高效液相色谱-电喷雾串联质谱法(LC-ESI-MS/MS)检测人血浆中罗红霉素浓度

目的: 建立人血浆中罗红霉素的高效液相色谱-电喷雾串联质谱(LC-ESI-MS/MS)检测方法。 方法: 血浆样品经乙腈沉淀蛋白后,反相液相色谱分离后进行质谱分析,采用选择反应监测模式(SRM)进行检测,罗红霉素和内标物克拉霉素的检测离子对分别为m/z 837.5→679.5和m/z 748.5→158.1。 结果: 罗红霉素在10~1,000 ng·mL^-1浓度范围内线性关系良好(r²>0.999),最低定量限(LLOQ)为10 ng·mL^-1。在人血浆基质中,高、中、低浓度及LLOQ (800,400,30和10 ng·mL^-1)4个添加水平下,罗红霉素的平均提取回收率为96.7%,日内及日间相对标准偏差分别为 ≤4.5%和 ≤8.2%。稳定性试验表明血浆样品室温下放置24 h或处理后放置12 h、-80 ℃放置3周及-80 ℃冻融3次对测定结果无影响。6份来自于不同受试者的血浆基质效应研究证实,该样品预处理方法对血浆中罗红霉素的测定无影响。 结论: 该方法样品处理简单、灵敏、特异性高,为罗红霉素临床药代动力学等研究提供了简捷、准确的分析测定方法。

Objective: To establish a method for determination of roxithromycin in human plasma by LC-ESI-MS/MS. Methods: The plasma samples were applied to a C₁₈column interfaced with a triple quadrupole tandem mass spectrometer after pretreated with acetonitrile.Positive electrospray ionization was employed as the ionization source.The analyte and IS clarithromycin were detected by the use of selected reaction monitoring (SRM) of the transitions m/z 837.5→m/z 679.5 for roxithromycin and m/z 748.5→m/z 158.1 for I.S,respectively. Results: The method was linear in the concentration range of 10-1,000 ng·mL^-1 for plasma roxithromycin samples.The lower limit of quantification (LLOQ) was 10 ng·mL^-1.The mean extraction recovery of roxithromycin was 96.7%.The intra-and inter-day relative standard deviation for roxithromycin plasma samples at LLOQ and three QC concentrations (10,30,400 and 800 ng·mL^-1) across six validation runs was less than 4.5% and 8.2%,respectively.Plasma matrix effect test shows endogenous matrix has no effect on the quantification of roxithromycin. Conclusion: A rapid and sensitive LC-MS/MS method was developed and validated for the determination of roxithromycin in human plasma.It can be applied to evaluate the pharmacokinetics of roxithromycin.