JP2010070541A - Tg synthesis inhibitor - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a TG synthesis inhibitor effective for prevention or amelioration of hyperlipidemia and obesity. <P>SOLUTION: The TG synthesis inhibitor comprises as an effective ingredient thereof ellagitannin, having a partial structure represented by formula (1) and a glucose residue in the molecule and having the 4- and 6-positions of the glucose residue substituted with a hexahydroxydiphenoyl group portion in the partial structure represented by formula (1), or a derivative thereof. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a triacylglycerol (hereinafter referred to as “TG”) synthesis inhibitor effective in preventing or improving hyperlipidemia, obesity, insulin resistance and diabetes.

  In recent years, the incidence of hyperlipidemia and obesity has increased due to the constant excessive intake of fat accompanying the westernization of dietary habits. Hyperlipidemia and obesity are also a major social problem in terms of maintaining and promoting the health of the people because they are associated with lifestyle-related diseases such as insulin resistance and diabetes.

  Hyperlipidemia, in particular, an increase in blood TG concentration, has been regarded as a problem as a risk of ischemic heart disease (Non-Patent Documents 1 to 3). In addition, serum lipids show an abnormal increase after meals, and the state in which this increase is prevalent is commonly called postprandial hyperlipidemia. Postprandial hyperlipidemia is associated with hypertension, ischemic heart disease, etc. It has been pointed out that there is a high risk of leading to the development of arteriosclerotic diseases (Non-Patent Documents 4 and 5).

  Obesity, particularly visceral fat-type obesity, is often associated with lifestyle-related diseases and has been shown to increase the risk of developing arteriosclerotic diseases (Non-Patent Documents 6 and 7). In adipocytes with excessive accumulation of fat, the production of adiponectin, which enhances the action of insulin, decreases, and plasminogen activator inhibitor (PAI-1) and tumor necrosis factor (TNF-α) that promote the onset of arteriosclerotic diseases Production of humoral factors is promoted, and this is considered to be associated with lifestyle-related diseases and onset of arteriosclerotic diseases (Non-patent Document 8).

Insulin resistance is a state in which insulin is sufficient in the blood even though there is sufficient insulin in the blood at the center of metabolic syndrome.
Insulin resistance is considered one of the causes of diabetes. Furthermore, insulin resistance and diabetes cause diabetic retinopathy, diabetic nephropathy, neuropathy, etc., as well as the risk of developing arteriosclerotic disease due to hyperlipidemia and hypertension. Has been shown by numerous epidemiological studies (Non-Patent Documents 9 and 10).
From the above, it is generally considered that prevention and / or improvement of hyperlipidemia, obesity, insulin resistance and diabetes is important for a healthy life.

  On the other hand, it is known that TG ingested as a meal is decomposed into 2-monoacylglycerol and fatty acid by the action of lipase in the small intestine and then absorbed into small intestinal epithelial cells. In addition, the absorbed lipid is regenerated into TG by the action of enzymes such as monoacylglycerol acyl substrate transferase (MGAT) and diacylglycerol acyltransferase (hereinafter referred to as “DGAT”) in small intestinal epithelial cells. Synthesized. Furthermore, the synthesized TG is released into the blood as chylomicron (CM) by associating with apoprotein B48 specifically produced by small intestinal cells. In this way, DGAT plays an important role in the uptake of TG from the small intestine into the body. In addition, DGAT is necessary for the synthesis and secretion of very low density lipoprotein (VLDL) in the liver and TG accumulation in adipose tissue. Therefore, inhibition of DGAT activity is considered to be an effective means for preventing and improving hyperlipidemia and obesity.

In addition, as a result of studies using knockout mice, DGAT-deficient mice have decreased body weight, as well as decreased blood glucose levels, improved insulin sensitivity, increased adiponectin expression in adipocytes, etc. Inhibiting the activity is considered to be a good means for preventing or improving insulin resistance and diabetes (Non-patent Document 11).
From the above, inhibiting DGAT activity makes it possible to prevent and improve hyperlipidemia, obesity, insulin resistance, and diabetes.

Representative examples of conventional approaches for the prevention and improvement of hyperlipidemia, obesity, insulin resistance and diabetes include diet therapy and appetite suppressants (Non-Patent Documents 12 and 13), lipid metabolism-improving drugs (Non-Patent Documents) A method using documents 14 and 15) can be mentioned. However, dietary therapy is very difficult to implement in the long term, and there are concerns about side effects when taking appetite suppressants and lipid metabolism improving drugs for long periods.
On the other hand, substances that inhibit DGAT activity have been searched for the purpose of preventing or improving hyperlipidemia, obesity, insulin resistance, and diabetes, and as a food material that inhibits DGAT activity, it has been included in citrus fruits so far. Flavonoids and tangeretin have been reported to inhibit DGAT activity of human liver-derived HepG2 cells (Non-patent Document 16).
Moreover, it is reported that the quinolone alkaloid contained in the fruit of the citrus plant Goshuyu (Evodia rutaecarpa) also has a DGAT inhibitory effect (Non-patent Document 17). However, the food materials found so far are not fully satisfactory in terms of their effects.

  In addition to food materials, phenoxyacetamide derivatives and oxadiazole derivatives have been disclosed as compounds that inhibit DGAT activity (Patent Documents 1 and 2), both of which have structures different from the ellagitannins used in the present invention. It is.

Elazitannin is a substance that exists widely in the plant kingdom and is known to have a strong antioxidant action. Lugosin A-G (Rugosin) is an ellagitannin identified from the Rosaceae plant Rosanas rugosa (Non-patent Documents 18 and 19). formosana) (Non-patent Document 20). Euspinin A is an ellagitannin identified from Euphorbia supina (Non-patent Document 21). Regarding lugosin, it has been reported that lugosin D has an anticancer activity, lugosin AG is effective in improving hair growth and acne (Patent Documents 3 and 4).
However, ellagitannin or a derivative thereof having a partial structure represented by the following formula in the molecule such as lugosin, isolugosin, and euppinein has an effect of preventing or improving hyperlipidemia, obesity, insulin resistance, and diabetes, or It has not been known so far that it has a DGAT inhibitory action and a TG synthesis inhibitory action.

International Publication No. 2006/082010 Pamphlet International Publication No. 2007/138304 Pamphlet JP-A-62-129220 Japanese Patent Laid-Open No. 3-188015

Carlson, L.A. et al., Acta. Med. Scand. 218, 207-211 (1985) Castelli, W.P.Am.Heart.J.112,432-437 (1986) Norio Tada, Clinical Medicine 21,215-224 (2005) NAKAMURA Haruo, History of Medicine 157,771-775 (1991) New Hyperglycerideemia Handbook, Junya, Pharmaceutical Journal (1998) Peiris, A. N., et al. Ann. Intern. Med. 110, 867-872 (1989) Matsuzawa, Y., et al. Obes. Res. 3 Suppl 2, 187S-194S (1995) Chan, J. C., et al. I, Int. J. Obes. Relat. Metab .Disord. 26, 994-1008, 2002. Lyon, C. J. and Hsueh, W. A., Am. J. Med. 115 Suppl 8A, 62S-68S, 2003. K. Hotta, K. Matsuzawa, Japanese Clinical 59, 481-486, 2001. Chen, H. C. et al., J. Clin. Invest. 11, 1715-1722 (2003) Inoue, S. et al., Am. J. Clin. Nutr. 55, 199S-202S (1992) Ryan, D.H. et al., Obes. Res. 3, 553S-559S (1995) Monk, J.P. and Todd, P.A., Drugs. 33, 539-576 (1987) Norio Tada, Ther. Res., 21, 2320-2325 (2000) Kurowska, E. M. et al., Lipids 39, 143-151 (2004) Ko, J. S. et al., Planta Med. 68, 1131-1133 (2002) Okuda, T. et al. Chem. Pharm. Bull. 30, 4230-4233 (1982) Okuda, T. et al. Chem. Pharm. Bull. 30, 4234-4237 (1982) Hatano, T. et al. Chem. Pharm. Bull. 36, 3920-3927 (1998) Agata, I. et al. Chem. Pharm. Bull. 39, 881-883 (1991)

  The present invention relates to providing a TG synthesis inhibitor effective for preventing or improving hyperlipidemia, obesity, insulin resistance and diabetes.

  As a result of searching for a substance having an action of inhibiting the activity of DGAT, the present inventors have found that ellagitannin or a derivative thereof has an excellent activity of inhibiting DGAT and can be used as a TG synthesis inhibitor or the like. .

That is, the present invention relates to the following 1) to 5).
1) Formula (1) in the molecule

The ellagitannin or derivative thereof having a partial structure represented by formula (1) and a glucose residue, wherein the hexahydrodiphenoyl group of the partial structure represented by formula (1) is substituted at the 4-position and the 6-position of the glucose residue A TG synthesis inhibitor containing as an active ingredient.
2) A hexahydrodipheno having a partial structure represented by the formula (1) and a glucose residue in the molecule, and a partial structure represented by the formula (1) at the 4th and 6th positions of the glucose residue. A prophylactic or ameliorating agent for hyperlipidemia comprising ellagitannin substituted with an yl group moiety or a derivative thereof as an active ingredient.
3) A hexahydrodipheno having a partial structure represented by the formula (1) and a glucose residue in the molecule, and a partial structure represented by the formula (1) at the 4-position and the 6-position of the glucose residue. An obesity preventive or ameliorating agent comprising ellagitannin substituted with an yl group moiety or a derivative thereof as an active ingredient.
4) Hexahydrodiphenoyl having a partial structure represented by the formula (1) and a glucose residue in the molecule, and a partial structure represented by the formula (1) at the 4th and 6th positions of the glucose residue An agent for preventing or improving insulin resistance comprising ellagitannin or a derivative thereof substituted in the base moiety as an active ingredient.
5) Hexahydrodiphenoyl having a partial structure represented by the formula (1) and a glucose residue in the molecule, and a partial structure represented by the formula (1) at the 4th and 6th positions of the glucose residue A preventive or ameliorating agent for diabetes comprising ellagitannin or a derivative thereof substituted in the base moiety as an active ingredient.

  Since the TG synthesis inhibitor, the hyperlipidemia preventive or ameliorating agent of the present invention, the obesity preventive or ameliorating agent, the insulin resistance preventing or ameliorating agent and the diabetes preventive or ameliorating agent have an excellent DGAT inhibitory action, hyperlipidemia It is useful as a pharmaceutical, food, etc. for preventing or improving obesity, insulin resistance and diabetes.

  In the present invention, ellagitannin means a generic name for tannin that has a hexahydrodiphenoyl group and that generates ellagic acid by hydrolysis and dehydration cyclization.

  In the present invention, the ellagitannin derivative means at least the following formula (1):

The ellagitannin related hydrolyzable tannin which has the partial structure represented by these.

Among these, from the viewpoint of DGAT inhibitory activity, preferred ellagitannin derivatives have a glucose residue in the partial structure represented by the formula (1), and the above formulas at the 4th and 6th positions of the glucose residue. What substituted the hexahydro diphenoyl group part of the partial structure represented by (1) is mentioned.
Further, ellagitannin also has a partial structure represented by the formula (1) and a glucose residue, and the hexahydrodi having a partial structure represented by the above formula (1) at positions 4 and 6 of the glucose residue. What substituted the phenoyl group part is preferable.

  Of the ellagitannins or derivatives thereof, those represented by the following formula (2) are preferred.

In formula (2), R ¹ represents a hydrogen atom or a galloyl group. In formula (2), “˜OR ¹ ” represents the presence of α-type and / or β-type.
In the formula (2), R ² and R ³ are the same or different and each represents a hydrogen atom or a galloyl group, or together represent a hexahydroxydiphenoyl group.
In the formula (2), R ⁴ is

[Wherein, R ⁵ and R ⁶ are the same or different and each represents a hydrogen atom or a galloyl group. ] Is represented.
In formula (2), n represents 0 or 1.

  The ellagitannin or derivative thereof of the present invention is preferably rugosin A, rugosin B, rugosin C, rugosin D, rugosin E, rugosin F, rugosin G, isolucosine A because it exists in edible plants and has high safety. And lugocin such as isorugosine B and isorugosine D; and juvenile pinins such as juspinin A. Among these, lugosin A, lugosin B, lugosin D and juspinin A are preferred because of their DGAT inhibitory activity.

The ellagitannins or derivatives thereof of the present invention include those obtained by a known chemical synthesis method, plant extracts mainly containing ellagitannins or their derivatives, or purified products thereof.
For example, as rugosin, it is possible to use an extract from the fruit or petal of Hermanus (Rosa rugosa) or Pink Rose (Rosa centifolia), or a concentrated or purified extract thereof. As a use pinin, Euphorbia supina ) Whole plant, pink crocodile (Rosa centifolia) fruit or petal, or a concentrated or purified extract thereof.

As an extraction solvent for extracting ellagitannin or a derivative thereof of the present invention, water; alcohols such as methanol, ethanol, propanol and butanol; organic solvents such as acetone are preferably used. More preferably, a solvent is used. These organic solvents may contain water.
Of these extraction solvents, it is preferable to use water and / or water-soluble organic solvents, and more preferably water, ethanol, water / ethanol mixed solution, acetone or water / acetone mixed solution, water / ethanol mixed solution or water / It is more preferable to use an acetone mixed solution. The ethanol or acetone concentration in the mixed solution is preferably 20 to 80% by volume, more preferably 30 to 70% by volume, and further preferably 40 to 60% by volume.
Moreover, you may use together the method of extracting in so-called non-oxidative atmosphere, ventilating inert gas, such as boiling deaeration and nitrogen gas, and removing dissolved oxygen.
Extraction is performed using, for example, 1 to 50 parts by weight of a solvent with respect to 1 part by weight of a plant containing the desired ellagitannin or a derivative thereof. It is preferable to immerse or heat reflux for 30 minutes to 15 days, and further for 2 hours to 10 days. Examples of the separation and purification means of the extract include activated carbon treatment, liquid-liquid distribution, column chromatography, liquid chromatography, gel filtration, and precision distillation.
Examples of stationary phases used in chromatography include strong and weak acidic ion exchange resins or strong and weak basic ion exchange resins; reverse phase resins such as octadecylated silica gel, octylated silica gel, butylated silica gel, and trimethylsilylated silica gel; silica gel Normal phase resins such as fluoridyl and alumina; aromatic synthetic resins such as styrene-divinylbenzene and methacrylic ester; modified dextran (eg, Sephadex (registered trademark) LH-20), hydrophilic vinyl polymer (For example, Toyopearl HW-40 etc.) Gel filtration chromatography filler; activated carbon and the like, among these, reverse phase resins and aromatic synthetic resins are preferred.

An example of extraction means for obtaining lugosin and youthpinin from pink rose (petal) is given, but the extraction means is not limited to this.
A water / ethanol extract is obtained from a pink rose (petal) using a water / ethanol mixture (preferably an ethanol concentration of 40 to 60% by volume). The water / ethanol extract is liquid-separated with a water / hexane solvent (preferably water: hexane (volume ratio) = 1: 3 to 3: 1), and then the aqueous layer is separated. Butanol is added to the aqueous layer, and after liquid-liquid separation with a water / butanol solvent (preferably water: butanol (volume ratio) = 1: 3 to 3: 1), the butanol layer is separated. This butanol layer is subjected to chromatography using an aromatic synthetic resin (preferably, a styrene-divinylbenzene aromatic synthetic resin), and a low-concentration ethanol water mixture (preferably an ethanol concentration of 10 to 30 volumes). %) To obtain a low-concentration ethanol-eluted fraction. Furthermore, this low-concentration ethanol elution fraction is subjected to gel filtration chromatography (preferably a modified dextran filler), washed with a methanol water mixture (preferably methanol concentration 40 to 60% by volume), ethanol, Elution with ethanol / acetone mixed solution (preferably ethanol: acetone (volume ratio) = 1: 3 to 3: 1) and containing rugosine B as an active ingredient; rugosine A, rugosine D And an ethanol / acetone fraction containing Eupinin A and containing these as active main components is obtained. The ethanol fraction and the ethanol / acetone fraction are each subjected to chromatography using a reverse-phase resin (preferably octadecylated silica gel reverse-phase resin), and the concentration of acetonitrile in the trifluoroacetic acid aqueous solution / acetonitrile mixture is one step. Elution is carried out with the above gradient, and rugosin B is isolated from the ethanol fraction, and rugosin A, rugosin D, and juspinin A are isolated from the ethanol / acetone fraction.

  The obtained extract or fraction of ellagitannin or its derivative may be used as it is, may be used as a diluted solution diluted with an appropriate solvent, or may be a concentrated extract or a dry powder, or prepared in a paste form But you can. Further, it can be freeze-dried and diluted with a solvent usually used for extraction at the time of use, for example, water, ethanol, water / ethanol mixed solution or the like. It can also be used by encapsulating in vesicles such as liposomes or microcapsules.

  The ellagitannin or derivative thereof of the present invention has an excellent DGAT inhibitory activity, as shown in Examples below. Further, a substance having DGAT inhibitory activity is suspected to have side effects if its enzyme inhibition is non-specific, but ellagitannin or a derivative thereof of the present invention is HMG-CoA reductase which is a major metabolic enzyme in the liver. It was determined that it had no effect on the activity and G6Pase activity, was specific to DGAT, and had few side effects on the human body. Therefore, the ellagitannin or a derivative thereof is a TG synthesis inhibitor, a hyperlipidemia prevention or improvement agent, an obesity prevention or improvement agent, an insulin resistance prevention or improvement agent, and a diabetes prevention or improvement agent (hereinafter referred to as “TG synthesis inhibitor”). Etc.) and can be used to produce these agents. At this time, as the TG synthesis inhibitor, etc., the ellagitannin or its derivative alone or in addition to this, a carrier that is appropriately selected as necessary, such as a carrier that is to be blended, is allowed in the following target to be blended. May be used. In addition, the said formulation can be manufactured by a conventional method according to the target object which should be mix | blended.

  Such TG synthesis inhibitors include DGAT inhibitory activity, TG synthesis inhibition, prevention or improvement of hyperlipidemia, prevention or improvement of obesity, prevention or improvement of insulin resistance or diabetes, quasi drugs It can be used as an active ingredient in foods or feeds. In addition, the TG synthesis inhibitor is based on the concept of DGAT inhibitory activity, TG synthesis inhibition, prevention or improvement of hyperlipidemia, prevention or improvement of obesity, prevention or improvement of insulin resistance or diabetes, if necessary It can be applied to foods that indicate this, functional foods, foods for the sick, and foods for specified health use.

  When the TG synthesis inhibitor of the present invention is used as an active ingredient of a pharmaceutical product, the pharmaceutical product can be administered in any dosage form. Examples of the dosage form include oral, enteral, transmucosal, injection and the like. Examples of the dosage form of the preparation for oral administration include tablets, capsules, granules, powders, and syrups. Examples of parenteral administration include intravenous injection, intramuscular injection, suppository, inhalation agent, transdermal absorption agent, eye drop, nasal drop and the like.

  In such a preparation, the TG synthesis inhibitor of the present invention may be used alone or in combination with other pharmaceutically acceptable carriers. Such carriers include, for example, excipients, binders, disintegrants, lubricants, diluents, osmotic pressure regulators, pH regulators, emulsifiers, preservatives, stabilizers, antioxidants, colorants, ultraviolet rays. Absorber, moisturizer, thickener, brightener, activity enhancer, anti-inflammatory agent, bactericidal agent, corrigent, flavoring agent, extender, surfactant, dispersant, buffer, preservative, fragrance, coating agent Etc.

  Of these dosage forms, oral administration is preferred, and the content of ellagitannin or a derivative thereof of the present invention in the preparation when used as an active ingredient of a preparation for oral administration is usually 0.01 to It is 80 mass%, and it is preferable that it is 0.5-80 mass%.

  The dosage of the above formulation may vary according to the patient's condition, weight, sex, age or other factors, but the daily dosage per adult (60 kg) for oral administration is usually ellagitannin or its The derivatives are more preferably 1 to 2000 mg, 10 to 1000 mg, and 50 to 500 mg. Moreover, although the said formulation can be administered according to arbitrary administration schedules, it is preferable to administer once to several times a day.

  When the TG synthesis inhibitor of the present invention is used as an active ingredient of food, the form of the food can be solid, semi-solid or liquid. Examples of food include breads, noodles, confectionery, jelly, dairy products, frozen foods, instant foods, processed starch products, processed meat products, other processed foods, beverages, soups, seasonings, nutritional supplements, etc. , And their raw materials. Further, like the above-mentioned oral administration preparation, it may be in tablet form, pill form, capsule form, liquid form, syrup form, powder form, granule form and the like.

  To prepare various forms of food, TG synthesis inhibitor alone or other food materials, solvents, softeners, oils, emulsifiers, preservatives, fragrances, stabilizers, colorants, UV absorption An agent, an antioxidant, a humectant, a thickener and the like can be used in appropriate combination.

  In addition, the content of ellagitannin or a derivative thereof in the food varies depending on the use form, but is usually 0.001 to 2% by mass, preferably 0.002 to 1% by mass in the beverage form, 0 0.01 to 0.5 mass% is more preferable. Moreover, in solid food forms, such as a tablet and processed food, it is 0.01-20 mass% normally, 0.02-10 mass% is preferable, and 0.1-5 mass% is more preferable.

In addition, when the TG synthesis inhibitor of the present invention is used as an active ingredient of feed, examples of the feed include livestock feed used for cattle, pigs, chickens, sheep, horses, etc., rabbits, rats, mice, etc. Examples include small animal feeds used for foods, fish foods used for tuna, eel, Thailand, yellowtail, shrimp, etc., pet foods used for dogs, cats, small birds, squirrels, and the like.
In addition, when producing feed, DGAT inhibitor alone or in addition, meat such as cattle, pigs, sheep, protein, grains, bran, moss, sugar, vegetables, vitamins, minerals In general, feed ingredients commonly used in foods, etc., and gelling agents, shape-preserving agents, pH adjusters, seasonings, preservatives, nutritional supplements, etc., commonly used in feeds are blended as necessary The feed can be processed and manufactured.
Moreover, although content of the ellagitannin or its derivative in a feed changes with the usage forms, it is 0.01-20 mass% normally, 0.02-10 mass% is preferable, 0.1-5 mass% Is more preferable.

  EXAMPLES Hereinafter, an Example is shown and this invention is demonstrated more concretely.

Production Example 1 Preparation of Test Substance For Lugosin, 20 g (12 liters) of 50 g of petal rose (Rosa centifolia) petals (hereinafter referred to as “%”) It was immersed in ethanol at room temperature for 14 days, insolubles were filtered off, and concentrated under reduced pressure to obtain an extract (solid content: 260 g). The obtained extract was liquid-partitioned in a hexane / water system, and the resulting aqueous layer was liquid-extracted with butanol. The butanol extract (51 g) obtained by concentration under reduced pressure was adsorbed on Diaion (registered trademark) HP-20 (styrene-divinylbenzene-based aromatic synthetic adsorbent) and eluted with 20% ethanol. The obtained elution fraction was concentrated under reduced pressure and freeze-dried to obtain a dried product. Furthermore, 22 g of this dried product was dissolved in 50% methanol and adsorbed on gel filtration chromatography using Sephadex (registered trademark) LH-20. After washing with 50% methanol, elution was performed with 99.5% ethanol and ethanol / acetone (1: 1). Each elution fraction was concentrated under reduced pressure and freeze-dried to obtain a 99.5% ethanol fraction (solid content 4 g) and an ethanol / acetone fraction (solid content 6.6 g).
Lugosin B (4.3 mg) was obtained by using 120 mg of the obtained 99.5% ethanol fraction and further fractionating by HPLC. Fractionation conditions are as follows.
Column: Inertsil ODS-3 (20x250mm, GL Science)
Solvent: A liquid 0.5% trifluoroacetic acid, B liquid acetonitrile A / B = 87/13 → (30 minutes) → 87/13 → (20 minutes) → 50/50
Temperature: 40 ° C., flow rate: 20 mL / min, detection: UV = 258 nm
In addition, 300 mg of the obtained ethanol / acetone fraction was used and further fractionated by HPLC to obtain Lugosin A (70 mg), Lugosin D (59 mg), and Eupinepin A (21 mg). Fractionation conditions are as follows.
Column: Inertsil ODS-3 (20x250mm, GL Science)
Solvent: A liquid 0.5% trifluoroacetic acid, B liquid acetonitrile A / B = 84/16 → (30 minutes) → 84/16 → (5 minutes) → 50/50
Temperature: 40 ° C., flow rate: 20 mL / min, detection: UV = 258 nm
In addition, when NMR analysis (JEOL-500 spectrometer; JEOL Ltd.) was performed about these, these were described in the literature, Lugosin A, Lugosin B (nonpatent literature 8), Lugosin D (nonpatent literature 9). ) And the signal of Youthpinin A (Non-Patent Document 11).

Example 1 : Evaluation of inhibition of DGAT activity As test substances, Lugosin A, Lugosin B, Lugosin D, and Eupinin A prepared in Production Example 1 above, and EGCG (epigallocatechin gallate: supplier Nagara Science Co., Ltd.) were used. Using.
From the mouse liver, a microsomal fraction was prepared as follows and used for DGAT activity measurement. The liver of C57BL / 6J mice (male, 6 weeks old) was homogenized in sucrose buffer (250 mM sucrose, 1 mM EDTA, 10 mM Tris-HCl (pH 7.5)), 4 ° C., 12,500 × g, The supernatant was collected by centrifugation for 15 minutes. The supernatant was further centrifuged at 4 ° C., 100,000 × g for 60 minutes, and the precipitate was resuspended in sucrose buffer and used as a liver microsome fraction.
50 μg of the liver microsome fraction was added to 200 μL of DGAT activity measurement buffer (250 mM sucrose, 10 mM Tris-HCl (pH 7.5), 10 mM MgCl ₂ , 0.8 mM) containing the test substances at the concentrations shown in Table 1. Incubation was carried out at 37 ° C. for 20 minutes in EDTA, 0.1% bovine serum albumin, 100 μM oleoyl-CoA (0.05 μCi / 200 μL, 55 mCi / mmol), 1.2 mM 1,2-diolein). The reaction was stopped by adding 1.5 mL of chloroform / methanol (1: 1) (volume ratio). Next, 250 μL of PBS was added and stirred, and separated into an aqueous layer and an oil layer (lipid extraction) by centrifugation. The lower chloroform layer was collected, dried with nitrogen gas, and redissolved in chloroform.
A part of the lipid dissolved in chloroform was subjected to a liquid scintillation counter and the radioactivity was measured. A portion of the extracted lipid of about 2000 dpm was fractionated using an HPTLC plate (20 × 10 cm Silica gel 60, Merck). As a developing solvent, hexane / diethyl ether / acetic acid (80: 20: 1) (volume ratio) was used. Fluorography and quantitative analysis were performed using BAS2500 (Fuji Film). The ratio of the radioactivity incorporated into TG to the total radioactivity incorporated into the extracted lipid was defined as DGAT activity.
Table 1 shows the DGAT activity in the presence of each test substance. The DGAT activity is expressed as a relative value with the solvent control DGAT activity as 100.
As shown in Table 1, Lugosin A, Lugosin B, Lugosin D and Euspinin A showed significant inhibition of DGAT activity. On the other hand, epigallocatechin gallate (EGCG) as a comparative control did not show an inhibitory effect on DGAT activity.
Thus, since ellagitannin or a derivative thereof exhibits an excellent inhibitory action on DGAT activity, a TG synthesis inhibitor, an antihyperlipidemic agent, an obesity prevention agent, an insulin resistance prevention agent, and a diabetes prevention agent It is considered useful as an agent.

Example 2 : Evaluation of inhibition of other enzyme activity The same test substance as in Example 1 was used.
Using the mouse liver microsome fraction prepared in the same manner as in Example 1, measurement of glucose 6-phosphate dephosphorylating enzyme (G6Pase) activity, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase Activity was measured.
In the measurement of G6Pase activity, a mouse liver microsome fraction (50 μg) was prepared by adding 100 μL of a reaction solution (50 mM HEPES buffer (pH 7.2), 100 mM KCl, 2.5 mM) in the presence of the test substance at the concentration shown in the table. EDTA, 2.5 mM MgCl ₂ , 1 m
(M DTT, 10 mM glucose 6 phosphate, 2 mM EDTA) at 37 ° C. for 30 minutes. Next, 400 μL of the reaction stop solution (0.42% ammonia moltbate tetrahydrate in 1N H ₂ SO ₄ , 10% S
DS, 10% ascorbic acid 6: 2: 1 mixture) (volume ratio) was added and incubated at 50 ° C. for 30 minutes. The resulting blue dye was subjected to absorbance measurement (OD ₈₂₀ ), and G6Pase activity was calculated by calculating the concentration of the resulting phosphoric acid from a standard curve prepared using an inorganic phosphoric acid solution.
HMG-CoA reductase activity, mouse liver microsomal fraction (50 [mu] g), in the presence of concentrations of the test substance shown in Table, the reaction solution containing ¹⁴ C-labeled HMG-CoA (0.128 mm HMG-CoA ( ¹⁴ C-HMG-CoA, 72 MBq / mmol), 1 mM NADPH, 10 mM DTT, 10 mM EDTA in 0.12 mM phosphate buffer (pH 7.2)), and incubated at 37 ° C. for 30 minutes. Next, one-tenth volume (20 μL) of 5N HCl was added and incubated for an additional 30 minutes (acid treatment). After adding ¹⁴ C-testosterone (0.08 nCi) as an internal standard, it was converted from the reaction product mevalonic acid of HMG-CoA by acid treatment, and the resulting mevalonolactone was extracted with an equal amount of ethyl acetate. A part of the extract was subjected to TLC fractionation using benzene-acetone (1: 1) (volume ratio) as a developing solvent. Fluorography and quantitative analysis were performed using BAS2500, and HMG-CoA reductase activity was calculated from the radioactivity contained in mevalonolactone.
Tables 2 and 3 show HMG-CoA reductase activity and G6Pase activity in the presence of each test substance, respectively. These enzyme activities are expressed as relative values with the enzyme activity of the solvent control as 100.
As shown in Tables 2 and 3, none of the DOG-type ellagitannins or derivatives thereof showed an inhibitory effect on HMG-CoA reductase activity and G6Pase activity.
Thus, ellagitannin or a derivative thereof has an excellent inhibitory action on DGAT activity, but does not show an inhibitory action on other enzyme activities, indicating that it is a compound having high specificity for DGAT activity. . This can be considered to mean that ellagitannin or a derivative thereof is a compound with few side effects.

Example 3
The following ingredients were mixed and then filled into a gelatin capsule to obtain a soft capsule of 250 mg per tablet.
(Composition) (%)
Lugosin B 2.5
Soy lecithin 10
Rape oil 87.5

Example 4
The following ingredients were mixed and then filled into a gelatin capsule to obtain a soft capsule of 250 mg per tablet.
(Composition) (%)
Lugosin B 0.5
Youth Pinin A 2.5
Soy lecithin 10
Safflower oil 87

Example 5
One tablet of 200 mg was produced according to a conventional method using the following components.
(Composition) (%)
Lugosin D or Youthpinin A 2.5
Hydroxypropylcellulose 45.5
Light anhydrous silicic acid 5
Lactose 13
Crystalline cellulose 13
Talc 13
Diacylglycerol 8

Example 6
One tablet of 200 mg was produced according to a conventional method using the following components.
(Composition) (%)
Lugosin B or Youthpinin A 5
Starch 90
Magnesium stearate 5

Example 7
30 g of green tea leaves were extracted with 1000 mL of water at 65 ° C. for 5 minutes and filtered to obtain a green tea liquid. The following composition was mixed using this green tea liquid, and then adjusted to pH 6.0 using sodium bicarbonate to produce a green tea drink for obesity / diabetes prevention / improvement.
(composition)
Lugosin B 50mg
Vitamin C 1000mg
Green tea liquid 1000mL

Example 8
The following composition was mixed to produce a juice drink for obesity / diabetes prevention / improvement.
(composition)
Lugosin B 50mg
Vitamin C 500mg
Lemon juice 5mL
475 mL of carbonated water
Fragrance Some amount Aspartame 5g

Example 9
The following composition was tableted to produce a chewable tablet food of 1000 mg per tablet.
(Composition) (%)
Youth Pinin A 2.5
Maltose 11
Lactose 30
Glucose 15
Vitamin C 20
Vitamin E 1
Cellulose 10
Xylitol 10
Fragrance 0.5

Claims (5)

In the molecule, the following formula (1)

The ellagitannin or derivative thereof having a partial structure represented by formula (1) and a glucose residue, wherein the hexahydrodiphenoyl group portion of the partial structure represented by formula (1) is substituted at the 4-position and the 6-position A TG synthesis inhibitor containing as an active ingredient. In the molecule, the following formula (1)

The ellagitannin or derivative thereof having a partial structure represented by formula (1) and a glucose residue, wherein the hexahydrodiphenoyl group portion of the partial structure represented by formula (1) is substituted at the 4-position and the 6-position An agent for preventing or improving hyperlipidemia, comprising In the molecule, the following formula (1)

The ellagitannin or derivative thereof having a partial structure represented by formula (1) and a glucose residue, wherein the hexahydrodiphenoyl group portion of the partial structure represented by formula (1) is substituted at the 4-position and the 6-position An obesity preventive or ameliorating agent comprising as an active ingredient. In the molecule, the following formula (1)

The ellagitannin or derivative thereof having a partial structure represented by formula (1) and a glucose residue, wherein the hexahydrodiphenoyl group of the partial structure represented by formula (1) is substituted at the 4-position and the 6-position of the glucose residue An agent for preventing or improving insulin resistance, comprising as an active ingredient. In the molecule, the following formula (1)

The ellagitannin or derivative thereof having a partial structure represented by formula (1) and a glucose residue, wherein the hexahydrodiphenoyl group portion of the partial structure represented by formula (1) is substituted at the 4-position and the 6-position Diabetes preventive or ameliorating agent containing as an active ingredient.