Abstract
Expanding metabolome coverage to include complex lipids and polar metabolites is essential in the generation of well-founded hypotheses in biological assays. Traditionally, lipid extraction is performed by liquid-liquid extraction using either methyl-tert-butyl ether (MTBE) or chloroform, and polar metabolite extraction using methanol. Here, we evaluated the performance of single-step sample preparation methods for simultaneous extraction of the complex lipidome and polar metabolome from human plasma. The method performance was evaluated using high-coverage Hydrophilic Interaction Liquid Chromatography-ESI coupled to tandem mass spectrometry (HILIC-ESI-MS/MS) methodology targeting a panel of 1159 lipids and 374 polar metabolites. The criteria used for method evaluation comprised protein precipitation efficiency, and relative MS signal abundance and repeatability of detectable lipid and polar metabolites in human plasma. Among the tested methods, the isopropanol (IPA) and 1-butanol:methanol (BUME) mixtures were selected as the best compromises for the simultaneous extraction of complex lipids and polar metabolites, allowing for the detection of 584 lipid species and 116 polar metabolites. The extraction with IPA showed the greatest reproducibility with the highest number of lipid species detected with the coefficient of variation (CV) < 30%. Besides this difference, both IPA and BUME allowed for the high-throughput extraction and reproducible measurement of a large panel of complex lipids and polar metabolites, thus warranting their application in large-scale human population studies.
Highlights
Blood plasma is one of the most commonly used biofluids for metabolic phenotyping, in human population studies
We demonstrate the performance of two single-step sample preparation methods, i.e., using BUME and IPA, for the simultaneous extraction of complex lipids and polar metabolites from human plasma
The ability ofrelative single‐step methods simultaneously extract signal, complex lipids polar was evaluated using abundance and to repeatability of metabolite against theand commonly metabolites was evaluated using relativewith abundance repeatability metabolite signal,for against applied protocols, a biphasic extraction
Summary
Blood plasma is one of the most commonly used biofluids for metabolic phenotyping, in human population studies. 70% of plasma metabolome diversity and so far, more than 600 distinct lipid molecular species have been detected in human plasma [2] These lipids can be classified, depending on their chemical structure, according to Lipid Maps consortium (http://www.lipidmaps.org) into sphingolipids, glycerophospholipids, fatty acids, glycerolipids, sterols, and prenols [3,4]. When compared to total lipid content, the total polar metabolite content (comprising mainly carbohydrates, amino acids, and other organic acids), represents a minor part of plasma metabolome [2]. These polar metabolites are known to regulate the intracellular energy metabolism, including glucose, lipid, and amino acid oxidation [6,7]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
