Study On Fermentation Conditions For Producing N-Butanol From Clostridium Acetobutylicum

Gradual depletion of non-renewable energy resources and the increasingly serious environmental pollution had been two serious issues which had direct impact on sustainable development of human being. So it is necessary for us to find alternative energy source that are environmentally friendly and renewable. Biobutanol, the principal product of ABE fermentation, is a kind of potential biofuel which has been gradually paid attention to by men for its unique advantage. This paper focuses on the disadvantage of traditional ABE fermentation and increases the yields of ABE, the key enzymes of the butanol formation pathway and the conditions for producing ABE from a good strain of Clostridium acetobutylicum in order to provide useful reference for the industrial production of ABE fermentation.This study made a preliminary choose by heat shocking from the original strain of Clostridium acetobutylicum,what we got from this study was a good strain which named S5 The identifications of cell shape and the physiological characteristics on the strain of S5 were done.The yield of acetone, butanol and starch utilization rate was 1.76g/L,3.52 g/L and 70.2%, increased by 7.32%and 4.0%than the original strain of S1, respectively.A NADH dependent butanol dehydrogenase which is one of the key enzymes of the butanol formation pathway in Clostridium acetobutylicum has been separated and purified by the methods of ammonium sulfate precipitation with saturation 20%to 60%and Sephadex G-75 chromatograph. The results showed that the enzyme activity reached 256.7U/mg, and it has been purified 12.7-fold, with a yield of 28.9%. The optimal temperature for the enzyme was 35℃and optimal pH was 6.0, respectively.On the basis of single factor tests, Plackett-Burman design were adopted to filter crucial factors from liquid volume, temperature, (NH4)SO4,ZnSO4·7H2O, FeSO4·7H2O, MgSO4·7H2O, Tween 80, CaCO3, Na2S2O3 during ABE fermentation. Then temperature, CaCO3 and (NH4)2SO4 were filtered as the three crucial factors as a result. Then the Response Surface Analysis (RSA) was applied to enhance the production of ABE fermentation. Under a extreme value condition, the predicted value of acetone is 3.24481 g/L and its experimental value is 3.06g/L. The yield of acetone and butanol increased from 1.76 g/L to 3.06 g/L and 3.52 g/L to 6.12 g/L respectively, was increased by 73.8%. It indicated RSA can well explain the relationship between ABE fermentation and effect factors, and it is feasible to optimize the ABE fermentation. The optimal conditions for ABE fermentation were also confirmed as follows:inoculums size 5%, inoculums age 18h, liquid volume 100ml, corn concentration 7%, CaCO3 0.55%, (NH4)2SO40.27%, the initial pH 7.0, and cultivation temperature 35.5℃.