Process for preparing alpha-1-antitrypsin

An anti-trypsin and IV-1 technology, applied in the field of preparation of α1-antitrypsin, can solve the problems of low product purity and low yield, and achieve the effects of improving economic benefits, reducing costs and improving efficiency

Active Publication Date: 2007-01-24
博晖生物制药(河北)有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Final product purity reaches about 60% and 45% productive rate, and the prod

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] The raw material for the preparation of α1-antitrypsin is the plasma component IV-1 prepared by Kong Shi (Cohn) 6 method, precipitation, and the precipitation of component IV-1 is prepared into a lysate, and the specific method is as follows:

[0027] The precipitate of component IV-1 was dissolved in 10 to 25 times the volume of Tris buffer (W / V) with a pH value of 9.0 to 9.8, stirred and dissolved at 2 to 10°C, about After 60-90 minutes, after mixing and dissolving, adjust the pH value or not adjust the pH value. If the pH value is adjusted, slowly add 0.5M NaOH solution until the pH value is 9.0-9.5. Heat at 40°C for 60-90 minutes to fully dissolve component IV-1 and activate α1-antitrypsin at the same time.

Embodiment 2

[0029] Component IV-1 solution contains various proteins, such as albumin, fibrinogen, globulin, transferrin and denatured protein, etc. α1-antitrypsin must be separated from other miscellaneous proteins, and the following steps can be used:

[0030] Cool the above Cohn component IV-1 solution to 2-10°C, then add 8-12% PEG4000 solution while stirring, after stirring for 30 minutes, adjust the pH value to 5.0 with 0.5M HCl solution ~6.0, continue to stir for 20~40 minutes, and stand at 2~8℃ for several hours or overnight to make it fully precipitate. PEG is a non-ionic precipitant, and its action is mild, so that the activity of α1-antitrypsin is not easily lost. PEG precipitation can remove foreign proteins and viruses; the precipitation is filtered or centrifuged (8000rpm, 30 minutes), the precipitation is discarded, and the PEG supernatant containing α1-antitrypsin is retained.

Embodiment 3

[0032] Adjust the pH value of the polyethylene glycol supernatant containing α1-antitrypsin to neutral with 0.5-1.0 M NaOH, then add Tween 80 / butyl triphosphate (S / D), and the final concentrations are respectively 1% and 0.3% at 24°C±1°C for 8 hours, this treatment method can effectively inactivate lipid enveloped viruses such as HBV, HCV and HIV. This method inactivates the virus and greatly improves the safety of the product.

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PUM

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Abstract

The present invention discloses safe and effective process of preparing alpha-1-antitrypsin through purifying human plasma component IV-1. The present invention extracts alpha-1-antitrypsin with high treating value from the waste human plasma component IV-1, and can raise the utilization rate of valuable human plasma, lower cost and raise economic benefit.

Description

technical field [0001] The present invention relates to a preparation method of α1-antitrypsin, in particular to a preparation method of purifying α1-antitrypsin from components separated from plasma or plasma protein by precipitation and chromatography. Background technique [0002] α1-antitrypsin is a glycoprotein with an approximate molecular weight of 52000-55000 Daltons (Da). This protein is a single chain protein covalently linked by several oligonucleotides, it is an endogenous protease inhibitor, such as trypsin, chymotrypsin, pancreatic elastase, renin, skin collagenase, urea Kinase and Segmented Nuclear Lymphocyte Protease. [0003] Recently, α1-antitrypsin has been used to treat genetic diseases caused by α1-antitrypsin deficiency and acquired α1-antitrypsin deficiency diseases. Administration of α1-antitrypsin can effectively inhibit lymphocyte elastase in the lung. Lymphocyte elastase can destroy foreign proteins in the lung. When α1-antitrypsin is missing, ...

Claims

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Application Information

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IPC IPC(8): C07K1/36C07K14/81
Inventor 卜凤荣宿艳笋陈祥松李昭
Owner 博晖生物制药(河北)有限公司
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