Zhao Liang, Li Wuhong, Li Yueyue, Xu Hao, Lv Lei, Wang Xinxia, Chai Yifeng, Zhang Guoqing
Department of Pharmacy, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai 200438, P.R. China School of Pharmacy, Second Military Medical University, Shanghai 200433, P.R. China.
School of Pharmacy, Second Military Medical University, Shanghai 200433, P.R. China.
J Chromatogr Sci. 2015 Aug;53(7):1185-92. doi: 10.1093/chromsci/bmu217. Epub 2015 Feb 5.
A liquid chromatography-mass spectrometry method has been developed and validated for rapid simultaneous determination of the oleanolic and ursolic acid contents in rat plasma with betulinic acid as the internal standard (IS). The plasma samples were prepared by a liquid-liquid extraction procedure. Chromatographic separation was performed with a Chromasil-C18 column (250 mm × 4.6 mm, i.d. 5 μm) with methanol-water as mobile phase at 1 mL/min. The detection was accomplished under selected-ion-monitoring mode with a negative electrospray ionization interface. Linear calibration curves were obtained between the range of 0.86-421.2 and 0.94-462.0 ng/mL for oleanolic and ursolic acids, with lower limits of quantification at 0.43 and 0.47 ng/mL, respectively. The extraction recovery exceeded 70% in plasma. The intra- and interday precision values were <9.8% with the accuracy as -7.0 to 9.9% at three different QC levels in both cases. The pharmacokinetic behaviors of oral dosage of QingGanSanJie decoctions were then studied in rats following the developed approach. The t1/2 values of the oleanolic and ursolic acids after oral administration of the monarch medicine extract were significantly different (P < 0.05) from other prescription extracts containing different herb pieces with different compatibilities. Bimodal phenomena appeared in every concentration-time curve for the oleanolic and ursolic acids at 3-8 h after administration. The minister, assistant and guide medicines in the formula could prolong the metabolism of the oleanolic and ursolic acids in vivo, providing an experimental basis for the slow onset and long action of the Traditional Chinese Medicine compound.
已建立并验证了一种液相色谱 - 质谱法，用于以桦木酸为内标（IS）快速同时测定大鼠血浆中齐墩果酸和熊果酸的含量。血浆样品通过液 - 液萃取程序制备。采用Chromasil - C18柱（250 mm×4.6 mm，内径5μm），以甲醇 - 水为流动相，流速为1 mL/min进行色谱分离。采用负电喷雾电离接口，在选择离子监测模式下进行检测。齐墩果酸和熊果酸在0.86 - 421.2和0.94 - 462.0 ng/mL范围内获得线性校准曲线，定量下限分别为0.43和0.47 ng/mL。血浆中的萃取回收率超过70%。在两种情况下，三个不同质量控制水平下的日内和日间精密度值均<9.8%，准确度为 - 7.0至9.9%。然后按照所建立的方法研究了大鼠口服清肝散结汤剂后的药代动力学行为。口服君药提取物后，齐墩果酸和熊果酸的t1/2值与其他含有不同药材配伍的方剂提取物相比有显著差异（P < 0.05）。给药后3 - 8小时，齐墩果酸和熊果酸的每条浓度 - 时间曲线均出现双峰现象。方剂中的臣药、佐药和使药可延长齐墩果酸和熊果酸在体内的代谢，为中药复方起效缓慢和作用持久提供了实验依据。
