Metabolomics: quantification of intracellular metabolite dynamics.

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