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A kind of derivative of 17-hydroxysteroid, detection reagent and preparation method

A technology for detecting reagents and steroids, which is applied in the field of detection reagents and preparation of derivatives of 17-hydroxysteroids, and can solve problems such as poor stability and low sensitivity

Active Publication Date: 2020-08-18
苏州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

At present, there are few detection reagents on the market, and there are still some defects, such as: poor stability, low sensitivity, not suitable for simultaneous detection of a large number of samples, etc.

Method used

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  • A kind of derivative of 17-hydroxysteroid, detection reagent and preparation method
  • A kind of derivative of 17-hydroxysteroid, detection reagent and preparation method
  • A kind of derivative of 17-hydroxysteroid, detection reagent and preparation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Example 1: Synthesis of 17-hydroxysteroid derivatives

[0072] The synthetic route of 17-hydroxysteroid derivatives is as follows:

[0073]

[0074] The preparation method of 17-hydroxy steroid derivatives, concrete steps are as follows:

[0075] (1) Synthesis of compound 2: Dissolve 5 g of compound 1 and 2.1 g of triethylamine (TEA) in dichloromethane (DCM) (50 mL), then add dihydrofuran-2,5- Diketone 1.8g, stirred at room temperature for 2 hours, thin layer chromatography (TLC) showed that after the reaction was complete, the reaction was quenched with 300mL purified water, the pH=3 was adjusted with 1N hydrochloric acid, and the reacted mixture was treated with 300mL DCM The extraction was repeated 3 times, the organic layers were combined, washed with purified water (200 mL) and brine (200 mL), dried and concentrated in vacuo to obtain 6 g of white solid, namely compound 2, yield: 94%;

[0076] (2) Synthesis of compound 3: Dissolve compound 2 (6g) obtained in s...

Embodiment 2

[0083] Example 2: Preparation of 17-hydroxysteroid immunogen and anti-17-hydroxysteroid specific antibody

[0084] The preparation steps of 17-hydroxysteroid immunogen are as follows:

[0085] (A1) Preparation of hemocyanin solution: 100 mg of hemocyanin was dissolved in 25 mL of 0.2 M, pH=8.5 phosphate buffer to obtain a hemocyanin solution;

[0086] (A2) Preparation of 17-hydroxysteroid derivative solution: 100 mg of the 17-hydroxysteroid derivative obtained in Example 1, 1.75 mL dimethylformamide, 1.75 mL ethanol, 3.5 mL, 10 mM, potassium phosphate at pH=5.0 Buffer, 100mg 1-ethyl-3-(-3-dimethylaminopropyl) carbodiimide, 25mg N-hydroxysulfosuccinimide were mixed, stirred and dissolved for 90min to obtain 17-hydroxysteroid derivatives solution;

[0087] (A3) Obtaining 17-hydroxysteroid immunogen: add the 17-hydroxysteroid derivative solution obtained in step (A2) to the hemocyanin solution obtained in step (A1), stir overnight at 4°C, and purify by dialysis , to obtain the 1...

Embodiment 3

[0092] Example 3: Preparation of 17-hydroxysteroid immunogen and anti-17-hydroxysteroid specific antibody

[0093] The preparation steps of 17-hydroxysteroid immunogen are as follows:

[0094] (A1) Preparation of thyroglobulin solution: 200 mg of thyroglobulin was dissolved in 50 mL, 0.2 M, pH=8.5 phosphate buffer to obtain thyroglobulin solution;

[0095] (A2) Preparation of 17-hydroxysteroid derivative solution: 200 mg of the 17-hydroxysteroid derivative obtained in Example 1, 3.5 mL of dimethylformamide, 3.5 mL of ethanol, 7.0 mL, 10 mM, potassium phosphate of pH=5.0 Buffer, 200mg 1-ethyl-3-(-3-dimethylaminopropyl) carbodiimide, 50mg N-hydroxysulfosuccinimide were mixed, stirred and dissolved for 90min to obtain 17-hydroxysteroid derivatives solution;

[0096] (A3) Obtaining 17-hydroxysteroid immunogen: add the 17-hydroxysteroid derivative solution obtained in step (A2) to the thyroglobulin solution obtained in step (A1), stir overnight at 4°C, and purify by dialysis , t...

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Abstract

The invention discloses a derivative of 17-hydroxysteroid, a detection reagent and a preparation method thereof, and relates to the technical field of biological detection. A 17-hydroxysteroid immunogen prepared by using the 17-hydroxysteroid derivative has high immunogenicity, and antibody specificity is strong and titer is high. A 17-hydroxyl steroid conjugate in a homogeneous enzyme immunoassayreagent prepared by using the derivative is linked to genetically engineered recombinant glucose-6-phosphate dehydrogenase. Thereby, detection sensitivity is significantly enhanced, and a sample withthe concentration being as low as 80 nmol / L can be detected effectively; and the detection reagent has strong specificity and has no cross reaction with common 92 interferents. High-throughput and rapid detection of the 17-hydroxysteroid content can be realized by a fully automatic biochemical analyser. The detection result is stable, accuracy is high, and the detection method is simple and is easy for implementation and promotion and application.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a derivative of 17-hydroxysteroid, a detection reagent and a preparation method. Background technique [0002] 17-hydroxysteroids (17-hydroxycorticosteroids, 17-OHCS), its structural formula is as shown in formula (IV): [0003] [0004] 17-Hydroxysteroids are sugar adrenal corticosteroids, which are hormones and their metabolites secreted by the adrenal cortex, mainly including cortisol, cortisol, tetrahydrocortisol and tetrahydrocortisol, etc., which are often excreted with urine. Therefore, the content of 17-hydroxysteroids in urine can reflect the secretion of cortisol by the adrenal cortex, and can assist in the diagnosis of related endocrine diseases. Decreased 17-hydroxysteroid levels are common in adrenal insufficiency, hypopituitarism, bilateral adrenalectomy, malnutrition, liver cirrhosis, tuberculosis, and certain chronic wasting diseases; 17-hydroxyst...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07J7/00G01N33/531G01N33/74
CPCC07J7/008G01N33/531G01N33/74
Inventor 虞留明张小可
Owner 苏州博源医疗科技有限公司
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