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Isoniazide derivative, homogeneous enzyme immunoassay reagent and preparation method

A homogeneous enzyme immunoassay and detection reagent technology, which is applied in the direction of immunoglobulin, chemical instruments and methods, measuring devices, etc., can solve complex operations, cannot meet the requirements of high-throughput and rapid clinical therapeutic drug monitoring, and take a long time And other issues

Active Publication Date: 2020-10-30
长沙博源医疗科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the detection results of these detection methods have certain problems in terms of accuracy and stability, and the operation is complicated and time-consuming, which cannot meet the high-throughput and rapid requirements of clinical therapeutic drug monitoring.

Method used

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  • Isoniazide derivative, homogeneous enzyme immunoassay reagent and preparation method
  • Isoniazide derivative, homogeneous enzyme immunoassay reagent and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Embodiment 1: the synthesis of isoniazid derivative

[0072] The synthetic route of isoniazid derivative is as follows:

[0073] .

[0074] The preparation method of isoniazid derivative, concrete steps are as follows:

[0075] Dissolve 2 g of compound 1 in 100 ml of toluene, then add 1.4 g of succinic anhydride to prepare a reaction mixture solution, reflux and stir the reaction mixture overnight, then concentrate the reaction mixture, and finally use flash chromatography to obtain 0.9 g of solid isofume Hydrazine derivatives.

[0076] passed 1 H NMR (Varian mercury plus 400 MHz) spectral scanning analysis (TMS as an internal standard) and LC-MS (Agilent 1200A) were used to identify the structure of the derivative. The results showed that the isoniazid derivative was represented by the formula (Ⅴ) The isoniazid derivatives shown.

[0077] Formula (Ⅴ).

Embodiment 2

[0078] Example 2: Preparation of Isoniazid Immunogen

[0079] The preparation method of isoniazid immunogen comprises the following steps:

[0080] (A1) Preparation of carrier solution: Dissolve carrier protein in 0.2M potassium phosphate buffer (pH=8.5), the final concentration of carrier protein is 4mg / mL, to obtain carrier solution;

[0081] (A2) Preparation of isoniazid derivative solution: 200 mg of the above-mentioned isoniazid derivative, 4 mL of dimethylformamide, 4 mL of ethanol, 5 mL of potassium phosphate buffer (10 mM, pH=5.0), 200 mg of 1-butane Base-3-(-3-dimethylaminopropyl) carbodiimide and 50 mg N-hydroxysulfosuccinimide were mixed, stirred and dissolved for 90 minutes to obtain a solution of isoniazid derivatives;

[0082] (A3) Synthesis of isoniazid immunogen: add the isoniazid derivative solution obtained in step (A2) to the carrier solution obtained in step (A1), stir overnight at 4°C, and purify by dialysis to obtain isoniazid Niacinamide immunogen.

Embodiment 3

[0083] Example 3: Preparation of Isoniazid Immunogen

[0084] The preparation method of isoniazid immunogen comprises the following steps:

[0085] (A1) Preparation of carrier solution: Dissolve carrier protein in 0.2M potassium phosphate buffer (pH=8.5), the final concentration of carrier protein is 3mg / mL to obtain carrier solution;

[0086] (A2) Preparation of isoniazid derivative solution: 100 mg of the above-mentioned isoniazid derivative, 2 mL of dimethylformamide, 2 mL of ethanol, 3 mL of potassium phosphate buffer (10 mM, pH=5.0), 100 mg of 1-butane Base-3-(-3-dimethylaminopropyl)carbodiimide and 20mg N-hydroxysulfosuccinimide were mixed, stirred and dissolved for 30 minutes to obtain a solution of isoniazid derivatives;

[0087] (A3) Synthesis of isoniazid immunogen: add the isoniazid derivative solution obtained in step (A2) to the carrier solution obtained in step (A1), stir overnight at 0°C, and purify by dialysis to obtain isoniazid Niacinamide immunogen.

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Abstract

The invention discloses an isoniazid derivative, a detection reagent and a preparation method, and relates to the technical field of biological detection. An isoniazid immunogen prepared from the isoniazid derivative has high immunogenicity, and an obtained antibody has strong specificity and high titer; an isoniazid enzyme-labeled conjugate in a homogeneous enzyme immunoassay reagent prepared from the derivative is connected with recombinant glucose-6-phosphate dehydrogenase modified by gene engineering; the detection sensitivity is remarkably improved, a sample with the concentration as lowas 5 ng / ml or below can be effectively detected, the specificity is high, and cross reaction with 62 common drugs is avoided; the homogeneous enzyme immunoassay reagent can realize high-throughput andrapid detection of isoniazid content on a full-automatic biochemical analyzer, and is stable in detection result, high in accuracy, simple in detection method and easy to realize, popularize and use.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to an isoniazid derivative, a homogeneous enzyme immunoassay reagent and a preparation method. Background technique [0002] Isoniazid (4-Pyridinecarboxylic acid hydrazide), the chemical name is 4-pyridinecarboxyhydrazide, and the trade name is Remifen. Its chemical structural formula is shown in formula (IV): [0003] Formula (Ⅳ). [0004] Isoniazid is the first-line drug for the treatment of tuberculosis. The mechanism of action of isoniazid as a drug for the treatment of tuberculosis is mainly through binding with the coenzyme of tuberculosis bacteria to prevent the catalysis of the corresponding enzyme, so as to achieve the branching of tuberculosis bacteria wall. Bacteric acid cannot be synthesized smoothly, which hinders the growth and reproduction of tuberculosis bacteria. Isoniazid is highly selective to Mycobacterium tuberculosis and has a strong antibacter...

Claims

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Application Information

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IPC IPC(8): C07D213/86C07K14/47C07K14/765C07K14/77C07K14/795C07K16/44G01N33/535G01N33/543
CPCC07D213/86C07K14/47C07K14/765C07K14/77C07K14/795C07K16/44G01N33/535G01N33/543
Inventor 张小可周伟峰张明张英蕾
Owner 长沙博源医疗科技有限公司
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