A rapid and quantitative LC-MS/MS method to profile sphingolipids
Sphingolipids comprise a highly diverse and complex class of molecules that serve not only as structural components of membranes but also as signaling molecules. To understand the differential role of sphingolipids in a regulatory network, it is important to use specific and quantitative methods. We...
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doaj-32593e1c467349f18bb081979309b2d72021-04-28T06:02:00ZengElsevierJournal of Lipid Research0022-22752010-07-0151720012011A rapid and quantitative LC-MS/MS method to profile sphingolipidsMax Scherer0Kerstin Leuthäuser-Jaschinski1Josef Ecker2Gerd Schmitz3Gerhard Liebisch4Institute for Clinical Chemistry and Laboratory Medicine, University of Regensburg, Regensburg, GermanyInstitute for Clinical Chemistry and Laboratory Medicine, University of Regensburg, Regensburg, GermanyInstitute for Clinical Chemistry and Laboratory Medicine, University of Regensburg, Regensburg, GermanyInstitute for Clinical Chemistry and Laboratory Medicine, University of Regensburg, Regensburg, GermanyTo whom correspondence should be addressed; Institute for Clinical Chemistry and Laboratory Medicine, University of Regensburg, Regensburg, GermanySphingolipids comprise a highly diverse and complex class of molecules that serve not only as structural components of membranes but also as signaling molecules. To understand the differential role of sphingolipids in a regulatory network, it is important to use specific and quantitative methods. We developed a novel LC-MS/MS method for the rapid, simultaneous quantification of sphingolipid metabolites, including sphingosine, sphinganine, phyto-sphingosine, di- and trimethyl-sphingosine, sphingosylphosphorylcholine, hexosylceramide, lactosylceramide, ceramide-1-phosphate, and dihydroceramide-1-phosphate. Appropriate internal standards (ISs) were added prior to lipid extraction. In contrast to most published methods based on reversed phase chromatography, we used hydrophilic interaction liquid chromatography and achieved good peak shapes, a short analysis time of 4.5 min, and, most importantly, coelution of analytes and their respective ISs. To avoid an overestimation of species concentrations, peak areas were corrected regarding isotopic overlap where necessary. Quantification was achieved by standard addition of naturally occurring sphingolipid species to the sample matrix. The method showed excellent precision, accuracy, detection limits, and robustness. As an example, sphingolipid species were quantified in fibroblasts treated with myriocin or sphingosine-kinase inhibitor. In summary, this method represents a valuable tool to evaluate the role of sphingolipids in the regulation of cell functions.http://www.sciencedirect.com/science/article/pii/S0022227520371224HILIChigh throughputESIfree sphingoid basemethylated sphingoid basehexosylceramide |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Max Scherer Kerstin Leuthäuser-Jaschinski Josef Ecker Gerd Schmitz Gerhard Liebisch |
spellingShingle |
Max Scherer Kerstin Leuthäuser-Jaschinski Josef Ecker Gerd Schmitz Gerhard Liebisch A rapid and quantitative LC-MS/MS method to profile sphingolipids Journal of Lipid Research HILIC high throughput ESI free sphingoid base methylated sphingoid base hexosylceramide |
author_facet |
Max Scherer Kerstin Leuthäuser-Jaschinski Josef Ecker Gerd Schmitz Gerhard Liebisch |
author_sort |
Max Scherer |
title |
A rapid and quantitative LC-MS/MS method to profile sphingolipids |
title_short |
A rapid and quantitative LC-MS/MS method to profile sphingolipids |
title_full |
A rapid and quantitative LC-MS/MS method to profile sphingolipids |
title_fullStr |
A rapid and quantitative LC-MS/MS method to profile sphingolipids |
title_full_unstemmed |
A rapid and quantitative LC-MS/MS method to profile sphingolipids |
title_sort |
rapid and quantitative lc-ms/ms method to profile sphingolipids |
publisher |
Elsevier |
series |
Journal of Lipid Research |
issn |
0022-2275 |
publishDate |
2010-07-01 |
description |
Sphingolipids comprise a highly diverse and complex class of molecules that serve not only as structural components of membranes but also as signaling molecules. To understand the differential role of sphingolipids in a regulatory network, it is important to use specific and quantitative methods. We developed a novel LC-MS/MS method for the rapid, simultaneous quantification of sphingolipid metabolites, including sphingosine, sphinganine, phyto-sphingosine, di- and trimethyl-sphingosine, sphingosylphosphorylcholine, hexosylceramide, lactosylceramide, ceramide-1-phosphate, and dihydroceramide-1-phosphate. Appropriate internal standards (ISs) were added prior to lipid extraction. In contrast to most published methods based on reversed phase chromatography, we used hydrophilic interaction liquid chromatography and achieved good peak shapes, a short analysis time of 4.5 min, and, most importantly, coelution of analytes and their respective ISs. To avoid an overestimation of species concentrations, peak areas were corrected regarding isotopic overlap where necessary. Quantification was achieved by standard addition of naturally occurring sphingolipid species to the sample matrix. The method showed excellent precision, accuracy, detection limits, and robustness. As an example, sphingolipid species were quantified in fibroblasts treated with myriocin or sphingosine-kinase inhibitor. In summary, this method represents a valuable tool to evaluate the role of sphingolipids in the regulation of cell functions. |
topic |
HILIC high throughput ESI free sphingoid base methylated sphingoid base hexosylceramide |
url |
http://www.sciencedirect.com/science/article/pii/S0022227520371224 |
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