The efficiency of anaerobic digestion (AD) is often constrained by the kinetic bottleneck of interspecies hydrogen transfer. This study investigates multilayer graphene (MG), a conductive carbon material, as a strategy to promote direct interspecies electron transfer (DIET). Batch AD experiments were conducted with MG dosages of 0, 1, 2.5, 5, and 10 g/L. All MG-amended reactors demonstrated accelerated volatile fatty acid (VFA) consumption and higher methane yields compared to the control. However, a saturation effect was observed above 2.5 g/L, with no statistically significant increase in methane production (p > 0.05), establishing this concentration as the most resource-efficient dosage under the tested conditions. Microbial community analysis revealed a mechanistic shift: MG addition suppressed the hydrogenotrophic methanogen Methanoculleus while enriching the DIET-associated syntroph Syntrophomonas and mixotrophic Methanosarcina . These findings suggest that MG redirects metabolism from a sensitive hydrogen-based syntrophy to a more direct and resilient DIET pathway, providing a strong proof-of-concept for enhancing AD performance. • MG selectively accelerated methanogenesis, not hydrolysis or protein degradation • A 2.5 g/L dose maximized CH 4 production rate and shortened lag phase • Methane yield increased by15%, with performance saturating at a 2.5 g/L dosage • MG enriched DIET-syntroph Syntrophomonas and mixotroph Methanosarcina
Luo et al. (Sun,) studied this question.