In vitro metabolism of alisol A and its metabolites’ identification using high-performance liquid chromatography–mass spectrometry

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2013-12, Vol.941, p.31-37
• Main Authors: Yu, Yue, Liu, Zhenzhen, Ju, Ping, Zhang, Yuanyuan, Zhang, Lunhui, Bi, Kaishun, Chen, Xiaohui
• Format: Article
• Language: English
• Subjects: Alisol A; Animals; Biotransformation; chemical inhibitors; Cholestenones - metabolism; Chromatography, High Pressure Liquid - methods; cytochrome P-450; Cytochrome P-450 Enzyme System - metabolism; enzyme kinetics; HPLC–QTOF MS; Human liver microsomes; humans; hydroxylation; In Vitro Techniques; isozymes; Kinetics; liquid chromatography; liver microsomes; mass spectrometry; Mass Spectrometry - methods; Metabolism; metabolites; Microsomes, Liver - enzymology; Microsomes, Liver - metabolism; Oxidation-Reduction; Rat liver microsomes; Rats
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2013.09.029

•The metabolites of alisol A were identified by HPLC–QTOF MS.•A total of 3 oxidative metabolites were found and identified in RLM incubation systems.•A total of 6 oxidative metabolites were found and 3 of which were identified in HLM incubation systems.•The formation of each metabolite of alisol A was mainly catalyzed by CYP3A4 enzyme. A liquid chromatography–mass spectrometry (LC–MS) method was developed and successfully applied to the study on the enzyme kinetics of alisol A in rat liver microsomes (RLM) and human liver microsomes (HLM) incubation systems, and employed for semi-quantitative determination of each metabolite of alisol A. The metabolites of alisol A in RLM, HLM and human recombinant CYP3A4 enzyme incubation systems were identified by high-performance liquid chromatography–quadrupole time-of-flight mass spectrometry (HPLC–QTOF MS). A total of 3 and 6 oxidative metabolites were found in RLM and HLM incubation systems, respectively. 3 metabolites found in both incubation systems were identified. The exact position of hydroxylation for the metabolites M1 and M2 could not be determined. Chemical inhibitors of cytochrome P450 (CYP450) and individual human recombinant CYP450 enzyme were used to identify the CYP450 isozymes involved in the formation of each metabolite of alisol A. The result indicated that the formation of each metabolite of alisol A was mainly catalyzed by CYP3A4 enzyme.