JPH09286737A - Glycerophosphate dehydrogenase inhibitor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To obtain a glycerophosphate dehydrogenase inhibitor containing an extract of white birch, having high safety and useful for the prevention and treatment of obesity. SOLUTION: White birch belonging to the genus Betula, family Betulaceae is extracted with water and/or an organic solvent and the extract is used as an active component. The extraction is carried out e.g. by extracting finely cut white birch chips with water, ethanol, etc., at refluxing temperature for 0.5-8hr or immersing the chips in water, ethanol, etc., at room temperature to 50 deg.C for 1-10 days. The inhibitor contains the active component in an amount of 0.3-15wt.% for oral administration and 0.01-10wt.% for parenteral administration. It is administered at a daily rate of 0.5-5,000mg for adult and 0.5-3,000mg for child in terms of the active component. The inhibitor may be compounded to a food in an amount of 0.001-15wt.%, preferably 0.01-10wt.% and used as a food composed mainly of cereal flour or starch such as bread, noodle, biscuit, hot cake or tablet candy, a dressing, a drink, etc.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a glycerophosphate dehydrogenase inhibitor, and more particularly to a glyceroline dehydrogenase capable of inhibiting the activity of glycerophosphate dehydrogenase, which is increased in the differentiation from preadipocytes to adipocytes. It relates to an acid dehydrogenase inhibitor. Further, the present invention relates to a food additive containing the glycerophosphate dehydrogenase inhibitor and a food containing the same.

[0002]

2. Description of the Related Art In recent years, westernization of eating habits has progressed in Japan,
Carbohydrate intake is decreasing and fat and protein intake is increasing. As a result, the problem of obesity has been taken up. Obesity causes adult illnesses such as high blood pressure, heart disease and diabetes, so it is necessary to work hard to maintain health. In considering the cause of obesity, the mechanism of differentiation and dedifferentiation into fat cells is important. In 1974, GREEN et al. Succeeded in establishing a cell line by cloning a mouse embryo-derived 3T3 fibroblast that differentiates into adipocytes, and produced a course of differentiation from preadipocytes into adipocytes. It became possible to carry out chemical research (GREEN, H. et al., CELL, VOL.1, pages 113-116 (1974); CELL, VOL.3, pages 127-133 (1974)). Ne'grel et al. Also differentiated from testicular adipose tissue of hereditary obese mice (C57BL / 6J ob / ob) into adipocytes (ob 17).
(Ne'grel, R. et al., PROC. NATL. ACA
D. SCI. USA, VOL. 75, 6054-6058 (1978). Biochemical studies have been carried out using these cells, and it has been revealed that almost no activity is detected in progenitor cells and that the enzyme increases by differentiation into adipocytes. For example, glycerophosphate dehydrogenase, lipoprotein lipase, diacylglycerol acyltransferase, etc. have been reported.
It is used as a marker enzyme for differentiation because the enzyme activity increases 000 times. Therefore, it is conceivable to inhibit differentiation into adipocytes by inhibiting glycerophosphate dehydrogenase. It is also conceivable to treat or prevent obesity with such an enzyme inhibitor. further,
A method in which an inhibitor for the enzyme is mixed in the food in advance is considered. As an inhibitor against glycerophosphate dehydrogenase, 5-alkylresorcinol (JP-A-6-
100440) is known, and the effect of preventing fat accumulation in animal cells has been clarified.

[0003]

An object of the present invention is to exhibit excellent inhibitory activity on glycerophosphate dehydrogenase,
Another object of the present invention is to provide a highly safe glycerophosphate dehydrogenase inhibitor. Another object of the present invention is to provide a food additive containing the glycerophosphate dehydrogenase inhibitor, and further to provide a food containing the glycerophosphate dehydrogenase inhibitor or the food additive.

[0004]

[Means for Solving the Problems] The inventors of the present invention eagerly searched for a substance having an inhibitory activity against glycerophosphate dehydrogenase, and found that an extract of birch water and / or an organic solvent showed a strong inhibitory activity. Found to show. Further, when the extract was divided into a water-soluble portion and a water-insoluble portion, it was found that both of them exhibited the same strong inhibitory effect, and the present invention was completed. Therefore, the present invention is a glycerophosphate dehydrogenase inhibitor containing an extract of birch water and / or an organic solvent as an active ingredient. The present invention also relates to a glycerophosphate dehydrogenase inhibitor containing, as an active ingredient, a water-soluble part of an extract of birch water and / or an organic solvent. The present invention further relates to a food additive containing the glycerophosphate dehydrogenase inhibitor, and a food containing the glycerophosphate dehydrogenase inhibitor or the food additive.

[0005]

BEST MODE FOR CARRYING OUT THE INVENTION Birch is a birch (Betulacea).
e), a plant belonging to the genus Betula, which is distributed all over the world. The birch used in the present invention is not particularly limited, and any birch can be used. Specific examples of white birch include B. ermanii Chamiss
o var. communis Koidz, B. platyphyl
la Sukatchev var. japonica Hara, B.
davurica Pall. (Yaegawa birch), B. schmidtii Re
gel (onore), B. globispica Shirai, B. costata Trautr.,
Betula alba L., Betulalenta L., Betula pendula R
oth. etc. As you can see, there are various trees called birch, and B. platyphylla Sukatchev var. Japo
nica Hara, which is the Japanese name for birch, is called birch, birch, birch, birch, birch. The traditional use of this birch is neuralgia for decoction of bark,
Take spills orally. Leaves are also decocted to make diuretics. Tar obtained by carbonizing the wood can be used externally for rheumatism and skin diseases. The main components of birch leaves, bark, xylem and their mixtures are betulin, betuloetic acid (betulo).
Retinic acid) mono-butyl ester, tannin, saponin, essential oil, nicotinic acid amide, guaiacol, creosote, cresol, and carboxylic acid are known. In the present invention, plants belonging to the family Birchaceae and genus Birch can be used, and are collectively referred to as "birch" in the present specification. The parts used for extraction include leaves, bark, roots or xylem,
Alternatively, a mixture thereof may be mentioned. Birch, especially birch Betula alba L. or Betula lenta L. (Betulace
The leaves, bark, and xylem of ae) are generally available on the market, and the active ingredient of the present invention may be one obtained by extracting such a commercially available product as an extraction raw material. It was not known until now that the extract of white birch inhibits glycerophosphate dehydrogenase.

The extraction solvent for obtaining the extract from white birch may be water, an organic solvent or a mixture thereof. Examples of the organic solvent include alcohols such as methanol, ethanol, propanol, propylene glycol and 1,3-butylene glycol, halogenated solvents such as chloroform, esters such as ethyl acetate and hexane. Preferred solvents include water, ethanol, and 70-90% hydrous ethanol. If extraction is performed using ethanol as a solvent, the water-soluble portion and the water-insoluble portion can be extracted at the same time. The water-soluble portion can be further extracted from such an extract to be used as an active ingredient. The extracted raw material can be dried and finely chopped. The amount of the solvent with respect to the extraction raw material is not particularly limited, and for example, about 5 times the amount of the solvent based on the weight of the extraction raw material is used. Extraction is heating under reflux extraction, dipping,
It can be carried out by any method such as stirring extraction,
Moreover, it may be carried out according to a conventional method. If it is a heating reflux extraction,
The temperature of the solvent is appropriately from room temperature to 40 to 50 ° C. for 1 to 10 days when immersed for about 0.5 to 8 hours at the reflux temperature. It can also be repeatedly extracted. The obtained extract can be concentrated under reduced pressure to obtain an extract as a dry solid. The extract may be used as it is as an active ingredient. Further, the water-soluble portion can be obtained by suspending the above dried product in water and filtering. A component that is soluble in ethanol and water is effective, and this fraction contains 280 nm
It is considered that white birch tannin is also an active ingredient, since UV absorption is observed in. The extract of white birch with water and / or organic solvent is generally available on the market, for example, an extract extracted with 90% ethanol is known. Birch extract is commercially available as a raw material for cosmetics. In the present invention, such a commercially available product can be used as an active ingredient.

[0007]

[Reference Example 1] Extraction of active ingredient from white birch 1 kg of birch (Betula alba L.) bark (Birch) was shredded, about 5 liters of 90% ethanol was added, and the mixture was immersed at room temperature for 7 days and filtered to extract Got The residue was similarly extracted with about 5 liters of 90% ethanol. All extracts were combined and concentrated under reduced pressure to obtain a concentrated dry solid (39 g).

Reference Example 2 20 g of the ethanol extract obtained in Reference Example 1 can be separated into a soluble portion and an insoluble portion by suspending in water and filtering. From the water-soluble part, freeze drying was performed to obtain a residue (14 g). From the insoluble portion, betulin could be obtained by an appropriate purification means such as column chromatography. It was identified by comparison with commercially available betulin.

[0008]

[Test Example] The glycerophosphate dehydrogenase inhibitory activity in the present invention was measured as follows according to the method of WISE etc. (WISE. LS et al., J. BIOL. CHEM., VOL. 254,
273-275 (1979)) 1.97 ml of 100 mM triethanol / hydrochloric acid buffer (pH 7.5) containing 2.5 mM EDTA in a 1 cm square cell, 6 mM dihydroxyacetone phosphate 0.3 ml, 1 mM β -Mercaptoethanol 0.3 ml, 1.2 mM NADH 0.3 m
Add l and 0.03 ml of the sample solution of each concentration and mix well.
Next, 0.15 ml of a 1 unit / ml glycerophosphate dehydrogenase (Sigma) solution was added, and after mixing, the absorbance at 340 nm was measured, and the inhibitory activity was determined from the absorbance after 10 minutes by the following formula. As a control, the sample solution was not added to the square cell in the same manner, and the absorbance at 340 nm was measured. Inhibition rate (%) = (A / B) x 100 A: Absorbance of various samples after 10 minutes-Absorbance after 10 minutes of control B: Absorbance after 0 minutes of control-Absorbance after 10 minutes of control The results , Showed a strong inhibitory activity as shown in the table below.

[0009]

[Table 1] ─────────────────────────────────── Sample name IC ₅₀ (μg / ml) ── ────────────────────────────────── White birch ethanol extract of Reference Example 0.49 White birch ethanol extract of Reference Example 2 Water-soluble part of 0.95 ───────────────────────────────────

The glycerophosphate dehydrogenase inhibitor of the present invention may contain additives in addition to the active ingredient. In addition, these active ingredients can be formulated into an arbitrary form with a suitable auxiliary agent to give a glycerophosphate dehydrogenase inhibitor which can be orally or parenterally administered. Hereinafter, the administration method, dosage, and formulation method of the glycerophosphate dehydrogenase inhibitor of the present invention will be described. The glycerophosphate dehydrogenase inhibitor of the present invention can be used for both oral and parenteral administration.When orally administered, it is administered as soft or hard capsules or tablets, granules, fine granules, and powders. You. When administered parenterally, it can be administered in the form of suppositories such as injections, drops, and solid or suspended viscous liquids so that continuous mucosal absorption can be maintained. External administration methods such as internal, subcutaneous, intramuscular and intravenous injections, topical application, spraying, suppositories, and intravesical injections can also be used.

The dose may vary depending on the administration method and the age, medical condition, general condition of the patient, etc.
The appropriate daily active ingredient is 0.5 to 5,000 mg, and the suitable amount for a child is 0.5 to 3,000 mg. The ratio of the active ingredient of the glycerophosphate dehydrogenase inhibitor of the present invention,
While it may vary depending on the dosage form, it is usually about 0.3 to 15.0% by weight when orally administered, and about 0.01 to 10% by weight when parenterally administered. In addition, in formulating the glycerophosphate dehydrogenase inhibitor of the present invention, it can be made into an aqueous solution, an oily formulation, or the like, and can be made into a formulation for subcutaneous or intravenous injection, or a formulation to be applied topically to the skin, etc. Or formulated into a dosage form such as capsules, tablets, fine granules and the like, and provided for oral use.

[0012] Further, in order to impart stability and acid resistance to long-term storage to the active ingredient and to completely maintain the medicinal effect, it is excellent if a pharmaceutically acceptable film is further formed into a formulation. It can be a glycerophosphate dehydrogenase inhibitor having stability. The surfactants, excipients, lubricants, adjuvants, pharmaceutically acceptable film-forming substances, and the like used in formulating the glycerophosphate dehydrogenase inhibitor of the present invention are as follows. . To improve the disintegration and elution of the glycerophosphate dehydrogenase inhibitor of the present invention,
Surfactants, for example, one or more of alcohols, esters, polyethylene glycol derivatives, fatty acid esters of sorbitan, sulfated fatty alcohols and the like can be added. In addition, as an excipient, for example, sucrose, lactose, starch, crystalline cellulose, mannitol, light anhydrous silicic acid, magnesium aluminate, magnesium metasilicate aluminate, synthetic aluminum silicate, calcium carbonate, sodium hydrogen carbonate, calcium hydrogen phosphate , Carboxymethylcellulose calcium and the like can be added alone or in combination of two or more.

As the lubricant, for example, magnesium stearate, talc, hydrogenated oil, etc. may be added alone or in combination of two or more, and as a corrigent and a flavoring agent, salt,
Saccharin, sugar, mannitol, orange oil, licorice extract, citric acid, glucose, menthol, eucalyptus oil,
Sweetening agents such as malic acid, flavors, coloring agents, preservatives and the like may be contained. Suspending agents, as adjuvants such as wetting agents, for example, coconut oil, olive oil, sesame oil, peanut oil,
It can contain calcium lactate, safflower oil, soybean phospholipids, and the like. As the film-forming substance, cellulose acetate phthalate (CAP) is used as a carbohydrate derivative such as cellulose or saccharide, and methyl acrylate / methacrylic acid is used as a polyvinyl derivative such as an acrylic acid copolymer or a dibasic acid monoester. Examples thereof include copolymers and methyl methacrylate / methacrylic acid copolymers. Also,
When coating the above film-forming substance, the properties of the film-forming agent are improved by adding a coating agent which is usually used, for example, a plasticizer, and various additives for preventing mutual adhesion of drugs during coating operation. Or, the coating operation can be made easier.

The glycerophosphate dehydrogenase inhibitor of the present invention can also be incorporated into foods and health foods, and can also be used as a component of food additives. When compounded in food, 0.001 to 15% by weight is suitable as an active ingredient to the food, and 0.01 to 10% by weight is particularly preferred. The type of food to which the glycerophosphate dehydrogenase inhibitor or food additive of the present invention is blended may be any type, for example, bread, noodles, biscuits, hot cakes, confectionery and other flour and starch. Foods, dressings, drinks and the like.
As a method of blending it into food, it may be appropriately blended in the manufacturing process according to various foods.

[0015]

The present invention will be described in more detail with reference to the following examples. Example 1 (tablet and tablet) 108 g of egg shell calcium, 2 g of ferric pyrophosphate, 40 g of ascorbic acid, 40 g of microcrystalline cellulose, 285 g of reduced maltose, and 25 g of ethanol extract of white birch were blended by a conventional method and then tableted. Then, tablet confectionery and tablets were produced. Example 2 (biscuits) 120 g of wheat flour, 1.2 g of water-soluble part of birch ethanol extract,
Betulin 1.2g, sugar 35g, shortening 15g, whole egg powder
Using 1.5 g, salt 1 g, sodium hydrogen carbonate 0.6 g, ammonia carbonate 0.75 g, and water 20 g, a dough is prepared by a conventional method,
It was molded and roasted to produce a biscuit.

Example 3 (Bread) Wheat flour 3 kg, birch water extract 3 g, yeast 60 g, yeast food 3 g, sugar 150 g, salt 60 g, shortening 150
g, 60 g of skim milk powder, and 2070 g of water were used to prepare a dough by a conventional method, and the dough was molded and baked to produce bread. Example 4 (Noodle) A mixture of semi-strong wheat flour, to which 1% by weight of a water-soluble portion of a birch ethanol extract, 34% by weight of water, 1% by weight of salt and 1% by weight of starch were added, After kneading for 12 minutes, the mixture was rolled and formed several times with a noodle machine to obtain raw noodle bands and raw noodle strings of Chinese noodles.

[0017]

Industrial Applicability The glycerophosphate dehydrogenase inhibitor of the present invention has high safety and high glycerophosphate dehydrogenase inhibitory activity. It can be easily added to foods and is useful for preventing or treating obesity.

─────────────────────────────────────────────────── ─── Continuation of the front page (51) Int.Cl. ⁶ Identification code Internal reference number FI Technical display location A23L 1/30 A23L 1/30 B C12N 9/99 C12N 9/99

Claims (4)

[Claims] 1. A glycerophosphate dehydrogenase inhibitor containing an extract of white birch as water and / or an organic solvent as an active ingredient. 2. A glycerophosphate dehydrogenase inhibitor containing, as an active ingredient, a water-soluble portion of an extract of birch water and / or an organic solvent. 3. A food additive containing the glycerophosphate dehydrogenase inhibitor according to claim 1 or 2. 4. A food containing the glycerophosphate dehydrogenase inhibitor according to claim 1 or 2 or the food additive according to claim 3.