Alternative autophagy activator and autophagy activator

Abstract

This alternative autophagy activator and this autophagy activator contain at least one selected from the group consisting of dehydrocafestol (DC), dehydrokahweol (DK), cafestol (C), and kahweol (K).

Description

Alternative autophagy activators and autophagy activators

[0001] The present invention relates to alternative autophagy activators and autophagy activators. The present invention also relates to food or beverage compositions for activating alternative autophagy and food or beverage compositions for activating autophagy.

[0002] Autophagy is known as an intracellular purification and recycling system present in all eukaryotes, including yeast and plants, and has been the subject of much research in recent years. As research into the molecular mechanisms of autophagy progresses, more than 30 molecules related to autophagy have been identified. Among these molecules, Atg5, Atg7, and LC3, in particular, are considered to be essential molecules for the expression of autophagy. Furthermore, recently, the existence of a novel type of autophagy has been discovered that does not require these molecules, originates from the Golgi apparatus and endosomes, and is regulated by molecules such as Rab9 (Non-Patent Literature 1). Such autophagy is called alternative autophagy to distinguish it from conventional autophagy that is dependent on Atg5, Atg7, LC3, etc. (sometimes simply called Atg5-dependent autophagy).

[0003] In recent years, it has become clear that decreased activity of autophagy and alternative autophagy leads to a decline in cell and tissue function. Non-patent document 2 discloses that enteritis is exacerbated in mice lacking Trim31, an alternative autophagy-related protein. Patent document 1 discloses alternative autophagy inducers and anticancer agents containing benzothiophene compounds as active ingredients. Patent document 2 discloses an anti-inflammatory agent for UV-induced inflammation containing an alternative autophagy inducer as an active ingredient.

[0004] International Publication No. 2013 / 118842, International Publication No. 2020 / 091070

[0005] Nishida et al., Nature, 2009, 641, 654-658. Yoichi Nibe et al., Grant-in-Aid for Scientific Research Report, 2021, Research Project / Area Number 19K17426.

[0006] The problem that this invention aims to solve is to provide alternative autophagy and novel components that can activate autophagy.

[0007] As a result of diligent research to solve the above problems, the inventors of the present invention discovered that dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), or kweol (K) activate alternative autophagy and autophagy, and thus completed the present invention.

[0008] The present invention is as follows: [1] An alternative autophagy activator containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K). [2] The alternative autophagy activator according to [1], used for at least one of the following uses selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [3] An autophagy activator containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K). [4] An autophagy activator according to [3], used for at least one of the following uses selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [5] A food or beverage composition for activating alternative autophagy, containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K). [6] A food or beverage composition for activating autophagy, containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K).[7] A food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K), for at least one selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [8] The food or beverage composition according to [7], which exhibits at least one effect selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating at least one alternative autophagy selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K). [9] The food or beverage composition according to [7], which exhibits at least one effect selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating at least one autophagy selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K).

[10] A method for producing a coffee residue extract containing at least one of dehydrocafestol (DC) and dehydrokaweol (DK), comprising obtaining coffee residue from coffee beans roasted at 160°C or higher, adjusting the moisture content of the obtained coffee residue to 15% or less, and obtaining a solvent extract of the coffee residue with adjusted moisture content.

[11] A method for producing DC or DK, comprising: obtaining coffee residue from coffee beans roasted at 160°C or higher; adjusting the moisture content of the obtained coffee residue to 15% or less; obtaining a pressed product of the coffee residue with adjusted moisture content; obtaining a solvent extract of the obtained pressed product; and purifying dehydrocafestol (DC) or dehydrokaweol (DK) from the obtained solvent extract.

[12] A method for producing a coffee residue extract containing at least one of dehydrocafestol (DC) and dehydrokaweol (DK), comprising: obtaining coffee residue from coffee beans roasted at 160°C or higher; and obtaining a solvent extract of the obtained coffee residue, but not adjusting the moisture content of the obtained coffee residue.

[0009] The inventions described in [1] to [9] above may also be described in [A1] to [D9] below.

[0010] [A1] A method for activating alternative autophagy using at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K). [A2] The method according to [A1], which activates alternative autophagy for at least one selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, improving memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [A3] A method for activating autophagy using at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K). [A4] The method according to [A3], which activates autophagy for at least one selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, enhancing motor function, maintaining skin moisture and elasticity, improving skin spots, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [A5] A method for activating alternative autophagy using a food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K). [A6] A method for activating autophagy using a food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K).[A7] A method for providing at least one of the following, selected from the group consisting of improving bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, improving memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, using a food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K). [A8] The method according to [A7], which uses a food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K) to activate alternative autophagy, thereby providing at least one selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin spots, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [A9] The method according to [A7], which uses a food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K) to activate autophagy, thereby providing at least one selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin spots, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells.

[0011] [B1] At least one use selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) for activating alternative autophagy. [B2] The use described in [B1] for activating alternative autophagy for at least one use selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [B3] At least one use selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) for activating autophagy. [B4] Use as described in [B3] to activate autophagy for at least one use selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [B5] Use of a food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) to activate alternative autophagy. [B6] Use of a food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) to activate autophagy.[B7] Use of a food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) for at least one selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [B8] Use according to [B7] to provide at least one selected from the group consisting of improving bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating at least one alternative autophagy selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K). [B9] Use according to [B7] to bring about at least one selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating autophagy with at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K).

[0012] [C1] At least one use selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) for the manufacture of an alternative autophagy activator. [C2] The use described in [C1] for the manufacture of an alternative autophagy activator, which activates alternative autophagy for at least one use selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [C3] At least one use selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) for the manufacture of an autophagy activator. [C4] The use described in [C3] for the manufacture of an autophagy activator, which activates autophagy for at least one use selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [C5] The use described in [C3] for the manufacture of a food or beverage composition, which activates alternative autophagy. [C6] Use of at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) for the manufacture of a food or beverage composition for activating autophagy.[C7] Use of at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kwaweol (K) for the manufacture of a food or beverage composition for at least one selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [C8] Use as described in [C7] for the manufacture of a food or beverage composition, which exhibits at least one effect selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating at least one alternative autophagy selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K). [C9] The use described in [C7] for the manufacture of a food or beverage composition, which exhibits at least one effect selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating autophagy with at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K).

[0013] [D1] Dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K) used to activate alternative autophagy. [D2] Dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K) as described in [D1], used to activate alternative autophagy for at least one use selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving athletic function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [D3] Dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K) used to activate autophagy. [D4] Dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K) as described in [D3], used to activate autophagy for at least one use selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [D5] A food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K), used to activate alternative autophagy. [D6] A food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K), which is used to activate autophagy.[D7] A food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K), which is used for at least one of the following purposes selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. [D8] The food or beverage composition according to [D7], used to provide at least one selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating at least one alternative autophagy selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K). [D9] The food or beverage composition according to [D7], used to provide at least one selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating autophagy with at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K).

[0014] In the present invention, and in particular in [A1] to [D9] above, the invention may be therapeutic or non-therapeutic. That is, it may be an invention of a therapeutic or non-therapeutic method, an invention of a therapeutic or non-therapeutic use, or an invention of a therapeutic or non-therapeutic substance. Furthermore, in the present invention, "non-therapeutic invention" means an invention that does not involve medical procedures when administered to a subject or when ingested by a subject, that is, an invention that does not involve methods of surgery, treatment, or diagnosis of a subject, especially a human being. Depending on the circumstances, the subject may also include animals that are not human. More specifically, "non-therapeutic invention" means an invention that does not involve methods of surgery, treatment, or diagnosis performed on a human being by a physician or a person under the direction of a physician. Examples of "non-therapeutic inventions" include inventions aimed at promoting health or inventions aimed at cosmetic purposes. Furthermore, "non-therapeutic inventions" may also be "preventive inventions," as long as they do not fall under the category of "therapeutic inventions."

[0015] The present invention makes it possible to provide alternative autophagy and novel components that can activate autophagy.

[0016] The results of evaluating alternative autophagy activity are shown. The results of evaluating autophagy activity are shown.

[0017] The following describes embodiments of the present invention (hereinafter referred to as "these embodiments"), but the scope of the present invention is not limited to these embodiments.

[0018] 1. Method for producing a coffee residue extract containing at least one of DC and DK. One embodiment of the present invention relates to a method for producing a coffee residue extract containing at least one of DC and DK, comprising: obtaining coffee residue from coffee beans roasted at 160°C or higher (coffee residue acquisition step); adjusting the moisture content of the obtained coffee residue to 15% or less (moisture content adjustment step); and obtaining a solvent extract of the coffee residue with adjusted moisture content (solvent extract acquisition step).

[0019] The structure of the DC is shown below.

[0020]

[0021] The structure of DK is shown below.

[0022]

[0023] The structure of C is shown below.

[0024]

[0025] The structure of K is shown below.

[0026]

[0027] In this specification, coffee residue extract means a component separated by extraction from coffee residue after extraction with a solvent such as water. Coffee residue may be, for example, ground coffee beans after extraction with a solvent such as water. Coffee beans are the seeds of plants belonging to the genus Coffea. In this embodiment, the coffee residue may be derived from only one species of the genus Coffea, or from two or more species.

[0028] 1-1. Coffee Residue Acquisition Process Coffee residue is obtained from coffee beans roasted at 160°C or higher. The roasting conditions for the coffee beans are not particularly limited as long as the roasting temperature is 160°C or higher, and can be appropriately determined according to the type of coffee beans and the desired degree of roasting. For example, roasting may be done at 160°C or higher, 170°C or higher, 180°C or higher, 190°C or higher, 200°C or higher, 210°C or higher, 220°C or higher, 230°C or higher, 240°C or higher, or 250°C or higher for 6 minutes or more, 8 minutes or more, 10 minutes or more, 12 minutes or more, 14 minutes or more, 16 minutes or more, 18 minutes or more, or 20 minutes or more. However, caution is necessary as high temperature and long roasting times may cause scorching and ignition. Alternatively, commercially available dark roast coffee beans (or their grounds) may be used. The extraction solvent for obtaining the coffee residue is not particularly limited, but water is preferred. The extraction conditions for obtaining coffee residue are not particularly limited, but for example, the extraction temperature can be 100°C or lower, and may be as low as 70-100°C or even 5-40°C.

[0029] 1-2. Moisture Content Adjustment Step The coffee residue obtained in 1-1 above may be adjusted to have a moisture content of 15% or less. The lower limit of the moisture content is not particularly limited and may be near 0%, or it may be above 0%. The moisture content may be adjusted to, for example, 10% or less, 5% or less, 3% or less, 2% or less, or 1% or less. Any method or apparatus known to those skilled in the art may be used to adjust the moisture content, for example, forced air drying, dry heat drying, vacuum drying, freeze drying, etc. In this step, it is preferable to dry at a low temperature or dry at a high temperature for a short time. While not particularly limited, for example, drying may be done at 100°C or below, 90°C or below, 80°C or below, 70°C or below, 60°C or below for 180 minutes or more, 200 minutes or more, 300 minutes or more, 400 minutes or more, 500 minutes or more, 600 minutes or more, 700 minutes or more, 800 minutes or more, 900 minutes or more, 920 minutes or more, 940 minutes or more, 960 minutes or more, 980 minutes or more, or 1000 minutes or more, or drying may be done at over 100°C, 200°C or above, 300°C or above, 400°C or above, 500°C or above, 550°C or above for 200 minutes or less, 150 minutes or less, 100 minutes or less, 50 minutes or less, 40 minutes or less, 30 minutes or less, 20 minutes or less, 10 minutes or less, or 5 minutes or less.

[0030] As one embodiment of the present invention, a method for producing a coffee residue extract containing at least one of DC and DK is also provided, which includes obtaining coffee residue from coffee beans roasted at 160°C or higher, and obtaining a solvent extract of the obtained coffee residue, but does not include adjusting the moisture content of the obtained coffee residue. By a method that does not include the step of adjusting the moisture content of the obtained coffee residue, it is possible to obtain a coffee residue extract containing at least one of DC and DK at a concentration equivalent to that obtained by a method that includes drying the obtained coffee residue at a low temperature or drying it at a high temperature for a short time.

[0031] 1-3. Solvent extraction step From the coffee residue whose moisture content was adjusted in 1-2 above, a solvent extract is obtained. The said solvent extract is a coffee residue extract and contains at least one of DC and DK. For obtaining the solvent extract, any known method or apparatus may be used. As the solvent, a solvent containing an organic solvent may be used, for example, an alcohol such as methanol or ethanol may be used. The concentration of the solvent is not particularly limited, but it may be 50-100%, or may be 60-99.5%, or 70-90%.

[0032] By the production method of the present embodiment, a coffee residue extract containing at least one of DC and DK at a high concentration can be obtained. As an embodiment of the present invention, an alternative autophagy activator and an autophagy activator containing a coffee residue extract containing at least one of DC and DK at a high content are provided.

[0033] The method for producing the coffee residue extract of the present embodiment may specifically be the following method. Roast coffee beans containing Arabica beans at 160-250 °C for 8 minutes or more. After grinding the roasted coffee beans with a coffee mill, drip water at a temperature of 75-95 °C onto the ground coffee beans using a filter to obtain coffee residue. The obtained coffee residue is dried at a temperature below 100 °C for 5-20 hours until the moisture content becomes 15% or less. Add 70-95 (v / v)% ethanol to the dried coffee residue, stir at 40-60 °C for 1-6 hours, filter using filter paper, and collect the filtrate. Repeat the same operation as above for the coffee residue remaining on the filter paper 1-3 times to collect the filtrate. Combine the collected filtrates, and concentrate the combined filtrates using an evaporator until ethanol is removed to obtain a coffee residue extract containing DC and / or DK.

[0034] The mode of the method for producing the above coffee residue extract is also applicable to the method for producing the following DC or DK.

[0035] 2. Method for Producing DC or DK One embodiment of the present invention relates to a method for producing DC or DK, which includes obtaining coffee residues from coffee beans roasted at 160 °C or higher (coffee residue obtaining step), adjusting the moisture content of the obtained coffee residues to 15% or less (moisture content adjusting step), obtaining an extract of the coffee residues with the adjusted moisture content (extract obtaining step), obtaining a solvent extract of the obtained extract (solvent extract obtaining step), and purifying dehydrocafestol (DC) or dehydrocarveol (DK) from the obtained solvent extract (purifying step).

[0036] 2-1. Coffee Residue Obtaining Step This is the same as the coffee residue obtaining step of 1-1 above.

[0037] 2-2. Moisture Content Adjusting Step This is the same as the moisture content adjusting step of 1-2 above.

[0038] 2-3. Extract Obtaining Step An extract is obtained from the coffee residues with the adjusted moisture content in 2-2 above. The extract is a liquid obtained after being squeezed and removing impurities. Any method or device known to those skilled in the art may be used to obtain the extract. Oil may be obtained as the extract of the coffee residues. The squeezing may be carried out only once or may be carried out two or more times.

[0039] 2-4. Solvent Extract Obtaining Step A solvent extract is obtained from the extract obtained in 2-3 above. Any known method or device may be used to obtain the solvent extract. As the solvent, a solvent containing an organic solvent may be used. For example, hexane, acetone, or an alcohol such as methanol or ethanol may be used. The concentration of the solvent is not particularly limited and may be 50-100%, 70-100%, 70-99.5%, or 80-99.5%.

[0040] 2-5. Purification Step DC or DK is purified from the solvent extract obtained in 2-4 above. Any known method or apparatus may be used to purify DC or DK, for example, by reversed-phase chromatography, ion exchange chromatography, column chromatography, high-performance liquid chromatography, etc. The solvent, carrier, mobile phase composition, temperature conditions, etc. used for purification can be appropriately set by those skilled in the art. As the mobile phase used for chromatography, for example, a solvent such as water, acetonitrile, or alcohol may be used, and one solvent may be used, a mixed solvent of two or more solvents may be used, or elution may be performed with a mobile phase under gradient conditions. DC and DK may be eluted separately by chromatography.

[0041] The method for producing DC or DK in this embodiment may be as follows: Coffee residue with a moisture content of 15% or less is obtained by the method described above. The obtained coffee residue is heated in a roasting kettle to 80-100°C, and the heated coffee residue is pressed 1-3 times to recover the oil. The recovered oil is filtered to obtain coffee residue pressed oil. Two to six times the amount of hexane is added to the coffee residue pressed oil and mixed, and then one to three times the amount of methanol is added and mixed. After standing for a while and confirming that it has separated into two layers, the methanol layer is recovered. An equal amount of methanol to the coffee residue pressed oil is added again to the recovered methanol layer and mixed. The above operation is repeated 1-3 times to obtain a methanol layer. Distilled water in an amount of 1 / 5 to 1 / 15 of the amount of coffee residue pressed oil is added to this methanol layer and mixed, and after standing and separation, the methanol layer is recovered. The recovered methanol layer is concentrated to dryness to obtain a dry product, and the obtained dry product is subjected to reverse-phase chromatography for fractionation. Using ultrapure water, acetonitrile, and isopropanol as the mobile phase, gradient elution was performed from ultrapure water-acetonitrile (95:5) to 100% acetonitrile, followed by the passage of isopropanol. The eluted fraction was concentrated to dryness to obtain a dry product. The dry product was subjected to HPLC to isolate DC and DK, respectively. Using ultrapure water and acetonitrile as the mobile phase, ultrapure water-acetonitrile (60:40) was passed through, and the eluates from each peak were collected. The components at each peak were identified, and the eluates identified as DC and DK were used as DC and DK isolates, respectively.

[0042] 3. Alternative Autophagy Activator and Autophagy Activator One embodiment of the present invention relates to an alternative autophagy activator and an autophagy activator that contains at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K).

[0043] In this embodiment, DC, DK, C, and K may be DC, DK, C, and K isolated from coffee residue extract, respectively, or they may be commercially available.

[0044] In this embodiment, the alternative autophagy activator and the autophagy activator may each contain at least one selected from the group consisting of DC, DK, C, and K in any amount, as long as they can exert alternative autophagy activating activity and autophagy activating activity, respectively. Other active ingredients may also be included, as long as they do not impair the effect. These other active ingredients may be other components known as alternative autophagy activators or autophagy activators, or other components not known to have or possess alternative autophagy activating activity or autophagy activating activity. The alternative autophagy activator and the autophagy activator may contain only at least one selected from the group consisting of DC, DK, C, and K, or they may contain only two or more.

[0045] In this embodiment, the alternative autophagy activator and the autophagy activator each contain at least one selected from the group consisting of DC, DK, C, and K, thereby enabling the activation of alternative autophagy and autophagy. In this embodiment, the alternative autophagy activator and the autophagy activator may each contain at least one selected from the group consisting of DC, DK, C, and K, which activates alternative autophagy and autophagy by promoting the fusion of lysosomal proteins and autophagosomes.

[0046] In this embodiment, the activation of alternative autophagy may, for example, induce or promote the degradation of intracellular proteins and other molecules, organelles, or bacteria and viruses that have invaded from outside the cell, without depending on Atg5, Atg7, and / or LC3. The activation of alternative autophagy may also be induced by the fusion of lysosomal proteins and autophagosomes, without depending on Atg5, Atg7, and / or LC3. That is, the activation of alternative autophagy may be induced by the fusion of lysosomal proteins and autophagosomes, without depending on at least one molecule selected from the group consisting of Atg5, Atg7, and LC3, or it may be induced by the fusion of lysosomal proteins and autophagosomes, without depending on all of the molecules of Atg5, Atg7, and LC3. In this embodiment, autophagy activation may be induced or promoted by, for example, Atg5, Atg7, and / or LC3, to degrade intracellular proteins and other molecules, organelles, and bacteria and viruses that have invaded from outside the cell. Autophagy activation may also be induced by the fusion of lysosomal proteins and autophagosomes, depending on Atg5, Atg7, and / or LC3. That is, autophagy activation may be induced by the fusion of lysosomal proteins and autophagosomes, depending on at least one molecule selected from the group consisting of Atg5, Atg7, and LC3, or by the fusion of lysosomal proteins and autophagosomes, depending on all of Atg5, Atg7, and LC3.

[0047] In this embodiment, the membrane of the autophagosome formed by the activation of alternative autophagy may be a membrane derived from the Golgi apparatus, late endosomes, etc. In this embodiment, the membrane of the autophagosome formed by the activation of autophagy may be a membrane derived from the endoplasmic reticulum, mitochondria, etc. For differences in the molecular mechanisms of alternative autophagy and autophagy, refer to Nishida et al., Nature, 2009, 641, 654-658. and Shigeomi Shimizu, Molecules and Cells, 2018, 41(1), 50-54.

[0048] The alternative autophagy activator in this embodiment may be an alternative autophagy activator that activates autophagy.

[0049] When alternative autophagy or autophagy is activated, substances within cells are broken down, and these broken-down substances are recycled, thus facilitating cellular metabolism. In addition, it can prevent the accumulation of abnormal proteins and maintain immune function by eliminating invaders from outside the body. Therefore, by activating alternative autophagy or autophagy, functions such as maintaining homeostasis in the body and eliminating abnormal molecules are improved, thereby improving the condition of all cells and organs in the body. Consequently, it becomes possible to prevent various diseases and infections and improve their symptoms. Specifically, the activation of alternative autophagy or autophagy brings about effects such as regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving athletic function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and / or improving the quality of germ cells.

[0050] As one embodiment of the present invention, an alternative autophagy activator and an autophagy activator are provided, which contain at least one selected from the group consisting of DC, DK, C, and K, and are used for at least one of the following uses selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. Specifically, the alternative autophagy activator, autophagy activator, or food or beverage composition in this embodiment, by containing at least one of DC, DK, C, and K, can activate alternative autophagy or autophagy, and can be used for the uses listed in Table 1, respectively.

[0051]

[0052] In this embodiment, the alternative autophagy activator and the autophagy activator may be fusion promoters of lysosomal proteins and autophagosomes, enhancers of alternative autophagy or autophagy degradation, or promoters of intracellular metabolism.

[0053] The activation of alternative autophagy or autophagy may be evaluated by methods known to those skilled in the art. For example, this could include evaluation of alternative autophagy activity or autophagy activity when autophagosomes are formed, evaluation of alternative autophagy activity or autophagy activity when autophagosomes fuse with lysosomes, or evaluation of these activities using fluorescence intensity or electron microscopy. Alternatively, alternative autophagy activity or autophagy activity may be measured by measuring the gene expression levels or protein expression levels of alternative autophagy-related proteins or autophagy-related proteins, respectively.

[0054] Alternative autophagy activity can be specifically evaluated by using cells deficient in Atg5, Atg7, and / or LC3 that express lysosomal proteins to which fluorescent proteins have been attached, as described in Patent Document 1, etc. Specifically, it can be evaluated using the fluorescence intensity generated when alternative autophagy is induced and lysosomal proteins aggregate as an indicator. The generated fluorescence intensity may also be quantified and used for evaluation. Furthermore, as described in Patent Document 1, etc., if the fluorescence intensity due to lysosomal protein aggregation is greater when the alternative autophagy activator of this embodiment is administered compared to when the alternative autophagy activator of this embodiment is not administered, it can be evaluated that alternative autophagy has been activated. Furthermore, alternative autophagy can be evaluated as activated when the fluorescence intensity is increased by, for example, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 300%, 400%, or 500% or more. Preferably, alternative autophagy can be evaluated as activated when the fluorescence intensity is increased by 10% or more.

[0055] Autophagy activity can be specifically measured by methods such as observing cells with a microscope, observing cells with a fluorescence microscope or measuring them with instruments (e.g., flow cytometer, fluorescence plate reader, etc.) after expressing a fluorescently labeled autophagy-related marker, or measuring autophagy-related markers with Western blotting, etc. More specifically, autophagy activity can be detected by methods such as RT-PCR, real-time RT-PCR, and immunological assays (e.g., chemiluminescence measurement, immunohistochemistry, ELISA, Western blotting, immunoprecipitation, etc.). If the chemiluminescence intensity measured by immunological assays is greater when the autophagy activator of this embodiment is administered compared to when the autophagy activator of this embodiment is not administered, it can be evaluated that autophagy has been activated. Furthermore, autophagy can be evaluated as activated when, for example, the chemiluminescence intensity is increased by 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 300%, 400%, or 500% or more. Preferably, autophagy can be evaluated as activated when it is increased by 10% or more. An example of an autophagy marker protein is LC3. LC3 (MAP1LC3: Microtubule-associated protein 1 light chain 3) is a ubiquitin-like molecule and a mammalian homolog of the molecule encoded by Atg8 (Autophagy-related protein 8) in yeast. The cytoplasmic form of LC3 (LC3-I) is known to be converted to LC3-II by the addition of phosphatidylethanolamine (PE) upstream of autophagy signaling, and then attracted to the autophagosome membrane. LC3 is specifically localized to the autophagy structure throughout the process from isolation membrane to lysosomal degradation and is used as a marker for autophagosomes. Because LC3-II is a lipidized form, it binds to both the outer and inner membranes of autophagosomes, and its amount is known to correlate with the number of autophagosomes. Therefore, by measuring the amount of LC3-II, the amount of autophagosomes, i.e., the activation of autophagy, can be measured.Subsequently, since LC-3 is degraded after fusing with lysosomes, the amount of LC3-II degradation can also be measured as an indicator of the activation of the autophagy recycling system.

[0056] Alternative autophagy activators and autophagy activators can exert their functions, for example, by being administered directly or indirectly to organs or cells of the body.

[0057] Alternative autophagy activators and autophagy activators may be administered in any dosage form as long as they can exert their function. For example, they may be administered in the form of oral preparations such as tablets, capsules, granules, fine granules, powders, liquids, syrups, chewables, lozenges, ointments, gels, creams, injections, sublingual preparations, inhalants, or suppositories. These dosage forms can be manufactured using conventionally known additives and are not particularly limited. For example, industry-known excipients, binders, diluents, disintegrants, buffers, thickeners, stabilizers, emulsifiers, dispersants, suspending agents, and / or preservatives may be used as needed.

[0058] The dosage and frequency of administration of alternative autophagy activators and autophagy activators may be appropriately determined according to the route of administration and various circumstances such as the age, weight, and symptoms of the recipient.

[0059] The target population for alternative autophagy activators and autophagy activators is preferably mammals, more preferably humans, monkeys, cats, pigs, horses, cattle, mice, rats, guinea pigs, dogs, and rabbits, and even more preferably humans.

[0060] As an embodiment of the present invention, a food, beverage, feed, cosmetic, pharmaceutical, or quasi-drug containing the alternative autophagy activator or autophagy activator of this embodiment is provided. The food, beverage, feed, cosmetic, pharmaceutical, or quasi-drug may be in the same dosage form as the alternative autophagy activator and autophagy activator described herein.

[0061] 4. Food or Beverage Compositions As another embodiment of the present invention, the present invention provides a food or beverage composition for activating alternative autophagy, which contains at least one selected from the group consisting of DC, DK, C, and K, and a food or beverage composition for activating autophagy.

[0062] Food or beverage compositions can be in forms suitable for the intended use, and are not particularly limited, but may include oral preparations such as tablets, capsules, granules, fine granules, powders, liquids, syrups, chewables, lozenges, ointments, gels, creams, injections, sublinguals, inhalants, or suppositories. Food or beverage compositions can be in forms suitable for consumption, such as solids, liquids, granules, powders, capsules, creams, pastes, or jelly-like forms. For example, when forming capsules, they may be coated with one or more layers of film as needed. These dosage forms and forms can be manufactured using conventionally known additives, and are not particularly limited, but may include excipients, binders, diluents, disintegrants, buffers, thickeners, stabilizers, emulsifiers, dispersants, suspending agents, and / or preservatives known in the food industry as needed. The food or beverage composition is not particularly limited, but may include, for example, health foods, foods with functional claims, foods for specified health uses, nutritional functional foods, health supplements, and supplements. Examples of food or beverage compositions include foods, beverages, seasonings, and food additives. Specific examples of foods include confectionery, dairy products, bread, noodles, rice, soups, and prepared foods. Specific examples of beverages include dairy beverages, soft drinks, fruit juices, carbonated drinks, sports drinks, nutritional drinks, and alcoholic beverages.

[0063] Food or beverage compositions may contain, as needed, colorants, flavorings, sweeteners, bittering agents, acidulants, preservatives, antioxidants, pH adjusters, and / or thickening stabilizers.

[0064] Food or beverage compositions may be provided after heat sterilization, or in a sealed form in a bag or container. Furthermore, at the time of provision, at least one statement selected from the group consisting of "improves digestive health," "maintains intestinal function," "rejuvenates the intestines," "improves cognitive function," "enhances memory," "removes waste products from the brain," "maintains healthy liver function," "improves motor function," "maintains skin moisture and elasticity," "improves skin blemishes, wrinkles and / or dullness," "improves hair color, moisture and / or shine," "extends healthy life expectancy," and "improves the quality of germ cells" may be attached.

[0065] The food or beverage composition in this embodiment, by containing at least one selected from the group consisting of DC, DK, C, and K, is capable of activating alternative autophagy or autophagy, and brings about at least one effect selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. Accordingly, this embodiment also provides a food or beverage composition containing at least one selected from the group consisting of DC, DK, C, and K, for at least one selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. The food or beverage composition in this embodiment may include at least one selected from the group consisting of DC, DK, C, and K, which activates alternative autophagy or autophagy, thereby improving digestive health, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells. Furthermore, by including at least one of each extract in the food or beverage composition in this embodiment, it becomes possible to activate alternative autophagy or autophagy, and each food or beverage composition containing at least one of each extract provides the effects described in Table 1 above. Accordingly, this embodiment also provides a food or beverage composition containing at least one selected from the group consisting of DC, DK, C, and K that provides the effects described in Table 1 above.

[0066] Food or beverage compositions can exert their alternative autophagy-activating effect or autophagy-activating effect when ingested orally.

[0067] The present invention will be described in more detail below with specific examples. However, the present invention is not limited thereto.

[0068] (1) Preparation of alternative autophagy activity measurement samples

[0069] (Caffeine) Caffeine (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd., model number: 031-06792) was used.

[0070] Chlorogenic acid 327-97-9 (manufactured by Tokyo Chemical Industry Co., Ltd., model number: C0181) was used.

[0071] (Caffeic acid) Caffeic acid standard (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd., model number: 036-23521) was used.

[0072] (Instant coffee) Instant coffee made by Nestlé was dissolved in hot water and prepared.

[0073] (Coffee Bean Hot Water Extract) 400g of roasted coffee beans (after grinding), a blend of Arabica and Robusta varieties, were mixed and steamed for 10 minutes with 1200g of water heated to 80°C. Then, 4800g of water heated to 80°C over 40 minutes was added and the coffee beans were brewed by dripping to obtain a coffee bean hot water extract. The coffee bean hot water extract was then freeze-dried to obtain 94g of the extract.

[0074] (Coffee residue pressed product (pressed oil)) Arabica and Robusta beans were blended and roasted at approximately 160°C to 220°C. The residue produced when the coffee beans (after grinding) were extracted with hot water was dried in an 80°C dryer for 30 hours to obtain coffee extract residue with a moisture content of 3.8%. 10 kg of coffee extract residue was heated to 100°C in a roasting machine, and after heating, it was pressed using an oil press (H-54, Hunder oil press) to obtain oil. In this process, it was pressed three times and a total of 700 g of oil was recovered. The recovered oil was filtered using No. 2 filter paper to finally obtain 425 g of coffee oil (coffee residue pressed product).

[0075] (Acetone extract from coffee grounds) 200 g of coffee extraction grounds was mixed with 800 mL of acetone. Extraction was carried out in a 30°C constant temperature bath using a stirrer for 1 hour, and filtration was performed using a Buchner funnel lined with No. 2 filter paper. The permeate after filtration was collected and added to a round-bottom flask, and the acetone was removed by vacuum concentration to obtain a concentrated solution.

[0076] (99.5 v / v% ethanol extract from coffee grounds) 900 mL of 99.5 v / v% ethanol was added to 100 g of coffee extraction grounds. Extraction was carried out in a 30°C constant temperature bath using a stirrer for 1 hour, and filtration was performed using a Buchner funnel lined with No. 2 filter paper. The permeate after filtration was collected and added to a round-bottom flask, and ethanol was removed by vacuum concentration to obtain a concentrate.

[0077] (99.5 v / v% ethanol extract of coffee beans) 900 mL of 99.5 v / v% ethanol was added to 100 g of coffee beans. Extraction was carried out in a constant temperature bath at 30°C using a stirrer for 1 hour, and filtration was performed using a Buchner funnel lined with No. 2 filter paper. The permeate after filtration was collected and added to a round-bottom flask, and ethanol was removed by vacuum concentration to obtain a concentrate.

[0078] (Cafe Stall (C) and Car Wear (K)) For C, we used Cafe Stall (manufactured by LKT, model number: C0020). For K, we used Car Wear (manufactured by LKT, model number: K0030).

[0079] (Dehydrocafestol (DC) (Standard)) Dehydrocafestol (manufactured by LKT, model number: D1731) was used.

[0080] (Dehydrocafestol (DC) and dehydrokaweol (DK)) The residue produced when roasted coffee beans (after grinding) blended from Arabica and Robusta varieties was extracted with hot water was dried at 80°C for 16 hours to adjust the moisture content of the coffee extraction residue to 2.5%. The dried coffee extraction residue was heated to 100°C in a roasting kettle, and after heating, it was pressed using an oil press (H-54, Hunder oil press). Three pressings were performed, and a total of 700g of oil was recovered. The recovered oil was filtered using No. 2 filter paper to obtain 425g of coffee residue pressed oil. 5g of coffee residue pressed oil was added to a separatory funnel, four times the amount of hexane was added and mixed, and then twice the amount of methanol was added and mixed. After standing for a while to confirm that it had separated into two layers, the methanol layer was collected. Equal amounts of coffee residue oil and methanol were added to the recovered methanol layer and mixed. This procedure was repeated twice to obtain a methanol layer. Distilled water equal to 1 / 10 the amount of coffee residue oil was added to this methanol layer and mixed, then allowed to stand and separate before the methanol layer was recovered. The recovered methanol layer was concentrated to dryness, yielding 0.42 g of dried material. This dried material was fractionated using a medium-pressure preparative liquid chromatograph (Yamazen Corporation EPCLC-W-Prep 2XY) under the following conditions. A UNIVERSAL Premium ODS-SM 30 μm 120A, 2 L size (3 x 20 cm, 51 g) column (Yamazen Corporation) was used, and ultrapure water, acetonitrile, and isopropanol were used as mobile phases. Linear gradient elution was performed at a flow rate of 20 mL / min from ultrapure water-acetonitrile (95:5) to 100% acetonitrile over 40 minutes, followed by 12 minutes of isopropanol elution, and fractions were divided into 10 mL portions. The fractions eluted between 37 and 45 minutes were concentrated to dryness. DC and DK were isolated from the dry material using preparative HPLC (Shimadzu Prominence) under the following conditions.A SunFire 5 μm (19 x 150 mm) column (Waters) was used, and ultrapure water and acetonitrile were used as the mobile phase. Ultrapure water-acetonitrile (60:40) was passed through the column at a temperature of 30°C and a flow rate of 17 mL / min for 160 minutes. The peaks that eluted between 116 and 126 minutes (component A) and between 128 and 141 minutes (component B) were separated to obtain 5 mg and 14 mg of concentrated dry matter of components A and B, respectively. NMR and mass spectrometry were performed on the isolated components A and B. For NMR measurements, an AVANCE NEO 400MHz instrument (Brker) was used, and for mass spectrometry, a Vanquish Flex liquid chromatograph (Thermo Fisher Scientific) and a ThermoFisher Scientific OrbiTrap Exploris 240 mass spectrometer (Thermo Fisher Scientific) were used to obtain the following data. Component A was identified as dehydrokaweol (DK) and component B as dehydrocafestol (DC). The concentrated dry products of components A and B were designated as in-house purified DK and DC, respectively. Commercially available DC was also used as a standard for evaluating alternative autophagy activity and autophagy activity.

[0081] The NMR and mass spectrometry results for DK are shown below. １ H-NMR (DMSO-d6, 400MHz, corrected with solvent): δ (ppm) 7.50 (1H, d, J = 1.5Hz, H19), 6.45 (1H, brs, H18), 6.29 (1H, d, J = 10.0Hz, H2), 5.93 (1 H, d, J=10.1Hz, H1), 5.34 (1H, s, H15), 4.65 (1H, brt, -OH, H17), 3.99 (2H, s, H17), 2. 54-2.50 (2H, overlapped, H5, 13), 2.01 (1H, d, J = 10.2Hz, H14a), 1.86 (1H, m, H6a), 1 .78-1.33 (9H, overlapped, H6b, 7a, 7b, 9, 11a, 11b, 12a, 12b, 14b), 0.94 (3H, s, H20) １３C-NMR (DMSO-d6, 100MHz, corrected for solvent): δ (ppm) = 149.37 (C3), 147.22 (C16), 141.72 (C19), 138.60 (C1), 133.60 (C15), 121.43 (C4), 115.03 (C2), 109.05 (C18), 59.05 (C17), 48.62 (C8), 44.43 (C14), 43.72 (C5), 41.23 (C10), 40.36 (C13), 40.00 (C9), 37.26 (C7), 24.86 (C12), 20.62 (C6), 19.02 (C11), 15.18 (C20) ESI MS (m / z) C ２０ H ２３ O([M+H-H2O] ＋ ), theoretical value 279.1743, experimental value 279.1742; C ２０ H ２５ O ２ ([M+H]) ＋ ) Theoretical value 297.1849, experimental value 297.1847

[0082] The results of DC NMR and mass analysis are shown below. １ ¹H-NMR (DMSO-d6, 400 MHz, corrected for solvent): δ (ppm) = 7.39 (¹H, d, J = 1.4 Hz, H19), 6.29 (¹H, d, J = 1.7 Hz, H18), 5.32 (¹H, s, H15), 4.62 (¹H, brt, -OH, H17), 3.99 (²H, brs, H17), 2.57–2.55 (²H, m, H2), 2.21 (¹H, dd, J = 12.5, 1 . 9Hz, H5), 2.11 (1H, d, J=10.2Hz, H14a), 2.03 (1H, dt , J=12.8, 4.2Hz, H1a), 1.79-1.33 (10H, overlapped , H6a, 6b, 7a, 7b, 11a, 11b, 12, 14b), 1.24-1.17 (1H, m, H1b), 1.14 (1H, d, J=8.4Hz, H9), 0.79 (3H, s, H20) １３C-NMR (DMSO-d6, 100MHz, corrected with solvent): δ (ppm) = 148.00 (C3), 147.04 (C16), 140. 84 (C19), 133.97 (C15), 119.88 (C4), 108.50 (C18), 59.08 (C17), 48.24 (C8) , 44.48 (C14), 43.62 (C9), 43.47 (C5), 40.48 (C13), 38.02 (C10), 37.81 (C7) , 35.17 (C1), 25.01 (C12), 21.56 (C6), 20.20 (C2), 18.95 (C11), 12.93 (C20) ESI MS (m / z) C ２０ H ２５ O([M+H-H2O] ＋ ), theoretical value 281.1900, measured value 281.1899; C ２０ H ２７ O ２ ([M+H] ＋ Theoretical value: 299.2006, Measured value: 299.2005

[0083] (DC and DK High-Content Residue Extract) 200 g (as dry weight) of coffee extraction residue adjusted to a moisture content of 2.5% as described above was mixed with 70 v / v% ethanol to a total volume of 1 L. The mixture was stirred at 250 rpm using a Three One Motor (BL-600, manufactured by Shinto Kagaku Co., Ltd.) at 50°C for 2 hours. After 2 hours of extraction, the mixture was filtered using No. 2 filter paper, and the filtrate and extraction residue on the filter paper were collected. 70 v / v% EtOH was added to the collected extraction residue on the filter paper to a total volume of 1 L, and the mixture was extracted again at 50°C for 1 hour while stirring. After 1 hour of extraction, the mixture was filtered using No. 2 filter paper, and the filtrate was collected. The filtrates from the first and second extractions were combined and concentrated using an evaporator until EtOH was removed, and the concentrated solution was freeze-dried. 7.9 g of the extract was obtained after freeze-drying. This extract was designated as the DC and DK high-content residue extract.

[0084] (2) Evaluation of alternative autophagy activity

[0085] DMEM (containing 4.5 g / L glucose) medium (Sigma Aldrich) was used, containing 10% FBS (Sigma Aldrich), 1 mM sodium pyruvate solution (Nacalai Tesque), 1 mM HEPES buffer (Nacalai Tesque), 0.1 mM non-essential amino acid solution (Fujifilm Wako Pure Chemical Industries), 0.1 mM MEM vitamin solution (Invitrogen), 100 U / mL penicillin-100 μg / mL streptomycin (Nacalai Tesque), and 0.05 mM 2-mercaptoethanol (Bio-Rad).

[0086] Based on Patent Document 1 (International Publication No. 2013 / 118842), ATG5-KO / GFP-Lamp1 was prepared as follows: Mizushima, N. et al. Dissection of autophagosome format using Apg5-deficient mouse embryonic stem cells. J. Cell Biol. 152, 657-668 (2001). Mouse embryonic fibroblasts (MEFs) established from 14.5-day-old embryonic ATG5-deficient C57 / B6J mice, created using the method described above, were immortalized by introducing a gene expressing the SV40T antigen into them using electroporation with the Nuclear Inspector system (Amaxa). Furthermore, a retrovirus created by introducing the MSCV-Lamp1-GFP (zeocin-resistant) vector into retroviral packaging cells, plate E cells, was used to introduce a gene expressing a Lamp1-GFP fusion protein (also referred to as "GFP-Lamp1"). Cells with high GFP-Lamp1 expression were then cloned and prepared as ATG5-KO / GFP-Lamp1.

[0087] ATG5-KO / GFP-Lamp1 at 4.0x10 ４ Seeds were seeded in 30 μL in each well of a 384-well plate (Greiner) at a concentration of cell / mL, and then stored at 37°C in 5% CO2. ２The cultures were incubated in an incubator for 24 hours. Furthermore, the following were added: caffeine, chlorogenic acid, caffeic acid, C, K, DC (standard), DC, DK, DC and DK high-content residue extract, instant coffee, soybean hot water extract, pressed oil, acetone extract, 99.5 v / v% EtOH extract (residue), and 99.5 v / v% EtOH extract (soybeans). After 24 hours, 12 μL of 4% paraformaldehyde-phosphate buffer (Fujifilm Wako Pure Chemical Industries) was added to each well, and the mixture was left to stand at room temperature for 15 minutes. After washing the cultures with HBSS (Fujifilm Wako Pure Chemical Industries), 30 μL of HBSS mixed with 30 μL of Hoechst (invitrogen) diluted 2000-fold was added to each well, and the mixture was left to stand at room temperature for 15 minutes. Microscopic images were then obtained using a confocal quantitative image cytometer CQ1 (Yokogawa Electric). Based on Patent Document 1, the cumulative fluorescence intensity of GFP-Lamp1 fluorescent granules per cell was output from a microscope image, and the alternative autophagy activity was evaluated using the value obtained by dividing this value by the vehicle group as an index. The results are shown in Figure 1. Similar experiments were also conducted with the extract disclosed in Patent Document 2, but the results showed that it did not have a sufficient effect to activate alternative autophagy (results not disclosed).

[0088] The results shown in Figure 1 confirm that C, K, DC, DK, and residue extracts with high concentrations of DC and DK have the function of activating alternative autophagy.

[0089] (3) DC and DK content under each drying condition. When roasted coffee beans (after grinding) blended from Arabica and Robusta varieties were extracted with hot water, the resulting coffee residue was dried under the conditions described in Table 2 to obtain dried coffee extract residue. 75 v / v% EtOH was added to each dried coffee extract residue for extraction, and the filtrate obtained by filtration was concentrated and freeze-dried to obtain a coffee residue extract. The changes in DC and DK content in the obtained coffee residue extract were investigated. The coffee residue obtained by the same procedure as above was freeze-dried to obtain dried coffee residue that had no heat history during drying. 75 v / v% EtOH was added to this dried coffee residue for extraction, and the filtrate obtained by filtration was concentrated and freeze-dried to obtain the coffee residue extract. The DC + DK content in the coffee residue extract obtained was set as 100%, and the DC + DK content of each dried coffee extract residue dried under the conditions described in Table 2 was investigated. As a result, as shown in Table 2, under drying conditions of 80°C or below, the DC + DK content was high, at 90% or more. While the DC+DK content decreased with drying at 160°C to 210°C for 200 minutes or more, the DC+DK content remained high at over 90% with shorter drying times (1 minute, 5 minutes) at even higher temperatures (550°C). These results indicate that coffee residue extracts containing high concentrations of DC and DK can be obtained by drying at low temperatures or for short periods at high temperatures.

[0090]

[0091] (4) Evaluation of autophagy activity Mouse embryonic fibroblasts (MEFs) established from 14.5-day-old embryonic wild-type C57 / B6J mice, prepared using the method described in Mizushima, N. et al. Dissection of autophagosome formation using Apg5-deficient mouse embryonic stem cells. J. Cell Biol. 152, 657-668 (2001), were immortalized by introducing a gene expressing the SV40T antigen into them using electroporation with the Nucleofector system (Amaxa). Furthermore, a GFP-LC3 expression vector, created by inserting rat LC3 cDNA into the BglII and EcoRI sites of the pEGFP-C1 vector (Clontech), was introduced into MEF cells using FuGENE 6 reagent, thereby introducing a gene that expresses the fusion protein of LC3 and GFP (referred to as "GFP-LC3"). Cells with high GFP-LC3 expression were then cloned and prepared as WT / GFP-LC3.

[0092] WT / GFP-LC3 1.2 x 10 ５ Seeds were sown in 90 μL each of a 96-well plate (Greiner) at a concentration of cell / mL, and stored at 37°C in 5% CO2. ２The cultures were incubated in an incubator for 24 hours. Furthermore, the following were added: caffeine, chlorogenic acid, caffeic acid, C, K, DC (standard), DC, DK, DC and DK high-content residue extract, instant coffee, soybean hot water extract, pressed oil, acetone extract, 99.5 v / v% EtOH extract (residue), and 99.5 v / v% EtOH extract (soybeans). After 24 hours, 30 μL of 4% paraformaldehyde-phosphate buffer (Fujifilm Wako Pure Chemical Industries) was added to each well, and the cultures were left to stand at room temperature for 15 minutes. After washing the cultures with HBSS (Fujifilm Wako Pure Chemical Industries), 100 μL of HBSS mixed with Hoechst (invitrogen) at a 2000-fold dilution was added to each well, and the cultures were left to stand at room temperature for 1 hour. Microscopic images were then obtained using a confocal quantitative image cytometer CQ1 (Yokogawa Electric). Based on Patent Document 1, the cumulative fluorescence intensity of GFP-LC3 fluorescent granules per cell was output from a microscope image, and autophagy activity was evaluated using the value obtained by dividing this value by the vehicle group as an index. The results are shown in Figure 2.

[0093] The results shown in Figure 2 confirm that C, K, DC, DK, and residue extracts with high concentrations of DC and DK have the function of activating autophagy.

Claims

1. An alternative autophagy activator containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokarweol (DK), cafestol (C), and karweol (K).

2. An alternative autophagy activator according to claim 1, used for at least one of the following uses selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells.

3. An autophagy activator containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K).

4. The autophagy activator according to claim 3, which is used for at least one of the following uses selected from the group consisting of regulating bowel function, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells.

5. A food or beverage composition for activating alternative autophagy, comprising at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K).

6. A food or beverage composition for activating autophagy, comprising at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kahweol (K).

7. A food or beverage composition containing at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K), for at least one selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells.

8. The food or beverage composition according to claim 7, which exhibits at least one effect selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating at least one alternative autophagy selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K).

9. The food or beverage composition according to claim 7, which exhibits at least one effect selected from the group consisting of regulating bowel movements, maintaining intestinal function, rejuvenating the intestines, improving cognitive function, enhancing memory, removing waste products from the brain, maintaining healthy liver function, improving motor function, maintaining skin moisture and elasticity, improving skin blemishes, wrinkles and / or dullness, improving hair color, moisture and / or shine, extending healthy lifespan, and improving the quality of germ cells, by activating autophagy with at least one selected from the group consisting of dehydrocafestol (DC), dehydrokaweol (DK), cafestol (C), and kweol (K).

10. A method for producing a coffee residue extract containing at least one of dehydrocafestol (DC) and dehydrokaweol (DK), comprising: obtaining coffee residue from coffee beans roasted at 160°C or higher; adjusting the moisture content of the obtained coffee residue to 15% or less; and obtaining a solvent extract of the coffee residue with adjusted moisture content.

11. A method for producing DC or DK, comprising: obtaining coffee residue from coffee beans roasted at 160°C or higher; adjusting the moisture content of the obtained coffee residue to 15% or less; obtaining a pressed product from the coffee residue with adjusted moisture content; obtaining a solvent extract from the obtained pressed product; and purifying dehydrocafestol (DC) or dehydrokaweol (DK) from the obtained solvent extract.

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