UNIVERSITY OF KABIANGA

SCHOOL OF SCIENCE AND TECHNOLOGY

DEPARTMENT OF BIOLOGICAL SCIENCES
BSC.BIOCHEMISTRY

FIELD ATTACHMENT REPORT SUBMITTED TO THE DEPARTMENT OF BIOLOGICAL

SCIENCES IN PARTIAL FULFILLMENT OF BACHELOR OF SCIENCE IN
BIOCHEMISTRY.INDUTRIAL ATTACHMENT AT KENYA BUREAU OF
STANDARDS COAST REGION.

PERIOD:
11 WEEKS FROM 16 APRIL TO 16 MAY AND FROM 16TH
th

JULY TO 24th AUGUST 2018

PRESENTED BY:
NAME: EVANS MAUTA ELCANAH
REG NO: BIO/006/16

DEPARTMENT: TESTING DEPARTMENT

CHEMISTRY LABORATORY

PRESENTED TO:
MR.CYPRIAN OMBATI

DATE: 15TH/OCTOBER/2018

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 DECLARATION
This project is my own original work and was not presented for judging in any institution.

Name of student………………………………………………………………..
Student Reg No: ........................... Signature: …………………………..........
Attachment Institution: ......................................................................................

Name of Industrial Supervisor..............................................................................

Signature: .............................................. Date.....................................................

Name of University Supervisor.............................................................................

Signature: ............................................... Date......................................................

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 ATTACHMENT DETAILS
INSTITUTION NAME: KENYA BUREAU OF STANDARDS
BRANCH: COAST REGION
DEPARTMENT: TESTING DEPARTMENT
LABORATORY: CHEMISTRY LAB

ATTACHEES’ RESPONSIBILITIES
(1). Perform any duty assigned by the supervisor or any other member of the staff.
(2). Sample housekeeping every morning, i.e.
a) Removing opened samples from sample arrival unit and transferring them to the unit for
sample undergoing analysis.
b) Transferring samples from reception desk to sample waiting analytical units,
c) Moving already analyzed samples to sample disposal units.
(3).Maintaining the working benches clean including Sample removal after analyses.
(4).Assist in packaging of samples that are being moved to other labs, including samples being
Moved to Nairobi KEBS HEAD OFFICE for more analyses.
(5).Serve as a messenger within the KEBS apartments when called upon to deliver official
Communications.

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 ACKNOWLEDGEMENT
I take this chance to thank the almighty God for enabling me secure this chance for industrial
attachment which provides meaningful “hands-on” training skills to supplement the theoretical
classwork and introduces me practical concepts. This golden opportunity provides diversified
experiences to students as much as possible not only in field of Biochemistry but also in wide
range of related fields.
My sincere gratitude goes to MR. CYPRIAN OMBATI, industrial attachment coordinator in the
department of Biological sciences, who on the behalf of the department assisted us in sourcing
for students’ placement from suitable industries, handled correspondence and students’
challenges concerning industrial attachment, organized supervision of students’ on attachment
and finally coordinated evaluation of students’ including reception and keeping of evaluation
materials. Feel much appreciated sir.
I also register my thanks to my industrial attachment supervisor, MR.TED OMOLLO for being
part and parcel in ensuring that I settled well and I was receiving the appropriate training. I also
wish to thank the entire analytical chemistry laboratory staff for assisting me learn as much as
possible in fields encompassing analytical skills, and team works.
Regards!

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 TABLE OF CONTENTS
CHAPTER ONE: INTRODUCTION
1.1 About BIO 320 (IDUSTRIAL ATTACHMENT).............................................................page 1
1.2 Abstract............................................................................................................................page 2
1.3 History of Kenya Bureau of Standards.............................................................................page 3
1.4 Main functions and core activities of KEBS.....................................................................page 4
1.5 Vision and mission statement of KEBS............................................................................page 5
1.6 Organizational structure of KEBS....................................................................................Page 6

CHAPTER TWO: KEBS TESTING DEPARTMENT

2.1The key functions and activities of Testing Department...................................................page 8
2.3 Specific activities in the field of Biochemistry...............................................................page 10

CHAPTER THREE: EVALUATION OF THE ATTACHMENT PERIOD

3.2 Challenges encountered during the attachment period.................................................page 14
3.3 How the challenges were overcome/solved....................................................................page 15
3.4 Conclusions and recommendations as to how the attachment exercise can be improved by
University of Kabianga..................................................................................................page 16

CHAPTER FOUR: BIBLIOGRAPHY AND REFERENCES

4.1 References.......................................................................................................................page 17
4.2 Appendices......................................................................................................................page 18
1.1 ABOUT BIO 320 (INDUTRIAL ATTACHMENT)
The aim of BIO 320 (industrial attachment) is to expose students to applications of Biochemistry
in research and industry. Third year students from the field of Biochemistry undertakes an eight
(8) week supervised industrial placement in selected work environments in the field of
Biochemistry or related areas. They will be required to participate in the technical activities and
operations of the institution/industry. Besides, they are expected to prepare an industrial
attachment/technical reports and keep logbooks of their activities at the station. The logbooks
detailing daily activities would therefore be submitted for grading.

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Industrial attachment is an attempt to expose students to the workings of industry and thus enable
them to:
 Familiarize themselves with industrial organization.
 Practice and enrich their technical skills in the field of Biochemistry.
 Appreciate the linkage between theory and practice.
 Develop desirable attitudes and work Habits in the field of Biochemistry.
 Familiarize themselves with technical problems with a view of developing techniques for
solving them.
 Appreciate the role played by the industry in human health, food security, environmental
management and other social economic activities.
To comply with the above mentioned requirement, I was attached in the Kenya Bureau of
Standards Coast Region for a period of nine (9) weeks. During this period, I achieved excellence
not only in the field of Biochemistry but also in analytical chemistry, most important being
nourished on how to promote Biochemistry through research, industrial teaching, consultancy
and community service.

1.2 ABSTRUCT
During the first week of my industrial attachment in KEBS, I was introduced to wines and spirits
analysis. The analytical parameters included ethyl alcohol, total acidity, volatile acidity, total
dissolved solids and labeling. Cereals (milled rice, shelled rice and beans etc) analysis for
parameters such as moisture content, grading, aflatoxins and total aflatoxins was done .Besides, I
was also shown the analysis of table sugars in parametric encompasses of moisture content,
Sulphur (iv) Oxide, invert sugar, matter insoluble in water, labeling and foreign matter content.
In the second week, I was shown how to analyze drinking water samples for total dissolved
solids, suspended matter, sulphates PH, conductivity, chlorides, alkalinity and fluorides. In

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addition, I was introduced to fats and oils and shown how to get acid value, free fatty acids,
volatile matter @ 105oC, refractive index, relative density, peroxide value, iodine value and
insoluble impurities.
During the third week, I was trained how to analyze table salt samples for parameters such as
moisture content, iodine value, salt alkalinity, sulphates, matter insoluble in water, chlorides,
magnesium, calcium and acid insoluble matter, carbonated beverages for PH and degree of Brix
content and finally tomato products (tomato source, ketchup, chili source) for PH value, total
solids, acidity and salt content. During the fourth week, I was introduced to yoghurt tests for
acidity, glucose energy drink for PH value, total solids and total ash .Cheese tests for moisture
content, color, texture, dry matter content and appearance. Finally, butter parametric analysis for
tests such as pH value, water content and salt content. In the fifth week, the tests were for jams
and jelly where I was shown the analysis of all physical tests, degree of Brix, acidity and PH,
pastries for total solid content, moisture content, total ash, PH of the aqueous extract, fat content
and fiber, food seasoning mixtures and spices for tests such as salt content, moisture content and
ash content. For washing bar soaps, the tests included grittiness, mush and alcohol insoluble
matter. Finally, parametric analysis for pasta products to test moisture content, total ash, cooking
test and fiber content.
In the sixth week, I was guided through laundry soaps and bar soaps analysis to test for total fatty
matter, matter insoluble in ethanol, free caustic alkali and grittiness test. For crisps (potato,
cassava and banana) the tests included taste & flavor, salt content and moisture content. Finally,
roasted ground nuts and soya beans were analyzed to determine the moisture content, defects,
foreign matter and salt content. In the seventh week, the analysis was targeted to canned meat for
tests such as moisture content, total ash, salt content and Sulphur (IV) Oxide. For canned fruits &
food stuff, tests were based on all physical parameters .The week ended with the analysis of
black tea and roasted ground coffee samples for tests such as water insoluble matter, alkalinity of
water soluble ash, total ash, water extract and moisture content. In the eighth week, I was taken
through bleaching agent/powder analysis to analyze available chlorine, free alkali and stability.
For detergents, I was shown how to test parameters such as total solids, stability, active
detergent, foam, odor, residual test, rinsing properties and marking. In my last week, I was
trained how to analyze beer to test ethanol content, volume of carbonation and original wort
extract. For honey analysis, the tests were targeted to insoluble matter, Fischer’s test and
reducing sugar content.

INTRODUCTION
1.3 History of Kenya Bureau of Standards.
Kenya Bureau of Standards (KEBS) is a statutory body established under the Standards Act (CAP
496) of the laws of Kenya. KEBS commenced its operations in July 1974.
The KEBS Board of Directors is known as the National Standards Council (NSC). It is the policy-
making body for supervising and controlling the administration and financial management of the

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Bureau. The Managing Director is the Chief Executive responsible for the day-to-day
administration of the Bureau within the broad guidelines formulated by the NSC.
The following chronological events led to establishment of The Kenya Bureau of Standards.
YEAR ACTIVITY
1974 KEBS was founded by the enhancement of the CAP 496 of the laws of Kenya.
1980 KEBS was moved its headquarters’ to South c (NAIROBI).
1981 Metrology Department was initiated.
1985 Testing Department was initiated.
KEBS COAST REGION was opened.
1995 KEBS KISUMU REGION was opened.
Legal inspection was commissioned under legal notice no.78.
2003 KEBS developed its first strategic plan.
2005 KEBS signs performance contract with the government.
PRE- EXPORT VERIFICATION OF CONFORMITY was initiated.

KEBS MANDATE
KEBS is mandated to provide standardization and conformity assessment services through:
 Promotion of standardization in commerce and industry.
 Provision of testing and calibration facilities.
 Product and system certification
 Undertaking an educational work in standardization and practical applications of
standards.
 Maintenance and dissemination of International System of Units (SI) of measurements.

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MAIN FUNCTIONS AND CORE ACTIVITIES OF KEBS.
1.4.1: STANDARDS DEVELOPMENT
Standards are documents that provide a common reference point for the assessment of the quality
of goods and services. They provide the starting point for fulfilling the KEBS mandate for
standardization and conformity of the assessment services. KEBS acts as the secretariat for
technical committees that develop standards in several sectors in food, textile and leather,
agriculture, electro-technical, chemical, mechanical, civil engineering, ICT and service sectors
such as tourism, education and financial services.
BENEFITS OF USE OF STANDARDS
a) Assure consumers that products are safe, reliable and of good quality.
b) Facilitate innovations, technology transfer and market access, provide transparency in the
exchange of products to enhance market access of the Kenyan products into local,
regional and international markets thus opening up export markets for products and
services.
c) Provide best practices, performance and safety reference points for regulators, researchers
and industry.
d) Help organizations meet their consumers’ needs while optimizing on company processes.
e) Drive efficiency in your business in your business operations and reduce waste.
f) Reduce technical barriers to trade.
g) Allow for a level playing field for all enterprises.
h) Improved credibility and consumer confidence for products and services delivered.
1.4.2: NATIONAL CODEX CONTACT POINT-KENYA.
Kenya is a member of an international codex alimentarious commission body (CAC) which is
the international body for developing food standards. KEBS is the National Codex Contact Point
for Kenya that links and coordinates Codex Alimentarious Commission activities to the country
through the National Codex committees. The CAC mandate is to ensure that food standards
developed are science- based, protect the health of consumer and provide fair food trade. CAC
works together with Codex three Expert advisors committees (JECFA, JMPR and JEMRA) on
food additives, contaminants, pesticide residues, veterinary residues in food to safe foods. The
National Codex committees comprises of 23 CAC mirror national committees from research
centers, universities, regulatory/supervisory bodies, food industry, food production, processing
and consumer protection associations, and professionals involved in food production, processing
and analysis.
1.4.3: QUALITY ASSURANCE AND INSPECTION
KEBS operates a product certification scheme in line with section 10 of the standards Act,
Cap.496 of the Kenyan constitution. This scheme leads to awards of permits to use product
certification marks of quality. This ensures health and safety of Kenyans as well as protecting the
environment during the exchange of goods and services .The aim of this scheme is to provide
evidence and regulators thereby facilitating trade and market access for Kenyan products.

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FOOD FORTIFICATION
Food fortification is the addition of one or more vitamins and/or mineral to a food to correct or
prevent a demonstrated micronutrient deficiency .It is one of the four key cost effective strategies
recommended by WHO in curbing micronutrient deficiency. The ministry of public health and
sanitation through a legal notice no.62 of 15th June 2012 has therefore declared it mandatory to
fortify the following food products.
 Wheat flour- zinc and iron.
 Dry milled maize products- B vitamins, iron, folic acid and zinc.
 Salt-iodine
 Vegetable fats and oils-vitamin A.
 Sugar-vitamin A.
1.4.4: ENFORCEMENT AND MONITORING (SURVEILLANCE).
KEBS monitors certified products through regular factory and market surveillance visits. During
these visits, KEBS officers evaluate compliance to the scheme of supervision of control (SSC)
signed between KEBS and the manufacturer as well as drawing samples for testing to determine
compliance to the relevant Kenya Standard or approved specification. Permit holders found to
have contravened the contract and/or offering for sale products not meeting the requirements of
the relevant Kenya Standards or approved specification may have their permits
suspended/withdrawn/cancelled.
1.4.5: PRE-EXPORT VERIFICATION OF CONFORMITY (PVoC).
KEBS offers pre-export conformity verification of conformity (PVoC) to standards at the
country of origin, so as to ensure that only quality goods are shipped into the Kenyan market .All
products imported into Kenya are required to meet the requirements of Kenya Standards or other
approved standards. To ensure worldwide coverage, KEBS has appointed agents to carry out
conformity assessment of regulated goods at the country of supply thus ensuring that sub-
standard products are stopped at the source. Consignments that are found to comply with the
requirements are issued with a certificate of conformity.
KEBS undertakes destination inspection on imports at all ports of entry. Goods accompanied by
certificate of conformity (COC) are permitted into the country without further testing. However,
goods not accompanied with COC are inspected on arrival at the point of entry after the
concerned importer has paid a penalty equivalent to 15% of the CIF value to KEBS. Such goods
will only be allowed into the country after satisfactory inspection and testing has been carried
out. KEBS also undertakes inspection of used motor vehicles to ensure that only vehicles that
comply with KS1515:2000 standard are allowed into the country. The key elements of the
standards are:
 Maximum of 8 years from the year of first registration.
 The vehicle must be right hand drive.
 The vehicle must have roadworthiness certificate issued by KEBS appointed Inspection
Company in the country of origin.

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INSURANCE OF ORDINARY CERTIFICATE OF ORIGIN
KEBS issues Ordinary Certificate of Origin to ensure traceability of exported products to the
country of origin (Kenya).It enables exporters to comply with the WTO rules of origin.
However, KEBS mandate is limited to selected export destinations, since the bulk of certificate
of origin for Kenya exports is issued by the Kenya Revenue Authority.

1.4.7: METROLOGY
Metrology (calibration of equipment) is the science of measurement and its application. KEBS
provides accurate and reliable measurements for use in trade. KEBS is the custodian of national
measurement standards and also provides the national measurement traceability to the
international system of units (SI) .Calibration services are offered in various fields of
measurements. Some of the sectors served
are:healthcare,communication,telecommunication,navigation,aviation,transport,construction,man
ufacturing,reseach and development, testing, education, agriculture, safety and ICT. Examples of
equipment calibrated are: weighing scales/balances, incubators, clinical thermometers, blood
pressure machines, acoustic/ultrasonic devices and X-ray machines. KEBS metrology
laboratories are accredited by the German Accreditation Service (DAKKS).The quality
management system has also been assessed and approved by the AFRIMETS.KEBS metrology
also provides measurement solutions to industry, specialized training and consultancy.

1.4.8: TESTING SERVICES

KEBS accredited laboratories test both locally manufactured and imported goods for compliance
with the Kenya standards or any other standards approved by the Kenya Bureau of Standards.
This provides quality assurance for goods traded in the country. KEBS testing laboratories are
well equipped with modern test facilities and have trained and experienced technical personnel
who carry out tests of raw materials, semi-finished and finished products. The products tested
include: food and agricultural products, electrical and electronic products, mechanical
engineering products, building and construction materials, textile products-fibres, yarn and
fabrics, organic products-petroleum, soaps, spirits and cosmetics, residues and other food and
environmental contaminants, plastics, paints, paper and rubber. The testing laboratories are
accredited by United Kingdom Accreditation Services (UKAS).KEBS strategic objective is to
transform its laboratories from offering routine testing services to become a Reference laboratory
that will provide traceability of measurements to other testing laboratories.

1.4.9: RESEACH AND DEVELOPMENT

The research and Development (R&D) department coordinates activities in standardization and
conformity assessment as well as improving the quality and comparability of chemical analytical
methods (testing method methods development and validation).R&D departments also networks

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and collaborates with academia, research institutions, and established National Metrology
Institute (NMIs) for capacity building. This enables the researchers and industry to harness
innovation and technological development in analytical fields with traceable measurement
techniques.

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 CHAPTER TWO: KEBS TESTING DEPARTMENT
The KEBS testing services department was established in 1981 with an aim of providing laboratory
facilities meant for examination and testing of commodities/materials whether in raw form, semi-
processed, or fully processed. The Testing services departments is one of the departments in
metrology, Testing and market surveillance division within the Kenya Bureau of Standards. The
department supports the KEBS mission by serving as the national reference laboratory for
measurements/testing in the chemical, biological and engineering/material sciences outlined in the
standards Act. The activities cover the following areas:
a) Conformity assessment based measurements (testing) for the composition, structure and
physical properties of various products in diverse such as chemical, food, industrial,
biological, environment, engineering and process raw materials.
b) The development and dissemination of reference test procedures and reference materials
c) Provision of proficiency testing and inter-laboratory comparison schemes.
d) Training consultancy and development of best practice guides that help assure
measurements quality.
e) Provision of contract testing services tailor made support specific client objectives.
The testing services department implements a quality management system based on ISO 17025
(General requirements for the competence of testing and calibration laboratories); all laboratories
in the department are accredited to ISO17025 and all laboratory operations implemented in line
with the requirements of the standards. The testing services department is structured (Provide a
link to laboratory structure) into three broad thematic areas namely engineering, chemistry biology
and the Sample Control Centre. Each field served by different specialized laboratories focused on
different product categories (Provide a link to the laboratory scope). The Laboratories are managed
by a highly trained and technically competent personnel supported by a team of skilled and
experienced technical staff. The technical staff comprise of university graduates (at undergraduate
and postgraduate levels) and technicians who hold national and higher national diplomas. The
department serves as a focal point for chemical metrology in the country and represents KEBS in
the Consultative Committee for Amount of Substance (CCQM) at the BIPM.

The primary products and services include:

 Sampling, testing and analysis (inside the lab).

 Performance Testing and validation of new products.
 Environmental monitoring (at client's premises).
 Proficiency testing.
 Reference materials.
 Training, research, advisory and consultancy services in testing, laboratory quality
management systems, quality controls etc.

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TESTING LABORATORIES.

1. Polymer Laboratory

Polymers laboratory is one testing laboratories in the Testing Services Department (TES). The role
of the laboratory is to carry out analysis on chemical and physical properties of material products
for the purpose of implementation of Kenya Standards by Quality Assurance, Import Inspection
and for Standards Development. Analysis is also carried out on request for external customers.
The laboratory analyses over 3000 samples annually. The laboratory also participates in research
and development work that deals with polymeric products .It also provides expertise in local,
regional and international technical committees. The laboratory also participates in proficiency
testing schemes; some of the PT schemes are in paper, leather and rubber products.
2. Organic Chemistry Laboratory.
Organic chemistry laboratory carries out analysis of consumer products whose ingredients are
predominantly organic compounds. These include cosmetic products, soaps and detergents,
petroleum products, alcoholic beverages, tests for air quality monitoring like car exhaust and flue
gas testing, determination of environmental pollutants like pesticides, food additives like
polycyclic aromatic hydrocarbons (PAHs), preservatives like benzoic acid, nutritional ingredients
like ascorbic acid (vitamin C) and caffeine in food products. The laboratory currently include
methods of test accredited by South Africa National Accreditation Services (SANAS). The scope
of accreditation includes:
a. Determination of Total Fatty Matter (TFM) in soaps.
b. Determination of matter insoluble in ethanol (MIE) in soaps.
c. Determination of free caustic alkali (FCA) in soaps.
d. Determination of total dissolved solids (TDS) in alcoholic beverages.
e. Determination of ethanol content in alcoholic beverages.
The laboratory participates in inter laboratory comparison comparisons (ILCs) and proficiency
testing (PT) schemes offered by international reputable and accredited PT provider like FAPAS,
ARA, NMISA, and BIPM among others. Participation in PT schemes is aimed at benchmarking
the laboratory, both locally and internationally and also a tool for quality (QC).
3. Molecular Biology Laboratory
The Molecular Biology Laboratory at KEBS is dedicated to pursuing proteomics and nucleic acid
analysis using the most resent techniques. The laboratory is well equipped with modern analytical
machinery that effectively and efficiently performs molecular analysis of proteins, DNA and RNA
in food, feeds and other organically derived products. Using the Mag MAX technology we have
developed a robust high throughput platform for nucleic acid extraction. Currently the laboratory
is able to perform the following tests.

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 a. Qualitative screening of GMO derived products using veriti® 96-well fast Thermal cycler
to perform the endpoint qualitative PCR analysis of GMO derived products.
b. Quantitative screening of GMO derived products using 7500 fast real time PCR system to
quantitatively measure the amplification of DNA using fluorescent dyes through a process
known as real time polymerase chain reaction.
c. Meat specification using gene specific oligonucleotide makers in the analysis of meat and
meat products derived from Gallus, porcine, Bovine and ovine among many other species.
d. Molecular diagnoses of horticultural plant pathogens using PCR based methods for early
detection of bacterial, viral and fungal infections in horticultural crops including flowers.
e. DNA fingerprinting of rice using simple sequence repeats (SSRs) microsatellites.
4. Microbiology Laboratory
The Microbiology laboratory has been in operation since 1985 and is domiciled under the Testing
services department of Kenya Bureau of Standards (KEBS). Testing laboratories offer a
comprehensive range of microbiological measurement solutions covering: food products, water
samples, environmental monitoring samples, cosmetics, hygiene monitoring samples and
antimicrobial agents. The laboratory uses validated methods to isolate, enumerate and identify
microorganisms.
5. Mechanical Engineering laboratory
This is one of the material engineering labs in the testing department. The lab is tasked with
carrying out tests to determine mechanical and physical properties of materials. The lab comprises
of two subdivisions namely destructive and non-destructive (NDT) labs. The destructive lab carries
out tests on construction materials, automotive parts, farm tools and household goods. Just as the
name suggests, once items have been tested, they cannot be reused since the tests are destructive.
On the other hand NDT involves testing products, parts or materials without destroying them hence
not affecting their future usefulness. The tests done on these products are either performance,
safety or conformity to various requirements as per local and international standards.
6. Inorganic Laboratory
Inorganic laboratory is one of the chemistry laboratories in testing department. The laboratory has
three sections: wet chemistry, electrochemical and spectrometry. The wet chemistry section uses
analytical methods such as colorimetry, titrimetry and gravimetry to analyse samples. Analytical
tests are performed after careful sample preparation, weighing and mixing of reagents for both
qualitative and quantitative measures. Electrochemical section deals with analysis of anions in
aqueous solution of samples using potentiometric techniques. Spectrometry uses a wide array of
techniques for analysis such as atomic absorption, atomic emission, atomic fluorescence and mass
spectrometry.

7. Electrical Testing Laboratory

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Electrical engineering lab in the testing department at Kenya Bureau of standards serves a broad
category of industries like Energy, Power, Metering and distribution, Telecommunication,
automotive and manufacturing sectors among others. It provides competent and efficient services
in conformity testing and assessment offering safety and performance tests of a full range of
household electrical appliances as well as industrial appliances. Safety tests are done to verify that
the user is not at risk in using the electrical device, while performance tests are carried out to verify
the appliance specifications.

KEBS VISION
To be a global leader in standards based solutions that deliver quality and confidence.
KEBS MISSION
To provide standards based solutions that provide innovation, trade and quality life.

KEBS ORGANIZATIONAL STRUCTURE:

COAST REGION TESTING SERVICES QUALITY MANAGEMENT SYSTEM

Region manager Chief Manager Manager

QUALITY ASSUARANCE TESTING SERVICES INSPECTION SERVICES

SERVICES Manager in charge Manager
Manager

CHEMISTRY LABORATORY MICROBIOLOGY SAMPLE CENTER

Head of laboratory LABORATORY Head of sample control center

LABOLATORY ANALYSTS LABORATORY ANALYSTS SAMPLE CONTROL

Laboratory technicians Laboratory technicians CUSTOMER CARE
Sample control staff

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ANALYTICAL TECHNIQUES.
During my industrial attachment at KEBS, I came across the following analytical techniques:
1. Gravimetric.
2. Physical analytical techniques.
3. Extraction/separation methods.
4. Potentiometric methods.
5. Analytical ash based techniques.
6. Titrimetric techniques.
7. Measurement techniques.
8. Biochemical techniques.

GRAVIMETRIC TECHNIQUES.
Gravimetry encompasses all techniques in which we measure mass or a change in mass.
Measuring mass is the most fundamental of all analytical measurements, and gravimetry is
unquestionably the oldest analytical technique. Test ways to use mass as an analytical signal, one
can measure an analyte’s mass directly by placing it on a balance and recording its mass. For
example, suppose you are to determine the total suspended solids in water released from a
sewage-treatment facility. Suspended solids is solid matter that has yet to settle out of its solution
matrix. You collect a sample and pass it through a preweighed filter that retains the suspended
solids. After drying to remove any residual moisture, you weigh the filter. The difference
between the filter’s original mass and final mass gives the mass of suspended solids.

Sometimes it is easier to remove the analyte and use a change in mass as the analytical signal.
Imagine how you would determine a food’s moisture content by a direct analysis. One possibility
is to heat a sample of the food to a temperature at which the water in the sample vaporizes. If we
capture the vapor in a preweighed absorbent trap, then the change in the absorbent’s mass
provides a direct determination of the amount of water in the sample. An easier approach,
however, is to weigh the sample of food before and after heating, using the change in its mass as
an indication of the amount of water originally present. Example of gravimetric techniques I
came across include:

(a).Moisture content in food samples.

This includes the moisture content in bread, cakes, sugar, salt, rice among other samples.

(i). moisture content analysis for bread samples.

Weigh 10 grams of the sample into an empty pre-weighed glass petri-dish. Incubate the sample
in an oven @ 120oC for two hours. Cool the samples in a desiccator to prevent moisture gain.
Weigh the samples in the analytical balance and calculate change in mass.

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(b). Matter volatile analysis.
Weigh 10 grams of the sample into an empty pre-weighed glass petri-dish. Incubate the sample
for one hour in an oven set @ 120oC. Cool the samples in a desiccator and weigh the samples to
get the mass difference.

(c).Sulphates analysis in water samples.

Measure 100ml of water sample into a clean conditioned conical flask. Add 2 drops of methyl
orange indicator. Add 5ml of 1:1 HCL: WATER, cover with aluminum foil and boil for 20
minutes. Add 10ml of Barium chloride whole boiling for another 20 minutes. While still
covered, cool overnight. Filter the solution through a pre-weighed no 3 grouch crucible. Dry the
crucible in an oven for 30 minutes and take the final crucible weight .Compute for the weight
increment margin.

(d). Bar soap mush test.

Measure 7.5cm long and 4.0cm width of the sample and wet the sample on one side via running
tap water. Cover the sample with a wet towel and enclose it in a vessel for six hours. Scrub
wetted side which was in contact with wet towel using a preweighed 30cm ruler and weigh the
ruler for weight difference to determine the mush content in the soap.

(e). Cook test.

Weigh 25 grams of the sample under analysis in a beaker. Transfer the weighed sample into a
glass beaker with 200 ml of boiling distilled water. Cook the sample at the cooking time duration
indicated on the label and add an extra of 2 minutes. Cool the beaker and extract 25 ml of the
gruel with a pipette into a preweighed Petri dish. Transfer in an oven for vaporization. Cool and
weigh the petri dish for the weight gain determination.

(f). Matter insoluble in ethanol

Weigh 0.5 grams of the sample into a glass beaker. Add 100 ml of ethanol and shake to dissolve
the sample. Warm gently if the sample does not dissolve completely. Filter the solution through a
preweighed number 4 gouch crucible. Condition the crucible in an oven for 30 minutes, cool and
take the final weight. Compute for the weight gain.

(g). Precipitation Gravimetry

Precipitation gravimetry is based on the formation of an insoluble compound following the
addition of a precipitating reagent, or precipitant, to a solution of the analyte. In most methods
the precipitate is the product of a simple metathesis reaction between the analyte and precipitant;
however, any reaction generating a precipitate can potentially serve as a gravimetric method.

(i).Free caustic alkali determination in soap samples.

Weigh 5 grams of the sample under analysis into a 250 ml conical flask. Add 100 ml of ethanol
and warm to dissolve. Add 3 drops of phenolphthalein indicator as it warms then 10% Bacl2
solution in excess. If pink color persists, titrate against 0.1N HCL.

(j). Determination of crude fibre content in cereal product and animal feeds.
Crude fibre content includes the insoluble and combustible organic residue including structural
polysaccharides such as cellulose, hemicelluloses, pectines and lignin which remains after

18
sequential treatment with acetone, boiling Sulphuric acid and dilute sodium hydroxide. After de-
fatting, boiling with Sulphuric acid (1.25% m/v), separation and washing of the insoluble
residue, then boiling this residue with 1.25% m/v sodium hydroxide, this is followed by
separation. The sample is washed using boiling distilled water, dried followed by weighing of the
insoluble residue then determination of loss of mass on incineration.
Samples should be prepared in such a way as to allow for maximum contact with the reagents
used. For fine samples, if the sample passes through a 1.00 mm sieve completely, mix
thoroughly with a spatula and scoop. For coarse samples, the sample is prepared by grinding and
blending to obtain a homogenous sample. Grinding is done in a well cleaned mill and then blend
using a laboratory blender until it passes through a 1.00 mm sieve completely .This is then kept
in a sample bottle. For samples that cannot be homogenized well due to high moisture contents,
preliminary drying is carried out @ 103oC for 4 hours. After blending in a blender, cool the
samples in a desiccator.

Procedure
Condition the glass crucibles of porosity number 2 by heating them in a muffle furnace at a
temperature of 200oC and increasing the temperature to 520oC. Heat at this temperature for 30
minutes before cooling and removing them at a 200oC in a desiccator and cool them to a room
temperature. Into each of the crucibles, put some conditioned celite. Record the serial number of
each of the crucibles. Weigh 1 gram of each sample into each of the crucible M1. Make sure that
one sample in the set has a duplicate. Put the crucibles in the holder. Mount the crucibles into the
cold extraction unit and de-fat each of the samples by spraying acetone gently until it covers the
sample using a spray bottle and filtering. This is repeated twice.

Acid Treatment
Fill reagent bottles using Sulphuric acid and place it on a hot plate. Heat the reagent to boiling.
Pour the hot acid into columns from the top using a funnel to the middle mark (150 ml). Add 3
drops of an antifoaming agent (n-octanol) into each column. Switch on the heater. When the
reagent starts boiling, time for 30 minutes. During the acid extraction time, heat distilled water in
a beaker and sodium hydroxide in a reagent bottle. At the end of the extraction time, wash the
sample 3 times using 50 ml of hot distilled water using a special water sprayer before continuing
with alkaline treatment.

Alkaline Treatment
Repeat the above procedure but instead of Sulphuric acid, use sodium hydroxide solution.

Drying
Using the crucible holder, transfer the crucibles with contents into an oven and dry them at
130oC for 2 hours. Allow them to cool in a desiccator and record the weight as M2. Transfer the
crucibles with dried residues into a muffle furnace at a temperature of 200oC when the furnace is
cold. Increase the temperature gradually to 500oC. Incinerate the sample fully to 530oC for at
least 3 hours or when the ash is white. Switch off the furnace to cool at 200oC and transfer the
crucibles to a desiccator. Cool them to room temperature and weigh, M3.

Calculation
Crude fibre content (%m/m) is given by:

19
 𝑀2−𝑀3
𝑀1
∗ 100

PHYSICAL METHODS
These are techniques that are not accompanied with chemical reactions. They include wheat
grading, rice grading, soap grittiness test.

(a).Wheat grading.
Take the weight of the empty bucket of the manual wheat grading machine. Fill the funnel of the
manual wheat grading machine with desired sample. Level the top of the bucket full with wheat
sample with a flat edged bar, in so doing to let the excess wheat overflow into the tray set below
the machine. Take the measurement of the filled and levelled bucket from the balance and note
the weight values .Grade the wheat as per set standards in correspondence with the weight
difference obtained.

(b). Rice grading

Weigh 25 grams of the sample under analysis and spread it on a clean dry working bench. Sort
the rice grains in consideration to broken grains, heat damaged, red streaked variety, husky,
organic and inorganic impurities, chalky variety among other parameters. Weigh each isolated
portion and multiply by a factor of four.

(c). Soap grittiness test.

Cut the washing bar soap sample a portion of 7cm by 4cm. Feel the texture of the sample by
rubbing it with your fingers through a running tap water. The texture should be of fine tilth.

EXTRACTION/SEPERATION METHODS
These techniques are based on physical separation of components from a mixture using the
components’ physical properties such as the degree of solubility in different solvents. They
include:

(a). Total fatty matter of soaps.

Soap is a sodium or potassium salts of various naturally occurring fatty acids. It is produced by
saponification or basic hydrolysis reaction of a fat or oil. Sodium carbonate or sodium hydroxide
is used to neutralize the fatty acid. The fatty acids, stearic, palmitic, myristic, lauric and oleic
acids, contribute to lathering and washing properties of the soap. The chemical characteristics of
soap depend on several factors: the strength and purity of alkali, the kind of oil used,
completeness of saponification

General Overall Hydrolysis Reaction

Fat + NaOH → glycerol + sodium salt of fatty acid

20
Soap is an anionic surfactant used in conjunction with water for washing and cleaning. Although
the reaction is shown as one step reaction, it is in fact two steps. The net effect as that the ester
bonds are broken. The glycerol turns back into an alcohol. The fatty acid portion is turned in to a
salt because of the presence of a basic solution of the NaOH.
In the carbonyl group, one oxygen now has a negative charge that attracts the positive sodium
ion. The fats and oils used in soap making come from animals or plant sources. Each fat or oil is
made up of a distinctive mixture of several different triglycerides. In a triglyceride molecule,
three fatty acid molecules are attached to one molecule of glycerin. There are many types of
triglycerides; each type consists of its own particular combination of fatty acid. Fatty acids are
the components of fats and oils that are used in soap making.

PROCEDURE
Grind the bar soap sample by scrubbing it against a special perforated steel plate. Weigh 5 grams
of the sample into a glass conical flask. Add 100 ml of distilled water and warm the sample to
dissolve. Cool and transfer the sample into a separating funnel. Add three drops of methyl orange
indicator then you acidify it by using 1:1 HCL: WATER solution. Add 50 ml of petroleum ether
and shake the tapped separating funnel once before opening the tap to release gas. Shake while
releasing the gas produced until the gas pressure release sound stops. Clamp the separating
funnel in a stand and allow for settling which will lead to phase separations. (The clear
petroleum ether liquid dissolves the fat from the sample and since it’s less dense than the colored
extract, it rests on its top). If the phases are not easily separated, add 5 ml of ethanol to cleave the
bonds at the interphase leading to a fast separation. Open the tap to release the dense colored
matrix into an empty clean and dried conical flask. Discard the clear liquid into a separate beaker
and return the colored sample matrix back to the separating funnel for second extraction. Add 50
ml of petroleum ether solution, shake several times while releasing the gas produced until the gas
pressure stabilizes. Also for phase separation before tapping the dense colored sample matrix.
Discard the clear liquid in the same vessel you had preserved the first extraction. Extract the fatty
matter one more time by adding 50 ml petroleum ether and repeated shaking while releasing
gases. Once this is done, place the preserved clear liquid back into the separating funnel and
rinse it with 70 ml of distilled water. Two phases separate which are both clear (petroleum ether
which dissolved soap fatty matter rests above a clear distilled water). Discard the dense phase
which is made of distilled water. Rinse the petroleum ether two more times with 30 ml of
distilled water each time and discarding the distilled water. Once this step is successful, pass the
petroleum ether through a filter funnel which is supporting crystals of anhydrous sodium
sulphate salt to trap any remaining water molecules in the petroleum ether-fatty matter complex.
This is tapped into a clean, conditioned and preweighed glass beaker. The liquid in the beaker is

21
transferred into an oven set @ 120oC for the petroleum ether to vaporize leaving behind the fatty
matter. This cooled beaker is cooled and weighed to establish the content of weight gain. The
amount of fatty matter is then expressed as a percentage.

(b). Fischer’s test for honey

Weigh one gram of the honey sample into a clean glass beaker and add 30 ml of petroleum ether
reagent. Transfer the content into a separating funnel. Shake and allow for settling which will
lead to formation of two phases. Extract and collect the upper layer. Place the beaker with the
extract in an oven all evaporate the petroleum ether until the sample remains to about 5 ml and
add 2 ml of Resoranal followed by 10 ml of concentrated HCL and observe in the first 30
seconds. If red, then it shows positive Fischer test. If it shows any other color, then its negative
Fischer test.

(c). Liquid detergent active detergent.

Weigh 5 grams of the sample into a 250 ml glass beaker. Add 100 ml of ethanol and warm to
dissolve. Decant and filter through a number 3 gouch crucible using filtration apparatus which
comprises of a vacuum pump. Collect the filtrate and transfer it to a separating funnel. Rinse the
flask three times with 40 ml of distilled water in each lot and add to the separating funnel.
Extract three times using 75 ml of petroleum ether in each cycle. Extract the upper organic layer.
Transfer to a preweighed glass beaker and condition it for 30 minutes. Cool and take the final
weight of the beaker. Calculate the weight difference to compute for the degree of active
ingredients.

POTENTIOMETRIC METHODS
These are analytical methods in which a measurement of potential, current, or charge in an
electrochemical cell serves as the analytical signal. Although there are only three principal
sources for the analytical signal—potential, current, and charge—a wide variety of experimental
designs are possible. The simplest division is between bulk methods, which measure properties
of the whole solution, and interfacial methods, in which the signal is a function of phenomena
occurring at the interface between an electrode and the solution in contact with the electrode. The
measurement of a solution’s conductivity, which is proportional to the total concentration of
dissolved ions, is one example of a bulk electrochemical method. A determination of pH using a
pH electrode is one example of an interfacial electrochemical method.

(a). Finding the PH of samples using a portable PH meter.

Weigh 10 grams of the sample into a glass conical flask for solid samples. Add 100 ml of
distilled water and let it soak for 10 minutes. Stir to dissolve using an electromagnetic stirrer.
Calibrate the PH meter using the buffer solutions (B4, B7 and B10). Measure the PH of the
sample @ 25oC each time rinsing the probe before measuring PH of a different sample. For
liquid samples, measure 50 ml of the sample and dip the PH meter probe into the sample and
read the PH value.

(b). Finding the concentration of total dissolved solids in samples.

22
Prepare the analytical samples using the above procedure. However, do not use the PH button
this time, use the TDS button to establish the concentration of total dissolved solids in parts per
million of a given sample @ 25oC.

(c). Finding the conductivity ash of sugar samples.

Weigh 31.3 grams of sugar sample into a 250 ml conical flask. Add 100 ml of distilled water and
shake to dissolve. Calibrate the portable meter with conductivity standard solutions (74 µM, 147
µM and 170 µM). Rinse the probe and measure the conductivity of the samples at the room
temperature.

(d). Finding the fluoride concentration in samples.

Prepare the analytical samples as in (a) above. Calibrate the fluoride ions analytical machine
using 0.1 and 0.001 fluoride standards. To 50 ml of test sample, add 5 ml of TIS SAP solution.
Stir the TIS SAP-sample solution complex with the probe and let it settle. Read the fluoride
concentration of the sample from the machine in parts per million.

ASHING TECHNIQUES
Ashing in analytical chemistry is defined as the heating of a substance to leave only
noncombustible ash, which is analyzed for its elemental composition. The sample preparation
techniques incorporating some form of 'ashing' are as follows: Dry Ashing is usually performed
by placing the sample in an open inert vessel and destroying the combustible (organic) portion of
the sample by thermal decomposition using a muffle furnace. Typical ashing temperatures are
450 to 550 °C. Magnesium nitrate is commonly used as an ashing aid. Charring the sample prior
to muffling is preferred. Charring is accomplished using an open flame. Sulfated Ashing
involves treatment of the sample after charring using an open flame with sulfuric acid (the char is
wetted using the minimum amount of sulfuric acid and then brought to dryness before muffling)
and then placing in a muffle furnace. Wet Ashing is treatment of the sample with a moderate
amount of sulfuric acid before charring. Charring is performed using an open flame. Liquid
samples tend to foam. After the excess sulfuric acid is driven off, the sample is muffled as above.
Low-temperature Ashing involves treatment of the sample at ~ 120 °C using activated (singlet
state) oxygen. Closed System Ashing involves thermal decomposition in oxygen in a closed
system such as a Schöniger flask or an oxygen Parr bomb.

General Dry Ashing Procedure

This procedure is used for a wide variety of sample types, which include organic polymers,
natural products such as agricultural materials, biological materials, petroleum products and
synthetic organic research materials.

Procedure:
Dry ashing procedures are typically and preferentially performed in Pt° crucibles. Glassy carbon
can be used but some attack may occur. Nickel and iron can also be used but the metal from the
crucible can cause significant spectral interference. A sample size ranging from a few milligrams
to 100 grams is weighed into the crucible. The crucible is placed on a hot plate and set on the
highest setting done in a Class-A hood due to highly toxic fumes. The use of a propane torch is

23
helpful in speeding up the process and is necessary for the ignition of certain sample types such
as polyethylene. As soon as fumes cease to evolve, the sample is placed in a muffle furnace at
450 - 500 °Celsius for one hour or until all of the carbon has been oxidized.

General Sulfated Ashing Procedure

This procedure is used for a wide variety of sample types which include organic polymers,
natural products such as agricultural materials, biological materials, petroleum products and
synthetic organic research materials. The sulfated ash is used over dry ashing when the analyst
needs to fix a material as the sulfate to prevent volatilization otherwise it has no real advantages
over dry ashing.

Procedure:
Sulfated ashing procedures are typically and preferentially performed in Pt crucibles. Glassy
carbon can be used but some attack may occur. Nickel and iron can also be used but the metal
from the crucible can cause significant spectral interference. A sample size ranging from a few
milligrams to 100 grams is weighed into the crucible. The crucible is placed on a hot plate and
set on the highest setting in a Class-A hood due to highly toxic fumes. The use of a propane torch
is helpful in speeding up the process and is necessary for the ignition of certain sample types
such as polyethylene. As soon as fumes cease to evolve, wet the char with concentrated sulfuric
acid. Typically a few drops are required. Continue to heat the sample until the white dense sulfur
trioxide fumes cease to evolve. The sample is placed in a muffle furnace at 450 - 500 °Celsius
for one hour or until all of the carbon has been oxidized.

Acid Insoluble Ash

To a sample already ashed through the dry weight ashing technique, add 5 ml of 1:1 HCL:
WATER and warm in a hot plate to dissolve. Filter mixture through a filter funnel and ash the
filter paper while in the same crucible. Cool and take the final weight for computation.

TITRIMETRY
Titrimetry is the measure of the volume of a reagent reacting stoichiometrically with the analyte.
Titrimetric methods are classified into four groups based on the type of reaction involved.
These groups are acid–base titrations, in which an acidic or basic titrant reacts with an analyte
that is a base or an acid; complexometric titrations involving a metal–ligand complexation
reaction; redox titrations, where the titrant is an oxidizing or reducing agent; and precipitation
titrations, in which the analyte and titrant react to form a precipitate.

ACID-BASE TITRATIONS.
For an acid–base titration, the equivalence point is characterized by a pH level that is a function
of the acid–base strengths and concentrations of the analyte and titrant. The pH at the end point,
however, may or may not correspond to the pH at the equivalence point. This due to fact that the
pH changes during a titration. Examples of acid-base titrations I came across include:

(1). Titration to find the saponification value of soaps:

24
Saponification value (SV) is the milligrams of potassium hydroxide required to saponify
completely 1 gram of fat. This covers neutral fat and FFA present, and obviously relates to the
molecular weights of the fatty acids involved. Where these molecular weights are low, the one
gram will require more potassium hydroxide (coconut oil SV = 250), and where they are high, it
will require less (rapeseed oil SV = 175).
Procedure.
Weigh 1 gram of the test sample into 250 ml conical flask. Add 25 ml of 0.1N NaOH followed
by 25 ml of ethanol. Add 3 drops of phenolphthalein indicator then reflux for one hour. Cool and
titrate against 1N HCl. Titrate alongside the reagent blank. Calculate the saponification value
using the formula:

Saponification value or number of fat = mg of KOH consumed by 1g of fat.

Weight of KOH = Normality of KOH * Equivalent weight* volume of KOH in litres

Volume of KOH consumed by 1g fat = [Blank –test] ml

(2). Titration for quantitative analysis of free fatty acids in samples.

Free fatty acids (FFA) are produced by the hydrolysis of oils and fats. The level of FFA depends
on time, temperature and moisture content because the oils and fats are exposed to various
environments such as storage, processing, heating or frying. Since FFA are less stable than neutral
oil, they are more prone to oxidation and to turning rancid. Thus, FFA is a key feature linked with
the quality and commercial value of oils and fats. Based on titration, oils or fats need to be
dissolved in hot neutralized ethanol or ethanol/diethyl ether using phenolphthalein as an end point
indicator.

Procedure
Grind the sample and weigh 5 grams of the sample into a conical flask. Add 50 ml of 1:1petroleum
ether: ethanol and shake to dissolve. Add 3 drops of phenolphthalein indicator and titrate against
0.1N NaOH to a pale pink color end point.

(3). Quantitative determination of acid value of samples.

A small quantity of free fatty acids is usually present in oils along with the triglycerides. The free
fatty acid content is known as acid number/acid value. It increases during storage. The keeping
quality of oil therefore relies upon the free fatty acid content. The free fatty acid in oil is estimated
by titrating it against KOH in the presence of phenolphthalein indicator. The acid number is
defined as the mg KOH required to neutralize the free fatty acids present in 1g of sample. However,
the free fatty acid content is expressed as oleic acid equivalents.

Procedure.
Grind the sample and weigh 5 grams of the sample into a conical flask. Add 50 ml of
1:1petroleum ether: ethanol and shake to dissolve. Add 3 drops of phenolphthalein indicator and
titrate against 0.1 KOH to a pale pink color end point.

Calculation

25
 Titrate value x Normality of KOH
Acid value (mg x 56.1
KOH/g) =
Weight of sample (g)

The free fatty acid is calculated as oleic acid using the equation
1mL N/10 KOH = 0.028g oleic acid

(4). Titratable acidity.

Titratable acidity deals with measurement of the total acid concentration contained within a food
(also called total acidity). This quantity is determined by exhaustive titration of intrinsic acids
with a standard base. Titratable acidity is a better predictor of acid’s impact on flavor than pH.

Procedure
Mix sample thoroughly by pouring it from one container to another. The temperature of the
sample should be near 20 degrees. Pipette 17.6 ml of milk or cream into a white cup. Readily
available 9 ml pipettes are also used but require application of a correction factor to the final
result. Add six drops of phenolphthalein indicator solution to milk, 10 drops if the product is
cream. Titrate the sample with the N/10 sodium hydroxide solution (0.1 Normal NaOH) while
stirring the sample with the glass rod. Look for the appearance of a faint pink color which signals
the endpoint. Add another drop or half a drop of NaOH if the pink color does not persist for 30 s.
Record the number of ml of NaOH used to reach the endpoint. This value is called the 'titre'.
Titratable acidity reported as percent lactic acid is dependent on the volume of sample.

For the 17.6 ml pipette, % Lactic acid = 0.5 x titre

For the 9.0 ml pipette, % Lactic acid = 0.98 x titre.
(5). Titration to determine the amount of sodium chloride content in samples.
For preservation and taste reasons, a knowledge of the concentration of salt (sodium chloride) in
foodstuffs is often required. Normally, it is sufficient to determine total chloride and express this
in terms of sodium chloride. Minor components of foods can also contribute chloride ions but
foods which require an analysis for salt are normally those in which salt is added as or significant
ingredient. Titrimetric methods are the most commonly used. Salt must first be extracted from
the food either by ashing at 500-550oC (alkali chlorides are relatively volatile at high
temperatures), followed by the dissolution of the ash, or by boiling the food stuff in diluted Nitric
(V) acid. In the absence of acid, the chloride ion can be determined by the Mohr procedure
involving direct titration with 0.1 N Silver Nitrate: or in the presence of acid by the volhard
procedure involving the addition of excess Silver Nitrate and titration with potassium
thiocyanate.
(a). Mohr method.
Wash the ash into a conical flask or white porcelain dish with minimal water. Add 1 ml of 5%
Potassium Chromate solution and titrate with 0.1 N Silver Nitrate solution to the first appearance
of an orange color. 1ml 0.1m AgNO3= 0.005844 g NaCl.
(b). Volhard method.

26
Accurately weigh about 5 grams of the prepared sample into a 250 ml conical flask, add 10 ml of
water and 25.0 ml of 0.1N Silver Nitrate solution (or sufficient excess to precipitate all the
chloride in the weighed sample). Add 10 ml of concentrated Nitric and some boiling granules.
Alternatively, dissolve the ash from a sample in the Nitric (V) acid, add water and Silver Nitrate.
Boil gently with a small funnel in the neck of the flask for about 10 minutes until the solution is
pale yellow. Cool and add 50 ml of water followed by 5 ml of saturated ammonium ferric
Sulphate solution and a few drops of Nitrobenzene and titrate with 0.1N Ammonium or
potassium thiocyanate solution until permanent reddish color persists for 15 seconds. 0.5 grams
of urea may be added to the hot solution to remove the yellow Nitrous fumes. Carry out a blank
determination on the reagent alone. The different between the titrations of the blank
determination and the test is equivalent to the chloride concentration.
(6). Determination of Total Alkali Content in the Soap Samples.
The alkalis used in soap making are NaOH (sodium hydroxide) and KOH (potassium hydroxide).
Sodium carboxylates are the common toilet soaps. Potassium carboxylates or potassium soaps
are obtained when saponification of a fat or oil is carried with potassium hydroxide. Potassium
soaps are softer than sodium soaps and they are used for special purposes when rapid solution is
desired eg: in making shaving creams or liquid soaps. The composition of sodium or potassium
carboxylates constituting soap depends on the percentage of fatty acids bonded to glycerol in the
original triglycerides. Solid fats give mixture with higher proportion of sodium or potassium salts
of higher fatty acids (palmitic acid, stearic acid) and give hard soaps. The vegetable oils give
mixtures with a greater proportion of unsaturated fatty acids (oleic acid and linoleic acid) and
give soft soaps.

Procedure.
5gm of soap sample is dissolved in 100ml hot water. About 40ml of 0.5N HNO3 is added to
make it acidic. The mixture is heated until fatty acids are floating as a layer above the solution. It
is cooled in ice water to solidify the fatty acids. The fatty acids were separated and the aqueous
solution was treated with 50ml chloroform to remove the remaining fatty acids. The aqueous
solution is measured and 10ml of it is titrated against 0.5N NaOH using methyl orange as
indicator and from the titer value the total alkali content is calculated using the following
method.
Calculation:
Total volume of the aqueous solution =V=__________ml
10 ml of aqueous solution require t ml of NaOH
V ml of aqueous solution requires = Vxt /10 = A ml.
Amount of NaOH required by acid in aqueous solution =A ml
Volume of HNO3 required, B ml =A x Normality of NaOH /Normality of HNO3
Volume of HNO3 required for neutralizing NaOH = C=40 – B
Amount of NaOH in 1000 cc of soap solution (E) = (C x 40 x Normality of HNO3 g) /1000
250 cc of soap solution contains (F) = (E x 250) / 1000 g
2 NaOH → Na2O + H2O
80 gram of NaOH 62 g of Na2O
F g of NaOH requires (Y) = (62 x F) / (80) g of Na2O
Weight of soap taken = 5 g
% of alkalinity = (Y x 100) / w = ______________.

27
REDOX TITRATIONS
Redox titrations are based on a reduction-oxidation reaction between an oxidizing agent and a
reducing agent. A potentiometer or a redox indicator is usually used to determine the endpoint of
the titration, as when one of the constituents is the oxidizing agent potassium dichromate. The
color change of the solution from orange to green is not definite, therefore an indicator such as
sodium diphenylamine is used. Analysis of wines for sulfur dioxide requires iodine as an
oxidizing agent. In this case, starch is used as an indicator; a blue starch-iodine complex is
formed in the presence of excess iodine, signaling the endpoint. Some redox titrations do not
require an indicator, due to the intense color of the constituents. For instance, in
permanganometry a slight persisting pink color signals the endpoint of the titration because of
the color of the excess oxidizing agent potassium permanganate. In iodometry, at sufficiently
large concentrations, the disappearance of the deep red-brown triiodide ion can itself be used as
an endpoint, though at lower concentrations sensitivity is improved by adding starch indicator,
which forms an intensely blue complex with triiodide.
(a). CHEMICAL OXYGEN DEMAND DETERMINATION IN WATER SAMPLES
The Chemical Oxygen Demand (COD) method determines the quantity of oxygen required to
oxidize the organic matter in a waste sample, under specific conditions of oxidizing agent,
temperature, and time. Since the test utilizes a specific chemical oxidation the result has no
definite relationship to the Biochemical Oxygen Demand (BOD) of the waste or to the Total
Organic Carbon (TOC) level. The test result should be considered as an independent
measurement of organic matter in the sample, rather than as a substitute for the BOD or TOC
test. The method can be applied to domestic and industrial waste samples having an organic
carbon concentration greater than 50 mg/L. For lower concentrations of carbon such as in surface
water samples, the Low Level Modification should be used. When the chloride concentration of
the sample exceeds 2000 mg/L, the modification for saline waters is required. Organic and
oxidizable inorganic substances in the sample are oxidized by potassium dichromate in 50%
sulfuric acid solution at reflux temperature. Silver sulfate is used as a catalyst and mercuric
sulfate is added to remove chloride interference. The excess dichromate is titrated with standard
ferrous ammonium sulfate, using orthophenanthroline ferrous complex as an indicator.
PROCEDURE
Place several boiling stones in the reflux flask, followed by 50.0 mL of sample or an aliquot
diluted to 50.0 mL and 1 g of HgSO4. Add 5.0 mL conc. H2SO4 ; swirl until the mercuric sulfate
has dissolved. Place reflux flask in an ice bath and slowly add, with swirling, 25.0 mL of 0.025
N K2Cr2O7. Now add 70 ml of sulfuric acid-silver sulfate solution to the cooled reflux flask,
again using slow addition with swirling motion. Caution: Care must be taken to assure that the
contents of the flask are well mixed. If not, superheating may result, and the mixture may be
blown out of the open end of the condenser. If volatile organics are present in the sample, use an
allihn condenser and add the sulfuric acid-silver sulfate solution through the condenser, while
cooling the flask, to reduce loss by volatilization. Apply heat to the flask and reflux for 2 hours.
Allow the flask to cool and wash down the condenser with about 25 mL of distilled water. If a
round bottom flask has been used, transfer the mixture to a 500 ml Erlenmeyer flask, washing
out the reflux flask 3 or 4 times with distilled water. Dilute the acid solution to about 300 ml with
distilled water and allow the solution to cool to about room temperature. Add 8 to 10 drops of

28
ferroin indicator to the solution and titrate the excess dichromate with 0.25 N ferrous ammonium
sulfate solution to the end point. The color change will be sharp, changing from a blue-green to a
reddish hue. Blank-Simultaneously run a blank determination following the details given in and
but using low COD water in place of sample.
(b).QUANTITATIVE DETERMINATION OF FREE CHLORINE IN WTER SAMPLES
Bleaching powder is a mixture of CaCl2, Ca(OCl)Cl and Ca(OCl)2 and is prepared by the
reaction of Cl2 and slaked lime, Ca(OH)2. The commercial bleaching powder consists essentially
of a mixture of calcium hypochlorite Ca(OCl)2 and the basic chloride CaCl2, Ca(OH)2, H2O and
some free slaked lime is usually present. The active constituent is the hypochlorite, which is
responsible for the bleaching action. Upon treating bleaching powder with hydrochloric acid,
chlorine is liberated.

ClO + Cl + 2H+ → Cl2 + H2O

The available chlorine refers to the chlorine liberated by the action of dilute acids on the
hypochlorite, and this is expressed as a percentage by weight in the case of bleaching powder.
Commercial bleaching powder contains 36-38% of available chlorine. The hypochlorite solution
or suspension is treated with excess solution of a solution of Potassium Iodide, and strongly
acidified with acetic acid.

ClO + 2I + 2H+→ Cl + I2 + H2O

The liberated iodine is titrated with standard sodium thiosulphate solution. The solution should
not be strongly acidified with hydrochloric acid, for the little calcium chlorate which is usually
present, by virtue of decomposition of the hypochlorite will react slowly with the Potassium
Iodide and liberate iodine.
ClO3- + 6I- + 3I2 + 3H2O
PROCEDURE
Standardize sodium thiosulphate solution by pipetting 20 ml of 0.1 N Potassium chromate
solution into a clean conical flask. Add 20 ml of Sulphuric (vi) acid followed by 10 ml of 15%
KI solution. Then titrate this against thiosulphate from the burette until it turns into pale yellow.
Add 1 ml of starch indicator and the liberated Iodine is immediately titrated against the
thiosulphate solution until the disappearance of blue color. Weigh 5 grams of the sample into a
mortar and add a little amount of water, grind the mixture to a suitable paste. Pour the milky
liquid suspension into 500 ml volumetric flask. Grind the residue in the mortar again with little
amount of water and repeat the above procedure until the whole suspension has been transferred
into the flask as a very fine suspension and the mortar is washed clean. Fill the volumetric flask
up to the mark and shake well. Transfer 50 ml of the turbid liquid into a iodine flask. Add 25 ml
of distilled water followed by 10 ml of 15% potassium Iodide solution then 10 ml of glacial
acetic acid. The liberated Iodine is titrated against 0.1 N Sodium Thiosulphate solution until it
turns to pale yellow. Add 1ml of starch indicator solution then titrate the liberated Iodine against
the thiosulphate solution until the disappearance of the blue color.

29
BACK TITRRATIONS
Back titration is a titration done in reverse; instead of titrating the original sample, a known
excess of standard reagent is added to the solution, and the excess is titrated. A back titration is
useful if the endpoint of the reverse titration is easier to identify than the endpoint of the normal
titration, as with precipitation reactions. Back titrations are also useful if the reaction between the
analyte and the titrant is very slow, or when the analyte is in a non-soluble solid.
(a). QUANTITATIVCE DETERMINATION OF THE INVERT SUGAR.
Sucrose, commonly known as table sugar, is a disaccharide composed of an alpha-D-glucose
molecule and a beta-D-fructose molecule linked by an alpha-1,4-glycosidic bond. When this
bond is cleaved in a hydrolysis reaction, an equimolar mixture of glucose and fructose is
generated. This mixture of monosaccharides is called invert sugar, which is derived from the fact
that sucrose rotates plane polarized light to the right i.e., dextrorotatory, +66.5º, whereas the
hydrolysis products rotates plane polarized light to the left i.e., levorotatory, -20º for the mixture
(+52.5º for D (+)-glucose and -92º for D (-)-fructose).
Sucrose can be hydrolyzed in the presence of an enzyme called invertase or sucrase.

Sucrose + H2O ---> glucose + fructose

The official name for invertase is beta-fructofuranosidase (EC3.2.1.26), which implies that the
reaction catalyzed by this enzyme is the hydrolysis of the terminal nonreducing beta-
fructofuranoside residues in beta-fructofuranosides. Alpha-D-glucosidase, which splits off a
terminal glucose unit, can also catalyze this reaction. Sucrose can be hydrolyzed relatively
easily; the reaction proceeds in an acidic environment without the aid of invertase. Invertase is
mainly used in the food (confectionery) industry where fructose is preferred over sucrose
because it is sweeter and does not crystallize as easily. However, the use of invertase is rather
limited because another enzyme, glucose isomerase, can be used to convert glucose to fructose
more inexpensively. For health and taste reasons, its use in food industry requires that invertase
be highly purified.
PROCEDURE
Weigh 10 grams of the sample into a 250 ml conical flask. Add 100 ml of distilled water
followed by 10 ml of the Miller’s solution. Incubate in a water bath for set @ 95oC for 10
minutes. Cool rapidly using ice bags placed in a water sink with tap water. Add 5 ml of 5 N
acetic acid. Add 20-25 ml of 0.03N of Iodine solution then titrate with 0.033 N Na2S2O3.
(b). Iodine Value Determination.
The iodine value or “iodine adsorption value” or “iodine number” or “iodine index” in chemistry
is the mass of iodine in grams that is consumed by 100 grams of a chemical substance. An iodine
solution is yellow/brown in color and any chemical group in the substance that reacts with iodine

30
will make the color disappear at a precise concentration. The amount of the substance thus
required to keep the iodine solution yellow/brown is a measure of the amount of the iodine
sensitive reactive groups.
PROCEDURE
Iodine solution is prepared by 13g of iodine crystals in a 1dm3 volumetric flask containing 25g
of potassium iodide crystals, and made up to the mark with deionized water. - Standardizing
iodine solution is done through taking 0.3g of primary standard arsenic (III) oxide (As2O3)
which is dried for an hour at room temperature and dissolved in deionized water. Few grains of
NaOH are added and 3.5g of sodium crystals added, and then titrated against the iodine solution.
10% Potassium iodide solution is prepared by dissolving 10g of potassium iodide crystals in
warm water and made up to the mark in 100cm3 volumetric flask. 0.1M Sodium thiosulphate
solution is prepared by dissolving 25g of sodium thiosulphate crystals in warm water and made
up to the mark in 1dm3 volumetric flask. Standardizing Sodium thiosulphate. Sodium
thiosulphate solution is run from the burette into the iodine solution until its original
yellow/brown color is pale yellow. Then a few drops of starch solution added, thereby producing
a dark brown coloration. Further drop by drop of thiosulphate solution is continued until it turns
into a pale purple color and then disappear. 2mg of the sample is mixed with 20cm3 wiji’s
solution and 10cm3 1,1,1-trichloroethane, then left in the dark for 30 minutes. Next, 15cm3 of
10% potassium iodide solution and 10cm3 of deionised water is added. This was then titrated
against 0.1M sodium thiosulphate (VI) solution. 1cm3 of 0.1M sodium thiosulphate solution is
equal to 0.01269g of iodine. The difference between a control titration and the titration with the
oil present multiplied by this factor, gives the mass of iodine absorbed by the sample.
(c).Quantitative determination of Sulphur (IV) oxide in sugar samples
The sample is diluted and treated with sodium hydroxide to release sulfur dioxide.
The solution is then acidified and the sulfurous acid determined by titration with a standard
iodine solution using starch as an end point indicator.
This method applies to corn syrups, corn sugars and other water soluble starch hydrolyzates that
contain sulfur dioxide.

PROCEDURE
Weigh accurately 100 g of sample into a 400 mL Erlenmeyer flask. Add sufficient purified water
to bring total weight to 200 g. Mix the sample and water until the solution is homogenous. For
low level samples: (less than 20 ppm), Cool to 10 °C or below. Place cold sample on magnetic
stirrer and stir at a rate sufficient to produce a small vortex at the solution surface. Add 10 mL of
cold 1.5 N sodium hydroxide solution and stir for 15 to 20 seconds. Add 10 mL of starch
indicator solution and 10 mL of cold 2.0 N sulfuric acid solution; titrate immediately with 0.005
N standard iodine solution until a light blue color persists for one minute. For high level samples:
(above 20 ppm), add 20 mL of 1.0 N sodium hydroxide solution and stir 5 minutes. Add 25 mL
of 1.0 N sulfuric acid solution, 10 mL starch indicator solution, and titrate with 0.05 N standard
iodide solution until a light blue color persists.
Perform a blank titration using 200 mL of purified water and all reagents.

31
(d). Quantitative Analysis of Sucrose in Confectionary
This analytical method is applied for the determination of sucrose contained in confectionery,
such as sugar confectionary, bakery products, chocolates and other products containing cocoa,
and boiled red beans, and the analysis is performed by titration method. Prepare analysis samples
appropriately depending on their conditions. For solid materials, grind them using a grinder or a
mixer. For pasty or wet materials, homogenize them in mortars. In any case, collect a relatively
large amount of the sample randomly and homogenize it by grinding and/or mixing to
uniformity. Accurately weigh a certain amount of the sample homogenized, remove fat if a large
amount of it is contained, and dissolve in water. Transfer the solution with about 200 mL of
water to a 250 mL volumetric flask, and shake well (about one hour) for extraction.
Add 20 mL each of deproteinizing solution if necessary, and shake well to mix. Dilute the
solution to volume with water and mix well again. After leaving to stand for 30 minutes, filter
the solution through filter paper. Use the filtrate as a test solution.

PROCEDURE
Put 5.0 mL of Fehling’s Solution A and 5 mL of Fehling’s Solution B into a 200 mL Erlenmeyer
flask containing a few glass beads and add from a 50 mL burette 15 mL of the sugar solution.
Boil the solution on an electric stove (heater) for 2 minutes and titrate. Subsequently, add from a
50 mL burette to another 200 mL Erlenmeyer flask the sugar solution within 1 mL of the
anticipated end point from the result of the preliminary titration above and titrate in the same
manner. Multiply the titer by the factor of the Fehling’s solution and find the concentration of
invert sugar.

(e). Peroxide value determination in samples.

Peroxide value is a measure of the peroxides contained in the samples such as oil samples.
During storage, peroxide formation is slow at first during an induction period that may vary from
a few weeks to several months according to the particular oil or fat, the temperature etc. Peroxide
value is always determined volumetrically methods. These methods depend on the reaction of
potassium iodide in acid solution with the bound oxygen followed by titration of the liberated
iodine with sodium thiosulphate. Chloroform is normally used as solvent.

Procedure
The test should be preferably carried out in subdued daylight. Weigh out 1 gram of the sample
into a clean dry boiling tube and while still liquid add 1 gram of powdered potassium iodide and
20 ml of solvent mixture. Place the tube in a boiling water so that the liquid containing 20 ml of
potassium iodide solution (5%), wash out the tube twice with 25 ml water and titrate with 0.02 N
Sodium thiosulphate using starch as an indicator. The endpoint is colorless solution.

COMPLEXOMETRIC TITRATIONS.
Complexometric titrations rely on the formation of a complex between the analyte and the titrant.
In general, they require specialized complexometric indicators that form weak complexes with
the analyte. The most common example is the use of starch indicator to increase the sensitivity of

32
iodometric titration, the dark blue complex of starch with iodine and iodide being more visible
than iodine alone. Other complexometric indicators are Eriochrome Black T for the titration of
calcium and magnesium ions, and the chelating agent EDTA used to titrate metal ions in
solution.
(a). QUANTITATIVE DETERMINATION OF MAGNESIUM IN SALT SAMPLES
This method is based on complexometric titration while using Eriochrome Black T indicator and
EDTA for salts as the chelating agent. Ammonium chloride-ammonium hydroxide is used as a
buffer.

Procedure
Measure 10 grams of the sample into a clean conical flask. Add 100 ml of distilled water. Draw
10 ml of the aliquot sample into another clean conical flask. Add 100 ml of distilled water. Add
10 ml of 1:1 NH4Cl: NH4OH buffer. Add 5 drops of Eriochrome black T indicator and titrate
against EDTA for salts to a pale blue end point.

(b). QUANTITATIVE DETERMINATION OF CALCIUM IN SALT SAMPLES.

This method is based on the complexometric titration while using calcein indicator under
alkaline medium initiated by 10% NaOH. EDTA for salts is still used as a chelating agent.

Procedure
Measure 10 grams of the salt sample into a clean conical flask. Add 100 ml of distilled water.
Draw 10 ml of the aliquot sample into another clean conical flask. Add 100 ml of distilled water.
Add 5 ml of 10% NaOH followed by 1 ml of calcein as the indicator. Titrate against EDTA for
salts as the chelating agent to a purple end.

7. MEASUEREMENT TECHNIQUES
These techniques are based on instrumentation to measure the degree of a parameter under
analysis. Some of them are:

(a). The degree of Brix in samples.

Brix is a scale based on the amount that light bends when it passes through a liquid. The brix
chart is the scale that anyone can use to determine poor, average, good or excellent in a
foodstuff. Brix measures the percent solids (TSS) in a given weight of plant juice. Brix is often
expressed as the percentage of sucrose. However, the "sucrose" can vary widely. Brix is actually
a sum of sucrose, fructose, vitamins, minerals, amino acids, proteins, hormones, and other solids
in one part of plant juice. Brix varies directly with plant quality. For instance, a poor, sour tasting
grape from worn-out land can test 8 or less brix. On the other hand, a full flavored, delicious
grape, grown on rich, fertile soil can test 24 or better brix.

Procedure
Switch on the refractometer from the power source. Open the cuvette and calibrate the machine
using the distilled water as a standard solution in which it should read zero from the lower scale.
Mount the sample and wipe any spillage on the machine before reading the Brix value. Adjust
the refractometer knobs such that the dark fields intersect. Read the value and read the Brix

33
correction values from the chat if the values are taken at any other temperature other than 20oC.
For sample’s refractive index, use the upper scale.

(b). Measurement of lather volumes in soaps.

Lather volume is the amount of foam that form from a soap when dissolved in a hard water and
agitated. Lather volume is used to establish the cleaning properties of soaps.

Procedure
Weigh 1 gram of the sample into a clean 250 ml measuring cylinder. Add 10 ml of hard water.
Shake well to mix. Let it settle for 5 minutes before adding 20 ml of hard water. Shake 12 times
to and from and the components to settle. Read the lather volume value.

BIOCHEMICAL TECHNIQUES
TOTAL AFLATOXIN ASSAY
(a). AFLATOXINS
Aflatoxins are chemical metabolites produced by a variety of molds such as Aspergillus flavus
and Aspergillus parasiticus. They are carcinogenic and can be present in grains, nuts, cotton
seeds and other commodities associated with human food or animal feeds. Crops may be
contaminated by one or more of sub-types of aflatoxins i.e. B1, B2, G1 and G2. Aflatoxin B1 is the
most toxic and frequently detected form. The other types present are significant danger if the
concentration is at high level. Aflatoxins have been implicated in human health disorders
including hepatocellular carcinoma, aflatoxicosis, Reye’s syndrome and chronic hepatitis.
Animals are exposed to aflatoxins by consumption of feeds that are contaminated by aflatoxin
producing fungal strains during growth, harvest or storage. Symptoms of toxicity in animals
range from death to chronic diseases, reproductive interference, immune suppression, decreased
milk and egg production. Most controlling government agencies worldwide have regulations
regarding the amount of aflatoxins allowable in human and animal feedstuffs. Accurate and rapid
determination of the presence of aflatoxin in commodities is of paramount importance. The
HELICA Total Aflatoxin Assay is a competitive enzyme-linked immunoassay intended for the
quantitative determination of aflatoxin B1, B2, G1 and G2 in grains, nuts, cotton seeds and other
commodities including animal feeds.

ASSAY PRINCIPLE
HELICA Total Aflatoxin Assay is a solid phase competitive inhibition enzyme immunoassay.
An aflatoxin specific antibody optimized to cross react with all the four sub-types of aflatoxin is
coated to a polystyrene microwell. Toxins are extracted from a ground sample with 70%
methanol. The extracted sample and the HRP- conjugated aflatoxin are mixed and added to the
antibody coated microwell. Aflatoxin from the extracted sample and the HRP- conjugated
aflatoxin compete to bind with the antibody coated to the microwell. Microwell contents are
decanted and non- specific reactants are removed by washing. An enzyme substrate (TMB) is
added and color (blue) develops. The intensity of the color is directly proportional to the amount

34
of bound conjugate and inversely proportional to the concentration of aflatoxin in the sample or
standard. Therefore, as the concentration of aflatoxin in the sample or standard increases, the
intensity of blue color will decrease. An acidic stop solution is added which changes the
chromogen color from blue to yellow. The microwells are measured optically by a microplate
reader with an absorbance of 450 nm. The optical densities of the samples are compared to the
optical densities of the kit standards and an interpretative result is determined.

Procedure
Grind sufficient sample using a blender (render the sample to particle size of fine instant
coffee).weigh five grams of the ground sample into a centrifugation tube. Add 25 ml of 70%
methanol solution and shake the tube for two minutes. Allow the particulate matter to settle then
filter 10 ml of the extract through a whatman #1 filter paper and collect the filtrate to be tested.
Bring all the reagents to room temperature before use. Reconstitute the PBS tween packet by
washing out the contents with a gentle stream of distilled water into a 1 litre container. Place one
mixing well in a microwell holder for each standard and sample to be tested. Place an equal
number of antibody coated microtiter well in another microwell holder. Dispense 200 µl of the
aflatoxin HRP conjugate into each mixing well. Using a new pipette tip for each sample, transfer
100 µl of each standard and sample to appropriate mixing well containing conjugate. Mix by
priming the pipettor at least 3 times. Using a new pipette tip for each, transfer 100 µl of content
from each mixing well to a corresponding antibody coated microtiter well. Incubate at room
temperature for 15 minutes. Decant the contents from the microwells into a discard basin. Wash
the microwells by filling each with PBS tween wash buffer, then decanting the buffer into
discard basin. Repeat the wash for a total of 5 washes. Tap the microwells (face down) on a layer
of absorbent towels to remove residual buffer. Measure the required volume of substrate reagent
and place it in a different container. Add 100 µl to each microwell. Incubate to room temperature
for 5 minutes while covering with aluminum foil to avoid direct light. Load your plate the
machine and run the test.

35
CHAPTER 4: EVALUATION OF THE ATTACHMENT PERIOD
4.1: CHALLENGES ENCOUNTERED DURING THE ATTACHMENT PERIOD

1. Difficulties during the instrumental analyses.

While running analytical samples, some machines were complex and sophisticated, hence
difficult to understand their working principles to basic levels. Such machines include the
HELICA microplate readers for aflatoxin analysis, milk cream extraction Gerber
centrifuges, Crude fibre extraction units, Refractomers among others.

2. Hazardous analytical reagents.

Some analytical reagents were hazardous and hence needed special handling during the
analytical procedures. These includes chemicals such as stock solutions, some analytical
reagents such as petroleum ether and diethyl ether. Some procedures also involved
emission of hazardous fumes such as when doing a sulphated ashing analysis for
confectionaries.

3. Depletion of analytical reagents.

In some cases, one would do all the sample preparation procedures i.e. grinding, blending
and weighing, only to realize that the analytical reagents were depleted. Stock solutions
could also be missing meaning that it needed to be picked from the stores. This in general
delayed the analytical processes although it was contained.

4. Confusing crucibles during ashing

During the ashing procedures, the ceramic crucibles don’t have serial numbers. Besides,
the marking using mark pens was inefficient due to less thermal stability at the furnace
temperatures. This led to difficulties in the process of tracing the change in mass as the
analytical signal.

5. Traffic jams during the peak hours.

KEBS COAST REGION is located at MAKADARA region, which is within the
Mombasa CBD. Many vehicles and motorcycles are entering and leaving the city per unit
time, especially during the rush hours. This leads to not only traffic jams but also
skyrocketing of bus fares hence general delays.

4.2: HOW THE CHALLENGES WERE SOLVED

1. Instrumentation

36
 The challenge of using complex and sophisticated machines was solved through guidance
by the industrial attachment supervisors, trainers and the entire analytical lab staff in case
of difficulties. They were free to attaches’ consultation schemes and responded
positively.

2. Analytical reagents
For the challenge of depletion of stock solutions and analytical solutions, a close
monitoring on the levels of analytical reagents was often done and topping up of reagents
done by the analysts in charge. Requisition of the depleted stock solutions from the
manufacturers was always done in time.

3. Hazards
For the chemical and any environmental hazards whole in the lab, protective clothing was
encouraged. This includes lab coats while in the lab, wearing gloves when dealing with
chemicals, wearing gas masks when the analytical procedure emits poisonous gas and
finally wearing closed shoes while in the lab.

4. Ashing crucibles
A new method of marking the crucibles was innovated. Marking of the ceramic based
ashing crucibles was done with a graphite pencil during each ashing analysis. Graphite is
thermal stable at the furnace temperature hence more efficient. The chief lab manager
promised to purchase crucibles with serial numbers in future.

5. Traffic jams
In case of delays due to traffic jams, it was encouraged that we inform our supervisors in
time. This was also the case when one fell sick or had emergency that made him/her not
to attend his/her assigned duty. Punctually was highly expected.

CONCLUSION
In conclusion, my attachment at KEBS COAST REGION was good and I leant a lot. Not only to
supplement my coursework but also the induction to the real industrial world. I familiarized
myself with industrial organization, Practiced and enriched my technical skills in the field of
Biochemistry as well as appreciating the linkage between theory and practice. I developed
desirable attitudes and work habits in the field of Biochemistry and familiarized myself with
technical problems with a view of developing techniques for solving them. Finally, I appreciated
the role played by the industry in human health, food security, environmental management and
other social economic activities.

RECOMMENDATIONS

37
For the betterment of the BIO 320 (industrial attachment) by the University of Kabianga, I
recommend the following:

1. Insurance covers
The Department of the Biological Sciences, through the attachment coordinator should
arrange for a group insurance covers. This is equally important not only for protection
during accidents while in the industry but also for easy security of attachments posts
since some institutions need insurance covers before they admit atachees.

2. Attachment Money allocation.

In most cases, the students are in financial constraints since every move in the
attachment needs money. These include bus fares, lunch, insurance fees as well as
attachment fees charged by some institutions. The money allocated to atachees is always
insufficient.

3. Increasing the assessment lecturers

Based on the university of Kabianga constitution, the attachment assessment should be
between 3rd to 6th week. However, this may not always the case due to insufficient
assessment personnel. In some rare cases one may not be assessed at all due to delays
and is forced to terminate his/her attachment as per requirements of the industry to pave
way to others, either assessed or not.

4. Curriculum adjustments
The Biochemistry curriculum should be reprogrammed in such a way to induce more
practical concepts alongside theoretical concepts. Most of students have difficulties
during industrial attachments due to weak basic practical skills. This is not only for the
industrial attachment benefits but will also encourage research and innovations while in
the universities.

38
CHAPTER FOUR:
4.1: BIBLIOGRAPHY AND REFERENCES
The following literature review was found useful:
1) Butler, J. N. Ionic Equilibria: A Mathematical Approach. Addison-Wesley: Reading,
MA, 1964.
2) Reading, MA, 1973.Chaston, S. “Calculating Complex Equilibrium Concentrations by a
Next Guess Factor Method,” J. Chem. Educ. 1993, 70, 622–624.
3) Currie, L. A., ed. Detection in Analytical Chemistry: Importance, Theory and Practice.
American Chemical Society: Washington, DC, 1988.
4) Kirchner, C. J. “Estimation of Detection Limits for Environmental Analytical
Procedures,” In Currie, L. A., ed. Detection in Analytical Chemistry: Importance, Theory
and Practice. American Chemical Society: Washington, DC, 1988.
5) Gy, P. ed. Sampling for Analytical Purposes. Wiley: New York, 1998. Karger, B. L.;
Snyder, L. R.; Harvath, C. An Introduction to Separation Science, Wiley-Interscience:
New York, 1973.
6) Bassett, J.; Denney, R. C.; Jeffery, G. H.; et al. Vogel’s Textbook of Quantitative
Inorganic Analysis, 4th ed. Longman: London, 1981.
7) Method 4500-ClA as published in Standard Methods for the Examination of Water and
Wastewater, 20th ed. American Public Health Association: Washington, DC, 1998, pp.
4–53 to 4–55.
8) J. David Anneken, Sabine Both, Ralf Christoph, Georg Fieg, Udo Steinberner, Alfred
Westfechtel “Fatty Acids” in Ullmann’s Encyclopedia of Industrial chemistry, Wiley-
VCH, Weinheim, 2006.
9) H. H. Uhlig and F.C. Duemmling, “An investigation of free alkali determinations in
soap”, Lever brothers Co. Cambridge, Mass., A.O.C.S. meeting, Chicago, pp.8-9, 1936.
10) Azizian, H., J. K. G. Kramer, S. Ehler, and J. M. Curtis. 2010. Rapid quantification of
fish oil fatty acids and their ethyl esters by FT-NIR models. European Journal of Lipid
Science and Technology 112: 452-462.
11) M.N. Boukhatem, F.M. Amine, A. Kameli, F. Saidi, K. Walid, S.B. Mohamed, Quality
assessment of the essential oil from Eucalyptus globules Labill of Blida (Algeria) origin,
International Letters of Chemistry, Physics and Astronomy, 17(3), 2014, 303-315.
12) P. Matthews, Advanced chemistry (Cambridge University Press: New York, 2011).
13) G.O. Ojokuku, Practical Chemistry for Schools and Colleges (Gbabeks Publishers
Limited: Ibadan- Nigeria, 2001).
14) Lane J. H., Eynon L. (1923) J. Soc. Chem. Ind. (Jan.26).
15) “Toxicological evaluation of some food additives including anticaking agents,
antimicrobials, antioxidants, emulsifiers, and thickening agents,” WHO Food Additives
Series No. 5, Geneva: World Health Organization (1974); 17th Report of the Joint

39
 FAO/WHO Expert Committee on Food Additives, WHO Tech Rep Ser No. 539 (1974);
FAO Nutrition Meetings Report Series No. 53 (1974)

4.2: APPENDICES

APPENDIX I (STANDARDS).

1) KEBS: Kenya Bureau of Standards.

2) SI: System of Units.
3) SIRC: Standards Information Resource Centre.
4) ISONET: International Organization for Standardization Information Network.
5) NEP: National Enquiry Point.
6) TBT: Technical Barriers to Trade.
7) CAC: Codex Alimentarious Commission.
8) NQI: National Quality Institute.
9) MOU: Memorandum Understanding.
10) SSC: Scheme of Supervision of Control.
11) PVoC: Pre-Export Verification of Conformity.
12) COC: Certificate of Conformity.
13) UKAS: United Kingdom Accreditation Services.
14) R&D: Research and Development.
15) NMIs: National Metrology Institute.
16) ISO: International Organization for Standardization.
17) ARSO: Africa Organization for Standardization.
18) AFSEC: African Electro technical Commission.
19) IEC: International Electro technical Commission.
20) EASC: East African Standards Committee.
21) AFRIMETS: Intra African Metrology System.
22) SIM: Inter American Metrology System.
23) APMP: Asian Pacific Metrology Programme.

APPENDIX II (ANALYTICAL TECHNIQUES)

1) PH meter: A potentiometer with an electrode whose potential depends on the amount of

H+ ion present in the solution. (This is an example of an ion-selective electrode.) The pH
of the solution is measured throughout the titration, more accurately than with an
indicator; at the endpoint there will be a sudden change in the measured pH.
2) Conductivity: A measurement of ions in a solution. Ion concentration can change
significantly in a titration, which changes the conductivity. (For instance, during an acid–
base titration, the H+ and OH− ions react to form neutral H2O.) As total conductance
depends on all ions present in the solution and not all ions contribute equally (due to
mobility and ionic strength), predicting the change in conductivity is more difficult than
measuring it.

40
 3) Color change: In some reactions, the solution changes color without any added indicator.
This is often seen in redox titrations when the different oxidation states of the product
and reactant produce different colors.
4) Precipitation: If a reaction produces a solid, a precipitate will form during the titration. A
classic example is the reaction between Ag+ and Cl− to form the insoluble salt AgCl.
Cloudy precipitates usually make it difficult to determine the endpoint precisely. To
compensate, precipitation titrations often have to be done as "back" titrations (see below).
5) Isothermal titration calorimeter: An instrument that measures the heat produced or
consumed by the reaction to determine the endpoint. Used in biochemical titrations, such
as the determination of how substrates bind to enzymes.
6) Thermometric titrimetry: Differentiated from calorimetric titrimetry because the heat of
the reaction (as indicated by temperature rise or fall) is not used to determine the amount
of analyte in the sample solution. Instead, the endpoint is determined by the rate of
temperature change.
7) Spectroscopy: Used to measure the absorption of light by the solution during titration if
the spectrum of the reactant, titrant or product is known. The concentration of the
material can be determined by Beer's Law.
8) Amperometry: Measures the current produced by the titration reaction as a result of the
oxidation or reduction of the analyte. The endpoint is detected as a change in the current.
This method is most useful when the excess titrant can be reduced, as in the titration of
halides with Ag+.

APPENDIX III STANDADIZATION OF ANALYTICAL REAGENTS

A. Fehling’s stock solution A
Made by dissolving ~70 g CuSO4·5H2O in DDI water and diluting to 1.000 L
B. Fehling’s stock solution B
Made by dissolving ~350 g KNaC4H4O6·4H2O (potassium sodium tartrate tetrahydrate)
and 100.0 g NaOH in DDI water and diluting to 1.000 L. After sitting overnight, any
sediment is filtered out.
C. Dextrose standard solution
Weigh accurately ~2.5 g of dried dextrose (i.e. glucose). Dissolve in DDI water and
dilute to 500.0 mL in a volumetric flask. Mix well.
D. Standard invert sugar solution
Accurately weigh 4.75 g of sucrose, transfer with 90 mL of water to a 500 mL volumetric
flask, and dissolve. Add 5 mL of hydrochloric acid (specific gravity: 1.18) to the flask
and store for three days at room temperature (20–30°C). Then, dilute the solution to
volume with water and keep in a cool dark place. Transfer a 50 mL portion of the
solution to a 200 mL volumetric flask, neutralize with 1 mol/L sodium hydroxide
aqueous solution using phenolphthalein as an indicator, and dilute to volume with water.
Use the solution, as a standard invert sugar solution for the standardization of Fehling’s
solution.
E. 1% soluble starch solution
Weigh 1 g of soluble starch in a beaker, disperse in about 30 mL of water and transfer to
about 70 mL of boiling water while stirring continually. Heat the mixed solution until it
becomes transparent, add 5 g of sodium chloride and dissolve. After cooling, store the
prepared solution in a refrigerator.

41
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APENDIX IV
KEBS ORGANIZATIONAL STRUCTURE:

COAST REGION TESTING SERVICES QUALITY MANAGEMENT SYSTEM

Region manager Chief Manager Manager

QUALITY ASSUARANCE TESTING SERVICES INSPECTION SERVICES

SERVICES Manager in charge Manager
Manager

CHEMISTRY LABORATORY MICROBIOLOGY SAMPLE CENTER

Head of laboratory LABORATORY Head of sample control center

LABOLATORY ANALYSTS LABORATORY ANALYSTS SAMPLE CONTROL

Laboratory technicians Laboratory technicians CUSTOMER CARE
Sample control staff

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