A core-shell structure Fe3O4@PDA@ZIF-67 peroxidase and a preparation method and application thereof

Abstract

The application relates to the field of nanomaterial structures and analysis sensing technology, and discloses a core-shell structure Fe3O4@PDA@ZIF-67 peroxidase, a preparation method and application thereof. The core-shell structure Fe3O4 nanoparticle is used as a core, the surface of the Fe3O4 nanoparticle is modified by dopamine to form a polydopamine layer, then ZIF-67 is modified on the surface of the Fe3O4@PDA as a shell, and finally the prepared Fe3O4@PDA@ZIF-67 is heat-treated, so that a stable structure is obtained. In the Fe3O4@PDA@ZIF-67 composite material, the ZIF-67 as the shell can adsorb a signal molecule TMB to form a confined reaction, reduce the loss of active oxygen in the catalytic process, improve the sensitivity of the peroxidase-like enzyme applied to hydrogen peroxide sensing analysis, and has certain guiding significance for the structure design of the peroxidase-like enzyme material.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of nanomaterial structure and analysis sensing technology, in particular to a core-shell structure Fe3O4@PDA@ZIF-67 peroxidase-like enzyme and a preparation method and application thereof. BACKGROUND

[0002] In the chemical reactions throughout the life cycle, oxygen undergoes various reductions, which helps to generate reactive oxygen species (ROS), including O 2- , ·OH, H2O2, etc. As one of the reactive oxygen species, H2O2 exists in the metabolic process of the human body, and plays a crucial role in signal transduction, cell metabolism and body immunity. Abnormal expression of H2O2 can destroy the intracellular homeostasis, leading to oxidative stress, and seriously damaging the structure and function of cell macromolecules. Therefore, oxidative stress is considered to be a common cause of many life-threatening diseases, including atherosclerosis, chronic fatigue syndrome, Parkinson's disease, diabetes, cancer and acute myocardial infarction. However, the current hydrogen peroxide detection methods generally have the defects of low sensitivity and poor accuracy, so it is of great significance to establish a rapid and quantitative method for detecting hydrogen peroxide.

[0003] Using horseradish peroxidase to catalyze hydrogen peroxide to produce reactive oxygen species, and further oxidizing the signal molecule to obtain the oxide of the signal molecule, the level of hydrogen peroxide can be evaluated. However, the natural enzyme has high cost, is unstable and cannot be reused, which limits its wide application. Compared with natural enzyme (HRP), nanoenzyme has more advantages: stable structure in extreme environment, simple synthesis process, low cost, and diversity of structure and morphology. In recent years, some peroxidase-like enzymes have been applied to the detection of hydrogen peroxide. However, due to the lower catalytic activity of nanoenzyme than HRP, the sensitivity of hydrogen peroxide detection based on nanoenzyme still needs to be improved.

[0004] As one of the top ten new technologies in 2019, MOFs are a kind of hybrid materials obtained by coordination of organic ligands and metal ions. The ligands and metal ions combine to show directional arrangement, which is reflected in the different framework voids of MOFs. The high uniform size and pore structure can provide larger specific surface area and more active sites, further promoting the catalytic reaction. Due to the alternating arrangement of organic units and metal particles of MOFs materials, the high utilization rate of metal centers as catalytically active sites is ensured; high porosity and large specific surface area are beneficial to efficient aggregation and mass transfer of target substances, which can effectively improve the detection sensitivity and further detect the substances. Therefore, by utilizing the adsorption performance of MOFs, the utilization rate of active oxygen generated in the catalytic process of enzyme-like enzyme is improved, and a peroxidase-like enzyme material with core-shell structure is developed, which is applied to the sensitive detection of H2O2. SUMMARY

[0005] The application provides a core-shell structure Fe3O4@PDA@ZIF-67 peroxidase and a preparation method and application thereof, and aims at problems in the prior art.

[0006] The application provides a core-shell structure Fe3O4@PDA@ZIF-67 peroxidase.

[0007] Further, the Fe3O4@PDA@ZIF-67 peroxidase is a three-layer core-shell structure.

[0008] The application further provides a preparation method of the core-shell structure Fe3O4@PDA@ZIF-67 peroxidase.

[0009] S1, Fe3O4 nanoparticles are prepared by a hydrothermal method, the Fe3O4 nanoparticles are dispersed in deionized water, hydrochloric acid dopamine and tris(hydroxymethyl) aminomethane are added to obtain a mixed solution, and stirring, washing and vacuum drying are carried out to obtain Fe3O4@PDA.

[0010] S2, the Fe3O4@PDA is dissolved in a methanol solution containing cobalt nitrate to obtain solution A; 2-methyl imidazole is dissolved in methanol to obtain solution B; the solution A is quickly added to the solution B, mechanical stirring is carried out, and magnetic separation is carried out after standing at room temperature for 20-26 h, then washing is carried out with methanol, vacuum drying is carried out, and carbonization is carried out to obtain Fe3O4@PDA@ZIF-67.

[0011] Further, in S2, the molar ratio of the cobalt nitrate to the 2-methyl imidazole is 1:5-1:10.

[0012] Further, in S2, the specific conditions of the carbonization are as follows:

[0013] The temperature is raised to 700-1100 DEG C at a temperature rising speed of 5 DEG C per minute under N2 atmosphere, and carbonization is carried out for 1.5-3 h.

[0014] Further, in S1, the mass ratio of the Fe3O4 nanoparticles, dopamine hydrochloride and tris(hydroxymethyl)aminomethane is 1:20:25.

[0015] Further, in S1, the pH value of the mixed solution is 8-10.

[0016] The application further provides an application of the core-shell structure Fe3O4@PDA@ZIF-67 peroxidase-like material in detection of hydrogen peroxide.

[0017] Theoretical explanation: the Fe3O4@PDA@ZIF-67 is prepared by using Fe3O4 nanoparticles as a core, polydopamine (PDA) as an intermediate layer and ZIF-67 as a shell by means of a layer-by-layer growth method. Since the peroxidase-like enzyme usually has higher catalytic activity under acidic conditions, but the MOF is not stable under acidic conditions, the Fe3O4@PDA@ZIF-67 is further subjected to high-temperature carbonization heat treatment to obtain a stable structure, and TMB is used as a signal molecule for sensitive detection of H2O2. The Fe3O4 nanoparticles have the advantages of stable structure performance, easy-to-control chemical and physical properties, good biocompatibility and the like of natural enzymes; the ZIF-67 is a material with good adsorption, and the porous structure therein is conducive to adsorption, loading and separation of target objects, the organic ligand therein has good optical, electrical and thermal properties and rich chemical modification functional groups, and the presence of the metal node therein provides more active sites for catalysis, thereby realizing high-sensitivity detection; the polydopamine (PDA) is an intermediate layer, which can avoid reduction of the peroxidase-like activity of the core due to direct covering of the core by the shell.

[0018] Advantages: compared with the prior art, the application has the following specific advantages:

[0019] (1) The core-shell structure material prepared by the application has a large specific surface area, which can reduce disordered accumulation of the material, increase the contact area with a substrate, and enhance the peroxidase-like activity of the core-shell structure material.

[0020] (2) The Fe3O4@PDA@ZIF-67 composite material prepared by the application has good peroxidase activity, and is applied to an H2O2-TMB colorimetric sensing system: in the system, the ZIF-67 as a shell can adsorb the signal molecule TMB to form a confined reaction, reduce loss of active oxygen in the catalytic process, and improve the sensitivity of the peroxidase-like enzyme applied to hydrogen peroxide sensing analysis, which has certain guiding significance for structural design of the peroxidase-like enzyme material. BRIEF DESCRIPTION OF DRAWINGS

[0021] Figure 1Structure schematic diagram of core-shell structure Fe3O4@PDA@ZIF-67 material;

[0022] Figure 2 TEM diagram of synthesis of core-shell structure Fe3O4@PDA@ZIF-67;

[0023] Figure 3 Peroxidase-like activity feasibility analysis of core-shell structure Fe3O4@PDA@ZIF-67: wherein, Figure 3 (a) represents the absorbance of the core-shell material without TMB, Figure 3 (b) represents the absorbance without core-shell material, Figure 3 (c) represents the absorbance of the core-shell material without hydrogen peroxide, Figure 3 (d) is the absorbance of the core-shell material in the presence of hydrogen peroxide and TMB, Figure 3 The upper right corner of the drawing corresponds to the change of the solution color under the conditions of (a) (b) (c) (d) respectively.

[0024] Figure 4 Absorbance value of colorimetric detection of hydrogen peroxide by core-shell structure Fe3O4@PDA@ZIF-67 with concentration change diagram: wherein, Figure 4 (a) is the diagram of the change of absorbance with concentration, Figure 4 (b) is the standard curve diagram of hydrogen peroxide detection, Figure 4 The upper right corner of the drawing corresponds to the change of the solution color under the conditions of (a) (b) (c) (d) respectively. DETAILED DESCRIPTION

[0025] The present application will be described in detail below in conjunction with the embodiments.

[0026] Embodiment 1:

[0027] The present embodiment provides a preparation method of core-shell structure Fe3O4@PDA@ZIF-67, and the specific steps are as follows:

[0028] First, 0.002 mol of FeCl3·6H2O and 0.0003 mol of sodium citrate were completely dissolved in 10 ml of ethylene glycol, 0.029 mol of anhydrous sodium acetate was completely dissolved in 10 ml of ethylene glycol, then the two were mixed, stirred for 10 min, transferred to a 25 ml reaction kettle, reacted at 190°C for 10 hours, and naturally cooled. Then washed with ethanol, water, and freeze-dried to obtain Fe3O4 nanoparticles. The Fe3O4 nanoparticles were configured into a Fe3O4 aqueous solution, 0.0005 mol of hydrochloric acid dopamine and 0.001 mol of tris (hydroxymethyl) aminomethane (Tris) were added to the Fe3O4 aqueous solution of 0.05 g / L, and the pH of the mixed solution was adjusted to 8. After mechanical stirring, the precipitate was washed with deionized water, vacuum dried to obtain Fe3O4@PDA; 50 mg of Fe3O4@PDA was dissolved in a methanol solution containing cobalt nitrate to obtain solution A, 2-methyl imidazole was dissolved in methanol to obtain solution B, and the molar ratio of cobalt nitrate to 2-methyl imidazole was 1:8. Solution A was quickly added to solution B, mechanically stirred, and placed at room temperature for 24 h, then magnetically separated, washed with methanol, vacuum dried, and carbonized at 1000°C for 2 h to obtain Fe3O4@PDA@ZIF-67.

[0029] Embodiment 2:

[0030] The embodiment provides a preparation method of a core-shell structure Fe3O4@PDA@ZIF-67, and the specific steps are as follows:

[0031] First, 0.002 mol of FeCl3·6H2O and 0.0003 mol of sodium citrate were completely dissolved in 10 ml of ethylene glycol, 0.029 mol of anhydrous sodium acetate was completely dissolved in 10 ml of ethylene glycol, then the two were mixed, stirred for 10 min, transferred to a 25 ml reaction kettle, reacted at 200°C for 10 hours, and naturally cooled. Then washed with ethanol, water, and freeze-dried to obtain Fe3O4 nanoparticles. The Fe3O4 nanoparticles were configured into a Fe3O4 aqueous solution, 0.0005 mol of hydrochloric acid dopamine and 0.001 mol of tris (hydroxymethyl) aminomethane (Tris) were added to the 0.05 g / L Fe3O4 aqueous solution, the pH of the mixed solution was adjusted to 8.5, and after mechanical stirring, the precipitate was washed with deionized water and vacuum dried to obtain Fe3O4@PDA; 50 mg of Fe3O4@PDA was dissolved in a methanol solution containing cobalt nitrate to obtain solution A, 2-methyl imidazole was dissolved in methanol to obtain solution B, and the molar ratio of cobalt nitrate to 2-methyl imidazole was 1:9. Solution A was quickly added to solution B, mechanically stirred, and placed at room temperature for 24 h before magnetic separation, washed with methanol, and vacuum dried. After carbonization at 900°C for 2 h, Fe3O4@PDA@ZIF-67 was finally obtained.

[0032] Embodiment 3:

[0033] The embodiment provides a preparation method of a core-shell structure Fe3O4@PDA@ZIF-67, and the specific steps are as follows:

[0034] First, 0.002 mol of FeCl3·6H2O and 0.0003 mol of sodium citrate were completely dissolved in 10 ml of ethylene glycol, 0.029 mol of anhydrous sodium acetate was completely dissolved in 10 ml of ethylene glycol, then the two were mixed, stirred for 10 min, transferred to a 25 ml reaction kettle, reacted at 220°C for 9 hours, and then naturally cooled. Then washed with ethanol, water, and freeze-dried to obtain Fe3O4 nanoparticles. The Fe3O4 nanoparticles were configured into a Fe3O4 aqueous solution, 0.0005 mol of hydrochloric acid dopamine and 0.001 mol of tris were added to the Fe3O4 aqueous solution of 0.05 g / L, and the pH of the mixed solution was adjusted to 9. After mechanical stirring, the precipitate was washed with deionized water and vacuum dried to obtain Fe3O4@PDA; 50 mg of Fe3O4@PDA was dissolved in a methanol solution containing cobalt nitrate to obtain solution A, 2-methyl imidazole was dissolved in methanol to obtain solution B, and the molar ratio of cobalt nitrate to 2-methyl imidazole was 1:10. Solution A was quickly added to solution B, mechanically stirred, and then placed at room temperature for 24 h before magnetic separation. After washing with methanol and vacuum drying, carbonization was carried out at 1100°C for 2 h, and finally Fe3O4@PDA@ZIF-67 was obtained.

[0035] Embodiment 4:

[0036] The peroxidase activity of the Fe3O4@PDA@ZIF-67 magnetic nanoscale enzyme prepared in the application was determined by TMB color reaction:

[0037] The peroxidase activity of Fe3O4@PDA@ZIF-67 was characterized by studying the oxidation ability of Fe3O4@PDA@ZIF-67 to TMB in the presence of H2O2. The experiment monitored the ultraviolet absorbance of TMB at 654 nm. TMB alcohol solution, H2O2 solution and Fe3O4@PDA@ZIF-67 solution were added to NaAc-HAc buffer solution (0.2 mol / L, pH=3.5), and reacted at room temperature for 10-20 min. Then the absorbance of the sample at 654 nm was measured, and each sample was measured in triplicate.

[0038] The test results are shown in Figure 3 As shown in Figure 3 (d), Fe3O4@PDA@ZIF-67 has the strongest absorption peak at 654 nm in the presence of H2O2 and TMB, indicating that Fe3O4@PDA@ZIF-67 has peroxidase-like properties and can trigger the oxidation of TMB. In contrast, Fe3O4@PDA@ZIF-67 has lower catalytic activity in the absence of H2O2 (as shown in Figure 3 (c).

[0039] Embodiment 5:

[0040] The peroxidase activity of Fe3O4@PDA@ZIF-67 prepared by the application was detected by hydrogen peroxide:

[0041] The alcohol solution of TMB was added to the NaAc-HAc buffer solution, and then different concentrations of H2O2 solution and alcohol solution of Fe3O4@PDA@ZIF-67 were added, and vortexed to mix thoroughly. After incubation at a certain temperature for 10-20 min, the absorbance value at 654 nm was measured in the enzyme marker. The concentration of hydrogen peroxide can be obtained by substituting the absorbance value into the established absorbance-concentration curve. Hydrogen peroxide is oxidized to blue TMB-ox by hydroxyl radicals generated under the catalysis of Fe3O4@PDA@ZIF-67 nanoscale enzyme. In theory, the higher the concentration of H2O2, the higher the absorbance value. The hydrogen peroxide concentration for constructing the standard curve for detecting hydrogen peroxide was 0, 0.4, 0.5, 1, 2, 5, 10, 30, 50, 60, and 100 mM.

[0042] The results are shown in Figure 4 Figure 4 (a) It can be observed that the absorbance detected by the enzyme marker is higher as the concentration of hydrogen peroxide increases, and the inset can also indicate that the color gradually deepens as the concentration of hydrogen peroxide increases; Figure 4 (b) It shows that both have a good linear relationship within the concentration range, and the linear correlation coefficient is 0.9515.

[0043] The above embodiments are only for illustrating the technical concept and characteristics of the application, and the purpose is to enable those skilled in the art to understand the content of the application and implement it, and cannot limit the protection scope of the application. Any equivalent transformation or modification made according to the spirit and essence of the application should be covered within the protection scope of the application.​

Claims

1. The use of a core-shell structure Fe3O4@PDA@ZIF-67 mimetic peroxidase, characterized in that, The core-shell structure Fe3O4@PDA@ZIF-67 peroxidase is used for detecting hydrogen peroxide, and the Fe3O4@PDA@ZIF-67 peroxidase takes Fe3O4 nanoparticles as a core, takes polydopamine as an intermediate layer, and takes ZIF-67 as a shell. 2.The application of the core-shell structure Fe3O4@ PDA@ ZIF-67 peroxidase according to claim 1, wherein: The Fe3O4@PDA@ZIF-67 peroxidase is a three-layer core-shell structure. 3.The application of the core-shell structure Fe3O4@ PDA@ ZIF-67 peroxidase-like enzyme according to claim 1, characterized in that, The specific preparation method of the core-shell structure Fe3O4@PDA@ZIF-67 peroxidase is as follows: S1, Fe3O4 nanoparticles are prepared by a hydrothermal method, the Fe3O4 nanoparticles are dispersed in deionized water, hydrochloric acid dopamine and tris(hydroxymethyl) aminomethane are added to obtain a mixed solution, stirring, washing, and vacuum drying are performed to obtain Fe3O4@PDA; S2, the Fe3O4@PDA is dissolved in a methanol solution containing cobalt nitrate to obtain solution A; 2-methyl imidazole is dissolved in methanol to obtain solution B; the solution A is quickly added to the solution B, mechanical stirring is performed, and magnetic separation is performed after being placed at room temperature for 20-26 h, then methanol is used for washing, vacuum drying, carbonization, and Fe3O4@PDA@ZIF-67 is obtained.

4. The use of the core-shell structure Fe3O4@PDA@ZIF-67 peroxidase-like enzyme according to claim 3, characterized in that: In S2, the molar ratio of the cobalt nitrate to the 2-methyl imidazole is 1:5-1:

10.

5. The use of the core-shell structure Fe3O4@PDA@ZIF-67 peroxidase-like enzyme according to claim 3, characterized in that: In S2, the specific conditions of the carbonization are as follows: The temperature is raised to 700-1100℃ at a temperature rising speed of 5℃ per minute under N2 atmosphere, and carbonization is performed for 1.5-3 h.

6. The use of the core-shell structure Fe3O4@PDA@ZIF-67 peroxidase-like enzyme according to claim 3, characterized in that: In S1, the mass ratio of the Fe3O4 nanoparticles, the hydrochloric acid dopamine, and the tris(hydroxymethyl) aminomethane is 1:20:

25.

7. The use of the core-shell structure Fe3O4@PDA@ZIF-67 peroxidase-like enzyme according to claim 3, characterized in that: In S1, the pH value of the mixed solution is 8-10.

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