Immobilization Of Laccase Via Biomimetic Mineralization And Its Application For Degradation Of Bisphenol A

Bisphenol A is an endocrine disruptor.When people ingest BPA,it will cause obesity and infertility,and more serious cases will cause cancer.However,it is also a very important intermediate in industrial production and is widely used in industrial production.In the process of production,it is discharged into the environment through waste liquid to cause water pollution.BPA in water causes serious safety hazards to human health through biological enrichment and biologic chain.Therefore,there is an urgent need to find an efficient and safe method for degrading bisphenol A.Laccase is a copper-containing oxidase that has a good degradation effect on phenols.However,the stability of laccase is poor,and its activity is easily affected by the external environment,which greatly limits its large-scale application in industrial production.In order to effectively solve the above problems,this paper is committed to immobilizing laccase by bionic mineralization method to improve its stability and catalytic activity.The specific experimental content is as follow:First,laccase@HKUST-1 biocomposite was synthesized in one step by using HKUST-1 containing laccase coenzyme factor as the immobilized laccase carrier.By testing them activity found that the activity of laccase@HKUST-1 was 1.5 times that of free laccase,which proved that HKUST-1 and laccase had synergistic catalytic performance.In addition,due to the HKUST-1 wrapped on the surface of laccase provided protection for laccase,the thermal stability,pH stability,cycle stability and organic solvent tolerance of laccase were greatly improved.Even after 10 cycles of BPA degradation,laccase@HKUST-1 still maintained the degradation efficiency of75.9%.The preparation of this biocomposite material breaked the concept that MOFs were only used as enzyme-immobilized carriers,and layed a foundation for the future selection of suitable carrier materials to improve enzyme activity while protecting enzyme.Secondly,we optimized the parameters of the experiment and prepared the laccase@HKUST-1 with a flower-like structure to make laccase@HKUST-1 expose more laccase active sites.Under optimal conditions,when the Cu²⁺concentration was62.5 mM,the H₃BTC concentration was 12.5 mM,the laccase concentration was 2mg/mL,the incubation temperature was 25 ^oC,and the incubation time was 8 h,the activity of laccase@HKUST-1 reached the highest value,134.9%.Then their catalytic activity was tested under their optimal reaction conditions,and it was found that the flower-like laccase@HKUST-1 retained activity were 3.04 times that of the globular structure.The kinetic study of the flower-shaped laccase@HKUST-1composite showed that its K_m value was 0.36 times that of spherical structure,while the V_maxax and K_catat of the flower-shaped material were 5.24 times and 4.59 times as large as spherical structure,respectively.It is proved that the flower-like structure increases the affinity and catalytic rate by exposing more active sites.Finally,based on the high adsorption capacity of the flower-like structure material and the strong affinity of streptavidin and biotin,the flower-like SA&laccase@HKUST-1/P1 composite material was prepared to achieve efficient degradation of bisphenol A without medium.The kinetic study of SA&laccase@HKUST-1/P1 showed that the catalytic degradation efficiency of SA&laccase@HKUST-1/P1 for bisphenol A was 4.43 times that of laccase@HKUST-1.The double-layer protection provided by HKUST-1 and P1further improves the stability of SA&laccase@HKUST-1/P1.During the process of degrading bisphenol A,the degradation rate of SA&laccase@HKUST-1/P1 was97.4%after 12 hours,while the degradation rates of free laccase and laccase@HKUST-1 were 17%and 37.1%,respectively.The synthesis of SA&laccase@HKUST-1/P1 composite material greatly improved the degradation efficiency of bisphenol A,laying a foundation for large-scale application of laccase.