| With increasing concerns on fossil energy shortage and environmental issues,biobutanol has gradually received more and more attention.When fermented on corn with Clostridium acetobutylicum,the cost of feedstock accounts for 70%of the total production cost,which also seriously affects the safety of human food supply.Therefore,there is an urgent need to find a cheaper,high-yield non-grain feedstock as a substitute for biobutanol production.In order to find a solution to the above problem,this paper used a kind of non-food crop fern root(FR)as the feedstock in butanol fermentation for the first time.In this paper,butanol fermentation test was carried out on different non-grain feedstocks by Clostridium acetobutylicum CGMCC 1.0134.The fermentation performance of corn and fern root was compared,and a two-level full factorial design and surface response experiments were designed to optimize the butanol fermentation process when fermented on fern root.To explore the mechanism of butanol fermentation,the transcription level of key enzyme genes and the metabolic flux in the metabolic pathway were measured.The paper proposed an effective optimization method for butanol fermentation on fern root,aiming at improving fermentation yield and butanol/acetone ratio,achieving effective replacement of traditional feedstocks with non-grain fern roots,and improving the overall butanol fermentation performance.The main results were summarized as follows:(1)The butanol fermentation performance of kitchen waste,apple pomace,kitchen waste and pomace mixture and fern root starch were studied separately.The butanol concentration(total solvent concentration)in the fermentation broth was 3.9 g/L(6.9 g/L),5.5 g/L(10.2 g/L),7.4 g/L(14.5 g/L)and 6.2 g/L(10.6 g/L),respectively;butanol yield(total solvent yield)was 0.026 g/g-substrate(0.046 g/g-substrate),0.025 g/g-substrate(0.051 g/g-substrate),0.044 g/g-substrate(0.085 g/g-substrate)and 0.069 g/g-substrate(0.12 g/g-substrate),respectively;butanol/acetone ratios was 1.8,1.7,1.6 and 2.5,respectively.(2)The butanol fermentation performance fern root and corn as feedstocks was studied.The results showed that the direct butanol fermentation on fern root showed a lower producton and longer fermentation time.In this study,after adding 3.5 g·L-broth-1 yeast extract solution(YE)to the fern root medium,the fermentation performance of fern root was significantly improved,the fermentation time was shortened,and the butanol production and productivity were 8.8 g/L and 0.09 g/L/h,respectively,which were 42%and 80%higher than that in control.The butanol production and productivity were 12 g/L and 0.13 g/L/h,respectively,after optimized by two-level full factorial design and response surface experiments.(3)The concentration of free amino acids in the fermentation broth and the transcription level of key enzyme genes in the metabolic pathway of Clostridium acetobutylicum were analyzed.It was found that after adding the yeast extract solution(YE)to the fern root medium,the amino acids of the aspartic acid families and the serine families were excessively secreted,and the expression level of the ctfB gene encoding the COA transferase was 13 times than the control at 72 h.(4)The butanol fermentation conducted on FR showed a higher butanol-acetone ratio than that of the butanol fermentation fermented on corn(58.9%).The results of the metabolic analysis indicated that the reabsorption pathway of the organic acid was weakened during the solvent production period when fermented on FR,and the acetone production pathway was also weakened.This was one reason of higher butanol-acetone ratio occurred during the butanol fermentation with FR as a substrate.According to the results of metabolic flux analysis,the synthesis rate of NADH in the butanol fermentation broth using FR as the substrate was higher compared to corn.Additionally,it is speculated that the higher NADH production rate was another reason for the higher butanol-acetone ratio occurred in butanol fermentation process when fermented on FR. |
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