INTRODUCTION:

Chemically Rizatriptan Benzoate is N,N-dimethyl-5(1H-1,2,4-triazol-1-ylmethyl)-1H-indole-3-ethanamine monobenzoate (Figure 1)¹ and is a selective 5-hydroxytriptamin (5-HT)_1B/1D receptor agonist. It has weak affinity for other 5-HT receptor subtype and was launched in 1998 for acute treatment of migraine in adults.¹ The suggested mechanism of anti-migraine action of rizatriptan Benzoate is through activation of post synaptic 5-HT _1B receptors within cerebral and dural vessel walls, causing vasoconstriction of perivascular nerve terminals.²

Analysis is an important aspect of formulation development of any drug molecule. A suitable and validated method has to be available for the analysis of drug in the bulk, in drug delivery systems, for release dissolution studies and estimation of drug in biological samples.

The literature survey reveals that HPLC, Electrospray Liquid chromatography –Mass spectrometry (LC-MS), NMR and Liquid chromatography-atmospheric pressure chemical ionization mass spectrometry.^3-4 methods are available for analysis of Rizatriptan Benzoate. Although Altinoz et al⁵have reported UV estimation of Rizatriptan benzoate in aqueous solution, estimation of the drug in other solvents has not been reported. Quantitative estimation of drug in other fluids is often necessary for release study.

The objective of the study was to develop a simple, sensitive, rapid, precise, cost effective and reproducible UV method for estimation of Rizatriptan benzoate in 0.1 N HCl, as per ICH guidelines.⁶

MATERIALS AND METHOD:

A Jasco V-630 UV/Visible double beam spectrophotometer (model) with 1 cm matched quartz cells was used for spectral measurement. Schimadzu AX200 analytical balance was used for weighing purposes. The reference standard of Rizatriptan benzoate was procured as a gift sample from Cipla Pharmaceuticals (India) as a gift sample with 99.9% w/w assay value and Rizatriptan benzoate tablets (RITZA, 10 mg) were utilized for the study. All chemicals and reagents used were of analytical grade. Rizatriptan benzoate tablets were purchased from market.

Selection of media:

Main criteria for media selection was solubility and stability, i.e. Rizatriptan benzoate should be soluble as well as stable for sufficient time in selected media. The media used in reported method was distilled water. Dissolution studies require use of 0.1 N HCl. Hence 0.1 N HCl was selected as analytical media for present work.

Preparation of standard stock solution:

Standard solution of drug (100 µg/ml) was prepared by dissolving 10mg Rizatriptan benzoate in 0.1 N HCl, ultrasonicating the solution for 10min.

Preparation of calibration curve:

Aliquots of 0.1 to 1 ml portions of the standard solution were transferred to a series of calibrated 10 ml volumetric flasks, and volume was adjusted with 0.1 N HCl. Solutions were scanned in the range of 200-400 nm against blank 0.1 N HCl. The absorption maxima of solutions was found to be at 225 nm the absorbance of solutions was measured at 225nm against blank (Table 1) and calibration curve was constructed (Figure 2). The optical characteristics were summarized in (Table 2)

Figure no. 1 Chemical structure of Rizatriptan Benzoate

Figure 2: Absorbance spectrum of Rizatriptan Benzoate

Preparation of sample solution:

The proposed method was applied to marketed Rizatriptan benzoate tablet (RITZA). Ten tablets of Rizatriptan benzoate were weighed and powdered in glass mortar. Powder equivalent to 10 mg of the drug was weighed accurately and transferred to 100 ml volumetric flask, dissolved in about 20ml of 0.1 N HCl with frequent shaking and made up the volume to the mark with 0.1 N HCl to obtain the concentration of 100 µg/ml. The solution was filtered through Whatmann filter paper No.41.The filtrate was diluted suitably with 0.1 N HCl to get the concentration of 5µg/ml. The absorbance of sample solution was measured at 225 nm and the amount of Rizatriptan benzoate present in tablet formulation was determined by extrapolating from the calibration curve. The results are shown in (Table 3).

RESULT AND DISCUSSION:

Accuracy (Recovery Test):^{7, 8}

In order to ascertain the suitability and reproducibility of the proposed method, recovery studies were carried out by adding known quantities of standard Rizatriptan benzoate (80,100,120%) to the tablet and the mixtures were analyzed by the proposed method. Three samples were prepared for each recovery level. The percentage recovery of Rizatriptan benzoate was found to be 99.7169 ± 0.7532% (Table 3) indicating that there is no interference by the excipients in the method.

Table 1 - Calibration curve of Rizatriptan Benzoate

Sr. no.	Concentration (µg/ml)	Absorbance	standard deviation
1	0	0	0
2	1	0.2763	± 0.01695
3	2	0.4676	± 0.01364
4	3	0.7083	± 0.00825
5	4	0.9149	± 0.01014
6	5	1.1441	± 0.00699
7	6	1.3439	± 0.00416
8	7	1.5565	± 0.00654
9	8	1.8086	± 0.01402
10	9	2.0412	± 0.00121
11	10	2.2827	± 0.00195

Precision:

Intra-day precision was evaluated by analyzing six test samples of Rizatriptan benzoate. The intermediate precision (inter-day precision) of the method was determined by evaluating the samples of Rizatriptan benzoate on different days and by two different analysts in the same laboratory. The assay and relative standard deviation (RSD) values are 99.668%, 0.8554 and 98.563%, 1.0603 respectively (Table4).

Linearity

Rizatriptan benzoate exhibits its maximum absorption at 225 nm and obeyed Beer's law in the range of 1-10 µg/ml. Linear regression of absorbance Vs concentration yielded equation y = 0.223x + 0.021 with a correlation coefficient of 0.999.

Figure 3: Calibration curve of Rizatriptan Benzoate

Limit of Detection (LOD) and Limit of Quantification (LOQ)

The LOD and LOQ of Rizatriptan benzoate were determined by using standard deviation of the response and slope approach as defined in International Conference on Harmonization (ICH) guidelines.⁷ The LOD and LOQ was found to be 0.31µg/ml and 0.94 µg/ml respectively. The proposed method of determination of Rizatriptan benzoate showed molar absorptivity of 8.9583×10⁴ l/mol.cm and Sandell's sensitivity ⁸ 0.004305 µg/cm² /0.001 absorbance units

Table 2 - Validation parameters

Sr. no.	Parameter	Result
1	Absorption maxima (nm)	225
2	Linearity Range (µg/ml)	1-10
3	Standard Regression Equation	y = 0.223x + 0.021
4	Correlation Coefficient (r² )	r² = 0. 999
5	Molar absorptivity	89583.325
6	A( 1% , 1cm )	233.13
7	Accuracy (% recovery ±SD)	99.7169 ± 0.7532%
8	Precision (%)	99.668, 98.563
9	Specificity	A 5ug/ml solution of drug in 0.1N HCl at UV detection λ of 225 nm shows an absorbance value of 1.1441± 0.00699
10	Sandell’s Sensitivity⁸ (ug/cm²/0.001 absorbance unit)	0.004305
11	LOD (µg/ml)	0.31
12	LOQ (µg/ml)	0.94

Table 3 - Determination of Accuracy by percentage recovery method

Ingredient	Tablet amount (µg/ml)	Level of addition (%)	Amount added (µg/ml)	Amount recovered (µg/ml)	%Recovery	Average % Recovery
Rizatriptan benzoate.	5	80	4	8.8967	98.8531%	99.7169 ± 0.7532%
	5	100	5	10.0236	100.2368%
	5	120	6	11.0067	100.0609%

Table 4 - Determination of Precision

Sample Number	Assay of Rizatriptan benzoate as % of labeled amount
Sample Number	Analyst-I (Intra-day precision)	Analyst-II (Inter-day precision)
1	99.842	97.360
2	98.190	97.124
3	100.323	99.493
4	100.559	99.431
5	99.860	98.732
6	99.239	99.239
Mean	99.668	98.563
Std. deviation	0.8554	1.0603

CONCLUSION:

The developed method was found to be simple, sensitive, accurate, precise, reproducible, and can be used for dissolution studies as well as routine quality control analysis of rizatriptan benzoate in bulk and tablet formulation.

ACKNOWLEDGEMENT:

We thank Padmashree Mrs. Fatma Rafiq Zakaria, Hon’ble Chairman of Maulana Azad Educational Trust, Dr. Rafiq Zakaria Campus, Aurangabad for providing the research facilities. We are also thankful to Cipla Pharmaceuticals. Mumbai (India) for providing us gift sample of Rizatriptan benzoate.

REFERENCES:

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4. Vishwanathan K, Bartlett MG, Stewart JT. Determination of Antimigraine Compounds Rizatriptan, Zolmitriptan, Nortriptan and Sumatriptan in Human Serum by Liquid Chromatography/ Electrospray Tandem Mass Spectrometry. Rapid Commun. Mass Spectrom. 2000; 14(13): 168–172.

5. Altinoz S, Ucar G. Determination of Rizatriptan in its tablet dosage forms by UV spectrophotometric and spectrofluorimetric methods. Analytical Letters.2002; 35: 2471–2485,

6. Validation of analytical Procedure: Methodology (Q2B) Harmonized tripartite Guidelines.1996:1.

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Received on 27.10.2009 Modified on 30.11.2009

Asian J. Research Chem. 3(1): Jan.-Mar. 2010; Page 175-177