H2 + CO2-based co-cultivation of D-lactate-producing Acetobacterium woodii and chain-elongating Clostridium drakei

Acetogen-based co-cultures represent a promising approach to integrate autotrophic carbon fixation with chain elongation. In this study, a D-lactate-producing Acetobacterium woodii strain was co-cultivated with Clostridium drakei SL1 T for butyrate and caproate production under continuous H 2 +CO 2 supply in a stirred-tank bioreactor. To anticipate co-culture performance, batch cultivations of C. drakei were conducted, revealing strong effects of H 2 availability and pH on growth, D-lactate utilization, and product formation. The A. woodii strain expressing a lactate dehydrogenase (LDHD) and pyruvate formate lyase (PFL) for D-lactate production was subsequently evaluated. Increased cultivation pressure and H 2 :CO 2 ratio did not increase D-lactate concentrations. Different co-cultivation strategies were assessed, comparing simultaneous and time-delayed inoculation at varying initial ratios with continuous gassing. During simultaneous inoculation, D-lactate consumption was delayed, whereas time-delayed inoculation of C. drakei enabled faster D-lactate conversion. Time-delayed inoculation at an initial 1:1 ratio resulted in no measurable D-lactate accumulation and rapid butyrate formation. Based on enhanced chain elongation activity observed with C. drakei mono-cultures at reduced pH, co-cultures were investigated at pH 7.0, and with a shift to pH 6.5 prior to C. drakei inoculation. Although final butyrate concentration (2.16 g L -1 ) after 96 h was higher at the constant pH configuration (pH 7.0), a configuration including a pH shift to 6.5 led to an increased caproate concentration (0.34 g L -1 ) and production rate (0.01 g L -1 h -1 ). These findings highlight the importance of carefully selecting process conditions and provide a framework for designing H 2 +CO 2 -based co-cultures for sustainable production of value-added chemicals from H 2 +CO 2 .