| γ-polyglutamic acid(y-PGA)is a multi-purpose,biodegradable biopolymer material,which is connected by repeating units of D-glutamic acid and L-glutamic acid or both through amide bonds Yes,it has broad development space in many industries such as environmental protection,agriculture,medicine,cosmetics and food.Currently,y-PGA is mainly obtained through fermentation production,and there are only a few companies in China that carry out small-scale production.In this paper,Bacillus licheniformis(strain number),which currently produces y-PGA strains in the laboratory,is used as the research object.The amount and proportion of yeast paste and peptone are optimized in a shaker and 5 L fermentation tank,and in a 200L tank,50T A scale-up production verification experiment was carried out on the tank.Finally,a metabolomics platform was established by GC-MS and protein profiling to analyze the metabolic differences after adding glycerol.The main experimental results of this paper are as follows:(1)Through the single factor experiment method,the yeast paste and peptone were optimized and screened,and finally the yeast paste SH3 and peptone SH were determined as the substitute objects.L,the concentration of peptone SH bovine bone is 4g/L,and at this addition ratio,the yield of polyglutamic acid reaches 36.7g/L.(2)On the basis of the above,the optimal concentration of K2SO4,glycerol and glucose was optimized by single factor experiment,and finally the concentration of K2SO4,glycerol and glucose was determined to be 15g/L,6%and 90g/L,respectively.(3)The response surface experiment was carried out by shaker fermentation,and the yield of y-PGA was used as the response value,and the fermentation medium was optimized by performing 4 factors and 3 levels experiment.After optimization,the shake flask output of y-PGA has been greatly improved,and the yield of y-PGA reached 39.1g/L in 72 hours.(4)The effects of three rotation speeds of 300 r/min,400 r/min and 500r/min on the fermentation process of 5 L fermentation tank were studied respectively.Finally,the stirring speed of 400 rpm was selected,and the ventilation rate was controlled by 1vvm.It was verified with a 200 L fermentor,and the final y-PGA output reached 35.6 g/L.(5)This paper explores the metabolic process changes of Bacillus licheniformis after adding glycerin through the mutual verification of GC-MS and protein profiles,and determines the accumulation of stearic acid,maleic acid,and ribic acid by the GC-MS analysis platform Compared with the control group,it has been greatly improved.The key enzymes in the protein profile analysis have determined that the glycolysis pathway and the biosynthesis and degradation of alanine have changed to varying degrees,promoting the TCA cycle and synthesizing polyglutamic acid.Provides more energy and premise material.These conclusions can provide a basis for further strain improvement and process optimization in the future.(6)Ten batches of industrial production scale-up experiments were carried out on a 50m3 fermentation tank,and the results showed that the production of y-PGA after fermentation and addition of glycerol increased by about 50%compared to before,reaching 41.4g/L. |
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