Development and validation of stability indicating method for determination of related substance in ribavirin drug substance

Author(s): Subbarao Eedara, Pradeep B.Kumar, B.M.Rao3,M.B.V.Narayana

A simple and accurate and precise reverse phase liquid chromatographic method was developed for the determination of the Assay and related substance in Ribavirindrug substance. Ribavirin is abroad spectrumAnti-Viral agentshows varying degrees of clinical efficacy in a variety of human viral infections including viral hepatitis, respiratory tract infections and haemorrhagic fevers. The Chromatographic separation between Ribavirin and its related substances was achieved using a YMC- Pack ODS-AQ; 150 x 4.6 mm, 3µm column, buffer contains 1.0g of anhydrous sodium sulfate and 2ml of 5%V/V Orthophosphoric acid and adjusted pH to 2.8with 5%v/ v orthophosphoric acid, mobile phase contains buffer as Mobile phase A and buffer: Acetonitrile (95:05 %v/v) as Mobile phase B using a binary gradient mode with flow rate of the mobile phase kept at 1.0 mL/min. The sample concentration was 0.5 mg/mL. The column temperature was maintained at 25°C and the detection wavelength was 220 nm. The injection volume was 5ìL. The resolution between the critical pair of peaks (Impurity- A&Ribavarin)was found to be greater than 2.0. The limit of detection (LOD) and limit of quantification (LOQ) of Impurity-A is 0.1ng/mL, and analyte were 0.3 ng/mL, for 5ìl injection volume. The test solution and mobile phase was observed to be stable up to 48 h after the preparation. The method was validated and found good results of precision, Selectivity and Solution Stability. The proposed method was found to be suitable and accurate for the quantitative determination of the assay and related substances during release and stability testing of Ribavirin active pharmaceutical ingredient.

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