Inula and sesame compositions and their use for repairing ultraviolet damage

Abstract

The present application provides a combination of Inula japonica and sesame and its use for repairing ultraviolet damage. Specifically, a combination of active ingredients is provided, which includes: an Inula japonica extract; and a sesame extract. The combination has a synergistic effect in preventing and / or repairing ultraviolet damage, anti-inflammation, etc.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of medicine, in particular to a combination of Inula japonica and Sesamum indicum and its use for repairing ultraviolet damage. BACKGROUND

[0002] Ultraviolet-induced skin problems are the main cause of exogenous skin damage and skin aging. Ultraviolet damage to the skin includes tanning, sunburn, causing collagen and elastin to decompose and denature, aggravating inflammation, inducing the formation of free radicals, inhibiting skin immunity, inducing photo-dermatosis, aggravating pigmentation, and inducing skin cancer, etc.

[0003] Ultraviolet damage can be divided into acute damage and chronic damage, wherein the acute damage includes sunburn, etc., and the skin ages or accelerates aging (i.e. photoaging) due to cumulative damage after long-term exposure to ultraviolet rays in sunlight.

[0004] Therefore, preventing and repairing ultraviolet-induced skin damage is an effective means to protect the skin and delay aging. There is a need in the art to provide more products with the effect of preventing and / or repairing ultraviolet-induced skin damage. SUMMARY

[0005] The purpose of the present application is to provide a composition capable of preventing and / or repairing ultraviolet damage.

[0006] In a first aspect of the present application, a combination of active ingredients is provided, the combination comprising:

[0007] (a) a first active ingredient: Inula japonica extract; and

[0008] (b) a second active ingredient: Sesamum indicum extract;

[0009] wherein the weight ratio of the first active ingredient to the second active ingredient is 1-10:1-20 by dry weight.

[0010] The weight ratio of the first active ingredient to the second active ingredient is 1-5:1-5, more preferably 1-2:1-2, for example, 1:5, 2:5, 3:5, 4:5, 5:5, 5:1, 5:2, 5:3, 5:4, or 5:5.

[0011] In another preferred embodiment, the Inula japonica extract is a flower extract.

[0012] In another preferred embodiment, the Sesamum indicum extract is a seed extract, preferably obtained from the residue after oil extraction from Sesamum indicum seeds.

[0013] In another preferred embodiment, the extract comprises a water extract, an alcohol-water solvent extract, an alcohol extract, or a combination thereof.

[0014] In another preferred embodiment, the extract is obtained by solvent extraction, extraction, and / or chromatography.

[0015] In another preferred embodiment, the extractant of the extract is selected from the group consisting of water, alcohol (preferably C1-C4 alcohol, such as methanol, ethanol, propanol), aqueous alcohol, or a mixture thereof.

[0016] In another preferred embodiment, the extract is obtained by the extraction method of the present application.

[0017] In another preferred embodiment, the extract of Inula japonica is prepared by a method comprising the steps of:

[0018] (I) extracting dry Inula japonica with 80-100 wt% aqueous alcohol for 1-5 times, combining the extract, and removing the solvent to obtain a crude extract; and

[0019] (II) resuspending the crude extract with water, extracting with an organic solvent, collecting the aqueous phase, and removing the solvent to obtain the extract of Inula japonica.

[0020] In another preferred embodiment, step (I) has one or more features selected from the group consisting of:

[0021] (1) the aqueous alcohol has C1-C6 alkyl alcohol, preferably methanol, ethanol, isopropanol, or a combination thereof, more preferably 90-95% ethanol-water;

[0022] (2) the ratio of the dry Inula japonica to alcohol is 1 g:2-10 mL, preferably 1 g:4-8 mL;

[0023] (3) the extraction method is selected from the group consisting of soaking, ultrasonic, shaking, or a combination thereof;

[0024] (4) the extraction temperature is 4-40°C, preferably 20±10°C;

[0025] (5) the extraction time is 1-48 h / time, preferably 12-36 h / time;

[0026] (6) the number of extractions is 1-5 times, preferably 2-4 times; and / or

[0027] (7) the solid-liquid separation is selected from the group consisting of filtration, centrifugation, or a combination thereof;

[0028] In another preferred embodiment, the method of removing the solvent is selected from the group consisting of reduced pressure evaporation, freeze-drying, or a combination thereof.

[0029] In another preferred embodiment, the removal of the solvent is partial (such as concentration) or complete.

[0030] In another preferred embodiment, in step (II), the organic solvent extraction comprises the steps of:

[0031] According to the polarity from low to high, 1-5 (e.g. 2-4) different polarity organic solvents (solvents which can be layered with water, such as petroleum ether, ethyl acetate, n-hexane, n-butanol) are used for sequential extraction, for example, petroleum ether, ethyl acetate and n-butanol are used for sequential extraction.

[0032] In another preferred embodiment, the organic solvent is immiscible with water or has a solubility in water of <10 wt%.

[0033] In another preferred embodiment, the sesame is selected from the group consisting of black sesame, white sesame, or a combination thereof.

[0034] In another preferred embodiment, the sesame extract is prepared by a method comprising the steps of:

[0035] (i) extracting the sesame residue after oil pressing with 80-100 wt% aqueous alcohol solvent for 1-5 times, combining the extract, and removing the solvent to obtain a crude extract; and

[0036] (ii) resuspending the crude extract with water, extracting with an organic solvent, and collecting the aqueous phase;

[0037] (iii) extracting the aqueous phase with n-butanol, collecting the organic phase, and removing the solvent to obtain the sesame extract.

[0038] In another preferred embodiment, step (i) has one or more features selected from the group consisting of:

[0039] (1) in the aqueous alcohol solvent, the alcohol is a C1-C6 alkyl alcohol, preferably ethanol, isopropanol, or a combination thereof, more preferably 80-100% ethanol;

[0040] (2) the use amount ratio of the sesame residue to the aqueous alcohol solvent is 1 g: 2-10 mL, preferably 1 g: 4-8 mL;

[0041] (3) the extraction method is selected from the group consisting of soaking, ultrasonic, shaking, or a combination thereof, preferably ultrasonic extraction;

[0042] (4) the extraction temperature is 4-60°C, preferably 20±10°C;

[0043] (5) the extraction time is 0.5-48 h / time, preferably 1-6 h / time, such as 2 h; and / or

[0044] (6) the number of extraction times is 1-5 times, preferably 2-4 times.

[0045] In another preferred embodiment, in step (II), the organic solvent extraction comprises the steps of:

[0046] According to the polarity from low to high, 1-5 (e.g., 2-4) different polar organic solvents are used for sequential extraction, for example, n-hexane, ethyl acetate are used for sequential extraction.

[0047] In a second aspect of the present application, a composition is provided, which comprises the combination of active ingredients as described in the first aspect of the present application, and a cosmetically or pharmaceutically acceptable carrier or excipient.

[0048] In another preferred embodiment, the composition is a cosmetic composition, wherein the mass percentage of the combination of active ingredients is 0.0001-15 wt%, preferably 0.0001-10 wt%, more preferably 0.001-5 wt%, based on the dry weight, relative to the total weight of the cosmetic composition.

[0049] In another preferred embodiment, the composition is a pharmaceutical composition, wherein the mass percentage of the combination of active ingredients is 0.001-50 wt%, preferably 0.01-20 wt%, based on the dry weight, relative to the total weight of the pharmaceutical composition.

[0050] In another preferred embodiment, the pharmaceutical or cosmetic composition further comprises an additional component selected from the group consisting of a whitening or freckle-removing ingredient, an anti-inflammatory ingredient, an antioxidant ingredient, an anti-UV ingredient, or a combination thereof.

[0051] In another preferred embodiment, the cosmetically acceptable carrier or excipient is selected from the group consisting of a humectant, an antioxidant, an anti-UV agent, a preservative, a film-forming agent, an oil-soluble gelation agent, an organically modified clay mineral, a resin, an antibacterial agent, a fragrance, a salt, a pH adjustor, a chelating agent, a cooling agent, an anti-inflammatory agent, a skin beautifying ingredient, a vitamin, an amino acid, a nucleic acid, a hormone, a clathrate compound, or a combination thereof.

[0052] In another preferred embodiment, the pharmaceutical composition comprises a powder, a granule, a capsule, an injection, a tincture, an oral solution, a tablet, or a lozenge.

[0053] In another preferred embodiment, the dosage form of the cosmetic composition is a solid dosage form, a semi-solid dosage form, or a liquid dosage form, such as a solution, a gel, a cream, an emulsion, a paste, a cream, a paste, a cake, a powder, a patch, and the like.

[0054] In a third aspect of the present application, the use of the combination of active ingredients as described in the first aspect of the present application or the composition as described in the second aspect of the present application is provided for the preparation of a cosmetic or a pharmaceutical for one or more purposes selected from the group consisting of (a) preventing and / or repairing UV damage; and / or (b) anti-inflammation.

[0055] In another preferred embodiment, the UV damage is UV-induced skin damage, preferably the UV-induced skin damage is selected from the group consisting of sunburn (e.g., UV-induced erythema), tanning (e.g., pigmentation), photoaging, phototoxic and photosensitizing reactions, skin tumors (e.g., squamous cell carcinoma, basal cell carcinoma), or a combination thereof.

[0056] In another preferred embodiment, the UV light is selected from the group consisting of UVA (320-400 nm), UVB (290-320 nm), or a combination thereof.

[0057] In another preferred embodiment, the sunburn comprises erythema, DNA damage, UV-induced epidermal cell death, skin surface roughness, increased keratin, impaired skin barrier, or a combination thereof.

[0058] In another preferred embodiment, the photoaging comprises wrinkles, skin laxity, telangiectasia, uneven skin pigmentation, breakdown of collagen and / or elastin.

[0059] In another preferred embodiment, the UV-induced skin damage comprises epidermal layer damage (e.g., stratum corneum) and dermal layer damage.

[0060] In another preferred embodiment, the prevention of UV damage is to enhance the tolerance of epidermal cells to UV light. (Compared to a control group (a group without using the combination of active ingredients of the present application, such as a vehicle control group, a group using only the extract of Inula japonica, or a group using only the extract of Sesamum indicum), the group treated with the combination of active ingredients of the present application has a cell activity that is 10% (i.e., 1.1 times or more), 20%, 30%, 40%, 50%, 60%, 80%, or 100% or more higher than the control group after being irradiated with the same intensity of UV light.

[0061] In another preferred embodiment, the epidermal cells are selected from the group consisting of keratinocytes, dendritic cells, or a combination thereof.

[0062] In another preferred embodiment, the pharmaceutical or cosmetic composition is used for sun protection and / or anti-photoaging.

[0063] In another preferred embodiment, the cosmetic or pharmaceutical is used for one or more purposes selected from the group consisting of:

[0064] (1) preventing UV-induced decrease in cell activity or increasing cell activity;

[0065] (2) inhibiting UV-induced increase in intracellular oxidative stress level;

[0066] (3) inhibiting UV-induced increase in intracellular active nitrogen level;

[0067] (4) repairing UV-induced intracellular DNA damage;

[0068] (5) inhibiting the increase of intracellular inflammatory factor IL-6 level induced by UV; and / or

[0069] (6) inhibiting the increase of intracellular inflammatory factor TNF-a level induced by UV.

[0070] In another preferred embodiment, the cell is an epidermal cell.

[0071] In another preferred embodiment, the epidermal cell comprises: a stratum corneum-forming cell, a dendritic cell, or a combination thereof.

[0072] In another preferred embodiment, the pharmaceutical or cosmetic composition is used for pre-sun prevention and / or post-sun repair.

[0073] In a fourth aspect, the present application provides a method for skin care, comprising the step of administering to a subject in need thereof the combination of active ingredients or a composition comprising the same according to the first aspect of the present application, thereby protecting the skin, and the skin care refers to (a) preventing and / or repairing UV damage; and / or (b) anti-inflammation.

[0074] In another preferred embodiment, the subject is a mammal, such as a human, a rat or a mouse.

[0075] In another preferred embodiment, the subject is a skin-sensitive population, such as an elderly, a child or a sensitive skin population.

[0076] It should be understood that, within the scope of the present application, each of the above technical features of the present application and each of the technical features specifically described below (such as in the examples) can be combined with each other to form a new or preferred technical solution. Due to the limited space, they are not listed one by one here. BRIEF DESCRIPTION OF DRAWINGS

[0077] Figure 1 The UV protection effect of different groups on cells is shown.

[0078] Figure 2 The effect of different groups on the oxidative stress level of cells is shown.

[0079] Figure 3 The effect of different groups on the active nitrogen level in cells is shown.

[0080] Figure 4 The effect of different groups on the inflammatory factor IL-6 level in cells is shown.

[0081] Figure 5 The effect of different groups on the inflammatory factor TNF-a level in cells is shown.

[0082] Figure 6 The effect of different groups on the DNA damage repair of cells is shown.

[0083] Figures 1-6 Each value is from the average of three independent experiments ± standard error, ****p<0.0001, ***p<0.001, **p<0.01. DETAILED DESCRIPTION

[0084] The present inventors have made extensive and in-depth research, and through a large number of screening and testing, provided a composition of Inula japonica extract and sesame extract. Surprisingly, the composition of the present application has a significant synergistic effect in improving cell activity reduction, reducing oxidative stress, active nitrogen, cell inflammatory factors, intracellular CPDs level and the like caused by ultraviolet radiation, and is very suitable for preparing cosmetics or drugs for sun protection, after-sun repair, and anti-inflammatory. On this basis, the present application is completed.

[0085] Terminology

[0086] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.

[0087] As used herein, the term "about," when used in reference to a particular recited numerical value, means that the value can vary from the recited value by not more than 1%. For example, as used herein, the expression "about 100" includes all values between 99 and 101 (e.g., 99.1, 99.2, 99.3, 99.4, etc.).

[0088] As used herein, the term "comprising" or "including" can be open, semi-closed and closed. In other words, the term also includes "consisting essentially of" or "consisting of".

[0089] As used herein, the term "room temperature" refers to a temperature of 4-40°C, preferably 25±5°C.

[0090] Active ingredients, combinations

[0091] The first active ingredient of the present application is Inula japonica extract. For a long time, Chinese medicine has used the flowers of Inula japonica to treat digestive system diseases, bronchitis and inflammation.

[0092] The second active ingredient of the present application is an extract of Sesamum indicum. In particular, the extract of the present application can be extracted from the residue left after the oil is pressed from Sesamum indicum. Sesamin and sesamolin are lignans unique to Sesamum indicum, and a considerable amount of lignans remains in the sesame cake produced after the sesame oil is prepared. Sesame has strong antioxidant activity, and this is due to the presence of two types of antioxidant substances, lignans and tocopherols.

[0093] The present application first discovered the synergistic effect of the extract of Inula japonica and the extract of Sesamum indicum in (a) preventing and / or repairing ultraviolet damage; and / or (b) anti-inflammation, etc. Thus, a composition comprising the active ingredients of the extract of Inula japonica and the extract of Sesamum indicum is provided.

[0094] Extract, preparation method

[0095] In the present application, the extract of Inula japonica is a flower extract.

[0096] In the present application, the extract of Sesamum indicum is a seed extract, preferably extracted from the residue left after the oil is pressed from Sesamum indicum seeds.

[0097] In a preferred embodiment of the present application, the preparation of the "extract" or "active ingredient" can be independently or jointly carried out by solvent extraction, extraction, supercritical extraction, and / or chromatography.

[0098] In the present application, the number of extractions can be one or more (e.g., 2, 3, 4, 5 times). When multiple extractions are performed, the multiple extracts can be combined and then subjected to subsequent processing.

[0099] In the present application, the solvent used in the solvent extraction method is not particularly limited, and representative examples include, but are not limited to, water, ethanol, methanol, and an alcohol-water (mixed) solvent.

[0100] In the present application, the alcohol-water solvent refers to a mixed solvent of water and another solvent such as an alcohol (particularly a C1-C4 alcohol such as methanol, ethanol, or isopropanol). Typically, the alcohol content is 40-100 wt%, preferably 60-95 wt%, such as 80 wt%, 90 wt%, 98 wt%, etc., based on the total weight of the alcohol-water solvent.

[0101] In the present application, the solvent used in the solvent extraction method is not particularly limited, and representative examples include, but are not limited to, one or a mixture of several solvents selected from the group consisting of n-butanol, dichloromethane, chloroform, C5-C7 alkanes, cyclohexane, and petroleum ether. The number of extractions can be one or more.

[0102] In the present application, the solvent used for supercritical extraction is not particularly limited, and representative examples include, but are not limited to, carbon dioxide or propane.

[0103] In the present application, the column chromatography is not particularly limited, and representative examples include, but are not limited to, one or a combination of several of activated carbon, silica gel, reversed-phase silica gel, macroporous resin, and Sephadex.

[0104] In a preferred embodiment of the present application, the preparation method of the Inula japonica extract is as follows: Inula japonica (dried flowers) is used as the raw material, and the raw material is extracted with an adequate amount of alcohol (e.g., 1-20 times the weight of water) or an alcohol-water mixture solvent (50-99% alcohol, preferably 80-95% ethanol) for 1-5 times, thereby obtaining the extract. In addition, in the present application, the fat-soluble components can be further separated (or removed) from the above-mentioned alcohol extract, thereby obtaining an Inula japonica extract with a higher content (or relative content) of water-soluble components, or an Inula japonica extract mainly containing water-soluble components.

[0105] In a preferred embodiment of the present application, the preparation method of the sesame extract is as follows: sesame (e.g., sesame residue after oil extraction) is used as the raw material, and the raw material is extracted with an adequate amount of alcohol (e.g., 1-20 times the weight of water) or an alcohol-water mixture solvent (50-99% alcohol, preferably 80-95% ethanol) for 1-5 times, thereby obtaining the alcohol extract. In addition, in the present application, the fat-soluble components (e.g., oil) can be further separated (or removed) from the above-mentioned alcohol extract, thereby obtaining a sesame extract with a higher content (or relative content) of active components.

[0106] Composition and use

[0107] As used herein, the term "pharmaceutical or cosmetic composition" includes (a) a combination of active ingredients according to the present application; and (b) a pharmaceutically or cosmetically acceptable carrier.

[0108] In addition, the cosmetic composition includes a skin care product.

[0109] It is possible to prepare a pharmaceutical composition, such as an ointment, a cream, a gel, a paste, a patch, etc., from the combination of active ingredients according to the present application. The pharmaceutical can be prepared by generally known preparation techniques, and suitable pharmaceutical additives can be added to the pharmaceutical.

[0110] Examples of the pharmaceutical additives include excipients, binders, disintegrants, lubricants, flow aids, suspending agents, emulsifiers, stabilizers, humectants (wetting agents), preservatives, solvents, solubilizers, preservatives, flavoring agents, sweetening agents, dyes, fragrances, propellants, etc., which can be selected and added in a suitable amount within a range that does not affect the effects of the present application.

[0111] It is possible to prepare the combination of the active ingredients of the present application into a cosmetic composition, a solid dosage form, a semi-solid dosage form, or a liquid dosage form such as a solution, a gel, a cream, an emulsion, a spray, a paste, a cream, a paste, a cake, a powder, a patch, and the like.

[0112] The first active ingredient and the second active ingredient of the present application can be prepared into separate dosage forms with a carrier, respectively, or into a single dosage form by mixing together.

[0113] In the cosmetic of the present application, other ingredients commonly used in cosmetics, such as a film-forming agent, an oil-soluble gelation agent, an organically modified clay mineral, a resin, a humectant, a preservative, an antibacterial agent, a perfume, a salt, an antioxidant, a pH adjustor, a chelating agent, a cooling agent, an anti-inflammatory agent, a skin beautifying ingredient (a whitening agent, a cell activity agent, a skin roughness improvement agent, a blood circulation promoter, a skin astringent, an anti-sebum agent, and the like), a vitamin, an amino acid, a nucleic acid, a hormone, a clathrate compound, and the like, can be added within a range not impairing the effects of the present application.

[0114] The oil-soluble gelation agent is a gelation agent selected from the group consisting of metal soaps such as aluminum stearate, magnesium stearate, zinc myristate, and the like; amino acid derivatives such as N-lauroyl-L-glutamic acid, α, γ-di-n-butylamine, and the like; cyclodextrin fatty acid esters such as cyclodextrin palmitate, cyclodextrin stearate, cyclodextrin 2-ethylhexanoate palmitate, and the like; sucrose fatty acid esters such as sucrose palmitate, sucrose stearate, and the like; benzaldehyde derivatives of sorbitol such as monobenzal sorbitol, dibenzal sorbitol, and the like; organically modified clay minerals such as dimethyl benzyl dodecyl ammonium montmorillonite clay, dimethyl octadecyl ammonium montmorillonite clay, and the like, and the like, and one or two or more kinds thereof can be used as needed.

[0115] The humectant is glycerin, sorbitol, propylene glycol, dipropylene glycol, 1,3-butanediol, glucose, xylitol, maltitol, polyethylene glycol, hyaluronic acid, chondroitin sulfate, pyrrolidone carboxylate, polyoxyethylene methyl glucoside, polyoxypropylene methyl glucoside, and the like.

[0116] The antibacterial preservative is alkyl parahydroxybenzoate, benzoic acid, sodium benzoate, sorbic acid, potassium sorbate, phenoxyethanol, and the like, and the antibacterial agent is benzoic acid, salicylic acid, phenol, sorbic acid, alkyl parahydroxybenzoate, p-chloro-m-cresol, hexachlorophene, benzalkonium chloride, chlorhexidine, trichloro-N-carbanilide, triclosan, luminous, phenoxyethanol, and the like.

[0117] Antioxidants are tocopherol, butylated hydroxyanisole, butylated hydroxytoluene, phytic acid, etc., pH adjusting agents are lactic acid, citric acid, glycolic acid, succinic acid, tartaric acid, dl-malic acid, potassium carbonate, sodium bicarbonate, ammonium bicarbonate, etc., chelating agents are alanine, sodium ethylenediaminetetraacetate, sodium polyphosphate, sodium metaphosphate, phosphoric acid, etc., cooling agents are L-menthol, camphor, etc., anti-inflammatory agents are allantoin, glycyrrhetinic acid, glycyrrhizic acid, tranexamic acid, azulene, etc.

[0118] Skin beautifying ingredients are placenta extract, arbutin, glutathione, saxifrage extract, etc. as whitening agents; royal jelly, melanopsin, cholesterol derivatives, calf blood extract, etc. as cell activators; skin roughness improvers; valinamide, nicotinyl benzoate, beta-butoxyethyl nicotinate, capsaicin, gingerone, cantharidin, ichthammol, caffeine, tannic acid, alpha-pinene, tocopheryl nicotinate, inositol hexanicotinate, cyclandelate, guaiazolene, tolazoline, acetylcholine, verapamil, stepholidine, gamma-oryzanol, etc. as blood circulation improvers; zinc oxide, tannic acid, etc. as skin astringents; sulfur, etc. as anti-sebum agents, vitamins are vitamin A oil, rosin oil, rosin oil acetate, rosin oil palmitate, etc. as vitamin A; riboflavin, riboflavin butyrate, flavin adenine nucleotide, etc. as vitamin B2; pyridoxine hydrochloride, pyridoxine dioctanoate, pyridoxine tripalmitate, etc. as vitamin B6, vitamin B12 and its derivatives, vitamin B15 and its derivatives, etc. as vitamin B; L-ascorbic acid, L-ascorbic acid dipalmitate, L-ascorbic acid-2-sodium sulfate, L-ascorbic acid phosphate dipotassium, etc. as vitamin C; ergocalciferol, cholecalciferol, etc. as vitamin D; alpha-tocopherol, beta-tocopherol, gamma-tocopherol, dl-alpha-tocopherol acetate, dl-alpha-tocopherol nicotinate, dl-alpha-tocopherol succinate, etc. as vitamin E; vitamin H; vitamin P; nicotinic acid, benzoyl nicotinate, nicotinamide, etc. as niacin; calcium pantothenate, D-panthenol, pantothenyl ethyl ether, acetyl pantothenyl ethyl ether, etc. as pantothenic acid; biotin, etc.

[0119] Amino acids are glycine, valine, leucine, isoleucine, serine, threonine, phenylalanine, arginine, lysine, aspartic acid, glutamic acid, cystine, cysteine, methionine, tryptophan, etc., nucleic acids are deoxyribonucleic acid, etc., hormones are estradiol, ethenyl estradiol, etc.

[0120] Preferred examples of the cosmetic of the present application include skin care cosmetics, makeup cosmetics, and ultraviolet protection cosmetics. For example, there are emulsions, creams, lotions, sunscreens, pack materials, face washes, essences, and the like as base cosmetics; foundations, face powders, blushes, and the like as makeup cosmetics, etc.

[0121] The form of the product is not particularly limited, and can be liquid, emulsion, cream, solid, paste, gel, powder, multi-layer, mousse, spray, or the like.

[0122] The present application also provides a skin care method, which comprises the step of administering the combination of active ingredients or the composition of the present application to a subject in need thereof. Typically, the method is a method for preventing and / or repairing ultraviolet-induced skin damage.

[0123] In another preferred embodiment, the effective concentration of the combination of active ingredients ranges from 1 μg / ml to 500 mg / ml, such as 5 μg / ml, 10 μg / ml, 20 μg / ml, 50 μg / ml, 100 μg / ml, or 200 μg / ml.

[0124] As used herein, the term "effective amount" means any amount of the combination or composition of the present application, either alone or in combination with another therapeutic agent or cosmetic, which when used in the treatment of a disease, results in the regression of the disease, as manifested by a decrease in severity of the disease symptoms, an increase in the frequency and duration of periods of disease symptom- free, or a prevention of a disorder or disability due to the disease. A "therapeutically effective amount" of the drug of the present application also includes a "prophylactically effective amount", which is any amount of the combination or composition of the present application, either alone or in combination with another therapeutic agent or cosmetic, which when administered to a subject at risk of developing a disease or symptom, or to a subject experiencing a recurrence of a disease or symptom, inhibits the development or recurrence of the disease or symptom.

[0125] In the present application, the use of the combination of active ingredients of the present application for the prevention and / or repair of sunburn, sunburn (such as ultraviolet-induced erythema), suntan (such as pigmentation), photoaging, phototoxic and photosensitive reactions, skin tumors, and the like, includes both prophylactic use and post-event improvement use. For example, for sunburn, the combination of active ingredients of the present application or a composition comprising the same is applied before, during, and / or after sun exposure to prevent and / or repair sunburn.

[0126] The main advantages of the present application include:

[0127] 1. The present application provides a combination of active ingredients for preventing and / or repairing ultraviolet damage and anti-inflammatory, which comprises an extract of Inula japonica and an extract of Sesamum indicum. Surprisingly, the extract of Inula japonica and the extract of Sesamum indicum of the present application have significant synergistic effects in preventing and / or repairing ultraviolet damage and anti-inflammatory.

[0128] 2. In the composition of the present application, the extract of Sesamum indicum can be extracted from the residue of sesame after oil extraction, which can be recycled and reused, thereby reducing the preparation cost and saving resources.

[0129] The application will be further described in conjunction with specific embodiments. It should be understood that these embodiments are intended to illustrate the application and are not intended to limit the scope of the application. The experimental procedures in the following examples, unless otherwise specified, were carried out under conventional conditions, for example, as described in Sambrook et al., Molecular Cloning: A Laboratory Manual (New York: Cold Spring Harbor Laboratory Press, 1989), or as recommended by the manufacturer. Percentages and parts are by weight unless otherwise specified.

[0130] Example 1

[0131] Preparation of plant extracts

[0132] 1.1 Preparation of sesame extract Dry raw black sesame seeds (SESAMUM INDICUM) were pressed to remove most of the oil. The sesame seed residue (100 g) after oil pressing was soaked in 95% ethanol (600 mL) and sonicated for 1 h. The extraction was repeated three times. The three ethanol extracts were combined and evaporated under reduced pressure to obtain the crude sesame ethanol extract. The crude sesame ethanol extract was resuspended in water and sequentially extracted with n-hexane (to retain the aqueous phase), ethyl acetate (to retain the aqueous phase) and n-butanol. The n-butanol extract was evaporated under reduced pressure to obtain the sesame extract used in the experiment.

[0133] 1.2 Preparation of inula extract Air-dried inula (INULA JAPONICA) (100 g) was soaked in 95% ethanol (600 mL) at room temperature for 24 h, followed by filtration. The residue was re-soaked and the extraction was repeated three times. The three filtrates were combined and evaporated under reduced pressure to obtain the crude inula ethanol extract. The crude inula ethanol extract was resuspended in water and sequentially extracted with petroleum ether, ethyl acetate and n-butanol. The aqueous solution remaining after extraction was evaporated under reduced pressure to obtain the inula extract used in the experiment.

[0134] Example 2

[0135] Cell culture, extract pretreatment and UVA irradiation

[0136] Human primary keratinocytes (purchased from ATCC (Rockville, MD, USA)). Only keratinocytes of passage 1-5 were selected for the experiment. The cells were cultured in a 5% carbon dioxide incubator at 37°C with dermal cell culture medium supplemented with keratinocyte growth kit. Twenty-four hours before UV irradiation, the culture medium of the experimental groups was replaced with the pre-prepared culture medium containing different concentrations of plant extracts (mixed extract was a mixture of sesame extract and inula extract at a ratio of 5:2), while the UV control group and the blank control group (protected from light) were replaced with an equal amount of fresh culture medium without any plant extract components.

[0137] UV radiation was performed using UVA fluorescent lamps (Vilber Lourmat, France) and the total UVA dose used in the experiment was 4.5 J / cm 2 Before UV irradiation, cells were washed with PBS solution and a small amount of PBS solution was retained to keep the cells moist during the experiment. All cells were treated identically throughout the experiment.

[0138] Example 3

[0139] Cell viability assay (CCK-8) method

[0140] After UVA irradiation, the remaining PBS solution was removed and the culture solution with 10% CCK-8 solution was added. The cells were returned to the incubator for further incubation. After 2 h of incubation, the absorbance was measured at 450 nm using a microplate reader.

[0141] The experimental results are shown in Table 1. Figure 1 As shown in Table 1, the use of Flos Inulae extract alone did not improve cell viability, but the use of sesame extract and the mixed extract of the present application for pretreatment of human primary keratinocytes before UV irradiation effectively improved cell viability. Compared with the use of sesame extract alone, the mixture of Flos Inulae extract and sesame extract had a significant synergistic effect on improving UV-induced cell viability.

[0142] Example 4

[0143] Cell oxidative stress (ROS) measurement

[0144] This method measures the level of intracellular ROS by DCFA and corrects by cell viability to accurately assess the damage caused to cells by UV (UVA) radiation. Human primary keratinocytes were seeded into 96-well plates at a density of 1.5 x 10 4 After 24 h of seeding, the culture solution of the experimental group was replaced with the previously prepared culture solution containing different concentrations of mixed extract, and the UV control group and the blank control group were replaced with an equal amount of fresh culture solution without any plant extract. After 24 h of pretreatment, the cells were washed three times with PBS solution and a small amount of PBS solution was retained to keep the cells moist during the experiment. The cells were irradiated with 4.5 J / cm 2 After UVA irradiation of the cells, 25 uM DCFA-containing PBS solution was added, and the cells were returned to the incubator for 30 min of incubation before fluorescence measurement. The excitation wavelength used was 485 nm and the emission wavelength was 535 nm.

[0145] The experimental results are shown in Table 1. Figure 2As shown, the mixed extract of the present invention, when used to pretreat human primary keratinocytes before ultraviolet radiation, can synergistically and effectively inhibit the increase in intracellular oxidative stress caused by ultraviolet radiation.

[0146] Example 5

[0147] Cellular reactive nitrogen (RNS) measurement

[0148] This method measures intracellular reactive nitrogen levels using a nitrotyrosine ELISA kit. Human primary keratinocytes were cultured at 1.5 × 10⁻⁶ cells / cells. 5 Cells were seeded at a density of cells / well in 24-well plates. After 24 h of seeding, cells were washed with PBS, leaving a small amount of PBS to keep them moist during the experiment. Cells were irradiated with 4.5 J / cm² UVA, and the remaining PBS was removed. Pre-prepared culture medium containing different concentrations of mixed extracts was added to the experimental group cells, while equal volumes of fresh culture medium without any plant extracts were added to the UV control and blank control groups. Cells were incubated for 16 h and then collected. The cell pellet was placed in extraction buffer and incubated on ice for 20 min. After centrifugation, the supernatant was transferred to clean tubes, and the protein concentration of the samples was measured using the BCA protein assay. 3 NT BSA was used as a standard positive control to validate the assay. Before loading, the protein concentration of the samples was adjusted to approximately the same level. Fifty μL of each diluted standard or sample was added to each well of the pre-coated plate along with the 2X HRP detector antibody and incubated at room temperature for 2 h. After washing four times and completely removing the liquid, 100 μL of HRP developing solution was added to each well. Measure the OD value and calculate the nitrotyrosine concentration based on the standard curve.

[0149] Experimental results are as follows Figure 3 As shown, treatment of human primary keratinocytes with a mixed extract after ultraviolet radiation can synergistically and effectively inhibit the increase in intracellular reactive nitrogen levels caused by ultraviolet radiation.

[0150] Example 6

[0151] Measurement of cellular inflammatory factors IL-6 and TNF-α

[0152] Cellular inflammatory factors were measured using an ELISA kit. Human primary keratinocytes were cultured at 1.5 × 10⁻⁶ cells / mL. 5 Cells were seeded at a density of [number] cells / well in 24-well plates. After 24 hours, cells were washed with PBS, leaving a small amount of PBS to keep the cells moist during the experiment. [Further details about the process are needed for accurate translation.] 2After UVA irradiation, the remaining PBS solution was removed, and the experimental group cells were added with the prepared culture solution containing different concentrations of mixed extracts, while the UV control group and the blank control group were added with the same amount of fresh culture solution without any plant extracts. The cells were placed back in the incubator for 24 h before collecting the cell culture solution. After centrifugation, the supernatant of the culture medium was taken as the sample, and 50 microliters of the sample or standard was added to the ELISA 96-well plate, while the same amount of antibody mixture was added to each well. After incubation at room temperature for 1 h, the wells were washed four times, 100 microliters of TMB substrate was added to each well and reacted for color development in the dark for 10 min, and then the termination solution was added. The OD value was measured and the concentration of cell inflammatory factors was calculated according to the standard curve.

[0153] The IL-6 level measurement results are shown in Table 2, and the mixed extract can effectively inhibit the increase of the cell inflammatory factor IL-6 level caused by ultraviolet radiation. Figure 4

[0154] The TNF-a level measurement results are shown in Table 3, and the mixed extract can effectively inhibit the increase of the cell inflammatory factor TNF-a level caused by ultraviolet radiation. Figure 5

[0155] Example 7

[0156] DNA damage CPDs measurement

[0157] The DNA damage level was determined using an ELISA kit. Human primary keratinocytes were seeded in 96-well plates at 3 x 105cells / well and incubated for 24 h. The cells were then irradiated with UVA at 0.5 J / cm2, and the cells were treated with the mixed extract at different concentrations for 24 h. The cells were then collected and centrifuged, and the supernatant was taken as the sample. The sample was added to the ELISA 96-well plate, and the same amount of antibody mixture was added to each well. After incubation at room temperature for 1 h, the wells were washed four times, 100 microliters of TMB substrate was added to each well and reacted for color development in the dark for 10 min, and then the termination solution was added. The OD value was measured and the concentration of cell inflammatory factors was calculated according to the standard curve. 5 ​​The cells were seeded at a density of 1 x 104cells / well into 12-well plates. After 24 h of seeding, the cells were washed with PBS solution and a small amount of PBS solution was retained to keep the cells moist during the experiment. After irradiating the cells with 4.5 J / cm2UVA, the remaining PBS solution was removed and the test cells were treated with the pre-prepared culture medium containing different concentrations of the mixed extract, while the UV control and blank control groups were treated with the same amount of fresh culture medium without any plant extract. After incubating the cells in the incubator for 1 h, the cells were treated with 0.05% trypsin and collected. The DNA samples were extracted using the DNeasy Blood and Tissue Kit. Each DNA sample was diluted to 4 μg / mL in cold AE buffer. The samples and standards were converted to single-stranded DNA by incubating the DNA at 95°C for 10 min and then quickly cooling on ice for 10 min. Fifty microliters of the sample or standard solution was added to the wells of a DNA high-binding plate. Fifty microliters of DNA binding solution was added to each well and incubated at room temperature overnight. After removing the DNA solution and washing twice with PBS, 200 microliters of Assay Diluent was added to each well and incubated at room temperature for 1 h. Then, 100 microliters of diluted anti-CPD antibody was added to all the wells and incubated at room temperature for 1 h. Subsequently, the wells were washed five times with washing buffer and 100 microliters of diluted secondary antibody-HRP conjugate was added to all the wells and incubated at room temperature for 1 h. After washing five more times, 100 microliters of TMB substrate was added to each well and incubated at room temperature for 15 min, followed by the addition of stop solution. The absorbance at 450 nm was read and the concentration of CPDs in the cells was calculated according to the standard curve.

[0158] The experimental results, as shown in Table 1, show that the repair treatment of human primary keratinocytes with the mixed extract after UV irradiation can effectively repair the DNA damage of the cells caused by UV irradiation. Figure 6

[0159] All the documents mentioned in the present application are incorporated herein by reference. Furthermore, it is to be understood that various alterations and modifications can be made to the application without departing from the scope thereof as defined by the appended claims.​

Claims

1. A composition for preventing or repairing ultraviolet-induced skin damage and / or anti-inflammation, characterized by, The composition consists of the following first active ingredient and second active ingredient: (a) first active ingredient: Inula japonica extract; and (b) second active ingredient: Sesamum indicum extract; wherein the weight ratio of the first active ingredient to the second active ingredient is 1-10:1-20 on a dry weight basis; wherein the Inula japonica extract is prepared by a method comprising the steps of: (I) extracting dry Inula japonica with 80-100 wt% aqueous alcohol solvent for 1-5 times, combining the extract, and removing the solvent to obtain a crude extract, wherein the alcohol is ethanol or isopropanol; and (II) resuspending the crude extract with water, extracting with an organic solvent, collecting the aqueous phase, and removing the solvent to obtain the Inula japonica extract; wherein the organic solvent extraction is sequential extraction using petroleum ether, ethyl acetate, and n-butanol; the Sesamum indicum extract is prepared by a method comprising the steps of: (i) extracting oil-extracted Sesamum indicum residue with 80-100 wt% aqueous alcohol solvent for 1-5 times, combining the extract, and removing the solvent to obtain a crude extract, wherein the alcohol is ethanol or isopropanol; and (ii) resuspending the crude extract with water, extracting with an organic solvent, and collecting the aqueous phase; wherein the organic solvent extraction is sequential extraction using n-hexane and ethyl acetate; (iii) extracting the aqueous phase with n-butanol, collecting the organic phase, and removing the solvent to obtain the Sesamum indicum extract.

2. The active composition of claim 1, wherein The Sesamum indicum is selected from the group consisting of black Sesamum indicum, white Sesamum indicum, or a combination thereof.

3. The composition of claim 1, wherein The weight ratio of the first active ingredient to the second active ingredient is 1-2:1-2.

4. The composition of claim 1, wherein The Inula japonica extract is a flower extract.

5. The composition of claim 1, wherein The Sesamum indicum extract is a seed extract.

6. The composition of claim 1, wherein Step (I) has one or more features selected from the following: (1) the aqueous alcohol solvent is 90-95 wt% ethanol-water; (2) the dry Inula japonica and alcohol are used in a ratio of 1 g:2-10 mL; (3) the extraction method is selected from the group consisting of soaking, ultrasonic, shaking, or a combination thereof; (4) the extraction temperature is 4-40℃; (5) the extraction time is 1-48 h / time; and / or (6) the solid-liquid separation is selected from the group consisting of filtration, centrifugation, or a combination thereof.

7. The composition of claim 1, wherein In step (I), the extraction is performed 2-4 times.

8. The composition of claim 1, wherein Step (i) has one or more features selected from the following: (1) the aqueous alcohol solvent is 80-100 wt% ethanol; (2) the Sesamum indicum residue and aqueous alcohol solvent are used in a ratio of 1 g:2-10 mL; (3) the extraction method is selected from the group consisting of soaking, ultrasonic, shaking, or a combination thereof; (4) the extraction temperature is 4-60℃; and / or (5) the extraction time is 0.5-48 h / time.

9. The composition of claim 1, wherein In step (i), the extraction is performed for 1-6 h / time.

10. The composition of claim 1, wherein In step (i), the extraction is performed 2-4 times.

11. A cosmetic composition characterized in that, Compositions as claimed in any one of claims 1-10, together with a cosmetically acceptable carrier or excipient. The mass percentage of the composition according to any one of claims 1-10 is 0.0001-15 wt% on a dry weight basis, based on the total weight of the cosmetic composition.

12. The cosmetic composition according to claim 11, wherein ​ 13. Use of a composition according to any one of claims 1 to 10 or a cosmetic composition according to claim 12 in the manufacture of a cosmetic for (a) the prevention and / or repair of ultraviolet-induced skin damage; and / or (b) anti-inflammation.

14. The use according to claim 13, characterized in that, The ultraviolet-induced skin damage includes epidermal layer damage and dermal layer damage.

15. A pharmaceutical composition comprising, including: A composition according to any one of claims 1 to 10, and a pharmaceutically acceptable carrier or excipient.

16. Use of a composition according to any one of claims 1 to 10 or a pharmaceutical composition according to claim 15 in the manufacture of a medicament for (a) the prevention and / or repair of ultraviolet-induced skin damage; and / or (b) anti-inflammation.

17. The use according to claim 16, characterized in that, The ultraviolet-induced skin damage includes epidermal layer damage and dermal layer damage.

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