Application of AuNCs@dmSiO2 composite materials in the detection of organophosphorus pesticide paraoxon

Abstract

The present invention discloses the application of AuNCs@dmSiO2 composite materials in the detection of organophosphorus pesticide paraoxon, belonging to the field of fluorescence sensing technology. The present invention combines mesoporous silica spheres with AuNCs, and through the spatial confinement effect, prepares AuNCs@dmSiO2 fluorescent composite materials with better fluorescence properties. 2+ The quenching effect of the fluorescent composite probe is caused by the reaction product of acetylcholinesterase and substrate acetylthiocholine (ATCh) binding to Cu 2+ This leads to the recovery of probe fluorescence, and organophosphorus pesticides can inactivate acetylcholinesterase, resulting in quenching of the fluorescence system. Therefore, the AuNCs@dmSiO2 composite material is used for the fluorescence detection of the organophosphorus pesticide paraoxon.

Description

Technical Field

[0001] The present invention belongs to the field of fluorescence sensing technology, and specifically relates to the application of AuNCs@dmSiO2 composite materials in the detection of organophosphorus pesticide paraoxon. Background Art

[0002] Organophosphate pesticides (OPs) are widely used in agriculture to control pests and increase crop yields. However, with increasing public concern about food safety, the issue of OP residues has garnered widespread attention. These residues enter the human body through ingestion, skin contact, and inhalation, potentially affecting the nervous, endocrine, and immune systems, manifesting as symptoms such as headaches, nausea, vomiting, and even potentially fatal poisoning. Furthermore, long-term exposure to OPs may increase the risk of cancer, reproductive system abnormalities, and neurological diseases. Therefore, accurate detection of OPs is crucial for maintaining food safety and public health.

[0003] Traditional techniques, such as chromatography and mass spectrometry, have been used to detect organophosphorus pesticide residues. However, these traditional techniques require expensive equipment, complex operating procedures, and are time-consuming. They also require skilled operators, limiting their field application in agricultural settings. Therefore, there is a need to develop efficient methods for detecting organophosphorus pesticides.

[0004] In recent years, fluorescence has emerged as a novel detection technology, demonstrating unique advantages in the detection of organophosphorus pesticide residues. These technologies offer the advantages of high sensitivity, rapid response, ease of operation, and real-time monitoring, and therefore hold broad application prospects and significant practical significance in the detection of organophosphorus pesticides. The development of high-performance fluorescent probes is crucial for improving sensor efficiency. Gold nanoclusters (AuNCs) have garnered significant attention due to their compelling physicochemical properties, including strong photoluminescence, nontoxicity, high water solubility, and excellent biocompatibility. However, the low quantum yield of AuNCs remains a major obstacle to their widespread application. Summary of the Invention

[0005] The present invention aims to provide an application of AuNCs@dmSiO2 composite material in the detection of organophosphorus pesticide paraoxon.

[0006] Based on the above objectives, the present invention adopts the following technical solutions:

[0007] Application of AuNCs@dmSiO2 composite materials (mesoporous silica spheres loaded with gold nanoclusters) in the detection of organophosphorus pesticide paraoxon.

[0008] The application method includes the following steps:

[0009] a. Mix the test solution containing organophosphorus pesticide paraoxon, acetylcholinesterase, acetylthiocholine and a portion of Tris-HCl buffer solution, incubate, and then add Cu 2+ solution and another part of Tris-HCl buffer solution to obtain a mixed solution;

[0010] b. Adding the AuNCs@dmSiO2 composite material aqueous solution to the mixed solution in step a, mixing to obtain a test solution, and performing fluorescence spectrum detection on the test solution;

[0011] Alternatively, the carrier loaded with the AuNCs@dmSiO2 composite material is placed in the mixed solution of step a, immersed, taken out, and the carrier is analyzed.

[0012] In step b, the carrier is a cotton swab. The cotton swab loaded with the AuNCs@dmSiO2 composite material is immersed in the mixed solution of step a. The cotton swab is photographed under ultraviolet light, and the color RGB value of the image is identified. The paraoxon concentration is determined by calculating the color signal ratio R / (G+B).

[0013] In step a, the concentration of Tris-HCl buffer solution is 17-25 mM, the pH value is 5-10, the concentration of acetylcholinesterase is 0.8-1.5 U / mL, the concentration of acetylthiocholine is 0.7-1.5 mM, and the concentration of Cu is 0. 2+ The concentration of the solution is 0.05-0.15 mM; in step b, the concentration of the AuNCs@dmSiO2 composite material aqueous solution is 1.0-1.5 mg / mL, Tris-HCl buffer solution, acetylcholinesterase, acetylthiocholine, Cu 2+ The volume ratio of the AuNCs@dmSiO2 composite material solution and the AuNCs@dmSiO2 composite material aqueous solution is (360-465): (10-15): (15-25): (50-80): (450-650); the incubation conditions are: the incubation temperature is 30-40 °C, and the incubation time is 30-50 min; the AuNCs@dmSiO2 composite material aqueous solution is prepared by dispersing the AuNCs@dmSiO2 composite material in water.

[0014] The preparation method of the AuNCs@dmSiO2 composite material comprises the following steps:

[0015] 1) Preparation of AuNCs solution: Mix glutathione aqueous solution and chloroauric acid aqueous solution, heat and stir, dialysis purification, freeze-dry, and disperse the resulting solid in water to obtain an AuNCs solution;

[0016] 2) Preparation of dmSiO2 dispersion: triethanolamine, hexadecyltrimethylammonium bromide, sodium salicylate, and water are mixed, heated and stirred, ethyl orthosilicate is added, heated and stirred to react, cooled, and the precipitate is collected. After washing and purification, the precipitate is dispersed in an organic solvent, and ammonia water and (3-aminopropyl)triethoxysilane are added. After stirring and reacting, the product is collected, washed, and freeze-dried to obtain mesoporous silica spheres. The mesoporous silica spheres are dispersed in water to obtain a dmSiO2 dispersion;

[0017] 3) Preparation of AuNCs@dmSiO2 composite material: The AuNCs solution from step 1) and the dmSiO2 dispersion from step 2) were mixed and freeze-dried to obtain the AuNCs@dmSiO2 composite material (mesoporous silica spheres loaded with gold nanoclusters).

[0018] In step 1), the molar ratio of glutathione in the glutathione aqueous solution to chloroauric acid in the chloroauric acid aqueous solution is (1-3):2; the heating and stirring conditions are: temperature of 65-75°C, stirring speed of 500-800 rpm, and time of 20-30 h.

[0019] In step 2), the amount ratio of triethanolamine, cetyltrimethylammonium bromide, sodium salicylate, ethyl orthosilicate, ammonia water and (3-aminopropyl)triethoxysilane is (60-80) mg: (360-400) mg: (140-180) mg: (3-5) mL: (2-4) mL: (0.5-1.5) mL. Triethanolamine, cetyltrimethylammonium bromide, sodium salicylate and water are mixed at a stirring speed of 500-800 rpm, stirred at 80-90°C for 1-2 h, ethyl orthosilicate is added, and the mixture is stirred at 70-85°C for 2-3 h. After cooling, the precipitate is collected, washed and purified, and then dispersed in an organic solvent. Ammonia water and (3-aminopropyl)triethoxysilane are added at a stirring speed of 500-800 rpm, and the mixture is stirred for 10-15 h. The organic solvent is ethanol.

[0020] In step 3), the mass ratio of dmSiO2 in the dmSiO2 dispersion and AuNCs in the AuNCs solution is (1-5):1, the mixing conditions are: temperature 20-35 °C, time 10-20 min; the particle size of the AuNCs@dmSiO2 composite material is 250-300 nm.

[0021] The preparation method of the cotton swab loaded with AuNCs@dmSiO2 composite material is as follows: placing the cotton swab in a solution containing the AuNCs@dmSiO2 composite material and gelatin, soaking it for 10-15 minutes, taking it out, and freeze-drying it; in the solution, the concentration of the AuNCs@dmSiO2 composite material is 1.0-1.5 mg / mL, and the concentration of gelatin is 4.0-7.0 mg / mL.

[0022] In step b, a cotton swab loaded with AuNCs@dmSiO2 composite material is placed in the mixed solution of step a, and each cotton swab is loaded with 0.50-0.75 mg of AuNCs@dmSiO2 composite material. One cotton swab can be used for 500 μL of the mixed solution. In step a, the concentration of Tris-HCl buffer solution is 17-25 mM, the pH value is 5-10, the concentration of acetylcholinesterase is 0.8-1.5 U / mL, the concentration of acetylthiocholine is 0.7-1.5 mM, and the concentration of Cu is 0. 2+ The concentration of the solution is 0.05-0.15 mM, Tris-HCl buffer solution, acetylcholinesterase, acetylthiocholine, Cu 2+ The volume ratio of the solution is (360-465): (10-15): (15-25): (50-80); incubation conditions: incubation temperature is 30-40 °C, and incubation time is 30-50 min.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] 1) In this invention, mesoporous silica spheres are combined with AuNCs to prepare AuNCs@dmSiO2 fluorescent composite materials with better fluorescence properties through spatial confinement effect. 2+ The quenching effect of the fluorescent composite probe is caused by the reaction product of acetylcholinesterase and substrate acetylthiocholine (ATCh) binding to Cu 2+ Leading to the recovery of probe fluorescence, and organophosphorus pesticides can inactivate acetylcholinesterase, leading to quenching of the fluorescence system, the sensing mechanism is as follows Figure 13 As shown, therefore, the AuNCs@dmSiO2 composite material was used for the fluorescence detection of organophosphorus pesticide paraoxon;

[0025] 2) The present invention prepares gold nanoclusters (AuNCs) and pomegranate-shaped mesoporous silica spheres (dmSiO2) with optimal fluorescence emission. The dmSiO2 dispersion and AuNCs solution are then mixed. Based on electrostatic interaction, the confinement effect of dmSiO2 is utilized to enhance the fluorescence of the gold nanoclusters, effectively overcoming the weak fluorescence intensity of AuNCs. The resulting AuNCs@dmSiO2 composite material exhibits optimal fluorescence intensity. This method is simple, rapid, and pollution-free.

[0026] 3) The AuNCs@dmSiO2 composite material of the present invention is applied to the fluorescence detection method of the organophosphorus pesticide paraoxon. The steps are simple and rapid, and it can achieve highly selective and specific quantitative detection of organophosphorus pesticides with a detection limit of 0.032 ng / mL.

[0027] 4) The present invention prepares a portable detection cotton swab by loading the AuNCs@dmSiO2 composite material on a cotton swab, which can detect the organophosphorus pesticide paraoxon, providing a promising portable technology for the qualitative identification and quantitative detection of the organophosphorus pesticide paraoxon on site. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] In order to more clearly illustrate the technical solution of the present invention, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without paying any creative work.

[0029] Figure 1 This is the fluorescence emission spectrum of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 1;

[0030] Figure 2 This is the fluorescence emission spectrum of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 2;

[0031] Figure 3 This is the fluorescence emission spectrum of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 3;

[0032] Figure 4 This is the fluorescence emission spectrum of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 4;

[0033] Figure 5 This is a fluorescence emission spectrum of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 5;

[0034] Figure 6 The fluorescence emission spectra of the gold nanoclusters, mesoporous silica spheres, and mesoporous silica sphere-loaded gold nanoclusters prepared in Example 5;

[0035] Figure 7 This is the SEM spectrum of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 5;

[0036] Figure 8 TEM spectrum of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 5;

[0037] Figure 9The AuNCs solution (gold nanoclusters) and AuNCs@dmSiO2 composite material (mesoporous silica spheres loaded with gold nanoclusters) prepared in Example 5 were used to measure the Cu 2+ Sensing test results, where 9A and 9B are the results of the AuNCs solution (gold nanoclusters) prepared in Example 5, 9A is the fluorescence spectrum, Figure 9 B is Cu 2+ The concentration is the horizontal axis, and the fluorescence intensity ratio signal (F / F0) is the vertical axis fitting straight line; 9C and 9D are the results of the AuNCs@dmSiO2 composite material prepared in Example 5, 9C is the fluorescence spectrum, and 9D is the Cu 2+ The concentration is the horizontal axis, and the fluorescence intensity ratio signal (F / F0) is the vertical axis fitting straight line;

[0038] Figure 10 10A is the fluorescence spectrum of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 5 detecting different concentrations of paraoxon, and 10B is a fitted straight line with the paraoxon concentration as the abscissa and the fluorescence intensity ratio signal (F / F0) as the ordinate, where F and F0 represent the fluorescence intensity in the presence and absence of paraoxon in the detection system, respectively;

[0039] Figure 11 The selectivity test results of the mesoporous silica sphere-loaded gold nanocluster material prepared in Example 5 for detecting organophosphorus pesticides;

[0040] Figure 12 middle, Figure 12 A is a schematic diagram of the preparation process of a cotton swab loaded with AuNCs@dmSiO2 composite material. Figure 12 B is a photo of a cotton swab taken under ultraviolet light containing different concentrations of the organophosphorus pesticide paraoxon. Figure 12 C is the fitting curve between the color signal ratio (R / (G+B)) and the paraoxon concentration;

[0041] Figure 13 The sensing mechanism of the AuNCs@dmSiO2 composite material of the present invention. DETAILED DESCRIPTION

[0042] To make the objectives, technical solutions, and advantages of the present invention more clear, the technical solutions of the present invention are described in detail below. However, the following embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other implementation methods obtained by those skilled in the art without making any creative work are within the scope of protection of the present invention.

[0043] Example 1

[0044] The preparation method of the AuNCs@dmSiO2 composite material comprises the following steps:

[0045] 1) Preparation of AuNCs solution: Dissolve glutathione and chloroauric acid in water to prepare 50 mM glutathione and 5 mM chloroauric acid solutions, respectively. Mix 0.6 mL of the glutathione solution and 4.0 mL of the chloroauric acid solution. Stir at 65°C and 600 rpm for 20 h. After purification by dialysis and freeze-drying, the resulting solid was dispersed in water to obtain a 1 mg / mL AuNCs solution.

[0046] 2) Preparation of dmSiO2 dispersion: 68 mg of triethanolamine, 380 mg of cetyltrimethylammonium bromide, and 168 mg of sodium salicylate were mixed with 25 mL of water at a stirring speed of 500 rpm. The mixture was stirred at 80 °C for 1 h. 4 mL of ethyl orthosilicate was added. The mixture was stirred at 70 °C for 2 h, and then cooled to room temperature. The precipitate was collected by centrifugation and washed three times with ethanol (analytical grade, the same below). The precipitate was dispersed in a mixture of 50 mL of methanol (analytical grade, the same below) and 3 mL of hydrochloric acid (mass concentration 38%, the same below), and refluxed at 60 °C for 6 h. The purified precipitate was dispersed in 100 mL of ethanol. 2.5 mL of ammonia (mass concentration 26%, the same below) and 1 mL of (3-aminopropyl)triethoxysilane (APTES, analytical grade, the same below) were added at a stirring speed of 800 rpm. The mixture was stirred at room temperature for 12 h. h, collect the product, wash it three times with ethanol, then wash it three times with deionized water, freeze-dry it for 12 h to obtain mesoporous silica spheres, and disperse the mesoporous silica spheres in water to obtain a 2 mg / mL dmSiO2 dispersion;

[0047] 3) Preparation of AuNCs@dmSiO2 composite material: Mix 2 mL of the AuNCs solution (1 mg / mL) prepared in step 1) and 1 mL of the dmSiO2 dispersion (2 mg / mL) prepared in step 2), stir at 25°C for 10 min, and freeze-dry to obtain the AuNCs@dmSiO2 composite material.

[0048] A certain amount of AuNCs@dmSiO2 was dispersed in water to obtain a 0.4 mg / mL AuNCs@dmSiO2 aqueous solution. Its fluorescence spectrum is shown in the figure below. Figure 1 shown.

[0049] Example 2

[0050] The difference from Example 1 is that in step 2), the mesoporous silica spheres are dispersed in water to prepare a 4 mg / mL dmSiO2 dispersion;

[0051] 3) Preparation of AuNCs@dmSiO2 composite material: Mix 2 mL of the AuNCs solution (1 mg / mL) prepared in step 1) and 1 mL of the dmSiO2 dispersion (4 mg / mL) prepared in step 2), stir at 25 °C for 10 min, and freeze-dry to obtain the AuNCs@dmSiO2 composite material.

[0052] A certain amount of AuNCs@dmSiO2 was dispersed in water to obtain a 0.6 mg / mL AuNCs@dmSiO2 aqueous solution. Its fluorescence spectrum is shown in the figure below. Figure 2 shown.

[0053] Example 3

[0054] The difference from Example 1 is that in step 2), the mesoporous silica spheres are dispersed in water to prepare a 6 mg / mL dmSiO2 dispersion;

[0055] 3) Preparation of AuNCs@dmSiO2 composite material: Mix 2 mL of the AuNCs solution (1 mg / mL) from step 1) and 1 mL of the dmSiO2 dispersion (6 mg / mL) from step 2), stir at 25 °C for 10 min, and freeze-dry to obtain the AuNCs@dmSiO2 composite material.

[0056] A certain amount of AuNCs@dmSiO2 was dispersed in water to obtain a AuNCs@dmSiO2 aqueous solution with a concentration of 0.8 mg / mL. Its fluorescence spectrum is shown in the figure below. Figure 3 shown.

[0057] Example 4

[0058] The difference from Example 1 is that in step 2), the mesoporous silica spheres are dispersed in water to prepare an 8 mg / mL dmSiO2 dispersion;

[0059] 3) Preparation of AuNCs@dmSiO2 composite material: Mix 2 mL of the AuNCs solution (1 mg / mL) prepared in step 1) and 1 mL of the dmSiO2 dispersion (8 mg / mL) prepared in step 2), stir at 25 °C for 10 min, and freeze-dry to obtain the AuNCs@dmSiO2 composite material.

[0060] A certain amount of AuNCs@dmSiO2 was dispersed in water to obtain a 1.0 mg / mL AuNCs@dmSiO2 aqueous solution. Its fluorescence spectrum is shown in the figure below. Figure 4 shown.

[0061] Example 5

[0062] The difference from Example 1 is that in step 2), the mesoporous silica spheres are dispersed in water to prepare a 10 mg / mL dmSiO2 dispersion;

[0063] 3) Preparation of AuNCs@dmSiO2 composite material: Mix 2 mL of the AuNCs solution (1 mg / mL) prepared in step 1) and 1 mL of the dmSiO2 dispersion (10 mg / mL) prepared in step 2), stir at 25°C for 10 min, and freeze-dry to obtain the AuNCs@dmSiO2 composite material.

[0064] A certain amount of AuNCs@dmSiO2 composite material was dispersed in water to obtain a AuNCs@dmSiO2 aqueous solution with a concentration of 1.2 mg / mL. Its fluorescence spectrum is shown in the figure below. Figure 5 shown.

[0065] Depend on Figure 1-5 It is found that under the same mass of AuNCs, the fluorescence intensity of the AuNCs@dmSiO2 composite material prepared in Example 5 is the highest.

[0066] The AuNCs solution and dmSiO2 dispersion prepared in Example 5 were subjected to fluorescence analysis. The fluorescence spectra of the AuNCs solution, dmSiO2 dispersion and AuNCs@dmSiO2 aqueous solution prepared in Example 5 are shown in Figure 2. Figure 6 shown.

[0067] from Figure 6 It can be seen from the figure that dmSiO2 has no fluorescence. Before and after dmSiO2 loading, the fluorescence of AuNCs is enhanced by 2.6 times, and the fluorescence emission peak is blue-shifted by 6 nm from 604 nm to 598 nm.

[0068] The SEM image of the AuNCs@dmSiO2 composite material (mesoporous silica spheres loaded with gold nanoclusters) prepared in Example 5 is shown in FIG. Figure 7 As shown in the TEM image Figure 8 shown.

[0069] from Figure 7 It can be seen that the mesoporous silica sphere-loaded gold nanocluster material of the present invention has a pomegranate-shaped morphology and a uniform size with a particle diameter of 250-300 nm.

[0070] from Figure 8 It can be seen that the mesoporous silica sphere-loaded gold nanocluster material has radial pore channels, which is more conducive to the enrichment of the substrate and improves the detection sensitivity.

[0071] Example 6

[0072] The preparation method of the AuNCs@dmSiO2 composite material comprises the following steps:

[0073] 1) Preparation of AuNCs solution: Glutathione and chloroauric acid were dissolved in water to prepare 50 mM glutathione and 5 mM chloroauric acid solutions, respectively. 0.4 mL of the glutathione solution and 4.0 mL of the chloroauric acid solution were mixed and stirred at 65°C and 500 rpm for 28 h. The mixture was then dialyzed and freeze-dried, and the resulting solid was dispersed in water to obtain the AuNCs solution.

[0074] 2) Preparation of dmSiO2 dispersion: 75 mg of triethanolamine, 360 mg of cetyltrimethylammonium bromide, and 180 mg of sodium salicylate were mixed with 30 mL of water at a stirring speed of 700 rpm, stirred at 90 °C for 2 h, 3 mL of ethyl orthosilicate was added, and the mixture was stirred at 85 °C for 3 h. After cooling to room temperature, the precipitate was collected by centrifugation and washed three times with ethanol. The precipitate was dispersed in a mixture of 55 mL of methanol and 4 mL of hydrochloric acid and refluxed at 60 °C for 5 h. The purified precipitate was dispersed in 110 mL of ethanol and stirred at 800 rpm. 3 mL of ammonia and 1.5 mL of (3-aminopropyl)triethoxysilane (APTES) were added. The mixture was stirred at room temperature for 15 h. The product was collected, washed three times with ethanol and three times with deionized water, and freeze-dried for 12 h. h, obtain mesoporous silica spheres, disperse the mesoporous silica spheres (dmSiO2) in water to obtain a dmSiO2 dispersion;

[0075] 3) Preparation of AuNCs@dmSiO2 composite material: The AuNCs solution from step 1) and the dmSiO2 dispersion from step 2) were mixed at a mass ratio of dmSiO2 to AuNCs of 2:1, stirred at 25°C for 10 min, and freeze-dried to obtain the AuNCs@dmSiO2 composite material.

[0076] A certain amount of AuNCs@dmSiO2 was dispersed in water to obtain an AuNCs@dmSiO2 aqueous solution.

[0077] Example 7

[0078] The preparation method of the AuNCs@dmSiO2 composite material comprises the following steps:

[0079] 1) Preparation of AuNCs solution: Glutathione and chloroauric acid were dissolved in water to prepare 50 mM glutathione and 5 mM chloroauric acid solutions, respectively. 0.2 mL of the glutathione solution and 4.0 mL of the chloroauric acid solution were mixed and stirred at 85°C and 800 rpm for 30 h. The mixture was then dialyzed for purification and freeze-dried. The resulting solid was dispersed in water to obtain the AuNCs solution.

[0080] 2) Preparation of dmSiO2 dispersion: 80 mg of triethanolamine, 400 mg of cetyltrimethylammonium bromide, and 180 mg of sodium salicylate were mixed with 35 mL of water at a stirring speed of 600 rpm, stirred at 80 °C for 2 h, 5 mL of ethyl orthosilicate was added, and the mixture was stirred at 70 °C for 3 h. After cooling to room temperature, the precipitate was collected by centrifugation and washed three times with ethanol. The precipitate was dispersed in a mixture of 60 mL of methanol and 5 mL of hydrochloric acid and refluxed at 60 °C for 6 h. The purified precipitate was dispersed in 120 mL of ethanol and stirred at 800 rpm. 4 mL of ammonia and 0.5 mL of (3-aminopropyl)triethoxysilane (APTES) were added and stirred at room temperature for 15 h. The product was collected, washed three times with ethanol and three times with deionized water, and freeze-dried for 12 h. h, obtaining mesoporous silica spheres, and dispersing the mesoporous silica spheres in water to obtain a dmSiO2 dispersion;

[0081] 3) Preparation of AuNCs@dmSiO2 composite material: The AuNCs solution from step 1) and the dmSiO2 dispersion from step 2) were mixed at a mass ratio of dmSiO2 to AuNCs of 5:1, stirred at 25°C for 20 min, and freeze-dried to obtain the AuNCs@dmSiO2 composite material.

[0082] A certain amount of AuNCs@dmSiO2 was dispersed in water to obtain an AuNCs@dmSiO2 aqueous solution.

[0083] Example 8

[0084] Preparation method of cotton swabs loaded with AuNCs@dmSiO2 composite materials: place a medical cotton swab in a solution containing the AuNCs@dmSiO2 composite material prepared in Example 5 above and gelatin, soak for 10 minutes, take out, and freeze-dry to obtain; in the solution, the concentration of AuNCs@dmSiO2 composite material is 1.2 mg / mL, and the concentration of gelatin is 5.0 mg / mL. Each cotton swab is loaded with 0.60 mg of AuNCs@dmSiO2 composite material.

[0085] Example 9 Sensitivity Detection Test

[0086] Similarly, 60 μL of different concentrations of Cu 2+The solution was mixed with 340 μL of Tris-HCl buffer (pH 6.0, 20 mM), 500 μL of AuNCs solution (0.2 mg / mL, prepared by diluting the AuNCs solution in Example 5), and 100 μL of H2O to obtain a mixed solution. The excitation wavelength was 405 nm, the emission wavelength was 604 nm, the excitation light source slit was 10 nm, and the emission light source slit was 5 nm. Fluorescence detection was performed using a 1 cm × 1 cm cuvette. The fluorescence spectrum is shown in Figure 9A. Figure 9 B is Cu 2+ The concentration is the horizontal axis and the fluorescence intensity ratio signal (F / F0) is the vertical axis for fitting the straight line.

[0087] 60 μL of Cu 2+ The solution was mixed with 340 μL of Tris-HCl buffer (pH 6.0, 20 mM), 500 μL of AuNCs@dmSiO2 aqueous solution (1.2 mg / mL, prepared by dispersing the AuNCs@dmSiO2 composite material from Example 5 in water), and 100 μL of H2O to obtain a mixed solution. The excitation wavelength was 405 nm, the emission wavelength was 598 nm, the excitation light source slit was 10 nm, and the emission light source slit was 5 nm. Fluorescence detection was performed using a 1 cm × 1 cm cuvette. The fluorescence spectrum is shown in Figure 9C. Figure 9 D is Cu 2+ The concentration is the horizontal axis and the fluorescence intensity ratio signal (F / F0) is the vertical axis for fitting the straight line.

[0088] Figure 9 Cu in A, 9B, 9C, and 9D 2+ The concentration of Cu in the mixed solution 2+ final concentration.

[0089] Figure 9 A sees that as Cu 2+ With the increase of concentration, the fluorescence of AuNCs gradually decreased; Figure 9 B can be seen that this method has a great 2+ The detectable range is 0.1-3.0 μM, and the linear equation is F / F0=1.004-0.147[Cu 2+ ], ng / mL, with a detection limit of 35 nM.

[0090] Figure 9 C shows that as Cu 2+ With the increase of concentration, the fluorescence intensity of AuNCs@dmSiO2 gradually decreased; Figure 9 D can be seen that this method has a great 2+The detectable range is 0.005-3.0 μM, and the linear equation is F0 / F=0.983-0.191[Cu 2+ ], ng / mL, with a detection limit of 2.8 nM. The Cu 2+ The detection limit was as low as 1 / 10 of that achieved using AuNCs as a probe. This suggests that the porous structure of the dmSiO2 support material provides a larger surface area, increasing the contact area between the probe and the target substance, making the probe more sensitive to the target substance. Furthermore, the richly porous dmSiO2 matrix acts as a high-performance adsorbent, effectively adsorbing and enriching the target substance, promoting its diffusion and transport, and thus improving the response sensitivity of the analyte.

[0091] Example 10 Detection Test of Organophosphorus Pesticide Paraoxon

[0092] The AuNCs@dmSiO2 composite material is used in the detection of the organophosphorus pesticide paraoxon. The application method includes the following steps:

[0093] a. 40 μL of the test solution containing different concentrations of the organophosphorus pesticide paraoxon was mixed with 12 μL of acetylcholinesterase (1.0 U / mL), 20 μL of acetylthiocholine (1.0 mM), and 28 μL of Tris-HCl buffer solution (pH 7.4, 20 mM). The mixture was incubated at 37°C for 40 min, and then 60 μL of Cu was added. 2+ solution (0.1 mM), 340 μL Tris-HCl buffer solution (pH 6.0, 20 mM) to obtain a mixed solution;

[0094] b. Add 500 μL of the AuNCs@dmSiO2 composite material aqueous solution (1.2 mg / mL) from Example 5 to the mixed solution in step a and mix to obtain a test solution. The excitation wavelength was 405 nm, the emission wavelength was 598 nm, the excitation light source slit was 10 nm, and the emission light source slit was 5 nm. Fluorescence spectroscopy of the test solution was performed using a 1 cm × 1 cm cuvette. The fluorescence spectrum is shown in Figure 10A. Figure 10 B is the fitted straight line with paraoxon concentration as the abscissa and fluorescence intensity ratio signal (F / F0) as the ordinate. The final paraoxon concentrations in each test solution were 0 ng / mL, 0.05 ng / mL, 0.1 ng / mL, 0.5 ng / mL, 1 ng / mL, 2 ng / mL, 4 ng / mL, 6 ng / mL, 8 ng / mL, 10 ng / mL, 15 ng / mL, 20 ng / mL, 25 ng / mL, 30 ng / mL, 40 ng / mL, and 50 ng / mL.

[0095] Figure 10 A shows that with the increase of paraoxon concentration, the fluorescence of AuNCs@dmSiO2 gradually decreases; Figure 10 As can be seen from B, the detectable range of this method for paraoxon is 0.05-25 ng / mL, the linear equation is F0 / F=1.061+0.046[paraoxon], ng / mL, and the detection limit is 0.032 ng / mL, thereby achieving sensitive detection of paraoxon.

[0096] Example 11 Systematic Method Selectivity Test for Organophosphorus Pesticides

[0097] 40 μL of other pesticides (propynyl, chlorpyrifos, imidacloprid, bifenthrin, cypermethrin) (0.625 μg / mL), 40 μL of ions (Na + Mg 2+ , K + 、Cl - 、NO3 - 、SO4 2- , I - ) (1.25 mM), 40 μL of amino acids (serine, methionine, aspartic acid, proline, glycine, valine, tyrosine) (1.25 mM), 40 μL of other biomolecules (creatinine, cholesterol, urea, glucose, sucrose, galactose) (1.25 mM) were mixed with 12 μL of acetylcholinesterase (1.0 U / mL), 20 μL of acetylthiocholine (1.0 mM), and 28 μL of Tris-HCl buffer solution (pH 7.4, 20 mM), and incubated at 37°C for 40 min. Then, 60 μL of Cu 2+ (0.1 mM), 340 μL Tris-HCl buffer solution (pH 6.0, 20 mM) and 500 μL AuNCs@dmSiO2 aqueous solution (1.2 mg / mL) of Example 5 were added, and fluorescence detection was performed on each test solution. The fluorescence spectrum is shown in Figure 2. Figure 11 shown.

[0098] Figure 11 It can be seen that the fluorescence intensity is significantly reduced only in the presence of the organophosphorus pesticide paraoxon, and there is no obvious change in the presence of other types of substances, which shows that the sensing system has high selectivity for organophosphorus pesticides.

[0099] Example 12: Cotton swabs loaded with AuNCs@dmSiO2 composite materials were used to detect organophosphorus pesticide paraoxon at different concentrations.

[0100] The application method is as follows: a. 40 μL of the test solution containing different concentrations of the organophosphorus pesticide paraoxon was mixed with 12 μL of acetylcholinesterase (1.0 U / mL), 20 μL of acetylthiocholine (1.0 mM) and 28 μL of Tris-HCl buffer solution (pH 7.4, 20 mM), incubated at 37 °C for 40 min, and then 60 μL of Cu was added. 2+ Solution (0.1 mM), 340 μL Tris-HCl buffer solution (pH 6.0, 20 mM), were added to prepare 8 mixtures containing different paraoxon concentrations; the final concentration of paraoxon in each mixture was 0 ng / mL, 2 ng / mL, 6 ng / mL, 10 ng / mL, 20 ng / mL, 40 ng / mL, 50 ng / mL, and 70 ng / mL;

[0101] b. Eight cotton swabs loaded with the AuNCs@dmSiO2 composite material from Example 8 were immersed in each of the eight portions of the mixed solution from step a. The cotton swabs were photographed under a 365 nm UV lamp. The RGB values ​​of the images were identified using a color recognition program (ColorCollection) on a smartphone. The paraoxon concentration was determined by calculating the color signal ratio R / (G+B).

[0102] The process diagram is as follows Figure 12 As shown in A, the test results are as follows Figure 12 As shown in B, Figure 12 C is the fitted straight line between the color signal ratio (R / (G+B)) and the paraoxon concentration. Figure 12 In B, from left to right and from top to bottom, the final concentrations of paraoxon in the mixed solution corresponding to the cotton swabs are 0 ng / mL, 2 ng / mL, 6 ng / mL, 10 ng / mL, 20 ng / mL, 40 ng / mL, 50 ng / mL, and 70 ng / mL.

[0103] Figure 12 As shown in B, the detection cotton swab has obvious fluorescence dimming and visible fluorescence changes under 365 nm ultraviolet light; Figure 12 C shows that the detection range of paraoxon by this detection method is 2-70 ng / mL, the linear equation is R / (G+B) = 0.656-0.00401[paraoxon], ng / mL, and the detection limit is 0.65 ng / mL, which provides a portable and simple method for the detection of organophosphorus pesticide paraoxon.

Claims

1. Application of AuNCs@dmSiO2 composite material in the detection of organophosphorus pesticide paraoxon, characterized by: The preparation method of the AuNCs@dmSiO2 composite material comprises the following steps: 1) Preparation of AuNCs solution: Mix glutathione aqueous solution and chloroauric acid aqueous solution, heat and stir, dialysis purification, freeze-dry, and disperse the resulting solid in water to obtain an AuNCs solution; 2) Preparation of dmSiO2 dispersion: triethanolamine, hexadecyltrimethylammonium bromide, sodium salicylate, and water are mixed, heated and stirred, ethyl orthosilicate is added, heated and stirred to react, cooled, and the precipitate is collected. After washing and purification, the precipitate is dispersed in an organic solvent, and ammonia water and (3-aminopropyl)triethoxysilane are added. After stirring and reacting, the product is collected, washed, and freeze-dried to obtain mesoporous silica spheres. The mesoporous silica spheres are dispersed in water to obtain a dmSiO2 dispersion; 3) Preparation of AuNCs@dmSiO2 composite material: The AuNCs solution prepared in step 1) and the dmSiO2 dispersion prepared in step 2) were mixed and freeze-dried to obtain the AuNCs@dmSiO2 composite material.

2. The use according to claim 1, characterized in that The application method includes the following steps: a. Mix the test solution containing organophosphorus pesticide paraoxon, acetylcholinesterase, acetylthiocholine and a portion of Tris-HCl buffer solution, incubate, and then add Cu 2+ solution and another part of Tris-HCl buffer solution to obtain a mixed solution; b. Adding the AuNCs@dmSiO2 composite material aqueous solution to the mixed solution in step a, mixing to obtain a test solution, and performing fluorescence spectrum detection on the test solution; Alternatively, the carrier loaded with the AuNCs@dmSiO2 composite material is placed in the mixed solution of step a, immersed, taken out, and the carrier is analyzed.

3. The use according to claim 2, characterized in that In step b, the carrier is a cotton swab. The cotton swab loaded with the AuNCs@dmSiO2 composite material is immersed in the mixed solution of step a. The cotton swab is photographed under ultraviolet light, and the color RGB value of the image is identified. The paraoxon concentration is determined by calculating the color signal ratio R / (G+B).

4. The use according to claim 2, characterized in that In step a, the concentration of Tris-HCl buffer solution is 17-25 mM, the pH value is 5-10, the concentration of acetylcholinesterase is 0.8-1.5 U / mL, the concentration of acetylthiocholine is 0.7-1.5 mM, and the concentration of Cu is 0. 2+ The concentration of the solution is 0.05-0.15 mM; in step b, the concentration of the AuNCs@dmSiO2 composite material aqueous solution is 1.0-1.5 mg / mL, Tris-HCl buffer solution, acetylcholinesterase, acetylthiocholine, Cu 2+ The volume ratio of the AuNCs@dmSiO2 composite material solution and the AuNCs@dmSiO2 composite material aqueous solution is (360-465): (10-15): (15-25): (50-80): (450-650); the incubation conditions are: the incubation temperature is 30-40 °C, and the incubation time is 30-50 min; the AuNCs@dmSiO2 composite material aqueous solution is prepared by dispersing the AuNCs@dmSiO2 composite material in water.

5. The use according to claim 1, characterized in that In step 1), the molar ratio of glutathione in the glutathione aqueous solution to chloroauric acid in the chloroauric acid aqueous solution is (1-3):2; the heating and stirring conditions are: temperature of 65-75°C, stirring speed of 500-800 rpm, and time of 20-30 h.

6. The use according to claim 1, wherein In step 2), the ratio of triethanolamine, cetyltrimethylammonium bromide, sodium salicylate, ethyl orthosilicate, aqueous ammonia, and (3-aminopropyl)triethoxysilane is (60-80) mg: (360-400) mg: (140-180) mg: (3-5) mL: (2-4) mL: (0.5-1.5) mL. Triethanolamine, cetyltrimethylammonium bromide, sodium salicylate, and water are mixed at a stirring speed of 500-800 rpm, stirred at 80-90°C for 1-2 h, and ethyl orthosilicate is added. The mixture is stirred at 70-85°C for 2-3 h, cooled, and the precipitate is collected. After washing and purification, the precipitate is dispersed in an organic solvent. Aqueous ammonia and (3-aminopropyl)triethoxysilane are added at a stirring speed of 500-800 rpm, and the mixture is stirred for 10-15 h.

7. The use according to claim 1, wherein In step 3), the mass ratio of dmSiO2 in the dmSiO2 dispersion to AuNCs in the AuNCs solution is (1-5):1, and the mixing conditions are: temperature of 20-35 °C and time of 10-20 min; The particle size of AuNCs@dmSiO2 composite materials is 250-300 nm.

8. The use according to claim 3, characterized in that The preparation method of the cotton swab loaded with the AuNCs@dmSiO2 composite material is as follows: placing the cotton swab in a solution containing the AuNCs@dmSiO2 composite material and gelatin, soaking it for 10-15 minutes, taking it out, and freeze-drying it; in the solution, the concentration of the AuNCs@dmSiO2 composite material is 1.0-1.5 mg / mL, and the concentration of gelatin is 4.0-7.0 mg / mL.

9. The use according to claim 8, characterized in that In step b, a cotton swab loaded with AuNCs@dmSiO2 composite material was placed in the mixed solution of step a, with each cotton swab loaded with 0.50-0.75 mg of AuNCs@dmSiO2 composite material.

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