Method for extracting superoxide dismutase from animal blood

A technology of superoxide and dismutase, applied in the direction of oxidoreductase, etc., can solve the problems of physical damage to operators, serious environmental pollution, complicated process, etc., and achieve the effects of low cost, increased ionic strength, and improved yield

Inactive Publication Date: 2013-12-11
浙江迈康生物工程有限公司
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  • Summary
  • Abstract
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  • Application Information

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Problems solved by technology

The disadvantage of this process is that the process is complicated, the cost is high, and organic solvents such as chloroform an

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0022] Example one

[0023] 1000L of fresh pig blood was added to 50L of 8% sodium citrate solution for anticoagulation, and 450L of red blood cells and 600L of plasma were separated by continuous flow centrifuge. The plasma is spray-dried to become plasma protein powder. The red blood cells are pumped into the reaction tank, while adding 1200L of distilled water and stirring with a mixer for 30 minutes, so that the red blood cells are fully broken and hemolyzed. After heating to 60°C for 15 minutes, add 37Kg of sodium chloride with stirring, heat to 65°C and keep for 30 minutes, then to 72°C, continue heating for 20 minutes, and quickly water-cool to below 30°C. Press filtration with 800 mesh filter cloth to obtain a total of 1300L of crude SOD extract with a unit activity of 220U / ml. The crude SOD extract is pumped into an ultrafiltration storage tank, and is pretreated by an ultrafiltration membrane to intercept macromolecular proteins and cell membrane fragments with a mole...

Example Embodiment

[0024] Example two

[0025] 2000L of fresh duck blood was anticoagulated with 100L of 10% sodium citrate solution, and 830L of red blood cells and 1270L of plasma were separated by continuous flow centrifuge. Place the red blood cells in a reaction tank, add 1660L of distilled water, and homogenize at high pressure for 20 minutes to completely break the red blood cells. Continue to stir and heat up to 60°C for 15 minutes, add 58Kg of sodium chloride, heat up to 65°C and keep holding for 30 minutes, then increase the temperature to 70°C, keep holding for another 20 minutes, and quickly discharging and cooling. 800 mesh filter cloth mechanical pressure filtration to obtain 1900L of crude SOD extract, unit activity 270U / ml. The crude SOD extract is pumped into an ultrafiltration storage tank, and is pretreated by an ultrafiltration membrane to intercept macromolecular proteins and cell membrane fragments with a molecular weight greater than 1 million, and then repeatedly ultrafilte...

Example Embodiment

[0028] Example three

[0029] A method for extracting superoxide dismutase from animal blood includes the following steps:

[0030] A. Blood separation: Take fresh chicken blood, add 10% sodium citrate solution by mass concentration, the volume of sodium citrate solution is 6.5% of the blood volume, after anticoagulation treatment, collect red blood cells by continuous flow centrifuge, pump Into the hydrolysis reaction tank;

[0031] B. Hemolysis and broken cells: add 1.5 times the volume of purified water to the red blood cells collected in step A, and homogenize the high pressure to completely break the red blood cells to obtain hemolysis of the red blood cells;

[0032] C. Thermal denaturation: After heating the erythrocyte hemolysis to 60°C for 10 minutes, add NaCl to increase the ionic strength of the solution to the erythrocyte hemolysis until the NaCl concentration is 0.55mol / L, and continue heating to 65°C. After holding for 50 minutes, the temperature is increased to 75°C fo...

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Abstract

The invention discloses a method for extracting superoxide dismutase from animal blood. The method comprises the following steps: A. blood separation; B. hemolytic cell breaking; C. heat denaturation; D. filter pressing; E. ultrafiltration and concentration; F. precipitation for removing hybrid protein. Such organic solvents as chloroform, ethanol and acetone, and the like, are completely eliminated, and a rough product SOD with high activity is simply and conveniently separated out through a heat denaturation and metal salt precipitation method according to the structure characteristics and physicochemical properties of SOD molecules, which is a revolutionary improvement of SOD production. Since no inorganic solvent is used, the method is low in cost and is more suitable for industrial production. Hemoglobin is removed by thermal denaturation in the invention, so that hemolysis can be realized at normal temperature, the operation is simpler and more convenient and the energy consumption is reduced.

Description

(1) Technical field [0001] The invention relates to a method for extracting superoxide dismutase from animal blood. (2) Background technology [0002] Superoxide dismutase (SOD) is a metalloenzyme ubiquitous in organisms that can scavenge superoxide anion free radicals. It has many functions such as anti-inflammation, anti-radiation, and anti-tumor. It has been widely used in medicine, food, health food and cosmetics. [0003] At present, the technological process of extracting superoxide dismutase from animal blood is mainly blood anticoagulation - washing with normal saline - low temperature hemolysis - chloroform ethanol extraction of hemoglobin - potassium phosphate extraction - heat denaturation - acetone precipitation - - ion exchange chromatography - molecular sieve column chromatography. The disadvantage of this process is that the process is complicated, the cost is high, and organic solvents such as chloroform and acetone have strong toxicity, which seriously pol...

Claims

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Application Information

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IPC IPC(8): C12N9/02
Inventor 龚兴国秦勇聂颢韩斌应育权
Owner 浙江迈康生物工程有限公司
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