Production Of L-tryptophan By Fermentation Of Escherichia Coli JN8

L-tryptophan is one of the limited essential amino acids, and it plays a vital role in themetabolism during growth and development of humans and animals. L-tryptophan is involvedin the synthesis of a variety of key biological active substances. It has important applicationvalues in many fields, such as pharmaceutical, food, animal feed, sewage disposal, etc.Highyields and high purity L-tryptophan can be obtained by direct microbial fermentation usingcheap raw materials simultaneously. Therefore, the direct microbial fermentation has potentialapplication value in the industrial production of L-tryptophan.In this thesis, the recombinant Escherichia coli JN8was used as the study object. Themain contents include medium components, fermentation conditions in shake-flask, feedingmethods in5L fermentor. The main results are as follows:The medium components for L-tryptophan production by E. coli JN8was optimized withsingle factor experiment and orthogonal design in6aspects. The optimum fermentationmedium consisted: glucose30g·L-1, soy peptone17.5g·L-1, NH4H2PO411.5g·L-1, KH2PO46g·L-1, MgSO4·7H2O2g·L-1, FeSO4·7H2O0.25g·L-1, Folic acid0.1mg·L-1, Biotin0.12mg·L-1, VB60.8mg·L-1, Tween-200.8g·L-1.The production of L-tryptophan reached3.33g·L-1, with2.8folds higher than that before optimization (1.20g·L-1).The optimum fermentation conditions were obtained through single factor experiment:optimum fermentation temperature is35℃, the optimum initial pH is7.0~7.2, the bestmedium volume was25mL/250mL, the optimum inoculation amount is10%(v/v). Theproduction of L-tryptophan reached3.59g·L-1, with3folds higher than that beforeoptimization (1.20g·L-1).It was found that the best time for feeding is about14h by comparing the concentrationof cell, the yield of L-tryptophan, the concentration of glucose and the accumulation ofmetabolic by-product acetic acid during the whole fermentation process of5L fermentorresearch.Finally, the fed-batch modes of5L fermentor producing L-tryptophan were studied. Thefeeding modes mainly include constant speed feeding, intermittent feeding, DO-Stat feedingand pH-Stat feeding. The main parameters including the maximum biomass, the maximumamount of L-tryptophan production, the conversion ratio of glucose and the concentration ofacetic acid in the fermentation broth were compared to determine the best feeding method.The optimum fed-batch mode was DO-Stat feeding. In this method, the production ofL-tryptophan reached24.62g·L-1, the conversion ratio of glucose30.15%higher than batchfermentation.