Electron Partitioning During Light- and Nutrient-Powered Hydrogen Production by Rhodobacter sphaeroides

Abstract

The partitioning of reducing power into different electron-accepting pathways was evaluated during growth and stationary phases of H2-producing Rhodobacter sphaeroides cultures. For this, an electron balance method was developed using the chemical oxygen demand concept to quantitatively analyze the partitioning of nutrient electrons into H2, cell biomass, polyhydroxybutyrate (PHB), and soluble microbial products (SMP). Overall, these four electron sinks were accounted for greater than 85% of the electrons provided by the nutrients. Glucose, lactate, succinate, fumarate, and pyruvate were individually provided as the main carbon source, and in all cases, glutamate was provided as a nitrogen source in order to enhance H2 production. About 25–35% of the electrons ended up in H2 during growth, while up to 60% of the electrons partitioned into H2 in some stationary phase cultures. The other two major electron sinks in the growth phase were cell mass and PHB, while in stationary phase, SMP were accounted for >30% of the substrate electrons utilized. In general, the largest portion of SMP comprised low-molecular weight (<3 kDa) compounds mostly produced during stationary phase, although larger-size molecules were also detected in both phases. Overall, the fractions of electrons that partitioned into H2 (0.21 to 0.35) and PHB (0.06 to 0.21) were highly correlated with the standard free energy change of the substrate oxidation half-reaction equation, normalized per electron equivalent. In a PHB(−) mutant, electron redistribution increased H2 production, the extent of which depended on the carbon source provided.