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Preparation method and application of aptamer-functionalized horseradish peroxidase@metal-organic framework materials

A technology of transforming horseradish peroxide and organic skeleton, applied in biochemical equipment and methods, redox enzymes, chemical instruments and methods, etc., can solve problems such as difficult recycling, improve catalytic activity, and solve problems with low catalytic activity Effect

Active Publication Date: 2021-12-21
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the presence of hydrogen peroxide, it has a strong ability to catalyze the oxidation of phenol and its derivatives, while free horseradish peroxidase has the disadvantages of fragility and difficulty in recovery in the actual catalytic environment.

Method used

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  • Preparation method and application of aptamer-functionalized horseradish peroxidase@metal-organic framework materials
  • Preparation method and application of aptamer-functionalized horseradish peroxidase@metal-organic framework materials
  • Preparation method and application of aptamer-functionalized horseradish peroxidase@metal-organic framework materials

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] (1) SA&HRP@ZIF-8 / BIOTIN-P1 material preparation

[0044] Add SA solution (5mg / mL, 15μL) and HRP solution (2mg / mL, 60μL) to zinc nitrate hexahydrate solution (50mM, 250μL), mix the solution for 15min; then add 2-methylimidazole solution to the mixed solution (50mM, 250μL), mix well, put the mixture in a constant temperature water bath at 30°C and let it stand for 16h, centrifuge to collect the solid product and wash it with deionized water for 3 times; disperse the above obtained solid in P1 solution (100μM, 60μL) , centrifuged to separate the solid material, and washed three times with deionized water, and the final solid product was SA&HRP@ZIF-8 / BIOTIN-P1.

[0045] (2) P&HRP@ZIF-8 material preparation

[0046] Add HRP solution (2mg / mL, 60μL) and aptamer (P) solution (100μM, 60mL) to zinc nitrate hexahydrate solution (50mM, 250μL), mix and react for 15min; Imidazole solution (50mM, 250 μL), mixed well, put the mixed solution in a constant temperature water bath at 30°...

Embodiment 2

[0056] Selectivity experiment of SA&HRP@ZIF-8 / BIOTIN-P1

[0057] Bisphenol A (BPA), phenol and p-chlorophenol with a concentration of 0.07 mg / mL were prepared with NaAc-HAc (0.2M) buffer solution at pH 7.0, and an equal amount of SA&HRP@ZIF-8 was added to three solutions of the same volume / BIOTIN-P1 (100μL, 0.33mg / mL) and 100μL H 2 o 2 (0.3% w / w), after reacting at room temperature for 5 minutes, filter the reacted system with a filter membrane, and then measure the ultraviolet absorbance values ​​of the obtained liquids at a wavelength of 277 nm. Such as Image 6 It was shown that under the same reaction conditions, both HRP@ZIF-8 and SA&HRP@ZIF-8 / BIOTIN-P1 had certain degradation performance on BPA, phenol and p-chlorophenol. Comparing the changes of absorbance values ​​after the reaction of the three substrates, it was found that the degradation ability of SA&HRP@ZIF-8 / BIOTIN-P1 on BPA was significantly higher than that of phenol and p-chlorophenol.

Embodiment 3

[0059] Enzyme performance experiment of SA&HRP@ZIF-8 / BIOTIN-P1

[0060] Dilute the 20mg / mL BPA solution with pH7.0 NaAc-HAc (0.2M) buffer solution to a concentration of 0.29mM, 0.28mM, 0.26mM, 0.25mM, 0.23mM, take 500μL of the diluted BPA solution, and add 100μL SA&HRP@ZIF-8 / BIOTIN-P1(0.33mg / mL) and 100μL HO 2 o 2 (0.3%w / w) solution, make it fully react 5min, filter the mixed solution after the reaction with filter membrane, measure the ultraviolet absorbance value of gained liquid at 277nm place, determine the initial degradation rate of BPA by the ultraviolet absorbance of measuring solution, then Plot and fit kinetic curves; fit data using Michaelis-Menten equation to obtain V max and K m . The same experiment was performed with HRP@ZIF-8 in the control experiment. Such as Figure 7 As shown, in the control experiment, the K of HRP@ZIF-8 m The value is 18.2mM, while the K of SA&HRP@ZIF-8 / BIOTIN-P1 m The value is 6.7mM, indicating that aptamer P1 increases the effect...

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Abstract

The purpose of the present invention is to provide a preparation method and application of aptamer functionalized horseradish peroxidase@metal-organic framework material. First, aptamer-based MOFs immobilized enzyme materials were prepared for the efficient catalytic degradation of phenolic pollutants (bisphenol A) in the environment. Then, based on the principle of complementary base pairing, magnetic aptamer materials were used to achieve the recovery and recycling of aptamer-functionalized horseradish peroxidase@metal-organic framework materials. The aptamer functionalized enzyme@metal-organic framework material involved in the present invention utilizes the high-efficiency recognition characteristics of bisphenol A aptamers to achieve self-enrichment of bisphenol A. At the same time, the immobilized enzyme material is recovered by utilizing the magnetic aptamer material capable of base complementary pairing. It has the characteristics of high catalytic efficiency, convenient operation, and easy recovery of immobilized enzyme materials.

Description

technical field [0001] The invention belongs to the technical field of new materials, and relates to the preparation of aptamer functionalized horseradish peroxidase @ metal-organic framework material, and its self-enrichment of bisphenol A to improve the degradation efficiency, and at the same time, it automatically complements the magnetic control after the degradation is completed Strand hybridization and preferential binding of bisphenol A to realize aptamer-functionalized horseradish peroxidase@metal-organic framework material for magnetically controlled complete recovery and continuous recycling of bisphenol A degradation. Background technique [0002] Phenolic wastewater in the environment is one of the most harmful and widely polluting industrial wastewater in the world today, and is an important source of water pollution in the environment. Phenolic compounds are contained in the wastewater discharged from many industrial fields such as metallurgy, machinery manufac...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N11/14C02F3/34C02F1/72C02F101/34
CPCC12N11/14C12N9/0065C02F3/342C02F1/725C12Y111/01007C02F2101/345
Inventor 王蕾兰会玲韩娟王赟刘园园王宇
Owner JIANGSU UNIV
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