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Comprehensive and robust analysis of the whole blood sphingolipidome by LC‐HRMS
Author(s) -
Wang Dezhen,
Xu Peining,
Mesaros Clementina
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.01935
Subject(s) - sphingolipid , ceramide , whole blood , lipidomics , chromatography , hemolysis , sphingomyelin , chemistry , analyte , sphingosine , metabolomics , biochemistry , cholesterol , biology , immunology , apoptosis , receptor
Plasma and serum are the most widely used blood‐derived biofluids for metabolomics and lipidomics assays, but many analytes that are present at high concentrations in blood cells cannot be measured and evaluated in those samples. Variable hemolysis during the pre‐process could introduce a bias and compromise the quantification. Compared with plasma or serum, whole blood may be a better alternative due to ease of process and integrality. In this study, we provide a comprehensive and robust method for quantification of the whole blood sphingolipidome. Combining a single‐phase extraction method with liquid‐chromatography high resolution mass spectrometry (R=120, 000), assisted by alkaline hydrolysis and an inclusion list based PRM/DIA, we were able to reliably identify and simultaneously quantify more than 150 sphingolipids, including ceramide, ceramide‐1‐P, hex‐ceramide, sphingomyelin, sphinganine (1‐P), sphingosine (1‐P), and gangliosides. The capabilities of different internal standards to minimize the matrix effects at different volumes were investigated, concluding that by choosing an appropriate internal standard, most of sphingolipids can achieve a good linearity (R 2 >0.98) between normalized peak intensity and blood volume. Furthermore, most of sphingolipids remained stable after freeze/thaw cycle, indicating the potential wide application in further sphingolipids research. Finally, we compared the sphingolipidome of whole blood and paired plasma samples and found higher concentration of most sphingolipids in whole blood than in the corresponding plasma. These findings demonstrate that whole blood could be a better alternative to plasma, and potentially guide the evaluation of sphinglipidome for biomarker discovery.