Novel cyclopeptide and use thereof

Abstract

The present invention relates to a novel cyclopeptide and uses thereof. It was confirmed that when preadipocytes were treated with the novel cyclopeptide according to the present invention, the fat content was significantly reduced compared to differentiation-induced preadipocytes. Therefore, a food composition, a health functional food, a pharmaceutical composition, a quasi-drug composition, a cosmetic composition, a feed composition, and a feed additive composition comprising the cyclopeptide as an active ingredient can be usefully used for preventing, alleviating, or treating obesity.

Description

Novel cyclopeptides and their uses

[0001] The present invention relates to a novel cyclopeptide and its use.

[0002]

[0003] Obesity generally refers to an excess of adipose tissue in the body. It occurs when the energy consumed through food is not balanced with the energy expended through physical activity, resulting in the accumulation of excess energy as body fat. Obesity is known to be caused by a variety of factors, including genetics; environmental factors such as irregular eating habits, lack of exercise, or a Westernized diet; psychological factors such as depression, boredom, and excessive stress; and pathological factors such as changes in thyroid or adrenal hormones. Recently, economic growth and lifestyle changes have led to significant changes in eating habits. Busy modern people are increasingly becoming overweight and obese due to high-calorie diets like fast food and low exercise levels. Furthermore, because obesity is caused by adipocyte hypertrophy, inhibiting adipocyte differentiation may be beneficial in the treatment of obesity. Therefore, research on the inhibition of adipocyte differentiation and lipid accumulation has been reported. Treatment includes regular exercise, lifestyle modifications such as behavioral therapy, and medications such as appetite suppressants and fat absorption inhibitors.

[0004] However, most of these anti-obesity drugs are psychotropic medications that act on the central nervous system to regulate appetite. Consequently, they can cause side effects such as headaches and vomiting, and pose concerns about abuse. Therefore, research is urgently needed to develop safe and effective anti-obesity agents that can address the side effects of these commercially available anti-obesity drugs.

[0005] Against this backdrop, the inventors of the present invention have conducted extensive research efforts to develop a natural material useful for the prevention or treatment of obesity, and have obtained a novel cyclopeptide and confirmed its ability to inhibit adipocyte differentiation, thereby confirming its applicability as an anti-obesity material, thereby completing the present invention.

[0006]

[0007] The present invention aims to solve the above-mentioned problems and other problems related thereto.

[0008] An exemplary object of the present invention is a cyclopeptide represented by the following general formula I:

[0009] Cyclo-(ABCDEFG) (general formula I)

[0010] (In the above formula, A, B, C, D, E, F, and G are each independently proline (Pro: Proline), valine (Val: Valine), phenylalanine (Phe: Phenylalanine), isoleucine (Ile: Isoleucine), serine (Ser: Serine), isoleucine (Ile: Isoleucine), or glycine (Gly: Glycine))

[0011] is to provide.

[0012] Another exemplary object of the present invention is to provide a food composition for preventing or improving obesity, comprising the cyclopeptide.

[0013] Another exemplary object of the present invention is to provide a health functional food for preventing or improving obesity, comprising the cyclopeptide.

[0014] Another exemplary object of the present invention is to provide a pharmaceutical composition for preventing or treating obesity, comprising the cyclopeptide.

[0015] Another exemplary object of the present invention is to provide a pharmaceutical composition for preventing or improving obesity, comprising the cyclopeptide.

[0016] Another exemplary object of the present invention is to provide a cosmetic composition for preventing or improving obesity, comprising the cyclopeptide.

[0017] Another exemplary object of the present invention is to provide a feed composition for preventing or improving obesity, comprising the cyclopeptide.

[0018] Another exemplary object of the present invention is to provide a feed additive composition for preventing or improving obesity, comprising the cyclopeptide.

[0019] Another exemplary object of the present invention is to provide a method for preventing or treating obesity, comprising administering to a subject a composition comprising the cyclopeptide.

[0020] The technical problem to be achieved according to the technical idea of ​​the invention disclosed in this specification is not limited to the problem to solve the above-mentioned problem, and other problems not mentioned will be clearly understood by those skilled in the art from the description below.

[0021]

[0022] This is explained in detail as follows. Meanwhile, each description and embodiment disclosed in this application can also be applied to each other description and embodiment. In other words, all combinations of the various elements disclosed in this application fall within the scope of this application. Furthermore, the scope of this application is not limited by the specific descriptions described below.

[0023] As one aspect for achieving the above object, the present invention provides a cyclopeptide represented by the following general formula I:

[0024] Cyclo-(ABCDEFG) (general formula I)

[0025] (In the above formula, A, B, C, D, E, F and G each independently provide proline (Pro: Proline), valine (Val: Valine), phenylalanine (Phe: Phenylalanine), isoleucine (Ile: Isoleucine), serine (Ser: Serine), isoleucine (Ile: Isoleucine) or glycine (Gly: Glycine)).

[0026] In the present invention, the cyclopeptide may be compound 1 represented by the following chemical formula 1:

[0027] [Chemical Formula 1]

[0028] .

[0029] In the present invention, the term "cyclopeptide (CP)" refers to a cyclic peptide. The peptide refers to a molecule formed by amino acid residues being linked to each other by peptide bonds.

[0030] In the present invention, the cyclopeptide can be obtained through a chemical synthesis process or separated from the fruit of Asimina triloba, and there is no significant limitation on the separation and preparation method of the compound.

[0031] In one embodiment of the present invention, 11 fractions were obtained by powdering, extracting and concentrating the fruit of Asimina triloba, and among these, a cyclopeptide composed of proline (Pro: Proline), valine (Val: Valine), phenylalanine (Phe: Phenylalanine), isoleucine (Ile: Isoleucine), serine (Ser: Serine), isoleucine (Ile: Isoleucine), and glycine (Gly: Glycine) was isolated from fraction 6.

[0032]

[0033] In another aspect for achieving the above object, the present invention provides a method for producing a total extract, comprising the steps of: powdering the fruit of Asimina triloba, extracting with ethanol and concentrating under reduced pressure to obtain an extract; fractionating the extract into CH2Cl2, EtOAc, n-BuOH, and H2O in that order, and obtaining 11 fractions from the EtOAc fraction using medium pressure liquid chromatography (C18 reverse phase column); and extracting one fraction from the 11 fractions using a RP-C18 column (MeCN-H2O, 45:55, v / v) at a flow rate of 2.5 mL min. -1 The present invention provides a method for producing a cyclopeptide, comprising the step of performing isocratic semipreparative HPLC at a flow rate of , thereby obtaining a cyclopeptide.

[0034]

[0035] As another aspect for achieving the above purpose, the present invention provides a food composition for preventing or improving obesity, comprising the cyclopeptide.

[0036] The above “cyclopeptide” and the like may be within the aforementioned range.

[0037] In the present invention, the term "obesity" means a state in which there is excessive fat tissue in the body, and may further include various complications resulting from obesity.

[0038] In the present invention, the term "prevention" means any act of suppressing or delaying obesity or various complications caused by obesity by administering the composition of the present invention, and "improvement" in the present invention means showing an effect of alleviating obesity or various complications caused by obesity by applying the composition of the present invention.

[0039] In the present invention, prevention, treatment or improvement of obesity specifically means prevention, treatment, improvement, etc. of various specific symptoms or indicators related to obesity, and as an example, may include reduction of body fat, weight loss, reduction of body fat mass or body fat percentage, reduction of fatty liver, reduction of blood lipids, etc., and may also include inhibition of fat accumulation, inhibition of fat differentiation, inhibition of expression of fat synthesis factors, and reduction of the size of fat cells in adipose tissue.

[0040] In the present invention, the cyclopeptide may inhibit differentiation of adipocytes.

[0041] In the present invention, the term "adipocyte" refers to a cell that mainly constitutes adipose tissue that stores energy as fat, and adipose tissue is a central metabolic organ that regulates energy balance and homeostasis.

[0042] In the present invention, the differentiation of adipocytes refers to the conversion of preadipocytes into adipocytes. Furthermore, in the present invention, the differentiation of adipocytes may refer to the accumulation of fat within adipocytes, but is not limited thereto.

[0043] The differentiation of the above adipocytes may be achieved by suppressing the expression of transcription factors that regulate the expression of a series of genes involved in adipogenesis, which is the process by which preadipocytes proliferate and differentiate into mature adipocytes, but is not limited thereto.

[0044] In the present invention, the cyclopeptide may reduce the fat content of the fat crater.

[0045] In one embodiment of the present invention, in order to confirm the adipogenesis inhibitory effect of cyclopeptide (CP, cyclopeptide), cyclopeptide (CP) was treated to differentiated 3T3-L1 cells and the fat content was measured. As a result, it was confirmed that the fat content of the fat crater in the control group, differentiated 3T3-L1 cells (C), increased by more than 2.6 times compared to undifferentiated cells (N), and when treated with cyclopeptide (CP), the fat content was significantly reduced compared to differentiated 3T3-L1 cells regardless of the concentration, confirming that cyclopeptide (CP) has an excellent adipogenesis inhibitory effect even at a low concentration of 1 μM or less.

[0046] The food composition of the present invention includes all forms, including functional foods, nutritional supplements, health foods, food additives, health functional foods, and feeds, and is intended for consumption by humans or animals, including livestock. The above-mentioned types of food compositions can be manufactured in various forms using conventional methods known in the art.

[0047] The above type of food composition can be manufactured in various forms according to conventional methods known in the art. General foods include, but are not limited to, beverages (including alcoholic beverages), fruits and processed foods thereof (canned fruits, bottled fruits, jams, marmalades, etc.), fish, meats and processed foods thereof (ham, sausages, corned beef, etc.), breads and noodles (udon, buckwheat noodles, ramen, spagate, macaroni, etc.), fruit juices, various drinks, cookies, taffy, dairy products (butter, cheese, etc.), edible plant oils, margarine, vegetable proteins, retort foods, frozen foods, various seasonings (soybean paste, soy sauce, sauces, etc.), etc., and the above-mentioned effective ingredient can be added thereto to manufacture the composition.

[0048] Additionally, nutritional supplements can be manufactured by adding the above-mentioned active ingredient to capsules, tablets, pills, etc. Furthermore, health functional foods, although not limited thereto, can be manufactured in the form of tea, juice, and drinks, and consumed as health beverages by liquefaction, granulation, encapsulation, and powderization. Furthermore, the composition can be manufactured by mixing with one or more known active ingredients known to be effective in improving obesity or various complications caused by obesity.

[0049] In addition to the above, the health functional food of the present invention may contain various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid, salts of pectic acid, alginic acid, salts of alginic acid, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, or carbonating agents.

[0050]

[0051] As another aspect for achieving the above purpose, the present invention provides a health functional food for preventing or improving obesity, comprising the cyclopeptide.

[0052] The above “cyclopeptide”, “obesity”, “prevention”, “improvement”, etc. may be within the aforementioned range.

[0053] The term "health functional food" as used herein refers to a food manufactured and processed using raw materials or ingredients with beneficial functionality for the human body. The term "functionality" refers to achieving beneficial health effects, such as regulating nutrients or physiological effects on the structure and function of the human body. The health functional food of the present invention can be manufactured using methods commonly used in the art, and can be manufactured by adding raw materials and ingredients commonly added in the art during manufacturing.

[0054] In addition, the formulation of the above health functional food can be manufactured without limitation as long as it is a formulation recognized as a health functional food, and non-limiting examples of the formulation include one or more formulations selected from the group consisting of health functional food preparations such as tablets, capsules, pills, granules, liquids, powders, flakes, pastes, syrups, gels, jellies, bars, beverages, gums, and candies, but are not particularly limited thereto.

[0055]

[0056] As another aspect for achieving the above purpose, the present invention provides a pharmaceutical composition for preventing or treating obesity, comprising the cyclopeptide.

[0057] The above “cyclopeptide”, “obesity”, “prevention”, etc. may be within the aforementioned range.

[0058] In the present invention, the composition may be treated to preadipocytes at 0.01 μM to 65 μM, specifically at 0.05 μM to 60 μM, and more specifically at 0.08 μM to 55 μM, but is not limited thereto.

[0059] In the present invention, “treatment” means any act in which obesity or obesity-related complications are improved, alleviated, or beneficially changed by administering the pharmaceutical composition of the present invention.

[0060] The pharmaceutical composition of the present invention may further comprise a pharmaceutically acceptable carrier. Examples of such pharmaceutically acceptable carriers include carriers for oral administration or carriers for parenteral administration. Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate, stearic acid, and the like.

[0061] Additionally, carriers for parenteral administration may include water, suitable oils, saline, aqueous glucose, and glycols. They may also contain stabilizers and preservatives. Suitable stabilizers include antioxidants such as sodium bisulfite, sodium sulfite, or ascorbic acid. Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben, and chlorobutanol.

[0062] The pharmaceutical composition of the present invention can be administered to mammals, including humans, by any method. For example, it can be administered orally or parenterally, and parenteral administration methods can include intravenous, intramuscular, intraarterial, central, intramedullary, intrathecal, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual, or rectal administration, and preferably, central administration is used, but is not limited thereto.

[0063] The pharmaceutical composition of the present invention may be formulated as a preparation for oral or parenteral administration, depending on the route of administration as described above. When formulated, it may be prepared using one or more buffers (e.g., saline or PBS (phosphate buffered saline)), antioxidants, bacteriostatic agents, chelating agents (e.g., EDTA or glutathione), fillers, bulking agents, binders, adjuvants (e.g., aluminum hydroxide), suspending agents, thickening agents, wetting agents, disintegrating agents, surfactants, diluents, or excipients.

[0064] Solid preparations for oral administration include tablets, pills, powders, granules, liquids, gels, syrups, slurries, suspensions, capsules, etc., and these solid preparations can be prepared by mixing the pharmaceutical composition of the present invention with at least one excipient, for example, starch (including corn starch, wheat starch, rice starch, potato starch, etc.), calcium carbonate, sucrose, lactose, dextrose, sorbitol, mannitol, xylitol, erythritol maltitol, cellulose, methyl cellulose, sodium carboxymethylcellulose, and hydroxypropylmethyl-cellulose, or gelatin. For example, tablets or sugar-coated tablets can be obtained by mixing an active ingredient with a solid excipient, grinding the mixture, adding a suitable auxiliary agent, and then processing the mixture into a granule mixture.

[0065] In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral administration include suspensions, solutions, emulsions, and syrups. In addition to the commonly used simple diluents such as water or liquid paraffin, various excipients such as wetting agents, sweeteners, flavoring agents, or preservatives may be included. In addition, cross-linked polyvinylpyrrolidone, agar, alginic acid, or sodium alginate may be added as disintegrants, and anticoagulants, lubricants, wetting agents, flavoring agents, emulsifiers, and preservatives may be additionally included.

[0066] When administered parenterally, the pharmaceutical composition of the present invention may be formulated in the form of injections, transdermal administration agents, and nasal inhalants together with a suitable parenteral carrier according to methods known in the art. The injections must be sterilized and protected from contamination by microorganisms such as bacteria and fungi. Examples of suitable carriers for injections include, but are not limited to, solvents or dispersion media containing water, ethanol, polyols (e.g., glycerol, propylene glycol, and liquid polyethylene glycol), mixtures thereof, and / or vegetable oils. More preferably, suitable carriers include Hanks' solution, Ringer's solution, PBS containing triethanolamine, or isotonic solutions such as sterile water for injection, 10% ethanol, 40% propylene glycol, and 5% dextrose. To protect the injections from microbial contamination, various antibacterial and antifungal agents such as parabens, chlorobutanol, phenol, sorbic acid, and thimerosal may be additionally included. Additionally, the above injections may in most cases additionally contain isotonic agents such as sugar or sodium chloride.

[0067] Transdermal administration includes ointments, creams, lotions, gels, topical solutions, pastes, liniments, and aerosols. "Transdermal administration" here refers to topically administering a pharmaceutical composition to the skin, thereby delivering an effective amount of the active ingredient contained in the pharmaceutical composition into the skin.

[0068] For inhalation administration, the compositions used according to the present invention may conveniently be delivered in the form of an aerosol spray from a pressurized pack or nebulizer using a suitable propellant, such as dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, or another suitable gas. For pressurized aerosols, the dosage unit may be determined by providing a valve to deliver a metered amount. For example, gelatin capsules and cartridges for use in inhalers or insufflators may be formulated to contain a powder mixture of the compound and a suitable powder base such as lactose or starch. Formulations for parenteral administration are described in the well-known prescription book of pharmaceutical chemistry (Remington's Pharmaceutical Science, 15th Edition, 1975 Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Blaug, Seymour).

[0069] The pharmaceutical composition of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy or biological response modifiers.

[0070] The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. The pharmaceutically effective amount refers to an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment and not causing side effects. The effective dosage level may be determined based on factors including the patient's health condition, type and severity of the disease, drug activity and sensitivity to the drug, administration method, administration time, administration route and excretion rate, treatment period, combination or concurrent drug use, and other factors well known in the medical field. The dosage and frequency of administration do not limit the scope of the present invention in any way.

[0071] The pharmaceutical composition of the present invention can be administered to mammals, including rats, dogs, cats, cows, horses, pigs, and humans, via various routes, preferably to humans. Any route of administration is conceivable, including, but not limited to, oral, intravenous, intramuscular, or subcutaneous injection.

[0072]

[0073] As another aspect for achieving the above purpose, the present invention provides a pharmaceutical composition for preventing or improving obesity, comprising the cyclopeptide.

[0074] The above “cyclopeptide”, “obesity”, “prevention”, “improvement”, etc. may be within the aforementioned range.

[0075] The quasi-drug composition of the present invention may contain, in addition to the cyclopeptide, ingredients commonly used in quasi-drug compositions, such as, but not limited to, an abrasive, a humectant, a binder, a foaming agent, a sweetener, a preservative, a pharmaceutically active agent, a flavoring agent, a pigment, a solvent, a whitening agent, a solubilizer, or a pH adjuster.

[0076] In the present invention, the type of the above-mentioned quasi-drug is not particularly limited and may be any quasi-drug commonly used in the relevant technical field. Non-limiting examples of the above-mentioned quasi-drug include one or more formulations selected from the group consisting of liquids, ointments, sprays, creams, gels, patches, and aerogels. These may be used alone or in combination of two or more.

[0077]

[0078] As another aspect for achieving the above purpose, the present invention provides a cosmetic composition for preventing or improving obesity, comprising the cyclopeptide.

[0079] The above “cyclopeptide”, “obesity”, “prevention”, “improvement”, etc. may be within the aforementioned range.

[0080] In addition to the cyclopeptide, the cosmetic composition according to the present invention may be composed by blending various components generally used in cosmetic compositions, such as water-soluble components, powder components, oils, surfactants, moisturizers, viscosity modifiers, preservatives, antioxidants, fragrances, pigments, etc., within a range that does not impair the effects of the present invention, as needed.

[0081] Non-limiting examples of the surfactants that can be used include anionic surfactants, cationic surfactants, nonionic surfactants, and amphoteric surfactants. More specifically, examples of the anionic surfactants include alkylbenzene sulfonates, polyoxyalkylene alkyl sulfate ester salts, alkyl sulfate ester salts, olefin sulfonates, alkyl phosphates, polyoxyalkylene alkyl ether phosphates, dialkyl sulfosuccinates, and fatty acid salts, and examples of the nonionic surfactants include polyoxyethylene alkyl ethers, polyoxyethylene fatty acid esters, polyhydric alcohol fatty acid partial esters, polyoxyethylene polyhydric alcohol fatty acid partial esters, polyglycerin fatty acid esters, polyoxyethylene hydrogenated castor oil derivatives, and fatty acid diethanolamides. In addition, examples of cationic surfactants include tertiary aliphatic amine salts, alkyltrimethylammonium halides, dialkyldimethylammonium halides, etc., and examples of amphoteric surfactants include amide betaine type, imidazolinium betaine type, sulfo betaine type, etc.

[0082] Examples of the above-mentioned moisturizers include glycerin, propylene glycol, 1,3-butylene glycol, dipropylene glycol, sorbitol, etc. Examples of the above-mentioned preservatives include benzoic acid, dehydroacetic acid, parahydroxybenzoic acid esters (methyl parahydroxybenzoate, butyl parahydroxybenzoate, etc.), phenoxyethanol, etc. In addition, examples of the above-mentioned antioxidants include ascorbic acid, BHA, etc. In addition, ultraviolet absorbers, anti-inflammatory agents, and refreshing agents, etc. can be added.

[0083] The cosmetic composition of the present invention can be prepared in a formulation selected from the group consisting of, but not limited to, a solution, an external ointment, a cream, a foam, a nourishing toner, an emollient toner, a pack, an emollient, a milky lotion, a makeup base, an essence, a soap, a liquid cleanser, a bath agent, a sunscreen cream, a sun oil, a suspension, an emulsion, a paste, a gel, a lotion, a powder, a soap, a surfactant-containing cleansing, an oil, a powder foundation, an emulsion foundation, a wax foundation, a patch, and a spray.

[0084] The cosmetic composition of the present invention may additionally include one or more carriers acceptable for cosmetic formulations, and may appropriately mix conventional ingredients such as oil, water, surfactants, moisturizers, lower alcohols, thickeners, chelating agents, pigments, preservatives, fragrances, etc., but is not limited thereto. Here, the term "acceptable carrier for cosmetic formulations" refers to a compound or composition already known and used, or a compound or composition to be developed in the future, that can be included in cosmetic formulations and that does not have toxicity, instability, or irritation beyond what the human body can adapt to when in contact with the skin. The carrier may be included in the composition of the present invention in an amount of about 1 wt% to about 99.99 wt% based on the total weight of the composition, preferably about 90 wt% to about 99.99 wt% of the weight of the composition. However, since the above ratio varies depending on the formulation in which the composition of the present invention is prepared, its specific application site (face, neck, etc.), its preferred application amount, etc., the above ratio should not be construed as limiting the scope of the present invention in any way.

[0085] In the cosmetic formulation included in the cosmetic composition of the present invention, acceptable carriers vary depending on the formulation of the cosmetic composition.

[0086] When the formulation of the present invention is an ointment, paste, cream, or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, etc. may be used as carrier components, but are not limited thereto. These may be used alone or in combination of two or more.

[0087] When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, etc. can be used as a carrier component, and especially in the case of a spray, a propellant such as chlorofluorohydrocarbon, propane / butane, or dimethyl ether can be additionally included, but is not limited thereto, and these can be used alone or in a mixture of two or more.

[0088] When the formulation of the present invention is a solution or emulsion, a solvent, a solubilizer or an emulsifier may be used as a carrier component, for example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyl glycol oil, etc. may be used, and in particular, cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil, glycerol fatty acid ester, polyethylene glycol or fatty acid ester of sorbitan may be used, but is not limited thereto, and these may be used alone or in a mixture of two or more.

[0089] When the formulation of the present invention is a suspension, liquid diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tragacanth may be used as carrier components, but are not limited thereto, and these may be used alone or in a mixture of two or more.

[0090] When the formulation of the present invention is soap, alkali metal salts of fatty acids, fatty acid hemiester salts, fatty acid protein hydrolysates, isethionates, lanolin derivatives, fatty alcohols, vegetable oils, glycerol, sugars, etc. may be used as carrier components, but are not limited thereto, and these may be used alone or in a mixture of two or more.

[0091]

[0092] As another aspect for achieving the above purpose, the present invention provides a feed composition for preventing or improving obesity, comprising the cyclopeptide.

[0093] The above “cyclopeptide”, “obesity”, “prevention”, “improvement”, etc. may be within the aforementioned range.

[0094] The above feed composition may include feed additives. The feed additives of the present invention fall under the category of supplementary feed under the Feed Management Act.

[0095] In the present invention, the term "feed" may mean any natural or artificial diet, meal, etc. or a component of said meal, especially for eating, ingesting, and digesting by an animal or suitable therefor.

[0096] The type of the above feed is not particularly limited, and feed commonly used in the relevant technical field can be used. Non-limiting examples of the above feed include plant-based feed such as grains, roots, food processing by-products, algae, fiber, pharmaceutical by-products, oils, starches, meal, or grain by-products; and animal-based feed such as proteins, inorganic substances, oils, minerals, oils, single-cell proteins, zooplankton, or food. These may be used alone or in combination of two or more types.

[0097]

[0098] As another aspect for achieving the above purpose, the present invention provides a feed additive composition for preventing or improving obesity, comprising the cyclopeptide.

[0099] The above “cyclopeptide”, “obesity”, “prevention”, “improvement”, “feed”, etc. may be within the aforementioned range.

[0100]

[0101] As another aspect for achieving the above object, the present invention provides a method for preventing or treating obesity, comprising a step of administering to a subject a composition comprising the cyclopeptide.

[0102] As another aspect for achieving the above object, the present invention provides a use of the cyclopeptide for preventing, improving or treating obesity.

[0103] As another aspect for achieving the above object, the present invention provides the use of the cyclopeptide for the manufacture of a drug for preventing or treating obesity.

[0104] At this time, the above “cyclopeptide”, “obesity”, “prevention”, “improvement”, “treatment”, etc. may be within the above-mentioned range.

[0105]

[0106] When the novel cyclopeptide according to the present invention is treated to preadipocytes, it was confirmed that the fat content is significantly reduced compared to the differentiation-induced preadipocytes. Therefore, a food composition, a health functional food, a pharmaceutical composition, a quasi-drug composition, a cosmetic composition, a feed composition, and a feed additive composition containing the cyclopeptide as an active ingredient can be usefully used for the prevention, improvement, or treatment of obesity.

[0107]

[0108] Figure 1 is a diagram showing the separation process and MS analysis results of the cyclopeptide of the present invention.

[0109] Figures 2a to 2f are diagrams showing the results of NMR analysis of the cyclopeptide of the present invention.

[0110] Figure 3 is a diagram showing the structure of the cyclopeptide of the present invention (A is the structure of the compound, B is the 2D NMR correlation of the compound).

[0111] Figure 4 is a diagram showing the results of measuring cell survival rate by treating preadipocytes with the cyclopeptide of the present invention.

[0112] Figure 5 is a diagram showing the results of evaluating the fat production inhibition effect by treating preadipocytes with the cyclopeptide of the present invention.

[0113]

[0114] The present invention will be described in more detail below through examples. However, these examples are intended to exemplify the present invention and the scope of the present invention is not limited to these examples.

[0115]

[0116] Example 1. Separation of cyclopeptide (CP)

[0117] Asimina triloba was collected from Bonghwang Healing Farm, Geochang-gun, Gyeongsangnam-do, Republic of Korea. The unpeeled fruit (7 kg) was powdered, extracted with 100% EtOH at room temperature, and concentrated under reduced pressure (700.2 g). The whole extract was fractionated with CH2Cl2, EtOAc, n-BuOH, and H2O to obtain 26.2 g, 13.5 g, 353.3 g, and 612.3 g of extract, respectively. The EtOAc phase extract (13.5 g) was subjected to medium-pressure liquid chromatography (on a C18 reversed-phase column) to obtain 11 fractions (F1-F11). The F6 fraction (754 mg) was purified by RP-C18 column (MeCN-H2O, 45:55, v / v) at a flow rate of 2.5 mL min. -1 After isocratic semipreparative HPLC, compound 1 (17 mg) was obtained (Fig. 1).

[0118] Compound 1 was isolated as a brown amorphous powder, and the molecular formula of compound 1 was determined to be C based on HRESIMS data and NMR analysis. 36 H 55 It was confirmed to be N7O8, and the degree of unsaturation was confirmed to be 8 (Fig. 1 and Fig. 2a to Fig. 2f). 1 The H NMR spectrum (DMSO-d6) was confirmed to exhibit seven exchangeable amide H signals (δH 7.0-9.0), six amino acid α-proton signals (δH 3.0-5.0), five aromatic signals (δH 6.0-8.0), and 27 aliphatic methine, methyl, and methyl signals (δH 0.0-3.0) (Table 1).

[0119] residuepositionδ C , mult.δ H (J in Hz)residuepositionδ C , mult.δ H　(J in Hz)PheCO171.24, qCILECO170.28, qCα55.31, CH4.53,mα59.23, CH4.21, t (10.6)β38.58, CH23.17,dd (14.0, 8.7),β38.39, CH1.66, m2.67,dd(14.0,7.5)γ24.99, CH21.39, mγ138.12, qC1.05, mortho129.23, CH27.24,d(7.0)δ15.58, CH30.87, t (7.2)meta128.77, CH27.17,mβ-CH310.61, CH30.83, t (8.6)para126.92, CH7.27,mNH8.00,d(10.6)NH7.46,d(9.5)SERCO170.85, qCValCO169.74, qCα53.97, CH4.60, mα55.98, CH4.47,dd(8.5,9.2)β64.39, CH24.25, mβ32.13, CH1.97,m3.73, mγ19.59, CH30.85,d(6.5)NH8.66,d(7.2)19.03, CH30.85,d(6.5)ILECO170.42, qCNH7.04,d(9.5)α60.71, CH3.71, t (8.0)ProCO171.68, qCβ35.34, CH1.50, mα61.55, CH4.12,mγ24.24, CH20.87, mβ29.58, CH22.12,m0.83, m1.82,mδ12.12, CH30.59, t (6.5)γ24.96, CH22.03,mβ-CH315.43, CH30.49, d (7.2)1.93,mNH8.17,d(8.6)δ48.54, CH23.71,m3.64,mGlyCO169.11, qCα42.993.31, m4.11(dd,16.8,8.4)NH8.84

[0120] 화합물 1에 대한 13The C NMR spectrum showed seven distinguishable signals for amide-type carbonyls (δC 170.5-171.5), indicating the peptide nature of compound 1. Integrated analysis of COSY, HMQC, and HMBC NMR data confirmed that compound 1 is composed of seven amino acids: two isoleucines (Ile), phenylalanine (Phe), valine (Val), proline (Pro), glycine (Gly), and serine (Ser) (Fig. 3). The amino acid sequence was confirmed by HMBC and ROESY analyses, and the planar structure of compound 1 was confirmed to be a cyclopeptide with the sequence cyclo-(PRO-VAL-PHE-ILE-SER-ILE-GLY).

[0121]

[0122] Example 2. Analysis of the inhibitory effect of cyclopeptide (CP) on adipocyte differentiation in vitro.

[0123] 2-1. Cell culture and differentiation induction

[0124] To confirm the inhibitory effect of the cyclopeptide (CP) obtained from Example 1 on adipocyte differentiation, 3T3-L1 preadipocyte cells were cultured and differentiation was induced. Specifically, 3T3-L1 cells were obtained from the American Type Culture Collection (ATCC, USA) and cultured in Dulbecco's Modified Eagle Medium (DMEM, Gibco, USA) containing 10% bovine serum (BS, Gibco, USA) and 1% penicillin-streptomycin (PS, Gibco, USA) at 5% CO2 and 37°C.

[0125] For differentiation induction, 80% confluent cells were seeded in 6 wells at 1 x 10 using trypsin-EDTA. 5 Cells were seeded at 1 cell / well. After 24 h, when the cells became post-confluent, differentiation was induced by treating them with 0.5 mM 3-isobutyl-1-methylxanthine (IBMX, Sigma, USA), 10 μg / mL insulin (Sigma, USA), 10 μM dexamethasone (Sigma, USA), and 2 μM rosiglitazone (Sigma, USA) (IBMX, a mixture of insulin, dexamethasone, and rosiglitazone: denoted as MDI) in DMEM supplemented with 10% fetal bovine serum (FBS, Gibco, USA). Two days after differentiation induction, the medium was replaced with 10% FBS DMEM containing 10 μg / mL insulin, and the same medium was replaced on days 4 and 6 of differentiation induction to induce adipocyte differentiation for a total of 8 days.

[0126] Afterwards, all experiments were expressed as the mean and standard error of the mean for three repeated experiments, and the significance between each group was verified using Graphpad PRISM 7 (Graph pad software, USA) using one-way ANOVA and Turkey method, and a p value less than 0.05 (p<0.05) was considered statistically significant.

[0127]

[0128] 2-2. Cytotoxicity analysis

[0129] To determine whether cyclopeptide (CP) exhibits toxicity to preadipocytes, cell viability was measured using the MTT assay. Specifically, 2 × 10 3T3-L1 cells (Natural, N) that had not been induced to differentiate were seeded in a 96-well plate. 4Cells were seeded per well and treated with MDI and cyclopeptide (CP) 24 h later. After 48 h, 5 mg / mL of 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT, Sigma, USA) was added. After 1 h, the medium was removed and dimethylsulfoxide (DMSO) was added to dissolve the formazan produced. The absorbance was measured at 570 nm to evaluate cell viability. 3T3-L1 cells (Natural, N) that had not been induced to differentiate were used as a control.

[0130] As a result, the cell viability in the group treated with cyclopeptide (CP) was similar to or higher than that of the control group regardless of the concentration, confirming that cyclopeptide (CP) did not exhibit cytotoxicity. In addition, 3T3-L1 cells were cultured at 2 X 10 4 When 3T3-L1 cells (Control, C) were seeded at cell / well and differentiated 24 hours later with MDI, and then treated with cyclopeptide (CP) and the cell viability was evaluated 8 days later, the cell viability of the group treated with cyclopeptide (CP) was similar to that of the control group, differentiation-induced 3T3-L1 cells, regardless of the concentration, confirming that cyclopeptide (CP) is not cytotoxic (Fig. 4).

[0131]

[0132] 2-3. Analysis of inhibition of lipogenesis

[0133] In order to confirm the inhibitory effect of cyclopeptide (CP) on adipogenesis, 3T3-L1 cells induced to differentiate in 2-1 above were treated with cyclopeptide (CP) and then the fat content was measured. Specifically, 3T3-L1 cells induced to differentiate in 2-1 above were treated with cyclopeptide (CP) at concentrations of 0.1 μM, 0.5 μM, 1 μM, 5 μM, 12.5 μM, 25 μM, and 50 μM, respectively, and on the 8th day, the medium was removed, washed with phosphate-buffered saline (PBS, Gibco, USA), and fixed with 3.6% formaldehyde for 15 minutes. After washing three times with distilled water, the cells were treated with the filtered Oil Red O solution and stained for 15 minutes. The stained cells were washed three times with distilled water and photographed. The stained lipids within the cells were eluted with 100% isopropanol and transferred to a 96-well plate. The absorbance was measured at 490 nm to evaluate the degree of lipid accumulation.

[0134] As a result, it was confirmed that the fat content of the adipose tissue of the control group, differentiation-induced 3T3-L1 cells (C), increased by more than 2.6 times compared to undifferentiated cells (N), and when treated with cyclopeptide (CP), the fat content was significantly reduced compared to differentiation-induced 3T3-L1 cells regardless of the concentration, confirming that cyclopeptide (CP) has an excellent fat production inhibitory effect even at a low concentration of 1 μM or less (Fig. 5).

[0135]

[0136] From the above description, those skilled in the art will understand that the present invention can be implemented in other specific forms without altering its technical spirit or essential characteristics. In this regard, it should be understood that the embodiments described above are illustrative in all respects and not restrictive. The scope of the present invention should be interpreted as encompassing all changes or modifications derived from the meaning and scope of the following claims and their equivalent concepts, rather than the detailed description above.

Claims

1. Cyclopeptide represented by the following general formula I: Cyclo-(ABCDEFG) (general formula I) (In the above formula, A, B, C, D, E, F and G are each independently proline (Pro: Proline), valine (Val: Valine), phenylalanine (Phe: Phenylalanine), isoleucine (Ile: Isoleucine), serine (Ser: Serine), isoleucine (Ile: Isoleucine) or glycine (Gly: Glycine)).

2. In paragraph 1, The above cyclopeptide is a cyclopeptide represented by the following chemical formula 1: [Chemical Formula 1] .

3. A food composition for preventing or improving obesity, comprising the cyclopeptide of claim 1 or 2.

4. In paragraph 3, A food composition wherein the above cyclopeptide inhibits differentiation of fat cells.

5. In paragraph 3, A food composition wherein the above cyclopeptide has the ability to inhibit fat production.

6. A health functional food for preventing or improving obesity, comprising the cyclopeptide of paragraph 1 or 2.

7. A pharmaceutical composition for preventing or treating obesity, comprising the cyclopeptide of claim 1 or 2.

8. A pharmaceutical composition for preventing or improving obesity, comprising the cyclopeptide of claim 1 or 2.

9. A cosmetic composition for preventing or improving obesity, comprising the cyclopeptide of claim 1 or 2.

10. A feed composition for preventing or improving obesity, comprising the cyclopeptide of claim 1 or 2.

11. A feed additive composition for preventing or improving obesity, comprising the cyclopeptide of claim 1 or 2.

12. A method for preventing or treating obesity, comprising administering to a subject a composition comprising the cyclopeptide of claim 1 or 2.

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