KR20240117026A - Composition for improvement, treatment or prevention of muscular disorders, or improvement of muscular functions comprising melinjo leaf - Google Patents

Abstract

The present invention relates to a composition for preventing and treating muscle disease, or improving muscle function, containing melinjo leaves, such as melinjo leaf powder or extract, as an active ingredient. Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract according to the present invention, reduce the mRNA expression of MuRF-1 and atrogin-1, which are involved in muscle protein degradation, and increase the activity of mTOR, which is involved in muscle protein synthesis. By increasing muscle function and muscle mass, it reduces sarcopenia, muscular atrophy, muscular dystrophy, atony, muscle degeneration, myasthenia, and cachexia. It can be useful in preventing, improving, or treating various muscle diseases.

Description

Composition for improvement, treatment or prevention of muscular disorders, or improvement of muscular functions comprising melinjo leaf}

The present invention relates to a composition for improving, treating or preventing muscle disease, or for improving muscle function, and more specifically, to a composition for improving muscle disease, such as melinjo leaf powder or melinjo leaf extract. It relates to a composition for improving, treating or preventing muscle disease, or improving muscle function, containing leaves as an active ingredient.

Muscle atrophy is caused by a gradual decrease in muscle mass and refers to muscle weakness and degeneration (Cell, 119(7): 907-910, 2004). Muscle atrophy is promoted by inactivity, oxidative stress, and chronic inflammation and weakens muscle function and exercise capacity (Clinical Nutrition, 26(5): 524-534, 2007). The most important factor determining muscle function is muscle mass, which is maintained by the balance of protein synthesis and breakdown. Muscular dystrophy occurs when protein degradation occurs more than synthesis (The International Journal of Biochemistry and Cell Biology, 37(10): 1985-1996, 2005).

Muscle size is regulated by intracellular signaling pathways that induce anabolism or catabolism within the muscle. If more signaling reactions that induce synthesis rather than decomposition of muscle protein occur, muscle protein synthesis increases, which is manifested as an increase in muscle size (hypertrophy) or an increase in the number of muscle fibers (hyperplasia) due to an increase in muscle protein (The Korea Journal of Sports Science, 20(3): 1551-1561, 2011).

Factors involved in muscle protein synthesis induce protein synthesis by phosphorylating downstream proteins starting from stimulation of the phosphatidylinositol-3 kinase (PI3K)/Akt pathway within muscle cells. Activation of mammalian target of rapamycin (mTOR) by PI3K/Akt signaling is recognized as a central growth signaling factor that integrates various growth signals within cells. mTOR induces muscle protein synthesis by activating two factors that initiate mRNA translation, 4E-binding protein (4EBP1) and phosphorylated 70-kDa ribosomal S6 kinase (p70S6K), thereby contributing to increased muscle mass (The Korea Journal of Sports Science, 20(3): 1551-1561, 2011; The International Journal of Biochemistry and Cell Biology, 43(9): 1267-1276, 2011). On the other hand, when the transcription factor forhead box (FoxO) moves from the cytoplasm into the nucleus, it increases the expression of atrogin-1 and MuRF1, E3 ubiquitin ligase factors involved in protein degradation (Disease Models and Mechanisms, 6: 25- 39, 2013). As their expression level increases, protein breakdown within the muscle is promoted, resulting in a decrease in muscle mass. Therefore, promoting the activity of mTOR and suppressing the expression of atrogin-1 and MuRF1 increases the amount of muscle protein and thus increases muscle mass.

Differentiation of muscle cells and muscle formation are regulated by various muscle regulatory factors. Among them, induction of myogenin expression by MyoD activity is the most important factor in the fusion of myogenic cells and is involved in the formation of myotube cells. Muscle fibers formed through this process form bundles and ultimately form muscles (Cellular and Molecular Life Sciences, 70: 4117-4130, 2013).

Melinjo is a plant belonging to the Gnetaceae family and its scientific name is Gnetum gnemon Linn. Melinjo is mainly produced in Thailand, Malaysia, Indonesia, and Fiji, and its leaves and seeds are mainly used as food. So far, various physiological activities such as antioxidant, anti-inflammatory, antibacterial, anticancer, antihypertensive, and antidiabetic have been reported in relation to Melinjo leaves (Journal of Plant Biochemistry and Biotechnology 20(2): 234-24, 2011; International Journal of Pharmacognosy and Phytochemical Research 7(3): 531-539, 2015; Food Sci. Campinas, 42, e100121, 2022).

However, prior to the present invention, nothing was known about the prevention and treatment of muscle disease or improvement of muscle function of Melinjo leaves.

Accordingly, the present inventors searched for natural substances that have excellent muscle function regulating activity and can be safely applied, and as a result, Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, were used to improve, treat, or prevent muscle diseases, or to improve muscle function. The present invention was completed by confirming that there was improvement activity.

Therefore, the object of the present invention is to provide a food composition for improving or preventing muscle disease or improving muscle function containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, as an active ingredient.

Another object of the present invention is to provide a pharmaceutical composition for treating or preventing muscle disease containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, as an active ingredient.

Another object of the present invention is to provide a cosmetic composition for improving or preventing muscle disease, or improving muscle function, containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, as an active ingredient.

Another object of the present invention is to provide a feed additive for improving or preventing muscle disease or improving muscle function containing Melinjo leaves as an active ingredient, such as Melinjo leaf powder or Melinjo leaf extract. will be.

To achieve the above object, the present invention provides a food composition for improving or preventing muscle disease, or improving muscle function, containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, as an active ingredient.

In addition, the present invention provides a pharmaceutical composition for treating or preventing muscle disease containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, as an active ingredient.

In addition, the present invention provides a cosmetic composition for improving or preventing muscle disease, or improving muscle function, containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, as an active ingredient.

In addition, the present invention provides a feed additive for improving or preventing muscle disease, or improving muscle function, containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, as an active ingredient.

Additionally, the present invention provides a method of treating muscle disease by administering the composition to humans or non-human animals.

In addition, the present invention provides new uses of Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, for manufacturing medicines for treating muscle diseases.

Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract according to the present invention, inhibit the mRNA expression of MuRF1 and atrogin-1, which are involved in muscle protein degradation, and increase the activity of mTOR, which is involved in muscle protein synthesis. It has excellent effects. Additionally, by increasing muscle function and muscle mass, it is possible to prevent, treat, or improve muscle function decline and muscle loss caused by various diseases. Since the present invention is a natural product, it can be used safely without side effects and can be usefully used in the production of food, medicine, cosmetics, or feed additives.

Figure 1 shows the results confirming the effect of reducing the mRNA expression levels of MuRF1 and atrogin-1 in C2C12 muscle cells following treatment with the Melinjo leaf hot water extract of Example 1.
Figure 2 shows the results confirming the effect of increasing mTOR activity in C2C12 muscle cells following treatment with the Melinjo leaf hot water extract of Example 1. Each graph represents the average value (means±SEM, n=3). ** means there is a significant difference at p<0.01 compared to the control group (Tukey's test).

Hereinafter, the configuration of the present invention will be described in detail.

The present invention provides a food composition for improving or preventing muscle disease of Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, or for improving muscle function; Pharmaceutical compositions for treating or preventing muscle diseases containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract; Or a cosmetic composition for improving or preventing muscle disease of Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, or for improving muscle function; or feed additives for improving or preventing muscle diseases of Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, or for improving muscle function; or a method of treating muscle disease comprising applying Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract, to a human or non-human mammal.

In this specification, 'Melinjo leaf' can be used as Melinjo leaf powder or Melinjo leaf extract. In the present invention, the Melinzo leaves can be purchased and used commercially, or those collected or cultivated in nature can be used.

In the present specification, 'Pulverized Melinjo leaves' refers to dried Melinjo leaves that can be easily ingested by mammals, including humans, according to a method that can be easily performed by a person skilled in the art to which the present invention pertains. It can be prepared and used in a form in which the active ingredient is easily released from the dry powder in the intestine after ingestion and can be easily absorbed into the mammalian body. The shape or size of the dried pulverized particles for this purpose is not limited, but the more the surface of the particles is maximized, the easier it is to release the active ingredients described above, so it is preferable to prepare them in the form of fine powder as much as possible. In one embodiment of the present invention, dried Melinjo leaves were pulverized with a mixer to prepare a pulverized product.

In this specification, 'Melinjo leaf extract' is obtained by extracting dried Melinjo leaves using a suitable solvent, and is an extract, a diluted or concentrated solution of the extract, a dried product obtained by drying the extract, or a crude or purified product thereof. Includes all shapes.

Specifically, the Melinjo leaf extract may be obtained by extracting Melinjo leaves with one or more solvents selected from water, an organic solvent having 1 to 6 carbon atoms, a subcritical fluid, and a supercritical fluid. The organic solvent having 1 to 6 carbon atoms is alcohol, acetone, ether, benzene, chloroform, ethyl acetate, and methylene chloride. It is preferable to use at least one selected from chloride, hexane, cyclohexane, and petroleum ether, but is not limited thereto. More preferred solvents for extracting the Melinjo leaf extract of the present invention are water, ethanol, ethyl acetate, hexane, subcritical fluid, or supercritical fluid.

As a specific example of the present invention, the extract may be prepared by mixing the Melinjo leaves with an extraction solvent at a weight ratio of 1:2 to 20, preferably 1:5 to 15, more preferably 1:8 to 12. there is. If the weight ratio of the Melinjo leaves and the extraction solvent is outside the above range, the active ingredient of the Melinjo leaves may be extracted in a small amount in the extract.

As another specific example, the Melinjo leaf extract of the present invention may be obtained by extracting Melinzoa leaves under ultra-high pressure conditions of 100 MPa or more. If necessary, it can be prepared by additionally including filtration and concentration steps according to methods known in the art.

The extraction may be performed by a heating extraction method, a cold needle extraction method, a reflux cooling extraction method, an ultrasonic extraction method, an ultra-high pressure extraction method, a subcritical fluid extraction method, or a supercritical fluid extraction method, and there is no limitation as long as it is an extraction method that is obvious to those skilled in the art. The extraction may be performed at room temperature, but for more efficient extraction, it may be performed under heated conditions, and extraction may be performed at a temperature of 20 to 100°C, preferably about 30 to 100°C, but is not limited thereto. The extraction time may be 1 to 10 hours, preferably 2 to 8 hours, and more preferably 3 to 5 hours, but is not limited thereto and may vary depending on conditions such as extraction solvent and extraction temperature. The extraction may be repeated one or more times to obtain a larger amount of the active ingredient, preferably 1 to 5 times, and more preferably 2 to 3 times.

The Melinjo leaf extract may be a crude extract of Melinjo leaves, or may be a solvent-soluble fraction obtained by refractionating the crude extract with a second solvent. The extract extracted by the above method or the soluble fraction of the extract can be used as is, but it can be used in the form of extract by filtering and concentrating, and after concentration, it can be prepared in powder form by additional processes such as reduced pressure distillation, freeze drying, or spray drying. You can use it.

In this specification, 'fraction' refers to a result obtained by a fractionation method that separates a specific component or specific group from a mixture containing various components. In the present invention, it refers to the result obtained by a fractionation method for separating a specific component or specific group from the Melinjo leaf extract prepared as described above.

In order to obtain the Melinjo leaf fraction according to the present invention, conventional fractionation solvents known in the art, such as water, polar solvents such as anhydrous or hydrous lower alcohols with 1 to 4 carbon atoms such as ethanol and methanol, and hexane and butanol , non-polar solvents such as ethyl acetate, chloroform, dichloromethane, or mixed solvents thereof may be used, but are not limited thereto.

The Melinjo leaf fraction of the present invention may also include one obtained by additionally applying a purification process. For example, the fraction obtained by passing the Melinjo leaf extract according to the present invention through an ultrafiltration membrane with a certain molecular weight cut-off value, subjected to various chromatography (size, charge, hydrophobicity or affinity) Fractions obtained through various purification methods additionally performed, such as separation by (produced for separation according to the present invention), are also included in the Melinjo leaf extract of the present invention.

In addition, fermentation products or extracts of fermentation products obtained by adding microorganisms (bacteria, yeast, mold, etc.) applicable to the human body to Melinjo leaf pulverization or Melinjo leaf extract can also be included in the Melinjo leaf extract.

In this specification, 'muscle' comprehensively refers to tendons, muscles, and tendons, and 'muscle function' refers to the ability to exert force through muscle contraction, and allows the muscle to exert maximum contractile force to overcome resistance. These include muscular strength, which is the ability to exert strength, muscular endurance, which is the ability to express how long or how many times a muscle can repeat contraction and relaxation with a given weight, and quickness, which is the ability to exert strong force in a short period of time. The term 'improvement of muscle function' in this specification refers to improving muscle function better by increasing muscle mass.

In a specific embodiment of the present invention, the present inventors extract Melinjo leaf extract using hot water, ethanol, ethyl acetate, or hexane as a solvent, or use ultra-high pressure extraction, subcritical fluid extraction, or supercritical fluid extraction. was prepared (Examples 1 to 7).

As a result of treating C2C12 muscle cells with the Melinjo leaf extract prepared as above, it was confirmed that the mRNA expression of MuRF-1 and atrogin-1, which are involved in muscle protein degradation, was significantly suppressed (Figure 1, Table 1) . In addition, as a result of treating C2C12 muscle cells with Melinjo leaf extract, it was confirmed that the activity of mTOR, which is involved in muscle protein synthesis, was significantly increased (Figure 2, Table 2). As a result of treating Melinjo leaf powder or Melinjo leaf extract using an immobilization muscle atrophy animal model using a skin stapler, muscle strength and muscle weight each significantly increased compared to the muscle atrophy group (Table 3).

When the composition of the present invention is a food composition for improving and preventing muscle disease or improving muscle function, it can be used to prevent or improve muscle disease caused by muscle wasting or degeneration. Muscle wasting and degeneration occur due to genetic factors, acquired factors, aging, etc. Muscle wasting is characterized by gradual loss of muscle mass and weakness and degeneration of muscles, especially skeletal or voluntary muscles and cardiac muscles. Examples of diseases related to this include sarcopenia, muscular atrophy, muscular dystrophy, atony, muscle degeneration, myasthenia, cachexia, etc. You can. The composition of the present invention has the effect of increasing muscle mass, and the type of muscle is not limited.

In this specification, 'prevention' means everything that suppresses or delays the muscle disease.

The term 'improvement' of the present invention means at least reducing the degree of symptoms, for example, parameters related to the condition being treated by administration of a composition comprising Melinjo leaf powder or Melinjo leaf extract of the present invention.

The food composition of the present invention includes all forms of functional food, nutritional supplements, health food, food additives, feed, etc., and includes all forms of food such as functional food, nutritional supplement, health food, food additives, feed, etc., and can be used for human or animals including livestock. It is intended for consumption. Food compositions of this type can be prepared in various forms according to conventional methods known in the art.

For example, as a health food, the pulverized Melinjo leaf or the Melinjo leaf extract of the present invention can be prepared and consumed in the form of tea, juice, and drink, or granulated, encapsulated, and powdered.

In addition, functional foods include beverages (including alcoholic beverages), fruits and their processed foods (e.g. canned fruit, bottled foods, jam, mamalade, etc.), fish, meat and their processed foods (e.g. ham, sausages, corned beef, etc.) etc.), bread and noodles (e.g. udon, buckwheat noodles, ramen, spagate, macaroni, etc.), fruit juice, various drinks, cookies, taffy, dairy products (e.g. butter, cheese, etc.), edible vegetable oil, margarine, vegetable oil It can be produced by adding the pulverized Melinjo leaf or extract of Melinjo leaf of the present invention to proteins, retort foods, frozen foods, various seasonings (e.g., soybean paste, soy sauce, sauce, etc.).

Additionally, in order to use the crushed Melinjo leaf or extract of Melinjo leaf of the present invention in the form of a food additive, it can be prepared and used in the form of a powder or concentrate.

Melinjo leaf powder or Melinjo leaf extract may be contained as an active ingredient in a food composition for improving and preventing muscle disease or improving muscle function, and the amount is sufficient to achieve the effect of improving and preventing muscle disease or improving muscle function. The effective amount is not particularly limited, but is preferably 0.01 to 100% by weight based on the total weight of the entire composition. The food composition of the present invention can be prepared by mixing Melinjo leaf powder or Melinjo leaf extract with other active ingredients known to be effective in improving and preventing muscle disease or improving muscle function.

Additionally, when the composition of the present invention is a pharmaceutical composition for treating or preventing muscle disease, it can be used to prevent or treat muscle disease caused by muscle wasting or degeneration. Muscle wasting and degeneration occur due to genetic factors, acquired factors, aging, etc. Muscle wasting is characterized by gradual loss of muscle mass and weakness and degeneration of muscles, especially skeletal or voluntary muscles and cardiac muscles. Examples of diseases related to this include sarcopenia, muscular dystrophy, muscular dystrophy, tension dystrophy, muscle degeneration, myasthenia gravis, and cachexia. The composition of the present invention has the effect of increasing muscle mass, and the type of muscle is not limited.

As used herein, 'prevention' means anything that suppresses or delays the muscle disease.

As used herein, ‘treatment’ means reversing, alleviating, inhibiting the progression of, or preventing the symptoms of the muscle disease.

Melinjo leaf powder or Melinjo leaf extract according to the present invention can be used as such or in a pharmaceutically acceptable form for the purpose of treating or preventing muscle diseases. In the above, 'pharmaceutically acceptable' means a non-toxic composition that is physiologically acceptable and does not inhibit the action of the active ingredient when administered to humans and does not usually cause gastrointestinal disorders, allergic reactions such as dizziness, or similar reactions. says

The pharmaceutical composition of the present invention may contain a pharmaceutically effective amount of Melinjo leaf powder or Melinzoa leaf extract alone or may include one or more pharmaceutically acceptable carriers. In the above, 'pharmaceutically effective amount' refers to an amount that shows a greater response than the negative control, and preferably refers to an amount sufficient to treat or prevent muscle disease.

The pharmaceutical composition of the present invention may contain 0.01 to 99.99% by weight of Melinjo leaf powder or Melinjo leaf extract, and the remaining amount may be accounted for by a pharmaceutically acceptable carrier.

The carrier includes all types of solvents, dispersion media, oil-in-water or water-in-oil emulsions, aqueous compositions, liposomes, microbeads and microsomes. As pharmaceutically acceptable carriers, those described in the following literature may be referred to (Remington's Pharmaceutical Sciences, 19th ed., Mack Publishing Company, Easton, PA, 1995).

Meanwhile, the route of administration of the pharmaceutical composition according to the present invention is not limited thereto, but may be administered orally or parenterally. Parenteral routes of administration include several routes, such as transdermal, intranasal, intraperitoneal, intramuscular, subcutaneous, or intravenous.

Additionally, the pharmaceutical composition of the present invention can be administered in combination with known compounds known to be effective in treating or preventing muscle diseases.

Additionally, the composition for improving and preventing muscle disease or improving muscle function of the present invention may be a cosmetic composition. The cosmetic composition of the present invention contains Melinjo leaf powder or Melinjo leaf extract as an active ingredient, and is used as a basic cosmetic composition (face wash such as lotion, cream, essence, cleansing foam, and cleansing water) along with dermatologically acceptable excipients. It can be manufactured in the form of color cosmetic compositions (foundation, lipstick, mascara, makeup base), hair product compositions (shampoo, rinse, hair conditioner, hair gel), soap, etc.

The excipients are not limited thereto, but may include, for example, skin emollients, skin penetration enhancers, colorants, fragrances, emulsifiers, thickeners, and solvents. In addition, it may additionally contain fragrances, pigments, disinfectants, antioxidants, preservatives, moisturizers, etc., and may include thickeners, inorganic salts, synthetic polymer materials, etc. for the purpose of improving physical properties.

The content of Melinjo leaf powder and Melinjo leaf extract contained in the cosmetic composition of the present invention is not limited thereto, but is preferably 0.001 to 10% by weight, more preferably 0.01 to 5% by weight, based on the total weight of the entire composition. . If the content is less than 0.001% by weight, the desired effect cannot be expected, and if the content is more than 10% by weight, there may be difficulties in safety or formulation manufacturing. The cosmetic composition of the present invention can be prepared by mixing Melinjo leaf powder or Melinjo leaf extract with other active ingredients known to be effective in improving and preventing muscle disease or improving muscle function.

Additionally, when the composition of the present invention is a feed additive for improving and preventing muscle disease or improving muscle function, it can be used to prevent or improve muscle disease caused by muscle wasting or degeneration in animals.

In this specification, 'feed additive' includes substances added to feed for various effects such as nutrient supplementation and weight loss prevention, improvement of digestibility of fiber in feed, improvement of oil quality, prevention of reproductive disorders and improvement of conception rate, and prevention of high temperature stress in summer. do. The feed additive of the present invention corresponds to supplementary feed under the Feed Management Act, and includes mineral preparations such as sodium bicarbonate, bentonite, magnesium oxide, and complex minerals, mineral preparations such as trace minerals such as zinc, copper, cobalt, and selenium, and kerotene. , vitamins such as vitamins A, D, E, nicotinic acid, and vitamin B complex, protective amino acids such as methionine and lysine, protective fatty acids such as fatty acid calcium salts, probiotics (lactic acid bacteria), yeast cultures, and live bacteria such as mold fermentation products, Yeast agents, etc. may be additionally included.

In this specification, 'feed' refers to any natural or artificial diet, meal, etc., or a component of the meal, for eating, ingestion, and digestion by animals, or a composition for treating or preventing muscle disease according to the present invention. Feed containing as an active ingredient can be manufactured from various types of feed known in the art, and may preferably include concentrated feed, roughage and/or special feed, but is not limited thereto.

The feed additive for improving and preventing muscle disease or improving muscle function according to the present invention can be prepared by adding Melinjo leaf powder or Melinjo leaf extract in an appropriate effective concentration range according to various feed manufacturing methods known in the art. You can.

In addition, the present invention provides a method of treating muscle disease comprising administering to a mammal a pharmaceutical composition containing an effective amount of Melinjo leaf powder or Melinzoa leaf extract as an active ingredient.

As used herein, the term 'mammal' refers to a mammal that is the subject of treatment, observation or experiment, and preferably refers to a human.

As used herein, the term 'effective amount' refers to the amount of an active ingredient or pharmaceutical composition that, as believed by a researcher, veterinarian, physician, or other clinician, induces a biological or medical response in a tissue system, animal, or human. Includes an amount that induces relief of symptoms of a disease or disorder. The effective amount and frequency of administration of the active ingredient of the present invention may vary depending on the desired effect.

The pharmaceutically effective amount of Melinjo leaf powder or Melinjo leaf extract according to the present invention is 0.001 to 100 mg/day/kg of body weight, preferably 0.01 to 10 mg/day/kg of body weight. However, the pharmaceutically effective amount may be adjusted according to various factors such as the disease and its severity, the patient's age, weight, health status, gender, administration route, and treatment period.

Hereinafter, the crushed Melinjo leaf or the Melinjo leaf extract according to the present invention will be described in detail through examples and test examples.

<Example>

Example 1: Preparation of Melinjo leaf hot water extract

The dried Melinjo leaves were pulverized with a mixer, and then 10 g of the crushed Melinjo leaf sample was added to 100 mL of water and extracted at 80°C for 3 hours. The extracted sample was filtered under reduced pressure with Whatman No. 1 filter paper, and the filtered extract was concentrated in a vacuum rotary concentrator to remove the solvent, thereby obtaining a melinjo leaf hot water extract.

Example 2: Preparation of Melinjo leaf ethanol extract

The dried Melinjo leaves were pulverized with a mixer, and then 10 g of the crushed Melinjo leaf sample was added to 100 mL of 100% ethanol and extracted at 40°C for 3 hours. The extracted sample was filtered under reduced pressure with Whatman No. 1 filter paper, the filtered extract was concentrated in a vacuum rotary concentrator, and the solvent was removed to obtain an ethanol extract of Melinjo leaf.

Example 3: Preparation of Melinjo leaf ethyl acetate extract

The dried Melinjo leaves were pulverized with a mixer, and then 10 g of the crushed Melinjo leaf sample was added to 100 mL of ethyl acetate and extracted at 40°C for 3 hours. The extracted sample was filtered under reduced pressure using Whatman No. 1 filter paper, and the filtered extract was concentrated using a vacuum rotary concentrator to remove the solvent, thereby obtaining an ethyl acetate extract from Melinjo leaf.

Example 4: Preparation of Melinjo leaf hexane extract

The dried Melinjo leaves were pulverized with a mixer, and then 10 g of the crushed Melinjo leaf sample was added to 100 mL of hexane and extracted at 40°C for 3 hours. The extracted sample was filtered under reduced pressure using Whatman No. 1 filter paper, and the filtered extract was concentrated using a vacuum rotary concentrator to remove the solvent, thereby obtaining a hexane extract of Melinjo leaf.

Example 5: Preparation of Melinjo leaf ultra-high pressure extract

After pulverizing the dried Melinjo leaves with a mixer, place 1g of the crushed Melinzoa leaves and 76 mL of 18% (v/v) ethanol in a polyethylene pack, seal it, and use an ultra-high pressure extraction device (Frescal MFP-7000; Mitsubishi Heavy). Industries) was used to extract the extract. Ultra-high pressure extraction conditions were extraction pressure of 320 MPa and extraction time of 5 min. The extracted sample was filtered through Whatman No. 2 filter paper, and the filtered extract was concentrated in a vacuum rotary concentrator to remove the solvent, thereby obtaining an ultra-high pressure extract of Melinjo leaf.

Example 6: Preparation of Melinjo leaf subcritical extract

The dried Melinjo leaves were pulverized with a mixer, and then 50 g of the pulverized Melinjo leaves were placed together with 1 L of water in the subcritical water reactor of a subcritical extraction device (Biovan, Gyeonggi, Korea) and sealed. After sealing, the temperature of the reactor was raised to 200°C, and when the temperature of the reactor reached 200°C, heating was stopped and the temperature was maintained for 20 minutes to perform extraction. After 20 minutes, the extract was transferred to a storage tank supplied with cooling water and rapidly cooled to 30°C. Then, it was centrifuged at 3,600 rpm for 30 minutes to separate the floating residue, and only the supernatant was collected. A subcritical extract of Melinjo leaves was obtained by removing all solvents using a freeze dryer (Ilshin Lab Co. Ltd., Seoul, Korea).

Example 7: Preparation of Melinjo leaf supercritical extract

The dried Melinjo leaves were pulverized with a mixer, and then 10 g of the crushed Melinjo leaves were charged into a sample cartridge and extracted using a supercritical fluid extraction device (SFX 3560, Isco Inc., Lincoln, NE, USA). The supercritical extraction conditions were extraction pressure of 300 bar, extraction temperature of 50°C, supercritical carbon dioxide flow rate of 60 mL/min, and extraction time of 2 hours. When supercritical fluid extraction was completed, the pressure of the extraction device was lowered to release the supercritical fluid state to obtain a Melinjo leaf supercritical extract.

<Test example>

Test Example 1: Effect of suppressing the expression level of muscle protein degradation biomarkers

Muscle cells, C2C12 myoblast (ATCC; Manassas, VA, USA) were cultured in a 6-well plate with Dulbecco's modified Eagle's media (DMEM; Hyclone) containing 10% fetal bovine serum (FBS; Hyclone, Logan, UT, USA). It was added to make 5×10 ⁵ cell/mL. When the cell density reached about 80-85%, the medium in the well was removed and the cells were treated with DMEM (Hyclone) containing 2% horse serum (HS; Hyclone) to induce myotube differentiation. Differentiation was carried out for a total of 4 days, with new medium replaced every 2 days. After differentiation, 10 μg/mL and 20 μg/mL of the Melinjo leaf hot water extract of Example 1 were added to DMEM (Hyclone) containing 10 μM of dexamethasone (Sigma-Aldrich, St. Louis, MO, USA). After melting, it was treated with cells. After 24 hours, total RNA was isolated using TRIzol reagent (Takara, Otsu, Japan). The isolated total RNA was quantified using NanoDrop 1000 (Thermo Fisher Scientific Inc., Waltham, MA, USA). 16 μL of quantified RNA was incubated at 42°C for 55 minutes and at 70°C for 15 minutes using Reverse Transcriptase Premix (ELPIS-Biotech) and a PCR machine (Gene Amp PCR System 2700; Applied Biosystems, Foster City, CA, USA). cDNA was synthesized under these conditions. Next, 1 μL of cDNA among the generated 16 μL of cDNA was amplified by PCR with PCR premix (ELPIS-Biotech) and the following specific primers (Bioneer, Daejeon, Korea). Amplification was performed with a total of 35 cycles of pretreatment at 95°C for 5 minutes, denaturation at 95°C for 30 seconds, annealing at 58°C for 15 seconds, and extension at 72°C for 45 seconds as 1 cycle, and 5 cycles at 72°C. A final extension was performed for 1 minute.

MuRF1

Forward primer: 5'-CCGGACGGAAATGCTATGGA-3' (SEQ ID NO: 1)

Reverse primer: 5'-AGCCTGGAAGATGTCGTTGG-3' (SEQ ID NO: 2)

Atrogin-1

Forward primer: 5'-GTTACTGCAACAAGGAGAATCTGTT-3' (SEQ ID NO: 3)

Reverse primer: 5'-CCGTATGAGTCTTATGTTTTGCTGG-3' (SEQ ID NO: 4)

β-Actin:

Forward primer: 5'-TGACAGGATGCAGAAGGAGATTAC-3' (SEQ ID NO: 5)

Reverse primer: 5’-TAAAACGCAGCTCAGTAACAGTC-3’ (SEQ ID NO: 6)

The PCR product was separated by electrophoresis using a 1.5% agarose gel, and the band was confirmed using the G;BOX EF imaging system (Syngene, Cambridge, UK). At this time, β-actin was used as an internal standard.

As a result, as shown in Figure 1, the mRNA expression of MuRF1 and atrogin-1, which are involved in muscle protein degradation, increased by dexamethasone (DEX), while when the hot water extract of Melinjo leaves of Example 1 was treated, MuRF1 and atrogin-1 It can be seen that the mRNA expression of 1 has decreased. This means that the Melinjo leaf hot water extract of the present invention has an excellent ability to inhibit the expression of biomarkers involved in muscle protein degradation in muscle cells.

Test Example 2: Inhibitory effect of muscle protein degradation of Melinjo leaf extract

The test was repeated a total of three times in the same manner as Test Example 1, except that C2C12 myogenic cells were treated with 20 μg/mL of Melinjo leaf extract according to Examples 2 to 7, respectively. Next, the band density of MuRF1 and atrogin-1 mRNA was measured using the Image J program (National Institute of Health, Bethesda, MD, USA), and then each measured value was calculated as a percentage (%) relative to the control group and averaged ± Expressed as standard deviation.

As a result, as shown in Table 1, the mRNA expression levels of MuRF1 and atrogin-1 increased significantly (##p < 0.01) compared to the control group as treated with dexamethasone, but 20 μg of the Melinjo leaf extract according to the example It can be seen that in the case of /mL treatment, the mRNA expression levels of both MuRF and Atrogin-1 were significantly (**p < 0.01) decreased compared to the dexamethasone treatment group. This means that the Melinjo leaf extract of the present invention has an excellent ability to inhibit muscle protein degradation within muscle cells.

division MuRF-1 expression level
(% compared to control group) Atrogin-1 expression level
(% compared to control group) control group 100±36.1 100±41.8 dexamethasone 598.4±42.1 ^## 392.2±32.8 ^## Example 2 216.9±34.6 ^** 140.8±44.3 ^** Example 3 255.0±33.9 ^** 151.3±39.0 ^** Example 4 201.5±44.1 ^** 134.1±28.1 ^** Example 5 239.7±42.6 ^** 152.0±40.8 ^** Example 6 260.7±32.1 ^** 160.3±28.1 ^** Example 7 209.5±36.0 ^** 129.9±45.0 ^**

Test Example 3: Effect of increasing muscle protein synthesis biomarker mTOR activity

It is known that when mTOR protein is phosphorylated and activated, it can induce the activation of proteins involved in muscle protein synthesis and muscle mass increase in the PI3K/Akt signaling pathway within muscle cells. Therefore, in order to confirm the muscle creation-inducing activity of the Melinjo leaf extract, the activity of mTOR was confirmed using the mTOR Sandwich ELISA kit (Cell Signaling Technology, Beverly, MA, USA).

Specifically, C2C12 myoblasts (ATCC; Manassas, VA, USA) were cultured in 6-well plates with Dulbecco's modified Eagle's media (DMEM; Hyclone) containing 10% fetal bovine serum (FBS; Hyclone, Logan, UT, USA). After seeding to reach 5X10 ⁵ cells/well, the cells were cultured for 24 hours. After culture, the medium in the well was removed, replaced with DMEM (Hyclone) containing 2% horse serum (HS; Hyclone), and cultured for an additional 4 days to differentiate C2C12 cells into myotube cells. Next, the Melinjo leaf hot water extract of Example 1 was dissolved in DMEM (Hyclone) at a concentration of 10 μg/mL and 20 μg/mL, then treated with cells and cultured for 12 hours. After culturing, cells were lysed by treatment with cell lysis buffer. The protein in the obtained cell lysate was quantified at a concentration of 1 mg/mL using Bradford (Bio-Rad Laboratories Inc., Hercules, CA, USA). 50 μL of cell lysate was dispensed into each microwell with anti-mTOR antibody attached and left at 37°C for 2 hours. After washing a total of 4 times with washing buffer, treatment with detection antibody and leaving at 37°C for 1 hour, washing again with washing buffer a total of 4 times, horseradish peroxidase ) conjugated secondary antibody was added and left at 37°C for 30 minutes. Finally, after washing a total of four times with the washing buffer solution, TMB substrate was added to each well, left at 37°C for 10 minutes, and stop buffer solution was added to stop the TMB reaction. After 2 minutes, the absorbance was measured at a wavelength of 450 nm to measure the level of mTOR in root canal cells treated with Melinjo leaf extract. The experiment was repeated a total of three times, and each measured value was calculated as a percentage (%) of the control group and expressed as mean ± standard deviation. Differences between groups were confirmed through Duncan multi-range analysis using one-way analysis of variance using the SPSS25.0 statistical package (SPSS Inc., Chicago, IL, USA). At this time, if the p value was less than 5%, it was indicated as statistically significant.

As a result, as shown in Figure 2, it can be seen that when C2C12 muscle cells were treated with the Melinjo leaf hot water extract of Example 1, mTOR activity was significantly (*p < 0.01) increased compared to the control group. This means that the Melinjo leaf hot water extract of the present invention has an excellent ability to promote muscle protein synthesis in muscle cells.

Test Example 4: Effect of Melinjo leaf extract on promoting muscle protein synthesis

mTOR activity was measured in the same manner as in Test Example 3, except that C2C12 myogenic cells were treated with 20 μg/mL of Melinjo leaf extract according to Examples 2 to 7, respectively.

As a result, as shown in Table 2, it was confirmed that the activity of mTOR was significantly (**p < 0.01) increased as compared to the control group that was not treated with the extract as the Melinjo leaf extract was treated according to the example of the present invention. You can. This means that the Melinjo leaf extract of the present invention has an excellent ability to promote muscle protein synthesis in muscle cells.

division mTOR activity
(% compared to control group) control group 100±3.8 Example 2 139.6±4.6 ^** Example 3 136.0±3.8 ^** Example 4 140.1±4.1 ^** Example 5 139.9±3.7 ^** Example 6 138.1±3.4 ^** Example 7 144.8±4.7 ^**

Test Example 5: Effect of Melinzoa leaf pulverization and extract on muscle function and muscle mass increase in immobilization animal model

As experimental animals, 7-week-old male mice (C57BL/6J; DBL, Korea) were purchased and tested. The breeding room environment for all animals was maintained at a temperature of 23 ± 2°C and a relative humidity of 55 ± 10%. Before starting the experiment, a total of 25 rats were randomly divided into 5 rats per group. After a week of adaptation, anesthesia was induced by intraperitoneal injection of 325 mg/kg tribromoethanol (Sigma-Aldrich). After anesthesia, the gastrocnemius muscle of the right hindlimb and the right sole of the rats in the muscle atrophy group and the sample administration group were stapled using a skin stapler (Unidus, Chungcheongbuk-do, Korea). The right hind leg was damaged and unable to move, and this condition was maintained for a week. One week later, the staple core fixed to the gastrocnemius muscle and sole was removed, and for another week, 500 mg/kg of the above-mentioned Melinjo leaf pulverization, 300 mg/kg of Melinjo leaf hot water extract of Example 1-1, Example 1-2 300 mg/kg of Melinjo leaf ethane extract was orally administered daily for 7 days. At this time, the normal group and the muscle atrophy group were orally administered saline solution instead of the sample.

After the oral administration period, the muscle strength of the rats was measured using a muscle strength meter (Panlab, Barcelona, Spain). The rat's tail was pulled with a constant force until the rat let go of the last part of the muscle strength meter, and a total of 5 consecutive tests were performed per animal.

After muscle strength measurements were completed, the experimental animals were anesthetized by intraperitoneal injection of 325 mg/kg of tribromomethanol (Sigma-Aldrich), and then sacrificed through cardiac blood sampling. After confirming that the heartbeat had stopped, the undamaged tibialis anterior muscle was removed from the right hind leg and its weight was measured.

As a result, as shown in Table 3, muscle strength ( ^# p < 0.05) and muscle weight ( ^## p < 0.01) were significantly decreased in the muscle atrophy group compared to the normal group, but in the group treated with crushed Melinjo leaf and extract. It was confirmed that strength and muscle weight increased significantly ( ^** p < 0.01) compared to the muscle atrophy group. This means that the Melinjo leaf powder and extract of the present invention are excellent in increasing muscle strength and muscle weight decreased due to muscle atrophy.

experimental group Muscle strength (g) Muscle weight (mg) Jeongsang-gun 245.0±13.1 58.4±5.1 Muscular atrophy group 162.5±12.7 ^## 42.7±3.8 ^## Melinjo leaf powder 188.8±10.9** 52.7±4.6** Melinzoa leaf thermal water extract 212.3±15.2** 53.2±5.1** Melinjo Leaf Ethanol Extract 226.1±13.0** 54.9±5.3**

Hereinafter, an example of manufacturing food, medicine or cosmetics containing Melinjo leaf powder and extract as an active ingredient according to the present invention will be described, but the present invention is not intended to be limited, but merely explained in detail. The food, pharmaceutical or cosmetic compositions of Preparation Examples 1 to 3 are conventionally prepared using Melinjo leaf powder or Melinjo leaf extract, which is excellent for preventing and treating muscle diseases or improving muscle function, according to the composition and composition ratio as follows. It was prepared according to the method.

<Production Example 1> Food

Production Example 1-1. health food

1000 mg of crushed Melinjo leaf or extract of Melinjo leaf of the present invention, 70 ug of vitamin A acetate, 1.0 mg of vitamin E, 0.13 mg of vitamin B1, 0.15 mg of vitamin B2, 0.5 mg of vitamin B6, 0.2 ug of vitamin B12, 10 ug of vitamin C mg, biotin 10 ug, nicotinic acid 1.7 mg, folic acid 50 ug, calcium pantothenate 0.5 mg, ferrous sulfate 1.75 mg, zinc oxide 0.82 mg, magnesium carbonate 25.3 mg, potassium phosphate monobasic 15 mg, calcium phosphate dibasic 55 mg. , can be prepared by mixing 90 mg of potassium citrate, 100 mg of calcium carbonate, and 24.8 mg of magnesium chloride, and the mixing ratio may be modified arbitrarily. After mixing the above ingredients according to a normal health food manufacturing method, Granules can be prepared and used to manufacture health food compositions according to conventional methods.

Production Example 1-2. health drink

Add purified water to 1000 mg of Melinzoa leaf extract of the present invention, 1000 mg of citric acid, 100 g of oligosaccharides, 2 g of plum concentrate, and 1 g of taurine to make a total of 900 mL. Mix the above ingredients according to a typical health drink manufacturing method, then add approx. After stirring and heating at 85°C for 1 hour, the resulting solution is filtered, placed in a sterilized 2 L container, sealed, sterilized, stored in the refrigerator, and then used to manufacture a health drink composition.

Production Example 1-3. chewing gum

Chewing gum was prepared in a conventional manner by mixing 20% by weight of gum base, 76.9% by weight of sugar, 1% by weight of flavor, 2% by weight of water, and 0.1% by weight of Melinjo leaf extract of the present invention.

Production Example 1-4. candy

Candy was prepared in a conventional manner by mixing 60% by weight of sugar, 39.8% by weight of starch syrup, 0.1% by weight of flavoring, and 0.1% by weight of the Melinjo leaf extract of the present invention.

Production Example 1-5. biscuit

25.59% by weight of strength grade 1, 22.22% by weight of gravity grade 1, 4.80% by weight of refined sugar, 0.73% by weight of table salt, 0.78% by weight of glucose, 11.78% by weight of palm shortening, 1.54% by weight of ammonium, 0.17% by weight of sodium bicarbonate, 0.16% by weight of sodium bisulfite. , 1.45% by weight of rice flour, 0.0001% by weight of vitamin B, 0.04% by weight of milk flavor, 20.6998% by weight, 1.16% by weight of whole milk powder, 0.29% by weight of powdered milk, 0.03% by weight of monobasic calcium phosphate, 0.29% by weight of spray salt and spray. Biscuits were prepared in a conventional manner by mixing 7.27% by weight of oil and 0.8301% by weight of Melinjo leaf powder or Melinjo leaf extract of the present invention.

<Manufacturing Example 2> Medicine

Manufacturing Example 2-1. Sanje

A powder was prepared by mixing 50 mg of the Melinjo leaf extract of the present invention and 2 g of crystalline cellulose and filling it into an airtight envelope according to a conventional powder production method.

Production Example 2-2. refine

Tablets were prepared by mixing 50 mg of Melinzoa leaf extract of the present invention, 400 mg of crystalline cellulose, and 5 mg of magnesium stearate and then tableting according to a conventional tablet manufacturing method.

Production Example 2-3. capsule

Capsules were prepared by mixing 30 mg of Melinzoa leaf extract of the present invention, 100 mg of whey protein, 400 mg of crystalline cellulose, and 6 mg of magnesium stearate and filling them into gelatin capsules according to a conventional capsule preparation method.

<Production Example 3> Cosmetics

Manufacturing Example 3-1. Nutritional lotion (milk lotion)

Nutritional lotion was prepared according to a conventional method using the Melinjo leaf extract of the present invention according to the nutritional lotion formulation ratio in Table 4 below.

Ingredients Preparation Example 3-1 (% by weight) Melinjo Leaf Extract 2.0 squalane 5.0 beeswax 4.0 Polysorbate 60 1.5 Sorbitan sesquioleate 1.5 liquid paraffin 0.5 Caprylic/Capric Triglyceride 5.0 glycerin 3.0 Butylene glycol 3.0 propylene glycol 3.0 Carboxy vinyl polymer 0.1 Triethanolamine 0.2 Preservatives, coloring, flavoring Appropriate amount Purified water to 100

Production Example 3-2. Soft lotion (skin lotion)

Soft lotion was prepared according to a conventional method using the Melinjo leaf extract of the present invention according to the softening lotion formulation ratio in Table 5 below.

Ingredients Preparation Example 3-2 (% by weight) Melinjo Leaf Extract 2.0 glycerin 3.0 Butylene glycol 2.0 propylene glycol 2.0 Carboxy vinyl polymer 0.1 PEG 12 nonylphenyl ether 0.2 Polysorbate 80 0.4 ethanol 10.0 Triethanolamine 0.1 Preservatives, coloring, flavoring Appropriate amount Purified water to 100

Manufacturing Example 3-3. Nutrition cream

A nutritional cream was prepared according to a conventional method using the Melinjo leaf extract of the present invention according to the nutritional cream formulation ratio in Table 6 below.

Ingredients Preparation Example 3-3 (% by weight) Melinjo Leaf Extract 2.0 Polysorbate 60 1.5 Sorbitan sesquioleate 0.5 PEG60 hydrogenated castor oil 2.0 liquid paraffin 10 squalane 5.0 Caprylic/Capric Triglyceride 5.0 glycerin 5.0 Butylene glycol 3.0 propylene glycol 3.0 Triethanolamine 0.2 antiseptic Appropriate amount pigment Appropriate amount Spices Appropriate amount Purified water to 100

Production Example 3-4. massage cream

Massage cream was prepared according to a conventional method by using the pulverized Melinjo leaf extract or Melinjo leaf extract of the present invention according to the massage cream formulation ratio in Table 7 below.

Ingredients Preparation Example 3-4 (% by weight) Melinjo leaf powder or Melinjo leaf extract 1.0 beeswax 10.0 Polysorbate 60 1.5 PEG 60 hydrogenated castor oil 2.0 Sorbitan sesquioleate 0.8 liquid paraffin 40.0 squalane 5.0 Caprylic/Capric Triglyceride 4.0 glycerin 5.0 Butylene glycol 3.0 propylene glycol 3.0 Triethanolamine 0.2 Preservatives, coloring, flavoring Appropriate amount Purified water to 100

Production Example 3-5. pack

A pack was prepared according to a conventional method using the pulverized Melinjo leaf or extract of Melinjo leaf of the present invention according to the pack formulation ratio in Table 8 below.

Ingredients Preparation Example 3-5 (% by weight) Melinjo leaf powder or Melinjo leaf extract 1.0 polyvinyl alcohol 13.0 Sodium Carboxymethylcellulose 0.2 glycerin 5.0 allantoin 0.1 ethanol 6.0 PEG 12 nonylphenyl ether 0.3 Polysorbate 60 0.3 Preservatives, coloring, flavoring Appropriate amount Purified water to 100

Production Example 3-6. gel

A gel was prepared according to a conventional method using the Melinjo leaf extract of the present invention according to the gel formulation ratio in Table 9 below.

Ingredients Preparation Example 3-6 (% by weight) Melinjo Leaf Extract 0.5 Sodium ethylenediamine acetate 0.05 glycerin 5.0 Carboxy vinyl polymer 0.3 ethanol 5.0 PEG 60 hydrogenated castor oil 0.5 Triethanolamine 0.3 Preservatives, coloring, flavoring Appropriate amount Purified water to 100

In the table above, 'appropriate amount' means that an appropriate amount can be added depending on preference, and 'to 100' means that the remaining weight% after subtracting the sum of the weight% of the remaining mixing ingredients excluding purified water from the total 100% by weight is added to the purified water. This means that it can be done in weight percent.

Although the present invention has been described in terms of the above-mentioned preferred embodiments, various modifications and variations can be made without departing from the gist and scope of the invention. Furthermore, the appended claims cover such modifications or variations as fall within the subject matter of the present invention.

As described above, the present invention provides a composition for improving, treating or preventing muscle disease, or improving muscle function, containing Melinjo leaves, such as Melinjo leaf powder or Melinjo leaf extract. More specifically, the Melinjo leaves of the present invention reduce the mRNA expression levels of MuRF1 and atrogin-1 and increase the activity of mTOR to improve muscle mass and strength, thereby improving, treating or preventing muscle diseases, or improving muscle function. It shows excellent effects. Therefore, the present invention has high industrial applicability because it can provide a composition that shows excellent effects for improving, treating or preventing muscle diseases, or improving muscle function.

Claims (8)

A food composition for improving or preventing muscle disease or improving muscle function containing Melinjo leaf as an active ingredient. According to clause 1,
The Melinjo leaves are the pulverized product of Melinjo leaves,
The Melinjo leaf extract is a food composition for improving or preventing muscle disease or improving muscle function, characterized in that it is obtained by extracting Melinjo leaves. According to clause 1,
The Melinjo leaf extract is obtained by extracting Melinjo leaves with one or more solvents selected from the group consisting of water, organic solvents having 1 to 6 carbon atoms, subcritical fluids, and supercritical fluids. Improving or preventing muscle disease. For, or food composition for improving muscle function. According to claim 3,
The organic solvent includes alcohol having 1 to 6 carbon atoms, acetone, ether, benzene, chloroform, ethyl acetate, methylene chloride, and hexane ( A food composition for improving or preventing muscle disease, or improving muscle function, characterized in that it is one or more solvents selected from the group consisting of hexane, cyclohexane, and petroleum ether. The method according to any one of claims 1 to 4,
The muscle disease is selected from the group consisting of sarcopenia, muscular atrophy, muscular dystrophy, atony, muscle degeneration, myasthenia, and cachexia. A food composition for improving or preventing muscle disease, or improving muscle function, characterized by one or more of the following: A pharmaceutical composition for treating or preventing muscle disease containing melinjo leaves as an active ingredient. A cosmetic composition for improving or preventing muscle disease or improving muscle function containing Melinjo leaves as an active ingredient. A feed additive for improving or preventing muscle disease or improving muscle function containing melinjo leaves as an active ingredient.