Analysis of the key enzymes of butyric and acetic acid fermentation in biogas reactors

This study aimed at the investigation of the mechanisms of acidogenesis, which is a key process during anaerobic digestion. To expose possible bottlenecks, specific activities of the key enzymes of acidification, such as acetate kinase (Ack, 0.23–0.99 U mg⁻¹ protein), butyrate kinase (Buk, < 0.03 U mg⁻¹ protein) and butyryl-CoA:acetate-CoA transferase (But, 3.24–7.64 U mg⁻¹ protein), were determined in cell free extracts of biogas reactor content from three different biogas reactors. Furthermore, the detection of Ack was successful via Western blot analysis. Quantification of corresponding functional genes encoding Buk (buk) and But (but) was not feasible, although an amplification was possible. Thus, phylogenetic trees were constructed based on respective gene fragments. Four new clades of possible butyrate-producing bacteria were postulated, as well as bacteria of the genera Roseburia or Clostridium identified. The low Buk activity was in contrast to the high specific But activity in the analysed samples. Butyrate formation via Buk activity does barely occur in the investigated biogas reactor. Specific enzyme activities (Ack, Buk and But) in samples drawn from three different biogas reactors correlated with ammonia and ammonium concentrations (NH₃ and NH₄⁺-N), and a negative dependency can be postulated. Thus, high concentrations of NH₃ and NH₄⁺-N may lead to a bottleneck in acidogenesis due to decreased specific acidogenic enzyme activities.