UPLC-MS/MS法测定大鼠血浆甘草酸及其代谢物的浓度及药物动力学研究

Determination of glycyrrhizic acid and its metabolites in rat plasma by UPLC-MS/MS and its pharmacokinetics

目的建立测定大鼠血浆中甘草酸及其代谢产物甘草次酸含量的超高效液相色谱-串联四极杆质谱(UPLC-MS/MS)方法,进行大鼠体内药物动力学研究。方法采用ACQUITY UPLC BEH C 18(2.1 mm×150 mm,1.7μm),以5 mmol/L乙酸铵(A)-乙腈(B)为流动相,梯度洗脱,流速0.3 mL/min,电喷雾离子源,负离子模式下,多反应监测(MRM)方式,以格列喹酮为内标;大鼠口服给药,取血后血样经甲醇沉淀蛋白,检测大鼠血药浓度。结果甘草酸与甘草次酸血浆浓度在0.0099~2.8990μg/mL与0.0400~26.8900μg/mL范围内线性关系良好,方法学各项指标均符合要求。药动学参数甘草酸T max为(1.83±0.41)h,C_(max)为(0.42±0.04)μg/mL,AUC_(0-∞)为(2.13±0.43)h·μg^(-1)·mL^(-1),t_(1/2)为(16.74±8.45)h。代谢产物甘草次酸T max为(7.5±1.64)h,C max为(3.58±0.65)μg/mL,AUC 0-∞为(25.24±0.86)h·μg^(-1)·mL^(-1),t_(1/2)为(8.97±3.22)h。结论该方法专属性好,准确可靠,线性范围广,适用于同时检测体内甘草酸与甘草次酸浓度用于药物动力学研究。

Objective To establish an UPLC-MS/MS method for simultaneous determination of glycyrrhizic acid and its metabolite glycyrrhetinic acid in rat plasma,and to determine its pharmacokinetics in rats.Methods ACQUITY UPLC BEH C 18(2.1 mm×150 mm,1.7μm),5 mmol/L ammonium acetate(A)-acetonitrile(B)as mobile phase,gradient elution,flow rate of 0.3 mL/min,electric spray ion source,negative ion mode,multiple reaction monitoring(MRM),and gliquidone as internal standard;After oral administration,the blood samples of rats were precipitated with methanol to detect the blood concentration of rats.Results The linearity of the plasma concentrations of glycyrrhizic acid and glycyrrhetinic acid is good within the range of 0.0099~2.8990μg/mL and 0.0400~26.8900μg/mL,respectively.All methodological indicators were in the line with requirements.The pharmacokinetic parameters of glycyrrhizic acid and glycyrrhetinic acid as following,glycyrrhizic acid T max(1.83±0.41)h,C max(0.42±0.04)μg/mL,AUC_(0-∞)(2.13±0.43)h·μg^(-1)·mL^(-1),t 1/2(16.74±8.45)h,glycyrrhetinic acid T max(7.5±1.64)h,C_(max)(3.58±0.65)μg/mL,AUC 0-∞(25.24±0.86)h·μg^(-1)·mL^(-1),t 1/2(8.97±3.22)h.Conclusion This method is specific,accurate and reliable with wide linear range.It is suitable for simultaneous determination of glycyrrhizic acid and glycyrrhetinic acid in vivo and pharmacokinetic research.