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CN-121994891-A - Vomitoxin rapid electrochemical detection method based on enzymatic electropolymerization signals

CN121994891ACN 121994891 ACN121994891 ACN 121994891ACN-121994891-A

Abstract

The application discloses a quick electrochemical detection method of vomitoxin based on an enzymatic electropolymerization signal, which comprises the steps of mixing a sample solution to be detected with enzyme-labeled vomitoxin with a preset concentration to form a competition reaction system, placing a working electrode in the competition reaction system for incubation to obtain a target working electrode, removing unbound substances on the target working electrode, placing the target working electrode in a detection solution containing electropolymerization monomers and enzyme-labeled substrates to form a polymer film, measuring electrochemical signal changes of the polymer film before and after formation by adopting an electrochemical method, and quantitatively detecting the vomitoxin concentration in the sample solution to be detected based on the electrochemical signal changes. The method skillfully integrates the advantages of high selectivity of molecular imprinting, signal amplification by enzyme catalysis and signal conversion by in-situ electropolymerization, does not need to depend on biological antibodies, and has the remarkable advantages of high sensitivity, good stability, simple and convenient operation, rapid detection and the like.

Inventors

  • Wang Jiarou
  • LI YUNPAN
  • LI JIANGYU

Assignees

  • 惠州市鼎硕实业有限公司

Dates

Publication Date
20260508
Application Date
20260228

Claims (10)

  1. 1. A method for rapid electrochemical detection of vomitoxin based on an enzymatic electropolymerization signal, comprising: S1, mixing a sample solution to be detected with enzyme-labeled vomitoxin with a preset concentration to form a competition reaction system, wherein enzyme in the enzyme-labeled vomitoxin can catalyze an enzyme-labeled substrate to generate an active substance for initiating monomers to carry out electropolymerization; S2, placing a working electrode in the competition reaction system for incubation, so that free vomitoxin in the sample solution to be detected and the enzyme-labeled vomitoxin compete for binding with a specific recognition site on the working electrode to obtain a target working electrode, wherein the working electrode is modified by using vomitoxin molecularly imprinted polymers; S3, removing unbound substances on the target working electrode, and then placing the target working electrode in detection liquid containing an electropolymerized monomer and an enzyme-labeled substrate, so that the enzyme-labeled vomitoxin on the target working electrode catalyzes the enzyme-labeled substrate to produce the active substances, and the active substances trigger the electropolymerized monomer to generate in-situ electropolymerization on the surface of the target working electrode to form a polymer film; and S4, measuring electrochemical signal changes of the polymer film before and after formation by adopting an electrochemical method, and quantitatively detecting the concentration of free vomitoxin in the sample solution to be detected based on the electrochemical signal changes.
  2. 2. The method for rapid electrochemical detection of vomitoxin based on enzymatic electropolymerization signals as recited in claim 1, wherein the sample solution to be tested in S1 is prepared based on the steps of: crushing a grain sample to be detected into sample powder; And adding a methanol aqueous solution or ethyl acetate into the sample powder, carrying out vortex mixing, sequentially carrying out vibration extraction for 3 min-5 min, carrying out centrifugal treatment for 1 min-3 min at 2000 rpm-4000 rpm, and collecting supernatant to obtain a sample solution to be detected.
  3. 3. The rapid electrochemical detection method of vomitoxin based on enzymatic electropolymerization signals of claim 1, wherein the enzyme-labeled vomitoxin in S1 is a glucose oxidase-labeled vomitoxin, the enzyme-labeled substrate is glucose, and the active substance is hydrogen peroxide.
  4. 4. The method for rapid electrochemical detection of vomitoxin based on an enzymatic electropolymerization signal of claim 3, wherein the predetermined concentration of the enzyme-labeled vomitoxin in S1 is 0.1 μg/mL to 5 μg/mL.
  5. 5. The method for rapid electrochemical detection of vomitoxin based on an enzymatic electropolymerization signal of claim 3, wherein the electropolymerization monomer in S3 is o-phenylenediamine, a phenylenediamine derivative, or pyrrole.
  6. 6. The method for rapid electrochemical detection of vomitoxin based on enzymatic electropolymerization signals of claim 1, wherein the working electrode in S2 is prepared based on the steps of: forming a molecularly imprinted polymer film on the surface of the electrode modified by the bare electrode or the nano material by taking vomitoxin as a template molecule and a functional monomer in an electropolymerization mode; eluting and removing the template molecules to form a cavity with specific recognition capability for vomitoxin, thereby obtaining the working electrode.
  7. 7. The rapid electrochemical detection method of vomitoxin based on an enzymatic electropolymerization signal of claim 6, wherein the functional monomer is one or more of arginine, acrylamide, o-phenylenediamine, dopamine, and pyrrole, and the nanomaterial is one or more of carboxylated carbon nanotubes, graphene oxide, and gold nanoparticles.
  8. 8. The method for rapidly detecting vomitoxin based on the enzymatic electropolymerization signal according to claim 1, wherein the incubation time in S2 is 5min to 20min, and the polymer film formation time in S3 is 1min to 10min.
  9. 9. The method for rapid electrochemical detection of vomitoxin based on an enzymatic electropolymerization signal of claim 1, wherein the electrochemical method in S4 is square wave voltammetry, differential pulse voltammetry, or alternating current impedance, and the electrochemical signal change is a decrease in peak current or an increase in electron transfer impedance.
  10. 10. The method for rapid electrochemical detection of vomitoxin based on an enzymatic electropolymerization signal of claim 1, wherein the step of quantitatively detecting the concentration of free vomitoxin in the sample solution to be tested based on the electrochemical signal change in S4 comprises: Determining the number of enzyme-labeled vomitoxins corresponding to the electrochemical signal change according to a preset corresponding relation between the electrochemical signal change and the number of enzyme-labeled vomitoxins, wherein the number of enzyme-labeled vomitoxins is the number of enzyme-labeled vomitoxins combined with a specific recognition site on a working electrode; determining the concentration of the free vomitoxin in the sample solution to be detected based on the linear relation between the number of enzyme-labeled vomitoxins and the concentration of the free vomitoxin, wherein the expression of the linear relation is as follows: ; To determine the free vomitoxin concentration in the sample solution, In order to compete for the concentration of the enzyme-labeled vomitoxin in the reaction system, For the total number of specific recognition sites on the working electrode, The amount of enzyme-labeled vomitoxin that binds to a specific recognition site on the working electrode.

Description

Vomitoxin rapid electrochemical detection method based on enzymatic electropolymerization signals Technical Field The application relates to the technical field of detection, in particular to a quick electrochemical detection method for vomitoxin based on an enzymatic electropolymerization signal. Background Vomitoxin (DON), also known as deoxynivalenol, is a common mycotoxin in cereals and their products, severely threatening food safety and human health. At present, the detection method of vomitoxin mainly comprises high performance liquid chromatography, liquid chromatography-mass spectrometry and enzyme-linked immunosorbent assay. The chromatographic method has the defects of high accuracy, expensive instrument, complex pretreatment, complex operation, long time consumption and the like, and is difficult to meet the requirement of on-site rapid screening. Although the immunological method is relatively simple to operate, the core of the immunological method depends on biological antibodies. The biological antibody has the inherent defects of long preparation period, high cost, large batch-to-batch difference, harsh storage conditions (cold chain transportation is required), easy inactivation in non-aqueous phase or organic solvent and the like, and the wide application of the biological antibody in complex food matrixes is limited. Disclosure of Invention The application provides a rapid electrochemical detection method for vomitoxin based on an enzymatic electropolymerization signal, which aims to solve the technical problem that an immunization method depends on a biological antibody. In a first aspect, the application provides a method for rapid electrochemical detection of vomitoxin based on an enzymatic electropolymerization signal, comprising: S1, mixing a sample solution to be detected with enzyme-labeled vomitoxin with a preset concentration to form a competition reaction system, wherein enzyme in the enzyme-labeled vomitoxin can catalyze an enzyme-labeled substrate to generate an active substance for initiating monomers to carry out electropolymerization; S2, placing a working electrode in the competition reaction system for incubation, so that free vomitoxin in the sample solution to be detected and the enzyme-labeled vomitoxin compete for binding with a specific recognition site on the working electrode to obtain a target working electrode, wherein the working electrode is modified by using vomitoxin molecularly imprinted polymers; S3, removing unbound substances on the target working electrode, and then placing the target working electrode in detection liquid containing an electropolymerized monomer and an enzyme-labeled substrate, so that the enzyme-labeled vomitoxin on the target working electrode catalyzes the enzyme-labeled substrate to produce the active substances, and the active substances trigger the electropolymerized monomer to generate in-situ electropolymerization on the surface of the target working electrode to form a polymer film; s4, measuring electrochemical signal changes of the polymer film before and after formation by adopting an electrochemical method, and quantitatively detecting the vomitoxin concentration in the sample solution to be detected based on the electrochemical signal changes. In some embodiments, the sample solution to be tested in S1 is prepared based on the following steps: crushing a grain sample to be detected into sample powder; And adding a methanol aqueous solution or ethyl acetate into the sample powder, carrying out vortex mixing, sequentially carrying out vibration extraction for 3 min-5 min, carrying out centrifugal treatment for 1 min-3 min at 2000 rpm-4000 rpm, and collecting supernatant to obtain a sample solution to be detected. In some embodiments, the enzyme-labeled vomitoxin in S1 is a glucose oxidase-labeled vomitoxin, the enzyme-labeled substrate is glucose, and the active substance is hydrogen peroxide. In some embodiments, the predetermined concentration of the enzyme-labeled vomitoxin in S1 is 0.1 μg/mL to 5 μg/mL. In some of these embodiments, the electropolymerized monomer in S3 is o-phenylenediamine, a phenylenediamine derivative, or pyrrole. In some embodiments, the working electrode in S2 is prepared based on the following steps: forming a molecularly imprinted polymer film on the surface of the electrode modified by the bare electrode or the nano material by taking vomitoxin as a template molecule and a functional monomer in an electropolymerization mode; eluting and removing the template molecules to form a cavity with specific recognition capability for vomitoxin, thereby obtaining the working electrode. In some embodiments, the functional monomer is one or more of arginine, acrylamide, o-phenylenediamine, dopamine, and pyrrole, and the nanomaterial is one or more of carboxylated carbon nanotubes, graphene oxide, and gold nanoparticles. In some embodiments, the incubation time in S2 is 5min to 20min, and the polymer film form