Kuklenyik Zsuzsanna, Martin Amy, Pau Chou-Pong, Holder Angela, Youngpairoj Ae S, Zheng Qi, Cong Mian-Er, Garcia-Lerma J Gerardo, Heneine Walid, Pirkle James L, Barr John R
Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA 30341,United States.
J Chromatogr B Analyt Technol Biomed Life Sci. 2009 Nov 1;877(29):3659-66. doi: 10.1016/j.jchromb.2009.09.007. Epub 2009 Sep 11.
We developed an automated on-line weak anion exchange (WAX) solid-phase extraction (SPE) method coupled with ion-pair (IP) chromatography-tandem mass spectrometry (MS/MS) detection for quantitatively measuring triphosphorylated metabolites of three reverse transcriptase inhibitors (RTI). The administered pro-drugs were Tenofovir disoproxil fumarate (TDF), Emtricitabine (FTC) and Lamivudine (3TC). Their intracellular metabolites Tenofovir-diphosphate (TFV-DP), Emtricitabine-triphosphate (FTC-TP), and Lamivudine-triphosphate (3TC-TP) were measured in peripheral blood mononuclear cells (PBMC). We coupled the WAX and IP chromatography systems using a combination of 6-port and 10-port switching valves, and we mixed the WAX elute with 1,5-dimethyl-hexyl-amine before IP chromatography separation. Multiple waste outlets allowed for eliminating potential matrix components interfering with MS/MS detection. Limits of detection were 9, 200 and 75 pg per sample for TFV-DP (448/176 m/z), FTC-TP (488/130 m/z) and 3TC-TP (468/119 m/z), respectively.
我们开发了一种自动化在线弱阴离子交换（WAX）固相萃取（SPE）方法，结合离子对（IP）色谱-串联质谱（MS/MS）检测，用于定量测定三种逆转录酶抑制剂（RTI）的三磷酸化代谢物。所使用的前体药物为替诺福韦酯富马酸盐（TDF）、恩曲他滨（FTC）和拉米夫定（3TC）。在人外周血单个核细胞（PBMC）中测定了它们的细胞内代谢物替诺福韦二磷酸（TFV-DP）、恩曲他滨三磷酸（FTC-TP）和拉米夫定三磷酸（3TC-TP）。我们使用6通和10通切换阀组合连接WAX和IP色谱系统，并在IP色谱分离前将WAX洗脱液与1,5-二甲基己胺混合。多个废液出口可消除干扰MS/MS检测的潜在基质成分。TFV-DP（448/176 m/z）、FTC-TP（488/130 m/z）和3TC-TP（468/119 m/z）的检测限分别为每个样品9、200和75 pg。
