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Theophylline conjugate and preparation method and application thereof

A conjugate, theophylline technology, applied in the field of clinical blood drug concentration monitoring

Inactive Publication Date: 2018-11-13
NANKAI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

To the best of our knowledge, there are currently no studies based on polyclonal antibody-based ELISA for the detection of theophylline

Method used

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  • Theophylline conjugate and preparation method and application thereof
  • Theophylline conjugate and preparation method and application thereof
  • Theophylline conjugate and preparation method and application thereof

Examples

Experimental program
Comparison scheme
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Embodiment example 1

[0039] (1) Preparation of solution A: Weigh theophylline-7-acetic acid 7.4mg, CDI 15mg, DMAP 0.38mg and dissolve in 1mL DMF in turn, stir at room temperature overnight in the dark, and set aside;

[0040] (2) Preparation of cBSA: Under ice bath conditions, slowly drop ethylenediamine into PBS buffer solution with a pH of 7.4 and a concentration of 0.01M, stir at 0-4°C, and adjust the pH of the solution to 7.4 with concentrated HCl; weigh 1g BSA (15μmol) and 56mg EDC (300μmol) were added to the above solution of ethylenediamine, stirred and reacted at 20°C for 2 hours; the reaction solution of ethylenediamine and BSA was mixed with PBS buffer at 0-4°C (0.01M, pH 7.4) dialyzed for 70 hours, and then dialyzed with distilled water for 24 hours, changing the dialysate every 6 hours; centrifuging the dialyzed solution at 13,000 rpm for 15 minutes at 0-4°C, and taking the supernatant solution; freeze-dried supernatant to obtain cBSA as a white powder solid, which was used for future ...

Embodiment example 2

[0046](1) Preparation of solution A: Weigh 7.4 mg of theophylline-7-acetic acid, 30 mg of CDI, and 0.38 mg of DMAP, and dissolve them in 1 mL of DMF in turn, stir overnight at room temperature in the dark, and set aside;

[0047] (2) Preparation of cBSA: Under ice bath conditions, slowly drop ethylenediamine into PBS buffer solution with a pH of 7.4 and a concentration of 0.01M, stir at 0-4°C, and adjust the pH of the solution to 7.4 with concentrated HCl; weigh 1g BSA (15μmol) and 56mg EDC (300μmol) were added to the above solution of ethylenediamine, stirred and reacted at 25°C for 4 hours; the reaction solution of ethylenediamine and BSA was mixed with PBS buffer at 0-4°C (0.01M, pH 7.4) dialyzed for 72 hours, then dialyzed with distilled water for 30 hours, changing the dialysate every 6 hours; centrifuge the dialyzed solution at 13,000 rpm for 15 minutes at 0-4°C, and take the supernatant solution; freeze-dried supernatant to obtain cBSA as a white powder solid, which was...

Embodiment example 3

[0052] (1) Preparation of solution A: Weigh 7.4 mg of theophylline-7-acetic acid, 30 mg of CDI, and 0.76 mg of DMAP, and dissolve them in 1 mL of DMF in turn, stir overnight at room temperature in the dark, and set aside;

[0053] (2) Preparation of cBSA: Under ice bath conditions, slowly drop ethylenediamine into PBS buffer solution with a pH of 7.4 and a concentration of 0.01M, stir at 0-4°C, and adjust the pH of the solution to 7.4 with concentrated HCl; weigh 1g BSA (15μmol) and 56mg EDC (300μmol) were added to the above solution of ethylenediamine, stirred and reacted at 25°C for 4 hours; the reaction solution of ethylenediamine and BSA was mixed with PBS buffer at 0-4°C (0.01M, pH 7.4) dialyzed for 80 hours, then dialyzed with distilled water for 20 hours, changing the dialysate every 6 hours; centrifuge the dialyzed solution at 13,000 rpm for 15 minutes at 0-4°C, and take the supernatant solution; freeze-dried supernatant to obtain cBSA as a white powder solid, which wa...

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Abstract

The invention discloses a theophylline conjugate and a preparation method and application thereof. The theophylline conjugate is as shown in general formula (I), wherein n is the molecular number of theophylline combined with one bovine serum albumin molecule and is an integer ranging from 1 to 20, BSA is bovine serum albumin, and the molecular weight range is 6.6-6.9KDa. The theophylline conjugate is formed by coupling theophylline hapten with carrier substance bovine serum albumin or ovalbumin generating immunogenicity. The preparation method includes: connecting and combining theophylline with the carrier substance generating immunogenicity to form the theophylline conjugate capable of inducing animal systems to generate antibodies. The theophylline conjugate immunizes New Zealand whiterabbits to prepare antiserum with the titer reaching 1:16000, and the lowest detection limit of the antiserum is 99ng / mL. The theophylline conjugate is fast, simple, accurate and high in specificityand lays a foundation for the enzyme linked immunosorbent assay kit of theophylline.

Description

technical field [0001] The invention relates to an anti-asthma drug conjugate and its preparation method and application, in particular to theophylline conjugate and its preparation method and application. The invention belongs to the field of clinical blood drug concentration monitoring. technical background [0002] The following titles involved in the present invention apply to the entire specification and claims: [0003] BSA:: Bovine Serum Albumin, product of Sigma [0004] PBS: Phosphate Buffered Saline (0.01M, pH=7.40) [0005] cBSA: bovine serum albumin modified with ethylenediamine [0006] Dialysis membrane: product of United Carbide [0007] EDC: Ethyl[3-(dimethylamino)propyl]carbodiimide, referred to as EDC, a product of Sigma [0008] Theophylline, also known as dioxodimethylpurine, is a methylpurine drug that has the effects of strengthening the heart, diuresis, dilating coronary arteries, and exciting the central nervous system. [0009] In recent decade...

Claims

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Application Information

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IPC IPC(8): C07K14/765C07K14/77C07K16/44C07K1/107
CPCC07K1/1077C07K14/765C07K14/77C07K16/44C07K19/00
Inventor 郗日沫宋兆瑞刘玮孟萌尹永梅董亚庆王玉芬
Owner NANKAI UNIV
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