JP2000239169A - Promotor agent for anti-carcinogenesis - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain the subject safe agent suppressing a carcinogenesis promoter activity, useful for preventing the carcinogenesis, and also having no adverse effect by containing a specific megastigmane glycoside. SOLUTION: This agent contains a megastigmane glycoside of the formula (R is H or OH; R1 is a monosaccharide or disaccharide), [concretely, roseoside, (6S,9R)-bomifoliol-9-O-β-D-apiofranosyl-(1→6)-β-D-glucopyranoside, or the like.]. The above megastigmane glycoside is preferably originated from an edible plant such as a loquat (Eriobotrya japonica) and Alangium premnifolium, and is obtained e.g. by extracting the plant such as the leaves of the loquat, or the like, with an organic solvent such as ethanol. As to its dosage, in the case of e.g. oral administration, usually approximately 10-1,000 mg/kg body weight is administered daily over one to several times.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to an anti-tumor promoter agent which comprises a megastigman glycoside as an active ingredient, suppresses the action of a tumor promoter on potential cancer cells, and is effective for preventing carcinogenesis.

[0002]

2. Description of the Related Art As a method for treating cancer, surgical removal of cancer cells, radiation therapy, chemotherapy using anticancer drugs, and immunostimulation therapy have been used. In Japan, cancer has been the leading cause of death since 1981 (Nutrition Journal Vol.54, No.3,12
1-127, 1998), prevention of carcinogenesis has become a very important issue for the health of the population.

[0003] Cancer is thought to occur through two distinct stages, initiation and promotion. That is, after the initiation of damage to the DNA of normal cells by initiators such as ultraviolet rays, radiation, and other mutagens, the resulting potential cancer cells undergo continuous stimulation by chemicals called promoters. It is thought that cancer will occur through the process of receiving promotion. Therefore, it is considered that carcinogenesis can be prevented by suppressing either the initiation or the promotion. However, initiation is an instantaneous reaction, and it is difficult to completely protect it. On the other hand, promotion requires a long-term continuous action by a promoter, and thus inhibiting this process is considered to be an effective method for preventing carcinogenesis. .36-38, Gakkai Shuppan Center, 1996).

[0004] In recent years, attention has been paid to the anti-carcinogenic effect of components derived from natural products, and epigallocatechin gallate contained in green tea (Pharmacia Vol. 34, No. 3, 223-225 (199)
8)), a triterpene compound derived from polia cocos (Japanese Patent Application Laid-Open No. 9-176184), a wasabi tree (Mor
inga oleifera) extract (Japanese Unexamined Patent Publication No. Hei 7-304684) and a ginkgo (Ginkgo biloba L.) leaf extract (Japanese Unexamined Patent Publication No.
-126180), Kaffir lime extract (Japanese Unexamined Patent Publication No.
JP-A-336364), a yellow leaf extract (Japanese Patent Laid-Open No. 6-2)
No. 47851), krill extract (JP-A-6-29)
No. 3647), sponge extract (JP-A-6-30597)
No. 0) and flavonoids contained in yellow ginger (Japanese Patent Application Laid-Open No. 5-25041) have been reported, but these have not yet been established as effective means for cancer prevention.

[0005]

SUMMARY OF THE INVENTION An object of the present invention is to provide a safe anti-cancer promoter agent which suppresses the action of a carcinogenesis promoter, is useful for preventing carcinogenesis, and has no side effects.

For this purpose, the present inventors have conducted intensive studies on natural components contained in edible plants, and have been used for edibles for a long time, and the safety of loquat (Eriob) has been established.
otrya japonica) and zinnia (Alangium premnifoliu)
m) and has been reported (Journal of Natural
Products, Vol.55, No.8, pp1025-1032, August 1992; Chem
Pharm Bull, Vol. 43 No. 5, 754-759 (1995)). So far, among the megastigmann glycosides, roseoside has been reported to have an antihistamine effect (Chem Pharm Bull, Vol. 45 No. 3, 464-469).
(1997)), however, there was no report on the anti-carcinogenic promotion effect of megastigman glycoside.

[0007]

That is, the present inventors have established safety by deriving from edible plants, and even if they are extracted from other plants or microorganisms, or if they are chemically synthesized products, It has been found that megastigman glycoside, which has no problem, has an anti-carcinogenic promotion effect.

Embedded image (In the formula, R represents hydrogen or a hydroxyl group, and R ¹ represents a monosaccharide or a disaccharide.) It has been found that the megastigman glycoside represented by the formula ( ¹ ) suppresses the action of a tumor promoter and is useful for preventing carcinogenesis. The present invention has been completed.

That is, the present invention relates to a compound represented by the general formula [I]:
R represents hydrogen or a hydroxyl group, and R ¹ represents a monosaccharide or disaccharide. ) Is an antitumor promoter agent containing the megastigman glycoside represented by the formula (1) as an active ingredient. Further, Megasuchiguman glycoside, in the general formula [I], R is OH, Roseoshido ^{R 1} is glucopyranose, R is OH, ^{R 1} is Apio furanosyl glucopyranose (6S, 9R)
-Bomifoliol-9-O-β-D-apiofuranosyl- (1 → 6) -β-D-glucopyranoside, where R is H,
R ¹ is a glucopyranose (6R, 9R) -3- oxo -α- Ioniru -9-O-β-D- glucopyranoside, R is H, ^{R 1} is xylopyranose pyranosyl glucopyranose (6R, 9R ) -3-Oxo-α-ionyl-9
-O-β-D-xylopyranosyl- (1 → 6) -β-D
-An antitumor promoter agent containing glucopyranosides as active ingredients.

[0009]

BEST MODE FOR CARRYING OUT THE INVENTION Megastigman glycoside represented by the general formula [I] can be extracted and separated from plants such as loquat (Eriobotrya japonica) leaves. Examples of the organic solvent used for the extraction include alcohol, ether, acetone, hexane, chloroform, toluene, ethyl acetate, tetrohydrofuran, and the like. Not something.
In practice, one or more of these solvents are selected and used, but from the viewpoint of safety, ethanol, acetone or ethyl acetate, etc., especially ethanol which is also used for drinking It is preferred to use More preferably, ethanol is used by mixing it with water at an appropriate ratio.

[0010] The plant may be subjected to the extraction as it is. However, in order to enhance the extraction efficiency, the plant is preferably cut or crushed before the extraction. The extraction method is as follows:
Add 1 to 50 liters, preferably 10 liters of extraction solvent per kg and leave at room temperature for 1 day to 1 month. If necessary, stirring may be carried out during the standing. Further, the extraction solvent can be used after being heated. After the extraction, the plant (extraction residue) and the extraction solvent (extract) are separated by filtration or sedimentation. Preferably, the same extraction operation is repeated about 2 to 4 times for the extraction residue. The whole extract is collected and concentrated and dried to obtain a crude extract.

Thereafter, the crude extract is separated and purified by using various types of chromatography to obtain a megastigman glycoside represented by the above general formula [I]. Specifically, in the general formula [I], R is OH, R ¹ is a roseoside of glucopyranose, R is H, and R ¹ is (6S, 9R) -bomifoliol-9-O of apioflanosylglucopyranose. −
β-D-apiofuranosyl- (1 → 6) -β-D-glucopyranoside, where R is H and R ¹ is glucopyranose (6
R, 9R) -3-oxo-α-ionyl-9-O-β-
(6R, 9R) -3-oxo-α of D-glucopyranoside, where R is H and R ¹ is xylopyranosylglucopyranose
-Ionyl-9-O-β-D-xylopyranosyl- (1
→ 6) β-D-glucopyranoside and the like are obtained.

Examples of the chromatography method include adsorption chromatography, liquid chromatography, ion exchange chromatography, partition chromatography, and liquid-liquid extraction. Although not particularly limited as long as the desired compound can be obtained, Diaion HP
-20, Toyopearl HW-40, MCI-g
el CHP-20P, Sephadex LH-2
0, a column packed with YMC-gel ODS AQ 120-50S or the like is preferably used. However, the present invention is not limited to these, and there is no problem even if it is extracted from other plants or microorganisms, or a chemically synthesized product.

The antitumor promoter agent of the present invention may be used directly as its active ingredient, the megastigman glycoside represented by the above general formula [I], but may be used as a pharmaceutically acceptable carrier or other preparation. Can be orally or parenterally administered and ingested in the form of a pharmaceutical formulation by mixing with a pharmaceutical additive or by mixing with a general quasi-drug or the like. Specific examples include tablets, capsules, powders, suppositories, rectal ointments, injections, lotions, hand creams, and the like, but are not necessarily limited to these forms.

[0014] The dose of the megastigman glycoside, which is the active ingredient of the present invention, can vary depending on the body weight of the subject, the mode of administration, and the site of administration. For example, in the case of oral administration, it is usually 10 to 1000 mg / mg. Administer about 1 kg of body weight once or several times a day. Use of concentrations lower than 10 mg / kg may be ineffective.

The present invention will be described in more detail with reference to the following Examples, which by no means limit the present invention.

Example 1 Preparation method of compound: 1 kg of fresh ground loquat leaves was immersed and extracted twice with 5 liters of 70% acetone water, and the extract was collected, concentrated under reduced pressure using a rotary evaporator, and then freeze-dried. , Loquat leaf crude extract 9
8 g were obtained. This crude loquat leaf extract was suspended in 1 liter of water, and liquid and extracted sequentially with equal amounts of chloroform, ethyl acetate and butanol. The butanol phase was concentrated under reduced pressure using a rotary evaporator, and then lyophilized to give a butanol fraction of 28 g. I got The butanol fraction was added to Diaion H.
P-20, Toyopearl HW-40, MCI-
gel CHP-20P, Sephadex LH-2
0, and sequentially subjected to force column chromatography using YMC-gel ODS AQ 120-50S to obtain roseoside (40 mg), (6S, 9R) -bomifoliol-
9-O-β-D-apioflanosyl- (1 → 6) -β-
D-glucopyranoside (25 mg), (6R, 9R)-
3-oxo-α-ionyl-9-O-β-D-glucopyranoside (55 mg), (6R, 9R) -3-oxo-
α-ionyl-9-O-β-D-xylopyranosyl-
Four kinds of megastigman glycosides of (1 → 6) -β-D-glucopyranoside (35 mg) were obtained.

[Test for Antitumor Promoter Activity] Example 1
The four megastigman glycosides obtained in the above were tested for the Epstein-Barr virus activation inhibition test method (Biochemical Experiment Method 38, Research Methods for Biological Function Modulating Substances in Foods, page 38-4).
1, Academic Press, 1996) was used to evaluate the antitumor promoter effect. This test method uses radio cells (DN
A cell line derived from Burkitt's lymphoma infected with the Epstein-Barr virus, a type A cancer virus), in the process of culturing in the presence of tetradecanoyl phorbol acetate (TPA), which is an oncogene promoter, to express early antigens. The purpose of the present invention is to evaluate the antigen expression suppressing effect of a test sample. PRMI16 as a culture solution of raji cells
40 plus fetal serum and antibiotics were used.
The spontaneous occurrence of Epstein-Barr virus early antigen under these culture conditions was 0.1% or less. 1 × 10 ⁶
/ Ml of raji cells prepared at a concentration of 4 mM n-butyric acid, 20 ng / ml (32 pmol) TPA, and
The cells were cultured in a culture solution containing a test substance at a concentration of 1000 mol ratio / TPA (1000 times the molar ratio with respect to TPA) at 37 ° C. and 5% CO ₂ for 48 hours.
The cells expressing the Epstein-Barr virus early antigen were detected by the indirect fluorescent antibody method using the serum of a nasopharyngeal pain patient, and the ratio of positive cells was calculated with respect to a control to which no test substance was added. Was determined to be the inhibitory activity. Further, the concentration of the test substance was set to 500 mol ra.
tio / TPA, 100 mol ratio / TPA, 1
The activity was measured by changing to 0 mol ratio / TPA. The results are shown in Table 1.

[0018]

[Table 1] Each of the megastigman glycosides exhibited an anti-carcinogenic promotion activity superior to epigallocatechin gallate, which has already been reported to have an anti-carcinogenic promotion activity.

Example 2 Tablets (1) Roseoside 5 g (2) Fine granules for direct hit No.209 (manufactured by Fuji Chemical Co., Ltd.) 7 g Magnesium aluminate metasilicate 20% Corn starch 30% Lactose 50% (3) Crystalline cellulose 6 g (4) CMC calcium 1.8 g (5) Magnesium stearate 0.2 g After uniformly mixing (1) to (4), (5) is added and further mixed, and the mixed powder is tableted. Then one tablet 2
A tablet of 00 mg was obtained. This tablet may be used, if necessary, with a commonly used gastrosoluble film coating agent (for example,
(Polyvinyl acetal diethylaminoacetate) or an edible colorant.

Example 3 Capsule (1) (6S, 9R) -Bomifoliol-9-O-β-D-apiofuranosyl- (1 → 6) -β-D-glucopyranoside 10 g (2) Lactose 9.6 g (3) 0.4 g of magnesium stearate The above components were uniformly mixed, and the mixed powder was filled into hard gelatin capsules in an amount of 2000 mg.

Example 4 Injection (1) (6R, 9R) -3-oxo-α-ionyl-9-O-β-D-glucopyranoside 100 mg (2) Glucose 100 mg (3) Water for injection 10 ml in total volume The solution obtained by dissolving (1) and (2) in (3) was filtered through a membrane filter, sterilized and filtered again, and the filtrate was aseptically dispensed into vials, filled with nitrogen gas, and then sealed. To give an intravenous injection.

Example 5 Lotion (1) (6R, 9R) -3-oxo-α-ionyl-9-O-β-D-xylopyranosyl- (1 → 6) -β-D-glucopyranoside 1 g (2 ) Paraoxybenzoate 0.5g (3) Propylene glycol 1g (4) Concentrated glycerin 1g (5) Sodium citrate 0.5g (6) Suitable amount of perfume (7) Purified water 95ml (1) to (6) ) To give a lotion.

[0023]

Industrial Applicability The megastigman glycoside of the general formula [I], which is an active ingredient of the present invention, has excellent anti-tumor promoting activity and can suppress the action of a carcinogenic promoter. It is useful for preventing carcinogenesis, and its safety has already been established.

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[Procedure amendment]

[Submission date] February 17, 1999 (1999.2.1
7)

[Procedure amendment 1]

[Document name to be amended] Statement

[Correction target item name] 0016

[Correction method] Change

[Correction contents]

Example 1 Preparation method of compound: 1 kg of fresh ground loquat leaves was immersed and extracted twice with 5 liters of 70% acetone water, and the extract was collected, concentrated under reduced pressure using a rotary evaporator, and then freeze-dried. , Loquat leaf crude extract 9
8 g were obtained. This crude loquat leaf extract was suspended in 1 liter of water, and liquid and extracted sequentially with equal amounts of chloroform, ethyl acetate and butanol. The butanol phase was concentrated under reduced pressure using a rotary evaporator, and then lyophilized to give a butanol fraction of 28 g. I got The butanol fraction was added to Diaion H.
P-20, Toyopearl HW-40, MCI-
gel CHP-20P, Sephadex LH-2
0, and sequentially subjected to column chromatography using YMC-gel ODS AQ 120-50S to give roseoside (40 mg), (6S, 9R) -bomifoliol-
9-O-β-D-apioflanosyl- (1 → 6) -β-
D-glucopyranoside (25 mg), (6R, 9R)-
3-oxo-α-ionyl-9-O-β-D-glucopyranoside (55 mg), (6R, 9R) -3-oxo-
α-ionyl-9-O-β-D-xylopyranosyl-
Four kinds of megastigman glycosides of (1 → 6) -β-D-glucopyranoside (35 mg) were obtained.

[Procedure amendment 2]

[Document name to be amended] Statement

[Correction target item name] 0018

[Correction method] Change

[Correction contents]

[0018]

[Table 1] Each of the megastigman glycosides exhibited an anti-carcinogenic promotion activity superior to epigallocatechin gallate, which has already been reported to have an anti-carcinogenic promotion activity.

[Procedure amendment 3]

[Document name to be amended] Statement

[Correction target item name] 0019

[Correction method] Change

[Correction contents]

Example 2 Tablets (1) Roseoside 5 g (2) Fine granules for direct hit No.209 (manufactured by Fuji Chemical Co., Ltd.) 7 g Magnesium aluminate metasilicate 20% Corn starch 30% Lactose 50% (3) Crystalline cellulose 6 g (4) CMC calcium 1.8 g (5) Magnesium stearate 0.2 g After uniformly mixing (1) to (4), (5) is added and further mixed, and the mixed powder is tableted. Then one tablet 2
A tablet of 00 mg was obtained. This tablet may be used, if necessary, with a commonly used gastrosoluble film coating agent (for example,
(Polyvinyl acetal diethylaminoacetate) or an edible colorant.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) C07H 15/18 C07H 15/18 (72) Inventor Naoki Kubo 941-6 Arashi Sozawa, Sagamiko-cho, Tsukui-gun, Kanagawa Pref. A-105 (72) Inventor Daigo Sugita 3-29-10 Asahicho, Nerima-ku, Tokyo (72) Inventor Susumu Shimura 3-2486, Moto-Hachioji-cho, Hachioji-shi, Tokyo (72) Inventor Yoshio Ito, Kiyose-shi, Tokyo 3-26-11 Noshio F term (reference) 4C057 AA06 BB03 CC01 DD01 JJ19 4C086 AA01 AA02 EA07 GA17 MA01 MA04 NA14 ZB26 4C088 AB12 AB51 BA13 BA32 ZB26

Claims (6)

[Claims] 1. A compound of the general formula [I] (Wherein, R represents a hydrogen or a hydroxyl group, and R ¹ represents a monosaccharide or a disaccharide). An antitumor promoter agent containing a megastigman glycoside represented by the following formula: 2. The antitumor promoter agent according to claim 1, wherein the megastigman glycoside is derived from edible plants such as loquat and zinnia. 3. The antitumor promoter according to claim 1, wherein the megastigman glycoside is roseoside. 4. The method of claim 1 wherein the megastigman glycoside is (6S, 9R)
The anticancer promoter agent according to claim 1 or 2, which is -bomifoliol-9-O-β-D-apiofuranosyl- (1 → 6) -β-D-glucopyranoside. 5. The method according to claim 5, wherein the megastigman glycoside is (6R, 9R)
The antitumor promoter agent according to claim 1 or 2, which is -3-oxo-α-ionyl-9-O-β-D-glucopyranoside. 6. The megastigman glycoside of (6R, 9R)
The antitumor promoter agent according to claim 1 or 2, which is -3-oxo-α-ionyl-9-O-β-D-xylopyranosyl- (1 → 6) -β-D-glucopyranoside.