Ethanol production from non-sterile methane fermentation residues for waste-to-biofuel conversion

Effective valorization of food waste is essential for sustainable resource management. Methane fermentation residues, a byproduct of biogas production from food waste, are typically used as liquid fertilizer. However, their full potential as bioresource substrates remains underexplored. This study investigated the feasibility of producing bioethanol from non-sterile methane fermentation residues mixed with beverage waste under realistic, low-input conditions. Four sucrose-fermenting yeast strains were evaluated for ethanol production in a non-sterile medium composed of residue and deteriorated beverage (1:1, v/v). Ethanol was detected only under the non-sterile condition, not in filter-sterilized controls, suggesting that native microbial populations did not inhibit—and may have enhanced—fermentation. Saccharomyces pastorianus NBRC 11024^T achieved the highest ethanol yield (27.4 g/L), reaching 89.4 % of the theoretical maximum within 48 h. Fermentation efficiency declined when the residue ratio exceeded 70 %, indicating possible inhibition due to nitrogen or volatile fatty acid accumulation. 16S rRNA amplicon sequencing revealed a marked shift in microbial community structure, with Leuconostoc species dominating at later stages. These findings suggest that methane fermentation residues can be used as functional co-substrates for ethanol production and may actively contribute to favorable fermentation conditions. In addition, commercially available beverages were shown to be a suitable model for waste beverages and provided practical insights into real-world applications. This approach offers a scalable, decentralized strategy for low-cost ethanol production and contributes to the development of circular bioresource systems.