| γ-aminobutyric acid (GABA) is a non-protein amino acid that is widely distributed in nature. GABA in the brain and spinal cord of mammals with a variety of effects in the central nervous system as inhibitory neurotransmitter function, GABA is a good medical drugs and raw materials for health care. GABA played a poultry heat stress and growth-promoting effect. In this paper, the E. coli (AS 1.505) which has high activity of glutamate decarboxylase as the originality strain.The strain transform L-glutamic acid as substrate to GABA. A series of scheme were used in ordre to achieve high yield, high concentration and high productivity of GABA production. The main elements of this thesis are as follows:1. Compositions of culture medium and culture conditions were studied to improve quantity of Glutamic acid decarboxylase (GAD) produced by AS 1.505 liquid culture. The optimized medium and conditions were as follows : glucose 1.0%, tryptone 3.0%, NaCl 0.3%, K2HPO4·3H2O 0.1% (w/v), MgSO4·7H2O 0.02% (w/v), L-glutamic acid 0.01% (w/v), corn steep liquor 1.5% (w/v), biotin 30μg·L-1, bran 4% (w/v), initial pH 7.0, 37℃, liquid volume 25 mL/250 mL baffled flask, for 20 hours. The activity of GAD was 1290 units of each milliliter broth under these conditions.2. The conditions of transformation from L-glutamic acid to GABA, including temperature, pH, buffer system, strain concentration, substrate concentration and reuse times of strain were optimized: temperature 37℃, pH 4.6, Na2HPO4- citric acid as the optimized buffer system, the ratio of volume of fermentation broth to transformation broth 2:1 defined as optimized strain concentration, substrate concentration 0.3mol·L-1, reuse within three times.3. Under the glucose-limited condition, the growth of GAD producer could be supposed by Monod equation withμmax = 0.543 h-1and Ks = 0.102 g·L-1 in the continuous culture process. The maximum yield of biomass and GAD product were Yx/s = 0.434 g·g-1, respectively. The maximum dilution rate was Dm = 0.45 h-1. The maximum biomass yield and the GAD yield are 1.785 g·(L·h)-1 and 119.34 U·(L·h·mL)-1, respectively.4. The effects of pH, ventilation volume, stirring speed and fed-batch on culture process and transformation were investigated in 5 L fermentor. When pH was controlled at 7.0, the ventilation volume, stirring speed, total concentration of glucose in culture medium and initial glucose concentration were 1.5 vvm, 600 rpm, 30.83 g·L-1 and 10.7 g·L-1, respectively. The residual glucose solution as fed-batch began feeding steadily after culture for 2 h. The fed-batch was ended after 5 h, and the biomass reached the highest value around 18.1 g·L-1 after culture for 11 h. Finally, 60 g·L-1 GABA was obtained from 88.2 g·L-1 L-glutamic acid after transformation for 16 h, conversion level reachs 100%. |
