JPS6212792A - Isolation of astragali radix saponin - Google Patents

Abstract

PURPOSE:To isolate a pure saponin mixture containing a novel saponin useful for preventing and treating arteriosclerosis, by concentrating an extract solution of Astragali Radix with a lower alcohol and treating the resultant extract by a special method. CONSTITUTION:Astragali Radix is first extracted with a lower alcohol preferably while warming or heating, and the resultant extract solution is concentrated and dissolved in a lower alcohol to give a solution, which is treated with an adsorbent, e.g. silicon gel. A fraction eluted from the above-mentioned adsorbent is subsequently subjected to chromatography, e.g. reversed phase silica gel chromatography, to isolate a saponin mixture. The above-mentioned mixture contains 10 or more novel compounds consisting of a compound expressed by formula I (R<2> is beta-D-glucopyranosyl, and R<3> is 2,3,4-tri-O-acetyl-beta- D-xylopyranosyl when R<1> is H) and a compound expressed by formula II.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to saponins isolated from Astragalus aspera and a method for isolating the same.

Leguminus (yellow g) referred to in this invention belongs to the leguminous family Leguminae.
Astragalusmembra of osae
means the root of naceus Bunge or other congenerous plants. Astragalus has been used as a tonic since ancient times as a tonic, cardiotonic, diuretic, antiperspirant, and antihypertensive agent. It is known that the ingredients of Astragalus include isoflavonic acids, isoflavanoic acids, betaine, piperidinic acid, and sucrose. However, it contains saponin glycosides:
It is completely unknown.

The inventors of this invention isolated a substantially pure saponin (see the margin below, continued on the next page) from Astrax, and further discovered that it contained at least 10 types of saponin unknown in the literature. .

Thus, according to the present invention, a substantially pure saponin mixture and its components of formula (1) and formula (If
) and salts thereof are provided.

[In the formula, when R1 is a hydrogen atom, R2 is a β-D glucopyranosyl group and R8 is a 2,3,4-tri0-7cetyl-β-D-xylopyranosyl group; R2 is a β-D-glucopyranosyl group and R3 is 2,3-di-0-acetyl-β-
D-xylopyranosyl group, R2 is β-D-glucopyranosyl group, R3 is 2,4-di〇-7 cetyl-β-D-xylopyranosyl group; R2 is β-D-glucopyranosyl group, R8 is 2-〇-7 cetyl-β -D-xylopyranosyl group, R2 is β-D-glucopyranosyl group and R11 is β
-D-cheleropyranosyl group, R2 is β-D-vlucopyranosyl group, R1+ is β-D-glucopyranosyl (1-2
) β-D-xylopyranosyl group or R2 is a hydrogen atom and R
8 is β-D-glucopyranosyl (1-2) β-D-xylopyranosyl group; When R1 is β-D-glucopyranosyl group, R2
is a hydrogen atom and R8 is β-D-glucopyranosyl (1-2
) β-D-xylopyranosyl group, or R2 is a β-monoglypyranosyl group, and 8 is β-D-xy The specific names of these saponins are as follows.

8-0-(2,8,4-tri〇-acetyl-β-D-
xylopyranosyl)-6-o-β-D-glucopyranosylcycloastragenol (referred to as acetyl astragaloside I), B-0-(2,8-cy0-7
'tryl-β-D-xylopyranosyl)-6-0-β
-D-glucopyranosyl-cycloastragenol
[Named as astragaloside■], 8-0-(2,4-di0-7cetyl-β-D-xylopyranosyl)-6-0-β-D-glucopyranosyl-cycloastragenol [isoastragalo Side I
], 3-0-(2-O-7 cetyl-β-D-xylopyranosyl)-6-0-β-D-glucopyranosyl-cycloastragenol [named as astragaloside■], 8-0 -(β-D-glucopyranosyl(1-2)β-D
-xylopyranosyl]-cycloastragenol [
Astragaloside ■], 3-0-β-D-xylopyranosyl-6-〇-β-D-glucopyranosyl-cycloastragenol [designated as Astragaloside ■], 8-0-( β-D-glucopyranosyl (1-2)
β-D-xylopyranosyl)-25-0-β-D-glucopyranosyl-cycloastragenol [referred to as astragaloside ■], a-O-(β-D-glucopyranosyl (1-2) β-D
-xylopyranosyl]-6-0-β-D-glucopyranosyl-cycloastragenol [referred to as astragaloside ■], 3-0-β-D-xylopyranosyl-6-〇-β-D-
Glucopyranosyl-25-〇-β-D-glucopyranosyl-cycloastragenol [Astragaloside■
], and 8-O-(a-L-rhamnopyranosyl(
1-2) β-D-xylopyranosyl (1-2) β-D-
Glucuronopyranosyl]-Sawyasapogenol B [named as Astragaloside ■].

The saponin of the present invention is substantially pure, and the term "substantially pure" means that it contains at least 90% or more, preferably 98% or more, of only saponin.

Furthermore, the present invention also relates to Astragali Ra.
Extract dix with a lower alcohol, concentrate the extract, treat the lower alcohol solution of this concentrate with an adsorbent,
There is then provided a method for isolating the novel saponin by subjecting the eluted fraction to at least one round of chromatography for purification and separation without or after esterification. This will be explained in detail below.

First, extract the Japanese Aspergillus with lower alcohol.

Examples of the lower alcohol include 99% or more methanol or ethanol. This extraction is preferably carried out at or under heating. It should be noted that the raw material, Aspergillus japonica, may be previously cut into pieces or defatted by a conventional method prior to extraction. The obtained extract is concentrated to obtain an extract.

This extracted extract is dissolved in lower alcohol, and the solution is sprinkled on silica gel, such as 60-230 mesh silica gel manufactured by Merck & Co., Ltd. Note that the concentration of the lower alcohol solution of the extracted extract is appropriately selected so as to easily coat the silica gel. This extract-adhered silica gel is laminated on a shell (LJ) filled with silica gel in advance. The weight of this pre-filled silica gel is 5 to 20 times that of the extract-attached silica gel. This silica gel column, for example chloroform:
Gradient elution with lower alcohol:water, preferably chloroform:methanol:water (10:3:1 (lower layer) - 6:4:1), and the eluate was fractionated into 6 fractions using thin layer chromatography (TLO) as an indicator. Concentrate and dry each fraction to obtain fractions 1 to 6.
get.

Among these fractions, fractions 1 and 5 were separated by reverse phase silica gel column chromatography [for example, the carrier is Bondapak C18, manufactured by Waters Co., Ltd., and the elution solvent is lower alcohol: water, preferably methanol: water (5:
4-5:1)], followed by silica gel column chromatography [for example, Merck & Co., Ltd. 60-230 mesh silica gel is used as a carrier,
Elution tlX is chloroform: lower alcohol;
Water preferably chloroform:methanol:water (10:B
:1. lower layer)] and purified and separated into fractions 2, 3 and 4.
is purified and separated by reverse phase silica gel chromatography similar to that used for fractions 1 and 5 above.

Further, for fraction 6, a saponin mixture obtained by subjecting it to reverse phase silica gel chromatography similar to that described above is dissolved in a lower alcohol, preferably methanol, and methyl esterified by adding a diazomethane-ether solution. Furthermore, silica gel column chromatography [for example, using 60 to 230 mesh silica gel manufactured by Merck as a carrier,
The mixture is separated using n-butanol:ethyl acetate:water (4:1:5.j: layer), and then purified and separated by alkali treatment (for example, 10% aqueous potassium hydroxide solution).

When fractions 1 to 6 are purified and separated as described above, acetyl astragaloside I and astragaloside I are extracted from fraction 1.
and isoastragaloside I, astragaloside ■ from fraction 2, astragaloside ■ from fraction 3, astragaloside ■ from fraction 4, astragaloside ■, astragaloside from fraction 5 side■ and astragaloside■, and astragaloside■ from fraction 6
and soya saponin ■ are each f-exclusive.

These saponins can be converted into salts if desired. Examples of the salt include alkali metal salts or alkaline earth metal salts, specifically sodium salts, potassium salts, calcium salts, magnesium salts, and the like. Also,
These salts are prepared by conventional methods.

The novel saponin thus obtained has the effect of suppressing the production of lipid peroxides, can be used for the prevention and treatment of arteriosclerosis, and is effective in preventing aging.

Next, the method for isolating saponin of the present invention will be explained with reference to Examples.

Example “°3(□°゛°′°”1°°”ゝ”= Cy (D
9 f m :/.

Extraction and isolation of Astragalus astragalus (Korean Astragalus, 8 kg) was cut into small pieces and heated under reflux in methanol (18#, 99% methanol, same hereinafter) for 5 hours. A methanol extract is obtained by filtration, and methanol (181) is added to the residue and extracted with heat. The same operation was performed a total of 5 times, the resulting methanol extracts were combined, and the solvent was distilled off under reduced pressure to obtain a methanol extract (1.9 kg).
)'z get le.

The methanol extract (20Of) was dissolved in methanol, and silica gel (60-230 mesh, manufactured by Merck & Co., Ltd.,
400F, the silica gel used in this example means this silica gel unless otherwise specified). After drying under reduced pressure, the layers were stacked on a column packed with silica gel (4 kg), and chloroform:methanol:water (10:8) was added.
:1 (lower layer) (20β), 7:8:1 (lower layer) (10d
).

65:35:10 (lower layer) (101), 6:4:1 (1
([')) using silica gel thin layer chromatography as an indicator, the eluate was fractionated into 6 fractions, and fraction 1 (2
2y), fraction 2 (7,5g), fraction 8 (10g), fraction 4
(7,51, fraction 5 (6,111) and fraction 6 (9,
2g) is obtained.

Fraction 1 (221) was subjected to reverse phase silica gel column chromatography [Carrier: Bondapak 018. Manufactured by Waters, 100F, elution solvent was methanol:water (5:4-5
:1)), and further separated by silica gel column chromatography [1 kg of silica gel, taloloform:methanol:water (10:8:1, lower layer)] to obtain acetyl astragaloside I (20011ty) and astragaloside I. (8,5F) and isostragaloside I (30011F) were obtained.

Fraction 2 (7,51) was separated and purified by reverse phase silica gel column chromatography similar to that used in the treatment of fraction 1 to obtain astragaloside I [(2,3M).

Astragaloside I (1.0 g) was obtained from fraction 3 (101i') by reverse-phase silica gel column chromatography similar to that used in the treatment of fraction 1.
Astragaloside Yi C0,8f) was obtained from (7,5f) by the same procedure as that for fraction 3.

After separating and purifying fraction 5 (6,8F) using reverse phase silica gel column chromatography similar to that used for fraction 1,
Silica gel column chromatography [silica gel, 70
Ofi chloroform:methanol:water (7:3:1.
lower layer)] and astragaloside 1100ml
F), Astragaloside ■ (300WLg), and Astragaloside ■ (1001Rg) were obtained.

Fraction 6 (9,2i was used for the treatment of fraction 1, separated and purified by reverse phase silica gel column chromatography similar to that of t2,
A saponin mixture (2.5 g) is obtained.

A saponin mixture (2.5 g) is dissolved in methanol and methyl esterified by adding diazomethane-ether solution. Silica gel column chromatography [silica gel 50
0g, n-butanol:ethyl acetate:water (4:1:5.
upper layer)] and then treated with alkali (10% aqueous potassium hydroxide solution) to astragaloside (600
(2) and soya saponin 1 (600+11) were obtained.

The physical properties of each saponin obtained in the above examples are as follows.

Acetyl astragaloside 1 1) mp280-281°C.

2) (a) D + 1-8° (C=1.0.m/-r) (7) Has optical rotation.

8) 04? H? It has a molecular composition of 4017.

4) Infrared absorption spectrum (KBr, an-')
C shows a characteristic absorption maximum at 8400 (broad) and 1750.1225°1030.

5) It does not exhibit ultraviolet absorption at wavelengths longer than 21071 Jl.

s) 18c nuclear magnetic resonance spectrum (ds-pyridine).

δC) is 170.1, 170.0, 169.5 (acetyl carbonyl C), 105.0.1014 (7 nomeric C), 89.5 (3-C), 87.8 (20-0)
, 82.1 (24-C), 79.8 (6-C), etc. are shown.

7) No odor, colorless needle-like crystals (crystallized from methanol).

8) Methanol, ethanol, n-butanol.

Easily soluble in pyridine and dimethyl sulfoxide, soluble in chloroform, ethyl acetate, and acetone, and insoluble in ether, benzene, and hexane.

9) Thin layer chromatography (TLC, carrier nib record silica gel 60 F264 plate.

0.251111. Manufactured by Merck; developing solvent: chloroform: methanol: water (7:8:1. lower layer)]
Indicates Rf=0.6.

When a 1% cerium sulfate-1G% sulfuric acid aqueous solution is sprayed on the TLC and heated, it becomes dark brown.

U yo 1 Astragalocide 1 1) mp: 184-186°C.

2) [α), 7) It has optical rotation of −12.7° (C=0.6.methanol).

3) It has a molecular composition of C45H72016.HzO.

4) Red-line absorption spectrum (KBr t cIR-”
') is 8400 (broad), 1784.1258°1
086.1045 shows a characteristic absorption maximum.

5) Does not exhibit ultraviolet absorption at wavelengths longer than 210M.

6) C nuclear magnetic resonance spectrum (aS-pyridine.

δC) is 170.6.169.8 (acetyl carbonyl C), 105.0, 104.1 (7 nomeric C), 8
9.4 (8-C), 87.4 (20-0), 82.2
Signals such as (24-C) and 79.4(6-() are shown.

7) No odor, colorless fine crystals (crystallized from acetone).

8) Methanol, ethanol, n-butanol.

Easily soluble in pyridine and dimethyl sulfoxide, soluble in chloroform, ethyl acetate, and acetone, and insoluble in ether, benzene, and hexane.

9) Thin layer chromatography (TLC, carrier nib record silica gel 60 F26. plate.

0.251j1M, manufactured by Merck; developing solvent: chloroform: methanol: water (7:8:1. lower layer)]
Indicates approximately f = 0.5.

When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed onto the TLO and heated, it becomes dark brown.

B Isostragaloside 1 1) mp 21g ~ 220'C.

2) It has an optical rotation of (a:l”+17.9° (Ci=1.0.methanol).

8) 04sH7zO16・HzO (D molecular composition)
.

4) Infrared absorption spectrum (KBr, cIR) is 840
0 (broad), shows a characteristic absorption maximum at 1740.1280°1050.

5) Ultraviolet Ii for wavelengths longer than 21071J! Shows no absorption.

6) C nuclear magnetic resonance spectrum (cls-pyridine.

δC) is 170.5, 170.2 (acetyl carbonyl C), 105.0, 104.4 (anomeric C), 8
9.8 (3-C), 87.2 (20-C), '82.2
(24-0), 79.5 (6-0), etc. are shown.

7) No odor, colorless fine crystals (crystallized from chloroform-methanol).

8) Methanol, ethanol, n-butanol.

Easily soluble in pyridine and dimethyl sulfoxide, soluble in chloroform, ethyl acetate, and acetone, and insoluble in ether, benzene, and hexane.

9) Thin layer chromatography (TLC, precoated silica gel 60 F254, 0.25 M, manufactured by Merck & Co., Ltd.,
Chloroform:methanol:water (7:3:1.lower layer)]
shows Rf=0.48.

When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed on the TLC and heated, a dark brown color appears.

10) Structural formula %formula% It has optical rotation of .

8) It has a molecular composition of 0aaEyoOt5.HzO.

4) The infrared absorption spectrum (KEr, cllt) is 3
400 (broad), 1789.12B6°1070.
1089 shows a characteristic absorption maximum.

5) Does not exhibit ultraviolet absorption at wavelengths longer than 210M.

6) C nuclear magnetic resonance spectrum (aS-pyridine.

δC) is 170.1 (acetylcarbonyl C), 105
．． 0.104.8 (Anomeric C), 89.2 (8-
C), 87.4 (20-0), 82.2 (24-C),
79.4 (6-c) and other signals are shown.

7) It has no odor and is colorless fine crystals (crystallized from chloroform-methanol).

8) Methanol, ethanol, n-butanol.

It is easily soluble in pyridine and dimethyl sulfoxide, slightly soluble in chloroform, ethyl acetate, and acetone, and insoluble in x-f, benzene, and hexane.

9) Thin layer chromatography (TLC, precoated silica gel 60 F254 +0.25ff, manufactured by Merck & Co., Ltd.
Chloroform:methanol:water (7:8:1.lower layer)]
shows Rf=0.45.

When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed onto the TLO and heated, it becomes dark brown.

10) Structural formula %formula% It has optical rotation of .

8) 041H6801a ・Hs+OO) Molecule 1a
It doesn't exist.

4) Infrared absorption spectrum (KEr, α-1) is 887
0 (broad), shows a characteristic absorption maximum at 1070.1080.

5) Does not exhibit ultraviolet absorption at wavelengths longer than 210%N.

6) C nuclear magnetic resonance spectrum (aS-pyridine.

δC) is 105.8, 105.4 (7 nomeric C),
88.8 (8-C'), 87.4 (20=c), 83
．． 1 (2'-C of xylose moiety), 82.2 (24
Indicates a signal such as -0>.

7) No odor, colorless needle-like crystals (crystallized from methanol).

8) Methanol, ethanol, n-butanol.

Soluble in pyridine, dimethyl sulfoxide, insoluble in ethyl acetate, acetone, ether, penzene, hexane.

9) Thin layer chromatography (TLC, precoated silica gel 60 F264, 0.25 mg, manufactured by Merck & Co., Ltd., chloroform:methanol:water (7:8:1. lower layer)
] indicates Rf=0.4.

When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed onto the TLO and heated, it becomes dark brown.

10) Structural formula Astragalocide Harr 1) mp 299-801°C.

2) (a) D+ It has an optical rotation of 24.4° (0=0.2.methanol).

8) It has a molecular composition of 041E68014/2H20.

4) Infrared absorption spectrum (KBr, m) is 388
0 (broad), showing a characteristic absorption maximum at 1065.1040.

5) Does not exhibit ultraviolet absorption at wavelengths longer than 2107LII.

e) 1 ac nuclear magnetic resonance spectrum (aS-pyridine.

δC) is 107.1, 105.0 (7 nomeric C),
88.7 (8-C!), 87.8 (20-C),
82. '0 (24-C), 79.2 (6-C> etc. signals are shown.

7) No odor, colorless needle-like crystals (crystallized from methanol).

8) Methanol, ethanol, n-butanol.

Soluble in pyridine, dimethyl sulfoxide, insoluble in ethyl acetate, acetone, ether, benzene, hexane.

9) Thin layer chromatography ('I'LC, precoated silica gel 60 Fzs 4.0.25+11. manufactured by Merck & Co., chloroform:methanol:water (7:3:1. lower layer)) shows Rf = 0.86.

When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed on the TLC and heated, a dark brown color appears.

10) Structural formula % Formula % 2) It has optical rotation of [α] + 7.2° (C = 1.0. methanol).

8) CuHysOts ・8HzO(7) molecule fi
There is a fl structure.

4) Infrared absorption spectrum (KBr, cIll) is 8
400 (broad), 1075.1085 shows a characteristic absorption maximum.

5) Does not exhibit ultraviolet absorption at wavelengths longer than 210M.

6) C nuclear magnetic resonance spectrum (aS-pyridine.

δC) is 105.7, 105.8, 98.7 (7 nomeric C), 88.6 (8-C), 87.2 (20-C)
, 88.0 (2-C of xylose moiety), 82.2 (2
4-0), 78.6 (25-0), etc. are shown.

7) No odor, colorless fine crystals (crystallized from methanol).

8) Methanol, ethanol, n-butanol.

Soluble in pyridine, dimethyl sulfoxide, insoluble in ethyl acetate, acetone, chloroform, ether, benzene, hexane.

9) Shows Rf=0.2 in thin layer chromatography (TLO, precoated silica gel 60F254, 0.25 m, manufactured by Merck & Co., chloroform:methanol:water (7:8:1, lower layer)).

When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed on the TLC and heated, a dark brown color appears.

10) Structural formula Astragalocide■ 1) mp 290-291'C.

2) [α), +17.8° (C=1°0.methanol)
It has optical rotation of .

8) C4yEyso 19 ・HgO(7) molecular composition.

4) Infrared absorption spectrum (KBr, α-1) is 840
0 (broad), showing a characteristic absorption maximum at 1075.1088.

5) It does not exhibit ultraviolet absorption at wavelengths longer than 2107!111.

6) C nuclear magnetic resonance spectrum (aS-pyridine.

δC) is 105.9, 105.2, 104.9 (7 nomeric C), 88.5 (3-0), 87.2 (20-0
), 8:13.5 (2-C of xylose moiety), 81.
8 (24-0'), 79.1 (6-0), etc. are shown.

? ) It has no odor and is colorless fine crystals (crystallized from methanol).

8) Methanol, ethanol, n-butanol.

Soluble in pyridine, dimethyl sulfoxide, insoluble in ethyl acetate, acetone, chloroporum, benzene, ether, hexane.

9) Rf=0.19 in thin layer chromatography (TLO, precoated silica gel 60 Fzs4+ 0.25 mm, manufactured by Merck & Co., chloroform:methanol:water (7:8:1, lower layer)).

When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed on the TLC and heated, a dark brown color appears.

10) Structural formula %formula% It has optical rotation of .

8) Allows the molecular composition of 0ayHfu8019・2■20.

4) Infrared absorption spectrum (1KBr, cm-”
, shows a characteristic absorption maximum at 8400 (broad) and 1070.1040.

5) 21M! It exhibits no ultraviolet absorption at longer wavelengths.

6) c nuclear m gas resonance spectrum ((15-pyridine.

δC) is 107.8, 104.8, 98.8 (7
Nomeric C), 88.6 (8-C), 87.2 (2
0-C), 82.2 (24-C), 79.1 (6-C
j'), 78.7 (25-Cj), etc. are shown.

7) No odor, colorless needle-like crystals (crystallized from methanol).

8) Methanol, ethanol, n-butanol.

Soluble in pyridine, dimethyl sulfoxide, insoluble in ethyl acetate, chloroform, acetone, ether, benzene, hexane.

9) Shows Rf=0.18 in 18-layer chromatography ('I'LO, pre-foot silica gel 60F254, 0.25 fin, manufactured by Merck & Co., chloroform; methanol: water (7:3:1. lower layer)).

'['When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed on LO and heated, it becomes dark brown.

10) Structural formula Astragalocide■ 1) mp 22a ~ 224°C.

2) [α) D-12,1° (C=1.0.methanol)
It has optical rotation of .

3) It has a molecular composition of C47H76017.H2O.

4) The infrared absorption spectrum (KBr + Bi-') shows characteristic absorption maxima at 3400 (broad) and 1725.1040.

5) 2107! l! ! It exhibits no ultraviolet absorption at longer wavelengths.

6) It has no odor and is colorless fine crystals (crystallized from methanol).

7) Easily soluble in methanol, pyridine, dimethyl sulfoxide, soluble in ethanol, n-butanol, water, chloroform, ethyl acetate.

Insoluble in acetone, benzene and hexane.

8) 47! Chromatography (TLC, precoated silica gel 60F254.0.25Mm, manufactured by Merck & Co., Ltd.,
Chloroform:methanol:water (7:3:1.lower layer)]
shows Rf=0.1.

When a 1% cerium sulfate-10% sulfuric acid aqueous solution is sprayed on the TLC and heated, a reddish-purple color appears.

9) Structural formula Next, the results of a pharmacological test of the saponin of the present invention to inhibit lipid peroxide production are shown.

Pharmacological test for inhibiting lipid peroxide production Adriamycin, an anti-inflammatory drug, binds to DNA and inhibits nucleic acid synthesis, and also inhibits lipid metabolism in the heart, causing lipid peroxide to accumulate and causing myocardial damage as a side effect. Are known.

The inventors of the present invention utilized this to investigate the effects of inhibiting lipid peroxide production on acetylametragalloside 11, astragaloside 1,1, ■, ■, ■, ■, ■, ■ and isostragaloside I. We tested the efficacy of saponi/@ contained in Astragalus as a lipid peroxide inhibitor and found that both had strong lipid peroxide inhibitory effects. The test results will be specifically explained below.

〔experimental method〕

1) Groups of 5 COF male mice (5 weeks old, 20-25f) were used, and adriamycin (manufactured by Kyowa Hakko Kogyo) was intraperitoneally administered to each mouse 2 at a dose of 15 da/kti (drug volume : 0.15 shoulders per 10 f of body weight1).

Table 1 below shows acetylametragalloside 11, astragaloside I,
■, ■, ■, ■, ■, ■ and Inas□ The results when trogalloside I was used are shown.

For each test drug, the body weight was 10 days before administration of Adriamycin.
Intraperitoneal administration was started at a rate of 50.10 gt/f and continued for 5 days. Immediately before each test drug is used, it is administered in 0.9% physiological saline or 1%7E78Q.
(Tween 3Q) was used after being suspended in 0.9% physiological saline. Each test drug was administered at noon every day, and only adriamycin was administered 3 hours after administration of the test drug.

The dose of each test drug was 20% for each astragaloside.
0W/9, and mice in the control group received 0.9% physiological saline. 2) For the measurement of lipid peroxide, each animal was killed by cervical dislocation at the 6th white eye, and the heart and After removing the liver and measuring its wet weight, a 2% homogenate solution was prepared using 0.9% physiological saline using a Botter type Teflon homogenizer under ice cooling. Using this as a test solution, the amount of lipid peroxide was measured using the following improved Yagi method, and the amount of lipid peroxide in the heart and liver was quantified and compared with the control group.

To 0.2 yttl of the above 2% homogenate solution, add 0.5 ytl of 3% lauryl sulfate solution, mix by shaking for 30 seconds, and add acetate buffer (pH 8,6) to this.
5 ml and 1.5 ml of 0.8% thiono-zeta rubituric acid solution
4. After forming OrlIl, shake well for 30 seconds and incubate at 95° for 60 minutes in an oil bath.
After heating at C, cool under running water for 5 minutes. Next, add 5.0 ml of 0.2 N hydrochloric acid and 1.0 ml of n-butanol/pyridine 7 (15:1) solution and blow vigorously! 15 after ll″1
The upper n-butanol layer was separated by centrifugation (8000 rpm) for 1 minute, and then measured using a fluorescence spectrophotometer (EX515 nm).
, Em 553 nm). Separately, a test identical to the present procedure was conducted using a malonaldehyde standard solution. A calibration curve showing the relationship between fluorescence and the amount of lipid peroxide was created, and the measured values were applied to this to determine the content.

〔Experimental result〕
In order to compare the effects of each test drug and each dose, the lipid peroxide production inhibition rate was calculated using the following formula, and the results are shown in Table 1.

(, -D Inhibition rate) = × 100 (, -A Lipid peroxide concentration in the group where Niadriamycin was not administered C Lipid peroxide concentration in the control group administered Niadriamycin D Niadriamycin and the test drug were administered Group lipid peroxide concentration

Claims (1)

[Claims] 1. Extract Astragali Radix with a lower alcohol, concentrate the extract, treat the lower alcohol solution of this concentrate with an adsorbent, and then elute the resulting fraction at least once by chromatography. 1. A method for isolating Astrax saponins, the method comprising isolating a substantially pure saponin mixture.