In the present study, a wastewater treatment system for the ethanol fermentation industry was developed by recycling distillery wastewater. The waste was able to be recycled for the next fermentation after being treated with bio-flocculation process. The bio-flocculation process contains three steps: screening, treatment with polyaspartic acid and filtration. When the filtrate from this process was recycled, the average ethanol production yield was very close to that in the conventional process using tap water. In contrast, the recycle of wastewater without flocculation and with chemical flocculation showed negative effects on ethanol yield as recycling was repeated. This new process was confirmed to have stable operation over ten recycles. Hazardous materials influencing distillery wastewater recycles on fermentation were also considered. It was found that the content of suspended solids (SS), volatile acid and Fe ions inhibited fermentation and resulted in a decreased ethanol yield. Bio-flocculation was shown to be an effective way to diminish the content of inhibitory compounds drastically when the waste was recirculated. (C) 2009 Elsevier B.V. All rights reserved.