JPS5855123B2 - RBA Noseizouhou - Google Patents

Description

[Detailed description of the invention] The present invention relates to a method for producing a novel physiologically active substance RBA.

The present inventors discovered that a substance with special physiological activity exists in rice bran, and as a result of various studies on the isolation of this substance, they succeeded in extracting and collecting the substance with a soluble solvent. The substance was named RBA.

The present invention was completed based on the above new findings,
This method of producing RBA is characterized by extracting rice bran with a solvent to collect RBA.

RBA obtained by the method of the present invention has an affinity for specific sugar chains, especially N-acetylglucosamine, and has a affinity for specific sugar chains, especially N-acetylglucosamine, and is suitable for cell surfaces [e.g., red blood cell surface, white blood cell (including lymphocyte) surface, free adipocyte surface, normal cell surface. It binds well to sugar chains on the surface of certain types of cells, particularly on the surface of certain types of cells, and has hemagglutinating effects, insulin-like effects, cancer cell aggregating effects, and cell growth-promoting effects. It is useful as a diabetes treatment agent, a cancer diagnostic agent, etc.

RBA obtained by the method of the present invention is a vegetable protein and has the following properties.

(1) The molecular weight is about 13,000 to 17,000, and 6M
- In systems such as guanidine-1M-propionic acid, it is no longer subunitized.

(2) It has two or more sugar binding sites.

(3) Equal points are 7-10.

(4) Sugar content measured by phenol-sulfuric acid method and gas chromatography is about 0.02-1.0%.

(5) The number of moles of each constituent amino acid per 100 moles of all the amino acids constituting RBA recovered by hydrolysis with 6N-hydrochloric acid at 110°C for 24 hours is as shown in the following table.

(6) Agglutinate rabbit blood cells with O,OO3~0.005 m9/'ml.

Rice bran is used as the raw material in the method of the present invention, and rice bran herein also includes rice bran-containing substances such as brown rice.

The method of the present invention is carried out by extracting and collecting rice bran with a solvent in which the target substance is soluble.

The rice bran may be subjected to the extraction and collection operation as it is, or may be ground if desired.

Further, more preferable results can be obtained if the sample is degreased with a lipophilic organic solvent such as acetone or chloroform before being subjected to the extraction and collection operation.

Any solvent may be used for extraction and collection as long as it has the ability to elute RBA, such as an aqueous solvent (e.g.
Examples include saline solution, phosphate buffer, etc.), polar organic solvents (eg, methanol, ethanol, gropanol, etc.), and water-containing polar organic solvents (eg, alcohol solution, dilute acetic acid, etc.).

The extraction solvent may further contain a solubilizing agent such as a surfactant (eg, sodium deoxycholate, sodium dodecyl sulfate, etc.).

Further, after extraction and collection with a solvent, purification treatment may be performed as necessary by a method known per se, such as an ion-exchange cellulose method, a gel filtration method, an affinity chromatography method, etc. In particular, ovomucoid, which is a type of glycoprotein, A method of purification by affinity chromatography using a chemically inactivated column support is advantageously used.

Furthermore, the extracted liquid may be salted out with ammonium sulfate or the like before being subjected to the purification treatment.
It is advantageous to separate the BA as a precipitate, dissolve it in a small amount of the aforementioned solvent, and then subject it to the purification treatment.

Example 1.5 liters of 0.9% saline solution (p
Add H5,0) and stir at 4°C overnight.

After removing the residue by filtration and centrifugation, add ammonium sulfate to the supernatant to 60% saturation.

After standing for several hours, centrifuge and collect the precipitate.

Dissolve the precipitate in 0.1M acetate buffer (pH 5,0),
This supernatant was placed on an Ovomucoid Sepharose column (10ml) that had been sufficiently buffered with the same buffer solution.
) and then washed with the same buffer.

After thoroughly washing away unadsorbed proteins, the adsorbed RBA was eluted with 0.1 M acetic acid, and a small amount of inactive protein was removed using CM-cellulose (pH 8,0) chromatography. of RBA was obtained.

The RBA thus obtained was subjected to disk electrophoresis (7.5%
Polyacrylamide gel, pH 4.0 for 60 minutes (577
1.A) Electrophoresis, RBA stained with Comassie Blue) showed uniform bands.

(Note) Preparation of ovomucoid sepharose; albumen, Journal of Biological Chemistry,
According to the method described in Vol. 171, No. 565-581 (1947), ovomucoid was obtained by fractionation with acidic acetone, and the ovomucoid was purified by Men's in Enzymology.
It was bound to Sepharose activated with cyanodiene bromide according to the method described in Vol. 22, pp. 345-378 (1972).

Claims (1)

[Claims] 1. A method for producing RBA, which comprises extracting rice bran with a solvent to collect RBA.