Kinetics and mechanism of acidic hydrolysis of nordazepam studied by high-performance liquid chromatography and fourth-order derivative ultraviolet spectrophotometry

Abstract

A reversed-phase HPLC method was developed to study the acid-catalysed hydrolysis of nordazepam in hydrochloric acid solutions of 0.01, 0.1 and 1.0 M. One intermediate was observed, which was isolated and identified. The mechanism of hydrolysis appeared to be biphasic, showing a consecutive reaction with a reversible first step. Initial breakage of the azomethine bond, followed by a slow hydrolysis of the amide bond resulted to creation of the benzophenone product in strongly acidic solutions. A fourth-order derivative method for monitoring the parent compound itself was also proposed and evaluated, as well. Relative standard deviation was less than 2% for the HPLC and less than 5% for the derivative method. Detection limits for nordazepam, intermediate and final degradation product were 1.8×10 −9 M, 2.1×10 −9 M and 2.0×10 −9 M, respectively, in the former method and 7.0×10 −7 M for nordazepam in the latter. Estimation of k 1, k −1 and k 2 values was tried and results of HPLC and fourth-order derivative methods were compared.