Conversion of artificial corn stalk wastewater, which was prepared according to the main composition of the corn stalk fermentation lixivium, into bio-hydrogen gas by mixed culture was performed in a 20 L half-continuous flow fermenter. The influences of several environmental factors on the bio-hydrogen production, such as HRT, C/N ratio, Fe2+ and artificial corn stalk wastewater concentration were discussed in the tests. The experimental results showed that HRT, C/N ratio, Fe2+ and artificial corn stalk wastewater concentration significantly affected the fermentation hydrogen production. The maximum H2 yield of 11.80 mol/kg, H2 concentration of 56% and hydrogen production rate of 8.81 L/(L x d) were obtained at HRT = 10 h, C/N = 100, Fe2+ concentration of 100 mg/L and substrate concentration of 12.5 g/L by mixed culture, respectively. In the fermentation hydrogen-producing process, the conversion efficiency of the substrate was more than 90%, and 39.40% of COD was removed from the reactor. The main by-products in the liquid phase were acetic acid, butyric acid, propionic acid and a little ethanol and butanol throughout this study.

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