JPH0646944B2 - Amylase inhibitor - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION (Field of Industrial Application) The present invention relates to a novel amylase inhibitor, and more particularly to an amylase inhibitor mainly composed of a solvent extract substance of bay leaves.

(Problems to be Solved by Conventional Techniques and Inventions) In recent years, various adult diseases have been increasing in developed countries due to causes such as overnutrition. Examples of such adult diseases include diabetes, obesity and arteriosclerosis.

Many of these adult diseases are caused by an increase in blood sugar due to excessive intake of starch and the like.

In general, the treatment of diabetes and obesity, which are such diseases, mainly involves dietary therapy, but in addition to causing mental distress to the patient, the burden of calorie calculation and the like is large. Therefore, as a simple method, when eating starchy foods such as rice and udon, which are the center of Japanese dietary habits, a method of taking an amylase inhibitor to inhibit the digestion of starch is considered. In other words, by inhibiting the action of amylase that digests starch, it is possible to suppress the digestive absorption of sugars, suppress the rise in blood sugar, and prevent or treat metabolic diseases such as diabetes, obesity, and arteriosclerosis. Based on the above, several kinds of amylase inhibitors have been discovered in recent years.

However, although these are oligosaccharide-based and peptide-based ones obtained from microorganisms or cereals, they have problems in terms of safety and efficacy, and most of them are still in the stage of practical application. I haven't arrived.

<Means for Solving the Problems> Therefore, the inventors of the present invention have diligently studied to find a novel amylase inhibitor having no problems in safety and efficacy for crude drugs that have been used for many years. As a result, they discovered that bay leaves contained an amylase inhibitor, and completed the present invention.

That is, the present invention firstly comprises an amylase inhibitor containing a hydrophilic solvent extract substance of bay leaves as an active ingredient.

Secondly, the present invention comprises an amylase inhibitor containing, as an active ingredient, a non-hydrophilic solvent-soluble portion obtained when a hydrophilic solvent extract substance of bay leaves is distributed with a non-hydrophilic solvent and water.

Thirdly, the present invention comprises an amylase inhibitor containing a non-hydrophilic solvent extract of bay leaves as an active ingredient.

Fourthly, the present invention comprises an amylase inhibitor containing, as an active ingredient, a substance obtained by removing a water-soluble portion from a substance extracted from a non-hydrophilic solvent of bay leaves.

i) Raw material The laurel leaf used in the present invention is an evergreen tree, Laurus nobilis, which is cultivated all over the world including the Mediterranean coast.
It is a leaf of L. <Lauraceae>.

Dried laurel leaves are called laurel and bay leaf, and are often used in meat dishes, soups, stews, and as a raw material for sauces.

For medicinal purposes, leaves are decoction and used for rheumatism and neuralgia. The leaves contain 1 to 3% of essential oil, the main component of which is cineol,
Eugenol and geraniol.

ii) Extraction The substance contained as an active ingredient in the amylase inhibitor of the present invention is characterized by α-amylase inhibitory activity. Therefore, extraction with water or an organic solvent, centrifugation or filtration is used as an indicator of these inhibitory activities. It can be isolated and purified using a suitable purification means of. These methods are
They can be applied alone or in combination in any order, or can be applied repeatedly as needed. Hereinafter, an example of a method for extracting a substance for extracting a hydrophilic solvent of bay leaves and a procedure for obtaining a non-hydrophilic solvent-soluble portion when the substance is distributed with a non-hydrophilic solvent and water will be described.

A) Degreasing the raw material by using a degreasing solvent such as hexane or ether at room temperature or by heating.

The amount of degreasing solvent used in this case, for bay leaves,
It is appropriate that the amount is 1 to 10 times, preferably 2 to 5 times. The degreasing time is 10 minutes to 72 hours, preferably 1 to 24 hours. The heating temperature is
Suitably up to 70 ° C, preferably up to 40 ° C. This extraction operation is preferably performed 2 to 5 times.

B) Air-drying or vacuum-drying the degreased raw material to remove the degreasing solvent.

C) The raw material obtained in b) is subjected to an extraction treatment by adding a hydrophilic organic solvent such as methanol or ethanol to the extraction raw material according to a conventional method. The amount of the hydrophilic organic solvent is appropriately 1 to 10 times, preferably 2 to 5 times the amount of the extraction raw material. Further, this extraction is usually carried out under boiling, but the active ingredient can be obtained even if it is carried out in a low temperature room at 4 ° C. The extraction time is appropriately 1 to 24 hours, preferably 2 to 10 hours. This extraction operation is preferably performed 2 to 5 times.

D) After the extract obtained in c) is concentrated to dryness, a non-hydrophilic organic solvent such as ethyl acetate or chloroform is added to give a suspension. The non-hydrophilic organic solvent is used in an amount of 1 to 20 times, preferably 3 to 10 times, the amount of the obtained solid matter.

A non-hydrophilic organic solvent is further added to the insoluble matter, and the mixture is stirred well, filtered, and combined with the previous filtrate. The stirring time is appropriately 10 minutes to 3 hours, preferably 30 minutes to 2 hours.

E) To this solution, add an equal amount of water and remove the water-soluble portion.

F) A brown powder is obtained as an active ingredient by concentrating and drying the non-hydrophilic organic solvent-soluble portion.

iii) Assay for α-amylase inhibitory activity A method for testing α-amylase inhibitory activity will be described.

As the substrate solution, a solution of soluble starch (Katayama Chemical Co., Ltd.) dissolved in 0.5% with 50 mM acetate buffer (pH 6.5) was used, and human saliva-derived α-amylase was used as the enzyme solution. did.

Enzyme solution 50μ, assay solution 50 to 0.25ml substrate solution
μ, 50 mM acetate buffer (pH 6.5) 50 μ and 50 containing 20 mM calcium chloride and 100 mM sodium chloride
Add 100 μm mM Acetate buffer (pH 6.5), and add 3 in a warm bath.
The reaction was carried out at 7 ° C for 15 minutes. After completion of the reaction, 5 ml of a 0.0017N hydrochloric acid aqueous solution containing 1.7 mM potassium iodide and 0.17 mM iodine was added, and the absorbance was measured at 700 nm.

The amylase inhibitor of the present invention comprises, as an active ingredient, a hydrophilic solvent extract of bay leaves; a non-hydrophilic solvent-soluble part obtained when the hydrophilic solvent extract is distributed with a non-hydrophilic solvent and water; bay leaves Or a substance obtained by removing the water-soluble part from the non-hydrophilic solvent-extracted substance. These active ingredients can be formulated into any form to be an amylase inhibitor which can be orally or parenterally administered.

The administration method, dose and formulation method are shown below.

i) Administration method The amylase inhibitor of the present invention can be used either orally or parenterally, and when orally administered, it is administered as soft or hard capsules or tablets, granules, fine granules or powders. When administered parenterally, it may be administered in a dosage form such as a suppository so that sustained mucosal absorption can be maintained as an injection, a drip, and a solid or suspension viscous liquid, but administration in a local tissue, External administration methods such as intradermal, subcutaneous, intramuscular, intravenous injection, topical application, spray, suppository, and intravesical injection can also be used.

ii) Dosage The dose is not constant depending on the administration method, the malignancy of the disease, the age of the patient, the condition and general condition of the patient, the degree of progression of the disease, etc. 5-
5,000 mg, usually 0.5-3,000 mg for dwarfs.

iii) Method of formulation The proportion of the active ingredient of the amylase inhibitor of the present invention can be changed depending on the dosage form, but when administered orally or by mucosal absorption, it is generally 0.3 to 15.0% by weight. When administered parenterally, approximately 0.01 to 10% by weight is suitable.

Further, in formulating the amylase inhibitor of the present invention,
According to a conventional method, an aqueous solution, an oily preparation and the like can be prepared into a preparation for subcutaneous or intravenous injection, or in the form of capsules, tablets, fine granules and the like for oral use.

Further, in order to impart stability and acid resistance to the active ingredient for long-term storage and to completely maintain the medicinal effect, if a formulation with a pharmaceutically acceptable film is further formulated, excellent stability is obtained. It can be an amylase inhibitor.

The surfactants, excipients, lubricants, adjuvants and pharmaceutically acceptable film-forming substances used for formulating the amylase inhibitor of the present invention are as follows.

In order to ensure good disintegration and dissolution of the inhibitor of the present invention,
Surfactants such as alcohols, esters, polyethylene glycol derivatives, fatty acid esters of sorbitan, and sulfated fatty alcohols can be added alone or in combination of two or more.

Examples of excipients include sucrose, lactose, starch,
Crystalline cellulose, mannite, light anhydrous silicic acid, magnesium aluminate, magnesium metasilicate aluminate, synthetic aluminum silicate, calcium carbonate, sodium hydrogencarbonate, calcium hydrogenphosphate, carboxymethylcellulose calcium, or a combination of two or more thereof. It can be added.

As the lubricant, for example, magnesium stearate, talc, hydrogenated oil, etc. may be added alone or in combination of two or more,
Further, as a corrigent and a flavoring agent, salt, saccharin, sugar,
Mannitol, orange oil, licorice extract, citric acid,
Glucose, menthol, eucalyptus oil, malic acid and other sweeteners, flavors, colorants, preservatives and the like may be contained.

Examples of adjuvants such as suspending agents and wetting agents include coconut oil, olive oil, sesame oil, peanut oil, calcium lactate,
Safflower oil, soybean phospholipid, etc. can be contained.

As film-forming substances, cellulose acetate phthalate (CAP) is used as a carbohydrate derivative such as cellulose and saccharides, and methyl acrylate / methacrylic acid is used as a polyvinyl derivative such as acrylic acid copolymers and dibasic acid monoesters. Examples thereof include copolymers and methyl methacrylate / methacrylic acid copolymers.

Further, when coating the above film-forming substance, the properties of the film-forming agent can be obtained by adding various additives for preventing mutual adhesion of chemicals during coating operation, in addition to coating aids that are usually used, such as plasticizers. Can be improved or the coating operation can be made easier.

Hereinafter, the present invention will be described in more detail with reference to examples.

Example 1 a) Dried bay leaves [Sold by: Uchida Wakan Yaku Co., Ltd.] 5
After immersing 00 g in 1500 ml of hexane and allowing it to stand at room temperature for 24 hours, hexane was removed by filtration. This operation was repeated 3 times to degrease. The yield of the hexane-soluble portion was 26.7 g.

(B) The defatted laurel leaves were air dried for 1 day.

C) Air-dried bay leaves were immersed in 1000 ml of methanol and extracted under boiling for 3 hours. This operation was repeated 3 times to collect the extract.

D) The obtained extract was concentrated under reduced pressure at 40 ° C. in a water bath with an evaporator and dried under vacuum to obtain a methanol extract substance (extract substance 1, yield 99.5 g).

500 ml of ethyl acetate was added thereto, and the mixture was stirred and filtered.
The insoluble matter was further added with 200 ml of ethyl acetate, stirred and filtered, and the filtrate was collected.

E) 500 ml of water was added to this filtrate for extraction. This operation was repeated 3 times to remove the water-soluble part (yield 19.1 g).

F) The ethyl acetate-soluble portion is water bath 40 ° C by an evaporator.
At reduced pressure under reduced pressure, and dried in vacuo to give a brown powder (extracted substance 2, yield 17.8g).

The α-amylase inhibitory activity at each stage of purification of the bay leaf extract was detected. The results are shown in Table 1 below.

Example 2 a) The air-dried bay leaves obtained in b) of Example 1 were immersed in 1000 ml of ethyl acetate and extracted under boiling for 3 hours. This operation was repeated 3 times to collect the extract.

(B) The obtained extract was concentrated and concentrated in an evaporator at a water bath of 40 ° C., and dried under vacuum to obtain an ethyl acetate extract substance (extract substance 3, yield 31.5 g).

500 ml of water was added thereto, and the mixture was stirred and filtered. 200 ml of water was further added to the insoluble matter, and the filtrate was collected. The filtrate was freeze-dried to obtain a water-soluble portion as a powder (yield 5.3 g).

(C) The insoluble matter in (b) was dried under vacuum to obtain a substance in which the water-soluble portion was removed from the extracted substance 3. (Extracted substance 4, yield 24.
3g).

The α-amylase inhibitory activity at each stage of purification of the bay leaf extract was detected. The results are shown in Table 2 below.

Comparative Example 1 The α-amylase inhibitory activity of a commercially available α-amylase inhibitor (manufactured by Sigma, derived from wheat germ) was assayed. The 50% inhibitory concentration (IC ₅₀ ) was 229.5 μg / ml.

The production examples of the amylase inhibitor of the present invention are shown below.

Extracted substances 1 to 4 obtained from Examples 1 and 2 above were used as active ingredients.

Formulation Example 1 (Injection / Drip) 5 g of powdered glucose was added to each vial aseptically so as to contain 500 mg of each of the above-mentioned extracted substances, and the vials were sealed and sealed with an inert gas such as nitrogen or helium. Store in a cool dark place. Before use, 0.85% saline 500m
10 to 500 as an intravenous injection by adding 1
ml is administered by intravenous injection or infusion depending on the symptoms.

Production Example 2 (Injection / Drip) An intravenous injection for mild disease was prepared in the same manner as in Preparation Example 1 except that 50 mg of each of the above extract substances was used, and 10 to 50 days a day.
0 ml is administered by intravenous injection or infusion depending on the symptoms.

Formulation Example 3 (Injection, Capsule) 30 mg of each of the above extract substances is dissolved in 1 g of refined sesame oil and 100 mg of aluminum stearate gel and sealed, and then sealed with an inert gas such as nitrogen and helium and stored in a cool dark place, subcutaneously. It should be a preparation for injection. Once a day depending on the symptoms,
Administer 1-10 ml by subcutaneous injection.

Further, 0.5 ml each of the above-mentioned preparation is dispensed into capsules to give oral capsules, and 1 to 10 capsules are orally administered per day depending on the symptoms.

Formulation Example 4 (Enteric-coated tablet) 1,000 enteric-coated tablets for adults (a) and dwarf (b) were prepared with the following composition.

Take each of the components of [A], mix well, press this directly or knead well, granulate through a screen of an extrusion type granulator, add sufficiently well dried one. Pressed to produce tablets.

Next, the base material of [B] well dissolved in the molded tablet is coated to form an enteric coated tablet.

About this tablet Japanese Pharmacopoeia (hereinafter referred to as "JP") disintegration test method, enteric coated artificial gastric juice (pH 1.2)
When the test was conducted, it did not disintegrate even after shaking for 1 hour, and disintegrated in 5 to 6 minutes in the artificial intestinal fluid (pH 7.5) test.

Formulation Example 5 (Enteric coated granules) 1,000 g of enteric coated granules were produced with the following components.

The components of [A] were taken, mixed well, and then formed into granules by a conventional method, dried well, and sieved to produce granules which were passed through pins, heat-sealed packaging and the like. next,
The granules for enteric coating are prepared by coating the base material [B], which is obtained by dissolving the granules in a floating flow. This granule had a pH of 1.2 when subjected to a disintegration test using a disintegration tester of Japanese Pharmacopoeia.
It does not disintegrate even when shaken in artificial gastric juice for 1 hour. The artificial gastric juice with pH 7.5 disintegrated in 5 minutes.

Formulation Example 6 (Enteric Coated Capsules) 1,000 enteric coated capsules were manufactured with the following ingredients.

An enteric-coated granule suitable for capsules was produced by the same method as described in Formulation Example 5 using the above components, and the capsule was filled into an enteric-coated capsule.

When this capsule was subjected to a disintegration test using a disintegration tester of Japanese Pharmacopoeia, no disintegration or elution was observed even if shaken for 1 hour in an artificial gastric juice of pH 1.2, and in 5 minutes in an artificial gastric juice of pH 7.5. Disintegration or total elution.

(Effects of the Invention) According to the present invention, an amylase inhibitor containing a solvent extract of bay leaves as a main component is provided. The amylase inhibitor obtained by the present invention is more effective than the conventional amylase inhibitors and is highly safe.

Claims (6)

[Claims] 1. An amylase inhibitor containing a substance extracted from a hydrophilic solvent of bay leaves as an active ingredient. 2. An amylase inhibitor containing, as an active ingredient, a non-hydrophilic solvent-soluble part obtained by partitioning a substance extracted from a hydrophilic solvent of bay leaves with a non-hydrophilic solvent and water. 3. The amylase inhibitor according to claim 1 or 2, wherein the hydrophilic solvent is methanol or ethanol. 4. An amylase inhibitor containing a non-hydrophilic solvent extract of bay leaves as an active ingredient. 5. An amylase inhibitor containing a substance obtained by removing a water-soluble part from a substance extracted from a non-hydrophilic solvent of bay leaves as an active ingredient. 6. The amylase inhibitor according to claim 2, 4 or 5, wherein the non-hydrophilic solvent is ethyl acetate.