Propiconazole hapten as well as preparation method and application thereof

A technology of propiconazole and hapten, which is applied in the biological field, can solve the problems of expensive instruments and cannot be widely used, and achieve the effects of rapid detection, good affinity and simple operation

Pending Publication Date: 2021-11-26
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Although the instrument detection method has the advantages of high precision and accuracy, the disadvantages such as expensive instruments and the need for professional and technical personnel to operate prevent it from being widely used

Method used

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  • Propiconazole hapten as well as preparation method and application thereof
  • Propiconazole hapten as well as preparation method and application thereof
  • Propiconazole hapten as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1: Synthesis of propiconazole hapten

[0039] Step 1) Add 1.52g of N-bromosuccinimide and 2g of 2-chloro-4-bromo-acetophenone into 12mL of acetonitrile, and stir the reaction at 12°C for 10 minutes. Then 2.95 g of p-toluenesulfonic acid was added to the above mixture, and the mixture was stirred and reacted at 15-80° C. for 1.17 hours. The product obtained by the reaction was purified by a silica gel column, eluted and separated with petroleum ether-ethyl acetate at a volume ratio of 100:1 to 10:1, and product 1 was obtained.

[0040] Step 2) 4g of product 1 and 2g of 1,2-pentanediol were added to 24mL of toluene, then 12mL of n-butanol and 1.1g of p-toluenesulfonic acid were added, and the above mixture was stirred and reacted at 135°C for 15h. After the reaction, the product was purified through a silica gel column and eluted with petroleum ether-ethyl acetate at a volume ratio of 100:1 to 10:1 to obtain product 2.

[0041] Step 3) 1.39g KOH and 0.73g 1H-1,2,4...

Embodiment 2

[0045] Example 2: Identification of propiconazole hapten

[0046] The results of NMR identification of propiconazole hapten: 1H NMR (500MHz, DMSO-d6) δ8.36(d, J=7.1Hz, 1H), 7.84(d, J=16.3Hz, 1H), 7.41(dd, J =14.9, 7.9Hz, 1H), 7.31(dd, J=6.9, 1.6Hz, 1H), 7.13(ddd, J=18.7, 8.0, 1.7Hz, 1H), 4.77-4.6(m, 2H), 3.91- 3.75(m, 2H), 3.15(q, J=6.6, 5.5Hz, 1H), 2.58(td, J=7.5, 3.3Hz, 2H), 2.23(t, J=7.3Hz, 2H), 2.07(s , 1H), 1.62-1.46 (m, 4H), 1.27 (tdt, J=25.7, 14.3, 7.1 Hz, 4H), 0.92-0.72 (m, 3H).

Embodiment 3

[0047] Embodiment 3: the preparation of propiconazole antigen

[0048] Step 1) Activation of hapten. Get 0.082g propiconazole hapten as claimed in claim 1 and dissolve in 1.5mL N,N-dimethylformamide, after adding 0.023g N-hydroxysuccinimide to the above solution, stir at room temperature for 15 minutes , and then 1 mL of N,N-dimethylformamide solution containing 0.041 g of N,N-dicyclohexylcarbodiimide was added to the mixture, and the reaction was stirred overnight.

[0049] Step 2) Coupling of the hapten to the carrier protein. Centrifuge the reaction product in step 1 at 4000 rpm for 5 minutes, take 1.25 mL of the supernatant and slowly add it dropwise to 5 mL of 10 mg / mL bovine serum albumin in phosphate buffer (0.01 mol / L, pH=8), and stir at room temperature in the dark After reacting for 4 hours, propiconazole immune antigen was obtained; another 1.25 mL of centrifuged product in step 1 was slowly added dropwise to 5 mL of 10 mg / mL chicken ovalbumin phosphate buffer (0....

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Abstract

The invention discloses a propiconazole hapten as well as a preparation method and application thereof. According to the propiconazole hapten, a de novo synthesis method is adopted, a connecting arm containing four C atoms is connected to a benzene ring of propiconazole, and a carboxyl group capable of being coupled with carrier protein is introduced to the tail end of the connecting arm. The monoclonal antibody prepared by using the hapten is high in sensitivity and strong in specificity. The sensitivity (IC50) measured by an indirect competitive enzyme-linked immunoassay (ic-ELISA) method is 2.33 [mu] g / L, and the detection range (IC10-IC90) is 0.26-21.28 [mu] g / L. The monoclonal antibody and the propiconazole analogue have no cross reaction (CR is less than 0.1%). In addition, the monoclonal antibody is used for preparing the propiconazole colloidal gold rapid detection test strip which is high in detection sensitivity, strong in specificity, simple, convenient, rapid and low in cost. The test strip can be used for detecting propiconazole residues in agricultural products, and the detection limit of the test strip reaches 5 [mu] g / L.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a propiconazole hapten and its preparation method and application. Background technique [0002] Propiconazole is a systemic triazole fungicide with both protective and therapeutic effects. It is widely used to prevent and control fungi such as powdery mildew, rust, and anthracnose on cereals, vegetables, and fruits. Its mechanism of action is to inhibit the growth and development of fungi by affecting the biosynthesis of fungal ergosterol and inhibiting the metabolism of fungal steroids. In addition, propiconazole, as a plant growth regulator, has a good effect of controlling growth and pressing seedlings. It is often used to control the appearance of leafy vegetables such as cabbage, Chinese cabbage, and Chinese kale, and promote thick stems and leaves and dark green leaves. ,increase output. [0003] The wide application of propiconazole has caused human beings to th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07D405/06C07K14/765C07K14/77C07K16/44G01N33/53G01N33/577G01N33/58
CPCC07D405/06C07K14/765C07K14/77C07K16/44G01N33/577G01N33/587G01N33/5308
Inventor 华修德王鸣华李姣丁园陈贺
Owner NANJING AGRICULTURAL UNIVERSITY
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