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Biomol Eng. 2002 Jun;19(1):5-15.

Metabolomics: quantification of intracellular metabolite dynamics.

Author information

1
Institute of Biotechnology 2, Forschungszentrum J├╝lich, Germany.

Abstract

The rational improvement of microbial strains for the production of primary and secondary metabolites ('metabolic engineering') requires a quantitative understanding of microbial metabolism. A process by which this information can be derived from dynamic fermentation experiments is presented. By applying a substrate pulse to a substrate-limited, steady state culture, cellular metabolism is shifted away from its metabolic steady state. With the aid of a rapid sampling and quenching routine it is possible to take 4-5 samples per second during this process, thus capturing the metabolic response to this stimulus. Over 30 metabolites, nucleotides and cofactors from Escherichia coli metabolism can be extracted and analysed using a range of different techniques, for example enzymatic assays, HPLC and LC-MS methods. Using different substrates as limiting and pulse-substrates (glucose, glycerol), different metabolic pathways and substrate uptake systems are investigated. The resulting plots of intracellular metabolite concentrations against time serve as a data basis for modelling microbial metabolic networks.

PMID:
12103361
DOI:
10.1016/s1389-0344(02)00003-5
[Indexed for MEDLINE]

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