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Preparation method of copper alginate loaded laccase and mediator ABTS composite biocatalyst

A biocatalyst, copper alginate technology, applied in biochemical equipment and methods, chemical instruments and methods, fixed on/in organic carriers, etc., can solve the problem that the mediator ABTS cannot be recycled and unfavorable for the green development of the surrounding environment , can not effectively catalyze the substrate and other problems, and achieve the effect of good industrial application prospect, excellent catalytic activity and low price

Pending Publication Date: 2021-06-01
WUHAN TEXTILE UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the prior art, the preparation and recycling process of biological enzyme-mediator ABTS (2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt) composite catalyst is complicated, The catalytic activity is low, and the residual ABTS has certain environmental toxicity, which is not conducive to the green development of the surrounding environment, and the prepared biological enzyme-mediator catalyst cannot be recycled for many times, and cannot effectively catalyze the substrate
However, during the treatment of organic pollutants in printing and dyeing wastewater, the ABTS mediator is in a free state, so the biological enzyme mediator ABTS needs to be added every time it is reused, causing the mediator ABTS to be unrecyclable and increasing production costs.

Method used

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  • Preparation method of copper alginate loaded laccase and mediator ABTS composite biocatalyst
  • Preparation method of copper alginate loaded laccase and mediator ABTS composite biocatalyst
  • Preparation method of copper alginate loaded laccase and mediator ABTS composite biocatalyst

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preparation example Construction

[0030] The invention provides a method for preparing a copper alginate-supported laccase and a mediator ABTS composite biocatalyst, comprising the following steps:

[0031] S1. Dissolving sodium alginate powder in deionized water to prepare an aqueous solution of sodium alginate with a mass fraction of 1.5wt%-3wt%, and then performing ultrasonic treatment; wherein, the ultrasonic treatment is performed at a temperature of 40-50°C and a frequency of 100Hz. Ultrasonic treatment time is 10-20min;

[0032] S2, dissolving anhydrous copper sulfate in deionized water to prepare a copper sulfate solution of 0.1mol / L-0.2mol / L;

[0033] S3. Adding the mediator substance ABTS with a concentration of 0.1mmol / L-5mmol / L to the sodium alginate aqueous solution obtained in step S1 to obtain solution A, and then performing ultrasonic treatment;

[0034] S4. Add laccase with a concentration of 2.5g / L-10g / L to the sodium alginate aqueous solution obtained in step S1 to obtain solution B, and th...

Embodiment 1

[0038] (1) Preparation of Alginate Copper Carrier Immobilized Mediator Substance ABTS

[0039] Add 2.5g of sodium alginate powder into 100mL of deionized water, stir vigorously at room temperature for about 30min until completely dissolved. Then, under the condition of a frequency of 100 Hz, it was ultrasonicated at room temperature for about 10 min to prepare a sodium alginate aqueous solution with a mass fraction of 2.5 wt%. Add the mediator substance ABTS (5mmol / L) to the above solution, and then, under the condition of frequency 50Hz, sonicate at room temperature for about 10min to obtain a uniform solution. The above solution was added dropwise into 100 mL of copper sulfate solution (0.1 mol / L), and stirred until the reaction was completed to obtain immobilized mediator substance ABTS beads.

[0040] (2) Preparation of copper alginate carrier immobilized laccase

[0041] Add 2.5g of sodium alginate powder into 100mL of deionized water, and stir vigorously at room temper...

Embodiment 2

[0043] Enzyme activity assay of immobilized laccase

[0044] Mix the buffer with ABTS, take 5g of copper alginate carrier to immobilize laccase, add 10mL of ABTS (0.1mmol·L-1) and 40mL of acetic acid-sodium acetate buffer solution (pH=4.6), stop in ice bath after 20min of reaction, The absorbance at 420 nm was measured after the supernatant was filtered.

[0045] The formula for calculating the enzyme activity of immobilized laccase is as follows:

[0046] U / mL=((ΔAV / εDt) / V0)×105.

[0047] ΔA=1.43 U=U / L=99.39 μmol / mg / min 99.39 / 422.23=23.54%.

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Abstract

The invention discloses a preparation method of a composite biocatalyst system composed of copper alginate loaded laccase and a mediator ABTS. A natural biomass material sodium alginate is adopted as a carrier, and free laccase protein molecules and mediator ABTS molecules are respectively immobilized by utilizing the coordination complexing action of a metal copper element, so that the composite biological enzyme catalyst is formed. After being immobilized, the macromolecular bio-enzyme and the mediator ABTS can be recycled, the service life is long, more than 80% of reactive brilliant blue KN-R degradation rate and more than 75% of Congo red degradation rate can still be maintained after 5 times of repetition, the production cost is reduced, and the method has a good industrial application prospect. The preparation method has the advantages of simple preparation process, mild reaction, easily available raw materials, low price and no pollution to the environment.

Description

technical field [0001] The invention belongs to the technical field of enzyme catalysts, and in particular relates to a method for preparing a copper alginate-loaded laccase and a mediator ABTS composite biocatalyst. Background technique [0002] Laccase is an environmentally friendly multi-copper oxidase, which has the advantages of strong catalytic ability, high efficiency and wide range of substrates. It is used in the processes of printing and dyeing wastewater decolorization, pulp bleaching, sewage detoxification and bioremediation. In the prior art, the preparation and recycling process of biological enzyme-mediator ABTS (2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt) composite catalyst is complicated, The catalytic activity is low, and the residual ABTS has certain environmental toxicity, which is not conducive to the green development of the surrounding environment, and the prepared biological enzyme-mediator catalyst cannot be recycled many t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N11/10C12N9/02C02F3/34C02F101/30C02F103/30
CPCC12N11/10C12N9/0061C12Y110/03002C02F3/342C02F2101/308C02F2103/30
Inventor 李玉广孙乾王鑫盛明月路兴杰蔡亚君王强
Owner WUHAN TEXTILE UNIV
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