design and synthesis of potential antimicrobial agents

290 Pages·2011·4.28 MB·English

DESIGN AND SYNTHESIS OF POTENTIAL ANTIMICROBIAL
AGENTS

SOUMICK SIKDAR

Doctor of Philosophy

ASTON UNIVERSITY

July 2010

This copy of the thesis has been supplied on condition that anyone who consults it is
understood to recognise that its copyright rests with its author and that no quotation
from the thesis and no information derived from it may be published without proper
acknowledgement.
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Abstracts 2010

ASTON UNIVRSITY

DESIGN AND SYNTHESIS OF POTENTIAL ANTIMICROBIAL AGENTS

A thesis submitted by Soumick Sikdar B.Pharm, for the degree of Doctor of Philosophy
June 2010

Abstract:

Chorismate mutase is one of the essential enzymes in the shikimate pathway and is key to
the survival of the organism Mycobacterium tuberculosis. The x-ray crystal structure of this
enzyme from Mycobacterium tuberculosis was manipulated to prepare an initial set of in
silico protein models of the active site. Known inhibitors of the enzyme were docked into the
active site using the flexible ligand / flexible active site side chains approach implemented in
CAChe Worksystem (Fujitsu Ltd). The resulting complexes were refined by molecular
dynamics studies in explicit water using Amber 9. This yielded a further set of protein models
that were used for additional rounds of ligand docking. A binding hypothesis was established
for the enzyme and this was used to screen a database of commercially available drug-like
compounds. From these results new potential ligands were designed that fitted appropriately
into the active site and matched the functional groups and binding motifs founds therein.
Some of these compounds and close analogues were then synthesized and submitted for
biological evaluation.

As a separate part of this thesis, analogues of very active anti-tuberculosis
pyridylcarboxamidrazone were also prepared. This was carried out by the addition and the
deletion of the substitutions from the lead compound thereby preparing heteroaryl
carboxamidrazone derivatives and related compounds. All these compounds were initially
evaluated for biological activity against various gram positive organisms and then sent to the
TAACF (USA) for screening against Mycobacterium tuberculosis. Some of the new
compounds proved to be at least as potent as the original lead compound but less toxic.

Keywords:

Mycobacterium tuberculosis, chorismate mutase, molecular modeling, molecular dynamics,
docking studies, heteroaryl carboxamidrazone

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Acknowledgements 2010

ACKNOWLEDGEMENTS

I offer countless thanks to Almighty God who has given me the ability, determination,
courage and will to complete this thesis. I offer my respect to my great grandfather Late
Nakul Chandra Sikdar and my grandfather Shri Shibdas Sikdar who taught me the meaning
and importance of life.

I offer sincere thanks to my supervisor Dr. Daniel Rathbone for being kind, patient and an
inspiring source through often difficult moments in writing my thesis. I acknowledge his
motivations and overall support that kept me enthusiastic throughout the entire period of this
work and I am grateful to have him.

Thanks to the School of Life and Health Sciences for the provision of the funding in this
project.

I would like to thank Dr. Peter Lambert for arranging the microbiological testing of all the
compounds which were prepared in this thesis. Many thanks to Dr Fraser in clarifying my
doubts associated with reaction mechanisms.

I would like to express my gratitude for the help and technical assistances given to me by Dr.
Mike Davis and Karen Farrow.

Many thanks to final year project students Jasjeet Singh and Pritpal Singh for their
assistance in preparing some of the carboxamidrazone. I would like to thank Ren, Mei,
Alexandra and Preeti for helping me to find the way through the complicated procedure of the
computer packages used in this thesis.

Finally I would like to thank everybody in my family, my dad, my mum, my sister for their
support and encouragement throught my time at university, and last but by no means least,
my wife Munmun, whose patience and understanding went beyond the call of duty.
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List of contents 2010

LIST OF CONTENTS

Page
TITLE PAGE 1
ABSTRACTS 2
ACKNOWLEDGEMENTS 3
LIST OF CONTENTS 4
LIST OF FIGURES 12
LIST OF TABLES 16
LIST OF SCHEMES 18
LIST OF GRAPHS 19
LIST OF EQUATIONS 21
LIST OF ABBREVIATIONS 23

CHAPTER 1 25
INTRODUCTION 25
1.1 Introduction to Tuberculosis: 25
1.1.1 Background: 26
1.1.2 Signs and symptoms in tuberculosis: 26
1.1.3 Aetiology: 27
1.1.4 Pathophysiology: 27
1.1.5 Diagnosis and Treatment: 28
1.1.5.1 Diagnosis of TB: 28
1.1.5.2 Isoniazid (INH): 30
1.1.5.3 Ethambutol (EMB): 30
1.1.5.4 Pyrazinamide (PZA): 31
1.1.5.5 Rifampacin (RMP): 31
1.1.5.6 Streptomycin (STM): 31
1.1.5.7 Polypeptides: 31
1.1.5.8 Fluoroquinolones: 32
1.1.5.9 Thioamides 32
1.1.5.10. Para amino salicylic acid class of drugs. 32
1.1.6 Newer Drugs: 32
1.1.7 Treatment regimen: 33
1.1.8 Drug resistance: 34
1.2 Biosynthetic pathway in bacteria. 35
1.2.1 Shikimic acid pathway: 35
1.2.2 Chorismate mutase: 36
1.2.3 Transition State Analogue (TSA): 37
1.3. Drug design methods: 37
1.3.1 Protein: 38
1.3.2. Protein crystal structures 39
1.4 Introduction to Molecular Modelling: 42
1.5 Molecular Mechanics: 42
1.5.1 Bond stretching: 43
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1.5.2 Bond bending: 44
1.5.3 Torsional energy: 45
1.5.4 Non bonded interactions: 46
1.5.4.1 Van der Waals interactions: 46
1.5.4.2. Electrostatic interactions: 47
1.6 Quantum mechanics: 48
1.6.1 Energy calculation using ab initio quantum mechanical methods: 49
1.6.2 Energy Calculations using semi empirical methods: 50
1.7 Geometry optimization: 50
1.7.1 Geometry optimization by molecular mechanic methods. 51
1.7.1.1 Minimisation: 52
1.7.1.2 Molecular dynamics: 52
1.7.2 Geometry optimization by semi-empirical methods. 53
1.7.3 Geometry optimization by ab initio methods. 53
1.8 Background and development of the Amber Force field. 54
1.8.1. Amber Project: 56
1.9. Selection of Algorithm: 56
1.9.1 Verlet algorithm: 57
1.9.2 Leap-frog algorithm 57
1.9.3 Velocity Verlet method 58
1.9.3 Beeman’s algorithm: 59
1.10. Starting, running, measuring and analysing simulation program: 59
1.10.1 Starting: 59
1.10.2 Running: 60
1.10.3 Measuring and analysing: 60
1.11. Validation of models using Ramachandran Plot: 61
1.12. Docking studies: 63
1.13.1. Docking 63
1.13.2. Scoring Methods: 66

CHAPTER 2 68
OPTIMIZATION OF AN INSILICO MODEL FOR MYCOBACTERIAL CHORISMATE
MUTASE: THE COMPUTATIONAL APPROACH 68
2.1 Aim: 68
2.2 Background: 68
2.3 Crystal structure of chorismate mutase from Mycobacterium tuberculosis: 68
2.3.1 Background of chorismate mutase: 68
2.3.2 Structure of Mycobacterium Chorismate Mutase: 69
2.4 Active site analysis: 71
2.5 The Ligands: 73
2.6 Computational approach: 77
2.6.1 Preparing the initial protein structure: 77
2.6.2 Preparing the initial Ligand structures: 78
2.6.3 Generating the model: 78
2.6.4 Phase I Study: 78
2.6.5 Phase II study: 83
2.6.6 Phase III study: 83
2.7 Analyzing the models: 85
2.7.1 Validation of models: 85
2.7.2 Ramachandran plot: 85
2.7.3 Analysis of the parameters. 89
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2.7.4 RMSD analysis: 100
2.8 Docking studies: 103
2.8.1 Docking scores results: 104
2.9 Identifying the interactions: 116
2.10 Discussion: 120
2.10.1 Model Hypothesis: 120
2.10.2 Molecular modelling results and discussions: 121
2.10.3 Selecting the targeted model: 127
2.10.4 Docking results and discussion: 127
2.10.5 Structural activity studies of Ligand 1: 128
2.10.6 Fragment based approach for Ligand 1: 130
2.10.7 Structural activity studies of Ligand 2: 131
2.10.8 Docking studies of the commercially available ligands from Maybridge: 132
2.11 Conclusions: 136

CHAPTER 3 137
SUBSTITUTED 3-NITRO 5-SULPHAMOYLBENZOIC ACIDS: DERIVATIVES AND
RELATED CHEMISTRY 137
3.1 Aim: 137
3.2 Background: 137
3.3 Substituted 3-sulphamoyl-5-nitro benzoic acid and related compounds: 138
3.4 Screening synthesisable compounds: 141
3.5 Varying the benzoic acids moiety: 145
3.6 Varying the phenylamines: 147
3.7 Varying the benzylhydrazine moiety: 147
3.8 Reactivity of the substituted benzoic acids: 148
3.9 Reactivity of the phenylamines: 150
3.10 Reactivity of the benzylhydrazines: 151
3.11 Reactivity of the benzoyl hydrazide: 152
3.12 Microbiological studies: 153
3.12.1 Dose Response screening: 153
3.13 Discussion: 154
3.13.1 Attempted synthesis of Compound 3.25 and other related compounds: 156
3.13.2 Attempted synthesis of Compound 3.20 and related compounds: 157
3.13.3 Attempted synthesis of 3-methoxy-2-hydroxy benzyl amine Compound 3.40: 157
3.13.4 Attempted synthesis of Compound 3.29: 158
3.13.5 Structural Activity Relationship studies: 159
3.14 Conclusion: 163

CHAPTER 4 164
MATERIALS AND METHODS 164
4.1. Chemicals: 164
4.2. Instrumentation: 164
4.3.1.1 Preparation of 4-chloro-3-chlorosulfonylbenzoic acid: Compound 3.30. 164
4.3.1.2 Preparation of 4-chloro-3-chlorosulfonyl-5-nitrobenzoic acid: Compound 3.31. 165
4.3.1.3 Preparation of 4-chloro-3-sulphamoyl-5-nitrobenzoic acid: Compound 3.27. 166
4.4.1 Preparation of 4-(3,4-dihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid :
Compound 3.2. 167
4.4.1.1. Preparation of DIEA salt of 4-(3, 4-dihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic
acid: 167
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4.4.1.2 Preparation of BOC substituted 4-(3,4-dihydroxyphenethylamino)-3-sulphamoyl-5-nitro
benzoic acid : 167
4.4 1.3. Preparation of 4-(3,4-dihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid : 168
4.5.1. Preparation of 4-(phenethylamino)-3-sulphamoyl-5-nitro benzoic acid: Compound 3.5. 168
4.5.1.1 Preparation of DIEA salt of 4-(phenethylamino)-3-sulphamoyl-5-nitro benzoic acid: 168
4.5.1.2 Preparation of BOC substituted 4-(phenethylamino)-3-sulphamoyl-5-nitro benzoic acid: 169
4.5.1.3. Preparation of 4-(phenethylamino)-3-sulphamoyl-5-nitro benzoic acid : 169
4.6.1. Preparation of 4-(4-hydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid: Compound
3.4. 170
4.6.1.1 Preparation of DIEA salt of 4-(4-hydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid:
170
4.6.1.2 Preparation of BOC substituted 4-(4-hydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic
acid: 170
4.6.1.3 Preparation of 4-(4-hydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid: 171
4.7.1 Preparation of 4-(3,4,5-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid :
Compound 3.3. 171
4.7.1.1 Preparation of DIEA salt of 4-(3,4,5-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic
acid: 171
4.7.1.2 Preparation of BOC substituted 4-(3,4,5-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro
benzoic acid: 172
4.7.1.3 Preparation of 4-(3,4,5-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid: 172
4.8.1 Preparation of 4-(3,4,6-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid :
Compound 3.7. 173
4.8.1.1 Preparation of DIEA salt of 4-(3,4,6-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic
acid: 173
4.8.1.2 Preparation of BOC substituted 4-(3,4,6-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro
benzoic acid: 173
4.8.1.3 Preparation of 4-(3,4,6-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid: 174
4.9.1 Preparation of 4-(3-methoxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid : Compound
3.6 174
4.9.1.1 Preparation of DIEA salt of 4-(3-methoxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid:
174
4.9.1.2. Preparation of BOC substituted 4-(3-methoxyphenethylamino)-3-sulphamoyl-5-nitro benzoic
acid: 175
4.9.1.3 Preparation of 4-(3,4,5-trihydroxyphenethylamino)-3-sulphamoyl-5-nitro benzoic acid: 175
4.10.1. Preparation of 4-(3,4-dihydroxyphenethylamino)-3,5-dinitro benzoic acid: Compound 3.8. 176
4.10.1.1 Preparation of DIEA salt of 4-(3,4-dihydroxyphenethylamino)-3,5-dinitro benzoic acid: 176
4.10.1.2 Preparation of BOC substituted 4-(3,4-dihydroxyphenethylamino)-3,5-dinitro benzoic acid:
176
4.10.1.3 Preparation of 4-(3,4-dihydroxyphenethylamino)-3,5-dinitro benzoic acid: 177
4.11.1 Preparation of 4-(3-methoxyphenethylamino)-3, 5-dinitro benzoic acid: Compound 3.12. 177
4.11.1.1 Preparation of DIEA salt of 4-(3-methoxyphenethylamino)-3,5-dinitro benzoic acid: 177
4.11.1.2 Preparation of BOC substituted 4-(3-methoxyphenethylamino)-3,5-dinitro benzoic acid: 178
4.11.1.3 Preparation of 4-(3-methoxyphenethylamino)-3,5-dinitro benzoic acid: 178
4.12.1. Preparation of 4-(4-hydroxyphenethylamino)-3,5-dinitro benzoic acid : Compound 3.9. 179
4.12.1.1. Preparation of DIEA salt of 4-(4-hydroxyphenethylamino)-3,5-dinitro benzoic acid: 179
4.12.1.2. Preparation of BOC substituted 4-(4-hydroxyphenethylamino)-3,5-dinitro benzoic acid: 179
4.12.1.3. Preparation of 4-(4-hydroxyphenethylamino)-3,5-dinitro benzoic acid: 180
4.13.1. Preparation of 4-phenethylamino-3,5-dinitro benzoic acid : Compound 3.11. 180
4.13.1.1. Preparation of DIEA salt of 4-phenethylamino-3,5-dinitro benzoic acid: 180
4.13.1.2. Preparation of BOC substituted 4-phenethylamine-3,5-dinitro benzoic acid: 181
4.13.1.3. Preparation of 4-phenethylamine-3,5-dinitro benzoic acid: 181
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4.14.1 Preparation of 4-(3,4,6-trihydroxyphenethylamino)-3,5-dinitro benzoic acid : Compound 3.13
182
4.14.1.1. Preparation of DIEA salt of 4-(3,4-dihydroxyphenethylamino)-3,5-dinitro benzoic acid: 182
4.14.1.2. Preparation of BOC substituted 4-(3,4,6-trihydroxyphenethylamino)-3,5-dinitro benzoic acid:
182
14.1.3. Preparation of 4-(3,4,6-trihydroxyphenethylamino)-3,5-dinitro benzoic acid: 183
4.15.1 Preparation of 4-(3,4,5-trihydroxyphenethylamino)-3,5-dinitro benzoic acid : Compound 3.9.
183
4.15.1.1. Preparation of DIEA salt of 4-(3,4,5-trihydroxyphenethylamino)-3,5-dinitro benzoic acid: 183
4.15.1.2. Preparation of BOC substituted 4-(3,4-dihydroxyphenethylamino)-3,5-dinitro benzoic acid:
184
4.15.1.3. Preparation of 4-(3,4,5-trihydroxyphenethylamino)-3,5-dinitro benzoic acid: 184
4.16.1. Preparation of 3-methoxy-4-hydroxybenzylhydrazine: Compound 3.33. 184
4.16.1.1 Preparation of 3-methoxy-4-hydroxybenzylidinehydrazine: 184
4.16.1.2 Preparation of 3-methoxy-4-hydroxybenzylhydrazine: 185
4.17.1. Preparation of 3,4-dimethoxybenzylhydrazine: Compound 3.32. 185
4.17.1.1 Preparation of 3,4-dimethoxybenzylidinehydrazine: 186
4.17.1.2 Preparation of 3,4-dimethoxybenzylhydrazine: 186
4.18.1. Preparation of 3,4-dihydroxybenzylhydrazine: Compound 3.24. 186
4.18.1.1 Preparation of 3,4-dihydroxybenzylidinehydrazine: 186
4.19.1. Preparation of 2-methoxy-4-((2-(4-methylbenzylidene) hydrazinyl) methyl) phenol:
Compound 3.36. 187
4.20.1. Preparation of 4-amino-3-(N-methylsulphamoyl)-5-nitrobenzoic acid: Compound 3.35. 188

CHAPTER 5 189
HETEROARYLCARBOXAMIDRAZONES: DERIVATIVES AND RELATED CHEMISTRY
189
5.1 Aim: 189
5.2 Background: 189
5.3 Structural activity relationship studies: 190
5.4 Optimisation of the lead compound: 191
5.4.1 Optimisation of lead compound following Compound 5.1: 191
5.5 Varying the aldehyde building blocks in the synthesis of benzylidine
carboxamidrazones: 199
5.6 Varying the aryl acid building blocks in the synthesis of carboxamidrazone amides:
201
5.7 Varying the Pyridyl-amine moiety in the synthesis of analogues of pyridine
carboxamidrazones: 203
5.8 Reactivity of the pyridyl-amine moiety. 204
5.9 Stability of the Pyridyl-amines: 207
5.10 Microbiological studies: 208
5.10.1 Initial Biological Screening: 208
5.10.2 TAACF screening: 211
5.10.2.1 Primary Screen (Dose Response): 211
5.10.2.2 Secondary Screen: 211
5.11 Results and Discussion: 215
5.11.1 Microbiological studies: 220
5.11.2 Toxicology: 222
5.11.3 Structural activity relationship studies: 223
5.12 Conclusion: 226

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CHAPTER 6 227
MATERIALS AND METHODS 227
6.1. Chemicals: 227
6.2. Instrumentation: 227
6.3.1. Preparation of Pyridine-4-methylhydrazone: Compound 5.87 227
6.3.2. Preparation of 4-[3,5-di-(tert-butyl)-2-OH-benzyl]-1-(4-pyridyl)-2, 3-diaza-1,3-butadiene:
Compound 5.17 228
6.3.3. Preparation of 4-[3, 5-di-(tert-butyl)-4-OH-benzyl]-1-(4-pyridyl)-2, 3-diaza-1,3-butadiene:
Compound 5.18 229
6.3.4. Preparation of 4-[3,5-di-iodo-2-OH-benzyl]-1-(4-pyridyl)-2,3-diaza-1,3-butadiene. Compound
5.19 229
6.3.5. Preparation of 4- [5-tert-butyl-2-OH-benzyl]-1-(4-pyridyl)-2, 3-diaza-1,3-butadiene Compound
5.22 230
6.3.6. Preparation of 4-[3-tert-butyl-2-OH-benzyl]-1-(4-pyridyl)-2, 3-diaza-1, 3-butadiene Compound
5.21 230
6.3.7. Preparation of 4-[3,5-di-bromo-2-OH-benzyl]-1-(4-pyridyl)-2,3-diaza-1,3-butadiene:
Compound 5.20 231
6.4.1 Preparation of 4-hydrazino pyridine: Compound 5.84. 231
6.4.2. Preparation of 3,5-di-tert butyl 2-OH benzylidine 4-hydrazinopyridine Compound 5.03 232
6.4.3. Preparation of 3,5-di-tert butyl 4-OH benzylidine 4-hydrazinopyridine. Compound 5.04 233
6.4.4. Preparation of 3,5-di-iodo 2-OH benzylidine 4-hydrazinopyridine Compound 5.05 233
6.4.5. Preparation of 3,5-di-bromo 2-OH benzylidine 4-hydrazinopyridine Compound 5.06. 234
6.4.6. Preparation of 3,5-di-tert-butyl benzylidine 4-hydrazinopyridine: Compound 5.09. 234
6.4.7. Preparation of 3-tert-butyl 2-OH benzylidine 4-hydrazinopyridine: Compound 5.07 235
6.4.8. Preparation of 5-tert-butyl 2-OH benzylidine 4-hydrazinopyridine Compound 5.08 235
6.5.1. Preparation of 4-[3, 5-di-(tert-butyl)-2-OH-benzylidine]-amino-pyridine: Compound 5.29. 236
6.5.2. Preparation of 4-[3, 5-di-iodo-2-OH-benzylidine]-amino-pyridine: Compound 5.31 236
6.5.3. Preparation of 4-[3, 5-di-(tert-butyl)-4-OH-benzylidine]-amino-pyridine: Compound 5.30 237
6.5.4. Preparation of 4-[3, 5-di-bromo-2-OH-benzylidine]-amino-pyridine: Compound 5.32 238
6.6.1. Preparation of 3,5-di-tert-butyl 2-OH benzylidine 2-hydrazinopyridine. Compound 5.10 238
6.6.2. Preparation of 3,5-di-tert-butyl-4-OH benzylidine-4-hydrazinopyridine: Compound 5.11. 239
6.6.3. Preparation of 3,5-diiodo-2-OH benzylidine-4-hydrazinopyridine Compound 5.12. 240
6.6.4. Preparation of 3,5-dibromo-2-OH benzylidine-2-hydrazinopyridine: Compound 5. 13. 240
6.6.5. Preparation of 3-tert-butyl-2-OH benzylidine-2-hydrazinopyridine: Compound 5.14. 241
6.6.6. Preparation of 5-tert-butyl-2-OH benzylidine-2-hydrazinopyridine: Compound 5.15. 241
6.6.7. Preparation of 3,5-di-tert-butyl-benzylidene-2-hydrazinopyridine: Compound 5.16. 242
6.7.1. Preparation of Pyridine-2-methylhydrazone: Compound 5.88. 242
6.7.2. Preparation of 4-[3,5-di-(tert-butyl)-2-OH-benzyl]-1-(2-pyridyl)-2, 3-diaza-1,3-butadiene:
Compound 5.23. 243
6.7.3. Preparation of 4-[3,5-di-(tert-butyl)-4-OH-benzyl]-1-(2-pyridyl)-2,3-diaza-1,3-butadiene.
Compound 5.24. 243
6.7.4. Preparation of 4-[3-(tert-butyl)-2-OH-benzyl]-1-(2-pyridyl)-2,3-diaza-1,3-butadiene:
Compound 5.25. 244
6.8.1 Preparation of Pyridine-3-methylhydrazone: Compound 5.86. 244
6.8.2. Preparation of 4-[3, 5-di-(tert-butyl)-2-OH-benzyl]-1-(3-pyridyl)-2, 3-diaza-1, 3-butadiene.
Compound 5.26 245
6.8.3. Preparation of 4-[3,5-di-iodo-2-OH-benzyl]-1-(3-pyridyl)-2,3-diaza-1,3-butadiene: Compound
5.27 246
6.8.4. Preparation of 4-[3,5-di-bromo-2-OH-benzyl]-1-(3-pyridyl)-2,3-diaza-1,3-butadiene:
Compound 5.28. 246
6.9.1. Preparation of 3,5-diiodo-2-OH benzylidine-3-aminopyridine: Compound 5.35. 247
6.9.2. Preparation of 3,5-di-(tert-Butyl)-2-OH benzylidine-3-aminopyridine: Compound 5.33. 247
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6.9.3. Preparation of 3,5-dibromo-2-OH benzylidine-3-aminopyridine: Compound 5.36. 248
6.9.4. Preparation of 5-(tert-Butyl)-2-OH benzylidine-3-aminopyridine: Compound 5.37. 248
6.9.5. Preparation of 3,5-di-(tert-Butyl)-4-OH benzylidine-3-aminopyridine: Compound 5.34. 249
6.9.6. Preparation of 3-bromo-5-Chloro-2-hydroxy benzylidine-3-aminopyridine: Compound 5.94.
249
6.9.7. Preparation of 3,5-dichloro-2-hydroxy benzylidine-3-aminopyridine: Compound 5.95. 250
6.9.8. Preparation of 3,5-difluoro-2-hydroxy benzylidine-3-aminopyridine: Compound 5.98. 250
6.9.9. Preparation of 5-Iodo-2-hydroxy benzylidine-3-aminopyridine: Compound 5.99. 251
6.9.10. Preparation of 3-chloro-2-hydroxy benzylidine-3-aminopyridine: Compound 5.97. 251
6.9.11. Preparation of 2-bromo-2-hydroxy benzylidine-3-aminopyridine: Compound 5.100. 252
6.9.12. Preparation of 5-chloro-2-hydroxy benzylidine-3-aminopyridine: Compound 5.96. 253
6.9.13. Preparation of 5-bromo-2-hydroxy benzylidine-3-aminopyridine: Compound 5.101. 253
6.10.1. Preparation of pyridine-2-carboxamidrazone: Compound 5.89. 254
6.10.2. Preparation of N1-(2-furoylacryl)-pyridine-2-carboxamidrazone: Compound 5.38. 254
6.10.3. Preparation of N1-2-(5-bromofuroyl)-pyridine-2-carboxamidrazone: Compound 5.41. 255
6.10.4. Preparation of N1-2-(4,5-dibromofuroyl)-pyridine-2-carboxamidrazone: Compound 5.42. 256
6.10.5. Preparation of N1-2-[3-(2-furoyl)-propyl]-pyridine-2-carboxamidrazone: Compound 5.44. 256
6.10.6. Preparation of N1-3-(2,5-dimethylfuroyl)-pyridine-2-carboxamidrazone: Compound 5.43. 257
6.10.7. Preparation of N1-3-furoyl-pyridine-2-carboxamidrazone: Compound 5.39. 257
6.10.8. Preparation of N1-2-benzofuroyl-pyridine-2-carboxamidrazone: Compound 5.45. 258
6.10.9. Preparation of N1-2-[5-(4-Nitrophenyl)furoyl]-pyridine-2-carboxamidrazone: Compound 5.40.
258
6.10.10. Preparation of N1-(thiophen-2-oyl)-pyridine-2-carboxamidrazone: Compound 5.51. 259
6.10.11. Preparation of N1-(3-bromothiophen-2-oyl)-pyridine-2-carboxamidrazone: Compound 5.54.
259
6.10.12. Preparation of N1-(5-bromothiophen-2-oyl)-pyridine-2-carboxamidrazone: Compound 5.57.
260
6.10.13. Preparation of N1-(5-chlorothiophen-2-oyl)-pyridine-2-carboxamidrazone: Compound 5.60.
260
6.10.14. Preparation of N1-(2-bromobenzoyl)-pyridine-2-carboxamidrazone: Compound 5.71. 261
6.10.15. Preparation of N1-(4-bromobenzoyl)-pyridine-2-carboxamidrazone: Compound 5.63. 261
6.10.16. Preparation of N1-(5-[4-bromophenyl]-furan-2-oyl)-pyridine-2-carboxamidrazone: Compound
5.66. 262
6.10.17. Preparation of N1-(5-[2-trifluoromethylphenyl]-furan-2-oyl)-pyridine-2-carboxamidrazone:
Compound 5.69. 262
6.11.1. Preparation of pyridine-4-carboxamidrazone: Compound 5.91. 263
6.11.2. Preparation of N1-(5-chlorothiophen-2-oyl)-pyridine-4-carboxamidrazone: Compound 5.59.
263
6.11.3. Preparation of N1-(3-bromothiophen-2-oyl)-pyridine-4-carboxamidrazone: Compound 5.53.
264
6.11.4. Preparation of N1-(5-bromothiophen-2-oyl)-pyridine-4-carboxamidrazone: Compound 5.56.
265
6.11.5. Preparation of N1-(4-bromobenzoyl)-pyridine-4-carboxamidrazone: Compound 5.62. 265
6.11.6. Preparation of N1-(thiophen-2-oyl)-pyridine-4-carboxamidrazone: Compound 5.50. 266
6.11.7. Preparation of N1-(5-[4-bromophenyl]-furan-2-oyl)-pyridine-4-carboxamidrazone: Compound
5.65. 266
6.11.8. Preparation of N1-(5-[2-trifluoromethylphenyl]-furan-2-oyl)-pyridine-4-carboxamidrazone:
Compound 5.68. 267
6.12.1. Preparation of pyridine-3-carboxamidrazone: Compound 5.90. 267
6.12.2. Preparation of N1-(3-bromothiophen-2-oyl)-pyridine-3-carboxamidrazone: Compound 5.55.
268
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