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Malathion hapten and preparation method thereof, and malathion artificial antigen

A malathion and artificial antigen technology, applied in the field of food analysis, can solve the problems that hinder the development of immunological detection methods for malathion, complicated synthesis steps, and unreasonable hapten structure

Inactive Publication Date: 2020-04-10
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the methods of malathion haptens, artificial antigens, antibodies and immunoassays based on them are immature. The existing haptens have unreasonable structures, complicated synthesis steps, and low affinity and specificity of antibodies, which seriously hinder the development of malathion. Development of Immunological Detection Method for Phosphorus

Method used

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  • Malathion hapten and preparation method thereof, and malathion artificial antigen
  • Malathion hapten and preparation method thereof, and malathion artificial antigen
  • Malathion hapten and preparation method thereof, and malathion artificial antigen

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0099] The preparation of embodiment 1 malathion hapten MLH (n=5)

[0100] 1. Experimental steps

[0101] The preparation of the malathion hapten MLH (n=5), the synthetic route is as follows:

[0102]

[0103] Specific steps include:

[0104] 1.1 take 1g P 2 S 5 Put it into the reaction bottle, add 1.2mL of toluene, stir, heat up to 45°C, start to add 1mL of methanol dropwise at 0.2mL / min, slowly drop into the reaction bottle through the dropper, keep at 55±5°C, react for 2h, When cooling to below 40°C, take out the colorless crude sulfide with pungent smell;

[0105] 1.2 Take 0.948g of the crude sulfide described in step 1.1, add 6mmoL of ethyl hydrogen maleate, and after the addition, keep the reaction at 55°C for 1h, and keep it at 70-75°C for 8h to obtain the intermediate product b;

[0106] 1.3 Take 0.302mg (0.83mmoL) of the intermediate product b described in step 1.2, dissolve it in 0.2mL DMF solution, add 1mmoL DCC and NHS solution respectively, react at 4°C fo...

Embodiment 2

[0113] The preparation of embodiment 2 malathion artificial antigen MLH-BSA

[0114] 1. Experimental steps

[0115] The preparation of malathion artificial antigen, concrete steps comprise:

[0116] 1.1 Dissolve 2mmoL of the malathion hapten (III) prepared in Example 1 in 200 μL of DMF, add 3mmoL of DCC and 3mmoL of NHS, and activate by shaking at 4°C for 12 hours in the dark to obtain an activated malathion derivative ;

[0117] 1.2 Centrifuge the malathion derivative described in step 1.1 at 12,000 rpm for 10 min, collect the supernatant, and obtain the malathion hapten derivative. Prepare a solution containing 5 mg / mL carrier protein BSA with a PBS buffer solution with a pH of 7.4;

[0118] 1.3 Add the hapten derivatives described in step 1.2 dropwise to the PBS buffer solution containing the carrier protein, so that the molar ratio of the malathion hapten to the carrier protein is 400:1, shake and react at 4°C for 10 h, That is to obtain the malathion artificial antige...

Embodiment 3

[0123] Example 3 Preparation of Malathion Polyclonal Antibody

[0124] 1. Experimental steps

[0125] Preparation of malathion polyclonal antibody, specific steps include:

[0126] 1.1 Mix and emulsify the prepared malathion artificial antigen MLH-BSA with an equal amount of complete adjuvant as an immunogen, select mice as experimental animals, and inject subcutaneously into the abdomen, back, and soles of each mouse at a rate of 50 μg ( Calculated by protein content) dose to immunize 6-week-old mice for initial immunization;

[0127] 1.2 The second immunization: Three weeks later, replace the complete adjuvant with incomplete adjuvant, and boost the immunization once, and the operation method is the same as step 1.1;

[0128] 1.3 The third booster immunization: booster immunization once every two weeks, the operation method is the same as step 1.2;

[0129] 1.4 Blood collection: blood was taken from the tail, and after standing at 37°C for 30 minutes, it was centrifuged a...

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Abstract

The invention discloses a malathion hapten and a preparation method thereof, and a malathion artificial antigen, wherein the structure of the malathion hapten is represented by a formula (I), and n isequal to 1-9. According to the invention, malathion hapten is synthesized by taking phosphorus pentasulfide as a starting raw material, then malathion artificial antigen is prepared, and the preparedartificial immunogen is used for immunizing animals, so that a specific antibody for malathion can be induced to be generated. According to the invention, the method has the advantages of simple andconvenient synthesis steps, readily available raw materials, mild reaction conditions, suitability for laboratory and factory production, wide application prospect and great popularization value, canbe applied to establishment of rapid malathion detection methods and development of immune rapid test kits, and meets the detection requirements of malathion residues.

Description

technical field [0001] The invention belongs to the technical field of food analysis. More specifically, it relates to a malathion hapten, an artificial antigen and a preparation method thereof. Background technique [0002] Malathion, also known as Marathon, 4049, is a colorless or yellow oily liquid with garlic and pungent odor. The molecular weight is 330, easily soluble in organic solvents such as alcohols, acetone, esters and aromatic hydrocarbons, slightly soluble in water and petroleum ether. As a broad-spectrum new organophosphorus insecticide, malathion has the characteristics of low toxicity and high efficiency, and is an ideal pesticide to replace highly toxic varieties. At present, it is widely used in agriculture and stored grain production, but its residues in the environment and food pose a potential threat to people's health to a certain extent. In some fruits, vegetables, and grains, malathion is one of the pesticides with a relatively high detection amou...

Claims

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Application Information

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IPC IPC(8): C07F9/17C07K14/765C07K14/77C07K14/795C07K16/44G01N33/53
CPCC07F9/17C07K14/765C07K14/77C07K14/795C07K16/44G01N33/53G01N33/184
Inventor 肖治理柳心梅田巍孙远明徐振林沈玉栋谢波
Owner SOUTH CHINA AGRI UNIV
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