高效液相色谱荧光法测定人血浆中普卢利沙星活性代谢产物NM394的浓度

HPLC determination of active metabolite of prulifloxacin:NM394 in human plasma with fluorescence detection

目的:建立普卢利沙星活性代谢产物 NM394血药浓度的高效液相色谱分析方法。方法:以妥舒沙星为内标,血浆样品用固相萃取,色谱柱:Waters Symmertry C_(18)色谱柱(4.6mm×250mm,5μm),在线过滤器;以0.1%三乙胺水溶液(用10%磷酸调 pH 为2.8)-乙腈(80:20)为流动相;激发波长为280nm,发射波长为425nm;流速1.0mL·min^(-1),柱温为25℃。结果:NM394在0.025～4.800μg·mL^(-1)的范围内有良好的线性关系(r=0.9999)最低检测浓度为0.025μg·mL^(-1),该方法的相对回收率为96.7%～104.7%(n=5);绝对回收率为76.1%～96.5%(n=5);日内 RSD 为0.8%～2.6%,日间 RSD 为1.4%～3.3%。结论:本方法简便、准确、灵敏,特异性强,重复性好,可用于血浆中普卢利沙星活性代谢产物 NM394的测定及人体内药代动力学研究。

Objective：To establish HPLC method for determination of NM394, active metabolite of prulifloxacin in human plasma. Methods： The method involves adopting of solid - phase extraction with by HPLC, using tosufloxacin as the internal standard. The Symmertry C18 （4. 6 mm × 250 mm,5 μm）was used as analytical column with a mobile phase consisted of 0. 1% triethylamine （adjusted to pH 2. 8 with phosphoric acid） -acetonitrile （80： 20）. The flow rate was of 1.0 mL · min^ - 1 with fluorescence detection set at λex 280 nm and λem 425 nm. Results ：The linear range of calibration curve was within drug plasma concentrations of 0. 025 ～ 4. 800 μg·mL^-1 （ r = 0. 9999 ）, The detection limit was 0. 025 μg·mL^-1, the absolute recovery was 96. 7% ～ 104. 7% （ n = 5 ） , the method recovery was 76. 1% ～96. 5% （n =5 ）,the within- day RSD and between- day RSD were 0. 8% ～2. 6% and 1.4% ～3.3% （ n = 5 ）, respectively. Conclusions： The method is shown to be sensitive, accurate, simple and reproducible for the determination of NM394 levels in human plasma. It is suitable for phannacokinetics study of NM394, active metabolite of prulifloxacin.