High-performance hydrophobic interaction chromatographic medium taking benzene methanamine as ligand, preparation method and application of medium to protein renaturation and purification

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

Active Publication Date: 2018-12-25
NORTHWEST UNIV
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  • 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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  • High-performance hydrophobic interaction chromatographic medium taking benzene methanamine as ligand, preparation method and application of medium to protein renaturation and purification
  • High-performance hydrophobic interaction chromatographic medium taking benzene methanamine as ligand, preparation method and application of medium to protein renaturation and purification
  • High-performance hydrophobic interaction chromatographic medium taking benzene methanamine as ligand, preparation method and application of medium to protein renaturation and purification

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Embodiment 1

[0037]A preparation method of a high-efficiency hydrophobic interaction chromatographic medium taking bianamine as a ligand, comprising 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, heat u...

Embodiment 2

[0042] A preparation method of a high-efficiency hydrophobic interaction chromatographic medium taking bianamine 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, warm...

Embodiment 3

[0047] A preparation method of a high-efficiency hydrophobic interaction chromatographic medium taking bianamine as a ligand, comprising 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, heat ...

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Abstract

The invention discloses a high-performance hydrophobic interaction chromatographic medium taking benzene methanamine as a ligand. The structural formula of the medium is as shown in the specification.The invention further discloses a preparation method of the high-performance hydrophobic interaction chromatographic medium. The preparation method includes the steps: (1) silica gel activation; (2)epoxy silica gel preparation; (3) epoxide ring-opening preparation of the high-performance hydrophobic interaction chromatographic medium. Besides, the invention further discloses an application of the high-performance hydrophobic interaction chromatographic medium to recombinant streptavidin renaturation and purification. Renaturation efficiency and active recovery rate are high, and recombinantstreptavidin after renaturation is high in purity.

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 amine as 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 molecular b...

Claims

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Application Information

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IPC IPC(8): B01J20/286B01J20/28B01J20/30B01D15/32C07K14/36C07K1/20
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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