Comprehensive characterization of metabolites and metabolic profiles in plasma has considerable sig-nificance in determining the efficacy and safety of traditional Chinese medicine(TCM)in vivo.However,this process is usually hindered by the insufficient characteristic fragments of metabolites,ubiquitous matrix interference,and complicated screening and identification procedures for metabolites.In this study,an effective strategy was established to systematically characterize the metabolites,deduce the metabolic pathways,and describe the metabolic profiles of bufadienolides isolated from Venenum Bufonis in vivo.The strategy was divided into five steps.First,the blank and test plasma samples were injected into an ultra-high performance liquid chromatography/linear trap quadrupole-orbitrap-mass spectrometry(MS)system in the full scan mode continuously five times to screen for valid matrix compounds and metabolites.Second,an extension-mass defect filter model was established to obtain the targeted precursor ions of the list of bufadienolide metabolites,which reduced approximately 39％of the interfering ions.Third,an acquisition model was developed and used to trigger more tandem MS(MS/MS)fragments of precursor ions based on the targeted ion list.The acquisition mode enhanced the acquisition capability by approximately four times than that of the regular data-dependent acquisition mode.Fourth,the acquired data were imported into Compound Discoverer software for identification of metabolites with metabolic network prediction.The main in vivo metabolic pathways of bufadienolides were elucidated.A total of 147 metabolites were characterized,and the main biotransformation reactions of bufadienolides were hydroxylation,dihydroxylation,and isomerization.Finally,the main prototype bufadienolides in plasma at different time points were determined using LC-MS/MS,and the metabolic profiles were clearly identified.This strategy could be widely used to elucidate the metabolic profiles of TCM preparations or Chinese patent medicines in vivo and provide critical data for rational drug use.

Wen-Long Wei;Hao-Jv Li;Wen-Zhi Yang;Hua Qu;Zhen-Wei Li;Chang-Liang Yao;Jin-Jun Hou;Wan-Ying Wu;De-An Guo

Shanghai Research Center for Modernization of Traditional Chinese Medicine,National Engineering Laboratory for TCM Standardization Technology,Shanghai Institute of Materia Medica,Chinese Academy of Sciences,Shanghai,201203,China;University of Chinese Academy of Sciences,Beijing,100049,China

药物分析学报（英文）

[1]Wen-Long Wei;Hao-Jv Li;Wen-Zhi Yang;Hua Qu;Zhen-Wei Li;Chang-Liang Yao;Jin-Jun Hou;Wan-Ying Wu;De-An Guo-.An integrated strategy for comprehensive characterization of metabolites and metabolic profiles of bufadienolides from Venenum Bufonis in rats)[J].药物分析学报（英文）,2022(01):136-144

bufadienolides,Venenum,Bufonis,bufadienolide,dihydroxylation

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