HPLC测定金丝桃苷的血药浓度及其在药动学研究中的应用

HPLC for Determination of Hyperoside in Rats Plasma and Its Application in Pharmacokinetics Study

关键词:  金丝桃苷  动力学  高效液相色谱法

Key words:  hyperoside  pharmacokinetics  HPLC

摘要:

目的建立HPLC测定大鼠血浆中金丝桃苷(槲皮素-3-O-半乳糖苷)浓度的方法,并用此方法对金丝桃苷在大鼠体内的药动学进行研究。方法金丝桃苷以12mg·kg-1的剂量经大鼠尾静脉注射,于不同时间点眼球静脉丛取血,甲醇直接沉淀蛋白,HPLC测定金丝桃苷的血药浓度。采用Shimadzu ODS(4.6mm×150mm,5μm)色谱柱,以甲醇∶水(40∶60)为流动相,检测波长360nm,流速为1.0mL·min-1,柱温25℃。乙酰苯胺为内标测定金丝桃苷的含量。结果金丝桃苷和乙酰苯胺能完全分离。线性范围为20～4888ng·mL-1,回归方程为Y=132.539X-8.736,r=0.9996。日内精密度RSD为0.1%～2.9%,日间精密度RSD为1.5%～7.3%;平均回收率为90.1%,RSD为0.9%。其体内过程符合三室模型,t1/2(p)为(1.62±0.54)min,t1/2(a)为(18.67±2.48)min,t1/2(b)为(660.15±155.56)min,AUC为(200229±4397.63)ng·min·mL-1。结论分析方法简便快速,专属好,精密度和回收率高,可满足动力学研究的一般要求。

Abstract:

OBJECTIVE To establish a HPLC for the determination of hyperoside (3-0-galactose-quercetin) in rats plasma and to study the pharmacokinetics of hyperoside in rats. METHODSBlood samples were collected via eyeball veniplex at different time intervals after injection of hyperoside through tail vein to rats at the doses of 12 mg·kg-1. The plasma protein was first precipitated directly with methanol and then the plasma concentrations of the hyperoside were analyzed by HPLC.A Shimadzu ODS column(4.6 mm×150 mm,5 μm)was used with a mobile phase consisting of methanol-water (40∶60).The detection wavelength was set at 360 nm. The mobile phase was run at a flow rate of 1.0 mL·min-1. The temperature of column was 25 ℃. Acetanilide was elected as the internal standard. RESULTSHyperoside and acetanilide were successfully separated. The linear range was 20-4 888 ng·mL-1, the regression equation of hyperoside was Y=132.539X-8.736,r=0.999 6. The precision of intra-assay and inter-assay was 0.1%-2.9% and 1.5%-7.3%. The average extraction recovery was 90.1% with RSD 0.9%. The results showed that the concentration-time curve of Hyperoside in rat plasma could be fitted to three-compartment open model and its pharmacokinetic parameters were as follows: t1/2(p)was (1.62±0.54)min,t1/2(a) was (18.67±2.48)min, t1/2(b) was (660.15±155.56)min, AUC was (200 229±4 397.63)ng·min·mL-1. CONCLUSIONThe HPLC method is simple and efficient with excellent specificity,sensitivity,precision,recovery and can be applied to pharmacokinetic investigation.