Mechanism Of Nitrogen Sources Regulating Butanol Production By Clostridium Acetobutylicum

Butanol is an important commodity chemical widely used in various fields.Biobutanol is a product of Clostridium Acetobutylicum that uses starch as material to be fermented by ABE.At present,cheaper carbon sources were found and used to reduce production costs at home and abroad.While,there is less research on nitrogen sources.Therefore,it was used as the basis to optimize the fermentation medium in order to reduce the production cost in this study.At the same time,it was used to regulate the mechanism of ABE fermentation and theoretical basis for the effect of nitrogen sources on butanol production was established.The Clostridum acetobutylicum was used as to investigate the mechanism of nitrogen sources regulating butanol production.To the end,the fermentation medium was optimized through single factor experiment and response surface experiment design.As a result:cassava 80 g/L,soyabeal 10.89 g/L,CaCO3 1.81 g/L,KH2PO4-K2HPO4(1:1)0.5 g/L,ZnSO4 0.34 g/L,.Under the optimum medium,the yields of butanol and total solvents reached 15.69 g/L and 26.32 g/L,which was 52.03%and 55.73%higher than that of control.In the previous,it was found that soyabeal can significantly increase the solvent yield,while ammonium acetate can effectively shorten the fermentation time.Therefore,it was explored whether the addition of soyabeal-ammonium acetate can further improve the productivity of ABE fermentation.However,the addition of complex nitrogen source inhibited butanol production by about 30%,whether it was in cassava or glucose.Through further fermentation experiments,NH4+ was found to be the main inhibitor.NH4+ reduced the yield of butanol and total solvent by 90.28%and 90.81%respectively,while the total solvent yield/glucose utilization was only 0.090 g/g,which severely affected the yield and productivity.To investigate the mechanism of NH4+ in ABE fermentation with the presence of organic nitrogen,we fully analyzed the inhibition mechanism of NH4+ from the microenvironment(NADH/NAD+,pH),enzymatic level and transcription level.Ammonium acetate can reduced the fermentation time as sole nitrogen source.While,in the presence of organic nitrogen,NH4+ will inhibit the use of NADH,the activity of butyrate acetoacetate CoA transferase and down-regulated expression of genes askA,buk,adc,ctfA,adhE,and bdhB,thereby inhibiting ABE fermentation.