AChE / AgNPs@PVA / RGO / SPE sensor and its preparation and application

Abstract

The present invention relates to the technical field of methyl parathion detection, and in particular to an AChE / AgNPs@PVA / RGO / SPE sensor and its preparation and application. Based on the inhibitory effect of methyl parathion on acetylcholinesterase, the present invention prepares a sensor for methyl parathion detection. The sensor has good electrocatalytic ability for the enzymatic oxidation of thiocholine, and exhibits excellent adsorption capacity and high SERS activity for methyl parathion; the inhibitory effect of methyl parathion on AChE is proportional to its concentration. The sensor is used for electrochemical detection and Raman spectroscopy detection of methyl parathion, and a linear relationship between the methyl parathion concentration and the current signal intensity and the Raman signal intensity is obtained. Compared with existing on-site rapid detection methods, the present invention has good stability, more SERS active hotspots, high sensitivity and is easy to carry, providing a new method for the detection of organophosphorus pesticides.

Description

Technical Field

[0001] The present invention relates to the technical field of methyl parathion detection, and in particular to an AChE / AgNPs@PVA / RGO / SPE sensor and preparation and application thereof. Background Art

[0002] Organophosphates (OPs) are widely used as pesticides due to their high toxicological profile. Unfortunately, this toxicity is not limited to the unique species to be eliminated. They are particularly toxic to humans and animals due to their high toxicity towards the enzyme acetylcholinesterase (AChE). In fact, inhibition of AChE activity by pesticides may lead to memory loss and impaired neuromuscular function, ultimately leading to death. AChE is an important enzyme in the brain. It hydrolyzes the neurotransmitter acetylcholine (ATC) into choline and acetic acid. Therefore, AChE activity can serve as a biomarker for exposure to this pesticide.

[0003] There are already technologies for detecting AChE inhibitors, including gas or liquid chromatography and mass spectrometry. However, the reported technologies involve delicate and complex sample pretreatment processes. In addition, most pollutants are specific to each pollutant and usually require a long analysis time. Various biosensing technologies based on AChE activity and different transduction technologies (electrochemical, fluorescent, colorimetric probes, etc.) have also been designed to provide rapid, simple and selective technologies for toxicity monitoring in environmental, agricultural, food or military applications. Before this pioneering work, many various detection methods based on AChE activity have been developed. However, these technologies have some disadvantages. Due to the single detection method, the detection process lacks reliability and accuracy of the detection results. At the same time, the preparation is complex, time-consuming, and the stability is poor, which limits their application. Summary of the Invention

[0004] In order to solve the above problems, the purpose of the present invention is to provide an AChE / AgNPs@PVA / RGO / SPE sensor and its preparation and application. Based on the inhibitory effect of methyl parathion on acetylcholinesterase, the present invention prepares a sensor for the detection of methyl parathion. The sensor has good electrocatalytic ability for the enzymatic oxidation of thiocholine, and exhibits excellent adsorption capacity and high surface enhanced Raman scattering (SERS) activity for methyl parathion; the inhibitory effect of methyl parathion on AChE is proportional to its concentration. The sensor is used for electrochemical detection and Raman spectroscopy detection of methyl parathion, and a linear relationship between the methyl parathion concentration and the current signal intensity and Raman signal intensity is obtained. Compared with existing on-site rapid detection methods, the present invention has good stability, more SERS active hotspots, high sensitivity and easy portability, providing a new method for the detection of organophosphorus pesticides.

[0005] In the present invention, the AChE / AgNPs@PVA / RGO / SPE sensor is also an acetylcholinesterase / silver particles@polyvinyl alcohol microspheres / graphene modified screen-printed electrode sensor.

[0006] The purpose of the present invention can be achieved by the following technical solutions:

[0007] The first object of the present invention is to provide a method for preparing an AChE / AgNPs@PVA / RGO / SPE sensor, comprising the following steps:

[0008] (1) Preparation of RGO / SPE electrode: The SPE electrode was placed in GO electrodeposition solution and reduced by cyclic voltammetry to obtain the RGO / SPE electrode;

[0009] (2) Preparation of AgNPs@PVA composites: PVA microgel spheres were prepared by microfluidics, and then silver nitrate was in situ reduced to AgNPs colloidal solution on the PVA microgel spheres by sodium citrate, and then post-treated to obtain AgNPs@PVA composites;

[0010] (3) Preparation of AgNPs@PVA / RGO / SPE composite working electrode: The AgNPs@PVA composite material prepared in step (2) was added dropwise to the surface of the RGO / SPE electrode prepared in step (1), and dried to obtain an AgNPs@PVA / RGO / SPE composite working electrode;

[0011] (4) Preparation of AChE / AgNPs@PVA / RGO / SPE sensor: AChE solution was added dropwise to the surface of the AgNPs@PVA / RGO / SPE composite working electrode prepared in step (3), and the mixture was dried to obtain an AChE / AgNPs@PVA / RGO / SPE sensor.

[0012] In one embodiment of the present invention, the GO electrodeposition solution is specifically prepared as follows: graphene oxide powder is mixed with a phosphate buffer solution and then incubated to activate the carboxyl groups on the graphene oxide, and then dispersed with Tris-HCl after centrifugation to obtain a GO electrodeposition solution.

[0013] In one embodiment of the present invention, in step (1), during the cyclic voltammetry, the potential range is -1.4 to +0.6 V, and the scan rate is 50 mV / s.

[0014] In one embodiment of the present invention, in step (2), the specific preparation method of PVA microgel spheres is as follows:

[0015] (2.1) PVA was mixed with potassium persulfate solution and sodium 2-acrylamide-2-methylpropanesulfonate to prepare an aqueous phase;

[0016] (2.2) Dichloromethane and n-butyl acetate were mixed to obtain a continuous oil phase containing TEMED;

[0017] (2.3) In a microfluidic system, a syringe pump is used to inject the aqueous phase obtained in step (1) and the oil phase obtained in step (2) into the vertical channel at a volume flow rate to form a PVA microgel.

[0018] In one embodiment of the present invention, in step (2), the usage ratio of PVA microgel beads, silver nitrate and sodium citrate is 0.1 g: 2 μmol: 200 μmol.

[0019] In one embodiment of the present invention, in step (2), the post-treatment is washing with deionized water and centrifugation.

[0020] In one embodiment of the present invention, in step (3), during the drying process, the drying temperature is 15-40° C. and the drying time is 2-4 hours.

[0021] In one embodiment of the present invention, in step (4), during the drying process, the drying temperature is 4° C. and the drying time is 2-4 h.

[0022] The second object of the present invention is to provide an AChE / AgNPs@PVA / RGO / SPE sensor prepared by the above method.

[0023] The third object of the present invention is to provide a method for electrochemically detecting methyl parathion in pesticides using an AChE / AgNPs@PVA / RGO / SPE sensor, comprising the following steps:

[0024] (1) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in an alkaline solution containing acetylthiocholine chloride (ATCl), and the initial response current signal intensity was measured by cyclic voltammetry;

[0025] (2) Prepare different concentrations of methyl parathion standard solutions as the experimental group and alkaline solution containing ATCl as the control group. Immerse the AChE / AgNPs@PVA / RGO / SPE sensor in the control group and the experimental group, and measure the response current signal intensity by cyclic voltammetry to obtain the response current signal intensity change value.

[0026] (3) Draw a standard curve with the concentration of methyl parathion standard solution as the horizontal axis and the change value of the response current signal intensity as the vertical axis;

[0027] (4) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in the pesticide, and the response current signal intensity was measured by cyclic voltammetry to obtain the response current signal intensity change value, which was substituted into the standard curve obtained in step (3) to obtain the concentration of methyl parathion in the pesticide.

[0028] A fourth object of the present invention is to provide a method for detecting methyl parathion in pesticides using an AChE / AgNPs@PVA / RGO / SPE sensor using a portable Raman spectrometer, comprising the following steps:

[0029] (1) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in an alkaline solution containing ATCl. After the reaction was protected from light, a portable Raman spectrometer was used to measure the initial Raman spectrum peak at 639 cm -1 Spectral signal;

[0030] (2) Prepare different concentrations of methyl parathion standard solutions as the experimental group and alkaline solution containing ATCl as the control group. Immerse the AChE / AgNPs@PVA / RGO / SPE sensor in the control group and the experimental group. After the reaction in the dark, a portable Raman spectrometer is used to measure the Raman spectrum peak 639 cm -1 Spectral signal to obtain spectral signal change value;

[0031] (3) Draw a standard curve with the concentration of methyl parathion standard solution as the horizontal axis and the spectral signal change value as the vertical axis;

[0032] (4) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in pesticides, and the Raman spectrum peak at 639 cm was measured using a portable Raman spectrometer. -1 The spectral signal is used to obtain the spectral signal change value, which is substituted into the standard curve obtained in step (3) to obtain the concentration of methyl parathion in the pesticide.

[0033] Compared with the prior art, the present invention has the following beneficial effects:

[0034] (1) The present invention successfully constructed an AChE / AgNPs@PVA / RGO / SPE sensor for the detection of methyl parathion based on AgNPs@PVA / RGO nanocomposite materials. The sensor combines electrochemical and SERS dual-mode strategies to perform reliable, cyclic, and highly sensitive detection of trace methyl parathion pesticides in water.

[0035] (2) The AgNPs@PVA in the AChE / AgNPs@PVA / RGO / SPE sensor has excellent conductivity, large surface area, and high viscosity, which can immobilize AChE on the AgNPs@PVA / RGO / SPE composite working electrode. At the same time, the AgNPs@PVA microgel has excellent SERS activity and can also absorb and enrich trace analytes in water by swelling. This makes the prepared AChE / AgNPs@PVA / RGO / SPE sensor have good stability, high electrochemical-SERS sensitivity, and easy portability.

[0036] (3) AgNPs@PVA / RGO in the AChE / AgNPs@PVA / RGO / SPE sensor improved the electron transfer rate, expanded the specific surface area, and had good electrocatalytic activity towards ATCl.

[0037] (4) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in a solution containing ATCl. Under the catalysis of AChE, ATCl was hydrolyzed to produce more thiocholine, resulting in an increase in the current signal intensity. When methyl parathion was added, methyl parathion inhibited the activity of AChE, reduced the catalytic effect of AChE on ATCl, thereby reducing the production of electroactive substances, and the current signal intensity decreased again.

[0038] (5) The AChE / AgNPs@PVA / RGO / SPE sensor can perform electrochemical determination and SERS trace detection of methyl parathion in water with good sensitivity, acceptable stability, and repeatability. It can also achieve on-site rapid quantitative detection and degradation of methyl parathion in water, with an electrochemical detection limit of 3 ng·L -1 , the SERS detection limit is 5 ng·L -1 The sensor is expected to become a new and effective method for on-site detection of high-performance organophosphorus pesticides. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 This is a physical picture and a partial enlarged picture of the AChE / AgNPs@PVA / RGO / SPE sensor prepared in Example 1 of the present invention;

[0040] Figure 2 is the electrochemical signal intensity graph of the sensor after incubation in acetylthiocholine chloride solution and acetylthiocholine chloride solution + methyl parathion solution;

[0041] Figure 3 This is a graph showing the change in electrochemical signal intensity after the sensor was incubated in a mixed solution of acetylthiocholine chloride solution and methyl parathion with different concentrations;

[0042] Figure 4 is the SERS spectra of the sensor after incubation in acetylthiocholine chloride solution and acetylthiocholine chloride solution + methyl parathion solution;

[0043] Figure 5 This is the SERS spectrum of the sensor after incubation in a mixed solution of acetylthiocholine chloride solution and methyl parathion with different concentrations. DETAILED DESCRIPTION

[0044] The present invention provides a method for preparing an AChE / AgNPs@PVA / RGO / SPE sensor, comprising the following steps:

[0045] (1) Preparation of RGO / SPE electrode: The SPE electrode was placed in GO electrodeposition solution and reduced by cyclic voltammetry to obtain the RGO / SPE electrode;

[0046] (2) Preparation of AgNPs@PVA composites: PVA microgel spheres were prepared by microfluidics, and then silver nitrate was in situ reduced to AgNPs colloidal solution on the PVA microgel spheres by sodium citrate, and then post-treated to obtain AgNPs@PVA composites;

[0047] (3) Preparation of AgNPs@PVA / RGO / SPE composite working electrode: The AgNPs@PVA composite material prepared in step (2) was added dropwise to the surface of the RGO / SPE electrode prepared in step (1), and dried to obtain an AgNPs@PVA / RGO / SPE composite working electrode;

[0048] (4) Preparation of AChE / AgNPs@PVA / RGO / SPE sensor: AChE solution was added dropwise to the surface of the AgNPs@PVA / RGO / SPE composite working electrode prepared in step (3), and the mixture was dried to obtain an AChE / AgNPs@PVA / RGO / SPE sensor.

[0049] In one embodiment of the present invention, the GO electrodeposition solution is specifically prepared as follows: graphene oxide powder is mixed with a phosphate buffer solution and then incubated to activate the carboxyl groups on the graphene oxide, and then dispersed with Tris-HCl after centrifugation to obtain a GO electrodeposition solution.

[0050] In one embodiment of the present invention, in step (1), during the cyclic voltammetry, the potential range is -1.4 to +0.6 V, and the scan rate is 50 mV / s.

[0051] In one embodiment of the present invention, in step (2), the specific preparation method of PVA microgel spheres is as follows:

[0052] (2.1) PVA was mixed with potassium persulfate solution and sodium 2-acrylamide-2-methylpropanesulfonate to prepare an aqueous phase;

[0053] (2.2) Dichloromethane and n-butyl acetate were mixed to obtain a continuous oil phase containing TEMED;

[0054] (2.3) In a microfluidic system, a syringe pump is used to inject the aqueous phase obtained in step (1) and the oil phase obtained in step (2) into the vertical channel at a volume flow rate to form a PVA microgel.

[0055] In one embodiment of the present invention, in step (2), the usage ratio of PVA microgel beads, silver nitrate and sodium citrate is 0.1 g: 2 μmol: 200 μmol.

[0056] In one embodiment of the present invention, in step (2), the post-treatment is washing with deionized water and centrifugation.

[0057] In one embodiment of the present invention, in step (3), during the drying process, the drying temperature is 15-40° C. and the drying time is 2-4 hours.

[0058] In one embodiment of the present invention, in step (4), during the drying process, the drying temperature is 4° C. and the drying time is 2-4 h.

[0059] The present invention provides an AChE / AgNPs@PVA / RGO / SPE sensor prepared by the above method.

[0060] The present invention provides a method for electrochemically detecting methyl parathion in pesticides using an AChE / AgNPs@PVA / RGO / SPE sensor, comprising the following steps:

[0061] (1) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in an alkaline solution containing ATCl, and the initial response current signal intensity was measured by cyclic voltammetry;

[0062] (2) Prepare different concentrations of methyl parathion standard solutions as the experimental group and alkaline solution containing ATCl as the control group. Immerse the AChE / AgNPs@PVA / RGO / SPE sensor in the control group and the experimental group, and measure the response current signal intensity by cyclic voltammetry to obtain the response current signal intensity change value.

[0063] (3) Draw a standard curve with the concentration of methyl parathion standard solution as the horizontal axis and the change value of the response current signal intensity as the vertical axis;

[0064] (4) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in the pesticide, and the response current signal intensity was measured by cyclic voltammetry to obtain the response current signal intensity change value, which was substituted into the standard curve obtained in step (3) to obtain the concentration of methyl parathion in the pesticide.

[0065] The present invention provides a method for detecting methyl parathion in pesticides using an AChE / AgNPs@PVA / RGO / SPE sensor and a portable Raman spectrometer, comprising the following steps:

[0066] (1) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in an alkaline solution containing ATCl. After the reaction was protected from light, a portable Raman spectrometer was used to measure the initial Raman spectrum peak at 639 cm -1 Spectral signal;

[0067] (2) Prepare different concentrations of methyl parathion standard solutions as the experimental group and alkaline solution containing ATCl as the control group. Immerse the AChE / AgNPs@PVA / RGO / SPE sensor in the control group and the experimental group. After the reaction in the dark, a portable Raman spectrometer is used to measure the Raman spectrum peak 639 cm -1 Spectral signal to obtain spectral signal change value;

[0068] (3) Draw a standard curve with the concentration of methyl parathion standard solution as the horizontal axis and the spectral signal change value as the vertical axis;

[0069] (4) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in pesticides, and the Raman spectrum peak at 639 cm was measured using a portable Raman spectrometer. -1 The spectral signal is used to obtain the spectral signal change value, which is substituted into the standard curve obtained in step (3) to obtain the concentration of methyl parathion in the pesticide.

[0070] The present invention will be described in detail below with reference to the accompanying drawings and specific embodiments.

[0071] In the following examples, unless otherwise specified, all reagents used are commercially available reagents; and all detection means and methods used are conventional detection means and methods in the art.

[0072] Example 1

[0073] This embodiment provides an AChE / AgNPs@PVA / RGO / SPE sensor and its preparation method.

[0074] (1) Preparation of RGO / SPE electrode: 500 μL of pH 7.4 phosphate buffer solution (containing 0.15 M EDC and 0.1 M NHS) was added to 5 mg of graphene oxide (GO) powder and incubated for 30 min to activate the carboxyl groups on GO. Then, Tris-HCl solution was added and the mixture was centrifuged three times (8000 rpm, 15 min). 5 mL of Tris-HCl was added to the resulting sediment and ultrasonically dispersed to form a uniform black suspension - GO electrodeposition solution.

[0075] The pretreated SPE electrode was immersed in GO electrodeposition solution and treated by cyclic voltammetry in the potential range of -1.4 to +0.6 V for 20 min (scan rate: 50 mV / s). At this time, GO was electrochemically reduced to RGO to obtain an RGO / SPE electrode; then it was rinsed with ultrapure water and dried with nitrogen, and the sensor was stored in a refrigerator at 4°C for use.

[0076] (2) Preparation of AgNPs@PVA composite material: 10 g of PVA was placed in a clean beaker, 0.16 g of potassium persulfate was taken, 3.5 g of deionized water was added to dissolve it, and then poured into the beaker containing PVA, 3 g of 2-acrylamide-2-methylpropanesulfonate sodium was added, and stirred for 20 min to prepare an aqueous phase; dichloromethane and n-butyl acetate were proportionally prepared to prepare a continuous oil phase containing TEMED; a syringe pump was used to inject the aqueous phase and oil phase into the cross-connected channel of the microfluidic chip according to the volume flow rate to form PVA microgel; the collected PVA microgel spheres were placed in a water bath for incubation, then washed with ethyl acetate / acetone, and stored in saline for later use;

[0077] 40 ml of glycerol and 60 ml of deionized water were refluxed under vigorous stirring until boiling, and then 0.1 g of PVA microgel and 2 ml of 0.005 M AgNO3 solution were added and stirred for 5 minutes. After the PVA microgel and the AgNO3 solution were fully in contact, 4 ml of 0.05 M sodium citrate solution was injected under vigorous stirring and refluxed for 60 minutes to obtain a yellow-green AgNPs colloidal solution. The obtained AgNPs colloidal solution was washed with deionized water and centrifuged to obtain a yellow-green AgNPs@PVA composite material, which was stored in a refrigerator at 4°C for later use.

[0078] (3) Preparation of AgNPs@PVA / RGO / SPE composite working electrode: 10 μL of AgNPs@PVA composite material was added to the surface of the GO / SPE electrode prepared in step (1) and dried naturally at room temperature for 2 h to obtain the AgNPs@PVA / RGO / SPE composite working electrode.

[0079] (4) Preparation of AChE / AgNPs@PVA / RGO / SPE sensor: 5 μL 0.02 U AChE was added to the surface of the AgNPs@PVA / RGO / SPE composite working electrode prepared in step (3), and the electrode was allowed to dry in a dry environment at 4 °C for 2 h to obtain an AChE / PVA@Ag / RGO / SPE sensor. Figure 1 shown.

[0080] Example 2

[0081] The prepared AChE / AgNPs@PVA / RGO / SPE sensor was first immersed in an alkaline solution (pH 7.5) containing 1.0MATCl and subjected to cyclic voltammetry (potential scanning range of 0.6 to 0.2 V; scanning rate of 50 mV / s; static time of 2 s) to obtain the initial response current signal intensity. The results are shown in Figure 2. Figure 2 shown.

[0082] Methyl parathion standard solutions of different concentrations (0.01, 0.1, 1, 10, and 100 μg·L-1) were prepared as experimental groups. The sensor was inserted into a mixture of a 1.0 M ATCl alkaline solution (control group) and a series of concentration gradient methyl parathion standard solutions. After reacting for 12 minutes, it was rinsed with PBS buffer (pH 7.5) for 1 minute. Finally, differential pulse voltammetry (DPV) measurements were performed again to obtain the change in the response current signal intensity. Figure 3 a, Shows the DPV effect of the AChE / AgNPs@PVA / RGO / SPE sensor in PBS solution containing 1.0 mM ATCl after incubation with different concentrations of methyl parathion under optimized conditions. Due to the inhibitory effect of methyl parathion on AChE, the peak current signal intensity decreases sharply with the increase of methyl parathion concentration. Figure 3 b is the relationship between the corresponding inhibition rate and the concentration of methyl parathion. The linear equation after analysis is y = 3.61x + 16.89, R 2 =0.99216, the detection limit of methyl parathion is 3 ng·L -1 .

[0083] For SERS testing, the prepared AChE / AgNPs@PVA / RGO / SPE sensor was first immersed in a test tube containing 1.0 MATCl alkaline solution (pH 7.5), and then the test tube was protected from light and placed in a constant temperature water bath for 10 minutes. Finally, the sensor was moved to measure the SERS spectrum. A 785 nm laser generator was used as the incident detection light source, with a laser power of 100% and an excitation time of 5 seconds. The results are shown in Figure 2. Figure 4 shown.

[0084] Methyl parathion standard solutions of different concentrations (0.01, 0.1, 1, 10, and 100 μg·L-1) were prepared as experimental groups. The sensor was inserted into a mixture of 1.0 M ATCl alkaline solution (control group) and a series of concentration gradient methyl parathion standard solutions. After a 12-minute reaction, the sensor was rinsed with PBS buffer (pH 7.5) for 1 minute. Finally, a portable Raman spectrometer was used to detect the spectral signal. The Raman peak at 639 cm -1 The intensity change is used as the characteristic peak to determine the presence of methyl parathion. Figure 5 a, Shows the Raman spectra of AChE / AgNPs@PVA / RGO / SPE after incubation with 1.0 mM ATCl and different concentrations of methyl parathion under optimized conditions. Due to the inhibitory effect of methyl parathion on AChE, the characteristic Raman peak signal intensity decreases sharply with the increase of methyl parathion concentration. Figure 5 b is the relationship between the corresponding Raman intensity change and the concentration of methyl parathion. The linear equation after analysis is y = -2.79x + 5.69, R 2 =0.97709, the detection limit of methyl parathion is 5 ng·L -1 .

[0085] Example 3

[0086] The AChE / AgNPs@PVA / RGO / SPE sensor prepared in Example 1 was used to detect methyl parathion in water.

[0087] (1) Electrochemical detection of methyl parathion;

[0088] The prepared AChE / AgNPs@PVA / RGO / SPE sensor was first immersed in an alkaline solution (pH 7.5) containing 1.0MATCl, and cyclic voltammetry was performed (potential scanning range of 0.6 to 0.2 V; scanning rate of 50 mV / s; static time of 2 s) to obtain the initial response current signal intensity.

[0089] The laboratory used sample water from the school's lake: 2 ml of lake water was mixed with 2 ml of 1.0 M ATCl. The sensor was then inserted into the mixture, reacted for 12 minutes, and then rinsed with PBS buffer (pH 7.5) for 1 minute. Finally, a DPV measurement was performed to obtain the response current signal intensity, which was compared with the standard curve of methyl parathion to detect methyl parathion in the lake water.

[0090] (2) SERS detection of methyl parathion;

[0091] The prepared AChE / AgNPs@PVA / RGO / SPE sensor was first immersed in an alkaline solution (pH 7.5) containing 1.0M ATCl, and then the test tube was protected from light and placed in a constant temperature water bath to react for 10 minutes. Finally, the sensor was moved to measure the SERS spectrum, using a 785nm laser generator as the incident detection light source, a laser power of 100%, and an excitation time of 5 seconds.

[0092] The laboratory used sample water from the school's lake: 2 ml of lake water was mixed with 2 ml of 1.0 M ATCl. The sensor was then inserted into the mixture, allowed to react for 12 minutes, and then rinsed with PBS buffer (pH 7.5) for 1 minute. Finally, the spectral signal was measured using a portable Raman spectrometer and compared with a standard curve for methyl parathion, enabling the detection of methyl parathion in the lake water.

[0093] Liquid chromatography-tandem mass spectrometry (LC-MS / MS) (GB / T 22975-2008) is a commonly used analytical method for methyl parathion. The lake water was also analyzed using LC-MS / MS, with the results shown in Table 1. As shown in Table 1, the AChE / AgNPs@PVA / RGO / SPE sensor prepared in Example 1 demonstrated good accuracy in detecting methyl parathion in water (electrochemical and SERS detection). The two methods can also be combined (electrochemical + SERS) to detect methyl parathion in water, demonstrating its potential as a rapid analytical method for methyl parathion in water.

[0094] Table 1 Comparison of detection results of electrochemical and SERS detection methods with HPLC

[0095]

[0096] The above description of the embodiments is intended to facilitate understanding and use of the invention by those skilled in the art. It will be apparent that those skilled in the art can readily make various modifications to these embodiments and apply the general principles described herein to other embodiments without requiring inventive effort. Therefore, the present invention is not limited to the above-described embodiments. Improvements and modifications made by those skilled in the art based on the disclosure of the present invention, without departing from the scope of the present invention, should be within the scope of protection of the present invention.

Claims

1. A method for preparing an AChE / AgNPs@PVA / RGO / SPE sensor, characterized in that: The steps include: (1) Preparation of RGO / SPE electrode: The SPE electrode was placed in GO electrodeposition solution and reduced by cyclic voltammetry to obtain the RGO / SPE electrode; (2) Preparation of AgNPs@PVA composite materials: PVA was mixed with potassium persulfate solution and sodium 2-acrylamide-2-methylpropanesulfonate to prepare an aqueous phase; dichloromethane and n-butyl acetate were mixed to obtain a continuous oil phase containing TEMED; a syringe pump was used in a microfluidic system to inject the aqueous phase and oil phase into the vertical channel at a volume flow rate to form a PVA microgel; Glycerol and deionized water were refluxed under stirring, and then PVA microgel and AgNO3 solution were added. After the PVA microgel and AgNO3 solution were fully in contact, sodium citrate solution was injected under stirring and refluxed to obtain a yellow-green AgNPs colloidal solution. The obtained AgNPs colloidal solution was washed with deionized water and centrifuged to obtain an AgNPs@PVA composite material. (3) Preparation of AgNPs@PVA / RGO / SPE composite working electrode: The AgNPs@PVA composite material prepared in step (2) was added dropwise to the surface of the RGO / SPE electrode prepared in step (1), and dried to obtain an AgNPs@PVA / RGO / SPE composite working electrode; (4) Preparation of AChE / AgNPs@PVA / RGO / SPE sensor: AChE solution was added dropwise to the surface of the AgNPs@PVA / RGO / SPE composite working electrode prepared in step (3), and the mixture was dried to obtain an AChE / AgNPs@PVA / RGO / SPE sensor.

2. The method for preparing an AChE / AgNPs@PVA / RGO / SPE sensor according to claim 1, wherein: In step (1), during the cyclic voltammetry, the potential range is -1.4 to +0.6 V, and the scan rate is 50 mV / s.

3. The method for preparing an AChE / AgNPs@PVA / RGO / SPE sensor according to claim 1, wherein: In step (2), the usage ratio of PVA microgel beads, silver nitrate and sodium citrate is 0.1 g: 2 μmol: 200 μmol.

4. The method for preparing an AChE / AgNPs@PVA / RGO / SPE sensor according to claim 1, wherein: In step (2), the post-treatment is washing with deionized water and centrifugation.

5. The method for preparing an AChE / AgNPs@PVA / RGO / SPE sensor according to claim 1, wherein: In step (3), during the drying process, the drying temperature is 15-40° C. and the drying time is 2-4 hours.

6. The method for preparing an AChE / AgNPs@PVA / RGO / SPE sensor according to claim 1, wherein: In step (4), during the drying process, the drying temperature is 4° C. and the drying time is 2-4 h.

7. An AChE / AgNPs@PVA / RGO / SPE sensor prepared by the method according to any one of claims 1-6.

8. A method for electrochemically detecting methyl parathion in pesticides using the AChE / AgNPs@PVA / RGO / SPE sensor according to claim 7, characterized in that: The following steps are involved: (1) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in an alkaline solution containing ATCl, and the initial response current signal intensity was measured by cyclic voltammetry; (2) Prepare different concentrations of methyl parathion standard solutions as the experimental group and alkaline solution containing ATCl as the control group. Immerse the AChE / AgNPs@PVA / RGO / SPE sensor in the control group and the experimental group, and measure the response current signal intensity by cyclic voltammetry to obtain the response current signal intensity change value. (3) Draw a standard curve with the concentration of methyl parathion standard solution as the horizontal axis and the change value of the response current signal intensity as the vertical axis; (4) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in the pesticide, and the response current signal intensity was measured by cyclic voltammetry to obtain the response current signal intensity change value, which was substituted into the standard curve obtained in step (3) to obtain the concentration of methyl parathion in the pesticide.

9. A method for detecting methyl parathion in pesticides using the AChE / AgNPs@PVA / RGO / SPE sensor according to claim 7 using a portable Raman spectrometer, characterized in that: The following steps are involved: (1) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in an alkaline solution containing ATCl. After the reaction was protected from light, a portable Raman spectrometer was used to measure the initial Raman spectrum peak at 639 cm -1 Spectral signal; (2) Prepare different concentrations of methyl parathion standard solutions as the experimental group and alkaline solution containing ATCl as the control group. Immerse the AChE / AgNPs@PVA / RGO / SPE sensor in the control group and the experimental group. After the reaction in the dark, a portable Raman spectrometer is used to measure the Raman spectrum peak 639 cm -1 Spectral signal to obtain spectral signal change value; (3) Draw a standard curve with the concentration of methyl parathion standard solution as the horizontal axis and the spectral signal change value as the vertical axis; (4) The AChE / AgNPs@PVA / RGO / SPE sensor was immersed in pesticides, and the Raman spectrum peak at 639 cm was measured using a portable Raman spectrometer. -1 The spectral signal is used to obtain the spectral signal change value, which is substituted into the standard curve obtained in step (3) to obtain the concentration of methyl parathion in the pesticide.

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