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Holistic Lipidomics of the Human Gut Phenotype Using Validated Ultra-High-Performance Liquid Chromatography Coupled to Hybrid Orbitrap Mass Spectrometry

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Laboratory of Chemical Analysis, Department of Veterinary Public Health and Food Safety, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, 9820 Merelbeke, Belgium
§ Inflammatory Bowel Disease Research Unit, Department of Internal Medicine, and §Department of Endocrinology, Ghent University Hospital, De Pintelaan 185, 9000 Ghent, Belgium
Cite this: Anal. Chem. 2017, 89, 22, 12502–12510
Publication Date (Web):October 20, 2017
https://doi.org/10.1021/acs.analchem.7b03606
Copyright © 2017 American Chemical Society

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    As lipids are assigned a plethora of biological functions, it is evident that dysregulated lipid metabolism signifies a key element in many pathological conditions. With this rationale, this study presents a validated lipidomics platform to map the fecal lipidome, which integrates unique information about host–gut microbiome interactions, gastrointestinal functionality, and dietary patterns. This particular method accomplished coverage across all eight lipid categories: fatty acyls, glycerolipids, phosphoglycerolipids, polyketides, prenols, saccharolipids, sphingolipids, and sterols. Generic extraction of freeze-dried feces was achieved by solid–liquid extraction using methanol and methyl tert-butyl ether. Extracted components were separated by liquid chromatography, whereby the selected ethylene-bridged hybrid phenyl ultra-high-performance liquid chromatography stationary phase allowed fast separation of both individual lipid species and categories. Detection was achieved by high-resolution full-scan Q-Exactive Orbitrap mass spectrometry and covered a broad m/z scan range (67–2300 Da). Method validation was performed in a targeted fashion to evaluate the analytical performance across all lipid categories, revealing excellent linearity (R2 ≥ 0.9921), acceptable repeatability (coefficients of variance ≤15.6%), and stable recovery (coefficients of variance ≤11.9%). Method suitability for untargeted fingerprinting was verified, demonstrating adequate linearity (R2 ≥ 0.90) for 75.3% and acceptable repeatability (coefficients of variance ≤30%) for 84.5% of about 9000 endogenous fecal compounds. Eventually, the potential of fecal lipidomics was exemplified within a clinical context of type 2 diabetes, thereby revealing significant perturbations [orthogonal partial least-squares discriminant analysis Q2(Y) of 0.728] in the fecal lipidome between participants with normal blood glucose levels (n = 26) and those with type 2 diabetes (n = 17).

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    The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.analchem.7b03606.

    • Five tables listing authentic reference standards, authentic analytical internal standards, parameter settings for extraction of features from full-scan MS data by use of Sieve 2.2 software, statistical output to evaluate factors’ significance, and validation results for targeted endogenous lipids (PDF)

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