JP2002080362A - Ppar-dependent gene transcription activator - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a PPAR-dependent gene transcription activator that has excellent PPAR-dependent gene transcription activity, high safety and is useful as a prophylactic and alleviating agent for obesity, diabetes, hyperglycemia, hyperlipemia and insulin resistance. SOLUTION: The objective PPAR(Peroxisome Proliferator Activated Receptor)-dependent gene transcription activator comprises the following substances (A) through (F): (A) gallic ester, (B) galloyl tannin, (C) quercetin or glycoside thereof, (D) flavone or its analogue, (E) isoflavone or its analogue and (F) a compound selected from catechins and epicatechins.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a peroxisome proliferator-activated receptor which is effective for activating lipid metabolism and preventing or improving obesity, diabetes, hyperglycemia, hyperlipidemia and insulin resistance. Recept
or: PPAR) -dependent gene transcription activator.

[0002]

2. Description of the Related Art Low molecular fat-soluble ligands such as steroids, thyroid hormones, retinoids, etc. are morphogenetic in ontogeny, cell proliferation and differentiation, and maintenance of homeostasis in the ontogeny via ligand-specific nuclear receptors. It is involved in the regulation of various physiological functions. PPAR is a kind of nuclear receptor, and was identified in 1990 as a protein that mediates the action of increasing peroxisome, an organelle involved in lipolysis, and means a receptor activated by a peroxisome proliferator. By Peroxisome Proliferato
r Activated Receptorα (peroxisome proliferator-activated receptor: PPARα). Thereafter, β / δ type and γ type were identified as isoform genes structurally similar to α type, and are known to consist of a total of three subtypes. The function of PPARα is widely considered to be involved in the energy metabolism and maintenance of homeostasis in living organisms, such as fatty acid synthesis, transport and secretion, ATP production in fat-consuming organs, and cell cycle regulation. In particular, it has been revealed that gene expression of β-oxidation-related enzymes important for fatty acid metabolism strongly depends on PPAR activation. PPA
Rγ, especially PPARγ2, is expressed with relatively strong specificity in adipocytes, and has been shown to play a central role in adipocyte differentiation (cells: 31 (6) 218).
-234, 1999, J. Lipid Res. 37, 907-925, 1996, Curr.
Opin. Lipidol. 10, 151-159, 1999).

[0003] Since it was revealed that fibrate compounds which are therapeutic agents for hyperlipidemia are agonists of PPARα, and that thiazolidine derivatives known as therapeutic agents for diabetes are agonists of PPARγ, these compounds have been followed by PPARα. The search for agonists has been advanced, and development as an agent for improving hyperlipidemia and diabetes has been attempted. So far, as PPAR agonists, in addition to thiazolidine derivatives, fibrate compounds, fatty acids, leukotriene B4, indomethacin, ibuprofen, fenoprofen, 15-deoxy-Δ-12, 14-PGJ2, etc. (Cell.
83, 813-819, 1995, J. Biol. Chem. 272 (6) 3406-3410,
1997, Proc. Natl. Acad. Sci. USA. 94, 4321-4327,
1997, J. Biol, Chem. 274 (10) 6718-6725, 1999, Mebio
Separate volume; Multiple Risk Factor Syndrome 2, 88-96, 199
9) has been reported. However, these compounds have problems such as side effects due to long-term ingestion.

[0004]

An object of the present invention is to provide a PPAR-dependent gene transcription activator which is excellent in safety and is effective in preventing and improving obesity, diabetes, hyperglycemia, hyperlipidemia and insulin resistance. Is to provide.

[0005]

Means for Solving the Problems The present inventors have searched for a PPAR-dependent gene transcription activator which is safe even if taken for a long time and has few side effects, and which is present in natural foods and has a long eating experience. It has been found that gallic acid derivatives, quercetins, flavone derivatives, isoflavone derivatives and catechins having the same have the action.

That is, the present invention provides the following (A) to (F): (A) gallic ester, (B) galloyl tannins,
(C) Quercetin (3, 3 ', 4', 5,7-pentah
ydroxyflavone) or its glycoside, (D) flavone or its analog, (E) isoflavone or its analog, (F) peroxisome proliferator-activated receptor-dependent gene transcription comprising a compound selected from catechin or epicatechin An activator is provided.

[0007]

BEST MODE FOR CARRYING OUT THE INVENTION The gallic acid ester (A) used in the present invention includes a linear or branched alkyl or alkenyl ester having 1 to 24 carbon atoms which may have a hydroxy group. . Particularly, those having 1 to 10 carbon atoms are preferable, and esters such as methyl, ethyl, propyl, butyl, and octyl are preferable.

[0008] The (B) galloyl tannins used in the present invention are tannins that generate gallic acid by hydrolysis, and more specifically, glucose, hamamelose, xylose, galactose, methyl glucoside, quinic acid, shikimi. Acids, catechins, gallocatechin, epigallocatechin and other hydroxyl groups galloylated to various degrees,
Examples thereof include various mixtures thereof called tannic acid. Examples of the galloyl tannin include 1, 2, 3,
6-tetragalloylglucose, 1,2,3,4,6-
Pentagalloylglucose, 3,4-digalloylshikimic acid, hamamelitannin, catechin gallate, epicatechin gallate, epigallocatechin gallate, assamicin A, assamicin B, theaflavin gallate, procyanidin gallate, eugenin, punica cortein A, punigluconin, steno Filanin A and the like. Particularly, epigallocatechin gallate and epicatechin gallate are preferable.

(C) Quercetin used in the present invention is a kind of flavonol, a yellow pigment widely distributed in the plant kingdom,
Many of them exist as glycosides.

[0010] Examples of the sugar residue of the quercetin glycoside include glucose, raffinose, rhamnose, galactose, maltose, fructose, N-acetylglucosamine and the like. Among them, glucose, raffinose and rhamnose are preferred. The number of sugar residues is 1 to
50, usually 1 to 20, and the solubility in water can be adjusted by the number of sugar residues. One example of quercetin glycoside is Rutin (Quercetin-3-rutinos)
ide), Quercitrin (Quercetin-3-ram)
unoside), Quercimeritrin (Que)
rcetin-7-glucoside), αG-rutin (Toyo Seishi Co., Ltd.) and the like.

Among the gallic esters, galloyl tannins and quercetin or glycosides thereof, particularly the galloyl tannins and quercetin or glycosides thereof are:
Since these compounds are contained in plants or crude drugs, these compounds may be contained in the PPAR-dependent gene transcription activator of the present invention by blending plants, crude drugs or extracts thereof containing them. . The crude drugs containing galloyl tannins include gallic and quintular, and the plants include tea, grape and the like. Here, the tea includes black tea and oolong tea in addition to green tea, and green tea is particularly preferable. In addition to grape fruits, grape seeds and pericarp containing a large amount of galloyl tannins, or extracts or dry powders thereof are particularly preferable. Examples of plants containing quercetin or its glycoside include Rutaceae (Ruta graveolens).
L.), soybeans (Sophora japonica L.), and soybean buckwheat (Fagopyrum esculentum Moench). It is particularly preferable that these florets contain about 20% by weight. These herbal medicines and plants may be used by directly drying and grinding as described above,
It may be used after extraction with an extraction solvent such as water, alcohols, ethers, and halogenated hydrocarbons.

The (D) flavone and its isomer (E) isoflavone used in the present invention are a kind of polyphenol, and usually have a hydroxyl group or a methoxy group added thereto, or are used as glycosides in the plant kingdom. Widely exist in.
These glycosides are also included in (D) and (E). (D) Flavone analogs include apigenin, acacetin, luteolin, chrysin, myricetin, and the like. (E) Isoflavone analogs include daidzein, genistein, prunetin, aflomosin, irigenin, formonetin, daidzin, genistin bio Canin, glycitin and the like. It is known that these various analogs are abundant in plants such as Rosaceae, Legumes and Iridaceae. In particular, isoflavones are contained abundantly in legumes, have many years of dietary experience, and are highly safe even if taken for a long time. Examples of plants or crude drugs containing flavone or an analog thereof, isoflavone or an analog thereof include parsley, soybean, perilla, and the like. A parsley extract and a perilla seed extract containing a large amount of flavonoids and a soybean extract containing a large amount of isoflavonoids are preferred. Specific products include, for example, Applephenon (Nikka Whiskey Co., Ltd.), Soyact (Kikkoman Co., Ltd.), and Geravinol (Kikkoman Co., Ltd.).

The catechin (3) used in the present invention (3)
5,7,3 ', 4'-Pentaoxyflavan) or epicatechin can be extracted from green teas such as sencha, incense tea, potted tea, semi-fermented tea, and fermented tea leaves with water or hot water. Good. Further, an organic acid such as sodium ascorbate or a salt thereof may be added in advance to the extraction water. A method of extracting under a non-oxidizing atmosphere from which dissolved oxygen in the extraction water has been removed by boiling degassing or passing an inert gas such as nitrogen gas may be used in combination. Instead of extracting from tea leaves, a concentrate of green tea extract may be dissolved in water and used, or an extract from tea leaves and a concentrate of green tea extract may be used in combination. Here, the concentrate of green tea extract is a concentrate obtained by extracting green tea leaves with hot water or a water-soluble organic solvent, and is, for example, disclosed in JP-A-59-219384 and JP-A-Hei. 20
No. 589, JP-A-5-260907, JP-A-5-306279, and the like. Commercially available Polyphenon, Mitsui Norin Co., Ltd., Theafran, Itoen Co., Ltd., Taiyo Chemical Co., Ltd.
A “Sanphenon” series or the like may be used. In addition, catechin or epicatechin is derived from other raw materials,
A column-purified product, a chemically synthesized product, or the like may be used.

The PPAR-dependent gene transcription activator of the present invention may be formulated in any form together with excipients and other additives used in pharmaceuticals and foods.

The PPAR-dependent gene transcription activator of the present invention can be administered or ingested to humans orally, enterally, transdermally, by injection, transmucosally or the like. The amount varies depending on various factors such as age, body weight, sex, and administration method. In the case of oral administration, the amount is usually 0.1 to 0.1 per adult.
It is preferred to administer the dose in the range of 5000 mg, especially 100 to 2000 mg, once to several times a day.

The dosage forms of the PPAR-dependent gene transcription activator of the present invention include those used in medicines and foods, such as tablets, powders, granules, capsules, troches, syrups, various forms of beverages, Snacks, dairy products, seasonings, processed starch products, processed meat products, and the like.

The PPAR-dependent gene transcription activator of the present invention is usually incorporated in the preparation in an amount of 0.001 to 80% by weight.

[0018]

EXAMPLES Example 1 PPAR-dependent gene transcription activation test: small intestinal epithelial cell line IEC-6 or hepatocyte cell line Hep
G2 is spread on a 12-well plate and cultured in DMEM (5% FCS) for 1 day. PPAR response sequence (underlined) (AACG TGAC
CTTTGTCCT GGTCAACG TGACCTTTGTCCT GGTC AACG TGACCTTTGTC
CT GGTC), SV40 promoter gene,
A PPAR reporter plasmid (PPAR-Luc) containing a firefly luciferase gene and a control plasmid (TK-Luc: Promeg) in which a thymidine kinase promoter gene is linked upstream of the Renilla luciferase gene
a) at the same time to give 0.5 μg / well of each transfection reagent (Superfect transfection reagent;
Promega). Thereafter, the culture solution was replaced with a DMEM (-FCS) medium containing a test substance, and the cells were further cultured for 24 hours. After washing with PBS, the cells were lysed using a dual luciferase assay system (Promega), a substrate solution containing luciferin was added to the lysate, and the firefly and Renilla luciferase activities were measured with a luminometer. By measuring the PPAR-dependent gene transcription activity (luciferase activity) in this experimental system,
An AR-dependent gene transcription activator was searched for. still,
The PPAR-dependent gene transcription activity (luciferase activity) was defined as follows.

PPAR-dependent gene transcription activity (luciferase activity) = (firefly luciferase activity by PPAR-Luc) / (renilla luciferase activity by TK-Luc)

Table 1 shows the ability of each test substance to activate PPAR-dependent gene transcription in IEC-6 and HepG2 cells. The PPAR-dependent transcription activity in the control is defined as 100, and the relative value is shown. Fenofibrate was used as a positive control.

[0021]

[Table 1]

Table 1 shows that gallic ester (1) and galloyl tannins are effective for PPAR-dependent gene transcription activation.

Example 2 A PPAR-dependent gene transcription activation test was performed in the same manner as in Example 1. In addition, IEC-6 was used as a cell and carbacycline was used as a positive control. Table 2 shows the results.

[0024]

[Table 2]

Table 2 shows that quercetin or its glycoside is effective for PPAR-dependent gene transcription activation.

Example 3 A PPAR-dependent gene transcription activation test was performed in the same manner as in Example 1. Table 3 shows the results.

[0027]

[Table 3]

As shown in Table 3, flavone or an analog thereof,
It can be seen that isoflavone or an analog thereof is effective in activating PPAR-dependent gene transcription.

Example 4 A tablet (200 mg / tablet) was produced according to a conventional method using the following components. (Composition) (g) Ethyl gallate or quercetin 1000 Hydroxypropylcellulose 800 Light silicic anhydride 200 Lactose 500 Crystalline cellulose 500 Talc 500 Diacylglycerol 300

Example 5 A tablet of 200 mg / tablet was prepared using the following components according to a conventional method. (Composition) (g) Propyl or rutin gallate 20 Starch 130 Magnesium stearate 10 Lactose 40

Example 6 After mixing the following ingredients, the mixture was filled into a gelatin capsule, and one tablet
50 mg of soft capsules were obtained. (Composition) (g) Propyl gallate or rutin 20 Epigallocatechin gallate or αG-rutin 20 Triglyceride (rapeseed oil) 60

Example 7 The following ingredients were mixed and filled into a gelatin capsule to obtain a soft capsule. (Composition) (g) Ethyl gallate or quercetin 20 Epigallocatechin gallate or αG-rutin 20 Soybean oil 200 γ-oryzanol 10 Caffeine anhydride 10 Docosahexaenoic acid 10 Eicosapentaenoic acid 10 α-Linolenic acid 10 Vitamin C 10

Example 8 After mixing the following ingredients, the mixture was filled in a gelatin capsule, and one tablet 2
50 mg soft capsules were obtained. (Composition) (g) Soybean extract (Soyact (Kikkoman Co., Ltd.)) 20 Tea extract (Theafuran 90S (Itoen Co., Ltd.)) 20 Gallic extract (alcohol extraction, dried product) 10 Triglyceride (seed oil) 50

[0034]

Industrial Applicability The PPAR-dependent gene transcription activator of the present invention has an excellent PPAR activating effect and is also highly safe, so that it is effective for obesity, diabetes, hyperglycemia, hyperlipidemia, insulin resistance. It is useful as a preventive or ameliorating agent.

[0035]

[Sequence List] SEQUENCE LISTING <110> KAO CORPORATION <120> Activators for PPAR-dependent gene transcription <130> P01591304 <140> <141> <150> JP 2000-186385 <151> 2000-6-21 <160> 1 <170> PatentIn Ver. 2.1 <210> 1 <211> 63 <212> DNA <213> RAT <400> 1 aacgtgacct ttgtcctggt caacgtgacc tttgtcctgg tcaacgtgac ctttgtcctg 60 gtc 63

Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat II (reference) A61K 35/78 A61K 35/78 K N Q X A61P 3/04 A61P 3/04 3/06 3/06 3/10 3 / 10 43/00 105 43/00 105 // C07D 311/30 C07D 311/30 311/36 311/36 C12N 15/09 ZNA C12N 15/00 ZNAA (72) Inventor Ichiro Tokimitsu Akabane, Kaimachi, Haga-gun, Tochigi Prefecture 2606 Kao Corporation Company F-term (reference) 4B024 AA11 CA02 FA02 GA11 HA20 4C062 EE49 EE53 EE76 4C086 AA01 AA02 BA08 MA09 NA14 ZA70 ZB21 ZC33 ZC35 4C088 AB38 AB40 AB43 AB45 AB56 AB59 AB61 AB62 NA14 ZA30 ZAA ZAB ZAB ZA30 ZAB DB41 MA13 NA14 ZA70 ZB21 ZC33 ZC35

Claims (5)

[Claims] 1. The following (A) to (F): (A) gallic ester, (B) galloyl tannins,
(C) quercetin or its glycoside, (D) flavone or its analog, (E) isoflavone or its analog, (F) a peroxisome proliferator-activated receptor-dependent receptor consisting of a compound selected from catechin or epicatechin Gene transcription activator. 2. A peroxisome proliferator-activated receptor-dependent gene transcription activity comprising a plant, a crude drug, or an extract thereof, which contains at least one compound of (A) to (F) according to claim 1. Agent. 3. The plant or herbal medicine containing at least one of (A), (B) and (C) according to claim 1 is Henruda, Enju, buckwheat, gallic, quintet, tea or grape. Item 4. The peroxisome proliferator-activated receptor-dependent gene transcription activator according to Item 2. 4. A plant or crude drug containing one or more compounds of (D) or (E) according to claim 1,
The peroxisome proliferator-activated receptor-dependent gene transcription activator according to claim 2, which is parsley, soybean or perilla. 5. The peroxisome proliferator-activated receptor-dependent gene transcription activator according to claim 2, wherein the plant containing (F) according to claim 1 is tea.