Abstract
In the field of metabolomics, GC-MS has rather established itself as a
tool for semi-quantitative strategies like metabolic fingerprinting or
metabolic profiling. Absolute quantification of intra- or extracellular
metabolites is nowadays mostly accomplished by application of diverse
LC-MS techniques. Only few groups have so far adopted GC-MS technology
for this exceptionally challenging task. Besides numerous and deeply
investigated problems related to sample generation, the pronounced
matrix effects in biological samples have led to the almost mandatory
application of isotope dilution mass spectrometry (IDMS) for the
accurate determination of absolute metabolite concentrations.
Nevertheless, access to stable isotope labeled internal standards
(ILIS), which are in many cases commercially unavailable, is quite
laborious and very expensive. Here we present an improved and simplified
gas chromatography-isotope dilution mass spectrometry (GC-IDMS) protocol
for the absolute determination of intra- and extracellular metabolite
levels. Commercially available C-13-labeled algal cells were used as a
convenient source for the preparation of internal standards. Advantages
as well as limitations of the described method are discussed. (C) 2011
Elsevier B.V. All rights reserved.
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