Preparation method and use method of an electrochemical molecular imprinting sensor for detecting DBP

Abstract

The application discloses a preparation method and use method of an electrochemical molecular imprinting sensor for detecting DBP. A PEDOT-PG nanocomposite is prepared through a liquid-liquid interface polymerization method, then the prepared PEDOT-PG is functionalized by using polylysine, so that the PEDOT / PG has good dispersity and stability, and good adhesion on the surface of the sensor. In the presence of DBP, a molecular imprinting film is prepared through a monomer electropolymerization method, so that the recognition response speed of the target molecule is improved, the molecular imprinting sensor has the advantages of structural stability, high mechanical strength, easy acquisition and low cost, and has the advantages of specificity for recognizing the target molecule.

Description

Technical Field

[0001] This invention relates to the field of electrochemical sensors, specifically to a method for preparing and using an electrochemically imprinted sensor for detecting DBP. Background Technology

[0002] Dibutyl phthalate (DBP), with the molecular formula C16H22O4, also known as dibutyl phthalate or 1,2-phthalate, is one of the most widely used phthalate esters (PAEs).

[0003] DBP is commonly used as a plasticizer to increase the flexibility, durability, and transparency of polymers such as PVC. Therefore, it is frequently used in items closely related to human life, such as plastic packaging, children's toys, medical devices, building materials, printing inks, cosmetics, and adhesives. However, because DBP molecules are linked to polymer carbon chains only by hydrogen bonds (or van der Waals forces) rather than chemical bonds, DBP is easily released and migrates into the environment, water bodies, and food through changes in external environmental factors such as temperature, water, and oils. As an endocrine disruptor, it can remain stable in the environment for a long time and is difficult to degrade. Through accumulation in the food chain, DBP can eventually accumulate in the human body, posing serious potential harm to human health. Numerous toxicological studies have shown that DBP has chronic and acute toxicity, adversely affecting the reproductive development and endocrine system of organisms, leading to metabolic disorders, malformations, cancer, and gene mutations. DBP is listed as a priority pollutant by the US Environmental Protection Agency (US-EPA), the European Union, and the China National Environmental Monitoring Center.

[0004] Existing methods for DBP detection include gas chromatography, high-performance liquid chromatography, gas chromatography-mass spectrometry, liquid chromatography-mass spectrometry, and enzyme-linked immunosorbent assay (ELISA). However, these methods require expensive equipment, toxic solvents, complex and time-consuming sample pretreatment and enrichment, and specialized and experienced researchers, all of which limit their widespread application. In recent years, rapid detection methods such as enzyme analysis and test strips have been developed, but they can only perform qualitative (or semi-quantitative) detection of the analyte, with lower sensitivity and accuracy than traditional detection methods. Furthermore, developed fluorescence and colorimetric methods are susceptible to signal interference from coexisting substances in the detection system, thus affecting the accuracy of the results. To overcome these shortcomings, there is an urgent need to develop simple, rapid, and sensitive sensing platforms for detecting DBP in samples. In addition, DBP has many homologues with similar molecular structures, such as DOP, DEHP, DEP, DINP, and DMP, requiring high specificity for the detection of DBP in test samples. Summary of the Invention

[0005] To address the problems mentioned in the background section, this invention provides a method for preparing an electrochemically imprinted sensor for detecting DBP, thereby solving the existing problems.

[0006] The technical solution adopted in this invention is:

[0007] A method for preparing an electrochemically imprinted sensor for detecting DBP includes the following steps:

[0008] S1 uses a liquid-liquid interface polymerization method to prepare PEDOT-PG nanocomposite material. Polylysine is used to functionalize the prepared PEDOT-PG, which gives PEDOT / PG good dispersibility and stability, as well as good adhesion to the sensor surface. The resulting material is used as the substrate of the sensor, which gives the sensor excellent sensitivity, thus obtaining PEDOT-PG-PLL nanocomposite material.

[0009] S2 used a drop-coating method to modify the PEDOT-PG-PLL nanocomposite material onto the surface of a bare glassy carbon electrode GCE, obtaining a modified electrode PEDOT-PG-PLL / GCE.

[0010] S3 uses DBP as a template molecule and O-PD as a polymer monomer to obtain pre-eluting molecularly imprinted sensors MIPs / PEDOT-PG-PLL / GCE on the surface of PEDOT-PG-PLL / GCE via electropolymerization. The use of electropolymerization molecular imprinting technology makes the prepared sensor specific for recognizing target molecules.

[0011] S4 elutes the DBP molecules in the molecularly imprinted membrane layer of the molecularly imprinted sensor MIPs / PEDOT-PG-PLL / GCE obtained in step S3 before elution, to obtain the DBP detection molecularly imprinted sensor MIPs-e / PEDOT-PG-PLL / GCE.

[0012] Further, in step S1, the method for preparing PEDOT-PG nanocomposite materials using liquid-liquid interfacial polymerization includes: using a volume of 1 ± 0.2 ml and a concentration of 1 mol L... -1 -3mol L -1 FeCl3 was added as an oxidant to a volume of 1 ± 0.2 ml and a concentration of 0.5 mg / ml. -1 -2.5mg.ml -1 After ultrasonic treatment, 2 ml of chloroform was slowly added to the PG dispersion. The resulting mixture was reacted at 45-70℃ for 8-12 hours under static conditions. After centrifugation, the resulting precipitate was washed multiple times with anhydrous ethanol and deionized water to generate PEDOT-PG nanocomposite material.

[0013] Furthermore, the concentration of EDOT in the chloroform is 15 mg / mL. -1 -30mg.ml -1 .

[0014] Further, the method for functionalizing the prepared PEDOT-PG with polylysine in step S1 includes: dispersing the prepared PEDOT-PG nanocomposite material in 1 ml of water, mixing it with a PEDOT-PG aqueous solution:PLL volume ratio of 3:1-1:1 and incubating it for 1-12 hours, and storing it under cold.

[0015] Furthermore, the ultrasonic treatment is performed for 10-30 minutes, followed by centrifugation at 12000-16000 rpm for 20-40 minutes. The resulting precipitate is then washed 2-3 times each with anhydrous ethanol and deionized water, and stored in a refrigerator at 2-5°C.

[0016] Further, in step S2, 5 μl of PEDOT-PG-PLL nanocomposite material is applied to the surface of the bare glassy carbon electrode GCE using a drop-coating method to obtain the modified electrode PEDOT-PG-PLL / GCE, which is then dried using infrared spectroscopy.

[0017] Further, the bare glassy carbon electrode GCE treatment step includes ultrasonically washing the glassy carbon electrode sequentially with nitric acid, ethanol, and deionized water for 1 minute each, polishing it on chamois leather with 0.3 μm polishing powder Al2O3, and then placing the electrode in 0.5 mmol / L... -1 K3[Fe(CN)6] and 0.1 mol L -1 In a KCl mixed solution, a stable cyclic voltammetric peak was obtained by cyclic voltammetry scanning at a scan rate of 50 mV / s in the potential range of -0.2 V to 0.6 V, with a potential difference of less than 80.

[0018] Further, in step S3, PEDOT-PG-PLL / GCE is placed in 0.01M PBS containing 1-5mM o-phenylenediamine and 0.5-3mM DBP, with the pH of the PBS being 7.4. Cyclic voltammetry scans are performed for 8-20 cycles in the range of 0-0.8V and a scan rate of 50mVs⁻¹ using an electropolymerization method. The electropolymerized electrode is then rinsed with deionized water and dried at room temperature to obtain the molecularly imprinted sensor MIPs / PEDOT-PG-PLL / GCE before elution.

[0019] Further, in step S4, the electrode is then immersed in a methanol-acetic acid mixture V. 甲醇 :V 20％乙酸 The sample was eluted in a 1:10 solution for 15-40 minutes to remove the template molecule DBP. The sample was then rinsed with deionized water to obtain the DBP detection molecularly imprinted electrochemical sensor MIPs-e / PEDOT-PG-PLL / GCE.

[0020] A method for using a molecularly imprinted sensor for detecting DBP includes placing the molecularly imprinted sensor in a PBS solution containing different concentrations of DBP at pH 2.5-6.5, stirring and adsorbing at room temperature for 10-30 min, and rinsing the electrode with water; placing the molecularly imprinted sensor in an electrolytic cell, wherein the detection base solution of the electrolytic cell is 0.5 mmol / L. -1 K3[Fe(CN)6] and 0.1 mol L - 1 A KCl mixed solution was used for differential pulse voltammetry detection by setting scanning parameters.

[0021] Compared with the prior art, the beneficial effects of the present invention are:

[0022] (1) This invention provides a method for preparing a molecularly imprinted sensor for detecting DBP. The molecularly imprinted electrochemical sensor combines the advantages of molecular imprinting technology and electrochemical sensing technology. It has both the fast response time and excellent sensitivity of electrochemical sensing technology and the specific recognition capability of molecular imprinting technology, providing a broad prospect for the development of highly sensitive and selective target detection and analysis methods.

[0023] (2) This invention provides a method for preparing a molecularly imprinted sensor for detecting DBP, thereby obtaining a molecularly imprinted sensor. The imprinted sites are located on the surface of the nanomaterial, which improves the recognition response speed of the target molecule and has specificity for recognizing the target molecule. Moreover, the molecularly imprinted sensor has the advantages of structural stability, high mechanical strength, easy availability, and low cost. Attached Figure Description

[0024] Figure 1 Cyclic voltammetry scans for different modified electrodes;

[0025] Figure 2 Nyquist plots for different modified electrodes;

[0026] Figure 3 The sensor's DPV response to different concentrations of DBP in K3[Fe(CN)6] and KCl electrolytes is shown in the graph.

[0027] Figure 4 This is a graph showing the linear relationship between the logarithm of DBP concentration and the change in DPV current.

[0028] Figure 5 This is the result of selective detection of DBP analogs by a molecularly imprinted sensor.

[0029] In the figure: a. PEDOT-PG-PLL / GCE; b. MIPs-e / PEDOT-PG-PLL / GCE; c. MIPs-e / PEDOT-PG-PLL / GCE (after DBP enrichment); d. MIPs / PEDOT-PG-PLL / GCE; e. GCE. Detailed Implementation

[0030] To clearly illustrate the technical features of this solution, the invention will be described in detail below through specific embodiments and in conjunction with the accompanying drawings. Many specific details are set forth in the following description to provide a thorough understanding of this application; however, this application may also be implemented in other ways different from those described herein. Therefore, the scope of protection of this application is not limited to the specific embodiments disclosed below.

[0031] Unless otherwise defined, all technical terms used herein have the same meaning as commonly understood by those skilled in the art. The technical terms used herein are for the purpose of describing particular embodiments only and are not intended to limit the scope of the invention.

[0032] Unless otherwise specified, all raw materials, reagents, instruments, and equipment used in this invention can be purchased commercially or prepared using existing methods.

[0033] Example 1

[0034] One embodiment of the present invention provides a method for preparing an electrochemically imprinted sensor for detecting DBP, comprising the following steps:

[0035] S1 prepared PEDOT-PG nanocomposite material by liquid-liquid interfacial polymerization, and functionalized the prepared PEDOT-PG with polylysine to obtain PEDOT-PG-PLL nanocomposite material.

[0036] Among them, PG is obtained by high-temperature etching of graphene with potassium hydroxide (KOH). Compared with two-dimensional graphene, PG has a three-dimensional uniform porous structure, a larger specific surface area and active sites, and better conductivity.

[0037] PEDOT, a polythiophene derivative, possesses unique structure and properties. The special ethylenedioxy groups at the 3- and 4-positions of thiophene not only lower the oxidation potential of the monomer but also prevent unwanted α,β- and β,β-couplings in the polymer backbone, giving the polymer high degree of conformation. In the doped state, it exhibits high conductivity, good stability, and superior transparency. It is considered one of the most promising conductive polymers in the field of electrochemistry and has received extensive research in recent years. The addition of inorganic materials (porous graphene) can, on the one hand, adjust the polymerization structure of PEDOT, improve carrier mobility, and expand the specific surface area. On the other hand, PEDOT helps form a uniform thin film with PG, improving the conductivity and stability of the sensor during operation.

[0038] Poly-L-lysine (PLL) is a cationic polymer with multiple amino groups. The multiple aminobutyl groups in PLL possess both the positive charge (-NH3+) of primary amines and the hydrophobic properties of the butyl spacers between primary amines. Therefore, using PLL for surface functionalization of materials can not only increase the conductivity of the material in aqueous solution but also enhance its dispersibility and stability. Furthermore, PLL itself has a certain viscosity, which can increase the adhesion of the composite material to the electrode surface.

[0039] S2 used a drop-coating method to modify the PEDOT-PG-PLL nanocomposite material onto the surface of a bare glassy carbon electrode GCE. After infrared drying, the modified electrode PEDOT-PG-PLL / GCE was obtained.

[0040] S3 uses DBP as a template molecule and O-PD as a polymer monomer to obtain molecularly imprinted sensors (MIPs) / PEDOT-PG-PLL / GCE on the surface of PEDOT-PG-PLL / GCE before elution by electropolymerization.

[0041] Among them, the poly(o-phenylene diamine) membrane has good insulation properties. After the template molecules are added, the carbonyl groups in the DBP molecules in the solution form hydrogen bonds with the amino groups in the o-phenylene diamine molecules and thus combine. Therefore, the DBP molecules are embedded in the polymer membrane during the electropolymerization of o-phenylene diamine.

[0042] S4 elutes the DBP molecules in the molecularly imprinted membrane layer of the molecularly imprinted sensor MIPs / PEDOT-PG-PLL / GCE obtained in step S3 before elution, to obtain the DBP detection molecularly imprinted sensor MIPs-e / PEDOT-PG-PLL / GCE.

[0043] Example 2

[0044] One embodiment of the present invention provides a method for preparing an electrochemically imprinted sensor for detecting DBP, comprising the following steps:

[0045] Preparation of S1.1 PEDOT-PG nanocomposite materials

[0046] 1 ml (1 mol L⁻¹) of FeCl₃ was added as an oxidant to 1 ml (2.0 mg / ml) of FeCl₃. -1 After sonicating the PG dispersion for 10 minutes, the above solution was slowly added to 2 ml of chloroform (containing 25 mg / mL of EDOT). -1 The resulting mixture was reacted at 50°C for 12 hours under static conditions, then centrifuged at 14,000 rpm for 30 minutes. The resulting precipitate was washed twice each with anhydrous ethanol and deionized water to generate PEDOT-PG nanocomposite material.

[0047] S1.2 Functionalization of the prepared PEDOT-PG with poly-L-lysine (PLL)

[0048] The PEDOT-PG nanocomposite material prepared above was dispersed in 1 ml of water. 300 μl of the dispersion was mixed with 200 μl of polylysine (PLL), and the mixture was shaken and incubated at room temperature for 2 hours. The mixture was then placed in a refrigerator to allow PLL and PEDOT-PG to react fully through non-covalent interaction, thus obtaining the PEDOT-PG-PLL nanocomposite material.

[0049] S2.1 Treatment and modification of the glassy carbon electrode surface

[0050] The treatment steps for the glassy carbon electrode included ultrasonically washing it sequentially with nitric acid, ethanol, and deionized water for 1 minute each, polishing it on chamois leather with 0.3 μm polishing powder (Al2O3), and then placing the electrode in 0.5 mmol L... -1 K3[Fe(CN)6] and 0.1 mol L -1 In a KCl mixed solution, a stable cyclic voltammetric peak was obtained by cyclic voltammetry scanning at a scan rate of 50 mV / s in the potential range of 0.2 V to 0.6 V, with a potential difference of less than 80, which proves that the electrode treatment is good.

[0051] S2.2 5 μl of PEDOT-PG-PLL nanocomposite material was drop-coated onto the surface of a bare glassy carbon electrode GCE to obtain a modified electrode PEDOT-PG-PLL / GCE, which was then dried by infrared at room temperature.

[0052] S3 placed the prepared PEDOT-PG-PLL / GCE in 0.01M PBS (pH=7.4) containing 2mM o-phenylenediamine and 0.8mM DBP, and electropolymerized it at 0.8V (scan rate 50mVs). 1The electrode was subjected to 12 cycles of cyclic voltammetry scanning within the range of electropolymerization. The electrode was then rinsed with deionized water and dried at room temperature to obtain the molecularly imprinted sensor MIPs / PEDOT-PG-PLL / GCE before elution.

[0053] S4. Immerse the above-obtained MIPs / PEDOT-PG-PLL / GCE in a methanol-acetic acid mixture (V... 甲醇 :V 20％乙酸 In a solution of 1:10, the sample was eluted at room temperature for 30 minutes to remove the template molecule DBP. The sample was then rinsed with deionized water to obtain the DBP detection molecular imprint sensor MIPs-e / PEDOT-PG-PLL / GCE.

[0054] S5 was characterized using cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS), such as... Figure 1-2 As shown.

[0055] Example 3

[0056] One embodiment of the present invention provides a method for using a molecularly imprinted sensor for detecting DBP, comprising: placing the DBP molecularly imprinted electrochemical sensor MIPs / PEDOT-PG-PLL / GCE (after elution) into a PBS solution (pH=5) containing different concentrations (1 fM-5 μM) of DBP, magnetically stirring for 15 min at room temperature, and rinsing the electrode with water; placing the molecularly imprinted sensor in an electrolytic cell, wherein the detection base solution of the electrolytic cell is 0.5 mmol L. -1 K3[Fe(CN)6] (redox probe) and 0.1 mol L -1 KCl (supporting electrolyte) mixed solution, scan parameters set (-0.2V-0.6V, scan rate 50mVs) -1 Differential pulse voltammetry (DPV) was used for detection, and the results are as follows: Figure 3 The display shows a linear correlation between the peak current difference of DPV measured by the molecularly imprinted sensor and the logarithm of the DBP concentration, as shown below. Figure 4 The linear equation is shown to be ΔI = 10.7275lgC. DBP +199.697, where the linear correlation coefficient is R. 2 =0.9932, the detection limit of this sensor is 0.88fM.

[0057] The selective detection of DBP analogs by the molecularly imprinted sensor yielded the following results: Figure 5 The results show that the sensor has a large current response to the experimental group containing DBP, indicating that the sensor has a specific and highly specific ability to identify DBP.

[0058] This is lower than any previously reported method, and the method is highly stable and reproducible, proving that the detection method established in this invention is practical and feasible, and can be successfully applied to the determination of DBP.

[0059] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from its spirit or essential characteristics. Therefore, the embodiments should be considered illustrative and non-limiting in all respects, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, all variations falling within the meaning and scope of equivalents of the claims are intended to be included within the present invention, and no reference numerals in the claims should be construed as limiting the scope of the claims.

Claims

1. A method for preparing an electrochemically imprinted sensor for detecting DBP, characterized in that, Includes the following steps: S1 uses a liquid-liquid interfacial polymerization method to prepare PEDOT-PG nanocomposites. The preparation steps include: With a volume of 1 ± 0.2 ml and a concentration of 1 mol L... -1 -3mol L -1 FeCl3 was added as an oxidant to a volume of 1 ± 0.2 ml and a concentration of 0.5 mg / ml. -1 -2.5mg.ml -1 After ultrasonic treatment, 2 ml of chloroform was slowly added to the above solution in the PG dispersion. The resulting mixture was reacted at 45-70°C for 8-12 hours under static conditions. After centrifugation, the precipitate was washed multiple times with anhydrous ethanol and deionized water to generate PEDOT-PG nanocomposite material. Then, the prepared PEDOT-PG was functionalized with polylysine to obtain the PEDOT-PG-PLL nanocomposite material. S2 used a drop-coating method to modify the PEDOT-PG-PLL nanocomposite material onto the surface of a bare glassy carbon electrode GCE, thereby obtaining a modified electrode PEDOT-PG-PLL / GCE; S3 uses DBP as a template molecule and O-PD as a polymer monomer to obtain molecularly imprinted sensors (MIPs) / PEDOT-PG-PLL / GCE on the surface of PEDOT-PG-PLL / GCE before elution by electropolymerization. S4 Immerse the uneluted molecularly imprinted sensors MIPs / PEDOT-PG-PLL / GCE obtained in step S3 in a methanol-acetic acid mixture V. 甲醇 :V 20%乙酸 In a 1:10 solution, elute for 15-40 minutes to remove the template molecule DBP, rinse with deionized water to obtain the DBP detection molecular imprint sensor MIPs-e / PEDOT-PG-PLL / GCE.

2. The method for preparing an electrochemically imprinted sensor for detecting DBP according to claim 1, characterized in that, The concentration of EDOT in the chloroform was 15 mg / mL. -1 -30mg.ml -1 .

3. The method for preparing an electrochemically imprinted sensor for detecting DBP according to claim 1, characterized in that, The method for functionalizing the prepared PEDOT-PG with polylysine in step S1 includes: dispersing the prepared PEDOT-PG nanocomposite material in 1 ml of water, mixing it with a PEDOT-PG aqueous solution:PLL volume ratio of 3:1-1:1 and incubating for 1-12 hours, and storing it under cold.

4. A method for preparing an electrochemically imprinted sensor for detecting DBP according to claim 1 or 3, characterized in that, The ultrasonic treatment is performed for 10-30 minutes, followed by centrifugation at 12000-16000 rpm for 20-40 minutes. The resulting precipitate is washed 2-3 times each with anhydrous ethanol and deionized water, and then stored in a refrigerator at 2-5°C.

5. The method for preparing an electrochemically imprinted sensor for detecting DBP according to claim 1, characterized in that, In step S2, 5µl of PEDOT-PG-PLL nanocomposite material is applied to the surface of a bare glassy carbon electrode GCE using a drop-coating method to obtain a modified electrode PEDOT-PG-PLL / GCE, which is then dried using infrared spectroscopy.

6. The method for preparing an electrochemically imprinted sensor for detecting DBP according to claim 5, characterized in that, The bare glassy carbon electrode GCE treatment steps include ultrasonically washing the glassy carbon electrode sequentially with nitric acid, ethanol, and deionized water for 1 minute each, polishing it on chamois leather with 0.3 µm Al2O3 polishing powder, and then placing the electrode in 0.5 mmol L... -1 K3 [Fe(CN)6] and 0.1 mol L -1 In a KCl mixed solution, a stable cyclic voltammetric peak was obtained by cyclic voltammetry scanning at a scan rate of 50 mV / s in the potential range of -0.2 V to 0.6 V, with a potential difference of less than 80.

7. The method for preparing an electrochemically imprinted sensor for detecting DBP according to claim 1, characterized in that, In step S3, PEDOT-PG-PLL / GCE is placed in 0.01M PBS containing 1-5mM o-phenylenediamine and 0.5-3mM DBP, with the pH of the PBS being 7.

4. Electropolymerization is performed by 8-20 cycles of cyclic voltammetry scans in the range of 0-0.8V and a scan rate of 50mVs⁻¹. The electropolymerized electrode is then rinsed with deionized water and dried at room temperature to obtain the molecularly imprinted sensor MIPs / PEDOT-PG-PLL / GCE before elution.

8. A method of using an electrochemically imprinted sensor for detecting DBP, characterized in that, The molecularly imprinted sensor was placed in a PBS solution containing different concentrations of DBP at pH 2.5-6.5 and stirred for 10-30 minutes at room temperature for adsorption. The electrode was then rinsed with water. The molecularly imprinted sensor was placed in an electrolytic cell containing a potassium ferricyanide solution of KCl, and the scanning parameters were set for differential pulse voltammetry detection.

Images