Hydrogen production from molasses wastewater was investigated in continuously-stirred tank reactor (CSTR). The CSTR was operated at an hydraulic retention time (HRT) of 6 hours and the temperature at 35°C. The highest gas production of 25.39 L/d using mixed culture fermentation, corresponding to a peak hydrogen production volume of 11.39 L/d were achieved in the CSTR reactor. While the variation of chemical oxygen demand (COD) ranged from 5500 to 8000 mg/L, the soluble end product system underwent a transition of fermentation type and the reactor gave ﬂuctuating and increasing of ORP. These experimental results demonstrate that the shock-loading and micro-anaerobic circumstance are important factors for enhancing and stabilizing H2 production.