Acidogenic gas utilization improves methane production in high-load digestion: Underlying mechanisms

High-load reactors face a significant challenge with fatty acid accumulation. Although injecting acidogenic gas (H2 and CO2) into methanogenic reactors improves biogas production, its mechanisms and stability enhancement remain unclear. Therefore, this study investigated the potential of using acidogenic gas to enhance methane production and system stability in a high substrate-loading co-digestion system with decreasing hydraulic retention time (HRT) and its regulatory role by considering biological pathways. Acidogenic gas injection effectively managed volatile fatty acid accumulation, with a concomitant increase in methane production from 0.042 to 0.066 L/L h as HRT decreased (16–8 days). However, the methane yield decreased from 611 to 460 mL/g volatile solids. Microbial community analysis revealed an increased abundance of hydrogenotrophic methanogens, along with several acidogenic and syntrophic fatty acid oxidizing consortia with decreasing HRT. The HRT10.67 sample exhibited the highest microbial species diversity and richness in the methanogenic reactor. Metabolomics analysis showed acidogenic gas reduced energy synthesis related to the tri-carboxylic acid cycle and oxidative phosphorylation while profoundly regulating the metabolism of amino acids, lipids, fatty acids, membrane transporters and inhibitory substances, thereby strengthening methane metabolism pathways. This study provides valuable insights into the roles of acidogenic gas in regulating energy metabolism to enhance methane production and offers potential opportunities for improving lipid and protein degradation. •H2 content increased from 30 to 53 % as HRT decreased from 16 to 8 days.•Organic matter removal was slightly decreased by acidogenic gas injection (AGI).•AGI improved CH4 production and yield, but the improvement decreased at lower HRT.•AGI increased abundance of hydrogenotrophic methanogens and syntrophic bacteria.•AGI triggered amino acid and lipid metabolism, reducing energy-related synthesis.