Method for portably and rapidly detecting ochratoxin A

An ochratoxin, rapid technique for use in analytical chemistry

An ochratoxin, rapid technique for use in analytical chemistry

CN103808716AActive Publication Date: 2014-05-21INST OF AGRI PROD QUALITY SAFETY & STANDARD JIANGXI ACAD OF AGRI SCI
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  • Method for portably and rapidly detecting ochratoxin A
  • Method for portably and rapidly detecting ochratoxin A
  • Method for portably and rapidly detecting ochratoxin A

Examples

Experimental program
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Effect test

Embodiment 1

[0032] A portable method for rapidly detecting ochratoxin A, the specific steps are as follows:

[0033]1. Prepare sucrose invertase-terminal alkyne-modified DNA: First, mix 30 μL of 0.3 mM thiol-terminal alkyne-modified DNA (thiol-terminal alkyne-modified DNA sequence from 5' to 3' end is: 5'-alkynyl- AAA AAA AAA AAA- thiol-3'), 2.0 μL 30 mM tris-(2-formylethyl)phosphine (TCEP) and 2.0 μL 1.0 M PBS buffer solution (a phosphate buffer solution prepared by sodium monohydrogen phosphate and sodium dihydrogen phosphate, pH is 5.5), mixed evenly, and incubated at room temperature for 1.0 hour to activate thiol-terminal alkyne modified DNA. After the end, an ultrafiltration centrifuge tube with a molecular weight of 10K was used to separate and wash 5 times. Next add 1.0 mg of sulfo-4-(N-maleimidomethyl)cyclohexane-1-carboxylate succinimidyl ester sodium salt (Sulfo-SMCC) to 400 μL of 20 mg / mL sucrose conversion In the enzyme solution, mix well and shake at room temperature for ...

Embodiment 2

[0039] The specificity of a portable rapid detection method for ochratoxin A, the specific steps are as follows:

[0040] Under the same experimental conditions as in Example 1, four other toxins were used instead of ochratoxin A to measure the blood glucose meter response values ​​of four other toxins respectively, and compared with the response values ​​of ochratoxin A, then The specificity of the method of the present invention was investigated. The four toxins are aflatoxin B 1 (AFB 1 ), aflatoxin M 1 (AFM 1 ), zearalenone (F-2), both at a concentration of 5.0 ng / mL.

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Abstract

The invention discloses a method for portably and rapidly detecting ochratoxin A. The method comprises the following steps: first, preparing sucrose invertase-terminal alkyne modified DNA; subsequently, combining biotin-nitrine modified DNA with the surface of a magnetic bead, adding the ochratoxin A with different concentration, and mixing; adding the previously-prepared sucrose invertase-terminal alkyne modified DNA, a CuSO4 solution and an ascorbic acid solution, wherein the ascorbic acid solution can be used for reducing Cu(II) to obtain a Cu(I) compound so as to catalyze the biotin-nitrine modified DNA and the sucrose invertase-terminal alkyne modified DNA to have a click chemistry reaction; taking a supernatant liquor after the reaction is ended, then adding the supernatant liquor into a sucrose solution, wherein the sucrose invertase can effectively catalyze the sucrose in the solution to be transformed into glucose; and finally, measuring by adopting a glucometer. The method is used for organically combining the rapid reaction advantage of the click chemistry reaction with the high specificity characteristic of an aptamer, and adopting the simple and portable glucometer to acquire data, thus the operation complexity and detection cost are greatly reduced, and the sensitivity and selectivity are improved.

Description

technical field [0001] The invention belongs to the field of analytical chemistry, and in particular relates to a portable and rapid detection method for ochratoxin A constructed based on a click chemical reaction. Background technique [0002] Ochratoxin A is one of the most common food contamination toxins, mainly produced by some Aspergillus and Penicillium fungi, which are easy to grow and multiply in cereal crops, fruits, beer and liquor. In animals, ochratoxin A is easy to be absorbed by the intestine, and it is easy to cause strong toxicity to the kidney and liver, and it also has carcinogenic, teratogenic, and immunosuppressive effects. The International Agency for Research on Cancer has listed ochratoxin A as a potential human carcinogen. The Food and Agriculture Organization of the United Nations / World Health Organization Joint Expert Committee on Food Additives (FAO / WHO JECFA) pointed out that 50% of ochratoxin A transmitted indirectly from food to humans comes f...

Claims

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Application Information

Patent Timeline
21 May 2014
Publication
CN103808716A
IPC
G01N21/78
Inventors
邱素艳; 罗林广