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l-Lauramide arginine hydrochloride ethanol ester artificial antigen, preparation method of specific antibody and use thereof

A technology of lauramide and arginine is applied in the field of immunochemical analysis to achieve the effects of avoiding organic solvents, facilitating on-site monitoring and low cost

Active Publication Date: 2022-07-19
NINGBO ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

But so far, the immunoassay method of LAE has not been reported at home and abroad.

Method used

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  • l-Lauramide arginine hydrochloride ethanol ester artificial antigen, preparation method of specific antibody and use thereof
  • l-Lauramide arginine hydrochloride ethanol ester artificial antigen, preparation method of specific antibody and use thereof
  • l-Lauramide arginine hydrochloride ethanol ester artificial antigen, preparation method of specific antibody and use thereof

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Experimental program
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Embodiment 1

[0030] 1. Synthesis of artificial antigens

[0031] Based on lauroyl arginine (LAS) is the main component in the molecular structure of LAE, and can fully expose the structural features and contain carboxyl groups that can be coupled with proteins, it is used as a hapten.

[0032] The structure of a hapten is as follows:

[0033]

[0034] 1.1 Synthesis and purification of immunizing antigens

[0035] The carbodiimide method was used for the synthesis of immune antigens: 71.3 mg of lauroyl arginine LAS (about 0.2 mmol) was dissolved in 1 mL of N,N-dimethylformamide DMF, and 81.6 mg of N-hydroxysuccinic acid was added. Imide NHS (about 0.6 mmol), stirred at room temperature for 15 min, and then added 72.9 mg of dicyclohexylcarbodiimide DCC (0.3 mmol), stirred at room temperature overnight, centrifuged, and slowly added 0.5 mL of the supernatant to the mixture. In 10 mL of 10 mg / mL bovine serum albumin carbonate buffer solution, the reaction was magnetically stirred for 5 ho...

Embodiment 2

[0047] 2. Synthesis of artificial antigens

[0048] Based on lauroyl arginine (LAS) is the main component in the molecular structure of LAE, and can fully expose the structural features and contain carboxyl groups that can be coupled with proteins, it is used as a hapten.

[0049] The structure of a hapten is as follows:

[0050]

[0051] 2.1 Synthesis and purification of immunizing antigens

[0052] The carbodiimide method was used for the synthesis of immune antigens: 107.0 mg of lauroyl arginine LAS (about 0.3 mmol) was dissolved in 2 mL of N,N-dimethylformamide DMF, and 122.4 mg of N-hydroxysuccinic acid was added. Imide NHS (0.9 mmol), stirred at room temperature for 15 min, then added 109.4 mg of dicyclohexylcarbodiimide DCC (0.45 mmol), stirred at room temperature overnight, centrifuged, and slowly added 0.8 mL of supernatant to 15 mL of 15 mg / mL of bovine serum albumin carbonate buffer solution, magnetic stirring for 5.5 hours. The solution after the reaction wa...

Embodiment 3

[0058] 3. Synthesis of artificial antigens

[0059] Based on lauroyl arginine (LAS) is the main component in the molecular structure of LAE, and can fully expose the structural features and contain carboxyl groups that can be coupled with proteins, it is used as a hapten.

[0060] The structure of a hapten is as follows:

[0061] 3.1 Synthesis and purification of immunizing antigens

[0062] The carbodiimide method was used for the synthesis of immune antigens: 142.6 mg of lauroyl arginine LAS (about 0.4 mmol) was dissolved in 1.5 mL of N,N-dimethylformamide DMF, and 163.2 mg of N-hydroxyl was added. Succinimide NHS (1.2 mmol) was stirred at room temperature for 15 min, and then 145.8 mg of dicyclohexylcarbodiimide DCC (0.6 mmol) was added. The reaction was stirred at room temperature overnight, centrifuged, and 1 mL of the supernatant was slowly added to 20 mL. 20mg / mL bovine serum albumin carbonate buffer solution, magnetic stirring for 6 hours. The solution after the r...

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Abstract

The invention discloses a preparation method of L-lauric arginine hydrochloride ethanol ester artificial antigen and specific antibody and use thereof, belongs to the technical field of immunochemical analysis, and is specially used for the preparation of L-lauric arginine-specific antibody and enzyme Combined immunoassay method, artificial antigens LAS-BSA and LAS-OVA were synthesized, and the specific polyclonal antibody of L-lauric amide arginine was obtained by immunizing rabbits. Finally, an enzyme-linked immunosorbent assay was established using the synthesized coating antigen and specific antibody. The method showed that the minimum detection limit of LAE was 0.005 mg / L, the recovery rate was 82.4-109.8%, and the average coefficient of variation was 3.2-5.8%. It had the characteristics of high accuracy and sensitivity, and met the residue analysis standards.

Description

technical field [0001] The invention belongs to the technical field of immunochemical analysis, and in particular relates to a preparation method and application of an antigen and a specific antibody of a novel food preservative L-lauric amide arginine hydrochloride ethanol ester. Background technique [0002] L-lauroyl arginine hydrochloride ethanol ester (ethyl lauroyl arginate (LAE)), the molecular formula is C 20 H 41 ClN 4 O 3 , the structure is as follows: [0003] [0004] LAE structure [0005] The appearance of LAE is white powder, the solubility in water at 20°C is 247g / kg, the chemical properties are stable in the pH range of 3-7, and it has high antibacterial activity, the melting point is 50-58°C, and its partition coefficient in water and oil greater than 10. That is, it mainly exists in the water phase. Compared with other preservatives with different chemical structures and similar functions, LAE can be more easily dissolved into water-based food and...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07C279/14C07K14/765C07K14/77C07K1/107C07K16/44G01N33/543G01N33/531
CPCG01N33/531G01N33/543C07C279/14C07K14/765C07K14/77C07K16/44C07K19/00
Inventor 付岩张艳张亮陈国吴银良
Owner NINGBO ACAD OF AGRI SCI
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