylidene]diethylammonium chloride.
monograph Cinnamon bark oil, Ceylon (1501).
Green crystals or reddish-violet powder, very soluble in water Content : minimum 96.0 per cent, calculated by the
and in ethanol (96 per cent). normalisation procedure.
Rhynchophylline. C22H28N2O4. (Mr 384.5). 1197800. Saikosaponin A. C42H68O13. (Mr 781). 1201900. [20736-09-8].
[76-66-4]. Methyl (16E)-17-methoxy-2-oxo-16,17-didehydro- 13,28-Epoxy-16β,23-dihydroxy-4α-olean-11-en-3β-yl
7β,20α-corynoxan-16-carboxylate. Methyl (16E)-16- 6-deoxy-3-O-β-D-glucopyranosyl-β-D-galactopyranoside.
(methoxymethylidene)-2-oxo-7β,20α-corynoxan-17-oate.
Saikosaponin D. C42H68O13. (Mr 781). 1201200. [20874-52-6].
Ribose. C5H10O5. (Mr 150.1). 1109600. [50-69-1]. D-Ribose. 13,28-Epoxy-16α,23-dihydroxy-4α-olean-11-en-3β-yl
Soluble in water, slightly soluble in ethanol (96 per cent). 6-deoxy-3-O-β-D-glucopyranosyl-β-D-galactopyranoside.
mp : 88 °C to 92 °C. Salicin. C13H18O7. (Mr 286.3). 1131300. [138-52-3].
Ricinoleic acid. C18H34O3. (Mr 298.5). 1100100. [141-22-0]. 2-(Hydroxymethyl)phenyl-β- D-glucopyranoside. Salicoside.
(9Z,12R)-12-Hydroxyoctadec-9-enoic acid. 12-Hydroxyoleic : − 62.5 ± 2.
acid. mp : 199 °C to 201 °C.
Yellow or yellowish-brown viscous liquid, consisting of a Assay. Liquid chromatography (2.2.29) as prescribed in the
mixture of fatty acids obtained by the hydrolysis of castor monograph Willow bark (1583) at the concentration of the
oil, practically insoluble in water, very soluble in anhydrous reference solution.
ethanol. Content : minimum 99.0 per cent, calculated by the
: about 0.942. normalisation procedure.

Salicylaldehyde. C7H6O2. (Mr 122.1). 1075400. [90-02-8]. : 6.7, determined with a solution in anhydrous ethanol.
2-Hydroxybenzaldehyde. bp19 mm : 218 °C to 220 °C.
Clear, colourless, oily liquid. mp : 96 °C to 98 °C.
: about 1.167. Sclareol used in the chromatographic profile test in the
: about 1.574. monograph Clary sage oil (1850) complies with the following
additional test.
bp : about 196 °C. Assay. Gas chromatography (2.2.28) as prescribed in the
mp : about − 7 °C. monograph Clary sage oil (1850).
Salicylaldehyde azine. C14H12N2O2. (Mr 240.3). 1075500. normalisation procedure.
[959-36-4]. 2,2′-Azinodimethyldiphenol.
Dissolve 0.30 g of hydrazine sulfate R in 5 mL of water R, add Scopoletin. C10H8O4. (Mr 192.2). 1158700. [92-61-5].
1 mL of glacial acetic acid R and 2 mL of a freshly prepared 7-Hydroxy-6-methoxy-2H-1-benzopyran-2-one.
20 per cent V/V solution of salicylaldehyde R in 2-propanol R. 7-Hydroxy-6-methoxycoumarin.
Mix, allow to stand until a yellow precipate is formed. Shake Faintly beige, fine crystals.
with two quantities, each of 15 mL, of methylene chloride R. mp : 202 °C to 208 °C.
Combine the organic layers and dry over anhydrous sodium
sulfate R. Decant or filter the solution and evaporate to SDS-PAGE running buffer. 1114900.
dryness. Recrystallise from a mixture of 40 volumes of Dissolve 151.4 g of tris(hydroxymethyl)aminomethane R,
methanol R and 60 volumes of toluene R with cooling. Dry the 721.0 g of glycine R and 50.0 g of sodium laurilsulfate R
crystals in vacuo. in water R and dilute to 5000 mL with the same solvent.
Immediately before use, dilute to 10 times its volume with
mp : about 213 °C. water R and mix. Measure the pH (2.2.3) of the diluted
Chromatography. Thin-layer chromatography (2.2.27) solution. The pH is between 8.1 and 8.8.
as prescribed in the test for hydrazine in the monograph
Povidone (0685) ; the chromatogram shows only one principal SDS-PAGE sample buffer (concentrated). 1115000.
spot. Dissolve 1.89 g of tris(hydroxymethyl)aminomethane R, 5.0 g
of sodium laurilsulfate R and 50 mg of bromophenol blue R in
Salicylic acid. 1075600. [69-72-7]. water R. Add 25.0 mL of glycerol R and dilute to 100 mL with
See Salicylic acid (0366). water R. Adjust the pH to 6.8 with hydrochloric acid R, and
Salvianolic acid B. C36H30O16. (Mr 719). 1184600. SDS-PAGE sample buffer for reducing conditions
[121521-90-2]. (2R)-2-[[(2E)-3-[(2S,3S)-3-[[(1R)-1- (concentrated). 1122100.
Carboxy-2-(3,4-dihydroxyphenyl)ethoxy]carbonyl]-2-(3,4-
dihydroxyphenyl)-7-hydroxy-2,3-dihydrobenzofuran-4- Dissolve 3.78 g of tris(hydroxymethyl)aminomethane R, 10.0 g
yl]prop-2-enoyl]oxy]-3-(3,4-dihydroxyphenyl)propanoic acid. of sodium dodecyl sulfate R and 100 mg of bromophenol blue R
in water R. Add 50.0 mL of glycerol R and dilute to 200 mL
Sand. 1075800. with water R. Add 25.0 mL of 2-mercaptoethanol R. Adjust to
pH 6.8 with hydrochloric acid R, and dilute to 250.0 mL with
White or slightly greyish grains of silica with a particle size water R.
between 150 µm and 300 µm.
Alternatively, dithiothreitol may be used as reducing
Sarafloxacin hydrochloride. C20H18ClF2N3O3. (Mr 421.8). agent instead of 2-mercaptoethanol. In this case
1181400. [91296-87-6]. 6-Fluoro-1-(4-fluorophenyl)-4-oxo- prepare the sample buffer as follows : dissolve 3.78 g of
7-piperazin-1-yl-1,4-dihydroquinoline-3-carboxylic acid tris(hydroxymethyl)aminomethane R, 10.0 g of sodium dodecyl
hydrochloride. sulfate R and 100 mg of bromophenol blue R in water R. Add
50.0 mL of glycerol R and dilute to 200 mL with water R. Adjust
Schisandrin. C24H32O7. (Mr 432.5). 1173800. to pH 6.8 with hydrochloric acid R, and dilute to 250.0 mL
[7432-28-2]. Schisandrol A. Wuweizichun A. with water R. Immediately before use, add dithiothreitol R to a
(6S,7S,12aRa)-5,6,7,8-Tetrahydro-1,2,3,10,11,12- final concentration of 100 mM.
hexamethoxy-6,7-dimethyldibenzo[a,c]cyclooctan-6-ol.
Selenious acid. H2SeO3. (Mr 129.0). 1100200. [7783-00-8].
White or almost white, crystalline powder. Deliquescent crystals, freely soluble in water.
Schisandrin used in the assay in the monograph Schisandra Storage : in an airtight container.
fruit (2428) complies with the following additional test.
Selenium. Se. (Ar 79.0). 1075900. [7782-49-2].
monograph Schisandra fruit (2428). Brown-red or black powder or granules, practically insoluble
in water and in ethanol (96 per cent), soluble in nitric acid.
normalisation procedure. mp : about 220 °C.
Storage : in an airtight container, at − 20 °C or below. Sennoside B. C42H38O20. (Mr 863). 1190400. [128-57-4].
(9R,9′S)-5,5′-Bis(β-D-glucopyranosyloxy)-4,4′-dihydroxy-
γ-Schisandrin. C23H28O6. (Mr 400.5). 1173900. 10,10′-dioxo-9,9′,10,10′-tetrahydro-9,9′-bianthracene-2,2′-
[61281-37-6]. Schisandrin B. Wuweizisu B. rac- dicarboxylic acid.
(6R,7S,13aRa)-1,2,3,13-Tetramethoxy-6,7-dimethyl-5,6,7,8- Pale yellow crystals, practically insoluble in water, very slightly
tetrahydrobenzo[3,4]cycloocta[1,2-f][1,3]benzodioxole. soluble in ethanol (96 per cent), soluble in dilute solutions of
White or almost white, crystalline powder. alkali hydroxides.
Storage : in an airtight container, at − 20 °C or below. mp : 180 °C to 186 °C.
Serine. 1076000. [56-45-1].
Sclareol. C20H36O2. (Mr 308.5). 1139900. [515-03-7].
(1R,2R,4aS,8aS)-1-[(3R)-3-Hydroxy-3-methylpent-4-enyl]- See Serine (0788).
2,5,5,8a-tetramethyldecahydronaphthalen-2-ol. Sialic acid. 1001100. [131-48-6].
Odourless crystals. See N-acetylneuraminic acid R.

Silibinin. C25H22O10. (Mr 482.4). 1151400. [22888-70-6]. Silica gel for chiral separation, vancomycin-bonded.
Silybin. (2R,3R)-3,5,7-Trihydroxy-2-[(2R,3R)-3-(4-hydroxy- 1205300.
3-methoxyphenyl)-2-(hydroxymethyl)-2,3-dihydro-1,4- High-purity silica gel chemically modified by the bonding of
benzodioxin-6-yl]-2,3-dihydro-4H-1-benzopyran-4-one. vancomycin through multiple covalent linkages.
White or yellowish powder, practically insoluble in water,
soluble in acetone and in methanol. Silica gel for chromatography. 1076900.
Silibinin used in the assay of Milk thistle fruit (1860) complies A very finely divided silica gel.
with the following additional test. Silica gel for chromatography, alkyl-bonded for use with
Assay. Liquid chromatography (2.2.29) as prescribed in the highly aqueous mobile phases. 1160200.
monograph Milk thistle fruit (1860). A very finely divided silica gel with bonded alkyl groups
Test solution. Dissolve 5.0 mg of silibinin, dried in vacuo, in suitable for use with highly aqueous mobile phases.
methanol R and dilute to 50.0 mL with the same solvent.
Silica gel for chromatography, alkyl-bonded for use with
Silibinin A and silibinin B content : minimum 95.0 per cent, highly aqueous mobile phases, end-capped. 1176900.
calculated by the normalisation procedure.
A very finely divided silica gel with bonded alkyl groups
Silica for chromatography, porous. 1207800. suitable for use with highly aqueous mobile phases. To
Porous silica with porous layer open tubular (PLOT) design. minimise any interaction with basic compounds it is carefully
end-capped to cover most of the remaining silanol groups.
Silica gel π-acceptor/π-donor for chiral separations.
1160100. Silica gel for chromatography, alkylsilyl, solid core,
A very finely divided silica gel for chromatography consisting
of spherical particles to which 1-(3,5-dinitrobenzamido)- Silica gel with spherical silica particles containing a
1,2,3,4-tetrahydrophenantrene has been covalently bound, non-porous solid silica core surrounded by a thin outer
showing both π-electron acceptor and π-electron donor porous silica coating with alkylsilyl groups. To minimise any
characteristics. interaction with basic compounds, it is carefully end-capped

to cover most of the remaining silanol groups.
Silica gel AGP for chiral chromatography. 1148700. Silica gel for chromatography, amidoalkylsilyl. 1205400.
See α1-Acid-glycoprotein silica gel for chiral separation R. A very finely divided silica gel, chemically modified at the
surface by the bonding of amidoalkylsilyl groups.
Silica gel, anhydrous. 1076100. [112926-00-8].
Silica gel for chromatography, amidohexadecylsilyl.
Partly dehydrated polymerised, amorphous silicic acid,
1170400.
absorbing at 20 °C about 30 per cent of its mass of water.
Practically insoluble in water, partly soluble in solutions A very finely divided silica gel with a fine particle size,
of sodium hydroxide. It contains a suitable indicator for chemically modified at the surface by the bonding of
detection of the humidity status, for which the colour change amidohexadecylsilyl groups.
from the hydrated to anhydrous form is given on the label. Silica gel for chromatography, amidohexadecylsilyl,
Silica gel BC for chiral chromatography. 1161300. end-capped. 1201100.
A very finely divided silica gel for chromatography (5 µm) A very finely divided silica gel, chemically modified at the
coated with β-cyclodextrin. Higher selectivity may be obtained surface by the bonding of amidohexadecylsilyl groups. To
when cyclodextrin has been derivatized with propylene oxide. minimise any interaction with basic compounds, it is carefully
Silica gel for chiral chromatography, urea type. 1181000.
Silica gel for chromatography, aminopropylmethylsilyl.
A very finely divided silica gel (5 μm) coated with the 1102400.
following derivative :
Silica gel with a fine particle size, chemically modified by
bonding aminopropylmethylsilyl groups on the surface.
Silica gel for chromatography, aminopropylsilyl. 1077000.
Silica gel with a fine particle size, chemically modified by
bonding aminopropylsilyl groups on the surface.
Silica gel for chromatography, aminopropylsilyl R1.
Silica gel for chiral separation, amylose derivative of. 1077001.
1171700.
Silica gel with a particle size of about 55 µm, chemically
Substituted amylose coated on a very finely divided silica gel modified by bonding aminopropylsilyl groups on the surface.
for chromatography.
Silica gel for chromatography, amylose derivative of.
Silica gel for chiral separation, cellulose derivative of. 1109800.
1110300. A very finely divided (10 µm) silica gel, chemically modified
Substituted cellulose coated on a very finely divided silica gel at the surface by the bonding of an amylose derivative.
for chromatography.
Silica gel for chromatography, butylsilyl. 1076200.
Silica gel for chiral separation, human albumin coated. A very finely divided silica gel, chemically modified at the
1138500. surface by the bonding of butylsilyl groups.
A very finely divided silica gel, chemically modified at the
surface by the bonding of human albumin. Silica gel for chromatography, butylsilyl, end-capped.
1170500.
Silica gel for chiral separation, protein derivative of. A very finely divided silica, chemically modified at the
1196300. surface by the bonding of butylsilyl groups. To minimise any
A very finely divided silica gel for chromatography consisting interaction with basic compounds, it is carefully end-capped
of spherical particles coated with a protein derivative. to cover most of the remaining silanol groups.

Silica gel for chromatography compatible with 100 per cent Silica gel for chromatography, hexadecylamidylsilyl.
aqueous mobile phases, octadecylsilyl. 1203900. 1162500.
A very finely divided silica gel with bonded octadecylsilyl A very finely divided (5 μm) silica gel, chemically
groups suitable for use with highly aqueous mobile phases modified at the surface by the introduction of
including 100 per cent aqueous phases. hexadecylcarboxamidopropyldimethylsilyl groups.
Silica gel for chromatography compatible with 100 per Silica gel for chromatography, hexadecylamidylsilyl,
cent aqueous mobile phases, octadecylsilyl, end-capped. end-capped. 1172400.
1188400.
A very finely divided silica gel with bonded octadecylsilyl A very finely divided (5 μm) silica gel, chemically
groups suitable for use with highly aqueous mobile phases modified at the surface by the introduction of
including 100 per cent aqueous phases. To minimise any hexadecylcarboxamidopropyldimethylsilyl groups. To
interaction with basic compounds it is carefully end-capped to minimise any interaction with basic compounds it is carefully
cover most of the remaining silanol groups. end-capped to cover most of the remaining silanol groups.
Silica gel for chromatography compatible with highly Silica gel for chromatography, hexylsilyl. 1077100.
aqueous mobile phases, octadecylsilyl diol, end-capped. A very finely divided silica gel, chemically modified at the
1207500. surface by the bonding of hexylsilyl groups.
A very finely divided silica gel, chemically modified at
the surface by the bonding of octadecylsilyl groups and Silica gel for chromatography, hexylsilyl, end-capped.
end-capping. Free diol groups are also present. For use with 1174400.
highly aqueous mobile phases. A very finely divided silica gel, chemically modified at the
Silica gel for chromatography, cyanopropylsilyl, surface by the bonding of hexylsilyl groups. To minimise any
end-capped, base-deactivated. 1194200. interaction with basic compounds it is carefully end-capped to
A very finely divided silica gel, pre-treated before the bonding cover most of the remaining silanol groups.
of cyanopropylsilyl groups by washing and hydrolysing
Silica gel for chromatography, human albumin coated.
most of the superficial siloxane bridges. To minimise any
1138500.
interaction with basic compounds, it is carefully end-capped
to cover most of the remaining silanol groups. A very finely divided silica gel, chemically modified at the
surface by the bonding of human albumin.
Silica gel for chromatography, cyanosilyl. 1109900.
A very finely divided silica gel chemically modified at the Silica gel for chromatography (hybrid material),
surface by the bonding of cyanosilyl groups. octadecylsilyl, ethylene-bridged, charged surface,
Silica gel for chromatography, cyanosilyl, end-capped.
1195000. Synthetic, spherical ethylene-bridged hybrid particles with a
charged surface, containing both inorganic (silica) and organic
A very finely divided silica gel chemically modified at the (organosiloxanes) components, chemically modified at the
surface by the bonding of cyanosilyl groups. To minimise any surface by the bonding of octadecylsilyl groups. To minimise
interaction with basic compounds it is carefully end-capped to any interaction with basic compounds they are carefully
cover most of the remaining silanol groups. end-capped to cover most of the remaining silanol groups.
Silica gel for chromatography, di-isobutyloctadecylsilyl.
1140000. Silica gel for chromatography (hybrid material),
octadecylsilyl, ethylene-bridged, end-capped. 1190500.
A very finely divided silica gel chemically modified at the
surface by the bonding of di-isobutyloctadecylsilyl groups. Synthetic, spherical, ethylene-bridged hybrid particles,
containing both inorganic (silica) and organic
Silica gel for chromatography, diisopropylcyanosilyl. (organosiloxanes) components, chemically modified
1168100. at the surface by the bonding of octadecylsilyl groups. To
A very finely divided silica gel chemically modified at the minimise any interaction with basic compounds, they are
surface by the bonding of diisopropylcyanosilyl groups. carefully end-capped to cover most of the remaining silanol
groups.
Silica gel for chromatography, 4-dimethylaminobenzylcar-
bamidesilyl. 1204000. Silica gel for chromatography (hybrid material),
A very finely divided silica gel, chemically modified at the phenylhexylsilyl, ethylene-bridged, charged surface,
surface by the bonding of 4-dimethylaminobenzylcarbamide end-capped. 1204100.
groups.
Synthetic, spherical ethylene-bridged hybrid particles with
Silica gel for chromatography, dimethyloctadecylsilyl. a charged surface, containing both inorganic (silica) and
1115100. organic (organosiloxanes) components, chemically modified
A very finely divided silica gel, chemically modified at the at the surface by the bonding of phenylhexylsilyl groups. To
surface by the bonding of dimethyloctadecylsilyl groups. minimise any interaction with basic compounds they are
carefully end-capped to cover most of the remaining silanol
Specific surface area : 300 m /g.
2
groups.
Silica gel for chromatography, diol. 1110000.
Silica gel for chromatography (hybrid material),
Spherical silica particles to which dihydroxypropyl groups are phenylsilyl, ethylene-bridged, end-capped. 1200700.
bonded. Pore size 10 nm.
Synthetic, spherical, ethylene-bridged hybrid particles,
Silica gel for chromatography, dodecylsilyl, end-capped. containing both inorganic (silica) and organic
1179700. (organosiloxanes) components, chemically modified
A very finely divided silica gel, chemically modified at at the surface by the bonding of phenylsilyl groups. To
the surface by the introduction of dodecylsilyl groups. To minimise any interaction with basic compounds, they are
minimise any interaction with basic compounds, it is carefully carefully end-capped to cover most of the remaining silanol
end-capped to cover most of the remaining silanol groups. groups.

Silica gel for chromatography (hybrid material), Silica gel for chromatography, octadecylsilyl, cross-linked,
polar-embedded, octadecylsilyl, ethylene-bridged, end-capped. 1204200.
end-capped. 1200800. A very finely divided silica gel, chemically modified at the
Synthetic, spherical, ethylene-bridged hybrid particles surface by the cross-linking and bonding of octadecylsilyl
containing both inorganic (silica) and organic groups. To minimise any interaction with basic compounds it
(organosiloxanes) components, chemically modified is carefully end-capped to cover most of the remaining silanol
at the surface by the bonding of polar-embedded octadecylsilyl groups.
groups. To minimise any interaction with basic compounds,
they are carefully end-capped to cover most of the remaining Silica gel for chromatography, octadecylsilyl, end-capped.
silanol groups. 1115400.
Silica gel for chromatography, hydrophilic. 1077200. A very finely divided silica gel, chemically modified at
the surface by the bonding of octadecylsilyl groups. To
A very finely divided silica gel whose surface has been minimise any interaction with basic compounds it is carefully
modified to provide hydrophilic characteristics. end-capped to cover most of the remaining silanol groups.
Silica gel for chromatography, nitrile. 1077300.
Silica gel for chromatography, octadecylsilyl,
A very finely divided silica gel, chemically modified at the end-capped R1. 1115401.
surface by the bonding of cyanopropylsilyl groups.
A very finely divided ultrapure silica gel, chemically modified
Silica gel for chromatography, nitrile R1. 1077400. at the surface by the bonding of octadecylsilyl groups. To
A very finely divided silica gel consisting of porous, spherical minimise any interaction with basic compounds it is carefully
particles with chemically bonded nitrile groups. end-capped to cover most of the remaining silanol groups.
Silica gel for chromatography, nitrile R2. 1119500. Silica gel for chromatography, octadecylsilyl, end-capped,
Ultrapure silica gel, chemically modified at the surface by the base-deactivated. 1108600.
introduction of cyanopropylsilyl groups. Less than 20 ppm A very finely divided silica gel, pre-treated before the bonding
of metals. of octadecylsilyl groups by washing and hydrolysing most
of the superficial siloxane bridges. To further minimise any
Silica gel for chromatography, nitrile, end-capped. 1174500. interaction with basic compounds, it is carefully end-capped
A very finely divided silica gel, chemically modified at the to cover most of the remaining silanol groups.
surface by the bonding of cyanopropylsilyl groups. To
minimise any interaction with basic components it is carefully

end-capped to cover most of the remaining silanol groups. Silica gel for chromatography, octadecylsilyl, extra-dense
bonded, end-capped. 1188500.
Silica gel for chromatography, 4-nitrophenylcarbamidesilyl. A very finely divided silica gel, chemically modified at the
1185200. surface by the extra-dense bonding of octadecylsilyl groups.
A very finely divided silica gel, chemically modified at the To minimise any interaction with basic compounds it is
surface by bonding of 4-nitrophenylcarbamide groups. carefully end-capped to cover most of the remaining silanol
groups.
Silica gel for chromatography, octadecanoylaminopropyl-
silyl. 1115200. Silica gel for chromatography, octadecylsilyl, for separation
A very finely divided silica gel, chemically modified at the of polycyclic aromatic hydrocarbons. 1202900.
surface by the bonding of aminopropylsilyl groups which are A very finely divided ultrapure silica gel, chemically modified
acylated with octadecanoyl groups. at the surface by the bonding of octadecylsilyl groups,
Silica gel for chromatography, octadecylphenylsilyl, optimised for the analysis of polycyclic aromatic hydrocarbons.
end-capped. 1199300. Silica gel for chromatography, octadecylsilyl, monolithic,
A very finely divided silica gel, chemically modified at the end-capped. 1154500.
surface by bonding of octadecylphenylsilyl groups. To
minimise any interaction with basic compounds it is carefully Monolithic rods of highly porous (greater than 80 per cent)
end-capped to cover most of the remaining silanol groups. metal-free silica with a bimodal pore structure, modified at the
surface by the bonding of octadecylsilyl groups. To minimise
Silica gel for chromatography, octadecylsilyl. 1077500. any interaction with basic compounds, they are carefully
A very finely divided silica gel, chemically modified at the end-capped to cover most of the remaining silanol groups.
surface by the bonding of octadecylsilyl groups. Silica gel for chromatography, octadecylsilyl,
Silica gel for chromatography, octadecylsilyl R1. 1110100. polar-embedded, encapsulated. 1206600.
A very finely divided ultrapure silica gel, chemically modified Silica gel chemically modified at the surface by the bonding
at the surface by the bonding of octadecylsilyl groups. Less of polar-embedded octadecylsilyl groups. To minimise any
than 20 ppm of metals. interaction with basic compounds, it is carefully encapsulated
Silica gel for chromatography, octadecylsilyl R2. 1115300.
A very finely divided (15 nm pore size) ultrapure silica Silica gel for chromatography, octadecylsilyl, polar
gel, chemically modified at the surface by the bonding of end-capped. 1205500.
octadecylsilyl groups (20 per cent carbon load), optimised for A very finely divided silica gel, chemically modified at the
the analysis of polycyclic aromatic hydrocarbons. surface by the bonding of octadecylsilyl groups. To minimise
any interaction with basic compounds it is carefully polar
Silica gel for chromatography, octadecylsilyl, end-capped to cover most of the remaining silanol groups.
base-deactivated. 1077600.
A very finely divided silica gel, pretreated before the bonding Silica gel for chromatography, octadecylsilyl, solid core.
of octadecylsilyl groups by careful washing and hydrolysing 1205600. Silica gel with spherical silica particles containing
most of the superficial siloxane bridges to minimise the a non-porous solid silica core surrounded by a thin outer
interaction with basic components. porous silica coating with octadecylsilyl groups.

Silica gel for chromatography, octadecylsilyl, solid core, Silica gel for chromatography, octylsilyl, extra-dense
end-capped. 1193900. bonded, end-capped. 1200900.
Silica gel with spherical silica particles containing a A very finely divided silica gel, chemically modified at the
non-porous solid silica core surrounded by a thin outer porous surface by the extra-dense bonding of octylsilyl groups. To
silica coating with octadecylsilyl groups. To minimise any minimise any interaction with basic compounds, it is carefully
interaction with basic compounds it is carefully end-capped to end-capped to cover most of the remaining silanol groups.
cover most of the remaining silanol groups.
Silica gel for chromatography, octylsilyl, with polar
Silica gel for chromatography, octadecylsilyl, with incorporated groups, end-capped. 1152600.
embedded polar groups, end-capped. 1177900. A very finely divided silica gel. The particles are based
A very finely divided silica gel. The particles are based on on silica, chemically modified with a reagent providing a
a mixture of silica chemically modified at the surface by surface with chains having polar incorporated groups and
the bonding of octadecylsilyl groups and silica chemically terminating octyl groups. Furthermore, the packing material
modified with a reagent providing a surface with chains is end-capped.
having embedded polar groups. Furthermore, the packing
material is end-capped. Silica gel for chromatography, oxypropionitrilsilyl.
1184700.
Silica gel for chromatography, octadecylsilyl, with extended A very finely divided silica gel chemically modified at the
pH range, end-capped. 1196700. surface by the bonding of oxypropionitrilsilyl groups.
surface by the bonding of octadecylsilyl groups resistant to Silica gel for chromatography, palmitamidopropylsilyl,
bases up to pH 11. To minimise any interaction with basic end-capped. 1161900.
compounds it is carefully end-capped to cover most of the A very finely divided silica gel, chemically modified at the
remaining silanol groups. surface by the bonding of palmitamidopropyl groups and
end-capped with acetamidopropyl groups.
Silica gel for chromatography, octadecylsilyl, with polar
incorporated groups, end-capped. 1165100. Silica gel for chromatography, pentafluorophenylpropyl-
A very finely divided silica gel. The particles are based on silyl, solid core, end-capped. 1207600.
silica, chemically modified with a reagent providing a surface Silica gel with spherical silica particles containing a
with chains having polar incorporated groups and terminating non-porous solid silica core surrounded by a thin outer porous
octadecyl groups. Furthermore, the packing material is silica coating with pentafluorophenylpropylsilyl groups. To
end-capped. minimise any interaction with basic compounds it is carefully
Silica gel for chromatography, octylsilyl. 1077700.
A very finely divided silica gel, chemically modified at the Silica gel for chromatography, phenylhexylsilyl. 1153900.
surface by the bonding of octylsilyl groups. A very finely divided silica gel, chemically modified at the
surface by the bonding of phenylhexyl groups.
Silica gel for chromatography, octylsilyl R1. 1077701.
A very finely divided silica gel, chemically modified at the Silica gel for chromatography, phenylhexylsilyl,
surface by the bonding of octylsilyl and methyl groups (double end-capped. 1170600.
bonded phase). A very finely divided silica gel, chemically modified at
the surface by the bonding of phenylhexylsilyl groups. To
Silica gel for chromatography, octylsilyl R2. 1077702. minimise any interaction with basic compounds it is carefully
Ultrapure very finely divided (10 nm pore size) silica gel, end-capped to cover most of the remaining silanol groups.
chemically modified at the surface by the bonding of octylsilyl
groups (19 per cent carbon load). Less than 20 ppm of metals. Silica gel for chromatography, phenylhexylsilyl, solid core,
Silica gel for chromatography, octylsilyl R3. 1155200.
Silica gel with spherical silica particles containing a
A very finely divided ultrapure silica gel, chemically modified non-porous solid silica core surrounded by a thin outer porous
at the surface by the bonding of octylsilyl groups and sterically silica coating with phenylhexylsilyl groups. To minimise any
protected with branched hydrocarbons at the silanes. interaction with basic compounds it is carefully end-capped to
cover most of the remaining silanol groups.
Silica gel for chromatography, octylsilyl, base-deactivated.
1131600. Silica gel for chromatography, phenylsilyl. 1110200.
A very finely divided silica gel, pretreated before the bonding A very finely divided silica gel, chemically modified at the
of octylsilyl groups by careful washing and hydrolysing most surface by the bonding of phenyl groups.
of the superficial siloxane bridges to minimise the interaction
with basic components.

Silica gel for chromatography, phenylsilyl, end-capped.
Silica gel for chromatography, octylsilyl, end-capped. 1154900.
1119600. A very finely divided silica gel, chemically modified at the
A very finely divided silica gel, chemically modified at the surface by the bonding of phenyl groups. To minimise any
surface by the bonding of octylsilyl groups. To minimise any interaction with basic compounds it is carefully end-capped to
interaction with basic compounds, it is carefully end-capped cover most of the remaining silanol groups.
Silica gel for chromatography, phenylsilyl, end-capped,
Silica gel for chromatography, octylsilyl, end-capped, base-deactivated. 1197900.
base-deactivated. 1148800. A very finely divided silica gel pre-treated before the bonding
A very finely divided silica gel, pre-treated before the bonding of phenylsilyl groups by washing and hydrolysing most of
of octylsilyl groups by washing and hydrolysing most of the superficial siloxane bridges, chemically modified at the
the superficial siloxane bridges. To further minimise any surface by bonding of phenyl groups. To further minimise any
interaction with basic compounds it is carefully end-capped to interaction with basic compounds it is carefully end-capped to
cover most of the remaining silanol groups. cover most of the remaining silanol groups.

Silica gel for chromatography, phenylsilyl, extra-dense Silica gel H. 1076500. [112926-00-8].
bonded, end-capped. 1207700. The particle size is of about 15 µm.
A very finely divided silica gel, chemically modified at the pH (2.2.3). Complies with the test prescribed for silica gel G R.
surface by the extra-dense bonding of phenylsilyl groups. To
minimise any interaction with basic compounds, it is carefully Silica gel H, silanised. 1076600.
end-capped to cover most of the remaining silanol groups. Preparation of a thin layer. See silanised silica gel HF254 R
Silica gel for chromatography, propoxybenzene, Chromatographic separation. Complies with the test
end-capped. 1174600. prescribed for silanised silica gel HF254 R.
Silica gel HF254. 1076700.
surface by the bonding of propoxybenzene groups.
Contains about 1.5 per cent of a fluorescent indicator having
Silica gel for chromatography, propylsilyl. 1170700. an optimal intensity at 254 nm. The particle size is about
A very finely divided silica gel, chemically modified at the 15 µm.
surface by the bonding of propylsilyl groups. pH. Complies with the test prescribed for silica gel G R.
Silica gel for chromatography, strong-anion-exchange. Fluorescence. Complies with the test prescribed for silica
1077800. gel GF254 R.
A very finely divided silica gel, chemically modified at the Silica gel HF254, silanised. 1076800.
surface by the bonding of quaternary ammonium groups. Contains about 1.5 per cent of a fluorescent indicator having
pH limit of use : 2 to 8. an optimal intensity at 254 nm.
Silica gel for chromatography, strong cation-exchange. Preparation of a thin layer. Vigorously shake 30 g for 2 min
1161400. with 60 mL of a mixture of 1 volume of methanol R and
2 volumes of water R. Coat carefully cleaned plates with a
A very finely divided silica gel, chemically modified at the layer 0.25 mm thick using a spreading device. Allow the
surface by the bonding of sulfonic acid groups. coated plates to dry in air and then heat in an oven at 100 °C
Silica gel for chromatography, trimethylsilyl. 1115500. to 105 °C for 30 min.
A very finely divided silica gel, chemically modified at the Chromatographic separation. Introduce 0.1 g each of methyl
surface by the bonding of trimethylsilyl groups. laurate R, methyl myristate R, methyl palmitate R and methyl
stearate R into a 250 mL conical flask. Add 40 mL of alcoholic
Silica gel for size-exclusion chromatography. 1077900. potassium hydroxide solution R and heat under a reflux
A very finely divided silica gel (10 μm) with a very hydrophilic condenser on a water-bath for 1 h. Allow to cool, transfer the
surface. The average diameter of the pores is about 30 nm. solution to a separating funnel by means of 100 mL of water R,
It is compatible with aqueous solutions between pH 2 and 8 acidify (pH 2 to 3) with dilute hydrochloric acid R and shake
and with organic solvents. It is suitable for the separation of with three quantities, each of 10 mL of chloroform R. Dry
proteins with relative molecular masses of 1 × 103 to 3 × 105. the combined chloroform extracts over anhydrous sodium
sulfate R, filter and evaporate to dryness on a water-bath.
Silica gel G. 1076300. [112926-00-8]. Dissolve the residue in 50 mL of chloroform R. Examine by
thin-layer chromatography (2.2.27), using silanised silica
Contains about 13 per cent of calcium sulfate hemihydrate. gel HF254 as the coating substance. Apply to the plate at each
The particle size is about 15 µm. of three separate points 10 µL of the chloroformic solution.
Calcium sulfate content. Place 0.25 g in a ground-glass Develop over a path of 14 cm with a mixture of 10 volumes of
stoppered flask, add 3 mL of dilute hydrochloric acid R glacial acetic acid R, 25 volumes of water R and 65 volumes of
and 100 mL of water R and shake vigorously for 30 min. dioxan R. Dry the plate at 120 °C for 30 min. Allow to cool,
Filter through a sintered-glass filter (2.1.2) and wash the spray with a 35 g/L solution of phosphomolybdic acid R in
residue. Carry out on the combined filtrate and washings the 2-propanol R and heat at 150 °C until the spots become visible.
complexometric assay of calcium (2.5.11). Treat the plate with ammonia vapour until the background
1 mL of 0.1 M sodium edetate is equivalent to 14.51 mg of is white. The chromatograms show four clearly separated,
CaSO4,1/2H2O. well-defined spots.
pH (2.2.3). Shake 1 g for 5 min with 10 mL of carbon

dioxide-free water R. The pH of the suspension is about 7. Silicotungstic acid. H4SiW12O40,xH2O. 1078000.
[11130-20-4].
Silica gel GF254. 1076400. [112926-00-8].
White or yellowish-white crystals, deliquescent, very soluble
Contains about 13 per cent of calcium sulfate hemihydrate in water and in ethanol (96 per cent).
and about 1.5 per cent of a fluorescent indicator having an
optimal intensity at 254 nm. The particle size is about 15 µm. Storage : in an airtight container.
Calcium sulfate content. Determine by the method prescribed Silicristin. C25H22O10. (Mr 482.4). 1151500. [33889-69-9].
for silica gel G R. (2R,3R)-3,5,7-Trihydroxy-2-[(2R,3S)-7-hydroxy-2-(4-
pH. Complies with the test prescribed for silica gel G R. hydroxy-3-methoxyphenyl)-3-hydroxymethyl-2,3-dihydro-1-
Fluorescence. Thin-layer chromatography (2.2.27) using benzofuran-5-yl]chroman-4-one.
silica gel GF254 R as the coating substance. Apply separately White or yellowish powder, practically insoluble in water,
to the plate at ten points increasing volumes from 1 µL to soluble in acetone and in methanol.
10 µL of a 1 g/L solution of benzoic acid R in a mixture of
10 volumes of anhydrous formic acid R and 90 volumes of Silidianin. C25H22O10. (Mr 482.4). 1151600. [29782-68-1].
2-propanol R. Develop over a path of 10 cm with the same (3R,3aR,6R,7aR,8R)-7a-Hydroxy-8-(4-hydroxy-3-
mixture of solvents. After evaporating the solvents examine methoxyphenyl)-4-[(2R, 3R)-3,5,7-trihydroxy-4-oxochroman-
the chromatogram in ultraviolet light at 254 nm. The benzoic 2-yl]-2,3,3a,7a-tetrahydro-3,6-methano-1-benzofuran-
acid appears as dark spots on a fluorescent background in the 7(6aH)-one.
upper third of the chromatogram for quantities of 2 µg and White or yellowish powder, practically insoluble in water,
greater. soluble in acetone and in methanol.

Silver diethyldithiocarbamate. C5H10AgNS2. (Mr 256.1). Sinensetin. C20H20O7. (Mr 372.4). 1110500. [2306-27-6].

1110400. [1470-61-7]. Silver diethylcarbamodithioate. 3′,4′,5,6,7-Pentamethoxyflavone.
Pale-yellow or greyish-yellow powder, practically insoluble White or almost white, crystalline powder, practically
in water, soluble in pyridine. insoluble in water, soluble in ethanol (96 per cent).
Storage below 8 °C is recommended. mp : about 177 °C.
It may be prepared as follows. Dissolve 1.7 g of silver nitrate R Absorbance (2.2.25). A solution in methanol R shows
in 100 mL of water R. Separately dissolve 2.3 g of sodium 3 absorption maxima, at 243 nm, 268 nm and 330 nm.
diethyldithiocarbamate R in 100 mL of water R. Cool both Assay. Liquid chromatography (2.2.29) as prescribed in the
solutions to 10 °C, then mix, and while stirring, collect the monograph Java tea (1229).
yellow precipitate on a sintered-glass filter (16) (2.1.2) and Content : minimum 95 per cent, calculated by the
wash with 200 mL of cold water R. Dry the precipitate in normalisation procedure.
vacuo for 10 h (2.2.32).
Sinomenine. C19H23NO4. (Mr 329.4). 1183400. [115-53-7].
Silver diethyldithiocarbamate solution. 1110401. 7,8-Didehydro-4-hydroxy-3,7-dimethoxy-17-methyl-
Prepare the solution immediately before use. Dissolve 9α,13α,14α-morphinan-6-one. Cucoline.
0.100 g of silver diethyldithiocarbamate R in pyridine R and Sirolimus. C51H79NO13. (Mr 914). 1205700. [53123-88-9].
dilute to 20.0 mL with the same solvent. Rapamycin.
Suitability test. The solution is clear (2.2.1). The absorbance mp : 183 °C to 185 °C.
(2.2.25) of the solution is maximum 0.20 at 450 nm,
maximum 0.01 at 510 nm and maximum 0.010 at 538 nm. Sitostanol. C29H52O. (Mr 416.7). 1140100. [19466-47-8].
Dihydro-β-sitosterol.
Silver manganese paper. 1078200. Content : minimum 95.0 per cent.
Immerse strips of slow filter paper into a solution containing β-Sitosterol. C29H50O. (Mr 414.7). 1140200. [83-46-5].
8.5 g/L of manganese sulfate R and 8.5 g/L of silver nitrate R. Stigmast-5-en-3β-ol. 22,23-Dihydrostigmasterol.
Maintain for a few minutes and allow to dry over an
appropriate desiccant, protected from acid and alkaline White or almost white powder, practically insoluble in water,
vapours. sparingly soluble in tetrahydrofuran.
Content : minimum 75.0 per cent m/m (dried substance).
Silver nitrate. 1078300. [7761-88-8]. Assay. Gas chromatography (2.2.28), as prescribed in the
See Silver nitrate (0009). monograph Phytosterol (1911).
Test solution. Dissolve 0.100 g of the substance to be
Silver nitrate reagent. 1078305. examined in tetrahydrofuran R and dilute to 10.0 mL
Prepare immediately before use. To a mixture of 3 mL with the same solvent. Introduce 100 µL of this solution
of concentrated ammonia R and 40 mL of 1 M sodium into a suitable 3 mL flask and evaporate to dryness under
hydroxide, add 8 mL of a 200 g/L solution of silver nitrate R, nitrogen R. To the residue add 100 μL of a freshly prepared
dropwise, with stirring. Dilute to 200 mL with water R. mixture of 50 µL of 1-methylimidazole R and 1.0 mL of
heptafluoro-N-methyl-N-(trimethylsilyl)butanamide R. Close
Silver nitrate solution R1. 1078301. the flask tightly and heat at 100 °C for 15 min. Allow to cool.
A 42.5 g/L solution of silver nitrate R. Injection : 1 µL of the test solution.
Storage : protected from light. Sodium. Na. (A 22.99). 1078500. [7440-23-5].
r
Silver nitrate solution R2. 1078302. A metal whose freshly cut surface is bright silver-grey. It
rapidly tarnishes in contact with air and is oxidised completely
A 17 g/L solution of silver nitrate R. to sodium hydroxide and converted to sodium carbonate. It
Storage : protected from light. reacts violently with water, yielding hydrogen and a solution
of sodium hydroxide ; soluble in anhydrous methanol, yielding
Silver nitrate solution, ammoniacal. 1078303. hydrogen and a solution of sodium methoxide ; practically
Dissolve 2.5 g of silver nitrate R in 80 mL of water R and insoluble in light petroleum.
add dilute ammonia R1 dropwise until the precipitate Storage : under light petroleum or liquid paraffin.
has dissolved. Dilute to 100 mL with water R. Prepare
immediately before use. Sodium acetate. 1078600. [6131-90-4].
See Sodium acetate trihydrate (0411).
Silver nitrate solution in pyridine. 1078304.
Sodium acetate, anhydrous. C2H3NaO2. (Mr 82.0). 1078700.
An 85 g/L solution of silver nitrate R in pyridine R. [127-09-3].
Storage : protected from light. Colourless crystals or granules, very soluble in water, sparingly
Silver oxide. Ag2O. (Mr 231.7). 1078400. [20667-12-3].
Disilver oxide. Loss on drying (2.2.32). Not more than 2.0 per cent,
determined by drying in an oven at 105 °C.
Brownish-black powder, practically insoluble in water and in
ethanol (96 per cent), freely soluble in dilute nitric acid and Sodium arsenite. NaAsO2. (Mr 129.9). 1165900. [7784-46-5].
in ammonia. Sodium metaarsenite.
Storage : protected from light. Sodium arsenite solution. 1165901.
Silver sulfate. Ag2SO4. (Mr 311.8). 1201000. [10294-26-5]. Dissolve 5.0 g of sodium arsenite R in 30 mL of 1 M sodium
hydroxide. Cool to 0 °C and add, while stirring, 65 mL of
Content : minimum 99.0 per cent. dilute hydrochloric acid R.
White or light grey powder, slightly soluble in water.
Sodium ascorbate solution. 1078800. [134-03-2].
mp : about 652 °C. Dissolve 3.5 g of ascorbic acid R in 20 mL of 1 M sodium
Storage : protected from light. hydroxide. Prepare immediately before use.

Sodium azide. NaN3. (Mr 65.0). 1078900. [26628-22-8]. Sodium citrate. 1079600. [6132-04-3].
White or almost white, crystalline powder or crystals, freely See Sodium citrate (0412).
soluble in water, slightly soluble in ethanol (96 per cent).
Sodium cobaltinitrite. Na3[Co(NO2)6]. (Mr 403.9). 1079700.
Sodium benzenesulfonate. C6H5SO3Na. (Mr 180.16). [13600-98-1]. Trisodium hexanitrocobaltate(III).
1196600. [515-42-4].
Orange-yellow powder, freely soluble in water, slightly soluble
White crystalline powder, soluble in water. in ethanol (96 per cent).
Sodium bicarbonate. 1081300. [144-55-8].
Sodium cobaltinitrite solution. 1079701.
See sodium hydrogen carbonate R.
A 100 g/L solution of sodium cobaltinitrite R. Prepare
Sodium bismuthate. NaBiO3. (Mr 280.0). 1079000. immediately before use.
[12232-99-4].
Content : minimum 85.0 per cent. Sodium decanesulfonate. C10H21NaO3S. (Mr 244.3). 1079800.
[13419-61-9].
Yellow or yellowish-brown powder, slowly decomposing when
moist or at a high temperature, practically insoluble in cold Crystalline powder or flakes, white or almost white, freely
water. soluble in water, soluble in methanol.
Assay. Suspend 0.200 g in 10 mL of a 200 g/L solution of Sodium decyl sulfate. C10H21NaO4S. (Mr 260.3). 1138600.
potassium iodide R and add 20 mL of dilute sulfuric acid R. [142-87-0].
Using 1 mL of starch solution R as indicator, titrate with 0.1 M
sodium thiosulfate until an orange colour is obtained. Content : minimum 95.0 per cent.
1 mL of 0.1 M sodium thiosulfate is equivalent to 14.00 mg White or almost white powder, freely soluble in water.
of NaBiO3.
Sodium deoxycholate. C24H39NaO4. (Mr 414.6). 1131800.
Sodium bromide. 1154300. [7647-15-6]. [302-95-4]. Sodium 3α,12α-dihydroxy-5β-cholan-24-oate.
See Sodium bromide (0190). Sodium deoxyribonucleate. (About 85 per cent has a relative
Sodium butanesulfonate. C4H9NaO3S. (Mr 160.2). 1115600. molecular mass of 2 × 107 or greater). 1079900. [73049-39-5].
[2386-54-1]. White or almost white, fibrous preparation obtained from calf
White or almost white, crystalline powder, soluble in water. thymus.
mp : greater than 300 °C. Test for suitability. Dissolve 10 mg in imidazole buffer solution
pH 6.5 R and dilute to 10.0 mL with the same buffer solution
Sodium calcium edetate. 1174000. [62-33-9]. (solution A). Dilute 2.0 mL of solution A to 50.0 mL with
See sodium calcium edetate (0231). imidazole buffer solution pH 6.5 R. The absorbance (2.2.25) of
Sodium carbonate. 1079200. [6132-02-1]. the solution, measured at 260 nm, is 0.4 to 0.8.
See Sodium carbonate decahydrate (0191). To 0.5 mL of solution A add 0.5 mL of imidazole buffer
solution pH 6.5 R and 3 mL of perchloric acid (25 g/L HClO4).
Sodium carbonate, anhydrous. Na2CO3. (Mr 106.0). A precipitate is formed. Centrifuge. The absorbance of the
1079300. [497-19-8]. Disodium carbonate. supernatant, measured at 260 nm using a mixture of 1 mL of
White or almost white powder, hygroscopic, freely soluble in imidazole buffer solution pH 6.5 R and 3 mL of perchloric acid
water. (25 g/L HClO4) as compensation liquid, is not greater than 0.3.
When heated to about 300 °C it loses not more than 1 per In each of two tubes, place 0.5 mL of solution A and 0.5 mL
cent of its mass. of a solution of a reference preparation of streptodornase
Storage : in an airtight container. containing 10 IU/mL in imidazole buffer solution pH 6.5 R.
To one tube add immediately 3 mL of perchloric acid (25 g/L
Sodium carbonate solution. 1079301. HClO4). A precipitate is formed. Centrifuge and collect
A 106 g/L solution of anhydrous sodium carbonate R. supernatant A. Heat the other tube at 37 °C for 15 min and add
3 mL of perchloric acid (25 g/L HClO4). Centrifuge and collect
Sodium carbonate solution R1. 1079302. supernatant B. The absorbance of supernatant B, measured at
A 20 g/L solution of anhydrous sodium carbonate R in 260 nm with reference to supernatant A is not less than 0.15.
0.1 M sodium hydroxide.
Sodium diethyldithiocarbamate. C5H10NNaS2,3H2O.
Sodium carbonate solution R2. 1079303. (Mr 225.3). 1080000. [20624-25-3].
A 40 g/L solution of anhydrous sodium carbonate R in White or almost white or colourless crystals, freely soluble in
0.2 M sodium hydroxide. water, soluble in ethanol (96 per cent). The aqueous solution
Sodium carbonate monohydrate. 1131700. [5968-11-6]. is colourless.
See Sodium carbonate monohydrate (0192). Sodium dihydrogen phosphate. 1080100. [13472-35-0].
Sodium cetostearyl sulfate. 1079400. See Sodium dihydrogen phosphate dihydrate (0194).
See Sodium cetostearyl sulfate (0847).
Sodium dihydrogen phosphate, anhydrous. NaH2PO4.
Sodium chloride. 1079500. [7647-14-5]. (Mr 120.0). 1080200. [7558-80-7].
See Sodium chloride (0193). White or almost white powder, hygroscopic.
Sodium chloride solution. 1079502. Storage : in an airtight container.
A 20 per cent m/m solution of sodium chloride R. Sodium dihydrogen phosphate monohydrate.
Sodium chloride solution, saturated. 1079503. NaH2PO4,H2O. (Mr 138.0). 1080300. [10049-21-5].
Mix 1 part of sodium chloride R with 2 parts of water R, White or almost white, slightly deliquescent crystals or
shake from time to time and allow to stand. Before use, granules, freely soluble in water, practically insoluble in
decant the solution from any undissolved substance and ethanol (96 per cent).
filter, if necessary. Storage : in an airtight container.

Sodium dioctyl sulfosuccinate. C20H37NaO7S. (Mr 444.6). Sodium hexanesulfonate monohydrate for ion-pair
1170800. [577-11-7]. Sodium 1,4-bis[(2-ethylhexyl)oxy]- chromatography. C6H13NaO3S,H2O. (Mr 206.2). 1182300.
1,4-dioxobutane-2-sulfonate. 1,4-Bis(2-ethylhexyl) [207300-91-2].
sulfobutanedioate sodium salt. Content : minimum 99.0 per cent.
White or almost white, waxy solid.
Sodium hydrogen carbonate. 1081300. [144-55-8].
Sodium dithionite. Na2S2O4. (Mr 174.1). 1080400. See Sodium hydrogen carbonate (0195).
[7775-14-6].
White or greyish-white, crystalline powder, oxidises in air, Sodium hydrogen carbonate solution. 1081301.
very soluble in water, slightly soluble in ethanol (96 per cent). A 42 g/L solution of sodium hydrogen carbonate R.
Storage : in an airtight container. Sodium hydrogen sulfate. NaHSO4. (Mr 120.1). 1131900.
Sodium dodecyl sulfate. 1080500. [151-21-3]. [7681-38-1]. Sodium bisulfate.
See Sodium laurilsulfate (0098). Freely soluble in water, very soluble in boiling water. It
decomposes in ethanol (96 per cent) into sodium sulfate and
free sulfuric acid.
Sodium edetate. 1080600. [6381-92-6]. mp : about 315 °C.
See Disodium edetate (0232).
Sodium hydrogensulfite. NaHO3S. (Mr 104.1). 1115700.
Sodium fluoresceinate. C20H10Na2O5. (Mr 376.3). 1080700. [7631-90-5].
[518-47-8]. White or almost white, crystalline powder, freely soluble in
Schultz No. 880. water, sparingly soluble in ethanol (96 per cent).
Colour Index No. 45350. On exposure to air, some sulfur dioxide is lost and the
Fluorescein sodium. Disodium 2-(3-oxo-6-oxido-3H- substance is gradually oxidated to sulfate.
xanthen-9-yl)benzoate.
Sodium hydroxide. 1081400. [1310-73-2].
Orange-red powder, freely soluble in water. Aqueous solutions
display an intense yellowish-green fluorescence. See Sodium hydroxide (0677).
Sodium fluoride. 1080800. [7681-49-4]. 2 M Sodium hydroxide. 3009800.

See Sodium fluoride (0514). Dissolve 84 g of sodium hydroxide R in carbon dioxide-free
water R and dilute to 1000.0 mL with the same solvent.
Sodium formate. CHNaO2. (Mr 68.0). 1122200. [141-53-7].
Sodium methanoate. 4 M Sodium hydroxide. 1081407.
White or almost white, crystalline powder or deliquescent Dissolve 168 g of sodium hydroxide R in carbon dioxide-free
granules, soluble in water and in glycerol, slightly soluble in water R and dilute to 1.0 L with the same solvent.
ethanol (96 per cent). Sodium hydroxide solution. 1081401.
mp : about 253 °C. Dissolve 20.0 g of sodium hydroxide R in water R and dilute
Sodium glucuronate. C6H9NaO7,H2O. (Mr 234.1). 1080900. to 100.0 mL with the same solvent. Verify the concentration
Sodium D-glucuronate monohydrate. by titration with 1 M hydrochloric acid, using methyl orange
solution R as indicator, and adjust if necessary to 200 g/L.
: about + 21.5, determined on a 20 g/L solution.
Sodium hydroxide solution, carbonate-free. 1081406.
Sodium glycocholate. C26H42NNaO6,2H2O.
(Mr 523.6). 1155500. [207300-80-9]. Sodium Dissolve sodium hydroxide R in carbon dioxide-free water R
[(3,7,12-trihydroxy-5-cholan-24-oyl)amino]acetate dihydrate. to give a concentration of 500 g/L and allow to stand.
N-[(3,5,7,12)-3,7,12-Trihydroxy-24-oxocholan-24-yl]glycine Decant the clear supernatant, taking precautions to avoid
monosodium salt dihydrate. the introduction of carbon dioxide.
Content : minimum 97 per cent of C26H42NNaO6,2H2O. Sodium hydroxide solution, dilute. 1081402.
Sodium heptanesulfonate. C7H15NaO3S. (Mr 202.3). 1081000. Dissolve 8.5 g of sodium hydroxide R in water R and dilute
[22767-50-6]. to 100 mL with the same solvent.
White or almost white, crystalline mass, freely soluble in Sodium hydroxide solution, methanolic. 1081403.
water, soluble in methanol. Dissolve 40 mg of sodium hydroxide R in 50 mL of water R.
Sodium heptanesulfonate monohydrate. C7H15NaO3S,H2O. Cool and add 50 mL of methanol R.
(Mr 220.3). 1081100. Sodium hydroxide solution, methanolic R1. 1081405.
Content : minimum 96 per cent (anhydrous substance). Dissolve 200 mg of sodium hydroxide R in 50 mL of
White or almost white, crystalline powder, soluble in water, water R. Cool and add 50 mL of methanol R.
very slightly soluble in anhydrous ethanol.
Sodium hydroxide solution, strong. 1081404.
Water (2.5.12) : maximum 8 per cent, determined on 0.300 g.
Dissolve 42 g of sodium hydroxide R in water R and dilute
Assay. Dissolve 0.150 g in 50 mL of anhydrous acetic acid R.
to 100 mL with the same solvent.
Titrate with 0.1 M perchloric acid, determining the end-point
potentiometrically (2.2.20). Sodium 2-hydroxybutyrate. C4H7NaO3. (Mr 126.1). 1158800.
1 mL of 0.1 M perchloric acid is equivalent to 20.22 mg of [19054-57-0]. Sodium (2RS)-2-hydroxybutanoate.
C7H15NaO3S.
Sodium hypobromite solution. 1081500.
Sodium hexanesulfonate. C6H13NaO3S. (Mr 188.2). 1081200. In a bath of iced water mix 20 mL of strong sodium hydroxide
[2832-45-3]. solution R and 500 mL of water R, add 5 mL of bromine
White or almost white powder, freely soluble in water. solution R and stir gently until solution is complete. Prepare
Sodium hexanesulfonate monohydrate. C6H13NaO3S,H2O.
(Mr 206.2). 1161500. [207300-91-2]. Sodium hypochlorite solution, strong. 1081600.
White or almost white powder, soluble in water. Content : 25 g/L to 30 g/L of active chlorine.

Yellowish liquid with an alkaline reaction. Sodium nitrite solution. 1082501.

Assay. Introduce into a flask, successively, 50 mL of water R, A 100 g/L solution of sodium nitrite R. Prepare immediately
1 g of potassium iodide R and 12.5 mL of dilute acetic acid R. before use.
Dilute 10.0 mL of the substance to be examined to 100.0 mL
with water R. Introduce 10.0 mL of this solution into the flask Sodium nitroprusside. Na2[Fe(CN)5(NO)],2H2O.
and titrate with 0.1 M sodium thiosulfate, using 1 mL of starch (Mr 298.0). 1082600. [13755-38-9]. Sodium
solution R as indicator. pentacyano-nitrosylferrate(III) dihydrate.
1 mL of 0.1 M sodium thiosulfate is equivalent to 3.546 mg Reddish-brown powder or crystals, freely soluble in water,
of active chlorine. slightly soluble in ethanol (96 per cent).
Storage : protected from light. Sodium octanesulfonate. C8H17NaO3S. (Mr 216.3). 1082700.
[5324-84-5].
Sodium hypophosphite. NaH2PO2,H2O. (Mr 106.0). 1081700.
[10039-56-2]. Sodium phosphinate monohydrate.
White or almost white, crystalline powder or flakes, freely
soluble in water, soluble in methanol.
crystals, hygroscopic, freely soluble in water, soluble in ethanol
(96 per cent). Absorbance (2.2.25) : maximum 0.10, determined at 200 nm
and maximum 0.01, determined at 250 nm using a 54 g/L
solution.
Sodium iodide. 1081800. [7681-82-5]. Sodium octanesulfonate monohydrate. C8H17NaO3S,H2O.
See Sodium iodide (0196). (Mr 234.3). 1176700. [207596-29-0].
Sodium laurilsulfate. 1081900. [151-21-3]. White or almost white powder.
See Sodium laurilsulfate (0098). Sodium octyl sulfate. C8H17NaO4S. (Mr 232.3). 1082800.
[142-31-4].
Sodium lauryl sulfate. 1081900. [151-21-3].
White or almost white, crystalline powder or flakes, freely
See Sodium laurilsulfate R. soluble in water, soluble in methanol.
Sodium laurylsulfonate for chromatography. C12H25NaO3S. Sodium oxalate. C2Na2O4. (Mr 134.0). 1082900. [62-76-0].
(Mr 272.4). 1132000. [2386-53-0].
White or almost white, crystalline powder, soluble in water,
White or almost white powder or crystals, freely soluble in practically insoluble in ethanol (96 per cent).
water.
Absorbance A15% Sodium oxidronate. CH4Na2O7P2. (Mr 236.0). 1194000.
cm (2.2.25), determined in water R : about 0.05
at 210 nm ; about 0.03 at 220 nm ; about 0.02 at 230 nm ; [14255-61-9]. Sodium hydroxymethylenediphosphonate.
about 0.02 at 500 nm. White or almost white powder or colourless crystals, very
soluble in water, very slightly soluble in ethanol (96 per cent),
Sodium metabisulfite. 1082000. [7681-57-4]. practically insoluble in methylene chloride.
See Sodium metabisulfite (0849).
Sodium pentanesulfonate. C5H11NaO3S. (Mr 174.2).
Sodium methanesulfonate. CH3SO3Na. (Mr 118.1). 1082100. 1083000. [22767-49-3].
[2386-57-4]. White or almost white, crystalline solid, soluble in water.
White or almost white, crystalline powder, hygroscopic.
Sodium pentanesulfonate monohydrate. C5H11NaO3S,H2O.
Storage : in an airtight container. (Mr 192.2). 1132100. [207605-40-1].
Sodium molybdate. Na2MoO4,2H2O. (Mr 242.0). 1082200. White or almost white crystalline solid, soluble in water.
[10102-40-6]. Disodium molybdate dihydrate. Sodium pentanesulfonate monohydrate R1.
White or almost white, crystalline powder or colourless C5H11NaO3S,H2O. (Mr 192.2). 1172500. [207605-40-1].
crystals, freely soluble in water. Content : minimum 99 per cent of C5H11NaO3S,H2O.
Sodium naphthoquinonesulfonate. C10H5NaO5S. (Mr 260.2). Sodium perchlorate. NaClO4,H2O. (Mr 140.5). 1083100.
1082300. [521-24-4]. Sodium 1,2-naphthoquinone-4- [7791-07-3].
sulfonate.
Content : minimum 99.0 per cent of NaClO4,H2O.
Yellow or orange-yellow, crystalline powder, freely soluble in
water, practically insoluble in ethanol (96 per cent). White or almost white, deliquescent crystals, very soluble in
water.
Sodium nitrate. NaNO3. (Mr 85.0). 1082400. [7631-99-4]. Storage : in a well-closed container.
White or almost white powder or granules or colourless,
Sodium periodate. NaIO4. (Mr 213.9). 1083200. [7790-28-5].
transparent crystals, deliquescent in moist air, freely soluble in
Sodium metaperiodate.
water, slightly soluble in ethanol (96 per cent).
White or almost white, crystalline powder or crystals, soluble
Sodium nitrite. NaNO2. (Mr 69.0). 1082500. [7632-00-0]. in water and in mineral acids.
Content : minimum 97.0 per cent. Sodium periodate solution. 1083201.
White or almost white, granular powder or a slightly yellow, Dissolve 1.07 g of sodium periodate R in water R, add
crystalline powder, freely soluble in water. 5 mL of dilute sulfuric acid R and dilute to 100.0 mL with
Assay. Dissolve 0.100 g in 50 mL of water R. Add 50.0 mL of water R. Use a freshly prepared solution.
0.02 M potassium permanganate and 15 mL of dilute sulfuric
acid R. Add 3 g of potassium iodide R. Titrate with 0.1 M Sodium phosphite pentahydrate. Na2HPO3,5H2O.
sodium thiosulfate, using 1.0 mL of starch solution R added (Mr 216.0). 1132200. [13517-23-2].
towards the end of the titration as indicator. White or almost white, crystalline powder, hygroscopic, freely
1 mL of 0.02 M potassium permanganate is equivalent to soluble in water.
3.450 mg of NaNO2. Storage : in an airtight container.

Sodium picrate solution, alkaline. 1083300. The solution should be colourless.

Mix 20 mL of picric acid solution R and 10 mL of a 50 g/L Sodium sulfide solution R1. 1083902.
solution of sodium hydroxide R and dilute to 100 mL with
water R. Prepare by one of the following methods.
Storage : use within 2 days. – Dissolve 5 g of sodium sulfide R in a mixture of 10 mL of
water R and 30 mL of glycerol R.
Sodium potassium tartrate. C4H4KNaO6,4H2O. (Mr 282.2). – Dissolve 5 g of sodium hydroxide R in a mixture of 30 mL
1083500. [6381-59-5]. of water R and 90 mL of glycerol R. Divide the solution
Colourless, prismatic crystals, very soluble in water. into 2 equal portions. Saturate 1 portion with hydrogen
sulfide R, with cooling. Mix the 2 portions.
Sodium 1-propanesulfonate. C3H9SO4Na. (Mr 164.2).
1197600. [304672-01-3]. Sodium propane-1-sulfonate Storage : in a well-filled container, protected from light ; use
monohydrate. within 3 months.
mp : about 250 °C. Sodium sulfite, anhydrous. 1084100. [7757-83-7].
Sodium pyrophosphate. Na4P2O7,10H2O. (Mr 446.1). See Sodium sulfite (0775).
1083600. [13472-36-1]. Tetrasodium diphosphate Sodium sulfite heptahydrate. 1084000. [10102-15-5].
decahydrate.
See Sodium sulfite heptahydrate (0776).
Colourless, slightly efflorescent crystals, freely soluble in water.
Sodium tartrate. C4H4Na2O6,2H2O. (Mr 230.1). 1084200.
Sodium pyruvate. C3H3NaO3. (Mr 110.0). 1204300. [6106-24-7]. Disodium (2R,3R)-2,3-dihydroxybutanedioate
[113-24-6]. 2-Oxopropanoic acid sodium salt. dihydrate.
White or faint yellow powder, soluble in water (100 mg/mL). White or almost white crystals or granules, very soluble in
mp : greater than 300 °C. water, practically insoluble in ethanol (96 per cent).
Sodium rhodizonate. C6Na2O6. (Mr 214.0). 1122300. Sodium taurodeoxycholate. C26H44NNaO6S,H2O.
[523-21-7]. [(3,4,5,6-Tetraoxocyclohex-1-en-1,2- (Mr 539.7). 1155600. [110026-03-4]. Sodium
ylene)dioxy]disodium. 2-[(3,12-dihydroxy-5-cholan-24-oyl)amino]ethanesulfonate
Violet crystals, soluble in water with an orange-yellow colour. monohydrate. 2-[[(3,5,12)-3,12-Dihydroxy-24-oxocholan-24-
Solutions are unstable and must be prepared on the day of use. yl]amino]ethanesulfonic acid monosodium salt monohydrate.
Content : minimum 94 per cent of C26H44NNaO6S,H2O.
Sodium salicylate. 1083700. [54-21-7].
See Sodium salicylate (0413). Sodium tetrahydroborate. NaBH4. (Mr 37.8). 1146900.
[16940-66-2]. Sodium borohydride.
Sodium stearyl fumarate. C22H39NaO4. 1195100. Colourless, hygroscopic crystals, freely soluble in water,
[4070-80-8]. soluble in anhydrous ethanol, decomposing at higher
See Sodium stearyl fumarate (1567). temperature or in the presence of acids or certain metal salts
forming borax and hydrogen.
Sodium sulfate, anhydrous. 1083800. [7757-82-6]. Storage : in an airtight container.
Ignite at 600 °C to 700 °C anhydrous sodium sulfate complying
with the requirements prescribed in the monograph on Sodium tetrahydroborate reducing solution. 1146901.
Anhydrous sodium sulfate (0099). Introduce about 100 mL of water R into a 500 mL
Loss on drying (2.2.32): maximum 0.5 per cent, determined by volumetric flask containing a stirring bar. Add 5.0 g
drying in an oven at 130 °C. of sodium hydroxide R in pellets and 2.5 g of sodium
tetrahydroborate R. Stir until complete dissolution, dilute
Sodium sulfate, anhydrous R1. 1083801. to 500.0 mL with water R and mix. Prepare immediately
Complies with the requirements prescribed for anhydrous before use.
sodium sulfate R with the following maximum contents.
Sodium tetraphenylborate. NaB(C6H5)4. (Mr 342.2).
Cl : 20 ppm. 1084400. [143-66-8].
Pb : 10 ppm. White or slightly yellowish, bulky powder, freely soluble in
As : 3 ppm. water and in acetone.
Ca : 50 ppm.
Sodium tetraphenylborate solution. 1084401.
Fe : 10 ppm.
Filter before use if necessary.
Mg : 10 ppm.
A 10 g/L solution of sodium tetraphenylborate R.
Sodium sulfate decahydrate. Na2SO4,10H2O. (Mr 322.2). Storage : use within 1 week.
1132300. [7727-73-3].
See Sodium sulfate decahydrate (0100). Sodium thioglycollate. C2H3NaO2S. (Mr 114.1). 1084500.
[367-51-1]. Sodium mercaptoacetate.
Sodium sulfide. Na2S,9H2O. (Mr 240.2). 1083900. White or almost white, granular powder or crystals,
[1313-84-4]. Disodium sulfide nonahydrate. hygroscopic, freely soluble in water and in methanol, slightly
Colourless, rapidly yellowing crystals, deliquescent, very soluble in ethanol (96 per cent).
soluble in water. Storage : in an airtight container.
Sodium thiosulfate. 1084600. [10102-17-7].
Sodium sulfide solution. 1083901. See Sodium thiosulfate (0414).
Dissolve 12 g of sodium sulfide R with heating in 45 mL
of a mixture of 10 volumes of water R and 29 volumes of Sodium thiosulfate, anhydrous. Na2S2O3. (Mr 158.1).
glycerol (85 per cent) R, allow to cool and dilute to 100 mL 1180700. [7772-98-7]. Disodium thiosulfate.
with the same mixture of solvents. Content : minimum 98.0 per cent.
Sodium tungstate. Na2WO4,2H2O. (Mr 329.9). 1084700. Starch, soluble. 1085100. [9005-84-9].

[10213-10-2]. Disodium tungstate dihydrate. White or almost white powder.
crystals, freely soluble in water forming a clear solution, Starch iodate paper. 1085101.
practically insoluble in ethanol (96 per cent). Immerse strips of filter paper in 100 mL of iodide-free
starch solution R containing 0.1 g of potassium iodate R.
Sorbitol. 1084800. [50-70-4]. Drain and allow to dry protected from light.
See Sorbitol (0435).
Starch iodide paper. 1085106.
Soya bean lecithin. 1196400. [8030-76-0]. Immerse strips of filter paper in 100 mL of potassium iodide
and starch solution R. Drain and allow to dry protected
Soya-bean oil, refined. 1201500.
from light.
See Soya-bean oil, refined (1473).
Test for sensitivity. Mix 0.05 mL of 0.1 M sodium nitrite
Sphingomyelin from egg yolk. 1199100. [85187-10-6]. with 4 mL of hydrochloric acid R and dilute to 100 mL with
(2R,3S,4E)-2-(Acylamino)-3-hydroxyoctadec-4-en-1-yl water R. Apply one drop of the solution to starch iodide
2-(trimethylazaniumyl)ethyl phosphate. paper ; a blue spot appears.
Squalane. C30H62. (Mr 422.8). 1084900. [111-01-3]. Starch solution. 1085103.

(6Ξ,10Ξ,15Ξ,19Ξ)-2,6,10,15,19,23-Hexamethyltetracosane. Triturate 1.0 g of soluble starch R with 5 mL of water R and
Perhydrosqualene. whilst stirring pour the mixture into 100 mL of boiling
Colourless, oily liquid, freely soluble in fatty oils, slightly water R containing 10 mg of mercuric iodide R.
soluble in acetone, in ethanol (96 per cent), in glacial acetic NOTE : commercially available reagents may be used ;
acid and in methanol. including mercury-free solutions or those containing
: 0.811 to 0.813. alternative preservatives.
: 1.451 to 1.453. Carry out the test for sensitivity each time the reagent is
used.
Stannous chloride. SnCl2,2H2O. (Mr 225.6). 1085000. Test for sensitivity. To a mixture of 1 mL of the starch
[10025-69-1]. Tin dichloride dihydrate. solution and 20 mL of water R, add about 50 mg of
Content : minimum 97.0 per cent of SnCl2,2H2O. potassium iodide R and 0.05 mL of iodine solution R1. The
Colourless crystals, very soluble in water, freely soluble in solution is blue.
ethanol (96 per cent), in glacial acetic acid and in dilute and Starch solution, iodide-free. 1085104.
concentrated hydrochloric acid.
Prepare the solution as prescribed for starch solution R
Assay. Dissolve 0.500 g in 15 mL of hydrochloric acid R in a omitting the mercuric iodide. Prepare immediately before
ground-glass-stoppered flask. Add 10 mL of water R and 5 mL use.
of chloroform R. Titrate rapidly with 0.05 M potassium iodate
until the chloroform layer is colourless. Starch solution R1. 1085105.
1 mL of 0.05 M potassium iodate is equivalent to 22.56 mg of Mix 1 g of soluble starch R and a small amount of cold
SnCl2,2H2O. water R. Add this mixture, while stirring, to 200 mL of
boiling water R. Add 0.25 g of salicylic acid R and boil for
Stannous chloride solution. 1085001. 3 min. Immediately remove from the heat and cool.
Heat 20 g of tin R with 85 mL of hydrochloric acid R until Storage : if long storage is required, the solution shall be
no more hydrogen is released. Allow to cool. stored at 4 °C to 10 °C. A fresh starch solution shall be
Storage : over an excess of tin R, protected from air. prepared when the end-point of the titration from blue to
colourless fails to be sharp. If stored under refrigeration,
Stannous chloride solution R1. 1085002. the starch solution is stable for about 2 to 3 weeks.
Immediately before use, dilute 1 volume of stannous Test for sensitivity. A mixture of 2 mL of starch solution R1,
chloride solution R with 10 volumes of dilute hydrochloric 20 mL of water R, about 50 mg of potassium iodide R and
acid R. 0.05 mL of iodine solution R1 is blue.
Stannous chloride solution R2. 1085003. Starch solution R2. 1085107.
To 8 g of stannous chloride R add 100 mL of a 20 per Triturate 1.0 g of soluble starch R with 5 mL of water R and
cent V/V solution of hydrochloric acid R. Shake until whilst stirring pour the mixture into 100 mL of boiling
dissolved, heating, if necessary, on a water-bath at water R. Use a freshly prepared solution.
50 °C. Pass a current of nitrogen R for 15 min. Prepare
immediately before use. Test for sensitivity. To a mixture of 1 mL of the starch
solution and 20 mL of water R, add about 50 mg of
Stanolone. C19H30O2. (Mr 290.4). 1154400. [521-18-6]. potassium iodide R and 0.05 mL of iodine solution R1. The
17β-Hydroxy-5α-androstan-3-one. solution is blue.
White or almost white powder. Stavudine. 1187000. [3056-17-5].
mp : about 180 °C. See Stavudine (2130).
Standard solution for the micro determination of water. Stearic acid. C18H36O2. (Mr 284.5). 1085200. [57-11-4].
1147300. Octadecanoic acid.
Commercially available standard solution for the coulometric White or almost white powder or flakes, greasy to the touch,
titration of water, containing a certified content of water in a practically insoluble in water, soluble in hot ethanol (96 per
suitable solvent. cent).
Staphylococcus aureus strain V8 protease, type XVII-B. mp : about 70 °C.
1115800. [66676-43-5]. Stearic acid used in the assay of total fatty acids in Saw palmetto
Microbial extracellular proteolytic enzyme. A lyophilised fruit (1848) complies with the following additional test.
powder containing 500 units to 1000 units per milligram of Assay. Gas chromatography (2.2.28) as prescribed in the
solid. monograph Saw palmetto fruit (1848).

Content : minimum 98 per cent, calculated by the Sucrose. 1085700. [57-50-1].

normalisation procedure. See Sucrose (0204).
Stearyl alcohol. C18H38O. (Mr 270.5). 1156400. [112-92-5]. Sudan orange. C16H12N2O. (Mr 248.3). 1110700. [842-07-9].
Octadecan-1-ol. Colour Index No. 12055.
mp : about 60 °C. 1-(Phenylazo)naphthalen-2-ol. Sudan I.
Content : minimum 95 per cent. Orange-red powder, practically insoluble in water, soluble in
methylene chloride.
Stigmasterol. C29H48O. (Mr 412.7). 1141400. [83-48-7].
(22E)-Stigmasta-5,22-dien-3β-ol. (22E)-24-Ethylcholesta- mp : about 131 °C.
5,22-dien-3β-ol. Sudan red G. C17H14N2O2. (Mr 278.3). 1085800.
White or almost white powder, insoluble in water. Schultz No. 149.
mp : about 170 °C. Colour Index No. 12150.
: about – 51, determined with a 20 g/L solution in Solvent Red 1. 1-[(2-Methoxyphenyl)azo]naphtalen-2-ol.
chloroform R. Reddish-brown powder, practically insoluble in water.
Streptomycin sulfate. 1085300. [3810-74-0]. Chromatography. Thin-layer chromatography (2.2.27) using
See Streptomycin sulfate (0053). silica gel G R as the coating substance : apply 10 µL of a 0.1 g/L
solution in methylene chloride R and develop over a path of
Strongly acidic ion-exchange resin. 1085400. 10 cm with the same solvent ; the chromatogram shows only
See ion-exchange resin, strongly acidic R. one principal spot.
Strontium carbonate. SrCO3. (Mr 147.6). 1122700. Sulfanilamide. C6H8N2O2S. (Mr 172.2). 1086100. [63-74-1].
[1633-05-2]. 4-Aminobenzenesulfonamide.
White or almost white, crystalline powder. White or almost white powder, slightly soluble in water, freely
soluble in boiling water, in acetone, in dilute acids and in
Content : minimum 99.5 per cent. solutions of the alkali hydroxides, sparingly soluble in ethanol
Strontium chloride hexahydrate. SrCl2,6H2O. (Mr 266.6). (96 per cent) and in light petroleum.
1167000. [10025-70-4]. mp : about 165 °C.
White or almost white crystals, very soluble in water. Sulfathiazole. C9H9N3O2S2. (Mr 255.3). 1086300. [72-14-0].
mp : about 115 °C (loss of water) and 872 °C. 4-Amino-N-(thiazol-2-yl)benzenesulfonamide.
Strontium selective extraction resin. 1167100. White or yellowish-white powder or crystals, very slightly
soluble in water, soluble in acetone, slightly soluble in ethanol
Commercially available resin prepared by loading a suspension (96 per cent). It dissolves in dilute mineral acids and in
of 4,4′(5′)-di-tert-butylcyclohexano-18-crown-6 (crown ether) solutions of alkali hydroxides and carbonates.
in octanol onto an inert chromatographic support. The bed
density of this resin is approximately 0.35 g/mL. mp : about 200 °C.
Strontium-85 spiking solution. 1166800. Sulfamic acid. H3NO3S. (Mr 97.1). 1085900. [5329-14-6].
Dilute strontium-85 standard solution R to a radioactivity White or almost white crystalline powder or crystals, freely
concentration of approximately 10 kBq/mL with a 0.27 g/L soluble in water, sparingly soluble in acetone, in ethanol
solution of strontium chloride hexahydrate R in a 1.03 g/L (96 per cent) and in methanol.
solution of hydrochloric acid R. mp : about 205 °C, with decomposition.
Strontium-85 standard solution. 1166900. Sulfan blue. C27H31N2NaO6S2. (Mr 566.6). 1086000.

[129-17-9].
A solution of strontium-85 in the form of Sr ions in a 51.5 g/L
2+
solution of hydrochloric acid R. Schultz No. 769.

Strychnine. C21H22N2O2. (Mr 334.4). 1190600. [57-24-9]. Acid Blue 1. Patent Blue VF. Disulfine blue. Blue VS.
(4aR,4bR,5aS,8aR,13aS,15aS)-2,4a,4b,5,5a,7,8,13a,15,15a- Sodium [[[(4-diethylamino)phenyl](2,4-disulfonatophenyl)-
Decahydro-4,6-methano-6H-indolo[3,2,1-ij]oxepino[2,3,4- methylene]cyclohexa-2,5-dien-1-ylidene]diethylammonium.
de]pyrrolo[2,3-h]quinolin-14-one. Strychnidin-10-one. Violet powder, soluble in water. Dilute solutions are blue and
White or almost white, crystalline powder, sparingly soluble turn yellow on the addition of concentrated hydrochloric acid.
in water.
Sulfanilic acid. C6H7NO3S. (Mr 173.2). 1086200. [121-57-3].
mp : about 285 °C. 4-Aminobenzenesulfonic acid.
Styrene. C8H8. (Mr 104.2). 1151700. [100-42-5]. Colourless crystals, sparingly soluble in water, practically
Ethenylbenzene. insoluble in ethanol (96 per cent).
bp : about 145 °C. Sulfanilic acid solution. 1086203.
Colourless, oily liquid, very slightly soluble in water. Dissolve 0.33 g of sulfanilic acid R in 75 mL of water R
Styrene-divinylbenzene copolymer. 1085500. heating gently if necessary and dilute to 100 mL with glacial
acetic acid R.
Porous, rigid, cross-linked polymer beads. Several grades are
available with different sizes of beads. The size range of the Sulfanilic acid solution R1. 1086201.
beads is specified after the name of the reagent in the tests Dissolve 0.5 g of sulfanilic acid R in a mixture of 75 mL of
where it is used. dilute acetic acid R and 75 mL of water R.
Succinic acid. C4H6O4. (Mr 118.1). 1085600. [110-15-6]. Sulfanilic acid solution, diazotised. 1086202.
Butanedioic acid.
Dissolve, with warming, 0.9 g of sulfanilic acid R in 9 mL
White or almost white, crystalline powder or colourless of hydrochloric acid R, and dilute to 100 mL with water R.
crystals, soluble in water and in ethanol (96 per cent). Cool 10 mL of this solution in iced water and add 10 mL
mp : 184 °C to 187 °C. of an ice-cold 45 g/L solution of sodium nitrite R. Allow to
stand at 0 °C for 15 min (if stored at this temperature, the To 50 g add 3 mL of nitric acid R and evaporate carefully
solution is stable for 3 days) and immediately before use until the volume is reduced to about 10 mL. Cool, add to the
add 20 mL of a 100 g/L solution of sodium carbonate R. residue 20 mL of water R and concentrate to 5 mL. Prepare
the standard using 1.0 mL of arsenic standard solution (1 ppm
Sulfomolybdic reagent R2. 1086400. As) R.
Dissolve about 50 mg of ammonium molybdate R in 10 mL Iron (2.4.9) : maximum 1 ppm.
of sulfuric acid R.
Dissolve the residue on ignition with slight heating in 1 mL of
Sulfomolybdic reagent R3. 1086500. dilute hydrochloric acid R and dilute to 50.0 mL with water R.
Dissolve with heating 2.5 g of ammonium molybdate R in Dilute 5 mL of this solution to 10 mL with water R.
20 mL of water R. Dilute 28 mL of sulfuric acid R in 50 mL Heavy metals (2.4.8) : maximum 2 ppm.
of water R, then cool. Mix the two solutions and dilute to Dilute 10 mL of the solution obtained in the test for iron to
100 mL with water R. 20 mL with water R. 12 mL of the solution complies with
Storage : in a polyethylene container. test A. Prepare the reference solution using lead standard
solution (2 ppm Pb) R.
Sulfosalicylic acid. C7H6O6S,2H2O. (Mr 254.2). 1086600. Residue on ignition : maximum 0.001 per cent, determined
[5965-83-3]. 2-Hydroxy-5-sulfobenzoic acid. on 100 g by evaporating cautiously in a small crucible over a
White or almost white, crystalline powder or crystals, very naked flame and igniting the residue to redness.
soluble in water and in ethanol (96 per cent). Assay. Weigh accurately a ground-glass-stoppered flask
mp : about 109 °C. containing 30 mL of water R, introduce 0.8 mL of the sulfuric
acid, cool and weigh again. Titrate with 1 M sodium hydroxide,
Sulfur. 1110800. [7704-34-9]. using 0.1 mL of methyl red solution R as indicator.
See Sulfur for external use (0953). 1 mL of 1 M sodium hydroxide is equivalent to 49.04 mg of
Sulfur dioxide. SO2. (Mr 64.1). 1086700. [7446-09-5]. H2SO4.
Sulfurous anhydride. Storage : in a ground-glass-stoppered container made of glass
A colourless gas. When compressed it is a colourless liquid. or other inert material.
5 M Sulfuric acid. 1086809.
Sulfur dioxide R1. SO2. (Mr 64.1). 1110900. [7446-09-5].
Dilute 28 mL of sulfuric acid R to 100 mL with water R.
Sulfuric acid, alcoholic, 2.5 M. 1086801.
Sulfuric acid. H2SO4. (Mr 98.1). 1086800. [7664-93-9].
Carefully and with constant cooling, stir 14 mL of sulfuric
Content : 95.0 per cent m/m to 97.0 per cent m/m. acid R into 60 mL of anhydrous ethanol R. Allow to cool
Colourless, caustic liquid with an oily consistency, highly and dilute to 100 mL with anhydrous ethanol R. Prepare
hygroscopic, miscible with water and with ethanol (96 per immediately before use.
cent) producing intense heat.
Sulfuric acid, alcoholic, 0.25 M. 1086802.
: 1.834 to 1.837.
Dilute 10 mL of 2.5 M alcoholic sulfuric acid R to 100 mL
A 10 g/L solution is strongly acid and gives the reactions of
with anhydrous ethanol R. Prepare immediately before use.
sulfates (2.3.1).
Appearance. It is clear (2.2.1) and colourless (2.2.2, Method II). Sulfuric acid, alcoholic solution of. 1086803.
Oxidisable substances. Pour 20 g cautiously, with cooling, Carefully and with constant cooling, stir 20 mL of sulfuric
into 40 mL of water R. Add 0.5 mL of 0.002 M potassium acid R into 60 mL of ethanol (96 per cent) R. Allow to cool
permanganate. The violet colour persists for at least 5 min. and dilute to 100 mL with ethanol (96 per cent) R. Prepare
Chlorides : maximum 0.5 ppm. immediately before use.
Pour 10 g, carefully and while cooling, into 10 mL of water R Sulfuric acid, dilute. 1086804.
and after cooling dilute to 20 mL with the same solvent. Contains 98 g/L of H2SO4.
Add 0.5 mL of silver nitrate solution R2. Allow to stand for Add 5.5 mL of sulfuric acid R to 60 mL of water R, allow to
2 min protected from bright light. The solution is not more cool and dilute to 100 mL with the same solvent.
opalescent than a standard prepared at the same time using
a mixture of 1 mL of chloride standard solution (5 ppm Cl) R, Assay. Into a ground-glass-stoppered flask containing
19 mL of water R and 0.5 mL of silver nitrate solution R2. 30 mL of water R, introduce 10.0 mL of the dilute sulfuric
acid. Titrate with 1 M sodium hydroxide, using 0.1 mL of
Nitrates : maximum 0.5 ppm. methyl red solution R as indicator.
Pour 50 g or 27.2 mL, carefully and while cooling, into 15 mL 1 mL of 1 M sodium hydroxide is equivalent to 49.04 mg
of water R. Add 0.2 mL of a freshly prepared 50 g/L solution of H2SO4.
of brucine R in glacial acetic acid R. After 5 min any colour
is less intense than that of a reference mixture prepared in Sulfuric acid, dilute R1. 1086810.
the same manner and containing 12.5 mL of water R, 50 g Contains 4.9 g/L of H2SO4.
of nitrogen-free sulfuric acid R, 2.5 mL of nitrate standard Prepared from sulfuric acid R.
solution (10 ppm NO3) R and 0.2 mL of a 50 g/L solution of
brucine R in glacial acetic acid R. Sulfuric acid-formaldehyde reagent. 1086805.
Ammonium : maximum 2 ppm. Mix 2 mL of formaldehyde solution R with 100 mL of
Pour 2.5 g, carefully and while cooling, into water R and dilute sulfuric acid R.
to 20 mL with the same solvent. Cool, and add dropwise Sulfuric acid, heavy metal-free. 1086807.
10 mL of a 200 g/L solution of sodium hydroxide R, followed by
1 mL of alkaline potassium tetraiodomercurate solution R. The Complies with the requirements prescribed for sulfuric
colour of the solution is less intense than that of a mixture of acid R with the following maximum contents of heavy
5 mL of ammonium standard solution (1 ppm NH4) R, 15 mL metals.
of water R, 10 mL of a 200 g/L solution of sodium hydroxide R As : 0.005 ppm.
and 1 mL of alkaline potassium tetraiodomercurate solution R. Cd : 0.002 ppm.
Arsenic (2.4.2, Method A) : maximum 0.02 ppm. Cu : 0.001 ppm.

Fe : 0.05 ppm. d 420 : about 0.837.

Hg : 0.005 ppm. nD20 : about 1.478.
Ni : 0.002 ppm. bp : about 174 °C.
Pb : 0.001 ppm. α-Terpinene used in gas chromatography complies with the
Zn : 0.005 ppm. following additional test.
Sulfuric acid, nitrogen-free. 1086806. Assay. Gas chromatography (2.2.28) as prescribed in the
Complies with the requirements prescribed for sulfuric monograph Tea tree oil (1837).
acid R with the following additional test. Content : minimum 90 per cent, calculated by the
Nitrates. To 5 mL of water R add carefully 45 mL of the normalisation procedure.
sulfuric acid, allow to cool to 40 °C and add 8 mg of γ-Terpinene. C10H16. (Mr 136.2). 1115900. [99-85-4].
diphenylbenzidine R. The solution is colourless or very pale 1-Isopropyl-4-methylcyclohexa-1,4-diene.
blue.
Oily liquid.
Sulfuric acid, nitrogen-free R1. 1086808. γ-Terpinene used in gas chromatography complies with the
Complies with the requirements prescribed for nitrogen-free following additional test.
sulfuric acid R. Assay. Gas chromatography (2.2.28) as prescribed in the
Content : 95.0 per cent m/m to 95.5 per cent m/m. monograph Peppermint oil (0405).
Sulfuric acid R1. H2SO4. (Mr 98.1). 1190900. [7664-93-9]. Test solution. The substance to be examined.
Content : 75 per cent V/V. Content : minimum 93.0 per cent, calculated by the
Sunflower oil. 1086900.
See Sunflower oil, refined (1371). Terpinen-4-ol. C10H18O. (Mr 154.2). 1116000. [562-74-3].
4-Methyl-1-(1-methylethyl)cyclohex-3-en-1-ol.
Swertiamarin. C16H22O10. (Mr 374.3). 1163600. p-Menth-1-en-4-ol.
[17388-39-5]. Swertiamaroside. (4R,5R,6S)-5-Ethenyl-6-(β-D- Oily, colourless liquid.
glucopyranosyloxy)-4a-hydroxy-4,4a,5,6-tetrahydro-1H,3H-
pyrano[3,4-c]pyran-1-one. Terpinen-4-ol used in gas chromatography complies with the
Tagatose. C6H12O6. (Mr 180.16). 1111000. [87-81-0]. Assay. Gas chromatography (2.2.28) as prescribed in the
D-lyxo-Hexulose. monograph Lavender oil (1338).
White or almost white powder. Test solution. The substance to be examined.
: − 2.3 determined on a 21.9 g/L solution. Content : minimum 90.0 per cent, calculated by the
mp : 134 °C to 135 °C. normalisation procedure.
Talc. 1087000. [14807-96-6]. α-Terpineol. C10H18O. (Mr 154.2). 1087300. [98-55-5].
See Talc (0438). (RS)-2-(4-Methylcyclohex-3-enyl)-2-propanol.
Tannic acid. 1087100. [1401-55-4]. Colourless crystals, practically insoluble in water, soluble in
Yellowish or light-brown, glistening scales or amorphous
powder, very soluble in water, freely soluble in ethanol (96 per : about 0.935.
cent), soluble in acetone. : about 1.483.
Storage : protected from light. : about 92.5.
Tanshinone IIA. C19H18O3. (Mr 294.3). 1184800. [568-72-9]. mp : about 35 °C.
1,6,6-Trimethyl-6,7,8,9-tetrahydrophenanthro[1,2-b]furan- It may contain 1 to 3 per cent of β-terpineol.
10,11-dione. α-Terpineol used in gas chromatography complies with the
following test.
Tartaric acid. 1087200. [87-69-4].
See Tartaric acid (0460). monograph Anise oil (0804).
Taxifolin. C15H12O7. (Mr 304.3). 1151800. [480-18-2]. Test solution. A 100 g/L solution in hexane R.
(2R,3R)-2-(3,4-Dihydroxyphenyl)-3,5,7-trihydroxy-2,3- Content : minimum 97.0 per cent, calculated by the
dihydro-4H-1-benzopyran-4-one. normalisation procedure.
White or almost white powder, slightly soluble in anhydrous
ethanol. Terpinolene. C10H16. (Mr 136.2). 1140400. [586-62-9].
Absorbance (2.2.25). A solution in anhydrous ethanol R shows p-Mentha-1,4(8)-diene. 4-Isopropylidene-1-
an absorption maximum at 290 nm. methylcyclohexene.
Clear, almost colourless liquid.
Tecnazene. C6HCl4NO2. (Mr 260.9). 1132400. [117-18-0].
d 420 : about 0.863.
bp : about 304 °C.
mp : 99 °C to 100 °C. nD20 : about 1.488.
A suitable certified reference solution (10 ng/µL in bp : about 184 °C.
cyclohexane) may be used. Terpinolene used in gas chromatography complies with the
trans-Terpin. C10H20O2. (Mr 172.3). 1205800. [565-50-4]. following additional test.
(1r,4r)-4-(2-Hydroxypropan-2-yl)-1-methylcyclohexan-1-ol. Assay. Gas chromatography (2.2.28) as prescribed in the
p-Menthane-1,8-diol. monograph Tea tree oil (1837).
mp : about 116 °C. Content : minimum 90 per cent, calculated by the
α-Terpinene. C10H16. ( Mr 136.2). 1140300. [99-86-5].
1-Isopropyl-4-methylcyclohexa-1,3-diene. Testosterone. 1116100. [58-22-0].
Clear, almost colourless liquid. See Testosterone (1373).
Testosterone propionate. 1087400. [57-85-2]. Tetrabutylammonium iodide. C16H36IN. (Mr 369.4).

See Testosterone propionate (0297). 1087900. [311-28-4].
1,2,3,4-Tetra-O-acetyl-β- D-glucopyranose. C14H20O10.
White or slightly coloured, crystalline powder or crystals,
(Mr 348.3). 1172600. [13100-46-4].
White or almost white powder, soluble in water with gentle Sulfated ash (2.4.14): maximum 0.02 per cent.
heating.
Assay. Dissolve 1.200 g in 30 mL of water R. Add 50.0 mL of
: + 11, determined on a 6 g/L solution in chloroform R. 0.1 M silver nitrate and 5 mL of dilute nitric acid R. Titrate the
mp : 126 °C to 128 °C. excess of silver nitrate with 0.1 M ammonium thiocyanate,
using 2 mL of ferric ammonium sulfate solution R2 as indicator.
1,3,4,6-Tetra-O-acetyl-β- D-mannopyranose. C14H20O10.
(Mr 348.3). 1174100. [18968-05-3]. 1 mL of 0.1 M silver nitrate is equivalent to 36.94 mg of
C16H36IN.
Colourless or white powder or crystals.
mp : 160 °C to 161 °C. Tetrachloroethane. C2H2Cl4. (Mr 167.9). 1088000. [79-34-5].
1,1,2,2-Tetrachloroethane.
: − 68, determined on a 7 g/L solution in methylene
chloride R. Clear, colourless liquid, slightly soluble in water, miscible with
Tetrabutylammonium bromide. C16H36BrN. (Mr 322.4). : about 1.59.
1087500. [1643-19-2]. : about 1.495.
White or almost white crystals. Distillation range (2.2.11). Not less than 95 per cent distils
mp : 102 °C to 104 °C. between 145 °C and 147 °C.
Tetrabutylammonium dihydrogen phosphate. C16H38NO4P. Tetrachlorvinphos. C10H9Cl4O4P. (Mr 366.0). 1132500.
(Mr 339.5). 1087600. [5574-97-0]. [22248-79-9].
White or almost white powder, hygroscopic. mp : about 95 °C.
pH (2.2.3) : about 7.5 for a 170 g/L solution. A suitable certified reference solution (10 ng/µL in iso-octane)
Absorbance (2.2.25) : about 0.10 determined at 210 nm using a may be used.
170 g/L solution. Tetracos-15-enoic acid methyl ester. C25H48O2. (Mr 380.7).
Storage : in an airtight container. 1144800. [2733-88-2]. 15-Tetracosaenoic acid methyl ester.
Methyl tetracos-15-enoate. Nervonic acid methyl ester.
Tetrabutylammonium dihydrogen phosphate solution.
1087601. Content : minimum 99.0 per cent, determined by gas
chromatography.
A 1.0 M solution of tetrabutylammonium dihydrogen
phosphate R. This solution is commercially available. Liquid.
Tetracycline hydrochloride. 1147000.
Tetrabutylammonium hydrogen sulfate. C16H37NO4S.
(Mr 339.5). 1087700. [32503-27-8]. See Tetracycline hydrochloride (0210).
Crystalline powder or colourless crystals, freely soluble in Tetradecane. C14H30. (Mr 198.4). 1088200. [629-59-4].
water and in methanol. n-Tetradecane.
mp : 169 °C to 173 °C. Content : minimum 99.5 per cent m/m.
Absorbance (2.2.25) : maximum 0.05, determined between A colourless liquid.
240 nm and 300 nm using a 50 g/L solution. : about 0.76.
Tetrabutylammonium hydrogen sulfate R1. 1087701. : about 1.429.
Complies with the requirements prescribed for bp : about 252 °C.
tetrabutylammonium hydrogen sulfate R with the following mp : about − 5 °C.
additional requirement. Tetradecylammonium bromide. C40H84BrN. (Mr 659).
Absorbance (2.2.25): maximum 0.02, determined between 1088300. [14937-42-9]. Tetrakis(decyl)ammonium bromide.
215 nm and 300 nm using a 50 g/L solution. White or slightly coloured, crystalline powder or crystals.
Tetrabutylammonium hydroxide. C16H37NO,30H2O. mp : 88 °C to 89 °C.
(Mr 800). 1087800. [147741-30-8].
Tetraethylammonium hydrogen sulfate. C8H21NO4S.
Content : minimum 98.0 per cent of C16H37NO,30H2O. (Mr 227.3). 1116200. [16873-13-5].
White or almost white crystals, soluble in water. Hygroscopic powder.
Assay. Dissolve 1.000 g in 100 mL of water R. Titrate mp : about 245 °C.
immediately with 0.1 M hydrochloric acid determining the
end-point potentiometrically (2.2.20). Carry out a blank Tetraethylammonium hydroxide solution. C8H21NO.
titration. (Mr 147.3). 1100300. [77-98-5].
1 mL of 0.1 M hydrochloric acid is equivalent to 80.0 mg of A 200 g/L solution.
C16H37NO,30H2O. Colourless liquid, strongly alkaline.
Tetrabutylammonium hydroxide solution (104 g/L). : about 1.01.
1087801. : about 1.372.
A solution containing 104 g/L of C16H37NO (Mr 259.5), HPLC grade.
prepared by dilution of a suitable reagent grade. Tetraethylene pentamine. C8H23N5. (Mr 189.3). 1102000.
Tetrabutylammonium hydroxide solution (400 g/L). [112-57-2]. 3,6,9-Triazaundecan-1,11-diamine.
1087802. Colourless liquid, soluble in acetone.
A solution containing 400 g/L of C16H37NO (Mr 259.5) of : about1.506.
a suitable grade. Storage : protected from humidity and heat.

Tetraheptylammonium bromide. C28H60BrN. (Mr 490.7). Tetramethylammonium hydroxide solution. 1088600.

1088400. [4368-51-8]. [75-59-2].
White or slightly coloured, crystalline powder or crystals. Content : minimum 10.0 per cent m/m of C4H13NO. (Mr 91.2).
mp : 89 °C to 91 °C. Clear, colourless or very pale yellow liquid, miscible with
Tetrahexylammonium bromide. C24H52BrN. (Mr 434.6). water and with ethanol (96 per cent).
1152500. [4328-13-6]. N,N,N-Trihexylhexan-1-aminium Assay. To 1.000 g add 50 mL of water R and titrate with 0.05 M
bromide. sulfuric acid, using 0.1 mL of methyl red solution R as indicator.
White or almost white, crystalline powder, hygroscopic. 1 mL of 0.05 M sulfuric acid is equivalent to 9.12 mg of
mp : about 100 °C. C4H13NO.
Tetrahexylammonium hydrogen sulfate. C24H53NO4S. Tetramethylammonium hydroxide solution, dilute.

(Mr 451.8). 1116300. [32503-34-7]. N,N,N-Trihexylhexan- 1088601.
1-aminium hydrogen sulfate. Dilute 10 mL of tetramethylammonium hydroxide
White or almost white crystals. solution R to 100 mL with aldehyde-free alcohol R. Prepare
mp : 100 °C to 102 °C. immediately before use.
Tetrahydrofuran. C4H8O. (Mr 72.1). 1088500. [109-99-9]. Tetramethylbenzidine. C16H20N2. (Mr 240.3). 1132600.

Tetramethylene oxide. [54827-17-7]. 3,3′,5,5′-Tetramethylbiphenyl-4,4′-diamine.
Clear, colourless, flammable liquid, miscible with water, with Powder, practically insoluble in water, very soluble in
ethanol (96 per cent). methanol.
: about 0.89. mp : about 169 °C.
Do not distil if the tetrahydrofuran does not comply with the 1,1,3,3-Tetramethylbutylamine. C8H19N. (Mr 129.3).
test for peroxides. 1141500. [107-45-9]. 2-Amino-2,4,4-trimethylpentane.
Peroxides. Place 8 mL of potassium iodide and starch solution R
in a 12 mL ground-glass-stoppered cylinder about 1.5 cm in
diameter. Fill completely with the substance to be examined, : about 0.805.
shake vigorously and allow to stand protected from light for : about 1.424.
30 min. No colour is produced. bp : about 140 °C.
Tetrahydrofuran used in spectrophotometry complies with the
following additional test. Tetramethyldiaminodiphenylmethane. C17H22N2.
Absorbance (2.2.25) : maximum 0.70 at 255 nm, 0.10 at (Mr 254.4). 1088700. [101-61-1]. 4,4′-Methylenebis-(N,N-
270 nm, 0.01 at 310 nm, determined using water R as dimethylaniline).
compensation liquid. White or bluish-white crystals or leaflets, practically insoluble
in water, slightly soluble in ethanol (96 per cent), soluble in
Tetrahydrofuran for chromatography. 1147100. mineral acids.
Complies with the requirements prescribed for mp : about 90 °C.
tetrahydrofuran R with the following additional requirements :
d 420 = 0.8892. Tetramethyldiaminodiphenylmethane reagent. 1088701.
bp : about 66 °C. Solution A. Dissolve 2.5 g of tetramethyldiaminodiphenyl-
methane R in 10 mL of glacial acetic acid R and add 50 mL
Content : minimum 99.8 per cent of C4H8O.
of water R.
Tetrahydropalmatine. C21H25NO4. (Mr 355.4). 1205900. Solution B. Dissolve 5 g of potassium iodide R in 100 mL
[2934-97-6]. (13aRS)-5,8,13,13a-Tetrahydro-2,3,9,10- of water R.
tetramethoxy-6H-dibenzo[a,g]quinolizine. Solution C. Dissolve 0.30 g of ninhydrin R in 10 mL of
α-Tetralone. C10H10O. (Mr 146.2). 1171800. [529-34-0]. glacial acetic acid R and add 90 mL of water R.
1-Oxotetraline. 3,4-Dihydronaphthalen-1(2H)-one. Mix solution A, solution B and 1.5 mL of solution C.
bp : about 115 °C.
Tetramethylethylenediamine. C6H16N2. (Mr 116.2). 1088800.
mp : about 5 °C. [110-18-9]. N,N,N’,N’-Tetramethylethylenediamine.
Tetramethylammonium bromide. C4H12BrN. (Mr 154.1). Colourless liquid, miscible with water and with ethanol
1156600. [64-20-0]. N,N,N-Trimethylmethanaminium (96 per cent).
bromide. : about 0.78.
White or slightly yellow crystals, freely soluble in water. : about 1.418.
mp : about 285 °C, with decomposition. bp : about 121 °C.
Tetramethylammonium chloride. C4H12ClN. (Mr 109.6).
1100400. [75-57-0]. Tetramethylsilane. C4H12Si. (Mr 88.2). 1088900. [75-76-3].
TMS.
Colourless crystals, soluble in water and in ethanol (96 per
cent). Clear, colourless liquid, very slightly soluble in water, soluble
in acetone and in ethanol (96 per cent).
: about 0.64.
Tetramethylammonium hydrogen sulfate. C4H13NO4S. : about 1.358.
(Mr 171.2). 1116400. [80526-82-5].
bp : about 26 °C.
Hygroscopic powder.
Tetramethylsilane used in nuclear magnetic resonance
mp : about 295 °C.
spectrometry complies with the following additional test.
Tetramethylammonium hydroxide. C4H13NO,5H2O. In the nuclear magnetic resonance spectrum of
(Mr 181.2). 1122800. [10424-65-4]. Tetramethylammonium an approximately 10 per cent V/V solution of the
hydroxide pentahydrate. tetramethylsilane in deuterated chloroform R, the intensity of
Suitable grade for HPLC. any foreign signal, excluding those due to spinning side bands
and to chloroform, is not greater than the intensity of the C-13 Thioacetamide reagent. 1089601.
satellite signals located at a distance of 59.1 Hz on each side of To 0.2 mL of thioacetamide solution R add 1 mL of a
the principal signal of tetramethylsilane. mixture of 5 mL of water R, 15 mL of 1 M sodium hydroxide
and 20 mL of glycerol (85 per cent) R. Heat in a water-bath
Tetrandrine. C38H42N2O6. (Mr 623). 1178500. [518-34-3].
for 20 s. Prepare immediately before use.
Tetrapropylammonium chloride. C12H28ClN. (Mr 221.8). Thioacetamide solution. 1089602.
1151900. [5810-42-4].
A 40 g/L solution of thioacetamide R.
White or almost white, crystalline powder, sparingly soluble
in water. Thiobarbituric acid. C4H4N2O2S. (Mr 144.2). 1111200.
mp : about 241 °C. [504-17-6]. 4,6-Dihydroxy-2-sulfanylpyrimidine.
Tetrapropylammonium hydrogen sulfate. C12H29NO4S. Thiodiethylene glycol. C4H10O2S. (Mr 122.2). 1122900.

(Mr 283.4). 1191300. [56211-70-2]. N,N,N-Tripropyl- [111-48-8]. Di(2-hydroxyethyl) sulfide.
propan-1-aminium hydrogen sulfate. Colourless or yellow, viscous liquid.
White or almost white, crystalline, hygroscopic powder. Content : minimum 99.0 per cent.
: about 1.18.
Tetrazolium blue. C40H32Cl2N8O2. (Mr 728). 1089000.
[1871-22-3]. 3,3′-(3,3′-Dimethoxy[1,1′-biphenyl]-4,4′- Thioglycollic acid. C2H4O2S. (Mr 92.1). 1089700. [68-11-1].
diyl)bis[2,5-diphenyl-2H-tetrazolium] dichloride. 2-Mercaptoacetic acid.
Yellow crystals, slightly soluble in water, freely soluble in Colourless liquid, miscible with water, soluble in ethanol
ethanol (96 per cent) and in methanol, practically insoluble (96 per cent).
in acetone.
Thiomalic acid. C4H6O4S. (Mr 150.2). 1161600. [70-49-5].
mp : about 245 °C, with decomposition. (2RS)-2-Sulfanylbutanedioic acid.
Tetrazolium bromide. C18H16BrN5S. (Mr 414.3). mp : 150 °C to 152 °C.
1152700. [298-93-1]. 3-(4,5-Dimethylthiazol-2-yl)-2,5- Thiomersal. C9H9HgNaO2S. (Mr 404.8). 1089800.
diphenyltetrazolium bromide. MTT. [54-64-8]. Sodium mercurothiolate. Sodium
Tetrazolium salt. C20H17N5O6S2. (Mr 487.5). 1174200. 2-[(ethylmercurio)thio]benzoate.
[138169-43-4]. 5-(3-Carboxymethoxyphenyl)-3-(4,5- Light, yellowish-white, crystalline powder, very soluble in
dimethylthiazol-2-yl)-2-(4-sulfophenyl)-2H-tetrazolium, water, freely soluble in ethanol (96 per cent).
inner salt. MTS.
Thiourea. CH4N2S. (Mr 76.1). 1089900. [62-56-6].
Thallous sulfate. Tl2SO4. (Mr 504.8). 1089100. [7446-18-6]. White or almost white, crystalline powder or crystals, soluble
Dithallium sulfate. in water and in ethanol (96 per cent).
White or almost white, rhomboid prisms, slightly soluble in mp : about 178 °C.
water, practically insoluble in ethanol (96 per cent).
Threonine. 1090000. [72-19-5].
Thebaine. C19H21NO3. (Mr 311.4). 1089200. [115-37-7]. See Threonine (1049).
(5R,9R,13S)-4,5-Epoxy-3,6-dimethoxy-9a-methylmorphina-
6,8-diene. Thrombin, bovine. 1090200. [9002-04-4].
White or pale yellow, crystalline powder, very slightly soluble A preparation of the enzyme, obtained from bovine plasma,
in water, soluble in hot anhydrous ethanol and in toluene. that converts fibrinogen into fibrin.
mp : about 193 °C. A yellowish-white powder.
Chromatography (2.2.27). Thin-layer chromatography (2.2.27) Storage : at a temperature below 0 °C.
as prescribed in identification test B in the monograph Raw Thrombin, human. 1090100. [9002-04-4].
opium (0777) : apply to the plate as a band (20 mm × 3 mm)
Dried human thrombin. A preparation of the enzyme which
20 μL of a 0.5 g/L solution ; the chromatogram shows an
converts human fibrinogen into fibrin. It is obtained from
orange-red or red principal band with an RF of about 0.5.
liquid human plasma and may be prepared by precipitation
Theobromine. 1138800. [83-67-0]. with suitable salts and organic solvents under controlled
See Theobromine (0298). conditions of pH, ionic strength and temperature.
Yellowish-white powder, freely soluble in a 9 g/L solution of
Theophylline. 1089300. [58-55-9]. sodium chloride forming a cloudy, pale yellow solution.
See Theophylline (0299). Storage : in a sealed, sterile container under nitrogen, protected
from light, at a temperature below 25 °C.
Thiamazole. C4H6N2S. (Mr 114.2). 1089400. [60-56-0].
Methimazole. 1-Methyl-1H-imidazole-2-thiol. Thrombin solution, human. 1090101.
White or almost white, crystalline powder, freely soluble Reconstitute human thrombin R as directed by
in water, soluble in ethanol (96 per cent) and in methylene the manufacturer and dilute to 5 IU/mL with
chloride. tris(hydroxymethyl)aminomethane sodium chloride buffer
mp : about 145 °C. solution pH 7.4 R.
2-(2-Thienyl)acetic acid. C6H6O2S. (Mr 142.1). 1089500. Thrombin solution, human R1. 1090102.

[1918-77-0]. Reconstitute human thrombin R as directed by the
Brown powder. manufacturer and dilute to 2.5 IU/mL with phosphate
buffer solution pH 6.5 R.
mp : about 65 °C.
Thrombin solution, human R2. 1090103.
Thioacetamide. C2H5NS. (Mr 75.1). 1089600. [62-55-5].
Reconstitute human thrombin R as directed by
Crystalline powder or colourless crystals, freely soluble in the manufacturer and dilute to 5 IU/mL with
water and in ethanol (96 per cent). tris(hydroxymethyl)aminomethane-EDTA buffer solution
mp : about 113 °C. pH 8.4 R1.

Thromboplastin. 1090300. Tin. Sn. (Ar 118.7). 1090800. [7440-31-5].

A preparation containing the membrane glycoprotein tissue Silvery-white granules, soluble in hydrochloric acid with
factor and phospholipid, either purified from animal brain release of hydrogen.
(usually rabbit) or human placenta or manufactured using Arsenic (2.4.2, Method A) : maximum 10 ppm, determined
recombinant DNA technology with added phospholipid. The on 0.1 g.
preparation is formulated for routine use in the prothrombin
time test and may contain calcium. Tin test kit, semi-quantitative. 1194100.
Commercially available set of reagents consisting of tin test
Thujone. C10H16O. (Mr 152.2). 1116500. [76231-76-0]. strips and a reagent mixture for the determination of tin in
4-Methyl-1-(1-methylethyl)bicyclo[3.1.0]hexan-3-one. aqueous solutions, in a range of 10-200 µg/mL.
Colourless or almost colourless liquid, practically insoluble Titan yellow. C28H19N5Na2O6S4. (Mr 696). 1090900.
in water, soluble in ethanol (96 per cent) and in many other [1829-00-1].
organic solvents.
Schultz No. 280.
Thymidine. C10H14N2O5. (Mr 242.2). 1158900. 1-(2-Deoxy-β- Colour Index No. 19540.
D-erythro-pentofuranosyl)-5-methylpyrimidine-2,4(1H,3H)- Thiazol yellow. Disodium 2,2′-[(1-triazene-1,3-diyl)di-4,1-
dione. phenylene]bis-[6-methylbenzothiazole-7-sulfonate].
Needles, soluble in water, in hot ethanol (96 per cent) and in A yellowish-brown powder, freely soluble in water and in
glacial acetic acid. ethanol (96 per cent).
Titan yellow paper. 1090901.
Thymine. C5H6N2O2. (Mr 126.1). 1090400. [65-71-4].
5-Methylpyrimidine-2,4(1H,3H)-dione. Immerse strips of filter paper in titan yellow solution R and
leave for a few minutes. Allow to dry at room temperature.
Short needles or plates, slightly soluble in cold water, soluble
in hot water. It dissolves in dilute solution of alkali hydroxides. Titan yellow solution. 1090902.
A 0.5 g/L solution of titan yellow R.
Thymol. 1090500. [89-83-8]. See Thymol (0791).
Test for sensitivity. To 0.1 mL of the titan yellow solution
Thymol used in gas chromatography complies with the following add 10 mL of water R, 0.2 mL of magnesium standard
additional test. solution (10 ppm Mg) R and 1.0 mL of 1 M sodium
Assay. Gas chromatography (2.2.28) as prescribed in the hydroxide. A distinct pink colour is visible by comparison
monograph Peppermint oil (0405). with a reference solution prepared in a similar manner
omitting the magnesium.
Test solution. Dissolve 0.1 g in about 10 mL of acetone R.
Content : minimum 95.0 per cent, calculated by the Titanium. Ti. (Ar 47.88). 1091000. [7440-32-6].
normalisation procedure. Content : minimum 99 per cent.
Metal powder, fine wire (diameter not more than 0.5 mm),
Thymol blue. C27H30O5S. (Mr 466.6). 1090600. [76-61-9]. sponge.
Thymolsulfonphthalein. 4,4′-(3H-2,1-Benzoxathiol-3-
mp : about 1668 °C.
ylidene)bis(2-isopropyl-5-methylphenol) S,S-dioxide.
Density : about 4.507 g/cm3.
Brownish-green or greenish-blue, crystalline powder, slightly
soluble in water, soluble in ethanol (96 per cent) and in dilute Titanium dioxide. 1117900. [13463-67-7].
solutions of alkali hydroxides. See Titanium dioxide (0150).
Thymol blue solution. 1090601. Titanium trichloride. TiCl3. (Mr 154.3). 1091200.
Dissolve 0.1 g of thymol blue R in a mixture of 2.15 mL [7705-07-9]. Titanium(III) chloride.
of 0.1 M sodium hydroxide and 20 mL of ethanol (96 per Reddish-violet crystals, deliquescent, soluble in water and in
cent) R and dilute to 100 mL with water R. ethanol (96 per cent).
Test for sensitivity. To 0.1 mL of the thymol blue solution mp : about 440 °C.
add 100 mL of carbon dioxide-free water R and 0.2 mL of Storage : in an airtight container.
0.02 M sodium hydroxide. The solution is blue. Not more
than 0.15 mL of 0.02 M hydrochloric acid is required to Titanium trichloride solution. 1091201.
change the colour to yellow. : about 1.19.
Colour change : pH 1.2 (red) to pH 2.8 (yellow) ; pH 8.0 A 150 g/L solution of titanium trichloride R in hydrochloric
(olive-green) to pH 9.6 (blue). acid (100 g/L HCl).
Titanium trichloride-sulfuric acid reagent. 1091202.
Thymolphthalein. C28H30O4. (Mr 430.5). 1090700.
[125-20-2]. 3,3-Bis(4-hydroxy-5-isopropyl-2-methylphenyl)- Carefully mix 20 mL of titanium trichloride solution R with
3H-isobenzo-furan-1-one. 13 mL of sulfuric acid R. Add sufficient strong hydrogen
peroxide solution R to give a yellow colour. Heat until white
White or yellowish-white powder, practically insoluble in fumes are evolved. Allow to cool. Dilute with water R
water, soluble in ethanol (96 per cent) and in dilute solutions and repeat the evaporation and addition of water R until
of alkali hydroxides. a colourless solution is obtained. Dilute to 100 mL with
water R.
Thymolphthalein solution. 1090701.
A 1 g/L solution of thymolphthalein R in ethanol (96 per TLC aluminium oxide G plate. 1165200.
cent) R. Support of metal, glass or plastic, coated with a layer of
aluminium oxide (particle size 5-40 μm) containing about
Test for sensitivity. To 0.2 mL of the thymolphthalein
10 per cent of calcium sulfate hemihydrate as a binder.
solution add 100 mL of carbon dioxide-free water R. The
solution is colourless. Not more than 0.05 mL of 0.1 M TLC cellulose plate. 1191400.
sodium hydroxide is required to change the colour to blue. Support of glass, metal or plastic, coated with a layer of
Colour change : pH 9.3 (colourless) to pH 10.5 (blue). cellulose.
TLC octadecylsilyl silica gel plate. 1148600. TLC silica gel GF254 plate. 1117000.
Support of glass, metal or plastic coated with a layer of Complies with the requirements prescribed for TLC silica gel
octadecylsilyl silica gel. The plate may contain an organic plate R with the following modifications.
binder. It contains calcium sulfate hemihydrate as binder and a
fluorescent indicator having a maximum absorbance at
TLC octadecylsilyl silica gel F254 plate. 1146600. 254 nm.
Support of glass, metal or plastic coated with a layer of Fluorescence suppression. Complies with the test prescribed
octadecylsilyl silica gel. for TLC silica gel F254 plate R.
It contains a fluorescent indicator having a maximum
absorbance in ultraviolet light at 254 nm. TLC silica gel plate for aminopolyether test. 1172700.
Immerse a TLC silica gel plate R in iodoplatinate reagent R1 for
TLC performance test solution. 1116600. 5-10 s. Dry at room temperature for 12 h, protected from light.
Prepare a mixture of 1.0 mL of each of the following solutions Storage : protected from light, in an open container ; use within
and dilute to 10.0 mL with acetone R : a 0.5 g/L solution of 30 days after preparation.
Sudan red G R in toluene R, a 0.5 g/L solution of methyl
orange R in ethanol R prepared immediately before use, a TLC silica gel plate for chiral separations, octadecylsilyl.
0.5 g/L solution of bromocresol green R in acetone R and a 1137700.
0.25 g/L solution of methyl red R in acetone R. Support of glass, metal or plastic, coated with a layer of
octadecylsilyl silica gel, impregnated with Cu2+ ions and
TLC silica gel plate. 1116700. enantiomerically pure hydroxyproline. The plate may contain
Support of glass, metal or plastic, coated with a layer of silica an organic binder.
gel of a suitable thickness and particle size (usually 2 µm to TLC silica gel, silanised plate. 1117100.
10 µm for fine particle size [High Performance Thin-Layer
Chromatography, HPTLC] plates and 5 µm to 40 μm for Support of glass, metal or plastic, coated with a layer of
normal TLC plates). If necessary, the particle size is indicated silanised silica gel of a suitable thickness and particle
after the name of the reagent in the tests where it is used. size (usually 2 µm to 10 µm for fine particle size [High
Performance Thin-Layer Chromatography, HPTLC] plates
The plate may contain an organic binder. and 5 μm to 40 μm for normal TLC plates). If necessary, the
Chromatographic separation. Apply to the plate an appropriate particle size is indicated after the name of the reagent in the
volume (10 µL for a normal TLC plate and 1 μL to 2 μL tests where it is used.
for a fine particle size plate) of TLC performance test The plate may contain an organic binder.
solution R. Develop over a pathlength two-thirds of the plate
Chromatographic separation. Introduce 0.1 g each of methyl
height, using a mixture of 20 volumes of methanol R and
laurate R, methyl myristate R, methyl palmitate R and methyl
80 volumes of toluene R. The plate is not satisfactory, unless
stearate R into a 250 mL conical flask. Add 40 mL of alcoholic
the chromatogram shows four clearly separated spots, the
potassium hydroxide solution R and heat under a reflux
spot of bromocresol green with an RF value less than 0.15, the
condenser on a water-bath for 1 h. Allow to cool, transfer
spot of methyl orange with an RF value in the range of 0.1 to
the solution to a separating funnel by means of 100 mL of
0.25, the spot of methyl red with an RF value in the range of
water R, acidify (pH 2 to 3) with dilute hydrochloric acid R
0.35 to 0.55 and the spot of Sudan red G with an RF value in
and shake with three quantitites each of 10 mL of methylene
the range of 0.75 to 0.98.
chloride R. Dry the combined methylene chloride extracts over
TLC silica gel F254 plate. 1116800. anhydrous sodium sulfate R, filter and evaporate to dryness
on a water-bath. Dissolve the residue in 50 mL of methylene
Complies with the requirements prescribed for TLC silica gel chloride R. Examine by thin-layer chromatography (2.2.27),
plate R with the following modification. using TLC silanised silica gel plate R. Apply an appropriate
It contains a fluorescent indicator having a maximum quantity (about 10 µL for normal TLC plates and about 1 µL
absorbance at 254 nm. to 2 μL for fine particle size plates) of the methylene chloride
Fluorescence suppression. Apply separately to the plate at five solution at each of three separate points. Develop over a
points increasing volumes (1 μL to 10 µL for normal TLC pathlength two-thirds of the plate height with a mixture of
plates and 0.2 µL to 2 µL for fine particle size plates) of a 10 volumes of glacial acetic acid R, 25 volumes of water R and
1 g/L solution of benzoic acid R in a mixture of 15 volumes 65 volumes of dioxan R. Dry the plate at 120 °C for 30 min.
of anhydrous ethanol R and 85 volumes of cyclohexane R. Allow to cool, spray with a 35 g/L solution of phosphomolybdic
Develop over a pathlength half of the plate height with the acid R in 2-propanol R and heat at 150 °C until the spots
same mixture of solvents. After evaporating the solvents become visible. Treat the plate with ammonia vapour until the
examine the chromatogram in ultraviolet light at 254 nm. For background is white. The chromatograms show four clearly
normal TLC plates the benzoic acid appears as dark spots separated, well-defined spots.
on a fluorescent background approximately in the middle of α-Tocopherol. 1152300. [10191-41-0].
the chromatogram for quantities of 2 µg and greater. For fine
particle size plates the benzoic acid appears as dark spots on See all-rac-α-Tocopherol (0692).
a fluorescent background approximately in the middle of the α-Tocopheryl acetate. 1152400. [7695-91-2].
chromatogram for quantities of 0.2 µg and greater.
See all-rac-α-Tocopheryl acetate (0439).
TLC silica gel F254, silanised plate. 1117200. o-Tolidine. C14H16N2. (Mr 212.3). 1123000. [119-93-7].
It complies with the requirements prescribed for TLC silanised 3,3′-Dimethylbenzidine.
silica gel plate R with the following modification. Content : minimum 97.0 per cent.
It contains a fluorescent indicator having a maximum Light brownish, crystalline power.
absorbance at 254 nm.
mp : about 130 °C.
TLC silica gel G plate. 1116900. o-Tolidine solution. 1123001.
Complies with the requirements prescribed for TLC silica gel Dissolve 0.16 g of o-tolidine R in 30.0 mL of glacial acetic
plate R with the following modification. acid R, add 1.0 g of potassium iodide R and dilute to
It contains calcium sulfate hemihydrate as binder. 500.0 mL with water R.

Toluene. C7H8. (Mr 92.1). 1091300. [108-88-3]. Toluidine blue. C15H16ClN3S. (Mr 305.8). 1091900. [92-31-9].
Methylbenzene. Schultz No. 1041.
Clear, colourless, flammable liquid, very slightly soluble in Colour Index No. 52040.
water, miscible with ethanol (96 per cent). Toluidine Blue O. 3-Amino-7-dimethylamino-2-
: 0.865 to 0.870. methylphenothiazin-5-ium chloride.
bp : about 110 °C. Dark-green powder, soluble in water, slightly soluble in
Toluene, sulfur-free. 1091301.
Complies with the requirements prescribed for toluene R Tosylarginine methyl ester hydrochloride.
with the following additional requirements. C14H23ClN4O4S. (Mr 378.9). 1092000. [1784-03-8].
N-Tosyl-L-arginine methyl ester hydrochloride. Methyl
Sulfur compounds. To 10 mL add 1 mL of anhydrous (S)-5-guanidino-2-(4-methylbenzenesulfonamido)valerate
ethanol R and 3 mL of potassium plumbite solution R and hydrochloride.
boil under a reflux condenser for 15 min. Allow to stand
for 5 min. No darkening is produced in the aqueous layer. : − 12 to − 16, determined on a 40 g/L solution.
Thiophen-related substances. Shake 2 mL with 5 mL of mp : about 145 °C.
isatin reagent R for 5 min and allow to stand for 15 min. Tosylarginine methyl ester hydrochloride solution.
No blue colour is produced in the lower layer. 1092001.
Toluenesulfonamide. C7H9NO2S. (Mr 171.2). To 98.5 mg of tosylarginine methyl ester hydrochloride R add
1091500. [70-55-3]. 4-Methylbenzenesulfonamide. 5 mL of tris(hydroxymethyl)aminomethane buffer solution
p-Toluenesulfonamide. pH 8.1 R and shake to dissolve. Add 2.5 mL of methyl red
Content : minimum 99.0 per cent. mixed solution R and dilute to 25.0 mL with water R.
White or almost white, crystalline powder, slightly soluble Tosyl-lysyl-chloromethane hydrochloride.
in water, soluble in ethanol (96 per cent) and in solutions of C14H22Cl2N2O3S. (Mr 369.3). 1092100. [4238-41-9].
alkali hydroxides. N-Tosyl-L-lysyl-chloromethane hydrochloride. (3S)-7-Amino-
mp : about 136 °C. 1-chloro-3-(4-methylbenzenesulfonamido)heptan-2-one
hydrochloride.
o-Toluenesulfonamide. C7H9NO2S. (Mr 171.2). 1091400. : − 7 to − 9, determined on a 20 g/L solution.
[88-19-7]. 2-Methylbenzenesulfonamide.
White or almost white, crystalline powder, slightly soluble
in water, soluble in ethanol (96 per cent) and in solutions of : 310 to 340, determined at 230 nm in water R.
alkali hydroxides. Tosylphenylalanylchloromethane. C17H18ClNO3S.
mp : about 156 °C. (Mr 351.9). 1092200. [402-71-1]. N-Tosyl-L-
phenylalanylchloromethane.
p-Toluenesulfonamide. 1091500. [70-55-3].
: − 85 to − 89, determined on a 10 g/L solution in ethanol
See toluenesulfonamide R. (96 per cent) R.
Toluenesulfonic acid. C7H8O3S,H2O. (Mr 190.2). 1091600. mp : about 105 °C.
[6192-52-5]. 4-Methylbenzenesulfonic acid. : 290 to 320, determined at 228.5 nm in ethanol (96 per
Content : minimum 87.0 per cent of C7H8O3S. cent) R.
White or almost white, crystalline powder or crystals, freely Toxaphene. 1132800. [8001-35-2].
soluble in water, soluble in ethanol (96 per cent). A mixture of polychloro derivatives.
Toluenesulfonylurea. C8H10N2O3S. (Mr 214.2). mp : 65 °C to 90 °C.
1177000. [1694-06-0]. 4-Methylbenzenesulfonylurea. A suitable certified reference solution (10 ng/µL in iso-octane)
p-Toluenesulfonylurea. (4-Methylphenyl)sulfonylurea. may be used.
Tragacanth. 1092300. [9000-65-1].
mp : 196 to 198 °C.
See Tragacanth (0532).
o-Toluidine. C7H9N. (Mr 107.2). 1091700. [95-53-4].
Triacetin. C9H14O6. (Mr 218.2). 1092400. [102-76-1].
2-Methylaniline.
Propane-1,2,3-triyl triacetate. Glycerol triacetate.
Pale-yellow liquid becoming reddish-brown on exposure to air
Almost clear, colourless to yellowish liquid, soluble in water,
and light, slightly soluble in water, soluble in ethanol (96 per
cent) and in dilute acids.
: about 1.16.
: about 1.01.
: about 1.43.
: about 1.569.
bp : about 260 °C.
bp : about 200 °C.
Storage : in an airtight container, protected from light. Triamcinolone. C21H27FO6. (Mr 394.4). 1111300. [124-94-7].
9-Fluoro-11β,16α,17,21-tetrahydroxypregna-1,4-diene-3,20-
o-Toluidine hydrochloride. C7H10ClN. (Mr 143.6). dione.
1117300. [636-21-5]. 2-Methylaniline hydrochloride. A crystalline powder.
2-Methylbenzenamine hydrochloride.
mp : 262 °C to 263 °C.
mp : 215 °C to 217 °C. Triamcinolone acetonide. 1133100. [76-25-5].
See Triamcinolone acetonide (0533).
p-Toluidine. C7H9N. (Mr 107.2). 1091800. [106-49-0].
4-Methylaniline. Tribromophenol. C6H3Br3O. (Mr 330.8). 1165300.
Lustrous plates or flakes, slightly soluble in water, freely [118-79-6]. 2,4,6-Tribromophenol.
soluble in acetone and in ethanol (96 per cent). Tributyl citrate. C18H32O7. (Mr 360.4). 1152800. [77-94-1].
mp : about 44 °C. Tributyl 2-hydroxypropane-1,2,3-tricarboxylate.
d 420 : about 1.043. Triethylamine. C6H15N. (Mr 101.2). 1093000. [121-44-8].

nD20 : about 1.445.
N,N-Diethylethanamine.
Colourless liquid, slightly soluble in water at a temperature
Tributyl phosphate. C12H27O4P. (Mr 266.3). 1179900. below 18.7 °C, miscible with ethanol (96 per cent).
[126-73-8]. Tributoxyphosphine oxide. Tributoxyphosphane : about 0.727.
oxide.
: about 1.401.
Colourless liquid, slightly soluble in water, soluble in the usual
organic solvents. bp : about 90 °C.
: about 0.976. Triethylamine R1. C6H15N. (Mr 101.2). 1093001.
: about 1.422. [121-44-8]. N,N-Diethylethanamine.
bp : about 289 °C, with decomposition. Complies with the requirements prescribed for
triethylamine R with the following additional requirements.
Tributylphosphine. C12H27P. (Mr 202.3). 1187100. [998-40-3].
Clear, colourless liquid. chromatography.
bp : about 240 °C.
Water : maximum 0.1 per cent.
Use freshly distilled or from a freshly opened container.
Trichloroacetic acid. C2HCl3O2. (Mr 163.4). 1092500.
[76-03-9]. Triethylamine R2. C6H15N. (Mr 101.2). 1093002.
[121-44-8]. N,N-Diethylethanamine.
Colourless crystals or a crystalline mass, very deliquescent,
very soluble in water and in ethanol (96 per cent). Complies with the requirements prescribed for
triethylamine R and with the following additional
requirements.
Trichloroacetic acid solution. 1092501. Content : minimum 99.5 per cent, determined by gas
Dissolve 40.0 g of trichloroacetic acid R in water R and chromatography.
dilute to 1000.0 mL with the same solvent. Verify the Water : maximum 0.2 per cent.
concentration by titration with 0.1 M sodium hydroxide It is suitable for gradient elution in liquid chromatography.
and adjust if necessary to 40 ± 1 g/L.
Use freshly distilled or from a freshly opened container.
1,1,1-Trichloroethane. C2H3Cl3. (Mr 133.4). 1092600.
[71-55-6]. Methylchloroform. Triethylenediamine. C6H12N2. (Mr 112.2). 1093100.
Non-flammable liquid, practically insoluble in water, soluble 1,4-Diazabicyclo[2.2.2]octane.
in acetone and in methanol. Crystals, very hygroscopic, sublimes readily at room
: about 1.34. temperature, freely soluble in water, in acetone and in
anhydrous ethanol.
: about 1.438.
bp : about 174 °C.
bp : about 74 °C.
mp : about 158 °C.
Trichloroethylene. C2HCl3. (Mr 131.4). 1102100. [79-01-6]. Storage : in an airtight container.
ethanol (96 per cent). Triethyl phosphonoformate. C7H15O5P. (Mr 210.2). 1132900.
: about 1.46. [1474-78-8]. Ethyl (diethoxyphosphoryl)formate.
: about 1.477. Colourless liquid.
bp12 mm : about 135 °C.
Trichlorotrifluoroethane. C2Cl3F3. (Mr 187.4). 1092700.
[76-13-1]. 1,1,2-Trichloro-1,2,2-trifluoroethane. Triflumuron. C15H10ClF3N2O3. (Mr 358.7). 1180800.
Colourless, volatile liquid, practically insoluble in water, [64628-44-0]. 1-(2-Chlorobenzoyl)-3-(4-triflumoromethoxy-
miscible with acetone. phenyl)urea.
: about 1.58. White or almost white crystalline powder, practically insoluble
Distillation range (2.2.11). Not less than 98 per cent distils in water, sparingly soluble in acetone and in methylene
between 47 °C and 48 °C. chloride.
Tricine. C6H13NO5. (Mr 179.2). 1138900. [5704-04-1]. Trifluoroacetic acid. C2HF3O2. (Mr 114.0). 1093200.
N-[2-Hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine. [76-05-1].
Use electrophoresis-grade reagent. Content : minimum 99 per cent.
mp : about 183 °C. Liquid, miscible with acetone and with ethanol (96 per cent).
: about 1.53.
Tricosane. C23H48. (Mr 324.6). 1092800. [638-67-5].
White or almost white crystals, practically insoluble in water, bp : about 72 °C.
soluble in hexane. Use a grade suitable for protein sequencing.
mp : about 48 °C. Storage : in an airtight container.
Tridecyl alcohol. C13H28O. (Mr 200.4). 1192500. [112-70-9]. Trifluoroacetic anhydride. C4F6O3. (Mr 210.0). 1093300.
Tridecanol. [407-25-0].
Tridocosahexaenoin. C69H98O6. (Mr 1023.5). 1144900. Colourless liquid.
[124596-98-1]. Triglyceride of docosahexaenoic acid : about 1.5.
(C22:6). Glycerol tridocosahexaenoate. Propane-1,2,3-triyl
3-Trifluoromethylaniline. C7H6F3N. (Mr 161.1). 1171900.
tri-(all-Z)-docosa-4,7,10,13,16,19-hexaenoate.
[98-16-8]. 3-(Trifluoromethyl)aniline. α,α,α-Trifluoro-m-
The reagent from Nu-Chek Prep, Inc. has been found suitable. toluidine. 3-(Trifluoromethyl)benzenamide.
Triethanolamine. 1092900. [102-71-6]. Colourless liquid.
See Trolamine (1577). Density : 1.30 g/cm3 (20 °C).

4-Trifluoromethylphenol. C7H5F3O. (Mr 162.1). 1161700. Trimethyltin chloride. C3H9ClSn. (Mr 199.3). 1170900.

[402-45-9]. [1066-45-1]. Chlorotrimethylstannane.
White or light yellow, crystalline solid or powder. 2,4,6-Trinitrobenzene sulfonic acid. C6H3N3O9S,3H2O.
mp : about 46 °C. (Mr 347.2). 1117500. [2508-19-2].
Trifluoropropylmethylpolysiloxane. 1171600. White or almost white, crystalline powder, soluble in water.
Polysiloxane substituted with trifluoropropyl groups and mp : 190 °C to 195 °C.
methyl groups. Triolein. C57H104O6. (Mr 885.4). 1168200. [122-32-7].
Triglycine. C6H11N3O4. (Mr 189.2). 1192600. [556-33-2]. Propane-1,2,3-triyl tris[(9Z)-octadec-9-enoate]. sn-Glyceryl
2-[[2-[(2-Aminoacetyl)amino]acetyl]amino]acetic acid. trioleate. Glycerol trioleate. Oleyl triglyceride.
Glycylglycylglycine. Content : minimum 99.0 per cent.
Trigonelline hydrochloride. C7H8ClNO2. (Mr 173.6). Triphenylmethanol. C19H16O. (Mr 260.3). 1093700.
1117400. [6138-41-6]. 3-Carboxy-1-methylpyridinium [76-84-6]. Triphenylcarbinol.
chloride. Nicotinic acid N-methylbetaine hydrochloride. Colourless crystals, practically insoluble in water, freely
Crystalline powder, very soluble in water, soluble in ethanol soluble in ethanol (96 per cent).
(96 per cent).
Triphenyltetrazolium chloride. C19H15ClN4. (Mr 334.8).
mp : about 258 °C. 1093800. [298-96-4]. 2,3,5-Triphenyl-2H-tetrazol-3-ium
1,2,4-Trimethylbenzene. C9H12. (Mr 120.2). 1188600. chloride.
[95-63-6]. Pseudocumene. Pale or dull-yellow powder, soluble in water, in acetone and in
Trimethylpentane. C8H18. (Mr 114.2). 1093400. [540-84-1].
Iso-octane. 2,2,4-Trimethylpentane. mp : about 240 °C, with decomposition.
Colourless, flammable liquid, practically insoluble in water, Storage : protected from light.
soluble in anhydrous ethanol. Triscyanoethoxypropane. C12H17N3O3. (Mr 251.3). 1093900.
: 0.691 to 0.696. 1,2,3-Tris(2-cyanoethoxy)propane.
: 1.391 to 1.393. Viscous, brown-yellow liquid, soluble in methanol. Used as a
Distillation range (2.2.11). Not less than 95 per cent distils stationary phase in gas chromatography.
between 98 °C and 100 °C. : about 1.11.
Trimethylpentane used in spectrophotometry complies with the Viscosity (2.2.9) : about 172 mPa·s.
1,3,5-Tris[3,5-di(1,1-dimethylethyl)-4-hydroxybenzyl]-
Absorbance (2.2.25) : maximum 0.01 from 250 nm to 420 nm,
1,3,5-triazine-2,4,6(1H,3H,5H)-trione. C48H69O6N3.
determined using water R as compensation liquid.
(Mr 784.1). 1094000. [27676-62-6].
Trimethylpentane R1. 1093401. White or almost white, crystalline powder.
Complies with the requirements prescribed for mp : 218 °C to 222 °C.
trimethylpentane R with the following modification.
Absorbance (2.2.25). Not more than 0.07 from 220 nm to Tris[2,4-di(1,1-dimethylethyl)phenyl] phosphite.
C42H63O3P. (Mr 647). 1094100. [31570-04-4].
360 nm, determined using water R as the compensation
liquid. White or almost white powder.
mp : 182 °C to 186 °C.
Trimethylpentane for chromatography. 1093402.
Complies with the requirements prescribed for Tris(hydroxymethyl)aminomethane. 1094200. [77-86-1].
trimethylpentane R with the following additional See Trometamol (1053).
requirement.
Tris(hydroxymethyl)aminomethane solution. 1094201.
Residue on evaporation : maximum 2 mg/L.
A solution containing the equivalent of 24.22 g of C4H11NO3
N,O-bis(Trimethylsilyl)acetamide. C8H21NOSi2. (Mr 203.4). in 1000.0 mL.
1093600. [10416-59-8].
Tris(hydroxymethyl)aminomethane solution R1.
Colourless liquid. 1094202.
: about 0.83. Dissolve 60.6 mg of tris(hydroxymethyl)aminomethane R
N-Trimethylsilylimidazole. C6H12N2Si. (Mr 140.3). 1100500. and 0.234 g of sodium chloride R in water R and dilute to
[18156-74-6]. 1-Trimethylsilylimidazole. 100 mL with the same solvent.
Colourless, hygroscopic liquid. Storage : at 2 °C to 8 °C ; use within 3 days.
: about 0.96. Tripotassium phosphate trihydrate. K3PO4,3H2O.
: about 1.48. (Mr 266.3). 1155300. [22763-03-7].
Storage : in an airtight container. White or almost white crystalline powder, freely soluble in
water.
N,O-bis(Trimethylsilyl)trifluoroacetamide. C8H18F3NOSi2.
(Mr 257.4). 1133200. [25561-30-2]. BSTFA. Trisodium phosphate dodecahydrate. Na3PO4,12H2O.
Colourless liquid. (Mr 380.1). 1094300. [10101-89-0].
: about 0.97. Colourless or white or almost white crystals, freely soluble in
: about 1.38. water.
bp12mm : about 40 °C Trometamol. 1094200. [77-86-1].
Trimethylsulfonium hydroxide. C3H10OS. (Mr 94.2). See Tris(hydroxymethyl)aminomethane R.
1145000. [17287-03-5]. Tropic acid. C9H10O3. (Mr 166.17). 1172000. [529-64-6].
: about 0.81. (2RS)-3-Hydroxy-2-phenylpropanoic acid.
Troxerutin. C33H42O19. (Mr 743). 1160300. [7085-55-4]. Valencene. C15H24. (Mr 204.4). 1152100. [4630-07-3].
Trihydroxyethylrutin. 3′,4′,7-Tris[O-(2-hydroxyethyl)]rutin. 4βH,5α-Eremophila-1(10),11-diene. (1R,7R,8aS)-
2-[3,4-Bis(2-hydroxyethoxy)phenyl]-3-[[6-O-(6-deoxy-α-L- 1,8a-Dimethyl-7-(1-methylethenyl)-1,2,3,5,6,7,8,8a-
mannopyranosyl)-β-D-glucopyranosyl]oxy]-5-hydroxy-7-(2- octahydronaphthalene.
hydroxyethoxy)-4H-1-benzopyran-4-one. Oily, colourless or pale yellow liquid, with a characteristic
mp : 168 °C to 176 °C. odour, practically insoluble in water, soluble in ethanol (96 per
cent).
Trypsin. 1094500. [9002-07-7].
: about 0.918.
A proteolytic enzyme obtained by activation of trypsinogen
extracted from the pancreas of beef (Bos taurus L.). : about 1.508.
White or almost white, crystalline or amorphous powder, bp : about 123 °C.
sparingly soluble in water. Valencene used in gas chromatography complies with the
Trypsin for peptide mapping. 1094600. [9002-07-7].
Trypsin of high purity treated to eliminate chymotryptic monograph Sweet orange oil (1811).
activity.
Tryptophan. C11H12N2O2. (Mr 204.2). 1094700. [73-22-3]. normalisation procedure.
White or yellowish-white, crystalline powder or colourless
Valerenic acid. C15H22O2. (Mr 234.3). 1165700. [3569-10-6].
crystals, slightly soluble in water, very slightly soluble in
(2E)-3-[(4S,7R,7aR)-3,7-Dimethyl-2,4,5,6,7,7a-hexahydro-1H-
inden-4-yl]-2-methylprop-2-enoic acid.
: about − 30, determined on a 10 g/L solution.
mp : 134 °C to 138 °C.
Typhaneoside. C34H42O20. (Mr 771). 1206000. [104472-68-6].
3-[6-Deoxy-α-L-mannopyranosyl-(1→2)-[6-deoxy-α- Valeric acid. C5H10O2. (Mr 102.1). 1095200. [109-52-4].
L-mannopyranosyl-(1→6)]-β-D-glucopyranosyloxy]-
Pentanoic acid.
5,7-dihydroxy-2-(4-hydroxy-3-methoxyphenyl)-4H-1- Colourless liquid, soluble in water, freely soluble in ethanol
benzopyran-4-one. (96 per cent).
: about 0.94.
Tyramine. C8H11NO. (Mr 137.2). 1117600. [51-67-2].
4-(2-Aminoethyl)phenol. : about 1.409.
Crystals, sparingly soluble in water, soluble in boiling bp : about 186 °C.
anhydrous ethanol. Valine. 1185300. [72-18-4].
mp : 164 °C to 165 °C. See Valine (0796).
Tyrosine. C9H11NO3. (Mr 181.2). 1094800. [60-18-4]. Vanillin. 1095300. [121-33-5].
2-Amino-3-(4-hydroxyphenyl)propionic acid.
See Vanillin (0747).
White or almost white, crystalline powder, or colourless
or white or almost white crystals, slightly soluble in water, Vanillin reagent. 1095301.
practically insoluble in acetone and in anhydrous ethanol, Carefully add, dropwise, 2 mL of sulfuric acid R to 100 mL
soluble in dilute hydrochloric acid and in solutions of alkali of a 10 g/L solution of vanillin R in ethanol (96 per cent) R.
hydroxides.
Umbelliferone. C9H6O3. (Mr 162.1). 1137500. [93-35-6].
7-Hydroxycoumarin. 7-Hydroxy-2H-1-benzopyran-2-one. Vanillin solution, phosphoric. 1095302.
Needles from water. Dissolve 1.0 g of vanillin R in 25 mL of ethanol (96 per
mp : 225 °C to 228 °C. cent) R. Add 25 mL of water R and 35 mL of phosphoric
acid R.
Undecanoic acid. C11H22O2. (Mr 186.29). 1195200.
[112-37-8]. Hendecanoic acid. Undecylic acid. Veratrole. C8H10O2. (Mr 138.2). 1165400. [91-16-7].
1,2-Dimethoxybenzene.
mp : about 30 °C.
: 1.085.
Content : minimum 97.0 per cent of C11H22O2.
: 1.534.
Uracil. C4H4N2O2. (Mr 112.1). 1161800. [66-22-8]. bp : about 206 °C.
Content : minimum 95.0 per cent. mp : about 22 °C.
Urea. 1095000. [57-13-6]. Verbenone. C10H14O. (Mr 150.2). 1140500. [1196-01-6].
See Urea (0743). (1S,5S)-4,6,6-Trimethylbicyclo[3.1.1]hept-3-en-2-one.
Uridine. C9H12N2O6. (Mr 244.2). 1095100. [58-96-8]. Oil with a characteristic odour, practically insoluble in water,
1-β-D-Ribofuranosyluracil. miscible with organic solvents.
White or almost white, crystalline powder, soluble in water. : about 0.978.
18
mp : about 165 °C. nD : about 1.49.
Ursolic acid. C30H48O3. (Mr 456.7). 1141600. [77-52-1]. : about + 249.6.
3β-Hydroxyurs-12-en-28-oic acid. bp : 227 °C to 228 °C.
White or almost white powder, practically insoluble in water, mp : about 6.5 °C.
sparingly soluble in methanol, slightly soluble in ethanol Verbenone used in gas chromatography complies with the
(96 per cent). following additional test.
[α]D21 : about 67.50, determined on a 10 g/L solution in a Assay. Gas chromatography (2.2.28) as prescribed in the
56.1 g/L solution of potassium hydroxide R in ethanol (96 per monograph Rosemary oil (1846).
cent) R. Content : minimum 99 per cent, calculated by the
mp : 285 °C to 288 °C. normalisation procedure.

Vinyl acetate. C4H6O2. (Mr 86,10). 1111800. [108-05-4]. Water. 1095500. [7732-18-5].

Ethenyl acetate. See Purified water (0008).
: about 0.930.
Water R1. 1095509.
bp : about 72 °C.
Prepared from distilled water R by multiple distillation.
Vinyl chloride. C2H3Cl. (Mr 62.5). 1095400. [75-01-4]. Remove carbon dioxide by boiling for at least 15 min before
Colourless gas, slightly soluble in organic solvents. use in a boiling flask of fused silica or borosilicate glass and
cool. Any other suitable method may be used. The boiling
Vinyl(1)phenyl(5)methyl(94)polysiloxane. 1100000. flask has been already used for the test or has been filled
Polysiloxane substituted with 1 per cent of vinyl groups, 5 per with water R and kept in an autoclave at 121 °C for at least
cent of phenyl groups and 94 per cent of methyl groups. 1 h prior to first use. When tested immediately before use,
water R1 is neutral to methyl red solution R, i.e. it shall
Vinyl polymer for chromatography, amino alkyl. 1191500. produce an orange-red (not a violet-red or yellow) colour
Spherical particles (5 μm) of a vinyl alcohol copolymer corresponding to pH 5.5 ± 0.1 when 0.05 mL of methyl red
chemically modified by bonding of amino alkyl groups. solution R is added to 50 mL of the water to be examined.
Conductivity : maximum 1 µS·cm− 1, determined at 25 °C by
Vinyl polymer for chromatography, octadecyl. 1155400. an in-line conductivity meter (see Purified water (0008)).
Spherical particles (5 μm) of a vinyl alcohol copolymer
chemically modified by bonding of octadecyl groups on the Water, ammonium-free. 1095501.
hydroxyl groups. To 100 mL of water R add 0.1 mL of sulfuric acid R. Distil
using the apparatus described for the determination of
Vinyl polymer for chromatography, octadecylsilyl. Distillation range (2.2.11). Reject the first 10 mL and collect
1121600. the following 50 mL.
Spherical particles (5 μm) of a vinyl alcohol copolymer bonded
to an octadecylsilane. Carbon content of 17 per cent. Water, carbon dioxide-free. 1095502.
Water R which has been boiled for a few minutes and
2-Vinylpyridine. C7H7N. (Mr 105.1). 1102200. [100-69-6]. protected from the atmosphere during cooling and storage
Yellow liquid, miscible in water. or deionised water R with a resistivity of not less than
: about 0.97. 0.18 MΩ·m, determined at 25 °C.
: about 1.549. Water, distilled. 1095504.
4-Vinylpyridine. C7H7N. (Mr 105.1). 1187200. [100-43-6]. Water R prepared by distillation.
4-Ethenylpyridine. Water, distilled, deionised. 1095508.
Clear, deep yellowish-brown liquid. Deionised water R prepared by distillation with a resistivity
bp : 58-61 °C. of not less than 0.18 MΩ·m, determined at 25 °C.
1-Vinylpyrrolidin-2-one. C6H9NO. (Mr 111.1). 1111900. Water for chromatography. 1095503.
[88-12-0]. 1-Ethenylpyrrolidin-2-one. Deionised water with a resistivity of not less than
Content : minimum 99.0 per cent. 0.18 MΩ·m, determined at 25 °C, prepared by distillation,
Clear colourless liquid. ion exchange, reverse osmosis or any other suitable
method, using water that complies with the regulations on
Water (2.5.12) : maximum 0.1 per cent, determined on water intended for human consumption, as laid down by
2.5 g. Use as the solvent, a mixture of 50 mL of anhydrous the competent authority.
methanol R and 10 mL of butyrolactone R.
Its quality is such that no significant interfering peaks or loss
Assay. Gas chromatography (2.2.28) : use the normalisation of sensitivity are observed when used in chromatography.
procedure. Isocratic elution with UV detection at low wavelengths
Column : (i.e. less than 230 nm), with evaporative detectors (e.g.
– material : fused-silica ; light scattering detector, particle counter detector, charged
– size : l = 30 m, Ø = 0.5 mm ; aerosol detector) or mass detectors, or gradient elution,
may require the use of water with a total organic carbon
– stationary phase : macrogol 20 000 R. content of maximum 5 ppb.
Temperature : Water for injections. 1095505.
See Water for injections (0169).
Time Temperature
(min) (°C)
Water, nitrate-free. 1095506.
Column 0-1 80
To 100 mL of water R add a few milligrams of potassium
1 - 12 80 → 190 permanganate R and of barium hydroxide R. Distil
12 - 27 190 using the apparatus described for the determination of
Distillation range (2.2.11). Reject the first 10 mL and collect
Injection port 190 the following 50 mL.
Detection : flame-ionisation. Water, particle-free. 1095507.
Injection : 0.3 µL of the substance to be examined. Filter water R through a membrane with a pore size of
Adjust the flow rate of the carrier gas so that the retention 0.22 μm.
time of the peak corresponding to 1-vinylpyrrolidin-2-one Weak cationic resin. 1096000.
is about 17 min.
Polymethacrylic resin, slightly acid, with carboxyl groups
Vitexin. C21H20O10. (Mr 432.4). 1133300. [3681-93-4]. present in a protonated form.
Apigenin 8-glucoside. Particle size : 75 µm to 160 µm.
Yellow powder. pH limits of use : 5 to 14.
Storage : in an airtight container, protected from light. Maximum temperature of use : 120 °C.
Wedelolactone. C16H10O7. (Mr 314.3). 1187300. Xylenol orange solution. 1096302.

[524-12-9]. 1,8,9-Trihydroxy-3-methoxy-6H-benzofuro[3,2- Dissolve 50.8 mg of xylenol orange R in water R and dilute
c][1]benzopyran-6-one. to 100.0 mL with the same solvent.
White beeswax. 1196500. Xylenol orange triturate. 1096301.
See White beeswax (0069). Triturate 1 part of xylenol orange R with 99 parts of
Xanthydrol. C13H10O2. (Mr 198.2). 1096100. [90-46-0]. potassium nitrate R.
9-Xanthenol. Test for sensitivity. To 50 mL of water R add 1 mL
Content : minimum 90.0 per cent. of dilute acetic acid R, 50 mg of the xylenol orange
triturate and 0.05 mL of lead nitrate solution R. Add
White or pale-yellow powder, very slightly soluble in water, hexamethylenetetramine R until the colour changes from
soluble in ethanol (96 per cent) and in glacial acetic acid. yellow to violet-red. After addition of 0.1 mL of 0.1 M
It is also available as a methanolic solution containing 90 g/L sodium edetate the colour changes to yellow.
to 110 g/L of xanthydrol.
mp : about 123 °C. Xylitol. C5H12O5. (Mr 152.1). 1190700. [87-99-0].
Assay. In a 250 mL flask dissolve 0.300 g in 3 mL of methanol R White or almost white, crystalline powder or crystals.
or use 3.0 mL of solution. Add 50 mL of glacial acetic acid R Content : minimum 96.0 per cent.
and, dropwise with shaking, 25 mL of a 20 g/L solution of
urea R. Allow to stand for 12 h, collect the precipitate on a Xylose. 1096400. [58-86-6].
sintered-glass filter (16) (2.1.2), wash with 20 mL of ethanol See Xylose (1278).
(96 per cent) R, dry in an oven at 100 °C to 105 °C and weigh.
Zinc. Zn. (Ar 65.4). 1096500. [7440-66-6].
1 g of precipitate is equivalent to 0.9429 g of xanthydrol.
Storage : protected from light. If a methanolic solution is
used, store in small sealed ampoules and filter before use if Silver-white cylinders, granules, pellets or filings with a blue
necessary. sheen.
Arsenic (2.4.2, Method A): maximum 0.2 ppm.
Xanthydrol R1. 1096101.
Dissolve 5.0 g in a mixture of the 15 mL of hydrochloric acid R
Complies with the requirements prescribed for and 25 mL of water R prescribed.
xanthydrol R with the following requirement.
Content : minimum 98.0 per cent of C13H10O2. Zinc, activated. 1096501.
Place the zinc cylinders or pellets to be activated in a conical
Xanthydrol solution. 1096102. flask and add a sufficient quantity of a 50 ppm solution of
To 0.1 mL of a 100 g/L solution of xanthydrol R in chloroplatinic acid R to cover the metal. Allow the metal to
methanol R add 100 mL of anhydrous acetic acid R and remain in contact with the solution for 10 min, wash, drain
1 mL of hydrochloric acid R. Allow to stand for 24 h before and dry immediately.
using.
Arsenic (2.4.2, Method A). To 5 g of the activated zinc add
Xylene. C8H10. (Mr 106.2). 1096200. [1330-20-7]. 15 mL of hydrochloric acid R, 25 mL of water R, 0.1 mL of
stannous chloride solution R and 5 mL of potassium iodide
Mixture of isomers. Clear, colourless, flammable liquid,
solution R. No colour is produced during the test.
practically insoluble in water, miscible with ethanol (96 per
cent). Activity. The requirements of the suitability test for arsenic
(2.4.2, Method A) are met.
: about 0.867.
: about 1.497. Zinc acetate. (C2H3O2)2Zn,2H2O. (Mr 219.5). 1102300.
bp : about 138 °C. [5970-45-6]. Zinc acetate dihydrate.
Bright white or almost white crystals, slightly efflorescent,
m-Xylene. C8H10. (Mr 106.2). 1117700. [108-38-3]. freely soluble in water, soluble in ethanol (96 per cent). It loses
1,3-Dimethylbenzene. its crystallisation water at 100 °C.
Clear, colourless, flammable liquid, practically insoluble in : about 1.735.
mp : about 237 °C.
: about 0.884.
: about 1.497. Zinc acetate solution. 1102301.
bp : about 139 °C. Mix 600 mL of water R with 150 mL of glacial acetic acid R,
mp : about − 47 °C. 54.9 g of zinc acetate R and stir to dissolve. Continue
stirring while adding 150 mL of concentrated ammonia R.
o-Xylene. C8H10. (Mr 106.2). 1100600. [95-47-6]. Cool to room temperature and adjust with ammonia R to
1,2-Dimethylbenzene. pH 6.4. Dilute the mixture to 1 L with water R.
Clear, colourless, flammable liquid, practically insoluble in Zinc chloride. 1096600. [7646-85-7].
See Zinc chloride (0110).
: about 0.881.
: about 1.505. Zinc chloride-formic acid solution. 1096601.
bp : about 144 °C. Dissolve 20 g of zinc chloride R in 80 g of an 850 g/L
mp : about − 25 °C. solution of anhydrous formic acid R.
Xylenol orange. C31H28N2Na4O13S. (Mr 761). Zinc chloride solution, iodinated. 1096602.

1096300. [3618-43-7]. Tetrasodium 3,3′-(3H-2,1- Dissolve 20 g of zinc chloride R and 6.5 g of potassium
benzoxathiol-3-ylidene)bis[(6-hydroxy-5-methyl-3,1- iodide R in 10.5 mL of water R. Add 0.5 g of iodine R and
phenylene)methyleneiminobisacetate] S,S-dioxide. shake for 15 min. Filter if necessary.
Reddish-brown crystalline powder, soluble in water. Storage : protected from light.

Zinc iodide and starch solution. 1096502. Zinc powder. Zn. (Ar 65.4). 1096800. [7440-66-6].
To a solution of 2 g of zinc chloride R in 10 mL of water R Content : minimum 90.0 per cent.
add 0.4 g of soluble starch R and heat until the starch has Very fine, grey powder, soluble in dilute hydrochloric acid R.
dissolved. After cooling to room temperature add 1.0 mL of
Zinc sulfate. 1097000. [7446-20-0].
a colourless solution containing 0.10 g zinc R as filings and
0.2 g of iodine R in water R. Dilute the solution to 100 mL See Zinc sulfate (0111).
with water R and filter. Zirconyl nitrate. A basic salt corresponding approximately to
Storage : protected from light. the formula ZrO(NO3)2,2H2O. 1097200. [14985-18-3].
A white or almost white powder or crystals, hygroscopic,
Test for sensitivity. Dilute 0.05 mL of sodium nitrite solution R
soluble in water. The aqueous solution is a clear or at most
to 50 mL with water R. To 5 mL of this solution add 0.1 mL of
slightly opalescent liquid.
dilute sulfuric acid R and 0.05 mL of the zinc iodide and starch
solution and mix. The solution becomes blue. Storage : in an airtight container.
Zirconyl nitrate solution. 1097201.
Zinc oxide. 1096700. [1314-13-2]. A 1 g/L solution in a mixture of 40 mL of water R and
See Zinc oxide (0252). 60 mL of hydrochloric acid R.
EUROPEAN PHARMACOPOEIA 4.1.2. Standard solutions for limit tests
01/2020:40102 Antimony standard solution (1 ppm Sb). 5000400.

Dissolve antimony potassium tartrate R equivalent to 0.274 g
of C8H4K2O12Sb2,3H2O in 20 mL of hydrochloric acid R1 and
dilute the clear solution to 100.0 mL with water R. To 10.0 mL
of this solution add 200 mL of hydrochloric acid R1 and dilute
4.1.2. STANDARD SOLUTIONS FOR to 1000.0 mL with water R. To 100.0 mL of this solution add
300 mL of hydrochloric acid R1 and dilute to 1000.0 mL with
LIMIT TESTS water R. Prepare the dilute solutions immediately before use.
Acetaldehyde standard solution (100 ppm C2H4O). Arsenic standard solution (10 ppm As). 5000500.
5000100. Immediately before use, dilute with water R to 100 times its
Dissolve 1.0 g of acetaldehyde R in 2-propanol R and dilute to volume a solution prepared by dissolving arsenious trioxide R
100.0 mL with the same solvent. Dilute 5.0 mL of the solution equivalent to 0.330 g of As2O3 in 5 mL of dilute sodium
to 500.0 mL with 2-propanol R. Prepare immediately before hydroxide solution R and diluting to 250.0 mL with water R.
use.
Arsenic standard solution (1 ppm As). 5000501.
Acetaldehyde standard solution (100 ppm C2H4O) R1. Immediately before use, dilute arsenic standard solution
5000101. (10 ppm As) R to 10 times its volume with water R.
Dissolve 1.0 g of acetaldehyde R in water R and dilute to
100.0 mL with the same solvent. Dilute 5.0 mL of the solution Barium standard solution (0.1 per cent Ba). 5000601.
to 500.0 mL with water R. Prepare immediately before use. Dissolve barium chloride R equivalent to 0.178 g of BaCl2,2H2O
in distilled water R and dilute to 100.0 mL with the same
Aluminium standard solution (200 ppm Al). 5000200. solvent.
Dissolve in water R a quantity of aluminium potassium Barium standard solution (50 ppm Ba). 5000600.
sulfate R equivalent to 0.352 g of AlK(SO4)2,12H2O. Add 10 mL
of dilute sulfuric acid R and dilute to 100.0 mL with water R. Immediately before use, dilute with distilled water R to
20 times its volume a solution in distilled water R containing
Aluminium standard solution (100 ppm Al). 5000203. barium chloride R equivalent to 0.178 g of BaCl2,2H2O in
Immediately before use, dilute with water R to 10 times its 100.0 mL.
volume a solution containing 8.947 g of aluminium chloride R Barium standard solution (2 ppm Ba). 5005600.
in 1000.0 mL of water R. Immediately before use, dilute barium standard solution
Aluminium standard solution (10 ppm Al). 5000201. (50 ppm Ba) R to 25 times its volume with distilled water R.
Immediately before use, dilute with water R to 100 times Bismuth standard solution (100 ppm Bi). 5005300.
its volume in a solution containing aluminium nitrate R Dissolve bismuth subnitrate R equivalent to 0.500 g of Bi in
equivalent to 1.39 g of Al(NO3)3,9H2O in 100.0 mL. 50 mL of nitric acid R and dilute to 500.0 mL with water R.
Dilute the solution to 10 times its volume with dilute nitric
Aluminium standard solution (5 ppm Al). 5006600. acid R immediately before use.
Immediately before use, dilute with water R to 100 times
its volume in a solution containing aluminium nitrate R Cadmium standard solution (0.1 per cent Cd). 5000700.
equivalent to 0.695 g of Al(NO3)3,9H2O in 100.0 mL. Dissolve cadmium R equivalent to 0.100 g of Cd in the
Alternatively, use a commercially available standard solution smallest necessary amount of a mixture of equal volumes of
containing a known amount of aluminium (5 ppm Al). hydrochloric acid R and water R and dilute to 100.0 mL with a
1 per cent V/V solution of hydrochloric acid R.
Aluminium standard solution (2 ppm Al). 5000202.
Immediately before use, dilute with water R to 100 times its Cadmium standard solution (10 ppm Cd) . 5000701.
volume a solution containing aluminium potassium sulfate R Immediately before use, dilute cadmium standard solution
equivalent to 0.352 g of AlK(SO4)2,12H2O and 10 mL of dilute (0.1 per cent Cd) R to 100 times its volume with a 1 per
sulfuric acid R in 100.0 mL. cent V/V solution of hydrochloric acid R.
Ammonium standard solution (100 ppm NH4). 5000300. Calcium standard solution (400 ppm Ca). 5000800.
Immediately before use, dilute to 25 mL with water R 10 mL Immediately before use, dilute with distilled water R to
of a solution containing ammonium chloride R equivalent to 10 times its volume a solution in distilled water R containing
0.741 g of NH4Cl in 1000 mL. calcium carbonate R equivalent to 1.000 g of CaCO3 and
23 mL of 1 M hydrochloric acid in 100.0 mL.
Ammonium standard solution (3 ppm NH4). 5006100.
Calcium standard solution (100 ppm Ca). 5000801.
Immediately before use, dilute with water R to 100 times Immediately before use, dilute with distilled water R to
its volume a solution containing ammonium chloride R 10 times its volume a solution in distilled water R containing
equivalent to 0.889 g of NH4Cl in 1000.0 mL. calcium carbonate R equivalent to 0.624 g of CaCO3 and 3 mL
Ammonium standard solution (2.5 ppm NH4). 5000301. of acetic acid R in 250.0 mL.
Immediately before use, dilute with water R to 100 times Calcium standard solution (100 ppm Ca) R1. 5000804.
its volume a solution containing ammonium chloride R Immediately before use, dilute with water R to 10 times its
equivalent to 0.741 g of NH4Cl in 1000.0 mL. volume a solution containing anhydrous calcium chloride R
equivalent to 2.769 g of CaCl2 in 1000.0 mL of dilute
Ammonium standard solution (1 ppm NH4). 5000302. hydrochloric acid R.
Immediately before use, dilute ammonium standard solution
(2.5 ppm NH4) R to 2.5 times its volume with water R. Calcium standard solution (100 ppm Ca), alcoholic.
5000802.
Antimony standard solution (100 ppm Sb). 5000401. Immediately before use, dilute with ethanol (96 per cent) R to
Dissolve antimony potassium tartrate R equivalent to 0.274 g 10 times its volume a solution in distilled water R containing
of C8H4K2O12Sb2,3H2O in 500 mL of 1 M hydrochloric acid and calcium carbonate R equivalent to 2.50 g of CaCO3 and 12 mL
dilute the clear solution to 1000 mL with water R. of acetic acid R in 1000.0 mL.

4.1.2. Standard solutions for limit tests EUROPEAN PHARMACOPOEIA
Calcium standard solution (10 ppm Ca). 5000803. Ferricyanide standard solution (50 ppm Fe(CN)6). 5001300.
Immediately before use, dilute with distilled water R to Immediately before use, dilute with water R to 100 times
100 times its volume a solution in distilled water R containing its volume a solution containing potassium ferricyanide R
calcium carbonate R equivalent to 0.624 g of CaCO3 and 3 mL equivalent to 0.78 g of K3Fe(CN)6 in 100.0 mL.
of acetic acid R in 250.0 mL.
Fluoride standard solution (10 ppm F). 5001400.
Chloride standard solution (50 ppm Cl). 5004100. Dissolve in water R sodium fluoride R previously dried at
Immediately before use, dilute with water R to 10 times its 300 °C for 12 h, equivalent to 0.442 g of NaF, and dilute to
volume a solution containing sodium chloride R equivalent to 1000.0 mL with the same solvent (1 mL = 0.2 mg F). Store in
0.824 g of NaCl in 1000.0 mL. a polyethylene container. Immediately before use, dilute the
solution to 20 times its volume with water R.
Chloride standard solution (8 ppm Cl). 5000900.
Immediately before use, dilute with water R to 100 times its Fluoride standard solution (1 ppm F). 5001401.
volume a solution containing sodium chloride R equivalent to Immediately before use, dilute fluoride standard solution
1.32 g of NaCl in 1000.0 mL. (10 ppm F) R to 10 times its volume with water R.
Chloride standard solution (5 ppm Cl). 5000901. Formaldehyde standard solution (5 ppm CH2O). 5001500.
Immediately before use, dilute with water R to 100 times its Immediately before use, dilute with water R to 200 times its
volume a solution containing sodium chloride R equivalent to volume a solution containing 1.0 g of CH2O per litre prepared
0.824 g of NaCl in 1000.0 mL. from formaldehyde solution R.
Chromium liposoluble standard solution (1000 ppm Cr). Germanium standard solution (100 ppm Ge). 5004400.
5004600.
Dissolve ammonium hexafluorogermanate(IV) R equivalent
A chromium (metal) organic compound in an oil. to 0.307 g of (NH4)2GeF6 in a 0.01 per cent V/V solution of
Chromium standard solution (0.1 per cent Cr). 5001002. hydrofluoric acid R. Dilute the clear solution to 1000 mL with
water R.
Dissolve potassium dichromate R equivalent to 2.83 g of
K2Cr2O7 in water R and dilute to 1000.0 mL with the same Glyoxal standard solution (20 ppm C2H2O2). 5003700.
solvent.
In a 100 mL graduated flask weigh a quantity of glyoxal
Chromium standard solution (100 ppm Cr). 5001000. solution R corresponding to 0.200 g of C2H2O2 and make up
to volume with anhydrous ethanol R. Immediately before
Dissolve potassium dichromate R equivalent to 0.283 g of use dilute the solution to 100 times its volume with the same
K2Cr2O7 in water R and dilute to 1000.0 mL with the same solvent.
solvent.
Glyoxal standard solution (2 ppm C2H2O2). 5003701.
Chromium standard solution (0.1 ppm Cr). 5001001.
Immediately before use, dilute glyoxal standard solution
Immediately before use, dilute chromium standard solution
(20 ppm C2H2O2) R to 10 times its volume with anhydrous
(100 ppm Cr) R to 1000 times its volume with water R.
ethanol R.
Cobalt standard solution (100 ppm Co). 5004300.
Hydrogen peroxide standard solution (2 ppm H2O2).
Dissolve cobalt nitrate R equivalent to 0.494 g of 5005200.
Co(NO3)2,6H2O in 500 mL of 1 M nitric acid and dilute the
clear solution to 1000 mL with water R. Dilute 10.0 mL of dilute hydrogen peroxide solution R to
300.0 mL with water R. Dilute 2.0 mL of this solution to
Copper liposoluble standard solution (1000 ppm Cu). 1000.0 mL with water R. Prepare immediately before use.
5004700.
Iodide standard solution (10 ppm I). 5003800.
A copper (metal) organic compound in an oil.
Immediately before use, dilute with water R to 100 times its
Copper standard solution (0.1 per cent Cu). 5001100. volume a solution containing potassium iodide R equivalent
Dissolve copper sulfate pentahydrate R equivalent to 0.393 g to 0.131 g of KI in 100.0 mL.
of CuSO4,5H2O in water R and dilute to 100.0 mL with the Iron standard solution (0.1 per cent Fe). 5001605.
same solvent.
Dissolve 0.100 g of Fe in the smallest amount necessary of a
Copper standard solution (0.1 per cent Cu) for ICP. mixture of equal volumes of hydrochloric acid R and water R
5006300. and dilute to 100.0 mL with water R.
A copper standard solution (1000 mg/L) suitable for
inductively coupled plasma (ICP) applications and traceable Iron standard solution (250 ppm Fe). 5001606.
to national or international standards. Immediately before use, dilute with water R to 40 times its
volume a solution containing 4.840 g of ferric chloride R in a
Copper standard solution (10 ppm Cu). 5001101. 150 g/L solution of hydrochloric acid R diluted to 100.0 mL.
Immediately before use, dilute copper standard solution
(0.1 per cent Cu) R to 100 times its volume with water R. Iron standard solution (20 ppm Fe). 5001600.
Copper standard solution (0.1 ppm Cu). 5001102. volume a solution containing ferric ammonium sulfate R
Immediately before use, dilute copper standard solution equivalent to 0.863 g of FeNH4(SO4)2,12H2O and 25 mL of
(10 ppm Cu) R to 100 times its volume with water R. dilute sulfuric acid R in 500.0 mL.
Ferrocyanide standard solution (100 ppm Fe(CN)6). Iron standard solution (10 ppm Fe). 5001601.
5001200. Immediately before use, dilute with water R to 100 times its
Immediately before use, dilute with water R to 10 times volume a solution containing ferrous ammonium sulfate R
its volume a solution containing potassium ferrocyanide R equivalent to 7.022 g of Fe(NH4)2(SO4)2,6H2O and 25 mL of
equivalent to 0.20 g of K4Fe(CN)6,3H2O in 100.0 mL. dilute sulfuric acid R in 1000.0 mL.

Iron standard solution (8 ppm Fe). 5001602. Magnesium standard solution (10 ppm Mg). 5001801.
Immediately before use, dilute with water R to 10 times its Immediately before use, dilute magnesium standard solution
volume a solution containing 80 mg of iron R and 50 mL of (100 ppm Mg) R to 10 times its volume with water R.
hydrochloric acid R (220 g/L HCl) in 1000.0 mL.
Magnesium standard solution (10 ppm Mg) R1. 5001802.
Iron standard solution (2 ppm Fe). 5001603. Immediately before use, dilute with water R to 100 times its
Immediately before use, dilute iron standard solution (20 ppm volume a solution containing 8.365 g of magnesium chloride R
Fe) R to 10 times its volume with water R. in 1000.0 mL of dilute hydrochloric acid R.
Iron standard solution (1 ppm Fe). 5001604. Manganese standard solution (1000 ppm Mn). 5005800.
Immediately before use, dilute iron standard solution (20 ppm Dissolve manganese sulfate R equivalent to 3.08 g of
Fe) R to 20 times its volume with water R. MnSO4,H2O in 500 mL of 1 M nitric acid and dilute the
Lead liposoluble standard solution (1000 ppm Pb). solution to 1000 mL with water R.
5004800. Manganese standard solution (100 ppm Mn). 5004500.
A lead (metal) organic compound in an oil.
Dissolve manganese sulfate R equivalent to 0.308 g of
Lead standard solution (0.1 per cent Pb). 5001700. MnSO4,H2O in 500 mL of 1 M nitric acid and dilute the clear
Dissolve lead nitrate R equivalent to 0.400 g of Pb(NO3)2 in solution to 1000 mL with water R.
water R and dilute to 250.0 mL with the same solvent. Mercury standard solution (1000 ppm Hg). 5001900.

Dissolve mercuric chloride R equivalent to 1.354 g of HgCl2

Lead standard solution (100 ppm Pb). 5001701. in 50 mL of dilute nitric acid R and dilute to 1000.0 mL with
Immediately before use, dilute lead standard solution (0.1 per water R.
cent Pb) R to 10 times its volume with water R.
Mercury standard solution (10 ppm Hg). 5001901.
Lead standard solution (10 ppm Pb). 5001702.
Immediately before use, dilute with water to 100 times its
Immediately before use, dilute lead standard solution (100 ppm volume a solution containing mercuric chloride R equivalent
Pb) R to 10 times its volume with water R. to 0.338 g of HgCl2 in 250.0 mL.
Lead standard solution (10 ppm Pb) R1. 5001706. Nickel liposoluble standard solution (1000 ppm Ni).
Immediately before use, dilute with water R to 10 times its 5004900.
volume a solution containing 0.160 g of lead nitrate R in
A nickel (metal) organic compound in an oil.
100 mL of water R, to which is added 1 mL of lead-free nitric
acid R and dilute to 1000.0 mL. Nickel standard solution (10 ppm Ni). 5002000.


Lead standard solution (2 ppm Pb). 5001703. volume a solution containing nickel sulfate R equivalent to
Immediately before use, dilute lead standard solution (10 ppm 4.78 g of NiSO4,7H2O in 1000.0 mL.
Pb) R to 5 times its volume with water R.
Nickel standard solution (5 ppm Ni). 5005900.
Lead standard solution (1 ppm Pb). 5001704. Immediately before use dilute nickel standard solution (10 ppm
Immediately before use, dilute lead standard solution (10 ppm Ni) R to twice its volume with water for chromatography R.

Nickel standard solution (0.2 ppm Ni). 5002002.

Lead standard solution (0.25 ppm Pb). 5006000. Immediately before use, dilute nickel standard solution
Immediately before use, dilute lead standard solution (1 ppm (10 ppm Ni) R to 50 times its volume with water R.
Pb) R to 4 times its volume with water R. Nickel standard solution (0.1 ppm Ni). 5002001.
Lead standard solution (0.1 ppm Pb). 5001705. Immediately before use, dilute nickel standard solution
Immediately before use, dilute lead standard solution (1 ppm (10 ppm Ni) R to 100 times its volume with water R.
Nitrate standard solution (100 ppm NO3). 5002100.
Lutetium standard solution (20 ppm Lu). 5006500. Immediately before use, dilute with water R to 10 times its
Immediately before use, dissolve 0.445 g of lutetium chloride volume a solution containing potassium nitrate R equivalent
hexahydrate R in a mixture of equal volumes of heavy to 0.815 g of KNO3 in 500.0 mL.
metal-free nitric acid R and water R, and dilute to 100.0 mL
with the same mixture of solvents. Nitrate standard solution (10 ppm NO3). 5002101.
Dilute 1.0 mL of this solution to 100.0 mL with water R. Immediately before use, dilute nitrate standard solution
(100 ppm NO3) R to 10 times its volume with water R.
Magnesium standard solution (0.1 per cent Mg). 5001803.
Dissolve magnesium sulfate R equivalent to 1.010 g of Nitrate standard solution (2 ppm NO3). 5002102.
MgSO4,7H2O in distilled water R and dilute to 100.0 mL with Immediately before use, dilute nitrate standard solution
the same solvent. (10 ppm NO3) R to 5 times its volume with water R.
Magnesium standard solution (1000 ppm Mg). 5006200. Palladium standard solution (500 ppm Pd). 5003600.
Dissolve 5.275 g of magnesium nitrate R in 16 mL of dilute Dissolve 50.0 mg of palladium R in 9 mL of hydrochloric
nitric acid R and dilute to 500.0 mL with water R. acid R and dilute to 100.0 mL with water R.
Standardisation : carry out the determination of magnesium
by complexometry (2.5.11). Palladium standard solution (20 ppm Pd). 5003602.
Dissolve 0.333 g of palladium chloride R in 2 mL of warm
Magnesium standard solution (100 ppm Mg). 5001800. hydrochloric acid R. Dilute the solution to 1000.0 mL with a
Immediately before use, dilute with water R to 10 times its mixture of equal volumes of dilute hydrochloric acid R and
volume a solution containing magnesium sulfate R equivalent water R. Immediately before use dilute to 10 times its volume
to 1.010 g of MgSO4,7H2O in 100.0 mL. with water R.

4.1.2. Standard solutions for limit tests EUROPEAN PHARMACOPOEIA
Palladium standard solution (0.5 ppm Pd). 5003601. Strontium standard solution (1.0 per cent Sr). 5003900.
Dilute 1 mL of palladium standard solution (500 ppm Pd) R Cover with water R, strontium carbonate R equivalent to
to 1000 mL with a mixture of 0.3 volumes of nitric acid R and 1.6849 g of SrCO3. Cautiously add hydrochloric acid R until
99.7 volumes of water R. all the solid has dissolved and there is no sign of further
effervescence. Dilute to 100.0 mL with water R.
Phosphate standard solution (200 ppm PO4). 5004200.
Dissolve potassium dihydrogen phosphate R equivalent to Sulfate standard solution (100 ppm SO4). 5002802.
0.286 g of KH2PO4 in water R and dilute to 1000.0 mL with Immediately before use, dilute with distilled water R to
the same solvent. 10 times its volume a solution in distilled water R containing
dipotassium sulfate R equivalent to 0.181 g of K2SO4 in
Phosphate standard solution (5 ppm PO4). 5002200.
100.0 mL.
Immediately before use, dilute with water R to 100 times
its volume a solution containing potassium dihydrogen Sulfate standard solution (10 ppm SO4). 5002800.
phosphate R equivalent to 0.716 g of KH2PO4 in 1000.0 mL. Immediately before use, dilute with distilled water R to
Platinum standard solution (30 ppm Pt). 5002300. 100 times its volume a solution in distilled water R containing
dipotassium sulfate R equivalent to 0.181 g of K2SO4 in
Immediately before use, dilute with 1 M hydrochloric acid 100.0 mL.
to 10 times its volume a solution containing 80 mg of
chloroplatinic acid R in 100.0 mL of 1 M hydrochloric acid. Sulfate standard solution (10 ppm SO4) R1. 5002801.
Potassium standard solution (0.2 per cent K). 5002402. Immediately before use, dilute with ethanol (30 per cent V/V) R
to 100 times its volume a solution containing dipotassium
Dissolve dipotassium sulfate R equivalent to 0.446 g of K2SO4 sulfate R equivalent to 0.181 g of K2SO4 in 100.0 mL of ethanol
in distilled water R and dilute to 100.0 mL with the same (30 per cent V/V) R.
solvent.
Sulfite standard solution (80 ppm SO2). 5005500.
Potassium standard solution (600 ppm K). 5005100.
Dissolve 3.150 g of anhydrous sodium sulfite R in freshly
prepared distilled water R and dilute to 100.0 mL with the
volume a solution containing dipotassium sulfate R equivalent
same solvent. Dilute 0.5 mL to 100.0 mL with freshly prepared
to 2.676 g of K2SO4 in 100.0 mL.
distilled water R.
Potassium standard solution (100 ppm K). 5002400.
Sulfite standard solution (1.5 ppm SO2). 5002900.
volume a solution containing dipotassium sulfate R equivalent Dissolve sodium metabisulfite R equivalent to 0.152 g of
to 0.446 g of K2SO4 in 100.0 mL. Na2S2O5 in water R and dilute to 100.0 mL with the same
solvent. Dilute 5.0 mL of this solution to 100.0 mL with
Potassium standard solution (20 ppm K). 5002401. water R. To 3.0 mL of the resulting solution, add 4.0 mL of
Immediately before use, dilute potassium standard solution 0.1 M sodium hydroxide and dilute to 100.0 mL with water R.
(100 ppm K) R to 5 times its volume with water R. Thallium standard solution (10 ppm Tl). 5003000.
Scandium standard solution (0.1 per cent Sc) for ICP. Dissolve thallous sulfate R equivalent to 0.1235 g of Tl2SO4 in
5006400. a 9 g/L solution of sodium chloride R and dilute to 1000.0 mL
A scandium standard solution (1000 mg/L) suitable for with the same solution. Dilute 10.0 mL of the solution to
inductively coupled plasma (ICP) applications and traceable 100.0 mL with the 9 g/L solution of sodium chloride R.
to national or international standards. Tin liposoluble standard solution (1000 ppm Sn). 5005000.
Selenium standard solution (100 ppm Se). 5002500. A tin (metal) organic compound in an oil.
Dissolve 0.100 g of selenium R in 2 mL of nitric acid R.
Tin standard solution (5 ppm Sn). 5003100.
Evaporate to dryness. Take up the residue in 2 mL of water R
and evaporate to dryness ; carry out three times. Dissolve the Dissolve tin R equivalent to 0.500 g of Sn in a mixture of 5 mL
residue in 50 mL of dilute hydrochloric acid R and dilute to of water R and 25 mL of hydrochloric acid R and dilute to
1000.0 mL with the same acid. 1000.0 mL with water R. Dilute the solution to 100 times its
volume with a 2.5 per cent V/V solution of hydrochloric acid R
Selenium standard solution (1 ppm Se). 5002501. immediately before use.
volume a solution containing selenious acid R equivalent to Tin standard solution (0.1 ppm Sn). 5003101.
6.54 mg of H2SeO3 in 100.0 mL. Immediately before use, dilute tin standard solution (5 ppm
Sn) R to 50 times its volume with water R.
Silver standard solution (5 ppm Ag). 5002600.
Immediately before use, dilute with water R to 100 times its Titanium standard solution (100 ppm Ti). 5003200.
volume a solution containing silver nitrate R equivalent to Dissolve 100.0 mg of titanium R in 100 mL of hydrochloric
0.790 g of AgNO3 in 1000.0 mL. acid R diluted to 150 mL with water R, heating if necessary.
Allow to cool and dilute to 1000 mL with water R.
Sodium standard solution (1000 ppm Na). 5005700.
Dissolve a quantity of anhydrous sodium carbonate R Vanadium standard solution (1 g/L V). 5003300.
equivalent to 2.305 g of Na2CO3 in a mixture of 25 mL of Dissolve in water R ammonium vanadate R equivalent to
water R and 25 mL of nitric acid R and dilute to 1000.0 mL 0.230 g of NH4VO3 and dilute to 100.0 mL with the same
with water R. solvent.
Sodium standard solution (200 ppm Na). 5002700. Zinc standard solution (5 mg/mL Zn). 5003400.
Immediately before use, dilute with water R to 10 times its Dissolve 3.15 g of zinc oxide R in 15 mL of hydrochloric acid R
volume a solution containing sodium chloride R equivalent to and dilute to 500.0 mL with water R.
0.509 g of NaCl in 100.0 mL.
Zinc standard solution (100 ppm Zn). 5003401.
Sodium standard solution (50 ppm Na). 5002701. Immediately before use, dilute with water R to 10 times its
Dilute the sodium standard solution (200 ppm Na) R to four volume a solution containing zinc sulfate R equivalent to
times its volume with water R. 0.440 g of ZnSO4,7H2O and 1 mL of acetic acid R in 100.0 mL.

Zinc standard solution (10 ppm Zn). 5003402. Zirconium standard solution (1 g/L Zr). 5003500.
Immediately before use, dilute zinc standard solution (100 ppm Dissolve zirconyl nitrate R equivalent to 0.293 g of
Zn) R to 10 times its volume with water R. ZrO(NO3)2,2H2O in a mixture of 2 volumes of hydrochloric
acid R and 8 volumes of water R and dilute to 100.0 mL with
Zinc standard solution (5 ppm Zn). 5003403.
the same mixture of solvents.
Immediately before use, dilute zinc standard solution (100 ppm
Zn) R to 20 times its volume with water R.

EUROPEAN PHARMACOPOEIA 4.1.3. Buffer solutions
01/2019:40103 0.1 M Phosphate buffer solution pH 3.0. 4011500.

Dissolve 12.0 g of anhydrous sodium dihydrogen phosphate R
in water R, adjust the pH with dilute phosphoric acid R1 and
Phosphate buffer solution pH 3.0. 4000500.
4.1.3. BUFFER SOLUTIONS
Mix 0.7 mL of phosphoric acid R with 100 mL of water R.
Buffered acetone solution. 4000100. Dilute to 900 mL with the same solvent. Adjust to pH 3.0 with
Dissolve 8.15 g of sodium acetate R and 42 g of sodium strong sodium hydroxide solution R and dilute to 1000 mL
chloride R in water R, add 68 mL of 0.1 M hydrochloric acid with water R.
and 150 mL of acetone R and dilute to 500 mL with water R. Phosphate buffer solution pH 3.0 R1. 4010000.
Buffer solution pH 2.0. 4000200. Dissolve 3.40 g of potassium dihydrogen phosphate R in
Dissolve 6.57 g of potassium chloride R in water R and add 900 mL of water R. Adjust to pH 3.0 with phosphoric acid R
119.0 mL of 0.1 M hydrochloric acid. Dilute to 1000.0 mL with and dilute to 1000.0 mL with water R.
water R. Phosphate buffer solution pH 3.2. 4008100.
0.125 M Phosphate buffer solution pH 2.0. 4015600. To 900 mL of a 4 g/L solution of sodium dihydrogen
Dissolve 17.0 g of potassium dihydrogen phosphate R and phosphate R, add 100 mL of a 2.5 g/L solution of phosphoric
17.8 g of anhydrous disodium hydrogen phosphate R in water R acid R. Adjust the pH if necessary.
and dilute to 1.0 L with the same solvent. If necessary adjust Phosphate buffer solution pH 3.2 R1. 4008500.
the pH with phosphoric acid R.
Adjust a 35.8 g/L solution of disodium hydrogen phosphate
Phosphate buffer solution pH 2.0. 4007900. dodecahydrate R to pH 3.2 with dilute phosphoric acid R.
Dilute 100.0 mL of the solution to 2000.0 mL with water R.
Dissolve 8.95 g of disodium hydrogen phosphate
dodecahydrate R and 3.40 g of potassium dihydrogen Phosphate buffer solution pH 3.25. 4014900.
phosphate R in water R and dilute to 1000.0 mL with the same
Dissolve about 1.36 g of potassium dihydrogen phosphate R in
solvent. If necessary adjust the pH with phosphoric acid R.
1000 mL of water R and adjust to pH 3.25 ± 0.05 with dilute
Sulfate buffer solution pH 2.0. 4008900. phosphoric acid R. Filter through a membrane filter (nominal
pore size 0.45 µm or finer).
Dissolve 132.1 g of ammonium sulfate R in water R and dilute
to 500.0 mL with the same solvent (Solution A). Carefully and Phosphate buffer solution pH 3.4. 4015800.
with constant cooling stir 14 mL of sulfuric acid R into about Dissolve 68.0 g of potassium dihydrogen phosphate R in
400 mL of water R ; allow to cool and dilute to 500.0 mL with water R and dilute to 1000.0 mL with the same solvent. Adjust
water R (Solution B). Mix equal volumes of solutions A and B. the pH with phosphoric acid R.
Adjust the pH if necessary.
Buffer solution pH 3.5. 4000600.
Dissolve 25.0 g of ammonium acetate R in 25 mL of water R
Mix 6.7 mL of phosphoric acid R with 55.0 mL of a 40 g/L and add 38.0 mL of hydrochloric acid R1. Adjust the
solution of sodium hydroxide R and dilute to 1000.0 mL with pH if necessary with dilute hydrochloric acid R or dilute
water R. ammonia R1. Dilute to 100.0 mL with water R.
Buffer solution pH 2.5. 4000300. Phosphate buffer solution pH 3.5. 4000700.
Dissolve 100 g of potassium dihydrogen phosphate R in 800 mL Dissolve 68.0 g of potassium dihydrogen phosphate R in
of water R ; adjust to pH 2.5 with hydrochloric acid R and water R and dilute to 1000.0 mL with the same solvent. Adjust
dilute to 1000.0 mL with water R. the pH with phosphoric acid R.
Buffer solution pH 2.5 R1. 4000400. Buffer solution pH 3.6. 4000800.
To 4.9 g of dilute phosphoric acid R add 250 mL of water R. To 250.0 mL of 0.2 M potassium hydrogen phthalate R add
Adjust the pH with dilute sodium hydroxide solution R and 11.94 mL of 0.2 M hydrochloric acid. Dilute to 1000.0 mL with
dilute to 500.0 mL with water R. water R.
0.2 M Phosphate buffer solution pH 2.5. 4014100. Buffer solution pH 3.7. 4000900.
Dissolve 27.2 g of potassium dihydrogen phosphate R in about To 15.0 mL of acetic acid R add 60 mL of ethanol (96 per
900 mL of water R, adjust to pH 2.5 with phosphoric acid R cent) R and 20 mL of water R. Adjust to pH 3.7 by the addition
and dilute to 1.0 L with water R. of ammonia R. Dilute to 100.0 mL with water R.
Phosphate buffer solution pH 2.8. 4010600. Buffered copper sulfate solution pH 4.0. 4001000.
Dissolve 7.8 g of sodium dihydrogen phosphate R in 900 mL of Dissolve 0.25 g of copper sulfate pentahydrate R and 4.5 g
water R, adjust to pH 2.8 with phosphoric acid R and dilute to of ammonium acetate R in dilute acetic acid R and dilute to
1000 mL with the same solvent. 100.0 mL with the same solvent.
Buffer solution pH 3.0. 4008000. 0.1 M Sodium acetate buffer solution pH 4.0. 4013800.
Dissolve 21.0 g of citric acid monohydrate R in 200 mL of 1 M Dissolve 822 mg of sodium acetate R in 100 mL of water R
sodium hydroxide and dilute to 1000 mL with water R. Dilute (solution A). Dilute 1.44 mL of glacial acetic acid R in 250 mL
40.3 mL of this solution to 100.0 mL with 0.1 M hydrochloric of water R (solution B). Titrate 100 mL of solution B using
acid. about 20 mL of solution A.
0.25 M Citrate buffer solution pH 3.0. 4012600. Acetate buffer solution pH 4.4. 4001100.
Dissolve 5.3 g of citric acid monohydrate R in 80 mL of Dissolve 136 g of sodium acetate R and 77 g of ammonium
water R. Adjust the pH with 1 M sodium hydroxide and dilute acetate R in water R and dilute to 1000.0 mL with the same
to 100.0 mL with water R. solvent ; add 250.0 mL of glacial acetic acid R and mix.

4.1.3. Buffer solutions EUROPEAN PHARMACOPOEIA
Phthalate buffer solution pH 4.4. 4001200. Sodium acetate buffer solution pH 5.0. 4015500.
Dissolve 2.042 g of potassium hydrogen phthalate R in 50 mL Dissolve 50.0 g of sodium acetate R in 10.0 mL of glacial
of water R, add 7.5 mL of 0.2 M sodium hydroxide and dilute acetic acid R and add water R. Adjust to pH 5.0 with a 4.2 g/L
to 200.0 mL with water R. solution of sodium hydroxide R or with glacial acetic acid R
and dilute to 1000.0 mL with water R.
Acetate buffer solution pH 4.5. 4012500.
Dissolve 77.1 g of ammonium acetate R in water R. Add 70 mL Dissolve 1.02 g of potassium hydrogen phthalate R in 30.0 mL
of glacial acetic acid R and dilute to 1000.0 mL with water R. of 0.1 M sodium hydroxide. Dilute to 100.0 mL with water R.
0.5 M Ammonium acetate buffer solution pH 4.5. 4014200. 0.067 M Phosphate buffer solution pH 5.4. 4012000.
Mix 14.3 mL of glacial acetic acid R and 470 mL of water R Mix appropriate volumes of a 23.99 g/L solution of disodium
and adjust to pH 4.5 with concentrated ammonia R. Dilute to hydrogen phosphate dodecahydrate R with a 9.12 g/L solution
500.0 mL with water R. of sodium dihydrogen phosphate monohydrate R to obtain
pH 5.4.
0.05 M Phosphate buffer solution pH 4.5. 4009000.
Acetate-edetate buffer solution pH 5.5. 4001900.
Dissolve 6.80 g of potassium dihydrogen phosphate R in
1000.0 mL of water R. The pH of the solution is 4.5. Dissolve 250 g of ammonium acetate R and 15 g sodium
edetate R in 400 mL of water R and add 125 mL of glacial
Sodium acetate buffer solution pH 4.5. 4010100. acetic acid R.
Dissolve 63 g of anhydrous sodium acetate R in water R, Buffer solution pH 5.5. 4001800.
add 90 mL acetic acid R and adjust to pH 4.5, and dilute to Dissolve 54.4 g of sodium acetate R in 50 mL of water R,
1000 mL with water R. heating to 35 °C if necessary. After cooling, slowly add 10 mL
of anhydrous acetic acid R. Shake and dilute to 100.0 mL with
Acetate buffer solution pH 4.6. 4001400. water R.
Dissolve 5.4 g of sodium acetate R in 50 mL of water R, add
2.4 g of glacial acetic acid R and dilute to 100.0 mL with Phosphate buffer solution pH 5.5. 4002000.
water R. Adjust the pH if necessary. Dissolve 13.61 g of potassium dihydrogen phosphate R in
water R and dilute to 1000.0 mL with the same solvent
Succinate buffer solution pH 4.6. 4001500. (solution A). Dissolve 35.81 g of disodium hydrogen phosphate
dodecahydrate R in water R and dilute to 1000.0 mL with the
Disssolve 11.8 g of succinic acid R in a mixture of 600 mL of
same solvent (solution B). Mix 96.4 mL of solution A and
water R and 82 mL of 1 M sodium hydroxide and dilute to
3.6 mL of solution B.
1000.0 mL with water R.
Phosphate-citrate buffer solution pH 5.5. 4008700.
Acetate buffer solution pH 4.7. 4001600. Mix 56.85 mL of a 28.4 g/L solution of anhydrous disodium
Dissolve 136.1 g of sodium acetate R in 500 mL of water R. Mix hydrogen phosphate R and 43.15 mL of a 21 g/L solution of
250 mL of this solution with 250 mL of dilute acetic acid R. citric acid monohydrate R.
Shake twice with a freshly prepared, filtered, 0.1 g/L solution of
dithizone R in chloroform R. Shake with carbon tetrachloride R Phosphate buffer solution pH 5.6. 4011200.
until the extract is colourless. Filter the aqueous layer to Dissolve 0.908 g of potassium dihydrogen phosphate R
remove traces of carbon tetrachloride. in water R and dilute to 100.0 mL with the same solvent
(solution A). Dissolve 1.161 g of dipotassium hydrogen
Acetate buffer solution pH 4.7 R1. 4013600. phosphate R in water R and dilute to 100.0 mL with the same
Dissolve 136.1 g of sodium acetate R in 500 mL of water R. Mix solvent (solution B). Mix 94.4 mL of solution A and 5.6 mL
250 mL of this solution with 250 mL of dilute acetic acid R. of solution B. If necessary, adjust to pH 5.6 using solution A
or solution B.
Acetate buffer solution pH 5.0. 4009100. Phosphate buffer solution pH 5.8. 4002100.
To 120 mL of a 6 g/L solution of glacial acetic acid R add Dissolve 1.19 g of disodium hydrogen phosphate dihydrate R
100 mL of 0.1 M potassium hydroxide and about 250 mL of and 8.25 g of potassium dihydrogen phosphate R in water R
water R. Mix. Adjust the pH to 5.0 with a 6 g/L solution of and dilute to 1000.0 mL with the same solvent.
acetic acid R or with 0.1 M potassium hydroxide and dilute to
1000.0 mL with water R. Acetate buffer solution pH 6.0. 4002200.
Dissolve 100 g of ammonium acetate R in 300 mL of water R,
Citrate buffer solution pH 5.0. 4010700. add 4.1 mL of glacial acetic acid R, adjust the pH if necessary
Prepare a solution containing 20.1 g/L of citric acid using ammonia R or acetic acid R and dilute to 500.0 mL with
monohydrate R and 8.0 g/L of sodium hydroxide R. Adjust the water R.
pH with dilute hydrochloric acid R. Diethylammonium phosphate buffer solution pH 6.0.
0.2 M Deuterated sodium phosphate buffer solution 4002300.
pH 5.0. 4013900. Dilute 68 mL of phosphoric acid R to 500 mL with water R.
To 25 mL of this solution add 450 mL of water R and 6 mL
Dissolve 2.76 g of sodium dihydrogen phosphate monohydrate R of diethylamine R, adjust to pH 6 ± 0.05, if necessary, using
in 90 mL of deuterium oxide R, adjust the pH with a deuterated diethylamine R or phosphoric acid R and dilute to 500.0 mL
solution of phosphoric acid R or a deuterated 1 M solution of with water R.
sodium hydroxide R, dilute to 100 mL with deuterium oxide R
and mix. 1 M Morpholinoethanesulfonate buffer solution pH 6.0.
4015900.
Phosphate buffer solution pH 5.0. 4011300. Dissolve 48.8 g of 2-[N-morpholino]ethanesulfonic acid R in
Dissolve 2.72 g of potassium dihydrogen phosphate R in 160 mL of water R and add 25 mL of 2 M sodium hydroxide R.
800 mL of water R. Adjust the pH with a 1 M potassium Adjust to pH 6.0 with 2 M sodium hydroxide R. Dilute to
hydroxide solution prepared from potassium hydroxide R and almost 250 mL with water R. Adjust the pH, if necessary, with
dilute to 1000 mL with water R. 2 M sodium hydroxide R and dilute to 250.0 mL with water R.

Phosphate buffer solution pH 6.0. 4002400. Phosphate buffer solution pH 6.8 R1. 4003400.
Mix 63.2 mL of a 71.5 g/L solution of disodium hydrogen To 51.0 mL of a 27.2 g/L solution of potassium dihydrogen
phosphate dodecahydrate R and 36.8 mL of a 21 g/L solution phosphate R add 49.0 mL of a 71.6 g/L solution of disodium
of citric acid monohydrate R. hydrogen phosphate dodecahydrate R. Adjust the pH if
necessary.
Phosphate buffer solution pH 6.0 R1. 4002500.
Dissolve 6.8 g of sodium dihydrogen phosphate R in water R
and dilute to 1000.0 mL with water R. Adjust the pH with 1 M Tris-hydrochloride buffer solution pH 6.8. 4009300.
strong sodium hydroxide solution R. Dissolve 60.6 g of tris(hydroxymethyl)aminomethane R in
Phosphate buffer solution pH 6.0 R2. 4002600. 400 mL of water R. Adjust the pH with hydrochloric acid R
and dilute to 500.0 mL with water R.
To 250.0 mL of 0.2 M potassium dihydrogen phosphate R add
28.5 mL of 0.2 M sodium hydroxide and dilute to 1000.0 mL Buffer solution pH 7.0. 4003500.
with water R.
To 1000 mL of a solution containing 18 g/L of disodium
Phosphate buffer solution pH 6.4. 4002800. hydrogen phosphate dodecahydrate R and 23 g/L of sodium
Dissolve 2.5 g of disodium hydrogen phosphate chloride R add sufficient (about 280 mL) of a solution
dodecahydrate R, 2.5 g of sodium dihydrogen phosphate R containing 7.8 g/L of sodium dihydrogen phosphate R and
and 8.2 g of sodium chloride R in 950 mL of water R. Adjust 23 g/L of sodium chloride R to adjust the pH. Dissolve in the
the pH of the solution to 6.4 with 1 M sodium hydroxide or solution sufficient sodium azide R to give a 0.2 g/L solution.
1 M hydrochloric acid, if necessary. Dilute to 1000.0 mL with Maleate buffer solution pH 7.0. 4003600.
water R.
Dissolve 10.0 g of sodium chloride R, 6.06 g of
0.5 M Phthalate buffer solution pH 6.4. 4009200. tris(hydroxymethyl)aminomethane R and 4.90 g of
Dissolve 100 g of potassium hydrogen phthalate R in water R maleic anhydride R in 900 mL of water R. Adjust the pH using
and dilute to 1000.0 mL with the same solvent. Adjust the pH a 170 g/L solution of sodium hydroxide R. Dilute to 1000.0 mL
if necessary, using strong sodium hydroxide solution R. with water R.
Storage : at 2 °C to 8 °C ; use within 3 days.
Dissolve 60.5 g of disodium hydrogen phosphate 0.025 M Phosphate buffer solution pH 7.0. 4009400.
dodecahydrate R and 46 g of potassium dihydrogen phosphate R Mix 1 volume of 0.063 M phosphate buffer solution pH 7.0 R
in water R. Add 100 mL of 0.02 M sodium edetate and 20 mg with 1.5 volumes of water R.
of mercuric chloride R and dilute to 1000.0 mL with water R.
Imidazole buffer solution pH 6.5. 4003000.
Dissolve 5.2 g of dipotassium hydrogen phosphate R in
Dissolve 6.81 g of imidazole R, 1.23 g of magnesium sulfate R 900 mL of water for chromatography R. Adjust the solution to
and 0.73 g of calcium sulfate R in 752 mL of 0.1 M hydrochloric pH 7.0 ± 0.1 using phosphoric acid R and dilute to 1000 mL
acid. Adjust the pH if necessary and dilute to 1000.0 mL with with water for chromatography R.
water R.
0.1 M phosphate buffer solution pH 6.5. 4010800.
Mix 34 mL of water R and 100 mL of 0.067 M phosphate buffer
Dissolve 13.80 g of sodium dihydrogen phosphate solution pH 7.0 R.
monohydrate R in 900 mL of distilled water R. Adjust the pH
using a 400 g/L solution of sodium hydroxide R. Dilute to 0.063 M Phosphate buffer solution pH 7.0. 4009500.
1000 mL with distilled water R.
Dissolve 5.18 g of anhydrous disodium hydrogen phosphate R
Phosphate buffer solution pH 6.5. 4012800. and 3.65 g of sodium dihydrogen phosphate monohydrate R in
950 mL of water R and adjust the pH with phosphoric acid R ;
Dissolve 2.75 g of sodium dihydrogen phosphate R and 4.5 g of
dilute to 1000.0 mL with water R.
sodium chloride R in 500 mL of water R. Adjust the pH with
phosphate buffer solution pH 8.5 R. 0.067 M Phosphate buffer solution pH 7.0. 4003800.
Buffer solution pH 6.6. 4003100. Dissolve 0.908 g of potassium dihydrogen phosphate R
To 250.0 mL of 0.2 M potassium dihydrogen phosphate R add in water R and dilute to 100.0 mL with the same solvent
89.0 mL of 0.2 M sodium hydroxide. Dilute to 1000.0 mL with (solution A). Dissolve 2.38 g of disodium hydrogen phosphate
water R. dodecahydrate R in water R and dilute to 100.0 mL with the
same solvent (solution B). Mix 38.9 mL of solution A and
0.1 M Phosphate buffer solution pH 6.7. 4014300. 61.1 mL of solution B. Adjust the pH if necessary.
Dissolve 15.6 g of sodium dihydrogen phosphate R in water R 0.1 M Phosphate buffer solution pH 7.0. 4008200.
and dilute to 1.0 L with the same solvent. Dissolve 17.8 g
of disodium hydrogen phosphate dihydrate R in water R and Dissolve 1.361 g of potassium dihydrogen phosphate R in
dilute to 1.0 L with the same solvent. Mix the solutions, check water R and dilute to 100.0 mL with the same solvent. Adjust
the pH and if necessary adjust to pH 6.7. the pH using a 35 g/L solution of disodium hydrogen phosphate
dodecahydrate R.
Phosphate buffered saline pH 6.8. 4003200.
Dissolve 1.0 g of potassium dihydrogen phosphate R, 2.0 g Phosphate buffer solution pH 7.0. 4003700.
of dipotassium hydrogen phosphate R and 8.5 g of sodium Mix 82.4 mL of a 71.5 g/L solution of disodium hydrogen
chloride R in 900 mL of water R, adjust the pH if necessary phosphate dodecahydrate R with 17.6 mL of a 21 g/L solution
and dilute to 1000.0 mL with the same solvent. of citric acid monohydrate R.
Phosphate buffer solution pH 6.8. 4003300. Phosphate buffer solution pH 7.0 R1. 4003900.
Mix 77.3 mL of a 71.5 g/L solution of disodium hydrogen Mix 250.0 mL of 0.2 M potassium dihydrogen phosphate R and
phosphate dodecahydrate R with 22.7 mL of a 21 g/L solution 148.2 mL of a 8 g/L solution of sodium hydroxide R, adjust the
of citric acid monohydrate R. pH if necessary. Dilute to 1000.0 mL with water R.

Phosphate buffer solution pH 7.0 R2. 4004000. Phosphate buffer solution pH 7.2. 4004200.
Mix 50.0 mL of a 136 g/L solution of potassium dihydrogen Mix 87.0 mL of a 71.5 g/L solution of disodium hydrogen
phosphate R with 29.5 mL of 1 M sodium hydroxide and dilute phosphate dodecahydrate R with 13.0 mL of a 21 g/L solution
to 100.0 mL with water R. Adjust the pH to 7.0 ± 0.1. of citric acid monohydrate R.
Phosphate buffer solution pH 7.0 R3. 4008600. Imidazole buffer solution pH 7.3. 4004500.
Dissolve 5 g of potassium dihydrogen phosphate R and 11 g Dissolve 3.4 g of imidazole R and 5.8 g of sodium chloride R in
of dipotassium hydrogen phosphate R in 900 mL of water R. water R, add 18.6 mL of 1 M hydrochloric acid and dilute to
Adjust to pH 7.0 with dilute phosphoric acid R or dilute sodium 1000.0 mL with water R. Adjust the pH if necessary.
hydroxide solution R. Dilute to 1000 mL with water R and mix.
Barbital buffer solution pH 7.4. 4004700.
Mix 50 mL of a solution in water R containing 19.44 g/L of
Dissolve 28.4 g of anhydrous disodium hydrogen phosphate R sodium acetate R and 29.46 g/L of barbital sodium R with
and 18.2 g of potassium dihydrogen phosphate R in water R 50.5 mL of 0.1 M hydrochloric acid, add 20 mL of an 85 g/L of
and dilute to 500 mL with the same solvent. sodium chloride R and dilute to 250 mL with water R.
Phosphate buffer solution pH 7.0 R5. 4011400. Buffer solution pH 7.4. 4004600.
Dissolve 28.4 g of anhydrous disodium hydrogen phosphate R Dissolve 0.6 g of potassium dihydrogen phosphate R, 6.4 g of
in 800 mL of water R. Adjust the pH using a 30 per cent m/m disodium hydrogen phosphate dodecahydrate R and 5.85 g of
solution of phosphoric acid R and dilute to 1000 mL with sodium chloride R in water R, and dilute to 1000.0 mL with the
water R. same solvent. Adjust the pH if necessary.
Phosphate buffered saline pH 7.4. 4005000.
Dissolve 3.56 g of disodium hydrogen phosphate dihydrate R
in 950 mL of water for chromatography R. Adjust the pH Dissolve 2.38 g of disodium hydrogen phosphate
with phosphoric acid R and dilute to 1.0 L with water for dodecahydrate R, 0.19 g of potassium dihydrogen phosphate R
chromatography R. and 8.0 g of sodium chloride R in water. Dilute to 1000.0 mL
with the same solvent. Adjust the pH if necessary.
Phosphate buffer solution pH 7.4. 4004800.
Dissolve 35 g of dipotassium hydrogen phosphate R in 900 mL
of water R, adjust to pH 7.0 with dilute phosphoric acid R and Add 250.0 mL of 0.2 M potassium dihydrogen phosphate R to
dilute to 1.0 L with water R. 393.4 mL of 0.1 M sodium hydroxide.
Potassium phosphate buffer solution pH 7.0. 4014700. Tris(hydroxymethyl)aminomethane buffer solution
Dissolve 10 mg of bovine albumin R and 68 mg of potassium pH 7.4. 4012100.
dihydrogen phosphate R in 30 mL of water R. If necessary, Dissolve 30.3 g of tris(hydroxymethyl)aminomethane R
adjust to pH 7.0 with potassium hydroxide R. Dilute to 50 mL in approximately 200 mL of water R. Add 183 mL of 1 M
with water R and filter. hydrochloric acid. Dilute to 500.0 mL with water R. Note :
the pH is 7.7-7.8 at room temperature and 7.4 at 37 °C. This
Sodium/calcium acetate buffer solution pH 7.0. 4014800. solution is stable for several months at 4 °C.
Dissolve 10 mg of bovine albumin R and 32 mg of calcium
acetate R in 60 mL of water R. Add 580 µL of glacial acetic Tris(hydroxymethyl)aminomethane sodium chloride
acid R and adjust to pH 7.0 with 2 M sodium hydroxide R. buffer solution pH 7.4. 4004900.
Dilute to 100 mL with water R and filter. Dissolve 6.08 g of tris(hydroxymethyl)aminomethane R, 8.77 g
of sodium chloride R in 500 mL of distilled water R. Add 10.0 g
Tetrabutylammonium buffer solution pH 7.0. 4010900. of bovine albumin R. Adjust the pH using hydrochloric acid R.
Dissolve 6.16 g of ammonium acetate R in a mixture of 15 mL Dilute to 1000.0 mL with distilled water R.
of tetrabutylammonium hydroxide solution (400 g/L) R and
185 mL of water R. Adjust the pH with nitric acid R. Tris(hydroxymethyl)aminomethane sodium chloride
buffer solution pH 7.4 R1. 4012200.
Buffered salt solution pH 7.2. 4004300.
Dissolve 0.1 g of bovine albumin R in a mixture containing
Dissolve in water R 8.0 g of sodium chloride R, 0.2 g of 2 mL of tris(hydroxymethyl)aminomethane buffer solution
potassium chloride R, 0.1 g of anhydrous calcium chloride R, pH 7.4 R and 50 mL of a 5.84 mg/mL solution of sodium
0.1 g of magnesium chloride R, 3.18 g of disodium hydrogen chloride R. Dilute to 100.0 mL with water R.
phosphate dodecahydrate R and 0.2 g of potassium dihydrogen
phosphate R and dilute to 1000.0 mL with water R. Tris-sodium acetate buffer solution pH 7.4. 4012900.
Buffer solution pH 7.2. 4004100. Dissolve 6.3 g of tris(hydroxymethyl)aminomethane R and
4.9 g of anhydrous sodium acetate R in 900 mL of water R.
To 250.0 mL of 0.2 M potassium dihydrogen phosphate R add Adjust to pH 7.4 with sulfuric acid R and dilute to 1000 mL
175.0 mL of 0.2 M sodium hydroxide. Dilute to 1000.0 mL with water R.
with water R. Adjust the pH if necessary.
Phosphate-albumin buffered saline pH 7.2. 4004400. Tris-sodium acetate-sodium chloride buffer solution
pH 7.4. 4013000.
dodecahydrate R, 7.6 g of sodium chloride R and 10 g of bovine Dissolve 30.0 g of tris(hydroxymethyl)aminomethane R, 14.5 g
albumin R in water R and dilute to 1000.0 mL with the same of anhydrous sodium acetate R and 14.6 g of sodium chloride R
solvent. Immediately before use adjust the pH using dilute in 900 mL of water R. Add 0.50 g of bovine albumin R. Adjust
sodium hydroxide solution R or dilute phosphoric acid R. to pH 7.4 with sulfuric acid R and dilute to 1000 mL with
water R.
Phosphate-albumin buffered saline pH 7.2 R1. 4009600.
Borate buffer solution pH 7.5. 4005200.
dodecahydrate R, 7.6 g of sodium chloride R and 1 g of bovine Dissolve 2.5 g of sodium chloride R, 2.85 g of disodium
albumin R in water R and dilute to 1000.0 mL with the same tetraborate R and 10.5 g of boric acid R in water R and dilute to
solvent. Immediately before use adjust the pH using dilute 1000.0 mL with the same solvent. Adjust the pH if necessary.
sodium hydroxide solution R or dilute phosphoric acid R. Storage : at 2 °C to 8 °C.

Buffer (HEPES) solution pH 7.5. 4009700. Buffer solution pH 8.0 R1. 4010400.

Dissolve 2.38 g of HEPES R in about 90 mL of water R. Adjust Dissolve 20 g of dipotassium hydrogen phosphate R in 900 mL
the pH to 7.5 with sodium hydroxide solution R. Dilute to of water R. Adjust the pH with phosphoric acid R. Dilute to
100 mL with water R. 1000 mL with water R.
0.05 M Phosphate buffer solution pH 7.5. 4014400. 0.02 M Phosphate buffer solution pH 8.0. 4006100.
Dissolve 0.89 g of disodium hydrogen phosphate dihydrate R To 50.0 mL of 0.2 M potassium dihydrogen phosphate R add
in about 80 mL of water R. Adjust to pH 7.5 with an 8.5 per 46.8 mL of 0.2 M sodium hydroxide. Dilute to 500.0 mL with
cent V/V solution of phosphoric acid R and dilute to 100.0 mL water R.
with water R.
0.2 M Phosphate buffer solution pH 7.5. 4005400. Dissolve 0.523 g of potassium dihydrogen phosphate R and
Dissolve 27.22 g of potassium dihydrogen phosphate R in 16.73 g of dipotassium hydrogen phosphate R in water R and
930 mL of water R, adjust to pH 7.5 with a 300 g/L solution of dilute to 1000.0 mL with the same solvent.
potassium hydroxide R and dilute to 1000.0 mL with water R.
1 M Phosphate buffer solution pH 8.0. 4007800.
0.33 M Phosphate buffer solution pH 7.5. 4005300. Dissolve 136.1 g of potassium dihydrogen phosphate R in
Dissolve 119.31 g of disodium hydrogen phosphate water R, adjust the pH with 1 M sodium hydroxide. Dilute to
dodecahydrate R in water R and dilute to 1000.0 mL with 1000.0 mL with water R.
the same solvent (solution A). Dissolve 45.36 g of potassium 0.02 M Sodium phosphate buffer solution pH 8.0. 4013700.
dihydrogen phosphate R in water R and dilute to 1000.0 mL
with the same solvent (solution B). Mix 85 mL of solution A Dissolve 0.31 g of sodium dihydrogen phosphate R in 70 mL of
and 15 mL of solution B. Adjust the pH if necessary. water R and adjust to pH 8.0 with 1 M sodium hydroxide, then
0.25 M Sodium phosphate buffer solution pH 7.5. 4016100.
1 M Tris-hydrochloride buffer solution pH 8.0. 4012700.
Dissolve 3.90 g of sodium dihydrogen phosphate R in 70 mL of
water R, adjust to pH 7.5 with a 300 g/L solution of sodium Dissolve 121.1 g of tris(hydroxymethyl)aminomethane R and
hydroxide R and dilute to 100 mL with water R. 1.47 g of calcium chloride R in 900 mL of water R. Adjust the
pH with hydrochloric acid R and dilute to 1000.0 mL with
0.05 M Tris-hydrochloride buffer solution pH 7.5. 4005600. water R.
Dissolve 6.057 g of tris(hydroxymethyl)aminomethane R in Tris-hydrochloride buffer solution pH 8.0. 4012300.
water R and adjust the pH with hydrochloric acid R. Dilute to Dissolve 1.21 g of tris(hydroxymethyl)aminomethane R and
1000.0 mL with water R. 29.4 mg of calcium chloride R in water R. Adjust the pH with
0.1 M Tris-hydrochloride buffer solution pH 7.5. 4016200. 1 M hydrochloric acid and dilute to 100.0 mL with water R.
Dissolve 3.03 g of tris(hydroxymethyl)aminomethane R in Tris-sodium acetate buffer solution pH 8.0. 4013100.
200 mL of water R, adjust to pH 7.5 with hydrochloric acid R Dissolve 6.3 g of tris(hydroxymethyl)aminomethane R and
and dilute to 250 mL with water R. 4.9 g of anhydrous sodium acetate R in 900 mL of water R.
1 M Tris-hydrochloride buffer solution pH 7.5. 4014500. Adjust to pH 8.0 with sulfuric acid R and dilute to 1000 mL
with water R.
Dissolve 12.11 g of tris(hydroxymethyl)aminomethane R in
90 mL of water R, adjust to pH 7.5 with hydrochloric acid R Tris-sodium acetate-sodium chloride buffer solution
and dilute to 100.0 mL with water R. pH 8.0. 4013200.
Tris(hydroxymethyl)aminomethane buffer solution Dissolve 30.0 g of tris(hydroxymethyl)aminomethane R, 14.5 g
pH 7.5. 4005500. of anhydrous sodium acetate R and 14.6 g of sodium chloride R
in 900 mL of water R. Add 0.50 g of bovine albumin R. Adjust
Dissolve 7.27 g of tris(hydroxymethyl)aminomethane R and to pH 8.0 with sulfuric acid R and dilute to 1000 mL with
5.27 g of sodium chloride R in water R, and adjust the pH if water R.
necessary. Dilute to 1000.0 mL with water R.
Tris(hydroxymethyl)aminomethane buffer solution
Tris(hydroxymethyl)aminomethane buffer solution pH 8.1. 4006200.
pH 7.5 R1. 4016400.
Dissolve 0.294 g of calcium chloride R in 40 mL of
Dissolve 1.21 g of tris(hydroxymethyl)aminomethane R in tris(hydroxymethyl)aminomethane solution R and adjust the
900 mL of water R and add 10 mL of 0.01 M calcium chloride pH with 1 M hydrochloric acid. Dilute to 100.0 mL with
solution R. Adjust the pH if necessary with sodium hydroxide water R.
solution R or hydrochloric acid R, and dilute to 1000.0 mL
with water R. Guanidine-tris(hydroxymethyl)aminomethane buffer
solution pH 8.3. 4016300.
Sodium citrate buffer solution pH 7.8 (0.034 M sodium Dissolve 1.21 g of tris(hydroxymethyl)aminomethane R in
citrate, 0.101 M sodium chloride). 4009800. 87.5 mL of a 764 g/L solution of guanidine hydrochloride R.
Dissolve 10.0 g of sodium citrate R and 5.90 g of sodium Adjust to pH 8.3 with hydrochloric acid R and dilute to 100 mL
chloride R in 900 mL of water R. Adjust the pH by addition of with water R.
hydrochloric acid R and dilute to 1000 mL with water R.
Tris-glycine buffer solution pH 8.3. 4006300.
0.0015 M Borate buffer solution pH 8.0. 4006000. Dissolve 6.0 g of tris(hydroxymethyl)aminomethane R and
Dissolve 0.572 g of disodium tetraborate R and 2.94 g of 28.8 g of glycine R in water R and dilute to 1000.0 mL with
calcium chloride R in 800 mL of water R. Adjust the pH with the same solvent. Dilute 1 volume to 10 volumes with water R
1 M hydrochloric acid. Dilute to 1000.0 mL with water R. immediately before use.
Buffer solution pH 8.0. 4005900. Tris-hydrochloride buffer solution pH 8.3. 4011800.
To 50.0 mL of 0.2 M potassium dihydrogen phosphate R add Dissolve 9.0 g of tris(hydroxymethyl)aminomethane R in 2.9 L
46.8 mL of 0.2 M sodium hydroxide. Dilute to 200.0 mL with of water R. Adjust the pH with 1 M hydrochloric acid. Adjust
water R. the volume to 3 L with water R.

Barbital buffer solution pH 8.4. 4006400. Buffer solution pH 9.0. 4007000.

Dissolve 8.25 g of barbital sodium R in water R and dilute to Dissolve 6.18 g of boric acid R in 0.1 M potassium chloride R
1000.0 mL with the same solvent. and dilute to 1000.0 mL with the same solvent. Mix 1000.0 mL
of this solution and 420.0 mL of 0.1 M sodium hydroxide.
Tris-EDTA BSA buffer solution pH 8.4. 4006500.
Dissolve 6.1 g of tris(hydroxymethyl)aminomethane R, 2.8 g Buffer solution pH 9.0 R1. 4007100.
of sodium edetate R, 10.2 g of sodium chloride R and 10 g Dissolve 6.20 g of boric acid R in 500 mL of water R and adjust
of bovine albumin R in water R, adjust to pH 8.4 using 1 M the pH with 1 M sodium hydroxide (about 41.5 mL). Dilute to
hydrochloric acid and dilute to 1000.0 mL with water R. 1000.0 mL with water R.
Tris(hydroxymethyl)aminomethane-EDTA buffer solution 0.05 M Tris-hydrochloride buffer solution pH 9.0. 4013500.
pH 8.4. 4006600. Dissolve 0.605 g of tris(hydroxymethyl)aminomethane R in
Dissolve 5.12 g of sodium chloride R, 3.03 g of water R. Adjust the pH with 1 M hydrochloric acid and dilute
tris(hydroxymethyl)aminomethane R and 1.40 g of to 100.0 mL with water R.
sodium edetate R in 250 mL of distilled water R. Adjust the
pH to 8.4 using hydrochloric acid R. Dilute to 500.0 mL with Tris(hydroxymethyl)aminomethane buffer solution
distilled water R. pH 9.0. 4015200.
Dissolve 1.21 g of tris(hydroxymethyl)aminomethane R in
Tris(hydroxymethyl)aminomethane-EDTA buffer solution 950 mL of water for chromatography R. Adjust to pH 9.0
pH 8.4 R1. 4015100. with acetic acid R and dilute to 1000.0 mL with water for
Dissolve 10.20 g of sodium chloride R, 6.10 g of chromatography R.
tris(hydroxymethyl)aminomethane R, 2.80 g of sodium
edetate R and 1.00 g of macrogol 6000 R or 2.00 g of bovine Tris(hydroxymethyl)aminomethane buffer solution
albumin R or of human albumin R in 800 mL of water R. pH 9.0 R1. 4016600.
Adjust to pH 8.4 with hydrochloric acid R and dilute to 1.0 L Dissolve 12.1 g of tris(hydroxymethyl)aminomethane R in
with water R. 950 mL of water R. Adjust to pH 9.0 with acetic acid R and
dilute to 1000.0 mL with water R.
Guanidine-tris(hydroxymethyl)aminomethane-EDTA
buffer solution pH 8.5. 4014600. Ammonium chloride buffer solution pH 9.5. 4007200.
Dissolve 1.0 g of sodium edetate R, 12.1 g of tris(hydroxy- Dissolve 33.5 g of ammonium chloride R in 150 mL of water R,
methyl)aminomethane R and 57.0 g of guanidine add 42.0 mL of concentrated ammonia R and dilute to
hydrochloride R in 35 mL of water R. Adjust to pH 8.5 with 250.0 mL with water R.
hydrochloric acid R and dilute to 100 mL with water R. Storage : in a polyethylene container.
Phosphate buffer solution pH 8.5. 4013300. Ammonium chloride buffer solution pH 10.0. 4007300.
Dissolve 3.5 g of dipotassium hydrogen phosphate R and 4.5 g Dissolve 5.4 g of ammonium chloride R in 20 mL of water R,
of sodium chloride R in 500 mL of water R. Adjust the pH add 35.0 mL of ammonia R and dilute to 100.0 mL with
with a mixture of equal volumes of dilute phosphoric acid R water R.
and water R.
Borate buffer solution pH 10.0. 4016000.
Tris acetate buffer solution pH 8.5. 4006700. Introduce 12.4 g of boric acid R into a 500.0 mL volumetric
Dissolve 0.294 g of calcium chloride R and 12.11 g of flask. Add 300 mL of water R to suspend the boric acid. Add
tris(hydroxymethyl)aminomethane R in water R. Adjust the 100 mL of a 56 g/L solution of potassium hydroxide R and mix
pH with acetic acid R. Dilute to 1000.0 mL with water R. to dissolve the boric acid. Adjust to pH 10.0 by slowly adding
a 56 g/L solution of potassium hydroxide R (about 60 mL is
Barbital buffer solution pH 8.6 R1. 4006900. usually needed). Mix. Dilute almost to volume with water R.
Dissolve in water R 1.38 g of barbital R, 8.76 g of barbital If necessary, adjust the pH with boric acid R or with a 56 g/L
sodium R and 0.38 g of calcium lactate pentahydrate R and solution of potassium hydroxide R. Dilute to 500.0 mL with
dilute to 1000.0 mL with the same solvent. water R.
Guanidine-tris(hydroxymethyl)aminomethane-EDTA Diethanolamine buffer solution pH 10.0. 4007500.
buffer solution pH 8.6. 4016500. Dissolve 96.4 g of diethanolamine R in water R and dilute to
Dissolve 0.018 g of sodium edetate R, 2.2 g of 400 mL with the same solvent. Add 0.5 mL of an 186 g/L
tris(hydroxymethyl)aminomethane R and 28.7 g of solution of magnesium chloride R and adjust the pH with 1 M
guanidine hydrochloride R in 20 mL of water R. Adjust to hydrochloric acid. Dilute to 500.0 mL with water R.
pH 8.6 with acetic acid R and dilute to 50 mL with water R.
0.1 M Ammonium carbonate buffer solution pH 10.3.
1.5 M Tris-hydrochloride buffer solution pH 8.8. 4009900. 4011900.
Dissolve 90.8 g of tris(hydroxymethyl)aminomethane R in Dissolve 7.91 g of ammonium carbonate R in 800 mL
400 mL of water R. Adjust the pH with hydrochloric acid R of water R. Adjust the pH with dilute sodium hydroxide
and dilute to 500.0 mL with water R. solution R. Dilute to 1000.0 mL with water R.
3 M Tris-hydrochloride buffer solution pH 8.8. 4015000. Ammonium chloride buffer solution pH 10.4. 4011000.
Dissolve 363.3 g of tris(hydroxymethyl)aminomethane R in Dissolve 70 g of ammonium chloride R in 200 mL of water R,
500 mL of water R. Adjust the pH with hydrochloric acid R add 330 mL of concentrated ammonia R and dilute to
and dilute to 1 L with water R. 1000.0 mL with water R. If necessary, adjust to pH 10.4 with
ammonia R.
Buffer (phosphate) solution pH 9.0. 4008300.
Dissolve 1.74 g of potassium dihydrogen phosphate R in 80 mL Borate buffer solution pH 10.4. 4011100.
of water R, adjust the pH with a 1 M potassium hydroxide Dissolve 24.64 g of boric acid R in 900 mL of distilled water R.
solution prepared from potassium hydroxide R and dilute to Adjust the pH using a 400 g/L solution of sodium hydroxide R.
100.0 mL with water R. Dilute to 1000 mL with distilled water R.

Ammonium chloride buffer solution pH 10.7. 4013400. (solution A). Dissolve 132 g of ammonium phosphate R
Dissolve 67.5 g of ammonium chloride R in water R, add in distilled water R and dilute to 1000.0 mL with the
570 mL of concentrated ammonia R and dilute to 1000.0 mL same solvent (solution B). To a suspension of 292 g of
with water R. (ethylenedinitrilo)tetra-acetic acid R in about 500 mL
of distilled water R, add about 200 mL of concentrated
Buffer solution pH 10.9. 4007600. ammonia R to dissolve. Adjust the pH to 6 to 7 with
Dissolve 6.75 g of ammonium chloride R in ammonia R and concentrated ammonia R. Dilute to 1000.0 mL with distilled
dilute to 100.0 mL with the same solvent. water R (solution C). Mix equal volumes of solution A, B,
and C and adjust to pH 7.5 with concentrated ammonia R.
Total-ionic-strength-adjustment buffer. 4007700.
Dissolve 58.5 g of sodium chloride R, 57.0 mL of glacial Buffer solution pH 11. 4014000.
acetic acid R, 61.5 g of sodium acetate R and 5.0 g of
Dissolve 6.21 g of boric acid R, 4.00 g of sodium hydroxide R
cyclohexylenedinitrilotetra-acetic acid R in water R and dilute
and 3.70 g of potassium chloride R in 500 mL of water R and
to 500.0 mL with the same solvent. Adjust to pH 5.0 to 5.5
dilute to 1000 mL with the same solvent.
with a 335 g/L solution of sodium hydroxide R and dilute to
1000.0 mL with distilled water R. 0.1 M Phosphate buffer solution pH 11.3. 4015400.
Total-ionic-strength-adjustment buffer R1. 4008800. Dissolve 17.4 g of dipotassium hydrogen phosphate R in about
Dissolve 210 g of citric acid monohydrate R in 400 mL 950 mL of water R, adjust to pH 11.3 using a 100 g/L solution
of distilled water R. Adjust to pH 7.0 with concentrated of potassium hydroxide R and dilute to 1.0 L with water R.
ammonia R. Dilute to 1000.0 mL with distilled water R Filter through a membrane filter (nominal pore size 0.45 µm).

EUROPEAN PHARMACOPOEIA 4.2. Volumetric analysis
4.2. VOLUMETRIC ANALYSIS

EUROPEAN PHARMACOPOEIA 4.2.1. Primary standards for volumetric solutions
07/2017:40201 Potassium bromate. KBrO3. (Mr 167.0). 2000300.

[7758-01-2].
Crystallise potassium bromate R from boiling water R. Collect
the crystals and dry to constant mass at 180 °C.
4.2.1. PRIMARY STANDARDS FOR Potassium hydrogen phthalate. C8H5KO4. (Mr 204.2).

2000400. [877-24-7]. Potassium 2-carboxybenzoate.
VOLUMETRIC SOLUTIONS
Recrystallise potassium hydrogen phthalate R from boiling
Primary standards for volumetric solutions are indicated water R, collect the crystals at a temperature above 35 °C and
by the suffix RV. Primary standards of suitable quality may dry to constant mass at 110 °C.
be obtained from commercial sources or prepared by the
following methods. Sodium chloride. NaCl. (Mr 58.44). 2000600. [7647-14-5].
For primary standards from commercial sources, a To 1 volume of the saturated sodium chloride solution R add
pre-treatment step may be necessary. Follow the supplier’s 2 volumes of hydrochloric acid R. Collect the crystals formed
instructions. and wash with hydrochloric acid R1. Remove the hydrochloric
A secondary standard may be used provided its traceability to acid by heating on a water-bath and dry the crystals to
a primary standard has been demonstrated. constant mass at 300 °C.
Arsenious trioxide. As2O3. (Mr 197.8). 2000100. [1327-53-3]. Sulfanilic acid. C6H7NO3S. (Mr 173.2). 2000700. [121-57-3].
Arsenic(III) oxide. 4-Aminobenzenesulfonic acid.
Sublime arsenious trioxide R in a suitable apparatus. Recrystallise sulfanilic acid R from boiling water R. Filter and
Storage : over anhydrous silica gel R. dry to constant mass at 100-105 °C.
Benzoic acid. C7H6O2. (Mr 122.1). 2000200. [65-85-0].
Trometamol. C4H11NO3. (Mr 121.1). 2001000.
Sublime benzoic acid R in a suitable apparatus. [77-86-1]. 2-Amino-2-(hydroxymethyl)propane-1,3-diol.
Ferrous ethylenediammonium sulfate. Tris(hydroxymethyl)aminomethane.
Fe(C2H10N2)(SO4)2,4H2O. (Mr 382.1). 2000900. [113193-60-5]. Content : minimum 99.5 per cent.
Ethylenediammonium iron(II) disulfate tetrahydrate.
Ethylenediammonium tetraaquabis(sulfato)iron(II). Zinc. Zn. (Mr 65.4). 2000800. [7440-66-6].
Content : minimum 99.5 per cent. Content : minimum 99.9 per cent.

EUROPEAN PHARMACOPOEIA 4.2.2. Volumetric solutions
07/2019:40202 Dilution. Use a diluted solution of sulfuric acid R (59 g/L

H2SO4) while cooling the solution.
0.1 M Ammonium thiocyanate. 3000500.
Dissolve 7.612 g of ammonium thiocyanate R in water R and
4.2.2. VOLUMETRIC SOLUTIONS dilute to 1000.0 mL with the same solvent.
Standardisation. To 20.0 mL of 0.1 M silver nitrate add 25 mL
Volumetric solutions are prepared according to the usual of water R, 2 mL of dilute nitric acid R and 2 mL of ferric
chemical analytical methods. The accuracy of the apparatus ammonium sulfate solution R2. Titrate with the ammonium
used is verified to ensure that it is appropriate for the intended thiocyanate solution until a reddish-yellow colour is obtained.
use.
The concentration of volumetric solutions is indicated in terms 0.1 M Barium chloride. 3000600.
of molarity. Molarity expresses, as the number of moles, the Dissolve 24.4 g of barium chloride R in water R and dilute to
amount of substance dissolved in 1 L of solution. A solution 1000.0 mL with the same solvent.
which contains x moles of substance per litre is said to be x M. Standardisation. To 10.0 mL of the barium chloride solution
Volumetric solutions do not differ from the prescribed add 60 mL of water R, 3 mL of concentrated ammonia R and
strength by more than 10 per cent. The molarity of the 0.5-1 mg of phthalein purple R. Titrate with 0.1 M sodium
volumetric solutions is determined by an appropriate number edetate. When the solution begins to decolorise, add 50 mL
of titrations. The repeatability does not exceed 0.2 per cent of ethanol (96 per cent) R and continue the titration until the
(relative standard deviation). blue-violet colour disappears.
Volumetric solutions are standardised by the methods
described below. When a volumetric solution is to be used 0.05 M Barium perchlorate. 3000700.
in an assay in which the end-point is determined by an Dissolve 15.8 g of barium hydroxide R in a mixture of 7.5 mL
electrochemical process (for example, amperometry or of perchloric acid R and 75 mL of water R, adjust the solution
potentiometry) the solution is standardised by the same to pH 3 by adding perchloric acid R and filter if necessary. Add
method. The composition of the medium in which a 150 mL of ethanol (96 per cent) R and dilute to 250 mL with
volumetric solution is standardised should be the same as that water R. Dilute to 1000.0 mL with buffer solution pH 3.7 R.
in which it is to be used. Standardisation. To 5.0 mL of 0.05 M sulfuric acid add 5 mL
Solutions more dilute than those described below are either of water R, 50 mL of buffer solution pH 3.7 R and 0.5 mL of
prepared by adapting the quantities stated or by dilution, with alizarin S solution R. Titrate with the barium perchlorate
carbon dioxide-free water R (unless otherwise prescribed), solution until an orange-red colour appears. Standardise
of a more concentrated solution that has been previously immediately before use.
standardised. In the first case, the correction factor is Dilution. Use buffer solution pH 3.7 R.
determined on the volumetric solution to be used in the
monograph. In the latter case, the correction factor of the 0.005 M Barium perchlorate. 3010200.
dilute solution is the same as that of the standardised solution
from which it was prepared. Dilute 10.0 mL of 0.05 M barium perchlorate to 100.0 mL with
a buffer solution prepared as follows : to 15.0 mL of acetic
Commercially available volumetric solutions traceable to acid R add 60.0 mL of 2-propanol R. Adjust to pH 3.7 with
a primary standard may be used provided their titre is ammonia R and dilute to 100.0 mL with water R.
determined or verified prior to first use.
Titres of volumetric solutions are verified at appropriate 0.004 M Benzethonium chloride. 3000900.
intervals that are defined in the quality system procedures. Dissolve in water R 1.792 g of benzethonium chloride R,
previously dried to constant mass at 100-105 °C, and dilute to
0.1 M Ammonium and cerium nitrate. 3000100. 1000.0 mL with the same solvent.
Shake for 2 min a solution containing 56 mL of sulfuric acid R Standardisation. Dissolve 0.350 g of the dried substance in
and 54.82 g of ammonium and cerium nitrate R, then add 5 35 mL of a mixture of 30 volumes of anhydrous acetic acid R
successive quantities, each of 100 mL, of water R, shaking after and 70 volumes of acetic anhydride R. Titrate with 0.1 M
each addition. Dilute the clear solution to 1000.0 mL with perchloric acid, using 0.05 mL of crystal violet solution R as
water R. Standardise the solution after 10 days. indicator. Carry out a blank titration.
Standardisation. Dissolve 0.300 g of ferrous ethylenediam- 1 mL of 0.1 M perchloric acid is equivalent to 44.81 mg of
monium sulfate RV in 50 mL of a diluted solution of sulfuric C27H42ClNO2.
acid R (49 g/L H2SO4). Titrate with the ammonium and cerium
nitrate solution, determining the end-point potentiometrically 0.01 M Bismuth nitrate. 3010000.
(2.2.20) or using 0.1 mL of ferroin R as indicator.
Dissolve 4.86 g of bismuth nitrate pentahydrate R in 60 mL of
1 mL of 0.1 M ammonium and cerium nitrate is equivalent to dilute nitric acid R and dilute to 1000.0 mL with water R.
38.21 mg of Fe(C2H10N2)(SO4)2,4H2O.
Standardisation. To 25.0 mL of the bismuth nitrate solution,
Storage : protected from light. add 50 mL of water R and titrate with 0.01 M sodium edetate
0.1 M Ammonium and cerium sulfate. 3000300. using 0.05 mL of a 1 g/L solution of xylenol orange R as
indicator.
Dissolve 65.0 g of ammonium and cerium sulfate R in a
mixture of 500 mL of water R and 30 mL of sulfuric acid R. 0.0167 M Bromide-bromate. 3001000.
Allow to cool and dilute to 1000.0 mL with water R. Dissolve 2.7835 g of potassium bromate RV and 13 g of
Standardisation. Dissolve 0.300 g of ferrous ethylenediam- potassium bromide R in water R and dilute to 1000.0 mL with
monium sulfate RV in 50 mL of a diluted solution of sulfuric the same solvent.
acid R (49 g/L H2SO4). Titrate with the ammonium and cerium
sulfate solution, determining the end-point potentiometrically 0.1 M Cerium sulfate. 3001100.
(2.2.20) or using 0.1 mL of ferroin R as indicator. Dissolve 40.4 g of cerium sulfate R in a mixture of 500 mL of
1 mL of 0.1 M ammonium and cerium sulfate is equivalent to water R and 50 mL of sulfuric acid R. Allow to cool and dilute
38.21 mg of Fe(C2H10N2)(SO4)2,4H2O. to 1000.0 mL with water R.

4.2.2. Volumetric solutions EUROPEAN PHARMACOPOEIA
Standardisation. Dissolve 0.300 g of ferrous ethylenediam- Standardisation. To 10.0 mL of the iodine solution add
monium sulfate RV in 50 mL of a diluted solution of sulfuric 1 mL of dilute acetic acid R and 40 mL of water R. Titrate
acid R (49 g/L H2SO4). Titrate with the cerium sulfate solution, with 0.1 M sodium thiosulfate, determining the end-point
determining the end-point potentiometrically (2.2.20) or potentiometrically (2.2.20) or using starch solution R as
using 0.1 mL of ferroin R as indicator. indicator.
1 mL of 0.1 M cerium sulfate is equivalent to 38.21 mg of Storage : protected from light.
Fe(C2H10N2)(SO4)2,4H2O.
0.01 M Iodine. 3002900.
0.02 M Copper sulfate. 3001200. Add 0.3 g of potassium iodide R to 20.0 mL of 0.05 M iodine
Dissolve 5.0 g of copper sulfate pentahydrate R in water R and and dilute to 100.0 mL with water R.
dilute to 1000.0 mL with the same solvent.
0.1 M Lanthanum nitrate. 3010100.
Standardisation. To 20.0 mL of the copper sulfate solution
add 2 g of sodium acetate R and 0.1 mL of pyridylazonaphthol Dissolve 43.30 g of lanthanum nitrate R in water R and dilute
solution R. Titrate with 0.02 M sodium edetate until the colour to 1000.0 mL with the same solvent.
changes from violet-blue to bright green. Titrate slowly Standardisation. To 20.0 mL of the lanthanum nitrate solution,
towards the end of the titration. add 15 mL of water R and 25 mL of 0.1 M sodium edetate.
Add about 50 mg of xylenol orange triturate R and about 2 g
0.1 M Ferric ammonium sulfate. 3001300. of hexamethylenetetramine R. Titrate with 0.1 M zinc sulfate
Dissolve 50.0 g of ferric ammonium sulfate R in a mixture of until the colour changes from yellow to violet-pink.
6 mL of sulfuric acid R and 300 mL of water R and dilute to 1 mL of 0.1 M sodium edetate is equivalent to 43.30 mg of
1000.0 mL with water R. La(NO3)3,6H2O.
Standardisation. To 10.0 mL of the ferric ammonium sulfate 0.1 M Lead nitrate. 3003100.
solution add 35 mL of water R, 3 mL of hydrochloric acid R
and 1 g of potassium iodide R. Allow to stand for 10 min. Dissolve 33 g of lead nitrate R in water R and dilute to
Titrate with 0.1 M sodium thiosulfate, determining the 1000.0 mL with the same solvent.
end-point potentiometrically (2.2.20) or using 1 mL of starch Standardisation. Take 20.0 mL of the lead nitrate solution
solution R as indicator. and carry out the determination of lead by complexometry
1 mL of 0.1 M sodium thiosulfate is equivalent to 48.22 mg (2.5.11).
of FeNH4(SO4)2,12H2O. 0.1 M Lithium methoxide. 3003300.
0.1 M Ferrous sulfate. 3001400. Dissolve 0.694 g of lithium R in 150 mL of anhydrous
methanol R and dilute to 1000.0 mL with toluene R.
Dissolve 27.80 g of ferrous sulfate R in 500 mL of dilute sulfuric
acid R and dilute to 1000.0 mL with water R. Standardisation. To 10 mL of dimethylformamide R add
0.05 mL of a 3 g/L solution of thymol blue R in methanol R
Standardisation. To 25.0 mL of the ferrous sulfate solution add and titrate with the lithium methoxide solution until a
3 mL of phosphoric acid R and titrate immediately with 0.02 M pure blue colour is obtained. Immediately add 0.100 g of
potassium permanganate. Standardise immediately before use. benzoic acid RV. Stir to effect solution and titrate with the
1 M Hydrochloric acid. 3001800. lithium methoxide solution until the pure blue colour is again
obtained. Protect the solution from atmospheric carbon
Dilute 103.0 g of hydrochloric acid R to 1000.0 mL with dioxide throughout the titration. From the volume of titrant
water R. used in the second titration ascertain the exact strength of the
Standardisation. Dissolve 0.950 g of trometamol RV in 50 mL lithium methoxide solution. Standardise immediately before
of water R. Titrate with the hydrochloric acid solution, use.
determining the end-point potentiometrically (2.2.20) or 1 mL of 0.1 M lithium methoxide is equivalent to 12.21 mg
using 0.1 mL of methyl orange solution R as indicator until a of C7H6O2.
yellowish-red colour is obtained.
1 mL of 1 M hydrochloric acid is equivalent to 121.1 mg of 0.1 M Magnesium chloride. 3003400.
C4H11NO3. Dissolve 20.33 g of magnesium chloride R in water R and dilute
to 1000.0 mL with the same solvent.
0.1 M Hydrochloric acid. 3002100. Standardisation. Carry out the determination of magnesium
Dilute 100.0 mL of 1 M hydrochloric acid to 1000.0 mL with by complexometry (2.5.11).
carbon dioxide-free water R.
1 M Nitric acid. 3003600.
Standardisation. Carry out the titration described for 1 M
hydrochloric acid using 95 mg of trometamol RV dissolved in Dilute 96.6 g of nitric acid R to 1000.0 mL with water R.
50 mL of water R. Standardisation. Dissolve 0.950 g of trometamol RV in 50 mL
1 mL of 0.1 M hydrochloric acid is equivalent to 12.11 mg of of water R. Titrate with the nitric acid solution, determining
C4H11NO3. the end-point potentiometrically (2.2.20) or using 0.1 mL of
methyl orange solution R as indicator until a reddish-yellow
0.5 M Iodine. 3009400. colour is obtained.
Dissolve 127 g of iodine R and 200 g of potassium iodide R in 1 mL of 1 M nitric acid is equivalent to 121.1 mg of C4H11NO3.
water R and dilute to 1000.0 mL with the same solvent.
0.1 M Perchloric acid. 3003900.
Standardisation. To 2.0 mL of the iodine solution add 1 mL of
Place 8.5 mL of perchloric acid R in a volumetric flask
dilute acetic acid R and 50 mL of water R. Titrate with 0.1 M
containing about 900 mL of glacial acetic acid R and mix. Add
sodium thiosulfate, using starch solution R as indicator.
30 mL of acetic anhydride R, dilute to 1000.0 mL with glacial
Storage : protected from light. acetic acid R, mix and allow to stand for 24 h. Determine the
water content (2.5.12) without addition of methanol and,
0.05 M Iodine. 3002700. if necessary, adjust the water content to 0.1-0.2 per cent by
Dissolve 12.7 g of iodine R and 20 g of potassium iodide R in adding either acetic anhydride R or water R. Allow to stand
water R and dilute to 1000.0 mL with the same solvent. for 24 h.

Standardisation. Dissolve 0.170 g of potassium hydrogen Standardisation. To 3.0 mL of the potassium iodate solution
phthalate RV in 50 mL of anhydrous acetic acid R, warming add 40.0 mL of water R, 1 g of potassium iodide R and 5 mL
gently if necessary. Allow to cool protected from air, and of dilute sulfuric acid R. Titrate with 0.1 M sodium thiosulfate,
titrate with the perchloric acid solution, determining the determining the end-point potentiometrically (2.2.20) or
end-point potentiometrically (2.2.20) or using 0.05 mL of using 1 mL of starch solution R, added towards the end of the
crystal violet solution R as indicator. Note the temperature of titration, as indicator.
the perchloric acid solution at the time of the titration. If the 1 mL of 0.1 M sodium thiosulfate is equivalent to 3.567 mg of
temperature at which an assay is carried out is different from KIO3.
that at which the 0.1 M perchloric acid has been standardised,
the volume used in the assay becomes : 0.001 M Potassium iodide. 3009200.
Dilute 10.0 mL of potassium iodide solution R to 100.0 mL
Vc = V [1 + (t1 - t2 )0.0011] with water R. Dilute 5.0 mL of this solution to 500.0 mL with
water R.
t1 = temperature during standardisation,
t2 = temperature during the assay, 0.02 M Potassium permanganate. 3005300.
Dissolve 3.2 g of potassium permanganate R in water R and
Vc = corrected volume, dilute to 1000.0 mL with the same solvent. Heat the solution
V = observed volume. for 1 h on a water-bath, allow to cool and filter through a
sintered-glass filter (2.1.2).
1 mL of 0.1 M perchloric acid is equivalent to 20.42 mg Standardisation. Dissolve 0.300 g of ferrous ethylenediammo-
of C8H5KO4. nium sulfate RV in 50 mL of a diluted solution of sulfuric acid R
Dilution. Use anhydrous acetic acid R. (49 g/L H2SO4). Titrate with the potassium permanganate
solution, determining the end-point potentiometrically
0.033 M Potassium bromate. 3004200.
(2.2.20) or by the colour of the solution changing to pink.
Dissolve 5.5670 g of potassium bromate RV in water R and Standardise immediately before use.
1 mL of 0.02 M potassium permanganate is equivalent to
0.1 M Potassium hydrogen phthalate. 3004700. 38.21 mg of Fe(C2H10N2)(SO4)2,4H2O.
In a conical flask containing about 800 mL of anhydrous acetic Storage : protected from light.
acid R, dissolve 20.42 g of potassium hydrogen phthalate RV. 0.1 M Silver nitrate. 3005600.
Heat on a water-bath until completely dissolved, protected
from humidity. Cool to 20 °C and dilute to 1000.0 mL with Dissolve 17.0 g of silver nitrate R in water R and dilute to
anhydrous acetic acid R. 1000.0 mL with the same solvent.
Standardisation. Dissolve 50 mg of sodium chloride RV
0.1 M Potassium hydroxide. 3004800. in water R, add 5 mL of dilute nitric acid R and dilute to
Dissolve 6 g of potassium hydroxide R in carbon dioxide-free 50 mL with water R. Titrate with the silver nitrate solution,
water R and dilute to 1000.0 mL with the same solvent. determining the end-point potentiometrically (2.2.20).
Standardisation. Dissolve 0.150 g of potassium hydrogen 1 mL of 0.1 M silver nitrate is equivalent to 5.844 mg of NaCl.
phthalate RV in 50 mL of water R. Titrate with the Storage : protected from light.
potassium hydroxide solution, determining the end-point
potentiometrically (2.2.20) or using 0.1 mL of phenolphthalein 0.1 M Sodium arsenite. 3005800.
solution R as indicator. Dissolve arsenious trioxide RV equivalent to 4.946 g of As2O3
1 mL of 0.1 M potassium hydroxide is equivalent to 20.42 mg in a mixture of 20 mL of strong sodium hydroxide solution R
of C8H5KO4. and 20 mL of water R, dilute to 400 mL with water R and add
dilute hydrochloric acid R until the solution is neutral to blue
0.5 M Potassium hydroxide in alcohol (60 per cent V/V). litmus paper R. Dissolve 2 g of sodium hydrogen carbonate R
3004900. in the solution and dilute to 500.0 mL with water R.
Dissolve 3 g of potassium hydroxide R in aldehyde-free
0.1 M Sodium edetate. 3005900.
alcohol R (60 per cent V/V) and dilute to 100.0 mL with the
same solvent. Dissolve 37.5 g of sodium edetate R in 500 mL of water R, add
100 mL of 1 M sodium hydroxide and dilute to 1000.0 mL
Standardisation. Dissolve 0.500 g of benzoic acid RV in 10 mL
with water R.
of water R and 40 mL of ethanol (96 per cent) R. Titrate with
the potassium hydroxide solution, determining the end-point Standardisation. Dissolve 0.120 g of zinc RV in 4 mL of
potentiometrically (2.2.20) or using 0.1 mL of phenolphthalein hydrochloric acid R1. Add dilute sodium hydroxide solution R
solution R as indicator. until the solution is weakly acid and carry out the assay of zinc
by complexometry (2.5.11).
1 mL of 0.5 M potassium hydroxide in alcohol (60 per cent V/V)
is equivalent to 61.06 mg of C7H6O2. 1 mL of 0.1 M sodium edetate is equivalent to 6.538 mg of Zn.
Storage : in a polyethylene container.
0.5 M Potassium hydroxide, alcoholic. 3005000.
Dissolve 3 g of potassium hydroxide R in 5 mL of water R and 1 M Sodium hydroxide. 3006300.
dilute to 100.0 mL with aldehyde-free alcohol R. Dissolve 42 g of sodium hydroxide R in carbon dioxide-free
Standardisation. Dissolve 0.500 g of benzoic acid RV in 10 mL water R and dilute to 1000.0 mL with the same solvent.
of water R and 40 mL of ethanol (96 per cent) R. Titrate with Standardisation. Dissolve 1.50 g of potassium hydrogen
the potassium hydroxide solution, determining the end-point phthalate RV in 50 mL of water R. Titrate with the
potentiometrically (2.2.20) or using 0.1 mL of phenolphthalein sodium hydroxide solution, determining the end-point
solution R as indicator. potentiometrically (2.2.20) or using 0.1 mL of phenolphthalein
1 mL of 0.5 M alcoholic potassium hydroxide is equivalent to solution R as indicator.
61.06 mg of C7H6O2. 1 mL of 1 M sodium hydroxide is equivalent to 204.2 mg of
Dilution. Use aldehyde-free alcohol R. C 8H5KO4.
If sodium hydroxide free from carbonate is prescribed, prepare
0.05 M Potassium iodate. 3005200. it as follows. Dissolve sodium hydroxide R in water R to give a
Dissolve 10.70 g of potassium iodate R in water R and dilute to concentration of 400-600 g/L and allow to stand. Decant the
1000.0 mL with the same solvent. clear supernatant, taking precautions to avoid the introduction

4.2.2. Volumetric solutions EUROPEAN PHARMACOPOEIA
of carbon dioxide, and dilute with carbon dioxide-free water R 0.1 M Sodium periodate. 3009500.
to the required molarity. The solution complies with the Dissolve 21.4 g of sodium periodate R in about 500 mL of
following test. Titrate 20.0 mL of hydrochloric acid of the water R and dilute to 1000.0 mL with the same solvent.
same molarity with the solution of sodium hydroxide, using
0.1 mL of phenolphthalein solution R as indicator. At the Standardisation. In a stoppered flask, introduce 5.0 mL of the
end-point add just sufficient of the acid to discharge the pink sodium periodate solution and add 100 mL of water R. Add
colour and concentrate the solution to 20 mL by boiling. 10 mL of potassium iodide solution R and 5 mL of hydrochloric
During boiling add just sufficient acid to discharge the pink acid R1, close, shake and allow to stand for 2 min. Titrate
colour, which should not reappear after prolonged boiling. with 0.1 M sodium thiosulfate until the yellow colour almost
The volume of acid used does not exceed 0.1 mL. disappears. Determine the end-point potentiometrically
(2.2.20) or add 2 mL of starch solution R and titrate slowly
0.1 M Sodium hydroxide. 3006600. until the colour is completely discharged.
Dilute 100.0 mL of 1 M sodium hydroxide to 1000.0 mL with 1 mL of 0.1 M sodium thiosulfate is equivalent to 2.674 mg of
carbon dioxide-free water R. NaIO4 or 0.125 mL of 0.1 M sodium periodate.
Standardisation. Carry out the titration described for 1 M
sodium hydroxide using 0.150 g of potassium hydrogen 0.1 M Sodium thiosulfate. 3007300.
phthalate RV in 50 mL of water R. Dissolve 25 g of sodium thiosulfate R and 0.2 g of sodium
1 mL of 0.1 M sodium hydroxide is equivalent to 20.42 mg of carbonate R in carbon dioxide-free water R and dilute to
C8H5KO4. 1000.0 mL with the same solvent.
Standardisation (for use in the assay of halide salts of organic Standardisation. To 10.0 mL of 0.033 M potassium bromate,
bases). Dissolve 0.100 g of benzoic acid RV in a mixture of add 40 mL of water R, 10 mL of potassium iodide solution R
5 mL of 0.01 M hydrochloric acid and 50 mL of ethanol (96 per and 5 mL of hydrochloric acid R1. Titrate with the sodium
cent) R. Carry out the titration (2.2.20), using the sodium thiosulfate solution, using 1 mL of starch solution R, added
hydroxide solution. Note the volume added between the towards the end of the titration, as indicator.
2 points of inflexion. 1 mL of 0.1 M sodium thiosulfate is equivalent to 2.783 mg of
1 mL of 0.1 M sodium hydroxide is equivalent to 12.21 mg of KBrO3 or 0.5 mL of 0.033 M potassium bromate.
C 7H 6O 2.
0.5 M Sulfuric acid. 3007800.
0.1 M Sodium hydroxide, ethanolic. 3007000. Dissolve 28 mL of sulfuric acid R in water R and dilute to
To 250 mL of anhydrous ethanol R add 3.3 g of strong sodium 1000.0 mL with the same solvent.
hydroxide solution R. Standardisation. Dissolve 0.950 g of trometamol RV in 50 mL
Standardisation. Dissolve 0.100 g of benzoic acid RV in 10 mL of water R. Titrate with the sulfuric acid solution, determining
of water R and 40 mL of ethanol (96 per cent) R. Titrate the end-point potentiometrically (2.2.20) or using 0.1 mL of
with the ethanolic sodium hydroxide solution, determining methyl orange solution R as indicator until the solution turns
the end-point potentiometrically (2.2.20) or using 0.2 mL reddish-yellow.
of thymolphthalein solution R as indicator. Standardise 1 mL of 0.5 M sulfuric acid is equivalent to 121.1 mg of
immediately before use. C4H11NO3.
1 mL of 0.1 M ethanolic sodium hydroxide is equivalent to
12.21 mg of C7H6O2. 0.1 M Tetrabutylammonium hydroxide. 3008300.
0.1 M Sodium methoxide. 3007100. Dissolve 40 g of tetrabutylammonium iodide R in 90 mL of
anhydrous methanol R, add 20 g of finely powdered silver
Cool 175 mL of anhydrous methanol R in iced water R and oxide R and shake vigorously for 1 h. Centrifuge a few
add, in small portions, about 2.5 g of freshly cut sodium R. millilitres of the mixture and test the supernatant for iodides.
When the metal has dissolved, dilute to 1000.0 mL with If a positive reaction is obtained, add an additional 2 g of silver
toluene R. oxide R and shake for a further 30 min. Repeat this procedure
Standardisation. To 10 mL of dimethylformamide R add until the liquid is free from iodides, filter the mixture through
0.05 mL of a 3 g/L solution of thymol blue R in methanol R, a fine sintered-glass filter (2.1.2) and rinse the reaction vessel
and titrate with the sodium methoxide solution until a and filter with three quantities, each of 50 mL, of toluene R.
pure blue colour is obtained. Immediately add 0.100 g of Add the washings to the filtrate and dilute to 1000.0 mL with
benzoic acid RV. Stir until dissolution and titrate with the toluene R. Pass dry carbon dioxide-free nitrogen through the
sodium methoxide solution until the pure blue colour is again solution for 5 min.
obtained. Protect the solution from atmospheric carbon Standardisation. To 10 mL of dimethylformamide R add
dioxide throughout the titration. From the volume of titrant 0.05 mL of a 3 g/L solution of thymol blue R in methanol R
used in the second titration ascertain the exact strength of the and titrate with the tetrabutylammonium hydroxide solution
sodium methoxide solution. Standardise immediately before until a pure blue colour is obtained. Immediately add 0.100 g
use. of benzoic acid RV. Stir to effect solution, and titrate with
1 mL of 0.1 M sodium methoxide is equivalent to 12.21 mg the tetrabutylammonium hydroxide solution until the pure
of C7H6O2. blue colour is again obtained. Protect the solution from
atmospheric carbon dioxide throughout the titration. From
0.1 M Sodium nitrite. 3007200. the volume of titrant used in the second titration ascertain the
Dissolve 7.5 g of sodium nitrite R in water R and dilute to exact strength of the tetrabutylammonium hydroxide solution.
1000.0 mL with the same solvent. Standardise immediately before use.
Standardisation. Dissolve 0.150 g of sulfanilic acid RV 1 mL of 0.1 M tetrabutylammonium hydroxide is equivalent
in 50 mL of dilute hydrochloric acid R and carry out the to 12.21 mg of C7H6O2.
determination of primary aromatic amino-nitrogen (2.5.8),
using the sodium nitrite solution and determining the 0.1 M Tetrabutylammonium hydroxide in 2-propanol.
end-point electrometrically. Standardise immediately before 3008400.
use. Prepare as described for 0.1 M tetrabutylammonium hydroxide
1 mL of 0.1 M sodium nitrite is equivalent to 17.32 mg of using 2-propanol R instead of toluene R and standardise as
C6H7NO3S. described.

0.05 M Zinc chloride. 3008500. 0.1 M Zinc sulfate. 3008600.

Dissolve 6.82 g of zinc chloride R, weighed with appropriate
precautions, in water R. If necessary, add dropwise dilute Dissolve 29 g of zinc sulfate R in water R and dilute to
hydrochloric acid R until the opalescence disappears. Dilute to 1000.0 mL with the same solvent.
Standardisation. To 20.0 mL of the zinc chloride solution add Standardisation. To 20.0 mL of the zinc sulfate solution add
5 mL of dilute acetic acid R and carry out the determination of 5 mL of dilute acetic acid R and carry out the determination of
zinc by complexometry (2.5.11). zinc by complexometry (2.5.11).

EUROPEAN PHARMACOPOEIA 5.1.4. Microbiological quality of non-sterile products for pharmaceutical use
04/2019:50104 – 101 CFU : maximum acceptable count = 20 ;

– 102 CFU : maximum acceptable count = 200 ;
– 103 CFU : maximum acceptable count = 2000, and so forth.
Table 5.1.4.-1 includes a list of specified micro-organisms for
which acceptance criteria are set. The list is not necessarily
5.1.4. MICROBIOLOGICAL QUALITY exhaustive and for a given preparation it may be necessary to
OF NON-STERILE PHARMACEUTICAL test for other micro-organisms depending on the nature of the
starting materials and the manufacturing process.
PREPARATIONS AND SUBSTANCES
If it has been shown that none of the prescribed tests will
FOR PHARMACEUTICAL USE(1) allow valid enumeration of micro-organisms at the level
prescribed, a validated method with a limit of detection as
◊This chapter does not apply to products containing viable close as possible to the indicated acceptance criterion is used.
micro-organisms as active ingredients.◊
In addition to the micro-organisms listed in Table 5.1.4.-1, the
The presence of certain micro-organisms in non-sterile significance of other micro-organisms recovered is evaluated
preparations may have the potential to reduce or even in terms of :
inactivate the therapeutic activity of the product and has
a potential to adversely affect the health of the patient. – use of the product : hazard varies according to the route of
Manufacturers therefore have to ensure a low bioburden of administration (eye, nose, respiratory tract) ;
finished dosage forms by implementing current guidelines – nature of the product : its ability to support growth, the
on Good Manufacturing Practice during the manufacture, presence of adequate antimicrobial preservation ;
storage and distribution of pharmaceutical preparations. – method of application ;
Microbial examination of non-sterile products is performed – intended recipient : risk may differ for neonates, infants,
according to the methods given in general chapters 2.6.12 and the debilitated ;
2.6.13. Acceptance criteria for non-sterile pharmaceutical
products based upon the total aerobic microbial count (TAMC) – use of immunosuppressive agents, corticosteroids ;
and the total combined yeasts/moulds count (TYMC) are – presence of disease, wounds, organ damage.
given in Tables 5.1.4.-1 and 5.1.4.-2. Acceptance criteria are Where warranted, a risk-based assessment of the relevant
based on individual results or on the average of replicate factors is conducted by personnel with specialised training in
counts when replicate counts are performed (e.g. direct microbiology and the interpretation of microbiological data.
plating methods). For raw materials, the assessment takes account of processing
When an acceptance criterion for microbiological quality is to which the product is subjected, the current technology of
prescribed it is interpreted as follows: testing and the availability of materials of the desired quality.
Table 5.1.4.-1. – Acceptance criteria for microbiological quality of non-sterile dosage forms
TAMC TYMC
Route of administration (CFU/g or (CFU/g or Specified micro-organisms
CFU/mL) CFU/mL)
Non-aqueous preparations for oral use 103 102 Absence of Escherichia coli (1 g or 1 mL)
Aqueous preparations for oral use 10 2

10 1
Absence of Escherichia coli (1 g or 1 mL)
Rectal use 10 3
10 2 -
Oromucosal use
Gingival use
Absence of Staphylococcus aureus (1 g or 1 mL)
Cutaneous use 102 101
Absence of Pseudomonas aeruginosa (1 g or 1 mL)
Nasal use
Auricular use
Absence of Pseudomonas aeruginosa (1 g or 1 mL)
Vaginal use 102 101 Absence of Staphylococcus aureus (1 g or 1 mL)
Absence of Candida albicans (1 g or 1 mL)
Transdermal patches (limits for one patch Absence of Staphylococcus aureus (1 patch)
102 101
including adhesive layer and backing) Absence of Pseudomonas aeruginosa (1 patch)
Absence of Staphylococcus aureus (1 g or 1 mL)
Inhalation use (special requirements apply to Absence of Pseudomonas aeruginosa (1 g or 1 mL)
102 101
liquid preparations for nebulisation) Absence of bile-tolerant gram-negative
bacteria (1 g or 1 mL)
♦Special Ph. Eur. provision for oral dosage Not more than 102 CFU of bile-tolerant gram-negative
forms containing raw materials of natural bacteria (1 g or 1 mL)
(animal, vegetal or mineral) origin for which
antimicrobial pretreatment is not feasible and 104 102 Absence of Salmonella (10 g or 10 mL)
for which the competent authority accepts Absence of Escherichia coli (1 g or 1 mL)
TAMC of the raw material exceeding 103 CFU/g Absence of Staphylococcus aureus (1 g or 1 mL)♦
or CFU/mL.
♦Special Ph. Eur. provision for premixes for Not more than 104 CFU of bile-tolerant gram-negative
medicated feeding stuffs for veterinary use bacteria (1 g or 1 mL)
105 104
using excipients of plant origin for which Absence of Escherichia coli (1 g or 1 mL)
antimicrobial treatment is not feasible. Absence of Salmonella (25 g or 25 mL)♦

5.1.4. Microbiological quality of non-sterile products for pharmaceutical use EUROPEAN PHARMACOPOEIA
Table 5.1.4.-2. – Acceptance criteria for microbiological quality

of non-sterile substances for pharmaceutical use
TAMC TYMC
(CFU/g or CFU/mL) (CFU/g or CFU/mL)
Substances for 103 102

pharmaceutical use
♦Recommended acceptance criteria for microbiological quality

of herbal medicinal products for oral use and extracts used in
their preparation are given in general chapter 5.1.8.♦

EUROPEAN PHARMACOPOEIA 5.4. Residual solvents
07/2018:50400 IMPURITIES : GUIDELINES

FOR RESIDUAL SOLVENTS
(CHMP/ICH/82260/2006)
1. INTRODUCTION
5.4. RESIDUAL SOLVENTS 2. SCOPE OF THE GUIDELINE

3. GENERAL PRINCIPLES
3.1. CLASSIFICATION OF RESIDUAL SOLVENTS BY RISK
LIMITING RESIDUAL SOLVENT LEVELS ASSESSMENT
IN ACTIVE SUBSTANCES, EXCIPIENTS 3.2. METHODS FOR ESTABLISHING EXPOSURE LIMITS
AND MEDICINAL PRODUCTS 3.3. OPTIONS FOR DESCRIBING LIMITS OF CLASS 2
SOLVENTS
3.4. ANALYTICAL PROCEDURES
3.5. REPORTING LEVELS OF RESIDUAL SOLVENTS
4. LIMITS OF RESIDUAL SOLVENTS
The International Council for Harmonisation of Technical
Requirements for Pharmaceuticals for Human Use (ICH) has 4.1. SOLVENTS TO BE AVOIDED
adopted Impurities Guidelines for Residual Solvents which 4.2. SOLVENTS TO BE LIMITED
prescribes limits for the content of solvents which may remain 4.3. SOLVENTS WITH LOW TOXIC POTENTIAL
in active substances, excipients and medicinal products after
processing. This guideline, the text of which is reproduced 4.4. SOLVENTS FOR WHICH NO ADEQUATE
below, excludes existing marketed products. The European TOXICOLOGICAL DATA WAS FOUND
Pharmacopoeia is, however, applying the same principles GLOSSARY
enshrined in the guideline to existing active substances,
excipients and medicinal products whether or not they are the APPENDIX 1. LIST OF SOLVENTS INCLUDED IN THE
subject of a monograph of the Pharmacopoeia. All substances GUIDELINE
and products are to be tested for the content of solvents likely
to be present in a substance or product. APPENDIX 2. ADDITIONAL BACKGROUND
A2.1. ENVIRONMENTAL REGULATION OF ORGANIC
VOLATILE SOLVENTS
A2.2. RESIDUAL SOLVENTS IN PHARMACEUTICALS
APPENDIX 3. METHODS FOR ESTABLISHING EXPOSURE
Where the limits to be applied comply with those given below, LIMITS
tests for residual solvents are not generally mentioned in
specific monographs since the solvents employed may vary 1. INTRODUCTION
from one manufacturer to another and the requirements of The objective of this guideline is to recommend acceptable
this general chapter are applied via the general monograph amounts of residual solvents in pharmaceuticals for the safety
on Substances for Pharmaceutical Use (2034). The competent of the patient. The guideline recommends the use of less toxic
authority is to be informed of the solvents employed during solvents and describes levels considered to be toxicologically
the production process. This information is also given acceptable for some residual solvents.
in the dossier submitted for a certificate of suitability of Residual solvents in pharmaceuticals are defined here as
the monographs of the European Pharmacopoeia and is organic volatile chemicals that are used or produced in the
mentioned on the certificate. manufacture of active substances or excipients, or in the
preparation of medicinal products. The solvents are not
completely removed by practical manufacturing techniques.
Appropriate selection of the solvent for the synthesis of active
substance may enhance the yield, or determine characteristics
Where only Class 3 solvents are used, a test for loss on drying such as crystal form, purity, and solubility. Therefore, the
may be applied or a specific determination of the solvent may solvent may sometimes be a critical parameter in the synthetic
be made. If for a Class 3 solvent a justified and authorised limit process. This guideline does not address solvents deliberately
higher than 0.5 per cent is applied, a specific determination of used as excipients nor does it address solvates. However, the
the solvent is required. content of solvents in such products should be evaluated and
justified.
Since there is no therapeutic benefit from residual solvents, all
residual solvents should be removed to the extent possible to
When Class 1 residual solvents or Class 2 residual solvents
meet product specifications, good manufacturing practices,
(or Class 3 residual solvents which exceed the 0.5 per cent)
or other quality-based requirements. Medicinal products
are used, the methodology described in the general method
should contain no higher levels of residual solvents than can
(2.4.24) is to be applied wherever possible. Otherwise an
be supported by safety data. Some solvents that are known
appropriate validated method is to be employed.
to cause unacceptable toxicities (Class 1, Table 1) should be
avoided in the production of active substances, excipients, or
medicinal products unless their use can be strongly justified
in a risk-benefit assessment. Some solvents associated with
less severe toxicity (Class 2, Table 2) should be limited in
When a quantitative determination of a residual solvent is order to protect patients from potential adverse effects.
carried out, the result is taken into account for the calculation Ideally, less toxic solvents (Class 3, Table 3) should be used
of the content of the substance except where a test for drying where practical. The complete list of solvents included in this
is carried out. guideline is given in Appendix 1.

5.4. Residual solvents EUROPEAN PHARMACOPOEIA
The lists are not exhaustive and other solvents can be used Class 3 solvents : solvents with low toxic potential
and later added to the lists. Recommended limits of Class 1 Solvents with low toxic potential to man ; no health-based
and 2 solvents or classification of solvents may change as new exposure limit is needed. Class 3 solvents have PDEs of 50 mg
safety data becomes available. Supporting safety data in a or more per day.
marketing application for a new medicinal product containing
a new solvent may be based on concepts in this guideline 3.2. METHODS FOR ESTABLISHING EXPOSURE LIMITS
or the concept of qualification of impurities as expressed in The method used to establish permitted daily exposures for
the guideline for active substances (Q3A, Impurities in New residual solvents is presented in Appendix 3. Summaries of the
Active Substances) or medicinal products (Q3B, Impurities in toxicity data that were used to establish limits are published in
New Medicinal Products), or all three guidelines. Pharmeuropa, Vol. 9, No. 1, Supplement April 1997.
3.3. OPTIONS FOR DESCRIBING LIMITS OF CLASS 2
2. SCOPE OF THE GUIDELINE SOLVENTS
Residual solvents in active substances, excipients, and in Two options are available when setting limits for Class 2
medicinal products are within the scope of this guideline. solvents.
Therefore, testing should be performed for residual solvents Option 1 : the concentration limits in parts per million stated
when production or purification processes are known to result in Table 2 can be used. They were calculated using equation (1)
in the presence of such solvents. It is only necessary to test below by assuming a product mass of 10 g administered daily.
for solvents that are used or produced in the manufacture
or purification of active substances, excipients, or medicinal 1000 ´ PDE (1)
product. Although manufacturers may choose to test the Concentration (ppm) =
medicinal product, a cumulative method may be used to dose
calculate the residual solvent levels in the medicinal product Here, PDE is given in terms of mg/day and dose is given
from the levels in the ingredients used to produce the in g/day.
medicinal product. If the calculation results in a level equal to
or below that recommended in this guideline, no testing of the These limits are considered acceptable for all substances,
medicinal product for residual solvents need be considered. If excipients, or products. Therefore this option may be applied
however, the calculated level is above the recommended level, if the daily dose is not known or fixed. If all excipients and
the medicinal product should be tested to ascertain whether active substances in a formulation meet the limits given
the formulation process has reduced the relevant solvent level in Option 1, then these components may be used in any
to within the acceptable amount. Medicinal product should proportion. No further calculation is necessary provided
also be tested if a solvent is used during its manufacture. the daily dose does not exceed 10 g. Products that are
administered in doses greater than 10 g per day should be
This guideline does not apply to potential new active considered under Option 2.
substances, excipients, or medicinal products used during the
clinical research stages of development, nor does it apply to Option 2 : it is not considered necessary for each component
existing marketed medicinal products. of the medicinal product to comply with the limits given in
Option 1. The PDE in terms of mg/day as stated in Table 2 can
The guideline applies to all dosage forms and routes of be used with the known maximum daily dose and equation (1)
administration. Higher levels of residual solvents may be above to determine the concentration of residual solvent
acceptable in certain cases such as short term (30 days or less) allowed in a medicinal product. Such limits are considered
or topical application. Justification for these levels should be acceptable provided that is has been demonstrated that the
made on a case by case basis. residual solvent has been reduced to the practical minimum.
See Appendix 2 for additional background information related The limits should be realistic in relation to analytical
to residual solvents. precision, manufacturing capability, reasonable variation
in the manufacturing process, and the limits should reflect
contemporary manufacturing standards.
3. GENERAL PRINCIPLES
Option 2 may be applied by adding the amounts of a residual
3.1. CLASSIFICATION OF RESIDUAL SOLVENTS BY RISK solvent present in each of the components of the medicinal
ASSESSMENT product. The sum of the amounts of solvent per day should be
The term “tolerable daily intake” (TDI) is used by the less than that given by the PDE.
International Program on Chemical Safety (IPCS) to describe
exposure limits of toxic chemicals and “acceptable daily intake” Consider an example of the use of Option 1 and Option 2
(ADI) is used by the World Health Organization (WHO) applied to acetonitrile in a medicinal product. The permitted
and other national and international health authorities and daily exposure to acetonitrile is 4.1 mg per day ; thus, the
institutes. The new term “permitted daily exposure” (PDE) Option 1 limit is 410 ppm. The maximum administered
is defined in the present guideline as a pharmaceutically daily mass of a medicinal product is 5.0 g, and the medicinal
acceptable intake of residual solvents to avoid confusion of product contains two excipients. The composition of the
differing values for ADI’s of the same substance. medicinal product and the calculated maximum content of
residual acetonitrile are given in the following table.
Residual solvents assessed in this guideline are listed in
Appendix 1 by common names and structures. They were Component Amount in Acetonitrile Daily
evaluated for their possible risk to human health and placed formulation content exposure
into one of three classes as follows : Active substance 0.3 g 800 ppm 0.24 mg
Class 1 solvents : solvents to be avoided Excipient 1 0.9 g 400 ppm 0.36 mg
Known human carcinogens, strongly suspected human Excipient 2 3.8 g 800 ppm 3.04 mg
carcinogens, and environmental hazards. Medicinal product 5.0 g 728 ppm 3.64 mg
Class 2 solvents : solvents to be limited
Excipient 1 meets the Option 1 limit, but the active substance,
Non-genotoxic animal carcinogens or possible causative
excipient 2, and medicinal product do not meet the Option 1
agents of other irreversible toxicity such as neurotoxicity or
limit. Nevertheless, the product meets the Option 2 limit of
teratogenicity.
4.1 mg per day and thus conforms to the recommendations
Solvents suspected of other significant but reversible toxicities. in this guideline.

Consider another example using acetonitrile as residual 4. LIMITS OF RESIDUAL SOLVENTS

solvent. The maximum administered daily mass of a 4.1. SOLVENTS TO BE AVOIDED
medicinal product is 5.0 g, and the medicinal product contains
Solvents in Class 1 should not be employed in the
two excipients. The composition of the medicinal product and
manufacture of active substances, excipients, and medicinal
the calculated maximum content of residual acetonitrile is
products because of their unacceptable toxicity or their
given in the following table.
deleterious environmental effect. However, if their use is
Component Amount in Acetonitrile Daily unavoidable in order to produce a medicinal product with
formulation content exposure a significant therapeutic advance, then their levels should
Active substance 0.3 g 800 ppm 0.24 mg be restricted as shown in Table 1, unless otherwise justified.
1,1,1-Trichloroethane is included in Table 1 because it is an
Excipient 1 0.9 g 2000 ppm 1.80 mg environmental hazard. The stated limit of 1500 ppm is based
Excipient 2 3.8 g 800 ppm 3.04 mg on a review of the safety data.
Medicinal product 5.0 g 1016 ppm 5.08 mg Table 1. – Class 1 solvents in pharmaceutical products
(solvents that should be avoided)
In this example, the product meets neither the Option 1 Solvent Concentration limit Concern
nor the Option 2 limit according to this summation. The (ppm)
manufacturer could test the medicinal product to determine Benzene 2 Carcinogen
if the formulation process reduced the level of acetonitrile. If
Carbon tetrachloride 4 Toxic and environmental hazard
the level of acetonitrile was not reduced during formulation
to the allowed limit, then the manufacturer of the medicinal 1,2-Dichloroethane 5 Toxic
product should take other steps to reduce the amount of
1,1-Dichloroethene 8 Toxic
acetonitrile in the medicinal product. If all of these steps fail
to reduce the level of residual solvent, in exceptional cases 1,1,1-Trichloroethane 1500 Environmental hazard
the manufacturer could provide a summary of efforts made
to reduce the solvent level to meet the guideline value, and 4.2. SOLVENTS TO BE LIMITED
provide a risk-benefit analysis to support allowing the product Solvents in Table 2 should be limited in pharmaceutical
to be utilised containing residual solvent at a higher level. products because of their inherent toxicity. PDEs are given to
3.4. ANALYTICAL PROCEDURES the nearest 0.1 mg/day, and concentrations are given to the
Residual solvents are typically determined using nearest 10 ppm. The stated values do not reflect the necessary
chromatographic techniques such as gas chromatography. analytical precision of determination. Precision should be
Any harmonised procedures for determining levels of residual determined as part of the validation of the method.
solvents as described in the pharmacopoeias should be used,
if feasible. Otherwise, manufacturers would be free to select Table 2. – Class 2 solvents in pharmaceutical products
the most appropriate validated analytical procedure for a Solvent PDE Concentration limit
particular application. If only Class 3 solvents are present, a (mg/day) (ppm)
non-specific method such as loss on drying may be used. Acetonitrile 4.1 410
Validation of methods for residual solvents should conform to Chlorobenzene 3.6 360
ICH guidelines “Text on Validation of Analytical Procedures” Chloroform 0.6 60
and “Extension of the ICH Text on Validation of Analytical
Procedures”. Cumene 0.7 70
3.5. REPORTING LEVELS OF RESIDUAL SOLVENTS Cyclohexane 38.8 3880

Manufacturers of pharmaceutical products need certain 1,2-Dichloroethene 18.7 1870
information about the content of residual solvents in excipients
or active substances in order to meet the criteria of this Dichloromethane 6.0 600
guideline. The following statements are given as acceptable 1,2-Dimethoxyethane 1.0 100
examples of the information that could be provided from a
supplier of excipients or active substances to a pharmaceutical N,N-Dimethylacetamide 10.9 1090
manufacturer. The supplier might choose one of the following N,N-Dimethylformamide 8.8 880
as appropriate :
1,4-Dioxane 3.8 380
– only Class 3 solvents are likely to be present. Loss on drying
is less than 0.5 per cent ; 2-Ethoxyethanol 1.6 160
– only Class 2 solvents X, Y, ... are likely to be present. All Ethyleneglycol 6.2 620
are below the Option 1 limit ; Formamide 2.2 220
(Here the supplier would name the Class 2 solvents Hexane 2.9 290
represented by X, Y, ...)
Methanol 30.0 3000
– only Class 2 solvents X, Y, ... and Class 3 solvents are
likely to be present. Residual Class 2 solvents are below 2-Methoxyethanol 0.5 50
the Option 1 limit and residual Class 3 solvents are below Methylbutylketone 0.5 50
0.5 per cent.
Methylcyclohexane 11.8 1180
If Class 1 solvents are likely to be present, they should be
identified and quantified. “Likely to be present” refers to the Methylisobutylketone 45.0 4500
solvent used in the final manufacturing step and to solvents N-Methylpyrrolidone 5.3 530
that are used in earlier manufacturing steps and not removed
consistently by a validated process. Nitromethane 0.5 50
If solvents of Class 2 or Class 3 are present at greater than Pyridine 2.0 200
their Option 1 limits or 0.5 per cent, respectively, they should Sulfolane 1.6 160
be identified and quantified.

Solvent PDE Concentration limit 4.4. SOLVENTS FOR WHICH NO ADEQUATE

(mg/day) (ppm) TOXICOLOGICAL DATA WAS FOUND
Tetrahydrofuran 7.2 720
The following solvents (Table 4) may also be of interest to
Tetralin 1.0 100 manufacturers of excipients, active substances, or medicinal
products. However, no adequate toxicological data on
Toluene 8.9 890
which to base a PDE was found. Manufacturers should
1,1,2-Trichloroethene 0.8 80 supply justification for residual levels of these solvents in
pharmaceutical products.
Xylene* 21.7 2170
Table 4. – Solvents for which no adequate toxicological data
*usually 60 per cent m-xylene, 14 per cent p-xylene, 9 per cent o-xylene was found
with 17 per cent ethyl benzene.
1,1-Diethoxypropane Methylisopropylketone
4.3. SOLVENTS WITH LOW TOXIC POTENTIAL
1,1-Dimethoxymethane Methyltetrahydrofuran
Solvents in Class 3 (shown in Table 3) may be regarded as less
toxic and of lower risk to human health. Class 3 includes no 2,2-Dimethoxypropane Petroleum ether
solvent known as a human health hazard at levels normally Isooctane Trichloroacetic acid
accepted in pharmaceuticals. However, there are no long-term
toxicity or carcinogenicity studies for many of the solvents in Isopropyl ether Trifluoroacetic acid
Class 3. Available data indicate that they are less toxic in acute
or short-term studies and negative in genotoxicity studies. It GLOSSARY
is considered that amounts of these residual solvents of 50 mg
per day or less (corresponding to 5000 ppm or 0.5 per cent Genotoxic carcinogens: carcinogens which produce cancer by
under Option l) would be acceptable without justification. affecting genes or chromosomes.
Higher amounts may also be acceptable provided they are LOEL : abbreviation for lowest-observed effect level.
realistic in relation to manufacturing capability and good Lowest-observed effect level : the lowest dose of substance in a
manufacturing practice. study or group of studies that produces biologically significant
increases in frequency or severity of any effects in the exposed
Table 3. – Class 3 solvents which should be limited by GMP or humans or animals.
other quality-based requirements
Modifying factor : a factor determined by professional
Acetic acid Heptane
judgement of a toxicologist and applied to bioassay data to
Acetone Isobutyl acetate relate that data safely to humans.
Anisole Isopropyl acetate Neurotoxicity : the ability of a substance to cause adverse
effects on the nervous system.
1-Butanol Methyl acetate NOEL : abbreviation for no-observed-effect level.
2-Butanol 3-Methyl-1-butanol No-observed-effect level : the highest dose of substance at which
Butyl acetate Methylethylketone
there are no biologically significant increases in frequency or
severity of any effects in the exposed humans or animals.
tert-Butylmethyl ether 2-Methyl-1-propanol PDE : abbreviation for permitted daily exposure.
Dimethyl sulfoxide Pentane Permitted daily exposure : the maximum acceptable intake per
day of residual solvent in pharmaceutical products.
Ethanol 1-Pentanol
Reversible toxicity : the occurrence of harmful effects that are
Ethyl acetate 1-Propanol caused by a substance and which disappear after exposure to
Ethyl ether 2-Propanol the substance ends.
Strongly suspected human carcinogen : a substance for which
Ethyl formate Propyl acetate
there is no epidemiological evidence of carcinogenesis but
Formic acid Triethylamine there are positive genotoxicity data and clear evidence of
carcinogenesis in rodents.
Teratogenicity : the occurrence of structural malformations in
a developing foetus when a substance is administered during
pregnancy.
APPENDIX 1. LIST OF SOLVENTS INCLUDED IN THE GUIDELINE
Solvent Other Names Structure Class
Acetic acid Ethanoic acid CH3COOH Class 3
Acetone 2-Propanone CH3COCH3 Class 3
Propan-2-one
Acetonitrile CH3CN Class 2
Anisole Methoxybenzene Class 3
Benzene Benzol Class 1
1-Butanol n-Butyl alcohol CH3[CH2]3OH Class 3

Butan-1-ol
2-Butanol sec-Butyl alcohol CH3CH2CH(OH)CH3 Class 3
Butan-2-ol
Butyl acetate Acetic acid butyl ester CH3COO[CH2]3CH3 Class 3


tert-Butylmethyl ether 2-Methoxy-2-methylpropane (CH3)3COCH3 Class 3
Carbon tetrachloride Tetrachloromethane CCl4 Class 1
Chlorobenzene Class 2
Chloroform Trichloromethane CHCl3 Class 2

Cumene Isopropylbenzene Class 2
(1-Methylethyl)benzene
Cyclohexane Hexamethylene Class 2
1,2-Dichloroethane sym-Dichloroethane CH2ClCH2Cl Class 1

Ethylene dichloride
Ethylene chloride
1,1-Dichloroethene 1,1-Dichloroethylene H2C=CCl2 Class 1
Vinylidene chloride
1,2-Dichloroethene 1,2-Dichloroethylene ClHC=CHCl Class 2
Acetylene dichloride
Dichloromethane Methylene chloride CH2Cl2 Class 2
1,2-Dimethoxyethane Ethyleneglycol dimethyl ether H3COCH2CH2OCH3 Class 2
Monoglyme
Dimethyl cellosolve
N,N-Dimethylacetamide DMA CH3CON(CH3)2 Class 2
N,N-Dimethylformamide DMF HCON(CH3)2 Class 2
Dimethyl sulfoxide Methylsulfinylmethane (CH3)2SO Class 3
Methyl sulfoxide
DMSO
1,4-Dioxane p-Dioxane Class 2
[1,4]Dioxane
Ethanol Ethyl alcohol CH3CH2OH Class 3

2-Ethoxyethanol Cellosolve CH3CH2OCH2CH2OH Class 2
Ethyl acetate Acetic acid ethyl ester CH3COOCH2CH3 Class 3
Ethyleneglycol 1,2-Dihydroxyethane HOCH2CH2OH Class 2
1,2-Ethanediol
Ethyl ether Diethyl ether CH3CH2OCH2CH3 Class 3
Ethoxyethane
1,1′-Oxybisethane
Ethyl formate Formic acid ethyl ester HCOOCH2CH3 Class 3
Formamide Methanamide HCONH2 Class 2
Formic acid HCOOH Class 3
Heptane n-Heptane CH3[CH2]5CH3 Class 3
Hexane n-Hexane CH3[CH2]4CH3 Class 2
Isobutyl acetate Acetic acid isobutyl ester CH3COOCH2CH(CH3)2 Class 3
Isopropyl acetate Acetic acid isopropyl ester CH3COOCH(CH3)2 Class 3
Methanol Methyl alcohol CH3OH Class 2
2-Methoxyethanol Methyl cellosolve CH3OCH2CH2OH Class 2
Methyl acetate Acetic acid methyl ester CH3COOCH3 Class 3
3-Methyl-1-butanol Isoamyl alcohol (CH3)2CHCH2CH2OH Class 3
Isopentyl alcohol
3-Methylbutan-1-ol
Methylbutylketone 2-Hexanone CH3[CH2]3COCH3 Class 2
Hexan-2-one


Methylcyclohexane Cyclohexylmethane Class 2
Methylethylketone 2-Butanone CH3CH2COCH3 Class 3

MEK
Butan-2-one
Methylisobutylketone 4-Methylpentan-2-one CH3COCH2CH(CH3)2 Class 2
4-Methyl-2-pentanone
MIBK
2-Methyl-1-propanol Isobutyl alcohol (CH3)2CHCH2OH Class 3
2-Methylpropan-1-ol
N-Methylpyrrolidone 1-Methylpyrrolidin-2-one Class 2
1-Methyl-2-pyrrolidinone
Nitromethane CH3NO2 Class 2

Pentane n-Pentane CH3[CH2]3CH3 Class 3
1-Pentanol Amyl alcohol CH3[CH2]3CH2OH Class 3
Pentan-1-ol
Pentyl alcohol
1-Propanol Propan-1-ol CH3CH2CH2OH Class 3
Propyl alcohol
2-Propanol Propan-2-ol (CH3)2CHOH Class 3
Isopropyl alcohol
Propyl acetate Acetic acid propyl ester CH3COOCH2CH2CH3 Class 3
Pyridine Class 2
Sulfonane Tetrahydrothiophene 1,1-dioxide Class 2
Tetrahydrofuran Tetramethylene oxide Class 2

Oxacyclopentane
Tetralin 1,2,3,4-Tetrahydronaphthalene Class 2
Toluene Methylbenzene Class 2
1,1,1-Trichloroethane Methylchloroform CH3CCl3 Class 1

1,1,2-Trichloroethene Trichloroethene HClC=CCl2 Class 2
Triethylamine N,N-Diethylethanamine N(CH2CH3)3 Class 3
Xylene* Dimethybenzene Class 2
Xylol
*usually 60 per cent m-xylene, 14 per cent p-xylene, 9 per cent o-xylene with 17 per cent ethyl benzene.
APPENDIX 2. ADDITIONAL BACKGROUND acceptable exposure levels. The goal is protection of human
health and maintenance of environmental integrity against
the possible deleterious effects of chemicals resulting from
long-term environmental exposure. The methods involved in
A2.1. ENVIRONMENTAL REGULATION OF ORGANIC the estimation of maximum safe exposure limits are usually
VOLATILE SOLVENTS based on long-term studies. When long-term study data
Several of the residual solvents frequently used in the are unavailable, shorter term study data can be used with
production of pharmaceuticals are listed as toxic chemicals modification of the approach such as use of larger safety
in Environmental Health Criteria (EHC) monographs and factors. The approach described therein relates primarily to
the Integrated Risk Information System (IRIS). The objectives long-term or life-time exposure of the general population in
of such groups as the International Programme on Chemical the ambient environment, i.e. ambient air, food, drinking
Safety (IPCS), the United States Environmental Protection water and other media.
Agency (USEPA) and the United States Food and Drug
Administration (USFDA) include the determination of

A2.2. RESIDUAL SOLVENTS IN PHARMACEUTICALS “safety factors” in Pharmacopoeial Forum. The assumption
Exposure limits in this guideline are established by referring of 100 per cent systemic exposure is used in all calculations
to methodologies and toxicity data described in EHC and regardless of route of administration.
IRIS monographs. However, some specific assumptions about
residual solvents to be used in the synthesis and formulation
of pharmaceutical products should be taken into account in The modifying factors are as follows :
establishing exposure limits. They are :
F1 = a factor to account for extrapolation between species :
1) Patients (not the general population) use pharmaceuticals
to treat their diseases or for prophylaxis to prevent infection F1 = 2 for extrapolation from dogs to humans ;
or disease.
F1 = 2.5 for extrapolation from rabbits to humans ;
2) The assumption of life-time patient exposure is not
necessary for most pharmaceutical products but may be F1 = 3 for extrapolation from monkeys to humans ;
appropriate as a working hypothesis to reduce risk to human
health. F1 = 5 for extrapolation from rats to humans ;
3) Residual solvents are unavoidable components in F1 = 10 for extrapolation from other animals to
pharmaceutical production and will often be a part of humans ;
medicinal products.
F1 = 12 for extrapolation from mice to humans.
4) Residual solvents should not exceed recommended levels
except in exceptional circumstances.
5) Data from toxicological studies that are used to determine F1 takes into account the comparative surface area : body
acceptable levels for residual solvents should have been weight ratios for the species concerned and for man. Surface
generated using appropriate protocols such as those described area (S) is calculated as :
for example, by OECD, EPA, and the FDA Red Book.
S = km0.67
APPENDIX 3. METHODS FOR ESTABLISHING EXPOSURE
LIMITS
in which m = body mass, and the constant k has been taken to
The Gaylor-Kodell method of risk assessment (Gaylor, D. W. be 10. The body weight used in the equation are those shown
and Kodell, R. L. Linear Interpolation algorithm for low dose below in Table A3.-1.
assessment of toxic substance. J. Environ. Pathology, 4, 305,
1980) is appropriate for Class 1 carcinogenic solvents. Only in Table A3.-1. – Values used in the calculations in this document
cases where reliable carcinogenicity data are available should
extrapolation by the use of mathematical models be applied to Rat body weight 425 g
setting exposure limits. Exposure limits for Class 1 solvents Pregnant rat body weight 330 g
could be determined with the use of a large safety factor (i.e.,
10 000 to 100 000) with respect to the no-observed-effect level Mouse body weight 28 g
(NOEL). Detection and quantification of these solvents should Pregnant mouse body weight 30 g
be by state-of-the-art analytical techniques.
Guinea-pig body weight 500 g
Rhesus monkey body weight 2.5 kg

Acceptable exposure levels in this guideline for Class 2
Rabbit body weight (pregnant or not) 4 kg
solvents were established by calculation of PDE values
according to the procedures for setting exposure limits in Beagle dog body weight 11.5 kg
pharmaceuticals (Pharmacopeial Forum, Nov-Dec 1989),
Rat respiratory volume
and the method adopted by IPCS for Assessing Human 290 L/day
Health Risk of Chemicals (Environmental Health Criteria Mouse respiratory volume 43 L/day
170, WHO, 1994). These methods are similar to those used
by the USEPA (IRIS) and the USFDA (Red Book) and others. Rabbit respiratory volume 1440 L/day
The method is outlined here to give a better understanding of Guinea-pig respiratory volume 430 L/day
the origin of the PDE values. It is not necessary to perform
these calculations in order to use the PDE values tabulated in Human respiratory volume 28800 L/day
Section 4 of this document. Dog respiratory volume 9000 L/day
Monkey respiratory volume 1150 L/day
PDE is derived from the no-observed-effect level (NOEL), or Mouse water consumption 5 mL/day
the lowest-observed effect level (LOEL), in the most relevant Rat water consumption 30 mL/day
animal study as follows :
Rat food consumption 30 g/day
NOEL ´ Weight Adjustment
PDE =
F1 ´ F2 ´ F3 ´ F4 ´ F5
F2 = a factor of 10 to account for variability between
The PDE is derived preferably from a NOEL. If no NOEL individuals.
is obtained, the LOEL may be used. Modifying factors A factor of 10 is generally given for all organic
proposed here, for relating the data to humans, are the same solvents, and 10 is used consistently in this guideline.
kind of “uncertainty factors” used in Environmental Health
Criteria (Environmental Health Criteria 170, World Health F3 = a variable factor to account for toxicity studies of
Organization, Geneva, 1994), and “modifying factors” or short-term exposure :

F3 = 1 for studies that last at least one half-lifetime calculation. It is recognised that some adult patients weigh less
(1 year for rodents or rabbits ; 7 years for cats, than 50 kg ; these patients are considered to be accommodated
dogs and monkeys) ; by the built-in safety factors used to determine a PDE. If the
solvent was present in a formulation specifically intended for
F3 = 1 for reproductive studies in which the whole paediatric use, an adjustment for a lower body weight would
period of organogenesis is covered ; be appropriate.
F3 = 2 for a 6 month study in rodents, or a 3.5 year As an example of the application of this equation, consider the
study in non-rodents ; toxicity study of acetonitrile in mice that is summarised in
Pharmeuropa, Vol. 9. No. 1, Supplement, April 1997, page S24.
F3 = 5 for a 3 month study in rodents, or a 2 year The NOEL is calculated to be 50.7 mg kg–1 day–l. The PDE for
study in non-rodents ; acetonitrile in this study is calculated as follows :
F3 = 10 for studies of a shorter duration. 50.7mg kg-1 day-1 ´ 50 kg
PDE = = 4.22 mg day-1
In all cases, the higher factor has been used for study durations 12 ´ 10 ´ 5 ´ 1 ´ 1
between the time points, e.g. a factor of 2 for a 9 month In this example,
rodent study.
F1 = 12 to account for the extrapolation from mice to
F4 = a factor that may be applied in cases of severe toxicity, humans ;
e.g. non-genotoxic carcinogenicity, neurotoxicity or
teratogenicity. F2 = 10 to account for differences between individual
In studies of reproductive toxicity, the following humans ;
factors are used : F3 = 5 because the duration of the study was only
13 weeks ;
F4 = 1 for foetal toxicity associated with maternal
toxicity ; F4 = 1 because no severe toxicity was encountered ;
F4 = 5 for foetal toxicity without maternal toxicity ; F5 = 1 because the no-effect level was determined.
F4 = 5 for a teratogenic effect with maternal toxicity ; The equation for an ideal gas, PV = nRT, is used to convert
concentrations of gases used in inhalation studies from units
F4 = 10 for a teratogenic effect without maternal of ppm to units of mg/L or mg/m3. Consider as an example
toxicity. the rat reproductive toxicity study by inhalation of carbon
tetrachloride (molecular weight 153.84) summarised in
F5 = a variable factor that may be applied if the no-effect Pharmeuropa, Vol. 9, No. 1, Supplement, April 1997, page S9.
level was not established.
When only a LOEL is available, a factor of up to 10 can be n P 300 ´ 10-6 atm ´ 153 840 mg mol-1
= =
used depending on the severity of the toxicity. V RT 0.082 L atm K -1mol-1 ´ 298 K
The weight adjustment assumes an arbitrary adult human
46.15 mg
body weight for either sex of 50 kg. This relatively low weight = = 1.89 mg / L
provides an additional safety factor against the standard 24.45 L
weights of 60 kg or 70 kg that are often used in this type of The relationship 1000 L = 1 m3 is used to convert to mg/m3.

EUROPEAN PHARMACOPOEIA 5.9. Polymorphism
01/2008:50900 If each crystalline form is the more stable within a given

corrected 10.0 temperature range, the change from one form to another is
reversible and is said to be enantiotropic. The change from one
phase to another is a univariate equilibrium, so that at a given
pressure this state is characterised by a transition temperature.
However, if only one of the forms is stable over the entire
5.9. POLYMORPHISM temperature range, the change is irreversible or monotropic.
Polymorphism (or crystal polymorphism) is a phenomenon Different crystalline forms or solvates may be produced by
related to the solid state ; it is the ability of a compound in varying the crystallisation conditions (temperature, pressure,
the solid state to exist in different crystalline forms having solvent, concentration, rate of crystallisation, seeding of
the same chemical composition. Substances that exist in a the crystallisation medium, presence and concentration of
non-crystalline solid state are said to be amorphous. impurities, etc.).
When this phenomenon is observed for a chemical element The following techniques may be used to study polymorphism :
(for example, sulfur), the term allotropy is used instead of – X-ray diffraction of powders (2.9.33),
polymorphism. – X-ray diffraction of single crystals,
The term pseudopolymorphism is used to describe solvates
– thermal analysis (2.2.34) (differential scanning calorimetry,
(including hydrates), where a solvent is present in the crystal
thermogravimetry, thermomicroscopy),
matrix in stoichiometric proportions ; the term may also be
extended to include compounds where the solvent is trapped – microcalorimetry,
in the matrix in variable proportions. However the term – moisture absorption analysis,
pseudopolymorphism is ambiguous because of its use in
– optical and electronic microscopy,
different circumstances. It is therefore preferable to use only
the terms “solvates” and “hydrates”. – solid-state nuclear magnetic resonance,
Where a monograph indicates that a substance shows – infrared absorption spectrophotometry (2.2.24),
polymorphism, this may be true crystal polymorphism, – Raman spectroscopy (2.2.48),
occurrence of solvates, allotropy or occurrence of the
amorphous form. – measurement of solubility and intrinsic dissolution rate,
The identity of chemical composition implies that all – density measurement.
crystalline and amorphous forms of a given species have the These techniques are often complementary and it is
same chemical behaviour in solution or as a melt ; in contrast, indispensable to use several of them.
their physico-chemical and physical characteristics (solubility, Pressure/temperature and energy/temperature diagrams based
hardness, compressibility, density, melting point, etc.), and on analytical data are valuable tools for fully understanding
therefore their reactivity and bioavailability may be different the energetic relationship (enantiotropism, monotropism) and
at the solid state. the thermodynamic stability of the individual modifications
When a compound shows polymorphism, the form for which of a polymorphic compound.
the free enthalpy is lowest at a given temperature and pressure
is the most thermodynamically stable. The other forms are For solvates, differential scanning calorimetry and
said to be in a metastable state. At normal temperature and thermogravimetry are preferable, combined with
pressure, a metastable form may remain unchanged or may measurements of solubility, intrinsic dissolution rate and
change to a thermodynamically more stable form. X-ray diffraction.
If there are several crystalline forms, one form is For hydrates, water sorption/desorption isotherms are
thermodynamically more stable at a given temperature and determined to demonstrate the zones of relative stability.
pressure. A given crystalline form may constitute a phase that In general, hydrates are less soluble in water than anhydrous
can reach equilibrium with other solid phases and with the forms, and likewise solvates are less soluble in their solvent
liquid and gas phases. than unsolvated forms.

EUROPEAN PHARMACOPOEIA 5.10. Impurities in substances for pharmaceutical use
04/2012:51000 organic and inorganic impurities that are relevant in view

corrected 10.0 of the sources of active substances in authorised medicinal
products.
Control of residual solvents is provided by the general
monograph Substances for pharmaceutical use (2034) and
general chapter 5.4. Residual solvents. The certificate of
5.10. CONTROL OF IMPURITIES suitability of a monograph of the European Pharmacopoeia
IN SUBSTANCES FOR for a given source of a substance indicates the residual solvents
that are controlled together with the specified acceptance
PHARMACEUTICAL USE criteria and the validated control method where this differs
from those described in general chapter 2.4.24. Identification
Preamble and control of residual solvents.
The monographs of the European Pharmacopoeia on Monographs on organic chemicals usually have a test entitled
substances for pharmaceutical use are designed to ensure “Related substances” that covers relevant organic impurities.
acceptable quality for users. The role of the Pharmacopoeia This test may be supplemented by specific tests where the
in public health protection requires that adequate control of general test does not control a given impurity or where
impurities be provided by monographs. The quality required there are particular reasons (for example, safety reasons) for
is based on scientific, technical and regulatory considerations. requiring special control.
Requirements concerning impurities are given in specific
monographs and in the general monograph Substances for Where a monograph has no Related substances (or equivalent)
pharmaceutical use (2034). Specific monographs and the test but only specific tests, the user of a substance must
general monograph are complementary : specific monographs nevertheless ensure that there is suitable control of organic
prescribe acceptance criteria for impurities whereas the impurities ; those occurring above the identification threshold
general monograph deals with the need for qualification, are to be identified (wherever possible) and, unless justified,
identification and reporting of any organic impurities that those occurring above the qualification threshold are to
occur in active substances. be qualified (see also under Recommendations to users of
monographs of active substances).
The thresholds for reporting, identification and qualification
contained in the general monograph Substances for Where the monograph covers substances with different
pharmaceutical use (2034) apply to all related substances. impurity profiles, it may have a single related substances test
However, if a monograph does not contain a related substances to cover all impurities mentioned in the Impurities section or
test based on a quantitative method, any new impurities several tests may be necessary to give control of all known
occurring above a threshold may be overlooked since the test profiles. Compliance may be established by carrying out only
is not capable to detect those impurities. the tests relevant to the known impurity profile for the source
of the substance.
The provisions of the Related substances section of the general
monograph Substances for pharmaceutical use (2034), notably Instructions for control of impurities may be included in the
those concerning thresholds, do not apply to excipients ; Production section of a monograph, for example where the
also excluded from the provisions of this section are : only analytical method appropriate for the control of a given
biological and biotechnological products ; oligonucleotides ; impurity is to be performed by the manufacturer since the
radiopharmaceuticals ; fermentation products and method is too technically complex for general use or cannot
semi-synthetic products derived therefrom ; herbal products be applied to the final drug substance and/or where validation
and crude products of animal and plant origin. Although of the production process (including the purification step) will
the thresholds stated in the general monograph do not apply, give sufficient control.
the general concepts of reporting, identification (wherever Impurities section in monographs on active substances
possible) and qualification of impurities are equally valid for
these classes. The Impurities section in a monograph includes impurities
Basis for the elaboration of monographs of the European (chemical structure and name wherever possible), which are
Pharmacopoeia usually organic, that are known to be detected by the tests
prescribed in the monograph. It is based on information
European Pharmacopoeia monographs are elaborated on available at the time of elaboration or revision of the
substances that are present in medicinal products that have monograph and is not necessarily exhaustive. The section
been authorised by the competent authorities of Parties to the includes specified impurities and, where so indicated, other
European Pharmacopoeia Convention. Consequently, these detectable impurities.
monographs do not necessarily cover all sources of substances
for pharmaceutical use on the world market. Specified impurities have an acceptance criterion not greater
than that authorised by the competent authorities.
Organic and inorganic impurities present in those substances
that have been evaluated by the competent authorities are Other detectable impurities are potential impurities with a
qualified with respect to safety at the maximum authorised defined structure but not known to be normally present above
content (at the maximum daily dose) unless new safety data the identification threshold in substances used in medicinal
that become available following evaluation justify lower limits. products that have been authorised by the competent
European Pharmacopoeia monographs on substances for authorities of Parties to the Convention. They are given in the
pharmaceutical use are elaborated by groups of experts and Impurities section for information.
working parties collaborating with national pharmacopoeia Where an impurity other than a specified impurity is found
authorities, the competent authorities for marketing in an active substance it is the responsibility of the user of the
authorisation, national control laboratories and the European substance to check whether it has to be identified/qualified,
Pharmacopoeia laboratory ; they are also assisted by the depending on its content, nature, maximum daily dose and
producers of the substances and/or the pharmaceutical relevant identification/qualification threshold, in accordance
manufacturers that use these substances. with the general monograph on Substances for pharmaceutical
Control of impurities in substances for pharmaceutical use use (2034), Related substances section.
The quality with respect to impurities is controlled by a set of It should be noted that specific thresholds are applied to
tests within a monograph. These tests are intended to cover substances exclusively for veterinary use.

5.10. Impurities in substances for pharmaceutical use EUROPEAN PHARMACOPOEIA
Interpretation of the test for related substances in the certain specified impurities only or to unspecified impurities
monographs on active substances and certain specified impurities, depending on the nature
A specific monograph on a substance for pharmaceutical use of the active substance and the applicable identification
is to be read and interpreted in conjunction with the general threshold. Pending editorial adaptation of already published
monograph on Substances for pharmaceutical use (2034). monographs using unequivocal terminology, the decision
Where a general acceptance criterion for impurities (“any tree (Figure 5.10.-1) may be used to determine the acceptance
criterion to be applied.
other impurity”, “other impurities”, “any impurity”) equivalent
to a nominal content greater than the applicable identification Recommendations to users of monographs of active
threshold (see the general monograph on Substances for substances
pharmaceutical use (2034)) is prescribed, this is valid only Monographs give a specification for suitable quality of
for specified impurities mentioned in the Impurities section. substances with impurity profiles corresponding to those
The need for identification (wherever possible), reporting, taken into account during elaboration and/or revision of the
specification and qualification of other impurities that occur monograph. It is the responsibility of the user of the substance
must be considered according to the requirements of the to check that the monograph provides adequate control of
general monograph. It is the responsibility of the user of the impurities for a substance for pharmaceutical use from a given
substance to determine the validity of the acceptance criteria source, notably by using the procedure for certification of
for impurities not mentioned in the Impurities section and for suitability of the monographs of the European Pharmacopoeia.
those indicated as other detectable impurities. A monograph with a related substances test based on a
Acceptance criteria for the related substances test are presented quantitative method (such as liquid chromatography, gas
in different ways in existing monographs ; the decision tree chromatography and capillary electrophoresis) provides
(Figure 5.10.-1) may be used as an aid in the interpretation adequate control of impurities for a substance from a given
of general acceptance criteria and their relation with the source if impurities present in amounts above the applicable
Impurities section of the monograph. identification threshold are specified impurities mentioned in
General acceptance criteria for “other” impurities are the Impurities section.
expressed in various ways in the monographs : “any other If the substance contains impurities other than those
impurity”, “other impurities”, “any impurity”, “any spot”, “any mentioned in the Impurities section, it has to be verified
band”, etc. The general acceptance criteria may apply to that these impurities are detectable by the method described
The general acceptance criterion applies to:

Does the Related substances – all unspecified impurities;
No
section of the monograph – specified impurities, except those that
Substances for pharmaceutical have their own specific acceptance
use (2034) apply?* criterion in the monograph.
Yes

– all unspecified impurities;
Is the general acceptance criterion Yes
less than or equal to the applicable – specified impurities, except those that
identification threshold? have their own specific acceptance
criterion in the monograph.
No

– specified impurities, except those that have
Yes their own specific acceptance criterion in the
Does the monograph have an monograph.
Impurities section?
For unspecified impurities, apply the Related
substances section of the monograph
Substances for pharmaceutical use (2034).**
No
Apply the Related substances section

of the monograph Substances
for pharmaceutical use (2034)**
* The requirements of this section apply to active substances with the exception of: biological and biotechnological products;
oligonucleotides; radiopharmaceuticals; products of fermentation and semi-synthetic products derived therefrom; crude
products of animal or plant origin; herbal products.
** To apply the Related substances section of the monograph Substances for pharmaceutical use (2034):
– an individual acceptance criterion must be defined for any impurity that may be present above the identification threshold;
– any impurity with an acceptance criterion above the identification threshold must wherever possible be identified;
– any impurity with an acceptance criterion above the qualification threshold must be qualified.
Figure 5.10.-1. – Decision tree for interpretation of general acceptance criteria for ‘other’ impurities in monographs

EUROPEAN PHARMACOPOEIA 5.10. Impurities in substances for pharmaceutical use
in the monograph, otherwise a new method must be GLOSSARY

developed and revision of the monograph must be requested. Disregard limit : in chromatographic tests, the nominal
Depending on the contents found and the limits proposed, content at or below which peaks/signals are not taken into
the identification and/or the qualification of these impurities account for calculating a sum of impurities. The numerical
must be considered. values for the disregard limit and the reporting threshold are
Where a single related substances test covers different impurity usually the same.
profiles, only impurities for the known profile from a single
source need to be reported in the certificate of analysis unless Identification threshold : a limit above which an impurity
the marketing authorisation holder uses active substances is to be identified.
with different impurity profiles. Identified impurity : an impurity for which structural
Identification of impurities (peak assignment) characterisation has been achieved.
Where a monograph has an individual limit for an impurity, Impurity : any component of a substance for pharmaceutical
it is often necessary to define means of identification, use that is not the chemical entity defined as the substance.
for example using a reference substance, a representative Nominal concentration : concentration calculated on the
chromatogram or relative retention. The user of the substance basis of the concentration of the prescribed reference and
may find it necessary to identify impurities other than those taking account of the prescribed correction factor.
for which the monograph provides a means of identification,
for example to check the suitability of the specification for Other detectable impurities : potential impurities with a
a given impurity profile by comparison with the Impurities defined structure that are known to be detected by the tests in
section. The European Pharmacopoeia does not provide a monograph but not known to be normally present above
reference substances, representative chromatograms or the identification threshold in substances used in medicinal
information on relative retentions for this purpose, unless products that have been authorised by the competent
prescribed in the monograph. Users will therefore have to authorities of Parties to the Convention. They are unspecified
apply the available scientific techniques for identification. impurities and are thus limited by a general acceptance
criterion.
New impurities/Specified impurities above the specified
limit Potential impurity : an impurity that theoretically can arise
Where a new manufacturing process or change in an during manufacture or storage. It may or may not actually
established process leads to the occurrence of a new appear in the substance. Where a potential impurity is known
impurity, it is necessary to apply the provisions of the general to be detected by the tests in a monograph but not known to
monograph on Substances for pharmaceutical use (2034) be normally present in substances used in medicinal products
regarding identification and qualification and to verify the that have been authorised by the competent authorities of
suitability of the monograph for control of the impurity. Parties to the Convention, it will be included in the Impurities
A certificate of suitability is a means for confirming for section under Other detectable impurities for information.
a substance from a given source that the new impurity is Qualification : the process of acquiring and evaluating data
adequately controlled or the certificate contains a method for that establishes the biological safety of an individual impurity
control with a defined acceptance criterion. In the latter case or a given impurity profile at the level(s) specified.
revision of the monograph will be initiated. Qualification threshold : a limit above which an impurity
Where a new manufacturing process or change in an is to be qualified.
established process leads to the occurrence of a specified
impurity above the specified limit, it is necessary to apply Related substances : title used in monographs for general
the provisions of the general monograph on Substances for tests for organic impurities.
pharmaceutical use (2034) regarding qualification. Reporting threshold : a limit above which an impurity is to
Expression of acceptance criteria be reported. Synonym : reporting level.
The acceptance criteria for related substances are expressed Specified impurity : an impurity that is individually listed and
in monographs either in terms of comparison of peak areas limited with a specific acceptance criterion in a monograph. A
(comparative tests) or as numerical values. specified impurity can be either identified or unidentified.
Chromatographic methods Unidentified impurity : an impurity for which a structural
General chapter 2.2.46. Chromatographic separation characterisation has not been achieved and that is defined
techniques deals with various aspects of impurities control. solely by qualitative analytical properties (for example, relative
Information is available via the EDQM website on commercial retention).
names for columns and other reagents and equipment found Unspecified impurity : an impurity that is limited by a general
suitable during monograph development, where this is acceptance criterion and not individually listed with its own
considered useful. specific acceptance criterion.

EUROPEAN PHARMACOPOEIA Substances for pharmaceutical use
01/2018:2034 The manufacture of active substances must take place under

conditions of good manufacturing practice.
The provisions of general chapter 5.10 apply to the control of
impurities in substances for pharmaceutical use.
SUBSTANCES Whether or not it is specifically stated in the individual

FOR PHARMACEUTICAL USE monograph that the substance for pharmaceutical use :
– is a recombinant protein or another substance obtained
Corpora ad usum pharmaceuticum as a direct gene product based on genetic modification,
where applicable, the substance also complies with the
DEFINITION requirements of the general monograph Products of
Substances for pharmaceutical use are any organic or inorganic recombinant DNA technology (0784) ;
substances that are used as active substances or excipients for – is obtained from animals susceptible to transmissible
the production of medicinal products for human or veterinary spongiform encephalopathies other than by experimental
use. They may be obtained from natural sources or produced challenge, where applicable, the substance also complies
by extraction from raw materials, fermentation or synthesis. with the requirements of the general monograph Products
This general monograph does not apply to herbal drugs, with risk of transmitting agents of animal spongiform
herbal drugs for homoeopathic preparations, herbal drug encephalopathies (1483) ;
preparations, herbal drug extracts, or mother tinctures – is a substance derived from a fermentation process,
for homoeopathic preparations, which are the subject of whether or not the micro-organisms involved are modified
separate general monographs (Herbal drugs (1433), Herbal by traditional procedures or recombinant DNA (rDNA)
drugs for homoeopathic preparations (2045), Herbal drug technology, where applicable, the substance also complies
preparations (1434), Herbal drug extracts (0765), Mother with the requirements of the general monograph Products
tinctures for homoeopathic preparations (2029)). It does not of fermentation (1468).
apply to raw materials for homoeopathic preparations, except
where there is an individual monograph for the substance in If solvents are used during production, they are of suitable
the non-homoeopathic part of the Pharmacopoeia. quality. In addition, their toxicity and their residual level
This monograph does not apply to chemical precursors are taken into consideration (5.4). If water is used during
for radiopharmaceutical preparations which are the production, it is of suitable quality.
subject of a separate monograph (Chemical precursors for The identity of elemental impurities derived from intentionally
radiopharmaceutical preparations (2902)). added catalysts and reagents is known, and strategies for
Where a substance for pharmaceutical use not described in controlling them should be established using the principles of
an individual monograph of the Pharmacopoeia is used in a risk management.
medicinal product prepared for the special needs of individual If substances are produced or processed to yield a certain
patients, the need for compliance with the present general form or grade, that specific form or grade of the substance
monograph is decided in the light of a risk assessment that complies with the requirements of the monograph. Certain
takes account of the available quality of the substance and its functionality-related tests may be described to control
intended use. properties that may influence the suitability of the substance
Where medicinal products are manufactured using substances and subsequently the properties of dosage forms prepared
for pharmaceutical use of human or animal origin, the from it.
requirements of chapter 5.1.7. Viral safety apply. Powdered substances may be processed to obtain a certain
Substances for pharmaceutical use may be used as such or degree of fineness (2.9.35).
as starting materials for subsequent formulation to prepare Compacted substances are processed to increase the particle
medicinal products. Depending on the formulation, certain size or to obtain particles of a specific form and/or to obtain
substances may be used either as active substances or as a substance with a higher bulk density.
excipients. Solid substances may be compacted, coated, Coated active substances consist of particles of the active
granulated, powdered to a certain fineness, or processed substance coated with one or more suitable excipients.
in other ways. A monograph is applicable to a substance
processed with an excipient only where such processing is Granulated active substances are particles of a specified
mentioned in the definition section of the monograph. size and/or form produced from the active substance by
granulation directly or with one or more suitable excipients.
If substances are processed with excipients, these excipients
Substance for pharmaceutical use of special grade. Unless comply with the requirements of the relevant monograph or,
otherwise indicated or restricted in the individual where no such monograph exists, the approved specification.
monographs, a substance for pharmaceutical use is intended
for human and veterinary use, and is of appropriate quality for Where active substances have been processed with excipients
the manufacture of all dosage forms in which it can be used. to produce, for example, coated or granulated substances,
the processing is carried out under conditions of good
manufacturing practice and the processed substances are
Polymorphism. Individual monographs do not usually specify regarded as intermediates in the manufacture of a medicinal
crystalline or amorphous forms, unless bioavailability is product.
affected. All forms of a substance for pharmaceutical use
comply with the requirements of the monograph, unless CHARACTERS
otherwise indicated. The statements under the heading Characters (e.g. statements
about the solubility or a decomposition point) are not to be
interpreted in a strict sense and are not requirements. They
PRODUCTION are given for information.
Substances for pharmaceutical use are manufactured by Where a substance may show polymorphism, this may be
procedures that are designed to ensure a consistent quality and stated under Characters in order to draw this to the attention
comply with the requirements of the individual monograph or of the user who may have to take this characteristic into
approved specification. consideration during formulation of a preparation.

Substances for pharmaceutical use EUROPEAN PHARMACOPOEIA
IDENTIFICATION The requirements above do not apply to biological and

Where under Identification an individual monograph biotechnological products, oligonucleotides, products of
contains subdivisions entitled ‘First identification’ and fermentation and semi-synthetic products derived therefrom,
‘Second identification’, the test or tests that constitute the to crude products of animal or plant origin or herbal products.
‘First identification’ may be used in all circumstances. The Elemental impurities. Permitted daily exposures for
test or tests that constitute the ‘Second identification’ may be elemental impurities (e.g. as included in the ICH Q3D
used in pharmacies provided it can be demonstrated that the guideline, the principles of which are reproduced in
substance or preparation is fully traceable to a batch certified general chapter 5.20. Elemental impurities) apply to the
to comply with all the other requirements of the monograph. medicinal product. Individual monographs on substances for
Certain monographs give two or more sets of tests for the pharmaceutical use therefore do not contain specifications for
purpose of the first identification, which are equivalent elemental impurities unless otherwise prescribed.
and may be used independently. One or more of these sets Residual solvents are limited according to the principles
usually contain a cross-reference to a test prescribed in the defined in chapter 5.4, using general method 2.4.24 or another
Tests section of the monograph. It may be used to simplify suitable method. Where a quantitative determination of a
the work of the analyst carrying out the identification and residual solvent is carried out and a test for loss on drying is
the prescribed tests. For example, one identification set not carried out, the content of residual solvent is taken into
cross-refers to a test for enantiomeric purity while the other account for calculation of the assay content of the substance,
set gives a test for specific optical rotation : the intended the specific optical rotation and the specific absorbance.
purpose of the two is the same, that is, verification that the Microbiological quality. Individual monographs give
correct enantiomer is present. acceptance criteria for microbiological quality wherever such
control is necessary. Table 5.1.4.-2. – Acceptance criteria
TESTS
for microbiological quality of non-sterile substances for
Polymorphism (5.9). If the nature of a crystalline or pharmaceutical use in chapter 5.1.4. Microbiological quality
amorphous form imposes restrictions on its use in of non-sterile pharmaceutical preparations and substances for
preparations, the nature of the specific crystalline or pharmaceutical use gives recommendations on microbiological
amorphous form is identified, its morphology is adequately quality that are of general relevance for substances subject to
controlled and its identity is stated on the label. microbial contamination. Depending on the nature of the
Related substances. Unless otherwise prescribed or justified substance and its intended use, different acceptance criteria
and authorised, organic impurities in active substances are may be justified.
to be reported, identified wherever possible, and qualified as Sterility (2.6.1). If intended for use in the manufacture of
indicated in Table 2034.-1 or in Table 2034.-2 for peptides sterile dosage forms without a further appropriate sterilisation
obtained by chemical synthesis. procedure, or if offered as sterile grade, the substance for
pharmaceutical use complies with the test for sterility.
Table 2034.-1. – Reporting, identification and qualification of
organic impurities in active substances Bacterial endotoxins (2.6.14). The substance for
pharmaceutical use complies with the test for bacterial
Use Maximum Report- Identification Qualification endotoxins if it is labelled as a bacterial endotoxin-free grade
daily ing threshold threshold or if it is intended for use in the manufacture of parenteral
dose threshold preparations or preparations for irrigation without a further
Human ≤ 2 g/day > 0.05 per > 0.10 per > 0.15 per appropriate procedure for the removal of bacterial endotoxins.
use or cent cent or a cent or a The limit, when not indicated in the individual monograph,
human daily intake daily intake is determined in accordance with the recommendations
and of > 1.0 mg of > 1.0 mg
veterinary (whichever is (whichever is of general chapter 5.1.10. Guidelines for using the test for
use the lower) the lower) bacterial endotoxins.
Human > 2 g/day > 0.03 per > 0.05 per > 0.05 per Pyrogens (2.6.8). If the test for pyrogens is justified rather
use or cent cent cent than the test for bacterial endotoxins and if a pyrogen-free
human grade is offered, the substance for pharmaceutical use
and complies with the test for pyrogens. The limit and test method
veterinary
use are stated in the individual monograph or approved by the
competent authority. Based on appropriate test validation
Veterinary Not > 0.10 per > 0.20 per > 0.50 per for bacterial endotoxins and pyrogens, the test for bacterial
use only applicable cent cent cent
endotoxins may replace the test for pyrogens.
Table 2034.-2. – Reporting, identification and qualification of Additional properties. Control of additional properties (e.g.
organic impurities in peptides obtained by chemical synthesis physical characteristics, functionality-related characteristics)
may be necessary for individual manufacturing processes
Reporting Identification Qualification or formulations. Grades (such as sterile, endotoxin-free,
threshold threshold threshold pyrogen-free) may be produced with a view to manufacture
> 0.1 per cent > 0.5 per cent > 1.0 per cent of preparations for parenteral administration or other dosage
forms and appropriate requirements may be specified in an
Specific thresholds may be applied for impurities known individual monograph.
to be unusually potent or to produce toxic or unexpected
pharmacological effects. ASSAY
Unless justified and authorised, contents of substances for
For DNA reactive impurities, the requirements of ICH pharmaceutical use are determined. Suitable methods are
Guideline M7 Assessment and Control of DNA Reactive used.
(Mutagenic) Impurities in Pharmaceuticals to Limit Potential
Carcinogenic Risk must be complied with for active substances LABELLING
to be used in medicinal products for human use, in cases In general, labelling is subject to supranational and national
defined in the scope of the guideline. regulation and to international agreements. The statements
If the individual monograph does not provide suitable under the heading Labelling therefore are not comprehensive
control for a new impurity, a suitable test for control must be and, moreover, for the purposes of the Pharmacopoeia only
developed and included in the specification for the substance. those statements that are necessary to demonstrate compliance

EUROPEAN PHARMACOPOEIA Substances for pharmaceutical use
or non-compliance with the monograph are mandatory. Any – compacted ;

other labelling statements are included as recommendations. – coated ;
When the term ‘label’ is used in the Pharmacopoeia, the
– granulated ;
labelling statements may appear on the container, the package,
a leaflet accompanying the package or a certificate of analysis – sterile ;
accompanying the article, as decided by the competent – free from bacterial endotoxins ;
authority. – free from pyrogens ;
Where appropriate, the label states that the substance is : – containing gliding agents.
– intended for a specific use ; Where applicable, the label states :
– of a distinct crystalline form ; – the degree of hydration ;
– of a specific degree of fineness ; – the name and concentration of any excipient.

EUROPEAN PHARMACOPOEIA Products of fermentation
04/2018:1468 All containers in a cell bank are stored under identical

conditions. Once removed from storage, the individual
ampoules, vials or culture straws are not returned to the cell
bank.
PRODUCTS OF FERMENTATION PROCESSES USING STAGED GROWTH IN CULTURES

The contents of a container of the working cell bank are
Producta ab fermentatione used, if necessary after resuspension, to prepare an inoculum
in a suitable medium. After a suitable period of growth,
This monograph applies to indirect gene products obtained by the cultures are used to initiate the fermentation process,
fermentation. It is not applicable to : if necessary following preculture in a prefermentor. The
– monographs in the Pharmacopoeia concerning vaccines for conditions to be used at each stage of the process are defined
human or veterinary use ; and must be met with each production run.
– products derived from continuous cell lines of human or
CHANGE CONTROL
animal origin ;
– direct gene products that result from the transcription and If the production process is altered in a way that causes a
translation from nucleic acid to protein, whether or not significant change in the impurity profile of the product, the
subject to post-translational modification ; critical steps associated with this change in impurity profile
are revalidated.
– products obtained by semi-synthesis from a product
of fermentation and those obtained by biocatalytic If a significant change has taken place in the micro-organism
transformation ; used for production that causes a significant change in the
– whole broth concentrates or raw fermentation products. impurity profile of the product, the critical steps of the
production process associated with this change, particularly
This monograph provides general requirements for the the procedure for purification and isolation, are revalidated.
development and manufacture of products of fermentation.
These requirements are not necessarily comprehensive in a Revalidation includes demonstration that new impurities
given case and requirements complementary or additional present in the product as a result of the change are adequately
to those prescribed in this monograph may be imposed in an controlled by the test procedures. If necessary, additional or
individual monograph or by the competent authority. alternative tests must be introduced with appropriate limits.
If the change in the process or in the micro-organism results
DEFINITION in an increase in the level of an impurity already present, the
For the purposes of this monograph, products of fermentation acceptability of such an increase is addressed.
are active or inactive pharmaceutical substances produced When a master cell bank is replaced, the critical steps of the
by controlled fermentation as indirect gene products. They production process must be revalidated to the extent necessary
are primary or secondary metabolites of micro-organisms to demonstrate that no adverse change has occurred in the
such as bacteria, yeasts, fungi and micro-algae, whether or quality and safety of the product. Particular attention must
not modified by traditional procedures or recombinant DNA be given to possible changes in the impurity profile of the
(rDNA) technology. Such metabolites include vitamins, amino product if a modified or new micro-organism is introduced
acids, antibiotics, alkaloids and polysaccharides. into the process.
They may be obtained by batch or continuous fermentation
processes followed by procedures such as extraction, RAW MATERIALS
concentration, purification and isolation. The raw materials employed in the fermentation and/or
down-stream processing are of suitable quality for the
PRODUCTION intended purpose. They are tested to ensure that they comply
Production is based on a process that has been validated and with written specifications. Special attention must be paid to
shown to be suitable. The extent of validation depends on the the levels of free histidine in fish peptones as the presence
critical nature of the respective process step. of free histidine may lead to histamine formation in certain
conditions.
CHARACTERISATION OF THE PRODUCER
MICRO-ORGANISM Levels of bioburden in media or in the inlet air for aeration are
reduced to an adequately low level to ensure that if microbial
The history of the micro-organism used for production is contamination occurs, it does not adversely affect the quality,
documented. The micro-organism is adequately characterised. purity and safety of the product. Addition of components such
This may include determination of the phenotype of the as nutrients, precursors, and substrates during fermentation
micro-organism, macroscopic and microscopic methods and takes place aseptically.
biochemical tests and, if appropriate, determination of the
genotype of the micro-organism and molecular genetic tests. IN-PROCESS CONTROLS
PROCESSES USING A SEED-LOT SYSTEM In-process controls are in place to ensure the consistency of the
conditions during fermentation and down-stream processing
The master cell bank is a homogeneous suspension or
and of the quality of the isolated product. Particular attention
lyophilisate of the original cells distributed into individual
must be paid to ensure that any microbial contamination that
containers for storage. The viability and productivity of the
adversely affects the quality, purity and safety of the product is
cells under the selected storage conditions and their suitability
detected by the controls applied.
for initiating a satisfactory production process after storage
must be demonstrated. Production conditions may be monitored, as appropriate, by
Propagation of the master cell bank may take place through a suitable procedures for example to control and check :
seed-lot system that uses a working cell bank. – temperature,
The working cell bank is a homogeneous suspension or – pH,
lyophilisate of the cell material derived from the master cell – rate of aeration,
bank, distributed in equal volumes into individual containers
for storage (for example, in liquid nitrogen). – rate of agitation,
Production may take place by batch or continuous culture and – pressure,
may be terminated under defined conditions. and to monitor the concentration of the required product.

Products of fermentation EUROPEAN PHARMACOPOEIA
DOWN-STREAM PROCESSING – residues from the producer micro-organism, culture media,

substrates and precursors,
At the end of fermentation, the producer micro-organism
is inactivated or removed. Further processing is designed – unwanted transformation products of substrates and
to reduce residues originating from the culture medium to precursors.
an acceptable level and to ensure that the desired product is If necessary, suitable tests are performed either as in-process
recovered with consistent quality. controls or on the isolated product of fermentation.
Various purification processes may be used, for example, IDENTIFICATION, TESTS AND ASSAY
charcoal treatment, ultrafiltration and solvent extraction. It The requirements with which the product must comply
must be demonstrated that the process or processes chosen throughout its period of validity, as well as specific test
reduce to a minimum or remove : methods, are stated in the individual monographs.

EUROPEAN PHARMACOPOEIA Pharmaceutical preparations
04/2019:2619 (e.g. the prescribing practitioners and/or the preparing

pharmacists) have, within their area of responsibilities, a duty
of care to the patient receiving the pharmaceutical preparation.
In considering the preparation of an unlicensed pharmaceutical
preparation, a suitable level of risk assessment is undertaken.
PHARMACEUTICAL PREPARATIONS The risk assessment identifies :
– the criticality of different parameters (e.g. quality of
Pharmaceutica active substances, excipients and containers ; design
of the preparation process ; extent and significance of
INTRODUCTION testing ; stability of the preparation) to the quality of the
This monograph is intended to be a reference source preparation ; and
of standards in the European Pharmacopoeia on active
– the risk that the preparation may present to a particular
substances, excipients and dosage forms, which are to be
patient group.
applied in the manufacture/preparation of pharmaceuticals,
but not a guide on how to manufacture as there is specific Based on the risk assessment, the person responsible for the
guidance available covering methods of manufacture and preparation must ensure, with a suitable level of assurance,
associated controls. that the pharmaceutical preparation is, throughout its
It does not cover investigational medicinal products, but shelf-life, of an appropriate quality and suitable and fit for
competent authorities may refer to pharmacopoeial standards its purpose. For stock preparations, storage conditions and
when authorising clinical trials using investigational medicinal shelf-life have to be justified on the basis of, for example,
products. analytical data or professional judgement, which may be based
on literature references.
DEFINITION
PRODUCTION
Pharmaceutical preparations are medicinal products generally
consisting of active substances that may be combined with Manufacture/preparation must take place within the
excipients, formulated into a dosage form suitable for the framework of a suitable quality system and be compliant with
intended use, where necessary after reconstitution, presented the standards relevant to the type of product being made.
in a suitable and appropriately labelled container. Licensed products must comply with the requirements of
their licence. For unlicensed products a risk assessment as
Pharmaceutical preparations may be licensed by the competent
outlined in the section ‘Ethical considerations and guidance
authority, or unlicensed and made to the specific needs of
in the preparation of unlicensed pharmaceutical preparations’
patients according to legislation. There are 2 categories of
is of special importance, as these products are not previously
unlicensed pharmaceutical preparations :
assessed by the competent authority.
– extemporaneous preparations, i.e. pharmaceutical Where pharmaceutical preparations are manufactured/
preparations individually prepared for a specific patient or prepared using materials of human or animal origin,
patient group, supplied after preparation ; the general requirements of general chapters 5.1.7. Viral
– stock preparations, i.e. pharmaceutical preparations safety and 5.2.6. Evaluation of safety of veterinary vaccines
prepared in advance and stored until a request for a supply and immunosera and of the general monograph Products
is received. with risk of transmitting agents of animal spongiform
In addition to this monograph, pharmaceutical preparations encephalopathies (1483) apply, where appropriate.
also comply with the General Notices and with the relevant Formulation. During pharmaceutical development or prior
general chapters of the Pharmacopoeia. General chapters are to manufacture/preparation, suitable ingredients, processes,
normally given for information and become mandatory when tests and specifications are identified and justified in order to
referred to in a general or specific monograph, unless such ensure the suitability of the product for the intended purpose.
reference is made in a way that indicates that it is not the This includes consideration of the properties required in order
intention to make the text referred to mandatory but rather to to identify whether specific ingredient properties or process
cite it for information. steps are critical to the required quality of the pharmaceutical
Where relevant, pharmaceutical preparations also comply preparation.
with the dosage form monographs (e.g. Capsules (0016), Active substances and excipients. Active substances
Tablets (0478)) and general monographs relating to and excipients used in the formulation of pharmaceutical
pharmaceutical preparations (e.g. Allergen products (1063), preparations comply with the requirements of the relevant
Herbal teas (1435), Homoeopathic preparations (1038), general monographs, e.g. Substances for pharmaceutical
Homoeopathic pillules, coated (2786), Homoeopathic use (2034), Essential oils (2098), Herbal drug extracts (0765),
pillules, impregnated (2079), Immunosera for human use, Herbal drugs (1433), Herbal drug preparations (1434),
animal (0084), Immunosera for veterinary use (0030), Live Herbal drugs for homoeopathic preparations (2045), Mother
biotherapeutic products for human use (3053), Monoclonal tinctures for homoeopathic preparations (2029), Methods of
antibodies for human use (2031), Radiopharmaceutical preparation of homoeopathic stocks and potentisation (2371),
preparations (0125), Vaccines for human use (0153), Vaccines Products of fermentation (1468), Products of recombinant DNA
for veterinary use (0062)). technology (0784), Vegetable fatty oils (1579).
In addition, where specific monographs exist, the quality of
ETHICAL CONSIDERATIONS AND GUIDANCE IN THE the active substances and excipients used complies with the
PREPARATION OF UNLICENSED PHARMACEUTICAL corresponding monographs.
PREPARATIONS Where no specific monographs exist, the required quality
The underlying principle of legislation for pharmaceutical must be defined, taking into account the intended use and
preparations is that, subject to specific exemptions, no the involved risk.
pharmaceutical preparation may be placed on the market When physicochemical characteristics of active substances
without an appropriate marketing authorisation. The and functionality-related characteristics (FRCs) of excipients
exemptions from the formal licensing requirement allow (e.g. particle-size distribution, viscosity, polymorphism) are
the supply of unlicensed products to meet the special needs critical in relation to their role in the manufacturing process
of individual patients. However, when deciding to use an and quality attributes of the pharmaceutical preparation, they
unlicensed preparation all health professionals involved must be identified and controlled.

Pharmaceutical preparations EUROPEAN PHARMACOPOEIA
Detailed information on FRCs is given in general chapter 5.15. The following tests are applicable to many preparations and
Functionality-related characteristics of excipients. are therefore listed here.
Microbiological quality. The formulation of the Appearance. The appearance (e.g. size, shape and colour) of
pharmaceutical preparation and its container must ensure that the pharmaceutical preparation is controlled.
the microbiological quality is suitable for the intended use. Identity and purity tests. Where applicable, the following
During development, it shall be demonstrated that the tests are carried out on the pharmaceutical preparation:
antimicrobial activity of the preparation as such or, if – identification of the active substance(s) ;
necessary, with the addition of a suitable preservative or – identification of specific excipient(s), such as preservatives ;
preservatives, or by the selection of an appropriate container,
provides adequate protection from adverse effects that may – purity tests (e.g. investigation of degradation products,
arise from microbial contamination or proliferation during residual solvents (2.4.24) or other related impurities,
the storage and use of the preparation. A suitable test method sterility (2.6.1)) ;
together with criteria for evaluating the preservative properties – safety tests (e.g. safety tests for biological products).
of the formulation are provided in general chapter 5.1.3. Elemental impurities. General chapter 5.20. Elemental
Efficacy of antimicrobial preservation. impurities applies to pharmaceutical preparations except
If preparations do not have adequate antimicrobial efficacy products for veterinary use, unlicensed preparations and other
and do not contain antimicrobial preservatives they are products that are excluded from the scope of this chapter.
supplied in single-dose containers, or in multidose containers For pharmaceutical preparations outside the scope of general
that prevent microbial contamination of the contents after chapter 5.20, manufacturers of these products remain
opening. responsible for controlling the levels of elemental impurities
In the manufacture/preparation of non-sterile pharmaceutical using the principles of risk management.
preparations, suitable measures are taken to ensure their If appropriate, testing is performed using suitable analytical
microbial quality ; recommendations on this aspect are procedures according to general chapter 2.4.20. Determination
provided in general chapters 5.1.4. Microbiological quality of elemental impurities.
of non-sterile pharmaceutical preparations and substances Uniformity (2.9.40 or 2.9.5/2.9.6). Pharmaceutical
for pharmaceutical use and 5.1.8. Microbiological quality of preparations presented in single-dose units comply with the
herbal medicinal products for oral use and extracts used in test(s) as prescribed in the relevant specific dosage form
their preparation. monograph. If justified and authorised, general chapter 2.9.40
Sterile preparations are manufactured/prepared using can be applicable only at the time of release.
materials and methods designed to ensure sterility and to Special uniformity requirements apply in the following cases :
avoid the introduction of contaminants and the growth
of micro-organisms ; recommendations on this aspect are – for herbal drugs and herbal drug preparations, compliance
provided in general chapter 5.1.1. Methods of preparation of with general chapter 2.9.40 is not required ;
sterile products. – for homoeopathic preparations, the provisions of general
chapters 2.9.6 and 2.9.40 are normally not appropriate,
Containers. A suitable container is selected. Consideration is however in certain circumstances compliance with these
given to the intended use of the preparation, the properties of chapters may be required by the competent authority ;
the container, the required shelf-life, and product/container
incompatibilities. Where applicable, containers for – for single- and multivitamin and trace-element
pharmaceutical preparations comply with the requirements preparations, compliance with general chapters 2.9.6 and
for containers (3.2 and subsections) and materials used for the 2.9.40 (content uniformity only) is not required ;
manufacture of containers (3.1 and subsections). – in justified and authorised circumstances, for other
preparations, compliance with general chapters 2.9.6 and
Stability. Stability requirements of pharmaceutical 2.9.40 may not be required by the competent authority.
preparations are dependent on their intended use and on the
desired storage time. Reference standards. Reference standards may be needed
at various stages for quality control of pharmaceutical
Where applicable, the probability and criticality of possible
preparations. They are established and monitored taking due
degradation products of the active substance(s) and/or
account of general chapter 5.12. Reference standards.
reaction products of the active substance(s) with an excipient
and/or the immediate container must be assessed. Depending ASSAY
on the result of this assessment, limits of degradation and/or
reaction products are set and monitored in the pharmaceutical Unless otherwise justified and authorised, contents of active
preparation. Licensed products require a stability exercise. substances and specific excipients such as preservatives are
determined in pharmaceutical preparations. Limits must be
Methods used for the purpose of stability testing for all defined and justified.
relevant characteristics of the preparation are validated as Suitable and validated methods are used. If assay methods
stability indicating, i.e. the methods allow the quantification of prescribed in the respective active substance monographs are
the relevant degradation products and physical characteristic used, it must be demonstrated that they are not affected by the
changes. presence of the excipients and/or by the formulation.
TESTS Reference standards. See Tests.
Relevant tests to apply in order to ensure the appropriate LABELLING AND STORAGE
quality of a particular dosage form are described in the specific
dosage form monographs. The relevant labelling requirements given in the general
dosage form monographs apply. In addition, relevant
Where it is not practical, for unlicensed pharmaceutical European Union or other applicable regulations apply.
preparations, to carry out the tests (e.g. batch size, time
restraints), other suitable methods are implemented to ensure GLOSSARY
that the appropriate quality is achieved in accordance with the Formulation : the designing of an appropriate formula
risk assessment carried out and any local guidance or legal (including materials, processes, etc.) that will ensure that the
requirements. patient receives the suitable pharmaceutical preparation in an
Stock preparations are normally tested to a greater extent than appropriate form that has the required quality and that will be
extemporaneous preparations. stable and effective for the required length of time.

EUROPEAN PHARMACOPOEIA Pharmaceutical preparations
Licensed pharmaceutical preparation : a medicinal product Reconstitution : manipulation to enable the use or application
that has been granted a marketing authorisation by a of a medicinal product with a marketing authorisation in
competent authority. Synonym : authorised pharmaceutical accordance with the instructions given in the summary of
preparation. product characteristics or the patient information leaflet.
Manufacture : all operations of purchase of materials and Risk assessment : the identification of hazards and the analysis
products, Production, Quality Control, release, storage, and evaluation of risks associated with exposure to those
distribution of medicinal products and the related controls. hazards.
Preparation (of an unlicensed pharmaceutical Unlicensed pharmaceutical preparation : a medicinal
preparation) : the ‘manufacture’ of unlicensed pharmaceutical product that is exempt from the need of having a marketing
preparations by or at the request of pharmacies or other authorisation issued by a competent authority but is made for
healthcare establishments (the term ‘preparation’ is used specific patients’ needs according to legislation.
instead of ‘manufacture’ in order clearly to distinguish it
from the industrial manufacture of licensed pharmaceutical
preparations).

EUROPEAN PHARMACOPOEIA Capsules
04/2018:0016 Uniformity of mass (2.9.5). Capsules comply with the test

for uniformity of mass of single-dose preparations. If the
test for uniformity of content is prescribed for all the active
substances, the test for uniformity of mass is not required.
Dissolution. Unless otherwise justified and authorised,
CAPSULES a suitable test is carried out, for example one of the tests
described in general chapter 2.9.3. Dissolution test for solid
dosage forms.
Capsulae Where a dissolution test is prescribed, a disintegration test
may not be required.
The requirements of this monograph do not necessarily apply
to preparations that are presented as capsules intended for STORAGE
use other than by oral administration. Requirements for such Store at a temperature not exceeding 30 °C.
preparations may be found, where appropriate, in other general
monographs, for example Rectal preparations (1145) and LABELLING
Vaginal preparations (1164). The label states the name of any added preservative.
DEFINITION Hard capsules
Capsules are solid preparations with hard or soft shells
of various shapes and capacities, containing a single dose DEFINITION
of active substance(s). They are usually intended for oral Hard capsules are solid single-dose preparations. They consist
administration. of a hard shell, the capacity of which can be varied, containing
The capsule shells are made of gelatin or other substances, a solid, semi-solid or liquid preparation. The shell is made of
the consistency of which may be adjusted by the addition gelatin or other substances, and consists of 2 prefabricated
of substances such as glycerol or sorbitol. Excipients such cylindrical sections that are open at one end and rounded and
as surface-active agents, opaque fillers, antimicrobial closed at the other. The contents are filled into one of the
preservatives, sweeteners, colouring matter authorised by the sections and the capsule is closed by slipping the other section
competent authority and flavouring substances may be added. over it.
The capsules may bear surface markings. PRODUCTION
The contents of capsules may be solid, semi-solid or liquid. The active substance(s), usually in solid form (powder or
They consist of one or more active substances with or granules), are filled into one of the sections that is then closed
without excipients such as solvents, diluents, lubricants and by slipping the other section over it. The security of the
disintegrating agents. The contents do not cause deterioration closure may be strengthened by suitable means.
of the shell. The shell, however, is attacked by the digestive
fluids and the contents are released. TESTS
Where applicable, containers for capsules comply with Disintegration (2.9.1). Hard capsules comply with the
the requirements of Materials used for the manufacture of test. Use water R as the liquid medium. When justified and
containers (3.1 and subsections) and Containers (3.2 and authorised, 0.1 M hydrochloric acid or artificial gastric juice R
subsections). may be used as the liquid medium. If the capsules float
on the surface of the water, a disc may be added. Operate
Several categories of capsules may be distinguished : the apparatus for 30 min, unless otherwise justified and
– hard capsules ; authorised.
– soft capsules ;
– gastro-resistant capsules ; Soft capsules
– modified-release capsules ; DEFINITION
– cachets. Soft capsules are solid single-dose preparations. They consist
of a soft shell, the capacity and shape of which can be varied,
PRODUCTION containing a semi-solid or liquid preparation. The shell is
made of gelatin or other substances and may have one or
In the manufacture, packaging, storage and distribution more solid active substances incorporated into it. The shell
of capsules, suitable measures are taken to ensure their is usually thicker than that of a hard capsule and consists
microbial quality ; recommendations on this aspect are of one part, as soft capsules are generally formed, filled and
provided in general chapter 5.1.4. Microbiological quality of sealed in a single operation.
non-sterile pharmaceutical preparations and substances for
pharmaceutical use. PRODUCTION
Soft capsules are usually formed, filled and sealed in one
TESTS operation, but for extemporaneous use the shell may be
Uniformity of dosage units. Capsules comply with the test prefabricated. The shell material may contain an active
for uniformity of dosage units (2.9.40) or, where justified and substance.
authorised, with the tests for uniformity of content and/or Liquids may be enclosed directly ; solids are usually dissolved
uniformity of mass shown below. Herbal drugs and herbal or dispersed in a suitable vehicle to give a solution or
drug preparations present in the dosage form are not subject dispersion of a paste-like consistency.
to the provisions of this paragraph. There may be partial migration of the constituents from the
Uniformity of content (2.9.6). Unless otherwise prescribed capsule contents into the shell and vice versa because of the
or justified and authorised, capsules with a content of active nature of the materials and the surfaces in contact.
substance less than 2 mg or less than 2 per cent of the fill mass
comply with test B for uniformity of content of single-dose TESTS
preparations. If the preparation has more than one active Disintegration (2.9.1). Soft capsules comply with the test. Use
substance, the requirement applies only to those ingredients water R as the liquid medium. When justified and authorised,
that correspond to the above conditions. 0.1 M hydrochloric acid or artificial gastric juice R may be used

Capsules EUROPEAN PHARMACOPOEIA
as the liquid medium. Add a disc to each tube. Liquid active TESTS
substances dispensed in soft capsules may attack the disc ; in Disintegration (2.9.1). Capsules with a gastro-resistant shell
such circumstances and where authorised, the disc may be comply with the test with the following modifications. Use
omitted. Operate the apparatus for 30 min, unless otherwise 0.1 M hydrochloric acid as the liquid medium and operate the
justified and authorised. If the capsules fail to comply because apparatus for 2 h, or other such time as may be authorised,
of adherence to the discs, the results are invalid. Repeat the without the discs. Examine the state of the capsules. The
test on a further 6 capsules omitting the discs. time of resistance to the acid medium varies according to the
formulation of the capsules to be examined. It is typically
Modified-release capsules 2 h to 3 h but even with authorised deviations it must not
be less than 1 h. No capsule shows signs of disintegration or
DEFINITION rupture permitting the escape of the contents. Replace the
Modified-release capsules are hard or soft capsules in which acid by phosphate buffer solution pH 6.8 R. When justified and
the contents or the shell or both contain special excipients authorised, a buffer solution of pH 6.8 with added pancreas
or are prepared by a special process designed to modify the powder (for example, 0.35 g of pancreas powder R per 100 mL
rate, the place or the time at which the active substance(s) of buffer solution) may be used. Add a disc to each tube.
are released. Operate the apparatus for 60 min. If the capsules fail to comply
Modified-release capsules include prolonged-release capsules, because of adherence to the discs, the results are invalid.
delayed-release capsules and pulsatile-release capsules. Repeat the test on a further 6 capsules omitting the discs.
Gastro-resistant capsules Cachets

DEFINITION DEFINITION
Gastro-resistant capsules are delayed-release capsules that are Cachets are solid preparations consisting of a hard shell
intended to resist the gastric fluid and to release their active containing a single dose of one or more active substances. The
substance or substances in the intestinal fluid. Usually they cachet shell is made of unleavened bread usually from rice
are prepared by filling capsule shells with granules or with flour and consists of 2 prefabricated flat cylindrical sections.
particles covered with a gastro-resistant coating. In other Before administration, the cachets are immersed in water for
cases, the shell of the capsule is covered with a gastro-resistant a few seconds, placed on the tongue and swallowed with a
coating or the shell itself has gastro-resistant properties. draught of water.

EUROPEAN PHARMACOPOEIA Eye preparations
01/2008:1163 Eye drops may contain excipients, for example, to adjust the
corrected 10.0 tonicity or the viscosity of the preparation, to adjust or stabilise
the pH, to increase the solubility of the active substance, or to
stabilise the preparation. These substances do not adversely
affect the intended medicinal action or, at the concentrations
used, cause undue local irritation.
EYE PREPARATIONS Aqueous preparations supplied in multidose containers
contain a suitable antimicrobial preservative in appropriate
concentration except when the preparation itself has adequate
Ophthalmica antimicrobial properties. The antimicrobial preservative
DEFINITION chosen must be compatible with the other ingredients of the
preparation and must remain effective throughout the period
Eye preparations are sterile liquid, semi-solid or solid of time during which eye drops are in use.
preparations intended for administration upon the eyeball
and/or to the conjunctiva, or for insertion in the conjunctival If eye drops do not contain antimicrobial preservatives
sac. they are supplied in single-dose containers or in multidose
containers preventing microbial contamination of the contents
Where applicable, containers for eye preparations comply after opening.
with the requirements of materials used for the manufacture
of containers (3.1 and subsections) and containers (3.2 and Eye drops intended for use in surgical procedures do not
subsections). contain antimicrobial preservatives.
Several categories of eye preparations may be distinguished : Eye drops that are solutions, examined under suitable
conditions of visibility, are practically clear and practically
– eye drops ; free from particles.
– eye lotions ; Eye drops that are suspensions may show a sediment that is
– powders for eye drops and powders for eye lotions ; readily redispersed on shaking to give a suspension which
– semi-solid eye preparations ; remains sufficiently stable to enable the correct dose to be
delivered.
– ophthalmic inserts.
Multidose preparations are supplied in containers that allow
PRODUCTION successive drops of the preparation to be administered. The
During the development of an eye preparation whose containers contain at most 10 mL of the preparation, unless
formulation contains an antimicrobial preservative, the otherwise justified and authorised.
necessity for and the efficacy of the chosen preservative shall TESTS
be demonstrated to the satisfaction of the competent authority.
A suitable test method together with criteria for judging the Particle size. Unless otherwise justified and authorised,
preservative properties of the formulation are provided in eye drops in the form of a suspension comply with the
chapter 5.1.3. Efficacy of antimicrobial preservation. following test : introduce a suitable quantity of the suspension
into a counting cell or with a micropipette onto a slide,
Eye preparations are prepared using materials and methods as appropriate, and scan under a microscope an area
designed to ensure sterility and to avoid the introduction corresponding to 10 μg of the solid phase. For practical
of contaminants and the growth of micro-organisms ; reasons, it is recommended that the whole sample is first
recommendations on this aspect are provided in chapter 5.1.1. scanned at low magnification (e.g. × 50) and particles greater
Methods of preparation of sterile products. than 25 μm are identified. These larger particles can then be
In the manufacture of eye preparations containing dispersed measured at a larger magnification (e.g. × 200 to × 500). For
particles, measures are taken to ensure a suitable and each 10 µg of solid active substance, not more than 20 particles
controlled particle size with regard to the intended use. have a maximum dimension greater than 25 µm, and not more
During development, it must be demonstrated that the than 2 of these particles have a maximum dimension greater
nominal contents can be withdrawn from the container of than 50 µm. None of the particles has a maximum dimension
liquid and semi-solid eye preparations supplied in single-dose greater than 90 µm.
containers.
LABELLING
TESTS The label states, for multidose containers, the period after
Sterility (2.6.1). Eye preparations comply with the test. opening the container after which the contents must not be
Applicators supplied separately also comply with the test. used. This period does not exceed 4 weeks, unless otherwise
Remove the applicator with aseptic precautions from its justified and authorised.
package and transfer it to a tube of culture medium so that it
is completely immersed. Incubate and interpret the results Eye lotions
as described in the test.
DEFINITION
STORAGE Eye lotions are sterile aqueous solutions intended for use in
Unless otherwise justified and authorised, store in a sterile, rinsing or bathing the eye or for impregnating eye dressings.
tamper-evident container. Eye lotions may contain excipients, for example to adjust the
tonicity or the viscosity of the preparation or to adjust or
LABELLING stabilise the pH. These substances do not adversely affect the
The label states the name of any added antimicrobial intended action or, at the concentrations used, cause undue
preservative. local irritation.
Eye lotions supplied in multidose containers contain a suitable
Eye drops antimicrobial preservative in appropriate concentration
except when the preparation itself has adequate antimicrobial
DEFINITION properties. The antimicrobial preservative chosen is
Eye drops are sterile aqueous or oily solutions, emulsions or compatible with the other ingredients of the preparation and
suspensions of one or more active substances intended for remains effective throughout the period of time during which
instillation into the eye. the eye lotions are in use.

Eye preparations EUROPEAN PHARMACOPOEIA
If eye lotions do not contain antimicrobial preservatives, they Semi-solid eye preparations are packed in small, sterilised
are supplied in single-dose containers. Eye lotions intended collapsible tubes fitted or provided with a sterilised cannula.
for use in surgical procedures or in first-aid treatment do not The containers contain at most 10 g of the preparation,
contain an antimicrobial preservative and are supplied in unless otherwise justified and authorised. The tubes must be
single-dose containers. well-closed to prevent microbial contamination. Semi-solid
Eye lotions, examined under suitable conditions of visibility, eye preparations may also be packed in suitably designed
are practically clear and practically free from particles. single-dose containers. The containers, or the nozzles of tubes,
The containers for multidose preparations do not contain are of such a shape as to facilitate administration without
more than 200 mL of eye lotion, unless otherwise justified and contamination.
authorised.
TESTS
LABELLING Particle size. Semi-solid eye preparations containing
The label states : dispersed solid particles comply with the following test :
– where applicable, that the contents are to be used on one spread gently a quantity of the preparation corresponding
occasion only ; to at least 10 µg of solid active substance as a thin layer.
– for multidose containers, the period after opening the Scan under a microscope the whole area of the sample. For
container after which the contents must not be used ; this practical reasons, it is recommended that the whole sample is
period does not exceed 4 weeks, unless otherwise justified first scanned at a small magnification (e.g. × 50) and particles
and authorised. greater than 25 µm are identified. These larger particles can
then be measured at a larger magnification (e.g. × 200 to
× 500). For each 10 μg of solid active substance, not more than
Powders for eye drops and powders 20 particles have a maximum dimension greater than 25 µm,
for eye lotions and not more than 2 of these particles have a maximum
dimension greater than 50 µm. None of the particles has a
DEFINITION maximum dimension greater than 90 µm.
Powders for the preparation of eye drops and eye lotions are
supplied in a dry, sterile form to be dissolved or suspended in LABELLING
an appropriate liquid vehicle at the time of administration. The label states, for multidose containers, the period after
They may contain excipients to facilitate dissolution or opening the container after which the contents must not be
dispersion, to prevent caking, to adjust the tonicity, to adjust used. This period does not exceed 4 weeks, unless otherwise
or stabilise the pH or to stabilise the preparation. justified and authorised.
After dissolution or suspension in the prescribed liquid, they
comply with the requirements for eye drops or eye lotions,
as appropriate. Ophthalmic inserts
TESTS DEFINITION
Uniformity of dosage units (2.9.40). Single-dose powders Ophthalmic inserts are sterile, solid or semi-solid preparations
for eye drops and eye lotions comply with the test or, where of suitable size and shape, designed to be inserted in the
justified and authorised, with the tests for uniformity of conjunctival sac, to produce an ocular effect. They generally
content and/or uniformity of mass shown below. Herbal drugs consist of a reservoir of active substance embedded in a matrix
and herbal drug preparations present in the dosage form are or bounded by a rate-controlling membrane. The active
not subject to the provisions of this paragraph. substance, which is more or less soluble in lacrymal liquid, is
Uniformity of content (2.9.6). Unless otherwise prescribed released over a determined period of time.
or justified and authorised, single-dose powders for eye Ophthalmic inserts are individually distributed into sterile
drops and eye lotions with a content of active substance less containers.
then 2 mg or less than 2 per cent of the total mass comply
with test B. If the preparation has more than one active PRODUCTION
substance, the requirement applies only to those substances In the manufacture of ophthalmic inserts, measures are taken
that correspond to the above condition. to ensure a suitable dissolution behaviour.
Uniformity of mass (2.9.5). Single-dose powders for eye drops
and eye lotions comply with the test. If the test for uniformity TESTS
of content is prescribed for all the active substances, the test Uniformity of dosage units (2.9.40). Ophthalmic inserts
for uniformity of mass is not required. comply with the test or, where justified and authorised, with
the test for uniformity of content shown below. Herbal drugs
Semi-solid eye preparations and herbal drug preparations present in the dosage form are
not subject to the provisions of this paragraph.
DEFINITION
Uniformity of content (2.9.6). Ophthalmic inserts comply,
Semi-solid eye preparations are sterile ointments, creams where applicable, with test A.
or gels intended for application to the conjunctiva or to the
eyelids. They contain one or more active substances dissolved LABELLING
or dispersed in a suitable basis. They have a homogeneous
appearance. The label states :
Semi-solid eye preparations comply with the requirements – where applicable, the total quantity of active substance per
of the monograph Semi-solid preparations for cutaneous insert ;
application (0132). The basis is non-irritant to the conjunctiva. – where applicable, the dose released per unit time.

EUROPEAN PHARMACOPOEIA Liquid preparations for oral use
01/2018:0672 In the manufacture of liquid preparations for oral use

containing dispersed particles, measures are taken to ensure
a suitable and controlled particle size with regard to the
intended use.
TESTS
LIQUID PREPARATIONS FOR
Uniformity of dosage units. Solutions, suspensions and
ORAL USE emulsions in single-dose containers comply with the test
for uniformity of dosage units (2.9.40) or, where justified
Praeparationes liquidae peroraliae and authorised, with the test for uniformity of content or
uniformity of mass shown below. Herbal drugs and herbal
Where justified and authorised, the requirements of this drug preparations present in the dosage form are not subject
monograph do not apply to liquid preparations for oral use to the provisions of this paragraph.
intended for veterinary use.
Uniformity of content (2.9.6). Unless otherwise prescribed
DEFINITION or justified and authorised, single-dose preparations that are
Liquid preparations for oral use are usually solutions, suspensions or emulsions comply with the following test.
emulsions or suspensions containing one or more active After shaking, empty each container as completely as possible
substances in a suitable vehicle ; they may, however, consist of and carry out the test on the individual contents. They
liquid active substances used as such (oral liquids). comply with test B for uniformity of content of single-dose
preparations.
Some preparations for oral use are prepared by dilution of
concentrated liquid preparations, or from powders or granules Uniformity of mass. Single-dose preparations that are
for the preparation of oral solutions or suspensions, for oral solutions comply with the following test : weigh individually
drops or for syrups, using a suitable vehicle. the contents of 20 containers, emptied as completely as
possible, and determine the average mass. Not more than 2 of
The vehicle for any preparation for oral use is chosen having the individual masses deviate by more than 10 per cent from
regard to the nature of the active substance(s) and to provide the average mass and none deviate by more than 20 per cent.
organoleptic characteristics appropriate to the intended use of
the preparation. Dose and uniformity of dose of oral drops. Into a suitable
Liquid preparations for oral use may contain suitable graduated cylinder, introduce by means of the dropping
antimicrobial preservatives, antioxidants and other excipients device the number of drops usually prescribed for one dose,
such as dispersing, suspending, thickening, emulsifying, or introduce by means of the measuring device the usually
buffering, wetting, solubilising, stabilising, flavouring and prescribed quantity. The dropping speed does not exceed
sweetening agents and colouring matter, authorised by the 2 drops per second. Weigh the liquid, repeat the addition,
competent authority. weigh again and carry on repeating the addition and weighing
until a total of 10 masses are obtained. No single mass deviates
Emulsions may show evidence of phase separation but are by more than 10 per cent from the average mass.
readily redispersed on shaking. Suspensions may show a
sediment, which is readily dispersed on shaking to give a If the labelling defines a mass, the total of 10 masses does
suspension that remains sufficiently stable to enable the not differ by more than 15 per cent from the nominal mass
correct dose to be delivered. of 10 doses defined on the labelling. If the labelling defines a
volume, measure the total volume of 10 doses. The volume
Where applicable, containers for liquid preparations for does not differ by more than 15 per cent from the nominal
oral use comply with the requirements of Materials used volume of 10 doses defined on the labelling.
for the manufacture of containers (3.1 and subsections) and
Containers (3.2 and subsections). Uniformity of mass of delivered doses from multidose
containers (2.9.27). Liquid preparations for oral use supplied
Several categories of preparations may be distinguished ;
in multidose containers comply with the test. Oral drops are
– oral solutions, emulsions and suspensions ; not subject to the provisions of this test.
– powders and granules for oral solutions and suspensions ;
– oral drops ; LABELLING
– powders for oral drops ; The label states the name of any added preservative.
– syrups ;
– powders and granules for syrups. Oral solutions, emulsions and suspensions
PRODUCTION DEFINITION
During development of a preparation for oral use whose Oral solutions, emulsions and suspensions are supplied
formulation contains an antimicrobial preservative, the in single-dose or multidose containers. Each dose from a
need for and the efficacy of the chosen preservative shall be multidose container is administered by means of a device
demonstrated to the satisfaction of the competent authority. suitable for measuring the prescribed volume. The device is
A suitable test method together with criteria for judging the usually a spoon or a cup for volumes of 5 mL or multiples
preservative properties of the formulation are provided in thereof or an oral syringe for other volumes.
general chapter 5.1.3. Efficacy of antimicrobial preservation.
During development, it must be demonstrated that the
nominal content can be withdrawn from the container, for
Powders and granules for oral solutions and
liquid preparations for oral use presented in single-dose suspensions
containers.
In the manufacturing, packaging, storage and distribution of DEFINITION
liquid preparations for oral use, suitable measures are taken Powders and granules for the preparation of oral solutions
to ensure their microbial quality ; recommendations on this or suspensions are intended to be reconstituted with the
aspect are provided in general chapter 5.1.4. Microbiological prescribed liquid to produce a liquid preparation for oral
quality of non-sterile pharmaceutical preparations and use. They may contain excipients, in particular to facilitate
substances for pharmaceutical use. dispersion or dissolution and to prevent caking.

Liquid preparations for oral use EUROPEAN PHARMACOPOEIA
After dissolution or suspension, they comply with the Uniformity of content (2.9.6). Unless otherwise prescribed
requirements for oral solutions or oral suspensions, as or justified and authorised, single-dose powders for oral drops
appropriate. with a content of active substance less than 2 mg or less than
2 per cent of the total mass comply with test B for uniformity
TESTS of content of single-dose preparations. If the preparation has
Uniformity of dosage units. Single-dose powders and more than one active substance, the requirement applies only
single-dose granules comply with the test for uniformity of to those substances that correspond to the above conditions.
dosage units (2.9.40) or, where justified and authorised, with Uniformity of mass (2.9.5). Single-dose powders for
the tests for uniformity of content and/or uniformity of mass oral drops comply with the test for uniformity of mass
shown below. Herbal drugs and herbal drug preparations of single-dose preparations. If the test for uniformity of
present in the dosage form are not subject to the provisions content is prescribed for all the active substances, the test for
of this paragraph. uniformity of mass is not required.
Uniformity of content (2.9.6). Unless otherwise prescribed or
justified and authorised, single-dose powders and single-dose Syrups
granules with a content of active substance less than 2 mg
or less than 2 per cent of the total mass comply with test B DEFINITION
for uniformity of content of single-dose preparations. If
Syrups are aqueous preparations characterised by a sweet
the preparation has more than one active substance, the
taste and a viscous consistency. They may contain sucrose
requirement applies only to those substances that correspond
at a concentration of at least 45 per cent m/m. The sweet
to the above conditions.
taste can also be obtained by using other polyols or
Uniformity of mass (2.9.5). Single-dose powders and sweetening agents. Syrups usually contain aromatic or other
single-dose granules comply with the test for uniformity of flavouring agents. Each dose from a multidose container is
mass of single-dose preparations. If the test for uniformity of administered by means of a device suitable for measuring the
content is prescribed for all the active substances, the test for prescribed volume. The device is usually a spoon or a cup
uniformity of mass is not required. for volumes of 5 mL or multiples thereof.
LABELLING LABELLING
The label states : The label states the name and concentration of the polyol or
– the method of preparation of the solution or suspension ; sweetening agent.
– the conditions and the duration of storage after
reconstitution. Powders and granules for syrups
Oral drops DEFINITION
Powders and granules for syrups are intended to be
DEFINITION reconstituted with the prescribed liquid to produce a liquid
Oral drops are solutions, emulsions or suspensions that are preparation for oral use. They may contain excipients to
administered in small volumes such as drops by means of facilitate dissolution.
a suitable device. After dissolution, they comply with the requirements for
LABELLING syrups.
The label states the number of drops per millilitre of TESTS
preparation or per gram of preparation if the dose is measured
in drops. Uniformity of dosage units. Single-dose powders and
granules for syrups comply with the test for uniformity of
dosage units (2.9.40) or, where justified and authorised, with
Powders for oral drops the tests for uniformity of content and/or uniformity of mass
DEFINITION shown below. Herbal drugs and herbal drug preparations
present in the dosage form are not subject to the provisions
Powders for the preparation of oral drops are intended to be
of this paragraph.
reconstituted with the prescribed liquid to produce a liquid
preparation for oral use. They may contain excipients to Uniformity of content (2.9.6). Unless otherwise prescribed
facilitate dissolution or suspension in the prescribed liquid or or justified and authorised, single-dose powders and granules
to prevent caking. for syrups with a content of active substance less than 2 mg
After dissolution or suspension, they comply with the or less than 2 per cent of the total mass comply with test B
requirements for oral drops. for uniformity of content of single-dose preparations. If
the preparation has more than one active substance, the
TESTS requirement applies only to those substances that correspond
Uniformity of dosage units. Single-dose powders for oral to the above conditions.
drops comply with the test for uniformity of dosage units Uniformity of mass (2.9.5). Single-dose powders and
(2.9.40) or, where justified and authorised, with the tests for granules for syrups comply with the test for uniformity of
uniformity of content and/or uniformity of mass shown below. mass of single-dose preparations. If the test for uniformity of
Herbal drugs and herbal drug preparations present in the content is prescribed for all the active substances, the test for
dosage form are not subject to the provisions of this paragraph. uniformity of mass is not required.

EUROPEAN PHARMACOPOEIA Parenteral preparations
04/2015:0520 Parenteral preparations are prepared using materials

corrected 10.0 and methods designed to ensure sterility and to avoid
the introduction of contaminants and the growth of
micro-organisms. Recommendations on this aspect are
provided in the text Methods of preparation of sterile products
(5.1.1).
PARENTERAL PREPARATIONS Water used in the manufacture of parenteral preparations
complies with the requirements of water for injections in bulk
stated in the monograph Water for injections (0169).
Parenteralia
TESTS
The requirements of this monograph do not necessarily apply
to products derived from human blood, to immunological Particulate contamination : sub-visible particles (2.9.19).
preparations, or radiopharmaceutical preparations. Special For preparations for human use, solutions for infusion or
requirements may apply to preparations for veterinary use solutions for injection comply with the test.
depending on the species of animal for which the preparation is In the case of preparations for subcutaneous or
intended. intramuscular injection, higher limits may be appropriate.
Radiopharmaceutical preparations are exempt from these
DEFINITION requirements. Preparations for which the label states that the
Parenteral preparations are sterile preparations intended for product is to be used with a final filter are exempt from these
administration by injection, infusion or implantation into the requirements, providing it has been demonstrated that the
human or animal body. filter delivers a solution that complies with the test.
Parenteral preparations may require the use of excipients, For preparations for veterinary use, when supplied in
for example to make the preparation isotonic with respect containers with a nominal content of more than 100 mL
to blood, to adjust the pH, to increase solubility, to prevent and when the content is equivalent to a dose of more than
deterioration of the active substances or to provide adequate 1.4 mL per kilogram of body mass, solutions for infusion or
antimicrobial properties, but not to adversely affect the solutions for injection comply with the test for particulate
intended medicinal action of the preparation or, at the contamination : sub-visible particles.
concentrations used, to cause toxicity or undue local irritation. Sterility (2.6.1). Parenteral preparations comply with the test
Containers for parenteral preparations are made as far as for sterility.
possible from materials that are sufficiently transparent
to permit the visual inspection of the contents, except for STORAGE
implants and in other justified and authorised cases. In a sterile, airtight, tamper-evident container.
Where applicable, the containers for parenteral preparations
comply with the requirements for Materials used for LABELLING
the manufacture of containers (3.1 and subsections) and The label states :
Containers (3.2 and subsections). – the name and concentration of any added antimicrobial
Parenteral preparations intended for chronic use or total preservative ;
parenteral nutrition should have appropriate limits for specific – where applicable, that the solution is to be used in
components or elements, taking long-term toxicity into conjunction with a final filter ;
account. – where applicable, that the preparation is free from bacterial
Parenteral preparations are supplied in glass containers endotoxins or that it is apyrogenic.
(3.2.1) or in other containers such as plastic containers (3.2.2,
3.2.2.1 and 3.2.9) and prefilled syringes. The tightness of the
container is ensured by suitable means. Closures ensure a Injections
good seal, prevent the access of micro-organisms and other DEFINITION
contaminants and usually permit the withdrawal of a part or
the whole of the contents without removal of the closure. The Injections are sterile solutions, emulsions or suspensions.
plastic materials or elastomers (3.2.9) used to manufacture the They are prepared by dissolving, emulsifying or suspending
closures are sufficiently firm and elastic to allow the passage of the active substance(s) and any added excipients in water, in
a needle with the least possible shedding of particles. Closures a suitable non-aqueous liquid, that may be non-sterile where
for multidose containers are sufficiently elastic to ensure that justified, or in a mixture of these vehicles.
the puncture is resealed when the needle is withdrawn. Solutions for injection, examined under suitable conditions of
Several categories of parenteral preparations may be visibility, are clear and practically free from particles.
distinguished : Emulsions for injection do not show any evidence of phase
– injections ; separation. Suspensions for injection may show a sediment
which is readily dispersed on shaking to give a suspension
– infusions ;
which remains sufficiently stable to enable the correct dose
– concentrates for injections or infusions ; to be withdrawn.
– powders for injections or infusions ; Multidose preparations. Multidose aqueous injections contain
– gels for injections ; a suitable antimicrobial preservative at an appropriate
– implants. concentration except when the preparation itself has adequate
antimicrobial properties. When a preparation for parenteral
PRODUCTION administration is presented in a multidose container, the
During the development of a parenteral preparation, the precautions to be taken for its administration and more
formulation for which contains an antimicrobial preservative, particularly for its storage between successive withdrawals
the effectiveness of the chosen preservative shall be are given.
demonstrated to the satisfaction of the competent authority. Antimicrobial preservatives. Aqueous preparations which
A suitable test method together with criteria for judging the are prepared using aseptic precautions and which cannot be
preservative properties of the formulation are provided under terminally sterilised may contain a suitable antimicrobial
Efficacy of antimicrobial preservation (5.1.3). preservative in an appropriate concentration.

Parenteral preparations EUROPEAN PHARMACOPOEIA
No antimicrobial preservative is added when : The volume of the infusion in the container is sufficient to
– the volume to be injected in a single dose exceeds 15 mL, permit the withdrawal and administration of the nominal dose
unless otherwise justified ; using a normal technique (2.9.17).
– the preparation is intended for administration by TESTS
routes where, for medical reasons, an antimicrobial
preservative is not acceptable, such as intracisternally, Bacterial endotoxins - pyrogens. They comply with a test
epidurally, intrathecally or by any route giving access to the for bacterial endotoxins (2.6.14) or, where justified and
cerebrospinal fluid, or intra- or retro-ocularly. authorised, with the test for pyrogens (2.6.8). For the latter
test inject 10 mL per kilogram of body mass into each rabbit,
Such preparations are presented in single-dose containers. unless otherwise justified and authorised.
PRODUCTION
In the manufacture of injections containing dispersed Concentrates for injections or infusions
particles, measures are taken to ensure a suitable and DEFINITION
controlled particle size with regard to the intended use.
Concentrates for injections or infusions are sterile solutions
Single-dose preparations. The volume of the injection in a intended for injection or infusion after dilution. They are
single-dose container is sufficient to permit the withdrawal diluted to a prescribed volume with a prescribed liquid
and administration of the nominal dose using a normal before administration. After dilution, they comply with the
technique (2.9.17). requirements for injections or for infusions.
TESTS TESTS
Uniformity of dosage units. Single-dose suspensions for Bacterial endotoxins - pyrogens. They comply with the
injection comply with the test for uniformity of dosage units requirements prescribed for injections or for infusions, after
(2.9.40) or, where justified and authorised, with the test for dilution to a suitable volume.
uniformity of content shown below. Herbal drugs and herbal
drug preparations present in the dosage form are not subject
to the provisions of this paragraph. Powders for injections or infusions
Uniformity of content (2.9.6). Unless otherwise prescribed or DEFINITION
justified and authorised, single-dose suspensions for injection
with a content of active substance less than 2 mg or less than Powders for injections or infusions are solid, sterile substances
2 per cent of the total mass comply with test A for uniformity distributed in their final containers and which, when shaken
of content of single-dose preparations. If the preparation with the prescribed volume of a prescribed sterile liquid
contains more than one active substance, the requirement rapidly form either clear and practically particle-free solutions
applies only to those substances that correspond to the above or uniform suspensions. After dissolution or suspension, they
conditions. comply with the requirements for injections or for infusions.
Freeze-dried products for parenteral administration are
Bacterial endotoxins - pyrogens. A test for bacterial
considered as powders for injections or infusions.
endotoxins (2.6.14) is carried out or, where justified and
authorised, the test for pyrogens (2.6.8). Recommendations PRODUCTION
on the limits for bacterial endotoxins are given in general
chapter 5.1.10. The uniformity of content and uniformity of mass of
freeze-dried products for parenteral administration are
Preparations for human use. The preparation complies with ensured by the in-process control of the amount of the
a test for bacterial endotoxins (2.6.14) or with the test for solution prior to freeze-drying.
pyrogens (2.6.8).
Preparations for veterinary use. When the volume to be TESTS
injected in a single dose is 15 mL or more and is equivalent Uniformity of dosage units. Powders for injections or
to a dose of 0.2 mL or more per kilogram of body mass, the infusions comply with the test for uniformity of dosage units
preparation complies with a test for bacterial endotoxins (2.9.40) or, where justified and authorised, with the tests for
(2.6.14) or with the test for pyrogens (2.6.8). uniformity of content and/or uniformity of mass shown below.
Any preparation. Where the label states that the preparation Herbal drugs and herbal drug preparations present in the
is free from bacterial endotoxins or apyrogenic, respectively, dosage form are not subject to the provisions of this paragraph.
the preparation complies with a test for bacterial endotoxins Uniformity of content (2.9.6). Unless otherwise prescribed
(2.6.14) or with the test for pyrogens (2.6.8), respectively. or justified and authorised, powders for injections or infusions
with a content of active substance less than 2 mg or less than
Infusions 2 per cent of the total mass, or with a unit mass equal to or
less than 40 mg comply with test A for uniformity of content
DEFINITION of single-dose preparations. If the preparation contains more
Infusions are sterile, aqueous solutions or emulsions with than one active substance, the requirement applies only to
water as the continuous phase. They are usually made isotonic those substances that correspond to the above conditions.
with respect to blood. They are principally intended for Uniformity of mass (2.9.5). Powders for injections or
administration in large volume. Infusions do not contain any infusions comply with the test for uniformity of mass of
added antimicrobial preservative. single-dose preparations. If the test for uniformity of content is
Solutions for infusion, examined under suitable conditions of prescribed for all the active substances, the test for uniformity
visibility, are clear and practically free from particles. of mass is not required.
Emulsions for infusion do not show any evidence of phase Bacterial endotoxins - pyrogens. They comply with the
separation. requirements prescribed for injections or for infusions, after
dissolution or suspension in a suitable volume of liquid.
PRODUCTION
In the manufacture of infusions containing dispersed particles, LABELLING
measures are taken to ensure a suitable and controlled particle The label states the instructions for the preparation of
size with regard to the intended use. injections and infusions.

EUROPEAN PHARMACOPOEIA Parenteral preparations
Gels for injections TESTS

DEFINITION A suitable test is carried out to demonstrate the appropriate
release of the active substance(s).
Gels for injections are sterile gels with a viscosity suitable to
guarantee a modified release of the active substance(s) at the
site of injection.
Implants
DEFINITION
Implants are sterile, solid preparations of a size and shape
suitable for parenteral implantation and release of the active
substance(s) over an extended period of time. Each dose is
provided in a sterile container.

EUROPEAN PHARMACOPOEIA Tablets
01/2018:0478 provided in general chapter 5.1.4. Microbiological quality of

non-sterile pharmaceutical preparations and substances for
pharmaceutical use.
Subdivision of tablets. Tablets may bear a break-mark or
break-marks for the purpose of being subdivided into parts,
TABLETS either to ease the intake of the medicinal product or to
deliver fractional doses. In cases where fractions of tablets
are necessary to deliver the intended dose stated on the label,
Compressi the efficacy of the breakmark is assessed during product
The requirements of this monograph do not necessarily apply development or for validation purposes by determining
to preparations that are presented as tablets intended for use the uniformity of mass of the sub-divided parts using the
other than by oral administration. Requirements for such following test.
preparations may be found, where appropriate, in other general Take 30 tablets at random, break them by hand and, from all
monographs ; for example Rectal preparations (1145), Vaginal the parts obtained from 1 tablet, take 1 part for the test and
preparations (1164) and Oromucosal preparations (1807). reject the other part(s). Weigh each of the 30 parts individually
This monograph does not apply to lozenges, oral pastes and and calculate the average mass. The tablets comply with the
oral gums. Where justified and authorised, the requirements test if not more than 1 individual mass is outside the limits of
of this monograph do not apply to tablets for veterinary use. 85 per cent to 115 per cent of the average mass. The tablets
Tablets for use in the mouth comply with the requirements of fail to comply with the test if more than 1 individual mass
the monograph Oromucosal preparations (1807). is outside these limits, or if 1 individual mass is outside the
limits of 75 per cent to 125 per cent of the average mass.
DEFINITION
Tablets are solid preparations each containing a single dose TESTS
of one or more active substances. They are obtained by Uniformity of dosage units (2.9.40). Tablets comply with
compressing uniform volumes of particles or by another the test or, where justified and authorised, with the tests for
suitable manufacturing technique, such as extrusion, uniformity of content and/or uniformity of mass shown below.
moulding or freeze-drying (lyophilisation). Tablets are Herbal drugs and herbal drug preparations present in the
intended for oral administration. Some are swallowed whole, dosage form are not subject to the provisions of this paragraph.
some after being chewed, some are dissolved or dispersed in Uniformity of content (2.9.6). Unless otherwise prescribed
water before being administered and some are retained in the or justified and authorised, tablets with a content of active
mouth where the active substance is liberated. substance less than 2 mg or less than 2 per cent of the total
The particles consist of one or more active substances with or mass comply with test A. If the preparation has more than
without excipients such as diluents, binders, disintegrating 1 active substance, the requirement applies only to those
agents, glidants, lubricants, substances capable of modifying substances that correspond to the above conditions.
the behaviour of the preparation in the digestive tract, Unless otherwise justified and authorised, coated tablets other
colouring matter authorised by the competent authority and than film-coated tablets comply with test A irrespective of
flavouring substances. their content of active substance(s).
Tablets are usually straight, solid cylinders, the end surfaces Uniformity of mass (2.9.5). Uncoated tablets and, unless
of which are flat or convex and the edges of which may be otherwise justified and authorised, film-coated tablets comply
bevelled. They may have break-marks and may bear a symbol with the test. If the test for uniformity of content is prescribed
or other markings. Tablets may be coated. or justified and authorised for all the active substances, the
Where applicable, containers for tablets comply with the test for uniformity of mass is not required.
requirements for materials used for the manufacture of Dissolution. Unless otherwise justified and authorised,
containers (3.1 and subsections) and containers (3.2 and a suitable test is carried out, for example one of the tests
subsections). described in general chapter 2.9.3. Dissolution test for solid
Several categories of tablets for oral use may be distinguished : dosage forms.
– uncoated tablets ; Where a dissolution test is prescribed, a disintegration test
– coated tablets ; may not be required.
– gastro-resistant tablets ;
– modified-release tablets ; Uncoated tablets
– effervescent tablets ; DEFINITION
– soluble tablets ; Uncoated tablets include single-layer tablets resulting from
– dispersible tablets ; a single compression of particles and multi-layer tablets
consisting of concentric or parallel layers obtained by
– orodispersible tablets ;
successive compression of particles of different composition.
– chewable tablets ; The excipients used are not specifically intended to modify the
– oral lyophilisates. release of the active substance in the digestive fluids.
Uncoated tablets conform to the general definition of tablets.
PRODUCTION
A broken section, when examined under a lens, shows either a
Tablets are usually prepared by compressing uniform volumes relatively uniform texture (single-layer tablets) or a stratified
of particles or particle aggregates produced by granulation texture (multi-layer tablets) but no signs of coating.
methods. In the manufacture of tablets, measures are taken
to ensure that they possess a suitable mechanical strength TESTS
to avoid crumbling or breaking on handling or subsequent Disintegration (2.9.1). Uncoated tablets comply with the
processing. This may be demonstrated using the tests test using water R as the liquid medium. Add a disc to each
described in general chapters 2.9.7. Friability of uncoated tube. Operate the apparatus for 15 min, unless otherwise
tablets and 2.9.8. Resistance to crushing of tablets. justified and authorised, and examine the state of the tablets.
In the manufacture, packaging, storage and distribution If the tablets fail to comply because of adherence to the discs,
of tablets, suitable measures are taken to ensure their the results are invalid. Repeat the test on a further 6 tablets,
microbiological quality ; recommendations on this aspect are omitting the discs.

Tablets EUROPEAN PHARMACOPOEIA
Coated tablets Modified-release tablets

DEFINITION DEFINITION
Coated tablets are tablets covered with one or more layers of Modified-release tablets are coated or uncoated tablets
mixtures of various substances such as natural or synthetic that contain special excipients or are prepared by special
resins, gums, gelatin, inactive and insoluble fillers, sugars, procedures, or both, designed to modify the rate, the place or
plasticisers, polyols, waxes, colouring matter authorised by the the time at which the active substance(s) are released.
competent authority and sometimes flavouring substances and
Modified-release tablets include prolonged-release tablets,
active substances. The substances used as coatings are usually
delayed-release tablets and pulsatile-release tablets.
applied as a solution or suspension in conditions in which
evaporation of the vehicle occurs. When the coating is a very
thin polymeric coating, the tablets are known as film-coated Effervescent tablets
tablets.
Coated tablets have a smooth surface, which is often coloured DEFINITION
and may be polished ; a broken section, when examined under Effervescent tablets are uncoated tablets generally containing
a lens, shows a core surrounded by one or more continuous acid substances and carbonates or hydrogen carbonates,
layers with a different texture. which react rapidly in the presence of water to release carbon
dioxide. They are intended to be dissolved or dispersed in
PRODUCTION water before administration.
Where justified, uniformity of mass or uniformity of content
of coated tablets other than film-coated tablets may be ensured TESTS
by control of the cores. Disintegration. Place 1 tablet in a beaker containing 200 mL
of water R at 15-25 °C ; numerous bubbles of gas are evolved.
TESTS When the evolution of gas around the tablet or its fragments
Disintegration (2.9.1). Coated tablets other than film-coated ceases the tablet has disintegrated, being either dissolved or
tablets comply with the test using water R as the liquid dispersed in the water so that no agglomerates of particles
medium. Add a disc to each tube. Operate the apparatus remain. Repeat the operation on 5 other tablets. The tablets
for 60 min, unless otherwise justified and authorised, and comply with the test if each of the 6 tablets used disintegrates
examine the state of the tablets. If any of the tablets has not in the manner prescribed within 5 min, unless otherwise
disintegrated, repeat the test on a further 6 tablets, replacing justified and authorised.
water R with 0.1 M hydrochloric acid. If 1 or 2 tablets fail to
disintegrate, repeat the test on 12 additional tablets.
Soluble tablets
The requirements of the test are met if not fewer than 16 of
the 18 tablets tested have disintegrated. DEFINITION
Film-coated tablets comply with the disintegration test Soluble tablets are uncoated or film-coated tablets. They are
prescribed above except that the apparatus is operated for intended to be dissolved in water before administration. The
30 min, unless otherwise justified and authorised. solution produced may be slightly opalescent due to the added
If coated tablets or film-coated tablets fail to comply because excipients used in the manufacture of the tablets.
of adherence to the discs, the results are invalid. Repeat the
test on a further 6 tablets, omitting the discs. TESTS
Disintegration (2.9.1). Soluble tablets disintegrate within
Gastro-resistant tablets 3 min, using water R at 15-25 °C as the liquid medium.
DEFINITION
Dispersible tablets
Gastro-resistant tablets are delayed-release tablets that are
intended to resist the gastric fluid and to release their active DEFINITION
substance(s) in the intestinal fluid. Usually they are prepared
by covering tablets with a gastro-resistant coating or from Dispersible tablets are uncoated or film-coated tablets
granules or particles already covered with a gastro-resistant intended to be dispersed in water before administration,
coating. giving a homogeneous dispersion.
Tablets covered with a gastro-resistant coating conform to the TESTS
definition of coated tablets.
Disintegration (2.9.1). Dispersible tablets disintegrate within
TESTS 3 min, using water R at 15-25 °C as the liquid medium.
Disintegration (2.9.1). Tablets covered with a gastro-resistant Fineness of dispersion. Place 2 tablets in 100 mL of water R
coating comply with the test with the following modifications. and stir until completely dispersed. A smooth dispersion is
Use 0.1 M hydrochloric acid as the liquid medium. Operate produced, which passes through a sieve screen with a nominal
the apparatus for 2 h, or another such time as may be justified mesh aperture of 710 μm.
and authorised, without the discs, and examine the state of
the tablets. The time of resistance to the acid medium varies
according to the formulation of the tablets to be examined. It
Orodispersible tablets
is typically 2 h to 3 h but even with authorised deviations is DEFINITION
not less than 1 h. No tablet shows signs of either disintegration
(apart from fragments of coating) or cracks that would allow Orodispersible tablets are uncoated tablets intended to be
the escape of the contents. Replace the acid by phosphate placed in the mouth where they disperse rapidly before being
buffer solution pH 6.8 R and add a disc to each tube. Operate swallowed.
the apparatus for 60 min and examine the state of the tablets.
If the tablets fail to comply because of adherence to the discs, TESTS
the results are invalid. Repeat the test on a further 6 tablets, Disintegration (2.9.1). Orodispersible tablets disintegrate
omitting the discs. within 3 min, using water R as the liquid medium.

EUROPEAN PHARMACOPOEIA Tablets
Chewable tablets the mouth where their contents are released in saliva and
swallowed or, alternatively, are intended to be dissolved or
DEFINITION dispersed in water before oral administration.
Chewable tablets are intended to be chewed before being PRODUCTION
swallowed.
Oral lyophilisates are obtained by freeze-drying
PRODUCTION (lyophilisation), involving division into single doses, freezing,
sublimation and drying of usually aqueous, liquid or
Chewable tablets are prepared to ensure that they are easily semi-solid preparations.
crushed by chewing.
TESTS
Oral lyophilisates Disintegration. Place 1 oral lyophilisate in a beaker
containing 200 mL of water R at 15-25 °C. It disintegrates
DEFINITION within 3 min. Repeat the test on 5 other oral lyophilisates.
Oral lyophilisates are solid single-dose preparations made by They comply with the test if all 6 have disintegrated.
freeze-drying of a liquid or semi-solid preparation. These Water (2.5.12). Oral lyophilisates comply with the test ; the
fast-releasing preparations are intended to be placed in limits are approved by the competent authority.

EUROPEAN PHARMACOPOEIA Semi-solid preparations for cutaneous application
04/2010:0132 use. Sterile semi-solid preparations for cutaneous application

corrected 10.0 are prepared using materials and methods designed to ensure
sterility and to avoid the introduction of contaminants and
the growth of micro-organisms ; recommendations on this are
provided in Methods of preparation of sterile products (5.1.1).
During development, it must be demonstrated that the
SEMI-SOLID PREPARATIONS FOR nominal content can be withdrawn from the container of
CUTANEOUS APPLICATION semi-solid preparations for cutaneous application presented
in single-dose containers.
Praeparationes molles ad usum dermicum In the manufacture of semi-solid preparations for cutaneous
application, suitable measures are taken to ensure that the
The requirements of this monograph apply to all semi-solid defined rheological properties are fulfilled. Where appropriate,
preparations for cutaneous application. Where appropriate, the following non-mandatory tests may be carried out :
additional requirements specific to semi-solid preparations measurement of consistency by penetrometry (2.9.9), viscosity
intended to be applied to particular surfaces or mucous (apparent viscosity) (2.2.10) and a suitable test to demonstrate
membranes may be found in other general monographs, for the appropriate release of the active substance(s).
example Ear preparations (0652), Nasal preparations (0676), In the manufacture of semi-solid preparations for cutaneous
Rectal preparations (1145), Eye preparations (1163) and application containing 1 or more active substances that are
Vaginal preparations (1164). not dissolved in the basis (e.g. emulsions or suspensions),
measures are taken to ensure appropriate homogeneity of the
DEFINITION preparation to be delivered.
Semi-solid preparations for cutaneous application are intended
In the manufacture of semi-solid preparations for cutaneous
for local or transdermal delivery of active substances, or for
application containing dispersed particles, measures are taken
their emollient or protective action. They are of homogeneous
to ensure a suitable and controlled particle size with regard to
appearance.
the intended use.
Semi-solid preparations for cutaneous application consist
of a simple or compound basis in which, usually, 1 or more TESTS
active substances are dissolved or dispersed. According to Uniformity of dosage units. Semi-solid preparations that are
its composition, the basis may influence the activity of the supplied either in single-dose containers that represent 1 dose
preparation. of medicinal product or in metered-dose containers, and that
The basis may consist of natural or synthetic substances are intended for transdermal delivery of the active substance(s)
and may be single phase or multiphase. According to the in view of a systemic effect, comply with the test for uniformity
nature of the basis, the preparation may have hydrophilic or of dosage units (2.9.40). Semi-solid preparations in which
hydrophobic properties ; it may contain suitable excipients the active substance(s) are dissolved comply with the test for
such as antimicrobial preservatives, antioxidants, stabilisers, mass variation ; semi-solid preparations in which the active
emulsifiers, thickeners and penetration enhancers. substance(s) are suspended comply with the test for content
Semi-solid preparations for cutaneous application intended uniformity. Follow the procedure described for liquid dosage
for use on severely injured skin are sterile. forms. Herbal drugs and herbal drug preparations present
Where applicable, containers for semi-solid preparations in the dosage form are not subject to the provisions of this
for cutaneous application comply with the requirements of paragraph.
Materials used for the manufacture of containers (3.1 and For semi-solid preparations presented in metered-dose
subsections) and Containers (3.2 and subsections). containers and in which the active substance(s) are dissolved,
Several categories of semi-solid preparations for cutaneous proceed as follows. Discharge once to waste. Wait for a
application may be distinguished : minimum of 5 s, shake for 5 s if necessary, and discharge again
– ointments ; to waste. Repeat this procedure for a further 3 actuations.
Weigh the container, discharge once to waste and weigh
– creams ; the container again. Calculate the difference between the
– gels ; 2 masses. Repeat the procedure for a further 9 containers.
– pastes ; Determine the mass variation (2.9.40).
– poultices ; For semi-solid preparations supplied in metered-dose
– medicated plasters ; containers and in which the active substance(s) are suspended,
– cutaneous patches. proceed as follows. Use an apparatus capable of quantitatively
retaining the dose leaving the metered-dose container. Shake
According to their structure, ointments, creams and gels
1 container for 5 s and discharge once to waste. Wait for a
generally show viscoelastic behaviour and are non-Newtonian
minimum of 5 s, shake for 5 s and discharge again to waste.
in character, e.g. plastic, pseudoplastic or thixotropic type
Repeat this procedure for a further 3 actuations. After 2 s, fire
flow at high shear rates. Pastes frequently exhibit dilatancy.
1 dose from the metered-dose container into the collecting
PRODUCTION vessel. Collect the contents of the collecting vessel by
During development of semi-solid preparations for cutaneous successive rinses. Determine the content of active substance
application whose formulation contains an antimicrobial in the combined rinses. Repeat the procedure for a further 9
preservative, the need for and the efficacy of the chosen containers. Determine the content uniformity (2.9.40).
preservative shall be demonstrated to the satisfaction of Sterility (2.6.1). Where the label indicates that the preparation
the competent authority. A suitable test method together is sterile, it complies with the test for sterility.
with criteria for judging the preservative properties of
the formulation are provided in Efficacy of antimicrobial STORAGE
preservation (5.1.3). In the manufacture, packaging, storage If the preparation contains water or other volatile ingredients,
and distribution of semi-solid preparations for cutaneous store in an airtight container. If the preparation is sterile, store
application, suitable measures are taken to ensure their in a sterile, airtight, tamper-evident container.
microbiological quality ; recommendations on this are
provided in 5.1.4. Microbiological quality of non-sterile LABELLING
pharmaceutical preparations and substances for pharmaceutical The label states :

Semi-solid preparations for cutaneous application EUROPEAN PHARMACOPOEIA
– the name of any excipient ; Pastes

– where applicable, that the preparation is sterile. DEFINITION
Pastes are semi-solid preparations for cutaneous application
Ointments containing large proportions of solids finely dispersed in the
basis.
DEFINITION
Poultices
An ointment consists of a single-phase basis in which solids
or liquids may be dispersed. DEFINITION
Hydrophobic ointments Poultices consist of a hydrophilic heat-retentive basis in which
solid or liquid active substances are dispersed. They are
Hydrophobic ointments can absorb only small amounts of usually spread thickly on a suitable dressing and heated before
water. Typical bases used for their formulation are hard, liquid application to the skin.
and light liquid paraffins, vegetable oils, animal fats, synthetic
glycerides, waxes and liquid polyalkylsiloxanes.
Medicated plasters
Water-emulsifying ointments
DEFINITION
Water-emulsifying ointments can absorb larger amounts
of water and thereby produce water-in-oil or oil-in-water Medicated plasters are flexible preparations containing 1 or
emulsions after homogenisation, depending on the nature of more active substances. They are intended to be applied to the
the emulsifiers : water-in-oil emulsifying agents such as wool skin. They are designed to maintain the active substance(s) in
alcohols, sorbitan esters, monoglycerides and fatty alcohols, or close contact with the skin such that these may be absorbed
oil-in-water emulsifying agents such as sulfated fatty alcohols, slowly, or act as protective or keratolytic agents.
polysorbates, macrogol cetostearyl ether or esters of fatty acids Medicated plasters consist of an adhesive basis, which may be
with macrogols may be used for this purpose. Their bases are coloured, containing 1 or more active substances, spread as a
those of the hydrophobic ointments. uniform layer on an appropriate support made of natural or
synthetic material. They are not irritant or sensitising to the
Hydrophilic ointments
skin. The adhesive layer is covered by a suitable protective
Hydrophilic ointments are preparations having bases that are liner, which is removed before applying the plaster to the
miscible with water. The bases usually consist of mixtures of skin. When removed, the protective liner does not detach the
liquid and solid macrogols (polyethylene glycols). They may preparation from the outer, supporting layer.
contain appropriate amounts of water. Medicated plasters are presented in a range of sizes directly
adapted to their intended use or as larger sheets to be cut
before use. Medicated plasters adhere firmly to the skin
Creams when gentle pressure is applied and can be peeled off without
causing appreciable injury to the skin or detachment of the
DEFINITION preparation from the outer, supporting layer.
Creams are multiphase preparations consisting of a lipophilic TESTS
phase and an aqueous phase.
Dissolution. A suitable test may be required to demonstrate
Lipophilic creams the appropriate release of the active substance(s), for example
Lipophilic creams have as the continuous phase the lipophilic one of the tests described in Dissolution test for transdermal
phase. They usually contain water-in-oil emulsifying agents patches (2.9.4).
such as wool alcohols, sorbitan esters and monoglycerides.
Hydrophilic creams Cutaneous patches
Hydrophilic creams have as the continuous phase the DEFINITION
aqueous phase. They contain oil-in-water emulsifying agents Cutaneous patches are flexible preparations containing 1 or
such as sodium or trolamine soaps, sulfated fatty alcohols, more active substances. They are intended to be applied to the
polysorbates and polyoxyl fatty acid and fatty alcohol esters skin. They are designed to maintain the active substance(s) in
combined, if necessary, with water-in-oil emulsifying agents. close contact with the skin such that these may act locally.
Cutaneous patches consist of an adhesive basis, which may
be coloured, containing 1 or more active substances, spread
Gels as a uniform layer on an appropriate support made of natural
or synthetic material. The adhesive basis is not irritant or
DEFINITION sensitising to the skin. The adhesive layer is covered by a
Gels consist of liquids gelled by means of suitable gelling suitable protective liner, which is removed before applying the
agents. patch to the skin. When removed, the protective liner does
not detach the preparation from the outer, supporting layer.
Lipophilic gels Cutaneous patches are presented in a range of sizes adapted
Lipophilic gels (oleogels) are preparations whose bases usually to their intended use. They adhere firmly to the skin when
consist of liquid paraffin with polyethylene or fatty oils gelled gentle pressure is applied and can be peeled off without
with colloidal silica or aluminium or zinc soaps. causing appreciable injury to the skin or detachment of the
preparation from the outer, supporting layer.
Hydrophilic gels
Hydrophilic gels (hydrogels) are preparations whose bases TESTS
usually consist of water, glycerol or propylene glycol gelled Dissolution. A suitable test may be required to demonstrate
with suitable gelling agents such as poloxamers, starch, the appropriate release of the active substance(s), for example
cellulose derivatives, carbomers and magnesium-aluminium one of the tests described in Dissolution test for transdermal
silicates. patches (2.9.4).

EUROPEAN PHARMACOPOEIA Abacavir sulfate
01/2017:2589 Time Mobile phase A Mobile phase B

(min) (per cent V/V) (per cent V/V)
0 - 25 100 0
25 - 27 100 → 0 0 → 100
27 - 37 0 100
ABACAVIR SULFATE
Abacaviri sulfas Detection : spectrophotometer at 286 nm.
Injection : 20 µL.
Identification of impurities : use the chromatogram supplied
with abacavir for system suitability CRS and the chromatogram
obtained with reference solution (a) to identify the peaks due
to impurities A and D.
Relative retention with reference to abacavir (retention
time = about 17 min) : impurity D = about 0.8 ;
impurity A = about 0.9.
C28H38N12O6S Mr 671 System suitability : reference solution (a) :
[188062-50-2] – resolution : minimum 1.5 between the peaks due to
DEFINITION impurities D and A ; minimum 1.5 between the peaks due
to impurity A and abacavir.
Bis[[(1S,4R)-4-[2-amino-6-(cyclopropylamino)-9H-purin-9-
yl]cyclopent-2-enyl]methanol] sulfate. Limit :
Content : 99.0 per cent to 101.0 per cent (anhydrous substance). – impurity A : not more than 3 times the area of the principal
peak in the chromatogram obtained with reference
CHARACTERS solution (b) (0.3 per cent).
Appearance : white or almost white powder. Related substances. Liquid chromatography (2.2.29). Prepare
the solutions immediately before use and transfer them to
Solubility : soluble in water, practically insoluble in ethanol low-adsorption, inert glass vials.
(96 per cent) and in methylene chloride.
Test solution. Dissolve 25 mg of the substance to be examined
IDENTIFICATION in water R and dilute to 100.0 mL with the same solvent.
A. Infrared absorption spectrophotometry (2.2.24). Sonicate until dissolution is complete.
Comparison : abacavir sulfate CRS. Reference solution (a). Dissolve 2.5 mg of abacavir for peak
identification CRS (containing impurities B and D) in 10.0 mL
B. Enantiomeric purity (see Tests). of water R.
C. Solution S (see Tests) gives reaction (a) of sulfates (2.3.1). Reference solution (b). Dilute 1.0 mL of the test solution to
TESTS 100.0 mL with water R. Dilute 1.0 mL of this solution to
Solution S. Dissolve 0.250 g in water R and dilute to 25.0 mL
with the same solvent. Column :
Enantiomeric purity. Liquid chromatography (2.2.29). – size : l = 0.15 m, Ø = 3.9 mm ;
Solution A. Mix 0.5 mL of trifluoroacetic acid R and 100 mL – stationary phase : end-capped octadecylsilyl silica gel for
of methanol R. chromatography R (5 µm) ;
Solution B. Mix 30 volumes of methanol R, 30 volumes of – temperature : 30 °C.
2-propanol R and 40 volumes of heptane R. Mobile phase :
Test solution. Dissolve 40 mg of the substance to be examined – mobile phase A : dilute 0.5 mL of trifluoroacetic acid R in
in 30 mL of solution A. Sonicate until dissolution is complete. 1000 mL of water R ;
Add 30 mL of 2-propanol R and dilute to 100.0 mL with
heptane R. – mobile phase B : water R, methanol R (15:85 V/V) ;
Reference solution (a). Dissolve 2 mg of abacavir for system Time Mobile phase A Mobile phase B
suitability CRS (containing impurities A and D) in 1.5 mL of (min) (per cent V/V) (per cent V/V)
solution A. Sonicate until dissolution is complete. Add 1.5 mL 0-5 95 5
of 2-propanol R and dilute to 5.0 mL with heptane R.
5 - 25 95 → 70 5 → 30
Reference solution (b). Dilute 1.0 mL of the test solution to
100.0 mL with solution B. Dilute 1.0 mL of this solution to 25 - 40 70 → 10 30 → 90
10.0 mL with solution B.
Column :
– size : l = 0.25 m, Ø = 4.6 mm ;
Injection : 20 µL.
– stationary phase : amylose derivative of silica gel for chiral
separation R (10 µm) ; Identification of impurities : use the chromatogram
– temperature : 30 °C. supplied with abacavir for peak identification CRS and the
chromatogram obtained with reference solution (a) to identify
Mobile phase : the peaks due to impurities B and D.
– mobile phase A : diethylamine R, 2-propanol R, heptane R Relative retention with reference to abacavir (retention
(0.1:15:85 V/V/V); time = about 22 min) : impurity D = about 1.04 ;
– mobile phase B : heptane R, 2-propanol R (50:50 V/V) ; impurity B = about 1.3.

Abacavir sulfate EUROPEAN PHARMACOPOEIA
System suitability : reference solution (a) :

– peak-to-valley ratio : minimum 3.0, where Hp = height
above the baseline of the peak due to impurity D and
Hv = height above the baseline of the lowest point of the
curve separating this peak from the peak due to abacavir.
Limits :
– impurity B : not more than twice the area of the principal
solution (b) (0.2 per cent) ;
– unspecified impurities : for each impurity, not more than the B. 6-(cyclopropylamino)-9-[(1R,4S)-4-[[(2,5-diamino-6-
area of the principal peak in the chromatogram obtained chloropyrimidin-4-yl)oxy]methyl]cyclopent-2-enyl]-9H-
with reference solution (b) (0.10 per cent) ; purine-2-amine,
– total : not more than 5 times the area of the principal peak
in the chromatogram obtained with reference solution (b)
(0.5 per cent) ;
– disregard limit : 0.5 times the area of the principal peak in
the chromatogram obtained with reference solution (b)
(0.05 per cent).
Water (2.5.32) : maximum 0.5 per cent, determined on
C. [(1S,4R)-4-(2,6-diamino-9H-purin-9-yl)cyclopent-2-
60.0 mg.
enyl]methanol,
Sulfated ash (2.4.14) : maximum 0.2 per cent, determined on
1.0 g.
ASSAY
Dissolve 0.300 g in 50 mL of water R. Titrate with 0.1 M sodium
hydroxide, determining the end-point potentiometrically
(2.2.20).
1 mL of 0.1 M sodium hydroxide is equivalent to 33.54 mg of
C28H38N12O6S. D. [(1R,4R)-4-[2-amino-6-(cyclopropylamino)-9H-purin-9-
yl]cyclopent-2-enyl]methanol,
IMPURITIES
Specified impurities : A, B.
Other detectable impurities (the following substances would,
if present at a sufficient level, be detected by one or other of
the tests in the monograph. They are limited by the general
acceptance criterion for other/unspecified impurities and/or
by the general monograph Substances for pharmaceutical use
(2034). It is therefore not necessary to identify these impurities
for demonstration of compliance. See also 5.10. Control of E. [(1R,3S)-3-[2-amino-6-(cyclopropylamino)-9H-purin-9-
impurities in substances for pharmaceutical use) : C, D, E, F. yl]cyclopentyl]methanol,
F. 6-(cyclopropylamino)-9-[(1R,4S)-4-[[(1,1-
A. [(1R,4S)-4-[2-amino-6-(cyclopropylamino)-9H-purin-9- dimethylethyl)oxy]methyl]cyclopent-2-enyl]-9H-
yl]cyclopent-2-enyl]methanol, purine-2-amine.

EUROPEAN PHARMACOPOEIA Aciclovir
01/2014:0968 Mobile phase :

– mobile phase A : acetonitrile R, phosphate buffer solution
pH 3.1 (1:99 V/V) ;
– mobile phase B : acetonitrile R, phosphate buffer solution
pH 2.5 (50:50 V/V) ;
ACICLOVIR
Time Mobile phase A Mobile phase B
Aciclovirum (min)
0-5
(per cent V/V)
100
(per cent V/V)
0
5 - 27 100 → 80 0 → 20
27 - 40 80 20

C8H11N5O3 Mr 225.2
[59277-89-3] Injection : 10 µL of the test solution and reference solutions (b),
(c) and (d).
DEFINITION Identification of impurities : use the chromatogram supplied
2-Amino-9-[(2-hydroxyethoxy)methyl]-1,9-dihydro-6H- with aciclovir for peak identification 1 CRS and the
purin-6-one. chromatogram obtained with reference solution (c) to identify
Content : 98.5 per cent to 101.0 per cent (anhydrous substance). the peaks due to impurities C and I ; use the chromatogram
supplied with aciclovir for peak identification 2 CRS and the
CHARACTERS chromatogram obtained with reference solution (d) to identify
Appearance : white or almost white, crystalline powder. the peaks due to impurities A, B, F, G, J, K, N, O and P.
Solubility : slightly soluble in water, very slightly soluble Relative retention with reference to aciclovir (retention
in ethanol (96 per cent), practically insoluble in heptane. time = about 13 min) : impurity B = about 0.4 ;
It dissolves in dilute solutions of mineral acids and alkali impurity P = about 0.7 ; impurity C = about 0.9 ;
hydroxides. impurity N = about 1.37 ; impurities O and Q = about 1.42 ;
impurity I = about 1.57 ; impurity J = about 1.62 ;
IDENTIFICATION impurity F = about 1.7 ; impurity A = about 1.8 ; impurities K
Infrared absorption spectrophotometry (2.2.24). and R = about 2.5 ; impurity G = about 2.6.
Comparison : aciclovir CRS. System suitability :
TESTS – resolution : minimum 1.5 between the peaks due to
impurity C and aciclovir in the chromatogram obtained
Appearance of solution. The solution is clear (2.2.1) and not with reference solution (c) ; minimum 1.5 between the
more intensely coloured than reference solution Y7 (2.2.2, peaks due to impurities F and A and minimum 1.5 between
Method II). the peaks due to impurities K and G in the chromatogram
Dissolve 0.25 g in a 4 g/L solution of sodium hydroxide R and obtained with reference solution (d).
dilute to 25 mL with the same solvent. Limits :
Related substances. Liquid chromatography (2.2.29). Prepare – correction factor : for the calculation of content, multiply
the solutions immediately before use. the peak area of impurity I by 1.5 ;
Solvent mixture : dimethyl sulfoxide R, water R (20:80 V/V). – impurity B : not more than 7 times the area of the principal
Phosphate buffer solution pH 2.5. Dissolve 3.48 g of peak in the chromatogram obtained with reference
dipotassium hydrogen phosphate R in 1000 mL of water R and solution (b) (0.7 per cent);
adjust to pH 2.5 with phosphoric acid R.
– sum of impurities O and Q : not more than 3 times the area
Phosphate buffer solution pH 3.1. Dissolve 3.48 g of of the principal peak in the chromatogram obtained with
dipotassium hydrogen phosphate R in 1000 mL of water R and reference solution (b) (0.3 per cent) ;
adjust to pH 3.1 with phosphoric acid R.
– sum of impurities K and R : not more than twice the area
Test solution. Dissolve 25 mg of the substance to be examined of the principal peak in the chromatogram obtained with
in 5.0 mL of dimethyl sulfoxide R and dilute to 25.0 mL with reference solution (b) (0.2 per cent) ;
water R.
– impurities A, G, J, N, P : for each impurity, not more than
Reference solution (a). Dissolve 5 mg of aciclovir for system twice the area of the principal peak in the chromatogram
suitability CRS (containing impurities A, B, J, K, N, O and obtained with reference solution (b) (0.2 per cent) ;
P) in 1 mL of dimethyl sulfoxide R and dilute to 5.0 mL with
water R. – impurities C, F, I : for each impurity, not more than the area
of the principal peak in the chromatogram obtained with
reference solution (b) (0.1 per cent) ;
100.0 mL with the solvent mixture. Dilute 1.0 mL of this
solution to 10.0 mL with the solvent mixture. – unspecified impurities : for each impurity, not more than
Reference solution (c). Dissolve the contents of a vial of 0.5 times the area of the principal peak in the chromatogram
aciclovir for peak identification 1 CRS (containing impurities C obtained with reference solution (b) (0.05 per cent) ;
and I) in 200 µL of dimethyl sulfoxide R and dilute to 1.0 mL – total : not more than 15 times the area of the principal peak
with water R. in the chromatogram obtained with reference solution (b)
Reference solution (d). Dissolve the contents of a vial of (1.5 per cent);
aciclovir for peak identification 2 CRS (containing impurities F – disregard limit : 0.3 times the area of the principal peak in
and G) in 1.0 mL of reference solution (a). the chromatogram obtained with reference solution (b)
Column : (0.03 per cent).
– size : l = 0.25 m, Ø = 4.6 mm ; Water (2.5.12) : maximum 6.0 per cent, determined on 0.500 g.
– stationary phase : end-capped octadecylsilyl silica gel for Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
chromatography R (5 μm). 1.0 g.

Aciclovir EUROPEAN PHARMACOPOEIA
Bacterial endotoxins (2.6.14, Method D): less than

0.50 IU/mg, if intended for use in the manufacture of
parenteral preparations without a further appropriate
procedure for the removal of bacterial endotoxins.
ASSAY
J. 9,9′-[ethylenebis(oxymethylene)]bis(2-amino-1,9-dihydro-
Dissolve 0.150 g in 60 mL of anhydrous acetic acid R. Titrate 6H-purin-6-one),
with 0.1 M perchloric acid, determining the end-point
potentiometrically (2.2.20). Carry out a blank titration.
1 mL of 0.1 M perchloric acid is equivalent to 22.52 mg
of C8H11N5O3.
IMPURITIES
Specified impurities : A, B, C, F, G, I, J, K, N, O, P, Q, R.
K. 2,2′-(methylenediimino)bis[9-[(2-hydroxyethoxy)methyl]-
Other detectable impurities (the following substances would, 1,9-dihydro-6H-purin-6-one],
for demonstration of compliance. See also 5.10. Control of
impurities in substances for pharmaceutical use) : L, M.
L. N-(9-acetyl-6-oxo-6,9-dihydro-1H-purin-2-yl)acetamide
(N2,9-diacetylguanine),
A. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl acetate,
M. 2-[[2-(acetylamino)-6-oxo-1,6-dihydro-7H-purin-7-
yl]methoxy]ethyl acetate,
N. unknown structure,
B. 2-amino-1,7-dihydro-6H-purin-6-one (guanine),
O. unknown structure,
C. 2-amino-7-[(2-hydroxyethoxy)methyl]-1,7-dihydro-6H-
purin-6-one, P. 2-amino-9-(2-hydroxyethyl)-1,9-dihydro-6H-purin-6-one,
F. N-[9-[(2-hydroxyethoxy)methyl]-6-oxo-6,9-dihydro-1H-
purin-2-yl]acetamide,
Q. mixture of 2-amino-9-[[2-(hydroxymethoxy)
ethoxy]methyl]-1,9-dihydro-6H-purin-6-one and
G. 2-[[2-(acetylamino)-6-oxo-1,6-dihydro-9H-purin-9- 2-amino-9-[[2-(hydroxyethoxy)methoxy]methyl]-1,9-
yl]methoxy]ethyl acetate, dihydro-6H-purin-6-one,
I. 2-amino-7-[[2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9- R. 9,9′-[methylenebis(oxyethyleneoxymethylene)]bis(2-
yl)methoxy]ethoxy]methyl]-1,7-dihydro-6H-purin-6-one, amino-1,9-dihydro-6H-purin-6-one).

EUROPEAN PHARMACOPOEIA Atazanavir sulfate
04/2019:2898 Reference solution (d). Dissolve 2.0 mg of atazanavir

corrected 10.1 impurity K CRS in 9 mL of the solvent mixture, sonicate for
3 min and dilute to 10.0 mL with the solvent mixture. Dilute
5.0 mL of the solution to 100.0 mL with the solvent mixture.
Dilute 3.0 mL of this solution to 20.0 mL with the solvent
mixture.
ATAZANAVIR SULFATE Column :
– size : l = 0.15 m, Ø = 4.6 mm ;
Atazanaviri sulfas – stationary phase : end-capped octadecylsilyl silica gel for
chromatography R (3.0 µm);
– temperature : 25 °C.
Mobile phase :
– mobile phase A : mix 25 volumes of acetonitrile R1 and
75 volumes of a freshly prepared 2.73 g/L solution of
potassium dihydrogen phosphate R previously adjusted to
pH 3.5 with dilute phosphoric acid R ;
– mobile phase B : mix 25 volumes of a freshly prepared
2.73 g/L solution of potassium dihydrogen phosphate R
previously adjusted to pH 3.5 with dilute phosphoric acid R,
and 75 volumes of acetonitrile R1 ;
C38H54N6O11S Mr 803 Time Mobile phase A Mobile phase B
[229975-97-7] (min) (per cent V/V) (per cent V/V)
0-5 100 0
DEFINITION
5 - 45 100 → 0 0 → 100
Methyl [(5S,10S,11S,14S)-11-benzyl-5-tert-butyl-10-
hydroxy-15,15-dimethyl-3,6,13-trioxo-8-[[4-(pyridin-2- Flow rate : 1.0 mL/min.
yl)phenyl]methyl]-2-oxa-4,7,8,12-tetraazahexadecan-14- Detection : spectrophotometer at 215 nm.
yl]carbamate sulfate.
Injection : 10 µL of test solution (a) and reference solutions (b)
Content : 98.0 per cent to 102.0 per cent (anhydrous substance). and (c).
CHARACTERS Identification of impurities : use the chromatogram
Appearance : white or pale yellow, slightly hygroscopic, supplied with atazanavir for system suitability CRS and the
crystalline powder that may contain agglomerates. chromatogram obtained with reference solution (c) to identify
the peak due to impurity F.
Solubility : slightly soluble in water, freely soluble in ethanol
Relative retention with reference to atazanavir (retention
(96 per cent), practically insoluble in heptane.
time = about 30 min) : impurity F = about 0.99.
IDENTIFICATION System suitability : reference solution (c) :
A. Specific optical rotation (see Tests). – resolution : minimum 1.5 between the peaks due to
B. Infrared absorption spectrophotometry (2.2.24). impurity F and atazanavir.
Comparison : atazanavir sulfate CRS. Calculation of percentage contents :
C. It gives reaction (a) of sulfates (2.3.1). – for each impurity, use the concentration of atazanavir
sulfate in reference solution (b).
TESTS Limits :
Specific optical rotation (2.2.7): − 44 to − 40 (anhydrous – unspecified impurities : for each impurity, maximum
substance), measured at 25 °C. 0.10 per cent ;
Dissolve 0.100 g in 8 mL of methanol R, using sonication if – total : maximum 0.5 per cent ;
necessary, and dilute to 10.0 mL with the same solvent. – reporting threshold : 0.05 per cent ; disregard any peak with
Related substances. Liquid chromatography (2.2.29). a relative retention with reference to atazanavir of less than
Solvent mixture. Mix equal volumes of acetonitrile R1 and a 0.2.
freshly prepared 2.73 g/L solution of potassium dihydrogen Impurity K. Liquid chromatography (2.2.29) as described in
phosphate R in water for chromatography R previously adjusted the test for related substances with the following modifications.
to pH 3.5 with dilute phosphoric acid R. Mobile phase :
Test solution (a). Dissolve 20.0 mg of the substance to be Time Mobile phase A Mobile phase B
examined in 40 mL of the solvent mixture, sonicate for 3 min (min) (per cent V/V) (per cent V/V)
and dilute to 50.0 mL with the solvent mixture. 0-5 95 5
Test solution (b). Dissolve 50.0 mg of the substance to be
5-8 95 → 0 5 → 100
examined in 40 mL of the solvent mixture, sonicate for 3 min
and dilute to 50.0 mL with the solvent mixture. 8 - 14 0 100
Reference solution (a). Dissolve 20.0 mg of atazanavir
sulfate CRS in 40 mL of the solvent mixture, sonicate for 3 min Injection : 20 µL of test solution (b) and reference solution (d).
and dilute to 50.0 mL with the solvent mixture. Identification of impurities: use the chromatogram obtained
Reference solution (b). Dilute 1.0 mL of test solution (a) to with reference solution (d) to identify the peak due to
100.0 mL with the solvent mixture. Dilute 1.0 mL of this impurity K.
solution to 10.0 mL with the solvent mixture. Relative retention with reference to atazanavir (retention
Reference solution (c). Dissolve 4 mg of atazanavir for system time = about 10 min) : impurity K = about 0.4.
suitability CRS (containing impurity F) in 8 mL of the solvent Calculation of percentage content :
mixture, sonicate for 3 min and dilute to 10 mL with the – for impurity K, use the concentration of impurity K in
solvent mixture. reference solution (d).

Atazanavir sulfate EUROPEAN PHARMACOPOEIA
Limit :
– impurity K : maximum 0.15 per cent.
Water (2.5.32) : maximum 2.5 per cent, determined on 0.100 g
by direct sample introduction.
1.0 g.
ASSAY
Liquid chromatography (2.2.29) as described in the test for
related substances with the following modifications.
D. (2S,3S)-3-amino-4-phenyl-1-[(E)-1-[[4-(pyri-
Mobile phase : solvent mixture. din-2-yl)phenyl]methyl]-2-[[4-(pyridin-2-yl)phenyl]meth-
Injection : test solution (a) and reference solutions (a) and (c). ylidene]hydrazin-1-yl]butan-2-ol,
Run time : 1.6 times the retention time of atazanavir.
Relative retention with reference to atazanavir (retention
time = about 9.5 min): impurity F = about 0.94.
System suitability : reference solution (c) :
– resolution : minimum 1.5 between the peaks due to
impurity F and atazanavir.
Calculate the percentage content of C38H54N6O11S using the
chromatogram obtained with reference solution (a) and taking
into account the assigned content of atazanavir sulfate CRS.
STORAGE
In an airtight container.
IMPURITIES E. methyl [(5S,10R,11S,14S)-11-benzyl-5-tert-butyl-10-

hydroxy-15,15-dimethyl-3,6,13-trioxo-8-[[4-(pyridin-2-
Specified impurities : K. yl)phenyl]methyl]-2-oxa-4,7,8,12-tetraazahexadecan-14-
Other detectable impurities (the following substances would, yl]carbamate,
by the general monograph Substances for pharmaceutical
use (2034). It is therefore not necessary to identify these
impurities for demonstration of compliance. See also 5.10.
Control of impurities in substances for pharmaceutical use) : A,
B, C, D, E, F, G, H, I, J.
A. 4-(pyridin-2-yl)benzoic acid, F. methyl [(5R,10S,11S,14S)-11-benzyl-5-tert-butyl-10-

hydroxy-15,15-dimethyl-3,6,13-trioxo-8-[[4-(pyridin-2-
yl)phenyl]methyl]-2-oxa-4,7,8,12-tetraazahexadecan-14-
yl]carbamate,
B. 4-(pyridin-2-yl)benzaldehyde,
G. methyl [(5S,10S,11S,14R)-11-benzyl-5-tert-butyl-10-
C. methyl [(5S,10S,11S,14S)-11-benzyl-5-tert-butyl-10- hydroxy-15,15-dimethyl-3,6,13-trioxo-8-[[4-(pyridin-2-
hydroxy-15,15-dimethyl-3,6,13-trioxo-2-oxa-4,7,8,12- yl)phenyl]methyl]-2-oxa-4,7,8,12-tetraazahexadecan-14-
tetraazahexadecan-14-yl]carbamate, yl]carbamate,

EUROPEAN PHARMACOPOEIA Atazanavir sulfate
H. methyl [(5S,10R,11R,14S)-11-benzyl-5-tert-butyl-10-
hydroxy-15,15-dimethyl-3,6,13-trioxo-8-[[4-(pyridin-2- J. tert-butyl 2-[(2S,3S)-3-(tert-butoxyformamido)-
yl)phenyl]methyl]-2-oxa-4,7,8,12-tetraazahexadecan-14- 2-hydroxy-4-phenylbutyl]-2-[[4-(pyridin-2-
yl]carbamate, yl)phenyl]methyl]hydrazine-1-carboxylate,
I. methyl [(2S)-1-[[(2S,3S)-3-hydroxy-1-phenyl-4-[(E)-
1-[[4-(pyridin-2-yl)phenyl]methyl]-2-[[4-(pyridin-2-
yl)phenyl]methylidene]hydrazin-1-yl]butan-2-yl]amino]-
3,3-dimethyl-1-oxobutan-2-yl]carbamate, K. (2S)-2-(methoxyformamido)-3,3-dimethylbutanoic acid.

EUROPEAN PHARMACOPOEIA Azithromycin
01/2018:1649 Reference solution (b). Dissolve the contents of a vial of

corrected 9.6 azithromycin for system suitability CRS (containing impurities
F, H and J) in 1.0 mL of the solvent mixture and sonicate for
5 min.
Reference solution (c). Dissolve 8.0 mg of azithromycin for
peak identification CRS (containing impurities A, B, C, E, F, G,
AZITHROMYCIN I, J, L, M, N, O and P) in 1.0 mL of the solvent mixture.
Column :
Azithromycinum – size : l = 0.25 m, Ø = 4.6 mm ;
– stationary phase : end-capped octadecylsilyl amorphous
organosilica polymer for chromatography R (5 µm) ;
Mobile phase :
– mobile phase A : 1.80 g/L solution of anhydrous disodium
hydrogen phosphate R adjusted to pH 8.9 with dilute
phosphoric acid R or with dilute sodium hydroxide
solution R ;
– mobile phase B : methanol R2, acetonitrile R1 (25:75 V/V);
0 - 25 50 → 45 50 → 55
C38H72N2O12,xH2O Mr 749 (anhydrous substance)
with x = 1 or 2 25 - 30 45 → 40 55 → 60
Azithromycin monohydrate : [121470-24-4] 30 - 80 40 → 25 60 → 75
Azithromycin dihydrate : [117772-70-0]
80 - 81 25 → 50 75 → 50
DEFINITION
81 - 93 50 50
(2R,3S,4R,5R,8R,10R,11R,12S,13S,14R)-13-[(2,6-Dideoxy-3-
C-methyl-3-O-methyl-α-L-ribo-hexopyranosyl)oxy]-2-ethyl- Flow rate : 1.0 mL/min.
3,4,10-trihydroxy-3,5,6,8,10,12,14-heptamethyl-11-[[3,4,6- Detection : spectrophotometer at 210 nm.
trideoxy-3-(dimethylamino)-β-D-xylo-hexopyranosyl]oxy]-1-
oxa-6-azacyclopentadecan-15-one. The degree of hydration is Injection : 50 µL.
1 or 2. Identification of impurities : use the chromatogram supplied
Semi-synthetic product derived from a fermentation product. with azithromycin for peak identification CRS and the
chromatogram obtained with reference solution (c) to identify
Content : 96.0 per cent to 102.0 per cent (anhydrous substance).the peaks due to impurities A, B, C, E, F, G, I, J, L, M, N, O
CHARACTERS and P ; use the chromatogram supplied with azithromycin for
system suitability CRS and the chromatogram obtained with
Appearance : white or almost white powder. reference solution (b) to identify the peak due to impurity H.
Solubility : practically insoluble in water, freely soluble in Relative retention with reference to azithromycin
anhydrous ethanol and in methylene chloride. (retention time = 45-50 min) : impurity L = about 0.29 ;
IDENTIFICATION impurity M = about 0.37 ; impurity E = about 0.43 ;
impurity F = about 0.51 ; impurity D = about 0.54 ;
Infrared absorption spectrophotometry (2.2.24). impurity J = about 0.54 ; impurity Q = about 0.54 ;
Comparison : azithromycin CRS. impurity I = about 0.61 ; impurity C = about 0.73 ;
If the spectra obtained in the solid state show differences, impurity N = about 0.76 ; impurity H = about 0.79 ;
prepare further spectra using 90 g/L solutions in methylene impurity A = about 0.83 ; impurity P = about 0.92 ;
chloride R. impurity O = about 1.23 ; impurity G = about 1.26 ;
impurity B = about 1.31.
TESTS
System suitability : reference solution (b) :
Solution S. Dissolve 0.500 g in anhydrous ethanol R and dilute – peak-to-valley ratio : minimum 1.4, where H = height
to 50.0 mL with the same solvent. p
above the baseline of the peak due to impurity J and
Appearance of solution. Solution S is clear (2.2.1) and Hv = height above the baseline of the lowest point of the
colourless (2.2.2, Method II). curve separating this peak from the peak due to impurity F.
pH (2.2.3) : 9.0 to 11.0. Limits :
Dissolve 0.100 g in 25.0 mL of methanol R and dilute to – correction factors : for the calculation of content,
50.0 mL with carbon dioxide-free water R. multiply the peak areas of the following impurities by
the corresponding correction factor : impurity F = 0.3 ;
Specific optical rotation (2.2.7): − 49 to − 45 (anhydrous impurity G = 0.2 ; impurity H = 0.1 ; impurity L = 2.3 ;
substance), determined on solution S. impurity M = 0.6 ; impurity N = 0.7 ;
Related substances. Liquid chromatography (2.2.29). – impurity B : not more than twice the area of the principal
Solvent mixture. Prepare a 1.73 g/L solution of ammonium peak in the chromatogram obtained with reference
dihydrogen phosphate R adjusted to pH 10.0 with ammonia R. solution (a) (2.0 per cent);
Transfer 350 mL of this solution to a suitable container. Add – impurities A, C, E, F, H, I, L, M, N, O, P : for each impurity,
300 mL of acetonitrile R and 350 mL of methanol R. Mix well. not more than 0.5 times the area of the principal peak in
Test solution. Dissolve 0.200 g of the substance to be examined the chromatogram obtained with reference solution (a)
in the solvent mixture and dilute to 25.0 mL with the solvent (0.5 per cent);
mixture. – sum of impurities D, J and Q : not more than 0.5 times the
Reference solution (a). Dilute 1.0 mL of the test solution to area of the principal peak in the chromatogram obtained
100.0 mL with the solvent mixture. with reference solution (a) (0.5 per cent) ;

Azithromycin EUROPEAN PHARMACOPOEIA
– impurity G : not more than 0.2 times the area of the

principal peak in the chromatogram obtained with
reference solution (a) (0.2 per cent) ;
– any other impurity : for each impurity, not more than
0.2 times the area of the principal peak in the chromatogram
obtained with reference solution (a) (0.2 per cent) ;
in the chromatogram obtained with reference solution (a)
(3.0 per cent) ;
the chromatogram obtained with reference solution (a) A. 6-demethylazithromycin,
(0.1 per cent) ; disregard the peaks eluting before impurity L
and after impurity B.
Water (2.5.12) : 1.8 per cent to 6.5 per cent, determined on
0.200 g.
1.0 g.
ASSAY
Liquid chromatography (2.2.29).
Solution A. Mix 60 volumes of acetonitrile R and 40 volumes
of a 6.7 g/L solution of dipotassium hydrogen phosphate R
adjusted to pH 8.0 with phosphoric acid R.
Test solution. Dissolve 53.0 mg of the substance to be
examined in 2 mL of acetonitrile R and dilute to 100.0 mL B. 3-deoxyazithromycin (azithromycin B),
with solution A.
Reference solution (a). Dissolve 53.0 mg of azithromycin CRS
in 2 mL of acetonitrile R and dilute to 100.0 mL with
solution A.
Reference solution (b). Dissolve 5 mg of the substance to be
examined and 5 mg of azithromycin impurity A CRS in 0.5 mL
of acetonitrile R and dilute to 10 mL with solution A.
Column :
– size : l = 0.25 m, Ø = 4.6 mm ;
– stationary phase : octadecylsilyl vinyl polymer for
chromatography R (5 μm) ;
C. 3″-O-demethylazithromycin (azithromycin C),
Mobile phase : mix 60 volumes of acetonitrile R1 and
40 volumes of a 6.7 g/L solution of dipotassium hydrogen
phosphate R adjusted to pH 11.0 with a 560 g/L solution of
potassium hydroxide R.
Injection : 10 µL.
Run time : 1.5 times the retention time of azithromycin.
Retention time : azithromycin = about 10 min.
impurity A and azithromycin.
Calculate the percentage content of C38H72N2O12 taking into D. 14-demethyl-14-(hydroxymethyl)azithromycin
account the assigned content of azithromycin CRS. (azithromycin F),
STORAGE
IMPURITIES
Specified impurities : A, B, C, D, E, F, G, H, I, J, L, M, N, O, P, Q.
Control of impurities in substances for pharmaceutical use): K. E. 3′-(N,N-didemethyl)azithromycin (aminoazithromycin),

EUROPEAN PHARMACOPOEIA Azithromycin
F. 3′-N-demethyl-3′-N-formylazithromycin,
K. C14,1″-epoxyazithromycin (azithromycin E),
G. 3′-N-demethyl-3′-N-[(4-methylphenyl)sulfonyl]- L. azithromycin 3′-N-oxide,

azithromycin,
M. 3′-(N,N-didemethyl)-3′-N-formylazithromycin,
H. 3′-N-[[4-(acetylamino)phenyl]sulfonyl]-3′-N-
demethylazithromycin,
N. 3′-de(dimethylamino)-3′-oxoazithromycin,
I. 3′-N-demethylazithromycin,
O. 2-desethyl-2-propylazithromycin,
J. 13-O-decladinosylazithromycin, P. unknown structure,

Azithromycin EUROPEAN PHARMACOPOEIA
Q. 3′-N-[[4-(acetylamino)phenyl]sulfonyl]-3′-(N,N-
didemethyl)azithromycin.

EUROPEAN PHARMACOPOEIA Chloroquine phosphate
01/2017:0544 the organic layers, wash with water R, dry over anhydrous
sodium sulfate R, evaporate to dryness and dissolve the
residues separately, each in 2 mL of methylene chloride R.
C. Dissolve 25 mg in 20 mL of water R and add 8 mL of picric
acid solution R1. The precipitate, washed with water R,
CHLOROQUINE PHOSPHATE with alcohol R and finally with methylene chloride R, melts
(2.2.14) at 206-209 °C.
Chloroquini phosphas D. Dissolve 0.1 g in 10 mL of water R, add 2 mL of dilute
sodium hydroxide solution R and shake with 2 quantities,
each of 20 mL, of methylene chloride R. The aqueous layer,
acidified by the addition of nitric acid R, gives reaction (b)
of phosphates (2.3.1).
TESTS
Solution S. Dissolve 2.5 g in carbon dioxide-free water R and
C18H32ClN3O8P2 Mr 515.9 Appearance of solution. Solution S is clear (2.2.1) and not
[50-63-5] more intensely coloured than reference solution BY5 or GY5
(2.2.2, Method II).
DEFINITION pH (2.2.3). The pH of solution S is 3.8 to 4.3.
Chloroquine phosphate contains not less than 98.5 per Related substances. Examine by thin-layer chromatography
cent and not more than the equivalent of 101.0 per cent of (2.2.27), using silica gel GF254 R as the coating substance.
N -(7-chloroquinolin-4-yl)-N ,N -diethylpentane-1,4-diamine
4 1 1
bis(dihydrogen phosphate), calculated with reference to the Test solution. Dissolve 0.50 g of the substance to be examined
dried substance. in water R and dilute to 10 mL with the same solvent.
Reference solution (a). Dilute 1 mL of the test solution to
CHARACTERS 100 mL with water R.
A white or almost white, crystalline powder, hygroscopic, Reference solution (b). Dilute 5 mL of reference solution (a)
freely soluble in water, very slightly soluble in alcohol and in to 10 mL with water R.
methanol. Apply to the plate 2 µL of each solution. Develop over a path
It exists in 2 forms, one of which melts at about 195 °C and the of 12 cm using a mixture of 10 volumes of diethylamine R,
other at about 218 °C. 40 volumes of cyclohexane R and 50 volumes of chloroform R.
Allow the plate to dry in air. Examine in ultraviolet light at
IDENTIFICATION 254 nm. Any spot in the chromatogram obtained with the test
First identification : B, D. solution, apart from the principal spot, is not more intense
Second identification : A, C, D. than the spot in the chromatogram obtained with reference
A. Dissolve 0.100 g in water R and dilute to 100.0 mL with the solution (a) (1.0 per cent) and not more than one such spot is
same solvent. Dilute 1.0 mL of this solution to 100.0 mL more intense than the spot in the chromatogram obtained
with water R. Examined between 210 nm and 370 nm with reference solution (b) (0.5 per cent).
(2.2.25), the solution shows absorption maxima at 220 nm, Loss on drying (2.2.32) : maximum 2.0 per cent, determined
235 nm, 256 nm, 329 nm and 342 nm. The specific on 1.000 g by drying in an oven at 105 °C.
absorbances at the maxima are respectively 600 to 660, 350
to 390, 300 to 330, 325 to 355 and 360 to 390. ASSAY
B. Examine by infrared absorption spectrophotometry Dissolve 0.200 g in 50 mL of anhydrous acetic acid R. Titrate
(2.2.24), comparing with the spectrum obtained with with 0.1 M perchloric acid determining the end-point
the base isolated from chloroquine sulfate CRS. Record potentiometrically (2.2.20).
the spectra using solutions prepared as follows : dissolve 1 mL of 0.1 M perchloric acid is equivalent to 25.79 mg of
separately 0.1 g of the substance to be examined and 80 mg C18H32ClN3O8P2.
of the reference substance in 10 mL of water R, add 2 mL
of dilute sodium hydroxide solution R and shake with 2 STORAGE
quantities, each of 20 mL, of methylene chloride R ; combine In an airtight container, protected from light.

EUROPEAN PHARMACOPOEIA Chloroquine sulfate
01/2017:0545 organic layers, wash with water R, dry over anhydrous

sodium sulfate R, evaporate to dryness and dissolve the
residues separately each in 2 mL of methylene chloride R.
C. Dissolve 25 mg in 20 mL of water R and add 8 mL of picric
acid solution R1. The precipitate, washed with water R,
CHLOROQUINE SULFATE with ethanol (96 per cent) R and finally with ether R, melts
(2.2.14) at 206 °C to 209 °C.
D. It gives reaction (a) of sulfates (2.3.1).
Chloroquini sulfas
TESTS
Solution S. Dissolve 2.0 g in carbon dioxide-free water R and
Appearance of solution. Solution S is clear (2.2.1) and not
more intensely coloured than reference solution BY5 or GY5
(2.2.2, Method II).
pH (2.2.3). The pH of solution S is 4.0 to 5.0.
C18H28ClN3O4S,H2O Mr 436.0 Related substances. Examine by thin-layer chromatography
(2.2.27), using silica gel GF254 R as the coating substance.
DEFINITION Test solution. Dissolve 0.50 g of the substance to be examined
Chloroquine sulfate contains not less than 98.5 per cent in water R and dilute to 10 mL with the same solvent.
and not more than the equivalent of 101.0 per cent of Reference solution (a). Dilute 1 mL of the test solution to
N4-(7-chloroquinolin-4-yl)-N1,N1-diethylpentane-1,4-diamine 100 mL with water R.
sulfate, calculated with reference to the anhydrous substance. Reference solution (b). Dilute 5 mL of reference solution (a)
to 10 mL with water R.
CHARACTERS Apply separately to the plate 2 μL of each solution. Develop
A white or almost white, crystalline powder, freely soluble in over a path of 12 cm using a mixture of 10 volumes of
water and in methanol, very slightly soluble in ethanol (96 per diethylamine R, 40 volumes of cyclohexane R and 50 volumes
cent). of methylene chloride R. Allow the plate to dry in air. Examine
It melts at about 208 °C (instantaneous method). in ultraviolet light at 254 nm. Any spot in the chromatogram
obtained with the test solution, apart from the principal
IDENTIFICATION spot, is not more intense than the spot in the chromatogram
obtained with reference solution (a) (1.0 per cent) and not
First identification : B, D. more than one such spot is more intense than the spot in the
Second identification : A, C, D. chromatogram obtained with reference solution (b) (0.5 per
A. Dissolve 0.100 g in water R and dilute to 100.0 mL with the cent).
same solvent. Dilute 1.0 mL of this solution to 100.0 mL Water (2.5.12) : 3.0 per cent to 5.0 per cent, determined on
with water R. Examined between 210 nm and 370 nm 0.500 g.
(2.2.25), the solution shows absorption maxima at 220 nm, Sulfated ash (2.4.14). Not more than 0.1 per cent, determined
235 nm, 256 nm, 329 nm and 342 nm. The specific on 1.0 g.
absorbances at the maxima are respectively 730 to 810, 430
to 470, 370 to 410, 400 to 440 and 430 to 470. ASSAY
B. Examine by infrared absorption spectrophotometry Dissolve 0.400 g in 50 mL of anhydrous acetic acid R. Titrate
(2.2.24), comparing with the spectrum obtained with with 0.1 M perchloric acid determining the end-point
the base isolated from chloroquine sulfate CRS. Record potentiometrically (2.2.20).
the spectra using solutions prepared as follows : dissolve 1 mL of 0.1 M perchloric acid is equivalent to 41.8 mg of
separately 0.1 g of the substance to be examined and of the C18H28ClN3O4S.
reference substance in 10 mL of water R, add 2 mL of dilute
sodium hydroxide solution R and shake with 2 quantities, STORAGE
each of 20 mL, of methylene chloride R ; combine the Store in an airtight container, protected from light.

EUROPEAN PHARMACOPOEIA Didanosine
01/2017:2200 – mobile phase B : mix 30 volumes of methanol R and

70 volumes of a 3.86 g/L solution of ammonium acetate R
adjusted to pH 8.0 with concentrated ammonia R ;
DIDANOSINE 0 - 18 100 0
Didanosinum 18 - 25 100 → 0 0 → 100
25 - 45 0 100
45 - 50 0 → 100 100 → 0
50 - 60 100 0

Injection : 20 µL.
Identification of impurities : use the chromatogram
C10H12N4O3 Mr 236.2 supplied with didanosine for system suitability CRS and the
[69655-05-6] chromatogram obtained with reference solution (c) to identify
DEFINITION the peaks due to impurities A to F and use the chromatogram
obtained with reference solution (d) to identify the peak due
9-(2,3-Dideoxy-β-D-glycero-pentofuranosyl)-1,9-dihydro-6H- to impurity G.
purin-6-one (2′,3′-dideoxyinosine).
Relative retention with reference to didanosine (retention
Content : 98.5 per cent to 101.0 per cent (anhydrous substance). time = about 13-15 min) : impurity A = about 0.3 ;
CHARACTERS impurity B = about 0.4 ; impurity C = about 0.44 ;
impurity D = about 0.48 ; impurity E = about 0.5 ;
Appearance : white or almost white, crystalline powder.
impurity F = about 0.8 ; impurity G = about 1.6.
Solubility : sparingly soluble in water, freely soluble in dimethyl
sulfoxide, slightly soluble in methanol and in ethanol (96 per System suitability : reference solution (c) :
cent). – resolution : minimum 2.5 between the peaks due to
impurity C and impurity D.
IDENTIFICATION Limits :
A. Infrared absorption spectrophotometry (2.2.24).
– impurity A : not more than the area of the principal peak
Comparison : didanosine CRS. in the chromatogram obtained with reference solution (b)
B. Specific optical rotation (2.2.7) : − 28.2 to − 24.2 (anhydrous (0.5 per cent);
substance). – impurities B, C, D, E, F, G : for each impurity, not more than
Dissolve 0.100 g in water R and dilute to 10.0 mL with the twice the area of the principal peak in the chromatogram
same solvent. obtained with reference solution (a) (0.2 per cent) ;
TESTS – unspecified impurities : for each impurity, not more than the
area of the principal peak in the chromatogram obtained
Related substances. Liquid chromatography (2.2.29). Prepare with reference solution (a) (0.10 per cent) ;
the solutions immediately before use.
Solvent mixture. Mix 8 volumes of mobile phase B and in the chromatogram obtained with reference solution (a)
92 volumes of mobile phase A. (1.0 per cent);
examined in 50.0 mL of the solvent mixture.
the chromatogram obtained with reference solution (a)
Reference solution (a). Dilute 1.0 mL of the test solution to (0.05 per cent).
solution to 10.0 mL with the solvent mixture. Water (2.5.12) : maximum 2.0 per cent, determined on 0.500 g.
Reference solution (b). Dissolve 5.0 mg of didanosine Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
impurity A CRS in the solvent mixture and dilute to 100.0 mL 1.0 g.
with the solvent mixture. Dilute 1.0 mL to 20.0 mL with the
solvent mixture. ASSAY
Reference solution (c). Dissolve 5 mg of didanosine for system Dissolve 0.200 g in 50 mL of glacial acetic acid R. Titrate
suitability CRS (containing impurities A to F) in the solvent with 0.1 M perchloric acid, determining the end-point
mixture and dilute to 10 mL with the solvent mixture. potentiometrically (2.2.20).
Reference solution (d). Dissolve 5 mg of didanosine 1 mL of 0.1 M perchloric acid is equivalent to 23.62 mg
impurity G CRS in the solvent mixture and dilute to 100 mL of C10H12N4O3.
with the solvent mixture. Dilute 1 mL to 20 mL with the
solvent mixture. IMPURITIES
Column : Specified impurities : A, B, C, D, E, F, G.
– size : l = 0.25 m, Ø = 4.6 mm ; Other detectable impurities (the following substances would,
– stationary phase : base-deactivated octadecylsilyl silica gel for
chromatography R (5 μm).
Mobile phase : by the general monograph Substances for pharmaceutical use
– mobile phase A : mix 8 volumes of methanol R and (2034). It is therefore not necessary to identify these impurities
92 volumes of a 3.86 g/L solution of ammonium acetate R for demonstration of compliance. See also 5.10. Control of
adjusted to pH 8.0 with concentrated ammonia R ; impurities in substances for pharmaceutical use) : H, I.

Didanosine EUROPEAN PHARMACOPOEIA
A. 1,7-dihydro-6H-purin-6-one (hypoxanthine),
F. 9-(2,3-dideoxy-β-D-glycero-pent-2-enofuranosyl)-
1,9-dihydro-6H-purin-6-one (2′,3′-dideoxy-2′,3′-
didehydroinosine),
B. 9-β-D-ribofuranosyl-1,9-dihydro-6H-purin-6-one
(inosine),
G. 9-(2,3-dideoxy-β-D-glycero-pentofuranosyl)-9H-purin-6-
amine (2′,3′-dideoxyadenosine),
C. 9-(2-deoxy-β-D-erythro-pentofuranosyl)-1,9-dihydro-6H-
purin-6-one (2′-deoxyinosine),
H. 9-(2,3,5-trideoxy-β-D-glycero-pentofuranosyl)-9H-purin-
6-amine (2′,3′,5′-trideoxyadenosine),
D. 9-(3-deoxy-β-D-erythro-pentofuranosyl)-1,9-dihydro-6H-
purin-6-one (3′-deoxyinosine),
E. 9-(2,3-anhydro-β-D-ribofuranosyl)-1,9-dihydro-6H-purin- I. 9-(2,3-dideoxy-β-D-glycero-pent-2-enofuranosyl)-9H-
6-one (2′,3′-anhydroinosine), purin-6-amine (2′,3′-dideoxy-2′,3′-didehydroadenosine).

EUROPEAN PHARMACOPOEIA Disulfiram
01/2017:0603 Reference solution (a). Dissolve 10 mg of disulfiram CRS in

ethyl acetate R and dilute to 5 mL with the same solvent.
Reference solution (b). Dilute 1 mL of test solution (b) to
20 mL with ethyl acetate R.
Apply to the plate 10 μL of each solution. Develop over a path
DISULFIRAM of 15 cm using a mixture of 30 volumes of butyl acetate R
and 70 volumes of hexane R. Allow the plate to dry in air
Disulfiramum and examine in ultraviolet light at 254 nm. Any spot in the
chromatogram obtained with test solution (a), apart from
the principal spot, is not more intense than the spot in the
chromatogram obtained with reference solution (b) (0.5 per
cent).
Diethyldithiocarbamate. Dissolve 0.20 g in 10 mL of
peroxide-free ether R, add 5 mL of buffer solution pH 8.0 R
C10H20N2S4 Mr 296.5 and shake vigorously. Discard the upper layer and wash
[97-77-8] the lower layer with 10 mL of peroxide-free ether R. Add to
the lower layer 0.2 mL of a 4 g/L solution of copper sulfate
DEFINITION pentahydrate R and 5 mL of cyclohexane R. Shake. Any yellow
Disulfiram contains not less than 98.5 per cent and colour in the upper layer is not more intense than that of a
not more than the equivalent of 101.0 per cent of standard prepared at the same time using 0.2 mL of a freshly
tetraethyldisulfanedicarbothioamide, calculated with prepared 0.15 g/L solution of sodium diethyldithiocarbamate R
reference to the dried substance. (150 ppm).
CHARACTERS Loss on drying (2.2.32). Not more than 0.5 per cent,
determined on 1.000 g by drying in vacuo at 50 °C.
A white or almost white, crystalline powder, practically
insoluble in water, freely soluble in methylene chloride, Sulfated ash (2.4.14). Not more than 0.1 per cent, determined
sparingly soluble in alcohol. on 1.0 g.
IDENTIFICATION ASSAY
First identification : A, B. Dissolve 0.450 g in 80 mL of acetone R and add 20 mL of a
Second identification : A, C, D. 20 g/L solution of potassium nitrate R. Titrate with 0.1 M
silver nitrate. Determine the end-point potentiometrically
A. Melting point (2.2.14): 70 °C to 73 °C. (2.2.20), using a silver electrode and a silver-silver chloride
B. Examine by infrared absorption spectrophotometry double-junction electrode saturated with potassium nitrate.
(2.2.24), comparing with the spectrum obtained with
1 mL of 0.1 M silver nitrate is equivalent to 59.30 mg of
disulfiram CRS. Examine the substances prepared as discs.
C10H20N2S4.
C. Examine the chromatograms obtained in the test for
related substances. The principal spot in the chromatogram STORAGE
obtained with test solution (b) is similar in position and Store protected from light.
size to the principal spot in the chromatogram obtained
with reference solution (a). IMPURITIES
D. Dissolve about 10 mg in 10 mL of methanol R. Add 2 mL
of a 0.5 g/L solution of cupric chloride R in methanol R. A
yellow colour develops which becomes greenish-yellow.
TESTS
Related substances. Examine by thin-layer chromatography
A. diethylthiocarbamic thioanhydride (sulfiram),
(2.2.27), using as the coating substance a suitable silica gel with
a fluorescent indicator having an optimal intensity at 254 nm.
Test solution (a). Dissolve 0.20 g of the substance to be
examined in ethyl acetate R and dilute to 10 mL with the same
solvent.
Test solution (b). Dilute 1 mL of test solution (a) to 10 mL
with ethyl acetate R. B. diethyldithiocarbamate.

EUROPEAN PHARMACOPOEIA Foscarnet sodium hexahydrate
01/2017:1520 Related substances. Liquid chromatography (2.2.29).

corrected 10.0 Test solution. Dissolve 25 mg of the substance to be examined
in the mobile phase and dilute to 10.0 mL with the mobile
phase.
Reference solution (a). Dilute 1.0 mL of the test solution to
50.0 mL with the mobile phase. Dilute 1.0 mL of this solution
FOSCARNET SODIUM HEXAHYDRATE to 10.0 mL with the mobile phase.
Reference solution (b). Dissolve 5 mg of foscarnet
Foscarnetum natricum hexahydricum impurity B CRS in the mobile phase, add 2 mL of the test
solution and dilute to 50 mL with the mobile phase.
Reference solution (c). Dissolve the contents of a vial of
foscarnet impurity mixture CRS (impurities A and C) in 1 mL
of mobile phase.
Column :
CNa3O5P,6H2O Mr 300.0
[34156-56-4] – size : l = 0.10 m, Ø = 4.6 mm ;
– stationary phase : octadecylsilyl silica gel for
DEFINITION chromatography R (3 µm).
Trisodium phosphonatoformate hexahydrate. Mobile phase : dissolve 3.22 g of sodium sulfate decahydrate R
Content : 98.5 per cent to 101.0 per cent (dried substance). in water for chromatography R, add 3 mL of glacial acetic acid R
and 6 mL of a 44.61 g/L solution of sodium pyrophosphate R
CHARACTERS and dilute to 1000 mL with water for chromatography R
Appearance : white or almost white, crystalline powder. (solution A) ; dissolve 3.22 g of sodium sulfate decahydrate R
Solubility : soluble in water, practically insoluble in ethanol in water for chromatography R, add 6.8 g of sodium acetate R
(96 per cent). and 6 mL of a 44.61 g/L solution of sodium pyrophosphate R
and dilute to 1000 mL with water for chromatography R
IDENTIFICATION (solution B). Mix about 700 mL of solution A and about
A. Infrared absorption spectrophotometry (2.2.24). 300 mL of solution B to obtain a solution of pH 4.4. To
Comparison : foscarnet sodium hexahydrate CRS. 1000 mL of this solution, add 0.25 g of tetrahexylammonium
hydrogen sulfate R and 100 mL of methanol R1.
B. It gives reaction (a) of sodium (2.3.1).
TESTS Detection : spectrophotometer at 230 nm.
Solution S. Dissolve 0.5 g in carbon dioxide-free water R and Injection : 40 µL.
Run time : 2.5 times the retention time of foscarnet.
Appearance of solution. Solution S is not more opalescent Identification of impurities : use the chromatogram supplied
than reference suspension I (2.2.1) and is colourless (2.2.2, with foscarnet impurity mixture CRS and the chromatogram
Method II). obtained with reference solution (c) to identify the peaks due
pH (2.2.3) : 9.0 to 11.0 for solution S. to impurities A and C ; use the chromatogram obtained with
Impurity D. Gas chromatography (2.2.28). reference solution (b) to identify the peak due to impurity B.
Test solution. Dissolve 0.250 g of the substance to be examined Relative retention with reference to foscarnet
in 9.0 mL of a 6 g/L solution of glacial acetic acid R using a (retention time = about 5 min) : impurity A = about 0.7 ;
magnetic stirrer. Add 1.0 mL of anhydrous ethanol R and mix. impurity B = about 1.5 ; impurity C = about 2.0.
Reference solution. Dissolve 25.0 mg of foscarnet System suitability : reference solution (b) :
impurity D CRS in anhydrous ethanol R and dilute to 100.0 mL – resolution : minimum 7.0 between the peaks due to
with the same solvent. Dilute 1.0 mL of this solution to foscarnet and impurity B.
10.0 mL with anhydrous ethanol R. Limits :
Column :
– impurities A, B, C : for each impurity, not more than the
– material : fused silica ; area of the principal peak in the chromatogram obtained
– size : l = 25 m, Ø = 0.31 mm ; with reference solution (a) (0.2 per cent) ;
– stationary phase : phenyl(5)methyl(95)polysiloxane R (film – unspecified impurities : for each impurity, not more
thickness 0.5 µm). than 0.25 times the area of the principal peak in the
Carrier gas : helium for chromatography R. chromatogram obtained with reference solution (a)
Split ratio : 1:20. (0.05 per cent) ;
Temperature : – total : not more than twice the area of the principal peak
Time Temperature (0.4 per cent);
(min) (°C)
0-8 100 → 180
Column
Injection port 200 (0.04 per cent); disregard any peak with a relative retention
Detector 250
less than 0.6.
Phosphate and phosphite. Liquid chromatography (2.2.29).
Detection : flame ionisation. Test solution. Dissolve 60.0 mg of the substance to be
Injection : 3 µL examined in water R and dilute to 25.0 mL with the same
Limit : solvent.
– impurity D : not more than the area of the principal peak Reference solution (a). Dissolve 28 mg of sodium dihydrogen
in the chromatogram obtained with the reference solution phosphate monohydrate R in water R and dilute to 100 mL
(0.1 per cent). with the same solvent.

Foscarnet sodium hexahydrate EUROPEAN PHARMACOPOEIA
Reference solution (b). Dissolve 43 mg of sodium phosphite ASSAY

pentahydrate R in water R and dilute to 100 mL with the same Dissolve 0.200 g in 50 mL of water R. Titrate with 0.05 M
solvent. sulfuric acid, determining the end-point potentiometrically
Reference solution (c). Dilute 1.0 mL of reference solution (a) (2.2.20) at the 1st point of inflexion.
and 1.0 mL of reference solution (b) to 25 mL with water R. 1 mL of 0.05 M sulfuric acid is equivalent to 19.20 mg
Reference solution (d). Dilute 3 mL of reference solution (a) of CNa3O5P.
and 3 mL of reference solution (b) to 25 mL with water R.
Column : STORAGE
– size : l = 0.05 m, Ø = 4.6 mm ; Protected from light.
– stationary phase : anion-exchange resin R. IMPURITIES
Mobile phase : dissolve 0.102 g of potassium hydrogen Specified impurities : A, B, C, D.
phthalate R in water for chromatography R, add 2.5 mL
of 1 M nitric acid and dilute to 1000 mL with water for
chromatography R.
Detection : spectrophotometer at 290 nm (indirect detection).
Injection : 20 µL of the test solution and reference solutions (c) A. disodium (ethoxycarbonyl)phosphonate,
and (d).
System suitability : reference solution (d) :
phosphate (1st peak) and phosphite ;
– signal-to-noise ratio : minimum 10 for the principal peak.
B. disodium (ethoxyoxydophosphanyl)formate,
Limits :
– phosphate : not more than the area of the corresponding
solution (c) (0.3 per cent) ;
– phosphite : not more than the area of the corresponding
solution (c) (0.3 per cent). C. ethyl sodium (ethoxycarbonyl)phosphonate,
Loss on drying (2.2.32) : 35.0 per cent to 37.0 per cent,
determined on 1.000 g by drying in an oven at 150 °C.
Bacterial endotoxins (2.6.14) : less than 83.3 IU/g, if intended
for use in the manufacture of parenteral preparations without
a further appropriate procedure for the removal of bacterial
endotoxins. D. ethyl (diethoxyphosphoryl)formate.

EUROPEAN PHARMACOPOEIA Ganciclovir
01/2017:1752 Detection : spectophotometer at 254 nm.

Injection : 20 µL.
Run time : 2.5 times the retention time of ganciclovir.
with ganciclovir impurity mixture CRS and the chromatogram
GANCICLOVIR obtained with reference solution (c) to identify the peaks due
to impurities A, B, C, D, E and F.
Ganciclovirum Relative retention with reference to ganciclovir
(retention time = about 14 min) : impurity A = about 0.6 ;
impurity B = about 0.67 ; impurity C = about 0.71 ;
impurity D = about 0.8 ; impurity E = about 0.9 ;
impurity F = about 2.0.
– peak-to-valley ratio : minimum 5, where Hp = height above
the baseline of the peak due to impurity E and Hv = height
above the baseline of the lowest point of the curve
C9H13N5O4 Mr 255.2 separating this peak from the peak due to ganciclovir.
[82410-32-0]
Limits :
DEFINITION – correction factors : for the calculation of content,
2-Amino-9-[[2-hydroxy-1-(hydroxymethyl)ethoxy]methyl]- multiply the peak areas of the following impurities by
1,9-dihydro-6H-purin-6-one. the corresponding correction factor : impurity B = 1.3 ;
Content : 99.0 per cent to 101.0 per cent (anhydrous substance). impurity F = 0.7 ;
– impurity F : not more than 4 times the area of the principal
CHARACTERS peak in the chromatogram obtained with reference
Appearance : white or almost white, hygroscopic, crystalline solution (a) (0.4 per cent);
powder. – impurity B : not more than twice the area of the principal
Solubility : slightly soluble in water, very slightly soluble in peak in the chromatogram obtained with reference
ethanol (96 per cent). It dissolves in dilute solutions of mineral solution (a) (0.2 per cent);
acids and alkali hydroxides. – impurities A, C, D, E : for each impurity, not more than
It shows polymorphism (5.9). 1.5 times the area of the principal peak in the chromatogram
obtained with reference solution (a) (0.15 per cent) ;
IDENTIFICATION
– unspecified impurities : for each impurity, not more than
Infrared absorption spectrophotometry (2.2.24). 0.5 times the area of the principal peak in the chromatogram
Comparison : ganciclovir CRS. obtained with reference solution (a) (0.05 per cent) ;
If the spectra obtained in the solid state show differences, – total : not more than 6 times the area of the principal peak
dissolve 0.10 g of the substance to be examined and the in the chromatogram obtained with reference solution (a)
reference substance separately in about 3.6 mL of water R at (0.6 per cent);
80 °C. Allow to cool in an ice-bath and filter the precipitate. – disregard limit : 0.3 times the area of the principal peak in
Dry in an oven at 105 °C for 3 h and record new spectra using the chromatogram obtained with reference solution (a)
the residues. (0.03 per cent).
TESTS Water (2.5.12) : maximum 4.0 per cent, determined on 0.300 g.
Appearance of solution. The solution is clear (2.2.1) and not Use methanol R as solvent. The substance to be examined has
more intensely coloured than reference solution Y5 (2.2.2, limited solubility in methanol. The sample will appear as a
Method II). slurry. Replace the solvent after each titration.
Dissolve 1.25 g in a 40 g/L solution of sodium hydroxide R and Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
dilute to 25 mL with the same solution. 1.0 g.
Related substances. Liquid chromatography (2.2.29). Bacterial endotoxins (2.6.14) : less than 0.84 IU/mg, if
Test solution. Dissolve 30 mg of the substance to be examined intended for use in the manufacture of parenteral preparations
in the mobile phase with the aid of ultrasound and dilute to without a further appropriate procedure for the removal of
50.0 mL with the mobile phase. bacterial endotoxins.
Reference solution (a). Dilute 1.0 mL of the test solution to ASSAY
100.0 mL with the mobile phase. Dilute 1.0 mL of this solution Dissolve 0.200 g in 10 mL of anhydrous formic acid R and
to 10.0 mL with the mobile phase. dilute to 60 mL with glacial acetic acid R. Titrate with 0.1 M
Reference solution (b). Dissolve 3 mg of ganciclovir CRS in the perchloric acid, determining the end point potentiometrically
mobile phase with the aid of ultrasound and dilute to 5.0 mL (2.2.20).
with the mobile phase. 1 mL of 0.1 M perchloric acid is equivalent to 25.52 mg of
Reference solution (c). Dissolve the contents of a vial of C9H13N5O4.
ganciclovir impurity mixture CRS (impurities A, B, C, D, E
and F) in 1.0 mL of reference solution (b). STORAGE
Column : In an airtight container.
– size : l = 0.25 m, Ø = 4.6 mm ; IMPURITIES
– stationary phase : strong cation-exchange silica gel for Specified impurities : A, B, C, D, E, F.
chromatography R (10 µm);
– temperature : 40 °C. if present at a sufficient level, be detected by one or other of
Mobile phase : mix equal volumes of acetonitrile R and a the tests in the monograph. They are limited by the general
0.05 per cent V/V solution of trifluoroacetic acid R. acceptance criterion for other/unspecified impurities and/or
Flow rate : 1.5 mL/min. by the general monograph Substances for pharmaceutical use

Ganciclovir EUROPEAN PHARMACOPOEIA

impurities in substances for pharmaceutical use) : H, I, J.
E. 2-amino-9-[[(2RS)-2,3-dihydroxypropoxy]methyl]-1,9-
dihydro-6H-purin-6-one,
A. 2-amino-9-[[(2-chloroprop-2-en-1-yl)oxy]methyl]-1,9-
dihydro-6H-purin-6-one,
F. 2-amino-1,9-dihydro-6H-purin-6-one (guanine),
H. 2-amino-7-[[2-hydroxy-1-(hydroxymethyl)ethoxy]-
methyl]-1,7-dihydro-6H-purin-6-one,
B. (2RS)-2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]-3-hydroxypropyl propionate,
I. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
C. 2-amino-9-[[(1RS)-2-chloro-1-(hydroxymethyl)ethoxy]- yl)methoxy]propane-1,3-diyl dipropanoate,
methyl]-1,9-dihydro-6H-purin-6-one,
D. 2-amino-9-[[[2-hydroxy-1-(hydroxymethyl)ethoxy]- J. 2-[2-(propanoylamino)-6-oxo-1,6-dihydro-9H-purin-9-
methoxy]methyl]-1,9-dihydro-6H-purin-6-one, yl]methoxy]propane-1,3-diyl dipropanoate.

EUROPEAN PHARMACOPOEIA Hydroxychloroquine sulfate
01/2017:2849 Column :
corrected 10.0 – size : l = 0.05 m, Ø = 2.1 mm ;
– stationary phase : end-capped ethylene-bridged octadecylsilyl
silica gel for chromatography (hybrid material) R (1.7 µm);
HYDROXYCHLOROQUINE SULFATE Mobile phase :
– mobile phase A : methanol R, buffer solution (10:90 V/V) ;
Hydroxychloroquini sulfas – mobile phase B : buffer solution, methanol R (15:85 V/V);
0-1 100 0
1 - 11 100 → 0 0 → 100

Injection : 4 μL of test solution (a) and reference solutions (a)
C18H28ClN3O5S Mr 434.0 and (b).
[747-36-4]
DEFINITION with hydroxychloroquine for system suitability CRS and the
chromatogram obtained with reference solution (a) to identify
2-[[(4RS)-4-[(7-Chloroquinolin-4-yl)amino]pentyl]- the peaks due to impurities B and C.
(ethyl)amino]ethan-1-ol sulfate.
Relative retention with reference to hydroxychloroquine
Content : 98.0 per cent to 102.0 per cent (dried substance). (retention time = about 6 min): impurity B = about 0.8 ;
impurity C = about 0.9.
CHARACTERS
Appearance : white or slightly yellowish, crystalline powder. – resolution : minimum 3.0 between the peaks due to
Solubility : freely soluble in water, practically insoluble in impurity C and hydroxychloroquine ; minimum 3.0
ethanol (96 per cent) and in methylene chloride. between the peaks due to impurities B and C.
Calculation of percentage contents :
IDENTIFICATION
– correction factor : multiply the peak area of impurity B by
A. Infrared absorption spectrophotometry (2.2.24). 1.6 ;
Comparison : hydroxychloroquine sulfate CRS. – for each impurity, use the concentration of
B. It gives reaction (a) of sulfates (2.3.1). hydroxychloroquine sulfate in reference solution (b).
Limits :
TESTS – impurity C : maximum 0.4 per cent ;
Appearance of solution. The solution is clear and not more – impurity B : maximum 0.15 per cent ;
intensely coloured than reference solution Y7 (2.2.2, Method I).
– unspecified impurities : for each impurity, maximum
Dissolve 1.0 g in water R and dilute to 10.0 mL with the same 0.10 per cent ;
solvent.
– total : maximum 0.6 per cent ;
Related substances. Liquid chromatography (2.2.29).
– reporting threshold : 0.05 per cent.
Solvent mixture. Mix 500 mL of water R, 500 mL of methanol R
Loss on drying (2.2.32) : maximum 2.0 per cent, determined
and 4 mL of a 10 per cent V/V solution of sulfuric acid R.
on 1.000 g by drying in an oven at 105 °C for 2 h.
Buffer solution. Dissolve 1.36 g of potassium dihydrogen
Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
phosphate R in 900 mL of water for chromatography R.
1.0 g.
Add 0.15 g of sodium heptanesulfonate R, adjust to pH 7.0
with triethylamine R and dilute to 1000 mL with water for ASSAY
chromatography R.
Test solution (a). Dissolve 25.0 mg of the substance to be related substances with the following modification.
examined in the solvent mixture and dilute to 50.0 mL with
the solvent mixture. Injection : 4 μL of test solution (b) and reference solution (c).
Test solution (b). Dilute 1.0 mL of test solution (a) to 10.0 mL Calculate the percentage content of C18H28ClN3O5S taking
with the solvent mixture. into account the assigned content of hydroxychloroquine
sulfate CRS.
Reference solution (a). Dissolve 5 mg of hydroxychloroquine
for system suitability CRS (containing impurities B and C) IMPURITIES
in the solvent mixture and dilute to 10 mL with the solvent Specified impurities : B, C.
mixture.
Reference solution (b). Dilute 1.0 mL of test solution (a) to if present at a sufficient level, be detected by one or other of
10.0 mL with the solvent mixture. Dilute 2.0 mL of this the tests in the monograph. They are limited by the general
solution to 100.0 mL with the solvent mixture. acceptance criterion for other/unspecified impurities and/or
Reference solution (c). Dissolve 25.0 mg of hydroxychloroquine by the general monograph Substances for pharmaceutical use
sulfate CRS in the solvent mixture and dilute to 50.0 mL with (2034). It is therefore not necessary to identify these impurities
the solvent mixture. Dilute 1.0 mL of the solution to 10.0 mL for demonstration of compliance. See also 5.10. Control of
with the solvent mixture. impurities in substances for pharmaceutical use) : A, D, E, F, G.

Hydroxychloroquine sulfate EUROPEAN PHARMACOPOEIA
A. mixture of diastereoisomers of 4-[(7-chloroquinolin- D. (4RS)-N4-(7-chloroquinolin-4-yl)-N1-ethylpentane-1,4-

4-yl)amino]-N-ethyl-N-(hydroxyethyl)pentan-1-amine diamine,
N-oxide,
E. (4RS)-4-[(7-chloroquinolin-4-yl)amino]pentan-1-ol,
B. 2-[[(4RS)-4-[(7-chloroquinolin-4-yl)amino]pentyl]-
(ethyl)amino]ethyl hydrogen sulfate,
F. 7-chloro-4-[(2RS)-2-methylpyrrolidin-1-yl]quinoline,
C. 2-[[(4RS)-4-[(7-chloroquinolin-4-yl)amino]pentyl]-
amino]ethan-1-ol, G. 4,7-dichloroquinoline.

EUROPEAN PHARMACOPOEIA Idoxuridine
01/2008:0669 Related substances. Examine by thin-layer chromatography

(2.2.27), using as coating substance a suitable silica gel with a
fluorescent indicator having an optimal intensity at 254 nm.
Test solution (a). Dissolve 0.20 g of the substance to be
examined in a mixture of 1 volume of concentrated ammonia R
IDOXURIDINE and 5 volumes of methanol R and dilute to 5 mL with the same
mixture of solvents.
Test solution (b). Dilute 1 mL of test solution (a) to 10 mL
Idoxuridinum with a mixture of 1 volume of concentrated ammonia R and
5 volumes of methanol R.
Reference solution (a). Dissolve 20 mg of 5-iodouracil R, 20 mg
of 2′-deoxyuridine R and 20 mg of 5-bromo-2′-deoxyuridine R
in a mixture of 1 volume of concentrated ammonia R and
5 volumes of methanol R and dilute to 100 mL with the same
mixture of solvents.
Reference solution (b). Dissolve 0.20 g of the substance to be
examined in 5 mL of reference solution (a).
Reference solution (c). Dissolve 20 mg of idoxuridine CRS
C9H11IN2O5 Mr 354.1
in a mixture of 1 volume of concentrated ammonia R and
[54-42-2]
5 volumes of methanol R and dilute to 5 mL with the same
DEFINITION mixture of solvents.
Idoxuridine contains not less than 98.0 per cent and not more Reference solution (d). Dilute 1 mL of test solution (b) to
than the equivalent of 101.0 per cent of 5-iodo-1-(2-deoxy- 20 mL with a mixture of 1 volume of concentrated ammonia R
β-D-erythro-pentofuranosyl)pyrimidine-2,4(1H,3H)-dione, and 5 volumes of methanol R.
calculated with reference to the dried substance. Apply separately to the plate 5 μL of each solution. Develop
twice over a path of 15 cm using a mixture of 10 volumes of
CHARACTERS concentrated ammonia R, 40 volumes of chloroform R and
A white or almost white, crystalline powder, slightly soluble 50 volumes of 2-propanol R, drying the plate in a current of
in water and in ethanol (96 per cent). It dissolves in dilute cold air after each development. Examine in ultraviolet light at
solutions of alkali hydroxides. 254 nm. In the chromatogram obtained with test solution (a) :
It melts at about 180 °C, with decomposition. any spots corresponding to 5-iodouracil, 2′-deoxyuridine
and 5-bromo-2′-deoxyuridine are not more intense than the
IDENTIFICATION corresponding spots in the chromatogram obtained with
First identification : A. reference solution (a) (0.5 per cent) ; any spot, apart from the
principal spot and the spots corresponding to 5-iodouracil,
Second identification : B, C, D. 2′-deoxyuridine and 5-bromo-2′-deoxyuridine, is not more
A. Examine by infrared absorption spectrophotometry intense than the spot in the chromatogram obtained with
(2.2.24), comparing with the spectrum obtained with reference solution (d) (0.5 per cent). The test is not valid
idoxuridine CRS. Examine the substances as discs prepared unless the chromatogram obtained with reference solution (b)
using 1 mg of the substance to be examined and of the shows four clearly separated spots.
reference substance each in 0.3 g of potassium bromide R. Iodide. Dissolve 0.25 g in 25 mL of 0.1 M sodium hydroxide,
B. Examine the chromatograms obtained in the test for related add 5 mL of dilute hydrochloric acid R and dilute to 50 mL
substances. The principal spot in the chromatogram with water R. Allow to stand for 10 min and filter. To 25 mL
obtained with test solution (b) is similar in position and of the filtrate add 5 mL of dilute hydrogen peroxide solution R
size to the principal spot in the chromatogram obtained and 10 mL of chloroform R and shake. Any pink colour in
with reference solution (c). the organic layer is not more intense than that in a standard
C. Heat about 5 mg in a test-tube over a naked flame. Violet prepared at the same time in the same manner using 1 mL
vapour is evolved. of a 0.33 g/L solution of potassium iodide R instead of the
substance to be examined (0.1 per cent).
D. Disperse about 2 mg in 1 mL of water R and add 2 mL
of diphenylamine solution R2. Heat in a water-bath for Loss on drying (2.2.32). Not more than 1.0 per cent,
10 min. A persistent light-blue colour develops. determined on 1.000 g by drying in vacuo at 60 °C.
Sulfated ash (2.4.14). Not more than 0.1 per cent, determined
TESTS on 1.0 g.
Solution S. Dissolve 0.500 g in 1 M sodium hydroxide and
dilute to 50.0 mL with the same solvent. ASSAY
Appearance of solution. Solution S is clear (2.2.1) and Dissolve 0.3000 g in 20 mL of dimethylformamide R. Titrate
colourless (2.2.2, Method II). with 0.1 M tetrabutylammonium hydroxide, determining the
end-point potentiometrically (2.2.20).
pH (2.2.3). Dissolve 0.10 g in carbon dioxide-free water R 1 mL of 0.1 M tetrabutylammonium hydroxide is equivalent to
and dilute to 100 mL with the same solvent. The pH of the 35.41 mg of C9H11IN2O5.
solution is 5.5 to 6.5.
Specific optical rotation (2.2.7) : + 28 to + 32, determined on STORAGE
solution S and calculated with reference to the dried substance. Store protected from light.

EUROPEAN PHARMACOPOEIA Indinavir sulfate
01/2017:2214 Reference solution (d). To 30 mg of the substance to be

examined add 0.25 mL of 2 M hydrochloric acid R and allow
to stand at room temperature for 1 h. Dilute to 100 mL with
a mixture of 2 volumes of acetonitrile R1 and 3 volumes
of mobile phase A and mix (in situ degradation to obtain
INDINAVIR SULFATE impurity D).
Column :
– size : l = 0.25 m, Ø = 4.6 mm ;
Indinaviri sulfas
– stationary phase : octadecylsilyl silica gel for
chromatography R (5 µm).
Mobile phase :
– mobile phase A : solution containing 0.27 g/L of potassium
dihydrogen phosphate R and 1.40 g/L of dipotassium
hydrogen phosphate R ; filter and degas ;
– mobile phase B : acetonitrile R1 ;
C36H49N5O8S,C2H6O Mr 758 0-5 80 20
[157810-81-6]
5 - 40 80 → 30 20 → 70
DEFINITION 40 - 45 30 70
(2S)-1-[(2S,4R)-4-Benzyl-2-hydroxy-5-[[(1S,2R)-2-hydroxy-
2,3-dihydro-1H-inden-1-yl]amino]-5-oxopentyl]-N-(1,1- 45 - 47 30 → 80 70 → 20
dimethylethyl)-4-(pyridin-3-ylmethyl)piperazine-2- 47 - 52 80 20
carboxamide sulfate ethanolate.
Content : 98.0 per cent to 102.0 per cent (anhydrous and Flow rate : 1.0 mL/min.
ethanol-free substance). Detection : spectrophotometer at 220 nm.
Injection : 20 µL.
PRODUCTION
A test for enantiomeric purity is carried out unless it has supplied with indinavir for system suitability CRS and the
been demonstrated that the manufacturing process is chromatogram obtained with reference solution (a) to identify
enantioselective for the substance. the peaks due to impurities B, C and E ; use the chromatogram
obtained with reference solution (d) to identify the peak due
CHARACTERS to impurity D.
Appearance : white or almost white, hygroscopic powder. Relative retention with reference to indinavir (retention
Solubility : freely soluble in water, soluble in methanol, time = about 25 min) : impurity A = about 0.2 ;
practically insoluble in heptane. impurity B = about 0.8 ; impurity C = about 0.98 ;
impurity D = about 1.1 ; impurity E = about 1.3.
IDENTIFICATION
A. Specific optical rotation (2.2.7): + 122 to + 129 (anhydrous
and ethanol-free substance), determined at 365 nm and at
impurity C and indinavir.
25 °C.
Limits :
Dissolve 0.500 g in water R and dilute to 50.0 mL with the
same solvent. – correction factor : for the calculation of content, multiply
the peak area of impurity D by 1.8 ;
B. Infrared absorption spectrophotometry (2.2.24).
– impurity A : not more than the area of the principal peak
Comparison : Ph. Eur. reference spectrum of indinavir in the chromatogram obtained with reference solution (c)
sulfate. (0.1 per cent);
C. It gives reaction (a) of sulfates (2.3.1). – impurity D : not more than twice the area of the principal
D. Ethanol (see Tests). peak in the chromatogram obtained with reference
solution (b) (0.2 per cent);
TESTS – impurities B, C, E : for each impurity, not more than the
Related substances. Liquid chromatography (2.2.29). area of the principal peak in the chromatogram obtained
Solution A. Thoroughly mix equal volumes of mobile phase A with reference solution (b) (0.1 per cent) ;
and acetonitrile R1. – unspecified impurities : for each impurity, not more than
Test solution. Dissolve 50.0 mg of the substance to be 0.5 times the area of the principal peak in the chromatogram
examined in solution A and dilute to 100.0 mL with the same obtained with reference solution (b) (0.05 per cent) ;
solution. – total : not more than 5 times the area of the principal peak
Reference solution (a). Dissolve 4 mg of indinavir for in the chromatogram obtained with reference solution (b)
system suitability CRS (containing impurities B, C and E) in (0.5 per cent);
solution A and dilute to 10 mL with the same solution. – disregard limit : 0.3 times the area of the principal peak in
Reference solution (b). Dilute 1.0 mL of the test solution to the chromatogram obtained with reference solution (b)
100.0 mL with solution A. Dilute 1.0 mL of this solution to (0.03 per cent).
10.0 mL with solution A. Ethanol. Gas chromatography (2.2.28).
Reference solution (c). Dissolve 5.0 mg of cis-aminoindanol R Internal standard solution. Dilute 1.0 mL of propanol R to
(impurity A) in solution A and dilute to 10.0 mL with the 200.0 mL with water R.
same solution. Dilute 1.0 mL of the solution to 100.0 mL with Test solution. Dissolve 0.400 g of the substance to be examined
solution A. Dilute 1.0 mL of this solution to 10.0 mL with in 50.0 mL of water R, add 8.0 mL of the internal standard
solution A. solution and dilute to 100.0 mL with water R.

Indinavir sulfate EUROPEAN PHARMACOPOEIA
Reference solution. Dilute 1.0 mL of anhydrous ethanol R to by the general monograph Substances for pharmaceutical
200.0 mL. Dilute 2.0 mL of this solution and 2.0 mL of the use (2034). It is therefore not necessary to identify these
internal standard solution to 25.0 mL with water R. impurities for demonstration of compliance. See also 5.10.
Column : Control of impurities in substances for pharmaceutical use) : F.
– material : fused silica ;
– size : l = 30 m, Ø = 0.53 mm ;
– stationary phase : macrogol 20 000 R (film thickness 1.0 μm).
Split ratio : 1:10. A. (1S,2R)-1-amino-2,3-dihydro-1H-inden-2-ol
Temperature : (cis-aminoindanol),
– column : 35 °C ;
– injection port : 140 °C ;
– detector : 220 °C.
Injection : 1.0 µL.
System suitability : reference solution :
– retention time : ethanol = 2 min to 4 min ;
– resolution : minimum 5.0 between the peaks due to ethanol B. (2S)-1-[(2S,4R)-4-benzyl-2-hydroxy-5-[[(1S,2R)-2-
and propanol. hydroxy-2,3-dihydro-1H-inden-1-yl]amino]-5-oxopentyl]-
N-(1,1-dimethylethyl)piperazine-2-carboxamide,
Calculate the percentage content of ethanol taking the density
(2.2.5) to be 0.790 g/mL.
Limit :
– ethanol : 5.0 per cent to 8.0 per cent m/m.
Water (2.5.12): maximum 1.5 per cent, determined on 0.500 g.
1.0 g.
ASSAY
C. (2S)-1-[(2R,4R)-4-benzyl-2-hydroxy-5-[[(1S,2R)-2-
Liquid chromatography (2.2.29). hydroxy-2,3-dihydro-1H-inden-1-yl]amino]-5-oxopentyl]-
Solution B. Add 20 mL of dibutylammonium phosphate for N-(1,1-dimethylethyl)-4-(pyridin-3-ylmethyl)piperazine-
ion-pairing R to 1000 mL of water R. Adjust to pH 6.5 with 2-carboxamide,
1 M sodium hydroxide.
examined in the mobile phase and dilute to 100.0 mL with
the mobile phase.
Reference solution. Dissolve 50.0 mg of indinavir CRS in the
mobile phase and dilute to 100.0 mL with the mobile phase.
Column :
– size : l = 0.25 m, Ø = 4.6 mm ; D. (3R,5S)-3-benzyl-5-[[(2S)-2-[(1,1-dimethylethyl)-
– stationary phase : base-deactivated octylsilyl silica gel for carbamoyl]-4-(pyridin-3-ylmethyl)piperazin-1-yl]methyl]-
chromatography R (5 μm) ; 4,5-dihydrofuran-2(3H)-one,
Mobile phase : acetonitrile R, solution B (45:55 V/V).
Injection : 10 µL.
Run time : twice the retention time of indinavir.
Retention time: indinavir = about 10 min.
declared content of indinavir CRS and multiplying by a
correction factor of 1.1598.
STORAGE
In an airtight container, protected from light. E. (2S)-1,4-bis[(2S,4R)-4-benzyl-2-hydroxy-5-[[(1S,2R)-2-
hydroxy-2,3-dihydro-1H-inden-1-yl]amino]-5-oxopentyl]-
IMPURITIES N-(1,1-dimethylethyl)piperazine-2-carboxamide,
Specified impurities : A, B, C, D, E.
acceptance criterion for other/unspecified impurities and/or F. 3-(chloromethyl)pyridine (nicotinyl chloride).

EUROPEAN PHARMACOPOEIA Lamivudine
01/2017:2217 Reference solution (d). Dissolve 5 mg of cytosine R and 5 mg of

uracil R in the mobile phase and dilute to 100.0 mL with the
mobile phase. Dilute 2.0 mL of the solution to 10.0 mL with
the mobile phase.
Reference solution (e). Dissolve 5 mg of lamivudine for system
LAMIVUDINE suitability 1 CRS (containing impurities A and B) in 2 mL of
the mobile phase. Add 1.0 mL of reference solution (d) and
dilute to 10.0 mL with the mobile phase.
Lamivudinum Column :
– size : l = 0.25 m, Ø = 4.6 mm ;
– stationary phase : base-deactivated octadecylsilyl silica gel for
chromatography R (5 µm) ;
Mobile phase : mix 5 volumes of methanol R and 95 volumes of
a 1.9 g/L solution of ammonium acetate R, previously adjusted
to pH 3.8 with glacial acetic acid R.
C8H11N3O3S Mr 229.3 Detection : spectrophotometer at 277 nm.
[134678-17-4]
Injection : 10 µL.
DEFINITION Run time : 3 times the retention time of lamivudine.
4-Amino-1-[(2R,5S)-2-(hydroxymethyl)-1,3-oxathiolan-5- Identification of impurities : use the chromatograms obtained
yl]pyrimidin-2(1H)-one. with reference solutions (b) and (e) to identify the peaks due
to impurities A, B, E, F and C.
Content : 97.5 per cent to 102.0 per cent (dried substance).
Relative retention with reference to lamivudine
CHARACTERS (retention time = about 9 min): impurity E = about 0.28 ;
impurity F = about 0.32 ; impurity A = about 0.36 ;
Appearance : white or almost white powder. impurity B = about 0.91 ; impurity J = about 1.45 ;
Solubility : soluble in water, sparingly soluble in methanol, impurity C = about 2.32.
slightly soluble in ethanol (96 per cent). System suitability : reference solution (e):
It shows polymorphism (5.9). – resolution : minimum 1.5 between the peaks due to
impurities F and A ; minimum 1.5 between the peaks due
IDENTIFICATION to impurity B and lamivudine.
First identification : B, C. Limits :
Second identification : A, B. – correction factors : for the calculation of content,
multiply the peak areas of the following impurities by
A. Specific optical rotation (2.2.7) : − 99 to − 97 (dried
substance). the corresponding correction factor : impurity E = 0.6 ;
impurity F = 2.2 ; impurity J = 2.2 ;
Dissolve 0.250 g in water R and dilute to 50.0 mL with the – impurity A : not more than 3 times the area of the principal
same solvent. peak in the chromatogram obtained with reference
B. Infrared absorption spectrophotometry (2.2.24). solution (a) (0.3 per cent);
Comparison : lamivudine CRS. – impurity B : not more than twice the area of the principal
If the spectra obtained in the solid state show differences, peak in the chromatogram obtained with reference
dissolve the substance to be examined and the reference solution (a) (0.2 per cent);
substance separately in methanol R, evaporate to dryness – impurity C : not more than the area of the principal peak
and record new spectra using the residues. in the chromatogram obtained with reference solution (b)
(0.1 per cent);
C. Enantiomeric purity (see Tests).
– any other impurity : for each impurity, not more than the
TESTS area of the principal peak in the chromatogram obtained
with reference solution (a) (0.1 per cent) ;
Absorbance (2.2.25) : maximum 0.3 at 440 nm, using a path
length of 4 cm.
Dissolve 1.00 g in water R, using sonication if necessary, and (0.6 per cent);
dilute to 20.0 mL with the same solvent. – disregard limit : 0.5 times the area of the principal peak in
Related substances. Liquid chromatography (2.2.29). the chromatogram obtained with reference solution (a)
Test solution. Dissolve 50.0 mg of the substance to be (0.05 per cent).
examined in the mobile phase and dilute to 100.0 mL with Enantiomeric purity. Liquid chromatography (2.2.29) : use
the mobile phase. the normalisation procedure.
Reference solution (a). Dilute 1.0 mL of the test solution to Test solution. Dissolve 25.0 mg of the substance to be
100.0 mL with the mobile phase. Dilute 1.0 mL of this solution examined in water R and dilute to 100.0 mL with the same
to 10.0 mL with the mobile phase. solvent.
Reference solution (b). Dissolve 5 mg of salicylic acid R in the Reference solution. Dissolve the contents of a vial of lamivudine
mobile phase and dilute to 100.0 mL with the mobile phase. for system suitability 2 CRS (containing impurity D) in 1.0 mL
Dilute 1.0 mL of the solution to 100.0 mL with the mobile of water R.
phase. Column :
Reference solution (c). Dissolve 50.0 mg of lamivudine CRS – size : l = 0.25 m, Ø = 4.6 mm ;
in the mobile phase and dilute to 100.0 mL with the mobile – stationary phase : silica gel BC for chiral chromatography R
phase. (5 μm) ;

Lamivudine EUROPEAN PHARMACOPOEIA
– temperature : maintain at constant temperature between

15 °C and 30 °C ; the temperature may be adjusted
to optimise the resolution between lamivudine and
impurity D ; a lower temperature favours improved
resolution.
Mobile phase : mix 5 volumes of methanol R and 95 volumes of
a 7.7 g/L solution of ammonium acetate R.
Flow rate : 1.0 mL/min. B. 4-amino-1-[(2RS,5RS)-2-(hydroxymethyl)-1,3-oxathiolan-
Detection : spectrophotometer at 270 nm. 5-yl]pyrimidin-2(1H)-one ((±)-trans-lamivudine),
Injection : 10 µL.
Run time : twice the retention time of lamivudine.
Relative retention with reference to lamivudine (retention
time = about 8 min) : impurity D = about 1.2 ; impurity B and
enantiomer = about 1.3 and 1.5. C. 2-hydroxybenzenecarboxylic acid (salicylic acid),
– peak-to-valley-ratio : minimum 15, where Hp = height above
the baseline of the peak due to impurity D and Hv = height
above the baseline of the lowest point of the curve
separating this peak from the peak due to lamivudine.
Calculate the sum of the percentage contents of all impurity
peaks with a relative retention from 1.2 to 1.5. Subtract the
percentage content of impurity B as obtained in the test for
related substances.
Limit : D. 4-amino-1-[(2S,5R)-2-(hydroxymethyl)-1,3-oxathiolan-
5-yl]pyrimidin-2(1H)-one,
– impurity D : maximum 0.3 per cent.
on 1.000 g by drying in an oven at 105 °C.
1.0 g.
ASSAY
Liquid chromatography (2.2.29) as described in the test for E. 4-aminopyrimidin-2(1H)-one (cytosine),
related substances with the following modification.
Injection : test solution and reference solution (c).
chromatograms obtained with the test solution and reference
solution (c) and the declared content of C8H11N3O3S in
lamivudine CRS.
F. pyrimidine-2,4(1H,3H)-dione (uracil),
STORAGE
Protected from light.
IMPURITIES
Specified impurities : A, B, C, D.
(2034). It is therefore not necessary to identify these impurities G. 4-amino-1-[(2R,3S,5S)-2-(hydroxymethyl)-1,3-oxathiolan-
for demonstration of compliance. See also 5.10. Control of 5-yl]pyrimidin-2(1H)-one S-oxide,
impurities in substances for pharmaceutical use) : E, F, G, H, I, J.
A. (2RS,5SR)-5-(4-amino-2-oxopyrimidin-1(2H)-yl)-1,3- H. 4-amino-1-[(2R,3R,5S)-2-(hydroxymethyl)-1,3-oxathiolan-
oxathiolane-2-carboxylic acid, 5-yl]pyrimidin-2(1H)-one S-oxide,

EUROPEAN PHARMACOPOEIA Lamivudine
I. 4-amino-1-[(2S,4S)-2-(hydroxymethyl)-1,3-dioxolan-4- J. 1-[(2R,5S)-2-(hydroxymethyl)-1,3-oxathiolan-5-
yl]pyrimidin-2(1H)-one, yl]pyrimidine-2,4(1H,3H)-dione.

EUROPEAN PHARMACOPOEIA Lopinavir
01/2017:2615 Reference solution (c). Dissolve 2.5 mg of lopinavir for

system suitability CRS (containing impurities A, B, C, F, G,
I, N, Q, R, S and T) in the solvent mixture and dilute to
5.0 mL with the solvent mixture.
Reference solution (d). Dissolve 2.5 mg of lopinavir for
LOPINAVIR peak identification CRS (containing impurities D and O) in
the solvent mixture and dilute to 5.0 mL with the solvent
mixture.
Lopinavirum Column :
– size : l = 0.25 m, Ø = 4.6 mm ;
– stationary phase : end-capped octadecylsilyl silica gel for
Mobile phase :
– mobile phase A : acetonitrile R1, phosphate buffer
solution (45:55 V/V) ;
– mobile phase B : phosphate buffer solution, acetonitrile R1
(25:75 V/V) ;
C37H48N4O5 Mr 629
[192725-17-0] Time Mobile phase A Mobile phase B
DEFINITION 0 - 60 100 0
(2S)-N-[(1S,3S,4S)-1-Benzyl-4-[[2-(2,6-dimethylphenoxy)- 60 - 61 100 → 0 0 → 100
acetyl]amino]-3-hydroxy-5-phenylpentyl]-3-methyl-2-[2-oxo-
tetrahydropyrimidin-1(2H)-yl]butanamide. 61 - 81 0 100
Content : 98.0 per cent to 102.0 per cent (anhydrous substance).
CHARACTERS Detection : spectrophotometer at 215 nm.
Appearance : white or yellowish-white, slightly hygroscopic Injection : 20 µL of test solution (a) and reference
powder. solutions (b), (c) and (d).
Solubility : practically insoluble in water, very soluble in Identification of impurities : use the chromatogram
methanol and in methylene chloride. supplied with lopinavir for system suitability CRS and the
It shows polymorphism (5.9). chromatogram obtained with reference solution (c) to
identify the peaks due to impurities A, B, C, F, G, I and
IDENTIFICATION N ; use the chromatogram supplied with lopinavir for
A. Specific optical rotation (see Tests). peak identification CRS and the chromatogram obtained
B. Infrared absorption spectrophotometry (2.2.24). with reference solution (d) to identify the peak due to
impurity D.
Comparison : lopinavir CRS.
Relative retention r (not rG) with reference to lopinavir
If the spectra obtained in the solid state show differences, (retention time = about 37 min): impurity A = about 0.03 ;
dissolve the substance to be examined and the reference impurity B = about 0.07 ; impurity C = about 0.10 ;
substance separately in methanol R, evaporate to dryness impurity D = about 0.13 ; impurity F = about 0.59 ;
and record new spectra using the residues. impurity G = about 0.62 ; impurity I = about 1.1 ;
TESTS impurity N = about 1.4.
System suitability : reference solution (c):
Specific optical rotation (2.2.7) : − 27.0 to − 22.0 (anhydrous
substance). – resolution : minimum 1.5 between the peaks due to
impurities F and G.
Dissolve 0.200 g in methanol R and dilute to 25.0 mL with the
same solvent. Calculation of percentage contents :
Related substances – for impurity A, multiply the peak area by the correction
factor 1.6 ;
A. Liquid chromatography (2.2.29).
– for impurity B, multiply the peak area by the correction
Solvent mixture : acetonitrile R1, water R (50:50 V/V). factor 1.3 ;
Phosphate buffer solution. Dissolve 0.9 g of dipotassium – for impurity C, multiply the peak area by the correction
hydrogen phosphate R and 2.7 g of potassium dihydrogen factor 1.5 ;
phosphate R in 900 mL of water R and mix well. Adjust
to pH 6.0 with phosphoric acid R, dilute to 1000 mL with – for impurity D, multiply the peak area by the correction
water R and filter. factor 1.3 ;
Test solution (a). Dissolve 50.0 mg of the substance to be – for each impurity, use the concentration of lopinavir in
examined in the solvent mixture and dilute to 100.0 mL reference solution (b).
with the solvent mixture. Limits :
Test solution (b). Dilute 5.0 mL of test solution (a) to – impurities B, I : for each impurity, maximum 0.2 per
100.0 mL with the solvent mixture. cent ;
Reference solution (a). Dissolve 50.0 mg of lopinavir CRS in – impurities A, C, D, F, G : for each impurity, maximum
the solvent mixture and dilute to 100.0 mL with the solvent 0.15 per cent ;
mixture. Dilute 5.0 mL of the solution to 100.0 mL with the – unspecified impurities : for each impurity, maximum
solvent mixture. 0.10 per cent ;
Reference solution (b). Dilute 5.0 mL of test solution (b) to – reporting threshold : 0.05 per cent ; disregard any peak
250.0 mL with the solvent mixture. eluting after impurity N.

Lopinavir EUROPEAN PHARMACOPOEIA
B. Liquid chromatography (2.2.29) as described in test A for impurities for demonstration of compliance. See also 5.10.
related substances with the following modifications. Control of impurities in substances for pharmaceutical use):
Mobile phase : mobile phase A, mobile phase B (30:70 V/V). E, H, J, K, L, M, N, P, S.
Run time : 8.3 times the retention time of lopinavir.
supplied with lopinavir for system suitability CRS and
the chromatogram obtained with reference solution (c)
to identify the peaks due to impurities Q, R, S and T ;
use the chromatogram supplied with lopinavir for peak
identification CRS and the chromatogram obtained
with reference solution (d) to identify the peak due to
impurity O.
Relative retention r (not rG) with reference to lopinavir A. (2S)-N-[(1S,3S,4S)-1-benzyl-4-amino-3-hydroxy-5-
(retention time = about 6 min) : impurity N = about 1.4 ; phenylpentyl]-3-methyl-2-[2-oxotetrahydropyrimidin-
impurity O = about 1.5 ; impurity Q = about 4.4 ; 1(2H)-yl]butanamide,
impurity R = about 6.0 ; impurity S = about 7.1 ;
impurity T = about 8.5.
impurities S and T.
– for impurity O, multiply the peak area by the correction
factor 1.3 ;
– for impurity Q, multiply the peak area by the correction B. (2S)-N-[(1S,3S,4S)-1-benzyl-4-(formylamino)-3-hydroxy-
factor 0.7 ; 5-phenylpentyl]-3-methyl-2-[2-oxotetrahydropyrimidin-
– for each impurity, use the concentration of lopinavir in 1(2H)-yl]butanamide,
reference solution (b).
Limits :
– impurities O, Q, R, T : for each impurity, maximum
0.15 per cent ;
0.10 per cent ;
– reporting threshold : 0.05 per cent ; disregard any peak
eluting before and including impurity N ;
– total of all impurities eluting before and including C. (2R)-N-[(1S,2S,4S)-1-benzyl-2-hydroxy-4-[[(2S)-3-methyl-
impurity N in test A and after impurity N in test B : 2-[2-oxotetrahydropyrimidin-1(2H)-yl]butanoyl]amino]-
maximum 0.7 per cent. 5-phenylpentyl]-3-methyl-2-[2-oxotetrahydropyrimidin-
Water (2.5.12): maximum 4.4 per cent, determined on 0.250 g. 1(2H)-yl]butanamide,
1.0 g.
ASSAY
Liquid chromatography (2.2.29) as described in test A for
Mobile phase : mobile phase A.
Injection : test solution (b) and reference solution (a).
Run time : 1.6 times the retention time of lopinavir. D. (1R,3R)-1-[(1R)-1-[[2-(2,6-dimethylphenoxy)acetyl]-
Calculate the percentage content of C37H48N4O5 taking into amino]-2-phenylethyl]-3-[[(2R)-3-methyl-2-[2-oxo-
account the assigned content of lopinavir CRS. tetrahydropyrimidin-1(2H)-yl]butanoyl]amino]-4-
phenylbutyl hydrogen sulfate,
STORAGE
IMPURITIES
Specified impurities : A, B, C, D, F, G, I, O, Q, R, T.
by the general monograph Substances for pharmaceutical E. N-[(1S,2S,4S)-4-amino-1-benzyl-2-hydroxy-5-
use (2034). It is therefore not necessary to identify these phenylpentyl]-2-(2,6-dimethylphenoxy)acetamide,

EUROPEAN PHARMACOPOEIA Lopinavir
L. N,N′-(Z)-ethene-1,2-diylbis[2-(2,6-dimethylphenoxy)-
acetamide],
F. N-[(1S,2S,4S)-1-benzyl-4-(formylamino)-2-hydroxy-5-
phenylpentyl]-2-(2,6-dimethylphenoxy)acetamide,
M. (2S)-N-[(1R,3R,4S)-1-benzyl-4-[[2-(2,6-dimethyl-
G. N-[(1S,2S,4S)-(4-acetylamino)-1-benzyl-2-hydroxy-5- phenoxy)acetyl]amino]-3-hydroxy-5-phenylpentyl]-
phenylpentyl]-2-(2,6-dimethylphenoxy)acetamide, 3-methyl-2-[2-oxotetrahydropyrimidin-1(2H)-yl]-
butanamide,
H. N-[(1S)-1-[(4S,6S)-4-benzyl-2-oxo-1,3-oxazinan-6-yl]-2-
phenylethyl]-2-(2,6-dimethylphenoxy)acetamide,
N. (2S)-N-[(1S,3R,4S)-1-benzyl-4-[[2-(2,6-dimethylphenoxy)-
acetyl]amino]-3-hydroxy-5-phenylpentyl]-3-methyl-2-[2-
oxotetrahydropyrimidin-1(2H)-yl]butanamide,
I. (2S)-N-[(1S,2S,4S)-1-benzyl-4-[[2-(2,6-dimethylphenoxy)-
O. (1S,3S)-1-[(1S)-1-[[2-(2,6-dimethylphenoxy)acetyl]-
amino]-2-phenylethyl]-3-[[(2S)-3-methyl-2-[2-oxo-
tetrahydropyrimidin-1(2H)-yl]butanoyl]amino]-4-
phenylbutyl (2S)-3-methyl-2-[2-oxotetrahydropyrimidin-
J. (2S)-N-[(1S,3S,4S)-1-benzyl-4-[[2-(2,4-dimethylphenoxy)- 1(2H)-yl]butanoate,
K. (2R)-N-[(1S,3S,4S)-1-benzyl-4-[[2-(2,6-dimethylphenoxy)- P. (2S)-N-[(1R,3S,4S)-1-benzyl-4-[[2-(2,6-dimethylphenoxy)-
acetyl]amino]-3-hydroxy-5-phenylpentyl]-3-methyl-2-[2- acetyl]amino]-3-hydroxy-5-phenylpentyl]-3-methyl-2-[2-
oxotetrahydropyrimidin-1(2H)-yl]butanamide, oxotetrahydropyrimidin-1(2H)-yl]butanamide,

Lopinavir EUROPEAN PHARMACOPOEIA
Q. N-[(1S,2S,4S)-1-benzyl-4-[[2-(2,6-dimethylphenoxy)-
acetyl]amino]-2-hydroxy-5-phenylpentyl]-2-(2,6-
dimethylphenoxy)acetamide,
S. (1S,3S)-1-[(1S)-1-[[2-(2,6-dimethylphenoxy)acetyl]-
amino]-2-phenylethyl]-3-[[(2S)-3-methyl-2-[2-oxo-
tetrahydropyrimidin-1(2H)-yl]butanoyl]amino]-4-
phenylbutyl 2-(2,6-dimethylphenoxy)acetate,
R. (2S)-N-[(1S,3S,4S)-1-benzyl-4-[[2-(2,6-dimethylphenoxy)-
acetyl]amino]-3-hydroxy-5-phenylpentyl]-2-[3-[2-(2,6-
dimethylphenoxy)acetyl]-2-oxotetrahydropyrimidin- T. N,N′-bis[(1S,3S,4S)-1-benzyl-4-[[2-(2,6-dimethyl-
1(2H)-yl]-3-methylbutanamide, phenoxy)acetyl]amino]-3-hydroxy-5-phenylpentyl]urea.

EUROPEAN PHARMACOPOEIA Nevirapine
01/2017:2255 Injection : 50 µL of test solution (a) and reference solutions (a)

corrected 9.3 and (b).
Run time : 10 times the retention time of nevirapine.
with nevirapine for peak identification CRS and the
chromatogram obtained with reference solution (b) to identify
NEVIRAPINE the peaks due to impurities A, B and C.
Relative retention with reference to nevirapine (retention
Nevirapinum time = about 8 min) : impurity B = 0.7 ; impurity A = 1.5 ;
impurity C = 2.8.
– resolution : minimum 5 between the peaks due to impurity B
and nevirapine.
Limits :
– impurities A, B, C : for each impurity, not more than
twice the area of the principal peak in the chromatogram
C15H14N4O Mr 266.3 obtained with reference solution (a) (0.2 per cent) ;
[129618-40-2] – unspecified impurities : for each impurity, not more than the
DEFINITION with reference solution (a) (0.1 per cent) ;
11-Cyclopropyl-4-methyl-5,11-dihydro-6H-dipyrido – total : not more than 6 times the area of the principal peak
[3,2-b:2′,3′-e][1,4]diazepin-6-one. in the chromatogram obtained with reference solution (a)
Content : 97.5 per cent to 102.0 per cent (dried substance). (0.6 per cent);
CHARACTERS the chromatogram obtained with reference solution (a)
Appearance : white or almost white powder. (0.05 per cent).
Solubility : practically insoluble in water, sparingly soluble Loss on drying (2.2.32) : maximum 0.5 per cent, determined
or slightly soluble in methylene chloride, slightly soluble in on 1.000 g by drying in an oven at 105 °C.
methanol. Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
1.0 g.
IDENTIFICATION
A. Infrared absorption spectrophotometry (2.2.24). ASSAY
Comparison : anhydrous nevirapine CRS. Liquid chromatography (2.2.29) as described in the test for
B. Loss on drying (see Tests).
Injection : 25 µL of test solution (b) and reference solution (c).
TESTS Calculate the percentage content of C15H14N4O from the
Related substances. Liquid chromatography (2.2.29). declared content of anhydrous nevirapine CRS.
Test solution (a). Dissolve 24.0 mg of the substance to be IMPURITIES
examined in a mixture of 4 mL of acetonitrile R and 80 mL of Specified impurities : A, B, C.
the mobile phase and sonicate until dissolution is complete.
Dilute to 100.0 mL with the mobile phase.
Test solution (b). Dilute 3.0 mL of test solution (a) to 25.0 mL
with the mobile phase.
Reference solution (a). Dilute 1.0 mL of test solution (a) to
100.0 mL with the mobile phase. Dilute 5.0 mL of this solution
to 50.0 mL with the mobile phase.
Reference solution (b). Add 2.0 mL of the mobile phase to A. 11-ethyl-4-methyl-5,11-dihydro-6H-dipyrido-
a vial of nevirapine for peak identification CRS (containing [3,2-b:2′,3′-e][1,4]diazepin-6-one,
impurities A, B and C), mix and sonicate for 1 min.
Reference solution (c). Dissolve 24.0 mg of anhydrous
nevirapine CRS in a mixture of 4 mL of acetonitrile R and
80 mL of the mobile phase and sonicate until complete
dissolution. Dilute to 100.0 mL with the mobile phase. Dilute
3.0 mL of this solution to 25.0 mL with the mobile phase.
Column : B. 4-methyl-5,11-dihydro-6H-dipyrido[3,2-b:2′,3′-e]-
– size : l = 0.15 m, Ø = 4.6 mm ; [1,4]diazepin-6-one,
– stationary phase : end-capped amidohexadecylsilyl silica gel
for chromatography R (5 µm);
Mobile phase : mix 20 volumes of acetonitrile R and 80 volumes
of a 2.88 g/L solution of ammonium dihydrogen phosphate R,
previously adjusted to pH 5.0 using dilute sodium hydroxide
solution R.
Flow rate : 1.0 mL/min. C. 4-methyl-11-propyl-5,11-dihydro-6H-dipyrido[3,2-b:2′,3′-
Detection : spectrophotometer at 220 nm. e][1,4]diazepin-6-one.

EUROPEAN PHARMACOPOEIA Nevirapine hemihydrate
01/2017:2479 Flow rate : 0.7 mL/min.

corrected 10.0 Detection : spectrophotometer at 282 nm.
Injection : 2.0 µL of the test solution and reference solutions (a)
and (b).
with nevirapine for peak identification CRS and the
NEVIRAPINE HEMIHYDRATE chromatogram obtained with reference solution (b) to identify
the peaks due to impurities A, B and C.
Nevirapinum hemihydricum Relative retention with reference to nevirapine
(retention time = about 3 min): impurity B = about 0.9 ;
impurity A = about 1.2 ; impurity C = about 1.3.
impurity B and nevirapine and minimum 5.0 between
the peaks due to nevirapine and impurity A in the
chromatogram obtained with reference solution (b) ;
C15H14N4O,½H2O Mr 275.3 – symmetry factor : maximum 1.7 for the peak due to
[220988-26-1] nevirapine in the chromatogram obtained with reference
solution (a).
DEFINITION Calculation of percentage contents :
11-Cyclopropyl-4-methyl-5,11-dihydro-6H-dipyrido[3,2- – for each impurity, use the concentration of nevirapine
b:2′,3′-e][1,4]diazepin-6-one hemihydrate. hemihydrate in reference solution (a).
Content : 97.5 per cent to 102.0 per cent (anhydrous substance). Limits :
CHARACTERS – impurities A, B, C : for each impurity, maximum 0.2 per
Appearance : white or almost white powder. cent ;
Solubility : practically insoluble in water, slightly soluble in – unspecified impurities : for each impurity, maximum
methanol and in methylene chloride. 0.10 per cent ;
IDENTIFICATION – reporting threshold : 0.05 per cent.
A. Infrared absorption spectrophotometry (2.2.24). Water (2.5.12) : 3.1 per cent to 3.9 per cent, determined on
Comparison : nevirapine hemihydrate CRS. 0.300 g.
B. Water (see Tests). Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
TESTS 1.0 g.
Related substances. Liquid chromatography (2.2.29). ASSAY
Test solution. Dissolve 20.0 mg of the substance to be Liquid chromatography (2.2.29) as described in the test for
examined in methanol R and sonicate until dissolution is related substances with the following modification.
complete. Dilute to 50.0 mL with methanol R. Injection : 2.0 µL of the test solution and reference solution (c).
Reference solution (a). Dilute 1.0 mL of the test solution to Calculate the percentage content of C15H14N4O taking into
100.0 mL with methanol R. Dilute 1.0 mL of this solution to account the assigned content of anhydrous nevirapine CRS.
10.0 mL with methanol R.
Reference solution (b). Add 1 mL of methanol R to a vial IMPURITIES
of nevirapine for peak identification CRS (containing Specified impurities : A, B, C.
impurities A, B and C), mix and sonicate for 1 min. Other detectable impurities (the following substances would,
Reference solution (c). Dissolve 20.0 mg of anhydrous if present at a sufficient level, be detected by one or other of
nevirapine CRS in methanol R and sonicate until dissolution is the tests in the monograph. They are limited by the general
complete. Dilute to 50.0 mL with methanol R. acceptance criterion for other/unspecified impurities and/or
Column :
– size : l = 50 mm, Ø = 2.1 mm ; impurities for demonstration of compliance. See also 5.10.
– stationary phase : end-capped octadecylsilyl silica gel for Control of impurities in substances for pharmaceutical use) : D.
chromatography R (1.8 µm) ;
Mobile phase :
– mobile phase A : dissolve 0.77 g of ammonium acetate R in
900 mL of water for chromatography R, adjust to pH 5.6
with acetic acid R and dilute to 1000 mL with water for
chromatography R ;
A. 11-ethyl-4-methyl-5,11-dihydro-6H-dipyrido[3,2-b:2′,3′-
– mobile phase B : acetonitrile R ; e][1,4]diazepin-6-one,
0 – 1.35 90 10
1.35 – 3.85 90 → 67 10 → 33
3.85 – 6.70 67 → 60 33 → 40
6.70 – 7.65 60 40 B. 4-methyl-5,11-dihydro-6H-dipyrido[3,2-b:2′,3′-

e][1,4]diazepin-6-one,

Nevirapine hemihydrate EUROPEAN PHARMACOPOEIA
D. 11,11′-dicyclopropyl-4,4′-dimethyl-5,5′,11,11′-tetrahydro-
C. 4-methyl-11-propyl-5,11-dihydro-6H-dipyrido[3,2-b:2′,3′- 6H,6′H-9,9′-bidipyrido[3,2-b:2′,3′-e][1,4]diazepine-6,6′-
e][1,4]diazepin-6-one, dione.

EUROPEAN PHARMACOPOEIA Oseltamivir phosphate
04/2011:2422 Flow rate : 1.5 mL/min.

corrected 10.0 Post-column split ratio : use a split ratio suitable for the mass
detector (e.g. 1:3).
Detection :
– mass detector : the following settings have been found to
be suitable and are given as examples ; if the detector has
OSELTAMIVIR PHOSPHATE different setting parameters, adjust the detector settings so
as to comply with the system suitability criterion :
Oseltamiviri phosphas – ionisation : ESI-positive ;
– detection m/z : 356.2 ;
– dwell : 580 ms ;
– gain EMV : 1 ;
– fragmentator voltage : 120 V ;
– gas temperature : 350 °C ;
– drying gas flow : 13 L/min,
C16H31N2O8P Mr 410.4 – nebuliser pressure : 345 kPa ;
[204255-11-8] – capillary voltage (Vcap) : 3 kV.
Injection : 1 µL of the test solution and reference solution (b).
DEFINITION
Run time : 3 min.
Ethyl (3R,4R,5S)-4-acetamido-5-amino-3-(1-ethylpropoxy)-
cyclohex-1-ene-1-carboxylate phosphate. System suitability : reference solution (b) :
Content : 98.0 per cent to 102.0 per cent (anhydrous substance). – repeatability : maximum relative standard deviation of
15 per cent determined on 6 injections.
Limit :
CHARACTERS – impurity B : not more than the area of the corresponding
Appearance : white or almost white powder. peak in the chromatogram obtained with reference
Solubility : freely soluble in water and in methanol, practically solution (b) (100 ppm).
insoluble in methylene chloride. Impurity H. Gas chromatography (2.2.28).
It shows polymorphism (5.9). Silylation reagent. Mix 1.0 mL of chlorotrimethylsilane R,
IDENTIFICATION 2.0 mL of hexamethyldisilazane R and 10.0 mL of anhydrous
pyridine R.
A. Specific optical rotation (see Tests).
Test solution. Introduce 15.0 mg of the substance to be
B. Infrared absorption spectrophotometry (2.2.24). examined into a 2 mL vial and add 1.0 mL of the silylation
Comparison : oseltamivir phosphate (impurity B-free) CRS. reagent. Close the vial, shake and heat at 60 °C for 20 min.
If the spectra obtained show differences, dissolve the Centrifuge and discard the precipitate.
substance to be examined and the reference substance Reference solution. Introduce 15.0 mg of oseltamivir
separately in methanol R, evaporate to dryness and record impurity H CRS into a 2 mL vial and add 1.0 mL of anhydrous
new spectra using the residues. pyridine R. Close the vial and shake (solution A). (Note :
C. Dissolve 200 mg in 10 mL of water R. It gives reaction (b) impurity H is hygroscopic.) Introduce 15.0 mg of the
of phosphates (2.3.1). substance to be examined into another 2 mL vial and add
1.0 mL of the silylation reagent. Close the vial, shake and heat
TESTS at 60 °C for 20 min. Centrifuge and discard the precipitate
Specific optical rotation (2.2.7): − 30.7 to − 32.6 (anhydrous (solution B). Introduce 10.0 µL of solution A and 10.0 µL of
substance), measured at 25 °C. solution B into a volumetric flask and dilute to 10.0 mL with
Dissolve 0.50 g in water R and dilute to 50.0 mL with the same anhydrous pyridine R.
solvent. Column :
Impurity B. Liquid chromatography (2.2.29) coupled with – material : fused silica ;
mass spectrometry (2.2.43). – size : l = 30 m, Ø = 0.32 mm ;
Test solution. Dissolve 0.100 g of the substance to be examined – stationary phase : methylpolysiloxane R (film thickness
in water for chromatography R and dilute to 10.0 mL with the 0.25 μm).
same solvent. Carrier gas : helium for chromatography R.
Reference solution (a). Dissolve 2.5 mg of oseltamivir Flow rate : 1.2 mL/min.
impurity B CRS in 5.0 mL of anhydrous ethanol R and dilute
to 50.0 mL with water for chromatography R. Dilute 2.0 mL of Split ratio : 1:50.
the solution to 100.0 mL with water for chromatography R. Temperature :
Reference solution (b). Dissolve 50.0 mg of oseltamivir Time Temperature
phosphate (impurity B-free) CRS in reference solution (a) and (min) (°C)
dilute to 5.0 mL with the same solution. Column 0-2 180
Column : 2 - 11 180 → 250
– size : l = 0.05 m, Ø = 3.0 mm ; 11 - 21 250
Injection port 260
chromatography R (5 μm) ; Detector 260
Mobile phase : mix 10 volumes of a 1.54 g/L solution of Detection : flame ionisation.
ammonium acetate R in water for chromatography R, Injection : 1 µL.
30 volumes of acetonitrile R1 and 60 volumes of water for Relative retention with reference to oseltamivir phosphate
chromatography R. (retention time = about 10 min) : impurity H = about 0.5.

Oseltamivir phosphate EUROPEAN PHARMACOPOEIA
System suitability : reference solution : STORAGE

– repeatability : maximum relative standard deviation of 5 per Protected from light.
cent for the peak due to impurity H after 6 injections.
Limit : IMPURITIES
– impurity H : not more than 1.5 times the area of the Specified impurities : B, C, H.
corresponding peak in the chromatogram obtained with Other detectable impurities (the following substances would,
the reference solution (0.15 per cent). if present at a sufficient level, be detected by one or other of
Related substances. Liquid chromatography (2.2.29). the tests in the monograph. They are limited by the general
Solvent mixture : acetonitrile R1, methanol R2, water for acceptance criterion for other/unspecified impurities and/or
chromatography R (135:245:620 V/V/V). by the general monograph Substances for pharmaceutical use
Test solution. Dissolve 50.0 mg of the substance to be for demonstration of compliance. See also 5.10. Control of
examined in the solvent mixture and dilute to 50.0 mL with impurities in substances for pharmaceutical use) : A, D, E, F, G.
the solvent mixture.
Reference solution (a). Dilute 1.0 mL of the test solution to
solution to 10.0 mL with the solvent mixture.
Reference solution (b). Dissolve 5 mg of oseltamivir
impurity A CRS and 5.0 mg of oseltamivir impurity C CRS in
mixture. Dilute 1.0 mL of the solution to 10.0 mL with the
solvent mixture.
Reference solution (c). Dissolve 50.0 mg of oseltamivir A. (3R,4R,5S)-5-acetamido-4-amino-3-(1-ethylpropoxy)-
phosphate (impurity B-free) CRS in the solvent mixture and cyclohex-1-ene-1-carboxylic acid,
dilute to 50.0 mL with the solvent mixture.
Column :
– size : l = 0.25 m, Ø = 4.6 mm ;
– stationary phase : end-capped octylsilyl silica gel for
chromatography R (5 μm) ;
Mobile phase : mix 135 volumes of acetonitrile R1,
245 volumes of methanol R2 and 620 volumes of a 6.8 g/L B. ethyl (1R,2R,3S,4R,5S)-4-acetamido-5-amino-2-azido-3-
solution of potassium dihydrogen phosphate R in water for (1-ethylpropoxy)cyclohexanecarboxylate,
chromatography R, adjusted to pH 6.0 with a 1 M potassium
hydroxide solution prepared from potassium hydroxide R.
Injection : 15 µL of the test solution and reference solutions (a)
and (b).
Run time : twice the retention time of oseltamivir phosphate.
Relative retention with reference to oseltamivir phosphate C. (3R,4R,5S)-4-acetamido-5-amino-3-(1-ethylpropoxy)-
(retention time = about 17 min) : impurity A = about 0.16 ; cyclohex-1-ene-1-carboxylic acid,
impurity C = about 0.17.
impurities A and C.
Limits :
– impurity C : not more than 0.3 times the area of the
corresponding peak in the chromatogram obtained with D. ethyl 4-acetamido-3-hydroxybenzoate,
reference solution (b) (0.3 per cent) ;
– unspecified impurities : for each impurity, not more than the
with reference solution (a) (0.10 per cent) ;
(0.7 per cent) ;
– disregard limit : 0.5 times the area of the principal peak in E. methyl (3R,4R,5S)-4-acetamido-5-amino-3-(1-ethyl-
the chromatogram obtained with reference solution (a) propoxy)cyclohex-1-ene-1-carboxylate,
(0.05 per cent).
Water (2.5.12) : maximum 0.5 per cent, determined on 0.500 g.
ASSAY
Injection : test solution and reference solution (c).
Calculate the percentage content of C16H31N2O8P
from the declared content of oseltamivir phosphate F. ethyl (3R,4R,5S)-4-acetamido-5-amino-3-(1-methyl-
(impurity B-free) CRS. propoxy)cyclohex-1-ene-1-carboxylate,

EUROPEAN PHARMACOPOEIA Oseltamivir phosphate
H. tributylphosphane oxide.
G. ethyl (3R,4R,5S)-5-acetamido-4-amino-3-(1-ethyl-
propoxy)cyclohex-1-ene-1-carboxylate,

EUROPEAN PHARMACOPOEIA Raltegravir chewable tablets
07/2018:2939 Column :
– size : l = 0.25 m, Ø = 4.6 mm ;
RALTEGRAVIR CHEWABLE TABLETS
Mobile phase :
– mobile phase A : mix 20 volumes of acetonitrile for
Raltegraviri compressi masticabiles chromatography R and 80 volumes of a 1.36 g/L solution of
DEFINITION potassium dihydrogen phosphate R previously adjusted to
pH 3.0 with phosphoric acid R ;
Raltegravir chewable tablets contain Raltegravir potassium
(2887). – mobile phase B : acetonitrile for chromatography R ;
The tablets comply with the monograph Tablets (0478) and Time Mobile phase A Mobile phase B
with the following additional requirements. (min) (per cent V/V) (per cent V/V)
0-2 100 0
Content : 95.0 per cent to 105.0 per cent of the content of
raltegravir (C20H21FN6O5) stated on the label. 2 - 27 100 → 50 0 → 50
IDENTIFICATION Flow rate : 1.0 mL/min.

Carry out either tests A, B or tests B, C. Detection : spectrophotometer at 220 nm.
A. Record the UV spectrum of the principal peak in the Injection : 15 µL of the test solution and reference solutions (b),
chromatograms obtained with the solutions used in (c) and (d).
the assay with a diode array detector in the range of Identification of impurities: use the chromatogram obtained
190-400 nm. with reference solution (d) to identify the peaks due to
Results : the UV spectrum of the principal peak in impurities C and D ; use the chromatogram obtained with
the chromatogram obtained with the test solution is reference solution (c) to identify the peak due to impurity E.
similar to the UV spectrum of the principal peak in the Relative retention with reference to raltegravir (retention
chromatogram obtained with reference solution (a). time = about 22 min) : impurity D = about 0.7 ;
B. Examine the chromatograms obtained in the assay. impurity C = about 0.8 ; impurity E = about 0.96.
Results : the principal peak in the chromatogram obtained System suitability : reference solution (c) :
with the test solution is similar in retention time and size – resolution : minimum 1.5 between the peaks due to
to the principal peak in the chromatogram obtained with impurity E and raltegravir.
reference solution (a).
C. Infrared absorption spectrophotometry (2.2.24).
– correction factors : multiply the peak areas of the following
Preparation : crush a tablet to a powder and homogenise. impurities by the corresponding correction factor :
Comparison : raltegravir potassium CRS. impurity C = 1.6 ; impurity D = 1.4 ;
Results : the spectrum obtained shows absorption maxima – for each impurity, use the concentration of raltegravir in
at about 1633 cm−1, 1515 cm−1, 1188 cm−1, 810 cm−1 and reference solution (b).
728 cm−1, similar to the spectrum obtained with raltegravir Limits :
potassium CRS.
– impurity C : maximum 0.3 per cent ;
Other absorption maxima may be present in the spectra.
– impurity D : maximum 0.2 per cent ;
TESTS – unspecified impurities : for each impurity, maximum 0.2 per
Related substances. Liquid chromatography (2.2.29). cent ;
Solvent mixture : acetonitrile R, water R (30:70 V/V). – total : maximum 0.8 per cent ;
Test solution. Place 10 tablets in an appropriate volumetric – reporting threshold : 0.1 per cent.
flask to obtain a concentration of 1 mg/mL of raltegravir and Dissolution (2.9.3, Apparatus 2). The tablets comply with the
make up to volume with the solvent mixture. Stir vigorously test, unless otherwise justified and authorised.
for 1 h. Centrifuge a portion of the solution and dilute 20.0 mL
Dissolution medium : water R, 900 mL.
of the clear supernatant to 200.0 mL with the solvent mixture.
Rotation speed : 50 r/min.
Reference solution (a). Dissolve 22.0 mg of raltegravir
potassium CRS in the solvent mixture and dilute to 200.0 mL Time : 15 min.
with the solvent mixture. Analysis. Liquid chromatography (2.2.29).
Reference solution (b). Dilute 1.0 mL of the test solution to Test solutions. Solutions from the dissolution test.
100.0 mL with the solvent mixture. Dilute 2.0 mL of this Reference solution. Using sonication if necessary, dissolve a
solution to 10.0 mL with the solvent mixture. suitable quantity of raltegravir potassium CRS in a suitable
Reference solution (c). Dissolve 2 mg of raltegravir quantity of a mixture of 30 volumes of acetonitrile R
impurity E CRS in the solvent mixture and dilute to 100.0 mL and 70 volumes of water R to obtain a concentration of
with the solvent mixture. Dilute 1.0 mL of the solution to raltegravir corresponding to the theoretical concentration
100.0 mL with the test solution. of raltegravir in the test solution, based on the labelled
Reference solution (d). In order to prepare impurities C and D content of the tablets.
in situ, dissolve 20 mg of raltegravir potassium R in a 40 g/L Column :
solution of sodium hydroxide R and dilute to 10 mL with the – size : l = 0.1 m, Ø = 4.6 mm ;
same solvent. Stir the solution for 2 h at room temperature.
To 5 mL of the solution add 5 mL of a 103 g/L solution of – stationary phase : end-capped monolithic octadecylsilyl
hydrochloric acid R and dilute to 50 mL with the solvent silica gel for chromatography R ;
mixture. – temperature : 40 °C.

Raltegravir chewable tablets EUROPEAN PHARMACOPOEIA
Mobile phase : mix 38 volumes of acetonitrile R and

62 volumes of a 1.36 g/L solution of potassium dihydrogen
phosphate R previously adjusted to pH 3.0 with phosphoric
acid R.
Injection : 30 µL. B. 2-[2-[(E)-[(dimethylamino)methylidene]amino]propan-
Run time : 1 min. 2-yl]-N-[(4-fluorophenyl)methyl]-5-hydroxy-1-methyl-6-
oxo-1,6-dihydropyrimidine-4-carboxamide,
– repeatability : maximum relative standard deviation of
1.5 per cent determined on 6 injections.
Calculate the amount of dissolved raltegravir, expressed as a
percentage of the content of raltegravir (C20H21FN6O5) stated
on the label, taking into account the assigned content of
raltegravir potassium CRS and a conversion factor of 0.9210.
Acceptance criterion :
– Q = 85 per cent after 15 min.
C. 2-[2-[2-(2-acetylhydrazin-1-yl)-2-oxoacetamido]propan-
ASSAY 2-yl]-N-[(4-fluorophenyl)methyl]-5-hydroxy-1-methyl-6-
Liquid chromatography (2.2.29) as described in the test for oxo-1,6-dihydropyrimidine-4-carboxamide,
Injection : test solution and reference solution (a).
Calculate the percentage content of raltegravir (C20H21FN6O5)
taking into account the assigned content of raltegravir
potassium CRS and a conversion factor of 0.9210.
IMPURITIES D. N-[2-[4-[[(4-fluorophenyl)methyl]carbamoyl]-5-hydroxy-
Specified impurities : C, D. 1-methyl-6-oxo-1,6-dihydropyrimidin-2-yl]propan-2-
yl]oxamic acid,
Other detectable impurities (the following substances would, if
present at a sufficient level, be detected by one or other of the
tests in the monograph) : A, B, E.
A. 2-(2-aminopropan-2-yl)-N-[(4-fluorophenyl)methyl]- E. N-benzyl-5-hydroxy-1-methyl-2-[2-[(5-methyl-1,3,4-
5-hydroxy-1-methyl-6-oxo-1,6-dihydropyrimidine-4- oxadiazol-2-yl)formamido]propan-2-yl]-6-oxo-1,6-
carboxamide, dihydropyrimidine-4-carboxamide.

EUROPEAN PHARMACOPOEIA Raltegravir potassium
04/2018:2887 Column :
corrected 10.0 – size : l = 0.15 m, Ø = 4.6 mm ;
– stationary phase : phenylsilyl silica gel for chromatography R
(3.5 μm) ;
RALTEGRAVIR POTASSIUM Mobile phase :
– mobile phase A : 0.1 per cent V/V solution of phosphoric
acid R ;
Raltegravirum kalicum
– mobile phase B : acetonitrile for chromatography R ;
0-2 75 25
2-5 75 → 60 25 → 40
5 - 10 60 → 55 40 → 45
C20H20FKN6O5 Mr 482.5
10 - 19 55 → 10 45 → 90
[871038-72-1]
19 - 22 10 → 5 90 → 95
DEFINITION
Potassium 4-[[(4-fluorophenyl)methyl]carbamoyl]-1-methyl- Flow rate : 1.0 mL/min.
2-[2-[(5-methyl-1,3,4-oxadiazol-2-yl)formamido]propan-2- Detection : spectrophotometer at 220 nm.
yl]-6-oxo-1,6-dihydropyrimidin-5-olate. Injection : 10 µL of the test solution and reference solutions (b),
Content : 98.0 per cent to 102.0 per cent (anhydrous substance). (c), (d) and (e).
Identification of impurities: use the chromatogram obtained
CHARACTERS
with reference solution (d) to identify the peak due to
Appearance : white or almost white powder. impurity C ; use the chromatogram obtained with reference
Solubility : soluble in water, very slightly soluble in ethanol solution (c) to identify the peak due to impurity E ; use
(96 per cent), practically insoluble in heptane. the chromatogram supplied with raltegravir for peak
It shows polymorphism (5.9). identification CRS and the chromatogram obtained with
reference solution (e) to identify the peaks due to impurities F
IDENTIFICATION and G.
A. Infrared absorption spectrophotometry (2.2.24). Relative retention with reference to raltegravir (retention
Comparison : raltegravir potassium CRS. time = about 10 min) : impurity C = about 0.7 ;
impurity E = about 0.95 ; impurity G = about 1.1 ;
If the spectra obtained in the solid state show differences, impurity F = about 1.15.
dissolve the substance to be examined and the reference
substance separately in methanol R, evaporate to dryness System suitability : reference solution (c) :
and record new spectra using the residues. – resolution : minimum 1.5 between the peaks due to
B. It gives reaction (b) of potassium (2.3.1). impurity E and raltegravir.
TESTS – correction factor : multiply the peak area of impurity C by
Related substances. Liquid chromatography (2.2.29). 1.6 ;
Solvent mixture : acetonitrile R, water R (25:75 V/V). – for each impurity, use the concentration of raltegravir
Test solution. Dissolve 25.0 mg of the substance to be potassium in reference solution (b).
examined in 100 mL of the solvent mixture using sonication Limits :
for 5 min. Add about 140 mL of the solvent mixture then – impurity C : maximum 0.3 per cent ;
dilute to 250.0 mL with the solvent mixture. – impurities E, F, G : for each impurity, maximum 0.15 per
Reference solution (a). Dissolve 25.0 mg of raltegravir cent ;
potassium CRS in 100 mL of the solvent mixture using – unspecified impurities : for each impurity, maximum
sonication for 5 min. Add about 140 mL of the solvent mixture 0.10 per cent ;
then dilute to 250.0 mL with the solvent mixture.
100.0 mL with the solvent mixture. Dilute 1.0 mL of this – reporting threshold : 0.05 per cent.
solution to 10.0 mL with the solvent mixture. Water (2.5.12) : maximum 0.6 per cent, determined on 0.500 g.
Reference solution (c). Dissolve 2 mg of raltegravir Use as the solvent a mixture of equal volumes of methanol R
impurity E CRS in the solvent mixture and dilute to 20.0 mL and formamide R.
with the solvent mixture. Dilute 1.0 mL of the solution to ASSAY
50.0 mL with reference solution (a).
Reference solution (d). In order to prepare impurity C in situ, Liquid chromatography (2.2.29) as described in the test for
dissolve 20 mg of the substance to be examined in a 40 g/L related substances with the following modification.
solution of sodium hydroxide R and dilute to 10 mL with the Injection : test solution and reference solution (a).
same solvent. Stir the solution for 30 min. To 5 mL of the Calculate the percentage content of C20H20FKN6O5 taking into
solution add 5 mL of a 103 g/L solution of hydrochloric acid R account the assigned content of raltegravir potassium CRS.
and dilute to 50 mL with the solvent mixture.
IMPURITIES
Reference solution (e). Dissolve 5 mg of raltegravir for peak
identification CRS (containing impurities F and G) in 20 mL Specified impurities : C, E, F, G.
of the solvent mixture using sonication for 5 min. Add about Other detectable impurities (the following substances would,
25 mL of the solvent mixture then dilute to 50 mL with the if present at a sufficient level, be detected by one or other of
solvent mixture. the tests in the monograph. They are limited by the general

Raltegravir potassium EUROPEAN PHARMACOPOEIA

impurities in substances for pharmaceutical use) : A, B, D, H.
E. N-benzyl-5-hydroxy-1-methyl-2-[2-[(5-methyl-1,3,4-
oxadiazol-2-yl)formamido]propan-2-yl]-6-oxo-1,6-
dihydropyrimidine-4-carboxamide,
A. 2-(2-aminopropan-2-yl)-N-[(4-fluorophenyl)methyl]-
5-hydroxy-1-methyl-6-oxo-1,6-dihydropyrimidine-4-
carboxamide,
F. ethyl (1E)-N-[[2-[4-[[(4-fluorophenyl)methyl]carbamoyl]-
5-hydroxy-1-methyl-6-oxo-1,6-dihydropyrimidin-2-
yl]propan-2-yl]oxamoyl]ethanehydrazonate,
B. 2-[2-[(E)-[(dimethylamino)methylidene]amino]propan-
2-yl]-N-[(4-fluorophenyl)methyl]-5-hydroxy-1-methyl-6-
G. ethyl (1Z)-N-[[2-[4-[[(4-fluorophenyl)methyl]carbamoyl]-
5-hydroxy-1-methyl-6-oxo-1,6-dihydropyrimidin-2-
yl]propan-2-yl]oxamoyl]ethanehydrazonate,
C. 2-[2-[2-(2-acetylhydrazin-1-yl)-2-oxoacetamido]propan-
D. N-[2-[4-[[(4-fluorophenyl)methyl]carbamoyl]-5-hydroxy- H. N,N′-bis[2-[4-[[(4-fluorophenyl)methyl]carbamoyl]-
1-methyl-6-oxo-1,6-dihydropyrimidin-2-yl]propan-2- 5-hydroxy-1-methyl-6-oxo-1,6-dihydropyrimidin-2-
yl]oxamic acid, yl]propan-2-yl]oxamide.

EUROPEAN PHARMACOPOEIA Raltegravir tablets
07/2018:2938 – stationary phase : end-capped octadecylsilyl silica gel for

Mobile phase :
– mobile phase A : mix 20 volumes of acetonitrile for
RALTEGRAVIR TABLETS chromatography R and 80 volumes of a 1.36 g/L solution of
potassium dihydrogen phosphate R previously adjusted to
Raltegraviri compressi pH 3.0 with phosphoric acid R ;
DEFINITION – mobile phase B : acetonitrile for chromatography R ;
Raltegravir tablets contain Raltegravir potassium (2887). Time Mobile phase A Mobile phase B
The tablets comply with the monograph Tablets (0478) and
0-2 100 0
with the following additional requirements.
Content : 95.0 per cent to 105.0 per cent of the content of 2 - 27 100 → 50 0 → 50
raltegravir (C20H21FN6O5) stated on the label.
IDENTIFICATION Detection : spectrophotometer at 220 nm.
Carry out either tests A, B or tests B, C. Injection : 15 µL of the test solution and reference solutions (b),
A. Record the UV spectrum of the principal peak in the (c) and (d).
chromatograms obtained with the solutions used in Identification of impurities: use the chromatogram obtained
the assay with a diode array detector in the range of with reference solution (d) to identify the peaks due to
190-400 nm. impurities C and D ; use the chromatogram obtained with
Results : the UV spectrum of the principal peak in reference solution (c) to identify the peak due to impurity E.
the chromatogram obtained with the test solution is Relative retention with reference to raltegravir (retention
similar to the UV spectrum of the principal peak in the time = about 22 min) : impurity D = about 0.7 ;
chromatogram obtained with reference solution (a). impurity C = about 0.8 ; impurity E = about 0.96.
B. Examine the chromatograms obtained in the assay. System suitability : reference solution (c) :
Results : the principal peak in the chromatogram obtained – resolution : minimum 1.5 between the peaks due to
with the test solution is similar in retention time and size impurity E and raltegravir.
to the principal peak in the chromatogram obtained with Calculation of percentage contents :
reference solution (a).
– correction factors : multiply the peak areas of the following
C. Infrared absorption spectrophotometry (2.2.24). impurities by the corresponding correction factor :
Preparation : crush a tablet to a powder and homogenise. impurity C = 1.6 ; impurity D = 1.4 ;
Comparison : raltegravir potassium CRS. – for each impurity, use the concentration of raltegravir in
Results : the spectrum obtained shows absorption maxima reference solution (b).
at about 1633 cm−1, 1515 cm−1, 1188 cm−1, 810 cm−1 and Limits :
728 cm−1, similar to the spectrum obtained with raltegravir – impurity C : maximum 0.5 per cent ;
potassium CRS.
– impurity D : maximum 0.3 per cent ;
Other absorption maxima may be present in the spectra.
– unspecified impurities : for each impurity, maximum 0.2 per
TESTS cent ;
Related substances. Liquid chromatography (2.2.29). – total : maximum 0.8 per cent ;
Solvent mixture : acetonitrile R, water R (30:70 V/V). – reporting threshold : 0.1 per cent.
Test solution. Place 10 tablets in an appropriate volumetric Dissolution (2.9.3, Apparatus 2). The tablets comply with
flask to obtain a concentration of 8 mg/mL of raltegravir and the test, unless otherwise justified and authorised. Use sinker
make up to volume with the solvent mixture. Stir vigorously devices.
for 1 h. Centrifuge a portion of the solution and dilute 5.0 mL Dissolution medium : water R, 900 mL.
of the clear supernatant to 100.0 mL with the solvent mixture. Rotation speed : 100 r/min.
Dilute 50.0 mL of this solution to 200.0 mL with the solvent Time : 15 min and 60 min.
mixture.
Analysis. Liquid chromatography (2.2.29).
Reference solution (a). Dissolve 22.0 mg of raltegravir
potassium CRS in the solvent mixture and dilute to 200.0 mL Test solutions. Solutions from the dissolution test.
with the solvent mixture. Reference solution. Using sonication, dissolve a suitable
Reference solution (b). Dilute 1.0 mL of the test solution to quantity of raltegravir potassium CRS in a suitable
100.0 mL with the solvent mixture. Dilute 2.0 mL of this quantity of a mixture of 30 volumes of acetonitrile R
solution to 10.0 mL with the solvent mixture. and 70 volumes of water R to obtain a concentration of
raltegravir corresponding to the theoretical concentration
Reference solution (c). Dissolve 2 mg of raltegravir of raltegravir in the test solution, based on the labelled
impurity E CRS in the solvent mixture and dilute to 100.0 mL content of the tablets.
with the solvent mixture. Dilute 1.0 mL of the solution to
100.0 mL with the test solution. Column :
Reference solution (d). In order to prepare impurities C and D – size : l = 0.1 m, Ø = 4.6 mm ;
in situ, dissolve 20 mg of raltegravir potassium R in a 40 g/L – stationary phase : end-capped monolithic octadecylsilyl
solution of sodium hydroxide R and dilute to 10 mL with the silica gel for chromatography R ;
same solvent. Stir the solution for 2 h at room temperature. – temperature : 40 °C.
To 5 mL of the solution add 5 mL of a 103 g/L solution of Mobile phase : mix 38 volumes of acetonitrile R and
hydrochloric acid R and dilute to 50 mL with the solvent 62 volumes of a 1.36 g/L solution of potassium dihydrogen
mixture. phosphate R previously adjusted to pH 3.0 with phosphoric
Column : acid R.
– size : l = 0.25 m, Ø = 4.6 mm ; Flow rate : 5.0 mL/min.

Raltegravir tablets EUROPEAN PHARMACOPOEIA

Injection : 10 µL.
Run time : 1 min.
B. 2-[2-[(E)-[(dimethylamino)methylidene]amino]propan-
Calculate the amount of dissolved raltegravir, expressed as a 2-yl]-N-[(4-fluorophenyl)methyl]-5-hydroxy-1-methyl-6-
percentage of the content of raltegravir (C20H21FN6O5) stated oxo-1,6-dihydropyrimidine-4-carboxamide,
on the label, taking into account the assigned content of
raltegravir potassium CRS and a conversion factor of 0.9210.
Acceptance criteria :
– 15-45 per cent after 15 min ;
– Q = 70 per cent after 60 min.
ASSAY
related substances with the following modifications. C. 2-[2-[2-(2-acetylhydrazin-1-yl)-2-oxoacetamido]propan-
Injection : test solution and reference solution (a). oxo-1,6-dihydropyrimidine-4-carboxamide,
Calculate the percentage content of raltegravir (C20H21FN6O5)
taking into account the assigned content of raltegravir
potassium CRS and a conversion factor of 0.9210.
IMPURITIES
Specified impurities : C, D. D. N-[2-[4-[[(4-fluorophenyl)methyl]carbamoyl]-5-hydroxy-
Other detectable impurities (the following substances would, if 1-methyl-6-oxo-1,6-dihydropyrimidin-2-yl]propan-2-
present at a sufficient level, be detected by one or other of the yl]oxamic acid,
tests in the monograph) : A, B, E.
A. 2-(2-aminopropan-2-yl)-N-[(4-fluorophenyl)methyl]- E. N-benzyl-5-hydroxy-1-methyl-2-[2-[(5-methyl-1,3,4-
5-hydroxy-1-methyl-6-oxo-1,6-dihydropyrimidine-4- oxadiazol-2-yl)formamido]propan-2-yl]-6-oxo-1,6-
carboxamide, dihydropyrimidine-4-carboxamide.

EUROPEAN PHARMACOPOEIA Ribavirin
01/2017:2109 – stationary phase : spherical end-capped octadecylsilyl silica

gel for chromatography R (3 µm) suitable for use with highly
aqueous mobile phases ;
Mobile phase :
RIBAVIRIN – mobile phase A : dissolve 1.0 g of anhydrous sodium sulfate R
in 950 mL of water for chromatography R, add 2.0 mL of
Ribavirinum a 5 per cent V/V solution of phosphoric acid R, adjust to
pH 2.8 with a 5 per cent V/V solution of phosphoric acid R
and dilute to 1000 mL with water for chromatography R ;
– mobile phase B : acetonitrile R1, mobile phase A (5:95 V/V) ;
0 - 15 100 0
15 - 25 100 → 0 0 → 100
25 - 35 0 100
C8H12N4O5 Mr 244.2
[36791-04-5] Flow rate : 1.0 mL/min.
DEFINITION Injection : 5 µL of the test solution and reference solutions (a)
1-β-D-Ribofuranosyl-1H-1,2,4-triazole-3-carboxamide. and (b).
Content : 98.0 per cent to 102.0 per cent (dried substance). Relative retention with reference to ribavirin (retention
time = about 6 min) : impurity A = about 0.8.
CHARACTERS System suitability : reference solution (a) :
Appearance : white or almost white, crystalline powder. – resolution : minimum 4.0 between the peaks due to
Solubility : freely soluble in water, slightly soluble in ethanol impurity A and ribavirin.
(96 per cent), slightly soluble or very slightly soluble in Limits :
methylene chloride. – correction factor : for the calculation of content, multiply
It shows polymorphism (5.9). the peak area of impurity A by 2.3 ;
– impurity A : not more than twice the area of the principal
IDENTIFICATION peak in the chromatogram obtained with reference
Infrared absorption spectrophotometry (2.2.24). solution (b) (0.2 per cent);
Comparison : ribavirin CRS. – unspecified impurities : for each impurity, not more than the
If the spectra obtained in the solid state show differences, area of the principal peak in the chromatogram obtained
dissolve the substance to be examined and the reference with reference solution (b) (0.10 per cent) ;
substance separately in methylene chloride R, evaporate to – total : not more than 3 times the area of the principal peak
dryness and record new spectra using the residues. in the chromatogram obtained with reference solution (b)
(0.3 per cent);
TESTS – disregard limit : 0.5 times the area of the principal peak in
pH (2.2.3) : 4.0 to 6.5. the chromatogram obtained with reference solution (b)
(0.05 per cent).
Dissolve 0.200 g in carbon dioxide-free water R and dilute to
10.0 mL with the same solvent. Loss on drying (2.2.32) : maximum 0.5 per cent, determined
on 1.000 g by drying in an oven at 105 °C for 5 h.
Specific optical rotation (2.2.7) : − 33 to − 37 (dried
substance). Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
1.0 g.
Dissolve 0.250 g in water R and dilute to 25.0 mL with the
same solvent. Determine the specific optical rotation within ASSAY
10 min of preparing the solution.
Related substances. Liquid chromatography (2.2.29). related substances with the following modification.
Test solution. Dissolve 50.0 mg of the substance to be Injection : test solution and reference solution (c).
examined in water for chromatography R and dilute to Calculate the percentage content of C8H12N4O5 from the
100.0 mL with the same solvent. declared content of ribavirin CRS.
Reference solution (a). In order to produce impurity A in
situ, mix 5.0 mL of the test solution and 5.0 mL of a 42 g/L STORAGE
solution of sodium hydroxide R and allow to stand for 90 min. Protected from light.
Neutralise with 5.0 mL of a 103 g/L solution of hydrochloric
acid R and mix well. IMPURITIES
Reference solution (b). Dilute 1.0 mL of the test solution to Specified impurities : A.
100.0 mL with water for chromatography R. Dilute 1.0 mL of Other detectable impurities (the following substances would,
this solution to 10.0 mL with water for chromatography R. if present at a sufficient level, be detected by one or other of
Reference solution (c). Dissolve 50.0 mg of ribavirin CRS in the tests in the monograph. They are limited by the general
water for chromatography R and dilute to 100.0 mL with the acceptance criterion for other/unspecified impurities and/or
same solvent. by the general monograph Substances for pharmaceutical use
Column : for demonstration of compliance. See also 5.10. Control of
– size : l = 0.15 m, Ø = 4.6 mm ; impurities in substances for pharmaceutical use) : B, C, D, F, G.

Ribavirin EUROPEAN PHARMACOPOEIA
D. 1H-1,2,4-triazole-3-carboxamide,
A. 1-β-D-ribofuranosyl-1H-1,2,4-triazole-3-carboxylic acid,
F. 1-(5-O-acetyl-β-D-ribofuranosyl)-1H-1,2,4-triazole-3-
B. 1-α-D-ribofuranosyl-1H-1,2,4-triazole-3-carboxamide carboxamide (5′-O-acetylribavirin),
(anomer),
G. 1-β-D-ribofuranosyl-1H-1,2,4-triazole-5-carboxamide
C. 1H-1,2,4-triazole-3-carboxylic acid, (N-isomer).

EUROPEAN PHARMACOPOEIA Ritonavir
01/2017:2136 Column :
– size : l = 0.15 m, Ø = 4.6 mm ;
– stationary phase : end-capped butylsilyl silica gel for
RITONAVIR – temperature : 60 °C.
Mobile phase :
Ritonavirum – mobile phase A : mix 5 volumes of butanol R, 8 volumes
of tetrahydrofuran R, 18 volumes of acetonitrile R and
69 volumes of a 4.1 g/L solution of potassium dihydrogen
phosphate R filtered through a 0.45 µm nylon membrane ;
– mobile phase B : mix 5 volumes of butanol R, 8 volumes
of tetrahydrofuran R, 40 volumes of a 4.1 g/L solution
of potassium dihydrogen phosphate R filtered through a
0.45 μm nylon membrane and 47 volumes of acetonitrile R ;
0 - 60 100 0
C37H48N6O5S2 Mr 721 60 - 120 100 → 0 0 → 100
[155213-67-5]
120 - 120.1 0 → 100 100 → 0
DEFINITION
120.1 - 155 100 0
Thiazol-5-ylmethyl [(1S,2S,4S)-1-benzyl-2-hydroxy-4-
[[(2S)-3-methyl-2-[[methyl[[2-(1-methylethyl)thiazol- Flow rate : 1.0 mL/min.
4-yl]methyl]carbamoyl]amino]butanoyl]amino]-5-
phenylpentyl]carbamate.
Content : 97.0 per cent to 102.0 per cent (anhydrous substance). Injection : 50 µL of test solution (a) and reference solutions (a)
and (b).
PRODUCTION Identification of impurities : use the chromatogram
The production method is validated to demonstrate suitable supplied with ritonavir for peak identification CRS and the
enantiomeric purity of the final product. chromatogram obtained with reference solution (a) to identify
the peaks due to impurities E, F, L, O and T.
CHARACTERS
Relative retention with reference to ritonavir (retention
Appearance : white or almost white powder. time = about 34 min): impurity E = about 0.39 ;
Solubility : practically insoluble in water, freely soluble in impurity F = about 0.40 ; impurity L = about 0.8 ;
methanol and in methylene chloride, very slightly soluble in impurity O = about 1.1 ; impurity T = about 2.6.
acetonitrile. System suitability : reference solution (a) :
It shows polymorphism (5.9).
IDENTIFICATION above the baseline of the peak due to impurity E and
Infrared absorption spectrophotometry (2.2.24). H v = height above the baseline of the lowest point of the
curve separating this peak from the peak due to impurity F.
Comparison : ritonavir CRS.
Limits :
If the spectra obtained in the solid state show differences
dissolve the substance to be examined and the reference – correction factors : for the calculation of content,
substance separately in methylene chloride R, evaporate to multiply the peak areas of the following impurities by
dryness and record new spectra using the residues. the corresponding correction factor : impurity F = 1.4 ;
impurity L = 1.9 ; impurity T = 1.4 ;
TESTS – impurities E, O : for each impurity, not more than 3 times
Related substances. Liquid chromatography (2.2.29). the area of the principal peak in the chromatogram
Solvent mixture. Mix equal volumes of acetonitrile R and a obtained with reference solution (b) (0.3 per cent) ;
4.1 g/L solution of potassium dihydrogen phosphate R. – impurity T : not more than twice the area of the principal
Test solution (a). Dissolve 10.0 mg of the substance to be peak in the chromatogram obtained with reference
examined in the solvent mixture and dilute to 10.0 mL with solution (b) (0.2 per cent);
the solvent mixture. Sonicate if necessary. – impurities F, L : for each impurity, not more than the area
Test solution (b). Dilute 5.0 mL of test solution (a) to 10.0 mL of the principal peak in the chromatogram obtained with
with the solvent mixture. Dilute 1.0 mL of this solution to reference solution (b) (0.1 per cent) ;
20.0 mL with the solvent mixture. – unspecified impurities : for each impurity, not more than the
Reference solution (a). Dissolve 5.0 mg of ritonavir for peak area of the principal peak in the chromatogram obtained
identification CRS (containing impurities E, F, L, O and T) with reference solution (b) (0.10 per cent) ;
in the solvent mixture and dilute to 5.0 mL with the solvent – total : not more than 10 times the area of the principal peak
mixture. Sonicate if necessary. in the chromatogram obtained with reference solution (b)
Reference solution (b). Dilute 1.0 mL of test solution (a) to (1.0 per cent);
10.0 mL with the solvent mixture. Dilute 1.0 mL of this – disregard limit : 0.5 times the area of the principal peak in
solution to 100.0 mL with the solvent mixture. the chromatogram obtained with reference solution (b)
Reference solution (c). Dissolve 10.0 mg of ritonavir CRS in (0.05 per cent).
mixture. Sonicate if necessary. Dilute 5.0 mL of this solution Water (2.5.12) : maximum 0.5 per cent, determined on 0.500 g.
to 10.0 mL with the solvent mixture. Dilute 1.0 mL of this Sulfated ash (2.4.14): maximum 0.2 per cent, determined on
solution to 20.0 mL with the solvent mixture. 1.0 g.

Ritonavir EUROPEAN PHARMACOPOEIA
ASSAY
Injection : test solution (b) and reference solution (c).
Calculate the percentage content of C37H48N6O5S2 from the
declared content of ritonavir CRS.
STORAGE
IMPURITIES E. thiazol-5-ylmethyl [(1S,2S,4S)-1-benzyl-2-hydroxy-
Specified impurities : E, F, L, O, T. 4-[[(2S)-2-[[[[2-(1-hydroxy-1-methylethyl)thiazol-4-
Other detectable impurities (the following substances would, yl]methyl]methylcarbamoyl]amino]-3-methylbutanoyl]-
if present at a sufficient level, be detected by one or other of amino]-5-phenylpentyl]carbamate,
Control of impurities in substances for pharmaceutical use) : A,
B, C, D, G, H, I, J, K, M, N, P, Q, R, S, U.
F. thiazol-5-ylmethyl [(1S,2S,4S)-1-benzyl-4-[[(2S)-
1-benzyl-2-hydroxy-4-[(4S)-4-(1-methylethyl)-2,5-
dioxoimidazolidin-1-yl]-5-phenylpentyl]carbamate,
A. (2S)-3-methyl-2-[[methyl[[2-(1-methylethyl)thiazol-4-
yl]methyl]carbamoyl]amino]butanoic acid,
G. thiazol-5-ylmethyl [(1S,2S,4S)-1-benzyl-4-[[(2S)-
2-[[[[2-(1-hydroperoxy-1-methylethyl)thiazol-4-
yl]methyl]methylcarbamoyl]amino]-3-methylbutanoyl]-
B. thiazol-5-ylmethyl [(1S,2S,4S)-4-[[(2S)-2-amino- amino]-2-hydroxy-5-phenylpentyl]carbamate,
3-methylbutanoyl]amino]-1-benzyl-2-hydroxy-5-
phenylpentyl]carbamate,
H. thiazol-5-ylmethyl (4S,5S)-4-benzyl-5-[(2S)-2-[(4S)-
4-(1-methylethyl)-2,5-dioxoimidazolidin-1-yl]-3-
C. thiazol-5-ylmethyl [(1S,2S,4S)-4-(acetylamino)-1-benzyl- phenylpropyl]-2-oxooxazolidine-3-carboxylate,
2-hydroxy-5-phenylpentyl]carbamate,
I. thiazol-5-ylmethyl [((1S,2S,4S)-1-benzyl-4-[[(2S)-
D. thiazol-5-ylmethyl [(1S,2S,4S)-1-benzyl-2-hydroxy-5- 2-[[[[2-ethylthiazol-4-yl]methyl]methylcarbamoyl]-
phenyl-4-[[(thiazol-5-ylmethoxy)carbonyl]amino]pentyl]- amino]-3-methylbutanoyl]amino]-2-hydroxy-5-phenyl-
carbamate, pentyl]carbamate,

EUROPEAN PHARMACOPOEIA Ritonavir
J. thiazol-5-ylmethyl [(1S,2S,4S)-1-benzyl-4-[[(1,1-
dimethylethoxy)carbonyl]amino]-2-hydroxy-5- O. thiazol-5-ylmethyl [(1S,2R,4S)-1-benzyl-2-hydroxy-4-
phenylpentyl]carbamate, [[(2S)-3-methyl-2-[[methyl[[2-(1-methylethyl)thiazol-
4-yl]methyl]carbamoyl]amino]butanoyl]amino]-5-
K. thiazol-5-ylmethyl (1S,2S,4S)-1-benzyl-2-hydroxy-
4-[[(2-methylpropoxy)carbonyl]amino]-5-
P. bis(thiazol-5-ylmethyl) [carbonylbis[imino[(2S,3S,5S)-3-
hydroxy-1,6-diphenylhexane-5,2-diyl]]]dicarbamate,
L. (4S,5S)-4-benzyl-5-[(2S)-2-[[(2S)-3-methyl-2-[[methyl[[2-
(1-methylethyl)thiazol-4-yl]methyl]carbamoyl]amino]-
butanoyl]amino]-3-phenylpropyl]oxazolidin-2-one,
Q. thiazol-5-ylmethyl [(1S,2R,4R)-1-benzyl-2-hydroxy-
4-[[(2S)-3-methyl-2-[[methyl[[2-(1-methylethyl)-
thiazol-4-yl]methyl]carbamoyl]amino]butanoyl]amino]-
5-phenylpentyl]carbamate,
M. 2-methylpropyl (2S)-3-methyl-2-[[methyl[[2-(1-methyl-
ethyl)thiazol-4-yl]methyl]carbamoyl]amino]butanoate,
N. thiazol-5-ylmethyl [(1S,3S,4S)-1-benzyl-3-hydroxy-4- R. thiazol-5-ylmethyl [(1S,2S,4R)-1-benzyl-2-hydroxy-4-

[[(2S)-3-methyl-2-[[methyl[[2-(1-methylethyl)thiazol- [[(2S)-3-methyl-2-[[methyl[[2-(1-methylethyl)thiazol-
4-yl]methyl]carbamoyl]amino]butanoyl]amino]-5- 4-yl]methyl]carbamoyl]amino]butanoyl]amino]-5-
phenylpentyl]carbamate, phenylpentyl]carbamate,

Ritonavir EUROPEAN PHARMACOPOEIA
U. (1S,3S)-3-[[(2S)-3-methyl-2-[[methyl[[2-(1-methylethyl)-
thiazol-4-yl]methyl]carbamoyl]amino]butanoyl]amino]-
S. thiazol-5-ylmethyl [(1S,2S,4S)-1-benzyl-4-[[(2S)-2- 4-phenyl-1-[(1S)-2-phenyl-1-[[(thiazol-5-ylmethoxy)-
[[[(1S,3S,4S)-1-benzyl-3-hydroxy-5-phenyl-4-[[(thiazol- carbonyl]amino]ethyl]butyl (2S)-3-methyl-2-[[methyl[[2-
5-ylmethoxy)carbonyl]amino]pentyl]carbamoyl]amino]- (1-methylethyl)thiazol-4-yl]methyl]carbamoyl]amino]-
3-methylbutanoyl]amino]-2-hydroxy-5-phenylpentyl]- butanoate.
carbamate,
T. (2S)-N-[(1S,2S,4S)-1-benzyl-2-hydroxy-4-[[(2S)-
3-methyl-2-[[methyl[[2-(1-methylethyl)thiazol-4-
yl]methyl]carbamoyl]amino]butanoyl]amino]-5-
phenylpentyl]-3-methyl-2-[[methyl[[2-(1-methylethyl)-
thiazol-4-yl]methyl]carbamoyl]amino]butanamide,

EUROPEAN PHARMACOPOEIA Saquinavir mesilate
01/2017:2267 Reference solution (c). Dissolve 30.0 mg of saquinavir

mesilate CRS in the solvent mixture, using sonication, and
Column :
– size : l = 0.15 m, Ø = 4.6 mm ;
SAQUINAVIR MESILATE – stationary phase : spherical end-capped octadecylsilyl silica
gel for chromatography R (3.5 μm).
Saquinaviri mesilas Mobile phase :
– mobile phase A : to 2.5 mL of strong sodium hydroxide
solution R add 900 mL of water for chromatography R,
adjust to pH 1.8 with perchloric acid R and dilute to
1000 mL with water for chromatography R ;
– mobile phase B : mobile phase A, acetonitrile R1
(38:62 V/V) ;
(min) (per cent) (per cent)
0-1 50 50
C39H54N6O8S Mr 767
[149845-06-7] 1 - 31 50 → 0 50 → 100
DEFINITION Flow rate : 1.0 mL/min.

(2S)-N1-[(1S,2R)-1-Benzyl-3-[(3S,4aS,8aS)-3-[(1,1- Detection : spectrophotometer at 210 nm.
dimethylethyl)carbamoyl]octahydroisoquinolin-2(1H)- Injection : 10 µL of the test solution and reference solutions (a)
yl]-2-hydroxypropyl]-2-[(quinolin-2-ylcarbonyl)amino]- and (b).
butanediamide methanesulfonate.
Content : 97.5 per cent to 102.0 per cent (anhydrous substance). supplied with saquinavir for system suitability CRS and the
PRODUCTION chromatogram obtained with reference solution (b) to identify
the peaks due to impurities A, B, C and D.
It is considered that alkyl methanesulfonate esters are
genotoxic and are potential impurities in saquinavir Relative retention with reference to saquinavir (retention
mesilate. The manufacturing process should be developed time = about 17 min) : impurity A = about 0.2 ;
taking into consideration the principles of quality risk impurity B = about 0.3 ; impurity C = about 0.5 ;
management, together with considerations of the quality impurity D = about 0.9.
of starting materials, process capability and validation. System suitability : reference solution (b) :
The general methods 2.5.37. Methyl, ethyl and isopropyl – peak-to-valley ratio : minimum 3, where Hp = height
methanesulfonate in methanesulfonic acid, 2.5.38. Methyl, above the baseline of the peak due to impurity D and
ethyl and isopropyl methanesulfonate in active substances and Hv = height above the baseline of the lowest point of the
2.5.39. Methanesulfonyl chloride in methanesulfonic acid are curve separating this peak from the peak due to saquinavir.
available to assist manufacturers. Limits :
CHARACTERS – correction factors : for the calculation of content,
Appearance : white or almost white, slightly hygroscopic multiply the peak areas of the following impurities by
powder. the corresponding correction factor : impurity A = 0.5 ;
impurity B = 0.5 ; impurity C = 2.5 ;
Solubility : practically insoluble in water, sparingly soluble in
methanol, slightly soluble in ethanol (96 per cent). – impurities A, B, C : for each impurity, not more than
1.5 times the area of the principal peak in the chromatogram
IDENTIFICATION obtained with reference solution (a) (0.15 per cent) ;
A. Specific optical rotation (see Tests). – unspecified impurities : for each impurity, not more than
B. Infrared absorption spectrophotometry (2.2.24). 0.5 times the area of the principal peak in the chromatogram
Comparison : saquinavir mesilate CRS. obtained with reference solution (a) (0.05 per cent) ;
TESTS in the chromatogram obtained with reference solution (a)
Specific optical rotation (2.2.7) : − 42.0 to − 35.0 (anhydrous (0.5 per cent);
substance). – disregard limit : not more than 0.3 times the area of
Dissolve 0.25 g in anhydrous methanol R and dilute to 50.0 mL the principal peak in the chromatogram obtained with
with the same solvent. reference solution (a) (0.03 per cent).
Related substances. Liquid chromatography (2.2.29). Water (2.5.12) : maximum 1.0 per cent, determined on 0.250 g.
Solvent mixture : water for chromatography R, acetonitrile R1 Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
(47:53 V/V). 1.0 g.
ASSAY
examined in the solvent mixture, using sonication, and dilute
to 100.0 mL with the same solvent. Liquid chromatography (2.2.29) as described in the test for
Reference solution (a). Dilute 1.0 mL of the test solution to related substances with the following modification.
100.0 mL with the solvent mixture. Dilute 1.0 mL of this Injection : 10 µL of the test solution and reference solution (c).
solution to 10.0 mL with the solvent mixture. Calculate the percentage content of saquinavir mesilate from
Reference solution (b). Dissolve the contents of a vial of the assigned content of saquinavir mesilate CRS.
saquinavir for system suitability CRS (containing impurities A,
B, C and D) in 1.0 mL of the solvent mixture and sonicate for STORAGE
2 min. In an airtight container, protected from light.

Saquinavir mesilate EUROPEAN PHARMACOPOEIA
IMPURITIES
Specified impurities : A, B, C.
impurities in substances for pharmaceutical use) : D, E, F, G, H. E. (3S)-4-[[(1S,2R)-1-benzyl-3-[(3S,4aS,8aS)-3-[(1,1-
dimethylethyl)carbamoyl]octahydroisoquinolin-2(1H)-
yl]-2-hydroxypropyl]amino]-4-oxo-3-[(quinolin-2-
ylcarbonyl)amino]butanoic acid,
A. (2S)-4-amino-4-oxo-2-[(quinolin-2-ylcarbonyl)amino]-
butanoic acid,
F. N-[(1S)-2-[[(1S,2R)-1-benzyl-3-[(3S,4aS,8aS)-3-[(1,1-
dimethylethyl)carbamoyl]octahydroisoquinolin-2(1H)-
yl]-2-hydroxypropyl]amino]-1-(cyanomethyl)-2-
oxoethyl]quinoline-2-carboxamide,
B. ethyl (2S)-4-amino-4-oxo-2-[(quinolin-2-ylcarbonyl)-
amino]butanoate,
G. methyl (3S)-4-[[(1S,2R)-1-benzyl-3-[(3S,4aS,8aS)-3-
[(1,1-dimethylethyl)carbamoyl]octahydroisoquinolin-
C. (3S,4aS,8aS)-2-[(2R,3S)-3-amino-2-hydroxy-4-phenyl- 2(1H)-yl]-2-hydroxypropyl]amino]-4-oxo-3-[(quinolin-2-
butyl]-N-(1,1-dimethylethyl)decahydroisoquinoline-3- ylcarbonyl)amino]butanoate,
carboxamide,
D. (2R)-N1-[(1S,2R)-1-benzyl-3-[(3S,4aS,8aS)-3-[(1,1- H. N-[(3S)-1-[(1S,2R)-1-benzyl-3-[(3S,4aS,8aS)-3-[(1,1-
dimethylethyl)carbamoyl]octahydroisoquinolin-2(1H)- dimethylethyl)carbamoyl]octahydroisoquinolin-2(1H)-
yl]-2-hydroxypropyl]-2-[(quinolin-2-ylcarbonyl)amino]- yl]-2-hydroxypropyl]-2,5-dioxopyrrolidin-3-yl]quinoline-
butanediamide (2-epi-saquinavir), 2-carboxamide.

EUROPEAN PHARMACOPOEIA Stavudine
01/2015:2130 Time Mobile phase A Mobile phase B

0 - 10 100 0
10 - 20 100 → 0 0 → 100
20 - 30 0 100
STAVUDINE
Stavudinum Detection : spectrophotometer at 254 nm.
Injection : 10 µL.
supplied with stavudine for system suitability CRS and the
chromatogram obtained with reference solution (c) to identify
the peaks due to impurities A, B, C, E and G.
Relative retention with reference to stavudine (retention
time = about 10 min) : impurity A = about 0.3 ;
C10H12N2O4 Mr 224.2 impurity B = about 0.50 ; impurity C = about 0.53 ;
[3056-17-5] impurity E = about 1.1 ; impurity G = about 1.9.
DEFINITION – peak-to-valley ratio : minimum 1.5, where Hp = height
1-(2,3-Dideoxy-β-D-glycero-pent-2-enofuranosyl)-5- above the baseline of the peak due to impurity C and
methylpyrimidine-2,4(1H,3H)-dione. Hv = height above the baseline of the lowest point of the
Content : 97.0 per cent to 102.0 per cent (anhydrous substance). curve separating this peak from the peak due to impurity B ;
minimum 1.5, where Hp = height above the baseline of the
CHARACTERS peak due to impurity E and Hv = height above the baseline
Appearance : white or almost white powder. of the lowest point of the curve separating this peak from
Solubility : soluble in water, sparingly soluble in ethanol the peak due to stavudine.
(96 per cent), slightly soluble in methylene chloride. Calculation of percentage contents :
It shows polymorphism (5.9). – correction factor : multiply the peak area of impurity A by
0.7 ;
IDENTIFICATION – for impurity A, use the concentration of stavudine in
A. Specific optical rotation (2.2.7): − 45.9 to − 39.5 (anhydrous reference solution (a);
substance). – for impurities other than A, use the concentration of
Dissolve 0.100 g in water R and dilute to 10.0 mL with the stavudine in reference solution (b).
same solvent. Limits :
B. Infrared absorption spectrophotometry (2.2.24). – impurity A : maximum 0.5 per cent ;
Comparison : stavudine CRS. – impurity G : maximum 0.2 per cent ;
If the spectra obtained show differences, dissolve the – unspecified impurities : for each impurity, maximum
substance to be examined and the reference substance 0.10 per cent ;
separately in anhydrous ethanol R, evaporate to dryness
and record new spectra using the residues.
– reporting threshold : 0.05 per cent.
TESTS Impurity I. Liquid chromatography (2.2.29). Prepare the
Related substances. Liquid chromatography (2.2.29). Prepare solutions immediately before use or store them at 2-8 °C until
the solutions immediately before use or store them at 2-8 °C use.
until use. Test solution. Dissolve 50.0 mg of the substance to be
Test solution. Dissolve 25.0 mg of the substance to be examined in the mobile phase and dilute to 50.0 mL with the
examined in water R and dilute to 50.0 mL with the same mobile phase.
solvent. Reference solution (a). Dissolve 2 mg of stavudine
Reference solution (a). Dilute 0.5 mL of the test solution to impurity I CRS in the mobile phase and dilute to 50.0 mL with
100.0 mL with water R. the mobile phase. Dilute 1.0 mL of the solution to 20.0 mL
Reference solution (b). Dilute 20.0 mL of reference solution (a) with the mobile phase.
to 100.0 mL with water R. Reference solution (b). Dilute 1.0 mL of the test solution to
Reference solution (c). Dissolve 5 mg of stavudine for system 100.0 mL with the mobile phase. Dilute 1.0 mL of this solution
suitability CRS (containing impurities A, B, C, E and G) in to 10.0 mL with the mobile phase.
water R and dilute to 10.0 mL with the same solvent. Column :
Column : – size : l = 0.25 m, Ø = 4.6 mm ;
– size : l = 0.25 m, Ø = 4.6 mm ; – stationary phase : end-capped octylsilyl silica gel for
– stationary phase : base-deactivated end-capped octadecylsilyl chromatography R (5 µm).
silica gel for chromatography R (5 μm). Mobile phase : mix 30 volumes of acetonitrile for
Mobile phase : chromatography R and 70 volumes of a 1.15 g/L solution of
ammonium dihydrogen phosphate R previously adjusted to
– mobile phase A : mix 35 volumes of acetonitrile for pH 6.8 with triethylamine R.
chromatography R and 965 volumes of a 0.77 g/L solution
of ammonium acetate R ; Flow rate : 1.0 mL/min.
– mobile phase B : mix 250 volumes of acetonitrile for
chromatography R and 750 volumes of a 0.77 g/L solution Injection : 20 µL.
of ammonium acetate R ; Run time : 7 times the retention time of stavudine.

Stavudine EUROPEAN PHARMACOPOEIA
Identification of impurities : use the chromatogram obtained IMPURITIES

with reference solution (a) to identify the peak due to Specified impurities : A, G, I.
impurity I.
Relative retention with reference to stavudine (retention if present at a sufficient level, be detected by one or other of
time = about 3 min) : impurity I = about 6.0. the tests in the monograph. They are limited by the general
System suitability : reference solution (b) : acceptance criterion for other/unspecified impurities and/or
– signal-to-noise ratio : minimum 40 for the principal peak. by the general monograph Substances for pharmaceutical
Calculation of percentage content : impurities for demonstration of compliance. See also 5.10.
– correction factor : multiply the peak area of impurity I by Control of impurities in substances for pharmaceutical use): B,
1.7 ; C, D, E, F, H.
– for impurity I, use the concentration of stavudine in
reference solution (b).
Limit :
– impurity I : maximum 0.15 per cent.
Water (2.5.12): maximum 0.5 per cent, determined on 0.500 g.
A. 5-methylpyrimidine-2,4(1H,3H)-dione (thymine),
1.0 g.
ASSAY
Liquid chromatography (2.2.29). Prepare the solutions
immediately before use or store them at 2-8 °C until use.
examined in water R and dilute to 100.0 mL with the same
solvent. Dilute 10.0 mL of the solution to 50.0 mL with B. 1-(2-deoxy-β-D-threo-pentofuranosyl)-5-
water R. methylpyrimidine-2,4(1H,3H)-dione (3′-epithymidine),
Reference solution (a). Dissolve 10.0 mg of stavudine CRS in
water R and dilute to 100.0 mL with the same solvent. Dilute
10.0 mL of the solution to 50.0 mL with water R.
Reference solution (b). Dissolve 5 mg of thymine R
(impurity A) and 7.5 mg of thymidine R (impurity C) in
water R and dilute to 100.0 mL with the same solvent. Dilute
10.0 mL of the solution to 50.0 mL with water R.
Column :
– size : l = 0.033 m, Ø = 4.0 mm ;
C. 1-(2-deoxy-β-D-erythro-pentofuranosyl)-5-
– stationary phase : base-deactivated end-capped octadecylsilyl methylpyrimidine-2,4(1H,3H)-dione (thymidine),
silica gel for chromatography R (3 μm).
Mobile phase : mix 5 volumes of acetonitrile for
chromatography R and 95 volumes of a 0.77 g/L solution of
ammonium acetate R.
Injection : 25 µL.
Run time : twice the retention time of stavudine. D. 1-[(2R)-5-oxo-2,5-dihydrofuran-2-yl]-5-methylpyrimi-
Identification of impurities : use the chromatogram obtained dine-2,4(1H,3H)-dione,
with reference solution (b) to identify the peaks due to
impurities A and C.
Relative retention with reference to stavudine
(retention time = about 4 min): impurity A = about
0.4 ; impurity C = about 0.6.
impurities A and C in the chromatogram obtained with
reference solution (b) ; E. 1-(2,3-dideoxy-α-D-glycero-pent-2-enofuranosyl)-5-
methylpyrimidine-2,4(1H,3H)-dione (stavudine anomer α),
– symmetry factor : maximum 1.6 for the peak due to
stavudine in the chromatogram obtained with reference
solution (a).
Calculate the percentage content of C10H12N2O4 using the
chromatograms obtained with the test solution and reference
solution (a) and taking into account the assigned content of
stavudine CRS.
STORAGE F. 1-(3,5-anhydro-2-deoxy-β-D-threo-pentofuranosyl)-5-
Protected from light and humidity. methylpyrimidine-2,4(1H,3H)-dione,

EUROPEAN PHARMACOPOEIA Stavudine
G. 1-[2-deoxy-5-O-[[(2S,5R)-5-[5-methyl-2,4-dioxo-3,4- I. 1-(5-O-benzoyl-2,3-dideoxy-β-D-glycero-pent-2-
dihydropyrimidine-1(2H)-yl]-2,5-dihydrofuran- enofuranosyl)-5-methylpyrimidine-2,4(1H,3H)-dione.
2-yl]methyl]-β-D-threo-pentofuranosyl]-5-
methylpyrimidine-2,4(1H,3H)-dione,
H. 1-[2-deoxy-5-O-(1-methylethyl)-β-D-threo-
pentofuranosyl]-5-methylpyrimidine-2,4(1H,3H)-dione,

EUROPEAN PHARMACOPOEIA Valaciclovir hydrochloride
07/2019:1768 Pretreatment : wash the plate with methanol R until the solvent
front has migrated over at least 4/5 of the plate ; allow the
plate to dry.
Mobile phase : concentrated ammonia R, tetrahydrofuran R,
methanol R, methylene chloride R (3:12:34:54 V/V/V/V) ; use
VALACICLOVIR HYDROCHLORIDE freshly prepared mobile phase.
Application : 4 µL of the test solution and reference
solutions (b), (c) and (d).
Valacicloviri hydrochloridum Development : over 4/5 of the plate.
Drying : in a current of air.
Detection : examine in ultraviolet light at 254 nm for
impurities E and G ; spray with a 0.1 g/L solution of
fluorescamine R in ethylene chloride R and examine in
ultraviolet light at 365 nm for impurity F.
Retardation factors : impurity A = about 0 ;
C13H21ClN6O4 Mr 360.8 impurity B = about 0.2 ; valaciclovir = about 0.3 ;
[124832-27-5] impurity C = about 0.5 ; impurity D = about 0.6 ;
impurity E = about 0.7 ; impurity F = about 0.75 ;
DEFINITION impurity G = about 0.79 ; impurity C is masked by the leading
2-[(2-Amino-6-oxo-1,6-dihydro-9H-purin-9-yl)- edge of the spot due to valaciclovir ; impurities F and G may
methoxy]ethyl L-valinate hydrochloride. co-elute, but this does not adversely affect their quantification
Content : 95.0 per cent to 102.0 per cent (anhydrous substance). because they are visualised differently.
System suitability : the chromatograms obtained with reference
CHARACTERS solutions (b), (c) and (d) each show 3 clearly separated spots
Appearance : white or almost white powder. when examined under ultraviolet light at 254 nm, due to
impurities D, E and G.
Solubility : freely soluble in water, slightly soluble in anhydrous
Limits :
ethanol.
– impurity E : any spot due to impurity E is not more intense
It shows polymorphism (5.9). than the corresponding spot in the chromatogram obtained
with reference solution (c) (0.2 per cent) ;
IDENTIFICATION
– impurity F : any spot due to impurity F is not more intense
Carry out either tests A, B, C, E or tests A, B, D, E. than the corresponding spot in the chromatogram obtained
A. Infrared absorption spectrophotometry (2.2.24). with reference solution (b) (0.3 per cent calculated as
Comparison : anhydrous valaciclovir hydrochloride CRS. hydrochloride salt) ;
If the spectra obtained in the solid state show differences, – impurity G : any spot due to impurity G is not more intense
dissolve the substance to be examined and the reference than the corresponding spot in the chromatogram obtained
substance separately in the minimum volume of anhydrous with reference solution (d) (0.05 per cent).
ethanol R and evaporate to dryness in a desiccator, under Related substances.
high vacuum, over diphosphorus pentoxide R. Record new A. Impurities A, B, I and R. Liquid chromatography (2.2.29) :
spectra using the residues. use the normalisation procedure.
B. It gives reaction (a) of chlorides (2.3.1). Test solution. Dissolve 50.0 mg of the substance to be
C. It complies with the limit for impurity R given in test A examined in a 0.5 per cent V/V solution of hydrochloric
for related substances. acid R and dilute to 100.0 mL with the same solution.
D. Optical rotation (2.2.7) : laevorotatory. Reference solution (a). Dissolve 2.5 mg of valaciclovir for
system suitability CRS (containing impurities A, B, C, D, H,
Dissolve 2.50 g in water R and dilute to 50.0 mL with the I, J, M and R) in a 0.5 per cent V/V solution of hydrochloric
same solvent. acid R and dilute to 5.0 mL with the same solution.
E. Water (see Tests). Reference solution (b). Dissolve 50.0 mg of anhydrous
valaciclovir hydrochloride CRS in a 0.5 per cent V/V
TESTS solution of hydrochloric acid R and dilute to 100.0 mL with
Impurities E, F and G. Thin-layer chromatography (2.2.27). the same solution.
Test solution. Dissolve 0.250 g of the substance to be examined Reference solution (c). Dilute 3.0 mL of the test solution to
in 2 mL of water R and dilute to 5.0 mL with ethanol (96 per 100.0 mL with a 0.5 per cent V/V solution of hydrochloric
cent) R. acid R. Dilute 1.0 mL of this solution to 100.0 mL with a
0.5 per cent V/V solution of hydrochloric acid R.
Reference solution (a). Dissolve 5 mg of valaciclovir
impurity D CRS, 5.0 mg of valaciclovir impurity E CRS, 5.0 mg Column :
of valaciclovir impurity G CRS and 8.4 mg of valaciclovir – size : l = 0.15 m, Ø = 4.0 mm ;
impurity F para-toluenesulfonate CRS in a mixture of 2 mL – stationary phase : crown-ether silica gel for chiral
of water R and 6 mL of ethanol (96 per cent) R, and dilute to separation R (5 µm) ;
10.0 mL with ethanol (96 per cent) R. – temperature : 10 °C.
Reference solution (b). Dilute 3.0 mL of reference solution (a) Mobile phase : perchloric acid R, methanol R, water for
to 10.0 mL with ethanol (96 per cent) R. chromatography R (0.5:5:95 V/V/V).
Reference solution (c). Dilute 2.0 mL of reference solution (a) Flow rate : 0.75 mL/min.
to 10.0 mL with ethanol (96 per cent) R. Detection : spectrophotometer at 254 nm.
Reference solution (d). Dilute 0.5 mL of reference solution (a) Injection : 10 µL of the test solution and reference
to 10.0 mL with ethanol (96 per cent) R. solutions (a) and (c).
Plate : TLC silica gel F254 plate R (2-10 μm). Run time : 1.5 times the retention time of valaciclovir.

Valaciclovir hydrochloride EUROPEAN PHARMACOPOEIA
Identification of impurities : use the chromatogram Relative retention with reference to valaciclovir
supplied with valaciclovir for system suitability CRS and (retention time = about 19 min) : impurity A = about 0.3 ;
the chromatogram obtained with reference solution (a) impurity B = about 0.4 ; impurity H = about 0.5 ;
to identify the peaks due to impurities A + B, C + R, D, I impurity C = about 1.06 ; impurity I = about 1.09 ;
and M. impurity D = about 1.2 ; impurity J = about 1.3 ;
Relative retention with reference to valaciclovir (retention impurity M = about 1.6.
time = about 21 min) : impurities A and B = about 0.2 ; System suitability : reference solution (a) :
impurity I = about 0.4 ; impurities C and R = about 0.6 ; – peak-to-valley ratio : minimum 2.5, where Hp = height
impurity D = about 0.7 ; impurity M = about 1.3. above the baseline of the peak due to impurity C and
System suitability : reference solution (a) : Hv = height above the baseline of the lowest point of
– peak-to-valley ratio : minimum 1.5, where Hp = height the curve separating this peak from the peak due to
above the baseline of the peak due to impurity D and valaciclovir ;
Hv = height above the baseline of the lowest point of – the chromatogram obtained is similar to the
the curve separating this peak from the peak due to chromatogram supplied with valaciclovir for system
impurities C and R. suitability CRS.
Limits : Limits :
– correction factor : for the calculation of content, multiply – impurity M : maximum 1.5 per cent ;
the peak area of impurities A and B by 0.7 ; – impurity D : maximum 0.5 per cent ;
– impurity R : maximum 3.0 per cent ; for the calculation, – impurity C : maximum 0.3 per cent ;
subtract the content of impurity C as determined in – impurities H, J : for each impurity, maximum 0.10 per
test B for related substances from the content of the cent ;
coeluting impurities C and R as determined in this test ;
– sum of impurities A and B : maximum 2.0 per cent ; 0.05 per cent ;
– impurity I : maximum 0.2 per cent ; – disregard limit : 0.6 times the area of the principal
– disregard limit : the area of the principal peak in the peak in the chromatogram obtained with reference
chromatogram obtained with reference solution (c) solution (b) (0.03 per cent) ; disregard the peaks due to
(0.03 per cent); disregard any peaks due to impurities impurities A, B and I.
other than A + B, C + R or I. Limit :
B. Liquid chromatography (2.2.29) : use the normalisation – total for tests A and B : maximum 5.0 per cent.
procedure. Use the solutions within 24 h of preparation.
Chloride : 9.4 to 9.9 per cent (anhydrous and solvent-free
Solvent mixture : ethanol (96 per cent) R, water R substance).
(20:80 V/V). Dissolve 0.350 g in 100 mL of water R and add 0.2 mL of nitric
Test solution. Dissolve 40 mg of the substance to be acid R. Carry out a potentiometric titration (2.2.20), using
examined in the solvent mixture and dilute to 100.0 mL 0.1 M silver nitrate. Use a silver indicator electrode and a
with the solvent mixture. silver-silver chloride reference electrode or a combined silver
Reference solution (a). Dissolve 2.5 mg of valaciclovir for electrode. Discard the result from the first titration, which is
system suitability CRS (containing impurities A, B, C, D, H, used to condition the electrodes. Carry out a blank titration.
I, J, M and R) in the solvent mixture and dilute to 5.0 mL 1 mL of 0.1 M silver nitrate is equivalent to 3.543 mg of Cl.
with the solvent mixture.
Water (2.5.12) : maximum 2.0 per cent, determined on 0.250 g.
100.0 mL with the solvent mixture. Dilute 1.0 mL of this Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
solution to 20.0 mL with the solvent mixture. 1.0 g.
Column : ASSAY
– size : l = 0.25 m, Ø = 4.6 mm ; Liquid chromatography (2.2.29) as described in test A for
– stationary phase : end-capped phenylhexylsilyl silica gel related substances with the following modification.
for chromatography R (5 µm) ; Injection : test solution and reference solution (b).
– temperature : 15 °C. Calculate the percentage content of C13H21ClN6O4 taking
Mobile phase : into account the assigned content of anhydrous valaciclovir
hydrochloride CRS.
– mobile phase A : trifluoroacetic acid R, water for
chromatography R (0.2:100 V/V) ; IMPURITIES
– mobile phase B : trifluoroacetic acid R, methanol R2 Specified impurities : A, B, C, D, E, F, G, H, I, J, M, R.
(0.2:100 V/V) ; Other detectable impurities (the following substances would,
Time Mobile phase A Mobile phase B if present at a sufficient level, be detected by one or other of
(min) (per cent V/V) (per cent V/V) the tests in the monograph. They are limited by the general
0-5 90 10
5 - 35 90 → 60 10 → 40 use (2034). It is therefore not necessary to identify these
Flow rate : 0.8 mL/min. Control of impurities in substances for pharmaceutical use) :
Detection : spectrophotometer at 254 nm. K, L, N, O, P, Q.
Injection : 10 µL.
supplied with valaciclovir for system suitability CRS and
to identify the peaks due to impurities A, B, C, D, H, I,
J and M. A. 2-amino-1,9-dihydro-6H-purin-6-one (guanine),

EUROPEAN PHARMACOPOEIA Valaciclovir hydrochloride
B. 2-amino-9-[(2-hydroxyethoxy)methyl]-1,9-dihydro-6H- J. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
purin-6-one (aciclovir), yl)methoxy]ethyl L-isoleucinate,
C. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9- K. 9-[(2-hydroxyethoxy)methyl]-2-[[[(6-oxo-6,9-dihydro-1H-
yl)methoxy]ethyl N-methyl-L-valinate, purin-2-yl)amino]methyl]amino]-1,9-dihydro-6H-purin-
6-one,
D. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl N-ethyl-L-valinate, L. 2,2′-(methylenediazanediyl)bis[9-[(2-hydroxye-
thoxy)methyl]-1,9-dihydro-6H-purin-6-one],
M. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
E. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9- yl)methoxy]ethyl N-formyl-L-valinate,
yl)methoxy]ethyl N-[(benzyloxy)carbonyl]-L-valinate,
F. 2-hydroxyethyl L-valinate,
N. 2-[[6-oxo-2-[[[(6-oxo-6,9-dihydro-1H-purin-2-
yl)amino]methyl]amino]-1,6-dihydro-9H-purin-9-
yl]methoxy]ethyl L-valinate,
G. N,N-dimethylpyridin-4-amine,
O. 2-[[2-[[[[9-[(2-hydroxyethoxy)methyl]-6-oxo-6,9-dihydro-
1H-purin-2-yl]amino]methyl]amino]-6-oxo-1,6-dihydro-
9H-purin-9-yl]methoxy]ethyl L-valinate,
H. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl L-alaninate,
I. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9- P. [methylenebis[azanediyl(6-oxo-1,6-dihydro-9H-purine-
yl)methoxy]ethyl acetate, 2,9-diyl)methyleneoxyethan-2,1-diyl]] di-L-valinate,

Valaciclovir hydrochloride EUROPEAN PHARMACOPOEIA
R. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl D-valinate.
Q. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl N-[[(6-oxo-6,9-dihydro-1H-purin-2-
yl)amino]methyl]-L-valinate,

EUROPEAN PHARMACOPOEIA Valaciclovir hydrochloride hydrate
07/2019:2751 Limits :
– impurity G : any spot due to impurity G is not more intense
than the corresponding spot in the chromatogram obtained
with the reference solution (0.05 per cent) ;
– impurity S : any spot due to impurity S is not more intense
VALACICLOVIR HYDROCHLORIDE than the corresponding spot in the chromatogram obtained
HYDRATE with the reference solution (0.05 per cent).
Related substances
Valacicloviri hydrochloridum hydricum A. Liquid chromatography (2.2.29) : use the normalisation
procedure.
examined in a 0.5 per cent V/V solution of hydrochloric
acid R and dilute to 100.0 mL with the same solution.
Reference solution (a). Dissolve 2.5 mg of valaciclovir for
system suitability CRS (containing impurities A, B, C, D,
C13H21ClN6O4,xH2O Mr 360.8 (anhydrous substance) H, M and R) in a 0.5 per cent V/V solution of hydrochloric
[1218948-84-5] acid R and dilute to 5.0 mL with the same solution.
DEFINITION Reference solution (b). Dissolve 50.0 mg of anhydrous
valaciclovir hydrochloride CRS in a 0.5 per cent V/V
2-[(2-Amino-6-oxo-1,6-dihydro-9H-purin-9-yl)methoxy]- solution of hydrochloric acid R and dilute to 100.0 mL with
ethyl L-valinate hydrochloride hydrate. the same solution.
Content : 95.0 per cent to 102.0 per cent (anhydrous substance). Reference solution (c). Dilute 3.0 mL of the test solution to
It contains a variable quantity of water. 100.0 mL with a 0.5 per cent V/V solution of hydrochloric
CHARACTERS acid R. Dilute 1.0 mL of this solution to 100.0 mL with a
0.5 per cent V/V solution of hydrochloric acid R.
Appearance : white or almost white powder, hygroscopic. Column :
Solubility : freely soluble in water, very slightly soluble in – size : l = 0.15 m, Ø = 4.0 mm ;
anhydrous ethanol, practically insoluble in acetonitrile.
– stationary phase : crown-ether silica gel for chiral
It shows polymorphism (5.9). separation R (5 µm) ;
IDENTIFICATION – temperature : 10 °C.
A. Infrared absorption spectrophotometry (2.2.24). Mobile phase : perchloric acid R, methanol R, water for
Preparation : dissolve the substance to be examined in the chromatography R (0.5:5:95 V/V/V).
minimum volume of water R, evaporate to dryness at room Flow rate : 0.75 mL/min.
temperature and record the spectrum using the residue. Detection : spectrophotometer at 254 nm.
Comparison : repeat the operations using anhydrous Injection : 10 µL of the test solution and reference
valaciclovir hydrochloride CRS. solutions (a) and (c).
B. It complies with the limit for impurity R (see test A for Run time : 1.5 times the retention time of valaciclovir.
related substances). Identification of impurities : use the chromatogram
C. Water (see Tests). supplied with valaciclovir for system suitability CRS and
D. It gives reaction (a) of chlorides (2.3.1). the chromatogram obtained with reference solution (a) to
identify the peaks due to impurities A + B, C + R, D and M.
TESTS
Relative retention with reference to valaciclovir (retention
Impurities G and S. Thin-layer chromatography (2.2.27). time = about 17 min): impurities A and B = about 0.2 ;
Test solution. Dissolve 0.250 g of the substance to be examined impurities C and R = about 0.6 ; impurity D = about 0.7 ;
in 2 mL of water R and dilute to 5.0 mL with ethanol (96 per impurity M = about 1.3.
cent) R. System suitability : reference solution (a) :
Reference solution. Dissolve 5.0 mg of valaciclovir – peak-to-valley ratio : minimum 1.5, where Hp = height
impurity G CRS and 5.0 mg of valaciclovir impurity S CRS in a above the baseline of the peak due to impurity D and
mixture of 2 mL of water R and 6 mL of ethanol (96 per cent) R Hv = height above the baseline of the lowest point of
and dilute to 10.0 mL with ethanol (96 per cent) R. Dilute the curve separating this peak from the peak due to
0.5 mL of the solution to 10.0 mL with ethanol (96 per cent) R. impurities C and R.
Plate : TLC silica gel F254 plate R (2-10 μm). Limits :
Pretreatment : wash the plate with methanol R until the solvent – correction factor : for the calculation of content, multiply
front has migrated over at least 4/5 of the plate ; allow to dry the peak area of impurities A and B by 0.7 ;
in air.
– impurity R : maximum 3.0 per cent ; for the calculation,
Mobile phase : concentrated ammonia R, tetrahydrofuran R, subtract the content of impurity C as determined in
methanol R, methylene chloride R (3:12:34:54 V/V/V/V) ; use test B for related substances from the content of the
freshly prepared mobile phase. coeluting impurities C and R as determined in this test ;
Application : 4 µL. – sum of impurities A and B : maximum 2.0 per cent ;
Development : over 4/5 of the plate. – disregard limit : the area of the principal peak in the
Drying : in a current of air. chromatogram obtained with reference solution (c)
Detection : examine in ultraviolet light at 254 nm. (0.03 per cent) ; disregard any peaks due to impurities
Retardation factors : valaciclovir = about 0.3 ; other than A + B and C + R.
impurity S = about 0.7 ; impurity G = about 0.8. B. Liquid chromatography (2.2.29) : use the normalisation
System suitability : the chromatogram obtained with the procedure. Use the solutions within 24 h of preparation.
reference solution shows 2 clearly separated spots due to Solvent mixture : ethanol (96 per cent) R, water R
impurities S and G. (20:80 V/V).

Valaciclovir hydrochloride hydrate EUROPEAN PHARMACOPOEIA
Test solution. Dissolve 80 mg of the substance to be Sulfated ash (2.4.14): maximum 0.1 per cent, determined on
examined in the solvent mixture and dilute to 100.0 mL 1.0 g.
ASSAY
Reference solution (a). Dissolve 1.6 mg of valaciclovir for
system suitability CRS (containing impurities A, B, C, D, Liquid chromatography (2.2.29) as described in test A for
H, M and R) in the solvent mixture and dilute to 2.0 mL related substances with the following modification.
with the solvent mixture. Injection : test solution and reference solution (b).
Reference solution (b). Dilute 1.0 mL of the test solution to Calculate the percentage content of C13H21ClN6O4 taking
100.0 mL with the solvent mixture. Dilute 1.0 mL of this into account the assigned content of anhydrous valaciclovir
solution to 20.0 mL with the solvent mixture. hydrochloride CRS.
Reference solution (c). Dissolve 2 mg of valaciclovir
impurity P CRS in the solvent mixture and dilute to 25.0 mL STORAGE
with the solvent mixture. Dilute 1.0 mL of the solution to In an airtight container.
100.0 mL with the solvent mixture.
IMPURITIES
Column :
– size : l = 0.25 m, Ø = 4.6 mm ; Specified impurities : A, B, C, D, G, H, M, P, R, S.
– stationary phase : end-capped phenylhexylsilyl silica gel Other detectable impurities (the following substances would,
for chromatography R (5 µm) ; if present at a sufficient level, be detected by one or other of
Mobile phase : by the general monograph Substances for pharmaceutical use
– mobile phase A : trifluoroacetic acid R, water for (2034). It is therefore not necessary to identify these impurities
chromatography R (0.2:100 V/V) ; for demonstration of compliance. See also 5.10. Control of
– mobile phase B : trifluoroacetic acid R, methanol R2 impurities in substances for pharmaceutical use) : I, J, N.
(0.2:100 V/V) ;
0-5 90 10
5 - 35 90 → 60 10 → 40
A. 2-amino-1,9-dihydro-6H-purin-6-one (guanine),
35 - 45 60 40

Injection : 10 µL.
supplied with valaciclovir for system suitability CRS and
B. 2-amino-9-[(2-hydroxyethoxy)methyl]-1,9-dihydro-6H-
the chromatogram obtained with reference solution (a) to
purin-6-one (aciclovir),
identify the peaks due to impurities A, B, C, D, H and M ;
use the chromatogram obtained with reference solution (c)
to identify the peak due to impurity P.
Relative retention with reference to valaciclovir
(retention time = about 20 min): impurity A = about 0.3 ;
impurity B = about 0.4 ; impurity H = about 0.5 ;
impurity C = about 1.06 ; impurity D = about 1.2 ;
impurity M = about 1.6 ; impurity P = about 2.0. C. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
System suitability : reference solution (a) : yl)methoxy]ethyl N-methyl-L-valinate,
above the baseline of the peak due to impurity C and
Hv = height above the baseline of the lowest point of
the curve separating this peak from the peak due to
valaciclovir.
Limits :
– impurity M : maximum 0.6 per cent ; D. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
– impurity D : maximum 0.3 per cent ; yl)methoxy]ethyl N-ethyl-L-valinate,
– impurity C : maximum 0.2 per cent ;
– impurities H, P : for each impurity, maximum 0.10 per
cent ;
0.05 per cent ;
G. N,N-dimethylpyridin-4-amine,
– disregard limit : 0.6 times the area of the principal
solution (b) (0.03 per cent); disregard the peaks due to
impurities A and B.
Limit :
– total for tests A and B : maximum 4.0 per cent.
Water (2.5.12) : 4.5 per cent to 11.0 per cent, determined on H. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
0.100 g. yl)methoxy]ethyl L-alaninate,

EUROPEAN PHARMACOPOEIA Valaciclovir hydrochloride hydrate
I. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl acetate,
P. [methylenebis[azanediyl(6-oxo-1,6-dihydro-9H-purine-
2,9-diyl)methyleneoxyethan-2,1-diyl]] di-L-valinate,
J. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl L-isoleucinate,
R. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl D-valinate,
M. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl)methoxy]ethyl N-formyl-L-valinate,
N. 2-[[6-oxo-2-[[[(6-oxo-6,9-dihydro-1H-purin-2-
yl)amino]methyl]amino]-1,6-dihydro-9H-purin-9- S. 2-[(2-amino-6-oxo-1,6-dihydro-9H-purin-9-
yl]methoxy]ethyl L-valinate, yl)methoxy]ethyl N-(tert-butoxycarbonyl)-L-valinate.

EUROPEAN PHARMACOPOEIA Zidovudine
01/2017:1059 Reference solution (b). Dissolve 5 mg of zidovudine for

system suitability CRS (containing impurities A, G and H)
in reference solution (a) and dilute to 5.0 mL with reference
solution (a).
Reference solution (c). Dilute 1.0 mL of test solution (a) to
ZIDOVUDINE 100.0 mL with the solvent mixture. Dilute 1.0 mL of this
solution to 10.0 mL with the solvent mixture.
Zidovudinum Reference solution (d). Dissolve 20.0 mg of zidovudine CRS
in the solvent mixture and dilute to 20.0 mL with the
solvent mixture. Dilute 10.0 mL of the solution to 50.0 mL
Reference solution (e). Dissolve 1 mg of zidovudine
impurity D CRS in a mixture of 4 volumes of acetonitrile R,
40 volumes of methanol R and 56 volumes of a 2 g/L
solution of ammonium acetate R previously adjusted to
pH 6.8 with dilute acetic acid R and dilute to 50.0 mL with
the same mixture of solvents. Dilute 5.0 mL of the solution
to 10.0 mL with the same mixture of solvents.
C10H13N5O4 Mr 267.2
[30516-87-1] Column :
– size : l = 0.25 m, Ø = 4.6 mm ;
DEFINITION – stationary phase : base-deactivated end-capped
1-(3-Azido-2,3-dideoxy-β-D-erythro-pentofuranosyl)-5- octadecylsilyl silica gel for chromatography R (5 μm).
methylpyrimidine-2,4(1H,3H)-dione. Mobile phase :
Content : 97.0 per cent to 102.0 per cent (dried substance). – mobile phase A : 2 g/L solution of ammonium acetate R
CHARACTERS previously adjusted to pH 6.8 with dilute acetic acid R ;
Appearance : white or slightly brownish powder. – mobile phase B : acetonitrile R ;
Solubility : sparingly soluble in water, soluble in anhydrous Time Mobile phase A Mobile phase B
ethanol, practically insoluble in heptane. (min) (per cent V/V) (per cent V/V)
0-3 95 5
It shows polymorphism (5.9). 3 - 18 95 → 85 5 → 15
IDENTIFICATION 18 - 28 85 → 30 15 → 70
A. Specific optical rotation (see Tests). 28 - 43 30 70
B. Infrared absorption spectrophotometry (2.2.24).
Comparison : zidovudine CRS.
If the spectra obtained in the solid state show differences,
dissolve the substance to be examined and the reference Injection : 20 µL of test solution (a) and reference
substance separately in the minimum volume of water R, solutions (b), (c) and (e).
evaporate to dryness in a desiccator, under high vacuum over Identification of impurities : use the chromatogram
diphosphorus pentoxide R and record new spectra using the supplied with zidovudine for system suitability CRS and
residues. the chromatogram obtained with reference solution (b) to
identify the peaks due to impurities A, B, C, G and H ; use
TESTS the chromatogram obtained with reference solution (e) to
Appearance of solution. The solution is not more opalescent identify the peak due to impurity D.
than reference suspension I (2.2.1) and not more intensely Relative retention with reference to zidovudine
coloured than reference solution BY5 (2.2.2, Method II). (retention time = about 16 min): impurity C = about 0.2 ;
Dissolve 0.5 g in 50 mL of water R, heating if necessary. impurity A = about 0.5 ; impurity H = about 0.95 ;
impurity B = about 1.05 ; impurity G = about 1.5 ;
Specific optical rotation (2.2.7) : + 60.5 to + 63.0 (dried
impurity D = about 2.0.
substance), measured at 25 °C.
Dissolve 0.50 g in anhydrous ethanol R and dilute to 50.0 mL
with the same solvent. – resolution : minimum 2.0 between the peaks due to
impurity H and zidovudine ; minimum 2.0 between the
Related substances peaks due to zidovudine and impurity B.
A. Liquid chromatography (2.2.29). Calculation of percentage contents :
Solvent mixture. Mix 4 volumes of acetonitrile R, – correction factor : multiply the peak area of impurity C
20 volumes of methanol R and 76 volumes of a 2 g/L by 0.6 ;
solution of ammonium acetate R previously adjusted to
pH 6.8 with dilute acetic acid R. – for each impurity, use the concentration of zidovudine
in reference solution (c).
Test solution (a). Dissolve 20.0 mg of the substance to be
examined in the solvent mixture and dilute to 20.0 mL with Limits :
the solvent mixture. – impurity G : maximum 0.5 per cent ;
Test solution (b). Dilute 10.0 mL of test solution (a) to – impurities A and C : for each impurity, maximum
50.0 mL with the solvent mixture. 0.15 per cent ;
Reference solution (a). Dissolve 2 mg of thymine R – unspecified impurities : for each impurity, maximum
(impurity C) and 2 mg of zidovudine impurity B CRS in 0.10 per cent ;
the solvent mixture and dilute to 50.0 mL with the solvent – reporting threshold : 0.05 per cent ; disregard any peak
mixture. Dilute 1.0 mL of the solution to 20.0 mL with the due to impurity D and any peak eluted after this
solvent mixture. impurity.

Zidovudine EUROPEAN PHARMACOPOEIA
B. Liquid chromatography (2.2.29). Other detectable impurities (the following substances would,
Test solution. Dissolve 0.5 g of the substance to be examined if present at a sufficient level, be detected by one or other of
in 10 mL of acetonitrile R1 and dilute to 100.0 mL with the the tests in the monograph. They are limited by the general
mobile phase. acceptance criterion for other/unspecified impurities and/or
Reference solution (a). Dissolve 5.0 mg of zidovudine use (2034). It is therefore not necessary to identify these
impurity D CRS in acetonitrile R1 and dilute to 10.0 mL impurities for demonstration of compliance. See also 5.10.
with the same solvent. Control of impurities in substances for pharmaceutical use) :
Reference solution (b). Dilute 1.0 mL of reference B, D, E, F, H, J, K.
solution (a) to 100.0 mL with the mobile phase.
Reference solution (c). Dilute 1.0 mL of reference
solution (a) to 50.0 mL with the test solution.
Column :
– size : l = 0.15 m, Ø = 4.6 mm ;
– stationary phase : base-deactivated end-capped
octadecylsilyl silica gel for chromatography R (5 μm).
A. 1-[(2R,5S)-5-(hydroxymethyl)-2,5-dihydrofuran-2-yl]-5-
Mobile phase : water for chromatography R, acetonitrile R1
methylpyrimidine-2,4(1H,3H)-dione,
(30:70 V/V).
Injection : 20 µL of the test solution and reference
solutions (b) and (c).
Run time : 10 times the retention time of zidovudine.
Identification of impurities : use the chromatogram obtained
with reference solution (b) to identify the peak due to
impurity D.
B. 1-(3-chloro-2,3-dideoxy-β-D-erythro-pentofuranosyl)-5-
Relative retention with reference to zidovudine (retention methylpyrimidine-2,4(1H,3H)-dione,
time = about 1.5 min) : impurity D = about 2.5.
zidovudine and impurity D.
– for each impurity, use the concentration of impurity D
in reference solution (b). C. 5-methylpyrimidine-2,4(1H,3H)-dione (thymine),
Limits :
0.10 per cent ;
– reporting threshold : 0.05 per cent ; disregard any peak
eluted before impurity D.
Limit :
– total for tests A and B : maximum 1.0 per cent. D. triphenylmethanol,
on 1.000 g by drying in an oven at 105 °C.
1.0 g.
ASSAY
Liquid chromatography (2.2.29) as described in test A for
Injection : test solution (b) and reference solution (d). E. 1-(2-deoxy-β-D-erythro-pentofuranosyl)-5-
Calculate the percentage content of C10H13N5O4 taking into methylpyrimidine-2,4(1H,3H)-dione (thymidine),
account the assigned content of zidovudine CRS.
STORAGE
IMPURITIES
Test A for related substances : A, B, C, E, F, G, H.
Test B for related substances : D, J, K. F. 1-(2-deoxy-β-D-threo-pentofuranosyl)-5-
Specified impurities : A, C, G. methylpyrimidine-2,4(1H,3H)-dione,

EUROPEAN PHARMACOPOEIA Zidovudine
J. 1-[3-azido-2,3-dideoxy-5-O-(triphenylmethyl)-β-D-
erythro-pentofuranosyl]-5-methylpyrimidine-2,4(1H,3H)-
dione (trityl zidovudine),
G. 1-[(2R,4S,5S)-4-azido-5-(hydroxymethyl)tetrahydrofuran-
2-yl]-3-[(2S,3S,5R)-2-(hydroxymethyl)-5-(5-methyl-2,4-
dioxo-3,4-dihydropyrimidin-1(2H)-yl)tetrahydrofuran-3-
yl]-5-methylpyrimidine-2,4(1H,3H)-dione,
K. 1,1′,1′′-(methoxymethanetriyl)tribenzene (methyl trityl
H. unknown structure, ether).


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EUROPEAN

Published in accordance with the

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© Council of Europe, 67075 Strasbourg Cedex, France - 2020

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Activity A becquerel Bq s− 1 1 Ci = 37·109 Bq = 37·109 s− 1

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REAGENTS Using the 5 standard solutions, prepare the following reference

Red solution. Dissolve 60 g of cobalt chloride R in about B9 1.0 99.0

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Table 2.2.2.-5. - Reference solutions GY Storage

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GY4 5.0 95.0

GY5 3.0 97.0

GY6 1.5 98.5

GY7 0.75 99.25

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07/2016:20203 Management of electrodes. The electrodes are stored

20 1.68 3.79 4.00 6.88 7.43 9.23 10.06 12.63

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30 1.68 3.55 3.77 4.02 6.85 7.40 9.14 9.97 12.29

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