目的 建立测定人血清中伏立康唑(voriconazole, VRCZ)浓度的高效液相色谱-紫外检测(HPLC-UV)方法。方法 200 μL血清样本采用400 μL二氯甲烷萃取,采用HPLC测定,以硝西泮为内标,色谱柱为kromasil(4.6 mm×150 mm,5 μm),流动相为水-乙腈(体积比60∶40),流速1.0 mL·min-1,柱温35 ℃,进样量20 μL,检测波长为254 nm。结果 VRCZ在0.2~16 μg·mL-1内线性关系良好(Y=0.073X+0.001 9,r =0.999 2),定量下限(LLOQ)为0.2 μg·mL-1;日内、日间精密度为1.23%~5.51%,准确度为88.5%~102.36%,提取回收率为92.0% ~106.5%,RSD<15%。血清样品在经历3次冻融(-20 ℃到室温)循环、冷藏放置2周和冷冻放置 30 d等条件下均稳定,处理后样品在室温和自动进样器中(4 ℃)各放置24 h均稳定(RSD<15%),VRCZ储备液于-20 ℃和工作液于4 ℃放置10个月,内标储备液于-20 ℃放置1.5个月稳定(RSD<15%)。测定9个临床样本,范围在0.81~7.96 μg·mL-1之间, 结论 该法操作简便、灵敏、准确,适用于VRCZ血药浓度的测定。
Abstract
OBJECTIVE To establish a HPLC-UV method for the determination of voriconzole in human serum. METHODS A Kromasil column (4.6 mm×150 mm,5 μm) was used, and the mobile phase consisted of H2O-acetonitrile (60∶40). The flow rate was 1.0 mL·min-1, the detection wavelength was set at 254 nm, and column temperature was maintainted at 35 ℃. RESULTS Voriconzole concentration had good linearity (r=0.999 1) in the range of 0.2-16 μg·mL-1, the regression equation was Y=0.073X+0.001 9,r =0.999 2. The limit of quantitation was 0.2 μg·mL-1 and the intra-day and inter-day precision is 1.23%-5.51%. The accuracy was 88.5%-102.36%, the extraction recovery was 92.0%-106.5%, and the RSD was <15%. The serum samples were stable after 3 freeze-thaw cycles (-20 ℃ to room temperature), being stored at 4 ℃ for 2 weeks and frozen for 30 days. The processed samples were stable at room temperature and autosampler (4 ℃) for 24 h (RSD<15%). The voriconazole stock solution and working solution were stable at -20 ℃ and 4 ℃ for 10 months, and the internal standard stock solution was stable at -20 ℃ for 1.5 months (RSD<15%). Nine samples were assayed, with the concentration of 0.81-7.96 μg·mL-1. CONCLUSION This method is simple, sensitive and accurate, and is suitable for the determination of the blood concentration of voriconzole.
关键词
伏立康唑 /
高效液相色谱-紫外检测法 /
方法学评价 /
人血清药物浓度
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Key words
voriconzole /
HPLC-UV /
method validation /
human serum drug concentration
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中图分类号：
R917
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参考文献
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脚注
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基金
北京市科学技术委员会“老年人多重用药管理模式的建立与临床应用研究”专项课题资助(D181100000218002);北京市卫生和计划生育委员会“老年重大疾病关键技术研究”项目资助(PXM2018_026283_000002)
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