Biohydrogen production via two-stage lactate-driven dark fermentation: A long-term operational performance study

Lactate-driven dark fermentation (LD-DF) can enhance hydrogen recovery during mixed-culture fermentation, but the long-term performance of continuous two-stage configurations remains underexplored. This study evaluated a two-stage LD-DF system treating synthetic food waste over 180 days of continuous operation. The system demonstrated stable H 2 production rates, ranging from 2.02 to 2.65 NL H 2 /L-d, and high resilience following a major process disturbance. The lactate-producing reactor accumulated up to 8.15 g/L of lactate but removed only 29% of the supplied carbohydrates, which achieved a high overall conversion rate. 16S rRNA gene analysis showed that the lactate-producing reactor was dominated by Lactobacillus , Fructilactobacillus , and Bacillus , whereas Prevotella and Clostridium were initially abundant in the hydrogen-producing reactor. Overall, this study shows that two-stage LD-DF can support long-term hydrogen production from organic residues but improving carbohydrate-to-lactate conversion in the first stage and preserving metabolic and ecological differentiation between reactors will be important for enhancing process performance.