Method of eliminating endotoxin from interferon preparation

A technology of interferon and endotoxin, which is applied in the field of recombinant protein drug production, can solve problems such as not being able to meet the production requirements of interferon preparations, and achieve the effects of improving drug production level, good specificity, and wide practical range

Inactive Publication Date: 2007-07-04
BEIJING KAWIN TECH SHARE HLDG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The methods of the prior art cannot meet the demand for the production of interferon preparations

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] Preparation of affinity membrane with chitosan as ligand:

[0016] Take 50 cellulose membranes with a diameter of 47 mm, rinse them with a large amount of water, place them in a 200ml beaker, add 40ml epichlorohydrin, 120ml 1M NaOH and 0.19gNaBH , mix well, and let stand overnight at room temperature. Activation; take the activated membrane the next day, wash it with a large amount of water to neutrality, drain, then add 150ml of 1% chitosan (molecular weight: 350,000) solution (prepared by 1% acetic acid solution), overnight at 60°C (16 Hours); the next day, take the activated membrane, wash it with a large amount of water until neutral, drain it, then add 100m10.25% sodium borohydride solution (prepared with 0.1M pH8.5 phosphate buffer), and reduce it for 4 hours at room temperature , take the reduced film, wash it with a large amount of water until neutral, drain and dry. The nitrogen content of the affinity membrane is 0.02%.

[0017] Application effect of affinit...

Embodiment 2

[0020] Preparation of affinity membrane with hexamethylenediamine as ligand:

[0021] Get 320 pieces of cellulose films with a diameter of 47 mm, rinse them with a large amount of water, put them in a 1000ml beaker, add 650ml of oxidizing solution (containing 2% NaIO ), and activate them overnight in a 30°C oven; the next day, take The activated membrane was washed with a large amount of water until it was neutral, drained, and then 800ml of 1% hexamethylenediamine (prepared in 0.1M pH=7 phosphate buffer) was added, and the oven was set at 30°C overnight for cross-linking reaction; the cross-linked membrane was taken the next day. The membrane was washed with a large amount of water to neutrality, drained, added 800ml of 0.6% NaBH4 (prepared with 0.1MpH=9 phosphate buffer) for reduction, and baked at 30° C. overnight. Wash again with plenty of water until neutral, drain and tumble dry. The nitrogen content of the affinity membrane is 2.2%.

[0022] Application effect of affi...

Embodiment 3

[0025] Preparation of affinity membrane with sodium deoxycholate as ligand:

[0026] Take 50 pieces of cellulose membranes with a diameter of 47mm, rinse them with a large amount of water, put them in a 200ml beaker, add 40ml of epichlorohydrin, 120ml of 1M NaOH and 0.19g of NaBH4, mix well, and let stand overnight at room temperature , to activate; the next day, take the activated membrane, wash it with a large amount of water until it is neutral, drain it, then add 125ml of 1% hexamethylenediamine solution (prepared with 1% pH9 phosphate buffer), overnight at 60°C (more than 16 hours) ); Take the membrane the next day, wash it with a large amount of water until it is neutral, drain it, then add 125ml of 0.1M pH6.0 MES buffer, then add 2.5g of sodium deoxycholate and 1.25g of EDC, mix well, and leave overnight at room temperature . Take the cross-linked film, wash it with plenty of water until neutral, drain and dry. That is, the affinity membrane with sodium deoxycholate a...

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PUM

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Abstract

The present invention relates to a production of recombinant protein medicine, in the concrete, it relates to a method for removing endotoxin from interferon preparation. It is characterized by that the chitosan or desoxycholate sodium or hexanediamine is used as affinity ligand, and can be covalently combined on the porous medium, then a dynamic filtration or static adsorption mode can be used to make operation so as to remove endotoxin from interferon preparation, in which the quantity of affinity ligand on the porous medium is 0.02%-3% nitrogen content. Said invention is high in endotoxin removing rate, high in interferon activity recovery and good in specificity.

Description

technical field [0001] The invention relates to the production of recombinant protein medicine, in particular to a method for removing endotoxin from interferon preparations. Background technique [0002] With the increasing maturity of genetic engineering technology, more and more recombinant protein drugs have been developed and utilized. Since the production process involves Gram-negative bacteria and the separation and purification process is complicated, it will cause endotoxin contamination. [0003] Affinity technology is an effective means of separating and purifying biomacromolecules, which mainly utilizes the principle of affinity between ligands and biomacromolecules; the ligands are covalently bonded to porous media to obtain affinity media, which can be used To selectively adsorb endotoxins and achieve the goal of removing endotoxins from recombinant protein solutions; existing affinity ligands and affinity media include: monoclonal antibodies, polyclonal antibo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/555C07K1/14
Inventor 李京华邵英光丛润滋王俊德
Owner BEIJING KAWIN TECH SHARE HLDG
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