INTRODUCTION:

Cetirizine hydrochloride is chemically known as [2-[4-[(4-chlorophenyl)phenylmethyl]-1piperazinyl] ethoxy acetic acid hydrochloride (fig.1). It is a white or almost white powder, freely soluble in water, ethanol, methanol, practically insoluble in acetone and dichloromethane¹. Cetirizine is a highly potent long-acting peripheral H1-receptor antagonist which acts both on the early and late allergic response ². Phenylpropanolamine hydrochloride is chemically known as 2-amino-1-phenylpropan-1-ol hydrochloride (fig.2). It is a white or almost white crystalline powder, soluble in water and ethanol (96%), practically insoluble in dichloromethane³ Phenylpropanolamine hydrochloride is an orally active sympathomimetic amine and exerts a decongestant action on the nasal mucosa.²

Cetirizine hydrochloride with Phenylpropanolamine hydrochloride is indicated for the relief of symptoms of allergic rhinitis. As per literature survey, no RP-HPLC method has been reported for the simultaneous estimation of Cetirizine hydrochloride and Phenylpropanolamine hydrochloride in combined dosage form. Although several UV ^4-12and HPLC^13-19, HPTLC²⁰methods have been reported for individual drug or in other combination.

Thus, focus of present study is to develop and validate a simple, rapid, accurate, precise RP-HPLC method for simultaneous estimation of Cetirizine hydrochloride and Phenylpropanolamine hydrochloride in tablet dosage form.

Fig .1 Structure of Cetirizine hydrochloride

Fig .2 Structure of Phenylpropanolamine hydrochloride

MATERIAL AND METHODS:

All chemicals and reagents used were of analytical grade and obtained from Loba chemie. Cetirizine hydrochloride and Phenylpropanolamine hydrochloride standards were procured as a gift sample from Cipla Ltd. Mumbai. Marketed formulation (Alerid D, Cipla India Ltd.) containing Cetirizine hydrochloride 5mg and Phenylpropanolamine hydrochloride 25mg was procured from local market and used for analysis.

The instrument used in analysis was SHIMADZU LC20A/10Avp/10A series High performance chromatograph.

Chromatographic condition

Column : Phenomenex 100-5 C18

Dimension : 250mm x 4.6mm

UV detector : SPD-20AD,

Pump : LC -20A

Stationary phase : C18 Phenomenex

Mobile phase :Water:methanol: triethylamine (20:80:0.5) pH 3.5 adjusted

with orthophosphoric acid

Detection Wavelength : 254 nm

Injection volume : 20 µL

Flow rate : 1 mL/min

Temperature : Ambient

Standard and sample preparations:

A. Standard Solutions:

i) Solution A (CET):

An accurately weighed quantity of 10 mg of CET was dissolved in methanol and volume was made up to 10 mL with same solvent. A 0.1 mL of resultant solution was diluted to 10 mL with methanol (Conc. 10 mg /mL).

ii) Solution B (PPA):

An accurately weighed quantity of about 10 mg of PPA was dissolved in methanol and diluted to 10 mL. A 0.5 mL of resultant solution was diluted to 10 mL with methanol (Conc.50 mg /mL).

iii) Mixed standard stock solution C:

An accurately weighed quantity of CET ( 10 mg) and PPA (50 mg) were transferred to 50 mL volumetric flask and dissolved in about 25 mL methanol and the volume was made up to the mark with methanol.

iv) Working mixed standard solution C1:

From the above solution 0.5 mL was further diluted to 10 mL by methanol to get concentration of 10 mg /mL and 50 mg /mL for CET and PPA respectively.

B. Sample preparations:

Twenty tablets were weighed and finely powdered. An accurately weighed quantity of tablet powder equivalent to about 5 mg of CET and 25 mg of PPA was transferred to 50 mL volumetric flask, sonicated with 25 mL methanol for 15 min. The volume was made up to the mark with methanol and filtered through whatman filter paper. Further dilutions were carried out to get final concentration 10 μg/mL of CET and 50 μg/mL of PPA was used as sample solution.

Analysis of Standard Laboratory Mixture:

Five replicate injections of mixed working standard solution C1 were injected and the chromatograms were recorded for both the drugs.

Analysis of Marketed Formulation:

Five replicate injections of sample solution were injected and the chromatograms were recorded. Typical chromatogram is shown in fig.3.

TABLE 1: ANALYSIS OF MARKETED FORMULATION

Sr. No.	Sample	Particulars	Estimation (%)
			CET	PPA
1.	Alerid-D	Mean (n=5)	99.50	99.46
		SD	0.037	0.041
		RSD(%)	0.0003	0.0004

CET : Cetirizine hydrochloride

PPA : Phenylpropanolamine hydrochloride

Fig 3 : Chromatogram of marketed formulation

Retention time:

4.607 for Cetirizine hydrochloride

2.808 for Phenylpropanolamine hydrochloride

Validation:²¹

The developed method was validated as per ICH and USP requirement in terms of linearity, accuracy, precision, limit of detection .limit of quantification, robustness.

Linearity:

Linearity evaluates the analytical method ability (within a given range) to obtain a response that directly proportional to concentration (amount) of analyte in the sample

Preparation of Calibration Curve:

The chromatographic conditions were set as per the optimized parameters and mobile phase was allowed to equilibrate with stationary phase as was indicated by the steady baseline. Standard solutions of concentration 5-40μg/mL for CET and 40-100 μg/mL for PPA were injected separately and the chromatograms were recorded. Peak areas were recorded for each injected concentration of drugs and the calibration curves, concentration Vs peak area were constructed for both the drugs.

Limit of Detection:

The limit of detection (LOD) was calculated using following formulae:

LOD = 3.3(SD)/S

Where

SD=standard deviation of response (peak area)

S= average of the slope of the calibration curve.

Limit of Quantitation:

The limit of quantitation (LOQ) was calculated using following formulae:

LOQ = 10 (SD)/S

Where

SD=standard deviation of response (peak area)

S= average of the slope of the calibration curve.

Accuracy:

Accuracy of proposed method was ascertained on the basis of recovery studies performed by standard addition method at different levels of labeled claim (i.e. 80 to 120 % of labeled claim). The percent recovery was calculated by using following formula:

Where,

A= Total amount of drug estimated, mg

B = Amount of drug contributed by tablet powder, mg

C = Amount of pure drug added, mg

Precision:

Precision of an analytical method is expressed as S.D. and % R.S.D. of series of measurement. It was ascertained by replicate estimation of both drugs by proposed method.

TABLE 2 : VALIDATION PARAMETER

Sr. no	Method characteristics	CET	PPA
1	Linearity performance parameter
	Linearity range	5-40 μg/ mL	40-100 μg/ mL
	Correlation coefficient	0.9995	0.9999
	LOD	0.00015 μg/ mL	0.00125 μg/ mL
	LOQ	0.00047 μg/ mL	0.0037 μg/ mL
2	Accuracy (% recovery)
	80%	99.95	100.00
	100%	99.98	99.99
	120%	99.98	100.00
3	Precision (% RSD)
3	Repeatability of peak area	0.0003	0.0004
4	Ruggedness (%)
	Intraday	100.04	100.13
	Interday	100.68	101.07
	Different analyst	99.92	99.76
5	Specificity (%)
	Normal	100.04	100.12
	0.1NHCl	86.33	82.14
	0.1N NaOH	72.07	69.08
	3% H₂O₂	91.28	90.05
	Thermal	57.15	51.09
6	System suitability parameter
	Retention time	4.604	2.812
	Theoretical plates	5059.96	2614.70
	Tailing factor	1.736	1.565

CET: Cetirizine hydrochloride

PPA: Phenylpropanolamine hydrochloride

Specificity:

The specificity studies were carried out by attempting deliberate degradation of the tablet sample with exposure to stress conditions like acidic (0.1 N HCl), alkaline (0.1 N NaOH), oxidizing (3% H₂O₂), heat (60 ⁰C).

Ruggedness:

The Ruggedness studies were carried out for different parameters i.e. different elapsed times (Intraday and Interday) and different analysts

RESULT AND DISCUSSION:

A RP-HPLC method was developed for CET and PPA, which can be conveniently employed for routine quality control in pharmaceutical dosage forms. The chromatographic conditioned were optimized in order to provide a good performance of assay. The mobile phase selected for each drug was selected based on its polarity. Different ratios of acetonirile : water: methanol, methanol : water, acetonitrile : water were tried and final working mobile phase water : methanol : triethylamine (20:80:0.5v/v/v) pH 3.5 by orthophosphoric acid was selected. The optimum wavelength selected was 254 nm. Under the chromatographic conditions, CET and PPA peaks were well resolved, retention times of CET and PPA was 4.604 and 2.812 min, respectively. The total run time was short for two drugs. Typical chromatogram is shown in fig.3.

The method were specific as none of the exicipient interfered with the analytes of interest. The results in Table 1 revealed that the percentage estimated in tablet dosage form was found to be 99.50 for CET and 99.46 for PPA respectively. All the validation parameter shown Table 2 to be within specified limit. Linear correlation obtained over the range of 5-40μg/mL for CET and 40-100 μg/mL for PPA with correlation coefficient 0.9995 for CET and 0.9999 for PPA. The limit of detection was found to be 0.00015 μg/mL for CET and 0.00125 μg/mL for PPA, whereas Limit of quantitation was found to be 0.00047 μg/mL for CET and 0.0037 μg/mL for PPA. The results in Table 2 revealed excellent accuracy and precision of assay method. The proposed method used for simultaneous determination of drug in combination from pharmaceutical dosage form after spiking with 80%, 100% and 120% of additional drug afforded recovery 99 - 100% as listed in Table 2 The ruggedness of the method was determined by intraday, interday and different analyst studies. The system suitability test for CET and PPA was found to be within specified limit.

CONCLUSION:

The developed RP-HPLC technique is simple, precise, specific and accurate and from statistical data, it is found that method is reproducible and selective for analysis of Cetirizine hydrochloride and Phenylpropanolamine hydrochloride in tablet dosage form.

AKNOWLEDGMENT:

The authors thank to M/S Cipla Ltd. Mumbai for providing the gift sample of pure Cetirizine hydrochloride and Phenylpropanolamine hydrochloride. The authors are grateful to the Principal Dr. K.P.Bhusari of Sharad Pawar College of Pharmacy Nagpur for providing all necessary facilities.

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Received on 26.06.2011 Modified on 30.06.2011

Research J. Pharm. and Tech. 4(9): Sept. 2011; Page 1471-1474

Mean (n=5)