Process Intensification of ABE Fermentation Through an Oxygen-Scavenging Microbial Co-Culture System

The development of scalable and economically viable biobutanol production remains challenging because conventional acetone–butanol–ethanol (ABE) fermentation depends on strict anaerobic conditions for Clostridium strains. Traditional anaerobiosis control requires continuous nitrogen sparging and chemical reducing agents, increasing operational costs and process complexity. This study proposes a microbial co-culture of Bacillus subtilis and Clostridium beijerinckii BA101 as an oxygen-scavenging strategy to simplify reactor operation and improve scalability. Growth kinetics were first evaluated in monoculture to define compatible operating conditions. Co-culture fermentations were then optimized by varying inoculation ratio, inoculation timing, and inoculum concentration. The best performance was achieved by inoculating C. beijerinckii 4 h after B. subtilis at a 30:70 inoculum ratio, producing 13.14 g L−1 of butanol, 26.6% higher than monoculture. This strategy was subsequently validated in a stirred tank bioreactor under pH-controlled conditions. B. subtilis depleted dissolved oxygen within 53 min without external gas sparging, establishing anaerobic conditions in situ. At pH 5.9, the co-culture reached 18.6 g L−1 butanol, a productivity of 0.194 g L−1 h−1, and 81.6% glucose conversion. These results demonstrate that microbial co-culture is an effective process-intensification strategy, simplifying anaerobic operation while enhancing fermentation performance and industrial scale-up potential.