Preparation of 1-amino hydantoin derivative hapten, antigen and antibody

A technology of aminohydantoin and its derivatives, applied in the field of immunochemistry, can solve the problems that there is no 1-aminohydantoin hapten, antigen and antibody, 1-aminohydantoin ELISA detection method has not been established, etc., and achieve good specificity , high potency effect

Inactive Publication Date: 2009-04-22
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Theoretically, the preparation of small molecule hapten antibodies of all original drugs can be realized, but it also requires specific and specific technical solutions to realize the preparation of specific antibodies according to the target drug. At present, there is no 1-amino hapten antibody that can be used for ELISA detection. Due to the related reports of haptens, antigens and antibodies, the ELISA detection method for 1-aminohydantoin has not yet been established

Method used

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  • Preparation of 1-amino hydantoin derivative hapten, antigen and antibody
  • Preparation of 1-amino hydantoin derivative hapten, antigen and antibody
  • Preparation of 1-amino hydantoin derivative hapten, antigen and antibody

Examples

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Effect test

Embodiment 1

[0034] Example 1 Preparation method of 1-aminohydantoin derivative hapten MCPSEM

[0035] Add 1.15g of m-carboxybenzaldehyde into a 50ml round bottom flask, slowly add methanol until the p-carboxybenzaldehyde is completely dissolved, add 0.3g of semicarbazide during stirring, and stir overnight at room temperature; after the reaction is completed, filter, wash twice with water, and wash twice with methanol Repeatedly, dry, obtain 0.62g white powder, its structure is as shown in formula (I). The APCI-MS molecular ion peak of hapten MCPSEM is 207, HNMR (600MHz, d 6 -DMSO, TMS): δ 10.35(s, 1H); 8.14(s, 1H); 8.03(d, J=7.8Hz, 1H); 7.93(s, 1H); 7.90(d, J=7.8Hz, 1H ); 7.52 (t, J=7.2Hz, 1H,). The above spectral data can be assigned correctly, which is consistent with the structure of MCPSEM, indicating that the hapten MCPSEM was synthesized successfully.

[0036]

Embodiment 2

[0037] Example 2 Preparation method of 1-aminohydantoin derivative hapten CEPSEM

[0038] Add 1.06g of 2-(4-formylphenoxy)acetic acid into the round bottom flask, slowly add methanol until the 2-(4-formylphenoxy)acetic acid is completely dissolved, add 0.4g semicarbazide during stirring, and stir at room temperature After overnight, the reaction was completed, filtered, washed twice with water and twice with methanol, and dried to obtain 0.90 g of white powder, the structural formula of which was shown in formula (II). CEPSEM spectral data: APCI-MSm / z: 237[M+H]+; 1H NMR (600MHz, d6-DMSO, TMS): δ 10.09(s, 1H); 7.78(s, 1H); 7.64(d, J = 9.0 Hz, 2H); 6.91 (d, J = 9.0 Hz, 2H); 4.71 (s, 2H). The above spectral data can be assigned correctly, which is consistent with the CEPSEM structure, indicating that the hapten CEPSEM was synthesized successfully.

[0039]

Embodiment 3

[0040] Example 3 Preparation method of immunogen CEPSEM-BSA

[0041] Take the hapten CEPSEM 0.1mmol and dissolve it in 2ml DMF, stir and add DCC 27.5mg and NHS14.4mg. Magnetically stir the reaction overnight at 4°C. After centrifugation, the supernatant night is liquid A; weigh 140 mg of BSA and dissolve it in 10 ml of PBS (pH 8.0) with a concentration of 0.1 mol / L, add 1 ml of DMF, stir and dissolve to prepare liquid B; under magnetic stirring , solution A was gradually dropped into solution B, and reacted at 4°C for 12 hours. After centrifugation, the supernatant was taken and dialyzed with normal saline at 4°C for 3 days, and the dialysate was changed 3 times a day. The obtained whole antigen was dispensed into 0.5ml centrifuge tubes at a concentration of 1 mg / ml, and frozen in a -20°C refrigerator for immunization.

[0042] The prepared semicarbazide derivative immunogen has a structure shown in formula (V):

[0043]

[0044] Immunogen identification: The carrier prot...

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Abstract

The invention discloses a method for preparing a hapten, an antigen and an antibody of a 1-amino-hydantoin derivative, wherein the hapten of a semicarbazide derivative is obtained after the aldehyde condensation between semicarbazide and carboxyl benzaldehyde ammonia, and the structure of the hapten is similar to that of the semicarbazide derivative, thus the antigenic determinant with the molecular specificity of a veterinary drug is highlighted, and the antigen is prepared from the coupled carrier protein by an active ester method. The antigen is used to immunize experimental animals to obtain the antibody which aims at the semicarbazide derivative and has high titer and good specificity. The antibody can be applied to the fast detection of semicarbazide residual quantity in animal derived food, and the lower limit of the detection can reach 0.09 nanogram per milliliter.

Description

technical field [0001] The invention belongs to the technical field of immunochemistry, and specifically relates to a method for preparing a hapten, an antigen and an antibody of a 1-aminohydantoin derivative, and an application of the prepared hapten, antigen, antibody and antibody. Background technique [0002] Nitrofurantoin is a broad-spectrum antibacterial drug, which has certain antibacterial effects on Gram-positive and negative bacteria, and has growth-promoting effects on meat-derived animals such as pigs and fish. Long-term studies have found that nitrofurantoin and its metabolites have carcinogenic effects, so many countries have banned their use in the breeding of meat-derived animals. Since 1995, the European Union has comprehensively banned the use of nitrofurantoin, and the detection limit of furan residues in food of animal origin is non-detectable. my country also stipulates that nitrofurantoin is listed as a banned drug in animal breeding. However, becaus...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D233/80C07K14/765C07K16/44C07K16/06G01N33/53
Inventor 沈玉栋张世伟孙远明雷红涛王弘肖治理杨金易蔡肇婷
Owner SOUTH CHINA AGRI UNIV
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