The Study On Optimization Of L-tryptophan Production By Escherichia Coli With Combination Of Nitrogen Source And Oxygen Uptake Rate Regulation

L-tryptophan is the second essential amino acid among the eight essential amino acids for humans and animals.The strain of recombinant Escherichia coli was used in this study.The following studies were carried out on the optimization and scale-up of the fermentation process.Firstly,a preliminary study on the basic fermentation medium was developped.On the basic optimization and screening of single factor,a series of statistical experiment design methods were used to obtain the combination of optimized fermentation medium.The best cultivation time of seeds was also studied.The yield of tryptophan reached 1451.30 mg/L,which was 94.5%higher than that of the original fermentation medium in shake flask.And the fermentation results of the optimized medium in the 5 L tank showed that the fermentation lag period was longer than that of the control.However,the yield of tryptophan was increased by 17.3%compared with the control group by using the optimized medium at the end of the fermentation process.The conversion rate of glucose was also 19.7%higher than that of the control group.Secondly,the regulation of nitrogen sources in fermentation process was studied.The shortage of ammonium ion and the limitation supply of oxygen may be the main reasons for the decrease of tryptophan specific synthesis rate during fermentation process.The effects of different nitrogen feeding modes on tryptophan fermentation by recombinant Escherichia coli were investigated.The results showed that a certain amount of ammonium sulfate supplementation could promote the growth of bacteria,but the increase of conductivity caused by excessive sulfate ion would seriously affect the activity of bacteria and the synthesis of tryptophan.The supplementation of yeast powder could maintain the viability of bacteria,but it was not conducive to the increase of tryptophan synthesis rate.The process results of feeding ammonium acetate and corn syrup showed that this method could shorten the lag phase of bacterial.Compared to the control,tryptophan production was increased by 35%and the conversion rate of glucose was increased by 27%with this method.The oxygen uptake rate(OUR)is a macroscopic metabolic character parameter that can be detected in real time,which can provide effective information for process scale-up and optimization.The effects of six different OUR control modes on fermentation were investigated by multi-parameter analysis.The results showed that in the 5 L tank fermentation,the OUR was slowly increased to 100 mmol/L/h before 24 h,and maintained at this level after 24 h was more conducive to the synthesis of tryptophan.