EA039773B1 - Personal care compositions with glutathione precursor comprising nicotinamide and amino acids - Google Patents

Abstract

Provided are topical personal care compositions, comprising: a. glutathione precursor comprising, by weight of the composition: i. from 0.001 to 2 wt.% of cystine or a salt or ester thereof; ii. from 0.01 to 5 wt.% of a glutamate source; iii. from 0.01 to 5 wt.% of glycine; and iv. from 0.01 to 5 wt.% of nicotinamide; and b. a cosmetically acceptable carrier in the form of a water and oil emulsion; wherein the pH of the composition is in the range of from 3.5 to 8.5; and wherein the weight ratio of nicotinamide to the total amount of amino acids is from 20:1 to 50:1. Said compositions are provided for potentiating glutathione synthesis within skin cells. Also provided are methods of improving the appearance of skin and attaining even skin color and reducing pigmentation, age spots and discoloration, comprising applying to the skin said compositions.

Description

Glutathione (GSH) is a tripeptide that consists of glutamate, cysteine and glycine. It is present in all tissues in mammals. It is the main antioxidant in a living organism: it protects cells from oxidation by quenching reactive oxygen species. GSH is believed to play an important role in protecting cells from the cytotoxic effects of ionizing radiation, heat, certain chemicals and, to a large extent, solar ultraviolet radiation (Tyrell et al., Photochem. Photobiol. 47: 405-412, 1988; Meister, J Biol Chem 263: 205-217, 1988; Meister, Science 200: 471-477, 1985). This statement is true for all parts of the body, but it is especially important for the skin, which is so susceptible to the damaging effects of radiation, especially ultraviolet radiation, and environmental factors. A decrease in the intracellular concentration of glutathione in the skin is associated with cell damage, inflammation, darkening of the skin, discoloration of the skin, the appearance of spots or freckles caused by exposure to ultraviolet radiation, physiological aging, and the like. Therefore, it is highly desirable to increase the production of glutathione in the skin.

A logical approach would seem to be to provide cells with an exogenous source of GSH (eg, by ingestion or topical application). Unfortunately, GSH is not bioavailable when administered exogenously, that is, once localized outside the cell, it undergoes cleavage into its constituent amino acids (glutamate, cysteine, and glycine) for uptake by the cell and synthesis of the GSH tripeptide. Thus, GSH cannot be directly transported into cells and therefore does not by itself lead to an intracellular increase in glutathione. GSH biosynthesis occurs in a highly regulated manner in the cell. The amount of glutathione in the cells largely depends on the presence of cysteine in the cells. Cysteine, a constituent amino acid of GSH, can increase GSH levels in the cell, but the open sulfhydryl group of cysteine makes it unstable and reactive, and also causes a strong malodor. Unlike cysteine, cystine can be administered safely; cystine is transported into the cell and converted to cysteine inside the cell, and then cysteine becomes available for intracellular production of GSH.

Topical formulations containing various amino acids and other skin care actives have been described - see e.g. Tanojo US7300649B, Laboratoire Filorga product, Schlachter WO 00/03689, Ermolin et al US2011183040, Garlen et al US4707354, Muller et al. US8361446, Hermann et al. US8241681. Compositions have been described to stimulate intracellular production of glutathione. See, for example, Chiba et al.: US Pat. Mammone: US Patent 6149925 and Perricone Publication US 20060063718.

Cystine usually enters the body from food. However, delivery of cystine from topical formulations is challenging due to its extremely low solubility in a biologically acceptable vehicle in the neutral pH range, which is exactly the pH range needed for topical application. The solubility of cystine in water is 0.112 mg/ml at 25°C; cystine is more soluble in aqueous solutions with pH less than 2 or pH above 8.

The present invention is based in part on the unexpected finding that the combination of amino acids with nicotinamide provides a synergistic increase in intracellular glutathione levels, thus neutralizing the low cystine solubility at neutral pH hindrance.

Brief description of the invention

In one embodiment, the skin care composition of the present invention comprises:

a) a glutathione precursor comprising, by weight of the composition:

i) from 0.001 to 2% by weight of cystine or a salt or ester thereof;

ii) from 0.01 to 5 wt.% source of glutamate;

iii) from 0.01 to 5% by weight of glycine; and iv) from 0.01 to 5% by weight of nicotinamide and

b) a cosmetically acceptable carrier;

while the pH of the composition is in the range from 3.5 to 8.5, and the mass ratio of nicotinamide to total amino acid content is from 20:1 to 50:1.

In one embodiment, the topical skin cosmetic composition is a no-rinse composition, in particular a no-rinse non-solid composition.

In one embodiment, the present invention provides a method for improving the appearance of skin, comprising applying said skin care composition to the skin.

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In one embodiment, the present invention provides a method for providing uniform skin color and reducing pigmentation, age spots, and discoloration of the skin, comprising applying said composition to the skin.

The compositions of the present invention enhance the synthesis of glutathione in skin cells and, therefore, they can be used to improve the appearance of skin with age-related aging or photoaging as a result of exposure to ultraviolet/sunlight or environmental pollutants. A preferred method of obtaining the benefits of the composition is regular/chronic topical application of the composition to prevent the development of skin damage, which can be caused even by ordinary exposure to ultraviolet light or other environmental influences that generate reactive oxygen species.

Detailed description of the invention

Throughout the description and claims, the following terms take on the meanings expressly associated with the present invention, unless the context clearly indicates otherwise.

The phrases in one implementation and in some implementations used herein do not necessarily refer to the same implementation variant(s), although they may. In addition, the phrases in another embodiment and in certain other embodiments used herein do not necessarily refer to another embodiment, although they may. Thus, as described below, the various embodiments of the invention can be easily combined without departing from the scope or spirit of the invention. In addition, each of the examples is given in connection with various embodiments of the invention, which are intended to be illustrative and not limiting.

Except in the examples or where expressly stated otherwise, all numerical values in the present description, indicating the amount of material or reaction conditions, the physical properties of the materials and/or application, should be understood as modified by the word approximately. All amounts are based on the weight of the final composition, unless otherwise indicated. The present invention is to be considered as embracing all embodiments that are defined in the claims as being multiple dependent on each other, regardless of the fact that the claims can be defined without multiple dependency or redundancy. In defining any concentration or amount range, any particular upper concentration may be associated with any particular lower concentration or amount.

The term "comprising" means including, but not necessarily consisting of or composed of. In other words, the listed steps or options are not exhaustive.

The term skin should be understood to include skin on the face, neck, chest, back, arms (including underarms), hands, legs, buttocks and scalp.

The term "leave-on" composition refers to a composition that is applied to the skin and is not intended to be rinsed off or rinsed off for a certain period of time, in particular hours, in contrast to compositions for cleansing or washing or rinsing the skin, which are removed by rinsing or rinsing off. immediately or a few minutes after application.

Non-solid in relation to the composition means that the composition has a measurable viscosity (measured, for example, using a Brookfield DV-I + viscometer (20 rpm (RPM), RV6 spindle, 30 s, 20 ° C) in the range from 1 Pa- s (Pas) up to 500 Pas, preferably from 2 to 100 Pas, more preferably from 3 to 50 Pas.

The term skin care composition refers to any product applicable to the human body for enhancing appearance, sun protection, cleansing, odor control, moisturizing, or general aesthetics. Non-limiting examples of skin care compositions include skin lotions, creams, gels, lotions, face masks, sticks, shampoos, conditioners, shower gels, toilet soaps, antiperspirants, deodorants, shaving creams, depilators, pomades, foundations, mascaras. for eyelashes, self-tanning products and sunscreen lotions.

The term skin cosmetic composition refers to any product applied to the human body to improve appearance, protect against the sun, reduce wrinkles or other signs of photoaging, control odor, brighten skin, even skin tone, or general aesthetics. Non-limiting examples of topical skin cosmetic compositions include skin lotions, creams, face masks, gels, sticks, antiperspirants, deodorants, lipsticks, foundations, mascaras, liquid or gel body washes, soaps, self-tanning creams, and sunscreen lotions.

The skin care composition according to the present technology is preferably a non-rinsing, non-solid skin cosmetic composition, since such compositions are the most difficult to incorporate cystine due to its low solubility.

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Glutathione Precursor (GSH)

The GSH precursor of the present invention contains amino acids (glutamate and cystine and optionally glycine) and 4-substituted resonicotinamidercinol. In one embodiment, the amino acids in the GSH precursor are a source of cystine and glutamate. In one embodiment, the amino acids in the GSH precursor are a source of cystine and glutamate and glycine.

The amino acids that are included in the composition according to the present invention are present as L-stereoisomers, since this is the most common and natural isomeric form found in nature. Since the building blocks of natural proteins found in human skin, hair and nails are amino acids in the L-isomer form, it is expected that the L stereoisomeric amino acids contained in the skin care products of the present invention may interact more with these proteins, that is, they are inherently more biocompatible compared to the D stereoisomeric form. In addition, the commercial production and supply of the L-stereoisomeric amino acids is much greater than that of the D stereoisomeric form. Finally, L-amino acid stereoisomers are more cost-effective to produce, more sustainable, more environmentally friendly, and available at a lower cost than D-amino acid stereoisomers.

Any of the amino acids included in the present invention may be in the form of a salt, ester, or salt thereof, and the term cystine, glutamate source, and glycine as used herein also encompasses salts, esters, and salts of such esters. The salt, ester, and salt of such an ester are not particularly limited as long as they are suitable for topical use. For example, salts formed with an inorganic acid or an organic acid or anionic surfactants may be mentioned. As the inorganic acid, for example, hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid, phosphoric acid and the like can be mentioned, and as the organic acid C _G C ₁₈ linear, branched or cyclic, saturated or unsaturated, unsubstituted or substituted with heteroatoms, e.g. formic acid, acetic acid, trifluoroacetic acid, propionic acid, lactic acid, tartaric acid, oxalic acid, fumaric acid, succinic acid, maleic acid, citric acid, malonic acid, methanesulfonic acid, stearic acid, oleic acid, 12 -hydroxystearic acid, ricinoleic acid and the like.

As the base salt, for example, alkali metal salts such as sodium salt, potassium salt and the like, alkaline earth metal salts such as calcium salt, magnesium salt and the like, and others can be mentioned.

As the base salt, for example, alkali metal salts such as sodium salt, potassium salt and the like, alkaline earth metal salts such as calcium salt, magnesium salt and the like, and others can be mentioned.

Amino acid esters are typically C1- _C8 esters or salts thereof, or alternatively C1- _C5 esters or alternatively C1- _C3 esters. Such esters may be straight or branched or cyclic. Esters of cystine are particularly suitable because of their increased solubility compared to cystine. The methyl and ethyl esters of cystine or their salts are most preferred due to their effectiveness in enhancing glutathione production and providing antioxidant activity. When ester salts are used, the same salts as listed above are suitable.

The source of glutamate may be present in the form of its functional equivalents - glutamine, glutamic acid and/or pyroglutamic acid and/or salts thereof may be used. Pyroglutamic acid (and/or salts thereof) is preferred because it is more stable than glutamine or glutamic acid. In one embodiment, the amino acids in the GSH precursor are cystine and pyroglutamic acid (and/or salts thereof). In one embodiment, the amino acids in the GSH precursor are cystine and pyroglutamic acid and glycine (and/or salts or esters thereof).

Nicotinamide is a component of the GSH precursor included in the present invention. The nicotinamide used in the present invention is described by formula I:

Formula I, where R is -CONH ₂ .

The nicotinamide compounds used in the present invention may form salts, complexes, hydrates and solvates. Suitable salts are selected from: fluoride, chloride, bromide, iodide, acetate, ascorbate, benzoate, carbonate, citrate, carbamate, formate, gluconate, lactate, fatty acid carboxylate

- 3 039773 acid, fatty acid dicarboxylate, oxalate, methyl bromide, methyl sulfate, nitrate, phosphate, diphosphate, succinate, sulfate or trifluoroacetate.

It has been found that due to the incorporation of amino acids and nicotinamide as described in the present invention, the production of glutathione is synergistically increased and thus the disadvantage of limited cystine solubility is overcome.

In one embodiment, the composition of the present invention is a leave-on, non-solid composition in the form of a topical skin emulsion, lotion, gel, cream, or rapidly absorbing cream containing a glutathione precursor that contains cystine, glutamate (particularly pyroglutamic acid or a salt thereof, e.g. sodium pyroglutamate) and nicotinamide, at pH 3.5 to 8.5.

In one embodiment, the composition of the present invention is a leave-on, non-solid composition in the form of a topical skin emulsion, lotion, gel, cream, or rapidly absorbing cream containing a glutathione precursor that contains cystine, glycine, and nicotinamide, with pH 3.5 to 8.5, in particular pH 5 to 8.

In one embodiment, the composition of the present invention is a leave-on, non-solid composition in the form of a topical skin emulsion, lotion, gel, cream, or rapidly absorbing cream containing a glutathione precursor that contains cystine, glutamate (particularly pyroglutamic acid or salts thereof, e.g. sodium pyroglutamate), glycine and nicotinamide at a pH of 3.5 to 8.5, in particular at a pH of 5 to 8.

quantities

In one embodiment, the glycine is present in an amount of 0.001 to 2%, or alternatively 0.005 to 1%, or 0.008 to 0.5%, or alternatively 0.008 to 0.4%. In one embodiment, the source of glutamate (preferably pyroglutamate) is present in an amount of 0.01 to 10%, or alternatively 0.01 to 5%, or 0.05 to 1%, or alternatively 0.05 up to 0.5%. In one embodiment, the glycine source is present in an amount of 0.01 to 10%, or alternatively 0.01 to 5%, or 0.05 to 1%, or alternatively 0.05 to 0.5 %. In one embodiment, the nicotinamide is present in an amount of 0.05 to 10% or alternatively 0.1 to 8% or 0.2 to 5%.

The amount of nicotinamide is significantly predominant compared to the total amount of amino acids in the composition. In one embodiment, the weight ratio of nicotinamide to total amino acids is between 20:1 and 50:1. In one embodiment, the weight ratio is between 25:1 and 45:1.

Carrier

The compositions of the present invention also contain a cosmetically acceptable carrier. The amount of carrier can be from 1 to 99.9%, preferably from 70 to 95%, optimally from 80 to 90%. Suitable carriers include water, emollients, fatty acids, fatty alcohols, thickeners, and combinations thereof. The carrier may be aqueous, anhydrous, or an emulsion. Preferably the compositions are aqueous, in particular water and oil emulsions of the water-in-oil or oil-in-water type, or multiple water-in-oil-in-water or oil-in-water-in-oil emulsions. In cases where water is used, its content may be from 5 to 95%, preferably from about 20 to about 70%, optimally from 35 to 60%, based on the weight of the composition.

Emollients may serve as cosmetically acceptable carriers. They may be in the form of silicone oils, natural or synthetic esters, hydrocarbons, alcohols and fatty acids. The amount of emollients may be from 0.1 to 95%, preferably from 1 to 50%, based on the weight of the composition.

Silicone oils can be divided into volatile and non-volatile. As used herein, the term volatile refers to those materials that have a measurable vapor pressure at ambient temperature. Volatile silicone oils are preferably selected from cyclic (cyclomethicone) or linear polydimethylsiloxanes containing 3 to 9, preferably 5 to 6 silicon atoms. Non-volatile silicone oils used as an emollient material include polyalkylsiloxanes, polyalkylarylsiloxanes, and polyethersiloxane copolymers. Essentially non-volatile polyalkylsiloxanes used here include, for example, polydimethylsiloxanes with a viscosity of 5x10'6 to 0.1 m ² /s at 25°C. Among the preferred non-volatile emollients used in the compositions according to the present invention are polydimethylsiloxanes having a viscosity of from 1x10'5 to about 4x10'4 m ² /s at 25°C.

Another class of non-volatile silicones are emulsifying and non-emulsifying silicone elastomers. Representative of this category is the Dimethicone/Vinyl Dimethicone crosspolymer available as Dow Corning 9040, General Electric SFE 839 and Shin-Etsu KSG-18. Silicone waxes such as Silwax WS-L (Dimethicone-Copolyol Laurate) can also be used.

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Essential emollients include:

a) alkyl esters of saturated fatty acids having from 10 to 24 carbon atoms. Examples of such substances include behenyl neopentanoate, isononyl isonanoate, isopropyl myristate and octyl stearate;

b) esters of alkoxyacids (ether-esters), such as esters of fatty acids and ethoxylated saturated fatty alcohols;

c) esters of polyhydric alcohols, esters of ethylene glycol and mono- and di-fatty acids, esters of diethylene glycol and mono- and di-fatty acids, esters of polyethylene glycol (200-6000) and mono- and di-fatty acids, esters of propylene glycol and mono- and di-fatty acids, polypropylene glycol 2000 monostearate, ethoxylated propylene glycol monostearate, esters of glycerol and mono- and di-fatty acids, polyglycerol fatty acid esters, ethoxylated glyceryl monostearate, 1,3-butylene glycol monostearate, 1,3-butylene glycol distearate, polyoxyethylene polyol ester and fatty acids, esters of sorbitan and fatty acids and esters of polyoxyethylene sorbitan and fatty acids are acceptable esters of polyhydric alcohols. Particularly suitable are the esters of C ₁ -C ₃₀ alcohols and pentaerythritol, trimethylolpropane and neopentyl glycol;

d) wax esters such as beeswax, spermaceti wax and tribegenin wax;

e) esters of sugar and fatty acids such as sucrose polybehenate and sucrose polycottonsedate.

Natural ester emollients are mainly based on mono-, di- and triglycerides. Exemplary glycerides include sunflower oil, cottonseed oil, borage oil, borage seed oil, evening primrose oil, castor and hydrogenated castor oil, rice bran oil, soybean oil, olive oil, safflower oil, shea butter, jojoba oil, and combinations thereof. Emollients of animal origin are represented by lanolin oil and lanolin derivatives. The content of natural esters can range from 0.1 to 20% by weight of the compositions.

Hydrocarbons which are suitable cosmetically acceptable carriers include petrolatum, mineral oil, C ₁₁ -C ₁₃ isoparaffins, polybutenes, and in particular isohexadecane commercially available as Permethyl 101A from Presperse Inc.

Fatty acids having from 10 to 30 carbon atoms may also be suitable as cosmetically acceptable carriers. Examples of this category are pelargonic, lauric, myristic, palmitic, stearic, isostearic, oleic, linoleic, linolenic, hydroxystearic and behenic acids and mixtures thereof.

Fatty alcohols containing from 10 to 30 carbon atoms are another category of cosmetically acceptable carriers suitable for use. Examples of this category are stearyl alcohol, lauryl alcohol, myristyl alcohol, oleyl alcohol and cetyl alcohol and mixtures thereof.

Thickeners or rheology modifiers may be used as part of a cosmetically acceptable carrier for the compositions of the present invention. Typical thickeners include cross-linked acrylates (eg Carbopol 982®), hydrophobically modified acrylates (eg Carbopol 1382®), polyacrylamides (eg Sepigel 305®), polymers and copolymers of acryloylmethyl propane sulfonic acid/salts thereof (eg Aristoflex HMB® and AVC®), cellulose derivatives and natural gums. Suitable cellulose derivatives include sodium carboxymethyl cellulose, hydroxypropyl metocellulose, hydroxypropyl cellulose, hydroxyethyl cellulose, ethyl cellulose and hydroxymethyl cellulose. Natural gums suitable for the present invention include guar, xanthan, sclerotium, carrageenan, pectin, and combinations of these gums. Inorganic substances can also be used as thickeners, especially clays such as bentonites and hectorites, colloidal silicas, talc, calcium carbonate and silicates such as magnesium aluminum silicate (Veegum®). The thickener content may be from 0.0001 to 10%, typically 0.001 to 1% or 0.01 to 0.5%.

Preferred are emollients that can be applied (especially for products intended for application to the face) to improve organoleptic properties and are selected from the group consisting of polypropylene glycol-14 butyl ether, also known as Tegosoft PBE, or PPG15 stearyl ether ( polypropylene glycol-15) such as Tegosoft E, other oils such as esters, in particular isopropyl myristate, isopropyl palmitate. Other oils may include castor oils and their derivatives.

As cosmetically acceptable carriers, humectants such as polyhydric alcohols can be used. Representative polyhydric alcohols include glycerol, polyalkylene glycols, and more preferably alkylene polyols and their derivatives, including propylene glycol, dipropylene glycol, polypropylene glycol, polyethylene glycol and its derivatives, sorbitol, hydroxypropyl sorbitol, hexylene glycol, 1,3-butylene glycol, isoprene glycol, 1,2,6-hexanetriol, ethoxylated glycerol,

- 5 039773 propoxylated glycerin and mixtures thereof. The amount of humectant may be from 0.5 to 50%, preferably from 1 to 15%, based on the weight of the composition.

Skin humectants such as hyaluronic acid and/or its precursor N-acetylglucosamine may be included in the composition. N-acetylglucosamine can be found in shark cartilage or shiitake mushrooms and is commercially available from Maypro Industries, Inc (New York). Other preferred wetting agents include hydroxypropyl-tri(C ₁ -C ₃ alkyl) ammonium salts. These salts can be obtained by various synthetic methods, in particular by hydrolysis of chlorohydroxypropyl-tri(C ₁ -C ₃ alkyl) ammonium salts. The most preferred representative is 1,2-dihydroxypropyltrimonium chloride, where C ₁ -C ₃ alkyl is a methyl group. The amount of salt may be from 0.2% to 30%, preferably from 0.5% to 20%, optimally from 1% to 12%, based on the weight of the topical composition, including all ranges included therein.

Typically the C1- _C3 alkyl component in the quaternary ammonium group will be methyl, ethyl, n-propyl, isopropyl or hydroxyethyl and mixtures thereof. Particularly preferred is the trimethylammonium group, known by the INCI nomenclature (International Cosmetic Ingredients Nomenclature) as the trimonium group. Any anion can be used in the quaternary salt. The anion may be organic or inorganic, provided the material is cosmetically acceptable. Typical inorganic anions are halides, sulfates, phosphates, nitrates and borates. Most preferred are the halides, especially chloride. Organic anionic counterions include methosulfate, toluoyl sulfate, acetate, citrate, tartrate, lactate, gluconate, and benzenesulfonate.

Also other preferred wetting agents that can be used, especially in combination with the aforementioned ammonium salts, include a substituted urea such as hydroxymethylurea, hydroxyethylurea, hydroxypropylurea; bis(hydroxymethyl)urea; bis(hydroxyethyl)urea; bis(hydroxypropyl)urea; N,N'-dihydroxymethylurea; N,N'dihydroxyethylurea; N,N'-dihydroxypropylurea; N,N,N'-trihydroxyethylurea; tetra(hydroxymethyl)urea; tetra(hydroxyethyl)urea; tetra(hydroxypropyl)urea; N-methylN'-hydroxyethylurea; N-ethyl-N'-hydroxyethylurea; N-hydroxypropyl-N'-hydroxyethylurea; and N,N'-dimethyl-N-hydroxyethylurea. Here, the term hydroxypropyl appears in the sense that is common to the radicals 3-hydroxy-n-propyl, 2-hydroxy-n-propyl, 3-hydroxyisopropyl or 2-hydroxyisopropyl. Most preferred is hydroxyethyl urea available as a 50% aqueous solution from the National Starch & Chemical Division ICI under the trademark Hydrovance. The amount of substituted urea that can be used in the topical composition of the present invention is in the range of 0.01 to 20%, or 0.5 to 15%, or 2 to 10%.

When an ammonium salt and a substituted urea are used, most preferably at least 0.01 to 25%, or 0.2 to 20%, or 1 to 15% of a humectant such as glycerol is used. Additional moisturizing agents for use in the present invention include petrolatum and/or various aquaporin-acting actives and/or oatmeal.

composition pH

In one embodiment, the skin care composition has a pH of 3.5 to 8.5. In some embodiments, the pH of the skin care composition is between 3.5 and 8. In some embodiments, the pH of the skin care composition is between 5 and 7.8. In some embodiments, the pH of the skin care composition is between 5 and 7.5.

Preferred Optional Ingredients

In some embodiments, the skin care composition, and especially the cosmetic skin care composition of the present invention, includes a sunscreen. Typically, this is a combination of organic and non-organic sunscreens. It is especially important to include both UVA and UVB sunscreens.

The UV-B sunscreen oil may be selected from the group of cinnamic acid, salicylic acid, diphenylacrylic acid, or derivatives thereof. The UV-B sunscreen oil may contain one or more of: octyl salicylate, 3,3,5-trimethylcyclohexyl-2-hydroxybenzoate, ethylhexyl salicylate, 2-ethylhexyl-2-cyano-3,3-diphenyl-2-propenoate, or 2-ethylhexyl - 4-methoxycinnamate (also known as octylmethoxycinnamate or OMC). Such UV-B sunscreen oils are commonly commercially available, such as Octisalate™ (octyl salicylate), Homosalate™ (3,3,5-trimethylcyclohexyl-2-hydroxybenzoate), NeoHeliopan™ (a range of organic UV filters including OMS (Neo Heliopan AV™ ) and ethylhexyl salicylate (Neo Heliopan OS™)), Octocrylene™ and Milestab 3039™ (2-ethylhexyl-2cyano-3,3-diphenyl-2-propenoate) or Parsol MCX™ (2-ethylhexyl-4-methoxycinnamate)). The amount of UVB sunscreen oil in the skin care composition may be from 0.1 to 20 wt.%, or from 0.2 to 10 wt.%, or from 0.5 to 7 wt.%, or from 2 to 6 wt. %.

The skin care composition may further comprise a UV-B sunscreen that is water soluble. The UV-B water-soluble sunscreen may also contain phenyl bemidazole sulfonic acid (also known as ensulizol), 4-aminobenzoic acid (also known as para-aminobenzoic acid or PABA), or both.

The skin care composition of any of the above embodiments may further comprise 0.1 to 10 wt % UV-A sunscreen oil. The UV-A sunscreen oil may contain one or more of: 4-tert-butyl-4'-methoxydibenzoylmethane (avobenzone), 2-methyldibenzoylmethane, 4-methyldibenzoylethane, 4-isopropyldibenzoylmethane, 4-tert-methane, butyldibenzoylmethane, benzoylmethane, benzoylmethane,

2,4-dimethyldibenzoylmethane, 2,5-dimethyldibenzoylmethane, 4,4'-diisopropyldi2-methyl-5-isopropyl-4'-methoxydibenzoylmethane, 2-methyl-5-tert-butyl-4'-methoxydi2,4-dimethyl-4 '-methoxydibenzoylmethane, 2,6-dimethyl-4-tert-butyl-4'-methoxydi benzoylmethane, diethylaminohydroxybenzoylhexylbenzoate, ecamsul or methyl anthranilate. The amount of UV-A sunscreen oil in the skin care composition may be 0.5 to 7% by weight, or 1 to 5% by weight.

Additional suitable sunscreen oils suitable for use in skin care compositions include those commercially available from BASF Corporation: Uvinul T-150 (ethylhexyl triazone; UV-B sunscreen oil), Uvinul A Plus (diethylaminohydroxybenzoylhexyl benzoate; UV-A sunscreen oil) , Tinosorb S (bis-ethylhexyloxyphenol methoxyphenyltriazine; UVA and UVB sunscreen oil), Tinosorb M (methylenebisbenzotriazolyltetramethylbutylphenol; UVA and UVB sunscreen oil). The composition of the skin care compositions may also contain bisdisulizone disodium.

A particularly preferred combination of UV-A and UV-B sunscreen oils is avobenzone and 2-ethylhexyl-4-methoxycinnamate.

In some embodiments, the sunscreen is an inorganic sunscreen. Examples of inorganic sunscreens suitable for use in the skin care composition of the present invention include, but are not limited to, micronized titanium dioxide, zinc oxide, polyethylene, and various other polymers. The term finely divided refers to particles with an average size in the range from 10 to 200 nm, alternatively from 20 to 100 nm.

The amount of sunscreen, when present in a skin care composition according to some embodiments of the present invention, may be 0.1% to 30%, alternatively 2% to 20%, alternatively 4% to 10%.

The composition of the present invention preferably contains a skin lightening compound in addition to the nicotinamide included in the present invention in order to achieve optimum skin lightening performance at an optimum cost. Examples of substances are placenta extract, lactic acid, resorcinols (4-substituted, 2,5-disubstituted, 4,5-disubstituted and 4,6-disubstituted, in particular 4-hexyl, 4-methyl, 4-butyl, 4-isopropyl, phenylethyl resorcinols ), arbutin, kojic acid, ferulic acid, hydroquinone, resorcinol derivatives, including disubstituted resorcinols, and combinations thereof. In one embodiment, such a skin lightening compound is a tyrosinase inhibitor, most preferably a compound selected from the group consisting of kojic acid, hydroquinone, and 4-substituted resorcinols. In addition, dicarboxylic acids represented by the formula HOOC-(CxHy)-COOH, where x=4 to 20 and y=6 to 40, such as azelaic acid, sebacic acid, oxalic acid, succinic acid, fumaric acid, octadecenedioic acid (eg Arlatone DC) or salts or mixtures thereof, most preferably fumaric acid or a salt thereof, especially the disodium salt. The combination of 12HSA (12-hydroxystearic acid) with fumaric acid or salts thereof has been found to be particularly advantageous, especially for skin lightening formulations. The content of these agents may be from 0.1 to 10%, preferably from 0.5 to 2% by weight of the composition. Preferably, 4-alkyl-resorcinol and/or 12-hydroxystearic acid is used as the skin-lightening co-active agent of the present invention.

Another preferred ingredient in the compositions of the present invention is retinoid. As used herein, the term retinoid includes all natural and/or synthetic analogs of vitamin A or retinol-like compounds that have the biological activity of vitamin A in the skin as well as geometric isomers and stereoisomers of these compounds. The retinoid is preferably retinol, retinol esters (e.g. C ₂ -C ₂₂ retinol alkyl esters including retinyl palmitate, retinyl acetate, retinyl propionate), retinal and/or retinoic acid (including all-trans retinoic acid and/or 13-cis- retinoic acid), more preferably retinoids other than retinoic acid. These compounds are well known in the art and are commercially available from a number of sources such as Sigma Chemical Company (St. Louis, MO) and Boerhinger Mannheim (Indianapolis, IN). Other retinoids that are applicable here are described in US patent No. 4677120, issued June 30, 1987 Parish et al.; U.S. Patent US 4,885,311 issued December 5, 1989 to Parish et al.; US Patent US 5,049,584 issued September 17, 1991 to Purcell et al.; US Patent No. 5,124,356 issued June 23, 1992 to Purcell et al.; and patent

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US Reissue No. 34075 issued September 22, 1992 to Purcell et al. Other suitable retinoids are tocopheryl retinoate [tocopherol ester of retinoic acid (trans- or cis)], adapalene {6-[3-(1-adamantyl)-4-methoxyphenyl]-2-naphthoic acid} and tazarotene (ethyl-6- [2-(4,4dimethylthiochroman-6-yl)ethynyl]nicotinate). Preferred retinoids are retinol, retinyl palmitate, retinyl acetate, retinyl propionate, retinal, and combinations thereof. The compositions of the present invention may contain a safe and effective amount of a retinoid such that the resulting composition is safe and effective in controlling the condition of keratin tissue, preferably in controlling visible and/or palpable irregularities in the skin, more preferably in controlling the signs of aging in the skin, even more preferably to regulate visible and/or tangible inhomogeneities in skin texture associated with aging. The compositions preferably contain 0.005 to 2% or 0.01 to 2% retinoid. Retinol is preferably used in an amount of 0.01 to 0.15%; retinol esters are preferably used in an amount of 0.01 to 2% (eg 1%); retinoic acids are preferably used in an amount of 0.01 to 0.25%; tocopheryl retinoate, adapalene and tazarotene are preferably used in an amount of 0.01 to 2%.

Various plant extracts may optionally be included in the compositions of this invention. Illustrative examples are pomegranate, white birch (Betula Alba), green tea, chamomile, licorice, and combinations of these extracts. The extracts can be either water soluble or water insoluble and contained in a solvent which is respectively hydrophilic or hydrophobic. Water and ethanol are the preferred extract solvents.

Materials such as resveratrol, alpha-lipoic acid, ellagic acid, kinetin, retinoxytrimethylsilane (available from Clariant Corp. under the trademark Silcare 1M-75), dehydroepiandrosterone (DHEA) and combinations thereof may also be included. Ceramides (including ceramide 1, ceramide 3, ceramide 3B, ceramide 6 and ceramide 7) and pseudoceramides can also be used in many of the compositions of the present invention, however, these materials can also be excluded. The amount of these materials can be from 0.000001 to 10%, preferably from 0.0001 to 1%.

The skin care composition may further comprise from about 0.1 to about 8 wt. % film-forming polymer. Such film-forming polymers include, but are not limited to, polyalkyleneoxy-terminated polyamides (e.g., INCI nomenclature: polyamide-3, polyamide-4), polyether polyamides (e.g., INCI nomenclature: polyamide6), acid-terminated mixed polyamides. groups (eg, INCI nomenclature: polyamide-7) and poly(ester-amides) with terminal esters (eg, INCI nomenclature: polyamide-8). Such film-forming polymers can be synthesized or commercially available, such as the Sylvaclear™ product line from Arizona Chemical Company, and the LLC and OleoCraft™ product line from Croda International PLC. Film forming polymers also include, but are not limited to, INCI polyester-5 (e.g., Eastman AQ™ 38S polymer), PPG17/IPDI/DMPA copolymer (e.g., Avalure™ UR 450 polymer), acrylate copolymer (e.g., Avalure™ polymer AC 120) and polysaccharides such as xylogel (tamarin gum), lotus bean gum, tara gum, beta-glucan, pullulan, carboxymethyl cellulose, hydroxypropyl cellulose, sodium alginate, potato starch, carrageenan. The film-forming polymer may contain combinations of any two or more of the polymers mentioned above. The amount of film-forming polymer in the skin care composition may be from 0.1 to 8% by weight.

Desirably, preservatives are included in the composition of the present invention to protect against the growth of potentially harmful microorganisms. Suitable conventional preservatives for the compositions of this invention are para-hydroxybenzoic acid alkyl esters. Other preservatives that have come into use recently include hydantoin derivatives, propionate salts, and various quaternary ammonium compounds. Cosmetologists are familiar with the appropriate preservatives and select them regularly to perform a preservative test and ensure product stability. Particularly preferred preservatives are iodopropynyl butyl carbamate, phenoxyethanol, caprylyl glycol, C1-6 parabens (especially methyl paraben and/or propyl paraben), imidazolidinyl urea, sodium dehydroacetate and benzyl alcohol. Preservatives should be selected taking into account the application of the composition and possible incompatibilities between preservatives and other ingredients in the emulsion. Preservatives are preferably used in amounts ranging from 0.01 to 2%. A particularly preferred combination is octocrylene and caprylyl glycol, since it has been disclosed that caprylyl glycol enhances UV-A and UV-B protection.

Suitable antifungal agents for inclusion in skin care compositions are well known to those skilled in the art. Examples include, but are not limited to, climbazole, ketoconazole, fluconazole, clotrimazole, miconazole, econazole, etaconazole, terbinafine, salts of any one or more of them (e.g., hydrochloride salts), zinc pyrithione, selenium disulfide, and combinations of any two or more of them.

- 8 039773

In some embodiments, the skin care compositions of the present invention include vitamins. Examples of vitamins are vitamin A (retinol), vitamin B2, vitamin B3 (niacin), vitamin B6, vitamin B12, vitamin C, vitamin D, vitamin E, vitamin K and biotin. You can also use derivatives of vitamins. For example, vitamin C derivatives include ascorbyl tetraisopalmitate, magnesium ascorbyl phosphate, and ascorbyl glycoside. Vitamin E derivatives include tocopheryl acetate, tocopheryl palmitate, and tocopheryl linoleate. You can also use DL-panthenol and its derivatives. In some embodiments, the B3 vitamin derivative is a nicotinamide riboside. In some embodiments, the vitamin B6 derivative is pyridoxine palmitate. Flavonoids can also be used, in particular glucosylhesperidin, rutin, soy isoflavones (including genistein, daidzein, equol and their glucosyl derivatives) and mixtures thereof. The total amount of vitamins or flavonoids, when present, can be in the range of 0.0001 to 10%, alternatively 0.001 to 10%, alternatively 0.01 to 10%, alternatively 0.1 to 10%, alternatively 1 up to 10%, alternatively from 0.01 to 1%, alternatively from 0.1 to 0.5%.

In some embodiments, the skin care compositions of the present invention comprise an enzyme, such as, for example, oxidases, proteases, lipases, and combinations thereof. In some embodiments, the skin care compositions of the present invention comprise superoxide dismutase, commercially available as Biocell SOD from Brooks Company, USA.

In some embodiments, the skin care compositions of the present invention comprise desquamation promoters. In some embodiments, the skin care compositions of the present invention comprise desquamation promoters at a concentration of 0.01 to 15%, alternatively 0.05 to 15%, alternatively 0.1 to 15%, alternatively 0.5 to 15%. %. Illustrative examples of desquamation promoters include monocarboxylic acids. Monocarboxylic acids may be substituted or unsubstituted with a carbon chain length of up to 16 carbon atoms. In some embodiments, the carboxylic acids are alpha hydroxy carboxylic acids, beta hydroxy carboxylic acids, or polyhydroxy carboxylic acids. The term acid means the content in the composition not only of the free acid, but also of its salts and C1- _C30 alkyl or aryl esters and lactones, formed upon removal of water to form cyclic or linear lactone structures. Typical acids include glycolic, lactic, malic and tartaric acids. In some embodiments, the salt is ammonium lactate. In some embodiments, the beta-hydroxycarboxylic acid is salicylic acid. In some embodiments, phenolic acids include ferulic acid, salicylic acid, kojic acid, and salts thereof.

In some embodiments, at least one additional component may be present from 0.000001 to 10%, alternatively from 0.00001 to 10%, alternatively from 0.0001 to 10%, alternatively from 0.001 to 10%, alternatively from 0.01 up to 10%, alternatively from 0.1 to 10%, alternatively from 0.0001 to 1%, based on the weight of the composition. Compositions according to the present invention may contain dyes, opacifiers or abrasives. Colorants, opacifiers or abrasives may be present in a concentration of 0.05 to 5%, alternatively 0.1 to 3% by weight of the composition.

In some embodiments, the personal care product of the present invention may also contain a peptide, such as, for example, the commercially available pentapeptide derivative, Matrixyl™, which is commercially available from Sederma, France. In another example, in some embodiments, the personal care product of the present invention may also contain carnosine.

Compositions according to the present invention may contain a wide range of other optional components. The CTFA Cosmetic Ingredient Handbook, Second Edition, 1992, which is incorporated herein by reference in its entirety, describes a wide range of non-limiting cosmetic and pharmaceutical ingredients commonly used in the cosmetic care industry. skin for topical application, which are suitable for use in the composition according to the present invention. Examples include antioxidants, binders, biological additives, buffering agents, dyes, thickeners, polymers, binders, fragrances, moisture-retaining agents, matting agents, conditioners, exfoliating agents, pH adjusters, preservatives, natural extracts, essential oils, skin sensitizers , skin soothing agents and skin healing agents.

Composition form

The compositions according to the present invention are preferably non-solid. The compositions according to the present invention are preferably non-rinsing compositions. The compositions of the present invention are preferably leave-on compositions intended to be left on the skin. These leave-in compositions are to be distinguished from compositions that are applied to the skin and subsequently removed either by washing, rinsing, rubbing or the like, either after or during application of the product. Surfactants commonly used for rinse-off compositions have fi

- 9 039773 zyco-chemical properties, which enable them to form a foam/foaming liquid when lightly rinsed; they may consist of mixtures of anionic, cationic, amphoteric and non-ionic surfactants. Surfactants used in leave-on compositions, on the other hand, need not have such properties. Rather, since the rinse-off compositions are not intended to be rinsed off, they must be non-irritating, and therefore it is necessary to minimize the total surfactant content and the total anionic surfactant content of the leave-on compositions. The total content of surfactant in the compositions according to the present invention is preferably from 1% to not more than 10%, more preferably less than 8%, most preferably not more than 5%, optimally not more than 3%.

In some embodiments, anionic surfactants are present in the leave-in skin care composition in an amount of 0.01 to no more than 5% by weight of the composition, alternatively 0.01 to 4% by weight of the composition. by weight of the composition, alternatively from 0.01 to 3 wt.% based on the weight of the composition, alternatively from 0.01 to 2 wt.% based on the weight of the composition, alternatively essentially absent (less than 1% or less than 0.1% or less than 0.01%). In some embodiments, the total surfactant content of the skin care compositions is no more than 10%, alternatively less than 8%, alternatively no more than 5%.

In some embodiments, the surfactant is selected from the group consisting of anionic, nonionic, cationic, and amphoteric active agents.

In some embodiments, the nonionic surfactants are C ₁₀ -C ₂₀ fatty alcohol or hydrophobic acid surfactants condensed with 2 to 100 moles of ethylene oxide or propylene oxide per mole of hydrophobic substance; C ₂ -C ₁₀ alkylphenols fused with 2 to 20 moles of alkylene oxide; esters of mono- and di-fatty acids and ethylene glycol; fatty acid monoglyceride; sorbitan, mono- and di-C ₈ C20 fatty acids; and polyoxyethylene sorbitan, as well as combinations thereof. In some embodiments, the nonionic surfactant is selected from the group consisting of alkyl polyglycosides, saccharide fatty amides (eg, methylgluconamides), and trialkylamine oxides.

Amphoteric surfactants suitable for skin care compositions according to some embodiments of the present invention include cocoamidopropyl betaine, C ₁₂ -C ₂₀ trialkyl betaines, sodium lauroamphoacetate, and sodium laurodiamphoacetate.

Suitable anionic surfactants for skin care compositions in accordance with some embodiments of the present invention include soap, alkyl ether sulfates and sulfonates, alkyl sulfates and sulfonates, alkyl benzene sulfonates, alkyl and dialkyl sulfosuccinates, C ₈ -C ₂₀ acyl isethionates, C ₈ - C ₂₀ alkyl ether phosphates, C ₈ -C ₂₀ sarcosinates, C ₈ -C ₂₀ acyllactylates, sulfoacetates, and combinations thereof.

Compositions according to the present invention are usually in the form of oil-in-water or water-in-oil type emulsions. In some embodiments, the skin care compositions are fast-absorbing creams and oil-in-water emulsion creams or lotions.

The basis for a fast-absorbing cream consists of 5-40% fatty acids and 0.1-20% soap. In such creams, the fatty acid is preferably essentially a mixture of stearic acid and palmitic acid, and the soap is preferably the potassium salt of a mixture of fatty acids, although other counterions and mixtures thereof may be used. The fatty acid in a fast absorbing cream base is often made using stearic acid, which is essentially (typically about 90 to 95%) a mixture of stearic acid and palmitic acid. Typical stearic acid (hysteric acid) contains approximately 52-55% palmitic acid and 45-48% stearic acid (hysteric acid) of the total palmitin-stearin mixture. Thus, the inclusion of stearic acid and its soap to provide a fast-absorbing cream base is within the scope of the present invention. It is particularly preferred that the composition contains more than 7%, preferably more than 10%, more preferably more than 12% fatty acids. A typical facial cream base is structured with a crystallization network and is sensitive to the addition of various ingredients.

In one embodiment, the skin care composition is formulated as an oil-in-water emulsion with cystine substantially solubilized in the aqueous phase. In one embodiment, the skin care composition is formulated as a water-in-oil emulsion with cystine in aqueous droplets, wherein at least 90% of the droplets have a diameter in the range of 100 nm to 20 µm, or alternatively 200 nm to 20 or up to 10 microns.

In some embodiments, in addition to containing the GSH precursor, the skin care composition is formulated as a face mask. In some embodiments, in addition to containing a GSH precursor, the skin care composition is formulated as a face mask according to the formulations described in US Pat. No. 5,139,771. In some embodiments, in addition to containing a GSH precursor, the skin care composition behind the skin is formulated as a face mask according to the formulations described in U.S. Patent No.

- 10 039773

4,933,177. In some embodiments, in addition to containing a GSH precursor, the skin care composition is formulated as a face mask according to the formulations described in US Pat. No. 6,001,367.

In some embodiments, in addition to containing a GSH precursor, the skin care composition is formulated as a shampoo. In some embodiments, the skin care compositions of the present invention are formulated as a deodorant. In some embodiments, in addition to containing a GSH precursor, the skin care composition is formulated as a deodorant according to the formulations described in US Pat. No. 7,282,471. In some embodiments, the skin care compositions of the present invention are formulated as an antiperspirant. In some embodiments, in addition to containing a GSH precursor, the skin care composition is formulated as an antiperspirant according to the formulations described in US Pat. No. 7,282,471.

In some embodiments, the skin care compositions of the present invention are formulated as a disposable skin care wipe. In some embodiments, in addition to containing a GSH precursor, the skin care composition is formulated as a disposable skin care wipe according to the formulations described in US Pat. No. 7,282,471.

In some embodiments, the skin care compositions of the present invention are in the form of a bar of soap. In some embodiments, in addition to containing a GSH precursor, the skin care composition is formulated as a bar of soap according to the formulations described in US Pat. No. 7,282,471.

Methods for preparing skin care compositions

In some embodiments, the skin care compositions of the present invention can be obtained by:

a) mixing all water-soluble ingredients, including preservatives, thickening polymer, optionally glycerin and water;

b) heating the mixture to a temperature of 70-90°C;

c) mixing all oil-soluble ingredients and the compound of formula (1) to a temperature of 70-90°C;

d) adding the mixed oil-soluble ingredients to the heated mixture of water-soluble ingredients and mixing by turbulence while maintaining the mixture at a temperature of 70-90°C; and

e) cooling the mixture to room temperature without stopping stirring.

In some embodiments, the skin care compositions of the present invention are prepared by emulsifying:

a) dissolving the required amount of cystine in a high pH aqueous solution (9 to 14 or 9 to 12),

b) obtaining a macroemulsion of this solution in oil with an emulsifier, then

c) adding, with stirring, an acidic aqueous solution to obtain an emulsion with a pH in the neutral range required for a topical formulation, and finally

d) subjecting the emulsion to high shear, homogenization or sonolysis, for example using a homogenizer such as a Nano DeBee homogenizer from BEE International (Massachusetts, USA) or a Sonolator homogenizer manufactured by Sonic Corporation (Connecticut, USA) to obtain the final emulsion with a homogeneous neutral pH range, with more than 90% of the droplets having a diameter in the size range from 100 nm to 20 μm.

Method of using skin care compositions

In some embodiments, the skin care composition is applied topically to a person's skin. In some embodiments, the skin care composition provides at least one benefit selected from the group consisting of: skin conditioning, skin smoothing, reduction of wrinkling or signs of skin aging, reduction of skin inflammation, reduction of dryness, reduction of age spots, reduction of sunburn. and skin lightening.

In some embodiments, a small amount of the skin care composition, e.g., 1 to 5 ml, is applied to exposed skin from a suitable container or applicator and, if necessary, then spread over the surface and/or rubbed into the skin using the hand or fingers. , or a suitable device.

Further reference is made to the following examples, which together with the above descriptions illustrate some embodiments of the invention in non-limiting examples.

Examples

Experimental Methods

Powders of the following amino acids (pyroglutamic acid and glycine) were purchased from Sigma and individual stock solutions of each amino acid were prepared by dissolving the powder in water (pH 7). All amino acids were L-stereoisomers. Stock solutions of cystine

- 11 039773 (Sigma) was obtained in 0.5 M sodium hydroxide (pH 12), since cystine is poorly soluble in neutral solutions. Pyroglutamic acid sodium salt (also known as sodium pyrrolidone carboxylate (NaPCA)) was supplied as a 50% solution in water from Ajinomoto. Cystine esters, dihydrobutyl-L-cystinate dihydrochloride (DTBC) and diethyl-L-cystinate dihydrochloride (DEC) were purchased from Bachem; L-cystine dimethyl ether dihydrochloride (CDME) from Sigma. To create a mixture that will hereinafter be described as GAP, appropriate volumes of three amino acids (source of glutamate (i.e. NaPCA), cystine and glycine) were combined in Hanks' balanced salt solution (Sigma) such that the source of glutamate, cystine and glycine were in a mass ratio of 0.5:1:1. The specific concentrations used in each test are given below.

Additional test reagents: nicotinamide and picolinamide (Sigma-Aldrich) and nicotinamide riboside (Chromadex). Nicotinamide and derivatives/related compounds are water soluble.

Glutathione Analysis

Aged human keratinocytes (Cascade Biologies, currently ThermoFisher Scientific, Waltham, MA) were cultured in EpiLife® medium containing 60 μM calcium chloride and 1% human keratinocyte growth supplement (both media components from ThermoFisher Scientific) in a humidified atmospheric incubator. containing 5% CO2 at 37°C. The medium was updated every 2-3 days. Cells were then trypsinized at 90% confluence and seeded at 1 x 10 ⁴ per well in 96-well white walled clear bottom plates. One or two days after seeding, the cell medium was replaced with Hanks' balanced salt solution (HBSS) and the keratinocytes were pretreated with GAP (or vehicle control - 0.12-0.240 mM sodium hydroxide, pH 7.5) or additional test reagents described above, separately or in combination, or with L-cystine esters alone for 1-2 hours. The concentrations of all amino acids constituting the GAP mixture used in this assay are indicated in the data tables and are fully described in the list of abbreviations below.

After one or two hours of pre-treatment with GAP (and additional test reagents where described), keratinocytes were then infected with 25 μM menadione (Sigma-Aldrich), a known oxidative stress inducer. After 18-20 hours, cells were harvested and total glutathione levels measured using a commercially available kit (GSH-GSSG GLo Assay, Promega): a luminescence-based system designed to detect and quantify total glutathione in cultured cells. Finally, after cell treatment, the medium was removed and replaced with a cell lysis reagent for 5 min at 20-25°C. Note that all remaining steps were also carried out at 20-25°C. The luciferase formation reagent was then added to each well and incubated for 30 min. After this step, the luciferin detection reagent was added to each well and the plates were incubated for 15 minutes. Finally, the plates were then read for luminescence in a FlexStation 3 plate reader (Molecular Devices). Total glutathione levels were calculated after interpolation of glutathione concentrations from a standard curve. Determinations under experimental conditions were carried out at least three times. Statistical significance was calculated using 1way ANOVA (one-way analysis of variance). Percent protection was calculated based on the calculated glutathione levels from each experiment according to the following equation: (test sample) - (vehicle control + vehicle control

X 100% = % pro tection

Analysis of reactive oxygen species (ROS)

Human keratinocytes (Promocell, Heidelberg, Germany) were cultured in EpiLife® keratinocyte medium containing 60 μM calcium chloride and 1% human keratinocyte growth supplement (HKGS) in a humidified incubator with an atmosphere containing 5% CO ₂ at 37°C. The medium was updated every other day.

The cells were then trypsinized at 90% confluence and seeded at a density of 1-2 x 10 ⁴ per well in 96-well black wall clear bottom plates. On the second day after seeding, the cell medium was replaced with Hanks' balanced salt solution (HBSS) with or without menadione (10-25 μM), GAP (usually with 49 μM pyroglutamic acid, 60 μM cystine, 194 μM glycine), vehicle for GAP (0 .2 mM NaOH, pH 7.5) with the additional test reagents described above, combinations of GAP with additional test reagents, or L-cysteine esters alone, and incubated for an additional 18 h. For ROS detection, CellROX green reagent (Life Tech, Thermo Fisher Scientific, Rockford, USA) was added to each well to a final concentration of 15-25 μM and incubated for an additional 2-4 hours at 37° C. in a humidified incubator. ROS fluorescence was recorded using a Tesap microplate reader (excitation/emission = 490/525). Relative ROS production was calculated with normalization based on vehicle control, the latter was set at 100%. Where indicated, percent protection was calculated, compared to menadione alone, using measured relative fluorescent units according to the following equation:

- 12 039773 (vehicle control + methadion) - (test sample) (vehicle control + methadion) = % protection

ROS levels were assessed along with cell viability assessment because it is possible to get a false negative result for ROS if the treatment is so toxic that the cells die during the assay (and there are no more detectable ROS). Reducing ROS levels makes sense if the cells are still viable.

Cell viability analysis

Human keratinocytes (Promocell, Heidelberg, Germany) were maintained in EpiLife® keratinocyte medium containing 60 μM calcium chloride and 1% human keratinocyte growth supplement (HKGS). The medium was updated every other day. Cells were then trypsinized at 90% confluence and plated at 2x104/well in 96-well transparent tissue culture plates. On the second day after seeding, the cell medium was changed using treatments as described above in the ROS analysis section. After 18 hours, cell culture kit reagent (CCK-8) (Dojindo, Kumamoto, Japan) was diluted 1:10 in HBSS and incubated with cells for 4 hours in a humidified incubator at 37°C. Optical absorbance at 450 nm was measured using a Tesap Safire2 plate reader. Relative viability was calculated with normalization based on vehicle control treatment, the latter was set at 100%.

Where indicated, percentage protection was calculated, compared to menadione alone treatment, using relative absorbance units according to the following equation: test sample - (vehicle control + m vehicle control + methadione - % protection

The examples use the following abbreviations:

mM = millimolar µM = micromolar µl = microliter

GAP = glutamate and a mixture of cystine and glycine

GAP 5 = 4 μM pyroglutamic acid sodium salt, 5 μM cystine, 16 μM glycine

GAP 10 = 8 μM pyroglutamic acid sodium salt, 10 μM cystine, 32 μM glycine

GAP 20 = 16 μM pyroglutamic acid sodium salt, 20 μM cystine, 64 μM glycine

DEC = diethyl-1_-cystinate dihydrochloride

CDME = L-cystine dimethyl ether dihydrochloride

DTBC = di-tert-butyl-1_-cystinate dihydrochloride

Example 1

Example 1 evaluated various combinations of nicotinamide (within the scope of the invention) or compounds outside the scope of the invention - either structurally similar (picolinamide) or derivatives of nicotinamide (nicotinamide riboside or N-methylnicotinamide) - in combination with amino acids that are building blocks GSH, to stimulate intracellular production of GSH or reduce reactive oxygen species. The results obtained are summarized in Tables 1A and 1B. Table 1C-1H also tested the efficacy of various cysteine esters.

Table 1A. Synthesis of glutathione

SAMPLE Average value (μM glutathione) % Protection BUT Carrier control 2.86 AT Carrier Control + Methadione 0.19 0 Samples tested With Nicotinamide riboside (0.5 mM) + Methadione 0.16 0

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D Nicotinamide (0.5 mM) + Metadione 0.17 0 E Nicotinamide (5 mM) + Methadione 0.14 0 F Nicotinamide (10 mM) + Methadione 0.20 one G Picolinamide (0.5 mM) + Metadione 0.23 2 H Picolinamide (5 mM) + Methadione 0.18 0 one Picolinamide (10 mM) + Methadione 0.16 0 J GAP 10+ Methadione 3.71 ¹ 123 ¹ To Nicotinamide riboside (0.5 mM) + GAP 10+ Methadione 3.80 ^{1 3} 126 ¹³ L Picolinamide (0.5 mM) + GAP 10+ Methadione 2.94 ^{15 96 1'5} M Picolinamide (5 mM) + GAP 10+ Methadione ^{2.11 1'5 67 1'5} N Picolinamide (10 mM) + GAP 10+ Methadione 2.84 ^{15 93 1'5} Q GAP 20+ Methadione 6.71 ^{12 228 1'2} one Nicotinamide (0.5 mM) + GAP 10+ Methadione ^{3.66 1'4} 121 ¹⁴ 2 Nicotinamide (5 mM) + GAP 10+ Methadione ^{4.60 1'2 4 154 1'2'4 _} 3 Nicotinamide (10 mM) + GAP 10+ Methadione 2.90 ^{14 95 1'4}

Statistically Significant Improvement

1p <0.05 compared to Vehicle Control + Methadione;

² p < 0.05 compared to GAP10 + Methadione;

³ p < 0.05 compared with Nicotinamide riboside (at an equivalent dose) + Methadione;

4p < 0.05 compared with Nicotinamide (at an equivalent dose) + Methadione;

⁵ p < 0.05 compared with Picolinamide (equivalent dose) + Methadione.

Table 1B - Reactive oxygen species and cell viability

Sample Production AFK (% of % protection Cell viability (% of % protection control sample) control sample) BUT Carrier control 100 100 AT Methadione + Carrier Control 280.60 80.50 Test samples With Methadione + GAP 190.80 ³ 32.00 ³ 98.70 ³ 22.52 ³ D Methadione + Nicotinamide (10 mM) 231.00 ³ 17.70 ³ 92.90 ³ 15.41 ³ one Methadione + OAR + Nicotinamide (10 mM) 137.92 ^{1 2 3} 60.90 ^{1 2} ' ³ 134.30 ^{123 66.79 1.2'3}

Statistically significant improvement:

¹ p<0.05 compared to GAP;

² p < 0.05 compared to nicotinamide;

³ p < 0.05 compared to Vehicle Control + Methadione.

- 14 039773

In tables 1A and 1B, the samples marked with alphabetical letters were outside the scope of the invention. The samples marked with numbers were within the scope of the invention. Sample Q in Table 1A was a positive control because it used a high concentration of GAP. Overall, the results in Tables 1A and 1B show that the combination of nicotinamide with GAP amino acids provided a synergistic increase in GSH synthesis and a synergistic decrease in reactive oxygen species compared to results obtained with either nicotinamide or GAP alone.

Table 1C. Synthesis of glutathione

Sample Average value (µM glutathione) % Defense (Upgrade vs. Carrier Control + Methadione) BUT Carrier Control 2.73 AT Carrier Control + Methadione 0.18 ¹ 0 With DEC (40 µM) + Methadione ^{0.54 1'2 13 1'2} D DEC (80 µM) + Methadione 2.72 ^{1 2 93 1'2}

Statistically significant decrease:

¹ p < 0.05 compared to vehicle control.

Statistically significant improvement:

² p < 0.05 compared to vehicle control + methadion.

Table 1D. Reducing the amount of reactive oxygen species

Sample ROS inhibition (% of vehicle control) % Protection (improved against "Vehicle Control + Methadion") Cell viability (% of vehicle control) % Protection (improvement vs vehicle control + methadion) Carrier control 100 100 Carrier Control + Methadione 375.5 ¹ 73.7 ¹ DEC (20 µM) + Methadione 232.8 ¹² 38.0 ¹² 109.8 ² 48.9 ² DEC (40 µM) + Methadione 211.9 ¹² 43.6 ¹² 127.3 ^{1 2} 72.7 ¹² DEC (80 µM) + Methadione 233.8 ¹² 37.7 ^{12 131.5 1'2} 78.3 ¹² DEC (160 µM) + Methadione 253.8 ¹² 32.4 ^{12 122.3 1'2 65.9 1'2}

Statistically significant difference:

1p < 0.05 compared to vehicle control.

Statistically significant difference:

² p < 0.05 compared to Vehicle Control + Methadione.

Table 1E. Synthesis of glutathione

Sample Average value (µM glutathione) % Protection (improvement against "Carrier control + Methadione") BUT Carrier Control 2.73 AT Control in the form carrier + Methadione 0.18 ¹ 0 With DTBC (40 µM) + Methadione 0.17 ¹ About ¹ D DTBC (80 µM) + Methadione 0.19 ¹ 1 ¹

Statistically significant decrease:

1p < 0.05 compared to vehicle control

Statistically significant improvement:

² p < 0.05 compared to Vehicle Control + Methadione

- 15 039773

Table 1F. Reducing the content of oxygen forms. Statistically significant difference

Sample ROS inhibition (% of vehicle control) % Defense (Upgrade vs. "Vehicle Control + Methadion") Cell viability (% of vehicle control) % Defense (Upgrade vs. "Vehicle Control + Methadion") Carrier Control 100 100 Vehicle Control* Methadione 375.5 ¹ 73.7 ¹ DTBC (20 µM) + Methadione 367.0 ¹ 2.2 78.1 ¹ 6.0 DTBC (40 μM) + Methadione 323.7 ^{1 2 13.8 1'2} 75.9 ¹ 3.0 DTBC (80 µM) + Methadione 268.7 ^{1 2} 28.4 ^{1 2} 73.6 ¹ -0.1 DTBC (160 µM) + Methadione 221,G ² 41, T' ² 104.2 ² 41.3 ²

'p < 0.05 compared to vehicle control.

Statistically significant difference: 2 - G/G NTL I TV D' and p < 0.05 compared to Vehicle Control + Methadione.

Table 1G. Synthesis of glutathione

Sample Average value (µM glutathione) % Protection (improved against "Carrier Control + Metadion") BUT Carrier control 2.96 AT Carrier Control* methadion 0.18 ¹ 0 With CDME (80 μM) + Methadione 1.09 ^{12 31 1'2}

Statistically significant decrease:

1p < 0.05 compared to vehicle control.

Statistically significant improvement:

² p < 0.05 compared to vehicle control + methadion.

Table 1H. Recovery of oxygen compounds

Sample ROS inhibition (% of vehicle control) % Protection (improvement relative to "vehicle + methadion control") Cell viability (% of vehicle control) % Protection (improvement relative to "vehicle + methadion control") Carrier Control 100 100 Vehicle Control* Methadione 131.8 ¹ 69.8 ¹ CDME (20 µM) + Methadione 107.8 ² 18.2 ^{2 90.9 1'2 30.2 1'2} CDME (40 µM) + Methadione ^{111.3 1'2 15.5 1'2} 93.1 ^{12 33.4 1'2} CDME (80 µM) + Methadione 114.3 ^{1 2 13.3 1'2} 87.0 ^{1 2} 24.6 ^1.2 CDME (160 µM) + Methadione 97.9 ² 25.7 ^{2 105.9 1'2} 51.7 ¹²

Statistically significant difference:

1p < 0.05 compared to vehicle control.

Statistically significant difference:

p < 0.05 compared to vehicle control + methadione.

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Example 2

In Example 2, various combinations of nicotinamide and GSH building block amino acids were evaluated to stimulate intracellular GSH. The results obtained are summarized in Table 2A.

Table 2A. Synthesis of glutathione

Sample Mean % Protection BUT Carrier Control 1.75 AT Carrier Control + Methadione 0.07 ¹ About ¹ Samples tested With GAP 10 + Methadione ^{1.34 1'2 72 1'2} D GAP 20 + Methadione 4.61 ^2.3 259 ^{2 3} E Nicotinamide (5µM) + Methadione 0.13 ¹ 3 ¹

F Nicotinamide (5 μM) + NaPCA + cystine + Methadione 0.77 ¹² 4012 G Nicotinamide (5μM) + glycine + cystine + Methadione ^{0.94 1'2} ₄₉ 1, ² H Nicotinamide (5µM) + cystine + Methadione ^{0.44 1'2 211.2} one Nicotinamide (5µM) + GAP 10 ^aPCA+glycine+cystine')+Methadione 1.66 ^{23 91 2'3}

Statistically significant decrease:

1p < 0.05 compared to Nicotinamide + GAP10 + Methadione.

Statistically significant improvement:

² p < 0.05 compared with Nicotinamide + Methadione; 3 p < 0.05 compared to GAP10 + Methadione.

Samples A to H were outside the scope of the invention. Sample D was a positive control because it used a high concentration of GAP. It can be seen from the results that only the combination of nicotinamide with all 3 amino acids (sample 1 of the present invention) provided a synergistic increase in GSH synthesis compared to GSH synthesis achieved with either nicotinamide or GAP alone. Sample 1 performed on par with the positive control (sample D), but at a lower concentration of GAP and therefore a lower concentration of cystine, which counterbalances the problem of having to dissolve relatively high amounts of cystine.

Example 3

The skin care compositions of the present invention are illustrated in the tables below. All numbers in the tables represent the weight percentages in the composition.

Table I. Oil-in-water formulations, lotions and creams

OW-1 OW-2 OW-3 OW-4 OW-5 Water Up to 100 Up to 100 Up to 100 Up to 100 Up to 100 Glycerol 0-40 1-40 1-5 1-10 1-40 propylene glycol 0-5 0-5 Butylene glycol 0-5 0-5 0-5 Carbomer 0-2 0.03-1 Acryloyldimethyl ammonium taurate/VP copolymer 0-1 0.03-1 0.01-1

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Styrene/acrylate copolymer 0-1 0.01-1 xanthan gum 0-1 0.01-1 EDTA 0.01- 0.01 0.01- 0.01 0.01-1 0.01-1 0.01-1 preservative 0.02-2 0.02-2 0.02-2 0.02-2 0.02-2 titanium oxide 0-10 0.01-10 0.01-10 0.01-10 0.01-10 Dye/Pigment 0-5 0-5 0-5 0-5 0-5 Triethanolamine/sodium hydroxide/potassium hydroxide 0-3 0.01-3 0.01-3 0.01-3 0.01-3 Stearic acid 0-5 0.01-5 0.01-5 0.01-5 0.01-5 Isopropyl myristate 0-10 0.01-10 Capric/Caprylic Triglyceride 0-10 0.01-10 C12-C15 alkyl benzoate 0-10 0.01-10 Mineral oil 0-10 0.01-10 Glyceryl stearate 0-5 0.01-5 Stearet-2 0-5 0.01-5 0.01-5 Stearet-21 0-5 0.01-5 PEG 100 stearate 0-5 0.01-2 0.01-5 Potassium cetyl phosphate 0-5 0.01-2 Twin20 0-5 0.01-5 cetyl alcohol 0-4 0.01-4 0.01-4 Dicaprylyl carbonate 0-5 0.01-5 Ethylhexylmethoxycinnamate 0-6 0.01-6 Butyl- methoxydibenzoylmethane 0-3 0.01-3 0.01-3 0.01-3 Ensulizol 0-4 0.01-4 Octinoxate 0-7.5 Octyl salicylate 0-5 0.01-5 0.01-5 Octocrylene 0-10 0.01-10 0.01-10 Homosalicylate 0-10 0.01-10 Dimethicone 0-10 0.01-10 0.01-10 Cyclomethicone 0-15 0.01-15 flavoring 0-2 0-2 0-2 0-2 0-2 Glutamine / NaPCA 0.01-10 0.01-10 0.01-10 0.01-10 0.01-10 Glycine 0.01-10 0.01-10 0.01-10 0.01-10 0.01-10 cystine 0.001-2 0.001-2 0.001-2 0.001-2 0.001-2 Nicotinamide 0.01-5 0.01-5 0.01-5 0.01-5 0.01-5

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Table II. Lotions and creams for topical water-in-oil applications

WO-1 WO-2 WO-3 WO-4 Water up to 100 up to 100 up to 100 up to 100 Glycerol 0-70 1-70 1-70 propylene glycol 0-5 0.01-5 Butylene glycol 0-5 0.01-5 0.01-5 Hectoritis dysteardimonia 0.01-1 0.01-1 EDTA 0.01-.01 0.01-1 0.01-1 0.01-1 preservative 0.02-2 0.02-2 0.02-2 0.02-2 TiO2 0-10 0.01-10 0.01-10 0.01-10 Dye/pigment 0-5 0-5 0-5 0-5 TEA/sodium hydroxide/potassium hydroxide 0-3 0.01-3 0.01-3 0.01-3 Stearic acid 0-5 0.01-5 Isopropyl myristate 0-10 Capric/Caprylic triglyceride 0-10 0.01-10 C12-C15 alkyl benzoate 0-10 0.01-10 Mineral oil 0-10 Glyceryl stearate 0-5 Dimethicone copolyol 0-5 0.01-5 0.01-5 Cetyl-PEG/PPG-IO/l Dimethicone 0-5 0.01-5 Stearet-2 0-2 Sucrose distearate 0-2 0.01-2 cetyl alcohol 0-2 0.01-2 0.01-2 Ethylhexylmethoxycinnamate 0-6 0.01-6 Butylmethoxydibenzoylmethane 0-3 0.01-3 0.01-3 0.01-3 Ensulizol 0-4 0.01-4 Octinoxate 0-7.5 Octyl salicylate 0-5 0.01-5 0.01-5 Octocrylene 0-10 0.01-10 0.01-10 Homosalicylate 0-10 0.01-10 Dimethicone 0-10 0.01-10 0.01-10 Cyclomethicone 0-40 0.01-40 0.01-10 caprili methicone 0-10 0.01-10 0.01-10 Dimethicone cross-polymer 0-90 0.01-90 0.01-90 C30-C45 alkyl cetearyl dimethicone cross polymer 0.01-90 Glycolic acid 0-10 0.01-10 KCI 0-5 0.01-5 0.01-5 0.01-5 flavoring 0-2 0-2 0-2 0-2 Glutamine / NaPCA 0.01-10 0.01-10 0.01-10 0.01-10 Glycine 0.01-10 0.01-10 0.01-10 0.01-10 cystine 0.001-2 0.001-2 0.001-2 0.001-2 Nicotinamide 0.01-5 0.01-5 0.01-5 0.01-5

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Table III. Fast absorbing creams

VC-1 VC-2 VC-3 VC-4 Water Up to 100 Up to 100 Up to 100 Up to 100 Glycerol 0-5 0.01-5 0.01-5 EDTA 0.01-0.1 0.01-0.1 0.01-0.1 0.01-0.1 preservative 0.02-2 0.02-2 0.02-2 0.02-2 TU2 0.01-10 0.01-10 0.01-10 0.01-10 Dye/Pigment 0-5 0.01-5 0.01-5 TEA/Sodium Hydroxide/Potassium Hydroxide 0-3 0.01-3 0.01-3 0.01-3 Stearic acid 0-30 0.01-30 0.01-30 0.01-30 Isopropyl myristate 0-5 0.01-10 0.01-10 C12-C15 alkyl benzoate 0-5 0.01-10 Bridge 35 0-5 0.01-5 Twin40 0-5 0.01-5 cetyl alcohol 0-2 0.01-2 0.01-2 Ethylhexylmethoxycinnamate 0-6 0.01-6 0.01-6 Butylmethoxydibenzoylmethane 0-3 0.01-3 0.01-3 0.01-3 Ensulizol 0-4 0.01-4 Octyl salicylate 0-5 0.01-5 Octocrylene 0-10 0.01-10 0.01-10 Dimethicone 0-5 0.01-5 Cyclomethicone 0-5 0.01-5 Dimethicone cross-polymer 0-4 0.01-4 Hydroxy stearic acid 0-5 0.01-5 0.01-5 0.01-5 flavoring 0-2 0-2 0-2 0-2 Glutamine / NaPCA 0.01-10 0.01-10 0.01-10 0.01-10 Glycine 0.01-10 0.01-10 0.01-10 0.01-10 cystine 0.001-2 0.001-2 0.001-2 0.001-2 Nicotinamide 0.01-5 0.01-5 0.01-5 0.01-5

CLAIM

Claims (13)

CLAIM 1. Composition for skin care, for local use, containing a) a glutathione precursor comprising, by weight of the composition: i) from 0.001 to 2% by weight of cystine or a salt or ester thereof; i) from 0.01 to 5% by weight of a source of glutamate; iii) 0.01 to 5% by weight of glycine; and iv) 0.01 to 5% by weight of nicotinamide; and b) a cosmetically acceptable carrier in the form of an emulsion of water and oil; while the pH of the composition is in the range from 3.5 to 8.5; and wherein the mass ratio of nicotinamide to total amino acids is from 20:1 to 50:1. 2. Composition according to claim 1 further containing nicotinamide riboside. 3. Composition according to claim 1 or 2, characterized in that said source of glutamate is selected from the group consisting of glutamine, glutamic acid, pyroglutamic acid and salts or mixtures thereof. 4. Composition according to claim 3, characterized in that said source of glutamate is pyroglutamic acid or its sodium salt. 5. Composition according to claim 1, characterized in that the cystine ester is selected from the group consisting of diethyl-E-cystinate dihydrochloride, L-cystine dimethyl ether dihydrochloride, di-tert-butyl L-cystinate dihydrochloride and mixtures thereof. 6. Composition according to claim 5, characterized in that the cystine ester is diethyl-Ecystinate dihydrochloride. 7. Composition according to any one of claims 1 to 6, characterized in that said composition is in the form of a water-in-oil emulsion containing cystine in the aqueous phase, where 90% of the water droplets have a diameter in the range from 100 nm to 20 µm. 8. The composition according to any one of claims 1 to 7, which is a non-rinsing non-solid cosmetic composition for the skin. 9. Composition according to any one of claims 1 to 8, which is a fast-absorbing cream. 10. Composition according to any one of claims 1 to 9, additionally containing 4-alkylresorcinol. 11. Composition according to any one of claims 1 to 10, additionally containing 12-hydroxystearic acid. 12. A method for improving the appearance of the skin, comprising applying to the skin a composition according to any one of claims 1-11. 13. A method of providing even skin color and reducing pigmentation, age spots and discoloration of the skin, including applying to the skin a composition according to any one of claims 1 to 11.