Autophagy inducer

Abstract

The present disclosure addresses the problem of providing a novel autophagy inducer.　The present disclosure pertains to an autophagy inducer containing a triterpene compound, and the like. In particular, the autophagy-inducing action of cells is effectively promoted by containing an oleanane-type triterpene having a five-ring structure, oleanolic acid, an olive leaf extract, a Ligustrum lucidum extract, and the like. Said autophagy inducer can be formulated as a solid composition, a food and drink composition, a cosmetic, or a medicine, and an excipient can also be used in combination.

Description

Autophagy inducers

[0001] The present disclosure relates to autophagy inducers and the like.

[0002] Autophagy is a general term for a system that degrades cytoplasmic components, including proteins, and parts of the cytoplasm, such as intracellular organelles like mitochondria. Autophagy plays a role in providing amino acids by degrading proteins under starvation conditions. Autophagy is a series of processes in which an isolation membrane formed between the rough endoplasmic reticulum and mitochondrial contact sites in the cell ingests substrates, forming a double-membrane structure called an autophagosome, which then fuses with a lysosome to form a single-membrane autophagolysosome, supplying degradative enzymes and degrading the internal substrates. Autophagy is classified into macroautophagy, microautophagy, and chaperone-mediated autophagy, but macroautophagy is usually referred to as autophagy. Effective ingredients for inducing autophagy are currently being explored. For example, Patent Document 1 discloses an autophagy derivative containing quercetin glycoside, and Patent Document 2 discloses an autophagy inducer containing fulvic acid. Novel autophagy inducers distinct from these autophagy inducers are desired.

[0003] JP 2021-104968 A JP 2022-108114 A

[0004] An object of the present disclosure is to provide a novel autophagy inducer.

[0005] As a result of extensive investigations aimed at solving the above problems, the present inventors discovered that triterpene compounds are useful for inducing autophagy, and thus completed the present disclosure.

[0006] That is, the present disclosure is as follows. [1] An autophagy inducer containing a triterpene compound. [2] The autophagy inducer according to [1], wherein the triterpene compound is a pentacyclic triterpene compound. [3] The autophagy inducer according to [1] or [2], wherein the triterpene compound contains an oleanane-type triterpene compound. [4] The autophagy inducer according to any of [1] to [3], wherein the triterpene compound contains an oleanolic acid compound. [5] The autophagy inducer according to [1], wherein the triterpene compound is contained in an olive leaf extract. [6] The autophagy inducer according to [1], wherein the triterpene compound is contained in an olive leaf extract and a Ligustrum lucidum extract. [7] A solid composition comprising the autophagy inducer according to any of [1] to [6]. [8] The solid composition according to [7], further containing an excipient. [9] A food or beverage composition comprising the autophagy inducer according to any of [1] to [6].

[10] A cosmetic comprising the autophagy inducer according to any one of [1] to [6].

[11] A pharmaceutical comprising the autophagy inducer according to any one of [1] to [6].

[12] Use of a solid composition containing a triterpene compound for inducing autophagy.

[13] The autophagy inducer according to any one of [1] to [6], wherein the triterpene compound is a plant-derived compound.

[14] The autophagy inducer according to any one of [1] to [6], wherein the triterpene compound is a compound derived from olive leaves.

[15] The autophagy inducer according to any one of [1] to [6], wherein the triterpene compound is a compound derived from Ligustrum lucidum.

[16] The autophagy inducer according to any one of [1] to [6], wherein the triterpene compound is extracted from a plant in an ethanol or ethanol-water mixture solution.

[17] A method for inducing autophagy, comprising administering an effective amount of a triterpene compound to a subject.

[18] Use of a triterpene compound for inducing autophagy.

[19] An autophagy inducer composition containing a triterpene compound and an excipient.

[20] A solid composition for inducing autophagy, comprising the autophagy inducer described in any one of [1] to [6].

[21] A food or drink composition for inducing autophagy, comprising the autophagy inducer described in any one of [1] to [6].

[22] A cosmetic for inducing autophagy, comprising the autophagy inducer described in any one of [1] to [6].

[23] A pharmaceutical for inducing autophagy, comprising the autophagy inducer described in any one of [1] to [6].

[0007] [1A] A solid composition comprising an autophagy inducer containing a triterpene compound as an active ingredient and an excipient. [2A] The solid composition according to [1A], wherein the triterpene compound is a five-ring triterpene compound. [3A] The solid composition according to [1A] or [2A], wherein the triterpene compound contains an oleanane-type triterpene compound. [4A] The solid composition according to any of [1A] to [3A], wherein the triterpene compound is an oleanolic acid compound. [5A] The solid composition according to any of [1A] to [4A], wherein the autophagy inducer is at least one autophagy inducer selected from the group consisting of olive leaf extract, Swertia japonica extract, and Ligustrum lucidum extract. [6A] The solid composition according to any of [1A] to [5A], wherein the autophagy inducer contains olive leaf extract and Ligustrum lucidum extract. [7A] The solid composition according to any one of [1A] to [6A], wherein the autophagy inducer is extracted from a plant in an ethanol or ethanol-water mixture solution. [8A] The solid composition according to any one of [1A] to [7A], further comprising a lubricant. [9A] The solid composition according to any one of [1A] to [8A], further comprising vitamins. [10A] The solid composition according to any one of [1A] to [9A], wherein the content of the triterpene compound is 1 to 10 mass% based on the total amount of the solid composition. [11A] The solid composition according to any one of [1A] to [10A], wherein the solid composition is prepared in the form of a tablet, pill, capsule, powder, granule, fine granule, or lozenge. [12A] The solid composition according to any one of [1A] to [11A], wherein the solid composition is for oral administration. [13A] The solid composition according to any one of [1A] to [12A], wherein the solid composition is further used in combination with an autophagy inducer other than the triterpene compound. [14A] A food or drink composition comprising the solid composition according to any one of [1A] to [13A]. [15A] The food or drink composition according to [14A], which is prepared as a supplement, a food with functional claims, a health food, an enteral nutritional food, a food for special dietary uses, a health supplement, a food for specified health uses, or a food with nutrient functions.

[0008] [1B] A solid composition comprising an autophagy inducer (1) containing a triterpene compound as an active ingredient and an autophagy inducer (2) containing a compound other than the triterpene compound as an active ingredient. [2B] The solid composition according to [1B], wherein the triterpene compound is a five-ring triterpene compound. [3B] The solid composition according to [1B] or [2B], wherein the triterpene compound contains an oleanane-type triterpene compound. [4B] The solid composition according to any of [1B] to [3B], wherein the triterpene compound is an oleanolic acid compound. [5B] The solid composition according to any of [1B] to [4B], wherein the autophagy inducer (1) is at least one autophagy inducer selected from the group consisting of olive leaf extract, Swertia japonica extract, and Ligustrum lucidum extract. [6B] The solid composition according to any of [1B] to [5B], wherein the autophagy inducer (1) contains olive leaf extract and Ligustrum lucidum extract. [7B] The solid composition according to any one of [1A] to [6A], wherein the autophagy inducer is extracted from a plant in an ethanol or ethanol-water mixture solution. [8B] The solid composition according to any one of [1B] to [7B], further comprising a lubricant. [9B] The solid composition according to any one of [1B] to [8B], further comprising vitamins. [10B] The solid composition according to any one of [1B] to [9B], wherein the content of the triterpene compound is 1 to 10% by mass, based on the total amount of the solid composition. [11B] The solid composition according to any one of [1B] to [10B], wherein the solid composition is prepared in the form of a tablet, pill, capsule, powder, granule, fine granule, or lozenge. [12B] The solid composition according to any one of [1B] to [11B], wherein the solid composition is for oral administration. [13B] The solid composition according to any one of [1B] to [12B], wherein the solid composition is further used in combination with an autophagy inducer other than the triterpene compound. [14B] A food or beverage composition containing the solid composition according to any one of [1B] to [13B].[15B] The food and beverage composition according to [14B], characterized in that it is prepared as a supplement, a functional food, a health food, an enteral nutritional food, a special purpose food, a health supplement, a food for specified health uses, or a food with nutrient functions.

[0009] [1C] A composition for inducing autophagy, comprising an olive leaf extract and a Ligustrum lucidum extract. [2C] The composition for inducing autophagy according to [1C], wherein the olive leaf extract and the Ligustrum lucidum extract contain a triterpene compound. [3C] The composition for inducing autophagy according to [2C], wherein the triterpene compound is a five-ring triterpene compound. [4C] The composition for inducing autophagy according to [2C] or [3C], wherein the triterpene compound contains an oleanane-type triterpene compound. [5C] The composition for inducing autophagy according to any of [2C] to [4C], wherein the triterpene compound is an oleanolic acid compound. [6C] The composition for inducing autophagy according to any of [1C] to [5C], wherein the olive leaf extract and the Ligustrum lucidum extract are extracted from ethanol or an ethanol-water mixed solution. [7C] The composition for inducing autophagy according to any of [1C] to [6C], further comprising an excipient. [8C] The composition for inducing autophagy according to any one of [1C] to [7C], further comprising a lubricant. [9C] The composition for inducing autophagy according to any one of [1C] to [8C], further comprising vitamins. [10C] The composition for inducing autophagy according to any one of [2C] to [9C], wherein the content of the triterpene compound is 1 to 10% by mass relative to the total amount. [11C] The composition for inducing autophagy according to any one of [1C] to [10C], which is prepared in the form of tablets, pills, capsules, powders, granules, fine granules, or lozenges. [12C] The composition for inducing autophagy according to any one of [1A] to [11A], which is for oral administration. [13C] The composition for inducing autophagy according to any one of [1C] to [12C], which is further used in combination with an autophagy inducer other than the olive leaf extract and Ligustrum lucidum extract. [14C] A food or drink composition containing the composition for inducing autophagy according to any one of [1A] to [13C].[15C] The food and beverage composition according to [14C], characterized in that it is prepared as a supplement, a functional food, a health food, an enteral nutritional food, a special purpose food, a health supplement, a food for specified health uses, or a food with nutrient functions.

[0010] According to the present disclosure, a novel autophagy inducer can be provided.

[0011] Figure 1 is a graph showing the autophagy effects of Examples 1 to 3 (oleanolic acid) and Comparative Examples 1 to 3 (resveratrol). Figure 2 is a graph showing the autophagy effects of Examples 4 and 5, Comparative Example 4, and Reference Examples 2 and 3.

[0012] The autophagy inducer of the present disclosure contains a triterpene compound. The triterpene compound includes triterpenes, their salts, their hydrates, and processed products thereof. The triterpene is a compound having a 30-carbon skeleton with six isoprene units, each of which has five carbon atoms. A representative example is a pentacyclic (five-ring) triterpene (hereinafter sometimes referred to as a pentacyclic triterpene).

[0013] Terpenes are biosynthesized via the mevalonate pathway, and terpenoids are known to exist widely in nature as secondary metabolic products of living organisms. Terpenes are composed of isoprene, a five-carbon unit (C5), and are classified into monoterpenes (C10), sesquiterpenes (C15), diterpenes (C20), sesterterpenes (C25), triterpenes (C30), tetraterpenes (C40), and the like.

[0014] Pentacyclic triterpenes (C30) are a type of triterpene, a pentacyclic compound consisting of six isoprene units. They basically have 30 carbon atoms, but may have different carbon atoms due to rearrangement, oxidation, elimination, or alkylation during the biosynthesis process. Pentacyclic triterpenes are generally classified by their skeleton. Pentacyclic triterpenes are not particularly limited, and examples include oleanane-type triterpenes, ursane-type triterpenes, lupane-type triterpenes, hopane-type triterpenes, selatan-type triterpenes, friedelane-type triterpenes, taraxerane-type triterpenes, taraxastan-type triterpenes, multiflorane-type triterpenes, and germanican-type triterpenes. Representative examples of oleanane-type triterpenes among pentacyclic triterpenes include oleanolic acid, β-amyrin, taraxerol, germanicol, and maslinic acid. Representative examples of ursane-type triterpenes include ursolic acid, α-amyrin, corosolic acid, tormentic acid, etc. Representative examples of lupane-type triterpenes include lupeol, betulinic acid, etc.

[0015] Oleanolic acid (molecular formula: C ３０ H ４８ O ３ , molecular weight: 456.7) has the structure of the following formula (1):

[0016]

[0017] Oleanolic acid has a basic structure of β-amyrin and is known to exist as a free form or as saponin in plants such as olive leaf (Olea europaea L.), Swertia japonica, clove, grape, beet, persimmon, plum, loquat, sea buckthorn, argan, ginseng, carrot, sugar beet, perilla, and ligustrum lucidum.

[0018] β-amyrin (molecular formula: C ３０ H ５０0, molecular weight: 426.7) is contained in, for example, Morus alba (herbal medicine name) and Acer nikoense. Morus alba (herbal medicine name) is the root bark of Morus alba L., a deciduous tall tree distributed in China and other regions, which belongs to the genus Mulberry in the family Mulberry. Acer nikoense is a deciduous tall tree belonging to the genus Acer in the family Amerasaceae.

[0019] Taraxerol (molecular formula: C ３０ H ５０ O, molecular weight: 426.7) is contained in plants of the Tilia family (Tilia cordata), Birch family (Alnus glutinosa), Asteraceae family (Teraxacum officinale), Lauraceae family (Litsea dealbata), etc.

[0020] Maslinic acid (molecular formula: C ３０ H ５０ O, molecular weight: 426.7) is found in, for example, olives, jujubes, almonds, banaba leaves, sage, apples, cranberries, and quince.

[0021] α-amyrin (molecular formula: C ３０ H ５０ O, molecular weight: 426.7) is found in a wide range of plants.

[0022] Ursolic acid (molecular formula: C ３０ H ５０ O, molecular weight: 426.7) is present in plants such as apples, basil, bilberries, cranberries, elderflowers, peppermint, rosemary, lavender, oregano, thyme, hawthorn, prunes, olives (leaves), almonds, raspberries, quince, guava, perilla leaves, blueberries, loquats, pomegranates, lemon balm, rose hips, persimmons, Swertia japonica, and bearberry.

[0023] Lupeol (molecular formula: C ３０ H ５０Lupeol (molecular weight: 426.7) is found in large amounts in legumes such as Lupinus luteus. Lupeol is also found in vegetables such as white cabbage, cucumber, and tomato, and in fruits such as mango, fig, strawberry, and red grape.

[0024] Betulinic acid (molecular formula: C ３０ H ５０ O, molecular weight: 426.7) is contained, for example, in chaga (obscured mushroom), a mushroom that parasitizes birch trees.

[0025] The triterpene compounds can be obtained by extraction and purification from the above-mentioned raw materials using known methods. Specifically, the triterpene compounds can be extracted from the above-mentioned raw materials using a lower alcohol (e.g., methanol, ethanol, n-propanol, isopropanol, n-butanol, etc.) from which the triterpene compounds can be extracted, or a hydrous alcohol thereof, followed by saponification, neutralization, etc., as necessary, and fractionation and purification using octadecylsilica group-bonded silica (ODS), silica gel, synthetic adsorbents, etc., as an adsorbent. Alternatively, the triterpene compounds can be obtained by chemical synthesis or as commercially available products. For example, commercially available oleanolic acid compounds are sold by, for example, Sigma-Aldrich, Fujifilm Wako Pure Chemical Industries, Ltd., Nacalai Tesque, Inc., Tokyo Chemical Industry Co., Ltd., and are easily available.

[0026] The autophagy inducers of the present disclosure include not only the triterpenes described above, but also their salts, hydrates, or processed products. The autophagy inducers of the present disclosure may be used in the form of a single triterpene, salt, hydrate, or processed product thereof, or in a combination of two or more forms.

[0027] The autophagy inducer of the present disclosure contains a triterpene or a processed triterpene as an active ingredient and is therefore capable of inducing autophagy. Note that the autophagy inducer can also be called an "autophagy activator."

[0028] The autophagy inducer of the present disclosure can also be a processed product of the triterpene. Examples of the processed product of the triterpene include triterpene derivatives. The triterpene derivatives can be produced from the triterpene using well-known techniques. When the triterpene compound is oleanolic acid, the oleanolic acid derivative may have a structure in which the hydroxyl group at position 3 in the beta configuration of formula (1) is changed to the alpha configuration. Furthermore, the carboxyl group at position 28 may be amidated, acylated, alkylated, or the like. These derivatives can be produced from oleanolic acid using well-known techniques.

[0029] The autophagy inducer of the present disclosure can also use triterpene salts.Triterpene salts generally refer to pharmacologically acceptable salts of triterpenes.Pharmacologically acceptable salts include, but are not limited to, metal salts such as sodium salt, potassium salt, calcium salt, magnesium salt, and aluminum salt; and salts with organic bases such as triethylamine, triethanolamine, and piperazine.

[0030] The autophagy inducer of the present disclosure may be a hydrate of a triterpene, a hydrate of a salt thereof (hydrate salt), or a hydrate of a processed product thereof. The hydrate is not particularly limited, and examples include a hemihydrate, and monohydrate to hexahydrate.

[0031] The autophagy inducer of the present disclosure may contain a triterpene compound, and thus the triterpene compound may be contained in the form of a plant or plant extract. When the triterpene compound is oleanolic acid, examples of plants containing oleanolic acid include olive leaf, Swertia japonica, Ligustrum lucidum, clove, grape, beet, persimmon, plum, loquat, sea buckthorn, argan, ginseng, carrot, sugar beet, and perilla. In other words, since the above plants contain triterpene compounds, plant extracts containing triterpene compounds can be used as the autophagy inducer of the present disclosure. The plant extract is not particularly limited as long as it contains a triterpene compound, and examples include extracts of the above plants. For example, when the triterpene compound is oleanolic acid, examples of plant extracts containing oleanolic acid include olive (leaf) extract, Swertia japonica extract, and Ligustrum lucidum extract. The triterpene compounds may be compounds, distillates, fractions, etc. obtained from plant extracts such as olive leaf extract, Swertia japonica extract, and Ligustrum lucidum extract. Distillates refer to portions separated by temperature ranges during distillation. Fractions refer to portions obtained by general separation procedures. In addition, the triterpene compounds may be compounds, extracts, distillates, fractions, etc. obtained from plants such as olive leaf extract, Swertia japonica, and Ligustrum lucidum.

[0032] Olive (leaf) extract can be obtained, for example, by the following method. Olives are evergreen trees of the genus Olea in the family Oleaceae. Olive varieties are not particularly limited, and olives can be used regardless of their origin (domestic or foreign), whether cultivated or used for oil extraction. Olive (leaf) includes olive fruit, olive leaves, olive stems, olive roots, etc. Among olive (leaf) extracts, those extracted solely from olive leaves are specifically referred to as "olive leaf extract." Olive leaf extract can be dried to produce olive leaf extract powder. The mass of oleanolic acid contained in the olive leaf extract powder is not particularly limited, but is, for example, 1-80%, preferably 5-70%, and more preferably 10-60%. Specifically, the olive leaf extract powder described in the following examples is a dried ethanol extract and contains approximately 55% oleanolic acid and other components in addition to oleanolic acid. Swertia japonica extract can be obtained, for example, by the following method. Swertia japonica is a biennial plant classified in the genus Swertia in the family Gentianaceae. It is used as a medicinal herb, and its herbal name and alternative name are "Touyaku." Ligustrum lucidum extract can be obtained, for example, by the following method. Ligustrum lucidum (scientific name: Ligustrum Lucidum Aiton) is an evergreen tall tree of the genus Ligustrum in the family Oleaceae. Ligustrum lucidum is primarily cultivated in China (Zhejiang, Jiangsu, Fujian, Hunan, Jiangxi, Guangxi, Sichuan, etc.). Dried mature fruits of Ligustrum lucidum are known as a medicinal herb called "Nüzhenzi" (nu zhenzi), and are said to have cardiotonic, diuretic, laxative, tonic, and aphrodisiac effects.

[0033] As used herein, the term "Ligustrum lucidum" refers to the plant body or a part thereof of the Ligustrum lucidum plant, unless otherwise specified. "A part thereof" includes organs or parts thereof (leaves, roots, stems, branches, flowers, stamens, pistils, or fragments thereof), fruits (sometimes referred to as "fruits"), seeds, cultured cells, callus, protoplasts, transformed plant cells, transformed plants, etc. The autophagy inducer of the present disclosure can be the entire Ligustrum lucidum plant, or any one or more of its organs. From the perspective of expected high efficacy, it is preferable to use the fruit or leaves. Fruits and leaves can also be used. As used herein, the term "Ligustrum lucidum fruit or leaves" refers to any one or more selected from the group consisting of Ligustrum lucidum fruit and Ligustrum lucidum leaves, unless otherwise specified.

[0034] The Ligustrum lucidum extract used in this specification can be obtained by extracting Ligustrum lucidum with an extraction solvent. The Ligustrum lucidum used as the extraction raw material may be fresh, dried, roasted, or the like. When dried, it may be naturally dried, dried under heated conditions, or frozen and dried. Before being subjected to the extraction step, pretreatment such as cutting or crushing may be performed to ensure efficient extraction. The Ligustrum lucidum extract used in the present disclosure can be prepared by a general method. For example, it can be prepared by immersing Ligustrum lucidum in a solvent. The extraction solvent is not particularly limited, and may be one or more selected from the group consisting of water, alcohols (monohydric alcohols such as methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, and 2-butanol; dihydric alcohols such as ethanediol, propylene glycol, 1,3-butanediol, and 1,4-butanediol; and trihydric alcohols such as glycerin), ketones (acetone, methyl ketone, and the like), ethers (diethyl ether, tetrahydrofuran, and the like), esters such as ethyl acetate, and hexane. The alcohols are preferably monohydric or dihydric alcohols. The number of carbon atoms in the alcohols is preferably about 1 to 5. Furthermore, water and / or ethanol are more preferred as the solvent, and ethanol and a water-ethanol mixed solution are particularly preferred.

[0035] Those skilled in the art can design the conditions for extraction as appropriate. For example, extraction can be performed by immersing Ligustrum lucidum in a solvent at room temperature or under heating for a period of 1 hour to several months. The ratio of raw Ligustrum lucidum to solvent can be designed as appropriate by those skilled in the art. For example, the amount of solvent per 1 part by mass of Ligustrum lucidum raw material may be an amount sufficient to fully immerse the raw material, and is preferably 0.5 parts by mass or more, more preferably 1 part by mass or more, and more preferably 2 parts by mass or more. The upper limit of the solvent amount can also be determined from an economical perspective. For example, the amount of solvent per 1 part by mass of raw material can be 100 parts by mass or less, preferably 50 parts by mass or less, and more preferably 25 parts by mass or less. The extraction temperature and period can also be designed as appropriate by those skilled in the art. Extraction can be performed by immersing Ligustrum lucidum in a solvent at room temperature or under heating for a period of 1 hour to several months.

[0036] A typical example of a method for preparing olive (leaf) extract and Swertia japonica extract that can be used in the present disclosure is as follows: The raw material is crushed to an appropriate size, soaked in 5 to 20 times the amount of 25 to 75% by volume ethanol at room temperature for one week, and the solid content is removed from the resulting soaking liquid. The resulting soaking liquid may be concentrated or dried, and can be obtained in powder form by drying.

[0037] In this disclosure, the term "extract" also includes distillates obtained by steam distillation. Steam distillation is a method in which steam is passed through natural raw materials, and the aroma components distilled along with the steam are condensed together with the steam. Methods include pressurized steam distillation, atmospheric steam distillation, reduced-pressure steam distillation, and gas-liquid multistage alternating current contact distillation (spinning cone column), and any of these can be used in this disclosure. Those skilled in the art can appropriately determine the pretreatment of raw materials for steam distillation, as well as the distillation time, temperature, and pressure, taking into account factors such as economic viability, based on the results of several experiments.

[0038] In this disclosure, unless otherwise specified, the term "extract" may refer to the liquid obtained from the extraction process (also referred to as "extract") itself (solids may be removed, if necessary), or the liquid may be concentrated, dried, or the like. The extract may be in any form, such as liquid, paste, gel, solid, or powder. To obtain a solid or powder form, the liquid may be subjected to a drying process and dried to a solid, or may be dried by spray drying to obtain a powder.

[0039] Unless otherwise specified, the term "agent" used in this disclosure may refer to the extract itself, or may refer to a combination of the extract and additives, such as a diluent, stabilizer, antioxidant, preservative, etc. The agent of the present disclosure does not include existing external skin compositions.

[0040] When the agent of the present disclosure contains a plant extract such as olive (leaf) extract, Swertia japonica extract, or Ligustrum lucidum extract, the plant extract can be contained in a solid content (amount equivalent to the dried matter obtained by the method shown in the examples of this specification) of preferably 0.00001% or more, more preferably 0.0001% or more, and even more preferably 0.001% or more. Note that, when the component content is expressed in % in this disclosure, it is % by mass (the rate of component mass per 100 g of total amount) unless otherwise specified.

[0041] The proportion of triterpene compounds contained in the autophagy inducer of the present disclosure can be, for example, 0.0001 to 99% by mass, 0.001 to 80% by mass, 0.01 to 70% by mass, 0.1 to 60% by mass, 0.1 to 50% by mass, 0.1 to 40% by mass, 0.1 to 30% by mass, 0.1 to 20% by mass, 1 to 10% by mass, etc. When the triterpene compounds contained in the autophagy inducer of the present disclosure are plant extracts, the proportion of the plant extract (e.g., olive leaf extract, Swertia japonica extract, Ligustrum lucidum extract, etc.) can be, for example, 0.0001 to 99% by mass, 0.001 to 80% by mass, 0.01 to 70% by mass, 0.1 to 60% by mass, 0.1 to 50% by mass, 0.1 to 40% by mass, 0.1 to 30% by mass, 0.1 to 20% by mass, 1 to 18% by mass, 2 to 15% by mass, etc.

[0042] When the triterpene compounds contained in the autophagy inducer of the present disclosure are plant extracts, they preferably contain an olive leaf extract and a Ligustrum lucidum extract. When both an olive leaf extract and a Ligustrum lucidum extract are contained, the content of the Ligustrum lucidum extract, in terms of triterpene compounds, is typically 0.01 to 10,000 parts by mass, preferably 0.1 to 1,000 parts by mass, more preferably 0.5 to 500 parts by mass, and even more preferably 1 to 300 parts by mass, per 100 parts by mass of the olive leaf extract. When both an olive leaf extract and a Ligustrum lucidum extract are contained, the content of the Ligustrum lucidum extract is typically 1 to 200 parts by mass, preferably 10 to 100 parts by mass, and more preferably 30 to 70 parts by mass, per 100 parts by mass of the olive leaf extract.

[0043] As described above, "autophagy (function)" refers to, for example, a system in which intracellular organelles or proteins are degraded via autophagosome formation within cells. Furthermore, activation of autophagy refers to a state in which the intracellular protein recycling mechanism is activated by the induction or promotion of autophagy within cells. Specifically, it refers to a state in which at least one of the following processes is induced or promoted: isolation membrane formation, isolation membrane elongation, autophagosome formation, and autophagosome-lysosome fusion. The autophagy inducers of the present disclosure can induce such autophagy (function). In this specification, induction is a concept that encompasses expression, improvement, enhancement, activation, and the like. Therefore, the present disclosure encompasses the use of triterpene compounds to induce autophagy.

[0044] Autophagy activity can be confirmed by conventionally known methods for measuring autophagy activity. Examples of such methods for measuring autophagy activity include methods for detecting autophagy marker proteins. Examples of autophagy marker proteins include rat microtubule-associated protein 1 light chain 3 (LC3). LC3 exists in two forms: LC3-I and LC3-II. LC3 exists in the cytoplasm as LC3-I, and upon autophagosome formation, LC3-I is converted to the lipid-bound form LC3-II, which is then incorporated into the inner and outer membranes of the autophagosome. Therefore, autophagy activity can be confirmed by detecting LC3-I and LC3-II and measuring the conversion rate to LC3-II (Kabeya Y., Mizushima N., Ueno T., Yamamoto A., Kirisako T., Noda T., Kominami E., Ohsumi Y., Yoshimori T.: LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing. The EMBO Journal, 19:5720-5728 (2000)).

[0045] Autophagy activity can also be confirmed by expressing tfLC3, a tandem fusion of LC3 with two fluorescent proteins (monomeric red fluorescent protein (mRFP) and enhanced green fluorescent protein (EGFP)) in cells (Kimura S., Noda T., Yoshimori T.: Dissection of the Autophagosome Maturation Process by a Novel Reporter Protein, Tandem Fluorescent-Tagged LC3. Autophagy 3:452-460 (2007)). According to this method, EGFP disappears in the (auto)lysosomal environment, and a lower EGFP / mRFP ratio in cells indicates more activated autophagy. The amount of fluorescence in cells can be measured using, for example, a fluorescence microscope, and the amount of fluorescence can be calculated using analytical software.

[0046] Uses The autophagy inducer of the present disclosure can be used as a solid composition, an oral composition, a food or drink composition, a pharmaceutical, or a cosmetic (hereinafter sometimes abbreviated as the "preparation of the present disclosure"), etc.

[0047] The autophagy inducer of the present disclosure may contain only a triterpene compound as an active ingredient, or may contain a triterpene compound (1) and an autophagy inducer other than the triterpene compound (2), and may further contain other ingredients as long as the effects of the present disclosure are not impaired.

[0048] The autophagy inducer (2) other than a triterpene compound is not particularly limited, and examples include known autophagy inducers such as rapamycin, torin-1, tRes (trans-resveratrol), metformin, lithium chloride, and carbamazepine. tRes is contained in grape skin extract powder. The autophagy inducer of the present disclosure preferably contains an autophagy inducer (1) containing one or more triterpene compounds and an autophagy inducer (2) other than the triterpene compounds, and more preferably contains an autophagy inducer (1) containing two or more triterpene compounds and an autophagy inducer (2) other than the triterpene compounds. When the autophagy inducer of the present disclosure contains triterpene compound (1) and an autophagy inducer (2) other than the triterpene compound (e.g., trans-resveratrol), the content of the autophagy inducer (2) other than the triterpene compound is typically 0.01 to 10,000 parts by mass, preferably 1 to 1,000 parts by mass, and more preferably 20 to 200 parts by mass, per 100 parts by mass of the triterpene compound (1).

[0049] The other components are not particularly limited, and examples thereof include pharmaceutically acceptable components (e.g., various organic or inorganic carrier substances commonly used as formulation materials, excipients, binders, disintegrants, lubricants, etc. in solid formulations; solvents, solubilizers, suspending agents, buffers, thickeners, emulsifiers, components known to have health benefits, etc. in liquid formulations). Furthermore, formulations of the present disclosure may also contain formulation additives such as colorants, sweeteners, and antioxidants, as needed. Furthermore, formulations of the present disclosure may be coated. The content of the other components in formulations of the present disclosure is not particularly limited, and is, for example, typically 5 parts by mass or more, preferably 10 to 100,000 parts by mass, and more preferably 100 to 10,000 parts by mass, per 100 parts by mass of the triterpene compound. Furthermore, the content of other components in the formulation of the present disclosure is not particularly limited, and is, for example, typically 1% by mass to 99.9% by mass or more, preferably 10% by mass to 98% by mass, and more preferably 30% by mass to 97% by mass.

[0050] Examples of excipients include dextrin, lactose, sucrose, D-mannitol, D-sorbitol, starch, pregelatinized starch, crystalline cellulose (e.g., microcrystalline cellulose, etc.), low-substituted hydroxypropyl cellulose, sodium carboxymethyl cellulose, gum arabic, pullulan, light anhydrous silicic acid, synthetic aluminum silicate, magnesium aluminometasilicate, etc. Among these, preferred excipients are dextrin, lactose, sucrose, D-mannitol, D-sorbitol, starch, pregelatinized starch, crystalline cellulose (e.g., microcrystalline cellulose, etc.), low-substituted hydroxypropyl cellulose, and sodium carboxymethyl cellulose, and more preferred are dextrin, lactose, sucrose, D-mannitol, D-sorbitol, starch, pregelatinized starch, and crystalline cellulose (e.g., microcrystalline cellulose, etc.). Examples of binders include pregelatinized starch, sucrose, gelatin, macrogol, gum arabic, methylcellulose, carboxymethylcellulose, sodium carboxymethylcellulose, crystalline cellulose, sucrose, D-mannitol, trehalose, dextrin, pullulan, hydroxypropyl cellulose (HPC), hydroxypropylmethylcellulose (HPMC), polyvinylpyrrolidone (PVP), etc. Examples of disintegrants include lactose, sucrose, starch, carboxymethylcellulose, calcium carboxymethylcellulose, crosslinked polyvinylpyrrolidone, carmellose sodium, croscarmellose sodium, sodium carboxymethyl starch, light anhydrous silicic acid, low-substituted hydroxypropyl cellulose, cation exchange resin, partially pregelatinized starch, corn starch, etc. Examples of lubricants include aerosil, sucrose fatty acid esters, stearic acid, magnesium stearate, calcium stearate, talc, waxes, colloidal silica, DL-leucine, sodium lauryl sulfate, magnesium lauryl sulfate, macrogol, etc. Among these, Aerosil, sucrose fatty acid ester, stearic acid, magnesium stearate, and calcium stearate are preferred as lubricants, and Aerosil, sucrose fatty acid ester, and calcium stearate are more preferred.

[0051] Examples of solvents include water for injection, physiological saline, Ringer's solution, alcohol, propylene glycol, polyethylene glycol, medium-chain triglycerides (MCT), vegetable oils (e.g., safflower oil, sesame oil, corn oil, olive oil, cottonseed oil, soybean lecithin, etc.), etc.

[0052] Examples of solubilizing agents include polyethylene glycol, propylene glycol, D-mannitol, trehalose, benzyl benzoate, ethanol, trisaminomethane, cholesterol, triethanolamine, sodium carbonate, sodium citrate, sodium salicylate, and sodium acetate.

[0053] Examples of suspending agents include surfactants such as stearyl triethanolamine, sodium lauryl sulfate, lauryl aminopropionic acid, lecithin, benzalkonium chloride, benzethonium chloride, and glycerin monostearate; hydrophilic polymers such as polyvinyl alcohol, polyvinylpyrrolidone, sodium carboxymethylcellulose, methylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, and hydroxypropylcellulose; polysorbates, and polyoxyethylene hydrogenated castor oil.

[0054] Examples of the buffering agent include buffer solutions such as phosphate, acetate, carbonate, and citrate.

[0055] Examples of thickeners include natural gums, cellulose derivatives, etc. Among them, natural gums are preferred as thickeners.

[0056] Examples of emulsifiers include fatty acid esters (e.g., sucrose fatty acid esters, glycerin fatty acid esters, sorbitan fatty acid esters, propylene glycol fatty acid esters, etc.), waxes (e.g., beeswax, hydrogenated rapeseed oil, hydrogenated safflower oil, hydrogenated palm oil, sitosterol, stigmasterol, campesterol, brassicasterol, cocoa butter powder, carnauba wax, rice wax, Japan wax, paraffin, etc.), and lecithins (e.g., egg yolk lecithin, soybean lecithin, etc.).

[0057] Examples of coloring agents include water-soluble food tar dyes (e.g., food dyes such as Food Red Nos. 2 and 3, Food Yellow Nos. 4 and 5, and Food Blue Nos. 1 and 2), water-insoluble lake dyes (e.g., aluminum salts of the above-mentioned water-soluble food tar dyes), and natural dyes (e.g., β-carotene, chlorophyll, red iron oxide, etc.).

[0058] Examples of sweeteners include sucrose, lactose, sodium saccharin, dipotassium glycyrrhizinate, aspartame, stevia, etc. Examples of antioxidants include sulfites, ascorbic acid, and alkali metal salts and alkaline earth metal salts thereof.

[0059] Tablets, granules, fine granules, etc. may be coated with a coating substrate by a conventional method for the purposes of taste masking, improving light stability, improving appearance, enteric coating, etc. Examples of such coating substrates include sugar coating substrates, water-soluble film coating substrates, and enteric film coating substrates.

[0060] Examples of sugar-coating bases include sucrose, which may be used in combination with one or more of talc, precipitated calcium carbonate, gelatin, gum arabic, pullulan, carnauba wax, and the like.

[0061] Examples of water-soluble film coating bases include cellulose polymers such as hydroxypropyl cellulose (HPC), hydroxypropylmethyl cellulose (HPMC), ethyl cellulose, hydroxyethyl cellulose, and methylhydroxyethyl cellulose; synthetic polymers such as polyvinyl acetal diethylaminoacetate, aminoalkyl methacrylate copolymer E (Eudragit E (trade name), Rohm Pharma), and polyvinylpyrrolidone; and polysaccharides such as pullulan.

[0062] Examples of enteric film coating bases include cellulose polymers such as hydroxypropyl methylcellulose phthalate, hydroxypropyl methylcellulose acetate succinate, carboxymethylethylcellulose, and cellulose acetate phthalate; acrylic acid polymers such as methacrylic acid copolymer L (Eudragit L (trade name), Rohm Pharma), methacrylic acid copolymer LD (Eudragit L-30D55 ​​(trade name), Rohm Pharma), and methacrylic acid copolymer S (Eudragit S (trade name), Rohm Pharma); and natural products such as shellac.

[0063] These coating bases may be used alone or in combination of two or more in an appropriate ratio, or two or more may be coated sequentially.

[0064] Further examples include ingredients acceptable for use as food (for example, minerals, vitamins (vitamin A; B vitamins (B1, B2, B6, B12, niacin, pantothenic acid, folic acid, biotin, etc.); vitamin C; vitamin D; vitamin E; vitamin K, etc.), flavonoids, quinones, polyphenols, amino acids, nucleic acids, essential fatty acids, cooling agents, binders, sweeteners, disintegrants, lubricants, colorants, flavorings, stabilizers, preservatives, sustained-release regulators, surfactants, solubilizers, humectants, etc.).

[0065] The content of the triterpene compound in the formulation of the present disclosure, relative to the entire formulation, can be, for example, 0.0001 to 99% by mass, 0.001 to 80% by mass, 0.01 to 70% by mass, 0.1 to 60% by mass, 0.1 to 50% by mass, 0.1 to 40% by mass, 0.1 to 30% by mass, 0.1 to 20% by mass, 1 to 10% by mass, etc. When the triterpene compound in the formulation of the present disclosure is a plant extract, the proportion of the plant extract (e.g., olive leaf extract, Swertia japonica extract, Ligustrum lucidum extract, etc.) can be, for example, 0.0001 to 99% by mass, 0.001 to 80% by mass, 0.01 to 70% by mass, 0.1 to 60% by mass, 0.1 to 50% by mass, 0.1 to 40% by mass, 0.1 to 30% by mass, 0.1 to 20% by mass, 1 to 10% by mass, 2 to 7% by mass, etc.

[0066] The method of administration of the formulation of the present disclosure is not particularly limited, and generally includes oral administration as described above. However, parenteral administration (intravascular (intravenous, intraarterial) administration, subcutaneous administration, rectal administration, etc.) may be appropriately selected depending on the condition, severity, etc. of the patient.

[0067] A preferred method of administration or ingestion of the formulation of the present disclosure is administration or ingestion at the time of, or before or after, a meal, although the method and form of administration or ingestion are not particularly limited.

[0068] The triterpene compound of the present disclosure can be used in an adult weighing 60 kg at a daily dose of 1 mg to 10 g, preferably 50 mg to 8 g, more preferably 150 mg to 3 g, and even more preferably 150 mg to 1.5 g, in terms of the amount of active ingredient. The dose can be appropriately set within the above range depending on factors such as the health condition of the person taking the compound, the administration method, and the combination with other agents.

[0069] The autophagy inducer of the present disclosure may be administered orally (ingested) or parenterally. As long as the daily amount of the active ingredient is within the above-mentioned range, the autophagy inducer of the present disclosure may be administered once a day or in multiple divided doses, such as twice or three times a day.

[0070] Because the autophagy-inducing effect is more pronounced, the autophagy inducer of the present disclosure is preferably orally administered once a day in an amount equivalent to 150 mg to 1.5 g of the active ingredient to an adult weighing 60 kg.

[0071] The target animals are preferably vertebrates such as humans, dogs, cats, parakeets, mynas, parrots, guinea pigs, rats, mice, pigs, sheep, goats, horses, cows, monkeys, rabbits, frogs, salamanders, goldfish, carp, and crucian carp, more preferably mammals, and particularly preferably humans.

[0072] The autophagy inducer of the present disclosure may be in any form, such as a solid (solid composition), liquid, or paste. The autophagy inducer of the present disclosure may be in the form of, for example, a tablet, pill, capsule (including, for example, hard capsules, soft capsules, and microcapsules), powder, granules, fine granules, lozenges, or liquid (including syrup, emulsion, or suspension), which may be sugar-coated or coated as necessary. Preferred forms are tablets, pills, capsules, powders, granules, fine granules, or lozenges, and more preferred forms are tablets or capsules.

[0073] The autophagy inducer of the present disclosure can be prepared, for example, by mixing the triterpene compound as the active ingredient with other ingredients as necessary, and molding the mixture into the above-mentioned dosage form.

[0074] The autophagy inducer of the present disclosure can be used as a pharmaceutical, quasi-drug, food or beverage composition, or cosmetic product itself, or can be added to a pharmaceutical, quasi-drug, food or beverage composition, or cosmetic product.

[0075] Here, the functions of "food" are classified into primary functions (nutritional functions that provide nutrition for life support), secondary functions (sensory functions that include taste, aroma, texture, etc. and allow for the enjoyment of food, which are taste and sensory functions: sensory functions), and tertiary functions (bioregulation functions that repair abnormalities in the physiological functions of the living body). In recent years, particularly from the perspective of disease and aging prevention, attention has been focused on the tertiary functions of "food," such as boosting immunity, preventing diseases such as high blood pressure, obesity, and diabetes, recovering from fatigue, and preventing aging by suppressing the production of active oxygen. Therefore, food and beverage compositions that emphasize the tertiary functions of "food" (physical condition regulation functions, i.e., autophagy induction functions, etc.) are preferred. Examples of food and beverage compositions that emphasize the tertiary functions of "food" include health foods, functional foods, dietary supplements, dietary supplements, and foods for specified health uses.

[0076] That is, food and beverage compositions containing the autophagy inducers of the present disclosure include autophagy-inducing foods, health foods, foods with functional claims (functional foods), nutritional supplements, dietary supplements, foods for specified health uses, etc. Food and beverage compositions include beverages and foods.

[0077] When the autophagy inducer of the present disclosure is used as a food or beverage composition itself or added to a food or beverage composition, the form of the food or beverage composition is not particularly limited, and may be, for example, beverages (coffee, juice, tea drinks, soft drinks such as jelly drinks, dairy drinks, lactic acid bacteria drinks, yogurt drinks, carbonated drinks, etc.), spreads (custard cream, etc.), pastes (fruit paste, etc.), Western confectionery (chocolate, donuts, pies, cream puffs, gum, jelly, candy, cookies, cakes, puddings, etc.), Japanese confectionery (daifuku, mochi, manju, castella, anmitsu, yokan, etc.), frozen desserts (ice cream, popsicles, sherbet, etc.), foods (curry, beef bowls, rice porridge, miso soup, soup, meat sauce, pasta, pickles, jam, etc.), or seasonings (dressings, furikake, umami seasonings, soup bases, etc.). Furthermore, the form of the food and drink composition may be, in addition to the forms of the food and drink composition described above, for example, plain tablets, sugar-coated tablets, granules, powders, tablets, capsules (hard capsules, soft capsules, seamless capsules, etc.), etc.

[0078] Pharmaceuticals, quasi-drugs, or food and beverage compositions comprising the autophagy inducers of the present disclosure, or pharmaceuticals, quasi-drugs, or food and beverage compositions containing the autophagy inducers of the present disclosure, may be used for inducing autophagy and may be labeled with a function claim such as "inducing autophagy." There are no particular limitations on where the function claim may be placed, and it may be, for example, on the product itself, container, packaging, instructions, package insert, or promotional materials.

[0079] The content of the autophagy inducer of the present disclosure in a pharmaceutical product, quasi-drug, or food or beverage composition may be appropriately determined depending on the type of pharmaceutical product, quasi-drug, or food or beverage composition, etc., so that the amount of active ingredient ingested per day falls within the above-mentioned range.

[0080] When the autophagy inducer of the present disclosure is used as a drug or quasi-drug itself, or added to a drug or quasi-drug, the form of the drug or quasi-drug is not particularly limited and may be, for example, a plain tablet, sugar-coated tablet, granule, powder, tablet, capsule (hard capsule, soft capsule, seamless capsule, etc.), etc.

[0081] The method for producing a pharmaceutical, quasi-drug, or food or beverage composition containing an autophagy inducer of the present disclosure is not particularly limited and can be any known method as appropriate. For example, the pharmaceutical, quasi-drug, or food or beverage composition used for the above-mentioned purposes can be obtained by mixing the autophagy inducer of the present disclosure with an intermediate or final product in the manufacturing process of the pharmaceutical, quasi-drug, or food or beverage composition.

[0082] The present disclosure is effective against autophagy-related diseases, such as type 2 diabetes, arteriosclerosis, infectious diseases, nephropathy, heart failure, various inflammations, Crohn's disease, metabolic syndrome, muscle atrophy / myopathy, anemia, neurodegenerative diseases, and cancer.

[0083] The present disclosure will be explained in more detail below using examples, but the technical scope of the present disclosure is not limited to these examples.

[0084] Examples 1 to 3 Commercially available oleanolic acid (Tokyo Chemical Industry Co., Ltd., purity 98%) was diluted with phosphate buffered saline (Nacalai Tesque, Inc.) to a final concentration of 22 μg / ml (Example 1), 11 μg / ml (Example 2), or 5.5 μg / ml (Example 3).

[0085] Comparative Examples 1 to 3 As a positive control, trans-resveratrol (manufactured by Tokyo Chemical Industry Co., Ltd.) (hereinafter referred to as "resveratrol") was used and diluted with phosphate buffered saline (manufactured by Nacalai Tesque, Inc.) to a final concentration of 25 μg / ml (Comparative Example 1), 12.5 μg / ml (Comparative Example 2), or 7 μg / ml (Comparative Example 3).

[0086] Reference Example 1 In Reference Example 1, dimethyl sulfoxide (DMSO) (Sigma-Aldrich) was used and diluted with phosphate buffered saline (Nacalai Tesque) to a concentration of 0.1%.

[0087] Test Example 1 <Evaluation of Autophagy Activity> According to the method of Bhargava et al. (Bhargava KH et al.: Structural basis for autophagy inhibition by the human Rubicon-Rab7 complex. Proceedings of the National Academy of Sciences of the United States of America 117: 17003-17010 (2020)), a tfLC3-stably expressing cell line was obtained from cervical epidermoid carcinoma HeLa cells (JCRB Cell Bank: JCRB9004) using a retrovirus. The obtained cells were suspended in growth medium (DMEM medium (catalog number D6546; manufactured by Sigma-Aldrich) supplemented with 10% FBS and 4 mM L-glutamine) to a cell concentration of 25,000 cells / mL, and 100 μL of the suspension was dispensed into each well of a 96-well plate (Cell Carrier 96 Ultra Black 96-well, Clear Bottom, with Lid; manufactured by PerkinElmer). This 96-well plate was incubated for 1 hour at 20°C for 1 hour at 20°C. ２ Place in an incubator at 37°C and 5% CO ２ The cells were cultured under +95% air conditions for 24 hours.

[0088] Diluted sample solutions were prepared by adding phosphate-buffered saline (PBS; manufactured by Nacalai Tesque, Inc.) as a solvent to oleanolic acid, resveratrol, or DMSO. The resulting sample solutions were added to the cell culture medium in each well to the final concentrations shown in Table 1. ２ Place in an incubator at 37°C and 5% CO ２The cells were cultured under +95% air conditions for 24 hours (n=3). In addition, as a negative control (blank), cells cultured in growth medium without the sample solution were cultured in a separate well under the same conditions as above.

[0089] After incubation, the culture medium was removed from each well, and the wells were washed twice by adding and removing phosphate-buffered saline (PBS). Subsequently, PBS containing 4% paraformaldehyde (Nacalai Tesque) containing Hoechst dye (Hoechst 33342; Dojindo Laboratories) was added to each well, and the wells were left to stand for 20 minutes to immobilize the cells.

[0090] The supernatant in each well was removed and washed twice with PBS. Cells were photographed using a confocal image cytometer (CQ1; Yokogawa Electric Corporation), and image data containing information on the fluorescence intensity of enhanced green fluorescent protein (EGFP) and monomeric red fluorescent protein (mRFP) was acquired. For EGFP fluorescence intensity measurement, the detection bandpass filter used was BP525 / 50, and the excitation wavelength was 488 nm. For mRFP fluorescence intensity measurement, the detection bandpass filter used was BP617 / 73, and the excitation wavelength was 561 nm. The ratio of EGFP fluorescence intensity to mRFP fluorescence intensity (EGFP / mRFP value) was calculated using image analysis software (CellPathfinder; Yokogawa Electric Corporation) for the acquired cell image data.

[0091] Table 1 and Figure 1 show the autophagy effect by showing the EGFP / mRFP values ​​in Examples 1 to 3 and Comparative Examples 1 to 3 as relative values ​​when the EGFP / mRFP value in Reference Example 1 is set to 1. Note that a lower EGFP / mRFP value indicates more activated autophagy.

[0092]

[0093] <Results> As shown in Table 1 and Figure 1, oleanolic acid was shown to have high autophagy activity at a lower concentration than resveratrol, which is known to have high autophagy activity. In other words, oleanolic acid showed high autophagy activity even at a lower concentration than resveratrol.

[0094] Examples 4 and 5 Olive leaf extract powder (manufactured by Fine Corporation, containing 55% oleanolic acid, ethanol extract) was used and diluted with phosphate buffered saline (manufactured by Nacalai Tesque, Inc.) to a final concentration of 10 μg / ml (Example 4) or 5 μg / ml (Example 5).

[0095] Comparative Example 4 Torin-1 (manufactured by ChemScene, CS-0237) was used as a positive control and diluted with phosphate-buffered saline (manufactured by Nacalai Tesque, Inc.) to a final concentration of 50 nM (Comparative Example 4). Torin-1 is a compound known as an autophagy activator.

[0096] Reference Examples 2 and 3 In Reference Example 2 (for evaluating Torin-1) and Reference Example 3 (for evaluating olive leaf extract powder), dimethyl sulfoxide (DMSO) (Sigma-Aldrich) was used and diluted to a concentration of 0.1% with phosphate buffered saline (Nacalai Tesque, Inc.).

[0097] Test Example 2 <Evaluation of Autophagy Activity> Diluted solutions of the samples were prepared by adding phosphate-buffered saline (PBS; manufactured by Nacalai Tesque, Inc.) as a solvent to olive leaf extract powder (solid at room temperature), Torin-1, or DMSO. Each of the obtained sample solutions was added to the cell culture medium in each well to achieve the final concentration shown in Table 2. An autophagy activity evaluation test was conducted in the same manner as in Test Example 1 above, except that the n number was set to 12.

[0098] Table 2 and Figure 2 show the autophagy effect by showing the EGFP / mRFP values ​​in Examples 4 and 5, Comparative Example 4, and Reference Examples 2 and 3 as relative values ​​when the EGFP / mRFP value in Reference Example 2 is set to 1. Note that a lower EGFP / mRFP value indicates more activated autophagy. Furthermore, Figure 2 states "GFP / RFP," which means "EGFP / mRFP."

[0099]

[0100] <Results> As shown in Table 2 and Figure 2, olive leaf extract powder containing 55% oleanolic acid at a concentration of 10 μg / ml exhibited higher autophagy activity than Torin-1, which is known to be an autophagy activator. This means that the autophagy effect can be achieved even with plants containing oleanolic acid.

[0101] <Formulation Examples> Formulation Example 1 (Capsules) 8% olive leaf extract (oleanolic acid content in olive leaf extract: 55%), 5% Ligustrum lucidum extract (oleanolic acid content in Ligustrum lucidum extract: approximately 1%), 9% resveratrol, 61% excipient, 8% lubricant, and 9% food ingredients (jobi lacryma-jobi, vitamin C, etc.) were mixed to obtain Mixture 1. Mixture 1 was encapsulated in a hard capsule according to a conventional method to produce a capsule. For example, three tablets of the above capsules are taken at a time with water or lukewarm water.

[0102] Formulation Example 2 (Tablets) 8% olive leaf extract (oleanolic acid content in olive leaf extract: 55%), 5% Ligustrum lucidum extract (oleanolic acid content in Ligustrum lucidum extract: approximately 1%), 9% resveratrol, 66% excipient (dextrin, etc.), 4% lubricant (sucrose fatty acid ester, etc.), and 8% food ingredient (vitamin, etc.) were mixed to obtain Mixture 2. Tablets were produced from Mixture 2 according to a conventional method. The above tablets are taken, for example, three tablets at a time with water or lukewarm water.

[0103] Formulation Example 3 (Granules) 8% olive leaf extract (oleanolic acid content in olive leaf extract: 55%), 5% Ligustrum lucidum extract (oleanolic acid content in Ligustrum lucidum extract: approximately 1%), 9% resveratrol, 70% excipient (dextrin, etc.), and 8% food ingredients (vitamins, etc.) were mixed to obtain Mixture 3. Granules were produced from Mixture 3 according to a conventional method. One packet of the above granules is taken with water or lukewarm water.

[0104] This disclosure claims priority from Japanese Patent Application No. 2024-139763, filed August 21, 2024. Japanese Patent Application No. 2024-139763 is incorporated herein by reference.

Claims

1. An autophagy inducer containing triterpene compounds.

2. The autophagy inducer of claim 1, wherein the triterpene compound is a five-ring triterpene compound.

3. The autophagy inducer of claim 1, wherein the triterpene compound contains an oleanane-type triterpene compound.

4. The autophagy inducer of claim 1, wherein the triterpene compound contains an oleanolic acid compound.

5. The autophagy inducer according to claim 1, wherein the triterpene compound is contained in an olive leaf extract.

6. The autophagy inducer according to claim 1, wherein the triterpene compound is contained in an olive leaf extract and a Ligustrum lucidum extract.

7. A solid composition comprising the autophagy inducer according to any one of claims 1 to 6.

8. The solid composition according to claim 7, further comprising an excipient.

9. A food or beverage composition comprising the autophagy inducer described in any one of claims 1 to 6.

10. A cosmetic comprising the autophagy inducer according to any one of claims 1 to 6.

11. A pharmaceutical comprising the autophagy inducer according to any one of claims 1 to 6.

12. Use of a solid composition containing a triterpene compound for inducing autophagy.

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