The optimistic impact of orthoferrite nanoparticles (NPs) on photo fermentation for biohydrogen production have been increased, and the mechanism still requires to be investigated more. Low hydrogen (H
2) yield often occurs, being mainly due to H
2 generation pathway shift. In present study, Terbium-iron oxide nanoparticles (TbFeO
3 NPs) were synthesized to investigate their impacts on bioH
2 production. The highest H
2 yield of 319.34 mL/g glucose was found at 100 mg/L of TbFeO
3, being 47.6% higher than that from the control (225.84 mL/g glucose). The relative abundance of
PNSB increased from 54.2% to 66.43%. The large specific surface area of TbFeO
3 provided appropriate sites for the colonization of
PNSB and enhanced the bacterial approach to nutrients. Additionally, the Tb
3+ gradually released from TbFeO
3 NPs raised microbial transmembrane transport capacity, promoting glycolytic efficacy and Fe availability, though improving
hydrogenase content, and shifting the bioH
2 evolution to butyrate pathway for more H
2. This provides the novelty for biochemical utilization of Tb and new insights into the improved H
2 yield amended with TbFeO
3.