| In this paper, based on the theory of fermentation process of metabolic control, the high yield strain of Glutamic acid was screened, then, the further research about optimization of fermentation conditions and production engineering was carried out, so that we raise the best program. In order to confirm the biotin content, the method of fluorescence spectrometry was utilized.Firstly, we choose the Corynebacterium glutamicum W3 as the starting strain, the reverse mutant W3-2-77 of Glutamic acid auxotroph, compared with the starting strain which enhanced the yield of Glutamic acid, was obtained after UV and NaNO2 mutagenesis. Secondly, the strain W3-2-77 treated with continuous UV, and we obtain the mutant strain W3-4-12 of Asp leakage and Gln resistance. The strain W3-2-77 had removed the feedback regulation in metabolic pathway, as a result, and the Glutamic acid yield got to 78 g/L, which raised 21% compared with the starting strain. Finally, we proved the stability of genetic traits for high-yield mutant after serial passage.The optimization of fermentation condition for the mutant W3-4-12 showed additive amount of urea had a apparent effect to the fermentation in seed medium, exactly speaking, we could get the biggest acid production, when the fermentation condition was in accordance with the follow principle that proportion of the urea was 0.6 %, the seed age was 11 h, 1st inoculating quantity was 25 mL/250 mL. In order to make sure the variety and the proportion of carbon source, nitrogen source, mineral salt, we made use of the method of several rounds of fermentation to compare Glutamic acid yield, and identify the best material in culture medium. Then, after the analysis of single factor experiment in medium, we gave the proper proportion of all kinds of single material. Finally, according to choosing four significant factors and designing orthogonal test of L9(34) to analyze, we obtained the best amount of glucose, urea, KCl, Na2HPO4. In addition, under different temperature and pH, the paper provided a concrete culture condition of fermentation stage to increase the Glutamic acid, and introduced the sub-flask concept that the fermentation was differentiated into two stages, that was"the growth term"and"production term", to boost the yield of the Glutamic acid. The early period condition of fermentation was 30 mL/500 mL for 15 h, and the late (latter) was 15 mL/500 mL for 38 h, which led to the Glutamic acid yield about 112 g/L and the convert ratio of glucose got to 62 %. Compared with the former fermentation, the yield of Glutamic acid heightened 44 %.We also took advantage of the way of combining 2nd inoculating with fed-batch fermentation to increase the Glutamic acid yield. According to the 2nd inoculating, we not only identified the inoculating quantity and seed age, but applied the 2nd-stage method of expansion of the seed culture fluid to increase the seed concentration, which overcame the weak point of the longer acid cycle and reduced the fermentation cycle to 34~36 h. At last, linking with the fed-batch fermentation, we supplied a complete fermentation process and the yield of Glutamic acid got to 117 g/L.After finishing fermentation, we detected the biotin amount in the fresh culture medium, the final fermentation fluid and the material. Then, the stable method of fluorescence spectrometry to detect the biotin amount was established. Besides, we proved the biotin amount was at"super sub-moderate"range in optimized medium and concluded formula of additive amount of corn syrup and molasses under different batches.
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