LC-MS法测定大鼠血浆中甘草次酸及其药动学研究

Method of LC-MS for the determination of glycyrrhetinic acid in rat plasma and pharmacokinetic study of glycy rrhetinic acid

【目的】建立液相色谱-质谱联用（LC—MS）的方法测定大鼠口服三拗汤后甘草次酸的药代动力学。【方法】采用Agilent ZorbaxSB-C18（4．6mm×lS0mm，5μm）色谱柱，流动相为乙腈-水（含10mmol／L甲酸铵），90％乙腈等度洗脱，流速0．3mL／min，柱温25℃，进样量20μL。采用电喷雾离子源，SIM负离子检测模式，甘草次酸和熊果酸（内标）定量离子分别为m／z469．4和m／z455．4。大鼠单剂量灌胃给予三拗汤浓缩液后，经液一液萃取法对大鼠血浆样品进行预处理，利用LC—MS法测定大鼠体内甘草次酸的血药浓度，并采用DAS（ver．1.0）软件计算药代动力学参数。【结果】甘草次酸的线性范围为5—10000μg，L，最低定量限为5μg几，内源性物质不干扰甘草次酸及内标的测定，方法精密度、准确度、回收率和稳定性均符合生物样品的测定要求。【结论】本方法具有专属性强、准确度高、灵敏度好的特点，可用于甘草次酸的药代动力学研究。

[Objective] To develop a LC-MS method for the pharmacokinetic study of glycyrrhetinic acid after administration of Sanao decoction. [Methods] The method of determination was performed on Agilent Zorbax SB- C18 （4.6 mm×l50 mm, 5μm） column using acetonitrile and water containing of 10 mM Ammonium formate as mobile phase in isocratic elution with 90：10. The flow rate was 0.3 mL/min and the temperature was 25℃. Electrospray ionization（ESI） source was applied and operated in the negative ion mode with SIM scanning mode. The quantitative ions of glycyrrhetinic acid and ursolic acid were m/z 469.4 and rn/z 455.4. Rats were oral administration with single dose Sanao decoction concentrated solution. The plasma samples were through liquid-liquid extraction pretreatment. Using the method of LC-MS for the determination of plasma concentration of glycyrrhetinic acid and calculate the pharmacokinetic parameters by DAS（ver. 1.0）.[Results] The linear rang of glycyrrhetinic acid was 5-10 000 ng/mL and it＇s limit of quantity was 5 ng/mL. Endogenous substances did not interfere with the determination of glycyrrhetinic acid and internal standard. The methods of precision, accuracy, recovery and stability are in line with the requirements of determination of biological samples and may be applied for glycyrrhetinic acid pharmacokinetic study. [Conclusion] The method has the characteristics of high specificity, high accuracy, good sensitivity, and can be used for glycyrrhetinic acid pharmacokinetic study.