| Summary: | Abstract Background Commercial biogas upgrading facilities are expensive and consume energy. Biological biogas upgrading may serve as a low-cost approach because it can be easily integrated with existing facilities at biogas plants. The microbial communities found in anaerobic digesters typically contain hydrogenotrophic methanogens, which can use hydrogen (H2) as a reducing agent for conversion of carbon dioxide (CO2) into methane (CH4). Thus, biological biogas upgrading through the exogenous addition of H2 into biogas digesters for the conversion of CO2 into CH4 can increase CH4 yield and lower CO2 emission. Results The addition of 4 mol of H2 per mol of CO2 was optimal for batch biogas reactors and increased the CH4 content of the biogas from 67 to 94%. The CO2 content of the biogas was reduced from 33 to 3% and the average residual H2 content was 3%. At molar H2:CO2 ratios > 4:1, all CO2 was converted into CH4, but the pH increased above 8 due to depletion of CO2, which negatively influenced the process stability. Additionally, high residual H2 content in these reactors was unfavourable, causing volatile fatty acid accumulation and reduced CH4 yields. The reactor microbial communities shifted in composition over time, which corresponded to changes in the reactor variables. Numerous taxa responded to the H2 inputs, and in particular the hydrogenotrophic methanogen Methanobacterium increased in abundance with addition of H2. In addition, the apparent rapid response of hydrogenotrophic methanogens to intermittent H2 feeding indicates the suitability of biological methanation for variable H2 inputs, aligning well with fluctuations in renewable electricity production that may be used to produce H2. Conclusions Our research demonstrates that the H2:CO2 ratio has a significant effect on reactor performance during in situ biological methanation. Consequently, the H2:CO2 molar ratio should be kept at 4:1 to avoid process instability. A shift toward hydrogenotrophic methanogenesis was indicated by an increase in the abundance of the obligate hydrogenotrophic methanogen Methanobacterium.
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