DEVELOPMENT OF VALIDATED RP-HPLC METHOD FOR THE ESTIMATION OF WITHAFERIN – A IN WITHANIA SOMNIFERA, Its EXTRACT AND POLYHERBAL FORMULATION

A simple, rapid and specific reversed phase HPLC method has been developed for the estimation ofWithaferin A in Polyherbalformulation containing Withaferin A and its extract. HPLC analysis was performed on C18 column using mixture of Acetonitrile and Methanol containing 0.05 % Triethylamine as isocratic mobile phase at a flow rate of 1.0 ml per minute at detection wavelength of 227 nm. The method was validated for accuracy, precision, linearity, specificity and sensitivity in accordance with International conference on Harmonization guidelines. Validation revealed the method is specific, accurate, precise, reliable and reproducible. Good linear correlation coefficients (r> 0.999) was obtained for calibration plots in the range of 5 – 30 μg/ml. Intraday and Interday RSD of retention times and peak areas were less than 2.0 %. Average Percent Recovery was 97.31 %. The method was successfully used for quantitative analysis of this marker compound in Polyherbal formulation.