Magnetic covalent organic framework material modified by gold nanoparticles as well as preparation method and application of magnetic covalent organic framework material

Abstract

The invention relates to the technical field of biological analysis and detection, in particular to a gold nanoparticle modified magnetic covalent organic framework material as well as a preparation method and application thereof. The magnetic covalent organic framework material modified by the gold nanoparticles has glucose oxidase-like activity and peroxidase-like activity at the same time under an acidic condition; when the substrate glucose appears, the gold nanoparticle modified magnetic covalent organic framework material is subjected to self-cascade catalysis to oxidize the glucose to generate hydrogen peroxide, then peroxidase-like activity is utilized to oxidize the chromogenic substrate TMB to generate oxTMB, and colorimetric detection is carried out according to the color depth of the substrate; in addition, the generated oxTMB and the fluorescence of the magnetic covalent organic framework material modified by the gold nanoparticles can generate an inner filtering effect, the fluorescence is quenched, and fluorescence detection can be carried out according to the fluorescence change. The glucose content is rapidly detected in a colorimetric / fluorescent dual mode, and a new way is provided for conveniently and accurately detecting the glucose content.

Description

Technical Field

[0001] The present invention relates to the technical field of bioanalysis and detection, and particularly relates to a gold nanoparticle-modified magnetic covalent organic framework material, a preparation method thereof, and an application thereof. Background Art

[0002] Disclosing the information of this background art section is only intended to enhance the overall understanding of the present invention, and it is not necessarily regarded as an admission or any form of implication that this information constitutes the prior art already known to those of ordinary skill in the art.

[0003] Glucose is an important energy source, but excessive intake or abnormal metabolism may lead to various diseases. The occurrence of various diseases such as diabetes, hypertension, obesity, and cardiovascular diseases is related to glucose. Therefore, measuring the concentration of glucose is of great significance for monitoring the occurrence of diseases.

[0004] Currently, commercial blood glucose monitors are mainly divided into two categories: electrochemical methods and optical reflection techniques. However, the electrochemical method has the disadvantages of a complex detection system, expensive required instruments, and the need for professional personnel for detection; the optical reflection technique has the disadvantages of poor anti-interference ability and insufficient accuracy of test strips. The enzyme-based cascade catalytic colorimetric method is a new glucose detection method, which has the advantages of simple operation and independence from specific instrument equipment, etc., but also faces the disadvantage of low detection sensitivity; this is because natural glucose oxidase or natural peroxidase is easily inactivated during the reaction process. Therefore, it is of great significance to synthesize substances with glucose oxidase-like or peroxidase-like activities.

[0005] In recent years, due to the designability and modifiability of covalent organic framework materials, they have attracted much attention in the field of enzyme mimetics. However, currently reported covalent organic framework material-based enzyme mimetic materials usually only possess one kind of enzyme mimetic activity, such as glucose oxidase-like activity or peroxidase-like activity, and there are few reports on materials with both glucose oxidase-like activity and peroxidase-like activity.

[0006] In addition, the enzyme-based cascade catalytic colorimetric method uses colorimetry for single-mode detection, which has the disadvantage of low precision. Compared with the single-mode detection strategy, the reliability of the dual-mode detection result is higher. Summary of the Invention

[0007] In order to overcome the above problems, the present invention provides a gold nanoparticle-modified magnetic covalent organic framework material, a preparation method thereof, and an application thereof.

[0008] To achieve the above technical objectives, the present invention adopts the following technical solutions: In the first aspect of the present invention, a gold nanoparticle-modified magnetic covalent organic framework material (Fe3O4@COF-Au NPs) is provided, which uses a magnetic covalent organic framework material (Fe3O4@COF NPs) as a matrix and is loaded with gold nanoparticles (Au NPs). Among them, the magnetic covalent organic framework material has a core-shell structure, with Fe3O4 nanoparticles (Fe3O4 NPs) as the core and a covalent organic framework material (COF) as the shell; the covalent organic framework material is a Schiff base-type covalent organic framework material formed by connecting 1,3,5-tris(4-aminophenyl)benzene and 2,5-dimethoxyterephthalaldehyde with a carbon-nitrogen double bond.

[0009] In one or more embodiments, the particle size of the gold nanoparticles (Au NPs) is 3 - 10 nm.

[0010] In one or more embodiments, the particle size of the Fe3O4 nanoparticles (Fe3O4 NPs) is 200 - 400 nm.

[0011] In one or more embodiments, the thickness of the shell layer covalent organic framework material is 100 - 300 nm.

[0012] In one or more embodiments, the covalent organic framework material (COF) has a repeating structural unit with the structure shown in formula (I):

[0013] Formula (I).

[0014] In the second aspect of the present invention, a preparation method of the gold nanoparticle-modified magnetic covalent organic framework material described in the first aspect is provided, including the following steps: (1) Dissolve 1,3,5-tris(4-aminophenyl)benzene and 2,5-dimethoxyterephthalaldehyde in a first organic solvent, mix evenly for the first time, then add Fe3O4 nanoparticles, mix evenly for the second time, add an acetic acid aqueous solution as a catalyst, and hydrothermally synthesize a magnetic covalent organic framework material; (2) Disperse the magnetic covalent organic framework material in a second organic solvent, add an HAuCl4 solution, mix evenly for the third time, then add a NaBH4 solution, and stir and react to obtain a gold nanoparticle-modified magnetic covalent organic framework material.

[0015] In one or more embodiments, in step (1), the preparation method of the Fe3O4 nanoparticles includes the following steps: Dissolve a ferric salt, sodium acetate, and sodium citrate in ethylene glycol, mix evenly, and hydrothermally synthesize Fe3O4 nanoparticles.

[0016] Preferably, the ferric salt includes ferric chloride, ferric sulfate or ferric acetate, preferably ferric chloride.

[0017] Preferably, the molar ratio of the ferric salt, sodium acetate and sodium citrate is (7-8):(40-50):(2-3), preferably 7.5:44:2.3.

[0018] Preferably, the concentration of the ferric salt in ethylene glycol is 0.117-0.133 mol / L, preferably 0.125 mol / L.

[0019] Preferably, the temperature of the hydrothermal reaction is 180-220 °C, preferably 200 °C; the time of the hydrothermal reaction is 12-15 h, preferably 14 h.

[0020] In one or more embodiments, in step (1), the molar ratio of 1,3,5-tris(4-aminophenyl)benzene to 2,5-dimethoxyterephthalaldehyde is 2:(2.5-4), preferably 2:3.

[0021] In one or more embodiments, in step (1), the first organic solvent is a mixed solution of butanol and 1,4-dioxane, wherein the volume ratio of butanol to 1,4-dioxane is (0.8-1.2):1, preferably 1:1.

[0022] In one or more embodiments, in step (1), the concentration of 2,5-dimethoxyterephthalaldehyde in the first organic solvent is 0.0125-0.02 mol / L, preferably, 0.015 mol / L.

[0023] In one or more embodiments, in step (1), the concentration of Fe3O4 nanoparticles in the first organic solvent is 2.5-3.5 g / L, preferably 3 g / L.

[0024] In one or more embodiments, in step (1), the temperature of the hydrothermal reaction is 65-80 °C, preferably 70 °C; the time of the hydrothermal reaction is 40-60 h, preferably 48 h. At this temperature, both the synthesis of the covalent organic framework material and the formation of the core-shell structured magnetic covalent organic framework material can be ensured.

[0025] In one or more embodiments, in step (2), the second organic solvent is tetrahydrofuran.

[0026] In one or more embodiments, in step (2), the concentration of the magnetic covalent organic framework material in the second organic solvent is 3-4 g / L, preferably 3.3 g / L.

[0027] In one or more embodiments, in step (2), the concentration (mass fraction) of the HAuCl4 solution is 0.08 - 0.12%, preferably 0.1%.

[0028] In one or more embodiments, in step (2), the temperature for adding the HAuCl4 solution is 38 - 42 °C, preferably 40 °C.

[0029] In one or more embodiments, in step (2), the concentration of the NaBH4 solution is 0.4 - 0.6 mM, preferably 0.5 mM.

[0030] In one or more embodiments, in step (2), when adding the NaBH4 solution, the stirring reaction time is 20 - 30 h, preferably 24 h.

[0031] The third aspect of the present invention provides the application of the gold nanoparticle-modified magnetic covalent organic framework material described in the first aspect or the gold nanoparticle-modified magnetic covalent organic framework material prepared by the preparation method described in the second aspect in detecting the glucose content.

[0032] In one or more embodiments, when detecting the glucose content, the pH value of the solution is 3 - 5; the temperature is 25 - 50 °C.

[0033] The fourth aspect of the present invention provides the application of the gold nanoparticle-modified magnetic covalent organic framework material described in the first aspect or the gold nanoparticle-modified magnetic covalent organic framework material prepared by the preparation method described in the second aspect in preparing a product for detecting the glucose content.

[0034] In one or more embodiments, the product includes a reagent or a kit.

[0035] The fifth aspect of the present invention provides a product for detecting the glucose content, including the gold nanoparticle-modified magnetic covalent organic framework material described in the first aspect or the gold nanoparticle-modified magnetic covalent organic framework material prepared by the preparation method described in the second aspect.

[0036] In one or more embodiments, the product includes a reagent or a kit.

[0037] The beneficial effects of the present invention are as follows: (1)The magnetic covalent organic framework material modified with gold nanoparticles (Fe3O4@COF-Au NPs) provided by the present invention has both glucose oxidase-like activity and peroxidase-like activity under acidic conditions; in the presence of the substrate glucose, Fe3O4@COF-Au NPs self-cascade catalyze the oxidation of glucose to hydrogen peroxide (H2O2), and then use the peroxidase-like activity to oxidize the chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) to generate oxTMB, and colorimetric detection can be carried out according to the depth of the substrate color; in addition, the generated oxTMB will have an inner filter effect with the fluorescence of Fe3O4@COF-Au NPs itself, quenching its own fluorescence, and fluorescence detection can be carried out according to the fluorescence change. Therefore, the magnetic covalent organic framework material modified with gold nanoparticles (Fe3O4@COF-Au NPs) provided by the present invention can realize a self-cascade reaction with dual enzyme activities at the same pH, and successfully realize colorimetric / fluorescence dual-mode rapid detection of glucose content, providing a new way for convenient and accurate detection of glucose content.

[0038] (2)The magnetic covalent organic framework material modified with gold nanoparticles (Fe3O4@COF-Au NPs) provided by the present invention realizes colorimetric / fluorescence dual-mode rapid detection of glucose content. The fluorescence and colorimetric linear ranges are 0.001 mM~2 mM and 0.05 mM~2.5 mM respectively, and the detection limits of fluorescence and colorimetric methods are as low as 106 nM and 5 μM. Therefore, it has broad application prospects in the detection of glucose content.

[0039] (3)The present invention first synthesized a magnetic covalent organic framework material with a core-shell structure (Fe3O4@COF NPs). There is a highly ordered porous structure on the surface of the covalent organic framework material (COF). Due to the pore size restriction effect on the surface of the covalent organic framework material (COF) carrier, and the surface -NH2 as the binding site of HAuCl4, the gold nanoparticles (Au NPs) obtained by the reduction of NaBH4 can be in-situ fixed to the shell layer of the magnetic covalent organic framework material (Fe3O4@COF NPs), and the size of the gold nanoparticles (Au NPs) is limited to less than 5 nm. At the same time, the gold nanoparticles (Au NPs) in-situ fixed to the magnetic covalent organic framework material (Fe3O4@COF NPs) are uniformly dispersed on the surface of the covalent organic framework material (COF); the uniformly dispersed and ultra-small-sized gold nanoparticles (Au NPs) are the prerequisite for having glucose oxidase-like activity. Compared with the method that requires adding a stabilizer or a protective agent to achieve the uniform dispersion of gold nanoparticles (Au NPs), the in-situ fixing method of the present invention does not limit the enzyme activity.

[0040] (4) Materials with only a single class of glucose oxidase activity usually have better activity under neutral or alkaline pH conditions, while materials with only a single class of peroxidase activity usually have better activity under acidic pH. A two-step colorimetric detection method is required. Compared with materials with only a single class of glucose oxidase activity or peroxidase activity, the gold nanoparticle-modified magnetic covalent organic framework material (Fe3O4@COF-Au NPs) provided by the present invention has both glucose oxidase-like activity and peroxidase-like activity under acidic conditions, and only one reaction is needed to achieve rapid and accurate detection of glucose content. Description of the Drawings

[0041] The specification drawings forming a part of the present invention are used to provide a further understanding of the present invention. The schematic embodiments and descriptions thereof of the present invention are used to explain the present invention and do not constitute an improper limitation of the present invention.

[0042] Figure 1 are the scanning electron microscope (SEM) images and transmission electron microscope (TEM) images of Fe3O4@COF-Au NPs, where A and B are the SEM images of Fe3O4@COF-Au NPs at different scales; C and D are the TEM images of Fe3O4@COF-Au NPs at different scales; Figure 2 are the Fourier transform infrared (FT-IR) spectrum, X-ray diffraction (XRD) pattern, and fluorescence spectrum of Fe3O4@COF-Au NPs, where A is the FT-IR spectrum, B is the XRD pattern, and C is the fluorescence spectrum; Figure 3 is the X-ray photoelectron spectroscopy (XPS) of Fe3O4@COF-Au NPs, where A is the survey spectrum of XPS, B is Figure 5 , 2+ , Figure 4 , , Figure 3 , , the high-resolution XPS spectrum of Fe, C is the high-resolution XPS spectrum of N 1s, and D is the high-resolution XPS spectrum of Au 4f; Figure 4 are the results of the peroxidase-like activity study of Fe3O4@COF-Au NPs, where A is the peroxidase-like activity of Fe3O4@COF-Au NPs; B is a comparison of the peroxidase-like activities of Fe3O4@COF, Fe3O4, and Fe3O4@COF-Au NPs; C is the fluorescence spectrum of the terephthalic acid (TA) trapping ·OH experiment; D is the ultraviolet-visible absorption spectrum of the tert-butanol (TBA), p-benzoquinone (PBQ), and tryptophan (Try) radical scavenging experiments; the insets in A, B, and D are the colorimetric solutions of different reaction systems, respectively; Figure 5To optimize the concentrations of Fe3O4@COF-Au NPs, TMB, and H2O2. Among them, A is to optimize the concentration of Fe3O4@COF-Au NPs in the range of (0 - 70 µg / mL) at the same concentrations of TMB and H2O2; B is the quantification of A; C is to optimize the concentration of TMB in the range of (0 - 0.5 mM) at the same concentrations of Fe3O4@COF-Au NPs and H2O2; D is the quantification of C; E is to optimize the concentration of H2O2 in the range of (0 - 20 mM) with Fe3O4@COF-Au NPs and TMB, and F is the quantification of E; Figure 6 For the kinetic test results of Fe3O4@COF-Au NPs. Among them, A is the Michaelis-Menten curve of Fe3O4@COF-Au NPs for H2O2, and B is the Lineweaver-Burk double reciprocal plot of Fe3O4@COF-Au NPs for H2O2; C is the Michaelis-Menten curve of Fe3O4@COF-Au NPs for TMB; D is the Lineweaver-Burk double reciprocal plot of Fe3O4@COF-Au NPs for TMB; Figure 7 For the results of the study on the glucose oxidase-like activity of Fe3O4@COF-Au NPs. Among them, A is the schematic diagram of the glucose oxidase-like activity of Fe3O4@COF-Au NPs; B is the ultraviolet-visible absorption spectrum of the glucose oxidase-like activity of Fe3O4@COF-Au NPs; The inset in B is the colorimetric solution for the study of glucose oxidase; Figure 8 For the effect of pH on the dual enzyme activities of Fe3O4@COF-Au NPs; Among them, A is the result of the effect on peroxidase-like activity, and B is the result of the effect on glucose oxidase-like activity; Figure 9 For the effect of temperature on the dual enzyme activities of Fe3O4@COF-Au NPs; Among them, A is the result of the effect on peroxidase-like activity, and B is the result of the effect on glucose oxidase-like activity; Figure 10 For the process of dual-mode detection of glucose by Fe3O4@COF-Au NPs; Figure 11Results of glucose detection by Fe3O4@COF-Au NPs. Among them, A is a schematic diagram of the cascade detection of glucose by Fe3O4@COF-Au NPs; B is the ultraviolet-visible absorption spectra after adding different concentrations of glucose (50 μM - 2.5 mM, from bottom to top); C is the linear relationship between the absorbance at 650 nm and the glucose concentration; the inset in B is the colorimetric solution with different concentrations of added glucose; the regression equation in C is: y = 0.2537x + 0.1455; Figure 12 Results of glucose detection by Fe3O4@COF-Au NPs. Among them, A is a schematic diagram of the fluorescence-mode detection of glucose by Fe3O4@COF-Au NPs; B is the fluorescence spectra of Fe3O4@COF-Au NPs after adding different concentrations of glucose (0 mM - 2 mM); C is the linear relationship between the fluorescence intensity of Fe3O4@COF-Au NPs at 510 nm and the glucose concentration, and the regression equation in C is: y = -738.2381x + 2486.5190. Detailed implementation manners

[0043] It should be noted that the following detailed description is exemplary and is intended to provide further explanation of the present invention. Unless otherwise specified, all technical and scientific terms used in the present invention have the same meaning as commonly understood by those of ordinary skill in the technical field to which the present invention belongs.

[0044] It should be noted that the terms used herein are only for describing specific implementation manners and are not intended to limit the exemplary embodiments according to the present invention. As used herein, unless the context clearly indicates otherwise, the singular form is also intended to include the plural form. In addition, it should be understood that when the terms "comprise" and / or "include" are used in this specification, they indicate the presence of features, steps, operations, devices, components, and / or combinations thereof.

[0045] In order to enable those skilled in the art to more clearly understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0046] Example 1 Synthesis of magnetic covalent organic framework materials modified with gold nanoparticles (Fe3O4@COF-Au NPs): 1.1 Synthesis of Fe3O4 nanoparticles (Fe3O4 NPs): Dissolve 2.04 g of FeCl3·6H2O, 3.6 g of anhydrous sodium acetate and 0.6 g of sodium citrate in 60 mL of ethylene glycol. After the mixed solution is stirred at 1200 rpm for 1.5 h, transfer it to a 100 mL stainless steel autoclave and heat it at 200 °C for 14 h; after cooling to room temperature, magnetically separate the product, wash it 3 times with anhydrous ethanol and deionized water respectively, and vacuum dry it at 30 °C for 12 h to obtain Fe3O4 nanoparticles (Fe3O4 NPs).

[0047] 1.2 Synthesis of magnetic covalent organic framework materials (Fe3O4@COF NPs): Dissolve 42 mg of 1,3,5-tris(4-aminophenyl)benzene (TAPB) and 34.8 mg of 2,5-dimethoxyterephthalaldehyde (DMTP) in a mixed solution of 8 mL of butanol and 8 mL of 1,4-dioxane, and ultrasonically mix for 5 min to make the solution uniformly mixed; add 36 mg of Fe3O4 NPs to the mixed solution, continue ultrasonically mixing for 5 min, then add 0.2 mL of acetic acid, stir at 1200 rpm at room temperature for 2 h, then add 1.8 mL of acetic acid aqueous solution (12 mol / L), and react the mixture at 70 °C for 48 h; after cooling to room temperature, magnetically separate the product, wash it three times with acetone and tetrahydrofuran respectively, and vacuum dry it at 30 °C for 12 h to obtain magnetic covalent organic framework materials (Fe3O4@COF NPs).

[0048] 1.3 Synthesis of gold nanoparticle-modified magnetic covalent organic framework materials (Fe3O4@COF-Au NPs): Dissolve 20 mg of Fe3O4@COF NPs in 6 mL of tetrahydrofuran (THF), and ultrasonically mix for 5 min to make the solution uniformly mixed; at 40 °C, dropwise add 1 mL of HAuCl4 (mass fraction 0.1%), and stir at 1500 rpm for 2 h; then add 2 mL of NaBH¬4 aqueous solution (0.5 mM); after stirring overnight, collect the product with a magnet, wash it three times with water and ethanol respectively, and vacuum dry it for 12 h to obtain gold nanoparticle-modified magnetic covalent organic framework materials (Fe3O4@COF-Au NPs).

[0049] Characterize the gold nanoparticle-modified magnetic covalent organic framework materials (Fe3O4@COF-Au NPs): Figure 1 In A and B are scanning electron micrographs of Fe3O4@COF-Au NPs at different scales. From Figure 1It can be seen that the magnetic covalent organic framework material (Fe3O4@COF NPs) has a core-shell structure, with Fe3O4 nanoparticles (Fe3O4 NPs) as the core and covalent organic framework material (COF) as the shell; the size of the internal core Fe3O4 is about 300 nm, and the outer COF shell layer is 200 nm. -NH2 on the COF surface serves as the binding site for HAuCl4, and Au NPs with small size and uniform distribution on the surface are obtained by NaBH4 reduction; in the transmission electron microscopy images ( Figure 1 C and D), it can be seen that the size of Au NPs is about 5 nm and they are uniformly distributed, which is a necessary condition for having glucose oxidase-like activity.

[0050] Figure 2 Figure Figure 2 6A shows the FI-TR spectrum, XRD pattern and fluorescence spectrum of Fe3O4@COF-Au NPs; in the FI-TR spectrum ( -1 Figure 6A), the strong absorption peak of Fe3O4@COF NPs at 577 cm -1 corresponds to the vibration of the Fe-O band, and the corresponding -C=N- characteristic absorption at 1622 cm -1 indicates that imine bonds are formed by the condensation of DMTP and TAPB; moreover, the -CHO of DMTP has weak stretching vibration bands at 1676 cm -1 and the -NH2 of TAPB has weak stretching vibration bands at 2870 cm -1 and 2934 cm Figure 2 Figure 6B shows that after the loading of Au NPs, new peaks at 38.35, 44.45, and 64.79 appear, corresponding to the (111), (200), and (220) diffraction peaks of Au, respectively. As can be seen from Figure 2 Figure 6C, Fe3O4@COF-Au NPs have strong fluorescence at 510 nm.

[0051] Figure 3 Figure Figure 3 7 shows the XPS spectrum of Fe3O4@COF-Au NPs. As can be seen from Figure 3 Figure 7B, Fe 2+ spectra indicate that the magnetic nanoparticles Fe3O4 are well encapsulated by COF; the two deconvoluted peaks of N 1s ( Figure 3 Figure 7C) at 398.10 and 398.93 are attributed to the residual -NH2 group and -C=N- group, respectively.Figure 3 The two peaks of D in correspond to 4f of Au(0) respectively 7 / 2 and 4f 5 / 2 .

[0052] Figures 1 to 3 The results in all prove the successful preparation of Fe3O4@COF-Au NPs.

[0053] Example 2 Study on the peroxidase-like activity of Fe3O4@COF-Au NPs Experimental method: 100 μL of Fe3O4@COF-Au NPs (1 mg / mL), 100 μL of TMB (10 mM), and 150 μL of H2O2 (100 mM) were added to 1650 μL of acetate buffer solution (pH = 4, 0.2 M), reacted for 3 min, and then the ultraviolet-visible absorption spectrum in the range of 500 - 800 nm and the absorbance at 652 nm were recorded.

[0054] In the presence of H2O2, the peroxidase-like activity of Fe3O4@COF-Au NPs was studied by oxidizing the chromogenic substrate TMB. As Figure 4 shown in A, only in the presence of H2O2, TMB was oxidized to the blue substrate oxTMB, which had an obvious ultraviolet absorption peak at 652 nm. Figure 4 It was found in B that Fe3O4 and Fe3O4@COF also had peroxidase-like activity, but the absorbance value of Fe3O4@COF-Au NPs at 652 nm was much larger than that of Fe3O4 and Fe3O4@COF, indicating that the activity of Fe3O4@COF-Au NPs was much higher than the former two. This was because the in-situ reduction formed small-sized Au NPs deposited on the surface of Fe3O4@COF provided more reactive sites. Generally, the oxidation of TMB was attributed to ROS generated by the reaction of H2O2 and Fe3O4@COF-Au NPs. Different reactive oxygen species scavengers were used to explore the types of ROS generated during the reaction. Figure 4 As shown in C, tert-butanol, p-benzoquinone, and tryptophan were used as hydroxyl radical (·OH), superoxide anion (O 2·- ), and singlet oxygen ( 1 1O2) scavengers respectively. As can be seen from Figure 4 D, after the appearance of tert-butanol and p-benzoquinone, the absorbance of oxTMB at 652 nm decreased significantly, and the solution color changed from dark blue to almost colorless. Then, terephthalic acid (TA) was used as a typical probe for ·OH, and 2-hydroxyterephthalic acid with a high blue fluorescence signal could be generated. As Figure 4As shown in C, 2-hydroxyterephthalic acid is generated only when Fe3O4@COF-AuNPs, H2O2, and TA are all present, and it has a fluorescence characteristic peak at 428 nm. The above results prove that after in-situ immobilizing Au NPs on the Fe3O4@COF shell by the NaBH4 reduction method, the peroxidase activity is improved, and it can catalyze H2O2 to generate the active intermediate ·OH, O 2·- .

[0055] Example 3 Kinetic test of Fe3O4@COF-Au NPs: Optimize the concentrations of Fe3O4@COF-Au NPs, TMB, and H2O2. In acetate buffer (pH = 4, 0.2 M), perform steady-state kinetic measurements by changing the TMB concentration and fixing the H2O2 concentration (and vice versa), and calculate and evaluate the catalytic rate of Fe3O4@COF-Au NPs.

[0056] By Figure 5 The most suitable concentrations of 40 µg / mL of Fe3O4@COF-Au NPs, 0.3 mM of TMB, and 10 mM of H2O2 can be obtained for the catalytic reaction.

[0057] Under these conditions, it is used for the calculation of steady-state kinetics. Through data fitting, a typical Michaelis-Menten curve of Fe3O4@COF-Au NPs is obtained, and the Michaelis constant ( K m ) and the maximum rate of the enzymatic reaction ( V max ) are calculated. Among them, K m represents the affinity of the enzyme for the substrate, and the lower the value, the stronger the affinity. Under different H2O2, the calculated K m and V max are 0.45 mM and 9.73×10 -8 M·s -1 ( Figure 6 A and B in).

[0058] Table 1 shows the comparison of kinetic parameters between Fe3O4@COF-Au NPs and HRP. It can be seen from Table 1 that compared with the K m of the natural enzyme HRP, Fe3O4@COF-Au NPs has a smaller K mand high adsorption capacity, which is 8 times that of HRP. High affinity indicates excellent peroxidase activity. Similarly, for different concentrations of TMB as the substrate, the calculated K m and V max are 0.25 mM and 9.83×10 -8 M·s -1 ( Figure 6 in C and D). And compared with other materials with peroxidase-like activity, it has comparable or better activity.

[0059] Table 1 Kinetic parameters of Fe3O4@COF-Au NPs and HRP

[0060] Example 4 Study on the glucose oxidase-like activity of Fe3O4@COF-Au NPs: Experimental method: 100 μL of Fe3O4@COF NPs-Au NPs (1 mg / mL) and 200 μL of glucose (0.1 M) were added to 1400 μL of acetate buffer solution (pH = 4, 0.2 M). After incubation at room temperature for 2 h, 200 μL of TMB (10 mM) and 100 μL of HRP (100 μg / mL) were added. Then, the absorbance of the mixture at 652 nm was recorded.

[0061] The results are as Figure 7 shown in B. In the presence of HRP and TMB, only the Fe3O4@COF NPs-Au NPs / glucose system has a high ultraviolet absorption peak at 650 nm. It is proved that Fe3O4@COF NPs-Au NPs exhibits glucose oxidase-like activity to catalyze glucose to produce H2O2, and then TMB is oxidized to oxTMB ( Figure 7 in A).

[0062] Example 5 The effects of pH and temperature on the dual enzyme activity of Fe3O4@COF NPs-Au NPs were studied. The results are as Figure 8 and 9 shown. As can be seen from Figure 8 A, Fe3O4@COF NPs-Au NPs have excellent peroxidase-like activity in a wide pH range of pH = 3 - 6. As can be seen from Figure 8As can be seen from Figure B, Fe3O4@COF NPs-Au NPs exhibit excellent glucose oxidase-like activity in the pH range of 3-5. According to the above results, the peroxidase-like activity and glucose oxidase-like activity of Fe3O4@COF NPs-Au NPs have an overlapping optimal pH range. At the same pH, the dual enzyme activities are exerted simultaneously, which avoids the errors brought by the two-step detection method, making the experimental results more accurate and the operation more convenient.

[0063] As can be seen from Figure 9 Figure C, Fe3O4@COF NPs-Au NPs have good peroxidase-like activity and glucose oxidase-like activity at temperatures of 40 °C and 25 °C.

[0064] Example 6 Dual-mode detection of glucose: The detection process is as Figure 10 shown.

[0065] 100 μL of Fe3O4@COF NPs-Au NPs (1 mg / mL) and 200 μL of glucose with different concentrations were added to 1500 μL of acetate buffer solution (pH = 4, 0.2 M). After incubation at room temperature for 2 h, 200 μL of TMB (10 mM) was added. After incubation at 40 °C for 20 min, the absorbance of the mixture at 652 nm and the fluorescence intensity at 510 nm (E x = 470 nm, E m = 510 nm) were recorded and used to plot the absorbance-concentration curve and fluorescence intensity-concentration curve.

[0066] Fe3O4@COF NPs-Au NPs can exert peroxidase-like activity and glucose oxidase-like activity at the same pH, and a self-cascade catalytic detection system for glucose can be established. Using the chromogenic substrate TMB, the content of the analyte glucose can be detected by the depth of the color of the system ( Figure 11 Figure A).

[0067] As Figure 11 shown in Figures B and C, as the glucose concentration increases, the absorbance at 650 nm gradually increases. When the glucose concentration is in the range of 0-2.5 mM, it shows a linear change with the absorbance, and the regression coefficient R 2 = 0.9903, and the detection limit can reach 5 μM (S / N = 3).

[0068] Similarly, in the fluorescence mode, as the glucose concentration increases, the quenching effect of the increase in oxTMB on Fe3O4@COF NPs-Au NPs increases, and the fluorescence intensity at 510 nm gradually decreases ( Figure 12A and B). When the glucose concentration is between 0.001 and 2 mM, it shows a linear change with the fluorescence intensity, and the regression coefficient R 2 = 0.9923, and the detection limit can reach 106 nM (S / N = 3) ( Figure 12 C).

[0069] These results indicate that Fe3O4@COF NPs-Au NPs have excellent dual-enzyme activities and excellent detection performance under fluorescence-UV dual-mode detection. Self-cascade catalysis can be achieved without the participation of natural enzymes, thus avoiding the harsh storage conditions of natural enzymes.

[0070] Example 7 Detection of actual samples: Glucose with different concentrations (0.05 mM, 0.50 mM, 2 mM) was added to Wahaha drinking water for colorimetric and fluorescence detection.

[0071] 100 μL of Fe3O4@COF NPs-Au NPs (1 mg / mL) and 200 μL of glucose with different concentrations were added to 1500 μL of acetate buffer solution (pH = 4, 0.2 M). After incubation at room temperature for 2 h, 200 μL of TMB (10 mM) was added. After incubation at 40 °C for 20 min, the absorbance of the mixture at 652 nm and the fluorescence intensity at 510 nm were recorded (E x = 470 nm, E m = 510 nm), The results are as described in Table 2. The recovery rates of the colorimetric method are between 98.75% and 109.45%, and the relative standard deviation is less than 3.27%. The recovery rates of the fluorescence method are between 103.28% and 107.85%, and the relative deviation is less than 2.56%. It shows that this method has high recovery rate, good selectivity and low relative standard deviation, and can be used for the analysis and detection of glucose content.

[0072] Table 2 Dual-mode detection of glucose by Fe3O4@COF NPs-Au NPs

[0073] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. For those skilled in the art, the present invention can have various changes and modifications. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A gold nanoparticle-modified magnetic covalent organic framework material, characterized in that, It uses a magnetic covalent organic framework material as the matrix and is loaded with gold nanoparticles; Among them, the magnetic covalent organic framework material has a core-shell structure, with Fe3O4 nanoparticles as the core and a covalent organic framework material as the shell; the covalent organic framework material is a Schiff base-type covalent organic framework material in which 1,3,5-tris(4-aminophenyl)benzene and 2,5-dimethoxyterephthalaldehyde are connected by carbon-nitrogen double bonds.

2. The magnetic covalent organic framework material modified with gold nanoparticles according to claim 1, characterized in that The particle size of the gold nanoparticles is 3 - 10 nm; The particle size of the Fe3O4 nanoparticles is 200 - 400 nm; The thickness of the shell layer covalent organic framework material is 100 - 300 nm.

3. The preparation method of the gold nanoparticle-modified magnetic covalent organic framework material described in claim 1 or 2, characterized in that, It includes the following steps: (1) Dissolve 1,3,5-tris(4-aminophenyl)benzene and 2,5-dimethoxyterephthalaldehyde in a first organic solvent, mix evenly for the first time, then add Fe3O4 nanoparticles, mix evenly for the second time, add an acetic acid aqueous solution as a catalyst, and hydrothermally synthesize the magnetic covalent organic framework material; (2) Disperse the magnetic covalent organic framework material in a second organic solvent, add an HAuCl4 solution, mix evenly for the third time, then add a NaBH4 solution, and stir to react to obtain the magnetic covalent organic framework material modified with gold nanoparticles.

4. The preparation method according to claim 3, characterized in that, In step (1), the preparation method of the Fe3O4 nanoparticles includes the following steps: Dissolve ferric salt, sodium acetate and sodium citrate in ethylene glycol, mix evenly, and hydrothermally synthesize Fe3O4 nanoparticles.

5. The preparation method according to claim 3, characterized in that, In step (1), the molar ratio of 1,3,5-tris(4-aminophenyl)benzene to 2,5-dimethoxyterephthalaldehyde is 2:(2.5 - 4); In step (1), the first organic solvent is a mixed solution of butanol and 1,4-dioxane, and the volume ratio of butanol to 1,4-dioxane is (0.8 - 1.2):1; In step (1), the concentration of 2,5-dimethoxyterephthalaldehyde in the first organic solvent is 0.0125 - 0.02 mol / L; In step (1), the concentration of Fe3O4 nanoparticles in the first organic solvent is 2.5 - 3.5 g / L; In step (1), the temperature of the hydrothermal reaction is 65 - 80 °C; the time of the hydrothermal reaction is 40 - 60 h.

6. The preparation method according to claim 3, characterized in that In step (2), the second organic solvent is tetrahydrofuran; In step (2), the concentration of the magnetic covalent organic framework material in the second organic solvent is 3 - 4 g / L; In step (2), the concentration of the HAuCl4 solution is 0.08 - 0.12%; In step (2), the temperature for adding the HAuCl4 solution is 38 - 42 °C; In step (2), the concentration of the NaBH4 solution is 0.4 - 0.6 mM; In step (2), after adding the NaBH4 solution, the stirring reaction time is 20 - 30 h.

7. Application of the magnetic covalent organic framework material modified with gold nanoparticles as described in claim 1 or 2 or the magnetic covalent organic framework material modified with gold nanoparticles prepared by the preparation method as described in any one of claims 3 - 6 in detecting the glucose content.

8. The application according to claim 7, wherein When detecting the glucose content, the pH value of the solution is 3 - 5; the temperature is 25 - 50 °C.

9. Use of the gold nanoparticle-modified magnetic covalent organic framework material described in claim 1 or 2, or the gold nanoparticle-modified magnetic covalent organic framework material prepared by the preparation method described in any one of claims 3 to 6, in the preparation of a product for detecting glucose content.

10. A product for detecting glucose content, characterized in that, Comprising the gold nanoparticle-modified magnetic covalent organic framework material described in claim 1 or 2, or the gold nanoparticle-modified magnetic covalent organic framework material prepared by the preparation method described in any one of claims 3 to 6.

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