Immobilized laccase and its prepn

A technology for immobilizing laccase and laccase, applied in directions such as oxidoreductase immobilized on/in an organic carrier, can solve problems such as exudation and high price of immobilized laccase, and achieve the effect of enhancing resistance

Inactive Publication Date: 2002-10-16
GUANGZHOU INST OF GEOCHEMISTRY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The technology adopted abroad is physical embedding, but in this method, the biological enzyme is in physical contact with the carrier, and it is easy to seep out of the carrier
What uses chemical method to immobilize laccase is D'Annibale A of Italy, and it utilizes epoxy-activated polyacrylic acid carrier Eupergit C to immobilize laccase, and this immobilized enzyme is used for sewage treatment, effectively removes the laccase in waste water. Phenolic harmful substances, but the immobilized laccase is more expensive

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] (1) 300 grams of polyvinyl alcohol solution with a mass percent concentration of 10% in a 250 milliliter round bottom flask, 227 grams of a 5 percent glutaraldehyde solution with a mass percent concentration, hydrochloric acid added dropwise to make the pH=1-2, violently Stir and react for 5 hours, wash, dry, and grind into 60-120 mesh particles.

[0032] (2) In a 250 ml three-necked flask equipped with a magnetic stirrer, a spherical condenser, and a thermometer, add 5 grams of the above-prepared granules. Dissolve 2.5 grams of succinic anhydride (carboxylate) in 80 milliliters of cyclohexane and dioxane in a mixed solvent with a volume ratio of 10:1, pour it into the above-mentioned three-necked flask, add 0.25 milliliters of pyridine, and reflux to stir the reaction. The reaction temperature was 82°C. After 24 hours of reaction, pour out and wash with 40 ml of acetone, water, and acetone successively, and dry naturally to constant weight. Carboxylated polyvinyl alc...

Embodiment 2

[0037] (1) as described in embodiment 1 (1), the difference is that the acid added dropwise is sulfuric acid.

[0038](2) In the same three-necked flask as in Example 1 (2), add 5 grams of the particles prepared above, 3.7 grams of phthalic anhydride (carboxylate), 80 milliliters of n-hexane: dioxane is 8: 2 The mixed solvent, phosphoric acid 0.3 ml, reflux and stir the reaction. After 36 hours of reaction, it was poured out and washed successively with 40 ml of acetone, water, and acetone respectively, and dried to constant weight. Carboxylated polyvinyl alcohol particles having an acid value of 1.50 were obtained.

[0039] (3) 0.9 gram of N-hydroxysuccinimide, 0.03 gram of 4-dimethylaminopyridine are dissolved in ethyl acetate, and the prepared pellets are added to the round bottom flask of Example 1 (3), and 0.9 g of N,N'-diisopropylcarbodiimide was dissolved in ethyl acetate and added dropwise, and reacted with magnetic stirring at 2°C for 8 hours, and the fine particles...

Embodiment 3

[0043] (1) With embodiment 1 (1).

[0044] (2) in the there-necked flask with embodiment 1 (2), add 5 gram above-mentioned granules that make, 3 gram succinic anhydrides (carboxylate), 80 milliliters of hexanaphthene and dioxane volume ratio are 8: 2 mixed solvents, 0.3 ml of pyridine, reflux and stir the reaction. After 36 hours of reaction, it was poured out and washed successively with 40 ml of acetone, water, and acetone respectively, and dried to constant weight. Carboxylated polyvinyl alcohol particles with an acid value of 2.12 were obtained.

[0045] (3) with embodiment 2 (3).

[0046] (4) Mix 0.2 g of the prepared immobilized carrier with 1 ml of laccase (enzyme activity 0.5 IU) at 40° C., add 4 ml of pH=4.0, 0.10 M buffer solution of sodium citrate-disodium hydrogen phosphate, and stir The immobilization reaction was 12 hours.

[0047] (5) with embodiment 1 (5). Its enzyme activity is 0.8IU / gram carrier.

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PUM

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Abstract

The present invention is immobilized laccase as one kind of redox enzyme and its preparation process. The immobilized laccase is immobilized onto PVA carrier with active ester radical through chemical linkage. The preparation process includes two steps of synthesis of the carrier and the immobilizing of the laccase, the synthesis of the carrier includes PVA crosslinking, dispersing, washing, grinding and the reaction with carboxylic acid derivative and compound containing N-succinimide radical; and the immobilizing of the laccase includes the mixing of the carrier and laccase, shaking reaction, washing and drying. The immobilized laccase has high stability, high activity and low cost.

Description

technical field [0001] The invention relates to a highly stable oxidoreductase-immobilized laccase and a preparation method thereof. technical background [0002] Laccase is a kind of phenol oxidase (p-diphenol oxidase EC.1.10.3.2), which was first discovered from the secretion of sumac tree. Later, it was found that many organisms, including plants, microorganisms (mainly fungi), and even insects have Laccase is present. Pulp bleaching is to remove the residual lignin in the pulp. The traditional pulp bleaching in my country is to use chlorine bleaching method. This method has brought serious environmental pollution. Chlorine bleaching wastewater is usually not recyclable, and chlorine bleaching wastewater contains Large quantities of highly toxic substances. In recent years, studies have found that laccases can destroy aromatic rings and C α -C β The demethylation of lignin, the initial step of lignin degradation, will eventually lead to the d...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/02C12N11/06C12N11/08
Inventor 刁颖辉付时雨
Owner GUANGZHOU INST OF GEOCHEMISTRY - CHINESE ACAD OF SCI
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