The Research Of Growth Characteristic For Saccharopolyspora Spinosa And Fermentation And Metabolic Regulation Of Spinosad

Spinosad,the macrolide secondary metabolites produced by Saccharopolyspora spinosa,is the active ingredients in a family of insect control agents.In this dissertation,the growth characteristic and inoculation standard for Saccharopolyspora spinosa,fermentation process,the batch fermentation kinetics of spinosad producing strain,process parameters controls and catabolism repression of spinosad were studied.The growth rule,cultivation process and inoculation standard of Saccharopolyspora spinosa were studied.The result showed that the cultivation of single colony on the plate for 7 days was the best inoculation standard of the slant strains.The optimized culture conditions of liquid seed were established as a combination of the glass bead of 30 grains in 500 mL shake flask,cultural temperature at 30 ? and initial pH of 8.5.The inoculation standard of liquid seed consisted of greater than 1.4 g/L of biomass,75.0-80.0 mg/L of amino nitrogen,less than 7.0 g/L of reduced sugar,less than 7.5 of pH and 32-36 h of inoculum age.Employing the new inoculation standard,the spinosad production is increased by 58.5%.Cottonseed meal,colza oil,phytic acid were screened by single factor experiment which contributed to inprove the production of spinosad.Corn flour hydrolysis sugar,fish peptone and phytic acid,which had significantly positive effect on spinosad production,were screened from 8 related factors by Plackeet-Burman design.The reasonable ranges of these three factors were determined by single factor experiment.ANN coupling PSO algorithm based on hybrid design was applied to predict these factors'mutual interactions for spinosad production.The optimal medium composition was determined as follows(g/L):glucose 55.0,hydrolysis sugar 8.5,yeast extract 10.0,peptone 7.5,cottonseed power 30.0,colza oil 30.0,MgSO4·7H2O 0.4,phytic acid 0.5.The optimized conditions allowed spinosad production to be increased 36.5 percent than before optimization after verification test.The batch fermentation kinetics of spinosad producing strain was studied.Mathematic models of mycelia growth,spinosad production and substrate consumption were established on the basis of the Logistic and Luedeking-Piret equations.Experimental values were compared fitted values by models with software origin 8.0 and they were well coincident.The proposed kinetic models could well describe the processing of batch fermentation process of spinosad production.The regulation of carbon and nitrogen metabolites repression on spinosad by Saccharopolyspora spinosa was studied.When the concentration of initial glucose was greater than 5.5%,or the concentration of glucose was greater than 4%or the concentration of amino nitrogen was greater than 140 mg/L in the rapid synthesis period of spinosad,the synthesis of spinosad would be restrained.Sucrose and beef extract were screened by single factor experiment which were the non-interfering carbon and nitrogen source of spinosad synthesis,the spinosad production was increased by 11.8 precent and 10.6 precent respectively.Employing the optimized medium feed process,the spinosad production reach to 1581.9 mg/L.The batch fermention process in a shake flask was studied in a 5-L fermentor,the spinosad production reach to 1520.5 mg/L.