摘要
目的研究呋喃香豆素活性成分欧前胡素在人和大鼠肝微粒体的代谢稳定性和酶促动力学,并分析细胞色素P450(CYP)酶的代谢表型。方法将欧前胡素分别与人和大鼠肝微粒体在37℃与不同辅酶因子孵育,应用液相色谱-串联质谱(LC-MS/MS)法测定孵育液中剩余的欧前胡素含量,分析其代谢稳定性及代谢消除反应类型,并计算酶促动力学参数——米氏常数(K_m)和最大反应速率(V_(max))。应用重组人源CYP同工酶(CYP1A2,CYP2B6,CYP2C8,CYP2C9,CYP2C19,CYP2D6和CYP3A4)及其特异性抑制剂,确定欧前胡素的CYP酶代谢表型。结果在人和大鼠肝微粒体中,欧前胡素主要依赖于还原性辅酶Ⅱ(NADPH)的Ⅰ相代谢消除,30 min代谢转化率分别为69.7%和94.5%,代谢消除半衰期t1/2分别为18.9±0.6和(2.8±0.4)min,经外推得到的肝清除率(Cl_h)分别是16.9±0.1和(51.9±0.4)m L·min^(-1)·kg^(-1),K_m分别为13.60±0.16和(14.00±0.24)μmol·L^(-1),V_(max)分别为2928±96和(8434±27)nmol·min^(-1)·g^(-1)。欧前胡素在大鼠肝微粒体的消除显著快于人肝微粒体(P<0.01)。欧前胡素在肝微粒体的Ⅰ相代谢是由多个CYP同工酶介导的,经整体归一化法评价得到CYP1A2,CYP2B6,CYP2C19和CYP3A4的代谢贡献率分别为20.4%,7.3%,10.5%和61.8%。结论欧前胡素在肝微粒体中主要发生多个CYP酶介导的Ⅰ相代谢,其中CYP3A4和CYP1A2的贡献率>20%。
OBJECTIVE To investigate enzyme kinetic characteristics of imperatorin in rat liver microsomes （RLM） or human liver microsomes （HLM）, and to identify the reaction phenotyping of human recombinant cytochrome P450 enzyme （CYP） mediated phase I metabolism. METHODS Imperatorin was incubated at 37℃ with HLM or RLM in the presence or absence of nicotinamide ade- nine dinucleotide phosphate （NADPH） or uridine 5＇-diphosphoglucuronic acid （UDGPA）. The con- centrations of imperatorin in the incubation systems were determined with LC-MS/MS to evaluate its met- abolic stability and enzymatic kinetics. The CYP phenotyping of imperatorin was identified using a panel of human recombinant CYP isoforms （CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP3A4） and also using a group of specific inhibitors in HLM. RESULTS Imperatorin was meta- bolically eliminated in the presence of NADPH in HLM or RLM. The elimination rates for HLM and RLM in 30 rain were 69.7% and 94.5%, respectively, and elimination half-life （t1/2）values were 18.9±0.6 and （2.8±0.4）min, respectively. The extrapolated hepatic clearance parameters （C/h） were 16.9--0.1 and （51.9±0,4）mL. min-1 · kg-1. The Michaelism-Menten parameters （Kr,） were 13.60±0.16 and （14.00±0.24）pmol·L-1, and maximum velocity （Vax） were （2928±96） and （8434±27）nmol·min-1·g-1 respec- tively. The metabolic elimination of imperatorin in RLM was quicker than in HLM. The results of CYP phenotyping indicated that CYP1A2, CYP2B6, CYP2C19 and CYP3A4 were the major CYP isoforms involved in the imperatorin metabolism. Their individual contributions assessed using the method of to- tal normalized rate were 20.4%, 7.3%, 10.5% and 61.8%, respectively. CONCLUSION Imperatorin is mainly eliminated by CYP mediated metabolism in HLM and RLM. CYP1A2 and CYP3A4 are the major responsible enzymes with a contribution rate above 20%.
出处
《中国药理学与毒理学杂志》
CAS
CSCD
北大核心
2016年第8期848-854,共7页
Chinese Journal of Pharmacology and Toxicology
基金
国家科技重大专项(2012ZX09301003-001)
国家科技重大专项(2015ZX09J15104)~~
