Optimization and Evaluation of Metabolite Extraction Protocols for Untargeted Metabolic Profiling of Liver Samples by UPLC-MS
Optimization and Evaluation of Metabolite Extraction Protocols for Untargeted Metabolic Profiling of Liver Samples by UPLC-MS
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DOI：
10.1021/ac101722e
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发表时间：
2010-09-15
影响因子：
7.4
通讯作者：
Want, Elizabeth J.
中科院分区：
文献类型：
--
作者：
Masson, Perrine;Alves, Alexessander Couto;Want, Elizabeth J.
在获得的代谢物特征的再现性和数量方面，评价了一系列六种方案用于基于UPLC-MS的肝脏提取物的非靶向代谢谱分析。这些方案设计用于提取极性和非极性代谢物，基于（i）两阶段提取方法或（ii）采用不同体积和提取与重悬溶剂组合在两相混合物中同时提取。开发了一种多元统计策略，以比较方法之间的多维变化。发现分析极性和非极性代谢物的最佳方案是用甲醇/水进行水提取，然后用二氯甲烷/甲醇进行有机提取，在UPLC-MS分析前将干燥提取物重悬于甲醇/水中。该方案导致以下实验重复中特征强度的中值CV：
A series of six protocols were evaluated for UPLC-MS based untargeted metabolic profiling of liver extracts in terms of reproducibility and number of metabolite features obtained. These protocols, designed to extract both polar and nonpolar metabolites, were based on (i) a two stage extraction approach or (ii) a simultaneous extraction in a biphasic mixture, employing different volumes and combinations of extraction and resuspension solvents. A multivariate statistical strategy was developed to allow comparison of the multidimensional variation between the methods. The optimal protocol for profiling both polar and nonpolar metabolites was found to be an aqueous extraction with methanol/water followed by an organic extraction with dichloromethane/methanol, with resuspension of the dried extracts in methanol/water before UPLC-MS analysis. This protocol resulted in a median CV of feature intensities among experimental replicates of