Object: To perform assay of isonicotinic acid hydrazide.
References:
1. Tiwari A, Kumar R. " A Practical book of medicinal chemistry" first edition 2019,
Nirali Prakashan. Pune, page no. 67.
2. Krishna Murti, C. R. (1974) Antibiotics, eds. J. W. Corcoran and F. E. Hahn, (North
Holland: Springer Verlag), Page no.- 623.
Ingredients table:
S. Ingredients Quantity
No. Taken
1 Isonicotinic Acid Hydrazide 0.25mg.
2 Potassium Bromate 0.55mg.
3 Potassium Iodide 3.0gm.
4 HCL 23.0ml.
5 Potassium Bromide 0.2gm.
6 Starch Solution 3.0ml
Requirements:
Chemical: Isonicotinic acid hydrazide tablets, 0.0167M Potassium bromate, Potassium
iodide, HCL, Potassium bromide, 0.1 M Sodium thiosulphate, Starch Solution.
Glassware: Conical flask, Beaker, Pipette, measuring cylinder, Weighing balance, Burette
stand, Burette, Funnel, Glass rod.
Reaction:
Standardization:
KBrO3 + 6 KI + 6 HCL KBr + 6 KCL + 3 I2 + 3H2O
I2 + 2 Na2S2O3 Na2S4O6 + 2 NaI
Assay:
KBrO3 + 6 KI + 6 HCL 3Br2 + 6 HCL + 3 H2O
Theory:
It is also known as isonicotyl hydride (INH), isonicotyl hydrazine or isonicotino hydrazide or
INHA. Is a hydride of isonicotinic acid and a derivative of pyridine.
Chemical formula of isonicotinic acid hydrazide is C 6H7N3O and molecular weight of
isonicotic acid hydrazide is 137.13
IUPAC name: 4- Pyridine carboxylic acid hydrazide Or Isonicotinyl hydrazide Or
Pyridine- 4- carbohydrazide Or Isonicotinic acid hydrazide
Drug Category:
Isonicotinic acid hydrazide is an anti-tubercular drug.
It is assayed by the direct titration of Potassium bromate with the addition of potassium
bromide in the presence of an acid medium using hydrochloric acid.
During oxidation reaction liberated bromine reacts with isoniazid in an aqueous solution to
form isonicotinic acid. Azo dye methyl red indicator is used in this titration, decolorized the
red color is the endpoint.
Properties:
Appearance: White crystalline powder
Odor: Odorless
Solubility: Soluble in water and alcohol; slightly soluble in chloroform
Melting point: 171.40C
Therapeutic indications:
1. It is the most effective antitubercular drug.
2. With Rifamycin or Ethambutol, it is used for the treatment of pulmonary and
extrapulmonary tuberculosis.
3. It is used for atypical types of mycobacteria (like M. avium, M. kansasii, and M.
xeniopi).
Isonicotinic acid hydrazide (Isoniazid) is an extremely valuable antitubercular agent in
chemotherapy. Most of the titrimetric and colorimetric procedures reported in the literature
have received little attention, because the commonly used excipients interfere. The hydrazino
group is susceptible to oxidation, and most titrimetric procedures are based on this property.
Titration with potassium bromate is considered to be the best method.
Pharmacodynamics:
Isoniazid is a bactericidal agent active against organisms of the genus Mycobacterium,
specifically M. tuberculosis, M. bovis and M. kansasii. It is a highly specific agent,
ineffective against other microorganisms. Isoniazid is bactericidal when mycobacteria grow
rapidly and bacteriostatic when they grow slowly.
Mechanism of Action:
Isoniazid is a prodrug and must be activated by bacterial catalase. Specifically, activation is
associated with reduction of the mycobacterial ferric KatG catalase-peroxidase by hydrazine
and reaction with oxygen to form an oxyferrous enzyme complex. Once activated, isoniazid
inhibits the synthesis of mycoloic acids, an essential component of the bacterial cell wall. At
therapeutic levels isoniazid is bactericidal against actively growing intracellular and
extracellular Mycobacterium tuberculosis organisms. Specifically, isoniazid inhibits InhA,
the enoyl reductase from Mycobacterium tuberculosis, by forming a covalent adduct with the
NAD cofactor. It is the INH-NAD adduct that acts as a slow, tight-binding competitive
inhibitor of InhA.
Observation table:
Titration of isonicotinic acid hydrazide against 0.0167M Potassium bromate using methyl red
as an indicator.
S. No. Volume of Burette reading (ml.) Volume(V) of
isonicotinic Potassium bromate
acid hydrazide solution used
solution Initial Final (Mean)
1 20ml. 0.0 11.5ml.
2 20ml. 0.0 12.1ml.
3 20ml. 0.0 12.5ml. Mean =
11.5+12.1+ 12.5
3
= 12.03ml.
Concordant Value = 12.03ml
Calculation:
Molecular formula of potassium bromate: KBrO3
Molecular weight of potassium bromate: 167 g/mol
Molecular formula of isonicotinic acid hydrazide: C6H7N3O
Molecular weight of isonicotinic acid hydrazide: 137.14 g/mol
2
1 mole KBrO3 ≅ mole Br2 ≅ 1 mole isoniazid ≅ 137 gm isoniazid
3
2
1 mole KBrO3 ≅ 1 mole Br2 ≅ mole isoniazid ≅ 205 gm isoniazid
3
1000ml 1 mole KBrO3 = 205.5 gm isoniazid
1ml 1 mole KBrO3 = 0.2055 gm isoniazid
1ml 0.0167mole KBrO3 = 0.2055 × 0.0167 = 0.003431 gm isonicotinic acid hydrazide
So, 12.03 ml 0.00167M KBrO3 = 12.03 × 0.003431
= 0.0412 gm isonicotinic acid hydrazide (In 20 ml dil.
solution)
= 0.0412 × 5 (For 20×5 = 100ml)
= 0.206gm isonicotinic acid hydrazide (In 100ml solution)
0.206 ×100
% purity of isonicotinic acid hydrazide =
0.25
=82.4%
Procedure:
Preparation of 0.0167M potassium bromate solution:
About 0.55mg of potassium bromate was dissolved in water and made up to 100ml with
distilled water.
Standardization of 0.0167 M Potassium bromate:
1. 20 ml of the above solution was transferred in a glass stopper flask and 3 g of
Potassium iodide and followed by 3 ml of HCl was added.
2. Allow to stand for 5 min, titrate liberated Iodine with 0.1 M sodium thiosulphate
adding 3 ml of starch solution TS and the endpoint is approached.
Assay:
1. Twenty tablets were weighed accurately and pulverized.
2. A weighed quantity of the tablet power equivalent to 0.25 mg isonicotinic acid
hydrazide was transferred into a clean and dry 100 ml volumetric flask, and then
sufficient amount of water was added to produce 100 ml.’
3. 20 ml of the above solution was taken.
4. Then 100 ml of water, 20 ml of hydrochloric acid and 0.2 gm of Potassium bromide
were added.
5. Then titrated slowly with continuous shaking with 0.0167M potassium bromate using
0.05ml of methyl red as an indicator until the red color disappears’
6. The above is repeated twice.
7. Then calculate the % purity of isonicotinic acid hydrazide.
Result:
The assay of isonicotinic acid hydrazide has been performed.
The % purity of isonicotinic acid hydrazide was found to be = 82.4%
