Development and validation of a high-throughput LC-MS/MS assay for routine measurement of molecular ceramides

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http://hdl.handle.net/10138/223986

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Kauhanen , D , Sysi-Aho , M , Koistinen , K M , Laaksonen , R , Sinisalo , J & Ekroos , K 2016 , ' Development and validation of a high-throughput LC-MS/MS assay for routine measurement of molecular ceramides ' , Analytical and Bioanalytical Chemistry , vol. 408 , no. 13 , pp. 3475-3483 . https://doi.org/10.1007/s00216-016-9425-z

Title: Development and validation of a high-throughput LC-MS/MS assay for routine measurement of molecular ceramides
Author: Kauhanen, Dimple; Sysi-Aho, Marko; Koistinen, Kaisa M.; Laaksonen, Reijo; Sinisalo, Juha; Ekroos, Kim
Contributor: University of Helsinki, Clinicum
Date: 2016-05
Language: eng
Number of pages: 9
Belongs to series: Analytical and Bioanalytical Chemistry
ISSN: 1618-2642
URI: http://hdl.handle.net/10138/223986
Abstract: Monitoring the levels of the ceramides (Cer) d18:1/16:0, Cer d18:1/18:0, Cer d18:1/24:0, and Cer d18:1/24:1 and ratios thereof in human plasma empowers the prediction of fatal outcome of coronary artery disease (CAD). We describe a validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) methodology for clinical-scaled measurement of the four distinct ceramides. Rapid plasma precipitation was accomplished in 96-well format. Excellent extraction recoveries in the range of 98-109 % were achieved for each ceramide. Addition of corresponding D-7-labeled ceramide standards facilitated precise quantification of each plasma ceramide species utilizing a novel short 5-min LC-MS/MS method. Neither matrix interference nor carryover was observed. Robust intra- and inter-assay accuracy and precision <15 % at five different concentrations were obtained. Linear calibration lines with regressions, R (2) > 0.99, were achieved for all analytes. Short-term bench top, long-term plasma, and extract stability demonstrated that the distinct ceramides were stable in the conditions evaluated. The validity of the methodology was demonstrated by determining the precise ceramide concentrations in a small CAD case-control study. Thus, our LC-MS/MS methodology features simple sample preparation and short analysis time for accurate quantification of Cer d18:1/16:0, Cer d18:1/18:0, Cer d18:1/24:0, and Cer d18:1/24:1, designed for routine analysis.
Subject: Mass spectrometry
Chromatography
Ceramides
Cardiovascular
Clinical
High throughput
TANDEM MASS-SPECTROMETRY
PERFORMANCE LIQUID-CHROMATOGRAPHY
ELECTROSPRAY-IONIZATION
QUANTITATIVE-ANALYSIS
LIPID ANALYSIS
PLASMA
SPHINGOLIPIDOMICS
METABOLITES
EXTRACTS
SAMPLES
1182 Biochemistry, cell and molecular biology
3121 General medicine, internal medicine and other clinical medicine
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