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Method for preparing N-desulfated heparin derivative affinity chromatographic materials

A technology of sulfated heparin and chromatographic materials, applied in the fields of peptide preparation methods, chemical instruments and methods, separation methods, etc.

Inactive Publication Date: 2014-12-31
FUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Preparation of de-2-O, de-6-O, de-N sulfated reacetylated heparin derivatives affinity chromatography materials has not been reported at home and abroad

Method used

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  • Method for preparing N-desulfated heparin derivative affinity chromatographic materials
  • Method for preparing N-desulfated heparin derivative affinity chromatographic materials
  • Method for preparing N-desulfated heparin derivative affinity chromatographic materials

Examples

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Embodiment 1

[0033] A method for preparing a de-2-O sulfated heparin derivative affinity chromatography material, the preparation method comprising the following steps:

[0034] (1) Activation of agarose gel particles: take 2 g of sepharose 4B gel particles (purchased from sigma company, the same below) after washing and drying, dissolve in 2.15 ml of 0.6M NaOH, and add 0.3 ml of epichlorohydrin , reacted in a water bath shaker at 40°C for 2 hours, then fully washed with water, drained, added 4ml of concentrated ammonia water, shaken in a 40°C water bath, 120rpm / min shaker for 2 hours, fully washed with water and drained for later use, to obtain activated agarose Gel Particles A.

[0035] (2) Cross-linking of activated agarose gel particles and de-2-O sulfated heparin derivatives: Take 1.30 g of the above activated agarose gel particles A and dissolve them in 2.6 ml of phosphate buffer with a pH of 6.8 and 0.2 M Add 37.0 mg of de-2-O sulfated heparin derivative to the solution, then add 6...

Embodiment 2

[0038] A method for preparing a de-6-O sulfated heparin derivative affinity chromatography material, the preparation method comprising the following steps:

[0039] (1) Activation of agarose gel particles: Take 2g of sepharose 4B gel particles after washing and drying, dissolve in 2.15ml of 0.6M NaOH, add 0.3ml of epichlorohydrin, and react in a water bath shaker at 40°C After 2 hours, fully wash with water, drain, add 4ml of concentrated ammonia water, shake in a 40°C water bath, 120rpm / min shaker for 2 hours, fully wash with water and drain to obtain activated agarose gel particles A.

[0040] (2) Cross-linking of activated agarose gel particles and de-6-O sulfated heparin derivatives: Take 1.28g of the above activated agarose gel particles A, dissolve in 2.6ml pH 8, 0.2M phosphoric acid Add 31.8 mg of de-6-O sulfated heparin derivatives to the buffer solution, then add 6 mg of sodium cyanoborohydride, shake vigorously at room temperature for 4 days, filter, and freeze-dry t...

Embodiment 3

[0043] A method for preparing a de-N-position sulfated reacetylated heparin derivative affinity chromatography material, the preparation method comprising the following steps:

[0044] (1) Activation of agarose gel particles: Take 2g of sepharose 4B gel particles after washing and drying, dissolve in 2.15ml of 0.6M NaOH, add 0.3ml of epichlorohydrin, and react in a water bath shaker at 40°C 2 hours, then fully washed with water, drained, added 4ml of concentrated ammonia water, shaken in a 40°C water bath, 120rpm / min shaker for 2 hours, fully washed with water and drained for later use, to obtain activated agarose gel particles A

[0045] (2) Cross-linking of activated agarose gel particles and de-N-position sulfated reacetylated heparin derivatives: Take 1.28g of the above activated agarose gel particles A, dissolve in 2.6ml pH 6.8, 0.2M Add 49.6 mg of de-N-position sulfated reacetylated heparin derivatives, and then add 6 mg of sodium cyanoborohydride, shake vigorously at ro...

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Abstract

The invention relates to a method for preparing a series of N-desulfated heparin derivative affinity chromatographic materials. The method comprises the following steps of: by respectively utilizing a de-2-O desulfated heparin derivative, a de-6-O desulfated heparin derivative and a de-N-desulfated and acetylated heparin derivative as ligands and using sepharose particles as a carrier, preparing a series of stable N-desulfated heparin derivative affinity chromatographic materials through the reductive amination reaction of the activated sepharose particles and all the heparin derivatives in the presence of a strong reducing agent. According to the method, heparin derivatives with different desulfated loci are used for preparing the affinity chromatographic materials, so that three heparin derivative affinity chromatographies with high stability and high activity are obtained; and the affinity chromatographic materials can be used for researching the structure and functions of heparin and separating and concentrating functional protein, tumor cells, virus and the like.

Description

technical field [0001] The invention relates to the preparation of a series of de-2-O sulfated heparin derivatives, de-6-O sulfated heparin derivatives, and de-N-position sulfated reacetylated heparin derivatives. The preparation of affinity chromatography materials belongs to the biological affinity The field of chromatography materials. Background technique [0002] Heparin is a highly sulfated glycosaminoglycan composed of repeating disaccharide units linked by hexuronic acid and glucosamine with 1-4 glycosidic linkages. The main disaccharide unit constituting heparin is uronic acid-glucosamine disaccharide containing three sulfate groups. This repeated arrangement of trisulfated disaccharides forms the so-called "hypersulfated region" of heparin, which is a major part of the heparin structure. Studies have shown that heparin has biological functions such as anti-smooth muscle hyperplasia, anti-coagulant activity, anti-tumor and anti-virus. It interacts with various pro...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): B01J20/281B01J20/30B01D15/38C07K1/22
Inventor 魏峥邹强林江慧魏可镁
Owner FUZHOU UNIV
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