title: An LC/MS/MS method for the simultaneous determination of individual sphingolipid species in B cells.
abstract: Comprehensive profiling of sphingolipids is of great importance for clinical and pharmaceutical studies. An LC/MS/MS method was established for the simultaneous separation and quantification of individual sphingolipid species including ceramides, dihydroceramides, glucosylceramides, sphingosine, sphingosine-1-phosphate, sphinganine and sphinganine-1-phosphate. All target individual sphingolipid species were separated and quantified in a single chromatographic run of <20min. Method validation results indicated that calibration curves were linear in the range of 2.5-10,000nM for ceramides and glucosylceramides, 10-10,000nM for dihydroceramides, 5-10,000nM for sphingosine, sphingosine-1-phosphate, sphinganine and sphinganine-1-phosphate, respectively. The limits of detection ranged from 0.5nM to 5nM. Accuracies of 92.5-113% with precisions of 0.3-8.0% RSD were obtained for all of the standards over a wide range of concentrations. The application of this method was demonstrated using B cells collected from Chronic Lymphocytic Leukemia patients (n=5) and healthy donors (n=4). 17 sphingolipid species were successfully characterized and quantified in the lipid extract. This is a rapid method that could be readily adapted to lipidomic investigations of sphingolipids in other bio-fluids and tissues.
authors.affiliation: Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB, Canada.
authors.forename: Si
authors.initials: S
authors.lastname: Mi
authors.affiliation: Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB, Canada.
authors.forename: Yuan-Yuan
authors.initials: YY
authors.lastname: Zhao
authors.affiliation: Department of Immunology, University of Manitoba, Winnipeg, Manitoba, Canada; Research Institute of Oncology and Hematology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
authors.forename: Rebecca F
authors.initials: RF
authors.lastname: Dielschneider
authors.affiliation: Department of Immunology, University of Manitoba, Winnipeg, Manitoba, Canada; Research Institute of Oncology and Hematology, Cancer Care Manitoba, Winnipeg, Manitoba, Canada.
authors.forename: Spencer B
authors.initials: SB
authors.lastname: Gibson
authors.affiliation: Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB, Canada. Electronic address: jcurtis1@ualberta.ca.
authors.forename: Jonathan M
authors.initials: JM
authors.lastname: Curtis
mesh_terms: D001402:B-Lymphocytes / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006801:Humans; D057230:Limit of Detection; D015203:Reproducibility of Results; D012189:Retrospective Studies; D013107:Sphingolipids / Q000378:metabolism*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D013107:Sphingolipids
keywords: B cells; Chronic Lymphocytic Leukemia; LC/MS/MS; Quantification; Sphingolipids; Validation