Oleanolic acid compositions for sebum control

Abstract

The invention provides a composition. The composition comprises oleanolic acid and salts, prodrugs and solvates of oleanolic acid, and one or more compounds selected from the group consisting of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, protocatechuic acid, and tyramine, and salts, prodrugs, and solvates thereof. Such compositions are useful for reducing lipid levels in sebaceous gland cells, and the compositions may find use in the treatment of acne vulgaris and rosacea.

Description

Technical Field

[0001] The present invention relates to a composition comprising oleanolic acid (OA) and one or more further specified compounds, and the use of the composition in a method for reducing lipid production in sebaceous gland cells. Also provided is the use of the composition in a cosmetic method for reducing lipid production in the skin of an individual, and the use of the composition in a method for treating a skin disease or disorder associated with increased lipid production in sebaceous gland cells. Background Art

[0002] Sebum is an oily substance secreted by the body's sebaceous glands. Sebum is produced by sebocytes, which are highly specialized epithelial cells commonly found in the skin that are associated with hair follicles, although sebaceous glands that are not associated with hair follicles also exist.

[0003] Adult sebum typically comprises triglycerides (~41%), wax monoesters (~25%), free fatty acids (~16%), and squalene (~12%) (Cheng et al., 2004). Other components, such as keratin and cellular material, may also be present.

[0004] Sebum forms an integral part of the epidermal barrier and the skin immune system. Sebum is a natural moisturizer of the epidermis that helps maintain the integrity of the epidermis. Sebum is important in maintaining the pH of the skin surface, which can play a role in protecting the skin from exogenous (pathogenic) microorganisms and promoting the growth of endogenous (resident) microbial communities. It is also believed that sebaceous gland secretions play an important role in thermoregulation together with apocrine (sweat) glands.

[0005] The formation of sebocytes is controlled by various molecular pathways (eg, Blimp1, Wnt, C-myc, Hedgehog), and sebum synthesis is strongly regulated by hormones, particularly by androgens such as testosterone.

[0006] Sebum is produced during holocytosis, in which sebocytes rupture and disintegrate as they release sebum along with cellular remnants. During terminal differentiation of sebocytes, metabolic activity is focused on the biosynthesis of lipids (lipogenesis), and in particular the neosynthesis of fatty acids and squalene.

[0007] Levels of sebum production vary from person to person and are affected by gender, age, physical activity, stress, some medications, and illness. Oily skin is common during adolescence due to hormonal changes that occur throughout puberty.

[0008] Excessive sebum production is associated with cosmetic problems such as oily or shiny skin and poor makeup retention as well as medical problems. Excessive sebum production is seen in acne vulgaris, one of the most common skin diseases. Excessive sebum secretion is a scalp problem caused by excessive sebum production. Immediate symptoms of excessive sebum secretion include scalp itching and pain, while later symptoms are hair loss. Individuals with hyperseborrheic skin typically exhibit greater than 200 μg cm measured on the forehead. -2 Sebum levels in the skin (as discussed, for example, in WO 2020 / 263188). Dysregulation of sebaceous gland cell differentiation is also a feature of some rare benign and malignant tumors.

[0009] Cosmetic treatments that target excess lipid production generally do not address the underlying cause. Rather, cosmetic treatments typically provide relief from immediate symptoms such as greasy, enlarged pores, acne-prone skin, and irregular skin texture. For example, a common method of treating oily or shiny skin is to use powders that provide an immediate concealing effect by absorbing excess sebum on the skin's surface. Alternatively, astringents and cleansers may be used.

[0010] Known methods for reducing lipids on the skin surface are limited and rarely produce sustainable visible results over extended periods of time. Long-term use of astringents and cleansers may exacerbate the condition.

[0011] Therefore, there is a need to develop compositions and methods for reducing lipid production in sebaceous gland cells and in the skin of an individual. Summary of the invention

[0012] Most generally, a first aspect of the invention relates to a composition comprising oleanolic acid (OA) and one or more compounds selected from the group consisting of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide (miquelianin, miguining), p-coumaric acid, pipecolic acid, protocatechuic acid and tyramine, and salts, prodrugs and solvates thereof. The inventors have found that such a composition provides an excellent lipid reduction effect in sebaceous gland cells.

[0013] Preferred features of the composition are described below.

[0014] The composition may comprise two or more compounds selected from said group, such as three or more compounds, for example wherein at least one compound is ellagic acid.

[0015] The molar ratio of oleanolic acid to the compound selected from the group may be 1:x, wherein x is in the range of 0.1 to 20.

[0016] The composition may include oleanolic acid and its salts and solvates; and a plant extract including one or more compounds. The plant extract may be a wild strawberry (Fragaria vesca) extract, a raspberry (Rubus idaeus) extract or a pineapple strawberry (Fragaria×ananassa) extract, such as a wild strawberry extract or a raspberry extract.

[0017] The inventors have found that the composition of the first aspect reduces lipid production in sebocytes.

[0018] Therefore, in a second aspect of the invention, there is provided a method for reducing lipid production in sebaceous gland cells, the method comprising contacting sebaceous gland cells with the composition of the first aspect. The method may be in vivo or ex vivo, for example in vitro.

[0019] The inventors have found that the composition of the first aspect can reduce or improve cosmetic problems associated with excessive lipid production in the skin, such as oily or shiny skin, oily hair, enlarged skin pores, undesirable body odor, and reduced retention of cosmetics on the skin.

[0020] Thus, in a third aspect of the invention there is provided a cosmetic method for reducing lipid production in the skin of an individual, the method comprising contacting the skin with a composition of the first aspect.

[0021] The inventors have found that the composition of the first aspect can be used to treat or prevent medical problems associated with overproduction of lipids in the skin, such as acne vulgaris and rosacea.

[0022] Thus, in a fourth aspect of the invention there is provided a composition of the first aspect for use in a method of treatment.

[0023] In a further related aspect of the invention, there is provided a composition of the first aspect for use in a method of treating a skin disease or disorder associated with overproduction of lipids in the skin, such as acne vulgaris and rosacea.

[0024] These and other aspects and embodiments of the invention are described in further detail below. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] The invention is described with reference to the drawings listed below.

[0026] Figure 1. In vitro lipid reduction percentage for adenine, betaine, choline, ellagic acid, and quercetin-3-O-glucuronide. Oleanolic acid (OA) was used at 1.25 µM, while the other compounds were used at 10 µM. The experiment was repeated to ensure reproducibility: (top) first run; (bottom) second run.

[0027] Figure 2 . In vitro lipid reduction percentage for p-coumaric acid, pipecolic acid, protocatechuic acid, and tyramine. Oleanolic acid (OA) was used at 1.25 µM, while the other compounds were used at 10 µM. The experiment was repeated to ensure reproducibility: (top) first run; (bottom) second run.

[0028] Figure 3 Mean fluorescence intensity of sebaceous gland cells treated with AdipoRed™ dye obtained by flow cytometry is shown. Cells were incubated with the test composition for 3 days. From left to right are DMSO (negative control), EGCG (positive control), oleanolic acid (OA), raspberry extract, two different concentrations of wild strawberry extract, a combination of oleanolic acid and raspberry extract according to an embodiment of the present invention, and two combinations of oleanolic acid and wild strawberry extract at two different wild strawberry concentrations according to an embodiment of the present invention.

[0029] Figure 4 Shown is the mean fluorescence intensity of sebaceous gland cells treated with AdipoRed™ dye obtained by flow cytometry. The cells were incubated with the test composition for 3 days. From left to right are DMSO (negative control), EGCG (positive control), salicylic acid (positive control), oleanolic acid, raspberry extract, and a combination of oleanolic acid and raspberry extract according to an embodiment of the present invention.

[0030] Figure 5 The mean fluorescence intensity of sebaceous gland cells treated with AdipoRed™ dye obtained by flow cytometry is shown. The cells were incubated with the test composition for 3 days. From left to right are DMSO (negative control), EGCG (positive control), oleanolic acid, wild strawberry extract, and a combination of oleanolic acid and wild strawberry extract according to an embodiment of the present invention.

[0031] In the figures, for comparisons in one-way analysis of variance (one-way ANOVA), the following notations are used:

[0032] * P value: <0.05 (statistically significant compared to DMSO)

[0033] ** P value: <0.01 (statistically significant compared to DMSO)

[0034] *** P value: 0.001 (statistically significant compared to DMSO)

[0035] **** P value: <0.0001 (statistically significant compared to DMSO)

[0036] # P value: <0.05 (statistically significant compared to oleanolic acid)

[0037] ## P value: <0.01 (statistically significant compared to oleanolic acid)

[0038] ### P value: <0.001 (statistically significant compared to oleanolic acid)

[0039] #### P value: <0.001 (statistically significant compared to oleanolic acid) DETAILED DESCRIPTION

[0040] The present invention relates to a composition comprising oleanolic acid (OA) and one or more compounds selected from the group consisting of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, protocatechuic acid and tyramine, and salts, prodrugs and solvates thereof; and the use of the composition in a method for reducing lipid production in sebaceous gland cells. Also provided is the use of the composition in a cosmetic method for reducing lipid production in the skin of an individual, and the use of the composition in a method for treating a skin disease or disorder associated with increased lipid production in sebaceous gland cells.

[0041] The following preferences are applicable to all aspects of the invention as described above. These preferences may be combined in any combination.

[0042] Oleanolic acid

[0043] The composition of the present invention contains oleanolic acid (OA) and one or more compounds selected from the group consisting of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, protocatechuic acid and tyramine, and salts and solvates thereof.

[0044] Oleanolic acid has the CAS Registry Number [508-02-1].

[0045] Reference to oleanolic acid also includes reference to its salts and solvates.

[0046] The compositions of the present invention may comprise an effective amount, such as a therapeutically or cosmetically effective amount, of oleanolic acid.

[0047] In some embodiments, for example, based on the total weight of the composition, the composition of the present invention comprises oleanolic acid in an amount of 0.0005% to 10% by weight. In a preferred embodiment, the composition of the present invention comprises oleanolic acid in an amount of 0.1% to 5% by weight, for example, 0.1% to 3% by weight, and for example, 0.2% to 3% by weight, and more preferably 0.2% to 2% by weight.

[0048] Oleanolic acid may be present in an amount of up to 2%, 3%, 5%, or 10% by weight.

[0049] Oleanolic acid may be present in an amount of at least 0.0005%, 0.001%, 0.005%, 0.01%, 0.05%, 0.1%, 0.5%, or 1.0% by weight.

[0050] In one embodiment, the composition comprises a plant extract comprising one or more compounds. Here, oleanolic acid (OA) may not be present in the plant extract used in combination with it, or may not otherwise be derived from (derived from) the plant extract. For example, in the case where the plant extract is a wild strawberry extract or a raspberry extract, these extracts are substantially free of oleanolic acid (OA). For example, oleanolic acid may be present in the plant extract in an amount of less than 1 wt %, such as less than 0.1 wt %, such as less than 0.01 wt %.

[0051] In one embodiment, triterpenoids may not be present in the plant extract used in combination therewith, or may not otherwise be derived from the plant extract. For example, where the plant extract is a wild strawberry extract or a raspberry extract, these extracts are substantially free of triterpenoids (OA). For example, the total amount of triterpenoids may be less than 1 wt %, such as less than 0.1 wt %, such as less than 0.01 wt % in the plant extract.

[0052] Compound

[0053] The composition of the present invention comprises one or more compounds selected from the group consisting of adenine, betaine, choline, ellagic acid, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, protocatechuic acid and tyramine.

[0054] Reference to each compound also includes reference to its salts and solvates.

[0055] The one or more compounds may be selected from the group consisting of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide and tyramine, and salts and solvates thereof.

[0056] The one or more compounds may be selected from the group consisting of ellagic acid, betaine, choline, quercetin-3-O-glucuronide and tyramine, and salts and solvates thereof.

[0057] The composition may comprise two or more, such as three or more, such as four or more compounds selected from any one of the above-mentioned groups.

[0058] At least one compound in the composition may be ellagic acid.

[0059] The composition may comprise a combination of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid and tyramine. Such a combination may be provided by raspberry plant extract. Optionally, the composition may additionally comprise protocatechuic acid.

[0060] The composition may comprise a combination of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid and pipecolic acid. Such a combination may be provided by a wild strawberry plant extract. Optionally, the composition may additionally comprise protocatechuic acid.

[0061] Ellagic acid has the CAS Registry Number [476-66-4].

[0062] Adenine has the CAS Registry Number [73-24-5].

[0063] Betaine has the CAS registry number [107-43-7].

[0064] Choline has the CAS Registry Number [62-49-7]. Choline is available as choline chloride.

[0065] Quercetin-3-O-glucuronide has the CAS registry number [22688-79-5].

[0066] p-Coumaric acid has the CAS Registry Number [7400-08-0].

[0067] Pipecoic acid has the CAS Registry Number [535-75-1].

[0068] Protocatechuic acid has the CAS registry number [99-50-3].

[0069] Tyramine has the CAS Registry Number [051-67-2].

[0070] In some embodiments, for example, based on the total weight of the composition, the composition of the present invention comprises a compound selected from the group in an amount of 0.0005% to 10% by weight. In a preferred embodiment, the composition of the present invention comprises a compound selected from the group in an amount of 0.1% to 5% by weight, for example, 0.1% to 3% by weight, and for example, 0.2% to 3% by weight, and more preferably 0.2% to 2% by weight.

[0071] The composition may include a compound selected from the group which may be present in an amount of up to 2%, 3%, 5%, or 10% by weight.

[0072] The composition may include a compound selected from the group that may be present in an amount of at least 0.0005 wt%, 0.001 wt%, 0.005 wt%, 0.01 wt%, 0.05 wt%, 0.1 wt%, 0.5 wt%, or 1.0 wt%.

[0073] In one embodiment, each compound selected from said group is present in the amount specified above.

[0074] In the composition of the present invention, the molar ratio of oleanolic acid to the compound selected from the group is 1:x, wherein x is in the range of 0.1 to 20. The molar ratio of oleanolic acid to the compound selected from the group may be 1:x, wherein x is in the range of 1 to 10, such as 2 to 10, such as 5 to 10, such as about 8.

[0075] In one embodiment, each compound selected from the group may be present in the above specified amounts relative to oleanolic acid.

[0076] Salts and solvates

[0077] Oleanolic acid (OA) may be provided in the form of a free base. Alternatively, oleanolic acid (OA) may be provided in the form of a salt (eg, a pharmaceutically or cosmetically acceptable salt).

[0078] Examples of pharmaceutically acceptable salts are discussed in Berge et al., 1977, “Pharmaceutically Acceptable Salts,” J. Pharm. Sci., Vol. 66, pp. 1-19.

[0079] In some embodiments, the compound (eg, oleanolic acid) is provided in anionic form (eg, deprotonated form) along with a suitable counter cation.

[0080] Suitable counter ions include both organic and inorganic cations. Suitable counter ions include both inorganic and organic cations. Examples of suitable inorganic cations include alkali metal ions such as Na+ and K + , alkaline earth metal cations such as Ca 2+ and Mg 2+ , and other cations such as Al 3+ Examples of suitable organic cations include ammonium ions (ie, NH4 + ) and substituted ammonium ions (e.g., NH3R + NH2R2 + 、NHR3 + NR4 + ). Examples of substituted ammonium ions include those derived from ethylamine, diethylamine, dicyclohexylamine, triethylamine, butylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, benzylamine, phenylbenzylamine, choline, meglumine, and tromethamine, and amino acids such as lysine and arginine. Common examples of quaternary ammonium ions are N(CH3)4 + .

[0081] In some embodiments, the compound (eg, choline) is provided in a cationic form along with a suitable counter cation.

[0082] Unless otherwise indicated, a reference to a particular compound also includes its salt forms.

[0083] The compound in the composition may be provided in the form of a solvate (a complex of a solute (eg, a compound, a salt of a compound) and a solvent). Examples of solvates include hydrates, such as monohydrates, dihydrates, and trihydrates.

[0084] The compound in the composition may be provided in a desolvated form (eg, in a dehydrated form).

[0085] Plant Extracts

[0086] The composition of the present invention comprises one or more compounds selected from the group consisting of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, protocatechuic acid and tyramine.

[0087] The one or more compounds may be derived or derivable from a plant extract.

[0088] In one embodiment, the composition of the present invention comprises:

[0089] Oleanolic acid and its salts and solvates; and

[0090] A plant extract comprising one or more of said compounds.

[0091] Thus, the compositions of the present invention may comprise plant extracts.

[0092] The plant extract may be selected from plant extracts belonging to the genera Fragaria and Rubus.

[0093] Suitable plant extracts are commercially available and may be purchased from, for example, BOC Sciences and BOCSCI (New York, USA).

[0094] Suitable species of plants belonging to the genus Rubus include Rubus idaeus (red raspberry). Raspberries are widely distributed throughout Europe and are native to, for example, the United Kingdom.

[0095] Suitable species of plants belonging to the genus Fragaria include Fragaria fusca (wild strawberry) and Fragaria oleracea (garden strawberry). Fragaria fusca is widely distributed throughout Europe and originates in, for example, the United Kingdom. Fragaria oleracea is widely cultivated in Europe, including the United Kingdom.

[0096] The plant extract may be derived from any suitable part of a plant. Suitable parts of a plant include roots, stems, leaves, flowers, fruits and seeds.

[0097] Examples of suitable fruit extracts include wild strawberry, raspberry and pineapple strawberry fruit extracts.

[0098] Typically, the plant extract is an aqueous extract of the plant or plant part. The aqueous extract can be used as a dried aqueous extract of the plant or plant part.

[0099] In a preferred embodiment, the plant extract is selected from the group consisting of strawberry fruit extract and raspberry fruit extract.

[0100] In a preferred embodiment, the plant extract is raspberry extract.

[0101] In a further embodiment, the plant extract is raspberry fruit extract.

[0102] In a preferred embodiment, the plant extract is wild strawberry extract.

[0103] In a further embodiment, the plant extract is Fragaria wild fruit extract.

[0104] In a preferred embodiment, the plant extract is pineapple and strawberry extract.

[0105] In a further embodiment, the plant extract is pineapple strawberry fruit extract.

[0106] As noted above, oleanolic acid may be substantially absent, eg, may be absent from, or not otherwise derived from, the plant extract with which it is used in combination.

[0107] The compositions of the present invention can include an effective amount (eg, a therapeutically or cosmetically effective amount) of a plant extract described herein.

[0108] The plant extract can be provided in an amount sufficient to produce one or more compounds in an amount as described above (eg, an effective amount, such as a therapeutically or cosmetically effective amount).

[0109] The raspberry extract may contain one or more (eg all) of the compounds given in Table 1 (see Examples section) in an amount within the given range. The extract here may be a dry extract.

[0110] The raspberry extract may be obtained or obtainable from raspberry fruit. The extract may be obtained by extraction with water, concentration and spray drying.

[0111] The wild strawberry extract may contain one or more (eg all) of the following compounds given in Table 2 (see Examples section) in amounts within the given ranges. The extract here may be a dry extract.

[0112] The wild strawberry extract may be obtained or obtainable from strawberry fruit. The extract may be obtained by extraction with water, concentration and spray drying.

[0113] In some embodiments, for example, based on the total weight of the composition, the composition of the present invention comprises a plant extract in an amount of 0.005% to 50% by weight. In a preferred embodiment, the composition of the present invention comprises a plant extract in an amount of 0.005% to 30% by weight, for example, 0.05% to 10% by weight, more preferably 0.1% to 3% by weight.

[0114] The plant extract may be present in an amount of up to 2%, 3%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 45%, or 50% by weight.

[0115] The plant extract may be present in an amount of at least 0.005%, 0.01%, 0.05%, 0.1%, 0.5%, or 1% by weight.

[0116] Composition

[0117] The compositions of the invention may be formulated for cosmetic or therapeutic use.Thus, the compositions of the invention may additionally comprise one or more pharmaceutical or cosmetic ingredients.

[0118] Pharmaceutically and cosmetically acceptable ingredients are those that are suitable, within the scope of sound judgment, for use in contact with the tissues of the subject under consideration (e.g., humans) without excessive toxicity, irritation, allergic response, or other problems or complications, commensurate with a reasonable benefit / risk ratio. Each ingredient must also be compatible with the other ingredients of the composition.

[0119] In one embodiment, the composition comprises one or more ingredients selected from the group consisting of solvents, oils, surfactants, thickeners, wetting agents, and preservatives.

[0120] Examples of suitable solvents include water; monohydric alcohols such as ethanol, isopropanol, benzyl alcohol and phenylethyl alcohol; polyhydric alcohols such as ethylene glycol, propylene glycol, butane-1,3-diol, mannitol, sorbitol, glycerol and erythritol; and glycol ethers such as ethylene glycol monomethyl ether and diethylene glycol monomethyl ether.

[0121] Examples of suitable oils include mineral oils, vegetable oils and waxes.

[0122] Examples of suitable vegetable oils include algae oil, annatto oil, argan oil, almond oil, apricot kernel oil, avocado oil, babassu oil, brazil nut butter, butter, cashew butter, castor oil, camellia oil, cherry seed oil, cocoa butter, coconut oil, corn oil, cottonseed oil, fish oil, grapeseed oil, gardenia oil, ghee, hazelnut oil, jatropha oil, jojoba oil, kokum oil, linseed oil, macadamia oil, corn oil, mango seed oil, mango butter, mineral oil, mink oil, olive oil, palm oil, palm kernel oil, peach kernel 5 oil, peanut butter, peanut oil, plum kernel oil, oil), pomegranate oil, rapeseed oil, rice bran oil, rosehip oil, saloil, sesame oil, shea butter, soybean oil, squalene, sunflower seed oil, tea seed oil, walnut oil.

[0123] Examples of suitable waxes include bayberry wax, beeswax, carnauba wax (palm wax), candelilla wax, ceresin, jojoba fat, lanolin wax, montan wax, ozokerite, polyglyceryl-3-beeswax, polyglyceryl-6-pentastearate, Japan wax, microcrystalline wax, paraffin, isoparaffin, petrolatum, squalene, oligomeric olefins, synthetic candelilla wax, synthetic carnauba wax, and synthetic beeswax.

[0124] Surfactants (surface active agents) can act as dispersants or wetting agents. Examples of suitable surfactants include anionic surfactants, cationic surfactants, nonionic surfactants, and amphoteric (zwitterionic) surfactants.

[0125] Examples of suitable anionic surfactants include ammonium lauryl sulfate, ammonium laureth sulfate, triethylamine lauryl sulfate, triethylamine laureth sulfate, triethanolamine lauryl sulfate, triethanolamine laureth sulfate, monoethanolamine lauryl sulfate, monoethanolamine laureth sulfate, diethanolamine lauryl sulfate, diethanolamine laureth sulfate, sodium lauric acid monoglyceride sulfate, sodium lauryl sulfate, sodium laureth sulfate, potassium lauryl sulfate, potassium laureth sulfate, sodium lauryl sarcosinate, sodium lauroyl sarcosinate, lauryl sarcosine, cocoyl sarcosine, ammonium cocoyl sulfate, ammonium lauroyl sulfate, sodium cocoyl sulfate, sodium lauroyl sulfate, potassium cocoyl sulfate, potassium lauryl sulfate, triethanolamine lauryl sulfate, triethanolamine lauryl sulfate, monoethanolamine cocoyl sulfate, monoethanolamine lauryl sulfate, sodium tridecylbenzene sulfonate, and sodium dodecylbenzene sulfonate.

[0126] Examples of suitable cationic surfactants include behentrimonium chloride, cocotrimonium chloride, cetethyldimonium bromide, dibehenyldimonium chloride, di(hydrogenated tallow)benzylmethyl ammonium chloride, disoybean dimethyl ammonium chloride, ditallow dimethyl ammonium chloride, hydroxycetyl hydroxyethyl dimethyl ammonium chloride, hydroxyethyl behenamidopropyl dimethyl ammonium chloride, hydroxyethyl cetyl dimethyl ammonium chloride, hydroxyethyl tallow dimethyl ammonium chloride, myristalkonium chloride, PEG-2 oleamonium chloride, PEG-5 stearyl methyl ammonium chloride, PEG-15 cocoyl quaternium salt 4, PEG-2 stearyl dimethyl benzyl ammonium 4, lauryl trimethyl ammonium chloride; quaternium-16; quaternium-18, lauralkonium chloride. chloride), olealkmonium chloride, cetylpyridinium chloride, polyquaternium-5, polyquaternium-6, polyquaternium-7, polyquaternium-10, polyquaternium-22, polyquaternium-37, polyquaternium-39, polyquaternium-47, cetyltrimonium chloride, dilauryldimonium chloride, cetalkonium chloride, dicetyldimonium chloride, soyaoiltrimonium chloride, and stearoctyldimonium methylsulfate.

[0127] Examples of suitable nonionic surfactants include fatty alcohol ethoxylates, such as octaethylene mono-n-dodecyl ester and pentaethylene glycol mono-dodecyl ether; alkylphenol ethoxylates (APE or APEO), such as nonoxynol-4, nonoxynol-7, nonoxynol-9, nonoxynol-14, nonoxynol-15, nonoxynol-18 and Triton X-100; glycerol fatty acid esters, such as glyceryl monostearate and glyceryl monolaurate; sorbitan fatty acid esters, such as sorbitan monolaurate, sorbitan monostearate, sorbitan tristearate, polysorbate (Tween) 20, polysorbate 40, polysorbate 60 and polysorbate 80.

[0128] Examples of suitable amphoteric (zwitterionic) surfactants include cocamidopropyl hydroxysultaine, cocamidopropyl betaine, lauryl betaine, dodecyldimethylamine oxide and myristylamine oxide.

[0129] Examples of suitable thickeners (rheology modifiers) include gums such as alginates, carrageenan, gum acacia, gum arabic, gum ghatti, gum karaya, gum tragacanth, guar gum; guar hydroxypropyltrimonium chloride, xanthan gum or gellan gum; cellulose derivatives such as sodium carboxymethylcellulose, hydroxyethylcellulose, hydroxymethylcarboxyethylcellulose, hydroxymethylcarboxypropylcellulose, ethylcellulose, sulfated cellulose, hydroxypropylcellulose, methylcellulose, hydroxypropylmethylcellulose, microcrystalline cellulose; agar; pectin; gelatin; starch and its derivatives; chitosan and its derivatives such as hydroxyethylchitosan; synthetic polymers such as polyvinyl alcohol, PVM / MA copolymer, PVM / MA decadiene crosspolymer, poly(ethylene oxide)-based thickeners.

[0130] The humectant can act as a hygroscopic agent, which increases the amount of water absorbed or retained by the composition. Examples of suitable humectants include acetamide MEA, ammonium lactate, chitosan and its derivatives, colloidal oats, galactoarabinan, glucose glutamate, glerecyth-7, glygeryth-12, glycereth-26, glyceryth-31, glycerol, lactamide MEA, lactamide DEA, lactic acid, methyl gluceth-10, methyl gluceth-20, panthenol, propylene glycol, sorbitol, polyethylene glycol, 1,3-butylene glycol, 1,2,6-hexanetriol, hydrogenated starch hydrolysate, inositol, mannitol, PEG-5 pentaerythritol ether, polyglyceryl sorbitol, xylitol, sucrose, sodium hyaluronate, and sodium PCA.

[0131] Examples of suitable preservatives include methylparaben, propylparaben and sorbic acid, methyldibromoglutaronitrile (MDBGN), benzyl alcohol, imidazolidinyl urea 1,3-bis(hydroxymethyl)-5,5-dimethyl-2,3-imidazolidinedione (DMDM hydantoin), methylchloroisothiazolinone and methylisothiazolinone, phenoxyethanol and sodium benzoate.

[0132] In one embodiment, the composition comprises one or more ingredients selected from the group consisting of emollients, anti-inflammatory agents, antioxidants, and UV blockers.

[0133] Examples of suitable emollients include fatty esters such as isopropyl myristate, isopropyl palmitate, caprylic / capric triglyceride, cetyl lactate, cetyl palmitate, hydrogenated castor oil, glyceryl esters, hydroxycetanol isostearate, hydroxycetanol phosphate, isopropyl isostearate, isostearyl isostearate, diisopropyl sebacate, PPG-5-ceteth-20, 2-ethylhexyl isononoate, 2-ethylhexyl stearate, C12 to C16 fatty alcohol lactates, isopropyl lanolate, and 2-ethyl-hexyl salicylate.

[0134] Examples of suitable anti-inflammatory ingredients include cyclooxygenase (e.g., COX-1 or COX-2) or lipoxygenase (e.g., LOX-5) enzyme inhibitors such as ascorbic acid, ascorbic acid derivatives, vitamin E, vitamin E derivatives, tocotrienols, rutin, quercetin, hesperidin (Citrus sinensis), hesperidin (Citrus sinensis), diosmin (Citrus sinensis), mangiferin (Mangifera indica), mangostin (Garcinia mangostana), cyanidin (Vaccinium myrtillus), astaxanthin (Haematococcus algae), lutein (Tagetes patula), lycopene (Lycopersicum esculentum), resveratrol (Polygonum cuspidatum), cuspidatum), tetrahydrocurcumin (Curcuma longa), rosmarinic acid (Rosmarinus officinalis), hypericin (Hypericum perforatum), ellagic acid (Punica granatum), chlorogenic acid (Vaccinium vulgaris), oleuropein (Olea europaea), alpha lipoic acid, glutathione, andrographolide (Andrographis paniculata), grape seed extract, green tea extract, polyphenols, pycnogenol (pine bark extract), white tea extract, black tea extract, (Andrographis paniculata), carnosine, and niacinamide.Further examples of suitable anti-inflammatory compositions may additionally be selected from horse chestnut extract (Aesculus hippocastanum extract), esculin, escin, yohimbine, Capsicum oleoresin, capsaicin, nicotinic acid, nicotinic acid esters, methyl nicotinate, benzyl nicotinate, ruscogenins (Butchers Broom extract; Ruscus aculeatus extract), diosgenin (Trigonel afoenumgraecum, Fenugreek), emblica extract (Phyllanthus emblica extract), asiaticoside (Centella asiatica extract), frankincense extract (Boswellia serrata extract), serrata), sericoside, visnadine, thiocolchicoside, grape seed extract, ginger root extract (Zingiber officianale), piperine, vitamin K, sweet osmanthus (Melilotus officinalis extract), glycyrrhetinic acid, ursolic acid, sericoside (Terminalia sericea extract), siegesbeckia glycoside (Siegesbeckiaorientalis extract), amni visnaga extract, grape leaf extract (Vitis vinifera), apigenin, soy extract (phytosan), luteolin.

[0135] Examples of suitable antioxidant ingredients include ascorbic acid, ascorbic acid derivatives, glucosamine ascorbate, arginine ascorbate, lysine ascorbate, glutathione ascorbate, niacinamide ascorbate, nicotinic acid ascorbate, allantoin ascorbate, creatine ascorbate, creatinine ascorbate, chondroitin ascorbate, chitosan ascorbate, carnosine ascorbate, vitamin E, vitamin E derivatives, tocotrienol, Phenol, rutin, quercetin, hesperidin (sweet orange), hesperidin (sweet orange), diosmin (sweet orange), mangiferin (mango), mangostin (mangosteen), cyanidin (European bilberry), astaxanthin (haematococcus), lutein (maraschino), lycopene (tomato), resveratrol (knotweed), tetrahydrocurcumin (turmeric), rosmarinic acid (rosemary), hypericin (Hypericum perforatum), ellagic acid (pomegranate), chlorogenic acid (Vaccinium vulgaris), oleuropein (Olea europaea), alpha lipoic acid, niacinamide lipoic acid, glutathione, andrographolide (Andrographis paniculata), carnosine, niacinamide, Potentilla erecta extract, polyphenols, grape seed extract, Pycnogenol (pine bark extract), pyridoxine, magnolol, honokiol, paeonol, resacetophenone, quinacetophenone, arbutin, and kojic acid.

[0136] Examples of suitable UV blockers (sunscreen actives) include octyl methoxycinnamate (ethylhexyl p-methoxycinnamate), octyl salicylate, oxybenzone (benzophenone-3), benzophenone-4, menthol anthranilate, dihydroxybenzone, aminobenzoic acid, amyl dimethyl PABA, diethanolamine p-methoxycinnamate, ethyl 4-bis(hydroxypropyl)aminobenzoate, 2-ethylhexyl 1-2-cyano-3,3-diphenylacrylate, homomenthyl salicylate, glyceryl aminobenzoate, dihydroxyacetone, octyl dimethyl PABA, 2-phenylbenzimidazole-5-sulfonic acid, triethanolamine salicylate, zinc oxide, titanium oxide, and mixtures thereof.

[0137] Further ingredients suitable for use in the composition include flavors, pH adjusters, pigments, odor absorbers, antimicrobial agents, antifungal agents, chelating agents, and sugars.

[0138] The compositions of the present invention are typically formulated for topical use.

[0139] The compositions of the present invention can be formulated as solutions, liquids, emulsions, creams, milks, dispersions, gels, or ointments. Examples of suitable emulsions include two-phase emulsions comprising an aqueous phase and an oil phase, such as oil-in-water (O / W) and water-in-oil (W / O), and multiple emulsions, such as triple emulsions (O / W / O and W / O / W).

[0140] The compositions and preparations can be prepared by any method well known in the art. Such method comprises the step of combining oleanolic acid and / or one or more compounds (the one or more compounds may be present in plant extracts) with a carrier constituting one or more auxiliary components. Generally speaking, preparations are prepared by uniformly mixing oleanolic acid and / or one or more compounds (the one or more compounds may be present in plant fungus extracts) with a carrier (e.g., a liquid carrier, a finely dispersed solid carrier, etc.).

[0141] The compositions of the present invention may be formulated both as cosmetics and as pharmaceuticals.

[0142] In one embodiment, a personal care product comprising a composition of the present invention is provided. Suitable personal care products include skin care products, hair care products, cleaning products, and cosmetic powders and liquids.

[0143] The example of suitable skin care products includes lotion / hand soap, skin cream / hand cream, skin ointment / hand ointment, skin cream / hand cream, lotion, shaving gel, shaving cream, sunscreen, deodorant, antiperspirant, sunscreen lotion, aftersun products, aftershave, body oil, bath oil and bubble bath. The example of suitable hair care products includes conditioner, hair detangling lotion, styling gel, styling cream, styling wax, styling lotion, mousse, gel water, hair tonic, spray and pomade. The example of suitable cleaning products includes liquid soap, bar soap, shower gel, cleanser and shampoo. The example of suitable cosmetic powder and liquid includes blush, talcum powder, contour powder, eye shadow, foundation, face powder, lip powder, makeup cake, liquid contour powder, eyeliner, lip gloss, lipstick and mascara.

[0144] Methods and uses for reducing lipid production

[0145] The compositions of the present invention reduce lipid production in sebaceous gland cells.

[0146] The present invention also provides a method for reducing lipid production in sebaceous gland cells, the method comprising contacting the sebaceous gland cells with a composition of the present invention.

[0147] Preferred features of the composition, such as oleanolic acid, the one or more compounds and the plant extract comprising the one or more compounds are as described above.

[0148] In one embodiment, the method for reducing lipid production in sebaceous gland cells is in vivo. In one embodiment, the method for reducing lipid production in sebaceous gland cells is ex vivo, such as in vitro.

[0149] In one embodiment, the method for reducing lipid production in sebaceous gland cells comprises contacting sebaceous gland cells with an effective amount of a composition of the present invention. The effective amount of the composition provides a detectable reduction in lipid production in sebaceous gland cells. The method for detecting and quantifying lipid production in sebaceous gland cells comprises, for example, using a suitable lipid detection dye, such as AdipoRed™ fluorescence assay.

[0150] In one embodiment, the method for reducing lipid production in sebocytes comprises contacting sebocytes with a composition of the present invention in an amount suitable for reducing lipid production by 10% or more, preferably 15% or more, more preferably 20% or more, even more preferably 20% or more, and most preferably 30% or more.

[0151] In one embodiment, the method for reducing lipid production in sebocytes reduces lipid production by 5% or more, preferably 10% or more, preferably 15% or more, more preferably 20% or more, even more preferably 20% or more, and most preferably 30% or more.

[0152] The invention also provides a composition of the invention for use in a method of reducing lipid production in sebaceous gland cells.

[0153] Preferred features of the composition are as described above. Preferred features of the method for reducing lipid production in sebaceous gland cells are also as described above.

[0154] The present invention also provides the use of the composition of the present invention for reducing lipid production in sebaceous gland cells.

[0155] Preferred features of the composition are as described above, such as oleanolic acid, the one or more compounds and plant extracts comprising the one or more compounds.Preferred features of the method for reducing lipid production in sebaceous gland cells also apply to the use of the composition for reducing lipid production in sebaceous gland cells.

[0156] Beauty methods and uses

[0157] The compositions of the present invention reduce lipid production in sebaceous gland cells, such as those in the skin of an individual. Thus, the compositions of the present invention can reduce or improve cosmetic problems associated with excessive production of lipids in the skin. Cosmetic problems associated with excessive production of lipids in the skin include oily or shiny skin, oily hair, enlarged skin pores, undesirable body odor, and reduced retention of cosmetics on the skin.

[0158] Accordingly, the present invention provides a cosmetic method for reducing lipid production in the skin of an individual, the method comprising contacting the skin with a composition of the present invention.

[0159] Preferred features of the composition are as described above.

[0160] In one embodiment, the cosmetic method for reducing lipid production in the skin of an individual is not a method of treatment. In one embodiment, the cosmetic method for reducing lipid production in the skin of an individual is not a method of treating the human or animal body by therapy.

[0161] In one embodiment, the cosmetic method reduces lipid production in sebaceous gland cells in the skin of a subject.

[0162] In one embodiment, the skin is skin on the head, such as skin on the face, mouth, neck, or scalp. In one embodiment, the skin is skin on the chest, back, arms, legs, or hands.

[0163] The beneficial effects of reducing lipid production in an individual's skin include reducing the oily appearance of the skin, controlling surface oil, minimizing skin pores, and reducing undesirable body odor. In one embodiment, the cosmetic method reduces lipid production in the skin, thereby achieving an effect selected from reducing the oily appearance of the skin, controlling surface oil of the skin, minimizing skin pores, and reducing undesirable body odor.

[0164] In one embodiment, the individual is in need of a cosmetic treatment. The individual in need of a cosmetic treatment may have a condition associated with an overproduction of lipids in the skin, such as oily or shiny skin, oily hair, enlarged skin pores, and undesirable body odor. In one embodiment, the individual has a cosmetic condition selected from oily or shiny skin, oily hair, enlarged skin pores, and undesirable body odor.

[0165] The present invention also provides a composition of the present invention for use in a cosmetic method of reducing lipid production in the skin of an individual.

[0166] Preferred features of the composition are as described above. Preferred features of the cosmetic method for reducing lipid production in the skin of an individual are also described above.

[0167] The present invention also provides the use of a composition of the present invention for reducing lipid production in the skin of a subject.

[0168] Preferred features of the composition are as described above.Preferred features of the cosmetic method for reducing lipid production in the skin of an individual also apply to the use of the composition for reducing lipid production in the skin of an individual.

[0169] Medical methods and uses

[0170] The compositions of the present invention reduce lipid production in sebaceous gland cells, such as those in the skin of an individual. Therefore, the compositions of the present invention can be used to treat or prevent (prevent) medical problems associated with excessive production of lipids in the skin. Medical problems associated with excessive production of lipids in the skin include acne vulgaris and rosacea.

[0171] Accordingly, the present invention provides a composition of the invention for use in a method of treatment, such as a method of treating the human or animal body by therapy.

[0172] In one embodiment, the method of treatment is a method of treating a disorder (eg, a disease) associated with overproduction of lipids in the skin. In one embodiment, the method of treatment is a method of treating a disorder associated with overproduction of lipids in sebaceous gland cells.

[0173] In one embodiment, the skin is skin on the head, such as skin on the face, mouth, neck, or scalp. In one embodiment, the skin is skin on the chest, back, arms, legs, or hands.

[0174] In one embodiment, the treatment is treatment of acne vulgaris. In one embodiment, the treatment is treatment of rosacea.

[0175] In one embodiment, treatment is administered to a subject in need of treatment.

[0176] The subject (patient) in need of treatment may be a mammal, such as a human.

[0177] The subject in need of treatment may be an adult or a teenager.

[0178] In a preferred embodiment, the subject in need of treatment is a human, more preferably an adult.

[0179] Alternatively, the subject in need of treatment is a non-human animal used in laboratory research.

[0180] In one embodiment, the treatment is administered by any convenient route of administration.In a preferred embodiment, the treatment is administered topically (ie, at the site of desired effect).

[0181] In one embodiment, the treating comprises administering to a subject in need of treatment a therapeutically effective amount of the composition.

[0182] It will be appreciated by those skilled in the art that the appropriate dosage of compositions as described herein may vary from patient to patient. Determining the optimal dosage will generally involve balancing the level of therapeutic beneficial effects with any risk or harmful side effects. The selected dosage level will depend on a variety of factors, including but not limited to the activity of oleanolic acid, the one or more compounds, or the plant extract comprising the one or more compounds, the route of administration, the time of administration, the excretion rate of oleanolic acid, the one or more compounds, or the plant extract comprising the one or more compounds, the duration of treatment, other drugs, compounds and / or materials used in combination, the severity of disorder, and the species, sex, age, body weight, illness, general health and previous medical history of the patient. The amount of oleanolic acid and the one or more compounds, or the plant extract comprising the one or more compounds, and the route of administration will ultimately be determined by a physician, a veterinarian, or a clinician, although the dosage is generally selected to achieve a local concentration at the site of action, which achieves the desired effect without causing substantial harmful or hazardous side effects.

[0183] Administration can be achieved in one dose (application), continuously or intermittently (e.g., in divided doses at appropriate intervals) throughout the course of treatment. Methods for determining the most effective mode of administration and dosage are well known to those skilled in the art and will vary with the formulation used for therapy, the purpose of the therapy, the one or more target cells being treated, and the subject being treated. Single or multiple administrations can be implemented according to the dosage level and pattern selected by the treating physician, veterinarian, or clinician.

[0184] Other Preferences

[0185] Each and every compatible combination of the above-described embodiments is expressly disclosed herein, as if each and every combination were individually and expressly recited.

[0186] Various other aspects and embodiments of the present invention will be apparent to those skilled in the art in view of this disclosure.

[0187] Where used herein, "and / or" should be considered as a specific disclosure of each of the two specified features or components, whether or not the other is present. For example, "A and / or B" is considered as a specific disclosure of each of (i) A, (ii) B, and (iii) A and B, just as if each were individually stated herein.

[0188] Unless the context dictates otherwise, the descriptions and definitions of the features set out above are not limited to any particular aspect or embodiment of the invention and apply equally to all aspects and embodiments described.

[0189] Example

[0190] Some aspects and embodiments of the present invention will now be described by way of example and with reference to the above-mentioned drawings.

[0191] Material

[0192] Dimethyl sulfoxide (DMSO), epigallocatechin gallate (EGCG), and salicylic acid (SA) were purchased from Sigma-Aldrich.

[0193] Oleanolic acid (OA) was purchased from Sabinsa Corporation.

[0194] Adenine, betaine, choline, ellagic acid, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, protocatechuic acid or tyramine were purchased from Sigma-Aldrich Pte Ltd at reagent grade.

[0195] Wild strawberry (FV) plant extract was purchased from BOC Sciences.

[0196] Hedera helix extract (HH), mangosteen (GM), Ilex aquifolium (European holly) extract (IA), and raspberry extract (RI) were purchased from BOCSCI (New York, USA).

[0197] Raspberry (RI) plant extract was purchased from BOC Sciences or BOCSCI (New York, USA).

[0198] Plant extracts can be obtained as described below.

[0199]

[0200] Primary sebaceous gland cells were derived from the forehead of a 48-year-old Chinese male. The same cell line was used in all experiments.

[0201] Raspberry Extract

[0202] The three raspberry solid plant extracts were deconvoluted using appropriate analytical techniques and the following compounds were identified in the extracts. The preparation of liquid samples for analysis from the solid extracts is described below.

[0203] Table 1. Components of raspberry solid plant extract

[0204]

[0205] Wild Strawberry Extract

[0206] The wild strawberry solid plant extract was deconvoluted using appropriate analytical techniques and the following compounds were identified in the extract. The preparation of liquid samples for analysis from the solid extract is described below.

[0207] Table 2. Composition of wild strawberry solid plant extract

[0208]

[0209] Analytical methods

[0210] Prepare liquid samples for analysis according to the protocol outlined below:

[0211] 1. Weigh 100 mg of dry powder of raspberry or wild strawberry extract sample and dissolve it in 1 mL of 50% ACN / water solution, then dilute 10X for LC-MS analysis;

[0212] 2. Dissolve the standard compound (adenine, betaine, choline, ellagic acid, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, protocatechuic acid, or tyramine) in 50% ACN / water solution in a 2 mL tube, vortex and sonicate for 10 minutes. Prepare a standard mixture with a final concentration of 1 ppm by combining each standard spiked with 10% (V / V) of an internal standard of known concentration (ISTD); and

[0213] 3. Inject 15 µL of sample and prepared standards into the LC-MS.

[0214] Liquid chromatography

[0215] Analyses were performed using a Waters Xevo® G2-S QToF and UltraPerformance LC® with an HSS T3 1.8um column and the following detection parameters:

[0216] 1. The mobile phase consisted of (A) water + 0.1% formic acid and (B) acetonitrile + 0.1% formic acid;

[0217] 2. The gradient was 0-1 min, 1% B isocratic; 1-11 min, 1-40% B linear; 11-13 min, 40-70% B linear; 13-15 min, 70-99% B linear; 15-16 min, 99% B isocratic; and 16-17 min, 99-1% B linear; the column was then re-equilibrated with 1% B isocratic for 3 min before the next run; and

[0218] 3. Maintain a constant column temperature of 40°C and autosampler temperature of 4°C throughout the 20-minute analysis. The sample injection volume is 15 µL and the sample loop is 2 µL.

[0219] Mass spectrometry

[0220] Mass spectrometry data were acquired in ESI positive ion mode and MS centroid acquisition mode using the following acquisition parameters to obtain product ion spectra:

[0221]

[0222] The amount of a given compound is then quantified by comparing its retention time and mass-to-charge ratio with those of standard compounds.

[0223] Characterization methods

[0224] Flow cytometry experiments were performed on a BD LSR Fortessa X-20 as follows: First, sebaceous gland cells stained with a lipid dye (AdipoRed™) were analyzed based on size and granularity, and only single cells were selected for AdipoRed™ signal intensity measurement. A 488 nm FITC blue laser was used to measure the AdipoRed™ signal, and the mean fluorescence intensity value of 10,000 cells was obtained. The mean fluorescence intensity is a measure of the amount of average lipid in 10,000 sebaceous gland cells. Cells treated with the composition of the present invention were compared with cells treated with a vehicle to determine the percentage of lipid reduction.

[0225] Primary sebaceous gland cell culture

[0226] Primary sebocytes were cultured to 80% confluence at 37°C with 5% CO2 in complete medium (CCMY) containing Y-27632, consisting of 3:1 DMEM (Gibco, 11995-065) / F12 (Gibco, 31765-035), supplemented with 10% fetal bovine serum (Hyclone, SV30160.03), 1x Penstrep (Gibco, 15140-122), 0.2 µg / mL epidermal growth factor (PeproTech, AF-100-15-1MG), 1 µg / mL hydrocortisone (Sigma-Aldrich, H0888), 10 -9M cholera toxin (Sigma-Aldrich, C8052-2MG) and 10 µM Y-27632 (Tocris, 1254 / 10). The cells were washed with PBS buffer [5 mL for a 10 cm plate] and then incubated with 0.125% trypsin-EDTA (Gibco, 15400054) [2 mL of trypsin for a 10 cm plate] at 37°C for 5-10 minutes. When all the cells were detached, neutralization medium consisting of DMEM containing 10% FBS and 1× Penstrep was added (2 mL of neutralization medium for a 10 cm plate), and the cells were transferred to a 15 mL conical tube and centrifuged at 1,000 rpm for 5 minutes.

[0227] After centrifugation, the cells were pelleted, the supernatant was discarded, and the cells were resuspended in culture medium.

[0228] Cells were cultured in a CelCulture CO2 Incubator 170L (Esco Lifesciences).

[0229] Cells were counted by loading onto a hemacytometer, visualized using a CKX41 inverted microscope (Olympus) and counted manually.

[0230] General test plan

[0231] The test composition was added to the wells of a 24-well plate to achieve the following final concentrations: 1.25 μM oleanolic acid (OA), 50 μM EGCG, 10 μM other compounds, and 31.25 μg / mL extract (4 μL of a 3,125 μg / mL stock solution made with DMSO in 396 μL medium). In some experiments, the plant extract was used at 62.5 μg / mL where indicated.

[0232] Primary sebaceous gland cells were cultured at 60 × 10 6cells (396 μL) were seeded into the culture medium. The cells were incubated at 37°C and 5% CO2 for 3 days. The culture medium was discarded, AdipoRed™ dye (0.06 %v / v in PBS) was added, and the cells were incubated at 37°C for 20 minutes. The staining solution was discarded, trypsin (0.125%; 250 μL) was added, and the cells were incubated at 37°C for 5-10 minutes. When all cells had detached, fixation solution (250 μL; MEM without phenol red, 10% fetal bovine serum, and 4% paraformaldehyde) was added, and the cells were transferred to a 5 mL polystyrene tube. The tubes were stored on ice prior to flow cytometry. The mean fluorescence intensity of the lipid signal was obtained from the flow cytometric graph, which was used to determine the inhibitory effect of the compound on sebaceous gland cells.

[0233] Sebum Reduction Example

[0234] The lipid-reducing effect of the composition of the present invention was measured compared to DMSO as a negative control and compared to the known lipid-reducing compound epigallocatechin gallate (EGCG) as a positive control. The results are shown in Tables 3 and 4 and Figures 1 to 3 middle.

[0235] Table 3. In vitro lipid reduction percentage for each entry. Note: OA was used at 1.25 µM, EGCG was used at 50 µM, and the other compounds were used at 10 µM.

[0236]

[0237] *Data measured in different experimental groups

[0238] A further series of experiments were subsequently conducted testing oleanolic acid alone and together with raspberry and wild strawberry plant extracts comprising components as shown in Tables 1 and 2, respectively, and the results are presented below. The experiments were performed as before, with the extracts and concentrations of compounds noted.

[0239] Table 4. In vitro lipid reduction percentage of each item

[0240]

[0241] As expected, the decrease in mean fluorescence intensity values ​​recorded between the experimental groups presented in Tables 3 and 4 showed some variation.

[0242] A further series of experiments were conducted using raspberry (RI) and wild strawberry (FV) extracts in combination with oleanolic acid (OA) compared to other plant extracts alone, including Hedera helix (HH), Ilex holly (IA) and Mangosteen (GM) and other plant extracts together with oleanolic acid. The activity was also compared with epigallocatechin gallate (EGCG) and salicylic acid (SA). The results are shown in Table 5 and Figure 4 and Figure 5 middle.

[0243] In these experiments, test compositions were added to wells of 24-well plates to achieve final concentrations of 10 μM compound (4 μL of a 10 mM stock solution made with DMSO in 396 μL of culture medium) or 31.25 μg / mL plant extract (4 μL of a 3,125 μg / mL stock solution made with DMSO in 396 μL of culture medium).

[0244] Table 5. In vitro lipid reduction percentage of each item

[0245]

[0246] The results show that the composition comprising oleanolic acid (OA) and plant extracts (raspberry extract (RI) and wild strawberry extract (FV)) provides improved lipid reduction compared to the individual compounds or plant extracts. The results show that the composition has a synergistic effect compared to the individual compounds and individual extracts.

[0247] For reference, data for other extracts and combinations are also provided.

[0248] References

[0249] Many publications are cited above to more fully describe and disclose the present invention and the prior art to which the present invention relates. The complete citations to these references are provided below. The entire contents of each of these references are incorporated herein.

[0250] Cheng et al., “Sebocytes, multifaceted epithelial cells: lipid production and holocrine secretion”, Int. J. Biochem. Cell Biol., 2010; Vol. 42, pp. 181-185.

[0251] WO 2020 / 263188

Claims

1. A composition comprising: Oleanolic acid and its salts and solvates; and One or more compounds selected from the group consisting of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, protocatechuic acid and tyramine, and salts, prodrugs and solvates thereof.

2. The composition of claim 1, wherein the one or more compounds are selected from the group consisting of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide and tyramine, and salts, prodrugs and solvates thereof.

3. The composition of claim 1, wherein the one or more compounds are selected from the group consisting of ellagic acid, betaine, choline, quercetin-3-O-glucuronide and tyramine, and salts, prodrugs and solvates thereof.

4. A composition according to any one of claims 1 to 3, comprising two or more compounds selected from said group, such as three or more compounds, such as wherein at least one compound is ellagic acid.

5. The composition according to claim 4, comprising: Oleanolic acid and its salts and solvates; and A combination of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid, pipecolic acid, and tyramine.

6. The composition according to claim 4, comprising: Oleanolic acid and its salts and solvates; and A combination of ellagic acid, adenine, betaine, choline, quercetin-3-O-glucuronide, p-coumaric acid, and pipecolic acid.

7. The composition according to any one of claims 1 to 6, wherein the molar ratio of oleanolic acid to the compound selected from said group is 1:x, wherein x is in the range of 0.1 to 20.

8. The composition according to claim 7, wherein the molar ratio of oleanolic acid to the compound selected from said group is 1:x, wherein x is in the range of 1 to 10, such as 2 to 10, such as 5 to 10, such as about 8.

9. A composition according to any one of claims 1 to 8, comprising: Oleanolic acid and its salts and solvates; and A plant extract comprising one or more of said compounds.

10. The composition according to claim 9, wherein the plant extract is a wild strawberry extract, a raspberry extract or a pineapple strawberry extract, such as a wild strawberry extract or a raspberry extract.

11. A skin care product selected from lotion / hand soap, cream / hand cream, ointment / hand cream, lotion, shaving gel, shaving cream, sunscreen, deodorant, antiperspirant, sunscreen, aftersun products, aftershave, body oil, bath oil and bubble bath, comprising a composition according to any one of claims 1 to 10.

12. A hair care product selected from the group consisting of conditioners, detangling lotions, styling gels, styling creams, styling waxes, styling lotions, mousses, gel waters, hair tonics, sprays and pomades, the hair care product comprising the composition according to any one of claims 1 to 10.

13. A cosmetic method for reducing lipid production in the skin of an individual, the method comprising contacting the skin with a composition according to any one of claims 1 to 10.

14. A method for reducing lipid production in sebaceous gland cells or in the skin of an individual, the method comprising contacting the skin with a composition according to any one of claims 1 to 10, wherein the method is not a therapeutic method.

15. A method for reducing lipid production in sebocytes, the method comprising contacting the sebocytes with a composition according to any one of claims 1 to 10.

16. A composition according to any one of claims 1 to 10 for use in reducing lipid production in sebocytes.

17. A composition according to any one of claims 1 to 10 for use in a method of treatment.

18. A composition according to any one of claims 1 to 10 for use in a method of treating a skin disease or disorder associated with overproduction of lipids in the skin.

19. A composition for use according to claim 18, wherein the disease or disorder is selected from acne vulgaris and rosacea.

20. Use of a composition according to any one of claims 1 to 10 for reducing lipid production in sebaceous gland cells.

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