Development and Validation of Four New Uv spectrometric Methods for the Determination of Isoniazid (Inh) In Pure and Tablet Dosage Forms

Development and Validation of Four New Uv spectrometric Methods for the Determination of Isoniazid (Inh) In Pure and Tablet Dosage Forms.

ABSTRACT

Tuberculosis (TB) is a serious infectious disease which is on increase since the emergence of the human immunodeficiency virus (HIV) leading to TB/HIV co-infection, causing complications and treatment failure. Isoniazid is a first-line drug for the treatment of TB therefore it is important to develop methods that will monitor its quality.

In this study, four rapid, simple, relatively accurate, and precise spectrophotometric methods for the quantitative determination of isoniazid in pure and tablet dosage form were developed and validated.

Methods 1 and 2 were based on dissolving standard isoniazid powder in distilled water and methanol respectively and scanning using UV-Vis spectrophotometer at a wavelength range of 200 – 400 nm to determine the wavelength of maximum absorption (λmax).

Method 3 was based on condensation reaction between isoniazid and 2,4-dinitrophenyl hydrazine (2,4 DNPH) and heating under acidic conditions using 2 mL of 85 % hydrochloric acid (HCl) to form an orange-red hydrazone which was allowed to stand for few minutes for complete color development.

Method 4 was based on the diazotization of isoniazid with sodium nitrite (NaNO2) and coupling with paraaminotoluidine under acidic conditions (2 % HCl) and gently heating to give a coloured chromogen (diazo dye) which was allowed to stand for few minutes for complete color development. 

TABLE OF CONTENT

Cover page.…………………………………………………………………………………………i
Title page ……………………………………………………………………………………………ii
Declaration………………………………………………….…………………………………….iii
Certification………………………………………………………………………………………iv
Acknowledgement…………………………………………………………………………………v
Abstract …………………………………………………………………………..………………vii
Table of contents………………………………………………………………..………………..ix
List of tables……………………………………………………………………..………………xiii
List of figures……………………………………………………………………………………xiv
List of appendices……………………………………………………………………………..xv
Abbreviation………………………………………………………..…………………………..xvi

CHAPTER ONE

1INTRODUCTION……….………………………………….…….…….……………………….1
1.1Theoretical Frame Work……………………………………….…………..……………….2
1.2 Statement of the Research …………………………………………………….……………..3
1.3Justification of the Research …………..……………………………….…….……………..4
1.4 Aim of the Research ……………………………………………….………….……………..4
1.4.1 Specific objectives………………………………………..……….….…….……………….4

CHAPTER TWO…………………………………………………………….……………………….5
2.0 LITERATURE REVIEW………………………………………………….………………..5
2.1 Chemistry of Isoniazid…………………….…..…………..…………..……….……………..5
2.1.1 Mechanism of action of isoniazid…………………………….…………..….…………………..5
2.1.2 Uses of isoniazid………………………………………………..……………………………6
2.2Chemistry of Schiff Base…………………………………….……………..…………………….6
2.3 Chemistry of Diazo dye……………………………………………………………………………………………..7
2.4 Analytical Methods for the Determination of Isoniazid…………………………………………8
2.4.1 UV spectrophotometric method using distilled water and methanol…………………………8
2.4.2 Colourometric method using 2,4-dinitrophenyl hydrazine and diazotization…………………..8
2.4.3Compedial method of analysis……………………………………………….………………8
2.4.4 Noncompedial methods of analysis.…………………………………………………………….8

CHAPTER THREE …………………………………………………………..………………..15
3.0 MATERIALS AND METHODS…………………………………….…..…………………15
3.1 Materials………………………………………………………….………..……………………15
3.1.1 Isoniazid analytical grade standard powder and tablet…………..….…..………………..15
3.1.2 Chemicals and reagents……………………………………….…………..………………..15
3.1.3 Glasswares and accessories………………………….………….….…..………………….15
3.1.4 Equipment……………………………………………..…………….……………………..16
3.2 Methods……………………..…………………………’…………………………………..16
3.2.1 Quality control of isoniazidstandard powder and tablets……..………..…………………16
3.2.2 Selection of solvents……………………………….……………….………………………18
3.2.3 Preparation of solutions and reagents..…………………….…..……………………………….18
3.2.4 UV spectrophotometric method using distilled water………………..……..………………18
3.2.5 UV spectrophotometric method using methanol ……………………….….………………19
3.2.6 Development of colorimetry method using 2,4dinitrophenylhydrazine and
optimization… ……………………………………………………………….………………….20
3.2.7 Development of colorimetric method using paraaminotoluidine (diazotization) and
optimization…………………………………………………………………..……………………..23
3.2.8Validation of the analytical methods………….…………………….……………………….25
3.2.9 Statistical analysis…………………………………………………………..…….………..28

CHAPTER FOUR…………………………………………………………………..…………29
4.0 RESULTS………………………………………………………………….……….……….29
4.1 Quality Control of Pure Isoniazid Standard Powder and Tablets………………………29
4.2Method Development………..…………………………….…………………………………33
4.2.1 UV spectrophotometric method using water and methanol………………………………..33
4.2.2 Colorometeric methods using 2,4dinitrophenylhydrazine and by diazotization………….33

CHAPTER FIVE……………………………………………………….…..….…….…………43
DISCUSSION………………………………………….……….…………..……..…………….43
CHAPTER SIX………………………………….…………………………..……………………..46
6.0 CONCLUSION AND RECOMMENDATIONS…………………………..……………..46
6.1 Conclusion…………………………………………………………………..…….…………46
6.2 Recommendations……………………………………..……………………………………46

REFERENCES…………………….…………………………………….……….…………….47

INTRODUCTION

Isoniazid (INH) also known as isonicotinyl, is a synthetic of isonicotinic hydrazide, chemically known as pyridine-4-carboxylic acid hydrazide (BP, 2009).

It is first-line drug in the of pulmonary and extrapulmonary in combination with other anti-tubercular drugs such as rifampicin, ethambutol, pyrazinamide, and streptomycin to avoid the development of resistance(WHO, 2010, Niraimathi et al., 2013).

Isoniazid is a pro-drug and requires activation by the bacterial catalase-peroxidase enzyme (BP, 2009). HPLC is the most sensitive and selective means of determining isoniazid in its pure and tablet dosage form and methods have been reported (Khuhawret al., 2005;Milan-Segovia et al., 2007;

Dhal and Sharma, 2009; Abdallah et al., 2016). However, the cost of procurement and the technical hands to operate the HPLC machine is a major challenge in most developing countries.

On the other hand, UV/Vis spectrophotometry is the most readily available instrument in research laboratories for the quantitative determination of drugs in their pure and tablet dosage form.

This could be due to its low cost, simplicity, relative accuracy, and precision (Raza et al.,2003, 2005a, 2005b). Several UV/Vis spectrophotometric methods for the determination of isoniazid in pure and tablet dosage form have also been reported.

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StudentsandScholarship Team.