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Method for preparing high-capacity protein chromatographic medium through atom transfer radical polymerization

A chromatographic medium and atom transfer technology, applied in the field of protein separation and purification, to achieve mild reaction conditions, mild elution conditions, and high biological activity

Inactive Publication Date: 2015-10-28
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Chinese invention patent 201410142858.1 discloses a method for preparing a polymer-grafted protein separation medium by grafting temperature-sensitive polymer brushes on the surface of ultra-large-porous polystyrene microspheres through ATRP technology. The advantage is that the medium is used to separate proteins very well Protein activity is maintained, but the saturated adsorption capacity of protein is only 37.1mg per gram of dry bulb

Method used

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  • Method for preparing high-capacity protein chromatographic medium through atom transfer radical polymerization
  • Method for preparing high-capacity protein chromatographic medium through atom transfer radical polymerization
  • Method for preparing high-capacity protein chromatographic medium through atom transfer radical polymerization

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Preparation of polymer grafted ion-exchange chromatography medium pGMA-M-DMCH

[0027] Weigh 4g of hydroxylated polyglycidyl methacrylate microspheres (pGMA) and place in a 100mL three-necked flask containing 40mL of n-hexane to prepare the suspension, then add 1.2mL of triethylamine to it; After ice bathing, continue to add dropwise a mixed solution of 1.0mL 2-bromoisobutyryl bromide and 5.0mL n-hexane to the suspension; after the above suspension was reacted in ice bath for 3h, it was left at room temperature to continue the reaction for 20h; the obtained bromide After the pGMA medium was washed several times with n-hexane, absolute ethanol and deionized water, the bromine content in the brominated pGMA microspheres was measured to be 38.0 μmol / g dry microspheres; the brominated pGMA medium was named pGMA-M.

[0028] Weigh 2 g of pGMA-M medium and place it in 30 mL of a mixed solution of isopropanol and water with a volume ratio of 1:1; then, add 64 mmol of ...

Embodiment 2

[0030] In Example 1, the addition amount of the monomer compound DMC is 32mmol, and the addition amounts of cuprous bromide, copper bromide, and bipyridine are 0.320mmol, 0.032mmol, and 0.640mmol respectively. It is a 5.8 μm polymer-grafted ion-exchange chromatography medium pGMA-M-DMCM, and its ion exchange capacity is 2496±117 μmol / g dry microspheres.

Embodiment 3

[0032] In Example 1, the addition amount of the monomer compound DMC is 10.8mmol, and the addition amount of cuprous bromide, copper bromide, and bipyridine becomes 0.110mmol, 0.011mmol, and 0.220mmol respectively. The polymer-grafted ion-exchange chromatography medium pGMA-M-DMCL with a particle size of 2.44 μm has an ion-exchange capacity of 1972±63 μmol / g dry microspheres.

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Abstract

The invention relates to a method for preparing high-capacity protein ion-exchange chromatographic medium through atom transfer radical polymerization. The method comprises a bromination procedure of coupling a glycidyl methacrylate microsphere with the microsphere of an initiator 2-bromoisobutyryl bromide and a synthesis procedure of initiating polymerization of monomeric compounds on the surface of the microspheres through free radical transfer so as to obtain the ion-exchange polymer grafted chromatographic medium. The method prepares the polymer grafted ion-exchange chromatographic medium through atom transfer radical polymerization technology; the synthesized polymer grafted ion-exchange chromatographic medium has controllable density and chain length of graft polymer; reaction conditions are mild; the selection range of the monomeric compounds used as monomer is wide; gamma globulin saturated adsorption capacity of the synthesized polymer grafted ion-exchange chromatographic medium can reach above 800 mg / g of a wet medium, so the chromatographic medium has excellent adsorptivity; and adsorption equilibrium can be realized within 5 min in a protein solution with a concentration of 1 mg / mL, so the chromatographic medium has wide application prospects.

Description

technical field [0001] The invention relates to a method for preparing a high-capacity protein ion-exchange chromatography medium based on atom transfer radical polymerization, which belongs to the protein separation and purification technology in the field of biotechnology. Background technique [0002] Protein ion exchange chromatography is the core technology for the production of genetically engineered protein drugs. The ion-exchange chromatography medium, which is the basis of this technology, has lagged behind the expression level and production capacity of genetically engineered proteins seriously in the past two or three decades. For example, the binding capacity of bovine serum albumin in conventional ion-exchange chromatography media is only 70 mg / mL or even lower. Therefore, improving protein binding capacity has become one of the important prerequisites for realizing high efficiency of protein ion exchange chromatography. In ion-exchange chromatography, the bin...

Claims

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Application Information

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IPC IPC(8): C08F265/04C08F220/34C08F220/60B01J20/286B01J20/30B01D15/36
Inventor 史清洪李舒余林玲孙彦
Owner TIANJIN UNIV
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