JP2007145782A - Blood sugar level depressant having momordica charantia extract as major component and food/pharmaceutical composition - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To obtain an extract having high blood sugar level depressing action from Momordica charantia of a natural plant, and to provide a blood sugar level depressant containing the same, and a food/pharmaceutical composition containing this extract. <P>SOLUTION: Three types of novel cucrubitane-type terpenoids (1-3) and eight types of known compounds (4-11) have been isolated by the fractionation which uses an index of bioactivity of a methanol extract of dried Momordica charantia. With 5,19-epoxy-3,25-dihydroxycucrubita-6,23-diene (4) and 3,7,25-trihydroxycucrubita-5,23-dien-19-al (5) which are the major components, a remarkable blood sugar level depressing activity has been observed in a male ddY mouse in an amount of 400 mg/kg. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to a compound contained in an extract of bitter gourd (scientific name: Mormodica charantia), a hypoglycemic agent based on the compound, and a food / pharmaceutical composition containing the compound.

  Diabetes is one of the most common chronic diseases in modern times and is one of the main causes of the occurrence of cardiovascular disease. Diabetes is caused by impaired normal activity of insulin, which consumes sugar in the diet. Diabetes is mainly classified into insulin-dependent diabetes and non-insulin-dependent diabetes. In recent years, the number of patients with non-insulin-dependent diabetes has increased rapidly due to lifestyle changes, increased obesity, and aging of the population. is doing.

  In general, in diabetes, even if the ability to produce insulin in the body is extremely reduced or made, the insulin is not effective due to a decrease in insulin sensitivity, etc., which causes an increase in blood sugar level, This causes various hyperglycemia symptoms. As a widely used treatment, blood glucose level is controlled by administration of a hypoglycemic agent in addition to insulin administration.

  On the other hand, in many countries including Asia, natural compounds derived from medicinal herbs are sometimes used as remedies for diseases. In these areas, there are customs of using these drugs since ancient times, and expensive synthetic drugs cannot be purchased. Recently, one of the research subjects of phytochemists is to try to find new drugs from these folk medicines. In addition, WHO research groups conclude that traditional and unique natural medicines should be used appropriately and rationally (Daniela Bagozzi, “Traditional medicine”, [online], March 2003, World. Health Organization, [September 24, 2005 search], Internet <http://www.who.int/mediaentre/factssheets/fs134/en/index.html>).

Plants known as bitter gourd or bitter gourd (scientific name Momordica charantia, Ayurvedic name: Carrera, hereinafter referred to as “nigauri”) have fruit that is effective in diabetes in Asia and some African countries Is known and is one of these natural therapeutic candidates.
More than a hundred scientific papers on bitter gourd describe their botanical and pharmacological properties. Various components such as cucurbitan type triterpene aglycone and / or glycoside have been isolated from bitter gourd. (Non-patent document 1, Non-patent document 2, Non-patent document 3, Non-patent document 4, Non-patent document 5, Non-patent document 6, Non-patent document 7). In addition, bitter taste components of bitter gourd have been reported as Mormodicides K and L, and mordicines I and II (Non-Patent Document 1, Non-Patent Document 4, etc.). However, there is no report about the concrete blood glucose level lowering effect with respect to each component in any literature.
Furthermore, some patent publications also provide a medicine or food containing a bitter gourd extract having a medicinal effect such as a blood glucose level lowering action. (Patent Document 1, Patent Document 2, Patent Document 3, Patent Document 4, etc.). However, for such an active ingredient having a blood glucose lowering action, Patent Document 3 shows that a low-molecular peptide of a water-soluble fraction shows its action, and Patent Document 4 by the present applicant shows that Ardono-water extracted from bitter gourd It has been disclosed that 1,4-lactones are effective ingredients of blood sugar level elevation inhibitors.

Okabe, H. et al., Chem. Pharm. Bull, 1982, Volume 30, p. 4334-4340 Okabe, H. et al., Tetrahedron Letters, 1982, Vol. 23, p. 77-80 Miyahara et al., Chem. Pharm. Bull. 1981, Vol. 29, p. 1561-1566 Yasuda et al., Chem. Pharm. Bull, 1984, vol. 32, p. 2044-2047 Murakami et al. Chem. Pharm. Bull, 2001, 49, p. 54-63. Begum et al., Phytochemistry, 1997, vol. 44, p. 1313-1320 Fatope et al., J. MoI. Nat. Prod. 1990, Vol. 53, p. 1491-1497 JP-A-8-92116 JP 2001-278804 A Special Table 2002-516608 gazette JP 2004-250363 A

  However, the clarification of the causative component having a blood glucose lowering action is insufficient in any literature, and all the ingredients having a blood glucose lowering action disclosed in several literatures are knowledge only about water-soluble components. It is.

  As a result of intensive studies, the present inventors have found that a compound in a specific extract of bitter gourd has a high blood glucose level lowering action, and completed the present invention.

  Accordingly, an object of the present invention is to provide an extract having a high blood glucose level-lowering action and a blood glucose level-lowering agent containing a certain compound that can be easily used from bitter gourd by a simple operation. Another object of the present invention is to provide a food / pharmaceutical composition having a hypoglycemic effect, comprising the extract or compound.

  That is, the present invention can solve the above-mentioned problem by providing a hypoglycemic agent characterized by containing an organic solvent extract of bitter gourd (scientific name: Mormorica charantia) as an active ingredient. It was.

  In the present invention, the organic solvent extract is preferably an alcohol extract. At this time, the alcohol is preferably a lower alcohol, and more preferably at least one alcohol selected from the group consisting of methanol, ethanol, propanol, and butanol.

  In the present invention, the organic solvent extract is preferably a fraction obtained by further extracting the organic solvent extract with ether or ethyl acetate.

  The present invention also provides a compound having a blood glucose lowering action represented by the following general formula 1, which is contained in an organic solvent extract of bitter gourd (scientific name: Mormorica charantia).

(In General Formula 1, R ₁ represents a hydrogen atom, a hydroxyl group, an alkyl group, an alkoxy group, or an allopyranosiloxy group, R ₂ forms a double bond in X or forms a cyclic ether with R ₅ , R ₃ represents a hydrogen atom, a hydroxyl group, an alkyl group, or an alkoxy group, R ₄ represents a hydrogen atom, a hydroxyl group, an alkyl group, or an alkoxy group, and R ₅ represents a hydrogen atom, a hydroxyl group, an alkyl group, an alkoxy group, or formyl. or represents a group, or a cyclic ether to form together with R _2, X double without including the R ₂ if either form a double bond, including R _2, or R ₂ forms a cyclic ether Represents a hydrocarbon group that forms a bond.)

  The present invention also provides a compound represented by the following structural formula 1.

  Furthermore, the present invention provides a compound represented by the following structural formula 2.

  Furthermore, the present invention provides a compound represented by the following structural formula 3.

  Furthermore, the present invention provides any of the compounds described above, which is contained in an organic solvent extract of bitter gourd (scientific name: Mormorica charantia) and has a blood glucose lowering effect. .

  The present invention also provides a compound having a blood glucose lowering effect represented by the following structural formula 4, which is contained in an organic solvent extract of bitter gourd (scientific name: Mormorica charantia).

  Furthermore, the present invention provides a compound having a blood glucose level lowering activity represented by the following structural formula 5, which is contained in an organic solvent extract of bitter gourd (scientific name: Mormorica charantia).

  In addition, the present invention is characterized by having a blood glucose level lowering action obtained by extracting bitter gourd (scientific name: Mormodica charantia) in methanol and further extracting the extract with ether or ethyl acetate. A bitter melon extract is provided.

  Furthermore, the present invention provides a food / pharmaceutical composition having any of the above-described hypoglycemic agent, compound and bitter gourd extract as an active ingredient and having a hypoglycemic effect.

  According to the present invention, an extract having a high blood glucose level-lowering action can be easily used by a simpler operation than a natural product bitter gourd.

  Among them, a specific compound can provide a hypoglycemic agent having a higher hypoglycemic effect.

  Moreover, the food / pharmaceutical composition containing the extract or compound can be easily administered to humans, and can provide a food / pharmaceutical composition for diabetic patients that is easy to take.

  Hereinafter, the present invention will be described in detail, but the present invention is not limited to the individual forms described below.

  Nigauri is an annual plant of the cucurbit family called scientific name: Momordica charantia (hereinafter also referred to as M. charantia), Ayurvedic name: Carrera. Native to India, it is cultivated in tropical Asia, China, and Japan (especially Okinawa in the west of Kanto). In general, it is cultivated for ornamental purposes, but in recent years it has been cultivated for food due to various medicinal effects. In particular, the unripe fruit is extremely bitter and the fruit part is oily, so it is served as edible as three cups of vinegar, pickles, and the ripe fruit is sweetened as the pulp that wraps the seeds. In China, the fruit has been used for medicinal purposes.

  In the present invention, there is no particular limitation on the part used as a raw material for extraction of bitter gourd, and it may be any part of whole grass, flowers, fruits, seeds, stems, etc. It is preferable to use it. The fruit part includes fruit skin, fruit juice, pulp, cotton, seeds, etc., but any part can be used partially or in combination.

  These raw materials for bitter melon extraction can be used as they are, but in order to increase the extraction efficiency, it is preferable to use a material that has been cut, pulverized, dried or the like. For example, the fruit part is cut or crushed to a certain size and then subjected to an extraction operation in a dry / undried state, or the dried fruit part is cut or pulverized to a certain size and subjected to an extraction operation, etc. It is preferable to adjust by. Although it does not specifically limit as a drying method, Hot air drying, vacuum freeze-drying, far-infrared drying, microwave heating drying, etc. are mentioned.

  An organic solvent is used as the extraction solvent in the present invention. Examples thereof include, but are not limited to, lower alcohols such as methanol, ethanol, propanol and butanol; esters such as ethyl acetate; glycols such as ethylene glycol, butylene glycol, propylene glycol and glycerine; diethyl ether and the like In addition, acetone, acetic acid, benzene, toluene, petroleum ether, hexane, etc. can be mentioned. These solvents can be used alone or in combination of two or more. In the present invention, alcohols, ethers and esters are preferably used, and more preferably lower alcohols, ethers and ethyl acetate. In the present invention, it is most preferable to first extract with methanol and re-extract from the extract with ether and ethyl acetate because of the high content of the compound according to the present invention.

  As the extraction method used in the present invention, a general method can be employed. For example, a raw fruit part or dried fruit part of bitter gourd as a raw material can be immersed in the above solvent as it is or as a cut and pulverized product under cooling, at ordinary temperature or under heating, with stirring or standing. . The obtained extraction mixture may be used as it is after removing solids by filtration or centrifugation, if necessary, or may be used after concentration of the solvent or drying. The extraction time is not particularly limited, and may be appropriately determined from 10 minutes to several tens of days according to the state of bitter gourd as a raw material.

  In particular, the extract in the present invention is preferably, for example, an ether fraction and an ethyl acetate fraction obtained from a methanol extract of bitter gourd.

  The compound of the present invention is isolated from the above extract and is represented by the following general formula 1.

In the above general formula 1, R ₁ represents a hydrogen atom, a hydroxyl group, an alkyl group, an alkoxy group, or an allopyranosiloxy group, R ₂ forms a double bond in X or forms a cyclic ether with R ₅ , R ₃ represents a hydrogen atom, a hydroxyl group, an alkyl group, or an alkoxy group, R ₄ represents a hydrogen atom, a hydroxyl group, an alkyl group, or an alkoxy group, and R ₅ represents a hydrogen atom, a hydroxyl group, an alkyl group, an alkoxy group, or formyl. or represents a group, or a cyclic ether to form together with R _2, X double without including the R ₂ if either form a double bond, including R _2, or R ₂ forms a cyclic ether It represents a hydrocarbon group that forms a bond.

Particularly preferably, R ₁ represents a hydroxyl group, a lower alkoxy group, or an allopyranosiloxy group, R ₃ represents a hydrogen atom, a hydroxyl group, or a lower alkoxy group, and R ₄ represents a hydroxyl group or a lower alkoxy group.

  Among the compounds of the general formula 1, compounds represented by the following structural formulas 1 to 5 are preferable.

  The food composition of the present invention can be applied particularly to diabetic foods. Blood sugar level control treatment can be easily performed by directly ingesting as a special purpose food such as a food for specified health use or a nutritional functional food containing the bitter gourd extract and / or compound of the present invention.

  Specifically, when used as a food composition, various foods and drinks (milk, soft drinks, fermented milk, yogurt, cheese, bread, biscuits, crackers, pizza crusts, prepared milk powder, liquid foods, food for the sick In addition, a bitter melon extract and / or a compound may be added to foods such as infant milk powder, foods such as breast milk powder, and nutritional foods. The present active ingredient can be used as it is, or can be used according to a conventional method for ordinary food compositions such as mixing with other foods or food ingredients. Moreover, about the property, the state of the food / beverage products normally used, for example, any of solid (powder, granule, etc.), paste, liquid or suspension may be sufficient.

Other ingredients are not particularly limited, but foods and drinks containing the bittern extract and / or compound of the present invention include water, proteins, carbohydrates, lipids, vitamins, minerals, organic acids, organic bases, fruit juices. Flavors and the like can be used as the main component. Examples of proteins include whole milk powder, skim milk powder, partially skim milk powder, casein, whey powder, whey protein, whey protein concentrate, whey protein isolate, α-casein, β-casein, κ-casein, β-lactoglobulin , Α-lactalbumin, lactoferrin, soy protein, chicken egg protein, meat protein and other animal and plant proteins, hydrolysates thereof; butter, milk minerals, cream, whey, non-protein nitrogen, sialic acid, phospholipid, lactose, etc. And various milk-derived components. Examples include saccharides, processed starch (in addition to text phosphorus, soluble starch, British starch, oxidized starch, starch ester, starch ether, etc.), dietary fiber, and the like. Examples of the lipid include animal oils such as lard, fish oil, etc., fractionated oils, hydrogenated oil, transesterified oil, etc .; palm oil, safflower oil, corn oil, rapeseed oil, coconut oil, fractionated oils thereof, Examples include vegetable oils such as hydrogenated oils and transesterified oils. Examples of vitamins include vitamin A, carotene, vitamin B group, vitamin C, vitamin D group, vitamin E, vitamin K group, vitamin P, vitamin Q, niacin, nicotinic acid, pantothenic acid, biotin, inositol, choline. Examples of minerals include calcium, potassium, magnesium, sodium, copper, iron, manganese, zinc, and selenium. Examples of the organic acid include malic acid, citric acid, lactic acid, and tartaric acid. These components can be used in combination of two or more, and synthetic products and / or foods containing a large amount thereof may be used.
Moreover, the normal intake when eating as a food composition containing the bitter gourd extract and / or compound of the present invention is preferably 0.1 mg to 2 g / kg as the bitter gourd extract and / or compound per day for adults, for example. 50 mg to 1 g / kg is more preferable.

The dosage of the blood glucose level-lowering agent containing the bitter gourd extract and / or compound of the present invention and the pharmaceutical composition containing this as an active ingredient varies depending on the administration route, the age, weight, symptoms, etc. of animals to be administered including humans. It can be set as appropriate in consideration of these factors. Although this invention is not limited to this, Preferably 0.1 mg-2 g / kg / day are suitable as an active ingredient. However, when ingested for therapeutic purposes over a long period of time, the amount may be smaller than the above range, and the active ingredient is derived from natural plants and has no safety problems. Even if you use a larger amount than the range, there is no problem.
Moreover, the hypoglycemic agent or pharmaceutical composition containing the bitter gourd extract and / or compound of the present invention can be administered either orally or parenterally (intramuscular, subcutaneous, intravenous, suppository, transdermal, etc.).

  Examples of the administration form of the hypoglycemic agent or pharmaceutical composition according to the present invention include oral administration such as tablets, coated tablets, capsules, granules, powders, solutions, syrups, and emulsions. These various preparations are prepared in accordance with conventional methods, the bittersweet extract and / or compound of the present invention, which is the excipient, binder, disintegrant, lubricant, coloring agent, flavoring agent, solubilizer, suspension, etc. It can be formulated using known adjuvants that can be usually used in the pharmaceutical preparation technical field such as pharmaceuticals and coating agents.

  The amount of the bitter gourd extract and / or compound of the present invention can be arbitrarily determined according to the purpose and application (pharmaceutical composition such as food composition, prophylactic agent, and therapeutic agent). Although this invention is not limited to this, As the content, 0.001 to 100% (w / w) normally with respect to the whole quantity, Especially 0.1 to 100% is preferable.

  EXAMPLES Hereinafter, although the Example of this invention is given and this invention is demonstrated further in detail, this invention is not limited to these specific Examples.

The measuring instruments and methods used in the following examples are as follows. Incidentally, each CDCl _3, C ₅ D ₅ N denotes chloroform, deuterated solvent pyridine.
Optical rotation: DIP1000 polarimeter (manufactured by JASCO, MeOH or CHCl ₃ solvent)
UV spectrum: UV-1650 PC apparatus (manufactured by Shimadzu Corporation, MeOH solvent)
IR spectrum: FT / IR-5300 spectrophotometer (manufactured by JASCO)
¹ H and ¹³ C NMR spectrum: Unity 600 NMR spectrometer (Varian, CDCl ₃ or C ₅ D ₅ N (glycoside) solvent, ¹ H: 600 MHz, ¹³ C: 150 MHz, internal reference substance ( ¹ H) tetramethylsilane) , ( ¹³ C) C ₅ D ₅ N
Mass spectrometry: JMS AX500 spectrometer (EI-MS, FAB-MS, manufactured by JEOL Ltd.), GC / MS (HP5890, SPB-1, Hewlett-Packard Company)
Crystal structure X-ray analysis system: (maXus (Bruker Nonius, DELFT & Mac Science, Japan))
Column chromatography: Sephadex-LH20 (18-111 μM, Amersham Pharmacia Biotech, CHCl ₃ -MeOH, 1: 1), silica gel Kieselgel 60 (Merck, 0.040-0.063 mm), DIAION column ( HP20 lot 2D503), manufactured by Mitsubishi Chemical Corporation)
Preparative HPLC: Cosomosil reverse phase column (10 × 250 mm 5C18-AR, Type Waters, Manuf. No. H14772, manufactured by Nacalai Tesque), JASCO880PU pump, JASCO875UV UV detector, and ERC7512 ERMA CR INC LC20 detector RP18F _254S (Merck)

  As the bittern used in this example, Indian dry bittern (M. charantia L.) was used.

(Liquid bitter gourd extraction)
20 kg of dried bittern was mechanically pulverized and extracted with methanol (MeOH) for 1 month at room temperature. The extract was filtered under reduced pressure and concentrated under reduced pressure. 415.8 g of methanol extract was then suspended in water (H ₂ O) and extracted sequentially with ether (Et ₂ O), ethyl acetate (EtOAc), and n-butanol, and 61 g of ether extract, extracted with ethyl acetate Extracts of 42 g of product and 298 g of n-butanol extract were obtained.

(Purification of bittern ether extract)
27 g of the ether extract obtained by the above operation was subjected to silica gel column chromatography using a hexane / EtOAc gradient (in order of increasing EtOAc concentration) and divided into 24 fractions (fractions 1 to 24). ). Fraction 5 was applied to a Sephadex LH-20 column to obtain three subfractions (fractions 5-1 to 5-3). This fraction 5-3 was subjected to HPLC with ODS RP-18 (Cosomosil reverse phase column) to obtain compounds 10 and 11 (53 mg). The Bun'eeki 9 Sephadex LH-20 to give to compound 9 (3.4 mg) purified by _{(MeOH:: CH 2 Cl 2} 1 1). Fraction 13 was crystallized in EtOAc to give compound 4 (178 mg). Fraction 17 was separated into three subfractions by Sephadex LH-20 chromatography (MeOH: CH ₂ Cl ₂ 1: 1) (fractions 17-1 to 17-3). This fraction solution 17-2 was subjected to silica gel column chromatography (hexane: EtOAc: Et ₂ O = 4: 3: 1) to obtain Compound 5 (175 mg). The fraction 20 was separated into three subfractions with Sephadex LH-20 (fractions 20-1 to 20-3). The fraction 20-2 was further purified by silica gel column chromatography (hexane: EtOAc = 6: 4-0: 10) to give compound 6 (25.2 mg), compound 1 (3.2 mg), compound 5 (76 mg). And compound 7 (21.3 mg) was obtained.

(Purification of extract of ethyl bitter acetate)
3 g of the ethyl acetate extract obtained by the above operation was subjected to silica gel column chromatography (CH ₂ Cl ₂ : MeOH: H ₂ O = 15: 3: 1-lower layer and EtOAc) to obtain 8 fractions. (Fractions 1 to 8). Compound 2 (9.3 mg) was obtained from fraction 1 by a combination of silica gel column chromatography and ODS RP-18 (Cosomosil reverse phase column) (n-hexane: EtOAc = 7: 3 to 0:10, and MeOH). Purification of fraction 3 with Sephadex LH-20 column and ODS RP-18 (Cosomosil reverse phase column) (CH ₃ CN: H ₂ O = 13: 7) gave compound 3 (7.8 mg).

(Purification of nigauri n-butanol extract)
15 g of the n-butanol extract obtained by the above operation was applied to a DIAION column and eluted with increasing amount of MeOH with a mixture of H ₂ O and MeOH, and further eluted with acetone (H ₂ O, 50% MeOH, MeOH and acetone are eluted in this order). Silica gel column chromatography (CH ₂ Cl ₂ : MeOH: H ₂ O = 15: 3: 1-lower layer; CH ₂ Cl ₂ : MeOH = 9: 1 and 4: 1) was applied to 16.2 g of the fraction eluted with MeOH. , ODS RP-18 (Cosomosil reverse phase column) (MeOH: H ₂ O: 9: 1) and silica gel column chromatography (CHCl ₃ : MeOH = 17: 3) in combination to give compound 8 (8.4 mg). Obtained.

  As described above, the size of the ether extract, the ethyl acetate extract, and the n-butanol extract of bitter gourd obtained in Example 1 was 11 by size exclusion, normal phase, reverse phase, ion exchange chromatography, recrystallization technique, and the like. A kind of compound (1-11) was obtained.

(Identification of the obtained compound)
(1) Compound 1
The high-resolution EI-MS result of Compound 1 supports the molecular formula C ₃₁ H ₅₂ O _{3. From} the ¹ H-NMR spectrum, ¹³ C-NMR spectrum, COSY, HMBC, and NOESY experiments, the structure of Compound 1 is It was determined to be 3β, 25-dihydroxy-7β-methoxycucurbita-5, (23E) -diene of the following structural formula 1.

3β, 25-dihydroxy-7β-methoxycucurbita-5, (23E) -diene (1):
Amorphous powder, [α] ²⁰ _D = 73.2 (c = 0.8, CHCl ₃ ), IR (KBr): 3336, 1449, 1068 cm ⁻¹ .
¹ H-NMR (600 MHz, CDCl ₃ ): 0.70, 0.92, 0.98, 1.04, 1.21 (each s and 3H, 30, 18, 19, 29, and 28-CH ₃ ) , 0.88 (d, J = 6.0 Hz, 3H, 21-CH ₃ ), 1.31 (6H, 26 and 27-CH ₃ ), 2.16 (m, 22-H), 2.28 ( dd, 1H, J = 12.5, 4.5 Hz, 10-H), 3.34 (s, 7-OCH ₃ ), 3.43 (dd, 1H, J = 5.4, 1.0 Hz, 7 -H), 3.51 (brs, 3-H), 5.57 (d, J = 15.8 Hz, 23-H), 5.59 (m, 6 and 22-H).
¹³ C-NMR data: shown in the table of FIG.
Positive-type high-resolution EI-MS m / z 472.3915 [ M] +, C 31 H 52 O 3, calculated value 472.3905.

(2) Compound 2
The molecular formula of Compound 2 was shown to be C ₃₂ H ₅₄ O ₃ by high-resolution EI-MS. From ¹ H and ¹³ C-NMR spectra, X-ray crystal structure analysis, etc., Compound 2 was obtained as 3β of the following Structural Formula 2. , 25-dimethoxy-7β-hydroxycucurbita-5, (23E) -diene.

3β, 25-dimethoxy-7β-hydroxycucurbita-5, (23E) -diene (2):
Amorphous powder; [α] ¹⁷ _D = 15.3 (c = 1.5, CHCl ₃ ), IR (KBr): 3453, 2943, 1467, 1032 cm ⁻¹ .
¹ H-NMR (600 MHz, CDCl ₃ ) .: 0.69, 0.92, 1.03, 1.20 (each s and 3H, 30, 18, 19, and 28-CH ₃ ), 1.24 (s , 6H, 26 and 27-CH ₃ ), 0.98 (s, 3H, 29-H), 0.89 (d, J = 6.0 Hz, 3H, 21-CH ₃ ), 2.18 (m, 22-H), 2.27 (dd , 1H, J = 12.5,4.5Hz, 10-H), 3.14 (s, 25-OCH 3), 3.34 (s, 7-OCH 3 ), 3.42 (brd, 1H, J = 5.4 Hz, 7-H), 3.51 (brs, 3-H), 5.37 (d, J = 15.8 Hz, 23-H), 5 .50 (ddd, J = 15.8, 8.7, 5.7 Hz, 24-H), 5.83 (d, J = 5.4 Hz, 6-H).
¹³ C-NMR data: shown in the table of FIG.
Positive high resolution EI-MS m / z 486.4051 [M] ⁺ , C ₃₂ H ₅₄ O ₃ , calculated 486.4073.
Crystal data: C ₃₂ H ₅₃ O ₃ , Mr = 4866.781, orthorhombic, P2 ₁ 2 ₁ 2 ₁ , a = 7.7220 (4) Å, b = 10.0960 (7) Å, c = 38 .762 (4) Å,. = 90.00 °, · = 90.00 °, · = 90.00 °, V = 3021.9 (4) Å ³ , Mo K · radiation, ··· 0.71073 DIP image plate, refined with F ² , full matrix least square refinement, R (gt) = 0.0738, wR (gt) = 0.2088, S (ref) = 1.043, 4333 reflection, 317 parameters, H Refinement of atomic coordinates only, calculation weight sigma, cell refinement: Scalepack (HKL), data reduction: maXus, program used for structure refinement: SHELXL-97.

(3) Compound 3
Positive high-resolution FAB-MS of compound 3 showed a quasimolecular ion peak (C ₃₆ H ₅₈ O ₉ K) based on [M + K] ⁺ at m / z 673.8391. ^{In the 13} C spectrum, a peak indicating monodesmosidic-3β, 25-dihydroxy-7β-methoxycucurbita-5 (23E) -diene, 19-al (5) and a peak indicating an allopyranosyl unit have been previously isolated. It was observed. From the data such as GC / MS, ¹ H-NMR spectrum and optical rotation, the structure of Compound 3 is 3β, 7β, 25-trihydroxycucurbita-5, (23E) -diene-19-al 3β-D-allopyranose Determined (Structural Formula 3).

3β, 7β, 25-trihydroxycucurbita-5, (23E) -diene-19-al, 3β-D-allopyranose (3):
_{^{[Α] 20 D = + 11.32}} (c = 0.9, MeOH), IR (KBr): 3378,2949,1464,1081cm -1.
¹ H-NMR (600 MHz, CD ₃ OD): 0.81, 0.91, 1.08, 1.21 (each s and 3H, 30, 18, 19, and 29-CH ₃ ), 0.94 ( d, J = 6.0Hz, 3H, 21-CH 3), 1.25 (s, 6H, 26 and _{27-CH 3), 1.31 (} 3H, 28-CH 3), 3.27 (dd, J = 7.9, 3.2 Hz, 1H, 7-H), 3.44 (dd, J = 9.4, 2.1, 1H, 4′-H), 3.63 (m, 2H, 6a) 'And 5'-H), 3.79 (d, J = 9.2, 1H, 6b'-H), 3.99 (d, 1H, J = 5.7 Hz, 7-H), 4.02. (D, J = 6.0, 2′-H), 4.60 (d, J = 7.6 Hz, 1′-H), 5.57 (m, 23-H), 5.87 (brd, J = 5.4 Hz, 6-H), 10.23 (s, 1 , 19-H). (300 MHz, C ₅ D ₅ N): 0.74, 0.84, 1.12 (each s and 3H, 30, 18 and 29-CH ₃ ), 0.96 (d, J = 6.2 Hz , 21-CH ₃ ), 1.55 (s, 6H, 26 and 27-CH ₃ ), 1.59 (s, 3H, 28-CH ₃ ), 3.87 (dd, J = 7.9, 3 .2 Hz, 1H, 7-H), 4.18 (dd, J = 9.4, 2.1, 1H, 4′-H), 4.35 and 4.49 (m, 2H, 6a ′ and 5 '-H), 5.34 (d, J = 7.8 Hz, 1'-H), 5.92 (brm, 23 and 24-H), 6.23 (brd, J = 5.4 Hz, 6- H), 10.60 (s, 19-H).
¹³ C-NMR data: shown in the table of FIG.
Positive type high resolution FAB-MS m / z 6733.8391 [M + K] ⁺ , C ₃₆ H ₅₈ O ₉ K, calculated value 6733.8405.

(4) Compound 4
The structure of Compound 4 was identified as 5,19 epoxy-3,25 dihydroxycucurbita-6, (23E) -diene ( ¹³ C-NMR data: shown in the table of FIG. 1) (Structural Formula 4). This compound 4 has already been obtained by hydrolysis of morphodicoside I (Mulholland et. Al. Physochemistry, Vol. 45, p. 391-395), but was first obtained from a bitter gourd extract. .

(5) Compound 5
The structure of Compound 5 was identified as 3β, 7β, 25-trihydroxycucurbita-5, (23E) diene-19-al (Structural Formula 5) by X-ray crystallography.

(6) Compounds 6-11
Compounds 6-9 have 5β, 19-epoxy cucurbita-6, (23E) -diene-3β, 19,25-, respectively, by comparing physical and spectroscopic data with those reported in the literature. It was identified as triol, 5β, 19-epoxy-19-methoxycucurbita-6, (23E) -diene-3β, 25-diol, paramethoxybenzoic acid.
Compounds 10 and 11 (isolated as a 1: 1 mixture) were identified as two aglycones of Charantin, sitosterol and stigmaster dienol.

(Antidiabetic activity test of Compound 4 and Compound 5)
Compounds 4 and 5 were then isolated in large quantities and tested for anti-diabetic activity in vivo.

  Male ddY mice weighing approximately 30 g were purchased from SLC (Shizuoka), and were bred for one week until the start of the experiment at a constant temperature of 21 ± 2 ° C., a humidity of 55 ± 15%, and a light / dark cycle of 12 hours.

Diabetes was induced by intravenous administration of alloxan (Sigma) at a dose of 40 mg / kg. Mice were fasted for 18 hours 5 days after induction of diabetes.
Each test sample was suspended in a 1% gum arabic solution. Glibenclamide (Sigma) was used as a positive control at a dose of 200 mg / kg. The blood glucose level was measured over time at Glucometer DEX (Buyer Healthcare, USA) immediately before sample administration, 2, 4 and 6 hours after sample administration. The control group of mice received 1% gum arabic solution instead of the sample. The test was performed using mice of each sample (Glibenclamide group: 4 mice, Compound 4 group: 3 mice, Compound 5 group: 3 mice).
The blood glucose level at each time point was calculated as a relative blood glucose level (%) with respect to the blood glucose level immediately before sample administration.

The obtained results are shown in FIG. A significant blood glucose lowering effect comparable to that of glibenclamide (200 mg / kg) was observed by intravenous administration of each extract (400 mg / kg).

  INDUSTRIAL APPLICABILITY According to the present invention, an extract having a high blood glucose level lowering action can be used easily by a simpler operation than a natural product bitter gourd. A price-lowering agent can be provided. By providing food / pharmaceutical compositions containing these extracts and compounds, it becomes possible to provide food / pharmaceutical compositions for diabetic patients that can be easily and easily administered to humans. The utility value of is high.

It is a table | surface which shows the < ¹³ > C-NMR spectrum data (150MHz) of the compounds 1-4. The chemical shift (ppm) corresponding to the position number of each compound is represented. The effect of the compound 4 and the compound 5 in an alloxan induced diabetes model mouse is shown. The vertical axis indicates the relative blood glucose level (%) with respect to the blood glucose level immediately before the sample administration, and the horizontal axis indicates the elapsed time after the sample administration. The symbols are: ◆: glibenclamide 200 mg / kg, ■: Compound 4 400 mg / kg, : Represents Compound 5 400 mg / kg.

Claims (19)

  A hypoglycemic agent characterized by containing an organic solvent extract of bitter gourd (scientific name: Mormodica charantia) as an active ingredient.   The blood sugar level-lowering agent according to claim 1, wherein the organic solvent extract is an alcohol extract.   The blood glucose level-lowering agent according to claim 2, wherein the alcohol is a lower alcohol.   The hypoglycemic agent according to claim 2, wherein the alcohol is at least one alcohol selected from the group consisting of methanol, ethanol, propanol and butanol.   The blood sugar level-lowering agent according to any one of claims 1 to 3, wherein the organic solvent extract is a fraction obtained by further ether-extracting the organic solvent extract.   The blood sugar level-lowering agent according to any one of claims 1 to 3, wherein the organic solvent extract is a fraction obtained by further extracting the organic solvent extract with ethyl acetate. A compound which is contained in an organic solvent extract of bitter gourd (scientific name: Mormorica charantia) and has a blood glucose level lowering activity represented by the following general formula 1.
(In General Formula 1, R ₁ represents a hydrogen atom, a hydroxyl group, an alkyl group, an alkoxy group, or an allopyranosiloxy group, R ₂ forms a double bond in X or forms a cyclic ether with R ₅ , R ₃ represents a hydrogen atom, a hydroxyl group, an alkyl group or an alkoxy group,, R ₄ represents a hydrogen atom, a hydroxyl group, an alkyl group, or an alkoxy group, R ₅ is a hydrogen atom, a hydroxyl group, an alkyl group, an alkoxy group or formyl, or represents a group, or a cyclic ether to form together with R _2, X double without including the R ₂ if either form a double bond, including R _2, or R ₂ forms a cyclic ether Represents a hydrocarbon group that forms a bond.)

A compound represented by the following structural formula 1.

A compound represented by the following structural formula 2.

A compound represented by the following structural formula 3.

  A compound according to any one of claims 7 to 9, which is contained in an organic solvent extract of bitter gourd (scientific name: Mormodica charantia) and has a hypoglycemic effect. A compound that is contained in an organic solvent extract of bitter gourd (scientific name: Mormorica charantia) and has a blood glucose level lowering action represented by the following structural formula 4.

A compound that is contained in an organic solvent extract of bitter gourd (scientific name: Mormorica charantia) and has a blood glucose level lowering activity represented by the following structural formula 5.

  A bitter gourd extract obtained by extracting bitter gourd (scientific name: Mormodica charantia) in methanol and further extracting the extract with ether, which has a blood glucose lowering effect.   A bitter gourd extract obtained by extracting bitter gourd (scientific name: Mormorica charantia) in methanol and further extracting the extract with ethyl acetate.   A food / pharmaceutical composition comprising the blood glucose level lowering agent according to any one of claims 1 to 5 as an active ingredient and having a blood glucose level lowering action.   A food / pharmaceutical composition containing the compound according to any one of claims 6 to 12 as an active ingredient and having a blood glucose lowering effect.   A food / pharmaceutical composition containing the bittern extract of claim 13 as an active ingredient and having a blood glucose lowering effect. A food / pharmaceutical composition comprising the bitter melon extract according to claim 14 as an active ingredient and having a blood glucose lowering effect.

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