Anti-inflammatory composition comprising lipid extract of oncorhynchus mykiss eggs as active ingredient

Abstract

The present invention relates to an anti-inflammatory composition comprising, as an active ingredient, a lipid extract of Oncorhynchus mykiss eggs. The anti-inflammatory activity of the lipid extract of Oncorhynchus mykiss eggs, of the present invention, was confirmed by analyzing the fatty acid composition and cytotoxicity of the lipid extract of Oncorhynchus mykiss eggs, and the effects thereof on inhibiting NO production, regulating the expression of cytokines, inhibiting NF-κB and MAPK activation, and inhibiting CD86 expression on the surface of immune cells.

Description

Anti-inflammatory composition containing rainbow trout roe lipid extract as an active ingredient

[0001] The present invention relates to an anti-inflammatory composition comprising rainbow trout roe lipid extract as an active ingredient.

[0002] One of the main components of fish roe is lipids and fatty acids, which provide metabolic energy and are found in fish cell membranes. Fish roe is generally known to contain high amounts of palmitic acid (C16:0), stearic acid (C18:0), oleic acid (C18:1 n-9), eicosapentaenoic acid (EPA, C20:5n-3), and docosahexaenoic acid (DHA, C22:6n-3). Furthermore, fish oil is known to play an important role in the treatment and prevention of cancer, diabetes, hypertension, arthritis, coronary artery disease, and other inflammatory and autoimmune diseases.

[0003] Furthermore, numerous animal and clinical studies have demonstrated that dietary supplementation with fish oil exhibits anti-inflammatory effects in conditions such as rheumatoid arthritis, asthma, inflammatory bowel disease, Crohn's disease, ulcerative colitis, psoriasis, lupus, multiple sclerosis, and migraines. Fish lipids are known to contain high levels of PUFAs, with the highest concentrations of DHA and EPA. These compounds have been shown to have beneficial effects on the host's defense system against infection, immune responses, and inflammatory processes.

[0004] Lipids extracted from natural sources have been demonstrated to exhibit antioxidant, antibacterial, antibiofilm, and anti-inflammatory activities. Many lipid extracts isolated from hard-shelled mussels, cyanobacteria, octopus, squid, Australian sardines, salmon, and school shrimp have demonstrated anti-inflammatory activity in vivo and in vitro, particularly in RAW264.7 macrophages, where they inhibited LPS-induced inflammatory responses.

[0005] In response to lipopolysaccharide (LPS), macrophages produce inflammatory cytokines, chemokines, nitric oxide (NO), and prostaglandin E2 (PGE2). Furthermore, they regulate the onset of inflammation through mediators such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and inflammatory cytokines such as interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor α (TNF-α). Furthermore, LPS stimulation induces inflammation through several intercellular signaling pathways, including nuclear factor-κB (NF-κB) and mitogen-activated protein kinases (MAPKs). The molecular mechanisms of anti-inflammatory effects may be activated by these pathways in in vitro animal cell systems.

[0006] Rainbow trout (Oncorhynchus mykiss) is a native fish found in marine and freshwater habitats of the northern Pacific Ocean, belonging to the salmonid family (Salmonidae) like Pacific salmon, and is frequently farmed for commercial aquaculture worldwide. They spend part of their lives in the ocean but return to lakes and rivers to spawn. Analysis of the fatty acid composition of O. mykiss eggs using solvent extraction revealed that the total lipid content of rainbow trout eggs consists of 46.4% polyunsaturated fatty acids (PUFAs) and 40.3% omega-3s. Most studies on rainbow trout have shown that dietary supplementation with natural extracts (n-3 highly unsaturated fatty acids (HUFAs), vitamin E, seaweed, mistletoe, Salvia officinalis oil, spearmint oil, and thyme oil) exhibits immunomodulatory, antioxidant, and immune-protective activities.

[0007] Additionally, rainbow trout peptides, hexane, histone H2A, and lipid extracts exhibited antioxidant, anticancer, antibacterial, and anti-obesity effects. However, the anti-inflammatory effects of rainbow trout lipids on immune cells have not yet been reported.

[0008] Therefore, the inventors of the present invention completed the present invention by analyzing the fatty acid composition, cytotoxicity, NO production inhibitory effect, cytokine expression regulating effect, NF-κB and MAPK activation inhibitory effect, and CD86 expression inhibitory effect on the surface of immune cells of the rainbow trout roe lipid extract, thereby confirming the anti-inflammatory activity of the rainbow trout roe lipid extract.

[0009] The purpose of the present invention is to provide an anti-inflammatory food composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0010] Another object of the present invention is to provide an anti-inflammatory health functional food composition comprising rainbow trout roe lipid extract as an active ingredient.

[0011] Another object of the present invention is to provide an anti-inflammatory pharmaceutical composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0012] Another object of the present invention is to provide an anti-inflammatory cosmetic composition comprising rainbow trout roe lipid extract as an active ingredient.

[0013] Another object of the present invention is to provide a method for preventing or treating inflammation, comprising a step of administering the pharmaceutical composition to a subject.

[0014] In order to achieve the above object of the present invention, the present invention provides an anti-inflammatory food composition comprising a rainbow trout roe lipid extract as an effective ingredient.

[0015] In addition, the present invention provides an anti-inflammatory health functional food composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0016] In addition, the present invention provides an anti-inflammatory pharmaceutical composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0017] In addition, the present invention provides an anti-inflammatory cosmetic composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0018] In addition, the present invention provides a method for preventing or treating inflammation, comprising a step of administering the pharmaceutical composition to a subject.

[0019] The present invention relates to an anti-inflammatory composition comprising a rainbow trout roe lipid extract as an active ingredient. By analyzing the fatty acid composition, cytotoxicity, NO production inhibitory effect, cytokine expression regulating effect, NF-κB and MAPK activation inhibitory effect, and CD86 expression inhibitory effect on the surface of immune cells of the rainbow trout roe lipid extract, the anti-inflammatory activity of the rainbow trout roe lipid extract of the present invention was confirmed, and the composition can be usefully used in related businesses.

[0020] Figure 1 is a diagram showing the cytotoxicity of a lipid extract of rainbow trout (O. mykiss) eggs in LPS-stimulated RAW264.7 cells.

[0021] Figure 2 is a diagram showing the change in NO production according to treatment with lipid extract of rainbow trout roe in LPS-stimulated RAW264.7 cells.

[0022] Figure 3 is a diagram showing changes in iNOS mRNA expression according to treatment with lipid extract of rainbow trout roe.

[0023] Figure 4 is a diagram showing the effect of lipid extract of rainbow trout roe on mRNA expression of TNF-α.

[0024] Figure 5 is a diagram showing the effect of lipid extract of rainbow trout roe on mRNA expression of IL-1β.

[0025] Figure 6 is a diagram showing the effect of lipid extract of rainbow trout roe on mRNA expression of IL-6.

[0026] Figure 7 is a diagram showing the effect of lipid extract of rainbow trout roe on mRNA expression of IL-10.

[0027] Figure 8 is a diagram showing the effect of lipid extract of rainbow trout roe on mRNA expression of IL-11.

[0028] Figure 9 is a diagram showing the effect of lipid extract of rainbow trout roe on mRNA expression of TGF-β.

[0029] Figure 10 is a diagram showing the results of Western blot analysis to confirm the effect of the lipid extract of rainbow trout roe on the phosphorylation of NF-κB and MAPK in LPS-stimulated RAW264.7 cells.

[0030] Figure 11 is a diagram showing the effect of specific inhibitors on TNF-α expression induced by lipid extract of rainbow trout roe in LPS-induced RAW264.7 cells.

[0031] Figure 12 is a diagram showing the effect of a lipid extract of rainbow trout roe on TNF-α expression in LPS-induced RAW264.7 cells by a specific inhibitor.

[0032] Figures 13 to 19 are diagrams showing the results of flow cytometry analysis on CD40 expression when rainbow trout roe lipid extract is treated in LPS-stimulated RAW264.7 cells.

[0033] Figure 20 is a diagram showing the mean fluorescence intensity (MFI) for CD40 expression according to flow cytometry.

[0034] Figures 21 to 27 are diagrams showing the results of flow cytometry analysis on CD486 expression when rainbow trout roe lipid extract is treated in RAW264.7 cells stimulated with LPS.

[0035] Figure 28 is a diagram showing the mean fluorescence intensity (MFI) for CD86 expression according to flow cytometry.

[0036] Hereinafter, the present invention will be described in detail with reference to the attached drawings, using exemplary embodiments of the present invention. However, the following exemplary embodiments are provided as illustrative examples of the present invention. If a detailed description of a technology or configuration well known to those skilled in the art is judged to unnecessarily obscure the gist of the present invention, such detailed description may be omitted, and the present invention is not limited thereby. The present invention is capable of various modifications and applications within the scope of the following claims and equivalents interpreted therefrom.

[0037] In addition, the terminology used in this specification is a term used to appropriately express preferred embodiments of the present invention, and this may vary depending on the intention of the user or operator, or the customs of the field to which the present invention belongs. Therefore, the definition of these terms should be determined based on the contents throughout this specification. Throughout the specification, when a part is said to "include" a certain component, this does not mean that other components are excluded, but rather that other components may be included, unless specifically stated otherwise.

[0038] The present invention provides an anti-inflammatory food composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0039] The extract according to the present invention can be obtained by extracting and separating from nature using extraction and separation methods known in the art, and the "extract" defined in the present invention is extracted from rainbow trout eggs using an appropriate solvent, and includes, for example, a crude extract, a polar solvent-soluble extract, or a non-polar solvent-soluble extract. Any pharmaceutically acceptable organic solvent may be used as a suitable solvent for extracting the extract from the rainbow trout eggs, and water or an organic solvent may be used, and is not limited thereto, for example, purified water, alcohols having 1 to 4 carbon atoms including methanol, ethanol, propanol, isopropanol, butanol, acetone, ether, benzene, chloroform, ethyl acetate, methylene chloride, hexane, and cyclohexane, and various solvents may be used alone or in combination. As an extraction method, any one of hot water extraction, cold immersion extraction, reflux cooling extraction, solvent extraction, steam distillation, ultrasonic extraction, dissolution, and pressing may be selected and used. Additionally, the desired extract may be subjected to additional conventional fractionation processes and purified using conventional purification methods.

[0040] There is no limitation on the method for preparing the extract of the present invention, and any known method can be used. For example, the extract included in the composition of the present invention can be prepared in a powder form by additional processes such as reduced pressure distillation and freeze drying or spray drying of the primary extract extracted by the above-mentioned hot water extraction or solvent extraction method. In addition, the primary extract can be further purified to obtain a fraction using various chromatography methods such as silica gel column chromatography, thin layer chromatography, high performance liquid chromatography, etc. Therefore, in the present invention, the extract is a concept that includes all extracts, fractions, and purified products obtained at each stage of extraction, fractionation, or purification, as well as their dilutions, concentrates, or dried products.

[0041] The food composition of the present invention may contain, in addition to containing the effective ingredient of the present invention, various flavoring agents or natural carbohydrates as additional ingredients, like conventional food compositions.

[0042] Examples of the above-mentioned natural carbohydrates include monosaccharides such as glucose, fructose, etc.; disaccharides such as maltose, sucrose, etc.; and polysaccharides such as dextrin, cyclodextrin, etc., and common sugars, and sugar alcohols such as xylitol, sorbitol, and erythritol. The above-mentioned flavoring agent can advantageously use natural flavoring agent (thaumatin), stevia extract (e.g., rebaudioside A, glycyrrhizin, etc.) and synthetic flavoring agent (saccharin, aspartame, etc.). The food composition of the present invention can be formulated in the same manner as the pharmaceutical composition and used as a functional food or added to various foods. Foods to which the composition of the present invention can be added include, for example, beverages, meat, chocolate, foods, confectionery, pizza, ramen, other noodles, gum, candy, ice cream, alcoholic beverages, vitamin complexes, and health supplements.

[0043] In addition, the food composition may contain, in addition to the active ingredient, various nutrients, vitamins, minerals (electrolytes), flavoring agents such as synthetic flavoring agents and natural flavoring agents, coloring agents and thickening agents (cheese, chocolate, etc.), pectic acid and its salts, alginic acid and its salts, organic acids, protective colloid thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonating agents used in carbonated beverages, etc. In addition, the food composition of the present invention may contain fruit pulp for producing natural fruit juice, fruit juice drinks, and vegetable drinks.

[0044] In one embodiment of the present invention, the rainbow trout egg lipid extract may be characterized in that it is extracted using a solvent selected from the group consisting of distilled water, C1 to C4 lower alcohol, lower alcohol aqueous solution, hexane, chloroform or acetic acid, but is not limited thereto.

[0045] In one embodiment of the present invention, the rainbow trout roe lipid extract may be characterized by including, but is not limited to, saturated fatty acids, monounsaturated fatty acids, or polyunsaturated fatty acids.

[0046] In one embodiment of the present invention, the saturated fatty acid may be at least one selected from the group consisting of myristic acid (C14:0), palmitic acid (C16:0), and stearic acid (C18:0), but is not limited thereto.

[0047] In one embodiment of the present invention, the monounsaturated fatty acid may be at least one selected from the group consisting of palmitoleic acid (C16:1n7), oleic acid (C18:1n9), and vaccenic acid (C18:1n7), but is not limited thereto.

[0048] In one embodiment of the present invention, the polyunsaturated fatty acid may be at least one selected from the group consisting of linoleic acid (C18:2n6), linolenic acid (C18:3n3), eicosadienonic acid (C20:2n6), eicosatrienonic acid (C20:3n3), eicosapentaenoic acid (EPA, C20:5n3, 7.77%), docosapentaenoic acid (DPA, C22:5n3), and docosahexaenoic acid (DHA, C22:6n3), but is not limited thereto.

[0049] In one embodiment of the present invention, the rainbow trout egg lipid extract may inhibit nitric oxide production, but is not limited thereto.

[0050] In one embodiment of the present invention, the rainbow trout egg lipid extract may suppress nitric oxide production by suppressing the expression of the iNOS gene, but is not limited thereto.

[0051] In one embodiment of the present invention, the rainbow trout roe lipid extract may suppress the production of inflammatory cytokines and promote the production of anti-inflammatory cytokines, but is not limited thereto.

[0052] In one embodiment of the present invention, the inflammatory cytokine may be at least one selected from the group consisting of tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), and interleukin-1 beta (IL-1β Interleukin-1 beta), but is not limited thereto.

[0053] In one embodiment of the present invention, the anti-inflammatory cytokine may be at least one selected from the group consisting of interleukin-10 (IL-10), interleukin-11 (IL-11), and transforming growth factor-beta (TGF-β), but is not limited thereto.

[0054] In one embodiment of the present invention, the rainbow trout egg lipid extract may have an activity of inhibiting a signal transduction pathway of Mitogen-Activated Protein Kinase (MAPK) or a signal transduction pathway of Nuclear Factor kappa B (NF-κB), but is not limited thereto.

[0055] In one embodiment of the present invention, the rainbow trout egg lipid extract may suppress the activation of immune cells, but is not limited thereto.

[0056] In one embodiment of the present invention, the rainbow trout egg lipid extract may suppress the activation of immune cells by reducing the expression of Cluster of Differentiation 86 (CD86) on the surface of immune cells, but is not limited thereto.

[0057] In addition, the present invention provides an anti-inflammatory health functional food composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0058] The functional food composition of the present invention can be manufactured and processed in the form of tablets, capsules, powders, granules, liquids, pills, etc. In the present invention, the term "health functional food composition" refers to a food manufactured and processed using raw materials or ingredients having functionality useful to the human body according to Act No. 6727 on Health Functional Foods, and means to be consumed for the purpose of obtaining a useful effect for health purposes such as regulating nutrients for the structure and function of the human body or physiological effects. The health functional food of the present invention may include conventional food additives, and whether it is suitable as a food additive is determined by the specifications and standards for the relevant item according to the general provisions and general test methods of the Food Additives Codex approved by the Ministry of Food and Drug Safety, unless otherwise specified. Items listed in the above "Food Additives Codex" include, for example, chemical compounds such as ketones, glycine, calcium citrate, nicotinic acid, and cinnamic acid; natural additives such as persimmon pigment, licorice extract, crystalline cellulose, sulphate pigment, and guar gum; Examples thereof include mixed preparations such as sodium L-glutamate preparations, noodle additive alkaline agents, preservative preparations, and tar color preparations. For example, health functional foods in tablet form can be prepared by mixing the active ingredient of the present invention with excipients, binders, disintegrants, and other additives, granulating the mixture using a conventional method, and then adding a lubricant, etc. to compress and molding the mixture, or directly compress and molding the mixture. In addition, the health functional foods in tablet form can contain a maturing agent, etc., if necessary. Among health functional foods in capsule form, hard capsules can be prepared by filling a mixture of the active ingredient of the present invention with additives such as excipients into a conventional hard capsule, and soft capsules can be prepared by filling a mixture of the active ingredient of the present invention with additives such as excipients into a capsule base such as gelatin. The soft capsules can contain a plasticizer such as glycerin or sorbitol, a coloring agent, a preservative, etc., if necessary.The ring-shaped health functional food can be prepared by molding a mixture of the active ingredient of the present invention with excipients, binders, disintegrants, etc. using a conventionally known method, and, if necessary, can be coated with white sugar or other coating agents, or the surface can be coated with a substance such as starch or talc. The granular health functional food can be manufactured into a granular form using a mixture of the active ingredient of the present invention with excipients, binders, disintegrants, etc. using a conventionally known method, and, if necessary, can contain a flavoring agent, a flavoring agent, etc.

[0059]

[0060] In addition, the present invention provides an anti-inflammatory pharmaceutical composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0061] The pharmaceutical composition of the present invention may further include an adjuvant in addition to the active ingredient. Any adjuvant known in the art may be used without limitation. However, for example, Freund's complete or incomplete adjuvant may be further included to increase immunogenicity.

[0062] The pharmaceutical composition according to the present invention can be prepared in a form in which the active ingredient is mixed with a pharmaceutically acceptable carrier. Here, the pharmaceutically acceptable carrier includes carriers, excipients, and diluents commonly used in the pharmaceutical field. Pharmaceutically acceptable carriers that can be used in the pharmaceutical composition of the present invention include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil.

[0063] The pharmaceutical composition of the present invention can be formulated and used in the form of oral formulations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, external preparations, suppositories, or sterile injection solutions, each according to a conventional method.

[0064] When formulated, it can be prepared using diluents or excipients such as fillers, bulking agents, binders, wetting agents, disintegrating agents, and surfactants that are commonly used. Solid preparations for oral administration include tablets, pills, powders, granules, and capsules, and such solid preparations can be prepared by mixing the active ingredient with at least one excipient, such as starch, calcium carbonate, sucrose, lactose, and gelatin. In addition to simple excipients, lubricants such as magnesium stearate and talc can also be used. Liquid preparations for oral administration include suspensions, oral solutions, emulsions, and syrups, and in addition to commonly used diluents such as water and liquid paraffin, various excipients such as wetting agents, sweeteners, fragrances, and preservatives can be included. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, and suppositories. Non-aqueous solvents and suspensions can include propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. Suppository bases include witepsol, Tween 61, cocoa butter, laurin, and glycerogelatin.

[0065] The pharmaceutical composition according to the present invention can be administered to a subject via various routes. All modes of administration are contemplated, including oral, intravenous, intramuscular, subcutaneous, and intraperitoneal injection.

[0066] The above pharmaceutical composition can be formulated into various oral or parenteral dosage forms.

[0067] Oral dosage forms include, for example, tablets, pills, hard and soft capsules, solutions, suspensions, emulsions, syrups, and granules. These dosage forms may further contain, in addition to the active ingredient, diluents (e.g., lactose, dextrose, sucrose, mannitol, sorbitol, cellulose, and / or glycine), lubricants (e.g., silica, talc, stearic acid and its magnesium or calcium salts, and / or polyethylene glycol). In addition, the tablets may contain binders such as magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose, and / or polyvinylpyrrolidine, and, if desired, disintegrants or effervescent mixtures such as starch, agar, alginic acid or its sodium salt, and / or absorbents, coloring agents, flavoring agents, and sweetening agents. The above formulation can be prepared by conventional mixing, granulating or coating methods.

[0068] In addition, representative parenteral administration formulations include injectable preparations, and solvents for injectable preparations include water, Ringer's solution, isotonic saline solution, or suspensions. Sterile fixed oils for the injectable preparations can be used as solvents or suspension media, and any non-irritating fixed oil, including mono- and di-glycerides, can be used for this purpose.

[0069] Additionally, the above injectable formulation may use a fatty acid such as oleic acid.

[0070]

[0071] In addition, the present invention provides an anti-inflammatory cosmetic composition comprising a rainbow trout roe lipid extract as an active ingredient.

[0072] The composition comprising the rainbow trout roe lipid extract of the present invention can be used in various ways, such as for the prevention or improvement of skin diseases. Products to which the composition can be added include, for example, cosmetics such as various creams, lotions, skin toners, and essences, as well as shampoos, rinses, cleansers, facial cleansers, soaps, treatments, packs, and beauty solutions.

[0073] The cosmetic of the present invention comprises a composition selected from the group consisting of water-soluble vitamins, oil-soluble vitamins, high molecular weight peptides, high molecular weight polysaccharides, sphingolipids, and seaweed extracts.

[0074] Any water-soluble vitamin that can be mixed into cosmetics may be used, but preferably, vitamin B1, vitamin B2, vitamin B6, pyridoxine, pyridoxine hydrochloride, vitamin B12, pantothenic acid, nicotinic acid, nicotinamide, folic acid, vitamin C, vitamin H, etc., and their salts (thiamine hydrochloride, sodium ascorbate, etc.) or derivatives (sodium ascorbate-2-phosphate, magnesium ascorbate-2-phosphate, etc.) are also included in the water-soluble vitamins that can be used in the present invention. Water-soluble vitamins can be obtained by conventional methods such as microbial transformation, purification from microbial culture, enzymatic method, or chemical synthesis method.

[0075] Any vitamin that can be incorporated into cosmetics may be used as the useful vitamin, but preferably vitamin A, carotene, vitamin D2, vitamin D3, vitamin E (d1-alpha tocopherol, d-alpha tocopherol, d-alpha tocopherol), and their derivatives (ascorbic acid palmitate, ascorbic acid stearate, ascorbic acid dipalmitate, dl-alpha tocopherol acetate, dl-alpha tocopherol nicotinate, vitamin E, DL-pantothenyl alcohol, D-pantothenyl alcohol, pantothenyl ethyl ether, etc.) are also included in the useful vitamins used in the present invention. The useful vitamins can be obtained by conventional methods such as microbial transformation, purification from a microbial culture, enzymatic or chemical synthesis, etc.

[0076] Any polymer peptide that can be incorporated into cosmetics may be used, but preferred examples include collagen, hydrolyzed collagen, gelatin, elastin, hydrolyzed elastin, and keratin. Polymer peptides can be purified and obtained by conventional methods such as purification from microbial cultures, enzymatic methods, or chemical synthesis methods, or can be purified and used from natural products such as the dermis of pigs or cows, or silkworm fibers.

[0077] Any polymer polysaccharide that can be incorporated into cosmetics may be used, but preferred examples include hydroxyethyl cellulose, xanthan gum, sodium hyaluronate, chondroitin sulfate, or a salt thereof (e.g., sodium salt). For example, chondroitin sulfate or a salt thereof can usually be purified from mammals or fish and used.

[0078] Any sphingolipid that can be incorporated into cosmetics may be used, but preferred examples include ceramide, phytosphingosine, and sphingoglycolipids. Sphingolipids can be purified by conventional methods from mammals, fish, shellfish, yeast, or plants, or obtained by chemical synthesis.

[0079] Any seaweed extract that can be mixed into cosmetics may be used, but preferably, brown seaweed extract, red seaweed extract, green seaweed extract, etc. are used. In addition, calagenan, arginic acid, sodium arginate, potassium arginate, etc. purified from these seaweed extracts are also included in the seaweed extract used in the present invention. Seaweed extract can be obtained by purifying seaweed using a conventional method.

[0080] In addition to the above essential ingredients, the cosmetic of the present invention may contain other ingredients commonly contained in cosmetics, if necessary. In addition, examples of ingredients that may be added include fat components, moisturizers, emollients, surfactants, organic and inorganic pigments, organic powders, ultraviolet absorbers, preservatives, bactericides, antioxidants, plant extracts, pH adjusters, alcohols, pigments, fragrances, blood circulation promoters, cooling agents, antiperspirants, purified water, and the like. Examples of fat components include ester-based fats, hydrocarbon-based fats, silicone-based fats, fluorine-based fats, animal fats, and plant fats.

[0081] As ester-based fats and oils, tri-2-ethylhexanoate glyceryl, cetyl 2-ethylhexanoate, isopropyl myristate, butyl myristate, isopropyl palmitate, ethyl stearate, octyl palmitate, isocetyl isostearate, butyl stearate, ethyl linoleate, isopropyl linoleate, ethyl oleate, isocetyl myristate, isostearyl myristate, isostearyl palmitate, octyldodecyl myristate, isocetyl isostearate, diethyl sebacate, diisopropyl adipate, isoalkyl neopentanoate, tri(caprylic, capric) glyceryl, trimethylolpropane tri-2-ethylhexanoate, trimethylolpropane triisostearate, tetra-2-ethylhexanoate pentaellistolate, cetyl caprylate, decyl laurate, Hexyl laurate, decyl myristate, myristyl myristate, cetyl myristate, stearyl stearate, decyl oleate, cetyl ricinoleate, isostearyl laurate, isotridecyl myristate, isocetyl palmitate, octyl stearate, isocetyl stearate, isodecyl oleate, octyldodecyl oleate, octyldodecyl linoleate, isopropyl isostearate, cetostearyl 2-ethylhexanoate, stearyl 2-ethylhexanoate, hexyl isostearate, ethylene glycol dioctanoate, ethylene glycol dioleate, propylene glycol dicaprate, di(caprylic, capric) propylene glycol, propylene glycol dicaprylate, neopentyl glycol dicaprate, neopentyl glycol dioctanoate, glyceryl tricaprylate, Glyceryl triundecyl acid, glyceryl triisopalmitate, glyceryl triisostearate, octyldodecyl neopentanoate, isostearyl octanoate, octyl isononanoate, hexyldecyl neodecanoate, octyldodecyl neodecanoate, isocetyl isostearate, isostearyl isostearate, octyldecyl isostearate, polyglycerol oleate ester, polyglycerol isostearate ester, triisocetyl citrate, triisoalkyl citrate, triisooctyl citrate, lauryl lactate, myristyl lactate, cetyl lactate, octyldecyl lactate, triethyl citrate, acetyl triethyl citrate, acetyl tributyl citrate, trioctyl citrate, diisostearyl malate, hydroxystearic acid 2-Ethylhexyl, di-2-ethylhexyl succinate, diisobutyl adipate, diisopropyl sebacate,Examples include esters such as dioctyl sebacate, cholesterol stearate, cholesteryl isostearate, cholesterol hydroxystearate, cholesterol oleate, dihydrocholesteryl oleate, phytosteryl isostearate, phytosteryl oleate, isocetyl 12-stearoylhydroxystearate, stearyl 12-stearoylhydroxystearate, and isostearyl 12-stearoylhydroxystearate.

[0082] Examples of hydrocarbon-based fats include squalene, liquid paraffin, alpha-olefin oligomer, isoparaffin, ceresin, paraffin, liquid isoparaffin, polybutene, microcrystalline wax, and vaseline.

[0083] Examples of silicone-based oils include polymethylsilicone, methylphenylsilicone, methylcyclopolysiloxane, octamethylpolysiloxane, decamethylpolysiloxane, dodecamethylcyclosiloxane, dimethylsiloxane and methylcetyloxysiloxane copolymers, dimethylsiloxane and methyl stearoxysiloxane copolymers, alkyl-modified silicone oils, and amino-modified silicone oils.

[0084] Examples of fluorine-based oils include perfluoropolyether.

[0085] Examples of animal or plant fats include avocado oil, almond oil, olive oil, sesame oil, rice bran oil, sunflower oil, soybean oil, corn oil, rapeseed oil, almond oil, palm kernel oil, palm oil, castor oil, sunflower oil, grape seed oil, cottonseed oil, coconut oil, kukui nut oil, wheat germ oil, rice germ oil, shea butter, colostrum oil, marc deimia nut oil, meadowsweet oil, egg yolk oil, beef tallow, horse oil, mink oil, orange rapeseed oil, jojoba oil, candelilla wax, carnauba wax, liquid lanolin, and hydrogenated castor oil.

[0086] Examples of moisturizers include water-soluble low-molecular-weight moisturizers, fat-soluble molecular-weight moisturizers, water-soluble polymers, and fat-soluble polymers.

[0087] Examples of water-soluble low-molecular-weight moisturizers include serine, glutamine, sorbitol, mannitol, sodium pyrrolidone-carboxylate, glycerin, propylene glycol, 1,3-butylene glycol, ethylene glycol, polyethylene glycol B (polymerization degree n = 2 or higher), polypropylene glycol (polymerization degree n = 2 or higher), polyglycerin B (polymerization degree n = 2 or higher), lactic acid, and lactate salts.

[0088] Examples of fat-soluble low-molecular-weight moisturizers include cholesterol and cholesterol esters.

[0089] Examples of water-soluble polymers include carboxyvinyl polymer, polyaspartate, tragacanth, xanthan gum, methylcellulose, hydroxymethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, carboxymethylcellulose, water-soluble chitin, chitosan, and dextrin.

[0090] Examples of lipid-soluble polymers include polyvinylpyrrolidone and eicosene copolymers, polyvinylpyrrolidone and hexadecene copolymers, nitrocellulose, dextrin fatty acid esters, and high molecular silicones. Examples of emollients include long-chain acylglutamic acid cholesteryl ester, cholesteryl hydroxystearate, 12-hydroxystearic acid, stearic acid, rosin acid, and lanolin fatty acid cholesteryl ester.

[0091] Examples of surfactants include nonionic surfactants, anionic surfactants, cationic surfactants, and amphoteric surfactants.

[0092] Nonionic surfactants include self-emulsifying monostearate glycerin, propylene glycol fatty acid ester, glycerin fatty acid ester, polyglycerin fatty acid ester, sorbitan fatty acid ester, POE (polyoxyethylene) sorbitan fatty acid ester, POE sorbitan fatty acid ester, POE glycerin fatty acid ester, POE alkyl ether, POE fatty acid ester, POE hydrogenated castor oil, POE castor oil, POE and POP (polyoxyethylene and polyoxypropylene) copolymer, POE and POP alkyl ether, polyether-modified silicone, lauric acid alkanolamide, alkylamine oxide, hydrogenated soybean phospholipid, etc.

[0093] Examples of anionic surfactants include fatty acid soaps, alpha-acyl sulfonates, alkyl sulfonates, alkyl allyl sulfonates, alkyl naphthalene sulfonates, alkyl sulfates, POE alkyl ether sulfates, alkyl amide sulfates, alkyl phosphates, POE alkyl phosphorus acids, alkyl amide phosphates, alkyloyl alkyl taurine salts, N-acyl amino acids, POE alkyl ether carboxylates, alkyl sulfosuccinates, sodium alkyl sulfoacetate, acylated hydrolyzed collagen peptide salts, and perfluoroalkyl phosphate esters.

[0094] Examples of cationic surfactants include alkyl trimethyl ammonium chloride, stearyl trimethyl ammonium chloride, stearyl trimethyl ammonium bromide, cetostearyl trimethyl ammonium chloride, distearyl dimethyl ammonium chloride, stearyl dimethyl benzylammonium chloride, behenyl trimethyl ammonium bromide, benzalkonium chloride, diethylaminoethyl amide stearate, dimethylaminopropyl amide stearate, and quaternary ammonium salts of lanolin derivatives. Examples of amphoteric surfactants include carboxy betaine type, amide betaine type, sulfo betaine type, hydroxysulfo betaine type, amide sulfo betaine type, phospho betaine type, aminocarboxylate type, imidazoline derivative type, and amide amine type.

[0095] Organic and inorganic pigments include inorganic pigments such as silicic acid, silicic anhydride, magnesium silicate, talc, sericite, mica, kaolin, bengala, clay, bentonite, titanium-coated mica, bismuth oxychloride, zirconium oxide, magnesium oxide, zinc oxide, titanium oxide, aluminum oxide, calcium sulfate, barium sulfate, magnesium sulfate, calcium carbonate, magnesium carbonate, iron oxide, ultramarine, chromium oxide, chromium hydroxide, calamine, and complexes thereof; Examples thereof include polyamide, polyester, polypropylene, polystyrene, polyurethane, vinyl resin, urea resin, phenol resin, fluororesin, silicon resin, acrylic resin, melamine resin, epoxy resin, polycarbonate resin, divinylbenzene and styrene copolymers, silk powder, cellulose, CI pigment yellow, CI pigment orange, and composite pigments of inorganic pigments and organic pigments.

[0096] Examples of organic powders include metal soaps such as calcium stearate; alkyl phosphate metal salts such as sodium zinc cetylphosphate, zinc laurylate, and calcium laurylate; acylamino acid polyvalent metal salts such as calcium N-lauroyl-beta-alanine, zinc N-lauroyl-beta-alanine, and calcium N-lauroylglycine; amide sulfonic acid polyvalent metal salts such as calcium N-lauroyl-taurine and calcium N-palmitoyl-taurine; N-acyl basic amino acids such as N-epsilon-lauroyl-L-lysine, N-epsilon-palmitoyl lysine, N-alpha-paritoyl olnitine, N-alpha-lauroyl arginine, and N-alpha-hydrogenated beef tallow fatty acid acylarginine; N-acyl polypeptides such as N-lauroyl glycyl glycine; Alpha-amino fatty acids such as alpha-aminocaprylic acid and alpha-aminolauric acid; polyethylene, polypropylene, nylon, polymethyl methacrylate, polystyrene, divinylbenzene and styrene copolymers, and ethylene tetrafluoride.

[0097] UV absorbers include para-aminobenzoic acid, para-aminobenzoic acid ethyl, para-aminobenzoic acid amyl, para-aminobenzoic acid octyl, ethylene glycol salicylate, phenyl salicylate, octyl salicylate, benzyl salicylate, butylphenyl salicylate, homomenthyl salicylate, benzyl cinnamic acid, 2-ethoxyethyl para-methoxycinnamic acid, octyl para-methoxycinnamic acid, mono-2-ethylhexaneglyceryl dipara-methoxycinnamic acid, isopropyl para-methoxycinnamic acid, diisopropyl and diisopropyl cinnamic acid ester mixtures, urocanic acid, ethyl urocanic acid, hydroxymethoxybenzophenone, hydroxymethoxybenzophenonesulfonic acid and its salts, dihydroxymethoxybenzophenone, sodium dihydroxymethoxybenzophenone disulfonate, dihydroxybenzophenone, Examples include tetrahydroxybenzophenone, 4-tert-butyl-4'-methoxydibenzoylmethane, 2,4,6-trianilino-p-(carbo-2'-ethylhexyl-1'-oxy)-1,3,5-triazine, and 2-(2-hydroxy-5-methylphenyl)benzotriazole.

[0098] Examples of disinfectants include hinokitiol, triclosan, trichlorohydroxydiphenyl ether, chlorhexidine gluconate, phenoxyethanol, resorcin, isopropylmethylphenol, azulene, salicylic acid, zincpyrithione, benzalkonium chloride, photosensitive agent No. 301, sodium mononitroguaiacol, and undecylenic acid.

[0099] Antioxidants include butylated hydroxyanisole, propyl gallic acid, and ellisorbic acid.

[0100] Examples of pH adjusters include citric acid, sodium citrate, malic acid, sodium malate, formaldehyde, sodium formaldehyde, succinic acid, sodium succinate, sodium hydroxide, and sodium hydrogen phosphate.

[0101] As for alcohol, we can mention high-grade alcohols such as cetyl alcohol.

[0102] In addition, the compounding ingredients that may be added are not limited to these, and any of the above ingredients may be compounded within a range that does not impair the purpose and effect of the present invention.

[0103] The cosmetic of the present invention can take the form of a solution, an emulsion, a viscous mixture, etc.

[0104] The ingredients included in the cosmetic composition of the present invention may include ingredients commonly used in cosmetic compositions as active ingredients, and include, for example, conventional auxiliary agents and carriers such as stabilizers, solubilizers, vitamins, pigments, and fragrances.

[0105] The cosmetic composition of the present invention can be manufactured in any formulation commonly manufactured in the art, and examples thereof include emulsion, cream, toner, pack, foundation, lotion, cosmetic solution, hair cosmetic, etc.

[0106] Specifically, the cosmetic composition of the present invention includes formulations of skin lotion, skin softener, skin toner, milk lotion, astringent, lotion, moisture lotion, nutrition lotion, massage cream, nutrition cream, moisture cream, hand cream, foundation, essence, nutrition essence, pack, soap, cleansing foam, cleansing lotion, cleansing cream, hair lotion, hair tonic, hair essence, hair shampoo, hair rinse, hair treatment, body lotion, and body cleanser.

[0107] When the formulation of the present invention is a paste, cream or gel, animal fiber, plant fiber, wax, paraffin, starch, tragacanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide may be used as a carrier component.

[0108] When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, and particularly in the case of a spray, a propellant such as chlorofluorohydrocarbon, propane / butane or dimethyl ether may be additionally included.

[0109] In the case where the formulation of the present invention is a solution or emulsion, a solvent, solvating agent or emulsifying agent is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or fatty acid ester of sorbitan.

[0110] 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.

[0111] When the formulation of the present invention is a surfactant-containing cleansing agent, aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyl taurate, sarcosinate, fatty acid amide ether sulfate, alkylamidobetaine, fatty alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, linolenic derivative, or ethoxylated glycerol fatty acid ester may be used as a carrier component.

[0112] In addition, the present invention provides a method for preventing or treating inflammation, comprising a step of administering the pharmaceutical composition to a subject.

[0113] The term "subject" as used in the present invention refers to a subject requiring a method for preventing, controlling, or treating a disease, and may be used without limitation as a human, dog, monkey, cat, rodent, such as a mouse, genetically modified mouse, etc. More specifically, it refers to a mammal such as a human or non-human primate, mouse, rat, dog, cat, horse, cow, etc.

[0114] The pharmaceutical composition of the present invention can be administered in a therapeutically effective amount or a pharmaceutically effective amount.

[0115] In the present invention, the term "therapeutically effective amount" means the amount of a pharmaceutically acceptable salt of a composition effective in preventing or treating a target disease, and the therapeutically effective amount of the composition of the present invention may vary depending on various factors, such as the administration method, the target site, the condition of the patient, etc. Therefore, the dosage for use in humans should be determined as an appropriate amount by taking both safety and efficacy into consideration. It is also possible to estimate the amount to be used in humans from the effective amount determined through animal testing. Such considerations in determining the effective amount are described, for example, in Hardman and Limbird, eds., Goodman and Gilman's The Pharmacological Basis of Therapeutics, 10th ed. (2001), Pergamon Press; and E.W. Martin ed., Remington's Pharmaceutical Sciences, 18th ed. (1990), Mack Publishing Co.

[0116] In the present invention, the term “pharmaceutically effective amount” means an amount sufficient to treat a disease at a reasonable benefit / risk ratio applicable to medical treatment and not causing side effects, and the effective dosage level can be determined according to factors including the patient’s health condition, type and severity of the disease, activity of the drug, sensitivity to the drug, administration method, administration time, administration route and excretion rate, treatment period, combination or concurrent use of drugs, and other factors well known in the medical field. The composition of the present invention can be administered as an individual therapeutic agent or in combination with other therapeutic agents, can be administered sequentially or simultaneously with conventional therapeutic agents, and can be administered singly or in multiple doses. Taking all of the above factors into consideration, it is important to administer an amount that can achieve the maximum effect with the minimum amount without side effects, and this can be easily determined by those skilled in the art.

[0117] Hereinafter, the present invention will be described in more detail through examples. These examples are intended to more specifically illustrate the present invention, and the scope of the present invention is not limited to these examples.

[0118]

[0119] <Preparation Example 1> Rainbow Trout Sample Preparation

[0120] Rainbow trout (Oncorhynchus mykiss) were purchased from a market in Pyeongchang, Gangwon Province, South Korea. The roe was harvested from the rainbow trout and immediately freeze-dried. The eggs were then immediately frozen and stored at -20 °C.

[0121]

[0122] <Preparation Example 2> Rainbow Trout Egg Lipid Extraction

[0123] In the above Preparation Example 1, the freeze-dried rainbow trout roe was powdered, and the lipids from the powdered frozen tissue (4.5 g) were extracted using a 1:2 mixture of chloroform and methanol. The extraction solution was homogenized, centrifuged, and filtered. The organic solvent was evaporated using a rotary evaporator (IKA RV 10-digital) and a nitrogen evaporator (12 position N-EVAP nitrogen evaporator, N-EVAP, USA), and the lipids to be used in cell culture experiments were extracted using a dimethyl sulfoxide (DMSO) solution.

[0124]

[0125] <Example 1> Analysis method

[0126] 1-1. Fatty acid composition analysis

[0127] To prepare fatty acid methyl esters (FAMEs), hydrolysis, extraction, and methylation procedures were performed. In addition, GC-FID analysis was performed using a 7890A (Agilent Technologies, CA, USA) coupled to a FID and an Agilent J&W GC column (30 m Х 0.32 mm ID, 0.25 μm film; Agilent Technologies, CA, USA).

[0128] 1 μL of the rainbow trout roe lipid extract sample obtained through Preparation Examples 1 and 2 was injected at a split ratio of 1:5, and the temperatures of the inlet and detector were maintained at 250°C. The initial oven temperature was maintained at 150°C for 2 min and then increased at a rate of 3.5°C / min to a final temperature of 230°C. FAMEs were identified by analyzing the retention times of standard compounds.

[0129]

[0130] 1-2. Cell culture and sample processing

[0131] RAW 264.7 cells (Korean Cell Line Bank, Korea) and macrophage cell lines were cultured in RPMI-1640 medium (Gibco, USA) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin / streptomycin (Welgene, Korea). RAW 264.7 cells were maintained at 37°C in a 5% carbon dioxide (CO2) incubator and passaged every 2–3 days. Rainbow trout roe lipid extract dissolved in DMSO, supplemented with 1% FBS and 1% penicillin / streptomycin, was diluted in RPMI 1640 medium without phenol red. RAW 264.7 cells were pretreated with various concentrations of rainbow trout roe lipid extract (100, 200, 300, and 400 μg / mL), and aspirin (200 μg / mL) was also pretreated as a positive control. After culturing the cells for 1 h, the cells were activated for an additional 24 h with or without LPS (1 μg / mL), and then evaluated for their anti-inflammatory capacity.

[0132]

[0133] 1-3. Cell proliferation analysis

[0134] 1 Х 10 6 The viability of cells treated with rainbow trout roe lipid extract samples at a density of cells / mL was measured using the EZ-Cytox Cell Viability Assay Kit (Daeil Lab Service Co., Korea). In this assay, the supernatant was removed, and 100 μL of WST solution was added to each well. The cells were incubated for 1 hour, and the absorbance was measured at 450 nm using an EPOCH 2 microplate reader from Agilent BioTek.

[0135]

[0136] 1-4. Measurement of NO production

[0137] To measure NO production, the Griess reagent (Promega, USA) was used. 100 μL of the supernatant from cells treated with rainbow trout roe lipid extract was transferred to another plate. Greiss reagent A (1% sulfanilamide in 5% phosphoric acid) and Greiss reagent B (0.1% N-1-napthylethylenediamine dihydrochloride in water) were added to each well and incubated for 10 minutes. Absorbance was measured at 540 nm using a microplate reader.

[0138]

[0139] 1-5. Quantitative real-time PCR (qRT-PCR)

[0140] Cell density 1 Х 10 6 Total RNA was extracted using TRI reagent (Molecular Research Center, Inc., Cincinnati, OH, USA) at 100 cells / mL. After RNA extraction, cDNA was synthesized using a High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems, Waltham, MA, USA). 20 μL of reaction solution was mixed with 10 μL of TB Green Premix Ex Taq II (Takara Bio Inc., Kusatsu, Japan), 6 μL of DEPC-treated water, 2 μL (5 ng) of cDNA, 0.4 μL of ROX Reference Dye, and 0.8 μL of forward and reverse primers, and qRT-PCR was performed for quantitative analysis. RNA amplification was performed using a QuantStudio 3 FlexReal-Time PCR System (Applied Biosystems, Waltham, MA, USA), and the expression of the target gene was normalized using β-actin as an internal standard.

[0141]

[0142] 1-6. Western Blot Analysis

[0143] After treatment with rainbow trout roe lipid extract or aspirin, cells were stimulated with LPS, and proteins were extracted using RIPA buffer (Tech & Innovation, Hebei, China) containing 0.5 mM EDTA solution and 0.1% protease and phosphatase inhibitor mixture (Thermo Fisher Scientific, Waltham, MA, USA). Equal amounts of proteins for each treatment group were separated using SDS-polyacrylamide gels and transferred to polyvinylidene fluoride membranes (Merck, Kenilworth, NJ, USA). The polyvinylidene fluoride membrane was incubated overnight at 4°C with primary antibodies specific for phospho-p44 / 42 MAPK (Erk1 / 2), phospho-SAPK / JNK, phospho-p38 MAPK, phospho-NF-κB p65 (Cell Signaling Technology, Danvers, MA, USA), and α-tubulin (Abcam, Cambridge, UK). After incubation with the primary antibodies, the membrane was incubated with secondary antibody, goat anti-rabbit IgG (H+L)-HRP (GenDEPOT, Katy, TX, USA), for 1 h. Protein bands were identified using Pierce ECL Plus Western Blotting Substrate (Thermo Fisher Scientific, Waltham, MA, USA), and immunofluorescence images were visualized using a ChemiDoc XRS + Imaging System (Bio-Rad, Hercules, CA, USA).

[0144]

[0145] 1-7. Path Obstruction Analysis

[0146] To identify the cell signaling pathway in LPS stimulation-induced activation of RAW264.7 cells, cells (1 Х 10 6 (cells / mL) were pretreated with inhibitors of NF-κB, ERK, JNK, and p38 before treatment with rainbow trout roe lipid extract (400 μg / mL) and LPS stimulation. After 24 h of incubation, the level of TNF-α expression was measured by real-time PCR to confirm the effect of rainbow trout roe lipid extract on the activation of RAW264.7 cells induced by LPS stimulation.

[0147]

[0148] 1-8. Statistical Analysis

[0149] Data were expressed as mean ± standard deviation (SD). Statistical analyses were performed using IBM SPSS statistical software (SPSS, Chicago, IL, USA) using one-way analysis of variance (ANOVA) and Duncan's multiple range test (p < 0.05).

[0150]

[0151] <Example 2> Analysis results

[0152] 2-1. Fatty acid profile

[0153] Lipids were extracted from rainbow trout (Onchorhynchus mykiss) egg samples according to Example 1-1. The extracted lipids had a yield of 14.22 ± 0.31% on a dry weight basis. The fatty acid composition of the extracted lipids was confirmed by GC-FID, and the fatty acid composition is shown in Table 1 below.

[0154]

[0155]

[0156] In total fatty acids, saturated fatty acids (SFAs) accounted for 29.83 ± 0.10%, monounsaturated fatty acids (MUFAs) accounted for 18.24 ± 0.59%, and polyunsaturated fatty acids (PUFAs) accounted for 51.92 ± 0.53%. The main saturated fatty acids, myristic acid (C14:0), accounted for 1.40%, palmitic acid (C16:0) accounted for 20.45%, and stearic acid (C18:0) accounted for 7.99%. Additionally, palmitoleic acid (C16:1n7), a major monounsaturated fatty acid, was found to account for 2.30%, oleic acid (C18:1n9) for 12.49%, and vaccenic acid (C18:1n7) for 3.45%. In the case of polyunsaturated fatty acids, linoleic acid (C18:2n6) accounted for 3.84%, linolenic acid (C18:3n3) accounted for 0.47%, eicosadienonic acid (C20:2n6) accounted for 1.37%, eicosatrienonic acid (C20:3n3) accounted for 2.45%, eicosapentaenoic acid (EPA, eicosapentaenoic acid C20:5n3) accounted for 7.77%, docosapentaenoic acid (DPA, docosapentaenoic acid C22:5n3) accounted for 2.36%, and docosahexaenoic acid (DHA, docosahexaenoic acid C22:6n3) accounted for 33.66%.

[0157]

[0158] 2-2. Cytotoxicity analysis of rainbow trout roe lipid extract

[0159] In order to analyze the cytotoxicity of the rainbow trout roe lipid extract according to the above Examples 1-2, the cell viability of RAW264.7 cells was confirmed when treated with rainbow trout roe lipid. Figure 1 shows the cell viability of RAW264.7 cells. As shown in Figure 1, the rainbow trout roe lipid extract did not significantly affect the viability of RAW264.7 cells, and cytotoxicity was found to increase in a concentration-dependent manner. LPS, used as a positive control, also showed concentration-dependent cytotoxicity, and aspirin, used as a positive drug, did not show cytotoxicity at any concentration.

[0160] Based on the above results, rainbow trout roe lipid extract was used at concentrations of 100, 200, 300, and 400 μg / mL, LPS was used as a positive control at concentrations of 1 μg / mL, and aspirin was used at concentrations of 200 μg / mL.

[0161]

[0162] 2-3. Analysis of NO production and iNOS expression

[0163] To confirm the effect of rainbow trout roe lipid extract on LPS-induced NO production, NO production was analyzed by measuring nitrate using Griess reagent (Promega, USA) according to the above Examples 1-3. Figure 2 shows the change in NO production according to the rainbow trout roe lipid extract treatment. As can be seen from Figure 2, it can be confirmed that the NO production was significantly reduced in a concentration-dependent manner by the rainbow trout roe lipid extract. The rainbow trout roe lipid extract at a concentration of 400 μg / mL showed a similar level of NO production to the control group.

[0164] In addition, the mRNA expression of iNOS, which produces NO, was confirmed according to the above Examples 1-5. Figure 3 shows the change in the mRNA expression level of iNOS. As shown in Figure 3, the iNOS mRNA expression level decreased as the concentration of rainbow trout roe lipid extract increased.

[0165] Through the above results, it was confirmed that rainbow trout roe lipid extract exhibited anti-inflammatory activity by suppressing NO production by regulating the transcription of the iNOS gene.

[0166]

[0167] 2-4. Analysis of inflammatory cytokine expression regulation

[0168] In order to confirm whether the rainbow trout roe lipid extract has the activity of regulating the gene expression of inflammatory or anti-inflammatory cytokines, the changes in the mRNA expression levels of TNF-α, IL-1β, IL-6, IL-10, IL-11, and TGF-β in RAW264.7 cells treated with the rainbow trout roe lipid extract according to Examples 1-5 were analyzed. Figures 4 to 9 show the changes in the mRNA expression levels of TNF-α, IL-1β, IL-6, IL-10, IL-11, and TGF-β. As shown in Figures 4 to 6, the rainbow trout roe lipid extract was found to significantly reduce the mRNA expression levels of TNF-α, IL-1β, and IL-6 in RAW264.7 cells stimulated with LPS in a concentration-dependent manner. However, looking at Figures 7 to 9, the expression levels of anti-inflammatory cytokines IL-10, IL-11, and transforming growth factor-beta (TGF-β) were found to increase in a concentration-dependent manner of rainbow trout egg lipid extract.

[0169] Through the above results, it was confirmed that rainbow trout roe lipid extract has the activity of regulating the gene expression of inflammatory cytokines and anti-inflammatory cytokines.

[0170]

[0171] 2-5. Analysis of NF-κB and MAPK activation

[0172] To confirm the anti-inflammatory effect of the rainbow trout roe lipid extract, NF-κB and MAPK activation were analyzed at the protein level in RAW264.7 cells using Western blotting according to Examples 1-6. The results of the Western blotting are shown in Figures 10 and 11. When treated with the rainbow trout roe lipid extract, the phosphorylation of NF-κB-p65 was significantly inhibited, and the inhibitory effect on the phosphorylation of NF-κB-p65 was found to be enhanced in a concentration-dependent manner by the rainbow trout roe lipid extract. In addition, when treated with the rainbow trout roe lipid extract, the phosphorylation of JNK, p38, and ERK1 / 2 was inhibited, and the inhibitory effect was enhanced as the concentration increased.

[0173] To further confirm the association between NF-κB and MAPK signaling molecules and the anti-inflammatory effect of rainbow trout roe lipid extract, the expression of TNF-α was analyzed using real-time quantitative PCR according to Examples 1-5 in a state where rainbow trout lipid extract was simultaneously treated with inhibitors of NF-κB, ERK, JNK or p38. The results of real-time quantitative PCR analysis are shown in Fig. 12. As shown in Fig. 12, LPS treatment increased phosphorylation of JNK, p38 and ERK1 / 2, but when rainbow trout roe lipid extract was treated together with inhibitors of NF-κB, ERK, JNK and p38, TNF-α expression was significantly reduced.

[0174] The above results confirmed that rainbow trout roe lipid extract has the activity of inhibiting NF-κB and MAPK activation.

[0175]

[0176] 2-6. Analysis of LPS-induced surface molecule expression

[0177] To confirm the effect of rainbow trout roe lipid extract on the expression of surface molecules induced by LPS, Flow Cytometry was performed according to Examples 1-8 above. The results of Flow Cytometry analysis are shown in Figs. 13 to 28. When macrophages were treated with LPS, the mean fluorescence intensity (MFI) of CD40 expression was 99.65 ± 0.17% compared to 2.61 ± 0.07% in RPMI, confirming that LPS has the effect of significantly increasing CD40 expression. However, when rainbow trout roe lipid extract was treated, the expression level of CD40 did not change, confirming that rainbow trout roe lipid extract is not related to the regulation of CD40 expression. However, when rainbow trout roe lipid extract was treated, the expression of CD86 was significantly reduced from 14.18 ± 0.13% to 11.35 ± 0.09%. The above results confirmed that rainbow trout roe lipid extract inhibited LPS-induced macrophage activation by suppressing CD86 expression.

[0178]

[0179] Through the above examples, the fatty acid composition, cytotoxicity, NO production inhibition effect, cytokine expression regulation effect, NF-κB and MAPK activation inhibition effect, and CD86 expression inhibition effect on the immune cell surface of the rainbow trout roe lipid extract were confirmed, and it was confirmed that the rainbow trout roe lipid extract can be used for anti-inflammatory purposes.

Claims

1. An anti-inflammatory food composition containing rainbow trout roe lipid extract as an effective ingredient.

2. In paragraph 1, A composition characterized in that the rainbow trout egg lipid extract is extracted using a solvent selected from the group consisting of distilled water, lower alcohols having C1 to C4, lower alcohol aqueous solutions, hexane, chloroform or acetic acid.

3. In paragraph 1, A composition characterized in that the rainbow trout roe lipid extract comprises a saturated fatty acid, a monounsaturated fatty acid or a polyunsaturated fatty acid.

4. In paragraph 3, A composition wherein the saturated fatty acid is at least one selected from the group consisting of myristic acid (C14:0), palmitic acid (C16:0), and stearic acid (C18:0).

5. In paragraph 3, A composition wherein the monounsaturated fatty acid is at least one selected from the group consisting of palmitoleic acid (C16:1n7), oleic acid (C18:1n9), and vaccenic acid (C18:1n7).

6. In paragraph 3, A composition wherein the polyunsaturated fatty acid is at least one selected from the group consisting of linoleic acid (C18:2n6), linolenic acid (C18:3n3), eicosadienonic acid (C20:2n6), eicosatrienonic acid (C20:3n3), eicosapentaenoic acid (EPA, C20:5n3), docosapentaenoic acid (DPA, C22:5n3), and docosahexaenoic acid (DHA, C22:6n3).

7. In paragraph 1, A composition wherein the rainbow trout roe lipid extract inhibits nitric oxide production.

8. In paragraph 7, A composition wherein the rainbow trout roe lipid extract inhibits nitric oxide production by inhibiting the expression of the iNOS gene.

9. In paragraph 1, A composition wherein the rainbow trout roe lipid extract inhibits the production of inflammatory cytokines and promotes the production of anti-inflammatory cytokines.

10. In paragraph 9, A composition wherein the above inflammatory cytokine is at least one selected from the group consisting of tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), and interleukin-1 beta (IL-1β).

11. In paragraph 9, A composition wherein the above anti-inflammatory cytokine is at least one selected from the group consisting of interleukin-10 (IL-10), interleukin-11 (IL-11), and transforming growth factor-beta (TGF-β).

12. In paragraph 1, A composition wherein the rainbow trout roe lipid extract has an activity of inhibiting the signal transduction pathway of Mitogen-Activated Protein Kinase (MAPK) or the signal transduction pathway of Nuclear Factor kappa B (NF-κB).

13. In paragraph 1, A composition wherein the rainbow trout roe lipid extract inhibits the activation of immune cells.

14. In paragraph 13, A composition wherein the rainbow trout roe lipid extract inhibits the activation of immune cells by reducing the expression of Cluster of Differentiation 86 (CD86) on the surface of immune cells.

15. An anti-inflammatory health functional food composition containing rainbow trout roe lipid extract as an effective ingredient.

16. An anti-inflammatory pharmaceutical composition comprising a rainbow trout roe lipid extract as an active ingredient.

17. An anti-inflammatory cosmetic composition comprising a rainbow trout roe lipid extract as an effective ingredient.

18. A method for preventing or treating inflammation, comprising administering to a subject the pharmaceutical composition of clause 16.

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