Immobilized laccase, preparation method and application thereof

A technique for immobilizing laccase and laccase, applied in chemical instruments and methods, immobilized on/in organic carriers, wastewater treatment in processing, etc., can solve difficult industrial production and practical application, complicated processing technology, Solve problems such as low immobilization efficiency, achieve stable properties, simple processing technology, and firm adsorption

Active Publication Date: 2010-12-15
合肥华纳生物医药科技有限公司
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  • Abstract
  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The bonding and cross-linking methods of immobilized laccase technology are the most widely studied, which provide an important theoretical basis for the wide application of laccase, but the existing laccase immobilization methods and carriers generally have low immobilization efficiency and poor processing technology. Complex, high production costs, unstable properties and other defects, it is difficult to industrial production and practical application

Method used

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  • Immobilized laccase, preparation method and application thereof
  • Immobilized laccase, preparation method and application thereof
  • Immobilized laccase, preparation method and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0020] The preparation of embodiment 1 iminocarboxylic acid type chelating resin immobilized laccase

[0021] (1) Soak 10g of commercially available iminocarboxylic acid type chelating resin (model LSC-100) in 500ml ethanol for 4h, after filtering off the ethanol, vacuum dry for 4h;

[0022] (2) Put the dried resin into 500ml of phosphate buffer solution containing 3000U laccase (pH=8.0), shake and adsorb at 4°C for 6-12h;

[0023] (3) Washing with a phosphate buffer solution of pH=8.0, and detecting the washing solution until no protein is detected in the washing solution.

[0024] (4) Measure the amount M1 of laccase added before fixation and the amount M2 of laccase remaining in the eluate after fixation by Coomassie brilliant blue staining method (Bradford method), and calculate the immobilization rate of laccase: the fixation rate of laccase Conversion rate=[(M1-M2) / M1]×100%. The immobilization rate of laccase was 92%.

Embodiment 2 3

[0025] Embodiment 2 Preparation of ternary carboxyl chloride resin immobilized laccase

[0026] (1) Soak 10g of commercially available ternary carboxyl-containing vinyl chloride resin (model C42) in 500ml of ethanol for 4h, filter off the ethanol, and vacuum-dry for 4h;

[0027] (2) Put the dried resin into 500ml of phosphate buffer containing 5000U laccase (pH=8.0), shake and adsorb at 4°C for 6-12h;

[0028] (3) Washing with a phosphate buffer solution of pH=8.0, and detecting the washing solution until no protein is detected in the washing solution.

[0029] (4) Measure the amount M1 of laccase added before fixation and the amount M2 of laccase remaining in the eluate after fixation by Coomassie brilliant blue staining method (Bradford method), and calculate the immobilization rate of laccase: the fixation rate of laccase Conversion rate=[(M1-M2) / M1]×100%. The immobilization rate of laccase was 86%.

Embodiment 3

[0030] The thermostability of embodiment 3 immobilized laccase

[0031] The immobilized laccase and free laccase in Example 1 were respectively placed in a 70°C water bath for incubation, and samples were taken regularly to detect their guaiacol oxidation activity. The results showed that after the free enzyme was incubated in a 70°C water bath for 1 hour, the enzyme activity Significantly decreased, and the loss of enzyme activity reached more than 99% after 4 hours. However, the immobilized laccase only lost 19% of its enzyme activity when incubated for 4 hours under the same conditions, and still retained 56.5% of its enzyme activity after 8 hours, showing that the thermal stability of the enzyme was significantly improved after immobilization. The immobilized laccase of Example 2 is similar to this.

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Abstract

The invention discloses an immobilized laccase. Laccase is immobilized on a carrier through absorption and chelation by using chelate resin as the carrier, and the chelate resin is bonded with an active functional group containing carboxyl. The immobilized laccase can be applied to treating wastewater containing phenols pollutants, and has the characteristics of simple production process, low cost, high laccase absorption rate, little laccase activity, stable property, and the like.

Description

technical field [0001] The invention relates to the technical field of enzyme immobilization, in particular to a chelating resin immobilized laccase, a preparation method thereof and an application in sewage treatment. Background technique [0002] Laccases, a protein that binds multiple copper ions, belong to the blue multi-copper oxidase family and are widely found in plants, fungi and even insects. Laccase can survive in the air and can catalyze a series of substrates including phenols and their derivatives, arylamines and their derivatives, carboxylic acids and their derivatives, steroid hormones and biochromes, metal organic compounds, and non-phenolic substrates. The oxidation reaction of the substrate, the only product after the reaction is water. It is a green, environmentally friendly, and mild biocatalyst. It has great potential in bio-papermaking, food production, environmental pollution control, organic synthesis, biological and immunological detection Strong ap...

Claims

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Application Information

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IPC IPC(8): C12N11/02C02F3/34C02F101/36C02F103/28C02F103/36
Inventor 刘迎春高源
Owner 合肥华纳生物医药科技有限公司
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