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目的:建立HPLC测定I相反应体系中蛇床子素及其代谢物。方法:采用鼠肝微粒体研究蛇床子素在I相反应体系的代谢情况,用高效液相色谱法(C18柱)测定其蛇床子素的含量,紫外检测波长为320nm,以7-羟基香豆素内标,线性梯度洗脱。采用HPLC-MS-MS分析蛇床子素及其代谢物。结果:回归方程为Y=0.0224X+7.0×10-5(r=0.9998),蛇床子素在0.47～60μmol.L-1范围内,线性关系良好;蛇床子素的最低检测量(LLOD)和最低定量限(LLOQ)分别是0.2,0.47μmol.L-1。方法回收率均大于98.70%,日内、日间RSD均小于5%。采用HPLC-MS/MS检测到5个代谢产物,推测可能是去甲基蛇床子素和蛇床子素去氢代谢产物的多个顺反异构体。结论:此法简便准确,可用于I相反应体系中蛇床子素及其代谢物的测定。
OBJECTIVE: To establish a method for the determination of Osthole and its metabolites in phase I reaction system by HPLC. Methods: Rat macrophages were used to study the metabolism of osthole in Phase I reaction system. The content of osthole was determined by high performance liquid chromatography (C18 column). The UV detection wavelength was 320 nm. Internal standard, linear gradient elution. Osthole and its metabolites were analyzed by HPLC-MS-MS. Results: The regression equation was Y = 0.0224X + 7.0 × 10-5 (r = 0.9998), and the osthole was linear in the range of 0.47 ~ 60μmol.L-1. The lowest detectable amount of osthole The lowest limit of quantitation (LLOQ) was 0.2, 0.47μmol.L-1, respectively. The recovery rates of the methods were both higher than 98.70%. The RSDs were less than 5% during the day and the day. Five metabolites were detected by HPLC-MS / MS. It is presumed that they may be cis-trans isomers of demethyl-osthole and osthole dehydrogenative metabolites. Conclusion: This method is simple and accurate and can be used for the determination of Osthole and its metabolites in Phase I reaction system.