Determination of warfarin alcohols by ultra-high performance liquid chromatography-tandem mass spectrometry: Application to in vitro enzyme kinetic studies
Determination of warfarin alcohols by ultra-high performance liquid chromatography-tandem mass spectrometry: Application to in vitro enzyme kinetic studies
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DOI：
10.1016/j.jchromb.2013.11.014
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发表时间：
2014-01-01
影响因子：
3
通讯作者：
Nolin, Thomas D.
中科院分区：
文献类型：
--
作者：
Alshogran, Osama Y.;Ocque, Andrew J.;Nolin, Thomas D.
开发并验证了一种灵敏、准确、重现性好的超高效液相色谱-串联质谱法，用于测定10 mM Tris-HCl孵育缓冲液（pH 7.4）中华法林及其醇代谢产物（RS/SR-和RR/SS-华法林醇）。样品制备涉及用4%甲酸酸化，然后用甲基叔丁基醚进行液-液萃取。使用Hypersil Gold C18（2.1 mm × 100 mm，1.9 μ m）分析柱，用溶剂A（含0.01%甲酸的水）和溶剂B（含0.1%甲酸的乙腈）进行梯度洗脱，实现色谱分离。流速为0.4 mL/min，总运行时间为5 min。使用加热电喷雾电离（负模式）和选择反应监测进行分析物检测。在华法林100- 10，000 ng/mL和华法林醇0.5-250 ng/mL的标准曲线浓度范围内，观察到所有分析物的良好线性。分析物的日内和日间准确度和精密度均在+/-10.0%范围内。观察到极好的回收率和可忽略的基质效应。该方法具有灵敏、准确、重现性好等优点，可用于华法林体外酶动力学研究。(C)2013爱思唯尔有限公司版权所有。
A sensitive, accurate, and reproducible ultra-high performance liquid chromatography-tandem mass spectrometry method was developed and validated for determination of warfarin and its alcohol metabolites (RS/SR- and RR/SS-warfarin alcohol) in 10 mM Tris-HCl incubation buffer (pH 7.4). Sample preparation involved acidification with 4% formic acid, followed by liquid-liquid extraction using methyl tert-butyl ether. Chromatographic separation was achieved using a Hypersil Gold C18 (2.1 mm X 100 mm, 1.9 mu m) analytical column with gradient elution of solvent A (water containing 0.01% formic acid) and solvent B (acetonitrile containing 0.1% formic acid). The flow rate was 0.4 mL/min and the total run time was 5 min. Detection of analytes was performed using heated electrospray ionization (negative mode) and selected reaction monitoring. Excellent linearity was observed for all analytes over the standard curve concentration ranges of 100-10,000 ng/mL for warfarin, and 0.5-250 ng/mL for warfarin alcohols. The intra- and inter-day accuracy and precision for analytes were within +/- 10.0%. Excellent recovery and negligible matrix effects were observed. The method is robust, sensitive, accurate and reproducible, and was successfully applied to in vitro enzyme kinetic studies of warfarin. (C) 2013 Elsevier B.V. All rights reserved.