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Novel immobilized oxidordeuctase electrode and preparation method thereof

A reductase and electrode technology, applied in the direction of oxidoreductase, biochemical equipment and methods, immobilized on or in the inorganic carrier, can solve the problems of low adsorption efficiency, difficult to put into production, etc., achieve good stability, enhance biological Activity, effect of improving repeated use stability

Pending Publication Date: 2019-12-20
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The oxidoreductase prepared by the invention adopts the adsorption method to reduce the influence of covalent binding on the enzyme activity, the operating conditions are mild, and the biocompatibility is good, but the adsorption efficiency is low and it is not easy to put into production

Method used

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  • Novel immobilized oxidordeuctase electrode and preparation method thereof
  • Novel immobilized oxidordeuctase electrode and preparation method thereof
  • Novel immobilized oxidordeuctase electrode and preparation method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0030] (1) horseradish peroxidase is modified:

[0031]Dissolve 10mmol of chloroacetic acid in 10mL of acetonitrile and add it to a dry round-bottomed flask. After the dissolution is complete, slowly add 1.1mmol of 1-butyl-2-methylimidazole dropwise, and reflux at 100°C for 10 hours with magnetic stirring. After the reaction, extract twice with anhydrous ether, remove the yellow oily substance in the lower layer, and dry it in vacuum at 70°C for 24 hours to obtain a yellow thick liquid 1-butyl-2-methyl-3-acetoxyimidazolium bromide modifier; Add 1mmol of 1-butyl-2-methyl-3-acetoxyimidazolium bromide to 1mmol of carbonyldiimidazole, stir magnetically under the action of 2mL of anhydrous dimethyl sulfoxide, react at room temperature for 8h and stop, after the reaction is complete, The active body was obtained; 90 μL of the active body was added to 5 mL of horseradish peroxidase solution with a concentration of 350 μM, and the reaction was stopped after 24 hours of magnetic stirri...

Embodiment 2

[0041] (1) modify laccase:

[0042] Dissolve 10mmol of bromopropionic acid in 10mL of acetonitrile and add it to a dry round-bottomed flask. After the dissolution is complete, slowly add 1.1mmol of 1,2-dimethylimidazole dropwise, and reflux at 60°C for 5 hours with magnetic stirring. After the end, extract twice with anhydrous ether, remove the yellow oily substance in the lower layer, and dry it in vacuum at 70°C for 24 hours to obtain a yellow thick liquid 1,2-methyl-3-propionyl imidazolium bromide modifier; take 1 mmol of 1 , 2-methyl-3-propionyl imidazolium bromide, add 1mmol carbonyldiimidazole, magnetically stir under the action of 2mL of anhydrous dimethyl sulfoxide, stop after 2h at room temperature, after the reaction is completed, the active body is obtained; Add 90 μL of the active substance into 5 mL of 150 μM laccase solution, stop the reaction after magnetic stirring for 8 hours at 0° C., and dialyze for 24 hours to obtain a methylimidazole acylated laccase solut...

Embodiment 3

[0049] (1) Modify polyphenol oxidase:

[0050] Dissolve 10mmol of bromopropionic acid in 10mL of acetonitrile and add it to a dry round bottom flask. After the dissolution is complete, slowly add 1.1mmol of 1-butyl-2-methylimidazole dropwise, and reflux at 80°C for 7 hours with magnetic stirring. After the reaction, extract twice with anhydrous ether, remove the yellow oily substance in the lower layer, and dry it in vacuum at 70°C for 24 hours to obtain a yellow thick liquid 1-butyl-2-methyl-3-propionyl imidazolium bromide modifier; Take 1mmol of 1-butyl-2-methyl-3-propionyl imidazolium bromide, add 1mmol of carbonyldiimidazole, stir magnetically under the action of 2mL of anhydrous dimethyl sulfoxide, stop the reaction at room temperature for 6h, after the reaction is completed, The active body was obtained; 90 μL of the active body was added to 5 mL of a polyphenol oxidase solution with a concentration of 200 μM, and the reaction was stopped after 12 hours of magnetic stirr...

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Abstract

The invention provides a novel immobilized oxidordeuctase electrode and a preparation method thereof. The preparation method comprises the steps: firstly, a modifier is prepared from halogenated carboxylic acid and an imidazole compound; the modifier and carbonyldiiazole are added into anhydrous dimethylsulfoxide in a mixed mode, and a magnetic stirring reaction is conducted to obtain an active body; the active body is added into oxidordeuctase liquid to be dialysed to obtain an acylated oxidordeuctase solution; then immobilized oxidordeuctase powder is prepared from a zinc nitrate solution, 2-methylimidazole, the acylated oxidordeuctase solution and a polyvinylpyrrolidone mixed solution; and finally, the oxidordeuctase electrode is prepared. The characteristics of large specific surface area, more types, large loading capacity, satble structure and the like of materials of metal-organic frameworks (MOFs) are combined, by improving a preparation process, modified enzymes with monomersare embedded in the MOFs, thus it can be guaranteed that the activity of the immobilized enzymes is not obviously lowered, and the reuse stability of the enzymes can further be improved.

Description

technical field [0001] The invention relates to the preparation technology of an immobilized oxidoreductase electrode, in particular to a novel immobilized oxidoreductase electrode and a preparation method thereof. Background technique [0002] As a biocatalyst, enzymes have high catalytic efficiency under mild reaction conditions, and have high specificity and selectivity. They are widely used in biomedicine, environmental protection, energy, food and chemical industries. Oxidoreductase is a high-efficiency biocatalyst with special catalytic function under normal temperature and pressure conditions, and its catalytic efficiency is 10 times higher than that of ordinary catalysts. 7 -10 13 Times, and specificity, many chemical reactions can use redox enzymes as catalysts, and can avoid side reactions. It not only catalyzes chemical reactions in living organisms, but also catalyzes non-in vivo chemical reactions. However, due to the composition and structure of oxidoreducta...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/00C12N11/14C12N11/08
CPCC12Q1/001C12N11/14C12N11/08C12N9/0065C12N9/0061C12N9/0059C12Y111/01007C12Y110/03002C12Y110/03001
Inventor 邹彬王鹏云夏姣姣张黎明
Owner JIANGSU UNIV
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