Preparation method and application of multifunctional molecularly imprinted electrochemical sensor with double amplification mechanism

Abstract

The application belongs to the field of sensing detection, and relates to a preparation method of a multifunctional molecular imprinting electrochemical sensor with a double amplification mechanism and application thereof. The multifunctional molecular imprinting electrochemical sensor is prepared by using amino-reduced graphene oxide with a large specific surface area as a carrier, diuron as a template, o-phenylenediamine with good affinity as a bulk functional monomer, 3,4-ethylenedioxythiophene with multiple functional groups as a cross-linking functional monomer, and silver nanowires as a dopant. The multifunctional molecular imprinting polymer film with three-dimensional cavities is synthesized on the surface of an electrode modified with the carrier by an electro-polymerization method. Compared with a traditional molecular imprinting polymer film, the multifunctional molecular imprinting electrochemical sensor can improve the specific recognition ability of diuron by playing a complementary function and a synergistic effect in molecular recognition from two functionalization ways of organic and inorganic, and realize double amplification of a detection signal. The sensor can be used for efficient and specific detection of diuron residues in cotton and soil, can be recycled, can reduce detection cost, and has good practical application value.

Description

TECHNICAL FIELD

[0001] The application belongs to the field of sensing detection, and relates to a preparation method of a multifunctional molecular imprinting electrochemical sensor with a dual amplification mechanism and application thereof. BACKGROUND

[0002] Diuron (DU) is a cotton defoliant with high efficiency at low temperature, which helps to unify the maturity of cotton before mechanical harvesting and reduces the impurity content of cotton. DU is widely used in cotton planting areas. It is reported that DU is harmful to human health even at a lower concentration, and DU is stable in the natural environment (can exist in the soil for about 300 days) and difficult to degrade. Therefore, it is necessary to detect DU quickly, sensitively and specifically. The developed detection methods are mainly some chromatography combined techniques and electrochemical methods. Electrochemical methods have attracted the attention of researchers due to their simplicity, low cost and high sensitivity. Based on the low electrochemical activity of DU, researchers have prepared various nanomaterials (such as carbon nanocomposites, metal-organic framework materials, covalent organic framework materials and metal oxides) to construct sensors for direct detection of DU, which exhibit good analytical performance. However, most of them lack specific recognition ability, resulting in limited selectivity and anti-interference performance, so it is necessary to further improve the selectivity and sensitivity (main purpose) of the sensor in the face of complex sample environment.

[0003] Molecular imprinting polymers (MIPs) are formed by polymerizing functional monomers with template molecules through molecular imprinting technology, and the corresponding imprint cavities are left in the polymer matrix after removing the template molecules. These cavities are unique to the template in shape structure and chemical functionality, which ensures the selectivity of MIPs. In addition, MIPs have the advantages of low cost and high chemical stability. In the preparation of MIPs, functional monomers play a key role in selectivity and sensitivity performance. Most of the current MIPs involve only one functional monomer, and the structure and functional groups are relatively simple, which can easily lead to polymer structure fracture and collapse during elution and repeated detection. In addition, the MIPs prepared by traditional electro-polymerization also have the problem of too dense structure and lack of effective recognition sites.

[0004] Therefore, it is necessary to innovate the structure of MIPs, increase specific recognition sites, enrich binding modes, and strengthen the transmission electron capability of the network interface of MIPs to further improve the selectivity and sensitivity of the sensor. SUMMARY

[0005] For the structure and functional groups of most MIPs are relatively single at present, the current functional MIPs are only simply from adding functional monomers or doping nanomaterials in any aspect, and the specificity recognition ability and the electronic transmission ability of MIPs are improved in a single way, although the method is effective, but there are still limitations in the single way improvement. In view of the deficiencies of the prior art, the present application intends to start from the organic and inorganic aspects, in the presence of the body monomer o-phenylenediamine (o-PD), the crosslinking monomer 3,4-ethylenedioxythiophene (EDOT) is added to prepare a bifunctional MIP with rich functional groups and excellent conductive performance; at the same time, doping silver nanowires (AgNWs) can not only increase the surface area of MIP and improve the imprinting efficiency, but also can be used as a wire to speed up signal transmission, and the dual-path functionalization plays a complementary function and synergistic effect in molecular recognition, and improves the detection performance of the sensor to DU.

[0006] The application provides a preparation method of a multifunctional molecular imprinting electrochemical sensor with a double amplification mechanism and application thereof.

[0007] In a first aspect, the application provides a preparation method of a molecular imprinting electrochemical sensor, comprising: loading a polymer containing a template molecule and doped with silver nanowires onto an amino-reduced graphene oxide modified electrode; removing the template molecule on the electrode to obtain the molecular imprinting electrochemical sensor.

[0008] In a second aspect, the application further provides a molecular imprinting electrochemical sensor, comprising an amino-reduced graphene oxide modified electrode and a multifunctional molecular imprinting polymer film prepared from silver nanowires and o-phenylenediamine and 3,4-ethylenedioxythiophene.

[0009] In a third aspect, the application further provides an application of a molecular imprinting electrochemical sensor, and the molecular imprinting electrochemical sensor is used for detecting pesticides.

[0010] In order to achieve the above technical purposes, the application adopts the following scheme:

[0011] A preparation method of a multifunctional molecular imprinting electrochemical sensor with a double amplification mechanism, specifically comprising the following steps:

[0012] (1) Preparation of NH2-rGO:

[0013] The reduced graphene oxide (rGO) is ultrasonically dispersed into an ethanol-water solution, then ammonia propyl triethoxysilane (APTES) and hydrochloric acid (HCl) are sequentially added, and after mixing and stirring, centrifugation is performed, and the precipitate is taken out, and then ethanol and water are sequentially used for cleaning (to remove unreacted residues), and finally the amino-reduced graphene oxide (NH2-rGO) solid is obtained after drying;

[0014] (2) Preparation of AgNWs:

[0015] First, ethylene glycol is heated to a certain temperature in an oil bath, polyvinylpyrrolidone (PVP) is added for the first heating reflux, then sodium chloride (NaCl) solution is added for the second heating reflux, and finally silver nitrate (AgNO3) solution is added for the third reflux heating. After heating treatment, the precipitate is collected by centrifugation, and washed with acetone and ethanol respectively. The washed product is the silver nanowire (AgNWs) solid, which is dispersed in aqueous solution to obtain an AgNWs suspension.

[0016] (3) The glassy carbon electrode (GCE) is polished and polished with aluminum oxide powder, and then ultrasonically cleaned in ultrapure water, ethanol, and ultrapure water and dried to obtain a pretreated glassy carbon electrode. Then the NH2-rGO solid prepared in step (1) is dispersed in ultrapure water to prepare an NH2-rGO dispersion. The NH2-rGO dispersion is uniformly modified to the surface of the pretreated GCE, and dried at room temperature to obtain an NH2-rGO / GCE.

[0017] (4) The NH2-rGO / GCE obtained in step (3) is immersed in a mixed solution of acetonitrile and acetate buffer solution containing o-phenylenediamine (o-PD, functional monomer), 3,4-ethylenedioxythiophene (EDOT, cross-linking monomer), AgNWs (dopant) and DU (template molecule), and a certain potential window is set for continuous cyclic voltammetry scanning. After scanning, the electrode is taken out and dried at room temperature to obtain a modified electrode, which is denoted as P-PD-EDOT@Ag / NH2-rGO / GCE.

[0018] (5) The modified electrode (P-PD-EDOT@Ag / NH2-rGO / GCE) obtained in step (4) is immersed in an elution solution and eluted (to remove the template molecule) under stirring. After elution, it is rinsed with ultrapure water and dried to obtain a multifunctional molecularly imprinted electrochemical sensor with a dual amplification mechanism, denoted as MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE.

[0019] Preferably, in step (1), the amount of rGO, ethanol-water solution, APTES, and HCl is 10 mg: 30 mL: 1.5 g: 50 μL; wherein the ethanol-water solution is a solution obtained by mixing ethanol and water in a volume ratio of 1:1; the ultrasonic dispersion time is 2 h, and the stirring time is 10 h; the drying temperature is 60°C, and the time is 15 h.

[0020] Preferably, in step (2), the volume ratio of ethylene glycol, polyvinylpyrrolidone, NaCl solution and AgNO3 solution is 30 mL:0.8 g:5 mL:10 mL, wherein the concentration of the NaCl solution is 1 mg / mL, and the concentration of the AgNO3 solution is 0.15 mol / L; the stirring speed is 25 rpm, the heating temperature is 160 DEG C, the first, second and third heating reflux times are 60 min, 5 min and 80 min respectively, and the centrifugal washing times are 3-5 times; the concentration of the AgNWs suspension is 5.3 mg / mL.

[0021] Preferably, in step (3), the diameter of the glassy carbon electrode (GCE) is 2 mm; the particle size of the aluminum oxide powder used is 0.05 μm; and the concentration of the NH2-rGO dispersion liquid is 1 mg / mL; the amount of the NH2-rGO dispersion liquid used for modification is 3.5 μL.

[0022] Preferably, in step (4), the volume ratio of acetonitrile and acetate buffer solution is 1:1, wherein the concentration of the acetate buffer solution is 0.1 mol / L, and the pH value is 5.0; the final concentrations of o-phenylenediamine (o-PD, functional monomer), 3,4-ethylenedioxythiophene (EDOT, cross-linking monomer), AgNWs (dopant) and DU (template molecule) in the mixed solution are 4 mmol / L, 6 mmol / L, 30 mg / L and 5 mmol / L respectively; the potential window for cyclic voltammetry scanning is -0.4-1.2 V, the scanning speed is 50 mV / s, and the number of cycles is 15.

[0023] Preferably, in step (5), the elution solution is a mixed solution of ethanol and acetic acid mixed at a volume ratio of 9:1, the stirring speed is 60 rpm, and the elution time is 90 s.

[0024] The multifunctional molecularly imprinted electrochemical sensor with a dual amplification mechanism prepared by the method is used for pesticide detection, and is particularly used for DU detection.

[0025] The detection steps are as follows:

[0026] (1) First, prepare DU standard solutions with different concentrations, then immerse the MIP (o-PD-EDOT@Ag) / NH2-rGO / GCE sensor in V1 volume of the DU standard solutions with different concentrations, and incubate under stirring, then take out the sensor, wash with ultrapure water to remove unbound DU molecules, and obtain the incubated MIP (o-PD-EDOT@Ag) / NH2-rGO / GCE.

[0027] The current signal of the sensor MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE is in one-to-one correspondence with the concentration of the DU standard solution, that is, the signal response of one sensor corresponds to one concentration of the DU standard solution;

[0028] (2) The MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE after incubation in step (1) is used as a working electrode, the Ag / AgCl electrode is used as a reference electrode, and the platinum wire electrode is used as a counter electrode for electrochemical detection; and the standard curve is established by taking the lg value of the concentration of the DU standard solution as the abscissa and the generated current signal as the ordinate;

[0029] (3) Detection of the actual sample DU: first, obtain the sample solution; then immerse the sensor MIP / NH2-rGO@AuNGs / GCE in V2 volume of the sample solution, incubate at room temperature, take out, wash with ultrapure water to remove unbound DU molecules, and then perform electrochemical detection to obtain the corresponding generated current signal; then the generated current signal is brought into the standard curve established in step (2), and the concentration of DU in the actual sample can be obtained, and finally the detection of DU in the actual sample is realized.

[0030] Preferably, the concentration of the DU standard solution in step (1) is 0.2 ng / mL to 10 μg / mL;

[0031] Preferably, the incubation time in steps (1) and (3) is 5 to 30 min, the volume ratio of V1 to V2 is 1:1, and the stirring speed is 50 rpm;

[0032] Preferably, the electrochemical detection instrument in steps (2) and (3) is a CHI852D (Shanghai Chenhua) electrochemical workstation, and the scanning voltage range during detection is 1.0-1.4 V.

[0033] The application prepares a multifunctional molecular imprinting-based electrochemical sensor with a double amplification mechanism. First, a high-conductivity amino-functionalized reduced graphene oxide composite (NH2-rGO) is prepared as a substrate to provide a large surface area for the formation of MIP. A good affinity o-PD is used as a bulk monomer, and a good conductivity EDOT is introduced as a cross-linking monomer, which is expected to improve the electron transmission capacity of MIP on the one hand, and to introduce more functional groups and form more imprint cavities after the addition of EDOT, thereby enhancing the binding capacity between the imprint sites and the target and improving the recognition efficiency of the target. Acetonitrile is added during the polymerization process, which not only improves the poor solubility of EDOT in the aqueous polymerization solution, but also acts as a pore-forming agent to increase the imprint sites. The doping of AgNWs first serves as a wire to bridge the electron transfer between the target in the MIP layer, and secondly, it can increase the roughness of the imprint film surface and increase the imprint sites. Compared with the traditional method, this one-step copolymerization process from two pathways overcomes the limitations of single functionalization and simultaneously improves the selectivity and sensitivity of the MIP sensor. To the best of our knowledge, this multifunctional MIP electrochemical sensor for DU detection has not been reported. Based on this, the application proposes a high-performance molecular imprinting electrochemical sensor with NH2-rGO as the substrate and multifunctional MIP (MIP (o-PD-EDOT@AgNWs)) as the recognition element, which has the advantages of simple production, low cost, strong selectivity, high sensitivity, high reuse rate and good stability, and realizes high specificity and high sensitivity analysis of DU residues in actual samples. This method can be used for large-scale production and commercial application, and provides a new way for the functionalization of MIP, which proposes a double signal amplification strategy from the organic and inorganic dimensions, and fills the gap in the existing DU detection technology.

[0034] The application has the following advantages:

[0035] (1) The application introduces another cross-linking monomer (EDOT) from the organic perspective and nanomaterials (AgNWs) from the inorganic perspective on the basis of the bulk functional monomer (o-PD), which not only enriches the types of functional groups in MIP and enhances the ability to bind target molecules, but also realizes complementary advantages under the synergistic action of the two effects and doubles the response signal. This double-path functionalization method simultaneously improves the selectivity and sensitivity of the MIP-based electrochemical sensor.

[0036] (2) The application adds acetonitrile to the polymerization solution, which not only improves the poor solubility of EDOT in the aqueous polymerization solution, but also acts as a pore-forming agent to increase the imprint sites, thereby increasing the number of bound target molecules and improving the analysis performance of the sensor.

[0037] (3) The multifunctional electrochemical sensor based on the electropolymerization molecular imprinting technology is used for detecting DU, has a wide linear range (0.2 ng / mL-10 μg / mL), a low detection limit of 89 pg / mL, high sensitivity (8.47 μA μM -1 cm -2 ), and strong specificity (an imprinting factor of 10.4). BRIEF DESCRIPTION OF DRAWINGS

[0038] Figure 1 Fig. 1 is a construction process and schematic diagram of the multifunctional molecular imprinting electrochemical sensor of the present application; the sub-figures (A) and (B) are current response schematic diagrams of the sensor after combining and eluting DU, respectively.

[0039] Figure 2 Fig. 2 is a SEM diagram of MIP (o-PD) / NH2-rGO / GCE (A), MIP (EDOT) / NH2-rGO / GCE (B), MIP (o-PD-EDOT) / NH2-rGO / GCE (C), and MIP (o-PD-EDOT@Ag) / NH2-rGO / GCE (D).

[0040] Figure 3 Fig. 3 is a current response diagram of different sensors to DU and an imprinting factor diagram (A), and a comparison diagram of the sensitivity of different sensors for detecting DU (B).

[0041] Figure 4 Fig. 4 is a linear relationship diagram between the concentration of the standard solution of DU and the current with 10 as the base (A), and the selectivity of the multifunctional molecular imprinting electrochemical sensor (B).

[0042] Figure 5 Fig. 5 is a reusability diagram of the multifunctional molecular imprinting electrochemical sensor (A), and an 8-day stability diagram of the multifunctional molecular imprinting electrochemical sensor (B). DETAILED DESCRIPTION

[0043] A number of illustrative embodiments of the present application are described below in detail. The detailed description is not intended to limit the application, but rather to explain certain aspects, features, and embodiments of the application.

[0044] It should be understood that the terms used herein are merely descriptive, but are not intended to limit the application. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the application pertains. Although preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application.

[0045] Many modifications and variations of the present application described in the specification are possible without departing from the scope or spirit of the application. Other implementations of the application will be apparent to those skilled in the art from consideration of the specification and practice of the application disclosed herein. The specification and examples are illustrative only.

[0046] Example 1:

[0047] The present embodiment provides a preparation method of a multifunctional molecularly imprinted electrochemical sensor with double amplification mechanism, according to the material preparation and sensor construction diagram shown in the figure, the specific steps are as follows: Figure 1

[0048] (1) 10 mg of reduced graphene oxide (rGO) was dispersed in 30 mL of an ethanol-water mixed solution with a volume ratio of 1:1 and ultrasonically dispersed for 2 h. Then 1.5 g of aminopropyltriethoxysilane (APTES) and 50 μL of hydrochloric acid (HCl) were added respectively, and stirred for 10 h. After centrifugation at 8000 rpm for 10 min, the precipitate was taken out, and then washed with ethanol and water three times to remove the residual unreacted substances. Finally, it was dried at 60°C for 15 h to obtain NH2-rGO solid.

[0049] The prepared NH2-rGO composite material is in the shape of a wrinkled two-dimensional nanosheet, has a large specific surface area and excellent conductivity; AgNWs are in the form of nanowires, with a micron-level length and a diameter of about 50 nm, which is conducive to the transmission of electrons as a wire.

[0050] (2) 30 mL of ethylene glycol was heated to 160°C in a 100 mL three-necked flask in an oil bath, 0.8 g of polyvinylpyrrolidone (PVP, M=4000) was added to the flask, and the temperature was kept at reflux for 1 h. Then 5 mL of 1 mg / mL sodium chloride (NaCl) solution was added, and the temperature was kept at reflux for 5 min. Finally, 10 mL of 0.15 mol / L silver nitrate (AgNO3) solution was slowly added, and the temperature was kept at reflux for 80 min (showing a stable silver-white color). Then the precipitate was collected by centrifugation, and washed with acetone and ethanol three times respectively to obtain silver nanowire (AgNWs) solid. The AgNWs solid was redispersed in an aqueous solution to obtain an AgNWs suspension with a concentration of 5.3 mg / mL, which was stored in a refrigerator at 4°C in the dark.

[0051] ​(3) The glassy carbon electrode (GCE, Φ = 2 mm) was polished and polished with 0.05 μm aluminum oxide powder, and then ultrasonically cleaned in ultrapure water, ethanol and ultrapure water in turn, and dried. Then cyclic voltammetry scanning was carried out in 5 mmol / L potassium ferricyanide solution containing 0.1 mol / L potassium chloride. When the peak potential difference was less than 90 mV, it was proved that the GCE pretreatment was completed, and the pretreated GCE was obtained.

[0052] (4) The NH2-rGO composite prepared in step (1) was ultrasonically dispersed in ultrapure water to prepare a 1 mg / mL NH2-rGO dispersion; 3.5 μL of the NH2-rGO dispersion was taken with a pipette gun and uniformly dropped on the surface of the pretreated GCE, and dried to obtain NH2-rGO / GCE.

[0053] (5) The acetonitrile and 0.1 mol / L acetate buffer solution (pH 5.0) were mixed according to the volume ratio of 1:1 to prepare the polymerization solvent, and then 4 mmol / L o-phenylenediamine (o-PD, functional monomer), 6 mmol / L 3,4-ethylenedioxythiophene (EDOT, crosslinking monomer), 30 mg / L silver nanowire (AgNWs, dopant) and 5 mmol / L diuron (DU, template molecule) were added to the mixed solution. The modified electrode was obtained by 15 times of continuous cyclic voltammetry scanning at a voltage range of-0.4-1.2 V and a scanning rate of 50 mV / s by electropolymerization, and then dried at room temperature, and was recorded as P-PD-EDOT@Ag / NH2-rGO / GCE.

[0054] (6) The modified electrode prepared in step (5) was immersed in a mixed solution of ethanol and acetic acid, and the volume ratio of ethanol and acetic acid was 9:1. The solution was stirred at a speed of 60 rpm for 90 s to elute the DU molecules, and then washed with ultrapure water and dried to obtain a multifunctional molecularly imprinted electrochemical sensor, which was recorded as MIP (o-PD-EDOT@Ag) / NH2-rGO / GCE.

[0055] Comparative Example 1:

[0056] Different molecularly imprinted based electrochemical sensors were prepared according to the steps of Example 1, and the difference was that the substances participating in the reaction in the polymerization solution in step (5) were different.

[0057] Only 4 mmol / L o-phenylenediamine (o-PD, functional monomer) and 5 mmol / L diuron (DU, template molecule) participated in the polymerization, and the eluted modified electrode was recorded as MIP (o-PD) / NH2-rGO / GCE.

[0058] Only 6 mmol / L of 3,4-ethylenedioxythiophene (EDOT, cross-linking monomer) and 5 mmol / L of diuron (DU, template molecule) were involved in polymerization, and the modified electrode after elution was denoted as MIP(EDOT) / NH2-rGO / GCE;

[0059] When 4 mmol / L of o-phenylenediamine (o-PD, functional monomer), 6 mmol / L of 3,4-ethylenedioxythiophene (EDOT, cross-linking monomer) and 5 mmol / L of diuron (DU, template molecule) were involved simultaneously, the modified electrode after elution was denoted as MIP(o-PD-EDOT) / NH2-rGO / GCE.

[0060] The above three different sensors were compared with MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE in Example 1 in terms of morphology, selectivity and sensitivity.

[0061] Figure 2 Figure (A) is the SEM image of MIP(o-PD) / NH2-rGO / GCE (denoted as MIP(o-PD) in the figure), and the surface retains the characteristic wrinkle nanosheet structure of the underlying graphene, and part of the cavities appears due to the presence of imprint cavities; Figure (B) is the SEM image of MIP(EDOT) / NH2-rGO / GCE (denoted as MIP(EDOT) in the figure), and a large number of holes are exposed on the surface due to the swelling behavior of the polymer after elution of the template molecule; Figure (C) is the SEM image of MIP(o-PD-EDOT) / NH2-rGO / GCE (denoted as MIP(o-PD-EDOT) in the figure), and the interface exhibits higher roughness under the action of two functional monomers, and a larger number of imprint cavities are exposed, which is conducive to the recombination of DU; Figure (D) is MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE (denoted as MIP(o-PD-EDOT@Ag) in the figure), and the nanowires are distributed on the surface due to the addition of AgNWs, which is conducive to the mass transfer and charge transfer of the interface as a conductor.

[0062] In addition, the imprint factor (IF) can be used to further characterize the specificity of the molecularly imprinted electrochemical sensor to the target substance, and the higher the value, the stronger the binding ability and the better the specificity. IF is the ratio of the signals of the imprinted sensor (MIP) and the non-imprinted sensor (NIP). From Figure 3As can be seen in the middle (A), the types of functional groups are enriched under the action of two functional monomers, the number of imprint cavities is increased, resulting in the MIP(o-PD-EDOT) / NH2-rGO / GCE having stronger specificity than the previous single functional monomer MIP sensor. After the introduction of AgNWs, it will provide a larger surface area for fixing DU molecules during polymerization to create more recognition sites, while increasing the roughness of the MIP, which is beneficial to binding more DU.

[0063] Figure 3 In the middle (B), the sensitivity of different sensors is compared. Compared with MIP(o-PD) / NH2-rGO / GCE, MIP(EDOT) / NH2-rGO / GCE shows higher sensitivity, which is mainly due to the fact that poly 3,4-ethylenedioxythiophene (PEDOT) has stronger charge transport ability than poly o-phenylenediamine (P-PD), resulting in a stronger current signal of DU. The addition of PEDOT also compensates for the conductivity of P-PD, and the MIP prepared by copolymerization of the two monomers can capture more DU molecules, so the sensitivity is further improved. The doping of AgNWs is beneficial to further promote the electronic transfer of the DU molecules bound in the MIP network, so that the signal is further amplified and the sensitivity is increased.

[0064] The above results prove that the MIP is functionalized from both organic and inorganic angles, and under the synergistic action of the two functionalization methods, the specificity and sensitivity of the sensor are simultaneously improved, which is beneficial to the practical application value in detection.

[0065] Example 2:

[0066] The MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE sensor provided in Example 1 was immersed in a DU standard solution containing different concentrations and stirred for 20 min to allow the DU molecules to recombine with the cavities; then the sensor surface was washed with ultrapure water to remove unbound DU molecules, and the resulting product was labeled as DU / MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE;

[0067] The concentrations of the DU standard solution were 0.0002, 0.0005, 0.001, 0.005, 0.01, 0.05, 0.1, 0.5, 1.0, 5.0, and 10.0 μg / mL, respectively.

[0068] The above product (DU / MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE) was used as the working electrode, an Ag / AgCl (saturated KCl) electrode as the reference electrode, and a platinum wire electrode as the auxiliary electrode, and the electrochemical signals were detected and recorded by an electrochemical workstation (Shanghai Chenhua, CHI852D). The differential pulse voltammetry method was used to collect the electrical signals, and the scanning potential was from 1.0 V to 1.4 V. The current signal of DU was used as the ordinate, and the lg value corresponding to the concentration of DU was used as the abscissa, and the corresponding standard curve was established for the concentration analysis of DU in the actual sample.

[0069] From Figure 4 As shown in (A), with the increase of the concentration of DU, the oxidation peak current gradually increased, and in the concentration range of 0.2 ng / mL to 10 μg / mL, the logarithmic value with 10 as the base and the current value had a good linear relationship, and the linear equation was I = 0.47lgC DU + 0.34 (R 2 = 0.996), and the detection limit was 89 pg / mL.

[0070] Comparative Example 2:

[0071] According to the same steps of Example 1, a non-imprinted polymer (NIP) was prepared, and the difference was that no template molecule DU was added in the electro-polymerization process of step (5), and the obtained sensor was recorded as NIP(o-PD-EDOT@Ag) / NH2-rGO / GCE.

[0072] The MIP (o-PD-EDOT@Ag) / NH2-rGO / GCE sensor provided in Example 1 (recorded as MIP) and the NIP (o-PD-EDOT@Ag) / NH2-rGO / GCE sensor in Comparative Example 2 (recorded as NIP) were subjected to selective test: wherein blank means no target substance, defined as blank sample; the interfering substances are chemical structural analogues of DU and other coexisting substances in actual samples, including isoproturon (ISO), ethephon (ETH), thiamethoxam (TMX), thidiazuron (TDZ), monuron (MU), carbendazim (CBZ), imidacloprid (IMC), benomyl (BM), and the corresponding concentrations of the above-mentioned interfering substances are all 1 μg / mL; Mix is a mixture of the above-mentioned interfering substances, and the corresponding concentrations of the above-mentioned interfering substances are all 1 μg / mL; DU is the target substance, and the corresponding concentration is 10 ng / mL; Mix+DU is a mixture of the interfering substances and DU, and in the mixture, the concentration of Mix is 1 μg / mL, and the concentration of DU is 10 ng / mL.

[0073] The sensors of Comparative Example 2 and Example 1 were respectively placed in the above-mentioned detection substances for incubation for 20 min, then taken out, and the electrodes were rinsed with ultrapure water. After the electrodes were dried, the detection was performed, and the electrical signals of the corresponding substances were obtained.

[0074] from Figure 4 (B) It was found that MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE(MIP) only showed a significant current signal when DU was present. However, due to the lack of a specifically identifiable imprint cavity, NIP(o-PD-EDOT@Ag) / NH2-rGO / GCE(NIP) did not show a significant difference in response to DU and other interfering substances.

[0075] The relevant current response values ​​are shown in Table 2.

[0076] Table 2

[0077]

[0078] Table 1 shows that MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE has the highest IF value for DU, at 10.4, which is much higher than other non-target substances. This proves that the sensor has a large number of imprinted cavities that can specifically identify DU, and has outstanding selectivity and high anti-interference ability for DU, making it suitable for the detection of real samples in complex environments.

[0079] Example 3:

[0080] Performance analysis of molecularly imprinted electrochemical sensors:

[0081] To investigate the reusability of this multifunctional molecularly imprinted electrochemical sensor, in Figure 5 In (A), the same MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE sensor was subjected to 10 consecutive cycles of DU testing and elution steps, and the changes in current response were observed. The experiment was performed in triplicate, with the signal response of DU being detected 10 times consecutively. Due to slight degradation of the imprinted holes, the peak current after the 10th repeated detection still maintained 89% of the initial response, and the relative standard deviation (RSD) was 3.1%, demonstrating good reusability.

[0082] Figure 5 (B) underwent a 7-day stability test. On the 7th day, the current response still retained 94% of the initial value, and the RSD was 3.8%, indicating that MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE has excellent stability.

[0083] Example 4:

[0084] The actual sample was analyzed using the multifunctional molecularly imprinted electrochemical sensor (MIP) customized for diuron, as follows:

[0085] (1) The cotton and soil collected in Tomshuk region of Xinjiang were used as actual samples to investigate the residual content of DU in them.

[0086] 1 g of cotton sample was weighed, 10 mL of methanol and 10 mL of ultrapure water were added, and ultrasonic treatment was performed for 20 min. After centrifugation at 8000 rpm for 15 min, the supernatant was collected, filtered with a 0.22 μm cellulose filter membrane, and the cotton actual sample extract was obtained.

[0087] The collected soil sample was air-dried, finely ground to 200 mesh nylon screen, and then collected for use. Then, 10 g of soil sample was transferred to a centrifuge tube, extraction solvent (5 mL of ultrapure water and 10 mL of acetonitrile) was added, and it was stirred vigorously in a vortex mixer for 10 min. Then, salting-out was performed with 3 g of NaCl and 4 g of MgSO4, and the extracted organic phase and aqueous phase were separated. After centrifugation at 5000 rpm for 5 min, the supernatant was collected and filtered with a cellulose membrane to obtain the soil actual sample extract, which was ready for analysis.

[0088] (2) The standard addition method was used to add 1 ng / mL, 10 ng / mL and 100 ng / mL of DU standard solution to the cotton extract and soil extract, respectively, and the samples after standard addition were detected. The recovery rate of DU in the cotton extract was calculated to be 94.3% to 100%, and the RSD was less than 4.00%; the recovery rate of DU in the soil extract was 94.9% to 101%, and the RSD was less than 4.70%, indicating that the sensor can be used for actual sample detection.

[0089] (3) The detection results of the constructed sensor were further verified by high performance liquid chromatography-mass spectrometry (HPLC-MS). The recovery rates of DU in the cotton extract and soil extract were 89.6% to 91.3% and 87.2% to 99.0%, respectively, and the detection values were basically consistent with the detection results of the constructed sensor, which verified that the constructed sensor had good reliability in actual sample analysis.

[0090] The relevant test results are shown in Table 1.

[0091] Table 1.

[0092]

[0093] It is explained that the above content can enable those skilled in the art to more fully understand the present application, and is not a limitation on the present application, and the present application is not limited to the above examples. Therefore, those skilled in the art, within the scope of the present application, make changes, modifications, additions or substitutions, and all technical solutions and improvements without departing from the spirit and technical essence of the present application, should belong to the protection scope of the present application.

Claims

1. A method for preparing a multifunctional molecularly imprinted electrochemical sensor with dual amplification mechanism, characterized in that, It comprises the following steps: (1) Preparation of NH2-rGO: The reduced graphene oxide is ultrasonically dispersed in an ethanol-water solution, then ammonia propyl triethoxysilane and hydrochloric acid are added in sequence, mixed and stirred, and then centrifuged, and the precipitate is taken out, washed with ethanol and water in sequence, and finally dried to obtain aminated reduced graphene oxide; (2) Preparation of AgNWs: First, ethylene glycol is heated to a certain temperature in an oil bath, polyvinylpyrrolidone is added for the first heating reflux, then sodium chloride solution is added for the second heating reflux, and finally silver nitrate solution is added for the third reflux heating, and the precipitate is collected after heating treatment, and then washed with acetone and ethanol, and the product after washing is a silver nanowire solid, which is dispersed in water to obtain an AgNWs suspension; (3) The glassy carbon electrode is polished and polished with aluminum oxide powder, and then ultrasonically cleaned in ultrapure water, ethanol and ultrapure water in sequence and dried to obtain a pretreated glassy carbon electrode; then the NH2-rGO solid prepared in step (1) is dispersed in ultrapure water to prepare an NH2-rGO dispersion; the NH2-rGO dispersion is uniformly modified on the surface of the pretreated GCE, and dried at room temperature to obtain an NH2-rGO / GCE; (4) The NH2-rGO / GCE obtained in step (3) is immersed in a mixed solution of acetonitrile and acetate buffer solution containing o-phenylenediamine, 3,4-ethylenedioxythiophene, AgNWs and diuron (DU), and a certain potential window is set for continuous cyclic voltammetry scanning; after scanning, the electrode is dried at room temperature, and the modified electrode obtained is marked as P-PD-EDOT@Ag / NH2-rGO / GCE; (5) The modified electrode P-PD-EDOT@Ag / NH2-rGO / GCE obtained in step (4) is immersed in an elution solution and eluted under stirring, and then washed with ultrapure water and dried to obtain a multifunctional molecularly imprinted electrochemical sensor with a dual amplification mechanism, marked as MIP(o-PD-EDOT@Ag) / NH2-rGO / GCE.

2. The method for preparing a multifunctional molecularly imprinted electrochemical sensor with dual amplification mechanism according to claim 1, characterized in that, In step (1), the amount ratio of the reduced graphene oxide, the ethanol-water solution, the ammonia propyl triethoxysilane, and the HCl is 10 mg:30 mL:1.5 g:50 μL; wherein the ethanol-water solution is a solution obtained by mixing ethanol and water in a volume ratio of 1:1; the ultrasonic dispersion time is 2 h, and the stirring time is 10 h; the drying temperature is 60°C, and the time is 15 h.

3. The method for preparing a multifunctional molecularly imprinted electrochemical sensor with dual amplification mechanism according to claim 1, characterized in that, In step (2), the volume ratio of ethylene glycol, polyvinylpyrrolidone, sodium chloride solution and silver nitrate solution is 30 mL:0.8 g:5 mL:10 mL, wherein the concentration of the sodium chloride solution is 1 mg / mL, and the concentration of the silver nitrate solution is 0.15 mol / L; the stirring speed is 25 rpm, the heating temperature is 160 ℃, the first, second and third heating reflux times are 60 min, 5 min and 80 min respectively, and the centrifugal washing times are 3-5 times; the concentration of the AgNWs suspension is 5.3 mg / mL.

4. The method for preparing a multifunctional molecularly imprinted electrochemical sensor with dual amplification mechanism according to claim 1, characterized in that, In step (3), the diameter of the glassy carbon electrode is 2 mm; the particle size of the aluminum oxide powder used is 0.05 μm; the concentration of the NH2-rGO dispersion liquid is 1 mg / mL; and the amount of the NH2-rGO dispersion liquid used for modification is 3.5 μL.

5. The method for preparing a multifunctional molecularly imprinted electrochemical sensor with dual amplification mechanism according to claim 1, characterized in that, In step (4), the volume ratio of acetonitrile and acetate buffer solution is 1:1, wherein the concentration of the acetate buffer solution is 0.1 mol / L, and the pH value is 5.0; the final concentrations of o-phenylenediamine, 3,4-ethylenedioxythiophene, AgNWs and diuron in the mixed solution are 4 mmol / L, 6 mmol / L, 30 mg / L and 5 mmol / L respectively; the potential window for cyclic voltammetry scanning is-0.4-1.2 V; the scanning speed is 50 mV / s; and the number of cycles is 15.

6. The method for preparing a multifunctional molecularly imprinted electrochemical sensor with dual amplification mechanism according to claim 1, characterized in that, In step (5), the elution solution is a mixed solution of ethanol and acetic acid with a volume ratio of 9:1; the stirring speed is 60 rpm; and the elution time is 90 s.

7. A multifunctional molecularly imprinted electrochemical sensor with a dual amplification mechanism prepared by the method according to any one of claims 1-6.

8. Use of the multifunctional molecularly imprinted electrochemical sensor with dual amplification mechanism according to claim 7 for detecting pesticides, characterized in that, The pesticide comprises diuron.

9. Use according to claim 8, characterized in that, The detection of diuron is as follows: (1) First, prepare diuron standard solutions with different concentrations, then immerse the MIP (o-PD-EDOT@Ag) / NH2-rGO / GCE sensor in V1 volume of diuron standard solution with different concentrations, and incubate under stirring, then take out the sensor and wash with ultrapure water to obtain the incubated MIP (o-PD-EDOT@Ag) / NH2-rGO / GCE; (2) Take the incubated MIP (o-PD-EDOT@Ag) / NH2-rGO / GCE in step (1) as the working electrode, the Ag / AgCl electrode as the reference electrode, and the platinum wire electrode as the counter electrode, and perform electrochemical detection; take the lg value of the diuron standard solution concentration as the abscissa and the generated current signal as the ordinate to establish a standard curve. (3) Detection of the actual sample DU: first, obtain the sample solution; then immerse the sensor MIP / NH2-rGO@AuNGs / GCE into V2 volume of the sample solution, incubate at room temperature, take out, wash with ultrapure water, and then perform electrochemical detection to obtain the corresponding generated current signal; then the generated current signal is brought into the standard curve constructed in step (2), and the concentration of diuron in the actual sample can be obtained, and finally the detection of diuron in the actual sample is realized.

10. Use according to claim 9, characterized in that, The concentration of the diuron standard solution in step (1) is 0.2 ng / mL-10 μg / mL; the incubation time in steps (1) and (3) is 5-30 min, the volume ratio of V1 and V2 is 1:1, and the stirring speed is 50 rpm; the scanning voltage range during electrochemical detection in steps (2) and (3) is 1.0-1.4 V.

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