【摘要】 目的 建立液相色谱串联质谱(LC-MS/MS)法测定人血浆中多黏菌素E1、E2浓度。方法 采用固相萃取法处理样品。采用Waters UPLC-API4000QTRAP进行测定，色谱柱采用Phenomenex Kinetex XB-C18柱(100 mm×2.1 mm I.D.，2.6 μm)，以0.1%甲酸溶液∶乙腈-甲醇(体积比1∶1)为流动相梯度洗脱，流速为0.4 mL/min，分析时间为3.5 min。多黏菌素E及内标多黏菌素B1的离子对分别为多黏菌素E1 m/z 390.7→101.3、多黏菌素E2 m/z 386.0→101.2、多黏菌素B1 m/z 402.3→101.2。结果 多黏菌素E1和E2的分离测定不受血浆中内源性物质影响。多黏菌素E1浓度在0.014 0～2.79 mg/L范围内线性关系良好，批内和批间精密度分别为2.7%～10.2%和2.5%～9.0%，批间准确度偏差为-4.5%～2.2%，内标归一化基质效应因子为94.8%～102.3%；多黏菌素E2浓度在0.0234～4.68 mg/L范围内线性关系良好，批内和批间精密度分别为2.8%～7.5%和2.9%～5.3%，批间准确度偏差为-4.8%～4.3%，内标归一化基质效应因子为93.1%～100.1%。血浆样品在-20℃存放3 d、-70℃存放71 d、反复冻融3 次、室温放置18 h及制备后样品4℃放置48 h均稳定。结论 建立了一种灵敏度高、分析时间短、选择性强的LC-MS/MS法，用于人血浆中多黏菌素E主要成分E1和E2的浓度测定，方法学验证结果满足生物样品定量分析方法验证指导原则的分析要求。该方法可为多黏菌素E临床药动学研究提供可靠检测方法。更多还原

【Abstract】 Objective To establish a sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of polymyxin E1 and E2 in human plasma.Methods The plasma samples were processed by solid phase extraction.The chromatographic separation was performed by Waters UPLC-API4000QTRAP system on a Phenomenex Kinetex XB-C18 (100 mm ×2.1 mm I.D.,2.6 μm) column.The mobile phase consisted of 0.1% formic acid in water and acetonitrile-methanol (v/v,1:1).The flow rate was 0.4 mL/min and the assay lasts 3.5 min per run.The multiple reaction monitoring (MRM) was performed at m/z 390.7→101.3 for polymyxin E1,m/z 386.0→101.2 for polymyxin E2,and m/z 402.3→101.2 for polymyxin B1.Results The endogenous substances in the plasma didn’t affect the analysis of polymyxin E1 and E2.For polymyxin E1,the calibration range was 0.014 0-2.79 mg/L.The intra-and inter-precision were 2.7%-10.2% and 2.5%-9.0%,respectively.The inter accuracy deviation was -4.5% to 2.2%.The internal standard-normalized matrix effect was 94.8%-102.3%.For polymyxin E2,the calibration range was 0.023 4-4.68 mg/L.The intra-and inter-precision were 2.8%-7.5% and 2.9%-5.3%,respectively.The inter accuracy deviation was -4.8% to 4.3%.The internal standard-normalized matrix effect was 93.1%-100.1%.The analyte was stable when the plasma samples were stored at-20℃ for 3 days,-70℃ for 71 days,after 3 freeze-thaw cycles,treated at room temperature for 18 h,and at 4℃ for 48 h after preparation.Conclusions The established LC-MS/MS method is highly sensitive,selective,rapid (shorter turnaround time),and reliable for the pharmacokinetics study of polymyxin to measure the concentrations of polymyxin E1 and E2 in human plasma.The results of methodological validation satisfy the requirements of the guidelines for validation of quantitative assay of biological samples.更多还原