Journal
METABOLIC ENGINEERING
Volume 6, Issue 1, Pages 44-58Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymben.2003.10.004
Keywords
heart; citric acid cycle; anaplerosis; glutamate; substrate oxidation
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Funding
- NATIONAL CANCER INSTITUTE [P30CA013148] Funding Source: NIH RePORTER
- NATIONAL HEART, LUNG, AND BLOOD INSTITUTE [R01HL067464, T32HL007703, R29HL048789, R01HL048789] Funding Source: NIH RePORTER
- NCI NIH HHS [CA-13148] Funding Source: Medline
- NHLBI NIH HHS [HL48789, 5T32HL07703, HL67464] Funding Source: Medline
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The aim of this article is to provide a guide for metabolic physiologists and bioengineers to the combined use of gas chromatography-mass spectrometry (GCMS) and nuclear magnetic resonance (NMR) in stable isotope investigations in any biological systems. Building on our past experience with these two techniques, as applied separately to the investigation of citric acid metabolism in the ex vivo perfused rat heart we initiated a collaborative study for their critical evaluation. This article, which expands on our previous work (Mol. Cel. Biol., 2003), directly compares GCMS- and NMR-determined C-13-isotopomer and flux data obtained from ex vivo rat heart perfusion studies with C-13-substrates. Overall we have found excellent agreement between the C-13-enrichments of GCMS- and NMR-determined citric acid cycle metabolites (citrate, 2-ketoglutarate, succinate and malate) and glutamate; however the unlabeled component (M) was consistently underestimated by NMR. Despite this discrepancy there was reasonably good agreement in the relative fluxes of C-13-substrates through the citric acid cycle determined by the two techniques. Nevertheless, further investigations appear necessary before maximal advantage can be taken of the complementary C-13-isotopomer and flux data of GCMS and NMR for probing the dynamics of cellular metabolism. (C) 2003 Elsevier Inc. All rights reserved.
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