The Study On Ethanol Production From Cheap Biomass By Thermoanaerobacterium Aotearoense Δldh

In the case of increasingly serious global problems, such as oil crisis, price fluctuations and greenhouse gases, biomass energy become an important part of sustainable power. Bioethanol is one of important green energy sources. However, limited plowland, population expansion and the increasingly serious world food provision calls for the development of an new generation of bioethanol to replace the traditional bioethanol which was produced using grain as the raw material. Cellulose is an important biomass resource and xylose accounted for one third in its hydrolysate, so the effective use of xylose is the key to promote a new method of producing bioethanol and successful industrial transformation of biomass resources.Thermoanaerobacterium aotearoense is a thermophilic bacteria which can use the xylose in the hydrolysate, and considered to be a potential strain which is able to turn xylose to ethanol. Our lab has got aΔldh strain of T.aotearoense, by knocking out of lactate dehydrogenase in a key branch pathway to accumulate the ethanol. This paper firstly optimized the medium composition to improve the ethanol production and then obtained the maximum amount of reducing sugar by optimazing cassava starch and cassava residue hydrolysis conditions. Finally, we researched the fermentation kinetics of the enigineering bacteria in fed-batch fermentation using the hydrolysis of carbon sources.Fermentation medium for ethanol production of the engineered bacteria T.aotearoenseΔldh was optimized using response surface methodlogy. A Plackett-Burman design was applied to screen potential key factors on the basis of single factors, and the steepest ascent method was adopted to approach the approximate optimal medium composition. Finally, central composite design and response surface methodology were applied to define the optimal composition of the fermentation medium more precisely. Our date revealed glucose, xylose and yeast extract to be the most influential parameters, and the optimized medium were: glucose 13.07 g/L, xylose 9.58 g/L, yeast extract 3.78 g/L, other components are the same with the MTC medium without Cysteine monohydrate and Ammonia pyridoxine hydrochloride. With the optimized medium, the ethanol production and cell dry weight were 6.102±0.21 g/L and 2.87±0.16 g/L respectively, as much as 1.46 and 1.53 times of that in primary medium.We explored the hydrolysis condition of cassava starch and cassava residue. In the premise of ensuring T.aotearoenseΔldh can grow in the hydrolyzate, hydrolysing for 30 min in 121℃with 0.1 mol/L of dilute HCl was set as the most optimal pretreatment conditions of cassava residue. In this pretreatment conditions the cavassa residue can product 50.2±1.5% reducing sugar. However, 98.1±2.1% reducing sugar can be produced in the condition of 0.05 mol/L dilute HCl, 121℃for 30 min.We added cassava starch hydrolysate at twice and total glucose is up to 94.0 g/L.After the fermentation for 132 h, T.aotearoenseΔldh produced 23.48 g/L ethanol when fed-batch fermentation used the cassava starch hydrolyzate as carbon source. With the consumption of the glucose 76.29 g/L from the hydrolyzate, the ethanol yield was 0.31 g/g sugar, 0.15 g/g cassava starch and the ethanol production yield rate was 0.24 g/(L·h). When cassava residue was used as carbon source, we added the hydrolyzate residue twice which contained 68.63 g/L glucose and 16.55 g/L xylose. After fermentation for 132 h, glucose consumption was 64.65 g/L and all the xylose was used up, the ethanol yield was 0.10 g/g cassava residue, ethanol yield rate was 0.18 g/(L·h). When glucose was used as sole carbon sources in fed-batch fermentation, after fermentation for 108 h ethanol was 25.43 g/L, ethanol yield was 0.31 g/g of glucose, ethanol yield rate was 0.30 g/(L·h) with glucose consumption of 81.38 g/L.When a mix sugar was used in fed-batch fermentation, glucose was comsumed 56.13 g/L and xylose 27.27 g/L, the final ethanol yield was 22.03 g/L, ethanol yield was 0.26 g/g of sugar and ethanol yield of 0.26 g/(L·h). The ethanol production when used cassava starch hydrolyzate as a carbon source was no significant decline compared with the fed-batch fermentation using the glucose as carbon source. The ethanol production was also no significant decrease when fed-batch fermentation used cassava residue as a carbon source compared fermentation with the glucose and xylose as the mixed carbon sources.