Magnetic composite microsphere immobilized laccase and preparation method thereof

A technology of magnetic composite microspheres and immobilized laccase, which is applied in the direction of immobilization on or in the inorganic carrier, can solve the problems of large loss of enzyme activity, large loss of enzyme activity, and shedding, so as to avoid chemical damage and benefit Effects of recovery and reuse, improvement of operational stability

Inactive Publication Date: 2011-10-05
BEIJING NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, the cross-linking method and the covalent bond method have relatively severe reaction conditions, and the loss of enzyme activity is relatively large; the encapsulation method has a large loss of enzyme activity because the substrate binding site and active center are blocked by the carrier; the adsorption method has mild conditions, The active part of the enzyme and its spatial conformation are not easy to be destroyed, but the enzyme immobilized by ionic bonds, hydrogen bonds, dipole bonds and hydrophobic bonds in the adsorption method is easily affected by the pH and ionic strength of the reaction medium and falls off from the carrier.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] A magnetic composite microsphere immobilized laccase, the preparation method mainly includes two steps:

[0024] Vector preparation

[0025] (1) Take FeCl with a molar ratio of 1:2 2 4H 2 O and FeCl 3 ·6H 2 The mixed powder of O was dissolved in 50 mL of sodium dodecylsulfonate solution with a mass fraction of 0.5%, and 2 mol / l NaOH solution was added dropwise with stirring to adjust the pH to about 12. The obtained product was matured in a water bath at 80°C for 30 minutes, washed repeatedly with distilled water until neutral, dried at 50°C, and ground to obtain ferric oxide powder;

[0026] (2) Take the Fe after grinding 3 o 4 Ultrasonic dispersion of 1.5g in 60ml of secondary water, while ultrasonic dispersion of 6.4ml tetraethoxysilane in 120ml of methanol, under mechanical stirring, the methanol solution of tetraethoxysilane was added to Fe 3 o 4 After stirring vigorously for 15 minutes, add 6ml of concentrated ammonia water, continue to stir and react at ...

Embodiment 2

[0032] The preparation method of the carrier is the same as Example 1, except that the concentration of polyacrylamide is 0.5 mg / L, the concentration of copper sulfate solution is 0.1 mol / L, and the shaking time is 12 hours. The laccase immobilization method is the same as Example 1, the difference is that 0.1g of the finished carrier is fully mixed with 5mL of free laccase dissolved in disodium hydrogen phosphate-citric acid buffer solution, and stirred at a pH of 3.0 and a temperature of 5°C for 3 Hour. Wherein the mixing ratio of the free laccase and the finished carrier is 20mg / 1g.

Embodiment 3

[0034] The carrier preparation method is as in Example 1, except that the concentration of polyacrylamide is 5 mg / L, the concentration of copper sulfate solution is 0.05 mol / L, and the shaking time is 8 hours; the laccase immobilization method is as in Example 1, but the difference is 0.1 After fully mixing the finished g carrier with 5 mL of laccase dissolved in disodium hydrogen phosphate-citric acid buffer solution, stir for 5 hours at a pH of 5.0 and a temperature of 15°C. Wherein the mixing ratio of the free laccase and the finished carrier is 50mg / 1g.

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Abstract

The invention relates to a magnetic composite microsphere immobilized laccase and a preparation method thereof, and belongs to the field of environmental microbiology. In the immobilized laccase, metal-chelated magnetic composite microspheres are taken as a carrier, and laccase is immobilized on the carrier through coordination bonding action. The preparation method comprises two steps of: preparing the carrier, namely synthesizing magnetic ferroferric oxide nanoparticles by using a chemical precipitation process, preparing core-shell magnetic silica by taking the nanoparticles as cores through a sol-gel method, grafting polymer containing carbonyl or amino on the surface of the magnetic silica by taking gamma-chloropropyltrimethoxysilane as a medium, and preparing a functional carrier capable of immobilizing the laccase by chelating transition metal ions (such as Cu2+. Zn2+, Ni2+ or Co2+); and immobilizing the laccase, namely performing coordination adsorption of the laccase on the surface of the carrier, washing, and drying to obtain the finished product of immobilized laccase. The immobilized laccase prepared by the method has high stability and activity and quick magnetic response, is easily recovered from a reaction system and can be repeatedly used; and the preparation process is simple and the cost is low.

Description

technical field [0001] The invention relates to a magnetic composite microsphere immobilized laccase and a preparation method of the immobilized laccase. More specifically, it is to prepare a magnetic composite microsphere immobilized laccase capable of rapid magnetic separation and good operational stability. Background technique [0002] Laccase is a copper-containing polyphenol oxidase that exists widely in various organisms. Laccase can catalyze oxygen in the air to directly oxidize and decompose various phenolic dyes, substituted phenols, chlorophenols, thiophenols, bisphenol A, aromatic amines, etc. In the presence of laccase mediators (ABTS, HBT, etc.), laccase It can also catalyze the degradation of diphenylmethane, N-substituted p-phenylenediamine, organophosphorus compounds (pesticides) and dioxins related to lignin. Due to its wide substrate specificity and good stability, laccase has important application value in paper industry, wastewater treatment, biologica...

Claims

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Application Information

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IPC IPC(8): C12N11/14
Inventor 王颖贺芙蓉沈珍瑶陈晓春于光认
Owner BEIJING NORMAL UNIVERSITY
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