普伐他汀在大鼠血浆中的固相提取和反相高效液相色谱法测定

Determination of Pravastatin in Rat Plasma by Solid Phase Extraction and RP - HPLC

目的:建立反相高效液相色谱法测定普伐他汀在大鼠血浆中的含量。方法:空白血浆加普伐他汀和内标,用Bond E-lut C18小柱进行固相提取,然后用色谱方法分析。采用Puroshper RP-C18反相色谱柱（5μm,250mm×4mm）,Lichrospher 100RP-18预柱（5μm,4mm×4mm）,流动相为0.035mmol·L-1磷酸二氢钠缓冲液（用磷酸调pH3.0）-乙腈（13:7）,流速1mL·min-1,检测波长239nm,柱温40℃,进样量100μL。结果:普伐他汀在血中的线性范围为10～400ng·mL-1,r=0.9997;最低检测限用标准液试验达2ng·mL（S/N=3）,用加标样血浆试验,方法定量限为10ng·mL-1血,RSD<20％;不同浓度固相提取回收率平均为71.48％;日内差平均为13％,日间差平均为19％。用上述色谱方法测定了普伐他汀在大鼠血浆中（Lewis大鼠,灌胃20mg·kg-1）的含量,并绘制药时曲线。结论:方法实用有效,可用于药代动力学研究。

Objective: To set up an assay method by solid phase extraction and RP - HPLC for the determination of pravastatin in rat plasma. Method: Pravastatin in an aliquot of 1 mL plasma, spiked with internal standard, was extracted by solid - phase extraction with Bond Elut C18 column ( Varian) , and determined by HPLC. The chromatography employed a Puroshper RP - C18 reverse - phase column (5μm, 250 mm × 4 mm) , a Lichrospher 100 RP - 18 pre -column (5μm, 4 mm × 4mm ) as the analytical column and the mixture of phosphate buffer (0.035 mmol·L-1 Na2HP04, pH adjusted to 3.0 with phosphoric acid) - acetonitrile (13:7) as the mobile phase with the flow rate of 1 mL·min-1. Pravastatin was detected with a UV detector at 239 nm. The column temperature was set at 40℃ . The sampling amount was 100μL. Results: The linearity range was 10-400 ng·mL-1 ( r = 0.999 7) with the limits of detection and quantification being 2 ng ?mL-1 ( signal - to - noise ratio of 3 ) and 10 ng ?mL-1 (RSD < 20 % ) , respec-tively. The average extraction recovery of pravastatin at different concentrations was 71.48% . The average intra - day and inter- day precision was 13% and 19% respectively. The method was applied to determine pravastatin in rat plasma and a time - plasma concentration curve was presented. Conclusion: The method is practical and can be used for the pharmacokinetic study of pravastatin.