HPLC测定唾液、龈沟液和血清中甲磺酸帕珠沙星药物浓度

Determination of pazufloxacin concentration in saliva,gingival crevicular fluid and serum of healthy adults using high-performance liquid chromatography

目的建立一种快速灵敏检测甲磺酸帕珠沙星在血清、龈沟液和唾液中药物浓度的HPLC方法。方法色谱柱为AgilentZorbax SB-C18(5μm,250 mm×4.6 mm),流动相为乙腈-磷酸三乙胺溶液(0.5%磷酸,1%三乙胺)=(155∶850),紫外检测波长为245 nm,流速为1.0 ml/min。样本处理采用甲醇沉淀蛋白,上清吹干,流动相复溶残渣,离心后上清20μl进样检测。结果样品中帕珠沙星色谱分离完全,无内源性物质干扰,在25-10 000 ng/ml内呈现良好的线性回归,线性方程为Area=0.10837903*Amt+0(r=0.99953)。检测灵敏度高,最低检测浓度可达到10 ng/ml,回收率,在91%以上。结论 HPLC可以迅速、准确地检测出甲磺酸帕珠沙星在血清、龈沟液和唾液样本中的含量。

Objective To establish a rapid and sensitive high-performance liquid chromatography （HPLC） method for detecting pazufloxacin concentrations in the saliva, gingival crevicular fluid and serum of healthy adults. Methods Samples of saliva, gingival crevicular fluid and serum were obtained from healthy adults receiving intravenous infusion of pazufloxacin. The concentrations of pazufloxacin in the samples were quantified by HPLC equipped with a reversed-phase column （Agilent Zorbax SB-C18 5 μm, 250 mm×4.6 mm）. The mobile phase for pazufloxacin was a mixture of acetonitrile and 0.5% phosphoric acid containing 1% triethylamine （155 ： 850）, and 20 μl of the resulting solution was injected into the HPLC system at a flow rate of 1.0 ml/min. The detection wavelength was set at 245 nm. The samples were first deproteinized by precipitation with methanol followed by supematant drying; the residue was reconstituted with the mobile phase and centrifuged, and the supernatants were directly injected into the HPLC system. Results Pazufloxacin in the samples were totally separated without interference by any endogenous substances. The calibration curves showed a good linear regression （r〉0.999）. The detection limit was 10 ng/ml with within-day and between-day coefficients of variation performance all below 5% and recovery rates all above 91%. Conclusion HPLC is both sensitive and selective for quantification of pazufloxacin in saliva, gingival crevicul fluid and serum.