Preparation Of Composite Carrier Of Enzyme Preparation And Its Remediation Effect On Petroleum Contaminated Soil

Petroleum hydrocarbon pollutant is a typical organic pollutant that is difficult to biodegrade in the environment.Its composition is complex and has obvious carcinogenic,teratogenic and mutagenic effects,which is easy to cause serious harm to the environment and human health.Laccase is a cuprine oxidase capable of using molecular oxygen for free radical catalytic reactions to oxidize a variety of aromatic and non-aromatic compounds.In view of the disadvantages of free laccase(Free Laccase,FL)having poor stability,easy deactivation and unrecovery in practical environmental remediation applications,improving the stability and catalytic activity of the enzyme is a very challenging and practical potential in the field of environmental remediation.In this study,eight inorganic carrier materials were screened,and expanded clay was selected as the inorganic carrier material for immobilized laccase.Expanded clay and rapeseed straw biochar were used to prepare a composite carrier of expanded clayrapeseed straw biochar immobilized laccase(Lac@CRBC).The immobilization conditions of expanded clay immobilized laccase(Lac@CER)and expanded clay-rapeseed straw biochar immobilized laccase(Lac@CRBC)were optimized.The immobilized laccase was characterized by physical and chemical indicators such as scanning electron microscopy(SEM),Fourier transform infrared spectroscopy(FTIR),and automated specific surface area and porosity analyzer(BET)analysis.The stability of laccase in different states and the degradation effect on petroleum-hydrocarbon-contaminated soil were studied and the soil enzyme activity during the remediation process was evaluated and analyzed.The main conclusions are as follows:(1)The effects of eight inorganic carriers on the immobilization of laccase are in the following order: ceramic > perlite > maifan stone> roseite > rainbow colored stone> carbon sphere > volcanic rock > white stone.Based on the analysis of SEM,FTIR,and BET results of the eight inorganic carriers,ceramic particles were selected as the optimal inorganic carrier material for immobilizing laccase.(2)The optimal immobilization conditions for Lac@CER are as follows: carrier particle size of 60 mesh,laccase addition of 10 m L,adsorption time of 3h,glutaraldehyde volume fraction of 2%,and cross-linking time of 3h.The optimal conditions for preparing Lac@CRBC are laccase addition of 10 m L,adsorption time of 1h,glutaraldehyde volume fraction of 4%,and cross-linking time of 2.5h.(3)The acid-base stability,thermal stability and storage stability of Lac@CER and Lac@CRBC were improved compared with FL.The relative enzyme activity of FL,Lac@CER and Lac@CRBC was the highest when p H was 3.At p H 8,the relative enzyme activity of Lac@CRBC was 34.75%,which was 21.47% and 13.87% higher than that of FL and Lac@CER,respectively.At 60℃,the relative enzyme activity of Lac@CRBC remained 97.1%,which was increased by 71.1% and 19.98% compared with FL and Lac@CER,respectively.When the storage time was 20 days,Lac@CRBC still retained 62.45% relative enzyme activity,which was 39.25% and 18.82% higher than FL and Lac@CER,respectively.(4)The degradation rates of petroleum hydrocarbons in contaminated soil in the five treatment groups during the remediation cycle in order from largest to smallest: Lac@CRBC(35.19%)> Lac@RBC(32.24%)> Lac@CER(28.85%)> FL(21.53%)> CK(17.7%).Compared with the free laccase-treated group,the immobilized laccase-treated group maintained higher stability and activity of catalase,dehydrogenase,polyphenol oxidase and laccase in the soil.