改良HPLC法测定大鼠血浆芍药苷浓度

目的:改良高效液相色谱(HPLC)法测定大鼠血浆芍药苷浓度。方法:采用乙酸乙酯提取法处理血浆,选咖啡因为内标物进行测定;色谱柱为Hypersil ODS2RP-C18(150mm×4.6mm,5μm),流动相为乙腈-水(16∶84,V/V),紫外检测波长为231nm,流速为1mL·min-1。结果:采用乙酸乙酯提取比用沉淀法处理的血浆内源性干扰物质少,在HPLC系统条件下,芍药苷和内标咖啡因均不受血浆内源性物质干扰,保留时间分别为6.8min和4.1min左右,定量下限为0.08μg·mL-1,芍药苷检测浓度在0.08～5.12μg·mL-1(r=0.9994)范围内与峰面积积分值呈良好线性关系。分析一个样品只需要7min,平均回收率为69.09%,低、中、高浓度(0.16、0.64、2.56μg·mL-1)的日内精密度和日间精密度分别为12.77%、4.47%、1.10%和8.39%、3.82%、4.27%。结论:本法简便、快速,可用于大鼠血浆芍药苷浓度的分析。

Concentration Determination of Paeoniflorin in Rat Plasma by Modified HPLC

OBJECTIVE： To modify HPLC method for the determination of paeoniflorin concentration in rat plasma. METHODS： Plasma sample was extracted by ethyl acetate using caffeine as internal standard. The chromatographic analysis was carried out using RP-C18 colunm with mobile phase consisted of acetonirile-water （16 ： 84, V/V） at flow rate of 1 mL· min^-1. UV detection wavelength was set at 231 nm. RESULTS： The rat plasma treated by ethyl acetate extraction was cleaner than precipitation method. Paeoniflorin and caffeine kept away from interference of endogenous plasma substance. The retention times of them were about 6.8 min and 4.1 min. The lowest limit of quantitation （LLQ） for paeoniflorin was 0.08 μg·mL^-1. The linear range of paeoniflorin was 0.08-5.12μg·mL^-1（r=0.999 4）. Sample analysis only needed seven minutes. The average recovery was 69.09%. The RSDs of intra-day and inter-day were 12.77% ,4.47%, 1.10% and 8.39% ,3.82% ,4.27% at concentrations of 0.16, 0.64, 2.56 μg·mL^-1. CONCLUSION： The method is simple and rapid for the analysis of paeoniflorin concentration in rat plasma.