A Metal Chelate Affinity Chromatography Medium Containing Rejection Layer

A technology of metal chelation and chromatography medium, which is applied in the field of metal chelation affinity chromatography medium, can solve the problems of unsatisfactory specific separation effect and cumbersome process, and achieve the reduction of non-specific adsorption, compact and solid core structure, The effect of high hardness

Active Publication Date: 2020-03-31
SUZHOU BOJIN BIOLOGICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the prior art, a hydrophilic substance polyethylene glycol (PEG) is grafted on the surface of the affinity medium matrix to form a semi-permeable protective barrier to size-exclude macromolecular impurities, thereby improving the separation and purification of small molecular polypeptides efficiency, but this method needs to convert the terminal hydroxyl groups of PEG into active groups such as aldehyde groups, carboxyl groups, halogens, and amino groups in advance to graft it on the matrix. The process is cumbersome, and the target small molecule The specific separation effect of peptides is not ideal

Method used

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  • A Metal Chelate Affinity Chromatography Medium Containing Rejection Layer

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] The preparation process is as follows:

[0021] (1) Soak 20 g of macroporous silica microspheres with a particle size of 50-100 μm and a pore size greater than 50 nm in a 0.5 mol / L sodium hydroxide solution, and stir at 50-55°C for 1-1.5 hours to make the macropores Silica microsphere activation, vacuum drying for 12 hours;

[0022] (2) Add 250mL of acetone solution with a volume fraction of 30% to the mixture of 60g sodium alginate, 54g chitooligosaccharide, 6g silicon nitride, and 10g sodium hydroxide, vigorously stir and ultrasonically form a uniform suspension; After the silica microspheres are taken out from the vacuum drying oven, they are quickly placed in the suspension, stirred at 45-50°C for 8-10 hours, filtered, rinsed with deionized water and dried to obtain the macroporous carbon dioxide filled with the mixture. Silicon microsphere core;

[0023] (3) Add 50 mL of acetone, 15 g of epichlorohydrin, and 5 g of sodium hydroxide to the inner core of the micros...

Embodiment 2

[0027] The preparation process is as follows:

[0028] The copper acetate solution in the step (5) of Example 1 is replaced with nickel acetate of the same concentration, and all the other preparation processes are the same as in Example 1.

Embodiment 5

[0042] Commercially available metal chelate affinity chromatography media that chelate copper ions.

[0043] In order to test the affinity adsorption efficiency of the metal chelate affinity chromatography medium prepared by the present invention to the small molecular polypeptide, and the repelling effect to the macromolecular protein, bovine serum albumin BSA is used as the macromolecular protein model here, it consists of Composed of 583 amino acid residues, there are a variety of amino acid sequences that are easy to chelate with metal ions, which are macromolecular protein impurities that need to be removed; a polypeptide fragment VSLPEW (Val-Ser-Leu- Pro-Glu-Try) as the target small molecule polypeptide, wherein the small molecule polypeptide has a good blood pressure lowering function, and the amino nitrogen or carboxyl oxygen in the VSLPEW polypeptide can be chelated with copper ions, thereby achieving separation; from the actual fermentation or enzyme When extracting ...

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Abstract

The invention relates to a metal-chelating affinity chromatography medium containing a resistant layer. The preparation method comprises the following steps: by taking macroporous silica microspheresas a core, filling a mixture composed of three components, namely sodium alga acid, chitosan oligosaccharide and silicon nitride, in pore channels of the microspheres, bonding epoxy chloropropane ontothe core surface, connecting the epoxy chloropropane and a ligand, and complexing the ligand and metal ions; and adhering amylose to form the resistant layer on the core surface, wherein a mass ratioof the epoxy chloropropane to the amylose is 1:(4-5). According to the resistant layer, blocking of nonspecific macromolecules is realized, nonspecific adsorption is reduced, and the adsorption and separation efficiency on target micro-molecule polypeptides is improved.

Description

technical field [0001] The invention relates to a chromatographic medium, in particular to a metal chelating affinity chromatographic medium containing a repelling layer. Background technique [0002] In recent years, with the continuous advancement of biotechnology, the rapid and effective separation and purification of peptides from complex natural mixtures has gradually become the key and difficulty in the development of new drugs. Therefore, whether it is possible to efficiently and stably separate and purify peptides has become the premise and basis for the research and application of peptides. With the rapid development of chromatography technology, immobilized metal chelate affinity chromatography, as an efficient separation technology, can be applied to the separation and purification of peptides. [0003] Immobilized metal chelation affinity chromatography is a separation technology based on the coordination between metal ions and amino acid residues. Due to its ad...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): B01J20/24B01J20/30B01D15/38C07K1/22
CPCB01D15/3804B01J20/103B01J20/24B01J2220/4806B01J2220/4812B01J2220/4825C07K1/22
Inventor 瞿欢欢朱至放
Owner SUZHOU BOJIN BIOLOGICAL TECH
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