Optimization Of Immobilization Conditions And Degradation Performance Of 17β-estradiol Degrading Bacteria

With the rapid development of intensive livestock and poultry farming in China,a large number of steroid estrogens in livestock excreta are polluting the environment.Biodegradation is one of the effective ways to remove estrogen,which is environmentally friendly and economical.However,the substrate of marsh irrigation area is complex,and the free functional strains need to adjust their environmental adaptability.Immobilized microbial technology can use the carrier to locate free strains in a limited space,improve the environmental adaptability of free strains,and significantly improve the repair efficiency.Therefore,the immobilization of Hyphomicrobium sp.to degrade estrogen was studied in this study.In this study,17β-estradiol（17β-E2）was used as the target material,sodium alginate embedding method and corncob adsorption method were respectively used to prepare the free strain with estrogen degradation function into immobilized strain.The removal rate of 17β-E2 was compared between the two immobilized methods in the same environment,and the microbial immobilized method with better degradation effect was selected.Response surface optimization design was carried out to investigate the effect of environmental factors on the degradation performance of immobilized strains,in order to improve the repair efficiency of free strains by immobilized strains.The main achievements are as follows:（1）Hyphomicrobium sp.SS-1,a strain of 17β-E2 specialized degradation bacteria screened from the soil in the marsh irrigation area,was immobilized by sodium alginate embedding method and corncob adsorption method respectively.Among them,the single factor experiment of sodium alginate embedding method determined that when the concentration of sodium alginate was 5%,the volume ratio of sodium alginate to bacterial liquid was 2:1,the concentration of Ca Cl₂·2H₂O was 3%,and the cross-linking time was6h,the removal effect of 17β-E2 was the best,and the degradation rate of immobilized strain was 84.18%.When the pyrolysis temperature of corn cob was 500℃,the number of corn cob mesh was 70,and the content of corn cob was 2%,the removal effect of 17β-E2 was the best,and the degradation rate of immobilized strain was 82.29%.Under the same environmental conditions,two methods were used to immobilize the strain.The removal effect of 17β-E2 by embedding method was generally superior to that by adsorption method,and the combination embedding method had the advantages of higher reaction stability,stronger strain binding ability than adsorption method,easy operation and low cost.Hyphomicrobium sp.SS-1 was immobilized with sodium alginate embedding method.（2）The factors affecting the degradation of 17β-E2 by immobilized strain by embedding method were analyzed by multi-factor response surface model fitting analysis.The optimal combination of factors for the degradation of 17β-E2 by sodium alginate embedding method was predicted as follows:the concentration of sodium alginate was5.17%,and the volume ratio of sodium alginate to bacterial liquid was 1.9:1.Ca Cl₂·2H₂O concentration was 3.87%,crosslinking time was 5.98h,and degradation rate of 17β-E2was 86.22%.（3）The degradation rate of 17β-E2 of immobilized strain was higher than that of free strain in different environment,especially in extreme environment.At 40℃,the degradation rate was 22.12%higher than that of the free strain,and the degradation rate of other strains also increased at different degrees.When the initial substrate concentration was 7mg/L and 10mg/L,the degradation rate was about 20%higher than that of free strain.When the inoculation amount was 1%-15%,the degradation rate increased by 5.5%-14.13%.Temperature,initial substrate concentration and inoculum amount had significant effects on the degradation performance of immobilized strains,and the degradation of 17β-E2 increased firstly and then decreased with the increase of temperature,initial substrate concentration and inoculum amount.The optimum environmental conditions for immobilized strain were temperature 30℃,initial substrate concentration 5mg/L and inoculation amount 5%.The degradation of 17β-E2 was in accordance with the first-order kinetic equation,and the shortest degradation half-life was2.39d.This study based on 17β-E2 obligate degradation bacteria comparison of two methods of microorganism immobilized,choice and its degradation characteristics,improve the application value in the free strain matrix in a complex environment,for livestock and poultry breeding surrounding ecological environment system of steroid estrogen pollution repair provides a effective way,and for the future of raw silk micro bacteria immobilized method provides a theoretical basis.