Figures

Figure 1. FT-IR Spectra of reference standard Phenytoin

82
 Figures

Figure 2. FT-IR spectra of working standard Phenytoin

83
 Figures

Figure 3. FT-IR spectra of Benzil

84
 Figures

Figure 4. FT-IR Spectra of reference standard Warfarin

85
 Figures

Figure 5. FT-IR Spectra of working standard Warfarin

86
 Figures

Figure 6. FT-IR Spectra of Benzalacetone

87
 Figures

Figure 7. FT-IR spectra of reference standard d Nicergoline

88
 Figures

Figure 8. FT-IR spectra of working standard Nicergoline

89
 Figures

Figure 9. FT-IR spectra of 5-Bromo Nicotinic Acid

90
 Figures

Figure 10. Typical chromatogram of standard Phenytoin and Benzil (10µg/ml)

91
 Figures

Figure 11. Typical mass spectra of standard Phenytoin and Benzil (10µg/ml)

Inten.(x1,000)
209

5.0

251
2.5

0.0
210.0 215.0 220.0 225.0 230.0 235.0 240.0 245.0 250.0 m/z

92
 Figures

Figure 12. Typical mass spectra of Phenytoin in scan negative mode.

Inten. (x100,000)
3.0

251
2.5

2.0

1.5

1.0

255
0.5
203

217

262

279
227
223

247
239

265
205

241
233

271
211

0.0
200 210 220 230 240 250 260 270 m/z

93
 Figures

Figure 13. Typical mass spectra of Phenytoin in SIM mode.

Inten. (x10,000)

251
1.00

0.75

0.50

0.25

0.00
250.000 250.250 250.500 250.750 251.000 251.250 251.500 251.750 m/z

94
 Figures

Figure 14. Typical mass spectra of Benzil in scan negative mode.

Inten. (x1,000)

228

244
6.0

252

256

269

279
211
5.0

4.0
209

260
245
3.0

230
2.0
218

237
201

221

1.0

0.0
200 210 220 230 240 250 260 270 m/z

95
 Figures

Figure 15. Typical mass spectra of Benzil in SIM mode.

Inten. (x10,000)

209
1.50

1.25

1.00

0.75

0.50

0.25

0.00
208.000 208.250 208.500 208.750 209.000 209.250 209.500 209.750 m/z

96
 Figures

Figure 16. Typical chromatogram of standard Warfarin and Benzalacetone.

97
 Figures

Figure 17. Typical chromatogram of Benzalacetone in formulation

98
 Figures

Figure 18. Typical chromatogram of Benzalacetone in bulk drug

99
 Figures

Figure 19. Typical mass spectra of Warfarin in scan mode.

Inten. (x1,000,000)

309
2.0

1.0

311

347
331

343
325
291

341
284

289

297
299

304

315

320

337
0.0
285.0 290.0 295.0 300.0 305.0 310.0 315.0 320.0 325.0 330.0 335.0 340.0 345.0 m/z

100
 Figures

Figure 20. Typical mass spectra of Warfarin and Benzalacetone in SIM mode.

Inten. (x10,000)
1.00

309
0.75
147

0.50

0.25

0.00
150.0 175.0 200.0 225.0 250.0 275.0 300.0 m/z

101
 Figures

Figure 21. Typical mass spectra of Benzalacetone in scan positive mode.

Inten. (x10,000)

147
5.0
130

2.5

148

188
179
139
132

173
143

181

195
122

156

163
165
127

169
0.0
130 140 150 160 170 180 190 m/z

102
 Figures

Figure 22. Typical chromatogram of Standard Nicergoline and 5-Bromo Nicotinic Acid (10µg/ml)

Detector A (277nm)
validation
assay001
0.06 Retention Time 0.06

0.04 0.04
Volts

Volts
2.958

0.02 0.02

4.633
0.00 0.00

0 1 2 3 4 5 6 7 8 9 10
Minutes

103
 Figures

Figure 23. Typical chromatogram of Sample Nicergoline and 5-Bromo Nicotinic Acid

Detector A (277nm)
AlkaliI 24Hr
Alkali 24 Hr 001
0.06 Retention Time 0.06

0.04 0.04
Volts

Volts
2.600
2.800
0.02 0.02
2.317

4.650
0.00 0.00

0 1 2 3 4 5 6 7 8 9 10
Minutes

104
 Figures

Figure 24. Typical mass spectra of Nicergoline scan positive mode.

Inten. (x100,000)

484
486
7.5
452

5.0

2.5

525
500
456

475

528
505
495

520

549
463

468

513

535

541
0.0
450 460 470 480 490 500 510 520 530 540 m/z

105
 Figures

Figure 25. Typical mass spectra of 5-Bromo Nicotinic Acid scan positive mode

Inten. (x10,000)
1.00

272

313
243
202
0.75

0.50

284
0.25

299
256
213

332
263

322

345
303
233
191
183
158
162

229
178

0.00
175.0 200.0 225.0 250.0 275.0 300.0 325.0 m/z
.

106
 Figures

Figure 26. Calibration curve of Benzil

107
 Figures

Figure 27. Calibration curve of Benzalacetone

108
 Figures

Figure 28. Calibration curve of 5-Bromo Nicotinic Acid

109
 Figures

Figure 29. Genotoxicity study control (Hydroxylamine 8% v/v)

110
 Figures

Figure 30. Genotoxicity study for Benzil

111
 Figures

Figure 31. Genotoxicity study for Benzalacetone

112
 Figures

Figure 32. Genotoxicity Report For5-Bromo Nicotinic Acidin terms of concentration

1 2 3 4 5 6 7

Well - 1: Plasmid alone, Well - 2: Quercetin (Positive control), Well- 3 : Fenton’s reagent (-ve control), Well - 4 : 6.5 µg/ ml
sample solution, Well - 5 : 100 µg/ml sample solution, Well - 6: 200 µg/ml sample solution, Well- 7 : 300µg/ml sample
solution

113
 Figures

Figure 33. genotoxicity report for 5-Bromo Nicotinic Acidin terms of time

1 2 3 4 5 6

Well-1: Plasmid alone, Well-2: 2 hrs incubation , Well-3: 4 hrs incubation , Well-4: 6 hrs incubation , Well-5:124 hrs
incubation and Well-6: 24 hrs incubation ,

114