Valorization of spent fermentation broth for enhanced photo-fermentative hydrogen production: Biochemical responses and preliminary economic and environmental screening

Recycling spent fermentation broth (FB) can reduce fresh nutrient demand in photo-fermentative hydrogen production (PFHP) while enabling resource recovery within circular biorefinery. Processed FB replaced 0-100% of inoculum cultivation medium for Rhodopseudomonas palustris in corn stover-derived PFHP. FB3 (75% FB) produced 533.71 ± 20.56 mL H 2 and a maximum 12 h rate of 29.88 ± 1.15 mL h −1 , 50.2% and 126.9% above FB0. Conversely, FB2 retained 5.55 ± 0.21 g L −1 reducing sugar and 1.89 ± 0.07 g L −1 total VFA at 72 h but produced only 2.65 ± 0.10 mL H 2 . FB3 combined substrate depletion, VFA turnover, high nitrogenase activity, and NADH/NAD + ratio, whereas FB2 showed higher ATP/ADP despite negligible H 2 . Gate-to-gate screening showed FB3 reduced fresh ICM demand by 83.4%, screening cost by 81.3%, and electricity-related GWP and water-use indicators by 33.4% versus FB0, identifying 75% FB as the best-performing substitution level and supporting circular PFHP recycling.