A polymer-grafted hydrophobic charge-induced chromatography medium and its preparation method

A hydrophobic charge and induction layer technology, applied in the field of protein chromatography separation technology, can solve problems such as no reports, and achieve the effects of high effective diffusion coefficient, convenient elution and high efficiency

Active Publication Date: 2016-08-17
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, the present invention is aimed at the special requirement of HCIC medium, adopts ARGET-ATRP new method, forms polyglycidyl methacrylate graft in the internal channel of porous microsphere matrix, and HCIC is coupled to polyglycidyl methacrylate On grafting, prepare polymer-grafted HCIC new medium, improve the coupling efficiency and density of ligands, improve the spatial arrangement of ligands in the matrix, increase the adsorption capacity and adsorption rate of antibodies, and facilitate the chromatography medium in High-efficiency antibody adsorption is achieved at high flow rates, and related content has not been reported at home and abroad

Method used

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  • A polymer-grafted hydrophobic charge-induced chromatography medium and its preparation method
  • A polymer-grafted hydrophobic charge-induced chromatography medium and its preparation method
  • A polymer-grafted hydrophobic charge-induced chromatography medium and its preparation method

Examples

Experimental program
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Effect test

Embodiment 1

[0029]Take 10 g of drained agarose gel, replace the contained water with acetone, then add 100 ml of anhydrous tetrahydrofuran, 5 ml of 2-bromoisobutyryl bromide, 5.5 ml of anhydrous triethylamine and 0.25 g of 4-dimethylaminopyridine, 0 Under ice bath for 2h at 30°C, then activate with magnetic stirring in a sealed three-neck flask for 12 hours at 30°C, filter with suction, wash with acetone and deionized water to obtain the activated matrix; then mix the activated matrix, 2ml glycidyl methacrylate, 100ml80% Mix isopropanol solution (v / v), 0.03g copper bromide, 0.06g ascorbic acid, and 0.06ml pentamethyldiethylenetriamine for surface-initiated free radical polymerization, and magnetically stir in a sealed three-necked flask at 50°C React for 12 hours, filter with suction, wash alternately with acetone and deionized water, remove the copolymer, and obtain a polymer-grafted matrix; then mix the grafted matrix with 0.8g 2-mercapto-1-methylimidazole and 1M sodium carbonate soluti...

Embodiment 2

[0031] Take 10 g of drained agarose gel, replace the contained water with acetone, add 100 ml of anhydrous tetrahydrofuran, 1 ml of 2-bromoisobutyryl bromide, 1.1 ml of anhydrous triethylamine and 0.05 g of 4-dimethylaminopyridine, 0 °C Under ice bath for 2h, then magnetically stirred and activated in a sealed three-neck flask at 30°C for 12 hours, suction filtered, washed with acetone and deionized water to obtain the activated matrix; then the activated matrix, 0.5ml glycidyl methacrylate, 100ml 80 % isopropanol solution (v / v), 0.01g of copper bromide, 0.02g of ascorbic acid and 0.02ml of pentamethyldiethylenetriamine were mixed to carry out surface-induced free radical polymerization, and magnetically Stir the reaction for 12 hours, filter with suction, and wash alternately with acetone and deionized water to remove the copolymer to obtain a polymer-grafted matrix; then mix the grafted matrix with 0.2g 2-mercapto-1-methylimidazole and 1M sodium carbonate Buffer solution (pH...

Embodiment 3

[0033] Take 10 g of drained agarose gel, replace the contained water with acetone, add 200 ml of anhydrous tetrahydrofuran, 1 ml of 2-bromoisobutyryl bromide, 1.1 ml of anhydrous triethylamine and 0.05 g of 4-dimethylaminopyridine, 0 °C Under ice bath for 3h, then magnetically stirred and activated in a sealed three-necked flask at 30°C for 24 hours, suction filtered, washed with acetone and deionized water to obtain the activated matrix; then the activated matrix, 2ml glycidyl methacrylate, 100ml 80% Mix isopropanol solution (v / v), 0.03g copper bromide, 0.06g ascorbic acid, and 0.06ml pentamethyldiethylenetriamine for surface-initiated free radical polymerization, and magnetically stir in a sealed three-necked flask at 50°C React for 12 hours, filter with suction, wash alternately with acetone and deionized water, remove the copolymer, and obtain a polymer-grafted matrix; then mix the grafted matrix with 0.8g 2-mercapto-1-methylimidazole and 1M sodium carbonate solution (pH 1...

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Abstract

The invention discloses a poly-glyceryl methacrylate grafted hydrophobic charge-induced chromatography medium and a preparation method thereof. The preparation method comprises the following steps: taking hydrophilic porous microspheres as a chromatography medium, activating the chromatography medium by virtue of an activating reaction, carrying out a poly-glyceryl methacrylate polymerization reaction on the surface of the activated chromatography medium, thereby obtaining a polymer grafted chromatography medium; and finally, coupling a hydrophobic charge-induced ligand, thereby obtaining the polymer grafting type hydrophobic charge-induced chromatography medium. The chromatography medium disclosed by the invention has the poly-glyceryl methacrylate grafting chain and hydrophobic charge-induced functional ligand, the ligand density is high, the dynamic loading capacity of target proteins such as antibodies is obviously improved at high flow velocity, the chromatography medium has salt-independent adsorption characteristics, and desorption and recovery can be realized by changing the pH value of the solution to be weakly acidic. The preparation process of the novel medium is simple and convenient, strict oxygen removal in the grafting reaction process is not needed, and the amount of catalysts and ligand is small.

Description

technical field [0001] The invention relates to a polymer-grafted hydrophobic charge-induced chromatographic medium and a preparation method thereof, belonging to free radical polymerization technology in the field of macromolecular chemistry and protein chromatography separation technology in the field of biochemical industry. Background technique [0002] With the advancement of upstream technologies such as hybridoma technology, cell biology and genetic engineering, antibodies and related medical proteins have been developed rapidly. Antibodies have the characteristics of strong targeting, high specificity, and low toxicity and side effects. They are widely used in the treatment of major diseases such as tumors and autoimmune diseases, in vitro diagnosis and detection, and have become the key development direction of biotechnology drugs. At present, through the culture of animal mammalian cells, the expression level of antibodies can reach more than 5g / L, but the downstre...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): B01J20/26B01J20/30B01D15/32C08J9/42C08J9/40C07K1/14
CPCB01D15/32B01J20/26B01J20/30C07K1/14C08J9/405C08J9/42C08J2301/02C08J2305/12
Inventor 林东强刘韬姚善泾
Owner ZHEJIANG UNIV
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