Biohydrogen production by a novel integration of dark fermentation and mixotrophic microalgae cultivation

Biohydrogen is usually produced via dark fermentation, which generates CO2 emissions and produces soluble metabolites (e.g., volatile fatty acids) with high chemical oxygen demand (COD) as the by-products, which require further treatments. In this study, mixotrophic culture of an isolated microalga (Chlorella vulgaris ESP6) was utilized to simultaneously consume CO2 and COD by-products from dark fermentation, converting them to valuable microalgae biomass. Light intensity and food to microorganism (F/M) ratio were adjusted to 150 μmol m−2 s−1 and F/M ratio, 4.5, respectively, to improve the efficiency of assimilating the soluble metabolites. The mixotrophic microalgae culture could reduce the CO2 content of dark fermentation effluent from 34% to 5% with nearly 100% consumption of soluble metabolites (mainly butyrate and acetate) in 9 days. The obtained microalgal biomass was hydrolyzed with 1.5% HCl and subsequently used as the substrate for bioH2 production with Clostridium butyricum CGS5, giving a cumulative H2 production of 1276 ml/L, a H2 production rate of 240 ml/L/h, and a H2 yield of 0.94 mol/mol sugar.