KR102504364B1 - Leucine derivatives, composition comprising the same, and uses thereof - Google Patents

Abstract

The present invention relates to a cosmetic composition, a pharmaceutical composition, a quasi-drug composition, and a food composition having excellent skin improvement effect, fatigue improvement effect, and/or female menopausal symptom relief effect. the glycyl-L-leucine peptide, the L-leucylglycine peptide, or both of the present invention; Or a composition containing a salt thereof as an active ingredient shows excellent effects in improving skin elasticity, moisturizing skin, preventing skin aging, improving fatigue, anti-inflammatory, antioxidant and/or alleviating female menopausal symptoms. the glycyl-L-leucine peptide, the L-leucylglycine peptide, or both of the present invention; Or a composition containing a salt thereof may be used as a cosmetic raw material, pharmaceutical ingredient, quasi-drug raw material, or functional food raw material that is safe for the skin and has excellent target effects, and may be included as a marker component or an active ingredient.

Description

Leucine derivatives, compositions comprising the same, and uses thereof {Leucine derivatives, composition comprising the same, and uses thereof}

The present invention relates to a leucine derivative, a composition containing the same, and a use thereof, and specifically, a novel cosmetic use, pharmaceutical use, food use, and a cosmetic composition containing the leucine derivative, a pharmaceutical composition, and a quasi-drug composition of the leucine derivative. and food compositions.

Aging is a phenomenon of life, and all living things start aging from birth. Improvements in hygiene and health environment have significantly extended human lifespan compared to the past, and the proportion of the elderly population is steadily increasing (Kang Tae-jin, BioWave 2011, Vol.13, No.3 (1)). Aging occurs throughout the body, and physiological functions of organs deteriorate, such as overall body function decline and lifestyle-related diseases such as cardiovascular disease and diabetes (Choi Hyun-cheol, Journal of the Korean Society of Dermatology, 1997; 35: 292-302).

The skin is the outermost part of the body, preventing loss of water, electrolytes, and proteins in the body, and is an important organ that simultaneously performs various roles such as temperature regulation, skin sensory function, and immune function.

Skin, like other organs, ages as we age. Skin aging consists of intrinsic skin aging and photoaging, and intrinsic skin aging is characterized by functional changes rather than macroscopic morphological changes. Photoaging due to chronic UV exposure overlaps with endogenous aging and results in marked structural and physiological changes, although not systemic or unavoidable. Aged epidermis not only loses its own functions such as wrinkles, loss of skin barrier, and immune abnormalities, but also aesthetically affects the appearance of the body. Although aging of the skin cannot be prevented, various attempts have been made to prevent and delay aging of the skin.

Collagen is a major matrix protein produced by fibroblasts of the skin and is present in the extracellular matrix. Its important functions include mechanical robustness of the skin, resistance and cohesiveness of connective tissue, support of cell adhesion, cell division and differentiation. (in the growth of organisms or wound healing), etc. are known. Such collagen is reduced by age and photoaging caused by ultraviolet irradiation, and collagen reduction is promoted by the activity of collagenase enzyme that decomposes collagen. It is known that it is closely related to the formation of wrinkles in the skin.

In addition, it forms elastin and is an important skin component for wrinkle formation involved in skin elasticity. Elastin fibers lack cross-linking with collagen and aggregate, and the marked increase in the activity of elastase, an elastin-degrading enzyme, has been found to be one of the factors for skin wrinkle formation. Elastase is the only enzyme capable of decomposing elastin, and inhibition of it is known to fundamentally reduce skin wrinkle improvement.

On the other hand, collagen and elastin fibers are matrix proteins that play an important role in retaining moisture in the dermal layer, and these matrix proteins adsorb moisture and increase the moisture retention within the structure they form so that the skin can maintain an appropriate moisture state. It is known to be involved in maintaining skin elasticity.

In addition, elastic fibers generated from fibroblasts of the skin form cross-linked bonds with collagen and are an important skin component for wrinkle formation involved in skin elasticity. Deficiency and decomposition of elastic fibers have been found to be one of the main causes of wrinkle formation and deterioration of skin elasticity.

Currently, retinoids, adenosine, animal placenta-derived proteins, chlorella extracts, and the like are known as wrinkle-improving cosmetics. The most well-known retinol is a substance that promotes collagen synthesis and inhibits the elastase enzyme, but it is unstable, and its use is limited due to safety issues such as irritation and redness when applied to the skin. It is known that it is difficult to expect wrinkle improvement effects.

Therefore, it is safe for the body, the active ingredient is safe for the skin, has high stability, and above all, improves skin elasticity, moisturizes, and prevents skin aging. development is desperately needed.

On the other hand, fatigue is one of the most common diseases of modern society and is a state in which physical ability is reduced (Uthayathas et al., 2007). Fatigue can be caused by excessive stress (Uthayathas et al., 2007) or abnormalities in the immune function and antioxidant system (Clear, 2003). As fatigue-related indicators, there are blood biomarkers such as blood urea nitrogen (BUN), alanine aminotransaminase (ALT), lactate dehydrogenase (LDH), and glucose (Klimas et al., 2012). LDH and creatine kinase (CK) values are generally known as markers associated with muscle damage (Coombes and McNaughton, 2000). Malic acid dehydrogenase (MDH) has also been implicated in the oxidative metabolism of fatigue (Callister et al., 2004). In addition, fatigue is induced through the occurrence of immune abnormalities in the oxidative stress pathway and various inflammatory pathways (Liu et al., 2000). Active oxygen generated after excessive exercise further aggravates fatigue (Powers et al., 1999). To reduce these active oxygen, superoxide dismutase (SOD) and catalase are produced (Liu et al., 2000). Nitric oxide (NO) is increased in patients with chronic fatigue (Suarez et al., 2010), and inflammatory cytokines are also related to fatigue symptoms (Liburt et al., 2010).

Currently, many studies are being conducted to treat fatigue, but a suitable method is still unknown.

The causes of fatigue are classified into deficiency or inability to use energy sources stored in the body, inflammatory reactions in the body due to metabolic actions, abnormalities in the antioxidant system, accumulation of fatigue substances, and loss of homeostasis in the body. It is known to be made by In order to recover from such fatigue, supply of sufficient energy sources, rest, suppression and removal of fatigue substances in the body, etc. are required. need is emerging.

On the other hand, menopause in women is a phenomenon of cessation of menstruation that occurs when the genetically pre-determined ovarian function of about 50 years after birth reaches the end of life. . Currently, the average life expectancy of Korean women is 81.2 years (2011: Statistics Korea), and assuming that the average age of menopause for Korean women defined by the Korean Society of Obstetrics and Gynecology is 50 years old, about 1/3 of a woman's life is lived in a state of estrogen depletion. (Pharmaceutical information center, Seongcheol Kim)

As women enter menopause, changes occur throughout the body, such as the vascular system, musculoskeletal system, genitourinary system, and cranial nerves, due to the imbalance and decrease in secretion of female hormones. In other words, vasomotor symptoms and psychological symptoms such as hot flushes, night sweats, sleep disorders, fatigue, depression, anxiety, difficulty concentrating, memory disorders and dyspareunia due to urogenital atrophy, frequent urination, loss of skin elasticity due to collagen reduction, and breast enlargement. It is accompanied by various diseases such as sagging, cardiovascular and musculoskeletal symptoms, and dementia. Although menopausal symptoms vary from person to person, it has been reported that the more menopausal symptoms are experienced, the more severe, and the longer the period, the lower the quality of life of women. It is likely to proceed

Hormone therapy, drug therapy, exercise therapy, and diet therapy can be applied to treat menopausal symptoms, but medically used female hormone therapy can increase the risk of breast cancer, etc., and long-term use can lead to uterine cancer, thrombotic vascular disease, and gallbladder disease , may increase the rate of hypertension.

Accordingly, studies on treatments that are safe for the human body and can achieve excellent therapeutic effects are continuously being conducted.

Therefore, the problem to be solved by the present invention is to solve the above problems, to provide a new active ingredient that is safe to the human body with few side effects and has excellent skin elasticity improvement, moisturizing or skin aging prevention effects, that is, such a useful use of the active ingredient. is to do

In addition, it is to provide a new active ingredient that is safe for the human body with few side effects and has excellent fatigue, inflammation, or antioxidant effects, that is, such a useful use of the active ingredient.

It is also to provide a novel active ingredient with excellent menopausal symptom improvement effect, that is, such a useful use of the active ingredient.

In other words, the problem to be solved by the present invention is to provide a composition containing an active ingredient with excellent efficacy, and furthermore, a composition containing the active ingredient with excellent efficacy as an active ingredient or indicator ingredient.

In order to solve the above problems, the present invention glycyl -L- leucine (Glycyl-L-Leucine) peptide, L- leucyl glycine (L-Leucylglycine) peptide or both; Or a composition for improving skin elasticity, moisturizing skin or preventing skin aging, comprising a salt thereof as an active ingredient, preferably a cosmetic composition, pharmaceutical composition, quasi-drug composition or food composition (preferably a functional food composition).

That is, the present invention relates to glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both; or a salt thereof as a cosmetic composition, pharmaceutical composition, quasi-drug composition, or food composition, and new uses such as improving skin elasticity, moisturizing skin, or preventing skin aging are provided.

The present invention glycyl -L- leucine (Glycyl-L-Leucine) peptide, L- leucyl glycine (L-Leucylglycine) peptide or both; Or it provides a method for improving skin elasticity, moisturizing the skin, or preventing skin aging by treating the skin with a salt thereof.

The glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide, or both of them may be active ingredients or indicator ingredients.

As a result of research efforts to develop a peptide effective for skin improvement or a natural material containing the same, the present inventors found that a composition containing glycyl-L-leucine peptide and/or L-leucyl glycine as an active ingredient improves skin elasticity and improves skin elasticity. It was confirmed that there is an effect on moisturizing and preventing skin aging, and the present invention was completed.

We find glycyl-L-leucine and/or L-leucylglycine; Or, it was experimentally confirmed that a composition containing a salt thereof as an active ingredient exhibits a skin improving effect by promoting hyaluronic acid synthesis, promoting procollagen synthesis, and inhibiting elastase.

In the present invention, 'improving skin elasticity' means alleviating the degree of sagging or sagging of the skin. In addition, the elasticity means maintaining the elasticity of the skin in a state in which elastin and collagen are sufficiently present.

In the present invention, 'skin moisturizing' means to increase the feeling of moisture in the skin and to maintain a moist state. The skin moisturizing effect can also help improve wrinkles and increase elasticity of the skin.

In the present invention, 'prevention of skin aging' means preventing and alleviating the occurrence of wrinkles and loss of elasticity in the skin. For example, skin aging can be inhibited by inhibiting the activity of elastase.

In another embodiment of the present invention, the present invention is glycyl -L- leucine (Glycyl-L-Leucine) peptide, L- leucyl glycine (L-Leucylglycine) peptide or both; Or a fatigue improvement, inflammation improvement or antioxidant composition comprising a salt thereof as an active ingredient, preferably a health functional food composition, a cosmetic composition, a pharmaceutical composition, or a quasi-drug composition is provided.

That is, the present invention relates to glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both; Or a new use of its salt as a health functional food composition, cosmetic composition, pharmaceutical composition, or quasi-drug composition and fatigue improvement, inflammation improvement or antioxidant use.

The present invention glycyl -L- leucine (Glycyl-L-Leucine) peptide, L- leucyl glycine (L-Leucylglycine) peptide or both; Or it provides a method for improving fatigue, improving inflammation or preventing oxidation by treating a salt thereof.

The glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide, or both of them may be active ingredients or indicator ingredients.

The inventors of the present invention glycyl -L- leucine (Glycyl-L-Leucine) peptide, L- leucyl glycine (L-Leucylglycine) peptide or both; Or, it was experimentally confirmed that a salt thereof prevented accumulation of lactic acid and creatine kinase (CK) in muscle in a mouse model in which fatigue was induced by exercise, and it was found that there was an effect of improving fatigue, and the present invention was completed. In addition, the glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both; Or its salt was experimentally confirmed to reduce the secretion of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in spleen cells, confirming the effect of improving inflammation, and superoxide dismutase in blood (superoxide dismutase, SOD) levels were experimentally confirmed to show antioxidant effects.

As used herein, the term 'fatigue improvement' means to relieve accumulated fatigue or remove fatigue symptoms, and the fatigue is a temporary decrease in the strength or sensitivity of cells, muscles, or organs due to physical or mental activities. means state. If you continue mental and physical activities in a state where you have not recovered from fatigue, the fatigue will overlap and lead to overwork or chronic fatigue, which can lead to movement disorders as well as health disorders. Such excessive physical activity causes fatigue. The physiological mechanism of fatigue is not limited to this, but it is because energy sources such as glucose and glycogen are consumed by physical activity. It accelerates the consumption of muscle glycogen and is relatively small at low intensity, but consumption increases over time even at low intensity exercise, which causes fatigue. Another cause of fatigue is the accumulation of intermediate products of metabolism such as lactic acid and carbon dioxide in the body. Carbohydrates used as an energy source for the body are broken down into glucose in the body, and the remaining excess is stored in the liver and muscles in the form of glycogen and used as an energy source when needed. As it decomposes, it generates a large amount of energy to sustain exercise, but when oxygen supply is insufficient, lactic acid is produced and accumulated. When lactic acid accumulates, the chemical reaction does not work well, causing fatigue. In the present invention, it was confirmed experimentally that the accumulation of lactic acid and creatine kinase (CK) in muscles was prevented in a mouse model in which fatigue was induced by exercise, and the fatigue improvement effect of the peptide was confirmed.

As used herein, the term 'improvement of inflammation' means not only suppressing inflammation but also removing and/or alleviating the inflammation that has occurred. It refers to a complex lesion in which three things are combined: circulatory failure, exudation, and tissue proliferation. More specifically, inflammation is part of innate immunity, and as in other animals, innate immunity in humans recognizes cell surface patterns that are specific to pathogens. Phagocytes recognize cells with such surfaces as non-self and attack the pathogen. If pathogens break through the body's physical barriers, an inflammatory response occurs. Inflammatory response is an immunological defense mechanism to recover and regenerate the damaged part when the body is damaged by various factors, but when it occurs excessively or continuously, acute or chronic inflammatory diseases are induced. This may later cause diseases such as circulatory disorders, multiple sclerosis, and Parkinson's disease. Inflammation occurs in various mechanisms in the body, such as hormone secretion, cytokines, and C-reactive protein (CRP), and there are many related substances. Among them, various cytokines secreted by immune cells regulate the immune system, and some promote inflammation. Therefore, the expression level of intracellular cytokines becomes an indicator of activation of the inflammatory response. In the present invention, it was experimentally confirmed that the secretion of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in splenocytes was reduced in a mouse model in which fatigue was induced by exercise, and the inflammation improvement effect of the peptide confirmed.

As used herein, the term 'antioxidation' means inhibiting oxidative stress. Substances that cause oxidative stress may occur in the human body. When oxygen used as an energy source in the body is incompletely reduced, active oxygen is liberated into the cytoplasm. These reactive oxygen species can damage surrounding cells and further promote tissue damage due to inflammation. However, these reactive oxygen species are generated even in normal circumstances, but if there is no antioxidant defense mechanism in the body, we cannot live. Therefore, various types of antioxidant defense systems are built in our body to counter normal oxidative stress. However, if oxidative stress exceeds normal levels, the body suffers serious damage, so countermeasures against oxidative stress are needed. Antioxidants are natural or synthetic substances that function to prevent or inhibit oxidative stress. In order to evaluate the antioxidant effect of these substances in the body, the contents of superoxide dismutase (SOD), catalase, peroxidase, and glutathione peroxidase (GPx), which are antioxidants in the body, were evaluated by various methods. And, these substances mainly exert the function of removing the active phase. In the present invention, it was experimentally confirmed that the level of superoxide dismutase (SOD) in the blood was increased in a mouse model in which fatigue was induced by exercise, and it was confirmed that the peptide exhibited an antioxidant effect.

In another embodiment of the present invention, the present invention is glycyl -L- leucine (Glycyl-L-Leucine) peptide, L- leucyl glycine (L-Leucylglycine) peptide or both; Or a composition for relieving, improving, or treating menopausal symptoms containing a salt thereof as an active ingredient, preferably a health functional food composition, a cosmetic composition, a pharmaceutical composition, or a quasi-drug composition is provided.

That is, the present invention relates to glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both; or a salt thereof as a health functional food composition, a cosmetic composition, a pharmaceutical composition, or a quasi-drug composition, and a new use for relieving, improving or treating menopausal symptoms.

The present invention glycyl -L- leucine (Glycyl-L-Leucine) peptide, L- leucyl glycine (L-Leucylglycine) peptide or both; Or it provides a method for alleviating, improving or treating menopausal symptoms by treating a salt thereof.

The glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide, or both of them may be active ingredients or indicator ingredients.

In the present invention, 'menopausal symptoms' collectively refers to symptoms and diseases that occur in women before and after menopause as estrogen secretion decreases due to ovarian aging and the like. It is also called 'menopausal syndrome' or 'menopausal symptoms'. Menopausal or menopausal symptoms include, for example, hot flashes, sweating, nervousness, depression, dizziness, fatigue, joint pain, muscle pain, headache, Garson's palpitations, drowsiness, sweating during sleep, trouble sleeping, dry skin, vaginal dryness, and vaginal atrophy. , lower urethral atrophy, sexual intercourse, vaginitis, cystitis, dysuria, urgency, difficulty concentrating, memory impairment, anxiety, nervousness, memory loss, dry skin, arthralgia, or osteoporosis, but is not limited thereto. In addition, it may be a cardiovascular abnormality such as heart disease, high blood pressure, or stroke.

In the present invention, 'prevention' refers to any activity that suppresses or delays desired symptoms by administering the composition of the present invention.

In the present invention, 'treatment' refers to all activities that improve or disappear desired symptoms or diseases by administering the composition of the present invention.

In the present invention, 'improvement' refers to any activity in which the desired symptoms are improved or advantageously changed by administering the composition of the present invention compared to before administration.

The present invention solves the above problems, and to achieve the object of the present invention, a compound represented by Formula 1, a compound represented by Formula 2, or both; or a salt thereof.

A compound represented by Formula 1 below, a compound represented by Formula 2, or both of them may be an active ingredient or a marker ingredient.

[Formula 1]

The compound represented by Chemical Formula 1 has a molecular formula of C ₈ H ₁₆ N ₂ O ₃ and a molecular weight of 188.22, and is named as glycyl-L-leucine. It is a dipeptide composed of two amino acids, glycine and L-leucine, and is abbreviated as Gly-Leu or GL.

The present invention is not particularly limited to the method for obtaining glycyl-L-leucine, and materials derived from natural products, chemically synthesized by methods known in the art to which the present invention pertains, or commercially available materials may be used. Preferably, a material extracted from the placenta of mammals such as pigs, cows, sheep, and horses may be used, and more preferably, a material extracted from the placenta of pigs may be used.

[Formula 2]

The compound represented by Chemical Formula 2 has a molecular formula of C ₈ H ₁₆ N ₂ O ₃ and a molecular weight of 188.22, and is named L-Leucylglycine. It is a dipeptide composed of two amino acids, L-Leucine and Glycine, and is abbreviated as Leu-Gly or LG.

The present invention is not particularly limited to the method for obtaining L-leucylglycine, and materials derived from natural products, chemically synthesized by methods known in the field to which the present invention pertains, or commercially available materials may be used. Preferably, a material extracted from the placenta of mammals such as pigs, cows, sheep, and horses may be used, and more preferably, a material extracted from the placenta of pigs may be used.

In the cosmetic composition, pharmaceutical composition, quasi-drug composition or food composition according to the present invention, the compound represented by Formula 1, the compound represented by Formula 2, or both; Or the content of the salt thereof is 0.00000001 to 100% by weight relative to the total weight of the cosmetic composition, pharmaceutical composition, quasi-drug composition, or food composition, that is, the compound represented by Formula 1, the compound represented by Formula 2, or both by themselves Intake, It is also possible to apply

Preferably, in the present invention, when the active ingredient is a mixture of the compound represented by Formula 1 and the compound represented by Formula 2, each component is mixed in a weight ratio of 1:0.05 to 20, preferably 1:0.1 to 10. It can be. The present inventors experimentally confirmed the synergistic effect by mixing the active ingredients at a weight ratio of 1: 1 included in the above range along with the effect of each component.

In the present invention, the compound represented by Formula 1, the compound represented by Formula 2, or both used as active ingredients may be used in the form of a salt. At this time, the composition according to the present invention can be used in the form of a salt that exhibits the desired effect of the present invention without causing health problems in the case of food, and in the case of a cosmetic composition, a cosmetically acceptable salt, pharmaceutical, In the case of a quasi-drug composition, it may be used in the form of a pharmaceutically acceptable salt. The compound represented by Chemical Formula 1, the compound represented by Chemical Formula 2, or a salt of both of them is a compound represented by Chemical Formula 1, a compound represented by Chemical Formula 2, or both of them with an acid (eg, an organic acid or an inorganic acid that has no or little toxicity). ) or a salt prepared by adding a base.

Examples of the salt include, but are not limited to, sodium salt, salicylate, benzoate, calcium salt, citrate, sulfite and hydrogen sulfite, aluminum salt, hydrochloride, permanganate, potassium salt, acetate, bromate, nitrate, carbonate. , hydrogen carbonate, thiocyanate, ferrous salt, thiosulfate, ferric salt, cupric salt, lactate, magnesium salt, sulfate, phosphate, bromide, zinc salt, iodine salt, succinate, etc. may be included.

a compound represented by Formula 1, a compound represented by Formula 2, or both of the present invention; Or a salt thereof may exist in unsolvated form as well as solvated form including hydrate, ethanolate and the like. a compound represented by Formula 1, a compound represented by Formula 2, or both of the present invention; Or a salt thereof may exist in crystalline or amorphous form, and all such physical forms are included in the scope of the present invention.

The present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a food composition for improving skin elasticity, moisturizing skin or preventing skin aging, comprising a salt thereof as an active ingredient.

The present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a food composition for fatigue improvement, inflammation improvement or antioxidant comprising a salt thereof as an active ingredient.

The present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a food composition for improving, preventing or treating menopausal symptoms comprising a salt thereof as an active ingredient. Preferably, the food composition may be a functional food.

As used herein, the term 'functional food' refers to a compound represented by Formula 1, a compound represented by Formula 2, or both, added to food materials such as beverages, teas, spices, chewing gum, confectionery, encapsulated, It is a food manufactured in powder form, suspension, etc., which means that it brings a specific effect on health when ingested. there is. Since the functional food of the present invention obtained in this way can be consumed on a daily basis, it is excellent for improving skin elasticity, moisturizing skin or preventing skin aging, relieving fatigue, relieving inflammation, antioxidant, and/or alleviating, preventing, or treating menopausal symptoms. It is very useful because you can expect the effect.

a compound represented by Formula 1, a compound represented by Formula 2, or both; Or when using a salt thereof as a food additive, the compound represented by the formula (1), the compound represented by formula (2), or both; Alternatively, a salt thereof may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to conventional methods. The mixing amount of the active ingredient may be appropriately determined depending on the purpose of use (prevention, health or therapeutic treatment). In general, when preparing food or beverage, the composition of the present invention is added in an amount of 15 parts by weight or less, preferably 10 parts by weight or less, based on the raw material. However, in the case of long-term intake for the purpose of health and hygiene or health control, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range. . There is no particular limitation on the type of food. Examples of foods to which the substance can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, There are alcoholic beverages, vitamin complexes, and the like, and includes all foods in a conventional sense.

If the food is a beverage, it may be included in a ratio of 1 to 30 g, preferably 3 to 20 g, based on 100 ml. In addition, the composition may include additional ingredients that are commonly used in food compositions to improve odor, taste, and visual properties. For example, vitamins A, C, D, E, B1, B2, B6, B12, niacin, biotin, folate, panthotenic acid, and the like may be included. In addition, minerals such as zinc (Zn), iron (Fe), calcium (Ca), chromium (Cr), magnesium (Mg), manganese (Mn), and copper (Cu) may be included. In addition, amino acids such as lysine, tryptophan, cysteine, and valine may be included. In addition, preservatives (potassium sorbate, sodium benzoate, salicylic acid, sodium dehydroacetate, etc.), disinfectants (bleaching powder, high bleaching powder, sodium hypochlorite, etc.), antioxidants (butylhydroxyanisole (BHA), butylhydroxytoluene ( BHT), etc.), coloring agents (tar color, etc.), coloring agents (sodium nitrite, sodium nitrite, etc.), bleaching agents (sodium sulfite), seasonings (MSG (sodium glutamate), etc.), sweeteners (dulcin, cyclemate, saccharin, sodium Food additives (e.g.), flavorings (vanillin, lactones, etc.), expanding agents (alum, D-potassium hydrogen tartrate, etc.), strengthening agents, emulsifiers, thickeners (thickeners), coating agents, gumicides, foam inhibitors, solvents, improvers, etc. food additives) may be added. The additive is selected according to the type of food and used in an appropriate amount. The ratio of these additives is not very important, but is generally selected in the range of 0.01 to 0.8 parts by weight per 100 parts by weight of the composition of the present invention.

The food of the present invention can be prepared by a method commonly used in the art, and can be prepared by adding raw materials and ingredients commonly added in the art during the preparation. In addition, unlike general medicines, there is an advantage in that there is no side effect that may occur when taking medicines for a long time by using food as a raw material, and it can be excellent in portability.

As another aspect, a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a cosmetic composition for improving skin elasticity, moisturizing skin or preventing skin aging comprising a cosmetically acceptable salt thereof as an active ingredient.

As another aspect, a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a cosmetic composition for fatigue improvement, inflammation improvement or antioxidant comprising a salt thereof as an active ingredient.

As another aspect, a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a cosmetic composition for improving, preventing or treating menopausal symptoms comprising a salt thereof as an active ingredient.

Cosmetics prepared by containing the composition as an active ingredient as a cosmetic composition may be prepared in the form of general emulsified formulations and solubilized formulations. For example, lotion such as softening lotion or nourishing lotion, emulsion such as facial lotion, body lotion, etc., cream such as nourishing cream, moisture cream, eye cream, essence, cosmetic ointment, spray, gel, pack, sunscreen, makeup base, liquid It may have formulations such as foundation, powder, cleansing cream, cleansing lotion, makeup remover such as cleansing oil, cleanser such as cleansing foam, soap, body wash, etc., such as type, solid type or spray type, but is not limited thereto.

The composition may contain a relatively high concentration of the composition of the present invention in the case of wash-off type cosmetics, such as makeup removers and detergents, in which active ingredients stay on the skin for a short period of time when commercialized as cosmetics. will be. On the other hand, in the case of leave-on type cosmetics such as lotion, lotion, cream, essence, etc., in which active ingredients stay on the skin for a long time, the composition of the present invention is contained at a lower concentration than wash-off type cosmetics. It will be fine even if you do.

In addition, the cosmetic may include, in addition to the composition of the present invention, a fatty substance, an organic solvent, a solubilizing agent, a thickening agent and a gelling agent, a softening agent, an antioxidant, a suspending agent, a stabilizer, a foaming agent, a fragrance, a surfactant, water, Ionic or nonionic emulsifiers, fillers, sequestering and chelating agents, preservatives, vitamins, blocking agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic actives, lipid vesicles or any commonly used in cosmetics It may contain adjuvants commonly used in the field of cosmetology, such as other ingredients of

In addition, the cosmetic composition of the present invention may further include one or more cosmetically acceptable carriers formulated in general skin cosmetics.

Cosmetically acceptable carriers included in the cosmetic composition of the present invention vary depending on the formulation. When the formulation of the present invention is an ointment, paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tracanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide or Mixtures of these may be used.

When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, or a mixture thereof may be used as a carrier component, and especially in the case of a spray, chloro propellants such as fluorohydrocarbons, propane/butane or dimethyl ether.

When the formulation of the present invention is a solution or emulsion, a solvent, solubilizing agent, or emulsifying agent is used as a carrier component, such as water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl benzoate, propylene glycol, 1,3 -There is butyl glycol oil, in particular cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan.

When the dosage form of the present invention is a suspension, a liquid diluent such as water, ethanol or propylene glycol, a suspending agent such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, a micro Crystalline cellulose, aluminum metahydroxide, bentonite, agar or tracanth and the like may be used.

When the formulation of the present invention is a soap, alkali metal salts of fatty acids, fatty acid hemiester salts, fatty acid protein hydrolyzates, isethionates, lanolin derivatives, aliphatic alcohols, vegetable oils, glycerol, sugars, etc. may be used as carrier components. can

In another aspect, the present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a pharmaceutical composition for improving skin elasticity, moisturizing skin or preventing skin aging comprising a pharmaceutically acceptable salt thereof as an active ingredient.

As another aspect, a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a pharmaceutical composition for fatigue improvement, inflammation improvement or antioxidant comprising a salt thereof as an active ingredient.

As another aspect, a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a pharmaceutical composition for improving, preventing or treating menopausal symptoms comprising a salt thereof as an active ingredient.

The composition is a solution, gel, emulsion, suspension, micro-emulsion, micro-capsule, liposome, cream, lotion, ointment, aerosol ( aerosol, spray, paste, and patch.

The pharmaceutical composition of the present invention can be used as a single formulation, or can be prepared and used as a combination formulation by further including a drug known to have a recognized anti-inflammatory or antioxidant effect, formulated using a pharmaceutically acceptable carrier or excipient. By doing so, it can be prepared in unit dose form or prepared by inserting it into a multi-dose container.

As used herein, the term "pharmaceutically acceptable carrier" may refer to a carrier or diluent that does not inhibit the biological activity and properties of the injected compound without irritating living organisms. The type of the carrier that can be used in the present invention is not particularly limited, and any carrier commonly used in the art and pharmaceutically acceptable can be used. Non-limiting examples of the carrier include saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol, and the like. These may be used alone or in combination of two or more. The carrier may include a non-naturally occurring carrier.

In addition, if necessary, other conventional additives such as antioxidants, buffers, and/or bacteriostatic agents may be added and used, and diluents, dispersants, surfactants, binders, lubricants, etc. may be additionally added to form main solutions such as aqueous solutions, suspensions, and emulsions. It may be formulated into a dosage form, pill, capsule, granule or tablet for use.

In addition, the pharmaceutical composition of the present invention may include a compound represented by Formula 1, a compound represented by Formula 2, or both of them in a pharmaceutically effective amount; or a pharmaceutically acceptable salt thereof. As used herein, the term "pharmaceutically effective amount" means an amount sufficient to induce skin elasticity improvement, skin moisturizing or skin aging prevention effect at a reasonable benefit/risk ratio applicable to medical treatment; or an amount sufficient to induce fatigue, inflammation, or antioxidant effects; Or an amount sufficient to alleviate, ameliorate or treat menopausal symptoms, generally in an amount of 0.001 to 1000 mg/kg, preferably in an amount of 0.05 to 200 mg/kg, more preferably in an amount of 0.1 to 100 mg/kg. It can be administered by dividing it into one to several times a day. However, for purposes of the present invention, a specific therapeutically effective amount for a particular patient is determined by the type and extent of the response to be achieved, the specific composition, including whether other agents are used as the case may be, the patient's age, weight, general state of health, It is preferable to apply differently according to various factors including gender and diet, administration time, administration route and secretion rate of the composition, treatment period, drugs used together with or concurrently used with a specific composition, and similar factors well known in the medical field.

In another aspect, the present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a quasi-drug composition for improving skin elasticity, moisturizing skin or preventing skin aging comprising a pharmaceutically acceptable salt thereof as an active ingredient.

In another aspect, the present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a quasi-drug composition for fatigue improvement, inflammation improvement or antioxidant comprising a pharmaceutically acceptable salt thereof as an active ingredient.

In another aspect, the present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a quasi-drug composition for relieving, improving or treating menopausal symptoms comprising a pharmaceutically acceptable salt thereof as an active ingredient.

As used herein, the term “quasi-drugs” refers to textiles, rubber products, or similar products used for the purpose of treating, alleviating, treating, or preventing diseases of humans or animals, devices that have weak effects on the human body or do not directly act on the human body, and devices or non-machines and similar items, products falling under one of the agents used for sterilization, insecticidal, and similar purposes to prevent infectious diseases, for the purpose of diagnosing, treating, mitigating, treating, or preventing human or animal diseases It may refer to items other than instruments, machines or devices among items used and items other than instruments, machines or devices among items used for the purpose of pharmacologically affecting the structure and function of humans or animals. In addition, the quasi-drugs may include external skin preparations and personal hygiene products. Preferably, it may be a disinfectant cleaner, shower foam, gargreen, wet tissue, detergent soap, hand wash, or ointment, but is not limited thereto.

When the composition according to the present invention is used as a quasi-drug additive, the composition may be added as it is or used together with other quasi-drugs or quasi-drug ingredients, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be appropriately determined depending on the purpose of use.

In addition, the present invention glycyl -L- leucine (Glycyl-L-Leucine) peptide, L- leucyl glycine (L-Leucylglycine) peptide or both; Or a composition for improving skin elasticity, moisturizing skin, and preventing skin aging, comprising a salt thereof as a marker component, preferably a cosmetic composition, pharmaceutical composition, quasi-drug composition, or food composition.

In another aspect, the present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a quasi-drug composition for fatigue improvement, inflammation improvement or antioxidant comprising a pharmaceutically acceptable salt thereof as a marker component.

In another aspect, the present invention relates to a compound represented by Formula 1, a compound represented by Formula 2, or both; Or it provides a quasi-drug composition for relieving, improving or treating menopausal symptoms comprising a pharmaceutically acceptable salt thereof as a marker component.

the glycyl-L-leucine peptide, the L-leucylglycine peptide, or both of the present invention; Or a composition containing a salt thereof as an active ingredient exhibits excellent effects in improving skin elasticity, moisturizing skin, and preventing skin aging.

Glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both of them; Or a composition containing a salt thereof exhibits excellent effects on fatigue improvement, inflammation improvement or antioxidant.

Glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both of them; Or a composition containing a salt thereof shows an excellent effect in relieving menopausal symptoms.

the glycyl-L-leucine peptide, the L-leucylglycine peptide, or both of the present invention; Or a composition containing a salt thereof may be used as a cosmetic raw material, a pharmaceutical ingredient, a quasi-drug raw material, or a food raw material that is safe to the skin and has an excellent skin condition improvement effect.

Glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both of them; Or a composition containing a salt thereof may be used as a cosmetic raw material, a pharmaceutical ingredient, a health functional food raw material or a feed raw material having excellent fatigue improvement, inflammation improvement or antioxidant effect while being safe.

Glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both of them; Or a composition containing a salt thereof can be used as a cosmetic raw material, a pharmaceutical ingredient, a health functional food raw material, or a feed raw material having excellent menopausal symptom alleviation effect while being safe.

1 is a graph showing the hyaluronic acid synthesis promoting effect of a glycyl-L-leucine peptide, an L-leucylglycine peptide, or both.
Figure 2 is a graph showing the procollagen synthesis promoting effect of glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both.
Figure 3 is a graph showing the elastase inhibitory activity of glycyl-L-leucine (Glycyl-L-Leucine) peptide, L-leucyl glycine (L-Leucylglycine) peptide or both of them.
Figure 4 is a graph showing the effect of improving inflammation and fatigue by inhibiting excessive secretion of prostaglandin E2 (PGE2) according to Examples and Comparative Examples of the present invention.
Figure 5 is a graph showing the antioxidant effect by DPPH free radical scavenging activity according to Examples and Comparative Examples of the present invention.
Figure 6 is a graph showing the fatigue improvement effect by preventing the accumulation of lactic acid in muscle according to Examples and Comparative Examples of the present invention.
7 is a graph showing the fatigue improvement effect by preventing the accumulation of creatine kinase (CK) in muscle according to Examples and Comparative Examples of the present invention.
Figure 8 is a graph showing the inflammation improvement effect by reducing the tumor necrosis factor-α (TNF-α) in the blood according to Examples and Comparative Examples of the present invention.
9 is a graph showing an effect of reducing inflammation by reducing interleukin-6 (IL-6) in blood according to Examples and Comparative Examples of the present invention.
Figure 10 is a graph showing the antioxidant effect by the increase of superoxide dismutase (SOD) in the blood according to Examples and Comparative Examples of the present invention.
11 is an experimental result confirming whether estrogen receptor activity is increased by treating Examples and Comparative Examples of the present invention. When the glycyl-L-leucine peptide and the L-leucyl glycine peptide were treated together, it was confirmed that the estrogen receptor activity was significantly increased. Through these results, it was confirmed that the glycyl-L-leucine peptide and/or the L-leucyl glycine peptide of the present invention can be effectively used for treating, improving, and/or preventing menopausal diseases.
12 is a result confirming the effect of inhibiting osteoclast differentiation by treating Examples and Comparative Examples of the present invention. Through these results, it can be confirmed that the glycyl-L-leucine peptide and/or the L-leucylglycine peptide of the present invention inhibits osteoclast differentiation and can be effectively used for the treatment, improvement, and/or prevention of postmenopausal osteoporosis. there was.

Hereinafter, examples and the like will be described in detail to aid understanding of the present invention. However, the embodiments according to the present invention can be modified in many different forms, and the scope of the present invention should not be construed as being limited to the following examples. Embodiments of the present invention are provided to more completely explain the present invention to those skilled in the art.

reference example One: Glysil -L- Leucine ( Glycyl -L- Leucine ) peptide substance information

[Formula 1]

Substance Name: Glycyl-L-Leucine

CAS No. : 869-19-2

Molecular formula: C ₈ H ₁₆ N ₂ O ₃

Molecular Weight: 188.22

Purchased from: Bachem AG (Bubendorf, Switzerland)

reference example 2: L- Leucylglycine (L- Leucylglycine ) peptide substance information

Substance Name: L-Leucylglycine

CAS No. : 686-50-0

Molecular formula: C ₈ H ₁₆ N ₂ O ₃

Molecular Weight: 188.22

Purchased from: Bachem AG (Bubendorf, Switzerland)

Experimental example 1: hyaluronic acid ( Hyaluronic acid) synthesis accelerating effect

As skin aging progresses, collagen and elastin, which are fibrous tissues that serve as a basic support for supporting the skin structure, harden and become insoluble. Substances that fill between cells and between fibers such as collagen and elastin are called matrices, and have a very strong ability to retain moisture. Representative examples of such matrices include hyaluronic acid and mucopolysaccharides. Hyaluronic acid is a polysaccharide widely distributed in intracellular tissues such as joint fluid and skin. As a polymer of hyaluronan in which N-acetylglucosamine and glucuronic acid are combined, it has moisturizing properties and viscosity. As aging progresses, when these substances decrease, skin dryness is induced and skin elasticity is weakened (Kim Yun-beom, Journal of Dong-A Physiopathology, 2010; 24(4): 533-542).

The present inventors cultured normal human epidermal keratinocytes and then treated the samples in order to confirm the hyaluronic acid synthesis effect of the Glycyl-L-Leucine peptide or the L-Leucylglycine peptide. After 24 hours, the medium of the cultured cells was collected, and the amount of hyaluronic acid produced from the culture medium was measured using a Hyaluronan DuoSet ELISA Kit (DY3614), and the amount of protein was corrected. As a positive control, retinoic acid was used. The test results are shown in FIG. 1 with the control group not treated as 100%.

As a result, it was confirmed that the Glycyl-L-Leucine peptide or L-Leucylglycine peptide increases hyaluronic acid production even at low concentrations, which means that it can improve skin condition by helping to moisturize the skin. In addition, when Glycyl-L-Leucine and L-Leucylglycine were treated together, it was confirmed that hyaluronic acid production increased more than when treated alone.

Experimental example 2: procollagen ( Procollagen ) Synthesis accelerating effect

Procollagen is a component of collagen, synthesized in fibroblast, osteoblast, etc., and finally converted into collagen by procollagen peptase. An increase in procollagen is related to the binding force and elasticity of the skin.

In order to confirm the procollagen synthesis promoting effect of Glycyl-L-Leucine peptide or L-Leucylglycine peptide, human-derived fibroblasts (Normal human primary dermal fibroblast-Neonatal) were cultured, and then samples were treated. After 24 hours, the medium of the cultured cells was collected, and the amount of procollagen was measured using a Procollagen type I-C peptide (PIP) kit. Cells adhering to the bottom were washed with DPBS and then lysed with 1N NaOH. The total amount of protein was measured using the BCA Protein Assay Kit, and the amount of procollagen synthesis per protein was obtained. TGF-β was used as a positive control. The test results are shown in FIG. 2 with the control group not treated with anything as 100%.

As a result, it was confirmed that Glycyl-L-Leucine and L-Leucylglycine peptides increased procollagen synthesis, thereby helping skin elasticity. In addition, when Glycyl-L-Leucine and L-Leucylglycine were treated together, it was confirmed that procollagen synthesis increased more than when treated alone.

Experimental example 3: elastase ( Elastase ) inhibitory activity

In the dermal tissue of the skin, collagen and elastin form a network structure to maintain skin elasticity. Due to internal and external stresses such as age and UV rays, elasticity and moisture decrease, and overexpressed elastase causes the network structure of elastin. As the skin breaks down, the skin sags and wrinkles form, resulting in skin aging. Therefore, skin aging can be inhibited by inhibiting the activity of elastase, an elastin degrading enzyme, which is one of the main causes of skin aging (Jaewon Lee, Journal of the Korean Society of Food and Nutrition, 2008, 21(2); 143-147).

After adding 1 U/ml porcine pancreatic elastase (Sigma, E1250) dissolved in 50 mM Tris-Hcl buffer (pH 8.0) and the sample to 96 wells, the mixture was reacted at room temperature for 30 minutes. After adding N-succinyl-(L-Ala)3-p-nitroanilide as a substrate, the amount of p-nitroanilide generated from the substrate was measured at 410 nm. The test results are shown in FIG. 3 based on the control group that was not treated with anything.

As a result, it was confirmed that Glycyl-L-Leucine peptide, L-Leucylglycine peptide, and mixture of Glycyl-L-Leucine peptide and L-Leucylglycine peptide (1:1) inhibit elastase activity, which is the main cause of skin aging. It was confirmed that skin aging can be inhibited by inhibiting the activity of phosphorus elastase.

Experimental example 4: In vitro experiment

The present inventors evaluated the fatigue improvement, inflammation improvement or antioxidant effect, and female menopausal symptom improvement effect of a composition containing glycyl-L-leucine, L-leucylglycine or both of them. In vitro experiments were conducted prior to the following. Glycyl-L-Leucine or L-Leucylglycine was provided by Bachem AG (Bubendorf, Switzerland). As shown in Table 1 below, a composite composition was prepared and used in the experiment.

Concentration (mg/ml) Glycyl-L-Leucine
(Glycyl-L-Leucine) L-Leucylglycine
(L-Leucylglycine) Comparative Example 1 - - Example 1 1 µg/ml - Example 2 10 µg/ml - Example 3 - 1 µg/ml Example 4 - 10 µg/ml Example 5 0.5 µg/ml 0.5 µg/ml Example 6 5 µg/ml 5 µg/ml

Experimental example 5: prostaglandin E2 ( PGE2 ) Inflammation improvement and fatigue improvement effect by inhibiting excessive secretion

Cytokines are proteins secreted mainly by monocytes, lymphocytes, T cells, B cells, natural killer cells, and fibroblasts during an inflammatory response. Like neurotransmitters, they are receptors on cell membranes. Acts on cells or, like hormones, acts on intracellular receptors to transmit information between cells. Cytokines that promote inflammation, represented by interleukin-1β, -2, -6 (IL-1β, -2, -6), IFN-γ, and TNF-α, activate cyclo-oxygenase-2 (COX-2) to prostaglandin E2 (prostaglandine E2, hereinafter referred to as PGE2) is increased and inflammatory cells are activated to induce an inflammatory process (Song Hu-rim et al., Korean Journal of Psychopharmacology 2013; 24: 5-10). The present inventors evaluated the anti-inflammatory effect through inhibition of prostaglandin E2 (PGE2) secretion.

First, in order to measure prostaglandin E2 (PGE2), mouse macrophage RAW 264.7 Mouse macrophage cell line (KCLB no 40071, Korean Cell Line Bank, Seoul, Korea) was purchased and 96 at a cell concentration of 2.5X10 ⁶ cells/ml. 100 μl was dispensed into each -well microplate. The medium was DMEM (GibcoBRL, Grand Island, NY, USA) supplemented with 10% FBS (GibcoBRL, Grand Island, NY, USA) and 1% penicillin/streptomycin (Thermo Scientific Hyclone, Waltham, MA, USA). , 5% CO ₂ , and cultured in a 37°C environment. Inflammation was induced by treating 1 μg/ml of Lipopolysaccharide (LPS) (Sigma, St. Louis, MO, US) to the cell culture solution in which the RAW 264.7 cells were dispensed, and treatment was performed according to Experimental Example 1. Thereafter, the cells were cultured for 24 hours, and the cell culture medium was recovered after centrifugation at 900 rpm and 4° C. for 5 minutes, and the secretion amount of prostaglandin E2 (PGE2) produced in the culture medium was confirmed. Prostaglandin E2 (PGE2) was measured using a commercially available PGE2 ELISA kit (MyBioSource Co., Ltd., San Diego, CA, USA) according to the manufacturer's instructions.

As a result, as shown in FIG. 4, prostaglandin E2 (PGE2) was found to be significantly increased in macrophages inflamed with LPS than in non-inflamed cells, but decreased again in the cells treated with the sample. 10 μg/ml more than when glycyl-L-leucine peptide or L-leucylglycine peptide was treated at 1 μg/ml (Example 1, Example 3), respectively (Example 2, Example 4), the concentration of prostaglandin E2 (PGE2) decreased. In particular, compared to cells treated with glycyl-L-leucine peptide or L-leucylglycine peptide (Examples 1 to 4) alone, the composite composition (Example 5, Execution It was confirmed that the secretion of prostaglandin E2 (PGE2) was reduced in the cells treated with Example 6).

The results of Experimental Example 2 show that glycyl-L-leucine, L-leucylglycine, or a composition containing both prostaglandin promotes inflammation in inflamed macrophages. By inhibiting E2 (PGE2) secretion, it showed an effective anti-inflammatory effect, further suggesting that it can exhibit fatigue improvement functionality.

Experimental example 6: DPPH Antioxidant effect by free radical scavenging activity

DPPH (2,2-Diphenyl-1-picrylhydrazyl) is a purple stable free radical that reacts with antioxidants to receive hydrogen and be reduced to decolorize. By comparing the DPPH free radical scavenging activity of samples using the same principle, their antioxidant ability can be evaluated. It can be interpreted that the higher the free radical scavenging activity (%), the higher the antioxidant capacity of the sample.

For DPPH (1,1-Diphenyl-2-picryl-hydrazyl) free radical scavenging activity, first, 4 mg of DPPH was dissolved in 100 ml of methanol to make a DPPH solution, and the composition of Experimental Example 1 was diluted in ethanol by concentration. 1ml of ethanol containing the composite composition and 1ml of DPPH solution were mixed well, and then allowed to stand at 37° C. for 30 minutes, and then absorbance was measured at 516nm. Ethanol was used as a control.

The antioxidant capacity by the DPPH free radical scavenging activity method was calculated as follows.

[Equation 1]

DPPH free radical scavenging activity = 100 - (absorbance of sample/absorbance of control) X 100

As a result, as shown in FIG. 5, DPPH free radical scavenging activity increased when the sample was treated. 10 μg/ml more than when glycyl-L-leucine peptide or L-leucylglycine peptide was treated at 1 μg/ml (Example 1, Example 3), respectively (Example 2, Example 4) DPPH free radical scavenging activity increased. In particular, composite compositions (Examples 5 and 6) compared to single samples of glycyl-L-leucine peptide or L-leucylglycine peptide (Examples 1 to 4) It was confirmed that DPPH free radical scavenging activity increased in the sample.

The results of Experimental Example 3 show that a composition containing Glycyl-L-Leucine, L-Leucylglycine or both can be used as an effective antioxidant functional material by suppressing oxidative stress. is to imply

Experimental example 7: Animal Testing

The present inventors purchased male ICR mice (4 weeks old) from Daehan Biolink (Daejeon, Korea) for animal experiments. Experimental animals were bred 5-10 animals per cage in a breeding room maintained at a temperature of 22±1° C. and a humidity of 55±10%. Glycyl-L-Leucine peptide or L-Leucylglycine peptide was provided by Bachem AG (Bubendorf, Switzerland) and used after being dissolved in distilled water. The control group was orally administered only distilled water, and the Glycyl-L-Leucine peptide or L-Leucylglycine peptide intake group was administered daily for 21 days using a sonde for oral administration. was administered orally once. The types and concentrations of the compositions administered to each group are shown in Table 2 below. Fatigue was induced by exercising mice in each group for 30 minutes once a week for 3 weeks. The exercise speed was 10 m/min for 10 minutes, 16 m/min for 10 minutes, and finally 21 m/min for 10 minutes. On the 21st day, they were exercised for 30 minutes while increasing the speed by 3 m/min every 3 minutes after 10 m/min for the first 5 minutes, and used for analysis after sacrifice.

Intake (mg/kg/day) Glycyl-L-Leucine
(Glycyl-L-Leucine) L-Leucylglycine
(L-Leucylglycine) Comparative Example 2 - - Example 7 1 mg/kg/day - Example 8 10 mg/kg/day - Example 9 - 1 mg/kg/day Example 10 - 10 mg/kg/day

Experimental example 8: Fatigue improvement effect by preventing accumulation of lactic acid in muscle

In situations where oxygen is sufficiently supplied to the cells, the TCA cycle proceeds smoothly, so the blood lactate concentration is maintained in the range of 0.56 - 2 mmol/L and does not accumulate beyond that. However, when the metabolic link between glycolysis and the TCA cycle does not progress in an equivalence ratio and the glycolysis is relatively active or the oxygen supply is insufficient compared to the amount of oxygen required by the cell, lactic acid is produced in the muscle. In addition, when the oxygen supply during high-intensity exercise does not meet the oxygen consumption of the muscles, the concentration of lactic acid in the muscle tissue increases, and the lactic acid generated at this time diffuses into the blood and is processed by the heart and liver. resulting in fatigue. Phosphorylase activity involved in carbohydrate metabolism during exercise is reduced, and as a result, glucose generation, which is a source of kinetic energy in an anaerobic state, is suppressed. The present inventors confirmed the inhibition of accumulation of lactic acid and creatine kinase (CK) in muscles representing a fatigue state among various fatigue improvement effect evaluations.

As a result, as shown in Figure 6, when glycyl-L-leucine (Glycyl-L-Leucine) peptide or L-leucylglycine (L-Leucylglycine) peptide was orally administered in a mouse model where fatigue was induced (Example 7 to 10) It was found that the concentration of lactic acid accumulated in muscle was significantly reduced compared to the control group (Comparative Example 2). In particular, when glycyl-L-leucine (Glycyl-L-Leucine) peptide or L-leucylglycine (L-Leucylglycine) peptide was ingested at 1 mg/kg/day (Examples 7 and 9), 10 mg/ It was confirmed that the concentration of lactic acid decreased when each kg was ingested (Example 8 and Example 10).

That is, the intake of Glycyl-L-Leucine peptide or L-Leucylglycine peptide in subjects induced by fatigue can reduce the accumulation of lactic acid in the muscle, thereby improving fatigue. confirmed that there is

Experimental example 9: intramuscular of creatine kinase (CK) Fatigue improvement effect by preventing accumulation

As an enzyme present in large amounts in muscle, when muscle cells are damaged, these enzymes are released into the blood to create an energy source. Creatine kinase (CK) plays a role in synthesizing creatine phosphate required for ATP resynthesis (a process of synthesizing ATP from ADP and phosphocreatine) during anoxia in muscle cells during exercise.

In order to measure intramuscular creatine kinase (CK), fatigue was induced in mice as in Experimental Example 4, and muscle tissue of each group of mice administered with the composition was collected and used for analysis. Creatine kinase (CK) was measured using an assay kit (Abcam, Cambridge, UK) according to the manufacturer's instructions.

As a result, as shown in FIG. 7, oral administration of glycyl-L-leucine peptide or L-leucylglycine peptide in a mouse model where fatigue was induced (Examples 7 to 7) 10), the concentration of creatine kinase (CK) accumulated in muscle was significantly reduced compared to the control group (Comparative Example 2). In particular, when glycyl-L-leucine (Glycyl-L-Leucine) peptide or L-leucylglycine (L-Leucylglycine) peptide was ingested at 1 mg/kg/day (Examples 7 and 9), each 10 mg It was confirmed that creatine kinase (CK) decreased when / kg was ingested (Example 8, Example 10).

That is, the intake of Glycyl-L-Leucine peptide or L-Leucylglycine peptide in fatigue-induced subjects reduces the accumulation of creatine kinase (CK) in muscle, improving fatigue It was confirmed that the enhancement effect can be shown.

Experimental example 10: Inflammation improvement effect by reduction of cytokines in blood

Cytokines include interleukin (IL), interferon (IFN), chemokine, tumor necrosis factor (TNF), and transforming growth factor (TGF). Among the pro-inflammatory cytokines are interleukin-1β, -2, -6 (IL-1β, -2, -6), IFN-γ, TNF-α, etc., and anti-inflammatory Cytokines include IL-4, IL-10, IL-11, IL-13, TGF-β, and the like. The present inventors confirmed the inhibition of the secretion of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), which are proteins that promote inflammation, among the evaluation of the inflammation improvement effect.

First, in order to measure tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in the blood, fatigue was induced as in Experimental Example 4, and blood was collected from mice for each group administered with the composition. . After leaving the collected blood at room temperature for 30 minutes, serum was separated by centrifugation at 6000 rpm for 20 minutes. The separated serum was filtered with a 0.45 μm syringe filter and used for analysis. Tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were tested using commercially available TNF-α and IL-6 ELISA kits (Thermo Scientific Co., Waltham, MA, USA). Measured according to the guidelines.

As a result, as shown in FIG. 8, oral administration of glycyl-L-leucine peptide or L-leucylglycine peptide in the fatigue-induced mouse model (Examples 7 to 8) 10), the concentration of tumor necrosis factor-α (TNF-α) in the blood was significantly reduced compared to the control group (Comparative Example 2). In particular, when glycyl-L-leucine (Glycyl-L-Leucine) peptide or L-leucylglycine (L-Leucylglycine) peptide was ingested at 1 mg / kg / day (Example 7, Example 9), each 10mg / kg / day intake (Example 8, Example 10) when the tumor necrosis factor -α (TNF-α) was reduced.

In addition, as shown in Figure 9, when glycyl-L-leucine (Glycyl-L-Leucine) peptide or L-leucylglycine (L-Leucylglycine) peptide was orally administered in a mouse model where fatigue was induced, the control group (Comparative Example) 2), the concentration of interleukin-6 (IL-6) in the blood was significantly reduced. In particular, when glycyl-L-leucine (Glycyl-L-Leucine) peptide or L-leucyl glycine (L-Leucylglycine) peptide was ingested 1 mg / kg (Example 7, Example 9), each 10mg / kg Interleukin-6 (IL-6) decreased when ingested (Example 8, Example 10).

That is, the intake of glycyl-L-leucine peptide or L-leucylglycine peptide in a fatigued subject reduces the concentration of inflammatory cytokines in the blood, thereby improving inflammation. confirmed that it can.

Experimental example 11: in the blood super oxide dismutase Antioxidant effect by increasing (SOD)

Superoxide dismutase (SOD) is an antioxidant enzyme that catalyzes a reaction that converts superoxide into oxygen and hydrogen peroxide (H ₂ O ₂ ), and serves to remove free radicals from the blood. An increase in blood SOD reduces intramuscular fatigue. The present inventors confirmed an increase in superoxide dismutase (SOD) responsible for antioxidant function in the body in order to evaluate the antioxidant effect.

In order to measure the level of superoxide dismutase (SOD) in blood, fatigue was induced as in Experimental Example 4, and serum obtained from mice of each group administered with the composition was used for analysis. Superoxide dismutase (SOD) was measured using an assay kit (Abcam, Cambridge, UK) according to the manufacturer's instructions.

As a result, as shown in FIG. 10, oral administration of glycyl-L-leucine peptide or L-leucylglycine peptide in the fatigue-induced mouse model (Examples 7 to 10) 10), it was found that the concentration of superoxide dismutase (SOD) in the blood significantly increased compared to the control group (Comparative Example 2). In particular, when glycyl-L-leucine (Glycyl-L-Leucine) peptide or L-leucylglycine (L-Leucylglycine) peptide was ingested at 1 mg/kg (Example 7, Example 8), each 10mg/kg When ingested (Example 8, Example 10), superoxide dismutase (SOD) increased.

In other words, the intake of Glycyl-L-Leucine peptide or L-Leucylglycine peptide in fatigued subjects increases the concentration of superoxide dismutase (SOD) in the blood, It was confirmed that it can exhibit an antioxidant effect.

Experimental example 12. Assessment of estrogen receptor activity

[0004] 'Menopausal symptoms' are caused by a decrease in the secretion of estrogen due to aging of the ovaries, and various symptoms are induced in women before and after menopause.

ERE (estrogen response element) reporter assay was performed to confirm the activity of two types of dipeptides of Chemical Formulas 1 and 2 on estrogen receptors. 293T cells were cultured in a 24 well plate for 24 hours using DMEM medium supplemented with 10% FBS, and then the medium was exchanged with 500 μl of 5% charcoal stripped FBS containing phenol red free medium per well, and transfection was performed. 0.1ug of ERa, 0.1ug of ERE, and 10ng of pRL-Tk DNA per well were transfected using Lipofectamine® reagents (Thermo Fisher Scientific). 4 hours after transfection, the composition of Experimental Example 1 was treated, and 1ppb (0.5 μl of DMSO) of 17β-estradiol (E2) was treated as a positive control group, and the same amount of DMSO (0.5 μl) was treated as a negative control group (control, ctrl). did After 24 hours of sample treatment, the luminescence was measured using the Dual-Luciferase® Reporter Assay System (Promega), and the negative control value was used as the standard (1.0). The measured values were corrected for firefly luciferase levels with renilla luciferase levels.

As a result, as shown in FIG. 11, estrogen receptor activity increased when the sample was treated. 10 μg/ml more than when glycyl-L-leucine peptide or L-leucylglycine peptide was treated at 1 μg/ml (Example 1, Example 3), respectively (Examples 2 and 4) increased estrogen receptor activity. In particular, composite compositions (Examples 5 and 6) compared to single samples of glycyl-L-leucine peptide or L-leucylglycine peptide (Examples 1 to 4) It was confirmed that estrogen receptor activity was increased in the sample.

Experimental example 13. Inhibition of osteoclast differentiation effect

When menopause occurs, female hormones that play an important role in bone metabolism are no longer secreted from the ovaries, so changes in bone replacement occur, and osteoblasts create new bones and fill the areas where osteoclasts have melted, but bone loss is not completely filled. this will come Bone loss gradually occurs after the age of 30. In the case of men, bone density gradually decreases, resulting in a loss of 20-30% of the maximum bone mass. Osteoporosis progresses more rapidly and severely in women than in men.

Osteoclasts can be generated from macrophage progenitor cells in vitro by activator of NF-κB ligand (RANKL, RANK ligand), a specific cytokine. It proliferates while maintaining its shape, and shows a negative reaction in TRAP staining, giving it a light brown or ocher color. In contrast, TRAP(+) multinucleated cells stained in dark brown to reddish brown were observed in RANKL-treated cells.

RAW264.7 cells cultured in Experimental Example 5 were used in the experiment to confirm the effect of two dipeptides of Chemical Formulas 1 and 2 on osteoclast differentiation induced by Receptor activator of NF-κB ligand (RANKL). . After dispensing 100 μl of RAW264 cells into a 96-well microplate at a cell concentration of 2.5 × 106 cells/ml and incubating them for 5 hours, when the cells are attached to the well, the medium is removed and osteoclast is placed in α-MEM medium supplemented with 10% FBS. A culture solution in which 50 ng/mL of RANKL, a cell differentiation factor, and the composition of Experimental Example 1 was mixed was dispensed and cultured for 7 days. In order to evaluate the effect on osteoclast differentiation, cultured cells were stained with tartrate-resistant acid phosphatase (TRAP), and red-stained cells were regarded as osteoclasts and their number was measured.

As a result, as shown in FIG. 12, osteoclast differentiation was inhibited when the sample was treated. 10 μg/ml more than when glycyl-L-leucine peptide or L-leucylglycine peptide was treated at 1 μg/ml (Example 1, Example 3), respectively (Example 2, Example 4) when osteoclasts were reduced. In particular, composite compositions (Examples 5 and 6) compared to single samples of glycyl-L-leucine peptide or L-leucylglycine peptide (Examples 1 to 4) It was confirmed that the differentiation of osteoclasts was inhibited in the sample.

The results of these Experimental Examples 12 and 13 show that the composition containing Glycyl-L-Leucine, L-Leucylglycine or both of them increases estrogen receptor activity, and menopausal It suggests that it can be effectively used for female menopausal symptoms through inhibition of osteoclast differentiation, which is a factor inducing later osteoporosis.

From the above description, those skilled in the art to which the present invention pertains will be able to understand that the present invention may be embodied in other specific forms without changing its technical spirit or essential features. In this regard, the embodiments described above should be understood as illustrative in all respects and not limiting. The scope of the present invention should be construed as including all changes or modifications derived from the meaning and scope of the claims to be described later and equivalent concepts rather than the detailed description above are included in the scope of the present invention.

Claims (23)

A food composition for improving skin elasticity through collagen synthesis, or for moisturizing the skin, comprising a compound represented by Formula 1 below, or a salt thereof.
[Formula 1]

A cosmetic composition for improving skin elasticity through collagen synthesis, or for skin moisturizing, comprising a compound represented by Formula 1, or a cosmetically acceptable salt thereof.
[Formula 1]

delete A quasi-drug composition for improving skin elasticity through collagen synthesis or for moisturizing the skin, comprising a compound represented by Formula 1 below or a pharmaceutically acceptable salt thereof.
[Formula 1]

delete A compound represented by Formula 1 below; Or a food composition for improving fatigue or antioxidant comprising a salt thereof.
[Formula 1]

A compound represented by Formula 1 below; Or a cosmetic composition for improving fatigue or antioxidant comprising a cosmetically acceptable salt thereof.
[Formula 1]

delete A compound represented by Formula 1 below; Or a quasi-drug composition for fatigue improvement or antioxidant comprising a pharmaceutically acceptable salt thereof.
[Formula 1]

delete A compound represented by Formula 1, a compound represented by Formula 2, or both; Or a functional food composition for preventing or improving female menopausal symptoms comprising a salt thereof.
[Formula 1]

[Formula 2]

A compound represented by Formula 1, a compound represented by Formula 2, or both; Or a cosmetic composition for preventing or improving female menopausal symptoms comprising a cosmetically acceptable salt thereof.
[Formula 1]

[Formula 2]

A compound represented by Formula 1, a compound represented by Formula 2, or both; Or a pharmaceutical composition for treating, preventing or improving female menopausal symptoms comprising a pharmaceutically acceptable salt thereof.
[Formula 1]

[Formula 2]

A compound represented by Formula 1, a compound represented by Formula 2, or both; Or a quasi-drug composition for the treatment, prevention or improvement of female menopausal symptoms comprising a pharmaceutically acceptable salt thereof.
[Formula 1]

[Formula 2]

delete The method according to any one of claims 11 to 14, wherein the female menopausal symptoms include hot flashes, sweating, nervousness, depression, dizziness, fatigue, joint pain, muscle pain, headache, palpitations, sweating during sleep, Sleep disorder, dry skin, vaginal dryness, vaginal atrophy, lower urethral atrophy, sexual intercourse, vaginitis, cystitis, dysuria, urgency, concentration disorder, memory disorder, anxiety, nervousness, memory loss, dry skin, joint pain, osteoporosis, heart disease, Composition, characterized in that any one selected from the group consisting of hypertension, and stroke. A food composition for moisturizing the skin comprising a compound represented by Formula 2, or a salt thereof.
[Formula 2]

A cosmetic composition for moisturizing skin comprising a compound represented by Formula 2 below, or a cosmetically acceptable salt thereof.
[Formula 2]

A quasi-drug composition for moisturizing skin comprising a compound represented by Formula 2 below or a pharmaceutically acceptable salt thereof.
[Formula 2]

A compound represented by Formula 2 below; Or a food composition for fatigue improvement, inflammation improvement or antioxidant comprising a salt thereof.
[Formula 2]

A compound represented by Formula 2 below; Or a cosmetic composition for fatigue improvement, inflammation improvement or antioxidant comprising a cosmetically acceptable salt thereof.
[Formula 2]

A compound represented by Formula 2 below; Or a pharmaceutical composition for improving inflammation comprising a pharmaceutically acceptable salt thereof.
[Formula 2]

A compound represented by Formula 2 below; Or a quasi-drug composition for fatigue improvement, inflammation improvement or antioxidant comprising a pharmaceutically acceptable salt thereof.
[Formula 2]

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