JP2002179592A - Composition for removing abnormal protein - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a composition for removing abnormal protein, or a composition capable of preventing diseases due to abnormal proteolysis accompanied by abnormal protein accumulation. SOLUTION: This composition for removing abnormal protein comprises as the active ingredient a compound having proteasome activity promotive action; wherein the compound is preferably a saponin derived from soybean. This composition is effective in preventing or treating diseases or disorders due to abnormal proteolysis including Alzheimer's disease, Parkinson's disease, Lewy bodies disease, triplet repeat disease, amyotrophic lateral sclerosis, cataract, arteriosclerosis, diabetic nephropathy and skin photoaging.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

[0001] The present invention relates to a composition for removing abnormal proteins comprising a compound having a proteasome activity promoting action as an active ingredient, and a composition for preventing a disease accompanied by a decrease in proteasome activity or accumulation of abnormal proteins.

[0002]

2. Description of the Related Art Abnormal proteins increase with age, and abnormal proteins accumulate in the living body, causing Alzheimer's disease, cataract,
It has been shown to be involved in many diseases such as wrinkles in the skin. At present, prevention and treatment of diseases caused by abnormal proteolysis are a major issue.
With respect to the protection of abnormal proteins from bioaccumulation, studies have been made to protect proteins from oxidative modification. That is, this is an attempt to eliminate active oxygen generated to the extent that the biological defense mechanism cannot cope with the oxidative stress by ingesting an antioxidant, thereby suppressing protein oxidation. Representative antioxidants include tocophenols, carotenoids, and flavonoids found in various plants.

[0003] However, ingestion of an antioxidant contributes to eliminating active oxygen generated in a living body, but does not contribute to removing abnormal proteins that have already accumulated. Therefore,
For the prevention and treatment of various diseases involving abnormal proteins accumulated in a living body with aging, removal of abnormal proteins is essential.

[0004]

SUMMARY OF THE INVENTION The present inventor has considered that removal of an abnormal protein is effective for prevention and treatment of a disease caused by accumulation of an abnormal protein in a living body. An object of the present invention is to provide a composition for removing abnormal proteins and to provide an anti-aging composition that contributes to prevention and treatment of diseases caused by abnormal protein degradation.

[0005]

Means for Solving the Problems In order to achieve the above object, the present inventors have paid attention to proteasome. The proteasome is a huge multi-component complex having an extremely complex molecular structure, and in recent years, attention has been paid to research on its physiological functions in vivo. Proteasomes are involved in the active removal of mutations, impaired proteins or abnormal proteins that have impaired normal folding or molecular assembly. That is, the proteasome is closely related to the protein quality control mechanism in the endoplasmic reticulum and cytoplasm and the stress response related thereto. As described above, the proteasome is a multifunctional proteolytic system that controls the active degradation of short-lived functional proteins in cells and abnormal proteins caused by mutations and stress, and is a central molecule that maintains and monitors cell homeostasis. Is known to be. However, there is no known technique for enhancing the activity of proteasome in vivo.

[0006] In the course of studying the proteasome activity promoting action, the present inventors have found that soybean-derived saponins have a proteasome activating effect and a concomitant effect of removing abnormal proteins. The present inventors have conducted further studies based on this fact and completed the present invention.

[0007] That is, the present invention is a composition for removing abnormal proteins comprising a compound having a proteasome activity promoting action as an active ingredient, wherein the compound having a proteasome activity promoting action is a soybean-derived saponin. It is a composition for use. Furthermore, the present invention is an anti-aging composition that contributes to the prevention and treatment of diseases caused by abnormal protein degradation, comprising a compound having a proteasome activity promoting effect as an active ingredient, and the compound having a proteasome activity promoting effect is derived from soybean. The anti-aging composition is saponin. Furthermore, the present invention relates to a cosmetic comprising a compound having a proteasome activity promoting action as an active ingredient, wherein the compound having a proteasome activity promoting action is soybean-derived saponin. Furthermore, the present invention relates to the cosmetic for anti-aging and the cosmetic for wrinkle suppression. Further, the present invention is an anti-aging composition comprising a compound having a proteasome activity promoting action and a compound having an antioxidant action.

[0008]

DESCRIPTION OF THE PREFERRED EMBODIMENTS The present invention will be described below in detail. In the present invention, the term "proteasome activity" refers to proteolytic activity and proteolytic activity of the proteasome. The abnormal protein generally refers to a protein that has undergone oxidation, saccharification, or aldehyde modification with aging. In the present invention, a compound having a proteasome activity promoting action is used as an active ingredient. Such compounds are not particularly limited, but preferred are soybean-derived saponins. Soybean-derived saponins are widely distributed in soybean seed coats, cotyledons, hypocotyls or the leaves, stems, roots, etc. of soybean plants. It is structurally similar to glycyrrhizin, but has a sugar chain consisting of 2 to 5 sugars in the triterpenoid skeleton.

In general, many saponins exhibit hemolytic properties. However, when the present inventor measured the hemolysis index of soybean saponins obtained from soybeans in a 2% blood suspension in rabbits, it was found to be 100 or less similarly to ginseng saponins, indicating no hemolysis.

[0010] The composition for removing abnormal proteins of the present invention can contain a compound having an antioxidant action together with a proteasome activator such as saponin. Compounds exhibiting antioxidant is not particularly limited, for example, various vitamins, various polyphenols, tocotrienols, natural ingredients and the like which contain coenzyme Q _10, and their.

The compound having a proteasome activity promoting action can be used for anti-aging. further,
A composition containing both a compound having a proteasome activity promoting action and a compound having an antioxidant action has an anti-aging action, and provides an anti-aging composition having a function of preventing abnormal protein accumulation and removing abnormal protein. Can be.

[0012] The compound having a proteasome activating action can be used as a cosmetic.
It is considered to have anti-aging and anti-wrinkle applications. The composition of the present invention can be administered orally or parenterally, such as those without hemolysis, which are administered as injections. When administered orally, it may be administered in the form of foods such as health foods and beauty foods.

The composition of the present invention may be, for example, an aqueous solution, an oil agent,
It can be applied in the form of liquid preparations such as emulsions and suspensions, semisolid preparations such as gels and creams, and solid preparations such as powders, granules, tablets and capsules. These forms can be prepared by a conventionally known method to prepare various dosage forms. Lotions, emulsions, creams, ointments, plasters, cataplasms, aerosols and the like are suitable as external skin preparations.

When the composition of the present invention is in the form of a food,
Additives such as starch commonly used in the field of foods can be blended. Granules, tablets, powders, capsules can be in the form of such a formulation,
It can be manufactured by commonly used formulation techniques.

The cosmetics of the present invention include oils and fats such as vegetable oils, higher fatty acids, higher alcohols, silicones, anionic surfactants, cationic surfactants, amphoteric surfactants,
Nonionic surfactants, preservatives, sugars, sequestering agents, polymers such as water-soluble polymers, thickeners, powder components,
An ultraviolet absorber, an ultraviolet blocker, a humectant such as hyaluronic acid, a fragrance, a pH adjuster and the like can be contained.
Vitamins, skin activators, blood circulation enhancers, resident bacteria control agents, active oxygen scavengers, anti-inflammatory agents, anti-cancer agents, whitening agents,
Other medicinal and physiologically active ingredients such as a bactericide can be contained.

As cosmetics, skin cosmetics such as lotions, emulsions, creams and packs, makeup base lotions, makeup creams, emulsions or creamy or ointment type foundations, hand creams, leg creams, body lotions Such as body cosmetics, etc.
It can be used as a bath additive or hair cosmetic. Usually, these dosage forms can be produced according to the formulation method used in cosmetics.

The composition for removing abnormal proteins and the composition for anti-aging according to the present invention contain saponin as an active ingredient.
If necessary, additives such as a pharmaceutically acceptable diluent or carrier can be contained. In addition, the composition of the present invention can contain other pharmaceutically active ingredients as required. The composition for removing abnormal proteins of the present invention,
It preferably contains about 0.1 to 99.5% by weight, more preferably about 0.5 to 95% by weight, of a compound having a proteasome activity promoting action. The dose and frequency of administration vary depending on individual differences (age, gender, body weight, etc.).
The compound having a proteasome activity promoting action per day may be appropriately adjusted in the range of 0.01 g to 9 g, and may be administered once or several times.

The abnormal protein originally produced in the living body is degraded by the proteasome, but when the proteasome activity decreases with aging, the abnormal protein is not decomposed, and the abnormal protein accumulates in the living body. Currently, accumulated abnormal proteins are involved in diseases such as Alzheimer's disease, Parkinson's disease, Lewy body disease, triplet repeat disease, amyotrophic lateral sclerosis, cataract, arteriosclerosis, diabetic nephropathy or skin photoaging It is known. Therefore, it is considered that the above-mentioned diseases can be prevented or treated by ingesting the composition for removing abnormal proteins of the present invention. (The FASEBJournal, Vol. 9, pp. 1173-1182, 1995
Year)

The composition of the present invention is useful as a cosmetic or health food for preventing aging and preventing aging, an anti-aging cosmetic or a beauty food, a rust prevention and anti-rust cosmetic or a health food. The composition for removing abnormal proteins of the present invention exhibits excellent effects on mammals and is highly safe.

[0020]

The present invention will be described in more detail with reference to the following examples, but the present invention is not limited to these examples. Example 1 [Abnormal protein removal effect test]
Once a day, 50 mg / kg of zinc chloride (manufactured by Wako Pure Chemical Industries, Ltd.) is injected subcutaneously to give oxidative stress, and soybean saponin (manufactured by Tokiwa Kampo Pharmaceutical Co., Ltd.) is administered twice a day according to the amount specified below.
Gavage oral administration by gastric sonde was performed for 10 days. All the groups were fed with CE-7 (CLEA Japan). The setting groups are shown below.

[0021]

[Table 1]

There was no difference in the change in weight of the rats among the five groups during the breeding period. After sacrifice on the 11th day of breeding, the liver was removed and the proteasome activity and the amount of oxidized protein in the liver were measured.

(1) Proteasome activity As a sample, known methods (Hayashi et al., Mechanisms of
Aging and Development, 102, 55-66, 1998)
The proteasome solution extracted from the liver was used.
N-CB as a substrate for measuring proteasome activity
Z-Leu-Leu-Glu-naphthylamine (Sigma) was used. 100 prepared in 0.1 M Tris buffer (pH 8.0)
μM N-CBZ-Leu-Leu-Glu-naphthylamine solution 10.5 μl
Was added with 39.5 μl of a proteasome solution prepared with 50 mM Tris buffer (pH 7.5), and the mixture was incubated at 37 ° C. for 30 minutes. The fluorescence intensity of the released β-naphthylamine was measured at Ex315 nm and Em420 nm. The result is shown in FIG. The greater the fluorescence intensity, the greater the proteasome activity. As is clear from FIG. 1, the proteasome activity significantly increased in the soybean saponin-administered group, indicating the proteasome activating effect of soybean saponin.

(2) Measurement of Oxidized Protein Amount 20 mg of liver was homogenized by adding 1 ml of 0.1 M phosphate buffer (pH 7.5), centrifuged at 1500 × g for 30 minutes, and the supernatant was analyzed. Dinitrophenylhydrazine (DNPH) conversion of protein is performed by a known method (Nakamura et al., Journal of Japan).
Biochemistry, 119, 768-774, 1996). The DNPH-converted protein was subjected to SDS-PAGE and transferred to a Polyvinylidene difluoride (PVDF) membrane using a protein transfer device. After the transfer, the membrane was blocked in a 3% skim milk solution for 2 hours, reacted with an anti-DNPH antibody for 2 hours, and treated with a 0.05%
After washing with Tween20 solution, it was reacted with [ ¹²⁵ I] -ProteinA. Further, the plate was washed with a 0.05% Tween 20 solution, exposed on an imaging plate, and subjected to image analysis. The result is shown in FIG. Oxidized protein content is detected as carbonylated protein content. As is clear from FIG. 2, the amount of oxidized protein significantly decreased in the soybean saponin-administered group, and the oxidized protein removal effect of soybean saponin was shown.

Example 2 [Cataract improvement effect test] A cataract model was prepared by administering naphthalene to rats. Specifically, 6
To give oxidative stress to week-old male BN rats, naphthalene (manufactured by Wako Pure Chemical Industries, Ltd.) was administered for 4 weeks. kg was administered orally by gavage twice a week according to the following dosage. Soybean saponin (manufactured by Tokiwa Kampo Pharmaceutical Co., Ltd.)
Administration was started at the same time as naphthalene, and oral gavage with a gastric tube was performed twice daily for 28 days. All groups were fed with CE-7 (CLEA Japan). The setting groups are shown below.

[0026]

[Table 2]

On the 29th day after naphthalene administration, the rats were sacrificed, the lenses were excised, and the proteasome activity and the amount of oxidized protein in the lenses were measured.

(1) Proteasome Activity As a sample, a proteasome solution extracted from a lens in the same manner as in Example 1 was used. Boc-Leu-Arg-Arg-MCA (manufactured by Peptide Research Laboratories) was used as a substrate for measuring proteasome activity, and prepared and reacted in the same manner as in Example 1. Ex380nm, Em
Measured at 440 nm. The result is shown in FIG. The greater the fluorescence intensity, the greater the proteasome activity. As is clear from FIG. 3, the proteasome activity significantly increased in the soybean saponin administration group, indicating the proteasome activating effect of soybean saponin.

(2) Measurement of Oxidized Protein Mass A 50 mM Tris buffer (pH 7.
5) Add 20 times the weight of the lens and homogenize to 1500
The mixture was centrifuged at xg for 30 minutes, and the supernatant was quantified for oxidized proteins in the same manner as in Example 1. The result is shown in FIG.
Oxidized protein content is detected as carbonylated protein content. As is clear from FIG. 4, the amount of oxidized protein in the lens of the cataract model rat was significantly reduced in the soybean saponin-administered group, indicating the effect of removing oxidized protein by soybean saponin.

Formulation Example 1 [Capsule] Composition Soybean saponin: 50 mg Tocotrienol: 30 mg Beeswax: 10 mg Grape seed oil: 110 mg The above ingredients were mixed and filled into a capsule base containing gelatin and glycerin to obtain soft capsules. Was.

[0031]

The use of the proteasome activator of the present invention can suppress the accumulation of abnormal proteins. Accordingly, the preparations of the present invention may be used for treating diseases or disorders caused by abnormal protein degradation (Alzheimer's disease, Parkinson's disease, Lewy body disease, triplet repeat disease, amyotrophic lateral sclerosis, cataract, arteriosclerosis, diabetic nephropathy, skin Is effective in preventing or treating diseases caused by abnormal protein degradation such as photoaging. Furthermore, it is also useful as an anti-aging cosmetic or food.

[Brief description of the drawings]

FIG. 1 is a graph showing the relationship between the dose of saponin and proteasome activity in the liver. The height of the graph is Means ± SE
(Mean ± standard error), and the ★ mark indicates that it is significant at the 5% risk rate by the t-test (p <0.05, n = 5)

FIG. 2 is a graph showing the relationship between the dose of saponin and the amount of carbonylated protein in the liver. The height of the graph is Means ± SE
(Mean ± standard error), and the ★ mark indicates that it is significant at the 5% risk rate by the t-test (p <0.05, n = 5)

FIG. 3 is a graph showing the relationship between the dose of saponin and the activity of proteasome in a lens. The height of the graph is Means ± SE
(Mean ± standard error), and the ★ mark indicates that it is significant at the 5% risk rate by the t-test (p <0.05, n = 5)

FIG. 4 is a diagram showing the relationship between the dose of saponin and the amount of carbonylated protein in the lens. The height of the graph is Means ±
Represents SE (mean ± standard error).
Significance is shown at the risk rate (p <0.05, n = 5)

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A61K 35/78 A61K 35/78 X 45/06 45/06 A61P 9/10 101 A61P 9/10 101 13 / 12 13/12 17/16 17/16 21/04 21/04 25/16 25/16 25/28 25/28 27/12 27/12 43/00 111 43/00 111 (72) Inventor Yasuhide Okuhara Kanagawa No. 12-13 Kaminoshino, Totsuka-ku, Yokohama-shi, Japan, inside FANCL Central Research Laboratory (72) Inventor Irihito Ono 12-13, Kaminoshino, Totsuka-ku, Yokohama-shi, Kanagawa Prefecture F-Fan Central Research Institute (reference) 4B018 LE01 LE02 LE03 MD43 MD58 ME10 4C083 AA111 AD391 BB47 BB51 CC02 EE13 4C084 AA02 AA17 AA20 MA02 MA52 NA14 ZA021 ZA161 ZA331 ZA451 ZA811 ZA891 ZA941 ZC521 4C088 AB59 AC04 MA07 Z03A03 MA03 ZA03 MA08 ZA94 ZC52

Claims (9)

[Claims] 1. A composition for removing abnormal proteins, comprising a compound having a proteasome activity promoting action as an active ingredient. 2. The composition for removing abnormal proteins according to claim 1, wherein the compound having a proteasome activity promoting action is a soybean-derived saponin. 3. An anti-aging composition comprising a compound having a proteasome activity promoting action as an active ingredient. 4. The anti-aging composition according to claim 3, wherein the compound having a proteasome activity promoting action is soybean-derived saponin. 5. A cosmetic comprising a compound having a proteasome activity promoting action as an active ingredient. 6. The cosmetic according to claim 5, wherein the compound having a proteasome activity promoting action is soybean-derived saponin. 7. The cosmetic according to claim 5, which is an anti-aging cosmetic. 8. The cosmetic according to claim 5, which is a cosmetic for suppressing wrinkles. 9. An anti-aging composition comprising a compound having a proteasome activity promoting action and a compound having an antioxidant action.