1. Reagents and Reactions
E. Udaya Rajitha
I M. Pharmacy
159V1S0404
1

2. CONTENTS
 Introduction
 Reactions
 Reagents
 Preparation
 Mechanism
 Applications
2

3. Introduction
Reagents are used for the qualitative and quantitative
determination of the pharmaceutical preparations, as they
react with particular groups and increase their detectability
by impart colour to the compounds.
3
Reaction Reagent
Diazo coupling reaction BM
Oxidative coupling reaction MBTH
Condensation reaction PDAB, PDAC
Oxidation / Reduction reaction FC

4. 4
Preparation: 100mg of BM reagent powder
100ml of mixture [ acetone : water (7:3) ]
Keep in dark place & protect from light
Mechanism:
Diazotization
Coupling
BM reagent:
The compounds which contains primary
aromatic amine groups are react with the BM regent and
form red colour azo dye which shows λmax at 550nm.
Diazo coupling reaction:

5. Diazotization:
5
NaNO2
HCL HNO2
HNO2 NH2
+
0-50C
+ Aromatic Ammonium molybdate+ Diazonium salt
NHCH2
CH2
NH2
Diazonium salt + Red azo dye
Coupling:
BM reagent

6. Applications:
6
Analysis of sulphonamides, paracetamol, dapsone,
isoniazid, procainamide, nifidipine, metronidazole…etc.,
OH
N NCl NHCH2
CH2
NH2 NHCH2
CH2
NH2
N OHN
+
+
Analysis of Paracetamol:
OH
NHCOCH3
OH
NH2
NaNO2
+HCL
OH
N NCl
Hydrolysis
+
Paracetamol
Step:1
Para amino phenol Diazonium salt
Step:2
BM reagent
Red azo dye

7. 7
Oxidative coupling reaction:
MBTH reagent:
Compounds like phenols, amines,
aldehydes, indoles, catechol amines reacts with the
MBTH reagent and forms orange-red colour product.
Preparation:
200mg of MBTH in 100ml of water
It gives 0.2% strength
Mechanism:
Oxidation
Coupling

8. 8
CH3
S
N-NH2
N
FeCl3
/CAS
CH3
S
N-NH
N
+
CH3
S
N-NH
N
OH
CH3
S
N
N-N O
+
+
Oxidation:
Coupling:
MBTH reagent
Phenol
Red azo dye

9. 9
CH3
S
N-NH2
N
FeCl3
/CAS
CH3
S
N-NH
N
+
CH3
S
N
N-N
CH3
S
N-NH
N HCHO
C
H
H
+
+
Applications:
Analysis of aldehydes, indoles, carbazoles,
nimesulide, diclophenac, oxiphenbutazole, primaquine,
metronidazole…etc.,
Analysis of Formaldehyde:
Step:1
Step:2
MBTH reagent
Red azo dye
Formaldehyde

10. Condensation reaction:
10
PDAB reagent:
Compounds like primary aromatic amines,
primary alkyl amines and compounds contains phenolic
–OH groups react with the PDAB reagent and give
yellow colour product.
Preparation:
Dissolve 125mg of PDAB reagent
cool
65ml of conc.H2SO4,
35ml of water

11. Mechanism:
11
NC
O
H
CH3
CH3
NC
CH3
CH3
H
Ar-NAr-NH2
H2
SO4
+
Schiff's base formation:
PDAB form coloured condensation products with
primary aromatic amines in the presence of conc.H2SO4.
Here C=O is converted to C=N.
PDAB reagent Schiff's base

12. 12
NC
O
H
CH3
CH3
NHNH2
NC
CH3
CH3
NHN
H
+
Analysis of phenyl hydrazine:
Applications:
Determination of phenyl hydrazine, nimesulide,
nifidipine, indole and pyrrole derivatives, amoxicillin,
metronidazole and estimation of urobilinogen…etc.,
PDAB reagent Phenyl hydrazine Schiff's base

13. Oxidation / Reduction reaction:
13
FC reagent:
10gm of sodium tungstate + 2.5gm of sodium molybdate
add 70ml of water and 10ml of conc.HCl
add 5ml of 80% phosphoric acid and 15g of lithium sulphate
reflux for 10 hrs
Add 5ml of water and 1 drop of bromine
reflux for 15 min
Cool to room temperature
Preparation:

14. 14
FC reagent is a oxidising agent, so it itself reduces and
oxidise the analyte, probably drug effects reduction of 1 or
more oxygen atoms from tungstate or molybdate in the FC
reagent, there by producing one or more possible reduced
species which have characteristic intense blue colour.
Mechanism:
Drug + FC reagent
Molybdenum blue /
Tungstic green
Alkali

15. Applications:
15
Estimation of Omeprazole, Aceclofenac, assay of Folic
acid, Acyclovir, Diclofenac, Antibiotics…etc.,
+
Omeprazole FC reagent
↓ Na2CO3
Blue colour complex

16. 16
Other reagents:
• Folins reagent
• Gibbs reagent
• NBS
• FeCl3, Potassium ferri cyanide, 1,10- Phenanthroline
• Froehde Reagent
• 4-(Dimethylamino) Cinnamaldehyde
• Phosphomolybdic acid
• Zwikker Reagent

17. 17
Common applications:
Reagents Drugs
BM, MBTH Procainamide
BM, PDAB Nifidipine, Sulphonamides,
Metronidazole
PDAB, MBTH Indole, Nimesulide
MBTH, Gibb’s Vitamin B6
Folins, PDAB, NBS Amoxicillin
BM, NBS, FeCl3, K.Fe(CN)6,
1,10-phenanthroline Paracetamol
PDAB, MBTH, FC, FeCl3,
K.Fe(CN)6, 1,10-phenanthroline Diclofenac

18. Conclusion
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19. References:
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• Text book of pharmaceutical analysis by P. D. Chaithanya sudha,
chapter VII (Reagents), Pg. no. 348-369.
• A review article on Chemical derivatization methodologies for
Uv‐Visible Spectrophotometric determination of pharmaceuticals by
Olajire A. Adegoke.
• A review article on Role of Chemical and Analytical Reagents in
Colorimetric Estimation of Pharmaceuticals by K. Sushma,
Somsubhra Ghosh et al.,

20. THANK YOU
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