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Simplified method for extracting purified superoxide dismutase

A technology of superoxide and extraction method, applied in the directions of oxidoreductase, peptide preparation method, chemical instrument and method, etc., can solve the problems of restricting SOD extraction and wide application, strong operation technology, high production cost, and achieve the operation method. Simple and easy to implement, short process flow and versatile effect

Inactive Publication Date: 2005-12-14
SHANDONG AGRICULTURAL UNIVERSITY
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Problems solved by technology

Because of the reported traditional method for preparing SOD from animal cells (erythrocytes, liver cells, etc.), after removing foreign proteins with organic solvents and thermal denaturation methods, SOD is precipitated with acetone, and the yield is low due to the small amount of precipitation.
In addition, the refining of the crude product should use ion exchange method, and the eluent should use ultrafiltration concentration method, resulting in high production cost and strong operation technology, which restricts the extraction and wide application of SOD

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  • Simplified method for extracting purified superoxide dismutase

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Embodiment

[0010] In a clean and sterilized container, add 1 liter of freshly collected blood per 100 ml of 3.8% (V / V) trisodium citrate aqueous solution, and mix immediately. Centrifuge at 3500r / min for 15min to collect blood cells. Add 2 times the volume of normal saline to float and wash the blood cells twice, and centrifuge at 3000r / min for 8 minutes to obtain blood cells. After freezing 1000ml of blood cells, add an equal volume of distilled water and stir for 1h (ice-water bath). Then add frozen 95% ethanol-chloroform (0.25-0.15 times) volume (500+300) ml mixed solution, continue stirring for 15 min, centrifuge at 4000 r / min for 30 min, and discard the precipitate. The collected supernatant is about 1150ml, adjust the pH to 5.5-6.0 with dilute hydrochloric acid, freeze to below 0°C, slowly add an equal volume of pre-cooled acetone under stirring, and then add zinc and magnesium ion mixture to a concentration of 1mol / L The final concentration is about 23ml of 1% (V / V), centrifuged...

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Abstract

For fresh anticoagulated blood, add twice acetone to the initial extract of one liter of blood, the amount of crude product precipitated is small, and the precipitation of superoxide dismutase (SOD) is not complete. Use a small amount of Zn 2+ , Mg 2+ Ions and adjust PI, add double acetone to precipitate superoxide dismutase (SOD), and dissolve the precipitate with weak alkaline aqueous solution. Add acetone to the water-soluble supernatant to precipitate SOD, and vacuum-dry it below 25°C to obtain a light green product. According to application requirements, this method can prepare products with different purity, and the yield is 1.1-1.5g / L red blood cells. This method avoids the complex processes of thermal denaturation, ion exchange, and ultrafiltration concentration, and the productivity is greatly improved compared with the traditional method. This method is simple and easy to implement, the product has high activity, wide application and remarkable benefit.

Description

(1) Technical field [0001] The invention relates to an extraction method for simple purification of superoxide dismutase (SOD). (2) Background technology [0002] Superoxide dismutase (SOD) is available in the United States, Germany, Australia and other countries. A series of studies on the chemistry, pharmacology, clinic and mechanism of SOD have found that the existence of SOD is related to anti-aging and tumorigenesis, autoimmune diseases and radiation protection. At present, it is mainly used for inflammation, rheumatoid arthritis, chronic polyarthritis and inflammation after radiation therapy. In my country, it is mostly used in cosmetics, alcohol, food, etc. Because of the reported traditional method for preparing SOD from animal cells (erythrocytes, hepatocytes, etc.), after removing foreign proteins with organic solvents and thermal denaturation methods, SOD is precipitated with acetone, and the yield is low due to the small amount of precipitation. In addition, t...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K1/30C12N9/02C12N9/08
Inventor 杨景芝孙衍华
Owner SHANDONG AGRICULTURAL UNIVERSITY