Accurate Assessment of Amino Acid Mass Isotopomer Distributions for Metabolic Flux Analysis

Metabolic flux analysis based on stable-isotope labeling experiments and analysis of mass isotopomer distributions (MID) of cellular metabolites is a tool of great significance for metabolic engineering and study of human disease. This method relies on accurate and precise measurements of mass isoto...

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Bibliographic Details
Published in:Analytical chemistry (Washington) 2007-10, Vol.79 (19), p.7554-7559
Main Authors: Antoniewicz, Maciek R, Kelleher, Joanne K, Stephanopoulos, Gregory
Format: Article
Language:English
Subjects:
Amino acids
Amino Acids - chemistry
Analytical chemistry
Chemistry
Chromatographic methods and physical methods associated with chromatography
Chromatography
Escherichia coli - chemistry
Exact sciences and technology
Gas chromatographic methods
Gas Chromatography-Mass Spectrometry
Isotopes
Mass spectrometry
Metabolism
Reproducibility of Results
Spectrometric and optical methods
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Summary:Metabolic flux analysis based on stable-isotope labeling experiments and analysis of mass isotopomer distributions (MID) of cellular metabolites is a tool of great significance for metabolic engineering and study of human disease. This method relies on accurate and precise measurements of mass isotopomers by gas chromatography/mass spectrometry. To improve flux estimates, we assessed potential errors in determining MID of tert-butyldimethylsilyl-derivatized amino acids, which were attributed to (i) the choice of integration algorithm, (ii) concentration effects, and (iii) overlapping fragments. We report 29 amino acid fragments that are useful for flux analysis and another 18 fragments that should be rejected, most importantly Val-302, Leu-200, Leu-302, Ile-302, Ser-302, and Asp-316. In addition, we provide a protocol to minimize errors for determining MID to less than 0.4 mol % for accepted fragments.
ISSN:0003-2700
1520-6882
DOI:10.1021/ac0708893