Stillage Backset And Its Impact On Continuous Ethanol Fermentation By The Flocculating Yeast

As a renewable and clean fuel, ethanol is an alternative to petroleum-based transportation fuels, and in the meantime alleviates environmental impact caused by the over-consumption of these products. At present, over90%fuel ethanol is produced from sugar-and starch-based feedstocks by microbial fermentations, and high production cost makes the industry heavily depended on governmental subsidies. While feedstock consumption is the major cost in the fuel ethanol production, energy consumption is the second largest, which mainly comes from ethanol distillation and stillage treatment, particularly for fuel ethanol production with grain-based feedstocks in which the multi-evaporation process is employed for stillage treatment. Apparently, stillage backset is an economically competitive strategy for reducing stillage as well as saving energy consumption for stillage treatment. In this work, a novel process has been developed for ethanol fermentation by the flocculating yeast under stillage backset conditions. Due to the self-immobilization of yeast cells within the fermentor through flocculation, harmful byproducts generated with the lysis of yeast cells during ethanol distillation were significantly reduced, making more stillage backset, even without discharge and all stillage backset.Impact on the fermentation process was investigated under all stillage backset conditions. Compared to ethanol fermentation without stillage backset, inhibition in yeast growth and ethanol fermentation was observed, the average concentration of ethanol and biomass were decreased by5.3%and10.9%respectivily. As a result, less biomass was accumulated within the fermentation system, and consequently more sugars were remained unfermented, with less ethanol produced. The analysis of byproducts indicated that except succinic acid and5-(hydroxymethyl) furfural, all other ten major byproducts and four metal ions accumulated significantly under the stillage backset condition. Compared to the control without stillage backset, their average concentrations increased at least3times. Exception of Ca2+, glycerol and propionic acid (were1126mg/L,15080mg/L and2100mg/L respectivily), concentrations of all other byproducts and metal ions were far less than1g/L. Metabolic flux analysis illustrated that the stillage backset had no significant impact on ratio of carbon flux distributions in the glycolysis and pentose phosphate pathway associated with yeast metabolism, but substrate uptake rate was inhibited, and the activities of rate-limiting enzymes hexokinase,6-phosphofructose and pyruvatekinae in the glycolysis were down-regulated, making the net carbon flow to ethanol production decreased33%at the end of ethanol fermentation with stillage backset.In order to identify the main inhibitors, effect of individual byproducts on the ethanol fermentation was studied by supplementing them into the medium and feeding into the fermentation system. When the concentration of citric acid, α-ketoglutaric acid, fumaric acid, lactic acid, pyruvic acid, succinic acid,2-phenylethanol, furfural and5-hydroxymethyl furfural were lower than1g/L, experimental results indicated that the growth and ethanol fermentation inhibition rate were lower than5%and4%respectively. And the yeast growth and ethanol fermentation were facilitated when metal ions were in a low concentration. However, the growth inhibition rate were lower than20.3%when1g/L propionic acid was supplemented. Furthermore, exception of propionic acid (MIC10:0.4g/L), concentrations detected within the fermentation system of all other byproducts and metal ions were much lower than their minimum inhibition concentrations (MIC10). When a mixture of these byproducts with concentrations of their averages detected within the fermentation system under the stillage backset condition was supplemented, no synergic effect was observed, and propionic acid was still the dominated inhibitor on yeast growth and ethanol fermentation.The results of effect of propionic acid on ethanol fermentation by the flocculating yeast indicated that the significant decrease of ehanol production, key enzyme activities and yeast flocculation were observed, When the propionic acid concentration was larger than IC50(40mmol/L). and propionic acid exhibited the strongest inhibition in yeast growth and ethanol fermentation with an inhibition rate of67%and66%respectively, when supplemented with the concentration of60mmol/L. and the size of flocs closes to50μm, its performance likes the free cells, which is absolutely a bad news for the process by using flocculating yeast.However, propionic acid was produced by the dehydration of pentose sugars liberated from hemicelluloses in the feedstock under high temperature and acidic conditions associated with mash liqufication, hydrolysate sterilization and ethanol distillation, but propionic acid production was negligible during mash saccharification at60℃. and a practical strategy was developed for ethanol fermentation by the flocculating yeast under stillage backset conditions Without fermentation medium was sterilized at high temperature. The result indicated that the propionic acid accumulation within the fermentation system decreased62%, and more sugars were fermented and the reducing sugar was decreased by40.1%, with an increase of7.4%and59.3%in ethanol production and biomass respectivily.No doubt, the research progress is significant for fuel ethanol production with more stillage backset to reduce the discharge and save energy consumption for stillage treatment.