Synergistic parameter optimization Unlocks High-Efficiency H2/CO2 Bio-methanation: Decoupling mass transfer enhancement from microbial Trade-offs.

The advancement of CO2 biomethanation represents a crucial pathway for renewable energy storage and carbon neutrality. However, its widespread application is often constrained by inefficient gas-liquid mass transfer of H2 and suboptimal operational parameters. This study investigates the synergistic effects and underlying mechanisms of three key engineering parameters-agitation intensity, gas recirculation rate, and H2/CO2 feed ratio-on the performance of an in-situ H2/CO2 biomethanation process at mesophilic temperature (37 ± 1℃). Enhanced agitation (up to 160 rpm) and increased gas recirculation (up to 1200 mL·min-1) significantly improved (p＜0.05) the volumetric mass transfer coefficient (kla) for H2, thereby boosting the maximum volumetric methane production (VMP) to 0.97 L·L-1·d-1. Optimizing the H2/CO2 feed ratio to 5:1 strengthened the thermodynamic driving force for hydrogenotrophic methanogenesis, increasing methane content by ∼15% compared to the stoichiometric 4:1 ratio. High-throughput sequencing revealed that intensified mass transfer selectively enriched dominant hydrogenotrophic methanogenic archaea phylum (e.g., Methanobacteriota reached 75.2% at an H2/CO2 feed ratio of 5:1), while suppressing hydrolytic and acidogenic bacterial phylum (e.g., Bacteroidetes decreased from 23.7% to 6.8%, under intensified agitation), indicating a functional trade-off between enhanced methanogenic efficiency and complex organic degradation. This work emphasized the long-term, integrated evaluation of hydrodynamics and microbial ecology of the biomethanation systems, leading to superior methane yield and system stability. The findings provide critical insights and practical guidance for scaling up efficient and stable biomethanation systems, addressing a key bottleneck in industrial Power-to-Gas applications.