Development and Validation for Reverse Phase High Performance Liquid Chromatographic Method for Simultaneously Determination for Diphenoxylic Acid, Nitrileamide and Bromdiphenyl Butoro Nitrile in Diphenoxylate Hydrochloride Bulk Drug

Abstract

Related substances by HPLC method was developed and validated for Diphenoxylate hydrochloride bulk drug and its three potential impurities i.e. Diphenoxylic acid (degradation product), Nitrile amide (precursor) and Bromo Diphenyl butro nitrile (precursor). Analysis was performed on AGILENT 1100 SERIES HPLC system with auto injector and binary gradient high pressure mixing pump on Inersil ODS-3 (RP C 18) column. Mobile phase of Acetonitrile and HPLC water pH adjusted to 2.3 with orthophosphoric acid in the ratio of 25:75 (v/v) with change in the ratio of mobile phase by linear gradient to 85: 25 (v/v) in 45 minute was pumped at 2mL/min and the detection was done at 210 nm. The parameter for which the method was validated included specificity, limit of detection and quantitation, linearity, precision, accuracy and robustness. The method was successfully used to quantitate the levels known impurities {i.e. Diphenoxylic acid (degradation product), Nitrileamide (precursor) and Bromo Diphenyl butro nitrile (precursor)} and unkown impurities in Diphenoxylate hydrochloride bulk drug.