INTRODUCTION:

Flunarizine dihydrochloride [FLU] chemically is (E)1[Bis(4fluorophenyl)methyl]-4-(3-phenyl-2-propenyl) piperazine dihydrochloride. It is a calcium channel blocker which reduces arterial and arteriolar smooth muscle spasm by reducing intracellular Ca2+ overload due to brain hypoxia. It is used in migraine prophylaxis and also as antihistaminic and sedative. FLU and PRO are official in B.P.¹ and I.P.² respectively. Propranolol HCl [PRO] chemically is (2RS)-1-[(1methylethyl) amino]-3-(1- naphthalenyloxy)-2-propanol hydrochloride. It is a non-selective beta blocker, that is, it blocks the action of epinephrine on both β1- and β2-adrenergic receptors. It is used for the treatment of angina pectoris, cardiac arrhythmia, hypertension, anxiety attacks, migraine prophylaxis and glaucoma. FLU and PRO these drugs are used in combination for migraine prophylaxis³. Literature survey reveals that various analytical methods like UV^4-5, HPLC^6-8, HPTLC⁹ and GC¹⁰ are reported for the individual drug and in combination with others.

Therefore, in the present work asuccessful attempt has been made to estimate both these drugs simultaneously by UV- spectrophotometric absorbance correction method using methanol and 0.1N HCl. The present paper describes a simple, accurate, precise and economic method for simultaneous determination of FLU and PRO in combined capsule dosage form.

MATERIALS AND METHOD:

Instrument, chemicals and reagents:

Pure drugs of FLU and PRO were procured as gift samples from FDC India Ltd. and Ipca Laboratories Ltd. Mumbai, India respectively. A.R. Grade Methanol from LOBA Chem, was used as the solvent for preparing solutions. The solution of 0.1N HCl was prepared in double distilled water as per IP 1996 procedure. The commercial formulation of FLU and PRO in ratio 1:4 (Betacap plus 10, FLU 10 mg and PRO 40 mg per capsule) were purchased from local market. The instrument used was Shimadzu double beam UV/Vis spectrophotometer model V- 1800. (UV Probe 2.32 software).Weighing was done on electronic single pan weighing balance (Make: Shimadzu Model: AX 200). Standard stock solutions 1 mg/ml of FLU and PRO were prepared separately in methanol. The aliquot portions of standard stock solutions of FLU and PRO were diluted with 0.1N HCl individually and combination to get series of concentration from 1-25 μg/ml for FLU and 4-100 μg/ml for PRO.

Procedure:

Absorbance correction method is based upon determination of identity, concentration and absorptivity of the absorbing interferents and finaly its contribution is calculated from the total absorbance of the mixture. Solutions of FLU and PRO (10μg/ml each) were prepared separately by appropriate dilution of standard stock solutions and scanned over the range of 400 to 200 nm.

Figure 1: Overlain spectra of FLU (2 μg/ml)and PRO (8 μg/ml)

From the overlain spectrum (figure1) the wavelengths selected for estimation of drugs were 288.8 nm as detecting wavelength for PRO and 253.2 nm as detecting wavelength for FLU. Both these drugs obeyed Beers law individually and in mixture within concentration range of 1-25 μg/ml for FLU and 4-100 μg/ml for PRO. The absorptivity values (A 1%, 1 cm) are recorded in Table 1. The sample absorbance, absorptivity and corrected absorbance were determined and finally the concentration of each drug was calculated using following equation

C=A/A (1%, 1 cm)

Where, C, A, A (1%, 1 cm) are the concentration, absorbance and absorptivity at selected wavelengths for selected drugs.

Application of proposed method to marketed formulation:

20 capsules were accurately weighed, average weight was calculated, finely powdered and mixed thoroughly. An accurately weighed capsule powder equivalent to about 2 mg of PRO was transferred to 25 ml volumetric flask. The contents were shaken for 15 minutes with methanol and volume was adjusted upto the mark with same. The solution was filtered through Whatman No.1 filter paper. An accurately measured 1 ml portion of filtrate was further diluted to 10 ml with 0.1 N HCl. The absorbance of the above solution was then measured at 288.8 nm and 253.2 nm in 1 cm cell against 0.1 N HCl as blank. Five replicate estimations were done in similar way. The contents of FLU and PRO were calculated and % label claim was determined. Observations and results of estimation of FLU and PRO in marketed formulation analysis are given in Table 2.

Table 1: (A 1%, 1 cm) values of FLU and PRO at selected wavelengths

Sr No.	FLU	PRO
Sr No.	253.2 nm	253.2 nm	288.8 nm
1	440	50	216.7
2	435	51.2	217.5
3	436.6	52.2	216.6
4	435	51.8	216.2
5	436	50.5	217
Mean± S.D.	436.52± 2.062038	51.14± 0.904434	216.8± 0.484768

Validation:

Validation of proposed method was carried out for the following parameter as per ICH/USP 16 guidelines. Accuracy of the proposed method was ascertained on the basis of recovery studies, performed by standard addition method. Accurately weighed quantities of preanalysed capsule content equivalent to 2 mg was taken in series of 25 ml volumetric flask and to them known amount of FLU and PRO were added at different concentration levels so as to produce solutions containing 80%, 100% and 120% of the label claim. Percentage recovery was calculated. The results are shown in Table 3. Precision of the above method was ascertained by replicate estimation of drugs for marketed formulation. The results are shown in Table 2.

The ruggedness of the method was studied under three different parameters. Intraday variation (samples were analysed at different times on the same day), Interday variation (samples were analysed at three different days 1^st, 3^rd and 5^thday) and different analyst (samples were analysed by three different analysts as per the proposed method). The results are shown in Table 4.

Table 2: Observations and results of marketed formulation analysis

Sr. No.	Wt of capsule content (gm)	Absorbance		% Label claim
		253.2 nm	288.8 nm	FLU	PRO
1	0.00745	0.127	0.173	99.86	99.86
2	0.00743	0.127	0.175	98.00	100.75
3	0.00740	0.124	0.171	96.25	99.39
4	0.00748	0.124	0.171	96.00	99.12
5	0.00741	0.122	0.170	95.25	98.26
Mean± S.D. R.S.D.				96.478± 1.03376 0.01071	99.40± 0.89647 0.00902

Table 3: Observations, results and statistical data for recovery studies

Sr. No.	Wt of capsule powder(gm)	Amount added μg/ml		Absorbance		% Recovery
		FLU	PRO	253.2 nm	288.8 nm	FLU	PRO
1	0.00748	1.6	6.4	0.153	0.312	99.38	99.19
2	0.00740	2	8	0.257	0.349	100.40	100.97
3	0.00742	2.4	9.6	0.281	0.381	97.67	99.90
Mean± S.D. R.S.D.						99.15± 1.379456 0.01391	100.02± 0.896047 0.00896

Table 4: Results of Ruggedness studies of proposed method

Parameters	FLU* (Mean± S.D.)	PRO* (Mean± S.D.)
Intraday	96.84±1.1806	100.18±0.6214
Interday	96.52±1.4813	98.97±0.5026
Different analyst	97.32±2.0468	99.72±1.7178

*Results are mean of three determinations, S.D. is standard deviation

For linearity and range accurately weighed quantities of capsule powder equivalent 80, 90, 100, 110, 120 % of label claim were taken and dilutions were made to obtain concentration in the range of 80-120% of test concentration. The absorbance of final solutions were read at 253.2 nm and 288.8 nm. Graph was plotted as % test concentration Vs absorbance shown in Figure2 FLU and PRO were found to be linear in the range 80-120% of the test concentration.

Figure 2: plot of linearity and range.

RESULTS AND DISCUSSION:

A quick and accurate absorbance correction method for simultaneous determination of FLU and PRO was developed. Accuracy of the proposed method was ascertained by recovery studies. % recovery with standard deviation were found to be 99.15±1.37 and 100.02±0.89 for FLU and PRO respectively. The results of marketed formulation analysis were in good agreement with labeled amounts indicating high degree of precision. The % RSD for ruggedness was less than 2% indicated the method was robust enough under different conditions.

CONCLUSION:

The applicability for the proposed method for quantitative estimation of FLU and PRO in commercial formulation gave accurate and precise results. Due to high sensitivity and simple sample preparation can be the part of undergraduate curriculum. Moreover spectrophotometric methods have obvious advantage over sophisticated instrumental analysis. Hence simple, economical and less time consuming spectrophotometric methods always have a role in routine pharmaceutical analysis.

ACKNOWLEDGEMENTS:

The authors are gratefully acknowledging FDC India Ltd. Jogeshwari (Mumbai, India) for providing the gift sample FLU and Ipca Laboratories Ltd. For providing the gift sample PRO. Authors are also thankful to Vidyabharati College of Pharmacy, Amravati for providing necessary facilities for the research work.

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Received on 12.04.2013 Modified on 19.04.2013

Asian J. Research Chem. 6(5): May 2013; Page 514-516