Simultaneous Spectrophotometric Estimation of Drotaverine Hydrochloride and Diclofenac Potassium in Combined Dosage Form

 

Snehal S. Ingale, Sumit A. Abnawe, Santosh N. Shinde, Archana S. More, Vishnu P. Choudhari* and Bhanudas S. Kuchekar

Pharmaceutical Analysis and Quality Assurance Department, MAEER’s Maharashtra Institute of Pharmacy, MIT Campus, Paud Road, Kothrud, Pune, 411038, MS, India.

*Corresponding Author E-mail: viraj1404@rediffmail.com

ABSTRACT:

Two simple, rapid, accurate and economic spectrophotometric methods are described for the determination of Drotaverine Hydrochloride (DRO) and Diclofenac Potassium (DIC) in combined dosage form. The first method is First Derivative and second is Absorption Corrected method. The amplitudes at 248.92 nm and 242.64 in the first derivative spectra were selected to determine Drotaverine Hydrochloride (DRO) and Diclofenac Potassium (DIC), respectively in combined formulation. The second method was based on the absorption corrected method in which DRO and DIC exhibit λ_max at 358.41 nm and 277.27 nm, respectively in double distilled water. DRO has some interference due to DIC at 277.27 nm, while DIC do not show any absorption at 358.41 nm. Quantitative estimation of DIC was carried out by subtracting the absorption due to DRO at 277.27 nm using experimentally calculated absorption factor. Beer’s law was obeyed in the concentration range of 8-40 μg mL^-1 for DRO and 5-25 μg mL^-1 for DIC for both the methods. The results of analysis have been validated statistically and recovery studies confirmed the accuracy and reproducibility of the proposed methods which were carried out by following ICH guidelines.

 

 

INTRODUCTION:

Drotaverine Hydrochloride, 1-[(3,4-diethoxy phenyl) methylene]-6,7-diethoxy-1,2,3,4-tetra hydro isoquinolene is an analogue of papaverine (Fig.1). It acts as an antispasmodic agent by inhibiting phosphodiesterase IV enzyme, specific for smooth muscle spasm and pain, used to reduce excessive labor pain¹. Diclofenac Potassium is chemically potassium (o-(2, 6-dichloroanilino) phenyl) acetate, a non steroidal anti-inflammatory drug (NSAID) exhibits anti-inflammatory and analgesic properties² (Fig.1). The exact mechanism of action of DIC is not entirely known, but it is thought that the primary mechanism responsible for its anti-inflammatory, antipyretic, and analgesic action is inhibition of prostaglandin synthesis by inhibition of cyclooxygenase (COX) and it appears to inhibit DNA synthesis³.

 

Literature survey reveals that various methods have been reported for estimation of DRO such as spectrophotometry, HPLC, HPTLC and voltammetry^1,4-6 individually and in combination dosage form. For DIC various analytical methods have been reported for its individual estimation and in combined dosage form and in human plasma which includes spectrophotometry, liquid chromatography with UV detection, HPLC with electrochemical detection^2,7-9.

 

Review of the literature revealed that no method has been reported for simultaneous analysis of DRO and DIC in its combination. Here an attempt has been made to develop simple, rapid and accurate First derivative and Absorption corrected spectroscopic methods for simultaneous estimation of DRO and DIC from its formulation. The proposed method was optimized and validated as per the International Conference on Harmonization (ICH) guidelines¹⁰.

 

MATERIALS AND METHODS:

Instrumentation:

An UV-Visible double beam spectrophotometer (Varian Cary 100) with 10 mm matched quartz cells was used. All weighing were done on electronic balance (Model Shimadzu AUW-220D). Ultrasonicator (Model 5.5 150H) was used for sample solution preparation.

 

        DICLOFENAC                             DROTAVERINE   

        POTASSIUM                             HYDROCHLORIDE

 Fig.1: Structures of the analytes

 

Reagents and chemicals:

Pure drug sample of DRO, % purity 99.86 and DIC, % purity 99.91 was kindly supplied as a gift sample by Alkem Pharmaceutical Pvt. Ltd. Mumbai and Aarti Drugs Pvt Ltd., Pune, respectively. These samples were used without further purification. Two tablet formulations (Lot 304 and 308) were supplied by JCPL Pharma Ltd., Jalgaon were used for analysis containing DRO 80mg and DIC 50mg per tablet. Spectroscopy grade methanol and double distilled water was used throughout the study. All the solvents and reagents used were purchased from LOBA Chemie Pvt. Ltd., Mumbai.

 

Preparation of Standard Stock Solutions and calibration Curve:

Standard stock solutions of pure drug containing 1000 μg mL^-1 of DRO and DIC were prepared separately in methanol. Working standard solutions of these drugs were obtained with suitable dilution with distilled water in the conc. range of 8 - 40μg/ml for DRO and 5 - 25μg/ml for DIC. Derivative amplitudes of spectrum, by using the above mentioned procedure and absorbance’s at selected wavelengths for absorbance corrected method, were used to prepare calibration curves for both the drugs.

 

Fig. 2 a:  Zero order UV spectra of solution containing   DRO (8 - 40 μg mL^-1) and DIC

(5 - 25μg mL^-1) in increasing and decreasing order in methanol/distilled water

 

PROCEDURE:

METHOD A: FIRST DERIVATIVE METHOD

Theoretical aspects:

To determine derivative amplitude for DRO and DIC, solution of increasing and decreasing concentrations of  DRO and DIC were prepared in combination and scanned on UV Spectrophotometer in the range 200 - 400 nm at 0.5 band width and 600 nm/min scan speed (Fig.2 a). These spectrums were converted to first order derivative spectra by using instrument mode with filter size 9 and interval 1.5 (Fig.2 b). Form the observations of the derivative spectrum, derivative amplitudes responsible for DRO and DIC were selected and wavelength for each amplitude was noted. These wavelengths were further confirmed by checking the first derivative amplitude of the mixed standard solutions of these drugs in the given ratio. Wavelengths 248.92 nm and 242.64 nm were selected for the quantification of DRO and DIC in mixture.^{11, 12, 14}

 

Fig 2 b.  First derivative spectra of zero order spectra shown in fig. 2 a.

 

METHOD B: ABSORPTION CORRECTED METHOD

λ_max of DRO and DIC was determined by scanning the drug solution in UV Spectrophotometer in the range 200 - 400 nm at 0.5 band width and 600 nm/min scan speed and was found to be at 358.41 nm and 277.27 nm respectively. DRO also showed absorbance at 277.27 nm, while DIC did not show any interference at 358.41 nm (Fig. 3). To construct Beer’s plot for DRO and DIC, stock solutions of 1000 μg/ml of both the drugs were prepared in methanol and working standard dilutions were made in double distilled water using stock solution of 1000 μg/ml. Also Beer’s plot was constructed for DRO and DIC in solution mixture at different concentration (8:5, 16:10, 24:15, 32:20, 40:25 μg/ml) levels. Both the drugs followed linearity individually and in mixture within the concentration range 8 - 40 μg/ml and 5 - 25 μg/ml for DRO and DIC respectively^{13, 14}.

 

Fig. 3: Overlay spectrum of DIC and DRO in methanol /distilled water. DRO (8 - 40µg   mL^-1) and DIC (5 - 25µg   mL^-1)

 

Determination of Absorption Factor at Selected Wavelengths:

DRO and DIC solution in double distilled water of known concentrations were scanned against blank on spectrophotometer. The value of absorption factor was found to be 0.98. Quantitative estimation of DRO and DIC was carried out using following equation:

Table 1: Optical characteristics of the proposed methods and results of formulation analysis &  precision study

Parameter		DRO		DIC
		Method A	Method B	Method A	Method B
λ (nm)		248.92	358.41	242.64	277.27
Beer’s law limit (μg mL-1)		8 - 40	8 - 40	5 - 25	5 - 25
Regression Equation (y = mx + c)	Slope (m)	0.164	0.02096	0.09916	0.03324
	Intercept (c)	0.112	-0.013	0.0448	-0.0145333
Correlation coefficient		0.9993	0.9993	0.9990	0.9993
Precision (%RSD)	Repeatability (n=6)	0.42	0.59	0.75	0.63
	Intra-day(3×3 times)	0.23	0.97	0.35	1.05
	Inter-day(3×5 days)	0.99	0.87	0.65	0.53
	Analyst	0.67	0.89	1.23	0.58
Formulation Analysis (%Assay, % RSD) n=6	Tablet  I	99.16, 0.87	98.18, 1.11	100.05, 1.05	101.02, 1.02
	Tablet  II	100.23, 0.73	99.51, 0.95	99.96,  0.54	101.08, 1.06

RSD is relative standard deviation

 

Table2 (A): Result of the recovery analysis by First Derivative Method (Method A)

Formulation	Recovery Level (%)	Amount  spiked (μg mL-1)		Recovery (%)		%RSD (n = 3)
		DRO	DIC	DRO	DIC	DRO	DIC
Tablet I	50	6	3.75	98.97	100.58	0.86	0.56
	100	12.0	7.5	101.04	99.70	1.23	0.91
	150	18.0	11.25	101.01	99.20	0.97	1.11
Tablet II	50	6.0	3.75	99.48	99.15	0.65	0.65
	100	12.0	7.5	99.73	99.63	0.78	0.73
	150	18.0	11.25	100.06	100.66	0.33	0.36

 

Table2 (B): Result of the recovery analysis by Absorbance corrected method (Method B)

Formulation	Recovery Level (%)	Amount  spiked (μg mL-1)		Recovery (%)		% RSD (n = 3)
		DRO	DIC	DRO	DIC	DRO	DIC
Tablet I	50	6	3.75	100.13	98.22	0.69	1.31
	100	12.0	7.5	100.87	99.20	1.51	1.63
	150	18.0	11.25	99.96	100.31	0.32	1.22
Tablet II	50	6.0	3.75	100.72	99.15	1.26	0.96
	100	12.0	7.5	99.16	98.72	0.37	0.79
	150	18.0	11.25	100.04	100.14	0.58	1.17

 

Corrected Absorbance of DIC at 277.27nm =

Abs_277.27 (DIC + DRO) – [(abs_277.27 (DIC)/ abs_358.41 (DRO)] × abs_358.41 (DRO) or

Corrected Absorbance of DIC at 277.27nm = abs_277.27 (DIC + DRO) – 0.98 × abs_358.41 (DRO)

Where; abs: Absorption value at given wavelengths.

 

Preparation of Sample Stock Solution and Formulation analysis:

Twenty tablets were weighed accurately and a quantity of tablet powder equivalent to 50 mg of DIC (80 mg of DRO) was weighed and dissolved in 40 mL of methanol with the aid of ultrasonicator for 5 min and solution was filtered through Whatman paper No. 41 into a 50 ml volumetric flask. Filter paper was washed with methanol, adding washings to the volumetric flask and volume was made up to mark. The solution was suitably diluted with distilled water to get of 15μg/ml of DIC (24 μg/ml of DRO) and proposed methods were followed for analysis.

 

Recovery studies:

The accuracy of the proposed methods was checked by recovery study, by addition of standard drug solution to preanalysed sample solution at three different concentration levels (50%, 100% and 150%) within the range of linearity for both the drugs. The basic concentration level of sample solution selected for spiking of the drugs standard solution was 12 μg/ml of DRO and 7.5 μg/ml of DIC for both the methods.

 

Precision of the Method:

Reparability of the methods was studied by repeating the methods six times. To study intraday precision, method was repeated 3 times in a day and the average % RSD was found to be 0.23 and 0.97 for DRO by method A and B, respectively; 0.35 and 1.05 for DIC by method A and B, respectively. Similarly the method was repeated on five different days and average % RSD was found to be 0.99 and 0.87 for DRO by method A and B, respectively; 0.65 and 0.53 for DIC by method A and B, respectively. These values confirm the intra and inter day precision.

 

RESULTS AND DISCUSSION:

Practically no interference from tablet excipients was observed in these methods. As their λmax differ more than 20 nm, absorption corrected method was tried for their simultaneous estimation in formulation. Quantitative estimation of DIC was carried out by subtracting interference of DRO using experimentally calculated absorption factor. Both the methods are accurate, simple, rapid, precise, reliable, sensitive, reproducible and economical as per ICH guidelines. The values of % RSD and correlation of coefficient were satisfactory and results of the formulation analysis in Table 1 and result of the recovery study in Table 2(A) and 2(B) indicates that there is no interference due to excipients present in the formulation. It can be easily and conveniently adopted for routine quality control analysis.

 

CONCLUSION:

The proposed methods are simple, precise, accurate and rapid for the determination of DRO and DIC in combined tablet dosage forms. Analysis of authentic samples containing DRO and DIC showed no interference from the common additives and excipients. Hence, recommended procedure is well suited for the assay and evaluation of drugs in pharmaceutical preparations. It can be easily and conveniently adopted for routine quality control analysis.

 

ACKNOWLEDGEMENT:

The authors are thankful to Alkem Pharmaceutical Pvt. Ltd., Mumbai and Aarti Drugs Pvt Ltd., Pune, India and for providing gift samples of Drotaverine Hydrochloride and Diclofenac Potassium, respectively. The authors are thankful to Management of MAEER’s Maharashtra Institute of Pharmacy, Pune for providing necessary facility for the work.

 

REFERENCES:

1.       Prasad Prakash Dahivelkar, Sanjay Baburao Bari, Suvarna Bhoir and Ashok Mukund Bhagwat. High Performance Liquid Chromatographic Estimation of Drotaverine Hydrochloride and Mefenamic Acid in Human Plasma. Iranian Journal of Pharmaceutical Research. 2009; 8 (3); 209-215.

2.       B.Gowramma, S. Rajan, S. Muralidharan, S. N. Meyyanathan and B. Suresh. A validated RP-HPLC method for simultaneous estimation of Paracetamol and Diclofenac Potassium in pharmaceutical formulation. Int.J. ChemTech Res. 2010; 2(1); 676-680.

3.       Dutta NK, Annadurai S, Mazumdar K, Dastidar SG, Kristiansen JE, Molnar J, Martins M, Amaral L. The anti-bacterial action of diclofenac shown by inhibition of DNA synthesis. Int. J. Antimicrob. Agents. 2000; 14 (3); 249–51.

4.       Kirti S Topagi, Rajesh M Jeswani, Purushotam K Sinha, Mrinalini C Damle. A validated normal phase HPLC method for simultaneous determination of drotaverine hydrochloride and omeprazole in pharmaceutical formulation. Asian Journal of Pharmaceutical and Clinical Research. 2010; 3(1); 20-24.

5.       L.P.Kothapalli, V.C. Dewoolkar, A.G. Banerjee, A.B. Thomas, R.K. Nanda, A.D. Deshpande, V.A.Hurne. Simultaneous Spectrophotometric estimations of Drotaverine Hydrochloride and Omeprazole. International Journal of ChemTech Research. 2010; 2(1); 493-498.

6.       S.I.M. Zayed, Y.M. Issa. Cathodic adsorptive stripping voltammetry of drotaverine hydrochloride and its determination in tablets and human urine by differential pulse voltammetry.  Bioelectrochemistry. 2009; 75; 9-12.

7.       Aleksandra Chmielewska, Lucyna Konieczna, Alina Plenis, Micha¬ Bieniecki and Henryk Lamparczyk. Determination of diclofenac in plasma by high-performance liquid chromatography with electrochemical detection. Biomed. Chromatogr. 2006; 20; 119–124.

8.       R. Sanjay Kumar, C. Karthikeyan, N. S. H. N. Moorthy and P. Trivedi. New Spectrophotometric methods Applied to the Simultaneous determination of Diclofenac Potassium and Tizanidine. Indian Journal of Pharmaceutical Sciences, May-June 2006; 68 (3); 317-322.

9.       G Subramanian, P Musmade, S Agarwal, N Udupa. Simultaneous RP HPLC estimation of tizanidine, diclofenac potassium and paracetamol in tablets. Indian Journal of Pharmaceutical Sciences. 2004; 66 (5); 694-696.

10.     ICH-Q2B Validation of Analytical Procedures: Methodology International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use, Geneva, Switzerland, 1996.

11.     Mannucci C, Bertini J, Cocchini A, Perico A, Salvagnini F, Triolo A. Simultaneous determination of otilonium bromide and diazepam by first-derivative spectroscopy. Indian Journal of Pharmaceutical Science 1992; 81(12); 1175-1177.

12.     Nanda RK, Gaikwad J and Prakash A. Simultaneous Spectrophotometric Estimation of Tolterodine in Pharmaceutical Dosage form. Research J. Pharm. and Tech. April.-June.2009; 2 (2); 312-314.

13.     PS Tarte, SP Wate, PB Khedikar and G Pawnikar. Absorption Correction Method for Estimation of Nebivolol and Hydrochlorothiazide in Combined Tablet Dosage Form.  Asian J. Research Chem. Oct.-Dec. 2008; 1(2); 74-76.

14.     Stenlake J. B. and Backett A. H.; Practical pharmaceutical Chemistry; C. B. S. Publishers and Distributors; New Delhi; 1997; 4th Ed., Part II, 281-306.

 

 

 

Received on 03.04.2010       Modified on 11.05.2010

Accepted on 31.05.2010      © RJPT All right reserved