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Co-crosslinking immobilization method of tannase

A technology of tannase and co-crosslinking, applied in the field of co-crosslinking and immobilization of tannase, can solve the problems of high reaction activity, excessive cross-linking of enzymes, blocking of mass transfer channels, etc. Improve catalytic performance and improve the effect of microenvironment

Active Publication Date: 2020-02-11
NINGBO UNIV
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AI Technical Summary

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 tannase
  • Co-crosslinking immobilization method of tannase
  • Co-crosslinking immobilization method of tannase

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Embodiment Construction

[0016] Enzyme immobilization

[0017] 1) Bisphenol A epoxy resin (brand name is E-51, epoxy value is 0.51, number average molecular weight is 392), three kinds of components of methanol and diethylenetriamine are mixed according to the mass ratio of 2: 2: 1, 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 tannase 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;

[0020...

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Abstract

The invention relates to a co-crosslinking immobilization method of tannase. Oil-soluble trimethylolpropane triacrylate is used as a crosslinking agent, reactants in a water phase are the tannase containing amino and a supramolecular complex formed by aminated epoxy resin and beta-cyclodextrin, through a Michael addition reaction of double bonds and the amino, a co-crosslinking polymerization reaction occurs at the low temperature, and immobilized tannase with different loading capacities is prepared. By controlling the cross-linking degree, dispersity is improved, the mass transfer microenvironment inside the tannase is improved, the immobilized tannase has high catalytic activity and has the highest activity when the loading capacity is 89 mg of an enzyme per gram of a carrier, and the activity reaches 92% 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 tannase. The novel immobilized tannase can be specially used to remove the bitter taste caused by tannin in natural food . Background technique [0002] Tannase (EC 3.1.1.20), also known as tanninyl hydrolase (isoelectric point 4.5), is a cell membrane-bound enzyme induced by microorganisms in the presence of tannins. Tannase is rich in sources, not only widely exists in tannin-rich plants in nature, but also exists in a large number of tannase-producing bacteria. The molecular weight of tannase is between 50 and 320kDa, and its quaternary structure is a heterohexamer or heterooctamer formed by two or more different subunits through disulfide bonds. Pure tannase is white or light black powder, insoluble in ethanol, soluble in water to form a colorless clear solution. The enzyme can catalyze the hydrolysis of...

Claims

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Application Information

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IPC IPC(8): C12N9/18C12N11/08
CPCC12N9/18C12Y301/0102C12N11/08Y02A40/90
Inventor 吴嘉沁张瑞丰李艳肖通虎龙能兵
Owner NINGBO UNIV
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