Compositions and methods for treating skin pigmentation disorders

Abstract

Provided herein are compositions for preventing and / or treating skin pigmentation disorders by degrading melanin in dermal fibroblasts. Also provided herein are methods and uses for treating skin pigmentation disorders and for degrading melanin in dermal fibroblasts, as well as non-therapeutic methods of lightening skin.

Description

[0001] field

[0002] The present disclosure relates to compositions for preventing and / or treating skin pigmentation disorders by degrading melanin in dermal fibroblasts. The present disclosure also relates to methods and uses for treating skin pigmentation disorders and for degrading melanin in dermal fibroblasts, as well as non-therapeutic methods for lightening skin.

[0003] background

[0004] Melanogenesis is a process that occurs in melanosomes by melanocytes, and the process involves a series of enzymatic reactions and chemical reactions to produce the pigment melanin, which mainly occurs in the epithelial layer of the skin. Skin hyperpigmentation is a common condition in humans, which is mainly caused by excessive melanin synthesis and storage through uncontrolled melanogenesis. Increased melanin production can be caused by many factors, including overexposure to sunlight, hormone changes, skin inflammation and aging, and many other biological processes (Nieuweboer-Krobotova et al., 2013).

[0005] Oxidative stress caused by the sun's UVA and UVB rays and high-energy visible light is the main cause of the problem of hyperpigmentation of the skin caused by sunlight, which results in small dark spots of the skin called sun spots. It is believed that when sex hormones (such as estrogen and progesterone) stimulate excessive production of melanin, larger hyperpigmentation spots (mainly on the face) known as melasma, which are particularly common in women, will occur, which can also be aggravated by sun exposure. Post-inflammatory hyperpigmentation refers to the darkening of the skin due to the uncontrolled melanin production process caused by inflammatory rashes or skin damage. These and other hyperpigmentation skin disorders can significantly affect the appearance of an individual and may cause emotional and psychological distress and reduce the quality of life.

[0006] Due to the cosmetically important problem of hyperpigmentation, there is an increasing demand for melanin production inhibitors. Existing topical hyperpigmentation treatments are based on accelerated epithelial turnover, which relies on removing melanin from the outer layer of the skin. Such treatments include exfoliants (e.g., AHAs or BHAs), anti-melanin production agents (including kojic acid and hydroquinone, which act to inhibit melanin synthesis), melanosome blockers (e.g., nicotinamide) (which act to block the transfer of melanosomes from melanocytes to keratinocytes) and anti-inflammatory agents (e.g., stearyl glycyrrhizinate) (which acts to block skin inflammation), and any combination thereof.

[0007] Although the topical treatments mentioned above are effective in lightening skin tone, they are less effective in treating hyperpigmentation disorders. This is due to their mode of action, which primarily acts on epithelial melanosis. In addition, many currently available products for treating hyperpigmentation may contain ingredients that have been associated with possible adverse effects, including hydroquinone and kojic acid. As an alternative to the above topical treatments, facial laser treatments (including Q-switched nanosecond pulse width technology) are considered to be effective methods for reducing or eliminating dermal melanosis, but these treatments are expensive and invasive (Watanabe, 2014).

[0008] Therefore, there remains a need for highly safe and effective anti-hyperpigmentation compositions that overcome the limitations and disadvantages of the prior art. SUMMARY OF THE INVENTION

[0010] It has now been found that dermal fibroblasts participate in the absorption of melanosomes, and that melanin can be retained in fibroblasts and can be degraded at a slow rate, resulting in a persistently darker skin tone. 3-O-ethyl ascorbic acid, an ascorbic acid derivative, has also been identified as being particularly effective in reducing melanin content in fibroblasts in a biomimetic dermal melanosis model, and provides a potent anti-hyperpigmentation effect in reducing skin melanin. It has further been found that the combination of 3-O-ethyl ascorbic acid together with certain anti-melanogenic agents provides superior anti-hyperpigmentation properties, providing superior results in reducing melanin content in fibroblasts.

[0011] Thus, in one aspect, there is provided a composition for preventing and / or treating a skin pigmentation disorder, the composition comprising:

[0012] a) ascorbic acid or its derivatives and / or salts; and

[0013] b) an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or a salt thereof;

[0014] Its amount effectively degrades melanin in dermal fibroblasts.

[0015] In some embodiments, component a) is selected from the group consisting of: 3-O-ethyl ascorbic acid, L-ascorbic acid, 2-O-ethyl ascorbic acid, ascorbyl glucoside, ascorbyl tetraisopalmitate, magnesium ascorbyl phosphate, sodium ascorbyl phosphate, 3-glyceryl ascorbate, diglyceryl ascorbate, hexyl 3-glyceryl ascorbate, myristyl 3-glyceryl ascorbate, and lauryl glyceryl ascorbate. In some embodiments, component a) is selected from the group consisting of: 3-O-ethyl ascorbic acid, L-ascorbic acid, 2-O-ethyl ascorbic acid, ascorbyl tetraisopalmitate, sodium ascorbyl phosphate, caprylyl-2-glyceryl ascorbate, diglyceryl ascorbate, hexyl 3-glyceryl ascorbate, myristyl 3-glyceryl ascorbate, and lauryl glyceryl ascorbate. In some embodiments, component a) is 3-O-ethyl ascorbic acid.

[0016] In some embodiments, component b) is resveratrol. In some embodiments, component b) is pterostilbene. In some embodiments, component b) is sclareolide. In some embodiments, component b) is 4-n-butylresorcinol.

[0017] In some embodiments, the amount of component a) ranges from 0.01% to 50% by weight of the composition, optionally 0.5% to 5% by weight of the composition, optionally 0.5% to 1.5% by weight of the composition.

[0018] In some embodiments, the amount of component b) ranges from 0.01% to 5% by weight of the composition, optionally 0.1% to 0.5% by weight of the composition, or optionally 0.2% to 0.35% by weight of the composition.

[0019] In some embodiments, the weight ratio of component a) to component b) ranges from 2:1 to 200:1, optionally 2:1 to 50:1, or optionally 3:1 to 5:1.

[0020] In some embodiments, the composition does not contain any anti-melanogenic agents other than a) and b).

[0021] In some embodiments, the composition comprises 3-O-ethyl ascorbic acid as component a) and resveratrol or 4-n-butylresorcinol as component b), and wherein the weight ratio of component a) to component b) ranges from 5:1 to 50:1.

[0022] In some embodiments, the composition is for topical administration.

[0023] In some embodiments, the composition comprises one or more of the following: an anti-inflammatory agent, a sunscreen, an antioxidant, an anti-aging peptide, a desquamating agent, and an exfoliating agent.

[0024] In some embodiments, the composition comprises one or more of the following: water, alkyl benzoate, PPG-3 myristyl ether, hydroxyacrylate / sodium acryloyldimethyl taurate copolymer, ethylhexylglycerin, phenoxyethanol, methylpropanediol, betaine, sclerotium gum, sodium polyacryloyldimethyl taurate, disodium EDTA, sodium hyaluronate, bis-PEG-18 methyl ether dimethyl silane, PEG-12 dimethicone, glycerin, chlorphenesin, sodium metabisulfite, lactic acid, dicaprylyl carbonate, and polyacrylate crosspolymer-6.

[0025] In some embodiments, the composition is in the form of a cream, lotion, paste, wax, liquid, semisolid composition, or a sprayable composition.

[0026] In another aspect, there is provided a method of preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual, or a method of degrading melanin in dermal fibroblasts in an individual, comprising administering to the individual an effective amount of a composition as defined herein.

[0027] In another aspect, there is provided use of a composition as defined herein for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual, or for degrading melanin in dermal fibroblasts of an individual.

[0028] In another aspect, there is provided a composition as defined herein for use in preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder.

[0029] In another aspect, there is provided a composition as defined herein for use in degrading melanin in dermal fibroblasts.

[0030] In some embodiments, the skin pigmentation disorder is selected from skin hyperpigmentation, melasma, solar lentigo, nevus of Ota, nevus of Ito, and Mongolian spots.

[0031] In some embodiments, the composition is topically applied to the skin of the individual.

[0032] In another aspect, a method of preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual, or degrading melanin in dermal fibroblasts of an individual is provided, comprising administering an effective amount of

[0033] a) ascorbic acid or its derivatives and / or salts,

[0034] Its amount effectively degrades melanin in dermal fibroblasts.

[0035] In some embodiments, ascorbic acid or a derivative and / or salt thereof is used in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0036] In another aspect, ascorbic acid or a derivative and / or salt thereof is provided for use in preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder, wherein the ascorbic acid or a derivative and / or salt thereof is administered in an amount effective to degrade melanin in dermal fibroblasts.

[0037] In some embodiments, ascorbic acid or a derivative and / or salt thereof is used in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0038] In another aspect, ascorbic acid or a derivative and / or salt thereof is provided for use in degrading melanin in dermal fibroblasts.

[0039] In some embodiments, ascorbic acid or a derivative and / or salt thereof is used in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0040] In another aspect, an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof, is provided for use in preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, wherein the anti-melanin production agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

[0041] In another aspect, an anti-melanin generating agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof, is provided for use in degrading melanin in dermal fibroblasts, wherein the anti-melanin generating agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

[0042] In another aspect, there is provided a use of ascorbic acid or its derivatives and / or salts for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, or for degrading melanin in dermal fibroblasts, wherein the ascorbic acid or its derivatives and / or salts are administered in an amount effective to degrade melanin in dermal fibroblasts.

[0043] In some embodiments, ascorbic acid or a derivative and / or salt thereof is administered in combination with an anti-melaninogenic agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0044] In another aspect, there is provided a use of an anti-melanin generating agent for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, or for degrading melanin in dermal fibroblasts, wherein the anti-melanin generating agent is selected from 4-n-butylresorcinol, resveratrol, pterostilbene and clareolide, or salts thereof, wherein the anti-melanin generating agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

[0045] In some embodiments, the skin pigmentation disorder is selected from skin hyperpigmentation, melasma, solar lentigo, nevus of Ota, nevus of Ito, and Mongolian spots.

[0046] In some embodiments, ascorbic acid, or a derivative and / or salt thereof, is topically applied to the skin of a subject.

[0047] In some embodiments, the anti-melanogenesis agent is topically administered to the skin of the individual.

[0048] In another aspect, there is provided a non-therapeutic method of lightening the skin of an individual, comprising administering to said individual an effective amount of a composition as defined herein.

[0049] In some embodiments, the composition is topically applied to the skin of a subject.

[0050] In another aspect, a non-therapeutic method of lightening the skin of an individual is provided, comprising administering to the individual an amount of ascorbic acid, or a derivative and / or salt thereof, effective to degrade melanin in dermal fibroblasts.

[0051] In some embodiments, ascorbic acid, or a derivative and / or salt thereof, is topically applied to the skin of the individual.

[0052] In some embodiments, ascorbic acid or a derivative and / or salt thereof is administered in combination with an anti-melaninogenic agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0053] In some embodiments, the anti-melanogenesis agent is topically administered to the skin of the individual. BRIEF DESCRIPTION OF THE DRAWINGS

[0055] Figure 1 provides the melanin content of dermal fibroblasts (HS68) treated with melanosome-conditioned medium derived from melanoma cells (B16-F10). Figure 1A ) and cell viability ( Figure 1B). Data are presented as mean ± SEM of three independent experiments (n=3). *p<0.05 when compared with untreated control by student t test.

[0056] FIG2 provides the melanin content of dermal fibroblasts (HS68) pretreated with melanosome-conditioned medium and co-cultured with reconstituted skin for a duration of 7 days ( Figure 2A ) and cell / tissue survival rate ( Figure 2B ). Data are presented as mean ± SEM of two independent experiments (n=4). *p<0.05 when compared with the control group by student t test.

[0057] Figure 3 Illustration showing the biomimetic dermal melanosis skin model.

[0058] Figure 4 Provides a graphical representation of the inhibitory effect of 3-O-ethylascorbic acid on melanin content in melanosome-internalizing fibroblasts. Data are presented as mean ± SEM of two independent experiments (n=2). *p<0.05 when compared to untreated controls by one-way ANOVA.

[0059] Specific description

[0060] definition

[0061] Unless specifically defined otherwise, all technical and scientific terms used herein should be considered to have the same meaning as commonly understood by one of ordinary skill in the art (eg, chemistry, biology, etc.).

[0062] The present disclosure relates to the entire contents of certain documents incorporated herein by reference. If there are any inconsistent teachings between the teachings of the present disclosure and the contents of those documents, the teachings of the present disclosure shall prevail.

[0063] It will be appreciated that if any prior art publication is referred to herein, this reference should not constitute an admission that the publication forms part of the common general knowledge in the art.

[0064] As used herein, the term "and / or", such as "X and / or Y", should be understood to mean "X and Y" or "X or Y", and should be considered to provide clear support for both meanings or either meaning.

[0065] As used herein, the term about refers to + / - 10% of the specified value unless otherwise stated.

[0066] Throughout this specification, references to single steps, compositions of matter, groups of steps, or groups of compositions of matter are to be considered to include both one and multiple (i.e., one or more) of those steps, compositions of matter, groups of steps, or groups of compositions of matter, unless expressly stated otherwise or the context requires otherwise. Thus, the singular forms "a," "an," and "the" as used herein include plural aspects unless the context clearly dictates otherwise. For example, reference to "a" includes a single as well as two or more; reference to "an" includes a single as well as two or more; reference to "the" includes a single as well as two or more, and so forth.

[0067] Unless otherwise indicated, terms such as "first," "second," and the like are used herein merely as labels and are not intended to impose order, position, or hierarchical requirements on the items to which these terms refer. In addition, reference to a "second" item does not require or exclude the presence of a smaller numbered item (e.g., a "first" item) and / or a larger numbered item (e.g., a "third" item).

[0068] As used herein, when used with a list of items, the phrase "at least one of..." means that different combinations of one or more of the listed items may be used and only one of the items in the list may be required. The item may be a specific object, thing, or category. In other words, "at least one of..." means that any combination or number of items from the list may be used, but all items in the list may not be required. For example, "at least one of item A, item B, and item C" may mean: item A; item A and item B; item B; item A, item B, and item C; or item B and item C. In some cases, "at least one of item A, item B, and item C" may mean, for example and without limitation, two items A, one item B, and ten items C; four items B and seven items C; or some other suitable combination.

[0069] As used herein, the word "comprise" and other forms of the word (such as "comprising" and "comprises") mean including but not limited to, and are not intended to exclude, for example, other additives, components, integers or steps.

[0070] As used herein, the term "subject" refers to any organism susceptible to a disease or condition. For example, the subject can be an animal, a mammal, a primate, a livestock animal (e.g., sheep, cattle, horses, pigs), a companion animal (e.g., dogs, cats), or a laboratory animal (e.g., mice, rabbits, rats, guinea pigs, hamsters). In one embodiment, the subject is a mammal. In one embodiment, the subject is a human. In one embodiment, the subject is a non-human animal.

[0071] As used herein, the term "treating" includes alleviating the symptoms associated with a particular disorder or condition.

[0072] As used herein, the term "prevention" includes preventing a particular disorder or condition.

[0073] The present disclosure also includes all steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations of said steps or features or any two or more thereof.

[0074] Each embodiment of the present disclosure applies mutatis mutandis to each and every other embodiment, unless expressly stated otherwise or the context requires otherwise.

[0075] Composition

[0076] As discussed above, the present disclosure relates to compositions and methods for preventing and / or treating skin pigmentation disorders by degrading melanin in dermal fibroblasts. The compositions have an anti-melanogenic agent that, when used in combination with ascorbic acid or its derivatives and / or salts, provides effective degradation of skin melanin.

[0077] Therefore, in one aspect, a composition for preventing and / or treating skin pigmentation disorders is provided, the composition comprising: (a) ascorbic acid or a derivative and / or salt thereof; and (b) an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and clareolide, or a salt thereof; in an amount effective to degrade melanin in dermal fibroblasts.

[0078] As used herein, the term "anti-melanin production agent" refers to a substance having melanin production inhibitory activity. Without intending to be limited by theory, in some embodiments the inhibitory activity may act by inhibiting tyrosinase activity. Tyrosinase is a copper-containing enzyme present in plant and animal tissues that catalyzes the production of melanin and other pigments from tyrosine by oxidation.

[0079] Ascorbic acid derivatives and / or salt forms thereof included in the present invention can be ascorbic acid or any known derivatives and / or salt forms thereof of ascorbic acid. Ascorbic acid derivatives and / or salt forms thereof can be chemically synthesized or extracted from any natural source known to those skilled in the art of synthetic organic chemistry, medicinal chemistry and phytochemistry. It will be appreciated by those skilled in the art that there are many methods known in the art and described in various patents and publications that can be used to obtain ascorbic acid derivatives and / or salt forms thereof to be used in practicing the present invention.

[0080] In some embodiments, the ascorbic acid derivative and / or its salt form is selected from: 3-O-ethyl ascorbic acid (3-EAA), L-ascorbic acid, 2-O-ethyl ascorbic acid, ascorbyl glucoside, ascorbyl tetraisopalmitate, magnesium ascorbyl phosphate, sodium ascorbyl phosphate, capryl-2-glyceryl ascorbate, diglyceryl ascorbate, 3-glyceryl ascorbate, hexyl 3-glyceryl ascorbate, myristyl 3-glyceryl ascorbate, and lauryl glyceryl ascorbate.

[0081] In some embodiments, the ascorbic acid derivative and / or its salt form is selected from: 3-O-ethyl ascorbic acid, L-ascorbic acid, 2-O-ethyl ascorbic acid, ascorbyl tetraisopalmitate, sodium ascorbyl phosphate, capryl-2-glyceryl ascorbate, 3-glyceryl ascorbate, diglyceryl ascorbate, hexyl 3-glyceryl ascorbate, myristyl 3-glyceryl ascorbate and laurylglyceryl ascorbate.

[0082] In some embodiments, the ascorbic acid derivative and / or its salt form is 3-O-ethyl ascorbic acid (3-EAA). The chemical structure of 3-O-ethyl ascorbic acid is described as follows:

[0083]

[0084] 3-O-ethyl ascorbic acid can be chemically synthesized or extracted from any natural source known to those skilled in the art of synthetic organic chemistry, medicinal chemistry and phytochemistry or obtained from any commercial source. Commercial sources for obtaining 3-O-ethyl ascorbic acid include, but are not limited to, Nippon Fine Chemical Co. Ltd. (trade name: ethyl ascorbic acid), Corum Inc. (trade name: Et-VCTM), Kimika, LLC (trade name: Neosome OL EAA) and Naturalis SRL (trade name: Nio-VCS).

[0085] The above anti-melanin production agents may also be chemically synthesized or extracted from any natural source known to those skilled in the art of synthetic organic chemistry or medicinal chemistry and phytochemistry or obtained from any commercial source.

[0086] In some embodiments, the anti-melanin production agent is resveratrol. The chemical structure of resveratrol is described as follows:

[0087]

[0088] Commercial sources for obtaining resveratrol include, but are not limited to, Spec-Chem Industry (trade names: Resveratrol), DSM Nutritional Products LLC (trade name: -Fade), Salvona Technologies Inc. (trade name: Salphere Resveratrol), Biogenics Inc. (trade name: BioGenic Resveratrol-200), and BCR Bio Component Research (trade name: Resveratrol BT).

[0089] In some embodiments, the anti-melanin production agent is pterostilbene. The chemical structure of pterostilbene is described as follows:

[0090]

[0091] Commercial sources for obtaining pterostilbene include, but are not limited to, Glanbia Nutritional (trade name: pTeroPure), Sabinsa Cosmetics (trade name: 90%) and Spec-Chem Industry (trade name: PTB).

[0092] In some embodiments, the anti-melanin production agent is sclareolide. The chemical structure of sclareolide is described as follows:

[0093]

[0094] Commercial sources for obtaining sclareolide include, but are not limited to, Symrise (trade name: 2036) and MMP International (trade name: Clary Sage FE).

[0095] In some embodiments, the anti-melanin generation agent is 4-n-butylresorcinol. The chemical structure of 4-n-butylresorcinol is described as follows:

[0096]

[0097] Commercial sources for obtaining 4-n-butylresorcinol include, but are not limited to, Kumar Organic Product Limited (trade name: Kopcino) and Biochempro Incorporation (trade name: Actosome Whitenol).

[0098] Ascorbic acid or its derivatives and / or salts and anti-melanin generation agents can be provided in any suitable form. For example, in some embodiments, one or more of the substances can be provided in the form of encapsulation. Encapsulation can be used to improve water dispersibility, chemical stability and bioavailability. In some other embodiments, some or all of the substances are not provided in the form of encapsulation.

[0099] The compositions of the present disclosure may be effective in degrading melanin in dermal fibroblasts in any combination providing component a) and an anti-melanogenic agent, component b). Beneficially, the combination of a) and b) has been found to act synergistically in providing enhanced melanin degradation relative to the individual components.

[0100] In some embodiments, component a) is 3-O-ethyl ascorbic acid and component b) is resveratrol. Beneficially, the inventors have demonstrated that 3-O-ethyl ascorbic acid and resveratrol work synergistically to inhibit melanin production activity in the epidermis to reduce the melanin content in fibroblasts of melanin internalization in the skin model of dermal melanosis. In some other embodiments, component a) is 3-O-ethyl ascorbic acid and component b) is pterostilbene. In other embodiments, component a) is 3-O-ethyl ascorbic acid and component b) is sclareolide. In some other embodiments, component a) is 3-O-ethyl ascorbic acid and component b) is 4-n-butylresorcinol.

[0101] The compositions of the present disclosure comprise component a) and component b) in an amount effective to degrade melanin in dermal fibroblasts. For example, the compositions of the present disclosure may comprise component a) in a range of 0.01% to 50%, or 0.01% to 40%, or 0.01% to 30%, or 0.01% to 20%, or 0.01% to 10%, or 0.01% to 5%, or 0.05% to 10%, or 0.15 to 10%, or 0.25 to 10%, or 0.50 to 10%, or 0.50 to 7.5%, or 0.5 to 5%, or 0.5 to 4%, or 0.5 to 3%, or 0.5 to 2%, or 1 to 2%, or 0.5 to 1.5% by weight of the composition.

[0102] In some embodiments, the composition comprises component a) in an amount ranging from 0.01% to 50% by weight of the composition, optionally 0.5% to 5% by weight of the composition, optionally 0.5% to 1.5% by weight of the composition.

[0103] In some embodiments, the compositions of the present disclosure comprise 3-O-ethyl ascorbic acid in a range of 0.01% to 50%, or 0.01% to 40%, or 0.01% to 30%, or 0.01% to 20%, or 0.01% to 10%, or 0.01% to 5%, or 0.05% to 10%, or 0.15 to 10%, or 0.25 to 10%, or 0.50 to 10%, or 0.50 to 7.5%, or 0.5 to 5%, or 0.5 to 4%, or 0.5 to 3%, or 0.5 to 2%, or 1 to 2%, or 0.5 to 1.5%, by weight of the composition.

[0104] The compositions of the present disclosure may comprise component b) in a range of 0.01 to 5%, or 0.01 to 4.5%, or 0.01 to 4%, or 0.01 to 3.5%, or 0.02 to 5%, or 0.03 to 5%, or 0.05 to 5%, or 0.05 to 4.5%, or 0.05 to 4%, or 0.05 to 3.5%, or 0.05 to 3%, or 0.05 to 2.5%, or 0.05 to 2.0%, or 0.05 to 1%, or 0.05 to 0.3%, or 0.1 to 2%, or 0.1 to 1%, or 0.1 to 0.5%, or 0.1 to 0.4%, or 0.1 to 0.3%.

[0105] In some embodiments, the composition comprises component b) in an amount ranging from 0.01% to 5% by weight of the composition, optionally 0.1% to 0.5% by weight of the composition, or optionally 0.2% to 0.35% by weight of the composition.

[0106] In some embodiments, the compositions of the present disclosure include resveratrol in an amount ranging from 0.01 to 5%, or 0.01 to 4.5%, or 0.01 to 5%, or 0.01 to 5%, or 0.02 to 5%, or 0.03 to 4%, or 0.05 to 3.5%, or 0.05 to 4.5%, or 0.05 to 4%, or 0.05 to 3.5%, or 0.05 to 3%, or 0.05 to 2.5%, or 0.05 to 2.0%, or 0.05 to 1%, or 0.05 to 0.3%, or 0.1 to 2%, or 0.1 to 1%, or 0.1 to 0.5%, or 0.1 to 0.4%, or 0.1 to 0.3%, or about 0.05%, or about 1%. In some embodiments, the resveratrol is encapsulated resveratrol.

[0107] In some embodiments, the compositions of the present disclosure comprise 4-n-butylresorcinol in an amount ranging from 0.01 to 5%, or 0.01 to 4.5%, or 0.01 to 4%, or 0.01 to 3.5%, or 0.02 to 5%, or 0.03 to 5%, or 0.05 to 5%, or 0.05 to 4.5%, or 0.05 to 4%, or 0.05 to 3.5%, or 0.05 to 3%, or 0.05 to 2.5%, or 0.05 to 2.0%, or 0.05 to 1%, or 0.05 to 0.3%, or 0.1 to 2%, or 0.1 to 1%, or 0.1 to 0.5%, or 0.1 to 0.4%, or 0.1 to 0.3%, or about 0.3%. In some embodiments, the 4-n-butylresorcinol is encapsulated 4-n-butylresorcinol.

[0108] In some embodiments, the compositions of the present disclosure comprise pterostilbene in an amount ranging from 0.01 to 5%, or 0.01 to 4.5%, or 0.01 to 4%, or 0.01 to 3.5%, or 0.02 to 5%, or 0.03 to 5%, or 0.05 to 5%, or 0.05 to 4.5%, or 0.05 to 4%, or 0.05 to 3.5%, or 0.05 to 3%, or 0.05 to 2.5%, or 0.05 to 2.0%, or 0.05 to 1%, or 0.05 to 0.3%, or 0.1 to 2%, or 0.1 to 1%, or 0.1 to 0.5%, or 0.1 to 0.4%, or 0.1 to 0.3%. In some embodiments, the pterostilbene is encapsulated pterostilbene.

[0109] In some embodiments, the compositions of the present disclosure comprise sclareolide in an amount ranging from 0.01 to 5%, or 0.01 to 4.5%, or 0.01 to 4%, or 0.01 to 3.5%, or 0.02 to 5%, or 0.03 to 5%, or 0.05 to 5%, or 0.05 to 4.5%, or 0.05 to 4%, or 0.05 to 3.5%, or 0.05 to 3%, or 0.05 to 2.5%, or 0.05 to 2.0%, or 0.05 to 1%, or 0.05 to 0.3%, or 0.1 to 2%, or 0.1 to 1%, or 0.1 to 0.5%, or 0.1 to 0.4%, or 0.1 to 0.3%. In some embodiments, the sclareolide is encapsulated sclareolide.

[0110] The composition of the present disclosure may comprise an amount of component a) and an amount of component b) in a weight ratio effective to degrade melanin in dermal fibroblasts. For example, the composition of the present disclosure may comprise an amount of component a) in a greater amount relative to component b), such as a weight ratio of 200:1, or 180:1, or 160:1, or 140:1, or 120:1, or 100:1, or 80:1, or 60:1, or 50:1, or 40:1, or 30:1, or 20:1, or 10:1, or 8:1, or 6:1, or 5:1, or 4:1, or 3:1, or 2:1, or greater than 1:1.

[0111] In some embodiments, the composition comprises component a) to component b) in a weight ratio ranging from 2:1 to 200:1, 2:1 to 100:1, 2:1 to 50:1, 2:1 to 30:1, 3:1 to 25:1, 10:1 to 25:1, 3:1 to 10:1, or 3:1 to 5:1.

[0112] In some embodiments, the composition comprises component a) to component b) in a weight ratio ranging from 2:1 to 200:1, optionally 2:1 to 50:1, or optionally 3:1 to 5:1.

[0113] In some embodiments, the composition comprises 0.5 to 5 wt % of 3-O-ethyl ascorbic acid as component a) and comprises 0.01 to 0.5 wt % of resveratrol as component b), and optionally wherein the weight ratio of component a) to component b) ranges from 3:1 to 25:1, or 4:1 to 25:1, or 10:1 to 25:1, or about 20:1. In some embodiments, the resveratrol is encapsulated resveratrol.

[0114] In some embodiments, the composition comprises 1 to 2 wt% of 3-O-ethyl ascorbic acid as component a) and comprises 0.1 to 0.2 wt% of resveratrol as component b), and optionally wherein the weight ratio of component a) to component b) ranges from 3:1 to 25:1, or 4:1 to 25:1, or 10:1 to 25:1, or about 20:1. In some embodiments, the resveratrol is encapsulated resveratrol.

[0115] In some embodiments, the composition comprises 0.5 to 5 wt % of 3-O-ethyl ascorbic acid as component a) and comprises 0.1 to 0.5 wt % of 4-n-butylresorcinol as component b), and optionally wherein the weight ratio of component a) to component b) ranges from 3: 1 to 10: 1, 4: 1 to 10: 1, or about 6.7: 1. In some embodiments, the 4-n-butylresorcinol is encapsulated 4-n-butylresorcinol.

[0116] In some embodiments, the composition comprises 1 to 2 wt% of 3-O-ethyl ascorbic acid as component a) and comprises about 0.3 wt% of 4-n-butylresorcinol as component b), and optionally wherein the weight ratio of component a) to component b) ranges from 3:1 to 10:1, or 4:1 to 10:1, or about 6.7:1. In some embodiments, the 4-n-butylresorcinol is encapsulated 4-n-butylresorcinol.

[0117] In some embodiments, the composition comprises 0.5 to 5 wt % of 3-O-ethyl ascorbic acid as component a) and comprises 0.01 to 0.5 wt % of sclareolide as component b), and optionally wherein the weight ratio of component a) to component b) ranges from 3: 1 to 25: 1, or 4: 1 to 25: 1, or 10: 1 to 25: 1, or about 20: 1. In some embodiments, the sclareolide is encapsulated sclareolide.

[0118] In some embodiments, the composition comprises 1 to 2 weight % of 3-O-ethyl ascorbic acid as component a) and comprises 0.1 to 0.2 weight % of sclareolide as component b), and optionally wherein the weight ratio of component a) to component b) ranges from 3: 1 to 25: 1, or 4: 1 to 25: 1, or 10: 1 to 25: 1, or about 20: 1. In some embodiments, the sclareolide is encapsulated sclareolide.

[0119] In some embodiments, the composition comprises 0.5 to 5 wt % of 3-O-ethyl ascorbic acid as component a) and comprises 0.01 to 0.5 wt % of pterostilbene as component b), and optionally wherein the weight ratio of component a) to component b) ranges from 3: 1 to 25: 1, or 4: 1 to 25: 1, or 10: 1 to 25: 1, or about 20: 1. In some embodiments, the pterostilbene is encapsulated pterostilbene.

[0120] In some embodiments, the composition comprises 1 to 2 weight % of 3-O-ethyl ascorbic acid as component a) and comprises 0.1 to 0.2 weight % of pterostilbene as component b), and optionally wherein the weight ratio of component a) to component b) ranges from 3: 1 to 25: 1, or 4: 1 to 25: 1, or 10: 1 to 25: 1, or about 20: 1. In some embodiments, the pterostilbene is encapsulated pterostilbene.

[0121] In some embodiments, the composition comprises 3-O-ethyl ascorbic acid as component a) and resveratrol or 4-n-butylresorcinol as component b), and wherein the weight ratio of component a) to component b) ranges from 5:1 to 50:1.

[0122] The synthesis and degradation of tyrosinase are closely related to its function, which is an important parameter in the regulation of melanin synthesis. Therefore, in some embodiments, the compositions of the present disclosure may also include additional substances from any natural or synthetic sources that act via any mechanism, including downregulating tyrosinase and its activity, to inhibit or reduce melanin production, including but not limited to p-coumaric acid; Biochanin A; Calycosin; TMBC; Taxifolin; Luteolin; 2,5-DHAP; Linderanolide B; and Subamolide A.

[0123] However, it has also been found that relatively simple formulations containing a) and b) provide unexpectedly effective results. Therefore, in some other embodiments, the composition does not contain any anti-melanin generation agent other than a) and b). In some embodiments, the composition consists of components a) and b) and one or more carriers and / or excipients or consists essentially of them.

[0124] The compositions can be formulated for any desired method of administration. For example, the compositions of the present disclosure can be formulated as topical compositions suitable for topical administration to the skin.

[0125] The compositions of the present disclosure may also contain various other additives in addition. For example, the compositions of the present disclosure may include any number of known additional cosmetic or pharmaceutical substances. In some embodiments, the composition is composed of components a) and b), one or more carriers and / or excipients, and optionally one or more additional cosmetic or pharmaceutical substances that are not anti-melanin generation agents or is essentially composed of them.

[0126] The CTFA Cosmetic Ingredient Handbook, 2nd edition, 1992 describes a variety of non-limiting cosmetic and pharmaceutical ingredients commonly used in the skin care industry that may be suitable for use in the compositions of the present invention. Examples of suitable formulation ingredients include antioxidants, binders, biological additives, buffers, colorants, thickeners, pearlescent agents, polymers, astringents, silicones, odor absorbers, gelling agents (e.g., hydrophilic or lipophilic gelling agents), fragrances, humectants, fillers, solvents, sunscreens, skin lighteners, skin tanning agents, fragrances, conditioners, exfoliants, solubilizers (e.g., methylpropanediol), sunscreens, vitamins, colorants, pH adjusters (e.g., sodium hydroxide), preservatives (e.g., ethylhexylglycerin, phenoxyethanol, or hydroxyacetophenone), rheology modifiers, foaming agents, surfactants, emollients, natural extracts, essential oils, skin sensates, scalp soothing agents, and scalp healing agents.

[0127] The compositions of the present disclosure may also include other skin conditioning agents, including but not limited to vitamins or vitamin derivatives and macro- or micro-capsules containing vitamins or their derivatives, cosmetic peptides, plant extracts, polyphenolic compounds, sunscreen additives, sunscreen agents, sensory additives or texturizing agents, natural extracts, and essential oils.

[0128] In some embodiments, the compositions of the present disclosure include one or more of the following: anti-inflammatory agents, sunscreens, antioxidants, anti-aging peptides, desquamation agents, and exfoliants. The compositions may be formulated with one or more other components (e.g., hyaluronic acid and / or aloe vera gel) that are beneficial for treating skin conditions.

[0129] In some embodiments, the composition comprises a pharmaceutically or cosmetically acceptable carrier or vehicle, and / or comprises one or excipient.

[0130] The pharmaceutically or cosmetically acceptable vehicle can be any pharmaceutically or cosmetically acceptable vehicle, including pharmaceutically and / or dermatologically acceptable carriers, including surfactants. Pharmaceutically and / or dermatologically acceptable carriers are preferably compatible with skin, nails, mucous membranes, tissues and / or hair, and can include any conventionally used pharmaceutical or dermatological carriers that meet these requirements. Carriers and / or vehicles can help ascorbic acid or its derivatives and / or salts or anti-melanin generating agents to be formulated into suitable medicines or cosmetic preparations. Those of ordinary skill in the art can easily select this carrier. Carriers include but are not limited to emollients, emulsifiers and rheology modifiers.

[0131] Suitable emollients for use in the vehicle or carrier of the compositions of the present invention include, for example, C 12-15 Alkyl benzoate, PPG-3 myristic ether, liquid paraffin and dimethicone. Emulsifiers suitable for use in the vehicle or carrier of the composition of the present invention include, for example, hydroxyethyl acrylate / sodium acryloyldimethyl taurate copolymer, Ceteareth-20 (cetostearyl alcohol, which is a polyethylene glycol ether of cetyl alcohol). Rheology modifiers suitable for use in the vehicle or carrier of the composition of the present invention include, for example, carbomer, polyacrylate glyceryl and acrylate / C 10-30 Alkyl acrylate crosspolymer. Other suitable carriers include, but are not limited to, mineral oil, liquid petroleum, white petroleum, propylene glycol, polyoxyethylene and / or polyoxypropylene compounds, emulsifying wax, sorbitan monostearate, polysorbate 60 (polyoxyethylene (20) sorbitan monostearate), cetyl esters wax, cetyl alcohol, 2-octyldodecanol, benzyl alcohol, sodium hyaluronate, hyaluronic acid and water.

[0132] For topical administration, the compositions of the present invention can be formulated as, for example, ointments, liquids, lotions, pastes, foams, sprays, serums, masks, gels, hydrogels or transdermal patches, leave-on and rinse-off lotions or toners, skin ampoules, skin cleansers, face and eye makeup removers, leave-on hair conditioners or hair vitamins, hair fixatives, shower gels / creams, bathroom bar soaps, antiperspirants, deodorants, depilatories, all color cosmetics including, but not limited to, lipsticks and foundations, sunless tanning agents or sunscreen lotions, shampoos, conditioners, hair tonics, mousses, gels, liposomes or other forms suitable for topical use.

[0133] For example, ointments and creams can be formulated with a water base or an oil base, and appropriate thickeners, emulsifiers and / or gelling agents (e.g., oil-in-water emulsions, water-in-oil emulsions, or water-in-silicon emulsions) can be added. Lotions can be formulated with a water base or an oil base, and generally also contain one or more emulsifiers, stabilizers, dispersants, suspending agents, thickeners, colorants, or preservatives (e.g., methylparaben, propylparaben, phenoxyethanol). Ointments, creams, lotions, etc. can also be formulated to contain C1-3 Alkoxylated oils and waxes (eg, ethoxylated vegetable oils, ethoxylated jojoba oil / wax, ethoxylated lanolin oil, ethoxylated coconut oil, ethoxylated cocoa butter).

[0134] Any cosmetic product containing water (i.e., creams, gels, lotions, etc.) has the potential for bacterial, yeast, and mold growth, and may require preservatives in some embodiments. Exemplary preservatives for use in the compositions of the present disclosure include ethylhexylglycerin, phenoxyethanol, chlorphenesin, and sodium metabisulfite. Exemplary emollients include C 12-15 Alkyl benzoate, PPG-3 myristyl ether, isopropyl lauroyl sarcosinate, glycerin, and di-PEG-18 methyl ether dimethyl silane. Exemplary humectants are 1,2-hexanediol or betaine. Exemplary rheology modifiers include hydroxyethyl acrylate, sodium acryloyldimethyl taurate copolymer, or mixtures thereof.

[0135] In some embodiments, the compositions of the present disclosure include one or more of the following: water, alkyl benzoate, PPG-3 myristyl ether, hydroxyacrylate / sodium acryloyldimethyl taurate copolymer, ethylhexylglycerin, phenoxyethanol, methylpropanediol, betaine, sclerotium gum, sodium polyacryloyldimethyl taurate, disodium EDTA, sodium hyaluronate, bis-PEG-18 methyl ether dimethyl silane, PEG-12 dimethicone, glycerin, chlorphenesin, sodium metabisulfite, lactic acid, dicaprylyl carbonate, and polyacrylate crosspolymer-6.

[0136] The composition of the present invention can be a composition suitable for topical use. In some embodiments, the composition is used for topical application. Beneficially, ascorbic acid or its derivatives and / or salts and anti-melanin generation agents can penetrate the stratum corneum to reach the basal layer of the epidermis (where melanocytes are located) to effectively inhibit melanin generation. Therefore, it is expected that the topical composition comprising 3-O-ethyl ascorbic acid and anti-melanin generation agents will improve skin hyperpigmentation clinically, for example, the condition of pigmentation such as acne-induced and aging-induced pigmentation.

[0137] Topical use can be, for example, by incorporating ascorbic acid or its derivatives and / or salts and anti-melanin generation agents into leave-on or rinse-off products that are applied to the human body primarily for improved skin appearance and general aesthetic benefits. As used herein, "skin" is intended to include the outer surface of mammals, especially humans, and includes skin, scalp, and hair. Use of the compositions of the present disclosure can be, for example, by incorporation into leave-on compositions.

[0138] The composition can be in the form of a liquid, lotion, cream, foam, scrub, gel, soap bar, toner, or applied with an appliance or via a mask, pad or patch in any material form. Non-limiting examples of such compositions include leave-on skin lotions and creams in the form of oil-in-water / silicone-in-water emulsions, water-in-oil / water-in-silicone emulsions, and composite emulsions, serums, shampoos, conditioners, shower gels, toilet bar soaps, antiperspirants, deodorants, depilatories, lipsticks, foundations, mascara, sunscreen lotions, ointments, sprays, pastes, mousses, foams, gels, liposomes, or other forms suitable for topical use.

[0139] In some embodiments, the compositions of the present disclosure are in the form of a cream, lotion, paste, wax, liquid, semisolid composition, or a sprayable composition.

[0140] In the case where the composition is in the form of a spray, the carrier is suitably water, and the composition may further comprise a preservative such as benzyl alcohol (e.g. in an amount of 0.1-5 wt %), a cosolvent such as propylene glycol (e.g. in an amount of 1-20 wt %) and a surfactant such as PEG-40 hydrogenated castor oil (e.g. in an amount of 1-20 wt %).

[0141] Where the composition is in the form of a lotion, it may further comprise water in the presence of a suitable emulsifier. Examples of suitable emulsifiers are cetearyl alcohol-20, PEG-100 stearate, glyceryl stearate, sodium cetearyl sulfate. The composition may further comprise emollients and fragrances in the form of esters and / or hydrocarbons and / or silicones and / or vegetable oils, preservatives (e.g. benzyl alcohol at a level of, for example, 0.1-10%), and humectants and / or excipients (e.g. glycerol at a level of 0.5-10% and cyclomethicone at a level of 0.1-20%).

[0142] When the composition is in the form of an ointment, it may further comprise a conventional ointment base to which the active ingredient is added. The ointment base may be a paraffin, such as soft paraffin, or a combination of soft paraffin and liquid paraffin. Other ointment bases may also be used, such as a polyalkylene glycol (such as polyethylene glycol or polypropylene glycol) base. Other possible ingredients of the ointment composition include emulsifying wax (e.g., in an amount of 1-40%, preferably 5-40%), and one or more preservatives optionally present.

[0143] When the composition is in the form of a liquid soap, any known liquid soap may be used. The key ingredient of the preferred liquid soap, saponified fatty acids, is derived from palm kernel oil or coconut oil. The pH of the liquid soap base may be adjusted to a level suitable for topical use.

[0144] When the composition is an anhydrous cream, any suitable cream base may be used. Preferred cream bases are petrolatum creams and / or natural vegetable oils and waxes which introduce a consistency factor.

[0145] When the composition is a shampoo or a conditioner, in addition to the active ingredients, it may further comprise one or more of a surfactant, a thickener, a pH adjuster / buffer, an aesthetic additive, water, a conditioning agent, a preservative, and a moisturizer / vitamin. Shampoo may also comprise other active ingredients known to those skilled in the art, which have some advantages when included in a shampoo formulation or in a pharmaceutical combination for treating hyperpigmentation disorders. Shampoo may comprise a primary surfactant to clean the hair by removing dirt and other impurities and providing a flash of foam. A secondary surfactant may be included to provide a stable foam and reduce the irritation of the primary surfactant. Surfactants may be used that comprise a charged hydrophilic head group and a long hydrophobic alkyl chain tail. Surfactants are formulated to reduce the surface tension of the intermediate phase between dirt and hair, allowing dirt to be transported to an aqueous medium to be rinsed off from the hair and scalp. According to certain embodiments, examples of surfactants that may be included in shampoos include sodium laureth sulfate, ammonium lauryl sulfate, and sodium cocoyl isethionate. Examples of cosurfactants include cocamide MEA and cocamidopropyl betaine. Suitable thickeners or suspending agents include carbomer and PEG 150 distearate. Thickeners may be included to stabilize the shampoo during storage and / or prevent precipitation or pouring of pigments and silicones. Suitable pH adjusters or buffers include citric acid, tartaric acid and sodium hydroxide. pH adjusters or buffers are formulated to make the shampoo mild to the skin. Lower pH can make the hair dense and shiny and protect the surfactant from being hydrolyzed, and so, pH.

[0146] Beneficially, ascorbic acid or its derivatives and / or salts or anti-melanin generating agents can be prepared into stable topical compositions and applied on the skin. Therefore, ascorbic acid or its derivatives and / or salts or anti-melanin generating agents can exist in the form of capsules in the composition. Beneficially, encapsulation technology can be used to protect and enhance the transdermal delivery of ascorbic acid or its derivatives and / or salts or anti-melanin generating agents. Suitable encapsulation technology can be selected from liposomes, vesicles (niosomes), nanoemulsions and cyclodextrin inclusion technology. Suitable encapsulation technology can be manufactured by any means known in the art and is not limited to the present disclosure.

[0147] Ascorbic acid or its derivatives and / or salts or anti-melanin generation agents can be encapsulated in an encapsulation material comprising at least one emulsifier and a stabilizer. Encapsulation materials can include emulsifiers and auxiliary emulsifiers. Emulsifiers can be selected from glycerides of different fatty chains, including but not limited to citric acid glycerides, lactic acid glycerides, myristic acid glycerides, lauric acid glycerides, linoleic acid glycerides, oleic acid glycerides, or any combination thereof or any mixed glycerides of fatty acids, including citric acid esters, lactic acid esters, myristic acid esters, lauric acid esters, linoleic acid glycerides and / or oleic acid esters. In one embodiment, the emulsifier is citric acid glycerides / lactic acid glycerides / linolenic acid glycerides / oleic acid glycerides (mixed esters of citric acid, lactic acid, linoleic acid and oleic acid with glycerol). In some embodiments, the auxiliary emulsifier is di- or polyglycerol esters, which are selected from but not limited to monooleic acid diglyceride, monooleic acid polyglyceride, monolinoleic acid polyglyceride or any combination thereof. In a specific embodiment, the auxiliary emulsifier is monooleic acid diglyceride.

[0148] When used, the stabilizer may, for example, be selected from any type of organic substance that can stabilize an emulsifier. Suitable examples include alcohols or glycols, including but not limited to ethanol, isopropanol, propylene glycol, pentylene glycol, caprylyl glycol, especially pentylene glycol.

[0149] Therapeutic and non-therapeutic methods and uses

[0150] The present disclosure further relates to methods and uses for treating skin pigmentation disorders and degrading melanin in dermal fibroblasts using compositions and substances described herein. The compositions can be used to treat, control, prevent and improve skin hyperpigmentation in a safe and non-invasive manner. As discussed above, it has been found that fibroblasts in the dermis of the skin participate in the absorption of melanosomes, and melanin can be retained in fibroblasts and can be degraded at a slower rate, thus resulting in a continuous darker skin tone. Beneficially, the novel discovery of the inventors has become a strategy for treating hyperpigmentation by promoting the removal of melanin in fibroblasts, which can be used to supplement existing anti-melanin production strategies. Therefore, 3-O-ethyl ascorbic acid (a kind of ascorbic acid derivative) has been identified to be particularly effective in reducing the melanin content in fibroblasts in a bionic dermal melanosis model, where it provides a powerful anti-pigmentation hyperpigmentation effect in reducing skin melanin. It has further been found that the combination of 3-O-ethyl ascorbic acid together with certain anti-melanogenic agents provides excellent anti-hyperpigmentation properties, providing excellent results in reducing melanin content in fibroblasts. For example, 3-O-ethyl ascorbic acid and resveratrol have been shown to synergistically inhibit melanogenic activity in the epidermis and reduce melanin content in melanin internalizing fibroblasts in a human skin model of dermal melanosis.

[0151] In addition to reducing melanin content in fibroblasts, compositions comprising ascorbic acid or its derivatives and / or salts and anti-melanogenic agents may also act to promote depigmentation via alternative mechanisms, including but not limited to autophagic activity (which accelerates melanin clearance in the epidermal and dermal skin layers), and phagocytic activity of melanophils (which aids in melanosome clearance in the epidermal and dermal skin layers).

[0152] According to recent studies conducted by Ando et al. (2020), melanosomes were shown to be internalized by dermal fibroblasts via an actin-dependent endocytosis process. Under normal skin physiology, internalized melanosomes in keratinocytes can be differentiated and removed or removed via exfoliants (such as glycolic acid, salicylic acid, etc.). However, unlike keratinocytes in the epidermis, internalized melanosomes in fibroblasts are retained in the dermis and degraded at a slower rate, which can lead to persistent darker skin spots. The inventors have therefore determined that promoting the clearance of melanin in fibroblasts has become a new dimension and strategy for treating hyperpigmentation.

[0153] Thus, in one aspect, there is provided a method of preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual, or a method of degrading melanin in dermal fibroblasts in an individual, comprising administering to the individual an effective amount of a composition as defined herein.

[0154] Beneficially, administration of the disclosed compositions in the methods defined herein exhibited minimal negative effects on tissue viability of reconstructed skin and cell viability of fibroblasts, indicating that the inhibitory activity associated with melanin reduction mediated by the disclosed compositions is not due to cytotoxic effects.

[0155] In another aspect, there is provided the use of a composition as defined herein for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual or for degrading melanin in dermal fibroblasts of an individual.

[0156] In another aspect, there is provided a composition as defined herein for use in preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder.

[0157] In another aspect, there is provided a composition as defined herein for use in degrading melanin in dermal fibroblasts.

[0158] The compositions of the present disclosure can be used to treat skin pigmentation disorders including, but not limited to, Nevus of Ota, Mongolian spots, Café-au-lait spots, melasma, post-inflammatory hyperpigmentation, solar lentigo, Nevus of Ota, Nevus of Ito, progressive pigmented purpura, age spots, acanthosis nigricans, and incontinence pigmenti.

[0159] In some embodiments, the skin pigmentation disorder is selected from skin hyperpigmentation, melasma, solar lentigo, nevus of Ota, nevus of Ito, and Mongolian spots.

[0160] In some embodiments, the composition is administered topically, for example, to the skin of a subject.

[0161] As discussed above, the compositions of the present disclosure comprising ascorbic acid or its derivatives and / or salts as the sole active agent potently reduce melanin content in dermal fibroblasts that internalize melanosomes.

[0162] Thus, in another aspect, a method of preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual, or a method of degrading melanin in dermal fibroblasts of an individual is provided, comprising administering an effective amount of

[0163] a) ascorbic acid or its derivatives and / or salts,

[0164] Its amount effectively degrades melanin in dermal fibroblasts.

[0165] In some embodiments, ascorbic acid or a derivative and / or salt thereof is used in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0166] In another aspect, ascorbic acid or a derivative and / or salt thereof is provided for use in preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder, wherein the ascorbic acid or a derivative and / or salt thereof is administered in an amount effective to degrade melanin in dermal fibroblasts.

[0167] In some embodiments, ascorbic acid or a derivative and / or salt thereof is used in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0168] In another aspect, ascorbic acid or a derivative and / or salt thereof is provided for use in degrading melanin in dermal fibroblasts.

[0169] In some embodiments, ascorbic acid or a derivative and / or salt thereof is used in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0170] In another aspect, an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or a salt thereof, is provided for use in preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, wherein the anti-melanin production agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

[0171] In another aspect, an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof, is provided for use in degrading melanin in dermal fibroblasts, wherein the anti-melanin production agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

[0172] In another aspect, there is provided a use of ascorbic acid or its derivatives and / or salts for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, or for degrading melanin in dermal fibroblasts, wherein the ascorbic acid or its derivatives and / or salts are administered in an amount effective to degrade melanin in dermal fibroblasts.

[0173] In some embodiments, ascorbic acid or a derivative and / or salt thereof is administered in combination with an anti-melaninogenic agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0174] In another aspect, there is provided a use of an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or a salt thereof, for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, or for degrading melanin in dermal fibroblasts, wherein the anti-melanin production agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

[0175] In some embodiments, the skin pigmentation disorder is selected from skin hyperpigmentation, melasma, solar lentigo, nevus of Ota, nevus of Ito, and Mongolian spots.

[0176] In some embodiments, the anti-melanogenesis agent is topically administered to the skin of the individual.

[0177] The present disclosure is also directed to non-therapeutic methods for lightening skin, for example, the compositions of the present disclosure may be used in cosmetic applications to lighten skin color.

[0178] Thus, in one aspect, there is provided a non-therapeutic method of lightening the skin of an individual comprising administering an effective amount of a composition as defined herein.

[0179] In some embodiments, the composition is topically applied to the skin of a subject.

[0180] In another aspect, a non-therapeutic method of lightening the skin of an individual is provided, comprising administering to the individual an amount of ascorbic acid, or a derivative and / or salt thereof, effective to degrade melanin in dermal fibroblasts.

[0181] In some embodiments, ascorbic acid, or a derivative and / or salt thereof, is topically applied to the skin of the individual.

[0182] In some embodiments, ascorbic acid or a derivative and / or salt thereof is administered in combination with an anti-melaninogenic agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

[0183] In some embodiments, the anti-melanogenesis agent is topically administered to the skin of the individual.

[0184] The compositions of the present disclosure may, for example, be topical compositions to be applied to an external surface of an individual. In certain embodiments, the external surface may be one or more of the scalp, face, arms, legs, or torso of an individual. Beneficially, the compositions of the present disclosure exhibit effective skin bioavailability, which enables skin penetration to lighten skin tone, which has been demonstrated in human skin penetration and efficacy studies and in a biomimetic skin model of dermal melanosis.

[0185] In some embodiments, the composition of the present disclosure is applied once a day, or twice a day, or three times a day, or four times a day, for example, for a period of time sufficient to cause the desired level of hyperpigmentation improvement. Alternatively, if the composition provides the desired effect, the composition can be used at a lower frequency. For example, it can be applied every other day or once, twice, three times, or four times a week, or temporarily or as needed.

[0186] For example, the user can topically apply the composition of the present invention directly to the skin where pigmentation needs to be reduced by gently massaging the composition of the present invention into the desired area. In some embodiments, the composition of the present invention can be left on the skin or other area where pigmentation needs to be reduced between applications occurring on the same day.

[0187] The compositions of the present disclosure may be applied topically for any suitable period of time. For example, within a few days to a few weeks or within a few days to 10 weeks of initial application, the user may notice a reduction in pigmentation. Beneficially, topical treatment with the compositions of the present disclosure exhibits anti-hyperpigmentation efficacy by significantly reducing the skin tone of identified acne-related post-inflammatory hyperpigmentation in, for example, as little as 2 weeks of product application, and significantly reducing the skin tone of other identified pigmented spots after 8 weeks of product application.

[0188] It will be appreciated that the frequency with which the compositions of the present invention should be applied will vary with the desired efficacy. In some embodiments, the degree of therapeutic and / or cosmetic enhancement will vary directly with the total amount of the composition used. Topical application of the composition may be applied in any amount sufficient to cause the desired level of improvement in hyperpigmentation.

[0189] For example, the composition can be administered in an amount ranging from about 0.5 g to about 50 g, or from about 0.5 g to about 40 g, or from about 0.5 g to about 30 g, or from about 0.5 g to about 20 g, or from about 0.5 g to about 15 g, or from about 0.5 g to about 10 g, or from about 0.5 g to about 5 g, or from about 0.5 g to about 2.5 g, or from about 0.5 g to about 1 g.

[0190] When applied topically, the composition of the present invention can be applied to the area to be treated (e.g., face) by spraying, dabbing, rubbing, kneading, or massaging. For example, in one embodiment, the composition of the present invention can be provided in a liquid, lotion, cream, foam, scrub, gel, soap bar, toner, or applied with an appliance or via a mask, pad or patch in any material form, or other leave-on product formulation, which can preferably be applied at least daily.

[0191] In some embodiments, the "individual" is a mammal. In some embodiments, the "individual" is a human, such as an adult human.

[0192] Preparation of composition

[0193] The compositions of the present disclosure can be prepared by any suitable method known to those skilled in the art.

[0194] In some embodiments, the composition is prepared as a composition suitable for topical use.

[0195] In some embodiments, the composition is prepared as an emulsion.For example, the composition of the present disclosure can be prepared by combining the aqueous phase and the lipid phase, followed by homogenization.

[0196] The composition may include suitable solvents or substances that function to enhance absorption of other ingredients into the skin, moisturizers and anti-irritants, gums for producing a gel base that may have skin smoothing and soothing properties, emulsion stabilizers and viscosity control agents, substances that function to prevent the formulation from spoiling, moisture absorbers, emollients, hair / skin conditioners, substances that help improve the overall texture and surface of the skin care formulation, gelling agents, preservatives, substances that reduce the appearance of large pores, age spots and hyperpigmentation, including lactic acid, together with ascorbic acid or its derivatives and / or salts and anti-melanin generation agents, and homogenizing the mixture. In some embodiments, the ascorbic acid or its derivatives and / or salts and anti-melanin generation agents may be in encapsulated form.

[0197] In some embodiments, the composition is prepared as a composition suitable for topical use by adding a suitable solvent (such as methylpropanediol) that can enhance the absorption of other ingredients into the skin, a humectant and anti-irritant (such as betaine), a natural gum (such as sclerotium gum), an emulsion stabilizer and viscosity agent (such as sodium polyacryloyldimethyl taurate), an anti-deterioration agent (such as disodium EDTA), a moisture absorbent (such as sodium hyaluronate), a water-dispersible silicone (such as di-PEG-18 methyl ether dimethyl silane, which can act as an emollient and have increased foaming properties), a hair conditioner / skin conditioning agent (such as PEG-12 dimethicone), other suitable emollients (selected from C 12-15 Alkyl benzoate, PPG-3 myristyl ether and glycerin), a gelling agent (selected from hydroxyethyl acrylate / sodium acryloyldimethyl taurate copolymer), a substance that reduces the appearance of large pores, age spots and hyperpigmentation (including lactic acid), a preservative (selected from ethylhexylglycerin and phenoxyethanol), an antioxidant (including sodium metabisulfite), a biocide (including chlorphenesin) is combined with deionized water and ascorbic acid or its derivatives and / or salts and anti-melanin generation agents, and the mixture is homogenized. In some embodiments, the ascorbic acid or its derivatives and / or salts and anti-melanin generation agents are in encapsulated form.

[0198] Those skilled in the art will appreciate that the disclosure herein is susceptible to changes and modifications other than those specifically described. It should be understood that the disclosure includes all such changes and modifications. Example

[0199] The present disclosure is further illustrated by the following non-limiting examples.

[0200] Example 1 - Development of a dermal melanosis model

[0201] 1.1 Purpose

[0202] To understand the ability of fibroblasts to internalize melanosomes and to establish a biomimetic skin model of dermal melanosis.

[0203] 1.2 Methods

[0204] 1.2.1 Establishment of fibroblasts with internalized melanosomes

[0205] The concentration was 5x 10 5 Mouse melanoma cells B16-F10 (ATCC, USA) with 100 cells were seeded in T75 flasks and maintained for 5 days in complete Dulbecco's modified Eagle's medium (DMEM, from Nacalai, Japan) supplemented with 10% FBS, sodium pyruvate (1 mM), L-glutamine (2 mM), streptomycin (50 μg / mL) and penicillin (50 U / mL) at 37°C and 5% CO2. Then, the culture medium was centrifuged at 300 x g to remove cell debris and collect the supernatant containing melanosomes (melanosome conditioned medium). Then, 7 x 10 5 Individual human skin fibroblast HS-68 cells (ATCC, USA) were seeded in T25 flasks and maintained in DMEM medium supplemented with 10% FBS, sodium pyruvate (1mM), L-glutamine (2mM), streptomycin (50μg / mL) and penicillin (50U / mL) at 37°C and 5% CO2 for 24 hours. Then, the medium was replaced with melanosome conditioned medium and cultured for another 24 hours. Finally, the cell viability of the treated fibroblasts was determined by using the MTT assay. The fibroblasts treated with melanosome conditioned medium were then treated with trypsin and harvested by centrifugation at 10,000xg for 10 minutes. The collected cell pellets were dissolved by resuspending with 1M NaOH (Merck, Germany) and cultured at 80°C for 1 hour. Finally, the dissolved samples were centrifuged at 3000xg for 5 minutes, and the supernatant was collected for melanin and protein quantification.

[0206] 1.2.2 Co-culture model of reconstituted skin and fibroblasts with internalized melanosomes

[0207] 1x 10 5Human skin fibroblast HS-68 cells were seeded in a 12-well hanging top dish at a cell concentration of cells / well and cultured for 24 hours. Then, the culture medium was replaced with melanosome conditioned medium and cultured for another 24 hours. On the same day, reconstructed skin (Melanoderm, from Mattek, USA) with melanocytes was maintained with a long-term maintenance medium (LLMM, from Mattek, USA) and cultured overnight at 37°C and 5% CO2. The next day, the melanosome conditioned medium was discarded and rinsed with PBS, and then the co-culture of Melanoderm skin was added to a 12-well hanging top dish containing fibroblasts treated with melanosome conditioned medium. The co-culture system was maintained in LLMM culture medium for a total duration of 7 days and the culture medium was renewed every 2 days. Finally, the fibroblasts were treated with trypsin and harvested for melanin quantification. The cell viability of the treated skin and fibroblasts was determined by using the MTT test.

[0208] 1.2.3 Melanin and protein quantification

[0209] In brief, the collected supernatant containing melanin of 150 μ L was processed with 45 μ L of hydrogen peroxide (Merck, Germany), and cultivated at room temperature in the absence of light. Then, the mixture was centrifuged for 5 minutes at 3000xg and the fluorescence intensity of the collected supernatant was measured by using Cytation 5 multifunctional detectors (Biotek Instrument, USA) at an excitation wavelength of 470nm and an emission wavelength of 550nm. The amount of melanin in the sample was calculated by the standard curve (0-100 μ g / mL) of the newly prepared melanin standard (Sigma, USA). The amount of quantitative protein was tested using Bradford. Finally, the melanin content was standardized with protein content, and represented by the total melanin (μ g) (μ g / mg protein) of each mg protein.

[0210] 1.3 Results and Discussion

[0211] To investigate the melanosome uptake capacity of fibroblasts, fibroblasts were treated with conditioned medium containing melanosomes. Figure 1A As shown in , fibroblasts treated with conditioned medium containing melanosomes showed a significant increase in melanin content when compared to a control group pretreated with medium alone. In addition, fibroblasts did not show significant cytotoxic effects after treatment with conditioned medium containing melanosomes ( Figure 1B ). This observation demonstrates the ability of fibroblasts to internalize melanosomes and is consistent with recent findings by Ando et al., (2020) showing that melanosomes are taken up by dermal fibroblasts via actin-dependent endocytosis.

[0212] A biomimetic skin dermal melanosis model was further established by co-culturing fibroblasts that internalized melanin with a reconstructed skin model for a duration of 7 days. Similarly, in the established co-culture system, a significant increase in melanin content was found in fibroblasts treated with conditioned medium, and there was no significant cytotoxic effect on fibroblasts and skin tissue (Figure 2). This skin model makes it possible to evaluate the product efficacy of topical preparations on the effects of melanin content in both the epidermis and dermis.

[0213] 1.4 Conclusion

[0214] Fibroblasts exhibit the ability to internalize melanosomes and increase their intracellular melanin content. A biomimetic skin model of dermal melanosis was developed to investigate the efficacy of topical formulations on melanosome-internalizing fibroblasts.

[0215] Example 2 - 3-O-Ethylascorbic acid (3-EAA) as a potent molecule for reducing melanin content in melanosome-internalizing fibroblasts.

[0216] 2.1 Purpose

[0217] To investigate the effect of 3-O-ethylascorbic acid (3-EAA) on reducing melanin content in melanosome-internalizing fibroblasts.

[0218] 2.2 Methods

[0219] The establishment of fibroblasts with internalized melanosomes was performed as described above in 1.2.1 with slight modifications. After incubation of fibroblasts with melanosome-conditioned medium for 24 hours, the conditioned medium was removed and replaced with complete medium containing 3-EAA at final concentrations of 500 μM and 1000 μM. For the control group, complete medium without 3-EAA was used for comparison.

[0220] 2.3 Results and Discussion

[0221] like Figure 4 As shown, 3-EAA treatment at both tested concentrations (500 μM and 1000 μM) significantly reduced melanin content in melanosome-internalizing fibroblasts when compared to the control group.

[0222] 2.4 Conclusion

[0223] 3-O-Ethyl ascorbic acid efficiently reduces melanin content in melanosome-internalizing fibroblasts.

[0224] Example 3 - Synergistic effect of 3-O-ethyl-ascorbic acid and resveratrol in reducing melanogenic activity in the epidermis and melanin content of melanoblasts internalizing in a skin model of dermal melanosis.

[0225] 3.1 Purpose

[0226] To investigate the synergistic effects of 3-EAA and resveratrol in reducing melanogenic activity in the epidermis and melanin content in melanoblasts internalizing melanin in a skin model of dermal melanosis.

[0227] 3.2 Methods

[0228] 3.2.1 Preparation of composition

[0229] Premix B was added to the aqueous phase (A) and homogenized at 2500 rpm for 5 minutes. Premixes C and D were prepared separately until completely dissolved. Then, premix C was added to the main body (A+B) and stirred at 100-200 rpm for 5 minutes. Finally, premix D was added to the main body (A+B+C) and stirred at 100-200 rpm for 30 minutes.

[0230] Table 1 - Formulations 1-4

[0231]

[0232] *Active substance level of the compound

[0233] 3.2.2 Local treatment of co-culture system

[0234] The establishment of fibroblasts with internalized melanosomes was performed as described in 1.2.2 with slight modifications to the topical preparation. The co-culture system of fibroblasts with internalized melanosomes and reconstituted skin was maintained in LLMM for a duration of 7 days. During the maintenance of the co-culture system, the LLMM culture medium was renewed, followed by topical application of the preparation to the skin for two hours and washing off. Both culture medium renewal and topical treatment were performed every 2 days. After completing 7 days of culture, the fibroblasts were treated with trypsin and harvested for melanin quantification. In addition, the treated skin was fixed in 4% polyoxymethylene for measurement of melanin index and skin tone using a Mexameter (Courage-Khazaka electronic GmbH, Germany). Finally, the cell / tissue viability of the treated skin and fibroblasts was determined by using the MTT assay.

[0235] 3.3 Results and Discussion

[0236] As shown in Table 2, formulation 2 containing 2% 3-EAA showed a significant reduction in the melanin content in fibroblasts, but the melanin index in the reconstructed skin was not significantly reduced. In contrast, formulation 3 containing 0.1% resveratrol showed a significant reduction in the melanin index in the reconstructed skin, but the melanin content in the fibroblasts was not significantly reduced. Interestingly, formulation 4 containing a relatively low dose of active substance level (1% 3-EAA and 0.05% resveratrol) showed a significant reduction in the melanin index in the reconstructed skin and the melanin content in the dermal fibroblasts. Compared to formulations 2 and 3, the inhibitory activity of formulation 4 is better, although the level of active substance is low. In addition, the tissue survival rate of reconstructed skin and fibroblasts exceeds 80%, indicating that the inhibitory activity related to the reduction of melanin mediated by the formulation is not due to cytotoxicity (data not shown). Overall, these data show that EAA and resveratrol both work synergistically by inhibiting the melanin production activity in the epidermis and reducing the formation of melanin in dermal fibroblasts.

[0237] Table 2 - Effect of selected formulations on skin lightness and melanin index of reconstructed skin and melanin content in melanosome-internalizing fibroblasts of a biomimetic skin model of dermal melanosis.

[0238]

[0239] All obtained data were standardized with placebo treated group (formulation 1) and the product efficacy of the formulation was explained as a percentage change relative to the placebo treated group (formulation 1). All data are expressed as mean ± SEM of two independent experiments (n = 2). When compared with the placebo treated group (formulation 1) by one-way analysis of variance, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

[0240] 3.4 Conclusion

[0241] 3-O-ethylascorbic acid and resveratrol act synergistically to inhibit melanogenic activity in the epidermis and to reduce melanin content in melanoblasts internalizing melanin in a skin model of dermal melanosis.

[0242] Example 4 - 4-n-Butylresorcinol as a functional enhancer of the activity of 3-O-ethylascorbic acid in reducing the melanin content of melanoblasts internalizing melanin in a skin model of dermal melanosis.

[0243] 4.1 Purpose

[0244] To investigate the potentiating effect of 4-n-butylresorcinol on the activity of 3-EAA in reducing melanin content in melanocortin-internalizing fibroblasts in a skin model of dermal melanosis.

[0245] 4.2 Methods 4.2.1 Preparation of composition

[0246] Premix B was added to the aqueous phase (A) and homogenized at 2500 rpm for 5 minutes. Premixes C and D were prepared separately until completely dissolved. Premix C was then added to the main body (AB) and stirred at 100-200 rpm for 5 minutes. Finally, premix D was added to the main body (AB) and stirred at 100-200 rpm for 30 minutes.

[0247] Table 3 - Formulations 1 and 5-7

[0248]

[0249] *Active substance level of the compound

[0250] 4.2.2 Local treatment of co-culture system

[0251] This is performed according to the operations listed in 3.2.2.

[0252] 4.3 Results and Discussion

[0253] As shown in Table 4, formulation 5 containing 1% 3-EAA showed a trend of decreasing melanin content in fibroblasts, but there was no trend of decreasing melanin index in reconstructed skin. In contrast, formulation 6 containing 0.3% 4-n-butylresorcinol showed a significant decrease in melanin index in reconstructed skin, but there was no significant decrease in melanin content in fibroblasts. Interestingly, formulation 7 containing a combination of two active substances (1% 3-EAA and 0.3% 4-n-butylresorcinol) showed a significant decrease in melanin index and melanin content in fibroblasts in reconstructed skin. Compared to formulations 5 and 6, the inhibitory activity of formulation 7 is superior in reducing melanin content in fibroblasts. The tissue viability of reconstructed skin and the cell viability of fibroblasts exceeded 80%, indicating that the inhibitory activity mediated by the formulation related to melanin reduction is not due to cytotoxicity (data not shown). Overall, these data show that 4-n-butylresorcinol is a powerful enhancer for the activity of 3-EAA in reducing melanin formation in dermal fibroblasts.

[0254] Table 4 - Effect of selected formulations on skin lightness and melanin index of reconstructed skin and melanin content in melanosome-internalizing fibroblasts of a biomimetic skin model of dermal melanosis.

[0255]

[0256] All obtained data were standardized with placebo treated group (formulation 1) and the product efficacy of the formulation was explained as a percentage change relative to the placebo treated group (formulation 1). All data are expressed as mean ± SEM of two independent experiments (n = 2). When compared with the placebo treated group (formulation 1) by one-way analysis of variance, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

[0257] 4.4 Conclusion

[0258] 4-n-Butylresorcinol potentiates the activity of 3-O-ethylascorbic acid in reducing the melanin content of melanoblasts internalizing melanin in a skin model of dermal melanosis.

[0259] Example 5 - Clinical anti-post-inflammatory hyperpigmentation efficacy of a topical composition comprising 3-EAA and resveratrol.

[0260] 5.1 Purpose

[0261] To clinically investigate the anti-post-inflammatory hyperpigmentation activity of a topical formulation containing 3-EAA and resveratrol.

[0262] 5.2 Methods

[0263] 5.2.1 Preparation of composition (Formulation 8)

[0264] Prepare premix A and premix B separately until completely dissolved. Then, add premix B to premix A and stir at 2500 rpm for 10 minutes. Prepare premix C by first warming to 40°C, then add to the main body (A+B), and stir the mixture at 2500 rpm until completely dispersed. Finally, add the ingredients in phase D to the main body (A+B+C) in sequence and stir at 100-200 rpm for 10 minutes.

[0265] Table 5 - Formulation 8

[0266]

[0267] 5.2.2 Clinical trials

[0268] The efficacy of the product in improving acne scars (post-inflammatory hyperpigmentation) was studied on a group of 12 females with visible acne scars ranging in age from 22 to 23 years old. In this study, the recruited panelists were instructed to topically apply Formulation 8 twice daily for a duration of two months. Then, the patients were evaluated at baseline (T0), 2 weeks after product application (T 2w ), 4 weeks (T 4w ) and 8 weeks (T 8w), by taking photos of the panelists using the VISIA Skin Color Analysis System (Canfield, USA). Finally, the skin tone of each acne scar was marked as an area of ​​concern and further analyzed by measuring its individual topological angle (ITA°).

[0269] 5.3 Results and Discussion

[0270] As shown in Table 6, the results show that topical treatment with Formulation 8 reduced the skin tone of identified acne-related post-inflammatory hyperpigmentation. This reduction was significantly demonstrated in as little as 2 weeks after product application. The product's progressive anti-hyperpigmentation effect was also demonstrated throughout the 8-week period of product application.

[0271] Table 6 - Effect of Formulation 8 on improving skin tone of acne scars (post-inflammatory hyperpigmentation) as measured using the Individual Topological Angle (ITA°).

[0272]

[0273] All values ​​are presented as mean ± SEM of the group of test panelists (n=12). *p<0.05 **p<0.01 ***p<0.001 indicated significant difference compared to baseline by using Wilcoxon Matched Pairs Signed Rank Test.

[0274] 5.4 Conclusion

[0275] A topical formulation comprising 3-EAA and resveratrol demonstrated clinical efficacy in reducing acne scars (post-inflammatory hyperpigmentation).

[0276] Example 6 - Clinical anti-hyperpigmentation efficacy of a composition comprising 3-EAA and 4-n-butylresorcinol.

[0277] 6.2 Purpose

[0278] To clinically validate the anti-hyperpigmentation activity of a topical formulation containing 3-EAA and 4-n-butylresorcinol.

[0279] 6.2 Methods

[0280] 6.2.1 Preparation of composition (Formulation 9)

[0281] Premix A (aqueous phase) and premix B (lipid phase) were prepared separately until completely dissolved. Premix B was then added to premix A and homogenized at 2500 rpm for 10 minutes. Premix C and D were also prepared separately. Premix C and D were then added to the main body (A+B) in sequence and homogenized at 2500 rpm for 5 minutes, followed by stirring at 100-200 rpm for 10 minutes.

[0282] Table 7 - Formulation 9

[0283]

[0284] *Active substance level of the compound

[0285] 6.2.2 Clinical trials

[0286] A group of 11 women aged between 22 and 35 years with noticeable skin pigmentation spots were evaluated for the efficacy of the product in improving hyperpigmentation. In this study, the recruited panelists were instructed to topically apply Formulation 9 twice daily for a duration of two months. 4w ) and 8 weeks (T 8w ), by taking photos of the panelists using the VISIA skin color analysis system (Canfield, USA). Finally, the skin tone of each pigmented spot was marked as an area of ​​interest and further analyzed by determining its individual topological angle (ITA).

[0287] 6.3 Results and Discussion

[0288] The results shown in Table 8 show that topical treatment with Formulation 9 reduced the skin tone of the identified pigmented spots and the reduction was significantly demonstrated after 8 weeks of product application.

[0289] Table 8 - Effect of Formulation 9 on improving skin tone of pigmented spots as measured using the Individual Topological Angle (ITA°).

[0290]

[0291] All values ​​are presented as mean ± SEM of the group of test panelists (n=11).**p<0.01 indicates significant difference compared to baseline by using Wilcoxon paired signed rank test.

[0292] 6.4 Conclusion

[0293] Topical Formulation 9 comprising 3-EAA and 4-n-butylresorcinol exhibited clinical efficacy in reducing pigmented spots (hyperpigmentation).

[0294] Those skilled in the art will appreciate that many changes and / or modifications may be made to the embodiments described above without departing from the broad overall scope of the present disclosure. Therefore, the present embodiments should be considered in all aspects as illustrative and not restrictive.

[0295] References

[0296] 1. Vashi, NA & Kundu, RV Facial hyperpigmentation: Causes and treatment. Br. J. Dermatol. 169, 41–56 (2013).

[0297] 2. Nieuweboer-Krobotova, L. Hyperpigmentation: Types, diagnostics and targeted treatment options. J. Eur. Acad. Dermatology Venereol. 27, 2–4 (2013).

[0298] 3. Watanabe, S. Facial Dermal Melanocytosis. Austin J Dermatolog 1, 1006 (2014).

[0299] 4. Kolbe, L. et al. 4-N-Butylresorcinol, a Highly Effective TyrosinaseInhibitor for the Topical Treatment of Hyperpigmentation. J. Eur. Acad. Dermatology Venereol. 27, 19–23 (2013).

[0300] 5. Yoon, HS, Hyun, CG, Lee, NH, Park, SS & Shin, DB Comparative depigmentation effects of resveratrol and its two methyl analogues in α-melanocyte stimulating hormone-triggered B16 / F10murine melanomacells. Prev. Nutr. Food Sci. 21, 155–159 (2016).

[0301] 6. Deguchi, T. et al. Anti-tyrosinase and Anti-oxidative Activities by Asana: the Heartwood of Pterocarpus marsupium. Nat. Prod. Commun. 14, (2019).

[0302] 7. Chen, S.J. et al. The anti-melanogenic effects of 3-O-ethyl ascorbic acid via Nrf2-mediated α-MSH inhibition in UVA-irradiated keratinocytes and autophagy induction in melanocytes. Free Radical Biology and Medicine 173, 151–169 (2021).

[0303] 8. Ando, H. et al. Dermal fibroblasts internalize phosphatidylserine-exposed secretory melanosome clusters and apoptotic melanocytes. Int. J. Mol. Sci. 21, 1–14 (2020).

Claims

1. A composition for preventing and / or treating skin pigmentation disorders, comprising: a) ascorbic acid or its derivatives and / or salts; and b) an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or a salt thereof; Its amount effectively degrades melanin in dermal fibroblasts.

2. The composition of claim 1 , wherein component a) is selected from the group consisting of 3-O-ethyl ascorbic acid, L-ascorbic acid, 2-O-ethyl ascorbic acid, ascorbyl glucoside, ascorbyl tetraisopalmitate, magnesium ascorbyl phosphate, sodium ascorbyl phosphate, capryl-2-glyceryl ascorbate, 3-glyceryl ascorbate, diglyceryl ascorbate, hexyl 3-glyceryl ascorbate, myristyl 3-glyceryl ascorbate and laurylglyceryl ascorbate.

3. The composition of claim 2, wherein component a) is selected from the group consisting of 3-O-ethyl ascorbic acid, L-ascorbic acid, 2-O-ethyl ascorbic acid, ascorbyl tetraisopalmitate, sodium ascorbyl phosphate, capryl-2-glyceryl ascorbate, 3-glyceryl ascorbate, diglyceryl ascorbate, hexyl 3-glyceryl ascorbate, myristyl 3-glyceryl ascorbate and laurylglyceryl ascorbate.

4. The composition according to claim 3, wherein component a) is 3-O-ethyl ascorbic acid.

5. The composition according to any one of claims 1 to 4, wherein component b) is resveratrol.

6. The composition according to any one of claims 1 to 4, wherein component b) is pterostilbene.

7. The composition according to any one of claims 1 to 4, wherein component b) is sclareolide.

8. The composition according to any one of claims 1 to 4, wherein component b) is 4-n-butylresorcinol.

9. The composition according to any one of claims 1 to 8, wherein the amount of component a) ranges from 0.01% to 50% by weight of the composition, optionally from 0.5% to 5% by weight of the composition, optionally from 0.5% to 1.5% by weight of the composition.

10. The composition of any one of claims 1 to 9, wherein the amount of component b) ranges from 0.01% to 5% by weight of the composition, optionally from 0.1% to 0.5% by weight of the composition, or optionally from 0.2% to 0.35% by weight of the composition.

11. The composition of any one of claims 1 to 10, wherein the weight ratio of component a) to component b) ranges from 2:1 to 200:1, optionally 2:1 to 50:1, or optionally 3:1 to 5:

1.

12. The composition according to any one of claims 1 to 11, wherein the composition does not contain any anti-melanogenesis agent other than a) and b).

13. The composition according to any one of claims 1 to 12, wherein the composition comprises 0.5 to 1.5 wt% of 3-O-ethyl ascorbic acid as component a) and comprises 0.2 to 0.35 wt% of resveratrol as component b), and wherein the weight ratio of component a) to component b) ranges from 3:1 to 5:

1.

14. The composition according to any one of claims 1 to 13, wherein the composition is for topical administration.

15. The composition according to any one of claims 1 to 14, wherein the composition comprises one or more of the following: an anti-inflammatory agent, a sunscreen, an antioxidant, an anti-aging peptide, a desquamating agent, and an exfoliating agent.

16. The composition of any one of claims 1 to 15, wherein the composition comprises one or more of the following: water, alkyl benzoate, PPG-3 myristyl ether, hydroxyacrylate / sodium acryloyldimethyl taurate copolymer, ethylhexylglycerin, phenoxyethanol, methylpropanediol, betaine, sclerotium gum, sodium polyacryloyldimethyl taurate, disodium EDTA, sodium hyaluronate, di-PEG-18 methyl ether dimethyl silane, PEG-12 dimethicone, glycerin, chlorphenesin, sodium metabisulfite, lactic acid, dicaprylyl carbonate and polyacrylate crosspolymer-6.

17. The composition according to any one of claims 1 to 16, wherein the composition is in the form of a cream, a lotion, a paste, a wax, a liquid, a semisolid composition or a sprayable composition.

18. A method for preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual, or a method for degrading melanin in dermal fibroblasts of an individual, comprising administering to the individual an effective amount of a composition according to any one of claims 1 to 17.

19. Use of a composition as claimed in any one of claims 1 to 17 for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual or for degrading melanin in dermal fibroblasts of an individual.

20. A composition according to any one of claims 1 to 17 for use in preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders.

21. The composition according to any one of claims 1 to 17, for use in degrading melanin in dermal fibroblasts.

22. The method, use or composition for use according to any one of claims 18 to 20, wherein the skin pigmentation disorder is selected from the group consisting of skin hyperpigmentation, melasma, solar lentigo, nevus of Ota, nevus of Ito and Mongolian spots.

23. The method, use or composition for use according to any one of claims 18 to 22, wherein the composition is topically applied to the skin of the individual.

24. A method for preventing, treating and / or alleviating symptoms associated with a skin pigmentation disorder in an individual, or degrading melanin in dermal fibroblasts of an individual, comprising administering an effective amount of b) ascorbic acid or its derivatives and / or salts, Its amount effectively degrades melanin in dermal fibroblasts.

25. The method according to claim 24, wherein the ascorbic acid or its derivatives and / or salts are used in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

26. Ascorbic acid or a derivative and / or salt thereof for use in preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, wherein the ascorbic acid or a derivative and / or salt thereof is administered in an amount effective to degrade melanin in dermal fibroblasts.

27. Ascorbic acid or its derivatives and / or salts for use according to claim 26, wherein the ascorbic acid or its derivatives and / or salts are used in combination with an anti-melanin production agent, the anti-melanin production agent is selected from 4-n-butylresorcinol, resveratrol, pterostilbene and clareolide, or salts thereof.

28. Ascorbic acid or a derivative and / or salt thereof for use in degrading melanin in dermal fibroblasts.

29. Ascorbic acid or its derivatives and / or salts for use according to claim 28, wherein the ascorbic acid or its derivatives and / or salts are used in combination with an anti-melanin production agent, the anti-melanin production agent being selected from 4-n-butylresorcinol, resveratrol, pterostilbene and clareolide, or salts thereof.

30. An anti-melaninogenic agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof, for use in preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, wherein the anti-melaninogenic agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

31. An anti-melaninogenic agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof, for use in degrading melanin in dermal fibroblasts, wherein the anti-melaninogenic agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

32. Use of ascorbic acid or its derivatives and / or salts for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, or for degrading melanin in dermal fibroblasts, wherein the ascorbic acid or its derivatives and / or salts are administered in an amount effective to degrade melanin in dermal fibroblasts.

33. The use according to claim 32, wherein the ascorbic acid or its derivatives and / or salts are administered in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

34. Use of an anti-melanin generating agent for the preparation of a medicament for preventing, treating and / or alleviating symptoms associated with skin pigmentation disorders, or for degrading melanin in dermal fibroblasts, wherein the anti-melanin generating agent is selected from 4-n-butylresorcinol, resveratrol, pterostilbene and clarylidene, or salts thereof, wherein the anti-melanin generating agent is administered in combination with ascorbic acid or a derivative and / or salt thereof.

35. The method, use or composition for use according to any one of claims 24-27, 30 or 32-34, wherein the skin pigmentation disorder is selected from hyperpigmentation of the skin, melasma, solar lentigo, nevus of Ota, nevus of Ito and Mongolian spots.

36. The method, use or composition for use according to any one of claims 24 to 35, wherein the composition is topically applied to the skin of the subject.

37. A non-therapeutic method of lightening the skin of an individual comprising administering to the individual an effective amount of a composition according to any one of claims 1 to 17.

38. The non-therapeutic method of claim 35, wherein the composition is topically applied to the skin of the individual.

39. A non-therapeutic method of lightening the skin of an individual comprising administering to the individual an amount of ascorbic acid, or a derivative and / or salt thereof, effective to degrade melanin in dermal fibroblasts.

40. The non-therapeutic method of claim 39, wherein the ascorbic acid or a derivative and / or salt thereof is topically applied to the skin of the individual.

41. The non-therapeutic method according to claim 39 or 40, wherein the ascorbic acid or its derivatives and / or salts are administered in combination with an anti-melanin production agent selected from 4-n-butylresorcinol, resveratrol, pterostilbene and sclareolide, or salts thereof.

42. The non-therapeutic method of claim 41, wherein the anti-melanogenesis agent is topically administered to the skin of the individual.

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