Characterization of Stress Degradation Products of Asenapine by LC-MS/MS and Elucidation of their Degradation Pathway

The investigation aimed to integrate Liquid Chromatography (LC) and Liquid Chromatography-Mass Spectrometry (LC–MS) for precise separation, identification, and characterization of asenapine’s Degradation Products (DPs) without isolating them from reaction mixtures. Asenapine underwent scrutiny under various stress conditions (oxidative, hydrolytic, thermal, and photolytic) following ICH Q1A(R2) guidelines. A total of five distinct degradation products emerged from these stress conditions. The separation of all degradation products was executed utilizing a BDS Y Persil C18 column employing with 0.1 % orthophosphoric acid, acetonitrile, and methanol in the ratio of 50:30:20 (v/v) as mobile phase as an isocratic mode with 231 nm. Elucidation of these degradation products was achieved by scrutinizing their fragmentation pattern and the masses discerned through LC–MS/MS and MSn analysis. These previously unidentified degradation products were ascertained to be 11-chloro-3a,12b-dihydro-1H-dibenzo[2,3:6,7] oxepino[4,5-c]pyrrole (DP 1), 3-(3-chloro-6-methylidenecyclohexa-2,4-dien-1-yl)-4-(cyclohexa-1,4-dien-1-yl)-1- methylpyrrolidine (DP-2), 5-chloro-2-methyl-2,3,3a,12b-tetrahydro-1H-dibenzo[2,3:6,7]oxepino[4,5-c]pyrrole 2-oxide (DP 3), 2-(1-methyl-4-phenylpyrrolidin-3-yl)cyclohexa-2,5-dien-1-ol (DP-4) and 13-(cyclohexa-1,3-dien-1-yl)-1-methyl- 4-(6-methylidenecyclohexa-1,4-dien-1-yl)pyrrolidine (DP-5). Based in findings, it was confirmed that the method was appropriate for resolution, identification and quantification of impurity 1 and 2 of asenapine along with its DPs.