JP2008115163A - Adiponectin production enhancer and stimulator - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide an enhancer and a stimulator for the natural product derived highly safe adiponectin production, and drugs, food and drink preparations containing them. <P>SOLUTION: An adiponectin production enhancer and stimulator having an effective ingredient of naringenin chalcone or its derivative. Drugs, and food and drink preparations containing the adiponectin production enhancer and stimulator. These agent, drugs, and food and drink preparations have potent adiponectin production enhancing and stimulating actions and are highly safe, and useful for prevention and improvement of arteriosclerosis or diabetes caused by insufficient secretion of adiponectin, and metabolic syndromes. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention relates to a novel adiponectin production enhancer / promoter, and pharmaceuticals and foods and drinks containing them.

  With recent rapid changes in lifestyle, obesity with visceral fat accumulation, or metabolic syndrome based on it (hereinafter abbreviated as “MS”) is steadily increasing. According to the results of the 2004 National Health and Nutrition Survey, the total number of MS prevalence and reserve soldiers between the ages of 40 and 74 is estimated to be 19.6 million.

  MS is a state in which hyperglycemia, hypertension, hyperlipidemia and the like are based on obesity accompanied by visceral fat accumulation, and the risk of developing arteriosclerotic disease is extremely high. Atherosclerotic disease is at the same level as a malignant neoplasm in Japanese cause-of-death statistics, and establishment of preventive and therapeutic measures for MS, which is the risk of developing it, is extremely important.

  Factors that cause the onset of MS include lifestyle disturbances such as lack of exercise and excessive lipid intake, and external stresses such as aging and smoking. The accumulation of these causes obesity accompanied by the accumulation of visceral fat. As this progresses, the balance of adipocytokines, which are bioregulators secreted from fat cells, is lost.

  Among adipocytokines, adiponectin is particularly closely related to MS, and it has been reported that the adiponectin concentration in coronary artery disease patients, type 2 diabetes patients, and obese patients is significantly lower than that in healthy individuals. In addition, in the analysis using adiponectin knockout mice, it has been reported that knockout mice are more prone to intimal thickening due to vascular injury than wild type and show stronger insulin resistance (for example, Non-Patent Documents 1 and 2). reference). From these facts, it has been found that adiponectin is an adipocytokine that suppresses the risk of type 2 diabetes with insulin resistance and arteriosclerosis.

Actually, when adiponectin is overexpressed in apoE knockout mice that develop atherosclerosis at a high rate, the progression of arteriosclerotic lesions is suppressed (for example, see Non-Patent Document 3), and the blood adiponectin concentration It has been reported that supplementation of physiological concentrations of adiponectin to KKA ^y and db / db mice, which are obese and type 2 diabetic mice exhibiting a low value compared to the wild type, improves insulin resistance (eg, Non-patent document 4). As described above, adiponectin has been found to improve MS component factors such as insulin resistance of type 2 diabetes, hypertension, lipid metabolism, anti-arteriosclerosis and anti-inflammatory effects. It was also confirmed to have liver fibrosis suppression and normal hepatocyte proliferation promoting effects (see, for example, Non-Patent Document 5).

  From these facts, adiponectin is the most important factor in considering the risk reduction by MS, and the enhancement and promotion of adiponectin production / secretion in adipocytes, which are specific expression tissues, is essential prevention of MS・ It can be said that it is useful as a treatment.

  There are thiazolidine derivatives that have been used as therapeutic agents for type 2 diabetes as known compositions / compounds having an effect of enhancing / promoting adiponectin production / secretion in adipocytes. In addition to safety issues, prescriptions are limited to type 2 diabetes patients under the control of doctors due to the fact that they are pharmaceuticals, and MS prevention for MS patients and reserves who have a relatively mild risk accumulation state・ There is a problem that it cannot be used for improvement. Therefore, there is a demand for the development of foods and drinks that are derived from natural products, have high safety, and are effective in enhancing and promoting adiponectin production and secretion.

  So far, it has been reported that hop-derived xanthohumol increases adiponectin in blood as a substance having an adiponectin regulating action derived from a natural product (see, for example, Patent Document 1). However, it is not clear that chalcones having a structure different from this can safely enhance or promote the production of adiponectin without showing side effects such as cytotoxicity in adipocytes where adiponectin is produced.

JP 2006-306800 A Matsuda et al. (M Matsuda, et al.), “Role of adiponctin in preventing vascular tenosis. The missing link of adipo-vascal axis. Bio, USA”. The American Society for Biochemistry and Molecular Biology), 2002, 277 (40), p. 37487-37491 Kondo et al. (H Kondo, et al.), “Association of adipectinintion with type 2 diabetics: a candidate gene for the insulin resistance, et al. 51 (7), p. 2325-2328 Okamoto et al. (Y Okamoto, et al.), “Adipectin Reduce Atherosclerosis in Apolipoprotein E-Defective Mice.” Circulation, (USA), American Heart Association (American). 2767-2770 ”Yamato et al. (T Yamauchi, et al.),“ The fat-derived hormone adiponectin reverse, insulative resistance associating with the Neptune, and the United States. ” (8), p. 941-946 J. et al., “The roles of leptin and adipontin: A novel paradigm in adiposito ur te ri ge o ri te ri t er e ri t er o t e n e, t e n e t e n e t e n e, ed. (American Society for Investigative Pathology), 2005, 166, p. 1655-1669

  The problem to be solved by the present invention is to provide an adiponectin production enhancing / promoting agent having a structure different from the conventional one, which is derived from a natural product and has high safety, and a pharmaceutical and a food or drink containing them.

  As a result of intensive studies to solve the above problems, the present inventors have found that naringenin chalcone or a derivative thereof has an action of enhancing and promoting the production of adiponectin, and based on these findings, the present invention has been completed. .

That is, the present invention
1) Adiponectin production enhancing / promoting agent comprising naringenin chalcone or a derivative thereof as an active ingredient.
2) The adiponectin production enhancing / promoting agent according to 1) above, wherein naringenin chalcone or a derivative thereof is contained in the tomato extract.
3) A pharmaceutical or a food or drink containing the agent according to 1) or 2) above.
Is to provide.

  The adiponectin production enhancing / promoting agent comprising naringenin chalcone or a derivative thereof of the present invention as an active ingredient is highly safe and enhances / promotes the production of adiponectin in adipocytes. As a result, it becomes possible to prevent or improve arteriosclerosis or diabetes caused by adiponectin secretion failure or metabolic syndrome. The phrase “caused by adiponectin secretion failure” means that secretion of adiponectin into the blood becomes abnormal due to some cause such as abnormal expression or production of adiponectin.

Hereinafter, the present invention will be described in detail.
The adiponectin production enhancing / promoting agent of the present invention comprises naringenin chalcone or a derivative thereof as an active ingredient. Naringenin chalcone (2′4′6′4-Tetrahydrylcharcone) is a substance represented by the following structural formula (II).
Structural formula (II)

  Naringenin chalcone may be a derivative in which a part of its structure is altered or modified as long as it has an adiponectin production enhancing / promoting action. Examples of derivatives of naringenin chalcone include pharmacologically acceptable salts, esters or prodrugs.

  Among the derivatives of naringenin chalcone, pharmacologically acceptable salts are not particularly limited. For example, alkali metals (sodium, potassium, etc.), alkaline earth metals (magnesium, calcium, etc.), hydroxides or carbonates thereof. And salts with alkali metal alkoxides (sodium methoxide, potassium t-ptoxide, etc.). Also, as salts, acid addition salts to which inorganic acids (hydrochloric acid, sulfuric acid, phosphoric acid, etc.) and organic acids (maleic acid, citric acid, fumaric acid, etc.) have been added, addition salts of amines, addition salts of amino acids, etc. Is mentioned. In addition, the hydrate of said salt is also contained in the salt here.

  Among the derivatives of naringenin chalcone, the ester is not particularly limited as long as it is an ester generated by an esterification reaction with an alcohol or a carboxylic acid. Examples of the alcohol include methanol, ethanol, 1-propanol, and 2-propanol, and examples of the carboxylic acid include formic acid, acetic acid, and lactic acid.

  Among the derivatives of naringenin chalcone, prodrug means a compound that changes to naringenin chalcone after being administered to a living body and exhibits an action as an adiponectin production enhancer / promoter. Derivatives referred to in the present invention also include naringenin chalcone prodrugated for the purpose of improving stability and absorbability and reducing side effects.

  Naringenin chalcone, which is an active ingredient of the adiponectin production enhancer / promoter of the present invention, may be produced by any method, such as a method for purification from an organism containing naringenin chalcone, a chemical synthesis method, a semi-synthetic method, etc. Can be widely adopted.

  Naringenin chalcone can be extracted from plants such as citrus fruits, vegetables, specifically tomatoes. The method for purifying naringenin chalcone from tomato is not particularly limited. For example, 10 to 15 times the amount of 50 to 90% ethanol is added to tomato peel or squeezed rice cake, and extraction is performed at 40 to 80 ° C. for 1 to 5 hours. Since an extract containing naringenin chalcone is obtained, the naringenin chalcone is obtained by subjecting the extract to silica gel using an organic solvent such as ethyl acetate and n-hexane as an eluent and an ODS reverse phase resin using an eluent as a water-acetonitrile solvent. Can be purified. The adiponectin production enhancer / promoter of the present invention is not limited to one containing only purified naringenin chalcone, and may be a crude product containing naringenin chalcone.

  Among the derivatives of naringenin chalcone, a pharmacologically acceptable salt can be produced by allowing an alkali metal, alkaline earth metal, alkali metal alkoxide, inorganic acid or organic acid to act on naringenin chalcone. Furthermore, among the derivatives of naringenin chalcone, an ester can be produced by allowing an alcohol or carboxylic acid to act on naringenin chalcone in the presence of an acid catalyst.

  The adiponectin production enhancing / promoting agent of the present invention exhibits adiponectin production enhancing / promoting activity in adipocytes. Therefore, it is useful for prevention and treatment of arteriosclerosis, diabetes and metabolic syndrome caused by adiponectin secretion failure. The adiponectin production enhancing / promoting agent can be used as it is or added to these products as a pharmaceutical, food / drink or cosmetic product having adiponectin production enhancing / promoting activity. The adiponectin production enhancer / promoter of the present invention may be used alone as a pharmaceutical product, food / beverage product or cosmetic, or may be used in combination with other adiponectin production enhancer / promoter.

<Pharmaceuticals>
The adiponectin production enhancer / promoter of the present invention can be used as a medicine as it is or by formulating it with a known pharmaceutical carrier. The adiponectin production enhancing / accelerating agent of the present invention can be formulated as a parenteral preparation such as an oral preparation such as a tablet, granule, powder or syrup, or a suppository or an external preparation. The pharmaceutical carrier is not particularly limited, and examples thereof include solid carriers (starch, lactose, carboxymethyl cellulose, etc.), liquid carriers (distilled water, physiological saline, aqueous glucose solution, ethanol, propylene glycol, etc.), oily carriers (various types). Animal and vegetable oils, white petrolatum, paraffin, etc.).

  The above pharmaceutical products can be used for humans and animals other than humans (pets, livestock). The dose of the above medicine may be appropriately set according to the symptoms, sex, and age of the patient or the like who uses it, for example, once or several times a day for each adult, 0.1 What is necessary is just to take so that about -1000 mg can be ingested.

<Food &Drink>
By adding the adiponectin production enhancing / promoting agent of the present invention to a food or drink, the adiponectin production enhancing or promoting activity can be imparted to the food or drink. The food and drink to be added is not particularly limited, and examples thereof include meat products, processed vegetables, side dishes, dairy products, confectionery, bread, soft drinks, fruit drinks, and alcoholic beverages. The mixing ratio of the adiponectin production enhancing / accelerating agent of the present invention to food is not particularly limited.

  Further, the adiponectin production enhancing / promoting agent of the present invention and other food materials may be mixed and formed into granules, powders, tablets, blocks, or the like, and used as food materials or health foods. Examples of other food materials include sugars, edible proteins, alcohols, vitamins, thickening polysaccharides, amino acids, calcium salts, pigments, fragrances, and preservatives.

<Cosmetics>
The adiponectin production enhancing / promoting agent of the present invention can be added to cosmetics to impart adiponectin production enhancing / promoting activity to the cosmetics. Examples of cosmetics include, but are not limited to, lotions, cosmetic creams, emulsions, foundations, lipsticks, hair styling agents, hair tonics, hair restorers, toothpastes, mouthwashes, shampoos, rinses, and the like. When preparing cosmetics, oils and fats such as vegetable oils, waxes such as lanolin and beeswax, hydrocarbons, fatty acids, higher alcohols, various surfactants, pigments, fragrances, vitamins, plant and animal extract components It can be produced by appropriately blending those used as usual cosmetic raw materials such as ultraviolet absorbers, antioxidants and preservatives.

  Hereinafter, the present invention will be specifically described with reference to examples. However, the technical scope of the present invention is not limited by these descriptions.

<Purification of Naringenin Chalcone>
20 kg of dried tomato peel was washed with 240 l of water at 50 ° C. for 2 hours to extract and remove impurities. Next, the extracted soot was recovered, and 240 l of 70% ethanol was added as a solvent, and extracted for 2 hours while heating at 70 ° C. The obtained extracted solution was filtered off, and the filtrate was concentrated under reduced pressure. The concentrate was purified with silica gel using an organic solvent such as ethyl acetate and n-hexane as an eluent, and purified with a reverse phase resin of ODS using a water-acetonitrile solvent as an eluent to obtain naringenin chalcone.

<Evaluation of adiponectin secretion promoting effect by cultured adipocytes>
The adiponectin production and secretion promoting action of various chalcones in mouse 3T3-L1 adipocytes was confirmed by ELISA. Mouse 3T3-L1 preadipocytes were seeded on a 24-well plate pre-collagenized so as to have 5 × 10 ⁴ cells per well, and cultured in DMEM containing 10% bovine serum. After the preadipocytes become confluent, the cells are further cultured for 2 days, cultured in DMEM containing 0.5 mM isobutylmethylxanthine, 1 μM dexamethasone, 10 μg / ml insulin, 10% fetal calf serum, and differentiated into adipocytes. Guidance was performed. Thereafter, the medium was replaced with DMEM containing 5 μg / ml insulin and 10% fetal bovine serum, and various chalcone DMSO solutions were added so that the DMSO concentration was 0.1%. The treatment concentrations of the various chalcones are as follows: control group (DMSO 0.1% only); Naringenin chalcone treatment group obtained in Example 1; 4-methylchalcone treatment group; 4'-methylchalcone treatment group; Trans-chalcone treatment group; Trans-4,4′-difluorochalcone treatment group; 2-Hydroxychalcone treatment group (all treated with 30 μg / ml). This medium was newly replaced every 2 days, and after 7 to 10 days from the start of differentiation induction, the culture supernatant was collected. The amount of adiponectin in the culture supernatant was measured with an adiponectin ELISA kit (Otsuka Pharmaceutical). The results are shown in FIG. In the chalcone treatment group other than naringenin chalcone, cell death due to cell injury was confirmed 24 hours after the start of treatment (micrograph is shown in FIG. 2), and adiponectin was not measured. From these results, it can be seen that, among various chalcones, only naringenin chalcone promotes the secretion of adiponectin safely without causing side effects of cytotoxicity.

<Evaluation of adiponectin gene expression enhancement by cultured adipocytes>
The adiponectin gene expression enhancing effect in mouse 3T3-L1 adipocytes cultured and differentiated in the same manner as in Example 2 was confirmed by quantitative RT-PCR. The mouse 3T3-L1 adipocytes from which the culture supernatant was collected in Example 2 were washed twice with PBS and then collected with 0.4 ml of trizol (Invitrogen) per well to extract RNA. DNA and protein were removed according to the attached instructions, and total RNA was recovered by isopropyl alcohol precipitation, then washed with 70% ethanol and air-dried. This RNA was converted to cDNA by performing a reverse transcription reaction with ExScript RT reagent Kit (TAKARA), and a quantitative PCR method was performed using this as a template, and the adiponectin gene expression level was measured. The primers used were as follows: mouse adiponectin; 5′-GATGGGCAGAGATGGCACTCC-3 ′ (SEQ ID NO: 1) and 5′-CTTGCCAGTGCTGCCGGTCAT-3 ′ (SEQ ID NO: 2), mouse cyclophilin as control; 5′-CAGAGCCCACTGTGCGCTTTT-3 '(SEQ ID NO: 3) and 5'-TGTCTTTGGAACTTTGCTCAA-3' (SEQ ID NO: 4). The quantitative PCR reaction was performed using MX3000P (STRATAGENE) and SYBR Premix Ex Taq kit (TAKARA) according to the attached instructions. The results are shown in FIG. It can be seen that adiponectin gene expression is enhanced by the addition of naringenin chalcone, and it can be seen that naringenin chalcone enhances and promotes adiponectin secretion and gene expression in combination with Example 2.

<Evaluation of blood adiponectin secretion promoting effect in obese / diabetic model animals>
The effect of promoting adiponectin secretion in vivo was confirmed using KK-Ay mice (CLEA Japan), which is a model animal for obesity and type 2 diabetes. 5-week-old male mice were pre-bred with MF diet (oriental yeast) for 1 week, and then grouped so that there was no difference between groups in blood glucose and body weight. Each group consisted of 5 animals, and grouping was performed as follows: control group (MF feed); 0.02% NGC group (mixed with MF feed so that Naringenin chalcone is 0.02%), C.I. 0.1% NGC group (mixed with MF feed so that Naringenin chalcone is 0.1%). Animals were reared for each individual, and water and food were freely consumed. Body weight and food consumption were recorded once every two days. There were no differences in body weight or food intake between groups during the study period. After breeding for 4 weeks, blood was collected from the heart under anesthesia, and the amount of adiponectin in the obtained serum was measured with an adiponectin ELISA kit (Otsuka Pharmaceutical). The results are shown in FIG. It can be seen that the concentration of adiponectin in the blood is increased depending on the intake of naringenin chalcone.

  The adiponectin production enhancing / promoting agent obtained in the present invention has an excellent adiponectin production enhancing / promoting action. Therefore, by using these, it becomes possible to provide a preventive / ameliorating agent for arteriosclerosis or diabetes caused by adiponectin secretion deficiency and metabolic syndrome. Furthermore, it becomes possible to provide pharmaceuticals, foods and drinks or cosmetics containing these.

The adiponectin concentration in the cell culture supernatant was measured by ELISA. (Example 2) The state of the cell 24 hours after the start of sample processing was photographed with a microscope. (Example 2) The expression of adiponectin gene in the cells was measured by quantitative PCR. (Example 3) The adiponectin in blood of KK-Ay mice, which are obesity / type 2 diabetes model animals, was measured by ELISA. Example 4

Claims (3)

  An adiponectin production enhancing / promoting agent comprising naringenin chalcone or a derivative thereof as an active ingredient.   The adiponectin production enhancing / promoting agent according to claim 1, wherein naringenin chalcone or a derivative thereof is contained in a tomato extract.   A pharmaceutical or a food or drink containing the agent according to claim 1 or 2.

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