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Co-crosslinking immobilization method of laccase

A technology of co-crosslinking and laccase, applied in immobilized enzymes, biochemical equipment and methods, immobilized on/in organic carriers, etc., can solve high reactivity, excessive enzyme crosslinking, blocking mass transfer channel and other issues to achieve the effect of improving catalytic reaction activity, improving catalytic performance and improving microenvironment

Active Publication Date: 2020-02-07
NINGBO UNIV
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Problems solved by technology

The most commonly used cross-linking agent is water-soluble glutaraldehyde, which has high reactivity and is difficult to control the dosage. It is easy to cause excessive cross-linking of the enzyme and cause a great loss of enzyme activity. In addition, the traditional cross-linking method often It is necessary to precipitate and aggregate enzyme molecules before cross-linking, which will not only cause waste of enzymes, but also block mass transfer channels, and cannot fully exert the catalytic efficiency of enzymes

Method used

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  • Co-crosslinking immobilization method of laccase
  • Co-crosslinking immobilization method of laccase
  • Co-crosslinking immobilization method of laccase

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Embodiment Construction

[0016] Enzyme immobilization

[0017] 1) Bisphenol A epoxy resin (brand name is E-44, epoxy value is 0.44, number average molecular weight is 454), three kinds of components of methanol and triethylenetetramine are mixed according to the mass ratio of 2: 2: 1.5, Stir and react in the range of 25-35°C for 4-5 hours, pour the mixture into water, wash the precipitate repeatedly with water to remove methanol and a small amount of amine, then put it in a vacuum oven and dry it at room temperature to obtain an aminated epoxy resin;

[0018] 2) Add epoxy resin amides and β-cyclodextrin into water at a molar ratio of 1:2.1 to 1:2.3, heat and stir until all epoxy resin amides are converted into molecular complexes and dissolved in water, and keep the The total mass concentration of the aqueous solution is in the range of 5 to 6wt.%.

[0019] 3) dissolving the laccase in a phosphate buffer solution with pH=6.5, and keeping the concentration of the enzyme in the range of 1.0-7.0 mg / mL; ...

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Abstract

The invention relates to a co-crosslinking immobilization method of laccase. Oil-soluble dipentaerythritol hexaacrylate is used as a cross-linking agent, reactants in a water phase are amino-containing laccase and a supermolecular complex formed by aminated epoxy resin and beta-cyclodextrin, a Michael addition reaction of double bonds and amino is utilized, a co-crosslinking polymerization reaction is carried out at a low temperature, and immobilized laccase with different loading capacities is prepared. By controlling the crosslinking degree, the dispersity is improved and the internal mass transfer microenvironment is improved. The immobilized enzyme has higher catalytic activity, and has the highest activity when the loading capacity is 77mg enzyme / g carrier, and the loading capacity reaches 88% of that of a free enzyme.

Description

technical field [0001] The invention relates to the technical field of immobilized enzyme biocatalysis, in particular to a method for co-crosslinking and immobilizing laccase. The novel immobilized laccase can be specially used for removing phenolic substances in water. Background technique [0002] Laccase (EC 1.10.3.2) is a copper-containing polyphenol oxidase (isoelectric point of 5.0), a typical oxidoreductase, which is widely distributed in the natural environment, mainly divided into fungal laccases And laccase laccase, its average molecular weight is between 50,000 and 100,000, and the peptide chain of laccase is usually composed of 500 to 550 corresponding amino acids. The difference between different types of laccases is generally due to the different content or types of sugar groups in the laccase proteins. There are 4 copper ions in a typical laccase, and these 4 copper ions constitute its catalytic center. Laccase catalyzes a wide range of substrates, and has g...

Claims

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Application Information

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IPC IPC(8): C12N11/10C12N11/096
CPCC12N11/10C12N11/08C12N9/0061C12Y110/03002
Inventor 吴嘉沁张瑞丰李艳肖通虎龙能兵
Owner NINGBO UNIV
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