Activation energy measurements to determine metabolic bottlenecks of a bioelectrochemical system studied using microfluidics and the Arrhenius equation

To target development of bioelectrochemical systems, we developed an advanced microfluidic method to identify reaction bottlenecks in the metabolic activity of a pure-culture Geobacter sulfurreducens electroactive biofilm (EAB). The microfluidic system was devised to include perpendicular flow orientation for improved boundary layer uniformity and was combined with an embedded 3-electrode system to accurately apply a constant potential during the entire experimental duration. A 3-sensor temperature control system provided the basis of accurate temperature pulsing, which modified the EAB metabolic activity over short time intervals relative to the bacterial doubling rate. The system, together with the unique ability to control hydrodynamic, electrochemical, and thermal conditions, was used as the basis for an Arrhenius approach to obtain activation energy barrier values at different growth times, acetate concentrations, and flow rates. The results indicated that bottlenecks in the overall metabolic activity after 1 month of growth time were related to electron transfer through extracellular cytochrome c. After the EAB further matured to 4 months old, the bottleneck appeared to switch to enzyme-driven acetate oxidation. Based on this hypothesis, we observed after 4-months, that strong increases in effective enzyme concentration were primarily obtained by increasing flow rate, and secondarily by increasing acetate concentration.