Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS)

Ceramide (CER) is an important signaling molecule involved in a variety of cellular processes, including differentiation, cell growth, and apoptosis. Currently, different techniques are applied for CER quantitation, some of which are relatively insensitive and/or time consuming. Tandem mass spectrom...

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Main Authors: G. Liebisch, W. Drobnik, M. Reil, B. Trümbach, R. Arnecke, B. Olgemöller, A. Roscher, G. Schmitz
Format: Article
Language:English
Published: Elsevier 1999-08-01
Series:Journal of Lipid Research
Subjects:
Online Access:http://www.sciencedirect.com/science/article/pii/S0022227520333988
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spelling doaj-251d9ad4e1864042961304aa2f824a502021-04-27T11:49:43ZengElsevierJournal of Lipid Research0022-22751999-08-0140815391546Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS)G. Liebisch0W. Drobnik1M. Reil2B. Trümbach3R. Arnecke4B. Olgemöller5A. Roscher6G. Schmitz7Institut für Klinische Chemie und Laboratoriumsmedizin, Klinikum der Universität Regensburg, Franz-Josef-Strauss-Allee 11, 93042 Regensburg, GermanyInstitut für Klinische Chemie und Laboratoriumsmedizin, Klinikum der Universität Regensburg, Franz-Josef-Strauss-Allee 11, 93042 Regensburg, GermanyInstitut für Klinische Chemie und Laboratoriumsmedizin, Klinikum der Universität Regensburg, Franz-Josef-Strauss-Allee 11, 93042 Regensburg, GermanyInstitut für Klinische Chemie und Laboratoriumsmedizin, Klinikum der Universität Regensburg, Franz-Josef-Strauss-Allee 11, 93042 Regensburg, GermanyLabor Becker, Olgemöller & Partner, München, GermanyLabor Becker, Olgemöller & Partner, München, GermanyKinderklinik, Klinische Chemie/Biochemie, Ludwig Maximilian Universität München, München, GermanyTo whom correspondence should be addressed.; Institut für Klinische Chemie und Laboratoriumsmedizin, Klinikum der Universität Regensburg, Franz-Josef-Strauss-Allee 11, 93042 Regensburg, GermanyCeramide (CER) is an important signaling molecule involved in a variety of cellular processes, including differentiation, cell growth, and apoptosis. Currently, different techniques are applied for CER quantitation, some of which are relatively insensitive and/or time consuming. Tandem mass spectrometry with its high selectivity and sensitivity is a very useful technique for detection of low abundant metabolites without prior purification or derivatization. In contrast to existing mass spectrometry methods, the developed electrospray tandem mass spectrometry (ESI-MS/MS) technique is capable of quantifying different CER species from crude cellular lipid extracts. The ESI-MS/MS is performed with a continuous flow injection and the use of an autosampler, resulting in a high throughput capability. The collision-induced fragmentation of CER produced, in addition to others, a characteristic fragment of m/z 264, making a precursor ion scan of 264 well suited for CER quantitation. Quantitation is achieved by use of a constant concentration of a non-naturally occurring internal standard C8-CER, together with a calibration curve established by spiking different concentrations of naturally occurring CER. The calibration curves showed linearity over a wide concentration range and sample volumes equivalent to 10 μg of cell protein corresponding to about 20,000 fibroblasts were sufficient for CER analysis. Moreover this assay showed a detection limit at the subpicomole level. In summary, this methodology enables accurate and rapid analysis of CER from small samples without prior separation steps, thus providing a useful tool for signal transduction research.—Liebisch, G., W. Drobnik, M. Reil, B. Trümbach, R. Arnecke, B. Olgemöller, A. Roscher, and G. Schmitz. Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS). J. Lipid Res. 1999. 40: 1539–1546.http://www.sciencedirect.com/science/article/pii/S0022227520333988ceramide quantitationelectrospray ionizationtandem mass spectrometry
collection DOAJ
language English
format Article
sources DOAJ
author G. Liebisch
W. Drobnik
M. Reil
B. Trümbach
R. Arnecke
B. Olgemöller
A. Roscher
G. Schmitz
spellingShingle G. Liebisch
W. Drobnik
M. Reil
B. Trümbach
R. Arnecke
B. Olgemöller
A. Roscher
G. Schmitz
Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS)
Journal of Lipid Research
ceramide quantitation
electrospray ionization
tandem mass spectrometry
author_facet G. Liebisch
W. Drobnik
M. Reil
B. Trümbach
R. Arnecke
B. Olgemöller
A. Roscher
G. Schmitz
author_sort G. Liebisch
title Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS)
title_short Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS)
title_full Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS)
title_fullStr Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS)
title_full_unstemmed Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS)
title_sort quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (esi-ms/ms)
publisher Elsevier
series Journal of Lipid Research
issn 0022-2275
publishDate 1999-08-01
description Ceramide (CER) is an important signaling molecule involved in a variety of cellular processes, including differentiation, cell growth, and apoptosis. Currently, different techniques are applied for CER quantitation, some of which are relatively insensitive and/or time consuming. Tandem mass spectrometry with its high selectivity and sensitivity is a very useful technique for detection of low abundant metabolites without prior purification or derivatization. In contrast to existing mass spectrometry methods, the developed electrospray tandem mass spectrometry (ESI-MS/MS) technique is capable of quantifying different CER species from crude cellular lipid extracts. The ESI-MS/MS is performed with a continuous flow injection and the use of an autosampler, resulting in a high throughput capability. The collision-induced fragmentation of CER produced, in addition to others, a characteristic fragment of m/z 264, making a precursor ion scan of 264 well suited for CER quantitation. Quantitation is achieved by use of a constant concentration of a non-naturally occurring internal standard C8-CER, together with a calibration curve established by spiking different concentrations of naturally occurring CER. The calibration curves showed linearity over a wide concentration range and sample volumes equivalent to 10 μg of cell protein corresponding to about 20,000 fibroblasts were sufficient for CER analysis. Moreover this assay showed a detection limit at the subpicomole level. In summary, this methodology enables accurate and rapid analysis of CER from small samples without prior separation steps, thus providing a useful tool for signal transduction research.—Liebisch, G., W. Drobnik, M. Reil, B. Trümbach, R. Arnecke, B. Olgemöller, A. Roscher, and G. Schmitz. Quantitative measurement of different ceramide species from crude cellular extracts by electrospray ionization tandem mass spectrometry (ESI-MS/MS). J. Lipid Res. 1999. 40: 1539–1546.
topic ceramide quantitation
electrospray ionization
tandem mass spectrometry
url http://www.sciencedirect.com/science/article/pii/S0022227520333988
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