InVO4 nano material and application thereof as mimic enzyme

Abstract

According to the InVO4 nanometer material and the application of the InVO4 nanometer material as mimic enzyme, the InVO4 nanometer material with a hollow cubic structure is successfully prepared, and the InVO4 nanometer material shows excellent mimic peroxidase catalytic activity, can rapidly detect H2O2 and has huge application potential in the fields of immunoassay and the like. In addition, the preparation process of the material is simple, convenient and feasible, the cost is low, and good repeatability is achieved.

Description

Technical Field

[0001] The present invention belongs to the technical field of simulated enzyme products and specifically relates to an InVO 4 Nanomaterials and their applications as enzyme mimics. Background Art

[0002] Peroxidase (Horseradish peroxidase, referred to as HRP) is a type of oxidoreductase produced by microorganisms or plants. It is an enzyme that catalyzes the oxidation of substrates using hydrogen peroxide as an electron acceptor and is often used to detect hydrogen peroxide. However, as a kind of natural enzyme, peroxidase is essentially a protein with a supramolecular structure. Although it has catalytic activity to specific substrates under mild conditions, its activity is easily affected by factors such as temperature and pH, and it is easily inactivated or denatured under extreme conditions such as high temperature, strong acid, and strong alkali. At the same time, the extraction process of peroxidase is complicated, the cost is high, and the yield is low. Therefore, the development of materials with stable catalytic performance and good catalytic activity to replace natural peroxidase has received extensive attention from researchers.

[0003] Among the many enzyme-mimicking materials, nanomaterials are very similar to natural enzymes in terms of size, shape, surface charge and other properties. Therefore, nanomaterials have a large specific surface area and abundant active sites, providing an opportunity to study efficient artificial enzyme-mimicking activities. At the same time, nanomaterials are inexpensive to synthesize, easy to store, and have excellent heat resistance and acid and alkali resistance. These advantages have greatly promoted the widespread application of nanomaterials in the field of enzyme-mimicking. Since Gao et al. first reported Fe 3 O 4 Since magnetic nanoparticles have highly efficient peroxidase mimicking catalytic activity, researchers have conducted in-depth research on nanomaterials mimicking peroxidase. Among them, vanadate nanomaterials have attracted much attention from scientific researchers due to their highly efficient catalytic activity. The preparation method of vanadate nanomaterials is relatively simple. Most vanadate materials can be synthesized by hydrothermal method. Moreover, vanadate has various morphologies, and some morphologies have large specific surface areas, which makes it possible to provide more reactive sites. Summary of the invention

[0004] The object of the present invention is to provide an InVO 4 Nanomaterials and their applications as enzyme mimics. The present invention successfully prepared InVO with a hollow cubic structure. 4 Nanomaterials exhibit excellent peroxidase-like catalytic activity and can rapidly detect H 2 O 2 , has great application potential in the fields of immunoassay, etc. In addition, the preparation process of this material is simple, easy, low-cost, and has good repeatability.

[0005] The InVO provided by the present invention 4 The enzyme-mimicking material is InVO with a hollow cubic structure 4 , with a diameter of about 300nm.

[0006] Provided by InVO 4 The enzyme-mimicking material is prepared as follows:

[0007] 1-5 mmol InCl 3 ·4H 2 O was dissolved in ultrapure water to obtain solution A. At the same time, 1-5 mmol NH 4 VO 3 The solution B is dissolved in ultrapure water; then the solution B is added dropwise into the solution A under stirring to obtain a suspension, the pH value of the suspension is adjusted to 1-7, and then stirring is continued for 30-60 minutes; after the stirring is completed, the mixed solution is placed in a reactor and heat-treated at 120-200°C for 12-48 hours; after the reaction is completed, the mixed solution is cooled to room temperature, filtered, washed, and dried at 40-80°C for 4-12 hours to obtain the enzyme simulation material.

[0008] The InCl 3 ·4H 2 O and NH 3 VO 4 The molar ratio is 1:1~2.

[0009] The pH value of the suspension is adjusted by using NH 3 ·H 2 O or NaOH solution.

[0010] The heat treatment conditions are as follows: temperature is 120-200° C., and time is 12-24 hours.

[0011] InVO of the present invention 4 The enzyme-mimicking material was used to catalyze the oxidation of the substrate 3,3',5,5'-tetramethylbenzidine (TMB) to achieve H 2 O 2 Rapid detection.

[0012] InVO of the present invention 4 The detection method of the liquid with the simulated enzyme material is as follows: phosphate buffered saline (PBS), the solution to be detected, the ethanol solution of TMB and InVO are added to the centrifuge tube in sequence. 4 The enzyme dispersion was simulated, and the color change of the solution was observed after 7 minutes of reaction, and the UV-visible absorption spectrum was recorded at 400-800 nm.

[0013] The beneficial effects of the present invention are:

[0014] 1) The present invention prepares InVO by a simple hydrothermal synthesis method 4 The enzyme-mimicking material has a simple preparation process, is easy to control and has low cost.

[0015] 2) InVO prepared by the present invention 4 The enzyme-mimicking material has a hollow cubic structure, a large specific surface area, and is easy to recycle and reuse.

[0016] 3) InVO prepared by the present invention 4 The enzyme-mimicking material has good catalytic performance of mimicking peroxidase and can quickly detect H 2 O 2 , and has good stability and reusability, and has potential application prospects in fields such as immunoassay. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 InVO prepared in Example 1 of the present invention 4 XRD patterns of enzyme-mimicking materials.

[0018] Figure 2 InVO prepared in Example 1 of the present invention 4 SEM photo of enzyme-mimicking material.

[0019] Figure 3 InVO prepared in Example 1 of the present invention 4 UV-visible absorption spectrum of the peroxidase reaction system simulated by the enzyme-mimicking material.

[0020] Figure 4 InVO prepared in Example 1 of the present invention 4 The absorbance value of the enzyme-mimicking material after 10 repeated enzyme-mimicking reactions. DETAILED DESCRIPTION

[0021] The present invention is further described below through specific examples, which are helpful for those skilled in the art to more fully understand the present invention, but are not intended to limit the present invention in any way.

[0022] Example 1: Preparation of hollow cubic InVO 4

[0023] 4.0 mmol InCl 3 ·4H 2 O was dissolved in 30 mL of ultrapure water and stirred magnetically to obtain solution A. At the same time, 4.0 mmol NH 4 VO 3Dissolve in 30 mL of ultrapure water and stir magnetically to dissolve to obtain solution B. Then, add the solution B dropwise into solution A under magnetic stirring to obtain a suspension. Then, add 1.0 mol / L NH 3 ·H 2 O solution to adjust the pH of the suspension to 4, and then continue stirring for 60 min.

[0024] After stirring, the suspension was transferred to a high-pressure reactor equipped with a polytetrafluoroethylene liner and placed in an electric constant temperature blast drying oven for heat treatment at 180°C for 24 hours. After the heat treatment, the reactor was cooled to room temperature, and the product was separated by suction filtration, washed with ultrapure water and anhydrous ethanol in turn, and then dried at 60°C for 6 hours to obtain InVO with a hollow cubic structure. 4 Nanomaterials.

[0025] Figure 1 This is the XRD pattern of the sample prepared in Example 1. As can be seen from the figure, the positions of all diffraction peaks are consistent with those of the orthorhombic phase InVO 4 The results are completely consistent with the standard card JCPDS No.48-0898, and no impurity phase appears. It can be determined that the sample prepared in Example 1 is pure InVO 4 In addition, it can be seen from the figure that the diffraction peak intensity of the sample is relatively large and the diffraction peak is relatively sharp, indicating that the prepared InVO 4 Has good crystallinity. Figure 2 This is a SEM photo of the sample prepared in Example 1. It can be seen from the figure that the prepared InVO 4 It is a hollow cubic structure with a size of about 300nm, and this structure has a large specific surface area.

[0026] Example 2: InVO 4 Simulated peroxidase

[0027] Take a 1.5mL centrifuge tube and add 400μL 50mmol / L phosphate buffer (PBS, pH=4.0), 100μL 20mmol / L H 2 O 2 solution, 100 μL of 8 mmol / L TMB ethanol solution and 330 μL of ultrapure water, and then 70 μL of InVO prepared in Example 1 was added. 4 dispersion (0.1 mg / mL), and after 7 min of reaction, observe the changes in the solution and record the UV-visible absorption spectrum at 400-800 nm ( Figure 3 ).

[0028] Figure 3 The following are the UV-visible absorption spectra of the four reaction systems. 652nm is the characteristic absorption peak of oxidized TMB.2 O 2 +InVO 4 、TMB+InVO 4 and TMB+H 2 O 2 The three systems have almost no absorption peaks, while TMB+H 2 O 2 +InVO 4 The system has an obvious absorption peak at 652nm with a good peak shape. 2 O 2 +InVO 4 、TMB+InVO 4 and TMB+H 2 O 2 The colors of the three systems were found to be colorless and transparent, with no significant changes, while TMB+H 2 O 2 +InVO 4 The solution in the system showed obvious blue color, indicating that InVO 4 In H 2 O 2 The blue oxide was generated by catalytic oxidation of TMB in the presence of . 4 The material has excellent catalytic activity of simulated peroxidase and can be regarded as a simulated enzyme material. 2 O 2 Under the condition of catalytic oxidation, TMB can be generated into blue oxidation product. Through this color reaction, H 2 O 2 , and has the potential to be widely used in fields such as biological immunoassay and environmental testing.

[0029] Example 3: InVO 4 Repeated application to simulate peroxidase

[0030] The InVO used in Example 2 4 The material was recovered by centrifugation, and then washed several times with ultrapure water and anhydrous ethanol, respectively, and dried at 60°C, and then subjected to a second round of simulated enzyme reaction according to the steps of Example 2, for a total of 10 rounds, while keeping other conditions unchanged. Figure 4 The results show that after 10 consecutive enzyme simulation experiments, the absorbance value of the reaction system did not decrease significantly, and the solution still showed a distinct blue color, demonstrating that InVO 4 The enzyme-mimicking material has excellent recycling effect and stability.

Claims

1. An InVO4 enzyme-mimicking material, characterized in that: The preparation method of the InVO4 enzyme-mimicking material is as follows: 1-5 mmol InCl3·4H2O is dissolved in ultrapure water to obtain solution A; at the same time, 1-5 mmol NH4VO3 is dissolved in ultrapure water to obtain solution B; then the above solution B is added dropwise to the above solution A under stirring conditions to obtain a suspension, the pH value of the suspension is adjusted to 1-7, and then stirring is continued for 30-60 minutes; after the stirring is completed, the mixed solution is placed in a reactor and heat-treated at 120-200°C for 12-48 hours; after the reaction is completed, the mixed solution is cooled to room temperature, filtered and washed, and then dried at 40-80°C for 4-12 hours to obtain a simulated enzyme material.

2. The InVO4 enzyme mimicking material according to claim 1, characterized in that The molar ratio of InCl3·4H2O to NH3VO4 is 1:1-2.

3. The InVO4 enzyme mimicking material according to claim 1, characterized in that The pH value of the suspension is adjusted by using NH3·H2O or NaOH solution with a concentration of 1.0-5.0 mol / L.

4. The InVO4 enzyme mimetic material according to claim 1, characterized in that The heat treatment conditions are as follows: temperature is 120-200°C and time is 12-24h.

5. Use of the InVO4 enzyme-mimicking material according to claim 1 in the preparation of products for detecting H2O2.

6. A detection product for detecting H2O2, characterized in that: The product contains the InVO4 enzyme mimicking material according to claim 1.

7. The detection product according to claim 6, characterized in that the product contains a substrate 3,3',5,5'-tetramethylbenzidine TMB.

8. A method for detecting H2O2, characterized in that: The method is to use the detection product described in claim 7 for detection.

9. The method according to claim 8, characterized in that The method comprises the following steps: Phosphate buffer, the solution to be tested, TMB ethanol solution and InVO4 simulated enzyme dispersion were added to the centrifuge tube in sequence. After the reaction, the color change of the solution was observed and the UV-visible absorption spectrum at 400-800 nm was recorded.

10. The method according to claim 9, characterized in that The final concentration of InVO4 is 7 μg / mL; the final concentration of TMB is 0.8 mmol / L.

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