高效液相色谱-荧光光谱法测定大鼠血浆中葛根素及其药动学研究

Determination of Puerarin in rat plasma by high performance liquid chromatography with fluorescence detection and its application to pharmacokinetic studies

目的建立高效液相色谱-荧光光谱法测定大鼠血浆中葛根素含量。方法色谱柱选用Hypersil ODS柱,流动相为甲醇∶水（含50 mmol/L醋酸铵）=23∶77（V/V）,流速为1 mL/min,荧光检测器参数：激发波长250 nm,发射波长480 nm,增益为11,柱温30℃。结果该方法专属性强,在0.16~120 mg/L浓度范围内线性关系良好,最低检测限为10μg/L,准确度介于99%~104%,日间、日内精密度均小于10%。该方法成功应用于大鼠静脉注射葛根素的药动学研究中,葛根素在大鼠体内符合开放二房室模型,主要的药动学参数如下：AUC0→t（41.94±12.90）mg/（L.h）,AUC0→∞（44.37±28.90）mg/（L.h）,MRT（0.97±0.37）h,T1/2（1.06±0.39）h,Vss（0.09±0.02）L,Vz（0.14±0.03）L,Cl（0.10±0.05）L/h。结论本方法无需柱后修饰,简单、可靠,适于大鼠血浆中葛根素的检测。

Objective It is to develop a method of high performance liquid chromatography （HPLC） with fluorescence detection to detect the level of puerarin in rat plasma. Methods Chromatography was performed on a Hypersil ODS column using a mobile phase consisting of methanol - 50 mmol/L ammonium acetate in water （23：77, v/v） with a flow rate of 1 mL/ rain. The fluorescence detector was set at excitation wavelength of 250 nm, emission wavelength of 480 nm and PMT- gain of 11. The column temperature was set at 30 ℃. Results The method was selective and sensitive with a detection limit of 10 μg/L, A good linear response was observed over a wide range （0.16-120.00 mg/L） in rat plasma. This validated method was applied successfully to a pharmacokinetic study in rat plasma after intravenous administration of puerarin. The main pharmacokinetic parameters were as follows： AUC0→t（41.94 ± 12.90） mg/（L/h）, AUC0→∞（44.37 ± 28.90）mg/（LAn）, MRT （0.97±0.37） h, T1/2α（0.13±0.02） h, T1/2β（1. 06 ± 0. 39） h, Vss （0.09±0.02） L, Vz （0.14±0.03） L, CI （0.10± 0.05 ） L/h. Conclusion A simple, rapid, sensitive and selective method of HPLC with fluorescence detection has been developed and validated for quantitative determination of puerarin in rat plasma.