Polyglutamic Acid Fermentation Process Optimization And Field Experiment

γ-polyglutamic acid(γ-PGA)is mainly a homopolymeric amino acid polymerization product produced by Bacillus subtilis with similar structure,which has good water solubility,biodegradability,water absorption and water retention characteristics,fertilizer synergy,etc.And is not toxic to the environment.The Zero Fertilizer Zero Growth Plan of the Ministry of Agriculture stipulates that the application of chemical fertilizers and the blind application of fertilizers should be gradually reduced to increase the utilization rate of fertilizers.According to the literature,γ-PGA can reduce the use of chemical fertilizer by 20%.It is a green and environmentally-friendly fertilizer synergist,so it has great development and utilization value.In the agricultural field,γ-PGA has received more and more attention.In this project,a strain of high-yield polyglutamic acid was screened by screening and rescreening the original strains and purchased strains,and identified as Bacillus subtilis by molecular biology 16 Sr DNA method.By consulting the literature,six polyglutamic acid basic fermentation media were selected and polyglutamic acid fermentation experiments were carried out.Based on the high and low yield of polyglutamic acid,a suitable basal medium for polyglutamic acid fermentation was determined.On the basis of polyglutamic acid basic medium,single factor experiments were carried out on the inoculum size,initial p H,liquid loading,fermentation temperature,etc.,and orthogonal test was performed based on the results of single factor fermentation to determine the most polyglutamic acid.Suitable fermentation process: glucose 33.97g/L,L-glutamic acid 35.34g/L,(NH4)Cl 1.08g/L,Mn SO4·H2O3g/L,Mg SO4 8g/L,K2HPO4·3H2O 20g/L,chlorination Sodium 10g/L;fermentation parameters: initial p H 8.0,culture temperature is 34 ℃,inoculum volume is 10%,liquid volume is 15%,rotation speed is 150 rev/min.The fermentation level of gamma-PGA reached 33.59g/L,which was 2.54 times of the initial fermentation level of 13.21g/L,and reached the level of industrial production.The solid liquid preservation experiment was carried out on polyglutamic acid.The liquid product was subjected to liquid preservation experiments by four kinds ofsodium benzoate,potassium sorbate,sorbitol and preservative of neostigmine,and the experiment showed that the preservation effect of chlorpheniramine on polyglutamic acid was found.Preferably,potassium sorbate and sorbitol are the second most effective effects on polyglutamic acid.In terms of concentration,0.1% of the new benzalkonium can exert a therapeutic effect on polyglutamic acid.Solid product: by adding maltodextrin to the fermentation broth material,followed by spray drying in an amount of 3.5%.The spray conditions were as follows: the inlet air temperature was175℃,the outlet air temperature was 85℃,and the centrifugal spray was dried at a rotation speed of 12,000 rpm.Can be discharged normally,but there is still a wall hanging phenomenon.The cyclone collects material 1 and the cyclone collects material2 as a different sample.Taking the cyclone separator as the example,the fermentation broth has a total of 2.5L and a solid content of 5%.200 g of starch is added for spray drying,and the yield is 28.9%.The accuracy,advantages and disadvantages of the three methods of turbidimetry,viscosity and liquid chromatography were compared by laboratory experiments under laboratory conditions.The optimal conditions for the determination of polyglutamic acid by CTAB method are detection wavelength of 250 nm,CTAB concentration of 5 g/L,reaction time of 3 min,and reaction temperature of room temperature.Although the CTAB method can quickly determine the content,the accuracy is relatively low,and the content can be determined more accurately by HPLC.Finally,liquid chromatography was chosen.Water absorption and water retention of polyglutamic acid: the absorption capacity of polyglutamic acid is 2.03 times that of the water retention agent;the soil moisture without polyglutamic acid reaches a balance before 15 h,and the soil with polyglutamic acid added After the equilibration time is 25 h,the retention time of polyglutamic acid can be extended by two-thirds than that without addition.Polyglutamic acid germination test: The addition of polyglutamic acid has little effect on the germination of wheat seeds.The germination rate of soybeans and the germination rate of baby foods were higher than those of the unused blank group,which were 3.4% and 29.6%,respectively.Carrot root irrigation test: Compared with the test group 2 without polyglutamic acid,the test group 4,test group 5,and test group 6 using polyglutamic acid showed a3.55% and 7.52 increase in weight from the point of fruit weight.%,13.67%,and their output has increased by 3.03%,7.38%,and 12.36% respectively.Wheat seed dressing test: after adding polyglutamic acid,the wheat stem height showed a certain increase,indicating that polyglutamic acid can be used in combination with azoxystrobin and thiamethoxam,and can play a role in promoting root growth and aerial parts.