“Cocktail”体外探针法评价补骨脂素对人肝微粒体P450酶亚型活性的影响

Evaluation of the effect of psoralen on human cytochrome P450 subunit activities by Cocktail invitro probe method

关键词:  Cocktail  补骨脂素  人肝微粒体P450酶  探针药物

Key words:  cocktail  psoralen  CYP450  probe drugs

摘要:

[目的]利用Cocktail体外探针法，测定补骨脂有效成分补骨脂素对人肝微粒体多种P450酶亚型活性的影响，为补骨脂临床联合用药安全提供参考。[方法]建立液质联用方法，同时测定人肝微粒体P450酶中各探针代谢产物6α-OH紫杉醇、α-OH咪达唑仑、4-OH甲苯磺丁脲、去甲基右美沙芬、7-OH香豆素、对乙酰氨基酚和6-OH氯唑沙宗的含量。通过Cocktail体外探针实验中补骨脂素对人肝微粒体P450酶各亚型的影响，证明补骨脂对人体肝脏的代谢有影响作用。[结果]所建立的液质联用方法精密可靠，符合生物样品测定要求，可用于人肝微粒体中各P450酶探针的代谢产物的定量分析。与对照组相比，补骨脂素组各底物的转化率均下降，其中香豆素的转化率下降最为显著，补骨脂素对其转化的抑制率为95.84%，而紫杉醇的转化的抑制最不明显，仅为10.39%。补骨脂素对其他几种底物的转化抑制率均在82%~91%之间。[结论]补骨脂素对人肝微粒体的CYP1A2、2A6、2C8、2C9、2D6、2E1和3A4亚型均有抑制，其中对CYP2A6、1A2和2D6 3种亚型的影响较为显著，表明补骨脂素对人肝微粒体P450酶不同亚型影响差异显著，长期给药补骨脂应关注药物联合用药安全。

Abstract:

[Objective] By using "Cocktail" probe method in vitro to determinate the effect of psoralen, which is the effective components of fructus psoraleae, on the activities of human liver microsomal enzymes subtype. It would provide strong evidence for the clinical safety of combination psoralen with other medicine.[Methods] Establish a method for simultaneous determination of seven types of the metabolites of probe drugs by liquid chromatography-mass spectrometry (LC-MS/MS). Throughing the effects of psoralen on CYP450 enzyme isoforms by cocktail probe experiments in vitro to investigate the effect of psoralen on metabolism of human liver.[Results] The established LC-MS/MS method was precise and credible; it was complied with the requirements for the determination of biological samples, and could be used for the quantitative analysis of the metabolites of CYP450 probe in human liver microsomes. Compared with the blank group, the transformation rate of the psoralen group on each CYP450 substrate decreased. The conversion rate of coumarin decreased most significantly and the inhibition rate was 95.84%. However, the decreasion of taxol was not obvious, and the inhibition rate was only 10.39%. The inhibition rate of psoralen on the other substrate was between 82%~91%.[Conclusion] Psoralen had inhibitory effects on subtypes of human liver microsomal CYP1A2, 2A6, 2C8, 2C9, 2D6, 2E1 and 3A4, the influence on CYP2A6, and the inhibition rate of 1A2 and 2D6 were much significantly. These results showed that psoralen had different effects on different subtypes of human liver microsomal enzymes, which indicated us to pay attention to the safety of drug combination when long-term administration of psoralen.