Optimizaition Of Fermentation Conditions And Kinetic Properties For New Pesticide- Alpha-Pinene

As an important chemical intermediate, a-pinene can be used to synthesize various substances, such as insecticides and medicines and so on. As a new natural botanic pesticide, alpha-pinene enjoys extremely broad market prospect. The study took the theory of metabolic control fermentation as the guidance, and made experiments on the optimization of shake-flask culture and fermentation conditions of bacterial strain YJM28 and on its fermentation kinetic properties. The main results were as follows:(1) Measured a-pinene content in fermentation liquor head space through GC methodology. The gas chromatographic condition was Innowax HP-1 column, high purity nitrogen as mobile phase, colum temperature was controled by temperature programmed chromatography as follows:maintained for 0.5min at 50℃, increased temperature to 70℃ at 4℃/min, then to 250℃ at 20℃/min, maintained for 10 min, the flow velocity was 1 mL/min, the detector was FID, and both the temperature and the temperature in vaporizing chamber were 200℃. a-pinene is within the range of 0.25-40 mg/L, and the linear regression was fair (R2=0.9993). The method had the features of simpleness, speediness and high accuracy, andwas suitable for the on-line analysis of a-pinene.(2) Made the focused research on shake flask fermentation conditions of bacterial strain YJM28. Optimize the matching ratio of fermentation culture medium through Central-Composed design and response surface analysis, and the optimized culture mediumwas as follows:2.47 g/L glucose,9 g/L peptone,63.78μL/100mL microelement solution,0.04 mmol/L IPTG concentration,0.15 g/L ammonium ferric citrate,8 mmol/L MgSO4 solution,4.9 g/L dipotassium hydrogen phosphate trihydrate, and 0.525 g/L citric acid monohydrate. Optimize the seed culture conditions and the fermentation conditions, and the optimized shake-flask culture and fermentation conditions were as follows:initial fermentation medium with pH6.4, anaerobic shake flask with loaded liquid of 100 mL per shake flask, rotating speed at 180 r/min, and fermentation temperature at 30℃. Under the conditions, ferment bacterial strain YJM28 in batches for 48h, and generate 33.26 mg/L a-pinene.(3) Made experiments on the fermentation of bacterial strain YJM28 in 590mL anaerobic bottle in batches, refered to a-pinene generation from bacterial strain YJM28 at different rotating speeds and constant pH, determined the optimal rotating speed of 180 r/min and the optimal pH of 6.4, at this point the a-pinene output capacity is 33.24 mg/L, made the kinetic research in fermentation process under the condition, and established the kinetic model of the bacterial strain fermentation. Those results showed that the established model can properly match the fermentation process of a-pinene, providing the theoretical basis for the further optimization and amplification.