Method for detecting paraquat based on GSH-Zn-Ag-In-S quantum dots

Abstract

The invention belongs to the technical field of fluorescent nano quantum dot material preparation and chemical analysis and detection, and particularly relates to a method for detecting paraquat based on GSH-Zn-Ag-In-S quantum dots. According to the method, the GSH-Zn-Ag-In-S quantum dots and paraquat are subjected to specific oxidation-reduction reaction, and an obvious aggregation phenomenon is generated, so that the fluorescence color and intensity are changed, and qualitative and quantitative detection of paraquat pesticide in food is realized. The method has the advantages of being good in specificity, high in sensitivity, high in response speed and the like, and a new means can be provided for paraquat content determination.

Description

Technical Field

[0001] The invention belongs to the technical field of fluorescent nano quantum dot material preparation and chemical analysis detection, and specifically relates to a method for detecting paraquat based on GSH-Zn-Ag-In-S quantum dots. Background Art

[0002] Paraquat, a bipyridine pesticide, was used extensively in my country in the early days because of its powerful weed-killing effect. Paraquat is extremely toxic and can cause damage to multiple organs in the human body if ingested by mistake. There is no effective treatment, and even a trace amount of ingestion can cause irreversible damage to the human body. Therefore, China has revoked the registration and production license of paraquat since July 1, 2014. It has been stopped from being sold in the Chinese market and is strictly prohibited from being used in fruit and vegetable cultivation. However, China still relies on foreign imports to varying degrees for some types of fruits and vegetables. In fact, more than half of the countries in the world, such as the United States, Japan, France and other countries, have the habit of using paraquat. Since fruits and vegetables are indispensable materials for supplementing essential vitamins and trace elements in daily life, and are also an important way for pesticides to invade the human body, the development of new methods for detecting paraquat is of great practical significance for regulating pesticide residues and maintaining public safety.

[0003] Common paraquat detection methods include chromatography, spectroscopy, and chromatography / mass spectrometry. Although these methods have the advantages of high sensitivity, good repeatability, and good stability, they still have the disadvantages of low detection efficiency, high instrument cost, complex operation, and inability to achieve rapid and instant detection outside the laboratory. In summary, given the defects of previous detection methods, designing and developing new paraquat detection methods has important practical significance.

[0004] In recent years, the fluorescent visualization array sensing method based on nanomaterials has been increasingly sought after by researchers for the detection of pesticides. Nanomaterials not only have the advantages of high stability, simple synthesis, and good biocompatibility, but also have unique quantum effects and size advantages. Therefore, further establishing a fluorescent sensing visualization method based on nanomaterials to identify paraquat can provide a new strategy for the rapid detection and real-time supervision of paraquat in food. Summary of the invention

[0005] In view of the shortcomings in the prior art, the purpose of the present invention is to provide a method for detecting paraquat using a nano fluorescent quantum dot colorimetric sensor, which utilizes the strong earthy yellow fluorescence of GSH-Zn-Ag-In-S quantum dots in the nano fluorescent quantum dot colorimetric sensor, and the fluorescence spectrum shows the presence of two emission peaks. GSH-Zn-Ag-In-S quantum dots undergo a specific redox reaction with paraquat and produce an obvious aggregation phenomenon, which changes the fluorescence color and the fluorescence intensity at 540±5nm, thereby achieving qualitative and quantitative detection of paraquat pesticides in samples to be tested, including food and agricultural products. The method has the advantages of simple operation, rapidity and sensitivity, and can achieve rapid and accurate identification of paraquat pesticides in food and agricultural products.

[0006] To achieve the above object, the technical solution adopted by the present invention is:

[0007] A method for detecting paraquat using a nano fluorescent quantum dot colorimetric sensor, wherein the nano fluorescent quantum dot colorimetric sensor comprises a GSH-Zn-Ag-In-S quantum dot stock solution and a Tris-HCl buffer solution, the nano fluorescent quantum dot colorimetric sensor is mixed with a sample solution to be tested, the mixture is allowed to stand for reaction, a fluorescent visualization image of paraquat is obtained by collecting the obtained detection system based on GSH-Zn-Ag-In-S quantum dots for identifying the paraquat, and the paraquat content in the sample solution to be tested is detected (determining whether the paraquat pesticide residue exceeds the standard) according to the fluorescent color of the detection system obtained after the reaction of the sample solution to be tested, wherein the paraquat concentration in the sample solution to be tested is preferably not more than 1 mg / L, more preferably 0.01 mg / L-1 mg / L, and the solution presents a fluorescent color difference from dark blue to blue-purple to khaki as the paraquat concentration decreases from high to low; and / or

[0008] The nano fluorescent quantum dot colorimetric sensor is mixed with the sample solution to be tested, allowed to stand for reaction, and the fluorescence emission spectrum of the resulting detection system is measured. If the fluorescence intensity ratio F0 / F at 540±5nm before and after the addition of the sample solution to be tested is greater than 2 (F0 refers to the fluorescence intensity before the addition of the sample solution to be tested, and F refers to the fluorescence intensity after the addition of the sample solution to be tested), it is determined that the sample solution to be tested contains paraquat; otherwise, the sample solution to be tested does not contain paraquat.

[0009] Among them, the optimal excitation wavelength of the fluorescence spectrum is 373nm.

[0010] Among them, the static reaction time is 3±0.5min.

[0011] The GSH-Zn-Ag-In-S quantum dots refer to quaternary Zn-Ag-In-S quantum dots capped with glutathione, which are synthesized by reacting reduced glutathione as a reducing agent and stabilizer with zinc acetate, silver nitrate, indium acetate and sodium sulfide. The molar ratio of zinc acetate, silver nitrate, indium acetate, reduced glutathione and sodium sulfide is 1.2:0.71:2:60:0.26, and the concentration of reduced glutathione in the reaction system is 0.055-0.060M.

[0012] The Tris-HCl buffer has a concentration of 0.2 M and a pH of 7.6.

[0013] The preparation method of the GSH-Zn-Ag-In-S quantum dot stock solution comprises the following steps:

[0014] (1) dissolving zinc acetate, silver nitrate, indium acetate and reduced glutathione in ultrapure water, mixing them evenly, adjusting the pH of the mixed solution to 8.5±0.1, and then adding sodium sulfide solution to the mixed solution to obtain a reaction system;

[0015] (2) The reaction system was heated to 100±5°C, refluxed in the dark for 4±0.5 h, cooled and centrifuged, filtered with a 0.2 μm filter membrane and dialyzed in the dark for 24-48 h to obtain a GSH-Zn-Ag-In-S quantum dot stock solution.

[0016] In step (1), the pH of the mixed solution is adjusted with a sodium hydroxide solution, wherein the concentration of the sodium hydroxide solution is 0.8-8 mol / L.

[0017] In step (2), the average particle size of the GSH-Zn-Ag-In-S quantum dots in the GSH-Zn-Ag-In-S quantum dot stock solution is 0.89±0.11 nm.

[0018] After the GSH-Zn-Ag-In-S quantum dot stock solution is diluted a certain multiple with ultrapure water (preferably, the GSH-Zn-Ag-In-S quantum dot stock solution is diluted 35-45 times), the diluted GSH-Zn-Ag-In-S quantum dot stock solution, Tris-HCl buffer and the sample solution to be tested are mixed, preferably, at a volume ratio of 1.5-2:1.5-3:3.

[0019] The detection limit of the nano fluorescent quantum dot colorimetric sensor for paraquat is 0.056 μg / L.

[0020] Furthermore, a method for detecting paraquat using a nano fluorescent quantum dot colorimetric sensor comprises the following steps:

[0021] S1. Draw a standard curve: accurately weigh the paraquat dichloride standard, prepare a paraquat standard stock solution with ultrapure water, and then dilute it with ultrapure water to different concentrations to obtain a paraquat linear solution. Evenly mix the dilution of the GSH-Zn-Ag-In-S quantum dot stock solution, Tris-HCl buffer and the paraquat linear solution, let it stand for reaction, measure the fluorescence emission spectrum of the detection system, and draw a standard curve with the concentration of paraquat as the horizontal coordinate and the fluorescence intensity value of 540±5nm as the vertical coordinate;

[0022] S2 pre-treats the sample to be tested to obtain a sample solution to be tested, mixes the diluted solution of the GSH-Zn-Ag-In-S quantum dot stock solution, Tris-HCl buffer and the sample solution to be tested, allows the reaction to proceed, measures the fluorescence intensity value of the detection system at 540±5nm, and calculates the paraquat concentration in the sample solution to be tested using the standard curve in S1.

[0023] The paraquat concentration in the sample solution to be tested is 0.001 mg / L to 0.01 mg / L.

[0024] The samples to be tested include food and agricultural products.

[0025] The sample to be tested is pre-treated to obtain the sample solution to be tested. For solid samples, it is first crushed, then added with pure water to squeeze juice or soak, filtered, and the filtrate is diluted with pure water until it is colorless; for liquid samples, it is shaken and filtered, and the filtrate is diluted with pure water until it is colorless.

[0026] Preferably, in steps S1-S2, the dilution of the GSH-Zn-Ag-In-S quantum dot stock solution is a 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution.

[0027] Preferably, in steps S1-S2, the volume ratio of the dilution solution of the GSH-Zn-Ag-In-S quantum dot stock solution, the Tris-HCl buffer and the paraquat linear solution / the sample solution to be tested is 2:3:3.

[0028] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0029] 1. The detection method of the present invention has high sensitivity, short detection time, simple operation, does not involve expensive instruments and reagents, and has low cost;

[0030] 2. The detection method of the present invention can be used to directly determine whether the paraquat pesticide residue exceeds the standard with the naked eye. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is a schematic diagram of the detection principle of the nano fluorescent quantum dot colorimetric sensor of the present invention;

[0032] Figure 2 The fluorescence spectra of the GSH-Zn-Ag-In-S quantum dots in the nano fluorescent quantum dot colorimetric sensor of the present invention before and after the reaction with paraquat (red line: GSH-ZAIS-QDs+ultrapure water; black line: GSH-ZAIS-QDs+PQCl2), the abscissa is the wavelength, and the ordinate is the fluorescence intensity;

[0033] Figure 3 TEM images and particle size distribution diagrams of GSH-Zn-Ag-In-S quantum dots before and after the reaction with paraquat, where (AB) is before the reaction, and (CD) is after the reaction;

[0034] Figure 4 This is the infrared spectrum of GSH-Zn-Ag-In-S quantum dots before and after the reaction with paraquat;

[0035] Figure 5 This is the pH optimization diagram for GSH-Zn-Ag-In-S quantum dots to detect paraquat, the horizontal axis is pH, and the vertical axis is the ratio of the fluorescence intensity change before and after the reaction to the fluorescence intensity value before the reaction, that is, (F0-F) / F0;

[0036] Figure 6 Fluorescence visualization comparison chart of optimized dilution factor for GSH-Zn-Ag-In-S quantum dots to detect paraquat;

[0037] Figure 7 This is the optimization diagram of the volume ratio of each solution for detecting paraquat by the nano fluorescent quantum dot colorimetric sensor. The horizontal axis is the reaction ratio, and the vertical axis is the ratio of the change in fluorescence intensity before and after the reaction to the fluorescence intensity value before the reaction, that is, (F0-F) / F0;

[0038] Figure 8 The specificity of the nano fluorescent quantum dot colorimetric sensor in detecting paraquat, the horizontal axis is the type of pesticide, and the vertical axis is the ratio of fluorescence intensity before and after the reaction, that is, F0 / F;

[0039] Fig. 9 The anti-interference performance of the nano fluorescent quantum dot colorimetric sensor in detecting paraquat, the horizontal axis is the type of interfering pesticides, and the vertical axis is the ratio of fluorescence intensity before and after the reaction, that is, F0 / F;

[0040] Fig.10 A visualization diagram of the colorimetric effect diagram after the reaction between the nano fluorescent quantum dot colorimetric sensor and paraquat after color extraction and refilling;

[0041] Fig.11After constructing the sensor of the present invention, a paraquat standard solution of known concentration is added to construct a fluorescence spectrum diagram (the horizontal axis is wavelength, the vertical axis is fluorescence intensity) and a linear regression diagram (the horizontal axis is concentration, the vertical axis is fluorescence intensity) of the standard curve.

[0042] In the present invention, GSH-Zn-Ag-In-S quantum dots, GSH-ZnAgInS QDs, GSH-ZAIS-QDs and ZAIS-QDs all have the same meaning. DETAILED DESCRIPTION

[0043] The applicant will further describe the present invention in conjunction with the embodiments and drawings of the specification below, but the protection scope of the present invention is not limited to these embodiments. Those skilled in the art should understand that equivalent replacements or corresponding improvements made to the technical features of the present invention still fall within the protection scope of the present invention.

[0044] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods; the materials and reagents used, unless otherwise specified, can be obtained from commercial sources. Among them, the standard product of paraquat dichloride was purchased from Sinopharm Group with a purity of 99.8% and a specification of 25 mg.

[0045] Example 1: Preparation of quantum dots

[0046] A method for preparing a colorimetric sensor based on glutathione-capped quaternary high-luminescence Zn-Ag-In-S quantum dots comprises the following steps:

[0047] 0.0022g (12μmol) of zinc acetate was added to 1mL of ultrapure water, 0.0012g (7.1μmol) of silver nitrate was added to 0.2mL of ultrapure water, 0.0058g (20μmol) of indium acetate was added to 2mL of ultrapure water, and 0.1843g (600μmol) of reduced glutathione was added to 5mL of ultrapure water. The above four solutions were mixed under stirring, and the pH of the above mixed solution was adjusted to 8.5 (about 0.1mL) with a sodium hydroxide solution having a concentration of 8mol / L. Then, 2mL of 1.3mmol / L sodium sulfide solution was added to the mixed solution to obtain a yellow clear solution. The mixed solution was heated to 100°C, refluxed in the dark for 4 hours, cooled, centrifuged at 10,000 r / min, filtered with a 0.2 μm filter membrane, and dialyzed in the dark for 24 hours to obtain a GSH-Zn-Ag-In-S quantum dot stock solution, which emitted a yellowish-brown fluorescence (Ex=373 nm).

[0048] The prepared GSH-Zn-Ag-In-S quantum dot stock solution was scanned by a transmission electron microscope, and the obtained TEM image was as follows: Figure 3As shown in A and B. From the TEM image, it can be seen that the quantum dots are uniform in size and are regular spherical, with an average particle size of 0.89±0.11nm.

[0049] Example 2: Analysis method establishment

[0050] Accurately weigh the paraquat dichloride standard and prepare a 10 mg / L paraquat standard stock solution with ultrapure water. Then dilute the paraquat standard stock solution with ultrapure water to obtain paraquat standard solutions of different concentrations.

[0051] A 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution prepared in Example 1, Tris-HCl buffer (0.2M, pH 7.6), and 1 mg / L paraquat standard solution were mixed at a volume ratio of 1:1:1. In the blank group, paraquat was replaced with an equal volume of ultrapure water. The fluorescence spectra of the two mixed solutions were collected using a fluorescence spectrophotometer. The results are shown in FIG. Figure 2 As shown. Figure 2 It can be seen that GSH-Zn-Ag-In-S quantum dots have two obvious fluorescence characteristic peaks in the buffer solution. After adding paraquat, the characteristic peak of GSH-Zn-Ag-In-S quantum dots at 540nm disappears.

[0052] The GSH-Zn-Ag-In-S quantum dot stock solution of Example 1, Tris-HCl buffer (0.2M, pH 7.6), and 1 mg / L paraquat standard solution were mixed at a volume ratio of 1:1:1, and a microscopic image of the mixed solution was taken using a transmission electron microscope. The results are as follows: Figure 3 As shown in C and D. Figure 3 It can be seen from C and D that the particle size of GSH-Zn-Ag-In-S quantum dots increased significantly after the reaction with paraquat, and the average particle size was 3.69±0.23nm.

[0053] Figure 4 The infrared spectra of GSH-Zn-Ag-In-S quantum dots before and after the reaction with paraquat. -1 、1680cm -1 、1590cm -1 、1400cm -1 、1300cm -1 and 1000cm -1 The characteristic peaks at 2990cm-1 in the FT-IR spectrum of paraquat correspond to the carboxyl group, amide bond, and thiol group in the chemical structure of GSH, respectively. -1、1640cm -1 、1560cm -1 、1440cm -1 、1180cm -1 The characteristic peaks are methyl CH stretching vibration, conjugated C=C stretching vibration, pyridine ring plane deformation vibration, 4,4-bipyridine skeleton CH stretching vibration, CN stretching vibration, which are consistent with the nitrogen methyl bipyridine structure of paraquat; and the FT-IR spectrum after the reaction of the two is 500-600cm -1 and 2880cm -1 New SS stretching vibration and methylene CH asymmetric stretching vibration characteristic peaks appeared at 1590 cm -1 The characteristic peak signal of CN stretching vibration at 1640cm -1 The characteristic peak signal of conjugated C=C stretching vibration weakened. It is inferred that a special chemical reaction occurred between GSH-Zn-Ag-In-S quantum dots and paraquat.

[0054] pH optimization of GSH-Zn-Ag-In-S quantum dots for detection of paraquat:

[0055] The following concentrations of paraquat standard solutions were taken respectively: 1, 0.5, 0.01, 0 mg / L (replaced with ultrapure water), and then GSH-Zn-Ag-In-S quantum dot stock solution, Tris-HCl buffer (0.2M, pH 7.0-8.0) of different pH values, and paraquat standard solutions of different concentrations were added in the order of 1:1:1 volume ratio, mixed evenly, reacted for 3 minutes, and the colorimetric effect diagram was taken by a camera under a 365nm ultraviolet lamp. The results are shown in FIG. Figure 5 As shown in A, a to f in the figure correspond to the colorimetric effects at pH 7.0, pH 7.2, pH 7.4, pH 7.6, pH 7.8, and pH 8.0, respectively; 4 samples in a to f: upper left: blank control, upper right: 0.01 mg / L, lower left: 0.5 mg / L, lower right: 1 mg / L; with pH as the abscissa, the ratio of the 540 nm fluorescence intensity change value before and after the reaction of GSH-Zn-Ag-In-S quantum dots with 0.5 mg / L paraquat standard solution to the fluorescence intensity value before the reaction, i.e. (F0-F) / F0, is plotted as the ordinate, see Figure 5 In B, it can be seen that when the pH of the buffer solution is 7.6, the change in (F0-F) / F0 before and after the reaction is the largest. Therefore, pH=7.6 was selected as the subsequent experimental condition.

[0056] Optimization of dilution factor for detection of paraquat by GSH-Zn-Ag-In-S quantum dots:

[0057] Take 1 mL of the GSH-Zn-Ag-In-S quantum dot stock solution prepared in Example 1, add 9 mL of ultrapure water and mix evenly to obtain a 10-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution; similarly, prepare a 20-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution and a 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution for later use. The following concentrations of paraquat standard solutions were taken respectively: 1, 0.8, 0.5, 0.1, 0.08, 0.05, 0.03, 0.01, 0 mg / L (replaced with ultrapure water), and then the dilutions of GSH-Zn-Ag-In-S quantum dot stock solutions of different dilution multiples, Tris-HCl buffer (0.2M, pH 7.6), and paraquat standard solutions of different concentrations were added in the order of 1:1:1 volume ratio, mixed evenly, reacted for 3 minutes, and the colorimetric effect diagram was taken by a camera under a 365nm ultraviolet lamp. The results are shown in FIG. Figure 6 As shown, A to C are the colorimetric effects of 10-fold, 20-fold, and 40-fold dilutions of the GSH-Zn-Ag-In-S quantum dot stock solution, respectively. There are nine samples in each of A to C: all from top to bottom, the first row from left to right: 1, 0.8, 0.5 mg / L, the second row from left to right: 0.1, 0.08, 0.05 mg / L, the third row from left to right: 0.03, 0.01, 0 mg / L. As shown in the figure: when 10 and 20 times dilutions of the GSH-Zn-Ag-In-S quantum dot stock solution were added, the fluorescence intensity of GSH-ZAIS-QDs after reacting with paraquat was high, the fluorescence color difference between high concentration and low concentration was large, and there was almost no fluorescence color difference between high concentrations, making it difficult to distinguish; when the GSH-Zn-Ag-In-S quantum dot stock solution was diluted 40 times, the fluorescence intensity of GSH-ZAIS-QDs after reacting with paraquat was moderate, and a clear and obvious fluorescence color change from blue to purple to yellow was shown as the paraquat concentration changed from high to low. Therefore, a 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution was selected for subsequent experiments.

[0058] Optimization of the ratio of each solution for GSH-Zn-Ag-In-S quantum dots detection of paraquat:

[0059] The following concentrations of paraquat standard solutions were taken respectively: 1, 0.8, 0.5, 0.1, 0.08, 0.05, 0.03, 0.01, 0 mg / L (replaced with ultrapure water), and then added in the order of 40-fold dilution of GSH-Zn-Ag-In-S quantum dot stock solution (V1), Tris-HCl buffer (0.2 M, pH 7.6) (V2), and paraquat standard solutions of different concentrations (V3). The volume ratios of V1:V2:V3 were 1:1:1, 1:1:2, 2:3:3, 1:2:1, 3:2:1, 3:3:1, and 2:1:1, respectively, and mixed evenly. The mixture was reacted for 3 minutes, and the colorimetric effect diagram was taken under 365 nm ultraviolet light using a camera. The results are shown in FIG. Figure 7 As shown in A, a to g in the figure correspond to the colorimetric effects under 1:1:1, 1:1:2, 2:3:3, 1:2:1, 3:2:1, 3:3:1, and 2:1:1, respectively. The 9 samples in a to g are from top to bottom, the first row from left to right: 1, 0.8, 0.5 mg / L, the second row from left to right: 0.1, 0.08, 0.05 mg / L, and the third row from left to right: 0.03, 0.01, 0 mg / L; with the reaction ratio as the horizontal axis, the ratio of the 540nm fluorescence intensity change value before and after the reaction of GSH-Zn-Ag-In-S quantum dots with 0.5 mg / L paraquat standard solution to the fluorescence intensity value before the reaction, that is, (F0-F) / F0, is plotted as the vertical axis, see Figure 7 In Figure B, it can be seen that when the volume ratio of V1:V2:V3 is 1:1:2, the reaction system (F0-F) / F0 change effect is the best. However, when the fluorescence ratio is similar, when the volume ratio of V1:V2:V3 is 2:3:3, the fluorescence color change of different concentrations of paraquat standard solution and GSH-ZAIS-QDs response is more obvious. At this volume ratio, a colorimetric effect diagram was taken by a camera under a 365nm ultraviolet lamp for color extraction and refilling. The colorimetric effect diagram is shown in the figure. Fig.10 As shown ( Fig.10 The concentration order is Figure 7 The concentration sequence in the equation is one-to-one). Fig.10 It can be seen that the colorimetric sensing system shows color changes from blue to purple and then to yellow according to the concentration of the paraquat standard solution from high to low, which can be clearly observed by naked eyes. The method of colorimetric sensing for identifying sulfur-containing pesticides constructed by the present invention is satisfactory.

[0060] The reaction principle is as follows: Paraquat aqueous solution is mainly composed of paraquat dichloride, which exists in the form of pyridine methyl nitrogen cations, and GSH-Zn-Ag-In-S quantum dots are encapsulated by glutathione containing two carboxyl groups. In aqueous solution, the carboxyl groups dissociate into negatively charged hydrogen ions, so the two attract each other and facilitate the reaction. Further reference shows that the nitrogen pyridine structure is easily reduced, while the thiol group of glutathione has strong reducing properties. Through the analysis of experimental results such as infrared spectroscopy, after the two react, new methylene structures and disulfide bonds appear, which conforms to the laws of chemical reactions. After the reaction, TEM shows that the particle size of the quantum dots increases to 3.69±0.23nm. This is because glutathione is encapsulated outside the quantum dots. When the reduction reaction occurs, the thiol groups between the quantum dots form disulfide bonds, causing the quantum dots to aggregate and increase in particle size. In addition, after the quantum dots reacted with paraquat, the emission peak at 540nm was quenched, and the solution showed a color change from dark blue to blue-purple and then to khaki as the concentration of paraquat changed from high to low (1mg / L-0.01mg / L). Quantitative and qualitative detection of low-concentration paraquat can be achieved.

[0061] Example 3: Validation of analytical method

[0062] (1) Linearity, limit of quantification, and limit of detection

[0063] Prepare linear solutions of paraquat: use the paraquat standard stock solution in Example 2, and dilute the stock solution with ultrapure water to 0.01 mg / L, 0.007 mg / L, 0.006 mg / L, 0.005 mg / L, 0.004 mg / L, 0.003 mg / L, 0.002 mg / L, and 0.001 mg / L, respectively.

[0064] A 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution prepared in Example 1, Tris-HCl buffer (0.2M, pH 7.6), and a linear solution of 0.01 mg / L to 0.001 mg / L paraquat were added to the EP tube in a volume ratio of 2:3:3; a 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution prepared in Example 1, Tris-HCl buffer (0.2M, pH 7.6), and ultrapure water were added to the EP tube in a volume ratio of 2:3:3 as a blank control. Mix well, react for 3 minutes, and collect the fluorescence spectrum of the reaction system. Fig.11 In A, record the fluorescence intensity at 540nm; draw a standard curve with the concentration of paraquat linear solution as the horizontal axis and the absorbance value at 540nm as the vertical axis, see Fig.11 B. The fluorescence intensity of the reaction system at 540nm shows a linear relationship with the paraquat concentration in the range of 10μg / L to 1μg / L. The standard curve equation of paraquat is as follows:

[0065] Y=-40.052193x+1199.45 (Formula 1)

[0066] Wherein x is the concentration of paraquat, and Y is the fluorescence intensity after the reaction of paraquat and GSH-Zn-Ag-In-S quantum dots.

[0067] The quantitative limit of the method was calculated according to the formula: LOQ = (10σ) / k, where σ is the standard deviation of the absorbance of the blank control measured 10 times, and k is the slope of the standard curve. The quantitative limit of paraquat was calculated to be 0.187 μg / L.

[0068] The detection limit of the method was calculated according to the formula: LOD = (3σ) / k, where σ is the standard deviation of the absorbance of the blank control measured 10 times, and k is the slope of the standard curve. The detection limit of paraquat was calculated to be 0.056 μg / L.

[0069] (2) Accuracy

[0070] About 5 g of green tea, apple, cabbage, and wolfberry samples were crushed, and 20 mL of ultrapure water was added to squeeze the juice (cabbage, apple) or soak (green tea, wolfberry, soak in boiling water for 5 minutes), filtered, and the filtrate was diluted 20 times with ultrapure water until it was colorless to obtain the food extract.

[0071] The paraquat standard stock solution in Example 2 was diluted with the above-mentioned food extract to three concentration levels of high (10 μg / L), medium (5 μg / L), and low (2 μg / L) to obtain a recovery determination solution; a reaction system was constructed according to the method in Example (1) (three groups of parallel reaction systems were set up), and the fluorescence intensity at 540 nm was measured; the fluorescence intensity value was substituted into the standard curve equation in (1) to obtain the test concentration of paraquat, and the recovery of paraquat in the actual sample was calculated according to the formula: spiked recovery = (test concentration / sample concentration) * 100%. The results are shown in Table 1 below.

[0072] Table 1

[0073]

[0074]

[0075] As shown in Table 1, the recoveries of paraquat at three concentration levels in apple, wolfberry, green tea and cabbage were all between 95% and 110%, and the RSDs were all less than 10%, which confirmed that the colorimetric sensor of the present invention had satisfactory detection effects in real samples.

[0076] (3) Specificity

[0077] The following pesticides were taken respectively, all with a concentration of 100 mg / L: triadimefon, glyphosate, chlorpyrifos, imidacloprid, acetamiprid, dipterex, cypermethrin, and cartap (all eight using ultrapure water as solvent); highly effective chlorflucythrinate, highly effective cypermethrin, quinalphos, dimethoate, and deltamethrin (all five using methanol as solvent); isoprocarb, procymidone, thiophanate-methyl, bispyribac, quinclorac, and bifenthrin (all six using ethanol as solvent), a total of 19 pesticide solutions, as well as 100 mg / L sodium chloride solution and 1 mg / L paraquat standard solution. Then, according to the order of adding the 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution prepared in Example 1, Tris-HCl buffer (0.2M, pH 7.6), and pesticides, the mixture was evenly mixed in a volume ratio of 2:3:3, and the fluorescence spectra of the above mixed solutions were collected by a fluorescence spectrophotometer. The ratio of the fluorescence intensity at 540 nm before and after the reaction of the GSH-Zn-Ag-In-S quantum dots with each pesticide, i.e., F0 / F, was used as the ordinate, and the type of pesticide was used as the abscissa to draw a bar graph, as shown in FIG. Figure 8 The results showed that only paraquat reacted with GSH-Zn-Ag-In-S quantum dots and the fluorescence intensity changed the most. Therefore, quantum dots have strong specificity in detecting paraquat.

[0078] (4) Anti-interference

[0079] In order to investigate the anti-interference performance of GSH-ZAIS-QDs in detecting PQCl2, a total of 20 pesticides including triadimefon, glyphosate, carbofuran, imidacloprid, acetamiprid, dipterex, cypermethrin, cartap, high-efficiency chlorflucythrinate, high-efficiency cypermethrin, quinalphos, dimethoate, deltamethrin, isoprocarb, procymidone, thiophanate-methyl, bispyribac, quinclorac, bifenthrin and sodium chloride were made into three interference mixed samples according to different solvents, and the pesticides in the same solvent were made into mixed samples with the concentration of each pesticide being 100 mg / L. Interference mixed sample group 1, i.e., the group using ultrapure water as the solvent, includes sodium chloride, triadimefon, glyphosate, chlorpyrifos, imidacloprid, acetamiprid, trichlorfon, cypermethrin, and cartap; interference mixed sample group 2, i.e., the group using methanol as the solvent, includes highly effective chlorflucythrinate, highly effective cypermethrin, quinalphos, dimethoate, and cypermethrin; interference mixed sample group 3, i.e., the group using ethanol as the solvent, includes isoprocarb, procymidone, methyl thiophanate, bispyribac, quinclorac, and cypermethrin.

[0080] Then, according to the order of adding 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution prepared in Example 1, Tris-HCl buffer (0.2M, pH 7.6), and interference mixed sample (or interference mixed sample containing 1 mg / L paraquat, or ultrapure water), after being evenly mixed at a volume ratio of 2:3:3, a fluorescence spectrophotometer was used to collect the fluorescence spectra of the GSH-Zn-Ag-In-S quantum dots before and after the reaction with each interference mixed sample, and the 540nm fluorescence intensity ratio of the GSH-Zn-Ag-In-S quantum dots before and after the reaction with the interference mixed sample, i.e., F0 / F, was used as the ordinate, and the type of interference mixed sample was used as the abscissa to plot the graph, as shown in FIG. Fig. 9 The results are as follows Fig. 9 As shown, in the complex system of mixed pesticides, the mixed sample containing paraquat standard solution still has the highest reactivity, which confirms that GSH-ZAIS-QDs has strong anti-interference performance in detecting paraquat.

Claims

1. A method for detecting paraquat using a nano fluorescent quantum dot colorimetric sensor, characterized in that: The GSH-Zn-Ag-In-S quantum dots in the nano-fluorescent quantum dot colorimetric sensor undergo a specific redox reaction with paraquat and produce an obvious aggregation phenomenon, which changes the fluorescence color and the fluorescence intensity at 540±5nm, thereby realizing the detection of paraquat pesticide in the sample to be tested.

2. The method according to claim 1, characterized in that The nano fluorescent quantum dot colorimetric sensor comprises a GSH-Zn-Ag-In-S quantum dot stock solution and a Tris-HCl buffer solution. The nano fluorescent quantum dot colorimetric sensor is mixed with a sample solution to be tested, and the mixture is allowed to stand for reaction. A fluorescent visualization image of paraquat is obtained by collecting the obtained detection system based on GSH-Zn-Ag-In-S quantum dots for identifying paraquat. The color difference of the fluorescent color response of paraquat at different concentrations is utilized. According to the fluorescent color of the detection system obtained after the reaction of the sample solution to be tested, the paraquat content in the sample solution to be tested is detected and whether the paraquat pesticide residue exceeds the standard is judged; and / or The fluorescence emission spectrum of the obtained detection system is measured. If the ratio of the fluorescence intensity at 540±5 nm before and after adding the test sample solution is F0 / F>2, it is determined that the test sample solution contains paraquat; otherwise, the test sample solution does not contain paraquat.

3. The method according to claim 2, characterized in that The GSH-Zn-Ag-In-S quantum dots are synthesized by reacting reduced glutathione as a reducing agent and stabilizer with zinc acetate, silver nitrate, indium acetate and sodium sulfide; preferably, the molar ratio of zinc acetate, silver nitrate, indium acetate, reduced glutathione and sodium sulfide is 1.2:0.71:2:60:0.26, and the concentration of reduced glutathione in the reaction system is 0.055-0.060M.

4. The method according to claim 2, characterized in that: The Tris-HCl buffer has a concentration of 0.2 M and a pH of 7.

6.

5. The method according to claim 3, characterized in that: The preparation method of the GSH-Zn-Ag-In-S quantum dot stock solution comprises the following steps: (1) dissolving zinc acetate, silver nitrate, indium acetate and reduced glutathione in ultrapure water, mixing them evenly, adjusting the pH of the mixed solution to 8.5±0.1, and then adding sodium sulfide solution to the mixed solution to obtain a reaction system; (2) The reaction system was heated to 100±5°C, refluxed in the dark for 4±0.5 h, cooled and centrifuged, filtered with a 0.2 μm filter membrane and dialyzed in the dark for 24-48 h to obtain a GSH-Zn-Ag-In-S quantum dot stock solution.

6. The method according to any one of claims 2 to 5, characterized in that: After the GSH-Zn-Ag-In-S quantum dot stock solution is diluted a certain number of times with ultrapure water, the dilution of the GSH-Zn-Ag-In-S quantum dot stock solution, Tris-HCl buffer and the sample solution to be tested are mixed; preferably, the GSH-Zn-Ag-In-S quantum dot stock solution is diluted 35-45 times; preferably, the dilution of the GSH-Zn-Ag-In-S quantum dot stock solution, Tris-HCl buffer and the sample solution to be tested are mixed at a volume ratio of 1.5-2:1.5-3:3; preferably, the static reaction time is 3±0.5min.

7. The method according to claim 2, characterized in that The steps include: S1. Draw a standard curve: accurately weigh the paraquat dichloride standard, prepare a paraquat standard stock solution with ultrapure water, and then dilute it with ultrapure water to different concentrations to obtain a paraquat linear solution. Mix the diluted solution of the GSH-Zn-Ag-In-S quantum dot stock solution, Tris-HCl buffer and the paraquat linear solution, let it stand for reaction, measure the fluorescence emission spectrum of the detection system, and draw a standard curve with the concentration of paraquat as the horizontal coordinate and the fluorescence intensity value of 540±5nm as the vertical coordinate; S2 pre-treats the sample to be tested to obtain a sample solution to be tested, mixes the diluted solution of the GSH-Zn-Ag-In-S quantum dot stock solution, Tris-HCl buffer and the sample solution to be tested, allows the reaction to proceed, measures the fluorescence intensity value of the detection system at 540±5nm, and calculates the paraquat concentration in the sample solution to be tested using the standard curve in S1.

8. The method according to claim 7, characterized in that The paraquat concentration in the sample solution to be tested is 0.001 mg / L to 0.01 mg / L.

9. The method according to claim 7, characterized in that: In steps S1-S2, the dilution of the GSH-Zn-Ag-In-S quantum dot stock solution is a 40-fold dilution of the GSH-Zn-Ag-In-S quantum dot stock solution; and / or The volume ratio of the diluent of the GSH-Zn-Ag-In-S quantum dot stock solution, the Tris-HCl buffer and the paraquat linear solution / the sample solution to be tested is 2:3:

3.

10. The method according to claim 7, characterized in that In step S2, the sample to be tested includes food and agricultural products; for solid samples, they are first crushed, then added with pure water to squeeze juice or soak, filtered, and the filtrate is diluted with pure water until it is colorless; For liquid samples, shake well and filter, then dilute the filtrate with pure water until it is colorless.

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