LC, LC-MS/TOF and MSn studies for the separation, identification and characterization of degradation products of ritonavir

The objective of the present study was to explore the decomposition behaviour of ritonavir under hydrolytic (acid, alkali and neutral), oxidative, photolytic and thermal stress conditions as advised in the International Conference on Harmonization (ICH) Q1A(R2) guideline. The drug was found to degrade in acidic, alkaline and neutral conditions, while it was stable under oxidative, thermal and photolytic stress conditions. A total of ten degradation products (DPs) were formed, which were separated on a C-18 column employing a gradient high-performance liquid chromatography (HPLC) method. To characterize the DPs, first a complete fragmentation pathway of the drug was established by carrying out mass spectrometry/time of flight (MS/TOF) and multistage-mass spectrometry (MSn) accurate mass studies. Then the stressed samples were subjected to liquid chromatography-mass spectrometry/time of flight (LC-MS/TOF) studies, which provided their fragmentation pattern and accurate masses. The elucidation of the structures of all the degradation products was achieved by comparison of MS/TOF data of the drug with those of the DPs. Finally the degradation pathway of the drug was established along with mechanisms of formation of the degradation products.