Preparation method and application of a BOS@BiOBr nanoenzyme catalyst

Abstract

This invention discloses a method for preparing and applying a BOS@BiOBr nanozyme catalyst, belonging to the field of nanozyme preparation and application technology. The method includes the following steps: (1) preparing a light yellow amorphous Bi2O3 precursor; (2) preparing a bismuth oxysulfate composite material BOS; and (3) preparing the BOS@BiOBr nanozyme catalyst. The BOS@BiOBr prepared in this invention introduces potassium bromide to generate BiOBr based on BOS. Firstly, BiOBr itself possesses peroxidase-like activity, further enhancing POD-like activity; secondly, the valence band position of BiOBr is conducive to H2O2 activation and •OH generation, while the conduction band position is conducive to O2 reduction to •O2. ‑ The dual reactive oxygen species (ROS) generation pathways enhance antibacterial / catalytic efficiency; finally, BiOBr, as an intermediate bandgap, regulates electron transport between the BOS core and the external environment, optimizing overall catalytic performance.

Description

Technical Field

[0001] This invention relates to the field of nanozyme preparation and application technology, specifically to a method for preparing and applying a BOS@BiOBr nanozyme catalyst. Background Technology

[0002] Technological advancements have fueled the continued expansion of the antibiotic market, but they have also exacerbated antibiotic overuse, leading to a significant increase in antibiotic residues in aquatic environments and making them a major source of water pollution. Tetracycline (TC), for example, possesses high water solubility (solubility > 1.3 g / L) and excellent chemical stability, resulting in widespread residues in water bodies at concentrations reaching micrograms per liter. Even more serious is the fact that tetracycline can penetrate every stage of the food chain through various pathways, creating a chain-wide residue risk: In aquaculture, tetracycline is often added in large quantities as a growth promoter to livestock and aquatic feed, directly causing excessive antibiotic residues in animal-derived foods such as meat, eggs, and milk; in agricultural production, wastewater containing tetracycline is used for farmland irrigation, leading to antibiotic accumulation in crops through root absorption; in food processing, contaminated water sources are used for raw material cleaning and processing, and coupled with lax supervision during withdrawal periods in aquaculture, this further amplifies the potential for tetracycline residues.

[0003] Tetracycline exhibits a significant dose-cumulative effect in its harmful effects on human health. Different exposure doses can cause varying degrees of health damage: high doses can directly lead to liver failure, and in severe cases, death; long-term low-dose exposure can slowly damage liver and kidney function, and may also induce liver cancer, cirrhosis, and other diseases, producing carcinogenic, teratogenic, and mutagenic effects; even trace residues can disrupt the homeostasis of the human gut microbiota and accelerate the spread of drug-resistant bacteria. Furthermore, tetracycline has extremely strong resistance to degradation; it is not only resistant to high temperatures (degradation rate during cooking <15%), but also resistant to common food processing sterilization techniques such as irradiation and pasteurization, making it difficult to remove effectively. Therefore, it persists in food and poses a long-term exposure risk.

[0004] In the detection of tetracycline (TC), the application of single nanomaterials has significant limitations, failing to meet the requirements for high efficiency and sensitivity. Semiconductor bismuth oxide, with its excellent specific surface area, provides abundant active sites and exhibits some potential for peroxidase-like catalytic colorimetric detection; however, its enzyme-catalyzed reaction time is long, making it difficult to apply in practical detection. While bismuth sulfide possesses unique photoelectric properties, the low peroxidase activity of a single Bi₂S₃ material greatly limits its practical application. Bismuth halide (BiOBr), due to its unique sheet-like structure which facilitates electron transport, lacks the excellent peroxidase-like activity of a single BiOBr, making it unsuitable for direct colorimetric detection of tetracycline. Therefore, there is an urgent need to develop a nanocomposite material with high peroxidase-like activity for efficient and sensitive detection, which has significant research value and practical significance for ensuring food safety and public health. Summary of the Invention

[0005] To address or partially address the problems existing in related technologies, this invention provides a method for preparing and applying BOS@BiOBr nanozyme catalysts. This method significantly enhances the catalytic activity of BOS@BiOBr nanozymes by combining Bi2O3 with Bi2S3 and BiOBr, enabling sensitive detection of tetracycline.

[0006] To achieve the above objectives, the technical solution of the present invention is as follows: a method for preparing BOS@BiOBr nanozyme catalyst, the innovation of which lies in the following steps: (1) Add anhydrous ethanol to the bismuth nitrate pentahydrate acetic acid solution to obtain a mixed solution, and add tetraethyl orthosilicate dropwise, stir quickly and evenly, heat to react, wash and dry after the reaction is completed, and calcine to obtain a light yellow amorphous Bi2O3 precursor. The calcination heating rate is a key kinetic factor affecting the crystal form of Bi2O3 precursors. Rapid heating (e.g., above 5–10 ℃ / min) causes violent thermal decomposition of the precursor, leaving insufficient time for atoms to diffuse and rearrange, easily forming amorphous or defective microcrystalline structures. In contrast, slow heating (e.g., below 1–3 ℃ / min) allows for uniform heating of the system and a gentle decomposition process, giving atoms sufficient time to overcome potential barriers and complete ordered arrangement, making it easier to generate single-crystal Bi2O3 with high crystallinity and few defects. This difference is essentially the result of a competition between kinetic and thermodynamic control. Subtle changes in the heating rate can directly alter the atomic diffusion and crystal nucleation processes, leading to different product crystal forms.

[0007] (2) The Bi2O3 precursor was dispersed in ultrapure water and subjected to continuous sonication to obtain a suspension. Polyvinylpyrrolidone was added to the suspension and mixed evenly. Then, thioacetamide aqueous solution was added and heated to react. After the reaction was completed, the mixture was centrifuged, washed, and dried to obtain bismuth oxysulfate composite material BOS. (3) The BOS powder obtained in step (2) is dispersed in a mixed solution of ultrapure water and ethylene glycol. Polyvinylpyrrolidone and potassium bromide are added in sequence to form a dispersion. After thorough stirring, the mixture is heated to react. After the reaction is completed, the mixture is washed and dried to obtain the BOS@BiOBr nanozyme catalyst.

[0008] Preferably, in step (1), the concentration of bismuth nitrate in the bismuth nitrate pentahydrate acetic acid solution is 0.22-0.67 mol / L; the volume ratio of the bismuth nitrate pentahydrate acetic acid solution to anhydrous ethanol is 1:3-7; and the molar ratio of bismuth nitrate pentahydrate to tetraethyl orthosilicate is 10:1-3.

[0009] Preferably, in step (1), the dropping rate of tetraethyl orthosilicate into the mixed solution is 0.4~10 mL / min.

[0010] Preferably, in step (1), the heating reaction conditions are: heating the reaction at 80℃-180℃ for 26-56h.

[0011] Preferably, in step (1), the calcination conditions are: heating to 330℃-470℃ at a heating rate of 5℃ / min, and calcination time is 1-5h.

[0012] Preferably, in step (2), the concentration of Bi2O3 precursor in the suspension is 1-20 g / L; the mass ratio of polyvinylpyrrolidone to Bi2O3 precursor is 1-2:2; the mass percentage of thioacetamide in the thioacetamide aqueous solution is 0.5%~10%; the volume ratio of the suspension to the thioacetamide aqueous solution is 1:1~2.5; and the heating reaction conditions are: heating at 100℃~200℃ for 2h~8h.

[0013] Preferably, in step (3), the volume percentage of ethylene glycol in the mixed solution of ultrapure water and ethylene glycol is 50%~80%; in the dispersion, the concentration of BOS is 1.33-3.33 mg / mL, the concentration of polyvinylpyrrolidone is 1.33-5 mg / mL, and the concentration of potassium bromide is 1.93-5.5 mg / mL; the heating reaction conditions are: heating reaction at 80-180℃ for 2-8 h.

[0014] This invention also provides an application of the BOS@BiOBr nanozyme catalyst prepared by the above method in tetracycline degradation. The innovation lies in preparing the BOS@BiOBr nanozyme catalyst into a BOS@BiOBr-ssDNA aptamer probe. The specific steps are as follows: TC-ssDNA is added to a buffer solution and then incubated; after incubation, the BOS@BiOBr nanozyme catalyst is added, and incubation continues; after incubation, the sample is washed with Tris-HCl solution to obtain the BOS@BiOBr-ssDNA aptamer probe, which is then applied to the detection of tetracycline.

[0015] Preferably, the buffer solution is composed of a mixture of tris(2-carboxyethyl)phosphine hydrochloride solution and Tris-HCl solution; the concentration of the tris(2-carboxyethyl)phosphine hydrochloride solution in the buffer solution is 10.0 mg / mL, the concentration of the Tris-HCl solution is 10.0 mmol / L, and the pH is 7.5; the volume ratio of the tris(2-carboxyethyl)phosphine hydrochloride solution to the Tris-HCl solution in the buffer solution is 0.2-1.5:0.5-2.0.

[0016] Preferably, the concentration of TC-ssDNA is 80.0-400.0 nmol / L, and the volume ratio of TC-ssDNA to buffer is 0.05-0.2:2.5-3.0; in the buffer after adding BOS@BiOBr nanozyme catalyst, the concentration of BOS@BiOBr nanozyme catalyst is 0.1-0.5 mg / mL.

[0017] The principle of this invention is as follows: Using amorphous bismuth oxide (Bi2O3) precursor as a substrate, a homogeneous composite bismuth oxide (BOS) material was obtained through a sulfidation reaction. Then, by introducing potassium bromide as a bromine source, a reaction was initiated on the surface of the bismuth oxide composite material BOS, converting a portion of Bi into BiOBr to generate BOS@BiOBr composite nanozymes. An interfacial effect was generated between the BOS composite material and BiOBr. Bi2O3, Bi2S3, and BiOBr have highly matched band structures. This band matching characteristic provides mechanical feasibility for charge transfer at the interface, thereby promoting the formation of a stable and tight contact interface. This tight contact interface is not only an efficient channel for charge transfer, but also a core carrier for constructing abundant catalytic active sites.

[0018] In this interfacial system, the interfacial effect achieves a synergistic enhancement of catalytic efficiency by precisely regulating the distribution and function of active sites. On the one hand, the sulfur vacancies on the Bi2S3 surface and the defective structure of BiOBr itself become highly efficient electron-capturing centers under the regulation of the interfacial effect, promoting the generation of free radicals with strong oxidizing activity. On the other hand, Bi2O3 exposes a large number of oxidizing active sites in the contact area through interfacial interactions. These sites enhance substrate adsorption and activation capabilities through synergistic effects, forming a functionally complementary catalytic system with the aforementioned electron-capturing centers, synergistically improving the catalytic reaction rate and significantly enhancing the activity of nanomaterial-based peroxidases. More importantly, the interfacial region formed by BOS and BiOBr has a specific surface electronic structure and spatial configuration, which can preferentially adsorb tetracycline molecules. The amino and hydroxyl functional groups in tetracycline molecules can form specific interactions with the interfacial active sites, thereby achieving selective enrichment of tetracycline in complex pollutant systems. This invention provides a method for preparing and applying BOS@BiOBr nanozyme catalysts, which have the following beneficial effects: (1) The BOS@BiOBr prepared in this invention introduces potassium bromide into BOS to generate BiOBr. First, BiOBr itself has peroxidase-like activity, which further enhances POD-like activity; second, the valence band position of BiOBr is conducive to H2O2 activation and •OH generation, and the conduction band position is conducive to O2 reduction to •O2. - The dual reactive oxygen species (ROS) generation pathways enhance antibacterial / catalytic efficiency; finally, BiOBr, as an intermediate bandgap, regulates electron transport between the BOS core and the external environment, optimizing overall catalytic performance.

[0019] (2) The BOS@BiOBr nanozyme catalyst prepared in this invention has the function of highly efficient detection of tetracycline. Based on the BOS@BiOBr-ssDNA aptamer probe, specific recognition and high-sensitivity colorimetric detection of tetracycline (TC) can be achieved. The modified aptamer probe has a high affinity for TC. After the aptamer binds to TC, it causes a change in the activity of the nanozyme in the solution, thereby inhibiting the efficiency of the nanozyme catalyzing the substrate reaction, thus achieving efficient, rapid and sensitive detection of TC. It has a fast response speed, a low limit of detection (LOD), a wide linear range, and excellent selectivity and anti-interference. It also shows excellent recovery rate according to the national standard detection method (high performance liquid chromatography).

[0020] (3) The BOS@BiOBr nanozyme catalyst prepared in this invention uses bismuth ions as a common metal source for Bi2O3, Bi2S3 and BiOBr materials to synergistically catalyze multiple matrix materials, generating more active oxygen and exhibiting excellent peroxidase-like activity, thus synergistically improving catalytic performance. Attached Figure Description

[0021] Figure 1 shows the scanning electron microscope (SEM) and transmission electron microscope (TEM) images of the BOS@BiOBr nanozyme catalyst prepared in Example 1 of the present invention. Figure 1(a) is a scanning electron microscope image of BOS@BiOBr, and Figure 1(b) is a high-resolution transmission electron microscope image of BOS@BiOBr.

[0022] Figure 2 This is the XRD spectrum of the BOS@BiOBr nanozyme catalyst prepared in Example 1 of this invention.

[0023] Figure 3 This is a comparison chart of the activities of BOS@BiOBr nanozyme catalyst prepared in Example 1 of this invention and BOS, bismuth oxide, bismuth sulfide, and BiOBr peroxidases.

[0024] Figure 4 This is a standard curve diagram of TC detection by colorimetric assay using the BOS@BiOBr-ssDNA aptamer probe prepared in Example 1 of this invention.

[0025] Figure 5 This is a comparison chart of the enzyme activity of the nanozyme prepared in Comparative Example 1 and the nanozyme prepared in Example 1 of this invention.

[0026] Figure 6 This is a comparison chart of the absorbance of TC in the prepared aptamer composite materials of Comparative Example 1 and Example 1 of the present invention.

[0027] Figure 7 This is the absorbance graph of the aptamer composite material prepared in Comparative Example 2 of this invention, showing the detection of different concentrations of TC.

[0028] Figure 8 This is a comparison chart of the absorbance of TC in the prepared aptamer composite materials of Comparative Example 4 and Examples of the present invention. Detailed Implementation

[0029] The present invention will be further described in detail below with reference to specific embodiments and accompanying drawings, but the scope of protection of the present invention is not limited to the content described.

[0030] Example 1 The specific steps for preparing the BOS@BiOBr nanozyme catalyst and the corresponding BOS@BiOBr-ssDNA aptamer probe in this embodiment are as follows: (1) Add 85.0 mL of anhydrous ethanol to 25 mL of 0.4 mol / L bismuth nitrate pentahydrate acetic acid solution to obtain a mixed solution, and add 0.5 mmol of tetraethyl orthosilicate dropwise at a rate of 4 mL / min. Stir quickly and evenly, and heat at 160 °C for 48 h. After the reaction is completed, wash and dry, and calcine at a rate of 5 °C / min to 400 °C. After calcining at 400 °C for 3 h, a light yellow amorphous Bi2O3 precursor is obtained. (2) 91 mg of amorphous Bi2O3 precursor was dispersed in 15 mL of ultrapure water and sonicated for 15 min to obtain a suspension. 84 mg of polyvinylpyrrolidone (PVP) was added to the suspension and mixed evenly. Then 22.5 mL of 5% (w / w) thioacetamide (TAA) aqueous solution was added and heated at 160 °C for 6 h. After the reaction was completed, the mixture was centrifuged, washed and dried to obtain bismuth oxysulfate composite material BOS. BOS is a homogeneous composite bismuth oxysulfate material (BOS is a homogeneous composite material of Bi2O3 and Bi2S3).

[0031] (3) Take 98 mg of the BOS powder obtained in step (2) and disperse it in a mixed solution of 60 mL of ultrapure water and ethylene glycol (60% by volume). Add 98 mg of polyvinylpyrrolidone and stir for 10 min. Then add 260 mg of potassium bromide (KBr) and stir thoroughly for 10 min. Heat the mixture at 180 °C for 3 h. After the reaction is complete, wash and dry to obtain the BOS@BiOBr nanozyme catalyst.

[0032] Using BOS@BiOBr nanozyme catalyst to prepare aptamer probes: 180 μL of TC-ssDNA with a concentration of 160 nmol / L was added to 6.0 mL of buffer (the buffer was a mixture of 0.55 mL of 10.0 mg / mL tris(2-carboxyethyl)phosphine hydrochloride solution and 5.45 mL of 10.0 mmol / L Tris-HCl solution, pH 7.5), and then incubated at 37 °C for 1 h; subsequently, 1.0 mg of BOS@BiOBr was added, and incubation was continued for 14 h. The sample was then washed three times with Tris-HCl to obtain the BOS@BiOBr-ssDNA aptamer probe.

[0033] The bromine content in the BOS@BiOBr nanozyme material prepared in this embodiment is approximately 43%.

[0034] Scanning electron microscope (SEM) and transmission electron microscope (TEM) images of the BOS@BiOBr nanozyme catalyst prepared in this embodiment are shown below. Figure 1 As shown, Figure 1As shown in (a), the BOS@BiOBr nanozyme prepared in this embodiment is spherical and has a flake-like morphology on its surface, with obvious porous and sheet-like stacked structure. Figure 1 As shown in (b), the interfaces are clearly intersecting, and the synergistic effect of the interfaces can generate a large number of reactive oxygen species that work together with the oxidative active sites.

[0035] The XRD spectrum of the BOS@BiOBr nanozyme catalyst prepared in Example 1 is shown below. Figure 2 As shown, Figure 2 It can be seen that the BOS@BiOBr nanozyme in this embodiment contains three phases (Bi2O3, Bi2S3 and BiOBr), with good crystallinity and high purity.

[0036] Example 1 tested the activities of the BOS@BiOBr nanozyme catalyst and BOS, bismuth oxide, bismuth sulfide, and BiOBr-type peroxidases. The testing procedure was as follows: First, an appropriate amount of nanozyme and a blank sample were added to a certain amount of NaAC-HAC buffer solution and mixed thoroughly. Then, an appropriate amount of TMB solution and H2O2 were added to the mixture, maintaining a consistent total volume. The mixture was incubated in the dark, and the solution gradually turned blue-green. After a certain reaction time, the reaction solution was centrifuged, and 3.0 mL of the supernatant was taken. The absorbance was measured at 652 nm using a UV-Vis spectrophotometer, and the changes in absorbance intensity between the experimental and control groups were observed and analyzed. The test results were compared as follows: Figure 3 As shown (a blank control is also included in the figure), Figure 3 In this embodiment, the BOS@BiOBr nanozyme exhibits superior peroxidase-like activity compared to bismuth oxide monomer, bismuth sulfide monomer, bismuth oxybromide monomer, and their BOS composite materials.

[0037] The BOS@BiOBr-ssDNA aptamer probe prepared in Example 1 was used for tetracycline detection. The procedure was as follows: 2.0 mg of the aptamer-modified material (BOS@BiOBr-ssDNA) was added to 2.31 mL of Tris-HCl (10.0 mmol / L, pH=7.5) in a series of freshly prepared tetracycline (TC) standard solutions of various concentrations. Subsequently, appropriate amounts of TMB (8.3 mmol / L) and H2O2 solution were added to the mixture, maintaining the total reaction volume at 3.1 mL. After incubation for a period of time, the absorbance value at 652 nm was recorded using a UV-Vis spectrophotometer for quantitative detection. All UV-Vis measurements were performed in triplicate to ensure reproducibility.

[0038] The detection results (standard curve of TC detection by BOS@BiOBr-ssDNA aptamer probe colorimetric assay) are as follows: Figure 4 As shown, by Figure 4 It was found that the BOS@BiOBr-ssDNA aptamer probe synthesized in Example 1 showed a significant response to 10.0 μg / L TC, reaching the response equilibrium point within 15 min. The detection range for TC was 0.3–500.0 μg / L, with a detection limit of 145 ng / L, exhibiting good stability, selectivity, and anti-interference properties. The synthesized BOS@BiOBr also demonstrated excellent nanozyme activity.

[0039] The BOS@BiOBr-ssDNA aptamer probe synthesized in this invention exhibits excellent performance in the detection of tetracycline (TC) within the range of 0–500.0 μg / L, such as… Figure 4 As shown, absorbance exhibits a good linear relationship with concentration (y = 4.60 × 10⁻⁶). -4 (x+0.18, R²=0.9986), conforming to the Lambert-Beer law, the high linearity and sensitivity indicate that the BOS@BiOBr-ssDNA aptamer probe of this invention is accurate and reliable, suitable for rapid quantitative detection of TC in environmental and biological samples, and has the advantages of simple operation and low cost.

[0040] The BOS@BiOBr composite nanozyme material prepared in this embodiment also demonstrates feasibility for monitoring tetracycline in actual milk samples. Table 1 shows a comparison of the recoveries of BOS@BiOBr prepared in Example 1 in actual milk samples detected by ultraviolet spectrophotometry (UV-Vis) and high-performance liquid chromatography (UHPLC). Table 1 In Table 1, the initial tetracycline concentration in the milk samples was "not detected," indicating a clean sample matrix. "Added" represents the addition of tetracycline standards at concentrations of 10 μg / L, 50 μg / L, and 300 μg / L, respectively. "Recovery" represents the recovery rate. Table 1 shows that in actual milk samples, the recoveries of both methods at the three spiking levels were within a reasonable range of 90% to 115%, and the results were close, indicating that the catalyst prepared in this invention has good accuracy for tetracycline detection in milk and is feasible in actual milk samples.

[0041] Example 2 The specific steps for preparing the BOS@BiOBr nanozyme catalyst and the corresponding BOS@BiOBr-ssDNA aptamer probe in this embodiment are as follows: (1) Add 45 mL of anhydrous ethanol to 15 mL of 0.22 mol / L bismuth nitrate pentahydrate acetic acid solution to obtain a mixed solution, and add 0.1 mmol of tetraethyl orthosilicate dropwise at a rate of 0.4 mL / min. Stir quickly and evenly, and heat at 80 °C for 56 h. After the reaction is completed, wash and dry, and heat to 330 °C at a rate of 5 °C / min. After calcining at 330 °C for 5 h, a light yellow amorphous Bi2O3 precursor is obtained. (2) 91 mg of amorphous Bi2O3 precursor was dispersed in 91 mL of ultrapure water and sonicated for 15 min to obtain a suspension. 46 mg of polyvinylpyrrolidone (PVP) was added to the suspension and mixed evenly. Then, 91 mL of 10% (w / w) thioacetamide (TAA) aqueous solution was added and heated at 100 °C for 8 h. After the reaction was completed, the mixture was centrifuged, washed, and dried to obtain bismuth oxysulfate composite material BOS.

[0042] (3) Take 98 mg of the BOS powder obtained in step (2) and disperse it in a mixed solution of 72 mL of ultrapure water and ethylene glycol (volume percentage of 50%). Add 98.76 mg of polyvinylpyrrolidone and stir for 10 min. Then add 139 mg of potassium bromide (KBr) and stir thoroughly for 10 min. Heat the mixture at 180 °C for 2 h. After the reaction is complete, wash and dry to obtain the BOS@BiOBr nanozyme catalyst.

[0043] (4) Add 0.084 mL of TC-ssDNA with a concentration of 80 nmol / L to 5.0 mL of buffer (the buffer is a mixture of 0.125 mL of tris(2-carboxyethyl)phosphine hydrochloride solution and 1.25 mL of Tris-HCl solution, pH 7.5), and then incubate at 37 °C for 1 h; then add 1.5 mg of BOS, continue incubation for 12 h, and wash three times with Tris-HCl to obtain the BOS@BiOBr-ssDNA aptamer probe.

[0044] The bromine content in the BOS@BiOBr nanozyme material prepared in this embodiment is about 17%. The activity of the BOS@BiOBr nanozyme catalyst prepared in this embodiment is similar to that in Example 1, and the detection effect of the synthesized BOS@BiOBr-ssDNA aptamer probe is similar to that in Example 1.

[0045] Example 3 The specific steps for preparing the BOS@BiOBr nanozyme catalyst and the corresponding BOS@BiOBr-ssDNA aptamer probe in this embodiment are as follows: (1) Add 105 mL of anhydrous ethanol to 15 mL of 0.67 mol / L bismuth nitrate pentahydrate acetic acid solution to obtain a mixed solution, and add 3 mmol of tetraethyl orthosilicate dropwise at a rate of 10 mL / min. Stir quickly until homogeneous, and heat at 180 °C for 26 h. After the reaction is complete, wash and dry, and heat to 470 °C at a rate of 5 °C / min. After calcining at 470 °C for 1 h, a light yellow amorphous Bi2O3 precursor is obtained.

[0046] (2) 91 mg of amorphous Bi2O3 precursor was dispersed in 5 mL of ultrapure water and sonicated for 15 min to obtain a suspension. 91 mg of polyvinylpyrrolidone (PVP) was added to the suspension and mixed evenly. Then 12.5 mL of 0.5% (w / w) thioacetamide (TAA) aqueous solution was added and heated at 200 °C for 2 h. After the reaction was completed, the mixture was centrifuged, washed, and dried to obtain bismuth oxysulfate composite material BOS.

[0047] (3) Take 98 mg of the BOS powder obtained in step (2) and disperse it in a mixed solution of 30 mL of ultrapure water and ethylene glycol (volume percentage of 80%). Add 150 mg of polyvinylpyrrolidone and stir for 10 min. Then add 165 mg of potassium bromide (KBr) and stir thoroughly for 10 min. Heat the mixture at 80 °C for 8 h. After the reaction is complete, wash and dry to obtain the BOS@BiOBr nanozyme catalyst.

[0048] (4) 0.1 mL of 400 nmol / L TC-ssDNA was added to 1.25 mL of buffer (the buffer was a mixture of 5.13 mL of tris(2-carboxyethyl)phosphine hydrochloride solution and 1.71 mL of Tris-HCl solution, pH 7.5), and then incubated at 37 °C for 1 h; subsequently, 0.2 mg of BOS@BiOBr was added, and incubation was continued for 12 h, followed by washing three times with Tris-HCl to obtain... BOS BiOBr-ssDNA aptamer probe.

[0049] The mass fraction of bromine in the BOS@BiOBr nanozyme material prepared in this embodiment is 57%. The activity of the BOS@BiOBr nanozyme catalyst prepared in this embodiment is similar to that in Example 1, and the detection effect of the synthesized BOS@BiOBr-ssDNA aptamer probe is similar to that in Example 1.

[0050] In Examples 1-3, the concentration of tris(2-carboxyethyl)phosphine hydrochloride solution was 10.0 mg / mL; the concentration of Tris-HCl solution was 10.0 mmol / L.

[0051] Comparative Example 1 The difference between this comparative example and the embodiment is that the heating conditions in step (2) are: heating the reaction at 80°C for 6 hours.

[0052] The BOS@BiOBr nanozyme material prepared in Comparative Example 1 was used to prepare the BOS@BiOBr-ssDNA aptamer probe using the same method as in Example 1.

[0053] Compared to the BOS@BiOBr nanozyme material prepared in Example 1, which contains approximately 3% sulfur, the BOS@BiOBr nanozyme material prepared in this comparative example showed that its peroxidase-like activity was not as good as that of the material in Example 1, as determined by the TMB colorimetric reaction. Furthermore, the absorbance of the synthesized BOS@BiOBr-ssDNA aptamer probe did not change significantly when placed in a reaction system containing TC. Figure 5 This indicates that the absorbance of the prepared BOS@BiOBr nanozyme material is not as high as that of the material prepared in Example 1. A comparison of the absorbance of the BOS@BiOBr-ssDNA aptamer probes prepared in Example 1 and Comparative Example 1 for detecting TC is shown in the figure below. Figure 6 As shown, by Figure 6 It can be seen that the BOS@BiOBr-ssDNA aptamer probe prepared by the BOS@BiOBr nanozyme material prepared in Example 1 has a better detection effect on TC.

[0054] Comparative Example 2 The methods and procedures for preparing BOS@BiOBr nanozyme materials in this comparative example and Example 1 are the same.

[0055] The process of preparing the BOS@BiOBr-ssDNA aptamer probe from the BOS@BiOBr nanozyme material prepared in Comparative Example 2 differs from that in the example as follows: 50 μL of TC-ssDNA with a concentration of 60.0 nmol / L was added to 1 mL of buffer (the buffer was a mixture of 0.5 mL tris(2-carboxyethyl)phosphine hydrochloride solution and 0.5 mL Tris-HCl solution), and then incubated at 37 °C for 1 h; subsequently, 1.0 mg of BOS@BiOBr was added, and incubation was continued for 12 h. The sample was then washed three times with Tris-HCl to obtain the BOS@BiOBr-ssDNA aptamer probe.

[0056] The absorbance results of the BOS@BiOBr-ssDNA aptamer probe prepared in this comparative example for detecting different concentrations of TC are as follows: Figure 7 As shown, Figure 7It can be seen that the BOS@BiOBr-ssDNA aptamer probe of Comparative Example 2 responded to 60.0 μg / L TC, but compared with Example 1, the response value changed less, especially under low concentration (e.g., 5 μg / L) conditions, and the detection response could not be directly obtained by the change in absorbance.

[0057] Comparative Example 3 BOS was prepared in this comparative example using the same method as in Example 1.

[0058] The process of preparing the BOS@BiOBr-ssDNA aptamer probe using BOS prepared in Comparative Example 2 was as follows: 50 μL of TC-ssDNA with a concentration of 80 nmol / L was added to 1 mL of buffer (the buffer was a mixture of 0.5 mL tris(2-carboxyethyl)phosphine hydrochloride solution and 0.5 mL Tris-HCl solution), and then incubated at 37 °C for 1 h; subsequently, 1.0 mg of BOS was added, and incubation was continued for 12 h, followed by washing three times with Tris-HCl to obtain the BOS-ssDNA aptamer probe.

[0059] Although the BOS-ssDNA aptamer probe prepared in this comparative example responded to 10.0 μg / L TC, the response value took 75 minutes to stabilize, a reaction time significantly longer than that of the BOS@BiOBr material in Example 1. Furthermore, its linear detection range for TC was limited to a detection limit of 0.41 μg / L, which was higher than the LOD of the examples. Compared to the examples, it lacked the ability to detect substances below the microgram level, making it less suitable for practical applications.

[0060] Comparative Example 4 The specific steps for preparing the BiOBr nanozyme catalyst and the corresponding BiOBr-ssDNA aptamer probe in this comparative example are as follows: (1) Dissolve 1.0 mmol of Bi(NO3)3·5H2O in 10.0 mL of ethylene glycol and sonicate continuously for 20 min. Then add 40.0 mL of ultrapure water and stir continuously for 10 min. Add 100 mg of KBr and 1.0 mmol of PVP to the above mixture and stir vigorously for 30 min. Transfer the resulting mixture to a stainless steel autoclave lined with polytetrafluoroethylene and react at 160 °C for 6 h. After the reaction is completed, centrifuge, wash with ethanol and water, and vacuum dry at 60 °C for 12 h to obtain white-gray BiOBr.

[0061] (2) Add 50 μL of TC-ssDNA with a concentration of 100 nmol / L to 1 mL of buffer (the buffer is a mixture of 0.5 mL tris(2-carboxyethyl)phosphine hydrochloride solution and 0.5 mL Tris-HCl solution), and then incubate at 37 °C for 1 h; then add 1.0 mg BiOBr, continue incubation for 12 h, and wash three times with Tris-HCl to obtain BiOBr-ssDNA aptamer probe.

[0062] The BiOBr prepared in this comparative example exhibited extremely low peroxidase activity, and the BiOBr-ssDNA aptamer probe prepared in this comparative example showed almost no response to 100.0 μg / L TC, with its absorbance value remaining almost identical to that of the blank control group. The results are as follows... Figure 8 As shown, by Figure 8 It can be seen that the BiOBr enzymes prepared in this comparative example have extremely weak activity and cannot detect tetracycline using the BiOBr-ssDNA aptamer probe.

[0063] In summary, the BOS@BiOBr composite nanozyme prepared by this invention exhibits excellent peroxidase-like activity; and the BOS@BiOBr-ssDNA aptamer probe synthesized by this invention can rapidly and specifically recognize TC. The BOS@BiOBr-ssDNA probe can directly capture TC and adsorb onto the active sites on the material surface, thereby reducing the nanozyme activity and realizing quantitative colorimetric detection of TC.

[0064] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A method for preparing a BOS@BiOBr nanozyme catalyst, characterized in that: Includes the following steps: (1) Add anhydrous ethanol to the bismuth nitrate pentahydrate acetic acid solution to obtain a mixed solution, and add tetraethyl orthosilicate dropwise, stir evenly, heat to react, wash and dry after the reaction is completed, and calcine to obtain a light yellow amorphous Bi2O3 precursor. (2) The Bi2O3 precursor was dispersed in ultrapure water and subjected to continuous sonication to obtain a suspension. Polyvinylpyrrolidone was added to the suspension and mixed evenly. Then, thioacetamide aqueous solution was added and heated to react. After the reaction was completed, the mixture was centrifuged, washed, and dried to obtain bismuth oxysulfate composite material BOS. (3) The BOS powder obtained in step (2) is dispersed in a mixed solution of ultrapure water and ethylene glycol. Polyvinylpyrrolidone and potassium bromide are added in sequence to form a dispersion. After thorough stirring, the mixture is heated to react. After the reaction is completed, the mixture is washed and dried to obtain the BOS@BiOBr nanozyme catalyst.

2. The method for preparing a BOS@BiOBr nanozyme catalyst according to claim 1, characterized in that: In step (1), the concentration of bismuth nitrate in the bismuth nitrate pentahydrate acetic acid solution is 0.22-0.67 mol / L; the volume ratio of the bismuth nitrate pentahydrate acetic acid solution to anhydrous ethanol is 1:3-7; and the molar ratio of bismuth nitrate pentahydrate to tetraethyl orthosilicate is 10:1-3.

3. The method for preparing a BOS@BiOBr nanozyme catalyst according to claim 1, characterized in that: In step (1), the dropping rate of tetraethyl orthosilicate into the mixed solution is 0.4~10 mL / min.

4. The method for preparing a BOS@BiOBr nanozyme catalyst according to claim 1, characterized in that: In step (1), the heating reaction conditions are: heating the reaction at 80℃-180℃ for 26-56h.

5. The method for preparing a BOS@BiOBr nanozyme catalyst according to claim 1, characterized in that: In step (1), the calcination conditions are: heating to 330℃-470℃ at a heating rate of 5℃ / min, and calcination time is 1-5h.

6. The method for preparing a BOS@BiOBr nanozyme catalyst according to claim 1, characterized in that: In step (2), the concentration of Bi2O3 precursor in the suspension is 1-20 g / L; the mass ratio of polyvinylpyrrolidone to Bi2O3 precursor is 1-2:2; the mass percentage of thioacetamide in the aqueous thioacetamide solution is 0.5%~10%; the volume ratio of the suspension to the aqueous thioacetamide solution is 1:1~2.5; and the heating reaction conditions are: heating at 100℃~200℃ for 2h~8h.

7. The method for preparing a BOS@BiOBr nanozyme catalyst according to claim 1, characterized in that: In step (3), the volume percentage of ethylene glycol in the mixed solution of ultrapure water and ethylene glycol is 50%~80%; in the dispersion, the concentration of BOS is 1.33-3.33 mg / mL, the concentration of polyvinylpyrrolidone is 1.33-5 mg / mL, and the concentration of potassium bromide is 1.93-5.5 mg / mL; the heating reaction conditions are: heating reaction at 80-180℃ for 2-8 h.

8. The application of the BOS@BiOBr nanozyme catalyst prepared by the method according to any one of claims 1-7 in the degradation of tetracycline, characterized in that: The BOS@BiOBr nanozyme catalyst was used to prepare the BOS@BiOBr-ssDNA aptamer probe. The specific steps were as follows: TC-ssDNA was added to a buffer solution and then incubated; after incubation, the BOS@BiOBr nanozyme catalyst was added and incubation continued; after incubation, the probe was washed with Tris-HCl solution to obtain the BOS@BiOBr-ssDNA aptamer probe, which was then applied to the detection of tetracycline.

9. The application of the BOS@BiOBr nanozyme catalyst according to claim 8 in tetracycline degradation, characterized in that: The buffer solution is composed of a mixture of tris(2-carboxyethyl)phosphine hydrochloride solution and Tris-HCl solution; the concentration of tris(2-carboxyethyl)phosphine hydrochloride solution in the buffer solution is 10.0 mg / mL, the concentration of Tris-HCl solution is 10.0 mmol / L, and the pH is 7.5; the volume ratio of tris(2-carboxyethyl)phosphine hydrochloride solution to Tris-HCl solution in the buffer solution is 0.2-1.5:0.5-2.

0.

10. The application of the BOS@BiOBr nanozyme catalyst according to claim 8 in tetracycline degradation, characterized in that: The concentration of TC-ssDNA is 80.0-400.0 nmol / L, and the volume ratio of TC-ssDNA to buffer is 0.05-0.2:2.5-3.0; in the buffer after adding BOS@BiOBr nanozyme catalyst, the concentration of BOS@BiOBr nanozyme catalyst is 0.1-0.5 mg / mL.

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