JP2000319192A - Enzyme inhibitor - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a safe and useful inhibitor against an enzyme. SOLUTION: After obtaining syrup having both tyrosinase-inhibiting activity and α-amylase-inhibiting activity by extraction-treating the rhizome of a mushroom (especially the rhizome of Grifola frondosa, i.e., Grifola frondosa mat) with an alcohol, the extracted syrup is heat-treated and is added to cosmetics and food to obtain a bleaching cosmetic, a moisturizing cosmetic, a bleaching bath salt, a moisturizing bath salt and/or diet food.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an enzyme inhibitor obtained by extracting a rhizome of mushrooms, especially a rhizome of Maitake (usually a non-edible part, called Maitake mat) with an organic solvent. Specifically, a mushroom rhizome using tyrosinase inhibitor and α-amylase inhibitor activity,
The present invention relates to an enzyme inhibitor derived from Maitake mat.

[0002]

BACKGROUND OF THE INVENTION Melanin, a pigment that causes sunburn, is a brown or black pigment that is formed through several intermediates when L-tyrosine is oxidized by tyrosinase in melanocytes. It is. From the medical point of view such as appearance of skin cancer as well as cosmetic point of view, suppression of melanin formation is considered preferable, and various sunscreen agents have been provided. However, there is still a need for the development of safe and effective sunscreen active ingredients. Obesity is also highly related to lifestyle-related diseases (adult diseases) such as diabetes,
The need to prevent or eliminate it has been pointed out. Current,
Various diet methods, including diet foods, have been proposed, but here again, development of active ingredients for safe and effective diet foods is required.

[0003] In general, plant enzyme inhibitors play a role as a source of protective substances, protecting themselves from pathogenic microorganisms and pests,
It has the function of regulating the activity of enzymes and protecting substances contained in the tissue, and it is said that where the enzyme is present, its inhibitor is present on the other hand.

[0004] One means of preventing the above-mentioned melanin formation and obesity can be achieved by using a substance that inhibits the activity of an enzyme involved in an in vivo reaction leading to them. That is, melanin is produced by an enzymatic reaction involving the oxidation of L-tyrosine by tyrosinase, whereas obesity originates, for example, from the digestion of starch by α-amylase. Therefore, it is considered that the above object can be achieved by using a tyrosinase inhibitor and an α-amylase inhibitor that inhibit the activity of these enzymes. Specifically, as an example of the former, substances such as kojic acid and arbutin are used as cosmetic additives (functional cosmetics, pages 213 to 223, edited by Japan Society for Cosmetic Science, CMC Publishing Co.), As an example of the latter, an extract from wheat is shown as an anti-obesity diet food (Shiraishi, Chemistry and Biology, 27, 491, 1989).
Year). However, in order to meet many demands, it is necessary to supply an even more effective and safe enzyme inhibitor in large quantities, and its development is awaited.

[0005] Furthermore, in addition to the above-mentioned cosmetics and diet foods, mushrooms, especially maitake mushrooms, as illness prevention and health foods, which are of great interest today, are cultivated and used in different amounts. Maitake foods are widely available on the market, and especially in order to supply the extract as a health food, enormous amounts of maitake must be used. The fact is that Maitakemat is being disposed of as industrial waste, which is confusing to dispose of it. The estimated amount is that the cultivation amount is several thousand tons per month, the waste part is estimated to be about one tenth of several hundred tons, and most of them are incinerated. It has been desired and has become a research topic for the parties concerned.

[0006] The present inventors have set out to solve the above-mentioned problems.
Focusing on the fact that tyrosinase and α-amylase are present more in the rhizome (maitake mat) of non-edible parts such as mushrooms, which are usually discarded, than in the edible part. As a result of intensive studies on the assumption that these inhibitors, that is, tyrosinase inhibitor and α-amylase inhibitor, are also present in a high proportion, the above object was achieved.

Further, the present inventors have found that the heat treatment of these extracts further increases the inhibitory activity thereof, and have completed the present invention.

[0008]

In order to achieve the above object, the invention according to claim 1 is characterized in that the rhizome of wild and / or cultivated mushrooms is extracted with an organic solvent, and the tyrosinase inhibitor and α-amylase are extracted. It is an enzyme inhibitor consisting of an inhibitor.

[0009] The invention according to claim 2 is an enzyme inhibitor comprising a tyrosinase inhibitor and an α-amylase inhibitor, wherein a rhizome (maitake mat) of wild and / or cultivated maitake is extracted with an organic solvent. is there.

A third aspect of the present invention is a tyrosinase inhibitor, wherein a rhizome of wild and / or cultivated mushrooms and a rhizome of wild and / or cultivated maitake (maitake mat) are extracted with an organic solvent. And α-amylase inhibitors.

[0011] The invention of claim 4 is characterized in that the extraction is performed using a solvent selected from methanol, ethanol, propyl alcohol, isopropyl alcohol and a mixture thereof or a mixture thereof with water. Item 14. The enzyme inhibitor according to any one of Items 1 to 3.

[0012] The invention of claim 5 is the enzyme inhibitor according to any one of claims 1 to 4, wherein the extract is heated or heat-treated after the extraction.

The invention according to claim 6 is the enzyme inhibitor according to claim 5, wherein the heat treatment is performed at 50 ° C to 120 ° C for 0.5 hours to 5 hours.

[0014] The invention of claim 7 is a cosmetic for whitening and / or moisturizing containing the enzyme inhibitor according to claims 1 to 6.

[0015] The invention of claim 8 is a whitening and / or moisturizing bath agent containing the enzyme inhibitor according to any one of claims 1 to 6.

A ninth aspect of the present invention is an anti-obesity food containing the enzyme inhibitor according to any one of the first to sixth aspects.

The enzyme inhibitor of the present invention comprises a tyrosinase inhibitor and an α-amylase inhibitor obtained by extracting the rhizomes of wild and / or cultivated mushrooms with an organic solvent. Inhibitors include extracts from rhizomes of any mushrooms containing enzyme inhibitory activity. A typical example is maitake. The rhizome of maitake is usually called a maitake mat in a non-edible portion, and as described above, in maitake, the enzyme inhibitor activity is higher in the non-edible portion than in the edible portion. Was found to have. It was found that these extracted fractions have a moisturizing effect in addition to the above-mentioned effects. At present, it is not clear whether the inhibitor active ingredient and the moisturizing ingredient are the same substance. Furthermore, it was found that by heating and heating these extracts, tyrosinase and α-amylase were further inactivated, and as a result, tyrosinase inhibitor and α-amylase inhibitor activities were further increased.

In order to obtain the enzyme inhibitor of the present invention, the rhizomes and / or maitake mats of wild and / or cultivated mushrooms are used as they are. All are cut into small pieces and extracted with a suitable solvent. For the purpose of the present invention, the rhizome of raw mushrooms,
In addition to maitake mats, their dried products can also be used. Further, the material may be pulverized using a mixer or the like in order to improve the extraction efficiency. Examples of the extraction solvent include alcohols such as methanol, ethanol, propyl alcohol and isopropyl alcohol, and a mixture thereof or a mixture thereof with water can be selected. It is preferable to use ethanol in consideration of the extraction efficiency and safety to the human body.

The water content of alcohols depends on the water content of the material. For example, when a dry product is used, 50 to 80%
Although it can be extracted with a certain amount of alcohol, it is preferable to extract with anhydrous alcohol when the rhizome of a high water content mushroom immediately after harvest and a maitake mat are the raw materials. The ratio of the extraction solvent to the material is about 300 ml to 1000 ml for 100 g of the material. The extraction temperature and time are not particularly limited, and are usually between room temperature and the boiling temperature of the extraction solvent. The period required for extraction also varies depending on the raw material, extraction temperature, type of solvent, amount used, amount of water contained in the solvent, etc., but at room temperature, usually about 1 to 10 days, and at the boiling temperature of the solvent, several times. Sufficient to several hours, usually about one hour, is sufficient. The solution thus extracted is roughly filtered with gauze or the like, and further filtered using a filter paper, a glass filter, and the like, and then the solvent is distilled off under reduced pressure to obtain a syrup of the target enzyme inhibitor. Can be This syrup can be used as it is or as a freeze-dried powder to produce a food or cosmetic product by blending it with cosmetic raw materials or food materials, but it can be further purified by chromatography or other means if necessary. it can.

Further, the above extract is heated or heated as required. Tyrosinase inhibitor and α-amylase inhibitor are extracted by the above steps,
Since some corresponding enzymes are also involved, these warming or heating deactivates the heat-labile corresponding enzymes.

When the syrup or freeze-dried powder obtained from the rhizome part of the mushroom or the maitake mat of the present invention is added to a food such as confectionery, a diet food is obtained by its α-amylase inhibitor activity. When added to emulsions, lotions, etc., whitening cosmetics and moisturizing cosmetics are obtained by their tyrosinase inhibitor activity, and various inorganic salts, liquid lanolin, hexadecyl alcohol, polyoxyethylene glycols, and hardened castor oil are used as bath agent components. When added, a tyrosinase inhibitor and a whitening bath agent, a moisturizing bath agent and the like due to moisturizing activity are obtained.

[0022]

BEST MODE FOR CARRYING OUT THE INVENTION

Embodiments of the present invention will be described below. Example 1 After 200 g of Maitake mat was cut finely, ethanol 1
1 and soak at room temperature for 5 days. This was filtered, the filtrate was concentrated under reduced pressure, and purified water 1 was added to the resulting tan syrup.
After addition of 00 ml, the mixture was heated at 50 ° C. for 30 minutes, filtered again, and the filtrate was concentrated under reduced pressure to obtain 2 g of the desired syrup. This product may be freeze-dried.

Example 2 After 200 g of Maitake mat was cut finely, ethanol 1
1 and left in a thermostat at 40 ° C. for 24 hours.
3g of the desired syrup can be obtained.

Example 3 After 200 g of Maitake mat was cut finely, ethanol 1
l, and left in a constant temperature bath at 40 ° C. for 24 hours, followed by filtration.
The filtrate was concentrated under reduced pressure, and purified water was added to the resulting syrup.
After dissolving in water, the solution is placed in a dialysis membrane and dialyzed with running water for 24 hours. The inner dialysis solution is heated at 70 ° C. for 30 minutes and then concentrated to dryness under reduced pressure to obtain 1 g of the desired syrup.

Example 4 Preparation of whitening cream 5 ml of the extract obtained in Example 1 was taken, the solvent was distilled off under reduced pressure, and then a whitening cream was prepared according to the following formula. Maitakemat Extract Extract 5 ml equivalent Stearyl alcohol 6.0 Stearic acid 1.0 Hydrogenated lanolin 5.0 Octyldodecyl alcohol 10.0 Polyoxyethylene (29 mol) monocetyl alcohol ether 4.0 Glycerin monostearate 2.0 Propylene glycol 5.0 Purified water 66.0 Perfume Appropriate amount Preservative Appropriate amount Thus, a whitening cream was obtained.

The extract prepared in the above example was tested for tyrosinase inhibitor activity. Test Example 1 Tyrosinase inhibitor activity of Maitakemat extract L-tyrosine aqueous solution (30 mg / 200 ml) 10 ml buffer (pH 6.8 McKilbain buffer) 10 ml distilled water 20 ml 1% solution of powder obtained in Example 1 0.9 ml After heating the solution having the above composition at 37 ° C. for 10 minutes, 0.1 ml of an aqueous tyrosinase solution (1 mg / ml, derived from mushroom, Wako Pure Chemical Industries) is added, and the enzyme reaction is performed for 30 minutes. As a control, an enzyme reaction is carried out using the same amount of distilled water instead of the unextracted extract. The amount of dopachrome (a substance at the previous stage when melanin is produced) produced by tyrosinase was determined by measuring the absorbance at 650 nm, and the tyrosinase inhibitor activity of the syrup solution extracted with Maytakemat was calculated by the following formula. Inhibitor activity = [(AB) / A] × 100 A: Absorbance of control solution B: Absorbance of extract-containing sample

The results are shown in Table 1.

[0027]

[Table 1] Maitake mat: This corresponds to the rhizome of Maitake and is usually discarded in the non-edible part. As seen in Table 1, the mat portion has a higher tyrosinase inhibitor activity than the edible portion. In addition, tyrosinase inhibitor activity increases by heating.

Example 5 Preparation of Cookies for Diet 300 g of shortening, 20 g of milk, 50 g of sugar and 5 g of aspartame are thoroughly mixed with a whisk. Add 60g of the egg little by little and mix well. Separately, 300 g of flour, 1 g of baking powder, and a solvent equivalent to 50 ml of the extract of Maitake mat obtained in Example 1 were completely distilled off under reduced pressure and kneaded well. This is mixed well with the previous mixture and then left in the refrigerator for 30 minutes. Next, this is molded into an appropriate mold and baked at about 170 ° C. for 20 minutes to obtain about 1,000 g of the desired cookie.

Test Example 2 .alpha.-Amylase Inhibitor Activity of Maitakemat Extract Nine test tubes were filled with 1 ml of water, and 1 ml of .alpha.-amylase (0.5% aqueous solution, derived from bacteria, Add light pure drug) and mix well. 1ml of them
Into a second test tube and mix. Add 1 ml of the mixture to a third test tube and mix. This operation is sequentially repeated to form a dilution series up to the ninth test tube. Discard 1 ml of the ninth liquid. A test tube containing only 1 ml of the amylase solution without water was also prepared and used as the 0th tube. The above is cooled in ice water. Next, 5 ml of a 1% soluble starch solution was added to each test tube, and after all the additions were completed, all at 40 ° C.
And perform enzyme reaction for 1 hour. Next, the test tube is placed in ice water and cooled. After cooling, test tube height 80
% Of water, and one drop of 0.1N iodine solution is added thereto, and the color (blue-violet to red) due to the iodine starch reaction is observed. A solution of 1 ml + 9 ml of the extract obtained in Example 1 was placed in each test tube instead of 1 ml of the above-mentioned water, and an enzymatic reaction was carried out in the same manner, and the coloration due to the iodine starch reaction was observed. The results are as shown in Table 1 below. In addition, ● represents iodine starch reaction positive, and ○ represents negative.

[0030]

[Table 2] As can be seen in Table 2, the mat part is α- compared to the edible part.
High amylase inhibitor activity. In addition, α
The amylase inhibitor activity is increased.

Example 6 Sodium sesquicarbonate 37.0% salt 10.0 Sodium sulfate 49.3 Maitake mat syrup 3.0 Liquid lanolin 0.5 Dye 0.2 By this, a bath agent for whitening and rich in moisture retention I got

Example 7 Hexadecyl alcohol 27.0% Polyethylene glycol 300 21.0 Polyoxyethylene (40 mol) Hydrogenated castor oil 7.0 Ethyl alcohol 40.0 Perfume Appropriate dye Appropriate UV absorber Appropriate Maitake mat syrup 0 By this, a bath agent for whitening and rich in moisture retention was obtained.

Test Example 3 Moisturizing Action of Bath Agent Added with Maitake Mat Extract Extract The moisturizing action of the bath agent to which the extract of Maitake mat obtained in Example 1 was added was described in the literature (Tezuka et al., West Japan Dermatological Magazine 8).
(P.02, 1975). The moisturizing effect of the bath agent added with the Maitakemat extract solution (0.5, 1.0, 2.0% solution) and the bath agent not added with the extract were compared. Fresh cow nose and hooves were obtained from the Meat Sanitation Laboratory (Yokohama), and after removing the subcutaneous tissue as much as possible, they were immediately cut into a size of about 1 cm ² and sliced to a thickness of 100 μm by a cryosection method. . Next, the specimen was immersed in each of the above-described sample solutions at room temperature for 18 hours, and then left in a desiccator at 25 ° C. and a relative humidity of 79% for 72 hours to determine a water retention amount (mg / 100 mg) of the section. The result

The results are shown in Table 3.

[0034]

[Table 3] From Table 3, it is clear that the bath agent added with the Maitake mat extraction solution of the present invention has a moisturizing effect that surpasses existing bath agents.

[0035]

As described above, according to the present invention, most of them use the rhizomes and / or maitake mats of mushrooms, which are usually incinerated and disposed of in large quantities. It is in line with government. In addition, by utilizing the tyrosinase inhibitor activity contained in these extracts, melanin can be suppressed and a whitening effect can be exerted, and it can be widely used in whitening cosmetics and whitening bath agents.

Further, by utilizing the α-amylase inhibitor activity contained in the rhizome part of the mushroom and / or the extract of Maitake mat, the action of α-amylase is suppressed, the anti-obesity effect is exerted, and it is widely used in diet foods. be able to.

Further, the moisturizing effect contained in the rhizome part of the mushroom and / or the extract of Maitake mat can be widely used for moisturizing cosmetics and moisturizing bath agents.

According to the first aspect of the present invention, the rhizome of wild and / or cultivated mushrooms can be widely used, and the enzyme inhibition comprising a tyrosinase inhibitor and an α-amylase inhibitor extracted with an organic solvent. An agent can be obtained, which has the above effects.

According to the second aspect of the present invention, it is possible to obtain an enzyme inhibitor comprising a tyrosinase inhibitor and an α-amylase inhibitor, which extract a wild and / or a sufficiently cultivated Maitake mat with an organic solvent. This has the above effects.

According to the third aspect of the present invention, a tyrosinase inhibitor and α which extract the rhizome of wild and / or cultivated mushrooms and the wild and / or cultivated Maitake mat with an organic solvent can be used. -It is possible to obtain an enzyme inhibitor comprising an amylase inhibitor, which has the above effects.

According to the fourth aspect of the present invention, the extraction is carried out using a solvent selected from methanol, ethanol, propyl alcohol, isopropyl alcohol and a mixture thereof or a mixture thereof with water. It is performed more appropriately and has the above effects.

According to the fifth aspect of the present invention, since the extract is heated or heat-treated after the extraction, the corresponding enzyme can be further deactivated, and the above-mentioned effect is obtained.

According to the sixth aspect of the present invention, the extract is subjected to a heat treatment at 50 ° C. to 120 ° C. for 0.5 hours to 5 hours after the extraction, so that the corresponding enzyme is more appropriately and reliably inactivated. Can have the above effects.

According to the seventh aspect of the present invention, since the enzyme inhibitor is used, there is an effect that a whitening cosmetic and a moisturizing cosmetic having a supple and moist feeling can be obtained.

Further, according to the invention of claim 8, since the enzyme inhibitor is used, there is an effect that a whitening bath agent and a moisturizing bath agent having a supple and moist feeling can be obtained.

According to the ninth aspect of the present invention, since the enzyme inhibitor is used, there is an effect that a diet food having an appropriate diet action can be obtained.

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Claims (9)

[Claims] 1. An enzyme inhibitor comprising a tyrosinase inhibitor and an α-amylase inhibitor, wherein rhizomes of wild and / or cultivated mushrooms are extracted with an organic solvent. 2. An enzyme inhibitor comprising a tyrosinase inhibitor and an α-amylase inhibitor, wherein the rhizome (maitake mat) of wild and / or cultivated maitake is extracted with an organic solvent. 3. A tyrosinase inhibitor and an α-amylase inhibitor, wherein the rhizomes of wild and / or cultivated mushrooms and the rhizomes of wild and / or cultivated maitake (Maitake mat) are extracted with an organic solvent. Enzyme inhibitors. 4. The extraction comprises methanol, ethanol,
The enzyme inhibitor according to any one of claims 1 to 3, wherein the reaction is performed using a solvent selected from propyl alcohol, isopropyl alcohol, a mixture thereof, and a mixture thereof with water. 5. The enzyme inhibitor according to claim 1, wherein the extract is heated or heat-treated after the extraction. 6. The heat treatment is performed at 50 ° C. to 120 ° C.
The method is performed for 0.5 to 5 hours.
An enzyme inhibitor according to the above. 7. A whitening and / or moisturizing cosmetic containing the enzyme inhibitor according to claim 1. 8. A whitening and / or moisturizing bath agent comprising the enzyme inhibitor according to claim 1. 9. An anti-obesity food containing the enzyme inhibitor according to claim 1.