CN1477197A

CN1477197A - Method for extracting superoxide dismutase from plant

Info

Publication number: CN1477197A
Application number: CNA021299129A
Authority: CN
Inventors: 秦和沅
Original assignee: Individual
Current assignee: Individual
Priority date: 2002-08-21
Filing date: 2002-08-21
Publication date: 2004-02-25

Abstract

The method for extracting superoxide dismutase from thallophyte is characterized by that its precipitating agrnt adopts the aqueous solution of sodium silicate (or sodium wolframate) and calcium carbonate, and its elutriant adopts distilled water and mild alcohol, and its purification adopts chromatographic equipment, and before precipitation of supernatant fluid an active carbon filtering process is added.

Description

A kind of method of from plant, extracting superoxide-dismutase

The present invention relates to a kind of method of from thallophyte, extracting superoxide-dismutase.

At present, domestic superoxide-dismutase biochemical product mainly is to extract from the red corpuscle of animal blood and animal livers.Extracting superoxide-dismutase from higher plant began one's study in nineteen nineties, from the Plantula Brassicae chinensis leaf, extract superoxide-dismutase as what " functional foodstuff " first roll of China Light Industry Press's publication was introduced, its method is plant to be pulverized the supernatant liquor that obtains after making beating is filtered add ammonium sulfate, leave standstill the back throw out that obtains of centrifugal treating and add potassium phosphate buffer again and carry out gradient elution, purifying with ion exchange column or gel column must the superoxide-dismutase goods after concentrated, freezing, drying.From plant, extract superoxide-dismutase, though it is more extensive than from blood, extracting raw material sources, method is fairly simple, and cost is low, but carry out gradient elution owing to overlong time with potassium phosphate buffer in the leaching process, and ammonium sulfate is bigger to the active ingredient damage of superoxide-dismutase, practicality is not strong, so rest on laboratory stage, fail to form the scale that batch production is produced always.

The object of the present invention is to provide and a kind ofly can shorten extraction time, can improve the superoxide-dismutase product activity again, and can adapt to the method for from plant, extracting superoxide-dismutase that batch production is produced.

The present invention extensively is present in the histocyte of animals and plants and microorganism according to superoxide-dismutase, it is a kind of attribute of water soluble protein, thereby can make water soluble protein solidify characteristics with water sepn according to sodium ion and calcium ion, and low alcohol has the principle of sterilization, sterilizing function, the technological design that to extract superoxide-dismutase from plant is: precipitation agent adopts water glass (or sodium wolframate) and calcium carbonate aqueous solution, eluent adopts distilled water to add low alcohol, and gradient elution is designed to carry out chromatography with chromatographic apparatus.That is: with plant pulverizing, pressure juice, in juice, add Acid precipitation, the separating plant leaf protein, obtain supernatant liquor, behind supernatant liquid filtering, Ghana adds calcium again, make the aqueous soluble protein matter precipitation in the supernatant liquor, the throw out centrifugal treating is gone to carry out sterilization and wash-out with low alcohol finishing distilled water behind the clear water, obtain rough superoxide-dismutase, thick enzyme carries out chromatography and separates through chromatographic apparatus, obtains refining superoxide-dismutase emulsion, through freezing impurity elimination, sublimation drying, what obtain is the refining finished product of superoxide-dismutase.

Since adopted technique scheme, the present invention have that technology is simple, cost is low, extraction yield is high, little to the active degree of injury of superoxide-dismutase product, process period short and characteristics such as suitable batch production production.After tested, reach more than every milligram 4000 unit, and functional, stable, edible safety all is better than the superoxide-dismutase that extracts from animal blood and liver with the superoxide-dismutase product activity of this explained hereafter.

Below the present invention is further illustrated: a kind of thallophyte is pulverized, presses juice, in juice, add acid and precipitate, the separating plant leaf protein, supernatant liquor; With the supernatant liquor active carbon filtration, filter partial impurities, pigment and inorganic salt; The aqueous solution that adds water glass (or sodium wolframate) and lime carbonate in the supernatant liquor after filtering, make aqueous soluble protein matter precipitation, the effect of water glass (or sodium wolframate) and lime carbonate is the activity that can make the aqueous soluble protein matter rapid solidification precipitation in the supernatant liquor and not damage aqueous soluble protein matter, with add ammonium sulfate and compare, sedimentation time can shorten about 2/3, and cost reduces; Throw out is left standstill centrifugal treating to be gone to carry out repeatedly rinsing with distilled water behind the clear water on upper strata, then with the low alcohol capable wash-out that progresses greatly, thereby obtain rough superoxide-dismutase, the distilled water rinsing does not damage the superoxide-dismutase product activity, low alcohol gets final product disinfection, but elution time is shortened in impurity elimination again, also do not damage the effective ingredient of superoxide-dismutase, and be suitable for batch production production with the elution process that distilled water adds low alcohol; Thick enzyme is carried out chromatography with chromatographic apparatus, obtain purified superoxide-dismutase emulsion, the chromatographic apparatus superoxide-dismutase product of can purifying also separablely goes out wherein contained multiple proteins; Purified superoxide-dismutase emulsion is carried out freezing impurity elimination, and sublimation drying promptly gets the finished product of superoxide-dismutase.

Embodiment: remaining supernatant liquor behind the utilization string leaf rosinweed extraction plant leaf protein, with adding concentration behind the active carbon filtration is that 1～3% water glass (or sodium wolframate) aqueous solution precipitates, adding concentration again and be 0.8～2% calcium carbonate aqueous solution leaves standstill after 8～15 minutes and removes clear water, the throw out that obtains carries out repeatedly rinsing with distilled water earlier, be 3～5% the following low alcohol of the 30 degree capable wash-out that progresses greatly with concentration then, get rough superoxide-dismutase, thick enzyme is carried out chromatography with chromatographic apparatus, get purified superoxide-dismutase emulsion, through freezing, impurity elimination, after the drying, get crystalline plant superoxide-dismutase finished product.

Claims

1, a kind of method of from plant, extracting superoxide-dismutase, complete processing precipitates for the supernatant liquor adding precipitation agent ammonium sulfate that thallophyte pulverizing making beating filtration is obtained produces it, throw out purified with ion exchange column (or gel column) add potassium phosphate buffer again and carry out gradient elution and get superoxide-dismutase emulsion, after concentrated, freezing, drying the superoxide-dismutase finished product, it is characterized in that precipitation agent adopts the aqueous solution of water glass (or sodium wolframate) and lime carbonate, elutriant adopts distilled water to add low alcohol, purifies and adopts chromatographic apparatus.

2, the method for extracting superoxide-dismutase from plant according to claim 1 is characterized in that once filtering with activated carbon before supernatant liquor adds the aqueous solution precipitation of water glass (or sodium wolframate) and lime carbonate.

3, according to claim 1, the 2 described methods of extracting superoxide-dismutase from plant, it is characterized in that the concentration of precipitation agent water glass (or sodium wolframate) aqueous solution is 1～3%, the concentration of calcium carbonate aqueous solution is 0.8～2%.

4, according to claim 1, the 2 described methods of extracting superoxide-dismutase from plant, the alcoholic strength that it is characterized in that the low alcohol of elutriant is 25～30 degree, and concentration is 3～5%.