Abstract
BackgroundAn indirect approach is usually used to estimate the metabolic fluxes of an organism: couple the available measurements with known biological constraints (e.g. stoichiometry). Typically this estimation is done under a static point of view. Therefore, the fluxes so obtained are only valid while the environmental conditions and the cell state remain stable. However, estimating the evolution over time of the metabolic fluxes is valuable to investigate the dynamic behaviour of an organism and also to monitor industrial processes. Although Metabolic Flux Analysis can be successively applied with this aim, this approach has two drawbacks: i) sometimes it cannot be used because there is a lack of measurable fluxes, and ii) the uncertainty of experimental measurements cannot be considered. The Flux Balance Analysis could be used instead, but the assumption of optimal behaviour of the organism brings other difficulties.ResultsWe propose a procedure to estimate the evolution of the metabolic fluxes that is structured as follows: 1) measure the concentrations of extracellular species and biomass, 2) convert this data to measured fluxes and 3) estimate the non-measured fluxes using the Flux Spectrum Approach, a variant of Metabolic Flux Analysis that overcomes the difficulties mentioned above without assuming optimal behaviour. We apply the procedure to a real problem taken from the literature: estimate the metabolic fluxes during a cultivation of CHO cells in batch mode. We show that it provides a reliable and rich estimation of the non-measured fluxes, thanks to considering measurements uncertainty and reversibility constraints. We also demonstrate that this procedure can estimate the non-measured fluxes even when there is a lack of measurable species. In addition, it offers a new method to deal with inconsistency.ConclusionThis work introduces a procedure to estimate time-varying metabolic fluxes that copes with the insufficiency of measured species and with its intrinsic uncertainty. The procedure can be used as an off-line analysis of previously collected data, providing an insight into the dynamic behaviour of the organism. It can be also profitable to the on-line monitoring of a running process, mitigating the traditional lack of reliable on-line sensors in industrial environments.
Highlights
An indirect approach is usually used to estimate the metabolic fluxes of an organism: couple the available measurements with known biological constraints
Flux Spectrum Approach (FSA) provides a prediction of the non-measured fluxes, and an indication of the reliability of this prediction. This example has shown that the procedure can be useful to estimate the evolution of the fluxes even when measurements are sufficient but uncertain, i.e. when the system is determined and redundant. This is the scenarios were the procedures based on the use of Metabolic Flux Analysis (MFA) are most successful, the use of FSA provides a more reliable estimation of the non-measured fluxes and offers an interesting approach to cope with inconsistency
The results obtained with the direct analysis are included there. In this contribution we have presented a new procedure to estimate the temporal evolution of the metabolic fluxes
Summary
An indirect approach is usually used to estimate the metabolic fluxes of an organism: couple the available measurements with known biological constraints (e.g. stoichiometry). This estimation is done under a static point of view. BMC Bioinformatics 2007, 8:421 http://www.biomedcentral.com/1471-2105/8/421 state, mass balances around each metabolite can be described by means of a homogeneous system of linear equations [5]. These equations can be considered as stoichiometric constraints. Thereby the imposed constraints define a space where every feasible flux distribution lives [7]. The environmental conditions given at a certain time instant will determine which of these flux distributions corresponds to the actual one [9]
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