Optimization of hydraulic retention time in continuous orange peel crude enzyme - mediated dark fermentation for sustainable biohydrogen production from synthetic swine wastewater

Orange peel crude enzyme (OPCE) effectively improves hydrogen production through dark fermentation by enhancing hydrogen generation processes. However, to facilitate the application of OPCE-mediated dark fermentation, an appropriate hydraulic retention time (HRT) should be explored. This study uses synthetic swine wastewater to compared hydrogen production with and without OPCE addition under three HRTs (18 h, 24 h, and 36 h). Results showed that the highest hydrogen yield (around 205 mL/g VS) and production rate (681.02 ± 22.72 mL H₂/L-d) were achieved at 24 h HRT in the OPCE group. Extending HRT in the control group facilitated hydrogen accumulation through butyrate-type fermentation, while OPCE addition enhanced hydrogen production through acetate-type fermentation. The addition of OPCE improved hydrogen yield by approximately 42.53 %, 26.34 %, and 14.86 % at 18 h, 24 h, and 36 h HRT, respectively. The more significant improvement was attributed to OPCE suppressing ethanol metabolism that were more likely to occur at short HRTs and enhancing substrate-microbial contact efficiency at both 18 h and 24 h HRT. Additionally, OPCE increased microbial diversity and abundance under the same HRT conditions, with Firmicutes , Actinobacteria , and Proteobacteria as the dominant phyla. • Adding orange peel crude enzyme (OPCE) enhanced H 2 yield under the same HRT. • The highest H 2 production was achieved at 24 h HRT with the addition of OPCE. • OPCE addition enhanced acetate-type fermentation but inhibited ethanol metabolism. • OPCE enhanced substrate-microbial interaction at both 18 h and 24 h HRT. • Dosing with OPCE improved microbial diversity and abundance under the same HRT.