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首页> 外文期刊>Journal of mass spectrometry: JMS >Specificity enhancement with LC-positive ESI-MS/MS for the measurement of nucleotides: application to the quantitative determination of carbovir triphosphate, lamivudine triphosphate and tenofovir diphosphate in human peripheral blood mononuclear cells
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Specificity enhancement with LC-positive ESI-MS/MS for the measurement of nucleotides: application to the quantitative determination of carbovir triphosphate, lamivudine triphosphate and tenofovir diphosphate in human peripheral blood mononuclear cells

机译：LC阳性ESI-MS / MS用于核苷酸测量的特异性增强：在人类外周血单核细胞中卡波韦三磷酸，拉米夫定三磷酸和替诺福韦二磷酸的定量测定中的应用

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摘要

Our previous negative ESI-LC-MS/MS method developed for nucleoside reverse transcriptase inhibitor (NRTI) triphosphate (-TP) measurements in human peripheral blood mononuclear cells (PBMC) encountered some specificity problems for several NRTI-TP and simultaneous endogenous nucleotide triphosphates analysis. As LC-MS/MS offers several possibilities to circumvent such problems, we have investigated the contribution of the positive electrospray ionization mode in enhancing the specificity of the intracellular analyses of triphosphate metabolites of lamivudine, abacavir, and tenofovir. For intracellular NRTI-TP analysis, after disruption of PBMCs, concentrated supernatants were directly injected into the LC-MS/MS system, dimethylhexylamine being used as ion-pairing agent to resolve NRTI-TP. MS/MS detection was performed after positive electrospray ionization. Total run time was 12 min instead of 26 min for NRTI-TP analysis. The validation parameters of the method met the international requirements, and endogenous chromatographic interferences were eliminated. The use of positive ESI, offering a better specificity and a slightly better sensitivity than the negative ESI mode for these compounds, resulted in specificity enhancement and more robust assay methods.
机译：我们先前为人类外周血单核细胞（PBMC）中的核苷逆转录酶抑制剂（NRTI）三磷酸（-TP）测量开发的阴性ESI-LC-MS / MS方法在几种NRTI-TP和同时进行内源核苷酸三磷酸分析时遇到了一些特异性问题。由于LC-MS / MS提供了几种解决此类问题的可能性，因此我们研究了正电喷雾电离模式在增强拉米夫定，阿巴卡韦和替诺福韦的三磷酸代谢物的细胞内分析特异性方面的贡献。对于细胞内NRTI-TP分析，在PBMC破裂后，将浓缩的上清液直接注入LC-MS / MS系统中，使用二甲基己胺作为离子配对剂来解决NRTI-TP。在正电喷雾电离后进行MS / MS检测。 NRTI-TP分析的总运行时间为12分钟，而不是26分钟。该方法的验证参数符合国际要求，消除了内源色谱干扰。对于这些化合物，使用阳性ESI比阴性ESI模式具有更好的特异性和稍微更好的灵敏度，从而导致特异性增强和更稳定的测定方法。

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