Extracellular Polymeric Substances from Geobacter sulfurreducens Biofilms in Microbial Fuel Cells

Bioelectrochemical systems (BESs) are hybrid systems using electroactive bacteria and solid electrodes, which serve as electron donor or acceptor for microorganisms. When forming a biofilm on the electrode, bacteria secrete extracellular polymeric substances (EPSs). However, EPS excretion of electroactive biofilms in BES has been rarely studied so far. Consequently, the aim of this study is to develop a routine including the electrochemical cultivation, biofilm harvesting, fractionation, and biochemical analysis of the EPS secreted by Geobacter sulfurreducens under electroactive conditions. G. sulfurreducens was cultivated in microbial fuel cell mode on graphite-based electrodes polarized to +400 mV versus Ag/AgCl for 8 d. A maximum current density of 172 ± 29 μA cm^-2 was reached after 7 d. The EPS secreted from the biofilms were harvested and fractioned into soluble, loosely bound, and tightly bound EPS and biochemically analyzed. Electroactive cultures secreted significantly more EPSs compared to cells grown under standard heterotrophic conditions (fumarate respiration). With 116 pg per cell, the highest amount of EPSs was measured for the soluble EPS fraction of G. sulfurreducens using anodic respiration, followed by the tightly bound (18 pg cell^-1) and loosely bound (11 pg cell^-1) fractions of the EPS. Proteins were found to dominate all EPS fractions of the biofilms grown under electrochemical conditions. To the best of the authors' knowledge, these experiments are the first approach toward a complete analysis of the main EPS components of G. sulfurreducens under anode-respiring conditions.