JPS61100524A - Method of separating saponin and flavone - Google Patents

Abstract

PURPOSE:To separate the titled compound efficiently, by treating a leguminous plant with protease so that an insoluble substance is separated efficiently, and treating the prepared liquid phase part by an ultrafilter. CONSTITUTION:At least part (e.g., flower, fruit, seed, wood part, rhizome, etc.) of leguminous plant is ground, extracted with a hot organic solvent such as n-hexane, etc., defatted, the dried defatted extract is treated with protease (e.g., one obtained from a variant of Bacillus subtilis) in a diluted aqueous solution (e.g., 7-10pH). After the treatment is over, the treated material is separated into insoluble substances containing fibers and slightly digestible protein and a liquid phase part by filtration, etc. The liquid phase part is separated into a concentrated part and a permeated part by an ultrafilter (e.g., cellulose nitrate, noylon, polystyrene, etc.), the permeated part is treated with a nonpolar or slightly polar adsorbing resin, saponins and flavones are eluted with a polar solvent such as methanol, etc., from the resin and obtained.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] This invention relates to an improvement in a method for separating saponins and flavones from leguminous plants.

[Conventional technology]

The main components of leguminous plants include fiber, saponins,
As a method for separating flavones, sugars and proteins, there is a method of extracting defatted soybeans with hot polar mM, concentrating the extract, re-extracting the concentrate with n-butanol, and concentrating this extract. , for example, JP-A No. 56-64000, No. 5
It has been proposed in No. 6-73025, No. 56-160981, etc.

However, these methods have not yet been satisfactory in terms of the purity of the extract and the economic efficiency of extraction.

As a result of various studies aimed at improving the extraction and separation method for the above-mentioned components, the inventors proposed a method that combines treatment with an adsorption resin in addition to treatment with an alkali (Japanese Patent Application Laid-open No. 33232/1983). As a result of further research, they discovered that by using protease during alkaline treatment and using a limit filter membrane before adsorption resin treatment, products with even higher purity could be obtained, and this led to the completion of this invention. be.

[Structure of the invention]

That is, the present invention involves treating at least a portion of a leguminous plant with a protease in a dilute alkaline aqueous solution after defatting if necessary, removing insoluble substances including fibers and indigestible proteins, and limiting the liquid phase. Saponins, characterized in that they are divided into a concentrated part and a permeated part by an outer membrane, the permeated part is treated with a non-polar or slightly polar adsorption resin, and the saponins and flavones adsorbed to the resin are eluted with a polar solvent. and a method for separating flavones.

The legume family plant or a part thereof used as a raw material may be flowers, flower buds, fruits, seeds, leaves, xylem, root violets, root nodules, etc., or the whole plant. Moreover, it may be fresh or dried. Usually, crushed plant matter is used as n-
Degrease by hot extraction with an organic solvent such as hexane, and use the dried product after defatting.

The saponins include those consisting of sapogenins such as triterpenoids or steroids and saccharides such as glucose, arabinose, galactose or glucuronic acid. An example of a typical legume saponin is glycyrrhizin (glycyrrhetinic acid +
glucuronic acid), soybean saponin contained in soybeans, and alfalfa saponin contained in cornstarch.

The above flavonoids include flavones, flavonols,
7 lavanones, flavanols, inflavones, and their hydroxy, methoxy, methyl or methylene dioxy derivatives, as well as their glycosides. An example of such 7-labonoids is platol (7-hydroxy-4'-methoxyflavone) contained in Akatsumegusa.
Acadiin (5,7-hydroxy-4'-methoxyflavone (acacetin)-7-
rhamno-glucoside) and robinetin (7,3,4゜5-te)-1-ni)'07 lavanol), liquiritin (7, easy hydroxy 7-ravanone (liquiritigenin)-glucoside) contained in Ganzo roots Genistin (5,7,4'-trihydroxyisoflavone (genistein)-glucoside) contained in soybeans, daidzin (7,4'-dihydroxyinflavone (daidzein)-glucoside) and tatoin (
5,4'-dihydroxy-8-methylisoflavone),
These include Φ-pabutigenin (7-hydroxy-3', 4'-methylenedioxyinflavone) contained in Baptisia tinctoria.

The method of this invention is preferably carried out as follows. First, the aforementioned plant (preferably a defatted and dried product) is treated with protease in a dilute alkaline aqueous solution (for example, pH 7-10). As the protease, one obtained from a mutant strain of Bacillus subtilis or a commercially available product is used. This treatment is preferably carried out for several hours under slight heating.

After treatment, the liquid phase is separated from insoluble substances containing fibers and indigestible proteins by suitable separation means such as filtration or centrifugation.

The liquid phase given here is then divided into a part that is concentrated by the ultrafiltration membrane and a part that passes through the membrane.

Examples of the ultrafiltration membrane include cellulose acetate (preferably with a degree of acetylation of about 40%), nylon, cellophane, polyvinyl alcohol (preferably heat-treated), polystyrene, polyethylene, aromatic polyamide, hydrazide copolymer, etc. Known ultrafiltration membranes and commercially available products are used. Ultra-retraction can be performed using commonly used equipment. - During filtration, a pressure of several atm, tens of atm or about one hundred atm is applied depending on the ultrafiltration membrane used. Appropriate concentration ratio is several times to several tens of times.

Next, the obtained permeable part is treated with a non-polar or slightly polar adsorption resin, either as it is or after being made acidic. As such an adsorption resin, for example, an Ichisaka product whose base material is a styrene/vinylbenzene copolymer or an acrylic ester polymer is used.

By treatment with an adsorption resin, saponins and flavones are adsorbed to the adsorption resin, and are eluted with a polar solvent. Methanol, ethanol, etc. are used as the polar solvent. Separation of saponins and flavones from the eluent is carried out by conventional methods such as concentration, crystallization, non-solvent treatment, etc.

Note that when the insoluble substance obtained after the protease treatment is pH adjusted to neutral, washed with water, and dried, a solid consisting of high-quality fiber and indigestible protein is obtained. This can be used, for example, as a raw material for producing diet foods.

In addition, when the concentrated portion obtained by the ultrafiltration membrane treatment is concentrated to dryness, a water-soluble protein hydrolyzate (molecule 41) 3
0000) is obtained. This is used as a taste improver.

Furthermore, sugars can be collected from the effluent that was not adsorbed during the adsorption resin treatment by concentrating it.

〔effect〕

According to this invention, the leguminous plant body or its defatted dried product is first treated with protease to hydrolyze digestible proteins, so that digestible proteins and indigestible proteins can be efficiently separated in the next step of separating insoluble substances. Can be divided.

Furthermore, when the liquid phase after removing insoluble substances is treated with a limit-filtration membrane, the protein hydrolyzate resulting from the protease treatment and the desired saponins and flavones are efficiently separated. Therefore, according to the present invention, digestible proteins, which were conventionally separated insufficiently and mixed with insoluble substances, saponins and flavones, causing harm such as browning as impurities, are efficiently removed, and the product can be continuously improved. In addition to improving the quality, the ultraviolet ray itself is collected as a concentration part through membrane filtration treatment, and is utilized as a useful substance. Thus, the present invention has excellent advantages.

〔Example〕

This invention will be explained below with reference to Examples.

Example 1 72% of water was immediately added with sodium hydroxide to adjust the pH to 7-10, the fatigue temperature was maintained at 50-60°C, and 8% of protease,
After adding 8.0 hours of defatted soybeans and reacting for 3 hours while stirring at 300 rpm, the liquid phase was separated into the same liquid using a separator [first, a screw press was used, and then the liquid phase was transferred to a centrifuge (aooo
rpm), and the solid phase is combined with the previous solid phase] to separate into a solid phase and a liquid phase. The solid phase (26,9S') has a pH of
After conditioning and washing with hot water (40.0 kg), dry. In this way, crude fiber/indigestible protein 2.0 K9 is obtained. The results of analyzing this product according to the food analysis method are as follows. Water 3.9%, protein I4O, 9c13, lipid 3.
7%, fiber 22.7%, carbohydrates 26.0%, ash 2.8%.

Next, an ultra-filter membrane DUY-H (0.7rIl, manufactured by Daicel Chemical Industries) was attached to the pleated test device 5R-5 (manufactured by Daicel Chemical Industries, Ltd.), and the liquid phase portion 92. 411 (a combination of the liquid phase of solid-liquid separation and hot water washing. This solution is once centrifuged (3000 rpm), the liquid phase is subjected to ultrapolar filtration, and the solid phase is combined with the above solid phase. ] was ultrafiltered.

Concentrate@(48,5~) is dried and crude hydrolyzed protein 1.
Gain 7 Kg. Analyzing this as above, we get
Water content is 3.2%, protein content is 59.1%, fat content is 20.7%, fiber content is 0.7%, carbohydrate content is 4.3%, and ash content is 10.2%.

10 liters of adsorption resin with nylbenzene as the resin matrix was dispersed in 15 liters of water, packed into a column with an inner diameter of 134 mm, and the above-mentioned passing liquid was injected from the top at a flow rate of 200 d/
Pass through and adsorb at a speed of 1 minute. Wash with water until the eluate becomes colorless and separate into adsorbed and non-adsorbed parts.

20 liters of methanol is passed through the adsorption section at a rate of 100 d/min to elute the adsorbed material, which is concentrated and dried under reduced pressure F to obtain crude flavones and crude saponins (44S').

Completion of elution was confirmed by thin layer chromatography (carrier Nisilica gel 60, developing solvent: lyloform/methanol/water-s).
:4:l, Detection: 105℃ after spraying 50% sulfuric acid aqueous solution
(heated for 5 minutes). Comparison with a standard product on a thin layer shows that these are mostly composed of flavones and saponins.

Next, the pH of the non-adsorbed area was adjusted to neutral using sodium hydroxide and acetic acid, and then concentrated to dryness under reduced pressure.
l Skewer Kp. The analysis of this stuff consists mostly of carbohydrates.

Example 2 A mixture of saponins and flavones 1 was prepared in the same manner as in Example 1 using shredded licorice (marked with l o) instead of defatted soybeans.
．． It's a disgrace to s Kp. This was washed with acetone and fractionated into soluble flavones and insoluble saponins.

Claims (3)

[Claims] (1) At least a part of the leguminous plant is defatted if necessary, and then treated with protease in a dilute alkaline aqueous solution to remove insoluble substances including fiber and indigestible proteins.
The liquid phase is divided into a concentrated part and a permeated part by an ultrafiltration membrane, the permeated part is treated with a non-polar or slightly polar adsorption resin, and the saponins and flavones adsorbed to the resin are eluted with a polar solvent. A method for separating saponins and flavones. (2) Adjust the insoluble material obtained after protease treatment to pH
The method according to claim 1, wherein fiber and indigestible protein are collected by conditioning, washing with water and drying. (3) The method according to claim 1, wherein the protein hydrolyzate is collected from the concentrated portion obtained by ultrafiltration membrane treatment.