UPLC-MS/MS测定大鼠血浆中柴胡皂苷B2、B4的含量及药动学研究

Development of UPLC-MS/MS Method for Determination of Saikosaponin B2 and B4 in Rat Plasma and Application in Pharmacokinetics Study

关键词:  柴胡皂苷B2  柴胡皂苷B4  药动学  UPLC-MS/MS

Key words:  saikosaponin B2  saikosaponin B4  pharmacokinetics  UPLC-MS/MS

摘要:

目的 建立UPLC-MS/MS测定大鼠血浆中柴胡皂苷B2、B4，并对柴胡皂苷B2、B4静脉给药后的药动学进行研究。方法 以柴胡皂苷F为内标，采用UPLC BEH C₁₈（2.1 mm×50 mm，1.7 μm）柱，柱温为40℃。流动相为乙腈-0.1%甲酸水溶液，梯度洗脱，流速为0.4 mL·min^-1，洗脱时间为4 min，使用氮气作为去溶剂化气体（800 L·h^-1）和锥形气体（50 L·h^-1）。MRM模式对柴胡皂苷B2 m/z 825.4→617.5、柴胡皂苷B4 m/z 957.6→649.6和柴胡皂苷F（内标）m/z 973.7→119.0进行定量分析。大鼠血浆用乙腈沉淀法去除蛋白。结果 在5~5 000 ng·mL^-1内，大鼠血浆中柴胡皂苷B2、B4线性良好（r>0.995），定量下限为5 ng·mL^-1。柴胡皂苷B2日内精密度RSD<12%，日间精密度RSD<15%。柴胡皂苷B4日内精密度RSD<14%，日间精密度RSD<15%。柴胡皂苷B2、B4的准确度范围为89.3%~110.2%。结论 该分析方法灵敏、快速、选择性良好，并成功应用于柴胡皂苷B2、B4静脉给药后的大鼠药动学研究。

Abstract:

OBJECTIVE To establish a UPLC-MS/MS method for the determination of saikosaponin B2 and B4 in rat plasma. The pharmacokinetics of saikosaponin B2 and B4 after intravenous administration in rats were studied. METHODS Saikosaponin F was used as internal standard, UPLC BEH C₁₈(2.1 mm×50 mm, 1.7μm) column was used and the column temperature was set at 40℃. The mobile phase was consisted of acetonitrile and water (containing 0.1% formic acid), gradient elution with the flow rate of 0.4 mL·min^-1 for 4 min. Nitrogen was used as desolvation gas (800 L·h^-1) and cone gas (50 L·h^-1). MRM was used to quantitatively analyze saikosaponin B2 m/z 825.4→617.5, saikosaponin B4 m/z 957.6→ 649.6 and saikosaponin F (internal standard) m/z 973.7→119.0. Rat plasma samples were treated by acetonitrile precipitation to remove protein. RESULTS In the range of 5-5 000 ng·mL^-1, the concentrations of saikosaponin B2 and B4 in rat plasma showed good linear relation ship(r>0.995), with the lower limit of quantification of 5 ng mL^-1. Saikosaponin B2 had an intra-day RSD<12% and inter-day RSD<15%. Saikosaponin B4 had an intra-day RSD<14%, inter-day RSD<15%. The accuracy of saikosaponin B2 and B4 ranged from 89.3% to 110.2%. CONCLUSION The method is sensitive, rapid and selective, and successfully applied to the pharmacokinetics study of saikosaponin B2 and B4 after intravenous administration.