Degradation Mechanism Of A Low Temperature And Efficient DMP-Degrading Bacterium Ensifer Species DNM-S1

Dimethyl phthalate(DMP)is one of the commonly used plasticizers.Due to its own characteristics and extensive use,phthalates have caused serious pollution in the natural environment.Because of its environmental estrogen effect,the harm to humans should not be underestimated.Based on the climatic characteristics of the black soil region in Northeast China,this study isolated the strain DNM-S1 from agriculture soil in a vegetable greenhouse in the suburb of Harbin which can remove DMP pollution at a low temperature.The DNM-S1 was used as the research object,and DMP was the target pollutant.Through the study of removing DMP from DNM-S1,the following results were obtained:(1)Five degrading bacteria were obtained from the greenhouse soil,namely DNM-S1,DNM-S2,DNM-S3,DNM-S4 and DNM-S5,only DNM-S1 can grow at 15? and utilize DMP as a carbon source.The degrading bacterium DNM-S1 is a Gram-negative bacterium;DNM-S1 is negative in the hydrolysis test of starch,gelatin and oil,indicating that the strain does not produce amylase,protease and lipase;in the IMVIC test(Indole,methyl-red,Volt-Pur,and Citrate tests)were positive,indicating that the strain produced tryptophanase and pyruvate after decomposing glucose.The H2S test results indicated that the degrading bacteria did not produce hydrogen sulfide when decomposing sulfur-containing organic matter.Molecular biological identification indicated that the DNM-S1 strain belongs to the bacterial world,Proteobacteria,Alphaproteobacteria,Rhizobiales,Rhizobiaceae,Ensifer.In combination with its morphology and physiological and biochemical properties,DNM-S1 is named Ensifer species DNM-S1.(2)Suitable culture conditions for Ensifer species DNM-S1 growth are 6-10 for the range of pH of the medium and 15-35? for culture environment temperature.Orthogonal test is used to optimize the culture conditions of Ensifer species DNM-S1:35?,pH=10,inoculum was 5%and the initial amount of DMP was 1500 mg-L-1.The effect of environmental factors on the growth of DNM-S1 is temperature>pH>accumulation.The growth of DNM-S1 under optimal growth conditions can be described by a logistic model.When using DEP,DBP and DEHP as carbon sources,the Logistic model can also be used for description.(3)After 72 h of culture,the content of DMP decreased from the initial 2000 to 516.65 mg-L-1;DMP was completely removed by DNM-S1 with initial concentration 1500 and 1000 mg-L-1 within 72 h;DMP with initial concentration of 500 mg·L-1 was completely removed by DNM-S1 within 24 h;DMP with initial concentration of 100 mg·L-1 was completely removed within 6 h,and it takes only 3 h to remove 50 mg·L-1 DMP completely.All above results indicated that Ensifer species DNM-S1 had good degradation effects on both low DMP concentration(50 mg-L-1)and high DMP concentration(2000 mg-L-1).DNM-S1 degraded different initial concentrations of DMP in accordance with the first-order reaction kinetics equation.Degradation of DEP,DBP and DEHP by DNM-S1 is also consistent with the description of the first-order reaction kinetic equation.On the fifth day,the degradation rate of DMP at initial concent'ration of 1500 mg L-1 was 25.73%at 10?;the degradation rate of DMP was 64,93%at 15?;the degradation rate of DMP was 86.6%at 25?.(4)After 72 h sampling,three substances were detected in DMP treatment at 35?,namely DMP,phthalic acid(PA)and protocatechuic acid(PCA).This indicates that Ensifer species DNM-S1 will produce PA and PCA during the degradation of DMP.Ensifer species DNM-S1 produced DMP,PA and PCA during degradation of DEP,DBP and DEHP at 35?.Ensifer species DNM-S1 treatment at 10? and 15? can only produce PA for metabolites of DMP.(5)The test results of cell surface hydrophobicity showed that DNM-S1 had a highly hydrophobic surface(more than 80%),and the effect of changing the type of PAEs or changing the medium type on CSH was not significant.Infrared spectroscopy studies have shown that the absorption peak of DNM-S1 at 1740 cm-1 indicates that the extracellular membrane lipopolysaccharide contains a large number of hydrophobic functional groups,which provide a site for binding to DMP.This indicates that the binding of DMP to DNM-S1 occurs on the extracellular motif of DNM-S1.Analysis of secondary metabolites showed that DNM-S1 produced carotenoids,which counteract the oxidative stress of DMP to DNM-S1,thus maintaining the integrity of DNM-S1 cells and allowing the degradation reaction to proceed smoothly.