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Prochloraz hapten, artificial antigen and antibody and preparation method and application thereof

An artificial antigen, prochloraz technology, applied in the preparation method of peptides, chemical instruments and methods, animal/human protein, etc., to achieve the effect of high purity and yield, high sensitivity, and low cross-reaction rate

Active Publication Date: 2022-07-08
BEIJING KWINBON BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, there is no relevant report on the prochloraz hapten in China

Method used

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  • Prochloraz hapten, artificial antigen and antibody and preparation method and application thereof
  • Prochloraz hapten, artificial antigen and antibody and preparation method and application thereof
  • Prochloraz hapten, artificial antigen and antibody and preparation method and application thereof

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preparation example Construction

[0034] In the second aspect, the present invention provides the preparation method of the above-mentioned prochloraz hapten, which comprises the following steps:

[0035] 1) 3-(3-hydroxy-2,4,6-trichloro)-propionic acid reacts with 1,2-dichloroethane to obtain Intermediate 1, which has the structural formula

[0036] 2) the intermediate 1 is reacted with n-propylamine to obtain the intermediate 2, and the intermediate 2 has the structural formula

[0037]

[0038] 3) The intermediate 2 is reacted with bis(trichloromethyl)carbonate and imidazole to obtain the prochloraz hapten.

[0039] Preferably, the step 1) includes the following steps: take 2.69 g of 3-(3-hydroxy-2,4,6-trichloro)-propionic acid, add 60 mL of 1,2-dichloroethane to dissolve, add 0.57 g of KOH , react at 60°C for 6 h, stop the reaction, add water to wash, and evaporate to dryness to obtain intermediate 1.

[0040] Preferably, the step 2) includes the following steps: adding 40 mL of n-propylamine, 0.31 g...

Embodiment 1

[0060] A kind of preparation method of prochloraz hapten, it comprises the steps:

[0061] 1) Take 2.69 g of 3-(3-hydroxy-2,4,6-trichloro)-propionic acid, add 60 mL of 1,2-dichloroethane to dissolve, add 0.57 g of KOH, react at 60°C for 6 h, stop the reaction, add water Washed and evaporated to dryness to obtain Intermediate 1;

[0062] 2) Add n-propylamine 40mL, anhydrous sodium iodide 0.31g, potassium carbonate 1.38g to intermediate 1, heat under reflux for 4h, stop the reaction, rotary steam, remove n-propylamine, add water and shake, extract with ethyl acetate, collect The organic phase was recrystallized with n-hexane-dichloromethane to obtain Intermediate 2;

[0063] 3) Dissolve 1.2 g of intermediate 2 in 80 mL of dichloromethane, add 1.1 g of bis(trichloromethyl)carbonate and 0.3 mL of triethylamine, stir at room temperature for 3 h, add 0.27 g of imidazole, continue to stir for 7 h to stop the reaction, Add water, extract and wash, evaporate to dryness to obtain a re...

Embodiment 2

[0065] A preparation method of prochloraz artificial antigen, the steps are as follows:

[0066] Dissolve 17 mg of the prochloraz hapten prepared in Example 1 in 1 mL of DMF, dissolve and clarify, add 15.3 mg of carbodiimide and 11.2 mg of N-hydroxysuccinimide, mix well, and react for 4 hours to obtain solution A ; Dissolve 50 mg of bovine serum albumin (BSA) in 0.05 mol / L PB buffer to obtain B solution; add A dropwise to B solution, react at 4°C for 24 h, and dialysis and purify with 0.02 mol / L PBS for 3 days , the medium was changed three times a day to obtain the prochloraz artificial antigen conjugated with bovine serum albumin, which was subpackaged and stored at -20°C.

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Abstract

The invention discloses a prochloraz hapten, an artificial antigen and an antibody as well as a preparation method and application thereof. The prochloraz hapten provided by the invention not only retains the characteristic structure of the prochloraz hapten to the greatest extent, and makes the immunization of the prochloraz hapten The originality is obviously enhanced, and it has a carboxyl group that can be coupled with the carrier protein; the prochloraz immunization antigen obtained by coupling the prochloraz hapten with the carrier protein is used to immunize animals, which is more conducive to stimulating the animal's immune response to produce specificity Stronger and more sensitive antibody, the sensitivity of prochloraz antibody can reach 0.1μg / L, and the cross-reaction rate with other pesticides is low, which provides a basis for the subsequent establishment of various immunoassay methods for prochloraz.

Description

technical field [0001] The invention belongs to the field of food safety detection. More specifically, the present invention relates to prochloraz haptens, artificial antigens and antibodies, and preparation methods and applications thereof. Background technique [0002] Prochloraz (Prochloraz) is a broad-spectrum fungicide that can prevent and control various diseases on field crops, fruits and vegetables, turf and ornamental plants; in addition, Prochloraz can also be used for citrus, bananas and other fruits after harvest. Product preservation has a wide range of applications. There are strict regulations on the dosage of prochloraz in various countries. For example, CAC, the European Union and Japan all stipulate the MRL value of prochloraz in raw milk, which are 0.05, 0.4, and 0.05 mg / kg, respectively. my country's national standard GB 2763-2016 The "National Food Safety Standard for Maximum Residue Limits of Pesticides in Foods" also stipulates the residue limits of p...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07D233/56C07K14/765C07K14/77C07K14/795C07K1/113C07K16/44G01N33/53G01N33/531G01N33/577
CPCC07D233/56C07K14/765C07K14/77C07K14/795C07K16/44G01N33/53G01N33/531G01N33/577
Inventor 万宇平崔廷婷崔海峰张瑜何方洋马玉华崔娜赵正苗
Owner BEIJING KWINBON BIOTECH
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