INTRODUCTION

1st YEAR CHEMISTRY PRACTICAL COURSE (CH110), 2017

1. INTRODUCTION_____________________________________________
In the B. Tech. Chemical Laboratory (R. No. 09/004) you will meet a wide range of techniques from the simple to those using complex instrumentation - with which to study the properties of
materials. During your time here, we want you to gain an understanding of experimental
techniques and methods. It is not easy to work in a laboratory; making precise measurements
requires practice, experience and judgment. In the practical course you will, we hope, acquire
these skills, which are a vital preparation for later research.

2. WORKING HOURS___________________________________________
You will work in the laboratory during a 14-week period between 2 p.m. and 5:00 p.m. on
Monday, Tuesday, Thursday or Friday. As you cannot work beyond 5:30 p.m., it is necessary for
you to plan in advance so that you are done by the time when the laboratory closes, without
disrupting your experiment. From 9 a.m. to 1 p.m., and all day on Wednesday the laboratory
remains closed for experiments, but you are permitted to record melting points to characterize
the samples which you will synthesize.

3. STARTING AN EXPERIMENT_________________________________
Prepare fully for an experiment by reading in advance the instructions and any appropriate
background material. If the experiment lies in an area not yet covered in lectures, you may need
to do some further reading, but in any case proper preparation before starting an experiment is
always time well spent. You will normally work on experiments as one of a pair.
You have one day to complete each of experiments. If you are unable to complete an experiment
through illness or other reasonable cause, see Dr. Neeladri Das or Dr. Amit Kumar who will
make arrangements for you to do the experiment at some other time.

4. NOTEBOOKS________________________________________________
Keep a record of experimental data in a note book. You must hand this in to Mrs. Antara Garai/
Mr. Imtiaz Ahmad after completing the experiments. Write your name on the spine/front of the
note book and maintain an index of experiments completed. As you do an experiment, record
measurements and results in your note book. Ask an instructor to countersign your results during
the time you are in the laboratory, to indicate that he is satisfied with the way you are working.
(This must be done while you are working, not later. Instructors will not sign your data once the
experiment is over, and it may not be possible to evaluate an experiment if the data are not
countersigned.)

5. REPORT FORMAT___________________________________________
Your first semester chemistry experiments are present in the manual. The instructions will make
clear where you are to do the write-up. All work should be written neatly in correct English;
sloppy or grammatically incorrect reports may not be accepted.

Introduction
6. SUBMISSION OF REPORTS___________________________________
The reports of each chemistry experiment must be submitted as soon as you complete it; there
should be no need to take the note book away for further embellishment. Take your note book to
a technical superintendent in the laboratory and ask to receive it.

7. APPARATUS_________________________________________________
Major apparatus for experiments is set out in various places in the laboratory. Glassware and
required chemicals are set out on the benches; for other requirements, see the technical
superintendents. Clean the apparatus and bench area when you finish an experiment. Rinse out
glassware and leave on the benches. If the apparatus is faulty in any way, report this to a
technical superintendent, even if you caused the problem. If not reported, a problem may remain
undetected until the next students attempt the experiment. Do not leave unlabelled liquids in
beakers or flasks - these may cause a hazard.

8. WEB SITE___________________________________________________
A web site is available for the practical course in chemistry at the following URL:
http://172.16.1.6/download/manuals/Web/. This contains safety and other information which you
may find of value.

9. QUESTIONS?_________________________________________________
Help on any aspect of the practical course in the Department of Chemistry, IIT Patna is always
available from Dr. Neeladri Das (Room No. 04/, neeladri@iitp.ac.in, 0612-3028023) and Dr.
Amit Kumar (Room No. 04/, amitkt@iitp.ac.in, 0612-3028124).

10. SAFETY_____________________________________________________

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You will need to work carefully in the B.Tech. Laboratory of IIT Patna, as in any other
laboratory. Special hazards associated with an experiment are explained in the instructions, but
laboratories are dangerous places, so you must always be alert.
If in doubt - ASK
Our experiments are drafted to take the regulations into account. If any particularly unpleasant or
dangerous chemical is used during an experiment, detailed precautions are given in the
instructions. However, remember that the prime responsibility for safety rests with you. The
guidelines are given separately in the website; PLEASE READ THEM BEFORE STARTING
WORK! Before starting work for the first time, check the location of emergency facilities. There
are two exits from the laboratory - make sure you know where they are. The laboratory has a
nearby emergency shower - find it! Check that you know where fire extinguishers are kept in
case you need to use one in case you discover (or create!) a fire. If you suffer from any condition
which may lead to sudden illness, or if you must take, even for a short time, drugs which may
affect your performance in the laboratory, obtain a medical record sheet from the medical officer.
In case of emergency, it may help if this has been completed and stuck inside your laboratory
data book.

SAFETY REGULATIONS
Before you begin any work in the laboratory, you should read and understand the rules below. If you do
not understand a rule, insist on a satisfactory explanation from your instructor or a faculty member. Keep
a copy of this document in your lab manual and refer to the safety rules frequently.

I have read and understand the safety rules (1-33) below and have had an opportunity to
question my instructor about them. I agree to follow these regulations when I am in the
chemistry laboratory.
Date of Safety Training:

Your Signature:

Chemistry Department Safety Rules

Comments and Examples
GENERAL RULES
1. Do not work in the laboratory unless your 1. A qualified person must be present to:
instructor is present to supervise your work.
(a) see that only safe procedures are used, and
(b) provide immediate aid in case of an accident.
2. Do not carry out any unauthorized experiment.

2. Perform only those experimental steps in the

printed manual, or those given directly to you by
your instructor.

3. Do not work under any condition that you 3. If such a condition exists (e.g., overcrowded area,
believe to be unsafe for you or others.
unsafe actions by another student), report it
immediately to your instructor or to a faculty
member in charge.

EYE PROTECTION
4. Wear approved eye protection at all times in the 4. Eyes are very susceptible to chemical injury and
laboratory. Approved eye protection means safety must be fully protected all the time. Even when you
goggles with indirect venting.
are not working, a person nearby may be carrying
out a chemical procedure that might affect you.
5. Contact lenses should not be worn in the 5. All types of contact lenses may trap a chemical
laboratory.
against the eye tissue and cause permanent eye
damage.
Check with your instructor if needed.
6. Do not work with a chemical above or near your 6. For example, holding a beaker up to look at what
face.
is in the bottom, or filling a burette which is higher
than eye-level, can result in a splash down onto your
face.

HANDLING CHEMICALS
7. Many chemicals are toxic and/or corrosive.

7. Chemical reagents require careful handling.

Safety Regulations
8. Do not taste or ingest any chemical in the
laboratory.
Do not take or keep food or drink items at your lab
bench.

8. It may be toxic. Even NaCl may be contaminated

and be unsafe. For the same reason, you can not
bring food or drink into the laboratory, or eat in the
laboratory (no chewing gum, tobacco, candy, bottled
water or drinks, etc.)

9. Never pipet by mouth.

9. Drawing up a liquid (e.g., into a pipet) should be

done only with a rubber bulb or water aspirator.

10. Never pipet directly from a reagent bottle.

10. Transfer only necessary amount of liquid

reagents to a secondary container, such as a clean,
dry beaker.

11. Avoid skin contact with any chemical.

11. Keep the outside of reagent containers, all of

your equipment and the desk top, free from chemical
spills. Wear gloves if instructed to do so.

12. Do not inhale reagent fumes.

12. Odour tests are to be made only when

specifically directed to do so. Use a waving motion
of your hand to bring the vapour near your nose (this
is wafting).

13. Fume hoods must be used whenever toxic or 13. Use the hood when directed to do so. If fumes
corrosive vapours are released during the work you develop unexpectedly, cover the container and take it
are doing.
to the hood at once. Work with concentrated
hydrochloric, nitric, or acetic acids, or with bromine,
chlorine, or hydrogen sulphide should be done only
in a fume hood.
14. Alkalis are particularly corrosive. Contact with
NaOH and other alkaline (basic) chemicals must be
avoided. Work with solid sodium or potassium
hydroxide, or with solutions of these more
concentrated than 0.1 molar, should be carried out
only under the direct supervision of the instructor.

14. Strong bases must be handled with great caution

because they attack tissues so rapidly. Using 0.1 M
sodium hydroxide in a titration also requires great
care. Your instructor will demonstrate proper
techniques in handling a base in the laboratory.

15. Do not heat a test tube containing a liquid over 15. To heat a test tube, hold it in a beaker of hot
an open flame or directly on a hot plate.
water. Liquids heated over an open flame may erupt
violently and splash onto you or a person nearby.
16. Do not add water to a concentrated reagent, 16. Keep the mixture as dilute as possible; add the
especially concentrated sulfuric acid.
reagent to water. Addition of concentrated sulfuric
acid to water causes much heat formation and may
result in spattering of this corrosive reagent.

18. Dry all wet glassware before storing or 18. Keep a cloth towel for drying glassware and one
returning it. If you drop a beaker, for example, do for wiping your hands. You may bring a towel from
not reach to catch it since this usually leads to home.
getting cut.

17. When pouring a liquid, grasp each container so

that drops cannot contact your fingers. When using a
flexible polyethylene bottle, think first; do not pour
from it or squeeze it in any manner that might result
in a stream of liquid getting on you, or someone
nearby.

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17. Handle liquid reagent containers with care.

Safety Regulations
HANDLING GLASS TUBING AND SHARPS
19. Carry glass tubing and glass thermometers 19. On impact, glass tubing can snap and become a
only in an upright position.
dagger. Do not run with it (or any other chemical
equipment).
20. To insert glass tubing or a thermometer into a 20.
rubber stopper:
(a) If needed, fire polish the ends of the tubing.
(a) After heating glass tubing, set it aside in a place
where you will remember that it is hot.
(b) Lubricate the stopper hole with water or (b) There should not be more than two inches
glycerine.
between the stopper and your fingertips on the
tubing or thermometer.
(c) Insert the tubing cautiously, using a towel to (c) If handled improperly glass tubing can break and
protect your hands.
become razor sharp when inserted into a stopper.
21. All broken glass laboratory waste must be 21. Only paper products go into the regular bins.
placed into the special black glass disposal tubs in There are several bins available near the slab in the
the lab.
chemistry laboratory.
22. Waste "Sharps" must be placed in the special 22. Examples of "Sharps" are: syringes, syringe
black tub provided in the lab.
needles, razor blades, and scalpels.

IN CASE OF ACCIDENT
Learn the basic laboratory first-aid measures.
23. (a) If a chemical splashes into your eye, get 23. (a) Seconds count! Immediate removal of the
help immediately. Shout out, "I have chemical chemical is necessary to prevent possible damage to
the eye.
in my eye!"
23. (b) If someone nearby gets a chemical in
23. (b) A person who has just gotten a chemical in
his/her eye, you should: (1) shout for help from the
his/her eye usually is frightened, confused, and may
instructor, (2) provide help if the instructor is not
be unable to help himself/herself.
there immediately.
23. (c) Wash the eye thoroughly with a stream 23. (c) After thorough washing (15 minutes is the
of water from the eye wash fountain, or any recommended time) the affected person must be
taken to get professional medical attention.
other water source. Hold the eyelids open.
24. Any chemical that comes in contact with your 24. This is especially important for concentrated
skin should be washed off with water right away.
reagents and organic liquids.
25. Know the location of fire extinguishers, fire 25. Use a wet towel to extinguish a small fire or the
blankets, and safety showers, in case of fire. Keep fire blanket if a person's clothes catch fire.
acetone and any other organic liquid at least ten
feet from an open flame.

27. When the fire alarm sounds, stop what you are
doing and immediately exit the lab, go down the
stairs and exit the building. Wait outside for
instructions.

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27. Know the exit route from your lab.

26. Proceed cautiously when handling hot objects. 26. In case of burn, immerse in water immediately.
Use a towel as a hot pad when handling hot objects. Notify your instructor. Apply clean moist cloth or
Hot glass looks just like cold glass.
bandage. Seek medical attention if any question
about treatment.

Safety Regulations
28. Immediately report any accident to your 28. Cuts, burns, chemical burns, and inhalation or
instructor no matter how minor it may seem to you. ingestion of chemicals should be treated as soon as
possible by a professional medical person. Neither
students nor chemistry staffs are qualified to make
medical decisions.

CLOTHING IN THE LAB

29. You must be covered continuously from
shoulders to below the knees and must wear shoes
that cover your feet. Bare feet, sandals, shorts,
sleeveless shirts, short shirts, and short skirts are
UNSAFE and must not be worn to laboratory. For
fire safety, flammable materials, loose clothes, ties
should not be worn, and long hair should be tied
back.

29. Full coverage by (cotton) clothing and leather

shoes offers the best protection against chemical
spills and fire. Older clothing is advised, as is the use
of lab coats or aprons. Open-toed shoes or sandals
should not be worn in the chemistry laboratory.

CHEMICAL WASTE DISPOSAL

30. Only neutral aqueous solutions go down the
sink drain. Waste determinations and disposal are
done by faculty and staff. Check with your
instructor before disposing of any chemical.

30. All chemical waste is to be sorted into the

appropriate waste container and the identity and
amount must be logged onto the accompanying
inventory sheet. Check with your instructor for
specific details.

CLEANING GLASSWARE
31. Clean all the glassware which you have used
a. Clean general glassware by using detergent
and brush kept near sink.
b. Clean the glassware with stains by using
chromic acid mixture kept in fume
cupboard.
c. Rinse with water and keep on drying racks.

31. Although it may not seem that important,

cleaning glassware is one of the most important tasks
that you will do in lab contaminated glassware
(along with contaminated solvents) are the two
biggest causes of reactions going bad!

LEAVING LABORATORY
32. Clean your work bench with a damp sponge. 32. Leave the area safe for the next person.
Neutralize all acid spills with sodium bicarbonate
and wash with a wet sponge. Shut gas jets
completely. Wash your hands.

Page

33. Do not take any chemical out of the laboratory 33. You may be liable if another person is injured
for any reason. It is illegal!
by a chemical (or unauthorized equipment) that you
remove from the laboratory.

List of Experiments
EXP. EXPERIMENT TITLE
NO.
1 SPECTROPHOTMETRIC DETERMINATION OF STOICHIOMETRY OF A
COMPLEX BY JOBS METHOD
2 DETERMINATION OF CONCENTRATIONS OF HYDROCHLORIC ACID
AND ACETIC ACID IN A MIXTURE CONDUCTOMETRICALLY
3 DETERMINATION OF THE CONCENTRATION AND THE
DISSOCIATION CONSTANT OF A WEAK ACID (USING pH METER)
4 STANDARDISATION OF KMNO4 SOLUTION BY OXALIC ACID
5 DETERMINATION OF THE TOTAL HARDNESS OF WATER BY
COMPLEXOMETRIC TITRATION
6 DETERMINATION OF NUMBER OF COMPONENTS IN AN ORGANIC
MIXTURE AND Rf OF EACH COMPONENT USING THIN LAYER
CHROMATOGRAPHIC TECHNIQUE
7

SYNTHESIS AND STUDY OF SILVER NANOPARTICLES

8 QUANTITATIVE ESTIMATION OF PROTEIN BY BIURET METHOD

9 SYNTHESIS AND CHARACTERISATION OF
TRIS(ACETYLACETONATO)MANGANESE (III)

EXPERIMENT NO.: 1
Spectrophotometric Determination of Stoichiometry of a Complex
by Jobs Method
1. Aim__________________________________________________________
In this experiment you will determine the composition of the complex formed by thiocyanate
and ferric ions, using Job's method of continuous variation.
2. Safety________________________________________________________
Do not pipette by mouth. Solutions of Hydrochloric acid at which you will be using it
(0.004M) present a negligible risk. Ferric nitrate nonahydrate is harmful if swallowed or inhaled,
causes irritation to skin, eyes and respiratory tract and affects the liver. Sodium thiocyanate is
harmful if swallowed, may be harmful by inhalation or through skin contact and is an irritant.
3. Requirements_________________________________________________
UV-Vis Spectrophotometer/ Colorimeter, 10 test tubes, test tube stand, 2 burettes (50mL), 2
stands, 2 burette clamps, wash bottle with distilled water, a pair of cuvettes, 410-3 M Ferric
nitrate and 410-3M Sodium thiocyanate in 410-3 M Hydrochloric acid solution, tissue paper.

4. Theory_______________________________________________________
Job's method of continuous variation is a simple method for finding the stoichiometry of a
complex. It is most effective when only a single complex is formed. The success of a Jobs
method experiment depends upon how well Beers law is followed. The absorbance A of a
sample (the fraction of incident light that it absorbs) is defined in terms of the light incident upon
it, Io, and that transmitted, I.
A = log10 Io / I
The absorbance is related to concentration of the solution, c, through Beers law, which is
A = .c.l.

is the molar extinction coefficient for a species and l is the optical path length.
Jobs method can be used to find the stoichiometry of the compound formed by two reacting
species. The spectra of solutions containing both species in varying proportions are recorded,
each solution containing the same total reagent concentration. In this experiment the reagents are
Fe3+ and SCN-, so
[SCN] + [Fe3+] = constant
(1)
A wavelength at which the complex absorbs strongly is selected and the absorbance of each
solution at this wavelength is determined. As the concentration of one of the reactants, say the

Experiment Number:1
Fe3+, increases from zero, so does the amount of complex, so that the absorbance rises.
Absorbance reaches a maximum in the solution in which metal ion and ligand are in the same
ratio as in the complex, since this solution will contain the highest concentration of complex.
Further additions of metal ion are balanced by a reduction in the amount of ligand because of the
need to satisfy equation (1), so absorbance due to the complex then falls.
A plot of absorbance as a function of the amount of added ligand should give two straight
lines, provided Beers law is obeyed. Extrapolation of the two lines (in the direction away from
zero concentration of each species) gives the stoichiometry of the complex directly, since, where
the two lines cross, ligand and metal are in the correct proportion to give maximum complex
formation.

5. Procedure____________________________________________________
Prepare the following solutions which will be used throughout the experiment.
A solution of 410-3M Fe3+ made by dissolving appropriate amount of Fe(NO3)3.9H2O in
410-3M HCl.
A solution of 410-3M SCN made by dissolving appropriate amount of NaSCN in 410-3 M
HCl.
Keep these solutions in two different burettes, after washing and rinsing. Mix them well as
follows in the test tubes.
Test Tube No.
Volume of Fe(NO3)3
Volume of NaSCN

1
1mL
9mL

2
2mL
8mL

3
3mL
7mL

4
4mL
6mL

5
5mL
5mL

6
6mL
4mL

7
7mL
3mL

8
8mL
2mL

9
9mL
1mL

Allow to stand for half an hour. Take distilled water in a clean cuvette and calibrate colorimeter
at 440nm. Check whether it is showing zero absorbance at this wavelength. At 440nm determine
the absorbance of each of the solutions prepared as in the above mentioned table.
6. Experimental Readings_________________________________________
Temperature of the experimental solution =
Concentration of Fe(NO3)3.9H2O
=
Concentration of NaSCN
=
Volume of SCN
solution (B)
9mL
8mL
7mL
6mL
5mL
4mL

Ratio of
volumes(=A/B)
0.111
0.250
0.429
0.667
1.000
1.500

Absorbance

Volume of Fe3+
solution (A)
1mL
2mL
3mL
4mL
5mL
6mL

Page

Test tube
no.
1.
2.
3.
4.
5.
6.

Experiment Number:1
Test tube
no.
7.
8.
9.

Volume of Fe3+
solution (A)
7mL
8mL
9mL

Volume of SCN
solution (B)
3mL
2mL
1mL

Ratio of
volumes(=A/B)
2.333
4.000
9.000

Absorbance

7. Calculation___________________________________________________
Plot the absorbance as a function of the composition (that is ratio of volumes of Fe3+ and
volumes of SCN). The ratio of the reagents corresponding to the maximum absorption in the
curve gives the stoichiometry of the complex.

Page

10

8. Results_______________________________________________________
Ratio of metal ion and the thiocyanate ligand in the complex
=
Expected formula of the complex formed
=

EXPERIMENT NO.: 2
Determination of Concentrations of Hydrochloric acid and Acetic
acid in a mixture conductometrically
1. Aim__________________________________________________________
In this experiment you will determine the concentrations of hydrochloric acid and acetic acid
in a mixture by titrating it against a solution of sodium hydroxide of known concentrations and
measuring the conductivities with the help of a conductivity meter.
2. Safety_________________________________________________________________
Hydrochloric acid and acetic acids are corrosive in nature; do not allow the chemicals to
come in contact of your skin. Do not pipette by mouth.

3. Requirements_________________________________________________
Conductivity meter with electrode, beaker (50mL), semi micro burette (10mL), stand, burette
clamp, pipette (20mL), pipette pump, wash bottle with distilled water, glass rod, mixture of HCl
and CH3COOH of unknown concentration, 0.5M NaOH solution, tissue paper.

4. Theory_______________________________________________________
In a mixture of HCl and CH3COOH, the dissociation of feebly dissociated CH3COOH will be
further suppressed due to the dissociation of HCl. So when a solution of NaOH is added to this
mixture the result is the replacement of H+ of HCl by Na+ and the conductivity of the mixture
decreases linearly as the volume of NaOH added increases.
Near about the point where the entire H+ of HCl is removed, the dissociation of CH3COOH
slightly increases. With the addition of more NaOH, more of the completely dissociated Sodium
acetate is formed. As a result the first inflexion point in rounded. Thereafter the conductivity
again increased linearly. Near about the second inflexion the curve is again rounded due to the
hydrolysis of CH3COO.
After that, as more NaOH is added the conductivity increases rapidly and linearly due to the
accumulation of free Na+ and OH. As a result the experimental conductivity versus volume of
NaOH added curve is expected to be of the following form:

Coductivity in Sm-1

Experiment Number:2

Volume of NaOH added in mL

The concentrations of the acids can be calculated by using A and B and the well known formula
N1V1=N2V2

5. Procedure____________________________________________________
Wash the electrode repeatedly with distilled water. Wash and rinse the semimicro burette
with standard NaOH solution. Fill the burette with the standard NaOH solution. Wash the beaker
and wash and rinse the pipette with the experimental solution. Pipette out 40mL of the
experimental solution in the beaker and measure the conductivity of the solution. Add NaOH
solution from the semi micro burette in lots of 0.5mL. After each addition mix the solution
thoroughly and wait for 2 minutes to allow temperature equilibrium, and then note the
conductivity. Continue adding NaOH solution and taking the conductivity readings until the
three limbs of the graph are traced fully.
6. Experimental Readings_________________________________________
Temperature of the experimental solution =
Volume of the mixture taken
=
Strength of NaOH
=

0.0
0.5
1.0
1.5
2.0
2.5
3.0

12

Conductivity (in Sm-1)

Total Volume of NaOH added (in mL)

Page

Obs. No.
1.
2.
3.
4.
5.
6.
7.

Experiment Number:2
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.

3.5
4.0
4.5
5.0
5.5
6.0
6.5
7.0
7.5
8.0
8.5
9.0
9.5
10.0

7. Calculation___________________________________________________
Plot the conductivity value against the corresponding volume of NaOH added. A curve with
three limbs will be obtained. The point A corresponds to the neutralization of the strong acid;
hence the volume of NaOH corresponding to the point A is the volume required for the
neutralization of HCl. Similarly the point B corresponds to the volume of NaOH required to
neutralize all the acids. Hence, volume corresponding to (B-A) gives the NaOH required to
neutralize the weak acid.
Concentration of HCl

Concentration of CH3COOH =

) (

(
)
(

)
)

8. Results_______________________________________________________
Concentration of HCl in the mixture
=

Page

13

Concentration of CH3COOH in the mixture =

EXPERIMENT NO.: 3
Determination of the concentration and the dissociation constant of
a weak acid (using pH meter)
1. Aim__________________________________________________________
In this experiment you will determine the dissociation constant of acetic acid (a weak acid)
by using pH meter.

2. Safety________________________________________________________
Do not pipette by mouth. Solutions of both acetic acid and sodium hydroxide are corrosive in
nature and irritant. Special care must be taken to avoid the contact of the solutions especially
with eyes.

3. Requirements_________________________________________________
pH meter, 100mL beakers (2 nos.), pipette (20mL), pipette pump, burette (50mL), Sodium
hydroxide (0.1M), unknown acetic acid solution, distilled water in a wash bottle, stand, burette
clamp.

4. Theory_______________________________________________________
The aqueous solution of acetic acid dissociates as shown below

CH3COOH + H2O

CH3COO + H3O+

..(1)

An appreciable quantity of undissociated acetic acid remains in solution. For the general
weak acid HA the dissociation reaction and the dissociation constant expressions are:

HA(aq) + H2O(l)
=

!"

#$

%# $

%$

and

H3O+ + A..(2)
'-$
&' = & *+,
.- $

By titrating a weak acid with a strong base and recording the pH versus the volume of base
added, the dissociation constant of the weak acid can be obtained.

5. Procedure____________________________________________________
Wash the electrode repeatedly with distilled water. Wash and rinse the burette with
standard NaOH solution. Fill the burette with the standard NaOH solution.
Wash the beaker with distilled water and wash and rinse the pipette with the experimental
acetic acid solution. Pipette out 20mL of the unknown acetic acid solution in the beaker and
measure the pH of the solution by dipping the glass electrode in the solution. Add NaOH
solution from the burette in lots of 1mL. After each addition mix the solution thoroughly and
wait for 2 minutes to allow temperature equilibrium, and then note the pH. Continue adding

Experiment Number:3
NaOH solution and taking the conductivity readings until a sharp change in the pH is observed.
This is the approximate neutralization point.
Repeat the process explained in the previous paragraph, this time adding NaOH solution
in 2mL portions till the volume of NaOH solution added is at least 2mL less than required to
reach the approximate neutralization point. Then add 0.5mL portions of NaOH so that the sharp
change in pH could be noted. After the neutralization point add 2mL portion of NaOH at least for
4 readings and note the pH after each addition.
Plot pH versus volume of NaOH added. Determine the neutralization point from the
curve and the pH of the solution at half-neutralisation point. This pH value corresponds to the
pKa of the acid. Calculate the dissociation constant.

6. Experimental Readings_________________________________________
Concentration of NaOH
Volume of unknown acetic acid solution
Titration No.
1.
2.
3.

=
= 20 mL

Volume of NaOH added (in mL)

pH(observed)

7. Calculation___________________________________________________
Plot the pH as a function of the volume of NaOH solution added. Find the neutralization
point from the graph.
Let the volume of NaOH added till the neutralization point= V
Also at neutralization point, number of moles of Acetic acid=number of moles of NaOH
Therfore,
/+012034536+0 +7 95:' ;+*<=2 +7 95:'
/+012034536+0 +7 -12361 -168 =
;+*<=2 +7 -12361 5168
0.1A ;
/+012034536+0 +7 -12361 5168 =
20=C
Also, the pH at half of neutralization point (V/2) is equal to pKa.
So, Acid dissociation constant, Ka= antilog(-pKa)

8. Results_______________________________________________________

15

=
=

Page

Concentration of Supplied Acetic acid solution

Dissociation constant of Acetic acid

EXPERIMENT NO.: 4
Standardization of KMnO4 solution by Oxalic acid
1. Aim__________________________________________________________
In this experiment you will determine the molar concentration of Potassium permanganate
solution by titrating it against standard oxalic acid solution.

2. Safety________________________________________________________
Do not pipette by mouth. Solution of potassium permanganate is irritant. It is readily
absorbed through skin and is harmful if swallowed. Sulphuric acid is extremely corrosive, causes
serious burns, it is highly toxic, harmful by inhalation, ingestion and through skin contact.
Ingestion may be fatal. Skin contact can lead to extensive and severe burns. Chronic exposure
may result in lung damage and possibly cancer. Oxalic acid is harmful if swallowed and in
contact with the skin. May cause burns on contact with the eyes. Special care must be taken to
avoid the contact of the solutions specially with eyes.

3. Requirements_________________________________________________
Burette (50mL), stand, burette clamp, 250ml conical flask (2 nos.), 10ml pipette, test tube,
pipette bulb, water bath, 0.1 (N) Oxalic acid in reagent bottle, unknown KMnO4 in volumetric
flask, H2SO4 (1:2) in a reagent bottle.

4. Theory_______________________________________________________
Commercially available potassium permanganate generally contains impurity. Thus it cannot
be used as primary standard. In order to make standard KMnO4 solution, it requires to be
standardized by a primary standard.
Equivalent weight of KMnO4 = [2 KMnO4/10] = 31.606,
which can be derived from the equation 2 KMnO4 = K2O.2MnO ;
i.e. Mn7+ +5e
Mn2+

5. Procedure____________________________________________________
Rinse a clean burette (capacity 50 mL) thrice with 5 mL portions of the KMnO4 solution. Fill
up the burette with KMnO4 solution up to the zero mark and note the upper meniscus. Examine
that the jet of the burette is completely filled with the solution and no air bubble is left behind.
Pipette out 10 mL of standard oxalic acid (0.1 M) into a 250 mL conical flask. Add 810mL of H2SO4 (1:2) and then add boiling water to dilute it to about 100 mL. Now titrate the
solution with KMnO4 solution. At first add KMnO4 solution small quantities at a time with
stirring; the pink colour of KMnO4 will take some time to discharge its colour at the beginning.
So initial addition should be very slow, when some KMnO4 solution has been added, the pink

Experiment Number:4
colour will be discharged quickly. Now add KMnO4 solution more quickly with stirring. Near
the end point when the rate of disappearance of the pink colour slows down, add KMnO4
solution drop wise with stirring until one drop makes the whole solution pink (the pink colour
persists for 30 seconds, after which the colour may be discharged again)
Note the volume of KMnO4 solution added. Repeat the operation thrice.

6. Experimental Readings_________________________________________
Concentration of Oxalic acid
Volume of Oxalic acid solution
Observation
No.
1.
2.
3.
4.

Volume of Oxalic acid

solution taken
25mL
25mL
25mL
25mL

= 0.1N
= 10 mL
Burette Readings
Initial
Final
Difference

Concurrent
Reading

7. Calculation___________________________________________________
Let volume of KMnO4 solution = VmL
Concentration of oxalic acid = N1
Therefore, Concentration of KMnO4 solution=
=

D EF

F.E EF

(N)

8. Results_______________________________________________________

Page

17

Concentration of Supplied KMnO4 solution =

EXPERIMENT NO.: 5
Determination of the total hardness of water by Complexometric
Titration
1. Aim__________________________________________________________
In this experiment you will determine the total hardness of normal tap water available in
laboratory by complexometric titration method.

2. Safety________________________________________________________
Do not pipette by mouth. Solution of EDTA is an eye irritant. Eriochrome Black-T is harmful
if swallowed. Avoid its contact with eyes, skin and cloth. Eye contact with ammonia containing
buffer can lead to serious eye damage. It is toxic if swallowed and harmful if inhaled.

3. Requirements_________________________________________________
50mL burette, stand, burette clamp, 25mL pipette, pipette bulb, 250mL conical flask,
Eriochrome Black-T in dropping bottle, standard EDTA solution in volumetric flask, wash
bottle, NH4Cl-NH4OH buffer in reagent bottle, 5mL measuring cylinder, conical funnel.

4. Theory_______________________________________________________
The hardness of water is generally due to dissolved calcium and magnesium salts mainly as
bicarbonates and can be determined by complexometric titration.
Complexometric titrations are those titrations in which the concentration of a metal ion is
determined titrimetrically by forming complex with a strong multidentate chelating ligand (e.g.
ethylenediaminetetraacetate = EDTA). In these kind of titrations, a metal ion is in the form of a
complex with an organic dye (indicator) having a characteristic colour. During the course of
titration, the ligand forms complex with the metal ion thereby setting the dye free from metal ion.
At the end point, when all the dye molecules are set free, the solution assumes the colour
characteristic of the dye.
When calcium ions are titrated with di-sodium salt of EDTA, a relatively stable complex is
formed.
Na2H2Y = 2Na+ + H2Y2Ca2+ + H2Y2- = CaY2- + 2H+
This titration is performed in alkaline buffer medium. In alkaline medium, the reaction is
driven towards the forward direction in which the H+ ions are consumed. Erichrome black-T is

Experiment Number:5
used as indicator. It is red when it forms complex with Ca2+ and blue in the free state. The end
point thus is indicated by the blue colour of the solution. The hardness of water is expressed in
parts per million unit.
1 mL of 0.01 M EDTA = 1 mg of CaCO3

5. Procedure____________________________________________________
Fill the cleaned burette with standard EDTA solution (supplied) after rinsing with same.
Pipette out 25 mL of water sample (supplied) into a 250 mL conical flask. Add 5 mL buffer
solution followed by 4 to 5 drops of Erichrome black-T solution. Titrate the solution with EDTA
solution from the burette till the colour of the solution changes purple red to blue. Note down the
volume of EDTA consumed.
Repeat the steps explained in the previous paragraph, till three concurrent readings are
obtained.

6. Experimental Readings_________________________________________
Temperature of the experimental solution
Strength of EDTA
Volume of water sample taken
Density of water
Observation
No.
5.
6.
7.
8.

Volume of Water
sample taken
25mL
25mL
25mL
25mL

=
=
=
=

Initial

Burette Readings
Final
Difference

Concurrent
Reading

7. Calculation___________________________________________________
Let the volume of EDTA required for 25mL water is V, and density of water is d g/mL
Amount of CaCO3 in 25mL of water = V mg

/+012034536+0 +7 /5/:G 60 H<&&*628 I5324(60 &&=) =

;
10G
25 8

8. Results_______________________________________________________

Page

19

Concentration of CaCO3 in sample of water =

EXPERIMENT NO.: 6
Determination of the number of components in an organic mixture
and Rf of each component using thin layer chromatographic
technique
1. Aim__________________________________________________________
In this experiment you will learn the technique of thin layer chromatography and determine
the number of components in a mixture and Rf values of few compounds.

2. Safety________________________________________________________
Dust of silica gel affects respiratory system and causes irritation to eyes. Wash hands (or
exposed area) with soap and water after use. Ethyl acetate is irritant (specially to eyes) and
harmful if swallowed. Its vapours may cause drowsiness. It is highly inflammable.

3. Requirements_________________________________________________
250mL beakers of tall form (4 nos.), petri dishes (4nos.), test tubes (3 nos.), capillary tubes
(3nos.), test tube stand, TLC Plates coated with slurry of silica gel (4nos.), unknown organic
compounds, solvents (S1, S2 and S3), Iodine crystals, 5mL measuring cylinder.

4. Theory_______________________________________________________
Liquid chromatography is a highly efficient technique used to identify number of
components in a mixture as well as to monitor the progress of a reaction. By this method, sharper
separation and detection of very small quantity of components can be achieved in a short time. In
thin layer chromatography (TLC), the stationary phase (silica gel or alumina mixed with a binder
like CuSO4) is mixed with a solvent to form a slurry and in then applied on a glass or plastic
plate to make a coating. The prepared thin layer on glass is called a chromoplate.
A small capillary tube is dipped inside the mixture and then taken out. A spot is made, at one
end of the chromoplate, with the mixture remaining inside the capillary tube. The chromoplate is
then placed inside a chamber containing the solvent. The solvent (eluent) moves up, along the
chromoplate, by capillary action through the spot. Solvent used for this purpose should be low
boiling so that the plate can dry quickly after removing from the chamber.
The ratio of the distance travelled by a compound (from the spot) to travelled by the solvent
front is called Rf value of the compound.

R =

LM

LM

N O PQ R

N O PQ R M N

The Rf value of a compound is a characteristic property of the compound for the particular
solvent used. This property is used to identify each component in a mixture. In general, a polar

Experiment Number:6
compound has a lower Rf than a non polar one. Written below are the trends of Rf values of some
compounds with increasing polar character.
Saturated hydrocarbon > alkenes, alkynes, aromatic hydrocarbons > esters, aldehydes and
ketones > amines, alcohols, thiols > carboxylic acids.
For a particular compound, the Rf value increases with increasing polarity of the solvent. The
polarity orders of some commonly used solvents are:
Hexane< Cyclohexane< Carbon tetrachloride< Trichloroethylene < Toluene,
Dichloromethane < Chloroform < Diethyl ether < Ethyl acetate < Acetone < Propanol < Ethanol
< Methanol.
To detect a spot on the plate several methods, such as visualisation under UV lamp (if the
compound is UV active), iodine vapour chamber, spraying with 10% sulfuric acid in methanol
etc. are used.

5. Procedure____________________________________________________

Page

21

Take a 510 cm coated TLC plate (Chromoplate). Make three spots 1cm above from the
bottom of the plate and make 1, 2 and 3 respectively on the top of the plate with a pin. Take 10
mg each of compounds A and B into two separate test tubes and dissolve in 2mL ethyl acetate. In
the third test tube, mix equal amounts of the solutions of A and B (1mL each). Take a thin
capillary tube and dip into the solution of A. Transfer the liquid from the loaded capillary to spot
number 1 by just touching the plate for a second. Repeat it for compound B into spot number 2
and for the mixture (A+B) for spot number 3. Put the TLC plate into a supplied solvent chamber
containing 50mL of solvent (labelled as S1), until the solvent level rises to 1 cm less than the top
of the plate. Remove the plate, quickly mark the upper limit of the solvent front with a pin and
allow it to dry at room temperature. Put the dry TLC plate into a chamber containing a few
crystals of Iodine. Leave it for a few minutes until Iodine vapours are adsorbed by the
compounds and distinct spots are visible on the plate. Remove the TLC plates from the chamber
and carefully encircle each darkened spot. Mark their mid points and measure the distance
between the midpoint of each spot and the starting point (that is spots 1, 2 and 3). Find out the Rf
values of the spots.
Repeat the above procedure for solvent systems S2 and S3 respectively. Find out the number
of components in A and B individually and compare with the number of components in the
mixture. Remember that the number of spots generated from each starting spot is equal to the
number of components in the mixture. You need to find out which one of the solvent system
gives better resolution. Determine the Rf value of each component.

Experiment Number:6
6. Experimental Readings_________________________________________
1

S1
Sovent
System

Solvent
front

S2
Distance travelled by
Alone
Mixture
A
B
A
B

S3
Rf
Alone
A

Mixture
A
B

S1
S2
S3

=
=
=
=
=
=
=

Page

Number of components in the mixture

Rf of A in S1
Rf of A in S2
Rf of A in S3
Rf of B in S1
Rf of B in S2
Rf of B in S3

22

7. Results_______________________________________________________

EXPERIMENT NO.: 7
Synthesis and study of Silver Nanoparticles
1. Aim__________________________________________________________
In this experiment you will synthesize silver nano particles and study its characteristics like
colour, spectrum and stability.

2. Safety________________________________________________________
AgNO3 is poisonous, skin and eye irritant and can lead to deposition of black silver stains on
the skin. Sodium boro hydride is toxic by ingestion and harmful if inhaled and in contact with
skin. The dilute solutions of both reagents do not cause any harm.

3. Requirements_________________________________________________
Silver Nitrate solution ( 10 mL of 1 mM), NaBH4 solution (ice cooled) (30 mL of 2 mM),
magnetic plate & magnetic bar, 50mL round bottomed flask, 50mL measuring cylinder, 100mL
Beaker (2 nos.), dropper (1 no.), test tube (2 nos.), test tube stand (1 no.), spectrophotometer
(Common to all).

4. Theory_______________________________________________________
Nanotechnology deals with processes that take place on nanometer scale, that is, from
approximately 1 to 100 nm. Properties of metal nanoparticles are different from those of bulk
materials made from the same atoms. For example, the striking effect of nanoparticles on color
has been known since antiquity when tiny metal particles were used to colour glass in church
windows. Silver particles stained the glass yellow, while gold particles were used to make rubycolored glass. In performing the experiment described here, you will observe the bright yellow
colour of silver nanoparticles compared to colorless silver nitrate solution and metallic bulk
silver.
The chemical reaction is the sodium borohydride reduction of silver nitrate:
AgNO3 + NaBH4

Ag + 1/2H2 + 1/2B2H6 + NaNO3

The method produces 12 2 nm particles. The plasmon absorbance is near 400 nm. The
wavelength of maximum absoption, max, depends upon the size of the particle. The particle size
and corresponding max are given below.

Experiment Number:7
Sl. No.
1.
2.
3.

Paricle size (in nm)

10-14
35-50
60-80

max (in nm)

395-405
420
438

5. Procedure____________________________________________________
Clean a 50mL round bottomed flask and a magnetic stirring bar. Take 30mL of 2.0mM
NaBH4 solution in the round bottomed flask and chill it in an ice bath. Add 10mL 1.0 mM silver
nitrate drop wise (about 1 drop/ second) to the solution taken in the round bottomed flask, kept
over magnetic stir plate. Continue stirring the reaction mixture vigorously. The solution will turn
light yellow after the addition of 2mL of silver nitrate and a brighter yellow when all of the silver
nitrate will be added. The entire addition should take about three minutes, after which stop the
stirring and remove the stirring bar. Note down the colour of the colloid.
Store the colloid in a transparent vial and observe the time for breakdown of colloid. If the
colloid doesnt breakdown even after 45 minutes, it may be considered stable. Meanwhile, take
about 3mL of the colloid in a clean cuvette and observe the Spectrum in 350-600nm range. Note
down the wavelength of maximum absorbance, max, and predict the approximate size of the
particles.

6. Experimental Readings_________________________________________

24

=
=
=

Page

Colour of the colloid

max
Corrosponding size of the particles

EXPERIMENT NO.: 8
Quantitative Estimation of Protein by Biuret Method
1. Aim__________________________________________________________
In this experiment you will determine the amount of protein by using Biuret method.

2. Safety________________________________________________________
Biuret reagent is a strong base (pH >12) and toxic due to the copper. Whenever you work
with Biuret Reagent solutions you must: wear eye protection, avoid skin contact, clean up any
spills, follow directions for proper disposal.
The protein standard solutions containing only BSA are harmless.

3. Requirements_________________________________________________
Test tubes (6 nos.), test tube stand, a pair of cuvettes, semi micro burette (2 nos.), 50 mL
burette, stands (3 nos.), burette clamps (3 nos.), UV-Vis spectrophotometer/Colorimeter, Biuret
solution in reagent bottle, BSA solution.

4. Theory_______________________________________________________
Proteins are known to be polymers of amino acids, which are linked serially by peptide bonds
(-CO-NH-). The Biuret method is the simplest method for estimating protein concentrations, and
is based on the fact that the CO-NH- group (present in all proteins) can form a colored complex
with copper(II) ions in an alkaline medium, whose absorbance value reached a maximum at 540
nm. The intensity of the color produced is proportional to the number of peptide bonds present in
the sample. The cupric ions cause chelation of the peptide bonds of proteins under alkaline
conditions resulting in the production of purple colored complex. Thus, molecules containing 2
or more peptide bonds associate with the cupric ions to form a coordination complex that imparts
a purple color to the solution with max = 540 nm. The purple color of the complex can be
measured independently of the blue color of the reagent itself with a spectrophotometer or
colorimeter.

Experiment Number:8

5. Procedure____________________________________________________
Preparation of Biuret Reagent
A formula for biuret reagent is (per liter final volume) 9g Sodium potassium tartrate (f.w.
282.22), 3g CuSO4.5H2O (f.w. 249.68), 5g Potassium iodide (f.w. 166.0), all dissolved in order
in 400 ml 0.2 M NaOH (f.w. 40.0) before bringing to final volume of 1 liter. The volume can be
scaled up or scaled down according to requirement. Discard a black precipitate, if formed.

Assay
Warm up the colorimeter 15 min. before use. Prepare stock solution of Bovine Serum
Albumin (BSA) containing 5 mg/ml in distilled water or millipore water. Set up a series of BSA
solutions in clean test tubes using 0.1, 0.2, 0.4, 0.6, 0.8 and 1 ml of the stock solution and using
distilled water, make up the volume to 4 ml. For the blank use distilled water. Prepare varying
solutions of the unknown as indicated in the table and take 4 ml of the diluted solution separately
in test tubes. Add 6 ml of Biuret solution in each tube, mix and incubate at 37C or room
temperature for 10 minutes. The appearance of the purple color indicates the presence of a
protein in the sample. Read the absorbance of the individual solutions at 540 nm after setting to
zero absorbance value with the blank. Calculate the protein concentration in the test sample by
comparing its absorbance value with those of the standard curve.

6. Experimental Readings_________________________________________
=

Concentration BSA

stock Distilled

Biuret

No.

ug/ml

solution(ml)

water (ml)

(ml)

125

0.1

3.9

6.0

250

0.2

3.8

6.0

soln. A540

26

Serial

Page

Temperature of the experimental solution

Experiment Number:8
3

500

0.4

3.6

6.0

750

0.6

3.4

6.0

1000

0.8

3.2

6.0

1250

1.0

3.0

6.0

UNKNOWN SAMPLE
7

Unknown

Unknown

Unknown

6.0

7. Calculation___________________________________________________
Prepare a standard curve of absorbance versus micrograms protein (or vice versa), and
determine amounts from the curve. Determine concentrations of original samples from the
amount of protein.

8. Results_______________________________________________________

Page

27

Concentration of BSA in unknown solution =

EXPERIMENT NO.: 9
Synthesis and Characterization of Tris(acetylacetonato)
Manganese (III)
1. Aim__________________________________________________________
In this experiment you will synthesize tris(acetylacetonato)manganese(III), characterize it by
determining melting point.

2. Safety________________________________________________________
KMnO4 is harmful if swallowed, irritant and readily absorbed through skin. Acetylacetone is
harmful if swallowed or inhaled and is irritant. It is flammable therefore it should be kept away
from flame.

3. Requirements_________________________________________________
100mL beaker, buchner funnel setup, watch glass, 5mL measuring cylinder, dropper, spatula,
glass rod, capillary tube, wash bottle, thermometer, melting point apparatus, filter paper,
KMnO4, Acetylacetone.

4. Theory_______________________________________________________
The complex manganese tris(acetylacetonate) or tris(acetylacetonato)manganese(III) is
synthesised by reacting KMnO4 with acetylacetone.
KMnO4+CH3COCH2COCH3

[Mn(acac)3]+(CH3CO)2CH-CH(COCH 3)2+K++H2O

The reaction is based on electron-transfer between Mn(VII) and acacH. No buffer is required.
The compound is obtained as dark brown-black crystals. Mn(acac)3 is monomeric in nature.
The oxidation state of manganese in the compound can be determined iodometrically by
reduction of a known amount of the compound with acidified potassium iodide solution followed
by titration of the liberated iodine with standard sodium thiosulphate solution. The redox titration
can be also used for quantitative determination of manganese content of the compound.

5. Procedure____________________________________________________
Take finely powdered KMnO4 (0.5 g, 3.2 mmol) in 5 mL of distilled water into a 100 mL
beaker. Dissolve it by slightly warming on a water-bath. To this solution add acetylacetone (2.3
mL, 22 mmol) in several portions in dropwise with time to time swirling. Stir the reaction
mixture for 5 min. Keep it again on a hot water-bath until dark brown-black shiny crystals

Experiment Number:9
appear from the reaction mixture. Remove the reaction mixture from the water-bath and allow
the mixture to cool for 20 min. Filter the reaction mixture with suction on a Buchner funnel and
wash the solid material with small amount of cold acetylacetone-water mixture (1:1, v/v, 1 mL).
Collect the dark-brown black crystals and dry in vacuo. Weigh the solid material. Calculate its
yield. Find out the melting point of the compound and submit the sample.

6. Experimental Readings_________________________________________
Weight of the sample prepared
Melting point of the compound

=
=

7. Calculation___________________________________________________
Calculate the theoretical yield, and find out the percentage yield of the product.

8. Results_______________________________________________________

29

=
=
=

Page

Weight of the sample

Percentage yield
Melting point