INTRODUCTION:

Chemically, Meclizine (MEC) is (RS)-1-(4-chlorobenzhydryl)-4-(3-methylbenzyl) piperazine hydrochloride ^1-4. It is a potent antihistaminic agent used in treatment of vertigo and motion sickness ⁵. Caffeine (CAF) is 3,7-dihydro-1,3,7-trimethyl-1H-purine-2,6-dione ^1-4. It is used as CNS stimulant. Structures of both the drugs (MEC and CAF) are shown in figure 1 and 2. Literature survey reveals that MEC and CAF are official in Indian Pharmacopoeia 2007 ¹, British Pharmacopoeia 2008 ²and United States Pharmacopoeia ³. Several analytical methods have been reported for estimation of MEC are spectrophotometry ^6-8, HPLC ^9-11, gas liquid chromatography ¹²and ion pair extraction ¹³. The analytical methods reported for estimation of CAF are spectrophotometry ^14-16, HPLC ^17-19, capillary electrophoresis ²⁰ and LC-MS ²¹. The present paper describes simple, accurate, specific and precise methods for simultaneous estimation of MEC and CAF in their combined tablet dosage form. The proposed methods are optimized and validated as per ICH guidelines ²².

In the present work, a successful attempt has been made to estimate both drugs simultaneously using two UV spectrophotometric methods (a) Absorbance Correction method and (b) Simultaneous equation method.

MATERIALS AND METHODS:

Instrument used was an UV-Visible double beam spectrophotometer (SHIMADZU UV-1800) with a pair of 1 cm matched quartz cells. All weighing was done on analytical balance (Shimadzu AU-220). A pure drug sample of MEC was obtained as gift sample from Sidmak Laboratories, Valsad. Combined tablet formulation (PREGNIDOXINE) was procured from local market. Methanol AR was used as solvent.

Preparation of standard stock solution:

Accurately weighed quantity of MEC (100 mg) and CAF (100 mg) was transferred to two separate 100 ml volumetric flasks, dissolved in methanol and diluted to the mark with same (stock solutions: 1000 μg/ml of MEC and 1000 μg/ml of CAF).

Preparation of working standard solution:

50 mg/ml of MEC solution was prepared by diluting 5 ml of stock solution upto 100 ml with methanol.

40 mg/ml of CAF solution was prepared by diluting 4 ml of stock solution upto 100 ml with methanol.

Figure 1: Chemical structure of Meclizine hydrochloride

Caffeine

Figure 2: Chemical structure of Caffeine

Absorbance correction method (method I):

Absorbance correction method uses the absorbances at two selected wavelengths, one at which both the drugs show considerable absorbance and other being the wavelength at which the first drug has practically nil absorbance. From the stock solutions, working standard solutions of MEC (15 μg/ml) and CAF (12 μg/ml) were prepared by appropriate dilution and were scanned in the entire UV range to determine the suitable wavelengths. Both the drugs were found to have considerable absorbance at 236 nm while at 288 nm only CAF has absorbance. The wavelengths selected for analysis were 236 nm and 288 nm for MEC and CAF respectively (Figure 3). A mixture of standard solutions ranging from 5-25 μg/ml of MEC and 4-20 μg/ml of CAF were prepared and the absorbances of solutions were measured at 236 nm and 288 nm. A calibration curve was prepared by plotting absorbance versus corresponding concentration of drug. The concentration of two drugs in sample solution was calculated by using following equations:

C_CAF= A₁/ a_x1……………………………..…………. (1)

……………………………….….... (2)

Where, A₁and A₂ are the absorbances of mixture at 288 nm and 236 nm respectively,

a_x1and a_x2 are absorptivities of CAF at 288 nm and 236 nm respectively,

a_y2isabsorptivity of MEC at 236 nm,

C_MECis concentration of MEC,

^CCAFis concentration of CAF.

Simultaneous Equation Method (method II):

From the stock solution of 1000 μg/ml MEC and 1000 μg/ml CAF in methanol, working standard solutions of the drugs were prepared by appropriate dilution. The solutions were scanned using the spectrum mode with medium scan speed in the range of 200-400 nm. From their spectra, 273 nm and 230 nm were selected for measurement of CAF and MEC respectively (Figure 4). Standard solutions were prepared having concentration 5-25 μg/ml for MEC and 4-20 μg/ml for CAF. The absorbances of solutions were measured at 230 nm and 273 nm using methanol as blank. A calibration curve was prepared by plotting absorbance against respective concentration.

The concentration of MEC and CAF can be obtained as

……………………………… (3)

……………..…………..…… (4)

Where, A₁and A₂ are the absorbances of mixture at 230 nm and 273 nm,

a_x1and a_x2 are absorptivities of CAF at 230 nm and 273 nm respectively,

a_y1anda_y2areabsorptivities of MEC at 230 nm and 273 nm respectively,

C_xis concentration of CAF,

^Cyis concentration of MEC,

Figure 3: Overlain zero order spectra of: (1) MEC and (2) CAF in methanol for Absorbance Correction Method.

Figure 4: Overlain spectra of: (1) MEC and (2) CAF in methanol for Simultaneous Equation Method.

Assay of tablet formulation by method I and II:

Twenty tablets were weighed and crushed to obtain a fine powder. An accurately weighed tablet powder equivalent to about 20 mg of CAF or 25 mg of MEC was transferred to 100 ml volumetric flask and dissolved in 50 ml of methanol. The volume was made up to the mark using methanol as solvent. The resulting solution was filtered through Whatman filter paper No 42 and 10 ml of filtrate was appropriately diluted with methanol to get concentration of 50 μg/ml of MEC and 40 μg/ml of CAF. This solution was further diluted with methanol to get concentration of 15 μg/ml of MEC and 12 μg/ml of CAF. Absorbance of sample solutions was measured at 236 and 288 nm for method I and 230 and 273 nm for method II. The concentration of two drugs in the sample was determined using Equations (1), (2), (3) and (4).

Method validation:

Linearity and range:

Aliquots of standard stock solutions of MEC and CAF were mixed in volumetric flasks and diluted with methanol to get final concentrations of mixtures in range of 5-25 μg/ml for MEC and 4-20 μg/ml for CAF. This calibration range was prepared five times and absorbances were measured at respective wavelengths for each drug separately.

Precision:

Precision of the methods was determined by performing interday variation, intraday variation and method repeatability studies. In interday variation, the absorbance of standard solutions of MEC (5-25 μg/ml) and CAF (4-20 μg/ml) were measured on five consecutive days. In intraday variation the absorbances were measured five times in a day. In repeatability study, three concentrations were analysed in triplicate.

Recovery studies:

To study the accuracy of the proposed methods, recovery studies were carried out by standard addition method at three different levels. A known amount of drug was added to preanalyzed tablet powder and percentage recoveries were calculated.

Ruggedness:

The data for ruggedness was obtained from two different analysts for MEC and CAF.

RESULTS AND DISCUSSION:

The proposed methods were validated according to ICH guidelines. The plot of absorbances versus respective concentrations of MEC and CAF were found to be linear in the concentration range of 5-25 μg/ml and 4-20 μg/ml respectively with correlation coefficient 0.9992 at 236 nm and 0.9995 at 288 nm for absorbance correction method (method I) (Fig-5 and 6). For Simultaneous equation method (method II) linearity range was same as for method I with correlation coefficient 0.9992 at 230 nm and 0.9997 at 273 nm. Precision was calculated as interday and intraday variations and % RSD was found to be less than 2 for both the methods (Table 1 and Table 2) (Fig-7 and 8). The accuracy of method was determined at 75, 100 and 125 % level. The % recovery ranges from 99.10 to 101.72 for both the methods (Table 3). The % RSD for ruggedness for MEC ranges from 0.15 to 0.73%, while for CAF it was found to be 0.20 to 1.93% for method I. For method II it ranges from 0.14 to 0.57% for MEC and 0.07 to 0.40% for CAF. The two _methods can be successfully _used for simultaneous estimation of MEC and C_{AF in their combined tablet dosage form.}Marketed tablets were analyzed and results obtained were in the range of 99-102 % (Table 4).

Figure 5: Calibration curve of standard MEC at 236 nm by Absorbance Correction Method

Figure 6: Calibration curve of standard CAF at 288 nm by Absorbance Correction Method

Table 1: Validation Parameters for Absorbance Correction Method

PARAMETERS	MEC	CAF
Linearity range	5-25 µg/ml	4-20 µg/ml
Correlation Coefficient	0.9992	0.9995
Precision	% RSD
Repeatability (n=3) Intraday (n=5) Interday (n=5)	0.10-0.13 0.10-0.52 0.09-0.42	0.32-0.40 0.23-0.75 0.25-1.23
% Recovery	99.10%-101.72	99.84-100.07
Ruggedness (% RSD)	0.15-0.73	0.20 – 1.93

*MEC- Meclizine; CAF-Caffeine; RSD-Relative Standard Deviation.

Table 2: Validation Parameters for Simultaneous Equation Method

PARAMETERS	MEC	CAF
Linearity range	5-25 µg/ml	4-20 µg/ml
Correlation Coefficient	0.9992	0.9995
Precision	% RSD
Repeatability (n=3) Intraday (n=5) Interday (n=5)	0.12-0.35 0.08-0.54 0.11-0.40	0.12-0.35 0.05-0.50 0.13-0.58
% Recovery	99.60-100.84	99.33-99.93
Ruggedness (% RSD)	0.14-0.57	0.07-0.40

*MEC- Meclizine; CAF-Caffeine; RSD-Relative Standard Deviation.

Figure 7: Calibration curve of standard MEC at 230 nm by Simultaneous Equation Method

Figure 8: Calibration curve of standard CAF at 273 nm by Simultaneous Equation Method

Table 3: Recovery studies

_{Name of Drug}	_{Amount of Drug Added (}µg/ml)	_Method I(AC)		Method II(SE)
_{Name of Drug}	_{Amount of Drug Added (}µg/ml)	_{% Rec} _overy*	_SD	_{% Rec} _overy*	_SD
MEC	15	101.21	0.0025	99.60	0.110
CAF	12	99.87	0.145	99.33	0.162
MEC	20	99.10	0.0045	99.70	0.140
CAF	16	100.07	0.080	99.93	0.173
MEC	25	101.72	0.0015	100.84	0.240
CAF	20	99.84	0.0737	99.90	0.194

*Mean of three estimations

AC-Absorbance Correction; SE-Simultaneous Equation;

MEC- Meclizine; CAF-Caffeine; SD-Standard Deviation.

Table 4: Results of simultaneous estimation of MEC and CAF in marketed Formulation by Method I and II.

Method	Labelled mg/tablet		Obtained % of label claim* + S.D.
Method	MEC	CAF	MEC	CAF
Method-I(AC)	25	20	101.80+0.251	99.10+0.368
Method-II(SE)	25	20	100.42+0.647	99.73+0.273

*Average of five determinations;

AC-Absorbance Correction; SE-Simultaneous Equation Method;

MEC- Meclizine; CAF-Caffeine; SD-Standard Deviation.

CONCLUSION:

The proposed methods give accurate and precise results for determination of MEC and CAF in marketed formulation (tablet) without prior separation and are easily applied for routine analysis. The most striking feature of both the methods is its simplicity and rapidity.

ACKNOWLEDGEMENT:

The authors are thankful to Sidmak Laboratories, Valsad for providing pure gift samples of meclizine hydrochloride. The authors are also thankful to the Principal, Maliba Pharmacy College for providing necessary facilities.

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Received on 05.05.2011 Modified on 17.05.2011

Asian J. Research Chem. 4(8): August, 2011; Page 1249-1253-706