Applying metabolic flux analysis to hydrogen fermentation using a metabolic network constructed for anaerobic mixed cultures.

In this study, a mixed-cultural metabolic network for anaerobic digestion that included the concept of a "universal bacterium" was constructed, and metabolic flux analysis (MFA) applying this network was conducted to evaluate the flow of electrons and materials during H2 fermentation under various conditions. The MFA results from two H2 fermenters feeding glucose with (GP) or without (GA) the addition of peptone suggest that hydraulic retention time (HRT) presents a significant impact on hydrogen production, and the reversed trends could be observed at HRTs below and above 4 h. From the MFA results of lactate/acetate-fed H2 fermenter, the highest flux of H2 production is associated with more significant acetate consumption and the following pathways toward the anaplerotic reactions cycle that produces NADH. The occurrence of acetogenesis in the H2 fermenters using various types of bioethanol-fermented residues (BEFRs) was also identified according to the MFA results. By analyzing the MFA results of all 49 sets of data from H2 fermenters via Pearson's correlation, it was revealed that the flux of H2 production positively correlates to the reduction of ferredoxin with pyruvate oxidation, acetate formation, and acetate emission when lactate was produced in the system. On the contrary, negative relationships were found between the flux of H2 production and these three fluxes. The extended application of MFA provides additional information, including the fluxes between intracellular metabolites, and the information has the potential to be used in decision-making systems during the future operation of anaerobic processes by connecting operational parameters.