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Zonisamide derivative, homogeneous enzyme immunoassay reagent and preparation method

A technology of homogeneous enzyme immunoassay and zonisamine enzyme is applied in the field of zonisamide derivatives, homogeneous enzyme immunoassay detection reagents and preparation, and can solve the problem of high professional requirements of operators, high price of high-precision instruments, and microscopic latex. It can improve the detection sensitivity, be easy to implement and popularize and use, and achieve the effect of high accuracy.

Pending Publication Date: 2022-05-06
安徽爱索特克医学检验实验室有限公司
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  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

High-performance liquid chromatography has high sensitivity and a wide detection range, but it has defects such as complicated sample processing, high-precision instruments are expensive, require specially trained technicians to operate, and high maintenance costs, making it difficult to be widely promoted in clinical practice
The advantages of ion chromatography-suppressed conductometric detection method are convenience, high sensitivity, and good selectivity, and ion chromatography-suppressed conductometric detection method can measure organic ions as quickly and conveniently as analyzing inorganic ions, and does not pollute the environment, but the detection method samples The pretreatment steps are relatively complicated, the detection cost is high, and the professional requirements for operators are high, so it is difficult to be widely used in clinical practice.
The advantages of latex-enhanced immunoturbidimetry are good precision, accuracy, sensitivity and stability, and easy operation, but the preparation of latex microspheres is complicated and costly

Method used

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

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Embodiment 1

[0075] Embodiment 1: Preparation of zonisamide derivatives

[0076] The synthetic route of zonisamide derivative is as follows:

[0077] .

[0078] The preparation method of described zonisamide derivative comprises the following steps:

[0079] Compound 1 (7 g) was weighed and dissolved in toluene (70 ml), then succinic anhydride (4 g) was added to prepare a reaction mixture solution, and the above reaction mixture solution was stirred overnight under reflux. After the reaction, the above reaction mixture solution was concentrated, and then purified by flash chromatography to obtain 4 g of zonisamide derivatives.

[0080] pass 1 H NMR (Varian mercury plus 400 MHz) spectral scanning analysis (TMS as internal standard) and LC-MS (Agilent 1200A) were used to identify the chemical structure of the obtained zonisamide derivatives. The results showed that: the zonisamide derivatives It is the zonisamide derivative with the structural formula shown in formula (V).

[0081] ...

Embodiment 2

[0083] Embodiment 2: Preparation of zonisamide immunogen

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

[0085] (A1) Preparation of carrier solution: Dissolve carrier protein in 0.2mol / L potassium phosphate buffer (pH=8.5), the final concentration of carrier protein is 5.5mg / mL to obtain carrier solution;

[0086] (A2) Preparation of zonisamide derivative solution: 200 mg of the above-mentioned zonisamide derivative, 4 mL of dimethylformamide, 4 mL of ethanol, 6 mL of potassium phosphate buffer (10 mmol / L, pH=7.0), 200mg of 1-ethyl-3-(-3-dimethylaminopropyl)carbodiimide and 60mg of N-hydroxysulfosuccinimide were mixed, stirred and dissolved for 120min to obtain a solution of zonisamide derivatives;

[0087] (A3) Synthesis of zonisamide immunogen: add the zonisamide derivative solution obtained in step (A2) to the carrier solution obtained in step (A1), stir overnight at 4°C, and purify by dialysis, The zonisamide immunogen was obtaine...

Embodiment 3

[0088] Example 3: Preparation of Anti-Zonisamide Specific Antibody

[0089] The preparation method of anti-zonisamide specific antibody comprises the following steps:

[0090] (B1) Dilute the above-mentioned zonisamide immunogen with 0.01mol / L sodium phosphate buffer (pH=6.5) to a final concentration of 2.0mg / mL to obtain an artificial antigen solution, and then mix the artificial antigen solution with an equal amount of Mixed with complete adjuvant, the experimental animal rabbits were injected at multiple points;

[0091] (B2) After 3 weeks, the same artificial antigen solution and the same amount of Freund's incomplete adjuvant were used to inject the above experimental animal rabbits at multiple points, and then injected once every 3 weeks, for a total of 5 injections;

[0092] (B3) Blood was collected from the experimental animal rabbit that had been injected in step (B2), separated and purified to obtain anti-zonisamide-specific antibodies.

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Abstract

The invention discloses a zonisamide derivative, a detection reagent and a preparation method, and relates to the technical field of biological detection. The zonisamide immunogen prepared from the zonisamide derivative is high in immunogenicity, and the obtained antibody is high in specificity and high in titer; a zonisamide enzyme-labeled conjugate in a homogeneous enzyme immunodetection reagent prepared from the derivative is connected with recombinant glucose-6-phosphate dehydrogenase modified by genetic engineering, so that the detection sensitivity is remarkably improved, a sample with the concentration as low as 2.00 g / mL or below can be effectively detected, the specificity is high, and no cross reaction with common 62 other drugs exists; the zonisamide content can be rapidly detected with high flux on a full-automatic biochemical analyzer, the detection result is stable, the accuracy is high, and the detection method is simple and easy to implement, popularize and use.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a zonisamide derivative, a homogeneous enzyme immunoassay reagent and a preparation method. Background technique [0002] Zonisamide (Zonisamide, ZNS) is a new type of antiepileptic drugs (antiepileptic drugs, AEDs), its chemical name is 1,2-benzisoxazole-3-methanesulfonamide, its molecular weight is 212.23, and its molecular formula is C 8 h 8 N 2 o 3 S, its chemical structural formula is shown in formula (IV): [0003] [0004] Formula (Ⅳ). [0005] Zonisamide exerts its effect mainly through three mechanisms: (1) altering the voltage-dependent ion channels related to action potential propagation or burst firing; (2) enhancing GABA-mediated inhibition; (3) interfering with amino acid-mediated Guided excitability. Main clinical applications: treatment of simple and complex partial seizures and secondary generalized tonic-clonic seizures, partial and general...

Claims

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Application Information

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IPC IPC(8): C07D261/20G01N33/535
CPCC07D261/20G01N33/94G01N33/535
Inventor 余琳赵玉冰李冬蔡江丽
Owner 安徽爱索特克医学检验实验室有限公司
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