Mechanistic Insights and Emerging Hybrid Strategies of Magnetite Nanoparticles for Enhanced Dark Fermentative Biohydrogen Production

Dark fermentative biohydrogen (bio-H2) production is a promising renewable energy technology for converting organic waste and wastewater into clean fuel. However, practical application is limited by low H2 yield, volatile fatty acid (VFA) accumulation, incomplete substrate utilization, electron diversion to competing pathways, prolonged lag phases, and inhibitory byproducts from lignocellulosic pretreatment. Magnetite nanoparticles (Fe3O4 NPs) have attracted attention as redox-active additives because of their electrical conductivity, reversible Fe2+/Fe3+ cycling, magnetic recoverability, biocompatibility, and microbial interaction potential. This review examines the physicochemical properties of Fe3O4 NPs and their proposed roles in dark fermentative bio-H2 production. Particular emphasis is placed on Fe3O4-mediated extracellular electron transfer (EET) in fermentative communities, which differs from direct interspecies electron transfer (DIET) in methanogenic systems. Fe3O4 NPs may enhance bio-H2 production by facilitating electron transfer, supporting hydrogenase activity, regulating redox balance, promoting acetate- and butyrate-type pathways, and enriching H2-producing bacteria such as Clostridium spp. Hybrid systems combining Fe3O4 with biochar, activated carbon, reduced graphene oxide, bimetallic nanocomposites, or immobilization matrices may further improve microbial retention and process stability. Remaining challenges include aggregation, dosage-dependent toxicity, recovery, environmental fate, mechanistic uncertainty, and scale-up feasibility.