超高效液相色谱-四极杆-飞行时间质谱法鉴定大鼠血浆中淫羊藿素代谢产物的研究

Identification of icaritin metabolites in rat plasma by UPLC-QTOF-MS

目的运用超高效液相色谱-四极杆-飞行时间质谱法(UPLC-QTOF-MS)鉴别淫羊藿素单体大鼠给药后的血浆代谢产物。方法大鼠血浆样品用乙腈沉淀蛋白,采用BEH C_(18)色谱柱(2.1 mm×50 mm,1.7μm),以10 mmol/L乙酸铵-乙腈作为流动相进行梯度洗脱,采用高分辨质谱分别在正、负离子模式进行检测,利用PALLAS软件进行代谢产物预测后,导入Waters UNIFI数据库进行检索鉴定。结果经腹腔注射给药后,大鼠血浆中鉴定出淫羊藿素经羟基化、氢化、环氧化、乙酰化、去甲基化、脱羟基化、磺酰化、葡萄糖醛酸化和糖苷化共24个代谢产物。结论UPLC-QTOF-MS法可快速有效地鉴定出大鼠血浆中淫羊藿素的代谢产物,为淫羊藿素临床前安全性评价提供了支持。

Objective To identify metabolites of icaritin in rat plasma by ultraperformance liquid chromatography quadrupole time of flight mass spectrometry（ UPLCQTOF-MS）. Methods Acetonitrile was employed to precipitate protein in rat plasma.The separation was carried out on a BEH C（18） column（ 2. 1 mm × 50 mm,1. 7 μm） using a gradient mobile phase of acetonitrile,water and ammonium formate. High resolution mass spectrometry was applied for qualitative analysis in positive and negative ion modes,respectively. The metabolites were predicted by the PALLAS software and identified in the UNIFI database. Results Twenty-four metabolites of icaritin in rat plasma which were generated by reactions of hydroxylation, hydrogenation, epoxidation, acetylation,demethylation,dehydroxylation,sulfation conjugation,glucuronidation and glycosylation,were identified. Conclusion A rapid and efficient UPLC-QTOF-MS method is established for the identification of icaritin metabolites in rat plasma,which provided support for the preclinical safety evaluation of icaritin.