| Fuel ethanol is a type of clean energy that can serve as a substitute for fossil fuels,thereby reducing our reliance on non-renewable resources.In China,biomass fuel ethanol is mainly produced from raw materials such as corn,aged grains,and cassava.However,when producing fuel ethanol from cassava,every ton of ethanol produced generates 8-12 tons of wastewater.Traditional methods of treating ethanol wastewater,such as solid-liquid separation and anaerobic digestion cannot meet emission standards.Currently,many companies utilize the sequencing batch reactor activated sludge process（SBR）to further treat ethanol wastewater,allowing for the discharge of SBR effluent into sewage networks after meeting quality standards.One of the pressing issues faced by the fuel ethanol industry is the comprehensive utilization of wastewater,as it presents a significant challenge in terms of both cost and environmental impact.To address this issue,this study proposes the recycling of SBR effluent in the process of ethanol production.The inhibitory components of the wastewater on ethanol fermentation were identified,and the inhibitory mechanism was analyzed.In order to achieve resource reuse,ozone oxidation was used to remove the inhibitory components of the SBR effluent.The main research findings are presented below.（1）The results showed that the direct use of SBR effluent for ethanol fermentation resulted in severe inhibition.Specifically,it was found that the SBR effluent caused a reduction of 46.2%in yeast cell after 24 hours of fermentation.The concentration of residual sugars was also found to be 52.7%higher than the control group,while the ethanol production decreased by 54.8%.The observed inhibition of yeast cells could be attributed to the presence of inhibitory compounds in the SBR effluent.These compounds may have induced stress response in yeast cells,leading to the activation of metabolic pathways involving trehalose and glycerol to counteract the environmental stress.These findings suggest that direct reuse of SBR effluent can impair yeast viability and growth,thereby inhibiting ethanol fermentation.（2）The water quality analysis of the SBR effluent revealed that the concentration of NO2-ions reached 104.83 mg·L-1,which had a strong inhibitory effect on fermentation.Chemical oxygen demand（COD）is a measure Indicators of organic pollutants in water,The COD of the SBR effluent was only 794.8 mg·L-1,and the conductivity was 6.36 ms·cm-1,indicating that the concentration of organic component was relatively low,but there was a high concentration of inorganic components.The SBR effluent was evaporated and divided into two parts:the evaporative condensate（mainly composed of volatile organic components）and the evaporative residue（mainly composed of inorganic and high-boiling-point organic components）.After reconstituting to the original volume,they were separately used in the ethanol fermentation process.The results showed that the evaporative condensate had no significant effect on fermentation,while the evaporative residue strongly inhibited ethanol fermentation.Through the determination of the inorganic ion content in the SBR effluent and the external addition experiment,it was confirmed that the NO2-ion content in SBR effluent reached 104.83 mg·L-1that inhibited yeast growth and ethanol fermentation,and thus was identified as the main inhibitory component in ethanol fermentation.（3）In order to remove inhibitory components,the ozone oxidation was used to treat SBR effluent.After 20 min of ozone oxidation treatment,the inhibitory effect of SBR effluent on ethanol fermentation was completely eliminated.Analysis of the inorganic ion content in SBR effluent showed that most inorganic ions did not change significantly after 20 min of ozone oxidation treatment,except for NO2-ions,which were effectively removed.Indicating that NO2-ion is the inhibitory component in SBR effluent,and ozone oxidation can detoxify SBR effluent.The factors affecting the removal efficiency of NO2-ions were investigated,and it was found that temperature and p H had no significant effect on the oxidation efficiency,but the oxidation efficiency was significantly enhanced with increasing ozone flow rate.（4）In order to investigate the toxicity mechanism of NO2-ion on ethanol fermentation,the physiological mechanism of yeast under NO2-ion stress was elucidated at the gene transcription level.The thesis found that NO2-ion had an inhibitory effect on ethanol fermentation at a concentration of 5 mg·L-1,reducing the yeast cell count by 31.6%and the final ethanol production by 1.7%after 24 h of fermentation.As the NO2-ion concentration increased,the inhibitory effect was significantly enhanced,and the death rate continued to increase with the increase of NO2-ion concentration.Transcriptomics results showed that under NO2-ion stress,some DEGs in pentose phosphate pathway（PPP）and tricarboxylic acid（TCA）cycle were downregulated.This suggests that the PPP and TCA cycle were inhibited after NO2-ion addition,thus affecting the growth of yeast and ethanol fermentation. |
