Simultaneous determination of elbasvir and grazoprevir in fixed-dose combination and mass spectral characterization of each degradation product by UHPLC-ESI-QTOF-MS/MS.

Zepatier® (Elbasvir and Grazoprevir) is a novel two-drug, fixed-dose combination product containing elbasvir and grazoprevir used for the treatment of chronic hepatitis C virus infection. Various forced degradation studies of the two drugs had been conducted in order to identify significant degradation products and establish the degradation pathway induced by thermal, photolytic, acid/base hydrolytic and/or oxidative stress conditions. A reversed phase C18 UHPLC-PDA method has been developed for the analysis of the stressed samples. Seven significant degradation products of elbasvir and five significant degradation products of grazoprevir were found and investigated further by high resolution ESI-QTOF-MS with high accurate mass measurement (-1.96 to 1.36 ppm). The chemical structures of each degradation product were proposed based on their relative MS/MS fragmentation spectra in comparison with the corresponding parent drugs and the chemical synthetic knowledge of process chemists. The validated stability-indicating UHPLC method can be used in routine analysis for the simultaneous determination of elbasvir and grazoprevir in pharmaceutical formulations. As more and more combination drugs will enter into the market, this study can also shed light on stability indicating method development for combined drugs at early development stage.