Profiling and identification of metabolites of isorhynchophylline in rats by ultra high performance liquid chromatography and linear ion trap Orbitrap mass spectrometry.

Abstract

The searching of potentially bioactive metabolites in the biological body is an interesting and meaningful work for the drug study. However, the structural clarification of possible metabolites is one of the most challenging tasks in drug metabolism studies because of the variety of metabolic reactions and complexity of metabolites in vivo. Here, an ultra high performance liquid chromatography/linear ion trap-Orbitrap mass spectrometry (U-HPLC/LTQ-Orbitrap-MS) with combination of data post-processing techniques, including extracted ion chromatogram (EIC) and multiple mass defect filters (MMDF), was established for profiling and identification of metabolites of isorhynchophylline (IR) in vivo and in vitro, and the possible metabolic pathways were subsequently proposed after the oral dose of 20mg/kg; A total of 47 metabolites of IR were tentatively identified, including 47, 21, 18, and 25 metabolites in rat urine, plasma, liver and rat liver microsomes (RLM) samples, respectively. To our knowledge, most of them were reported for the first time. Seven metabolic pathways, including dehydrogenation, oxidation, hydrolysis, reduction, demethylation, hydroxylation and glucuronide conjugation were involved in the metabolism. Among them, dehydrogenation, hydrolysis, hydroxylation and oxidation were considered as the main metabolic pathway of metabolism according to metabolic profile of in vivo and in vitro. The relative percentage of each metabolite and main metabolite types were also determined to better understand the metabolic behavior of IR in rats. The newly discovered IR metabolites significantly expanded our understanding and were going to be greatly helpful for the further pharmacokinetic study of IR in vivo.