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Method for purifying superoxide dismutase from tilapia mossambica blood

A technology of superoxide and tilapia, applied in the direction of oxidoreductase, etc., to achieve the effect of short production cycle, low price and simple purification process

Inactive Publication Date: 2013-01-23
SOUTH CHINA SEA FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Common SOD purification techniques include thermal denaturation, salting-out, organic solvent and chromatography, etc., but the many SOD purification processes and parameters reported so far on different raw materials are not the same. Therefore, it is necessary to develop a method suitable for tilapia blood. The comprehensive utilization of SOD purification process route is particularly important

Method used

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Examples

Experimental program
Comparison scheme
Effect test

example 1

[0017] Take 100ml of clean tilapia red blood cells, add 100ml of double distilled water and mix gently, put it into the ultrasonic instrument, adjust the power to 300W, let it act for 15min, centrifuge at 5000×g for 15min, and collect the upper red blood cell liquid. Slowly add 15ml of absolute ethanol and 10ml of chloroform to the erythrocyte solution successively, stir while adding, after stirring for 15min, let it stand for 30min, centrifuge at 10000×g for 15min, collect the supernatant, and then use a nitrogen blower to remove ethanol and Chloroform.

[0018] The supernatant obtained in the previous step was placed in a constant temperature water bath at 60°C for 15 minutes, cooled rapidly and then centrifuged at 10,000×g for 15 minutes, and the supernatant was collected again.

[0019] Prepare 500ml pH8.0 20mmol / L Tris-HCl buffer as the ion exchange starting buffer (solution A), prepare 1000ml pH8.0 20mmol / L Tris-HCl buffer (containing 1mol / L NaCl) as ion exchange Exchan...

example 2

[0021] Take 100ml of clean tilapia red blood cells, add 200ml of double distilled water and mix gently, put it into the ultrasonic instrument, adjust the power to 400W, let it act for 20min, centrifuge at 5000×g for 15min, and collect the upper red blood cell liquid. Slowly add 10ml of absolute ethanol and 15ml of chloroform to the erythrocyte solution successively, stir while adding, after stirring for 20min, let it stand for 30min, centrifuge at 10000×g for 15min, collect the supernatant, and then use a nitrogen blower to remove ethanol and Chloroform.

[0022] The supernatant obtained in the previous step was placed in a constant temperature water bath at 60°C for 20 minutes, cooled rapidly and then centrifuged at 10,000×g for 15 minutes, and the supernatant was collected again.

[0023] Prepare 500ml of pH8.0 20mmol / L Tris-HCl buffer as the ion exchange starting buffer (solution A), prepare 500ml of pH8.0 20mmol / L Tris-HCl buffer (containing 1mol / L NaCl) as the ion exchang...

example 3

[0026] Take 1000ml of clean tilapia red blood cells, add 2000ml of double distilled water and mix gently, put it into an ultrasonic instrument, adjust the power to 400W, let it act for 20min, centrifuge at 5000×g for 15min, and collect the upper layer of red blood cell fluid.

[0027] Slowly add 100ml of absolute ethanol and 100ml of chloroform to the red blood cell solution while stirring. After stirring for 20 minutes, let it stand for 30 minutes, centrifuge at 10,000×g for 15 minutes, collect the supernatant, and then use a nitrogen blower to remove ethanol and chloroform. .

[0028] The supernatant obtained in the previous step was placed in a constant temperature water bath at 60°C for 15 minutes, cooled rapidly and then centrifuged at 10,000×g for 15 minutes, and the supernatant was collected again.

[0029] Prepare 5000ml of pH8.0 20mmol / L Tris-HCl buffer as the ion exchange starting buffer (solution A), and prepare 5000ml of pH8.0 20mmol / L Tris-HCl buffer (containing 1...

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Abstract

The invention discloses a method for purifying superoxide dismutase (SOD) from tilapia mossambica blood. The method comprises the following steps of: diluting the collected clean tilapia mossambica blood red cell with double distilled water, carrying out membrane rupture treatment by using ultrasonic accessory cells; successively adding absolute ethyl alcohol and chloroform sediment protein, centrifuging, collecting supernate and removing an organic solvent with a pressure blowing concentrator; heating the collected supernate at 60 DEG C, centrifuging and collecting the supernate; rapidly purifying a liquid chromatogram system control by using multifunction protein; and purifying the SOD liquid obtained in the step with anion exchange to obtain high-purity SOD liquid. The purification method is short in time consumption and simple in operating unit and is suitable for industrial production. By the method, the problem of environmental pollution caused by tilapia mossambica blood discharge can be solved, and the additional value of enterprises can be remarkably increased.

Description

technical field [0001] The invention relates to a method for deep processing and utilization of aquatic fish, in particular to a method for purifying superoxide dismutase from tilapia blood. Background technique [0002] Superoxide dismutase (superoxide dismutase, SOD) is an oxygen free radical scavenger derived from the organism, which can maintain the dynamic balance of superoxide anion in the organism and prevent the 2 ﹣ Adverse reactions caused by excessive concentration, known as "human garbage scavenger", has shown unique functions in anti-aging, anti-inflammation, anti-oxidation, anti-radiation and prevention and treatment of tumors, and has been widely used in food, Pharmaceutical and daily chemical industries. At present, there are many researches at home and abroad on the development of SOD by using terrestrial animals and plants (especially pig, cattle, sheep blood and garlic) and microorganisms, but there is little research on the development of SOD from aquati...

Claims

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Application Information

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IPC IPC(8): C12N9/02
Inventor 李来好岑剑伟杨贤庆郝淑贤刘在军吴燕燕黄卉魏涯林婉玲邓建朝周婉君石红
Owner SOUTH CHINA SEA FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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