Preesented by

Rohit U. Bharati

T.Y. B. Pharm.

GUIDE

Ms. Vandana T. Gawande

M.Pharm. (Q.A.)

DATE : 21st March 2012 TIME : 02 : 45 p.m.

1

Sr. No. Particulars

1 AIM & OBJECTIVES

2 INTRODUCTION

3 SELECTION OF DRUG

4 DRUG PROFILE

5 DEVELOPMENT OF ASSAY METHOD

6 VALIDATION METHODS

7 OBSERVATION

8 CONCLUSION

9 BIBLIOGRAPHY

2

Part A: To developed analytical (UV) method for determination of TOPIRAMATE in bulk and in oral solid dosage form.

Part B: To validate developed method as per ICH guidelines for following parameters:

a) Accuracy

b) Precision

d) Linearity and Range

e) Limit of Detection

f) Limit of Quantitation

g) Robustness

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COLORIMETRY

It is the science & technology used to quantify & describe physically the Human color perception.

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INSTRUMENTATION

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1. Drugs which not have strong UV absorbance.

2. Drugs for which Colorimetric methods are not

available.

3. Drugs for which methods are available but they are

time consuming & complex.

Eg. Dicloxacillin, Topiramate etc.

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TOPIRAMATE

Chemical Name 2, 3, 4, 5-bis-o-(1-methylidene)-betaD-fructopyranose sulfamate

Solubility freely soluble in acetone, chloroform

and ethanol. water it is 9.8 mg/ml.

Its saturated solution has a pH of 6.3.

Category Anticonvulsant & Antimigraine Drug

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1. Preparation of std. stock solution.

2. Development of colour.

3. Selection of analytical wavelength.

4. selection of analytical concentration Range.

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10 mg of topiramate + distilled water

Ultrasonicated (10 min. )

Vol. adjusted with Distilled water

(upto 10ml)

concentration 1mg/ml 12

Dilutions of topiramate in Distilled water + 1ml 10% pyridine solution + 1ml 2% ninhydrin solution

20 mins.

Cooling at R.T.

violet color

spectra taken (λ =568nm)

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Spectra Of Colour Complex

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1. LINEARITY

2. ACCURACY

3. PRECISION

4. LIMIT OF DETECTION (L.O.D.)

5. LIMIT OF QUANTIFICATION (L.O.Q.)

6. RANGE

7. ROBUSTNESS15

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y = 0.0019x + 0.2184R² = 0.9963

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

0 50 100 150 200 250 300 350

Abs.

abs

Linear (abs)

Conc.

Conc. (µg/ml) Absorption

50 0.2999

100 0.413

150 0.5072

250 0.6988

300 0.7679

Fig. Calibration Curve for TOPIRAMATE at 568 nm

Table : Linearity of TOPIRAMATE AT 568 nm

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Level of recovery

Conc. Of drug

Amt. added

absorbance amt.recovered

% recovery

80 0.32 0.4 0.5613 0.321 80.47

80 0.32 0.4 0.5595 0.318 79.50

80 0.32 0.4 0.5599 0.319 79.75

100 0.4 0.4 0.6004 0.404 101.00

100 0.4 0.4 0.5998 0.402 100.50

100 0.4 0.4 0.5988 0.400 100.00

120 0.48 0.4 0.6365 0.480 120.00

120 0.48 0.4 0.6366 0.480 120.00

120 0.48 0.4 0.6370 0.481 120.25

Table: Statistical treatment of Recovery data for TOPIRAMATE

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Parameters Level of recovery Level of recovery Level of recovery

80% 100% 120%

Concentration added

(μg/ml)

0.32 0.4 0.48

Mean of Absorbance

0.5602 0.5996 0.6367

Mean Conc.

Recovered (μg/ml)

0.319 0.402 0.480

Mean % recovery

79.90 100.66 120.15

% RSD0.4785 0.4975 0.1201

Table: Recovery studies of TOPIRAMATE

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System precision :

Replicate Absorbance of TOPI

1 0.4383

2 0.4256

3 0.4204

4 0.4185

5 0.4130

6 0.4231

Mean 0.42315

SD 0.008576

% RSD 1.99

Table: System precision for TOPIRAMATE

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Conc.

(µg/ml)

Absorbance Mean Std dev % RSD

Trial 1 Trial 2 Trial 3

100 0.4886 0.4658 0.4557 0.4767 0.00182 0.3826

150 0.4919 0.4779 0.4740 0.4812 0.00941 1.9550

250 0.5855 0.5782 0.5722 0.5786 0.00666 1.1509

Table: Intra-day variability for TOPIRAMATE

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Conc.

(µg/ml)

Absorbance Mean Std dev % RSD

Day 1 Day 2 Day 3

100 0.4923 0.4687 0.4476 0.4695 0.00476 0.476

150 0.5019 0.4799 0.4777 0.5547 0.00133 0.275

250 0.5995 0.5812 0.5811 0.5872 0.00105 0.180

Table: Inter-day variability of TOPIRAMATE

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Limit of Detection (LOD)- 1.823 μg/ ml

3.3 σ

S

Limit of Quantification (LOQ)- 5.52 μg/ ml

10 σ

S

Range- 50 - 300 μg/ ml

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A. Analysis of tablet

Conc. Of drug (µg) Absorbance Amt. added µg/ml

Amt. found in %

100 0.4254 0.4 98.95

100 0.4320 0.4 102.02

100 0.4251 0.4 98.80

100 0.4292 0.4 100.76

100 0.4329 0.4 102.05

100 0.4331 0.4 102.01

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Conc. of drug Wavelength at 568 nm Wavelength at 570 nm

100 µg 0.4383 0.5602

100 µg 0.4256 0.5611

100 µg 0.4204 0.5615

100 µg 0.4185 0.5613

100 µg 0.4130 0.5612

Mean 0.42316 0.56106

Std deviation 0.009588 0.000503

% RSD 2.02 0.089

ROBUSTNESS

Table: Robustness studies

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1. Colorimetric method was developed and validated as per ICH

guidelines for estimation of Topiramate in tablets.

2. Development of color is by reaction of amino group of drug

with Ninhydrin reagent in presence of pyridine.

3. The method was found to be simple, accurate, precise and

specific.

So, the proposed method can be used for the routine quality

control analysis of the bulk drug as well as oral dosage forms.

Parameters Result

Linearity > 0.99

% Recovery 98 – 102 %

% RSD < 2

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1. Davidson, A.G., (2002), Ultraviolet-Visible absorption spectrophotometry, In; Beckett,

A.H., Stenlake, J.B., Practical pharmaceutical chemistry, 4 th edi., Part-II, CBS

Publishers and Distributors, New Delhi, 278- 82.

2. USP 24/ NF 19,The United states pharmacopoeia 24 and National formulary 19,

(2000), the United States Pharmacopoeial Convention, Rockville, 1767-69.

3. National Organization for Drug Control and Research, 6 Abou Hazem Street,

Pyramids Ave, P.O.Box 29 Giza, `Egypt. (NODCAR)

Nahla Nour El-Din Ahmed Salama* et al /International Journal Of Pharmacy.

4. Sanket S.D.1*, Madhuri V. G.1, Shilpa V.1, Mahesh M.D.2, Vikas V.G. International

Journal of Pharmaceutical Sciences and Nanotechnology, Volume 2 Issue 4 January –

March 2010

5. Davidson,A.G.(2002),Ultraviolet-Visible-absorption spectrophotometry, In; Beckett,

A.H., Stenlake, J.B., Practical pharmaceutical chemistry, 4 th edi., Part-II, CBS

Publishers and Distributors, New Delhi, 275- 78

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