Butanol is a promising biofuel due to its high energy density, compatibility with existing fuel infrastructure, and efficient conversion into sustainable aviation fuel. Butanol can be produced via acetone-butanol-ethanol (ABE) fermentation; however, its efficiency is limited by carbon loss in the form of CO₂ and H₂ and microbial inhibition from fermentation byproducts. To improve bioproduct yields and maximize carbon utilization from lignocellulosic biomass, this study investigates a co-culture fermentation strategy integrating Clostridium beijerinckii (Cb), a sugar-fermenting microorganism, with C. carboxidivorans (Cc), a gas-fermenting species. This approach enables simultaneous utilization of sugars and fermentation-produced gases, enhancing solvent production while reducing CO₂ emissions. Switchgrass hydrolysates (SH), a common lignocellulosic feedstock, contain microbial inhibitors that impact fermentation efficiency. To assess microbial tolerance and productivity under realistic conditions, monoculture experiments were conducted in buffered P2 and P11 media with glucose concentrations ranging from 20-60 g/L, gas ratios (H₂:CO₂ of 3:1 and 0.67:1), and temperatures of 28°C and 37°C in 280 mL bottle bioreactors. Results demonstrated that Cc efficiently consumed CO₂ at 28°C in P11 medium under all conditions, while Cb exhibited stable performance across media and buffer systems at 37°C. Based on these findings, co-culture fermentations were performed in P11-acetate buffered medium with 20 g/L glucose at 28°C and 37°C. Preliminary results showed that co-culture fermentation significantly reduced CO₂ emissions, utilizing 16% CO₂ and 37% H₂ at 37°C, while producing 4-fold higher acid yields compared to Cb monoculture. Similarly, at 28°C, the co-culture consumed 19% CO₂ and 54% H₂, resulting in 4.5-fold higher total acid production. To further align with sustainable biomass conversion goals, ongoing experiments focus on optimizing co-culture fermentation with detoxified and non-detoxified switchgrass hydrolysates. Additionally, the role of CO and H₂ supplementation in enhancing alcohol production and further reducing CO₂ emissions is under investigation. These findings highlight the potential of co-culture biomanufacturing as a scalable and sustainable strategy for improving bioproduct formation, reducing carbon loss, and increasing lignocellulosic biomass utilization efficiency in biorefinery processes.
