Screening And Fermentation Conditions Optimization Of 2,3-butanediol Producing Strain

2,3-butanediol (2,3-BDO) is one of the important chemical intermediate, which can be used in many fields such as food, chemistry, fuel. A Bacillus sublitis strain producing 2,3-BDO from glucose as substrate was isolated from different samples. The fermentation medium and fermentation conditions were investigated. On the baisis of medium and fermentation conditons optimization, preliminary test was performed in 5 L fermentor. The main research contents and results are follows:(1) 42 strains capable of producing 2,3-BDO from glucose were isolated from different natural environment samples. The strain 6-7 which could grow on medium containing glucose had a higher 2,3-BDO converting capability. After preliminary fermentation test, the concentration of 2,3-BDO reached 48.7 g/L. In addition to general morphaological and biochemical characteristics, the strain 6-7 was identified by 16S rDNA sequence and systematic analysis. The results showed that 16S rDNA sequence of strain 6-7 had similarity of 99% with Bacillus subtilis strain BIHB332, suggesting that the strain 6-7 is one of subspecied of Bacillus subtilis.(2) Effect of carbon source, nitrogen source, mineral salt and metal ion on 2,3-BDO production of Bacillus subtilis 6-7 were investigated in the shaking-flask. Glucose, corn steep liquor, (NH₄)₂SO₄, K₂HPO₄ were the siginificant factors based on the results of single factor experiments. The optimum fermentation medium compositions were determined as follows(g/L): glucose 150; corn steap liquor 10; (NH₄)₂SO₄ 6; CH3COONa 3; KH₂PO₄ 6; K₂HPO₄ 12; citrate sodium 6.0; MgSO₄ 0.05; ZnSO₄ 0.01; FeSO₄ 0.05; CaCl₂ 0.05; NiCl₂ 0.05.(3) Fermentation conditons of Bacillus subtilis 6-7 were optimized in the shaking-flask from following aspects: pH, temperature, rotation speed and so on. The best fermentation conditions were described as: initial pH 6.5, temperature37℃, the amount of inoculation 5%(V/V), the liquid volume 60 mL(in 250 mL, flask), rotation speed 160 rpm. Under optimum conditions, afer the strain 6-7 was cultured 72 hours, it produced 55.2 g/L 2,3-BDO, which increased about 13.3% more than before.(4) In order to increase the 2,3-BDO production in 5 L fermentor, effect of pH, stirrer speed and air flow on the batch fermentation were studied. The results showed that high 2,3-BDO accumulation was acheieved at the condition follow as: pH 6.5, stirrer speed 300 r/min, air flow 1 L/min. To reduce substrate inhibition and increase 2,3-butanediol yield, the fed-batch fermentation in 5 L fermentor was studied, the final 2,3-butanediol production was 72.6 g/L.