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Index

Serial no Name of experiment Page no

01 Preparing a phosphate (Sorensen’s) buffer pH 7.4 01-04

02 Preparation of citro-phosphate buffer pH 4.5 at laboratory 10-12

03 Dissolution testing of Ibuprofen tablet 12-14

04 Dissolution testing of paracetamol tablet 15-19

Experiment no. : 01
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Name of the experiment: Preparing a phosphate (Sorensen’s) buffer pH 7.4

Objectives:
pH buffer is used to maintain pH of the solution in chemical analyses, syntheses, and calibration
of pH meters .Consequently no significant fluctuation of pH of solution in which buffer solution
is used occurs.
Principle:
pH is a measure of hydrogen ion concentration, a measure of the acidity & alkalinity of a
solution. pH usually range from 0 to 14.buffers are solutions which can resist changes in pH
when acid or alkali is added .
Example: Phosphate buffer, bicarbonate buffer
An acid will dissociate in water to a conjugate base & proton. Consequently, acids are typically
thought of as proton donors;

[HA] Ka [H+] + [A-] …………………. (1)

→

Acid proton conjugate base

Ka = ¿ ¿ ………………………………………………….. (2)
Ka is equilibrium constant
pH = -log [H+] or, 10-pH = [H+] …………………………………… (3)
PKa= -log (Ka) or, 10-pka = Ka …………………………………… (4)
By substituting equations 3& 4into equation 2, we can derive the Handerson-Hasselbalch
equation -
10-pka = 10-pH [A-]/ [HA]
[HA]/ [A-] =10-pH /10-pka
[HA]/ [A-] = 10Pka-pH
[Acid] / [base] = 10pka-pH
(According to Handerson–Hasselbalch buffer relationship, pH, pka & buffer component
concentration for a weak acid)
Here, Acid = proton donor (HA)
Base = conjugate base (A-)

 If the pka is greater than pH, there will be more of the acid form in the solution.
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 If the pka is equal to the pH, there will be an equal amount of acid and base in solution.
 If the pka is less than the pH, there will be more of the base form in the solution

The lower of the pka of the acid, the stronger acid

For base,
[B] + H2O = [B-H+] + [OH-]
Base water conjugate acid hydroxide ion
pOH = -log [OH-] ; pOH = 14-pH
pkb = -log [Kb] ; pka = 14-pkb
The higher the pka of a base, the stronger the Base.
Chemicals:

1. Sodium phosphate monobasic

2. Sodium phosphate dibasic
3. Citric acid
4. pH standardizing buffers

Equipment:

 pH meter
 mixing paste & magnetic stir bar
 100ml graduated cylinder
 250 ml volumetric flask
 250ml beaker
 50 ml burette, burette clamp
 Retort stand

Reagent preparation:
Preparing 250 ml of a 0.2M solution of sodium phosphate monobasic (SPM)
Step 1:
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1 M SPM contains - 119.98 g SPM

0.2M ” ” - 199.98×0.2
= 23.996 g
Step 2:
1000 ml of 0.2M solution contains - 23.996 g SPM
23.996
1 ml ” ” ” ” - g
1000
23.996× 250
250 ml ” ” ” ” - g
1000
=5.99 g SPM
Same procedure can be applied for others

Working procedure:
pH meter standardizing buffers (pH 7.4)
Part A: Preparing Sorensen’s buffer

 Preparing 250ml of a 0.2M solution of sodium phosphate monobasic

 Preparing 250ml of a 0.2M solution of sodium phosphate dibasic
 Loading a burette with 50ml of the 0.2M sodium phosphate monobasic solution
 Setting up the burette on a retort stand with a burette clamp.
 Measuring out 97ml of sodium phosphate dibasic using a 100ml graduated cylinder. pour
into a 250ml beaker.
 Calibrate pH meter with the standard solutions provided
 Mixing slowly
 Slowly titrate the sodium phosphate dibasic solution with the sodium phosphate
monobasic solution until attain a pH of 7.4
 Recording the volume of sodium phosphate monobasic added
 Transferring the titrated mixture to a 250ml beaker. Adding an equal volume of water to
create a 0.1 M Sorensen’s buffer.
 Sealing the buffer with the aluminum foil & store it for next lab

Precaution:
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 Proper safety concern should be maintained; such as hand gloves, mask apron etc.
 Volume adjustment should be done carefully. Solution should not cross meniscus level
 pH should be calibrated before taking the reading.
 Careful measurement of pH.

Experiment no. : 02
Name of the experiment: Preparation of citro-phosphate buffer pH 4.5 at laboratory.
Objectives:
 To understand the nature of buffer solution.
 To learn how to prepare buffer.
Principle:
Buffer solution is an aqueous solution of a weak acid and its conjugate base or vice versa which
has the capacity of showing resistance in pH change. Its pH change is very little; actually
negligible when small amount of acid or base is added. The main significance of buffer solution
is to maintain a certain pH of formulation.
Buffer solutions are of two types-
1. Acidic buffer: acidic buffer is the buffer solution having pH less than 7 and consist of a
weak acid and its one of salt. Examples ; a mixture of ethanoic acid and sodium ethanoate
in solution

CH3 COOH CH3 COO- + H+

CH3 COONa CH3 COO- + Na+
2. Alkaline buffer: an alkaline buffer is the buffer solution having pH greater than 7 and
consist of weak base and its one of salt. Examples; mixture of weak base ammonium
hydroxide and its salt in solution.

NH4OH NH4 + + OH-

NH4Cl NH4 + + Cl-
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The basic principle of preparing citro-phosphate buffer includes preparing a solution of weak
citric acid and salt disodium hydrogen orthophosphate in distilled water which is followed by pH
adjustment to 4.5 by using weak citric acid or NaOH base.
Materials and reagent
Materials:
a) 1000ml beaker
b) 1000ml volumetric flask
c) weight balance
d) pH meter
e) spatula
f) glass rod
Reagent:
1. Di-sodium hydrogen ortho-phosphate (Na2HPO4.2H2O)
2. Citric acid
3. Distilled water
4. NaOH solution (0.1 M)
Reagent preparation:
Follow rules given in experiment one to prepare the molar solution.
Procedure:
According to British Pharmacopeia-
 prepare 800 ml of distilled water in a 1000 ml of volumetric flask
 18.15gm of disodium hydrogen orthophosphate is added to the water with stirring
 9.336gm of citric acid is added to the solution & stir until dissolve
 Final solution volume is adjusted to 1000ml with distilled water
 pH is measured by a pH meter & adjusted to pH 4.5 with addition of citric acid solution
(0.1M)
Calculation: 50 ml 0.1 M citric acid solution is prepared 0.96gm citric acid is required which is
calculated by following-
Molecular weight of citric acid = 192.124gm
Now, 1000ml 1M citric acid contains 192.124gm
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192.124 ×50 ×0.1

∴50ml 0.1M citric acid contains =0.96gm
1000
Result: Finally we prepared 0.1M citro-phosphate buffer at pH 4.5
Discussion:
pH of final solution must be adjusted to 4.5. After pH adjustment, the solution must be kept in
proper conditions.
Precautions:
 Proper safety concern should be maintained; such as hand gloves, mask apron etc.
 Volume adjustment should be done carefully. Solution should not cross meniscus level
 pH should be calibrated before taking the reading

Experiment no. : 03
Experiment Name: Dissolution testing of Ibuprofen tablet.
Principle:
Dissolution is pharmaceutically defined as the rate of mass transfer from a solid surface into the
dissolution medium or solvent under standardized conditions of liquid/solid interface,
temperature and solvent composition. It is a dynamic property that changes with time and
explains the process by which a homogenous mixture of a solid or a liquid can be obtained in a
solvent.
In the pharmaceutical industry, drug dissolution testing is routinely used to provide critical in
vitro drug release information for both quality control purposes, to assess batch-to-batch
consistency of solid oral dosage forms such as tablets, and drug development to predict in vivo
drug release profiles. In vitro drug dissolution data generated from dissolution testing
experiments can be related to in vivo pharmacokinetic data by means of in vitro-in vivo
correlations (IVIVC). A well- established predictive IVIVC model can be very helpful for drug
formulation design and post- approval manufacturing changes.
The main objective of developing and evaluating an IVIVC is to establish the dissolution test as
a surrogate for human bioequivalence studies, as stated by the Food and Drug Administration.
Analytical data from drug dissolution testing are sufficient in many cases to establish safety and
efficacy of a drug product without in vivo tests, the dissolution testing which is conducted in
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dissolution apparatus must be able to provide accurate and reproductive results. Several
dissolution apparatuses exist. In United States Pharmacopeia (USP), there are four dissolution
apparatus standardized and specified they are:
1. USP Dissolution Apparatus 1 - Basket (37°C)
2. USP Dissolution Apparatus 2 - Paddle (37°C)
3. USP Dissolution Apparatus 3 - Reciprocating Cylinder (37°C)
4. USP Dissolution Apparatus 4 - Flow-Through Cell (37°C)
5. USP Dissolution Apparatus 2 is the most widely used apparatus among these
four.
Ibuprofen tablet is a painkiller. It is more potent than aspirin and paracetamol. It has anti-
inflammatory and anti- platelet action also. It is a potent inhibitors of cyclooxygenase. If onset of
action is 30 minutes and chemical formula is C13H18O2.It is a drug of NSAIDs class.

Figure: Structure of ibuprofen

Apparatus:
● Dissolution tester with paddle.
● Beaker 100 ml
● Volumetric flask
● Glass rod
● UV-Spectrometer
Chemical Ingredient:
1. Ibuprofen tablet (Flamex 400mg)
2. Distilled water
3. Buffer solution
4. Standard Ibuprofen solution (Profen 400mg)

Procedure:
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Preparation of buffer solution:

● 28.80 g of disodium hydrogen orthophosphate is weighted.
● 11.45 g of potassium dihydrogen orthophosphate is weighted.
● They are mixed in sufficient amount of water to create a clear solution of phosphate
buffer.
● Then their PH is measured to be 6.8 according to British pharmacopeia.

Dissolution test of Ibuprofen tablet:

 Each of dissolution vessels are filled with phosphate buffer solution (pH 6.8) to 900 ml
mask.
 Temperature of the dissolution medium checked to 37+- 5° C.
 One ibuprofen tablet is placed into each drug vessel.
 The stirring speed of paddle is set to 150 rpm and then the operation is started.
 After 30 minutes, 10ml sample is withdrawn from each vessel for analysis
 A standard solution of ibuprofen is prepared by diluting 10mg of standard ibuprofen to
50ml with dissolution media
 2 ml sample solution & 2ml of standard solution are diluted with 25ml of dissolution
medium in separate volumetric flask
 Finally absorption of solution is measured in a 1cm cell at 221nm wavelength of UV
spectrometer.
Result:

Absorbance of standard ibuprofen solution 1.

at wavelength 221 nm 2.
3.
Absorbance of the sample solution of 1.
ibuprofen at wavelength 221 nm 2.
3.

Precaution:
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 Disodium hydrogen orthophosphate & potassium dihydrogen phosphate should be

handled carefully
 pH buffer solution should be accurate
 Dissolution tester should be carefully handled
 Temperature & paddle rotation should be carefully settled
 We should wear protective wear during this whole procedure

Experiment no. : 04

Name of the experiment: Dissolution testing of Paracetamol tablet.

Principle:
Dissolution is pharmaceutically defined as the rate of mass transfer from a solid surface into the
dissolution medium or solvent under standardized conditions of liquid/solid interface,
temperature and solvent composition. It is a dynamic property that changes with time and
explains the process by which a homogenous mixture of a solid or a liquid can be obtained in a
solvent.
In the pharmaceutical industry, drug dissolution testing is routinely used to provide critical in
vitro drug release information for both quality control purposes, to assess batch-to-batch
consistency of solid oral dosage forms such as tablets, and drug development to predict in vivo
drug release profiles. In vitro drug dissolution data generated from dissolution testing
experiments can be related to in vivo pharmacokinetic data by means of in vitro-in vivo
correlations (IVIVC). A well- established predictive IVIVC model can be very helpful for drug
formulation design and post- approval manufacturing changes.
The main objective of developing and evaluating an IVIVC is to establish the dissolution test as
a surrogate for human bioequivalence studies, as stated by the Food and Drug Administration.
Analytical data from drug dissolution testing are sufficient in many cases to establish safety and
efficacy of a drug product without in vivo tests, the dissolution testing which is conducted in
dissolution apparatus must be able to provide accurate and reproductive results. Several
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dissolution apparatuses exist. In United States Pharmacopeia (USP), there are four dissolution
apparatus standardized and specified they are:

1. USP Dissolution Apparatus 1 - Basket (37°C)

2. USP Dissolution Apparatus 2 - Paddle (37°C)
3. USP Dissolution Apparatus 3 - Reciprocating Cylinder (37°C)
4. USP Dissolution Apparatus 4 - Flow-Through Cell (37°C)
5. USP Dissolution Apparatus 2 is the most widely used apparatus among these
four.

Apparatus:
● Dissolution tester.
● Beaker 100 ml
● Volumetric flask
● Glass rod
● UV-Spectrometer
Chemical Ingredient:

1. Paracetamol tablet ( Renova 500mg)

2. Distilled water
3. Buffer solution
4. Standard Paracetamol solution( Ace 500mg)

Procedure:
Preparation of buffer solution:
● 28.80 g of disodium hydrogen orthophosphate is weighted.
● 11.45 g of potassium dihydrogen orthophosphate is weighted.
● They are mixed in sufficient amount of water to create a clear solution of phosphate
buffer.
● Then their PH is measured to be 6.8 according to British pharmacopeia.
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Dissolution test of Paracetamol tablet:

 Each of dissolution vessels are filled with phosphate buffer solution (pH 6.8) to 900 ml
mask.
 Temperature of the dissolution medium checked to (37± 5)° C.
 One Paracetamol tablet is placed into each drug vessel.
 The stirring speed of paddle is set to 150 rpm and then the operation is started.
 After 30 minutes, 10ml sample is withdrawn from each vessel for analysis
 A standard solution of Paracetamol is prepared by diluting 10mg of standard Paracetamol
to 50ml with dissolution media
 2 ml sample solution & 2ml of standard solution are diluted with 25ml of dissolution
medium in separate volumetric flask
 Finally absorption of solution is measured in a 1cm cell at 243nm wavelength of UV
spectrometer.
Result:

Absorbance of standard paracetamol 1.

solution at wavelength 243 nm 2.
3.
Absorbance of the sample solution of 1.
paracetamol at wavelength 243 nm 2.
3.

Precaution:
 Disodium hydrogen orthophosphate & potassium dihydrogen phosphate should be
handled carefully
 pH buffer solution should be accurate
 Dissolution tester should be carefully handled
 Temperature & paddle rotation should be carefully settled
 We should wear protective wear during this whole procedure