Benefits of prolonged gradient separation for high-performance liquid chromatography-tandem mass spectrometry quantitation of plasma total 15-series F2-isoprostanes
Benefits of prolonged gradient separation for high-performance liquid chromatography-tandem mass spectrometry quantitation of plasma total 15-series F2-isoprostanes
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DOI：
10.1016/j.ab.2005.12.003
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发表时间：
2006-03-01
影响因子：
2.9
通讯作者：
Traber, MG
中科院分区：
文献类型：
--
作者：
Taylor, AW;Bruno, RS;Traber, MG
F-2-异前列腺素是前列腺素F-2α(PGF(2α))的立体异构体花生四烯酸(AA)在自由基作用下氧化的产物。本文介绍了一种高效液相色谱-串联质谱法测定多种15系列前列腺素F异构体(15-PGF)和AA的方法。血浆样品经碱解、酸化后，用有机溶剂提取总(游离+酯化)15-PGF和AA。分析物用梯度反相高效液相色谱分离，在三重四极杆质谱仪上进行多反应监测，使用氚内标物进行定量。8-iso-PGF(2α)在色谱柱上的线性范围为1-40pg(2α)，可定量至40pg/mL(0。11 nM)。结果显著相关(p<；0.0001)与通过气相色谱-质谱仪、气相色谱-质谱仪或酶联免疫吸附试验获得的数据相关。所有血浆15-前列腺素F异构体随着体外香烟烟雾暴露时间的延长而增加，并相互关联(p<；0.0001)。在健康的青年受试者的血浆中也观察到了同样强的15-PGF间相关性。该方法的变异系数(24-32%)小于气相色谱-质谱法(53%)。因此，与现有方法相比，高效液-质联用法有可能提供更高的精密度，并允许以更简单的样品制备来定量更多的化合物。我们的方法是第一个应用于血浆总15-PGF的高效液相色谱-串联质谱仪定量分析方法。(C)2005 Elsevier Inc.保留所有权利。
The F-2-isoprostanes are products of free-radical -induced oxidation of arachidonic acid (AA) that are stercoisomers of prostaglandin F-2 alpha (PGF(2 alpha)). We describe a method for quantitation of several 15-series PGF isomers (15-PGFs) and AA by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS-MS). Plasma samples were subjected to alkaline hydrolysis and acidified, and total (free + esterified) 15-PGFs and AA were extracted with organic solvents. The analytes were separated by gradient reverse-phase HPLC and detected by multiple reaction monitoring on a triple-quadrupole mass spectrometer, using deuterated internal standards for quantitation. The assay had a linear range of 1-40 pg of 8-iso-PGF(2 alpha) on column and can quantify as little as 40 pg/mL (0. 11 nM) in plasma. Outcomes significantly correlated (p < 0.0001) with data obtained by gas chromatography-mass spectrometry GC MS or enzyme-linked immunosorbent assay. All plasma 15-PGF isomers increased over time with in vitro cigarette smoke exposure and correlated (p < 0.0001) with each other. The same strong inter-15-PGF correlations were observed in plasma from healthy young adult subjects. The coefficients of variation of HPLC-NIS-NIS measurements (24-32%) were smaller than those obtained by GC-MS (53%). Thus, HPLC-MS-MS potentially offers greater precision and allows quantitation of more compounds with simpler sample preparation than existing methods. Ours is the first validated quantitative assay using HPLC-tandem MS applied to plasma total 15-PGFs. (c) 2005 Elsevier Inc. All rights reserved.