Development and Validation of Derivative Spectrophotometric Method for Simultaneous Estimation of Diclofenac and Pantoprazole in Combined Capsule Dosage Form

Khushbu Naik*, Gunjansinh Parmar, Jalpesh Ahiya, Kunjan Bodiwala, Shailesh Shah

Department of Quality Assurance, Maliba Pharmacy College, Bardoli, Gujarat, India

*Corresponding Author E-mail: naikkhushbu20@gmail.com

ABSTRACT:

Derivative spectroscopy offers a useful approach for the analysis of drugs in multi-component mixtures. In this study a first-derivative spectroscopic method was used for the simultaneous determination of diclofenac and pantoprazole in combined capsule dosage form using the zero-crossing technique. The wavelengths 290.2 nm and 276.2 nm of first derivative spectrum were selected for the estimation of diclofenac and pantoprazole, respectively without mutual interference. The method was linear in the concentration range 15-75μg/ml and 5-25μg/ml for diclofenac and pantoprazole, respectively. Validation studies confirmed the accuracy and precision of the proposed method. The result of the formulation analysis shows that the proposed method can be successfully used for the simultaneous estimation of both the drugs in their combined capsule dosage form.

KEYWORDS: Diclofenac, Pantoprazole, Simultaneous determination, Derivative spectrophotometry, Zero crossing point.

INTRODUCTION:

Diclofenac sodium (DS); 2-[(2, 6-dichlorophenyl) amino] phenyl] acetate (Fig. 1) is a widely prescribed as non-steroidal anti-inflammatory agent^1,2. Several methods have been reported for the determination of DS in pharmaceutical preparations and biological fluids. These methods include: spectrophotometry, HPLC and HPTLC^3-7.

Pantoprazole (PAN); 5- (difluoromethoxy) - [[(3,4- dimethoxy-2-pyridiynyl) methyl] sulphinyl] - 1H benzimidazole (Fig. 2) is a gastric proton pump inhibitor^1,2. The methods reported for quantitative determination of pantoprazole in bulk or pharmaceutical formulations include colorimetry, spectroscopy, HPLC and HPTLC^8-14.

No spectrophotometric method was found in literature for the simultaneous estimation of Diclofenac and pantoprazole. The present paper describes a simple, rapid, accurate and precise first order derivative spectrophotometric method for the simultaneous estimation of DS and PAN in their combined capsule dosage form.

Fig. 1. Chemical structure of diclofenac

Fig. 2. Chemical structure of pantoprazole

MATERIALS AND METHODS:

Apparatus

Instrument used was UV-Visible double beam spectrophotometer, Shimadzu (model UV-1800) with a pair of 1 cm matched quartz cells. All weighing was done on analytical balance, Shimadzu (Model AU-220). Calibrated glass wares were used to prepare the solutions.

Reagents and chemicals

All reagents and solvents were of Analytical Reagent grade.

Both DS and PAN were kindly gifted by Torrent Research Centre, Ahmadabad. Marketed formulation, Dufex capsules containing 75 mg DS and 20 mg PAN, were procured from local pharmacy.

Preparation of standard solutions

Accurately weighed DS (25 mg) and PAN (25 mg) were transferred to separate 25 ml volumetric flasks, dissolved in and diluted to the mark with methanol to obtain standard solutions having concentration 1000 μg /ml.

Selection of analytical wavelengths

Standard solutions of DS (15, 30, 45, 60 and 75μg/ml) and PAN (5, 10, 15, 20 and 25μg/ml) were scanned in the range of 200 to 400 nm. Zero crossing points (ZCP) of both drugs were selected by converting both the spectra into first order derivative. ZCP of DS and PAN were found to be 276.2 nm and 290.2 nm, respectively (Fig. 3).

Fig. 3 First order derivative overlain spectra of DS(15μg/ml) and PAN (5μg/ml).

Stability of standard solutions

Stability of standard solutions, DS(15, 30, 45, 60 and 75μg/ml) and PAN (5, 10, 15, 20 and 25μg/ml) was checked by measuring their absorbance after 8 hrs of preparation at room temperature (25^o C).

Validation of Method¹⁵

The method was validated as per ICH guidelines Q2 (R1).

RESULTS AND DISCUSSION:

Zero-order absorption spectra of diclofenac and pantoprazole sodium showed overlapping peaks that interfere with their simultaneous determination in formulation. First derivative spectroscopy, based on a mathematical transformation of the zero-order spectra into the derivative spectra, allows a fast, accurate and precise resolution of a multicomponent mixture.

Stability of standard solutions

The standard solutions were found to be stable for the period of 8 hrs after preparation at room temperature (25^o C).

Validation of the proposed method

Linearity

The calibration curves were plotted over a concentration range of 15-75μg/ml for DS and 5-25μg/ml for PAN. The absorbance of solutions was measured at 290.2 nm (ZCP of PAN) and at 276.2 nm (ZCP of DS) for estimation of diclofenac and pantoprazole, respectively (Fig. 4 and 5).

Fig. 4. Calibration curve of DS at 290.2 nm (ZCP of PAN)

Fig. 5. Calibration curve of PAN at 276.2 nm (ZCP of DS)

Table 1: Recovery data of proposed method

Drug	Amount taken from sample (μg/ml)	Amount of Standard added (μg/ml)	% Recovery ± SD (n=3)
DIC	30	24	100.43 ± 0.68
	30	30	101.16 ± 0.56
	30	36	99.54 ± 0.74
PAN	10	08	99.23 ± 0.79
	10	10	101.52 ± 0.92
	10	12	99.28 ± 0.87

Table 2: Summary of Validation Parameters

Parameters	DS	PAN
Linearity Range (μg/ml)	15-75	5-25
Correlation coefficient	0.9953	0.9959
Precision (% RSD) Repeatability (n=6) Intraday (n=3) Interday (n=3)	0.46 0.34-1.18 0.42-1.47	0.62 0.38-1.12 0.54-1.72
Accuracy (% recovery)	99.54 -101.16%	99.23 - 101.52 %
LOD (μg/ml)	0.58	0.38
LOQ (μg/ml)	1.75	1.15

Accuracy

The accuracy of the method was determined by standard addition method. Known amount of standard solutions of DS and PAN were added to pre-quantified sample solutions of combined capsule dosage form. The amounts of DS and PAN were estimated by applying obtained values to the respective regression line equations and percent recovery of each drug was determined. Results of the recovery data are given in Table 1.

Method precision (repeatability)

The precision of the method was checked by repeated scanning and measurement of the absorbance of six different solutions of DS (45μg/ml) and PAN (15μg/ml) without changing the parameters of the proposed method (Table 2).

Intermediate precision (reproducibility)

The intraday and inter day precision of the proposed method was determined by estimating the corresponding responses 3 times on the same day and on 3 different days over a period of one week for 3 different concentrations of standard mixture solutions containing DS (30, 45, 60μg/ml) and PAN (10, 15, 20μg/ml) (Table 2).

Limit of detection and Limit of quantification

The limit of detection (LOD) and the limit of quantification (LOQ) of the drug were obtained using the following equations designated by (ICH) guidelines.

LOD = 3.3 × s/S

LOQ = 10 × s/S

Where,

s = the standard deviation of the response and

S = mean slope of 5 calibration curves.

Results of the validation parameters are presented in Table 2.

Assay of marketed formulation by proposed method

Twenty capsules were individually weighed, emptied and their contents were mixed. The powder equivalent to 75 mg DS or 20 mg PAN was accurately weighed and transferred to 100 ml volumetric flask. To this, about 50 ml of methanol was added and sonicated for 15 min. The volume was made up to the mark with methanol and mixed well. The solution was filtered through watman filter paper and 1 ml filtrate was diluted to 10 ml with methanol to give a solution containing 75μg/ml DS and 40μg/ml PAN. From this solution, 4 ml was further diluted to 10 ml with methanol to give a solution containing 30μg/ml DS and 16μg/ml PAN which was used for the estimation of DS and PAN. The results obtained for DS and PAN were compared with the corresponding labelled amounts (Table 3).

CONCLUSION:

The present study describes a simple, precise and accurate first derivative UV-Vis Spectroscopy method for the simultaneous estimation of diclofenac and pantoprazole. The method was successfully validated and applied for the estimation of the two drugs in their combined capsule dosage form.

ACKNOWLEDGEMENT:

The authors are thankful to Torrent Research Center, Ahmadabad and the Principal, Maliba Pharmacy College for providing all the resources for the experiment.

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Received on 21.01.2013 Modified on 08.02.2013

Asian J. Research Chem. 6(2): February 2013; Page 155-157

Formulation	Drug	Label claim (mg)	Amount found (mg) ± SD (n=3)	% Label claim ± SD (n=3)
Dufex Capsules	DS	75	74.42 ± 0.54	98.22 ± 0.78
Dufex Capsules	PAN	20	19.76 ± 0.38	98.80 ± 0.92