Membrane biofilm reactor (MBfR) enabling optimal H2 delivery and supporting minimal nutrient supplementation to hydrogenotrophic biofilms

• A new MBfR design optimizing H 2 mass transfer to operate optimal biomethanation. • Long-term operation of the MBfR for 435 days under minimal nutrient provision. • High-purity methane (≈95%) was produced from H 2 /CO 2 without gas-liquid mixing. • A cell-dense biofilm developed, rich in Methanothermobacter (CH 4 yield >95%). • First multidisciplinary study of an energy-efficient, gas-fed methanogenic MBfR. The hydrogenotrophic conversion of carbon dioxide (CO 2 ) and hydrogen (H 2 ) gases to methane (CH 4 ), presents a viable alternative to fossil-based natural gas. While different bioreactors have been investigated, the key challenge in these systems is overcoming mass transfer limitations of H 2 to hydrogenotrophic archaea due to its low solubility. Ideally, optimal H 2 bioavailability is achieved at thermophilic temperatures, directly feeding the gas to the biomass. In this study, a membrane biofilm reactor (MBfR) with gas-permeable polydimethylsiloxane (PDMS) membranes was designed to support methanogenic biofilm formation and enhance CO 2 and H 2 conversion under minimal mixing conditions, in a closed-loop and without additional nutrients supplementation. Over the 437-day operation, a thick, proteins-rich biofilm developed on the membrane. High-purity methane (≈95%) was produced, and a >95% conversion efficiency to methane was achieved for CO 2 and H 2 . The MBfR sustained high methane production 397 days without additional nutrient supply, and reach the highest methane production rate of 24.3 L·m -2 ·day -1 after trace element supplementation, likely replenishing molybdenum. In both biofilm and mixed liquor, the microbial community was dominated by the hydrogenotrophic methanogen Methanothermobacter together with a new Hydrothermae spp. bacterium, likely utilizing H 2 and CO 2 for homoacetogenesis and/or fermenting detrital biomass. The stable microbial community functions ensured a complete anaerobic digestion metabolic chain, allowing for nutrients recycling and metals mobilization. The MBfR design presented in this study offers a promising strategy for scalable, energy-efficient biological methanation optimizing H 2 utilization with minimal nutrient input.