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Technique for purifying recombined human interferon alpha 1b

A technology of recombinant human interferon and interferon α, which is applied in the separation and purification of protein and the separation and purification process of recombinant human interferon α1b, which can solve the problems of low production efficiency, reduced total yield, complex process, etc., and achieve production cost reduction , The effect of improving production efficiency and shortening the process cycle

Active Publication Date: 2006-02-01
SHENZHEN KEXING PHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It can be seen that the whole purification process is quite complicated, especially the preliminary purification part, with many steps
[0008] Due to the numerous steps, complex process, and strong acid and strong alkali treatment, the existing production process has the following problems: in the process of protein purification, there will be losses in each step, which will inevitably lead to a reduction in the total yield; long-term treatment , leading to low production efficiency, coupled with a large number of open operations in the preliminary purification, it is easy to cause product pollution and affect product quality; too many steps and a large number of manual operations lead to high material costs and labor costs, and poor process economy
[0011] Is it possible to apply expanded bed adsorption technology to the purification process of interferon? How can the expanded bed adsorption technology be applied to the purification process of interferon to achieve the purpose of simplifying the process and improving production efficiency? The prior art does not seem to take this into account, nor does it provide a corresponding solution

Method used

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  • Technique for purifying recombined human interferon alpha 1b
  • Technique for purifying recombined human interferon alpha 1b
  • Technique for purifying recombined human interferon alpha 1b

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1: Expanded bed adsorption gel selection and chromatography parameter optimization

[0039] The operation of EBA requires special chromatography columns to produce a stable expanded bed. In the initial stage of process development, experiments were first carried out in the form of packed beds. The reason for using a packed bed for gel selection and parameter optimization is that although the adsorption effect of the expanded bed is better, the conditions required for gel adsorption in the two methods are consistent and have no effect on the selection of gels and chromatographic parameters. And it can save the investment of small-scale expansion bed system. With a packed bed, since particulate impurities cannot be removed, the sample needs to be centrifuged first to obtain clarified material.

[0040] the gel

Experiment number

Column

mm*cm

buffer system

Loading amount

Elution point

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type

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Embodiment 2

[0044] Example 2: Exploration of interferon purification process using STREAMLINE DEAE packed bed column

[0045] On the laboratory scale, use XK16×10 column in the form of packed bed, choose STREAMLINE DEAE and Tris buffer solution with pH 7.4, and use the same batch of bacterial homogenate as the old process to produce pure interferon along the new process route. In the comparison between the new and the old process, the data of the old process is taken from the actual production.

[0046] Experimental Materials:

[0047] 1. Equipment: chromatography column: XK16×10, gel: STREAMLINE DEAE, chromatography system: Pharmacia GP-250 gradient controller

[0048] 2. Buffer: Equilibrium: pH7.4 50mM Tris-HCl, eluent: 0-0.5MNaCl pH7.450mM Tris-HCl Experimental procedure:

[0049] 1. Packing: Pack the column in the equilibrium buffer system at a flow rate of 20ml / min, and the volume of the column is 20ml

[0050] 2. Equilibrium: use 10ml / min flow rate, equilibrate the column bed wit...

Embodiment 3

[0058] Example 3: Establishment of expanded bed adsorption manual purification system

[0059] In the process of scaling up from the experimental scale to the production scale, it is faced with the problem of establishing an expanded bed adsorption purification system. Based on the consideration of reducing investment, we worked with the chromatography column supplier to establish a manual purification system for expanded bed adsorption.

[0060] According to the principle diagram of the expanded bed adsorption operation in Figure 2, the steps of the expanded bed adsorption include equilibrium expansion, sample loading, column cleaning, sedimentation and elution, and column regeneration. In order to realize the two functions of expansion and chromatography at the same time, a double pump system must be used. One pump is responsible for the expansion and settling of the column bed, and the other pump is responsible for equilibration, sample loading, elution, and regeneration. ...

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Abstract

A process for purifying recombinant human interferon alpha 16 includes such steps as breaking the engineered bacteria able to express human interferon alpha 16 in buffering Tris-HCl solution, adsorbing on expanding bed by use of anionic gel, buffering Tris-HCl solution and the eluting liquid containing NaCl and Tris-HCl, affinity chromatography by use of the gel coupled by its monoclonal antibody, PBS as balancing liquid and washing liquid, Gly-HCl as eluting liquid, Tris-HCl as regeneration liquid and NaAc-Hac as buffering liquid, and gel filtering by use of Sephacryl gel and PBS as buffering system.

Description

technical field [0001] The present invention relates to a pharmaceutical preparation containing peptides, in particular to a method for separating and purifying proteins, in particular to a process for separating and purifying recombinant human interferon α1b. Background technique [0002] Interferon has a wide range of antiviral activities. In 1957, Isaacs and Lindenmann discovered this substance when they were studying the interference phenomenon of influenza virus: after acting on cells with inactivated influenza virus, the cells will produce a soluble substance that makes the live virus Cell reproduction is disturbed, so it is called interferon (Interferon, abbreviated as IFN). Interferon has three major functions: inhibiting virus reproduction activity, inhibiting cell division activity and inhibiting immunomodulatory activity. [0003] Interferons are a class of proteins that regulate cellular function. According to the different antigen specificity and molecular str...

Claims

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Application Information

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IPC IPC(8): C07K14/56C07K1/36
Inventor 何询于德强张为
Owner SHENZHEN KEXING PHARMA CO LTD
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