A silver-based metal-organic framework nanoscale enzyme with multi-enzyme-like activity loaded with silver nanoparticles and preparation and application thereof

Abstract

The application discloses a silver-based metal organic framework nano-enzyme loaded with silver nanoparticles and having multi-enzyme-like activity, and a preparation and application thereof. The application uses 2-methyl imidazole as an organic ligand, triethylamine as a deprotonation reagent and a basic environment provider, methanol as a solvent and a silver ion reducing agent, and obtains a silver-based metal organic framework by using the coordination of the deprotonated 2-methyl imidazole and the silver ion. Meanwhile, the silver ion adsorbed on the surface of the silver-based metal organic framework is reduced into silver nanoparticles by using methanol under the alkaline condition, and a silver-based metal organic framework nano-enzyme loaded with silver nanoparticles is prepared. The nano-enzyme can exhibit multi-enzyme-like activities such as oxide enzyme-like activity, peroxide enzyme-like activity and laccase enzyme-like activity under different conditions, and can realize the colorimetric detection of a plurality of trace analysis objects in different types of complex samples by using different enzyme-like activities under different detection conditions.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of synthesis and application of nanomaterials, and particularly relates to a silver-based metal organic framework nanomolecule loaded with silver nanoparticles with multi-enzyme-like activity and a preparation and application thereof. BACKGROUND

[0002] Metal organic frameworks (MOFs) are a kind of crystalline porous inorganic-organic hybrid materials with periodic network structure, which are formed by self-assembly of organic ligands and metal ions as secondary structural units. Due to the outstanding design flexibility, excellent surface area and diversity of topological structure, MOFs exhibit a wide range of potential applications in gas adsorption / separation, catalysis, sensing, chromatographic separation and energy storage, etc. At present, the research focus of MOFs has shifted from early synthesis to optimization of performance and expansion of application scenarios. In the field of nanomolecule research, MOFs exhibit incomparable advantages over other traditional materials due to their unique properties. Compared with natural enzymes, MOFs can not only provide more active sites for nanomolecules, endowing them with excellent enzyme-like catalytic activity, but also have the advantages of low cost, simple preparation, high stability and durability.

[0003] Most of the reported MOF nanomolecules only have single enzyme-like activity, while MOF materials with multi-enzyme-like activity are rarely reported, and their preparation methods are complex and require high conditions. Compared with single enzyme-like nanomolecules, multi-enzyme-like nanomolecules have many excellent properties, including synergistic effect, cascade reaction and different response activities in different environments, which make them have a wider application in practice. Based on this, the present application uses 2-methyl imidazole as an organic ligand, triethylamine as a deprotonation reagent and provides an alkaline environment, methanol as a solvent and a reducing agent for silver ions, and uses the coordination between deprotonated 2-methyl imidazole and silver ions to obtain a silver-based metal organic framework. At the same time, under alkaline conditions, methanol is used to reduce silver ions adsorbed on the surface of the silver-based metal organic framework into silver nanoparticles, and a silver-based metal organic framework nanomolecule loaded with silver nanoparticles is prepared. The preparation method of the present application is simple and easy to operate, and the synthesis conditions are mild. It can be synthesized directly at room temperature without high temperature and pressure, and does not require expensive instruments. The nanomolecule can exhibit three different enzyme-like activities, namely, oxidation enzyme, peroxidase and laccase, and can realize simple, rapid and low-cost multi-target colorimetric detection in different response scenarios, which has very important application value in the fields of biological analysis and colorimetric sensing. SUMMARY

[0004] The purpose of the present application is to provide a silver nanoparticle loaded silver-based metal organic framework (AgNPs@Ag-MOF) nanoscale enzyme with multi-enzyme activity, and to realize colorimetric detection application in different application scenarios. The present application improves the coordination bond strength between silver ions and nitrogen atoms on the metal organic framework by deprotonation, and realizes the preparation of AgNPs@Ag-MOF nanoscale enzyme by reducing silver ions in situ on the silver-based metal organic framework through the reducing agent methanol under alkaline conditions. On the one hand, the imidazole ring of 2-methyl imidazole has deprotonizable ammonia nitrogen, and the protons on the ammonia nitrogen are removed by the proton extraction reaction of triethylamine and 2-methyl imidazole to make it carry a negative charge, thereby enhancing the coordination ability of silver ions and ammonia nitrogen. At the same time, 2-methyl imidazole is a molecule with a π-π conjugated system, and changing the electron distribution in the conjugated system can enhance the coordination ability of the nitrogen atom. The present application uses a solvent with weak polarity (methanol) to reduce the attraction of the solvent to the electrons around the nitrogen atom, thereby increasing the electron density of the nitrogen atom on the imidazole ring, making the nitrogen atom more easily coordinate with silver ions in Lewis base-Lewis acid coordination, enhancing the coordination ability between 2-methyl imidazole and silver ions, and improving the enzyme-like activity of the material. On the other hand, methanol will generate alkoxide under alkaline conditions, and act as a reducing agent for silver ions, reducing silver ions in situ on the silver-based metal organic framework to generate silver nanoparticles; the silver nanoparticles generated in situ on the silver-based metal organic framework further improve the enzyme-like activity of the material. AgNPs@Ag-MOF nanoscale enzyme can directly oxidize the substrate 3,3',5,5'-tetramethyl benzidine (TMB) to the blue oxidation product oxTMB, exhibiting oxidase-like activity; can oxidize the substrate 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) to the green product ABTS·+ in the presence of hydrogen peroxide (H2O2), exhibiting peroxidase activity; can also catalyze phenolic substances 2,4-dichlorophenol (2,4-DCP) to 2,4-dichlorobenzoquinone, and form a pink coupling product with 4-aminoantipyrine (4-AAP), exhibiting laccase-like activity.

[0005] The present application proposes a mild preparation method of silver nanoparticle loaded silver-based metal organic framework nanoscale enzyme, and exhibits multi-enzyme activity of the new nanoscale enzyme, further developing the application of metal organic framework materials in the fields of biological analysis and colorimetric detection.

[0006] To achieve the above purpose, the present application adopts the following technical solutions:

[0007] The preparation method of the silver nanoparticle loaded silver-based metal organic framework nanoscale enzyme comprises the following steps:

[0008] (1) Preparation of deprotonated 2-methylimidazole methanol solution: 2-methylimidazole was used as the organic ligand, methanol was used as the solvent (the solvent methanol was also used as the reducing agent of silver ions in the next step), and triethylamine was used as the deprotonating agent and provided the basic environment required for the reduction reaction.

[0009] (2) Preparation of silver nitrate aqueous solution with the same concentration as the deprotonated 2-methylimidazole methanol solution, and then slowly drop the same amount of silver nitrate aqueous solution into the deprotonated 2-methylimidazole methanol solution, continuously stirring until the addition is complete, and then reacting at room temperature for 2-24 hours in the dark to obtain a uniform milky white suspension; then centrifugation, the obtained solid product was washed with water and ethanol for 3 times respectively, and dried in a vacuum drying oven at 60°C for 24 hours to obtain AgNPs@Ag-MOF nanoscale enzyme.

[0010] Multi-enzyme activity display of silver nanoparticle-loaded silver-based metal organic framework nanoscale enzyme:

[0011] (1) Oxidase-like activity: AgNPs@Ag-MOF was used as nanoscale enzyme, and color developing substrate TMB was added, and the solution changed from colorless to blue, and combined with UV-Vis absorption spectrum, the oxidase-like activity was exhibited. The specific steps are as follows: 50 μL of AgNPs@Ag-MOF dispersion (final concentration 25 μg / mL) and 50 μL of TMB solution (final concentration 0.5 mM) were added to 1900 μL of acetic acid-sodium acetate (HAc-NaAc) solution (12 mM, pH 5.0), and mixed uniformly, and then reacted at 20°C for 20 minutes, while the TMB solution and AgNPs@Ag-MOF dispersion were used as control groups, and the absorbance value at wavelength 650 nm (characteristic peak position of oxidized TMB (oxTMB)) was recorded by UV-Vis absorption spectrometer. The results showed that the control group solutions were not blue, only when AgNPs@Ag-MOF and TMB were present at the same time, the solution turned blue, and a characteristic absorption peak appeared at 650 nm, which proved that AgNPs@Ag-MOF nanoscale enzyme had oxidase-like activity.

[0012] (2) Peroxidase-like activity: In the presence of H2O2, AgNPs@Ag-MOF as nanoscale enzyme, the colorless solution changed to green after adding the chromogenic substrate ABTS, combined with UV-vis absorption spectrum, showing its peroxidase-like activity. The specific steps are as follows: 100 μL AgNPs@Ag-MOF dispersion (final concentration 20 μg / mL), 50 μL ABTS solution (final concentration 0.5 mM) and 50 μL H2O2 solution (final concentration 0.1 mM) are added to 1800 μL HAc-NaAc solution (10 mM, pH 3.0), mixed uniformly, and reacted at 37.5°C for 40 minutes, while ABTS solution, H2O2 solution+ABTS solution, AgNPs@Ag-MOF dispersion+ABTS solution, H2O2 solution+AgNPs@Ag-MOF dispersion are used as control groups, and the absorbance value at 414 nm (characteristic peak position of ABTS·+) is recorded by UV-vis absorption spectrometer. It is found from the results that the control group solution does not change green, only when AgNPs@Ag-MOF, H2O2 and ABTS exist at the same time, the solution changes to green, and a characteristic absorption peak appears at 414 nm, which proves that AgNPs@Ag-MOF nanoscale enzyme has peroxidase-like activity.

[0013] (3) Laccase-like activity: AgNPs@Ag-MOF as nanoscale enzyme, after adding phenolic substance 2,4-DCP and chromogenic substrate 4-AAP, the colorless solution changed to pink, combined with UV-vis absorption spectrum, showing its laccase-like activity. The specific steps are as follows: 100 μL AgNPs@Ag-MOF dispersion (final concentration 250 μg / mL), 200 μL 2,4-DCP solution (final concentration 2.5 mM) and 200 μL 4-AAP solution (final concentration 0.6 mM) are added to 1500 μL 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES) solution (20 mM, pH 6.0), mixed uniformly, and reacted at 40°C for 30 minutes, and the supernatant was taken after centrifugation, while 2,4-DCP solution, 2,4-DCP solution+AgNPs@Ag-MOF dispersion, 4-AAP solution, 4-AAP solution+AgNPs@Ag-MOF dispersion are used as control groups, and the absorbance value at 510 nm (characteristic peak position of pink coupling product) is recorded by UV-vis absorption spectrometer. It is found from the results that the control group solution does not change pink, only when AgNPs@Ag-MOF, 2,4-DCP and 4-AAP exist at the same time, the solution changes to pink, and a characteristic absorption peak appears at 510 nm, which proves that AgNPs@Ag-MOF nanoscale enzyme has laccase-like activity.

[0014] The significant advantages of the present application are:

[0015] Compared with traditional nanocomposites, the silver-based metal organic framework nanozyme loaded with silver nanoparticles provided by the application is simple to prepare, low in cost, does not require expensive instruments, and is easy to popularize.

[0016] The silver-based metal organic framework nanozyme loaded with silver nanoparticles provided by the application can exhibit different enzyme-like activities in different substrates under different conditions in different use scenarios, so that the actual use scenarios of the nanozyme are no longer single.

[0017] The silver-based metal organic framework nanozyme loaded with silver nanoparticles provided by the application can be combined with colorimetric detection and other methods to realize its wide application in the fields of biological analysis and colorimetric sensing. BRIEF DESCRIPTION OF DRAWINGS

[0018] Figure 1 The transmission electron microscope morphology diagram of the silver-based metal organic framework nanozyme loaded with silver nanoparticles prepared in the application.

[0019] Figure 2 The high-angle annular dark field (HAADF) scanning image and the particle size distribution diagram of the silver-based metal organic framework nanozyme loaded with silver nanoparticles prepared in the application.

[0020] Figure 3 The X-ray photoelectron spectroscopy characterization diagram of the silver-based metal organic framework nanozyme loaded with silver nanoparticles prepared in the application.

[0021] Figure 4 The X-ray powder diffraction characterization diagram of the silver-based metal organic framework nanozyme loaded with silver nanoparticles prepared in the application.

[0022] Figure 5 The oxygen-like oxidase activity diagram of the silver-based metal organic framework nanozyme loaded with silver nanoparticles prepared in the application.

[0023] Figure 6 The peroxide-like oxidase activity diagram of the silver-based metal organic framework nanozyme loaded with silver nanoparticles prepared in the application.

[0024] Figure 7 The laccase-like oxidase activity diagram of the silver-based metal organic framework nanozyme loaded with silver nanoparticles prepared in the application.

[0025] Figure 8 The working curve of colorimetric detection of different concentrations of ascorbic acid (AA) standard solution by the silver-based metal organic framework nanozyme loaded with silver nanoparticles prepared in the application.

[0026] Figure 9is a working curve of the silver-based metal organic framework loaded with silver nanoparticles prepared by the present application as a nano-enzyme for colorimetric detection of different concentrations of H2O2 standard solution.

[0027] Figure 10 is a working curve of the silver-based metal organic framework loaded with silver nanoparticles prepared by the present application as a nano-enzyme for colorimetric detection of different concentrations of adrenaline standard solution. DETAILED DESCRIPTION

[0028] In order to make the content of the present application more convenient to understand, the technical solutions of the present application are described below in combination with specific embodiments. Obviously, the described embodiments are part of the embodiments of the present application, rather than all the embodiments.

[0029] Embodiment 1:

[0030] The preparation of the silver-based metal organic framework loaded with silver nanoparticles as a nano-enzyme includes the following steps:

[0031] (1) Preparation of deprotonated organic ligand solution: 1 mmol of triethylamine and 2 mmol of 2-methylimidazole are added to 10 mL of methanol, and ultrasonic dissolution is performed at room temperature to obtain a deprotonated organic ligand solution.

[0032] (2) Preparation of silver-based metal organic framework loaded with silver nanoparticles as a nano-enzyme: 2 mmol of silver nitrate is dissolved in 10 mL of double-distilled water, and the obtained silver nitrate aqueous solution is slowly added dropwise into the deprotonated organic ligand solution of step (1) while stirring until the addition is complete. Reaction is carried out at room temperature in the dark for 12 hours to obtain a uniform milky white suspension. The solid product is collected by centrifugation, and the solid product is washed with water and ethanol for 3 times respectively, and then dried in a vacuum drying oven at 60°C for 24 hours to obtain the silver-based metal organic framework loaded with silver nanoparticles (AgNPs@Ag-MOF) as a nano-enzyme.

[0033] The silver-based metal organic framework loaded with silver nanoparticles (AgNPs@Ag-MOF) as a nano-enzyme prepared according to the above steps is characterized by transmission electron microscopy (TEM) and X-ray photoelectron spectroscopy (XPS). Figure 1 The TEM morphology of the nano-enzyme can be seen from the figure, and a large number of silver nanoparticles are distributed in the nano-enzyme material. Figure 2 The figure shows Figure 1 The high-angle annular dark field (HAADF) scanning image of the morphology figure can more clearly observe the morphology of the silver nanoparticles, and Figure 2 The particle size distribution of the silver nanoparticles is also shown in the figure, and it can be found that the average particle size is 5.9 nm. The above results prove the successful synthesis of the AgNPs@Ag-MOF nano-enzyme. Figure 3This is the full-band XPS energy spectrum of the nanozyme material, from which the four element signals of C, N, O, and Ag can be clearly seen, which also proves the successful synthesis of AgNPs@Ag-MOF nanozyme.

[0034] Example 2:

[0035] The preparation of silver-based metal-organic framework nanozymes loaded with silver nanoparticles includes the following steps:

[0036] (1) Preparation of deprotonated organic ligand solution: 2 mmol of triethylamine and 2 mmol of 2-methylimidazole were added to 10 mL of methanol and dissolved under ultrasonication at room temperature to obtain a deprotonated organic ligand solution.

[0037] (2) Preparation of silver-based metal-organic framework nanozymes loaded with silver nanoparticles: 2 mmol of silver nitrate was dissolved in 10 mL of secondary water, and the resulting silver nitrate aqueous solution was slowly dripped into the deprotonated organic ligand solution of step (1) and continuously stirred until the addition was complete. The mixture was reacted in the dark at room temperature for 16 hours to obtain a uniform milky white suspension; the solid product was collected by centrifugation, and the solid product was washed three times with water and three times with ethanol, and then dried in a vacuum drying oven at 60°C for 24 hours to obtain silver-based metal-organic framework (AgNPs@Ag-MOF) nanozymes loaded with silver nanoparticles.

[0038] The silver nanoparticle-loaded silver-based metal organic framework (AgNPs@Ag-MOF) nanozyme prepared according to the above steps was characterized by X-ray powder diffraction (XRD). Figure 4 As shown, the main diffraction peaks show several very sharp peaks at 2θ = 16.90°, 22.45°, 28.00° and 28.70°, which are respectively related to (011), (200), The results show that the nanozyme has good crystallinity and is a typical polycrystalline material, which also proves the successful synthesis of AgNPs@Ag-MOF nanozyme.

[0039] Example 3:

[0040] (1) The AgNPs@Ag-MOF prepared in Example 1 was used as nanozyme to investigate the oxidase-like activity.

[0041] The experiment uses ultraviolet-visible absorption spectrometer as a detection instrument to prove that AgNPs@Ag-MOF as a nano-enzyme has oxidase-like activity. 50 μL of AgNPs@Ag-MOF dispersion liquid (final concentration of 25 μg / mL) and 50 μL of TMB solution (final concentration of 0.5 mM) are added to 1900 μL of acetic acid-sodium acetate (HAc-NaAc) solution (12 mM, pH 5.0), mixed uniformly, and reacted at 20°C for 20 minutes. Meanwhile, the TMB solution and the AgNPs@Ag-MOF dispersion liquid are used as control groups, and the absorbance value at a wavelength of 650 nm is recorded. The results are shown in Figure 5 The control groups do not have characteristic absorption peaks at 650 nm. Only when AgNPs@Ag-MOF and TMB exist simultaneously, a characteristic absorption peak appears at a wavelength of 650 nm, proving that the AgNPs@Ag-MOF nano-enzyme has oxidase-like activity.

[0042] (2) AgNPs@Ag-MOF prepared in Example 1 is used as a nano-enzyme to investigate peroxidase-like activity.

[0043] The experiment uses ultraviolet-visible absorption spectrometer as a detection instrument to prove that AgNPs@Ag-MOF as a nano-enzyme has peroxidase-like activity. 100 μL of AgNPs@Ag-MOF dispersion liquid (final concentration of 20 μg / mL), 50 μL of ABTS solution (final concentration of 0.5 mM), and 50 μL of H2O2 solution (final concentration of 0.1 mM) are added to 1800 μL of HAc-NaAc solution (10 mM, pH 3.0), mixed uniformly, and reacted at 37.5°C for 40 minutes. Meanwhile, the ABTS solution, the H2O2 solution + ABTS solution, the AgNPs@Ag-MOF dispersion liquid + ABTS solution, and the H2O2 solution + AgNPs@Ag-MOF dispersion liquid are used as control groups, and the absorbance value at a wavelength of 414 nm is recorded. The results are shown in Figure 6 The control group H2O2 solution + ABTS solution has a small peak at 414 nm due to the thermal decomposition of hydrogen peroxide at 37.5°C to produce ABTS·+. Only when AgNPs@Ag-MOF is added, AgNPs@Ag-MOF, H2O2, and ABTS exist simultaneously, and a clear characteristic absorption peak appears at a wavelength of 414 nm, proving that the AgNPs@Ag-MOF nano-enzyme has oxidase-like activity.

[0044] (3) AgNPs@Ag-MOF prepared in Example 1 is used as a nano-enzyme to investigate laccase-like activity.

[0045] The experiment used a UV-visible absorption spectrometer as a detection instrument to prove that AgNPs@Ag-MOF as a nano-enzyme has laccase-like activity. 100 μL of AgNPs@Ag-MOF dispersion (final concentration 250 μg / mL), 200 μL of 2,4-DCP solution (final concentration 2.5 mM) and 200 μL of 4-AAP solution (final concentration 0.6 mM) were added to 1500 μL of 4-hydroxyethylpiperazine ethanesulfonic acid (HEPES) solution (20 mM, pH 6.0), mixed uniformly, and after reaction at 40°C for 30 minutes, the supernatant was centrifuged, and at the same time, 2,4-DCP solution, 2,4-DCP solution + AgNPs@Ag-MOF dispersion, 4-AAP solution, and 4-AAP solution + AgNPs@Ag-MOF dispersion were used as control groups, and the absorbance value at a wavelength of 510 nm was recorded. The results are shown in Figure 7 The control groups did not appear characteristic absorption peaks at 510 nm, and only when AgNPs@Ag-MOF, 2,4-DCP and 4-AAP were present at the same time, a characteristic absorption peak appeared at a wavelength of 510 nm, proving that AgNPs@Ag-MOF nano-enzyme has laccase-like activity.

[0046] Example 4:

[0047] AgNPs@Ag-MOF prepared in Example 1 was used as a nano-enzyme, and ascorbic acid (AA) with reducing property was added to the color developing system of the pseudo-oxidase, in which blue oxTMB was reduced to colorless TMB by AA, and by comparing the color changes produced by adding different concentrations of AA, the absorbance change difference was recorded to realize colorimetric detection of AA. The specific steps are as follows:

[0048] 50 μL of AgNPs@Ag-MOF dispersion (final concentration 25 μg / mL) and 50 μL of TMB solution (final concentration 0.5 mM) were added to 1900 μL of HAc-NaAc solution (12 mM, pH 5.0), mixed uniformly, and after reaction at 20°C for 20 minutes, the absorbance value at 650 nm was recorded; then 50 μL of ascorbic acid standard solution with different concentrations (0.001-3.0 mM) was added to the above solution, and after standing for 5 minutes, the absorbance value at 650 nm was recorded. By detecting the absorbance difference produced by different concentrations of AA solution, colorimetric detection of AA was realized.

[0049] AgNPs@Ag-MOF, as a nanozyme with oxidase-like activity, has a high affinity for the substrate 3,3',5,5'-tetramethylbenzidine (TMB). The amount of blue oxTMB produced by the oxidase-like reaction mediated by it decreases after the addition of AA solution. Combined with UV-visible spectrophotometer as a readout device, a simple and rapid AA colorimetric detection method was established. By comparing the absorbance difference produced by AA solutions of different concentrations, a working curve ( Figure 8 ). The linear equation of the curve is Δ 650nm =0.40433C AA +0.05249(R 2 =0.992), with a linear range of 0.001 to 3 mM and a detection limit of 0.3 μM. These results demonstrate that the silver nanoparticle-loaded silver-based metal-organic framework nanozyme (AgNPs@Ag-MOF) provided by the present invention can be used as an oxide-like nanozyme and has good application prospects in the field of colorimetric detection.

[0050] Example 5:

[0051] Using the AgNPs@Ag-MOF prepared in Example 2 as a peroxidase-like nanozyme, the concentration of the product ABTS·+ was changed by changing the concentration of H2O2, and the absorbance value was recorded by using the color change to achieve colorimetric detection of H2O2. The specific experiment is as follows:

[0052] 100 μL AgNPs@Ag-MOF dispersion (final concentration of 20 μg / mL), 50 μL ABTS solution (final concentration of 0.5 mM) and 50 μL H2O2 solution of different concentrations (0.004-4.5 mM) were added to 1800 μL HAc-NaAc solution (10 mM, pH 3.0), mixed evenly, and reacted at 37.5°C for 40 minutes. The absorbance value at a wavelength of 414 nm was recorded using a UV-visible absorption spectrometer.

[0053] AgNPs@Ag-MOF, as a nanozyme with peroxidase-like activity, has a high affinity for the substrate 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) in the presence of hydrogen peroxide. The amount of green ABTS·+ produced by the peroxidase-like reaction mediated by it increases with the increasing concentration of H2O2. Combined with a UV-visible spectrophotometer as a readout device, a simple and rapid H2O2 colorimetric detection method was established. Figure 9 This is the working curve for colorimetric detection of H2O2 solutions with different concentrations. The linear equation of this curve is Δ 650nm =0.19884+0.68586C H2O2 (R 2= 0.998), the linear range is 0.004-4.5 mM, and the detection limit is 2.0 μM. The above results show that the AgNPs@Ag-MOF provided by the application can be used as a peroxidase-like nanoenzyme and has good practical application prospects in the field of colorimetric detection.

[0054] Example 6:

[0055] The AgNPs@Ag-MOF prepared in Example 2 was used as a laccase-like nanoenzyme, and adrenaline was used as an analysis object to investigate its application in colorimetric detection. The specific experiment is as follows:

[0056] 100 μL (final concentration 0.25 mg / mL) of AgNPs@Ag-MOF dispersion solution and 200 μL of adrenaline solution with different concentrations (final concentration 6-50 μM) were added to 1700 μL (20 mM, pH = 6.0) of HEPES solution, mixed at 40°C for 30 minutes, and then centrifuged at 8000 rpm for 3 minutes. The supernatant 1.5 mL was taken, and the absorbance value at 510 nm was recorded by a UV-visible absorption spectrometer.

[0057] The AgNPs@Ag-MOF can be used as a laccase-like nanoenzyme to directly catalyze adrenaline, and the pink substance produced by the laccase-like activity mediated by the AgNPs@Ag-MOF increases with the increase of the amount of adrenaline added. In combination with a UV-visible spectrophotometer as a readout device, a simple and rapid colorimetric detection method for adrenaline was established. Figure 10 is the working curve for colorimetric detection of adrenaline solutions with different concentrations. The linear equation of the curve is Δ 510nm = 0.13157 + 0.00242C EPi (R 2 = 0.998), the linear range is 6-50 μM, and the detection limit is 2 μM. The above results show that the AgNPs@Ag-MOF provided by the application can be used as a laccase-like nanoenzyme and has good application prospects in the field of colorimetric detection.

[0058] The above only describes the preferred embodiments of the application, and any equivalent changes and modifications made within the scope of the application should be included in the scope of the application.

Claims

1. A method for preparing a silver-based metal-organic framework nanoscale enzyme loaded with silver nanoparticles with a multi-enzyme-like activity, characterized in that: The method comprises the following steps: 1) adding triethylamine and 2-methylimidazole into methanol, ultrasonic dissolving at room temperature to obtain a deprotonated organic ligand solution; 2) dissolving silver nitrate in double distilled water, slowly dropping the obtained silver nitrate aqueous solution into the deprotonated organic ligand solution and continuously stirring until the dropping is completed, reacting at room temperature for 2-24 hours in the dark to obtain a uniform milky white suspension; centrifuging to collect the solid product, washing the solid product with water and ethanol respectively for 3 times, and drying in a vacuum drying oven at 60℃ for 24 hours to obtain a silver-based metal organic framework nanoscale enzyme loaded with silver nanoparticles with multi-enzyme-like activity.

2. The method of claim 1, wherein: The molar ratio of the triethylamine, 2-methylimidazole and silver nitrate is 1-2:2:

2.

3. The method of claim 1, wherein: The multi-enzyme-like activity includes oxidase-like activity, peroxidase-like activity and laccase-like activity.

4. A silver nanoparticle loaded silver-based metal-organic framework nanoszyme with a pseudo-multienzyme activity, characterized in that: Prepared by the method of any one of claims 1-3.

5. The silver nanoparticles loaded silver-based metal-organic framework nanoenzyme with multi-enzyme-like activities according to claim 4, characterized in that: The multi-enzyme-like activity includes oxidase-like activity, peroxidase-like activity and laccase-like activity.

6. Application of the silver-based metal organic framework nanoscale enzyme loaded with silver nanoparticles with multi-enzyme-like activity of any one of claims 4-5 in the field of biological analysis and colorimetric detection.

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