Volume 358 of the series Methods in Molecular Biology™ pp 177-197

Determination of Metabolic Flux Ratios From 13C-Experiments and Gas Chromatography-Mass Spectrometry Data

Protocol and Principles
  • Annik NanchenAffiliated withInstitute for Molecular Systems Biology, ETH Zürich
  • , Tobias FuhrerAffiliated withInstitute for Molecular Systems Biology, ETH Zürich
  • , Uwe SauerAffiliated withInstitute for Molecular Systems Biology, ETH Zürich

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Network topology is a necessary fundament to understand function and properties of microbial reaction networks. A valuable method for experimental elucidation of metabolic network topology is metabolic flux ratio analysis, which quantifies the relative contribution of two or more converging pathways to a given metabolite. It is based on 13C-labeling experiments, gas chromatography-mass spectrometry analysis, and probabilistic equations that relate mass distributions in proteinogenic amino acids to pathway activity. Here, we describe the protocol for sample generation and illustrate the principles underlying the calculation of metabolic flux ratios with three examples. These principles are also implemented in the publicly available software FiatFlux, which directly calculates flux ratios from the mass spectra of amino acids.