丹参酮Ⅱ_A在大鼠肝微粒体酶中的代谢动力学

Metabolic kinetics of tanshinone Ⅱ_A in rat liver microsomal enzyme

目的研究丹参酮ⅡA在大鼠肝微粒体酶中的代谢,以及选择性细胞色素P450(CYP)酶抑制剂对其代谢的影响。方法超速离心法制备大鼠肝微粒体,采用高效液相-质谱联用方法测定孵育液中丹参酮ⅡA原形药物的浓度。研究丹参酮ⅡA的酶促动力学,推导出药物米氏常数(Km)和最大反应速度(Vmax);并计算体外酶对药物的清除率(CLint)。同时观察不同浓度和不同种类的CYP酶特异性抑制剂对丹参酮ⅡA代谢的影响。结果丹参酮ⅡA在大鼠肝微粒体酶中的Vmax为(1.20±0.18)nmol/(min·mg);Km为(4.35±0.67)nmol/mL;CLint为(0.28±0.06)mL/(min·mg)。噻氯匹啶和酮康唑能够显著抑制丹参酮ⅡA的代谢,奎尼丁对丹参酮ⅡA的代谢也有一定的抑制作用,而其他CYP酶抑制剂对丹参酮ⅡA的代谢无明显影响。结论CYP2C19和CYP3A1主要参与了丹参酮ⅡA的代谢,CYP2D6也起到了部分代谢作用;这些CYP酶的抑制剂可能使丹参酮ⅡA的代谢受到抑制,造成药物药效或毒性的增加。

Objective To study the metabolic kinetics of tanshinone ⅡA and the effects of selective CYP450 inhibitors on the metabolism of tanshinone Ⅱ A in rat liver microsomal enzyme. Methods Rat liver microsomes were prepared by using uhracentrifugation method. Tanshinone ,Ⅱ A concentration in the incubation pool was determined by LC/LC/MS method. The Michaelis-Menten parameters Km and Vmax in rat liver microsomes were initially estimated by analyzing Lineweave-Brurk plot. The clearance （CLint） was calculated for in vitro enzyme to tanshinone Ⅱ A. Various selective CYP inhibitors were used to investigate their inhibitory effects on the metabolism of tanshinone Ⅱ A and the principal CYP isoforms involved in tanshinone ⅡA metabolic ring. Results The Vmax, Km, and Cint, of tanshinone Ⅱ A were （1. 20±0. 18） nmol/（min · mg）, （4. 35±0. 67） nmol/mL, and （0. 28±0. 06） mL/（min · mg）, respectively. The metabolism of tanshinone Ⅱ A was significantly inhibited by ticlopidine and ketoconazole. Quinidine could inhibit the metabolism of tanshinone Ⅱ A, while the other inhibitors showed little inhibitory effect on the metabolism of tanshinone ⅡA. Conclusion It is shown that CYP2C19 and CYP3A1 are involved in the metabolism of tanshinone Ⅱ A and CYP2D6 contribute to the metabolism to some extent. Their selective CYP inhibitors may have potential inhibition on metabolism of tanshinone Ⅱ A and make the efficacy or toxicity increased.