UPLC-MS/MS法测定人尿液中替芬泰及其代谢物浓度  

A validated UPLC-MS/MS method for determination of Y101 and its metabolites in human urine

作　　者：刘筱雪[1] 黄晨蓉 薛领[1] 许青青[1] 张诗超[1] 张华[1] 钱丽芳[1] 蒋坤 吴贵辉 李星 夏文 缪丽燕[1] LIU Xiao-xue;HUANG Chen-rong;XUE Ling;XU Qing-qing;ZHANG Shi-chao;ZHANG Hua;QIAN Li-fang;JIANG Kun;WU Gui-hui;LI Xing;XIA Wen;MIAO Li-yan(Department of Clinical Pharmacology,The First Affliated Hospital of Soochou University,Suzhou 215006,China;Bailing Enterprise Group Pharmaceutical Co.,Ld.,Anshun 561000,China)

机构地区：[1]苏州大学附属第一医院临床药理研究室,苏州215006 [2]贵州百灵企业集团制药股份有限公司,安顺561000

出　　处：《中国新药杂志》2020年第14期1628-1635,共8页Chinese Journal of New Drugs

基　　金：国家科技重大专项课题资助项目(2015ZX09101012);国家自然科学基金青年科学基金资助项目(81703619);贵州省科技计划课题资助项目[黔科合重大专项字(2015)6002号];江苏省药学会奥赛康医院药学基金资助项目(A201723)。

摘　　要：目的:建立人尿液中替芬泰(Y101)及其代谢物M8和M9浓度的超高效液相色谱-串联质谱(UPLC-MS/MS)分析方法并用于Y101在人尿液中排泄的研究。方法:采用甲醇沉淀法进行尿液前处理并稀释后分析,色谱柱为ACQUITY UPLC BEH C18(50 mm×2.1 mm,1.7μm),流动相为5 mmol·L^-1醋酸铵水溶液(含0.1%甲酸)-甲醇=57∶43(v∶v),流速为0.3 mL·min^-1,柱温为40℃,进样量为2μL。质谱采用电喷雾离子源(electrospray ionization,ESI),正离子模式,多反应监测(multiple reaction monitoring,MRM)方式。Y101、代谢物M8、M9和帕拉米韦(内标)的监测离子对为m/z490.1→339.0,m/z357.2→105.2,m/z373.1→105.1和m/z329.1→270.2。结果:该方法选择性良好,人尿液中原型Y101、代谢物M8和M9分别在00200~20.0,0.200~100和0.0300~10.0μg·mL^-1范围内线性良好,平均回收率分别为95.9%~102.2%,973%~99.3%和98.1%~98.5%,内标归一化基质因子分别为095~104,098~102和099~100,批内、批间精密度均<15%。同时,对Y101、代谢物M8和M9浓度分别为1500,4000和3000μg·mL^-1的样本采用空白基质进行10倍稀释的稀释可靠性考察,结果表明分析物准确度为920%~1070%,精密度<5%。结论:所建立的UPLC-MS/MS分析方法专属性强、灵敏度高、重现性好,适用于人尿液样本Y101及其代谢物M8和M9的定量分析。Objective:To establish an ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)method for determination of Y101 and its metabolites M8 and M9 in human urine for the study ofits excretion.Methods:Urine samples were pretreated by protein precipitation and analyzed after dilution.Thechromatography was performed on an ACQUITY UPLC BEH C18(50 mm×2.1 mm,1.7μm,Waters)column usinga mixture of 0.1%formic acid water,5 mmol·L^-1 um acetate and methanol(57∶43,v∶v)as the mobilephase at a flow rate of 0.3 mL·min^-1 ammoni.The injection volume was 2μL.The column temperature was 40℃.Positiveelectrosmpray ionization(ESI)detection was performed in multiple reaction monitoring(MRM)mode using targetions mat/z490.1→339.0 for Y101,m/z357.2→105.2 for metabolite M8,m/z373.1→105.1 for metabolite M9 and/z329.1→270.2 for peramivir.Results:The specificity of the method was good.The calibration curve exhibited excellent linearity over the range of 0.0200~20.0,0.200~100 and 0.0300~10.0μg·mL-1 forY101,metabolite M8,and M9,respectively.The average recovery rate was 95.9%~102.2%,97.3%~99.3%and 98.1%~98.5%and the internal standard normalized matrix factors were 0.95~1.04,0.98~1.02 and 0.99~1.00 for Y101,metabolite M8,and M9,respectively.The intra-assay and inter-assay precision RSDs were lessthan 15%.Meanwhile,the dilution i n1 tegrity of 10 times dilution of samples with Y101,M8 and M9 concentrationof 150.0,400.0 and 30.00μg·mL^-1 was investigated.The results showed that the accuracy of the analytes was 92.0%~107.0%,and the precision was less than 5%.Conclusion:The established UPLC-MS/MS method hashigh specificity,sensitivity and reproducibility.It is suitable for the quantitative analysis of Y101 and its metabolitesM8 and M9 in human urine.

关 键 词：替芬泰 代谢物 尿液 超高效液相色谱-串联质谱 定量分析 

分 类 号：R917[医药卫生—药物分析学]