Metabolites and metabolic pathways of mesaconitine in rat liver microsomal investigated by using UPLC-MS/MS method in vitro.
- Author:
Yun-Feng BI
;
Shu LIU
;
Rui-Xing ZHANG
;
Feng-Rui SONG
;
Zhi-Qiang LIU
- Publication Type:Journal Article
- MeSH:
Aconitine;
analogs & derivatives;
metabolism;
Animals;
Chromatography, High Pressure Liquid;
Cytochrome P-450 CYP3A;
metabolism;
Cytochrome P-450 CYP3A Inhibitors;
Cytochrome P-450 Enzyme Inhibitors;
Cytochrome P-450 Enzyme System;
metabolism;
Enzyme Inhibitors;
pharmacology;
Ketoconazole;
pharmacology;
Male;
Metabolic Networks and Pathways;
Microsomes, Liver;
enzymology;
metabolism;
Quinine;
pharmacology;
Rats;
Rats, Sprague-Dawley;
Sulfaphenazole;
pharmacology;
Tandem Mass Spectrometry
- From:
Acta Pharmaceutica Sinica
2013;48(12):1823-1828
- CountryChina
- Language:Chinese
-
Abstract:
Mesaconitine was incubated with rat liver microsomes in vitro. The metabolites of mesaconitine in rat liver microsomes were identified by ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method with high resolution power. A typical reaction mixture of 100 mol L-1 Tris-HCI buffer (pH 7.4) containing 0.5 gL-1 microsomal protein and 50 micro molL-1 mesaconitine was prepared. The above reaction mixture was divided into six groups, and the volume of each group was 200 micro L. The incubation mixture was pre-incubated at 37 degrees C for 2 min and the reactions were initiated by adding NADPH generating system. After 90 min incubation at 37 degrees C, 200 micro L of acetonitrile was added to each group to stop the reaction. The metabolites of mesaconitine were investigated by UPLC-MS/MS method. Mesaconitine and 6 metabolites M1-M6 were found in the incubation system. The structures were characterized according to the data from MS/MS spectra and literatures. The metabolic reactions of mesaconitine in rat liver microsomes included the demethylation, deacetylation, dehydrogenation and hydroxylation. The major metabolic pathways of mesaconitine in rat liver microsomes were determined by UPLC-MS/MS on multiple reaction monitoring (MRM) mode combined with specific inhibitors of cytochrome P450 (CYP) isoforms, including alpha-naphthoflavone (CYP1A2), quinine (CYP2D), diethyldithiocarbamate (CYP2E1), ketoconazole (CYP3A) and sulfaphenazole (CYP2C), separately. Mesaconitine was mainly metabolized by CYP3A. CYP2C and CYP2D were also more important CYP isoforms for the metabolism reactions of mesaconitine, but CYP1A2 and CYP2E1 haven't any contribution to MA metabolism in rat liver microsomes.
