A high-efficiency hydrophobic interaction chromatography medium with benzylamine as a ligand, its preparation method and its application in protein refolding and purification

A technology of hydrophobic interaction and chromatographic medium, which is applied in the application fields of high-efficiency hydrophobic interaction chromatographic medium and its preparation, renaturation and purification, can solve the problems of cumbersome steps and large losses, and achieves high purity, high renaturation efficiency, The effect of high activity recovery rate

Active Publication Date: 2021-06-11
NORTHWEST UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The steps of using this method are cumbersome, and the loss in the process is relatively large

Method used

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  • A high-efficiency hydrophobic interaction chromatography medium with benzylamine as a ligand, its preparation method and its application in protein refolding and purification
  • A high-efficiency hydrophobic interaction chromatography medium with benzylamine as a ligand, its preparation method and its application in protein refolding and purification
  • A high-efficiency hydrophobic interaction chromatography medium with benzylamine as a ligand, its preparation method and its application in protein refolding and purification

Examples

Experimental program
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Effect test

Embodiment 1

[0037]A kind of preparation method using benzylamine as a high-efficiency hydrophobic interaction chromatographic medium of ligand, comprises the following steps:

[0038] (1) Silica gel activation: Add unactivated silica gel into hydrochloric acid solution with a mass concentration of 20% according to the mass volume ratio of 1g:15mL, reflux and stir at 120°C for 2h, wash with distilled water and filter until neutral, Dry in a vacuum oven for 24 hours to obtain activated silica gel; wherein, the unactivated silica gel has a pore size of 80-200 Å and a particle size of 5-10 μm spherical silica gel;

[0039] (2) Preparation of epoxy silica gel: according to 1g: 50mL, add the activated silica gel obtained in step (1) into anhydrous toluene, sonicate for 3 minutes to disperse the activated silica gel evenly, and then slowly add silane distillate into it under stirring condition Coupling agent, the mass volume ratio of the activated silica gel and silane coupling agent is 1g:1mL, ...

Embodiment 2

[0042] A preparation method of a high-efficiency hydrophobic interaction chromatographic medium using benzylamine as a ligand, comprising the following steps:

[0043] (1) Silica gel activation: add unactivated silica gel into hydrochloric acid solution with a mass concentration of 20% according to the mass volume ratio of 1g:20mL, reflux and stir at 120°C for 6h, wash with distilled water and filter until neutral, Dry in a vacuum oven for 12 hours to obtain activated silica gel; wherein, the unactivated silica gel has a pore size of 80-200 Å and a particle size of 5-10 μm spherical silica gel;

[0044] (2) Preparation of epoxy silica gel: According to 1g: 50mL, add the activated silica gel obtained in step (1) into anhydrous toluene, sonicate for 5 minutes to disperse the activated silica gel evenly, and then slowly add silane distillate into it under stirring condition Coupling agent, the mass volume ratio of the activated silica gel and silane coupling agent is 1g:2 mL, war...

Embodiment 3

[0047] A kind of preparation method using benzylamine as a high-efficiency hydrophobic interaction chromatographic medium of ligand, comprises the following steps:

[0048] (1) Silica gel activation: Add unactivated silica gel into hydrochloric acid solution with a mass concentration of 20% according to the mass volume ratio of 1g:10mL, reflux and stir at 120°C for 5h, wash with distilled water and filter until neutral, Dry in a vacuum oven for 18 hours to obtain activated silica gel; wherein, the unactivated silica gel has a pore size of 80-200 Å and a particle size of 5-10 μm spherical silica gel;

[0049] (2) Preparation of epoxy silica gel: according to 1g: 50mL, add the activated silica gel obtained in step (1) into anhydrous toluene, sonicate for 4 minutes to disperse the activated silica gel evenly, and then slowly add silane distillate into it under stirring condition Coupling agent, the mass volume ratio of the activated silica gel and silane coupling agent is 1g:1mL,...

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Abstract

The invention discloses a high-efficiency hydrophobic interaction chromatographic medium with amine as a ligand, and its structural formula is:; the invention also discloses a preparation method of the high-efficiency hydrophobic interaction chromatographic medium, which includes the following steps: (1) Silica gel activation; (2) Preparation of epoxy silica gel; (3) Preparation of high-efficiency hydrophobic interaction chromatography medium by epoxy ring opening; at the same time, the present invention also discloses the refolding and reconstitution of the high-efficiency hydrophobic interaction chromatography medium with recombinant streptavidin The application of purification has high renaturation efficiency, high activity recovery rate, and high purity of recombinant streptavidin after renaturation.

Description

technical field [0001] The invention belongs to the technical field of high-performance liquid chromatography, and specifically relates to a high-efficiency hydrophobic interaction chromatography medium with benzylamine as a ligand and a preparation method thereof, and its use in high-performance liquid chromatography for protein recombinant streptavidin Renaturation and purification applications. Background technique [0002] Streptavidin (STV) is an exocrine protein produced during the cultivation of actinomycetes, which can highly specifically bind water-soluble vitamins (D-biotin) to form a biotin-streptavidin system (Biotin -streptavidinbinding system, BSA). This system does not affect the activity of biological macromolecules, and has significant advantages in practical applications, such as strong affinity, high sensitivity, strong specificity, good stability, etc., and has the function of signal amplification step by step. It has a wide range of applications in mol...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K1/14B01J20/286B01J20/28B01J20/30B01D15/32C07K14/36C07K1/20
CPCB01D15/327B01J20/28011B01J20/286B01J20/30C07K1/20C07K14/36
Inventor 王骊丽王思尧李维敏王志颖张念喆
Owner NORTHWEST UNIV
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