Arborescence half antigen capable of directly detecting furaltadone metabolite AMOZ, arborescence antigen and application of arborescence half antigen

A technology of furaltadone and hapten, which is applied in the field of dendritic hapten to achieve the effects of high sensitivity, enhanced immunogenicity and strong specificity

Active Publication Date: 2014-07-30
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The technical problem to be solved by the present invention is to overcome the defect that the traditional AMOZ immunoassay technology needs to be derivatized, and to provide a dendritic hapten, artificial dendritic antigen

Method used

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  • Arborescence half antigen capable of directly detecting furaltadone metabolite AMOZ, arborescence antigen and application of arborescence half antigen
  • Arborescence half antigen capable of directly detecting furaltadone metabolite AMOZ, arborescence antigen and application of arborescence half antigen
  • Arborescence half antigen capable of directly detecting furaltadone metabolite AMOZ, arborescence antigen and application of arborescence half antigen

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] The preparation of embodiment 1 AMOZ glyoxylic acid derivative I:

[0046] Take 604mg (3mmol) of AMOZ and 266.5mg (3.6mmol) of glyoxylic acid and dissolve them in 2mL of absolute ethanol respectively, add the ethanol solution of glyoxylic acid to the ethanol solution of AMOZ dropwise during the stirring process, and stir at room temperature overnight. After the reaction was completed, vacuum filtration was performed, and the filter cake was washed twice with 2 mL of ethanol and diethyl ether, and the precipitate was collected to obtain 540.2 mg of the target product as a white solid with a yield of 70%. ESI-MS analysis (positive) m / z 258.7 [M+H] + ; 1 H NMR (600MHz, d5-Pyridine, TMS): δ 7.56 (s, 1H), 4.99-4.97 (m, 1H), 4.16 (t, J=8.8Hz, 1H), 3.79 (dd, J1=6.7, J2 =8.8, 1H), 3.68-3.64 (m, 4H), 2.66 (dd, J1=6.1, J2=13.5, 1H), 2.62 (dd, J1=5.5, J2=13.4, 1H), 2.51-2.46 (m , 4H). The structural formula of AMOZ glyoxylic acid derivative I is shown in formula (I).

Embodiment 2

[0047] The preparation method of embodiment 2 four-branched polyamide-amine II

[0048] Ethylenediamine and methyl acrylate are used as raw materials, and the reaction follows Michael addition and amidation condensation reactions. With ethylenediamine as the core, one terminal amino group is protected by tert-butoxycarbonyl (Boc), and Michael addition reaction is carried out with methyl acrylate to obtain Boc-Eddp-OCH 3 ; One terminal amino group of ethylenediamine is protected with fluorenylmethoxycarbonyl (Fmoc), and Boc-Eddp-OCH3 is subjected to amidation condensation reaction with excess Fmoc-protected ethylenediamine to obtain Boc-Eddp-(Fmoc-Ethylenediamine) 2 , go to Boc protection to get H-Eddp-(Fmoc- Ethylenediamine) 2 ; H-Eddp-(Fmoc- Ethylenediamine) 2 Further with Boc-Eddp-OCH 3 Carry out amidation condensation reaction, obtain Boc-Eddp-(Eddp) 2 -(Fmoc-Ethylenediamine) 4 , go to Fmoc protection to get Boc-Eddp-(Eddp) 2 -(Ethylenediamine) 4 , that is, the f...

Embodiment 3

[0049] Example 3 Preparation method of AMOZ dendritic hapten HⅢ

[0050] The four-branched polyamide-amine prepared in Example 2 and the glyoxylic acid derivative of AMOZ prepared in Excessive Example 1 were subjected to amidation condensation reaction to obtain Boc-Eddp-(Eddp) 2 -(AMOZ-2-iminoacetyl-Ethylenediamine) 4 , remove the Boc amino protecting group to obtain Eddp-(Eddp) 2 -(AMOZ-2-iminoacetyl-Ethylenediamine) 4 , the target AMOZ dendritic hapten HⅢ. The structure was identified by ESI-MS, and the purity was determined by HPLC. The structural formula of AMOZ dendritic hapten HIII is shown in formula (III).

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Abstract

The invention discloses an arborescence half antigen capable of directly detecting furaltadone metabolite AMOZ, an arborescence antigen and application of the arborescence half antigen. The arborescence half antigen is obtained by coupling glyoxylic acid derivative of AMOZ to an active amino terminal of four-branch polyamide-amine, an artificial antigen is prepared based on the arborescence half antigen, and then an antibody is prepared by virtue of the artificial antigen; By adopting the artificial antigen disclosed by the invention, a high-quality antibody directly aiming at AMOZ can be obtained, valence of the antibody is 1:64000, the lowest detection limit of a chemiluminiscence enzyme linked immunosorbent assay (CLEIA) analytical method established based on the antibody is 0.112ng / ml, 50% inhibiting concentration is 15.26ng / ml, the CLEIA analytical method can be directly applied to detection of AMOZ, and the defects of the traditional scheme that an antibody prepared by virtue of an AMOZ half antigen and the artificial antigen is poor in antibody specificity and can not directly identify the AMOZ. The arborescence half antigen, the artificial antigen and the antibody have a wide application prospect in detection of AMOZ residue in food, and a preparation thinking of the half antigen can provide a reference for design of half antigens of other similar small molecule compounds.

Description

technical field [0001] The invention belongs to the technical field of food safety immunoassay, and more specifically relates to a dendritic hapten and a dendritic antigen for directly detecting furaltadone metabolite AMOZ and applications thereof. Background technique [0002] Nitrofuran antibiotics have the advantages of strong bactericidal ability, broad antibacterial spectrum, and low price, and are widely used in clinical and animal breeding industries. In recent years, studies have shown that nitrofuran drugs and their metabolites are very toxic to humans. The European Union banned the use of nitrofuran antibiotics in food animals in 1995, and in 1997 all nitrofuran antibiotics were banned. Listed as prohibited drugs, my country also issued a ban on the use of nitrofuran antibiotics in 2002. Due to the rapid metabolism of nitrofuran drugs, they can be degraded within a few hours in the body, and the metabolites can combine with cell membrane proteins to form stable con...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D263/26C07K14/765C07K14/795C07K14/77G01N33/544
CPCC07D263/26C07K14/765C07K14/77C07K14/795C07K19/00G01N33/53G01N33/531
Inventor 徐振林沈玉栋孙远明刘凤银肖治理杨金易雷红涛王弘
Owner SOUTH CHINA AGRI UNIV
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