Simultaneous Estimation of Cal DL-2-Hydroxy,4-methyl thio-butyrate and Ca-2-oxo3-phenyl Propionate in Alpha-Ketoanalogues Tablets by Validated RP-HPLC Method

Abstract

Introduction: Alpha-Ketoanalogues is a dietary supplement drug containing Calcium-DL- 2-hydroxy-4-(methylthio) butyrate (CHMTB) and Calcium-2-oxo-3-phenyl propionate (COPP) for the management of chronic renal disease.Objectives: In this study, a simple RP-HPLC method has been developed to simultaneously determine Calcium-DL-2-hydroxy-4-(methylthio) butyrate (CHMTB) and Calcium-2-oxo-3-phenyl propionate (COPP) in Alpha-Ketoanalogues Tablets.Materials and Methods: The compounds were separated in a single run using a Waters XBridge 250 mm x 4.6 mm C18 column with 5μm particle size and 130 Å pore size. Elution was carried out via gradient mode with the UV detector wavelength set at 210 nm. The mobile phase selected was a binary mixture of ammonium hydrogen sulphate buffer (pH 7.0) and acetonitrile at a 1.0 mL/min flow rate.Results: The retention time of CHMB and COPP was found to be about 7 min and 20 min, respectively. The chromatographic method shows detector linearity in 50-150% of the operating range with the square of correlation coefficient at 0.9998 and 0.9999 for these two compounds, respectively. The parentage recovery of both CHMB and COPP was within 98-102% range. The method validation was carried out per the USP (chapter 1225) and ICH guidelines (Q2-R2). The developed method was found to be specific, precise, linear, robust, and accurate.Conclusion: This method could be utilized to assay tablet formulation of CHMB and COPP during the in-process and finished product quality control.

Keywords: Alpha-Ketoanalogues; Chronic kidney disease; Chromatography; Method Validation; Simultaneous estimation; Formulation