A kind of affinity chromatography medium with glutathione as ligand

A chromatographic medium and glutathione technology, applied in the field of chromatographic medium, can solve the problems of low density of active epoxy groups, less coupling of glutathione, and high requirements for column filling technology, and achieve the coupling site Increased, increased epoxy group density, low production cost

Active Publication Date: 2020-07-31
SUZHOU BOJIN BIOLOGICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The active group comes from one of epichlorohydrin or allyl glycidyl ether, the density of active epoxy groups on the surface of the substrate is small, the amount of coupled glutathione is small, and the separation efficiency is low. The preparation process of affinity chromatography media with glutathione as a ligand is complex, expensive, and requires high column packing technology, and the technology is monopolized by a few companies

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] The preparation process is as follows:

[0019] 1) Soak 20g of agarose gel microspheres with a particle size of 30-100μm in 0.5L of deionized water, stir magnetically at 45-50°C for 15 minutes, then add 1.5g of nano-sized boron nitride to it, Stir magnetically at ~55°C for 1.5 to 2 hours, cool to room temperature, filter, wash and dry;

[0020] 2) Add 0.8g sodium hydroxide, 30mL acetonitrile, 40mL (0.5mol) epichlorohydrin, 55mL (0.3mol) resorcinol diglycidyl ether to the microspheres obtained in step 1), place the suspension in In a constant temperature oscillator at 40-45°C, react at a speed of 240r / min for 10 hours, filter after the reaction, wash the microspheres repeatedly with 20wt% ethanol solution and deionized water, and dry them with a vacuum pump;

[0021] 3) Add 30mL of coupling reaction solution consisting of 0.1mol / L glutathione, 0.1mol / L sodium phosphate, and 1mmol / L tetrasodium ethylenediaminetetraacetic acid dihydrate to the activated microspheres obtai...

Embodiment 2

[0024] The preparation process is as follows:

[0025] The amount of resorcinol diglycidyl ether added in step 2) of Example 1 was changed to 36 mL (0.2 mol) respectively, and the rest of the preparation process was the same as in Example 1.

Embodiment 3

[0027] The preparation process is as follows:

[0028] The amount of resorcinol diglycidyl ether added in step 2) of Example 1 was changed to 73 mL (0.4 mol) respectively, and the rest of the preparation process was the same as in Example 1.

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Abstract

The invention relates to an affinity chromatography medium with glutathione as a ligand. Agarose gel microspheres with boron nitride dispersed on surfaces are taken as a core, the surfaces of the agarose gel microspheres are activated by a first crosslinking agent and a second crosslinking agent, both the first crosslinking agent and the second crosslinking agent are coupled with glutathione as the ligand, the first crosslinking agent adopts epichlorohydrin, and the second crosslinking agent adopts resorcinol diglycidyl ether or neopentyl glycol diglycidyl ether; meanwhile, acetonitrile whichis cheaper and more environmentally friendly is selected as a solvent for coupling with glutathione, so that the dissolution degree of the crosslinking agents is increased, and the agarose gel microspheres are activated sufficiently. The affinity chromatography medium has the advantages that the density of active epoxy groups is high, the amount of coupled glutathione is large, the separation efficiency is higher, a preparation process of the affinity chromatography medium with glutathione as the ligand is simple, the production cost is lower, and large-scale popularization and application arefacilitated.

Description

technical field [0001] The invention relates to a chromatographic medium, in particular to an affinity chromatographic medium with glutathione as a specific ligand. Background technique [0002] The separation and purification of proteins are widely used in the research of biochemistry, biomedicine, biohealth food and other fields. As one of the important means of protein separation and purification, affinity chromatography medium is used to separate target proteins or other macromolecular substances that can specifically bind ligands in protein mixtures by covalently linking specific ligands on the chromatography medium. [0003] Glutathione (GSH) is one of the commonly used specific ligands. It is a tripeptide containing γ-amide bonds and sulfhydryl groups composed of glutamic acid, cysteine ​​and glycine. Glutathione can specifically bind glutathione-S-transferase (GST), glutathione-dependent proteins and glutathione-S-transferase recombinants. At the same time, due to ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): B01J20/26B01J20/28B01J20/30B01D15/38C07K1/22
CPCB01D15/3804B01J20/0259B01J20/267B01J20/28011C07K1/22
Inventor 瞿欢欢朱至放
Owner SUZHOU BOJIN BIOLOGICAL TECH
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