FR2915378A1 - Use of an active ingredient obtained from hydrolyzed soy, in a cosmetic composition or for preparing a pharmaceutical composition comprising the ingredient or composition to activate aconitase and protect mitochondrion - Google Patents

Abstract

Use of an effective quantity of at least an active ingredient (I) obtained from hydrolyzed soy (Glycine max L.), in a cosmetic composition or for preparing a pharmaceutical composition comprising (I) or composition to activate aconitase and protect mitochondrion, is claimed. ACTIVITY : Dermatological; Muscular-Gen; Neuroprotective; Cardiant; Antidiabetic; Nootropic. MECHANISM OF ACTION : Aconitase activator. The ability of (I) to activate the acotinase enzyme was tested in human fibroblasts. The result showed that (I) exhibited increased activity of 55%.

Description

The present invention is in the field of cosmetics and

  pharmaceutical, and more particularly in the field of dermatology. The present invention relates to the use in a cosmetic composition or for the preparation of a pharmaceutical composition, of an effective amount of active ingredient derived from soy (Glycine max L.); the active ingredient, or the composition containing it, being intended to protect mitochondria and to activate aconitase. Preferably, the active ingredient is derived from the hydrolysis of soy protein and contains mainly polypeptides and peptides. It can be used alone or in combination with at least one other active ingredient. The invention also relates to a cosmetic treatment method intended to protect the skin and the integuments from external aggressions and to fight against skin aging. Said active ingredient, mitochondrial protector and activator of aconitase, can also be used to prepare pharmaceutical compositions intended to prevent or fight against pathologies related to mitochondrial dysfunctions; for example certain neuromuscular or cardiac degenerations, type II diabetes or certain pathologies of aging. The term dander according to the invention encompasses all the keratinous appendages present on the surface of the body, in particular the hairs, the eyelashes, the eyebrows, the nails and the hair. Mitochondria are the organelles of eukaryotic cells specialized in the production of energy. They have their own DNA (mtDNA). The energy produced by the respiratory chain is stored as ATP. Under certain cellular stress conditions, electrons, released by the oxidative phosphorylation respiratory chain, produce free radicals that can damage mitochondrial structures. These stressors can be intrinsic or external. Among these are: UV radiation, toxins, radiation, food oxidants. Thus, it has been possible to show a progressive decline in mitochondrial functions with age, probably related to the accumulation of mutations on mtDNA (Singh, K., N.Y. Acad Sci, 1019, 2004). These mutations in mtDNA can be induced by repeated UV radiation exposures, and would even be a control of photo-aging (M. Bernburg et al., J. of Invest Dermatol 122 (5), 2004). Mitochondria also play a central role in apoptosis. Mitochondrial degradations can thus be a triggering factor of the reaction cascade -2 leading to apoptosis in many cell types (G. Kroemer et al., FASEB J. 9: 1277-87, 1995). In mitochondria, natural mechanisms of repair have been identified. Aconitase (aconitate hydratase or Iron Regulatory Protein, EC 4.2.1.3) has thus been shown to be essential for maintaining the integrity of mtDNA, regardless of its catalytic activity. Aconitase was first identified as a mitochondrial enzyme involved in the Krebs cycle; it catalyzes the stereospecific conversion of citrate to isocitrate and thus participates in the energy production of the cell. It has an active site with an iron-sulfur (4Fe-4S) cluster with pseudocubic geometry. Recently, Dr. Butow's team (X.J. Chen et al., Science 307, 2005) has shown that aconitase plays a key role in preserving the integrity of mitochondrial DNA. It would interact with the DNA within regulatory regions suggesting a role in compaction of the mitochondrial genome in the manner of a protein of the HSP 70 family. The transition from the DNA binding form to the active form of the enzyme would be based on the assembly or disassembly of the iron-sulfur cluster. Due to its dual functionality, aconitase establishes a direct link between the mitochondrial energy-generating activity and the conservation of mtDNA. Aconitase could thus constitute a therapeutic target to prevent or fight against pathologies related to mitochondrial dysfunctions, for example, type II diabetes, certain neuromuscular disorders, certain pathologies of aging. The search for compounds that can stimulate or protect mitochondria is a concern of medical research and cosmetics. With regard to the skin, these new compounds could be useful to prevent or fight against the signs of skin aging, but also to protect or repair the skin after burns, exposure to UV radiation or radiation, or exposure to toxins or to pollutants. In order to protect the mitochondria and to fight against the associated pathologies or the effects of skin aging, solutions have been proposed such as the supply of substances involved in energy metabolism, and more particularly intermediates or cofactors of the skin cycle. Krebs such as fumarate, L-malate, acetyl CoA (WO 02064129) or the treatment of the skin with substances capable of reducing free radicals, such as vitamin C (US 2004/0086526) or L -ergothioneine (WO 9836748). But, to the knowledge of the applicant, the use of active principles derived from the hydrolysis of soybeans, capable of activating aconitase, in order to protect mitochondria and fight against associated pathologies and / or the effects of skin aging, has never been described. The main objective of the present invention is the use of an active ingredient derived from the hydrolysis of soybeans, capable of protecting the skin against external aggressions and of combating cutaneous aging. Said active ingredient may be used alone or in combination with at least one other active ingredient. The inventors have indeed demonstrated a therapeutic activity, and more particularly a dermatological and cosmetic activity, of an active ingredient of a peptide nature, derived from the hydrolysis of soybeans, capable of activating aconitase. In particular, it has been demonstrated that this active ingredient, when applied to the skin, significantly promotes the mitochondrial activity, demonstrated by an increased activity of the enzyme. The newly identified properties of this mitochondrial protective active ingredient thus open up new therapeutic and cosmetic perspectives.

  Protective active ingredient is understood to mean mitochondria or capable of protecting mitochondria, any substance of vegetable origin, and more particularly derived from soy (Glycine max L.), capable of limiting the functional or structural alterations of cell or tissue mitochondria. subject to physico-chemical or environmental stress. The term "active ingredient capable of activating aconitase" means any substance of plant origin, and more particularly derived from soy (Glycine max L.), capable of increasing cytoplasmic or mitochondrial aconitase, or by the activation of the protein synthesis of aconitase (by direct or indirect modulation of the gene expression of aconitase), either by increasing the enzymatic activity of aconitase or by other biological processes such as stabilization of the aconitase protein or the stabilization of the messenger RNA transcripts. Preferably, the active ingredient capable of activating the aconitase according to the invention is an extract of peptide nature and comes from the hydrolysis of soy proteins (Glycine max L.). By peptide nature is meant a mixture of compounds predominantly represented by peptides or polypeptides.

  The term peptide denotes a sequence of two or more amino acids linked together by peptide bonds or by modified peptide bonds; the term polypeptide designating a peptide of larger size. By biologically active expression is meant that which has an activity in vivo or in vitro characteristic of the activity of the active ingredient according to the invention. The term "hydrolyzate or derived from hydrolysis" refers to any substance or mixture of substances, or isolated preparation, obtained after hydrolysis of plant material. The active ingredient according to the invention can be obtained by extraction of proteins of vegetable origin, followed by controlled hydrolysis which liberates biologically active peptide fragments. Many proteins found in plants are likely to contain biologically active peptide fragments within their structure. The controlled hydrolysis makes it possible to release these peptide fragments. It is possible, but not necessary to carry out the invention, to extract either the proteins concerned first and then hydrolyze them, or to carry out the hydrolysis first on a crude extract and then to purify the peptide fragments. . It is also possible to use certain hydrolysed extracts without purifying the peptide fragments corresponding to the biologically active peptides according to the invention, but nevertheless ensuring the presence of said fragments by appropriate analytical means. To carry out the extraction, one can use the whole plant. or a specific part of the plant (leaf, grain, etc.). More particularly according to the invention, one of the many plants of the Fabaceae family, of the genus glycine, is used; soybean or soybean meal and preferably of the species Glycine max L. In a first step, the plant is milled using a plant mill. The powder thus obtained may subsequently be "delipidated" with the aid of a conventional organic solvent (such as, for example, an alcohol, hexane or acetone). The proteins of the plant are then extracted according to the conventional method (Osborne, 1924) modified; the plant mash is suspended in an alkaline solution containing an insoluble polyvinylpolypyrrolidone adsorbent material (PVPP) (0.01-20%); in fact it has been observed that subsequent hydrolysis and purification operations are facilitated by this means. The concentration of phenolic-type substances interacting with the proteins is thus reduced. The soluble fraction is collected after centrifugation and filtration steps, this crude solution then constituting a first form of the extract containing the proteins, carbohydrates and possibly lipids. The proteins are then precipitated by varying the ionic strength by acidifying the medium, which eliminates soluble components and nucleic acids. The precipitate is then washed with an organic solvent such as, for example, ethanol or methanol and the solvent is evaporated by drying in vacuo. The protein-rich precipitate is dissolved in water or other solvent and is then a more purified form of the hydrolyzate.

  The extraction can also be carried out in neutral or acidic medium always in the presence of polyvinylpolypyrrolidone. After a filtration step, the precipitation step is then carried out using a conventional precipitation agent such as salts (sodium chloride, ammonium sulfate) or an organic solvent (alcohol, acetone). The precipitate obtained can be separated from the precipitating agents by dialysis after redissolving in water or another solvent. The isolated protein fraction according to the invention is then hydrolyzed under mild conditions to generate polypeptides and soluble peptides. Hydrolysis is defined as a chemical reaction involving the cleavage of a molecule by water, this reaction being possible in a neutral, acidic or basic medium. According to the invention, the hydrolysis is carried out chemically and / or advantageously by proteolytic enzymes. We can then mention the use of endoproteases of plant origin (papain, bromelain, ficin) and microorganisms (Aspergillus, Rhizopus, Bacillus, etc.). For the same reasons as above during this hydrolysis step, a quantity of polyvinylpolypyrrolidone is added to the reaction medium. After filtration, the solution obtained constitutes the active hydrolyzate. The active hydrolyzate can be further purified in order to select the molecular weights and the nature of the peptides generated. The fractionation can advantageously be carried out by ultrafiltration and / or by a chromatographic type method. Any of the more or less purified forms of the hydrolyzate is then solubilized in water or in any mixture containing water, and then sterilized by ultrafiltration. The vegetable hydrolyzate obtained according to the invention is analyzed qualitatively and quantitatively for its physico-chemical characteristics and its content of protein and peptide compounds. Peptide-like compounds are understood to mean protein fragments, peptides and free amino acids present in the mixture. The peptides, amino acids and protein fragments are determined according to conventional techniques, which are well known to those skilled in the art. Thus, according to an advantageous embodiment of the invention, the active plant hydrolyzate has a pH of between 4 and 7, and preferably between 5 and 6. A solids content of between 1 and 8 g / l and preferably between 2 and 5 g / 1, its content of peptide compounds is between 0.1 and 5g / 1 and preferably between 0.5 and 2 g / 1 and its sugar content is 0.5 to 2.5 g / l.

  According to an advantageous embodiment of the invention, the active principle according to the invention is solubilized beforehand in one or more cosmetically or pharmaceutically acceptable solvents, conventionally used by those skilled in the art, such as water, glycerol, ethanol, propylene glycol, butylene glycol, dipropylene glycol, ethoxylated or propoxylated diglycols, cyclic polyols, petroleum jelly, a vegetable oil or any mixture of these solvents. According to yet another advantageous embodiment of the invention, the active principle according to the invention is solubilized beforehand in a cosmetic or pharmaceutical vector such as liposomes or adsorbed on powdery organic polymers, mineral supports such as talcs and bentonites, and more generally solubilized in, or attached to, any cosmetically or pharmaceutically acceptable vector. It is understood that the active ingredient according to the invention can be used alone or in combination with at least one other active ingredient, in a cosmetic composition or for the preparation of a pharmaceutical and / or dermatological composition. The compositions according to the invention may be applied by any appropriate route, in particular oral, parenteral or external topical, and their formulation will be adapted by those skilled in the art, in particular for cosmetic or dermatological compositions. Advantageously, the compositions according to the invention are intended for topical administration to the skin. These compositions must therefore contain a cosmetically and / or dermatologically acceptable medium, that is to say compatible with the skin and superficial body growths, and cover all cosmetic or dermatological forms. These compositions may especially be in the form of creams, oil-in-water or water-in-oil emulsions, multiple emulsions, solutions, suspensions, gels, milks, lotions, sticks or else powders, suitable for application on the skin, lips and / or integuments. These compositions comprise the excipients necessary for their formulation, such as solvents, thickeners, diluents, surfactants, antioxidants, dyes, preservatives, perfumes. Of course, those skilled in the art will take care to choose any additional compounds, active or non-active, and / or their quantity, so that the advantageous properties of the mixture are not impaired by the addition envisaged. The composition that may be used according to the invention may in particular consist of a composition for hair care, and in particular a shampoo, a conditioner, a setting lotion, a treatment lotion, a cream or a styling gel, a restructuring lotion for hair, a mask, etc. The cosmetic composition according to the invention can be used in particular in treatments using an application that is followed or not followed by rinsing, or in the form of shampoo. It can also be in the form of dye or mascara to be applied with a brush or a comb, in particular on eyelashes, eyebrows or hair.

  Advantageously, the compositions that can be used according to the invention also contain various active principles intended, in particular, for the prevention and / or treatment of disorders associated with photoaging. Thus, the composition according to the invention can associate, with the active ingredient capable of activating the aconitase according to the invention, other active ingredients promoting its action. For example, it may be added active ingredients having an anti-radical or antioxidant action, selected from vitamin C, vitamin E or coenzyme Q10 or polyphenol extracts of plants. By anti-radical active ingredients is meant any compound capable of trapping free radicals. These active principles are capable of blocking free radical chain reactions before the ultimate stages of degradation of the biological constituents of the skin and thus have an antioxidant activity. The composition according to the invention may also associate, with the active ingredient activator of aconitase, active principles stimulating the synthesis of dermal macromolecules (laminin, fibronectin, collagen), for example the collagen peptide sold under the name Collaxyl by the Vincience company.

  The composition according to the invention may also associate, with the active ingredient activator of aconitase, active ingredients stimulating energy metabolism, such as the active ingredient marketed under the name GP4G by Vincience. In another aspect, the composition according to the invention may be a solar composition, that is to say a composition that helps to protect against solar radiation. Thus, it can be advantageously added, to the composition according to the invention, active assisting solar protection such as, for example, sunscreens.

  It is obvious that the invention is directed to mammals in general, and more particularly to humans. The effective amount of active ingredient according to the invention corresponds to the amount necessary to obtain the desired result, namely: activate aconitase, protect mitochondria and thus protect the skin from external aggression and fight against skin aging. According to an advantageous embodiment of the invention, the active ingredient derived from soy is present in the compositions of the invention at a concentration of from 0.0001% to about 20%, and preferably at a concentration of between 0.05%. and about 5% based on the total weight of the final composition.

  These compositions may especially be in the form of an aqueous solution, hydroalcoholic or oily; an oil-in-water, water-in-oil emulsion or multiple emulsions; they may also be in the form of creams, suspensions or powders, suitable for application to the skin, mucous membranes, lips and / or integuments. These compositions may be more or less fluid and have the appearance of a cream, lotion, milk, serum, ointment, gel, paste or paste. a foam. They can also be in solid form, as a stick, or be applied to the skin in the form of an aerosol. They can be used as a care product and / or as a make-up product for the skin. These compositions additionally comprise any additive commonly used in the field of application envisaged as well as the adjuvants necessary for their formulation, such as solvents, thickeners, diluents, antioxidants, dyes, sunscreens, self-tanning principles, pigments, fillers, preservatives, perfumes, odor absorbers, cosmetic or pharmaceutical active ingredients, essential oils, vitamins, essential fatty acids, surfactants, film-forming polymers, etc. . In all cases, those skilled in the art will ensure that these adjuvants and their proportions are chosen in such a way as not to harm the desirable advantageous properties of the composition according to the invention. These adjuvants may, for example, correspond to 0.01 to 20% of the total weight of the composition. When the composition of the invention is an emulsion, the fatty phase may represent from 5 to 80% by weight and preferably from 5 to 50% by weight relative to the total weight of the composition. The emulsifiers and co-emulsifiers used in the composition will be chosen from those conventionally used in the field under consideration. For example, they can be used in a proportion ranging from 0.3 to 30% by weight, relative to the total weight of the composition. By its particular activities, the active ingredient according to the invention can be used advantageously in a cosmetic composition or for the preparation of a pharmaceutical composition.

  In particular, the active ingredient according to the invention may advantageously be used in a cosmetic composition intended to fight in a preventive and / or curative manner against the manifestations of skin aging and, more specifically, in order to fight against and / or prevent aging. photo-induced (photo-aging). Skin manifestations of aging means any changes in the external appearance of the skin due to aging such as, for example, wrinkles and fine lines, wilted skin, soft skin, thinned skin, lack of elasticity and / or or skin tone, dull and lackluster skin or skin pigmentation spots, but also any internal changes in the skin that do not systematically result in a modified external appearance such as, for example, any internal degradation of the skin. skin following exposure to ultraviolet (UV) radiation. The active ingredient according to the invention, or the composition containing it, will make it possible, in particular, to combat the loss of elasticity and firmness of the skin. The active ingredient according to the invention makes it possible to protect the skin and integuments against all types of external aggressions. The use of the active ingredient, or a composition containing it, will allow the skin and integuments to be protected and to better withstand environmental stresses. By the term external aggression, we mean the aggressions that the environment can produce. By way of example, mention may be made of aggressions such as pollution, UV, or irritating products such as surfactants, preservatives or perfumes. Pollution is understood to mean both external pollution, eg due to diesel particles, ozone or heavy metals, as well as internal pollution which may be due to solvent emissions from paints, adhesives or paper. (such as toluene, styrene, xylene or benzaldehyde), or even cigarette smoke. The active ingredient according to the invention can be advantageously used in a cosmetic composition or for the preparation of a pharmaceutical composition, as a photo-protective active ingredient and, more particularly, as a photoprotective active ingredient said Secondary. In fact, the primary photoprotective active principles are distinguished from the secondary photoprotective active ingredients. Primary photoprotective active ingredients are substances that exercise physical power: they are able to absorb UV radiation and return it as heat to protect the skin. Secondary photoprotective active ingredients are substances that usually have a biological effect; these are, for example, the active ingredients capable of limiting damage to DNA and membranes by the penetration of UV radiation into the skin. The subject of the invention is also the use in a cosmetic composition, or for the preparation of a pharmaceutical composition, of an effective quantity of active principle as described above, the active ingredient, or the composition containing it, being intended to to prevent damage to the skin caused by exposure to the sun or exposure to ionizing radiation during radiation therapy. The subject of the invention is also the use in a cosmetic composition, or for the preparation of a pharmaceutical composition, of an effective quantity of active principle as described above, the active ingredient, or the composition containing it, being intended to to protect mitochondria, especially on areas of the body exposed to UV radiation. The invention also relates to the use in a cosmetic composition, or for the preparation of a pharmaceutical composition, of an effective amount of active ingredient as described above, the active ingredient, or the composition containing it, being intended to to protect the skin from damage caused by free radicals. The invention also consists in a pharmaceutical composition characterized in that the active principle according to the invention is formulated to attenuate a pathology related to mitochondrial dysfunctions, for example certain neuromuscular or cardiac degenerations, type II diabetes, certain pathologies of the aging.

  Particular embodiments of this cosmetic treatment method also result from the foregoing description. Other advantages and characteristics of the invention will appear better on reading the examples given for illustrative and not limiting. -11-Example 1: Preparation of active ingredient from sofa cake (Glycine max L.) Cakes are the solid residues obtained after oil extraction from soybeans. They represent 50 to 75% of the seed mass. The active principle is obtained from a plant extract of the species Glycine max L. Of course the extract can be prepared from plants of at least one of the many varieties and species belonging to the genus Glycine . In a first step, 1 kg of soybean meal is milled in a cereal mill the flour obtained is delipidated by the action of an organic solvent, hexane. After filtration and drying under vacuum the powder obtained is suspended in an aqueous alkaline solution (1/10 dilution) pH 10, containing 1% of polyvinylpolypyrrolidone (Polyclar V ISP). This mixture is stirred for a time long enough to allow the solubilization of the soluble fractions. The extraction temperature is variable (between 4 and 80 C); preferably the operation will be performed cold. After this extraction phase, the medium is clarified by centrifugation and then filtered through a plate filter. This filtrate, which contains the soluble fractions of soybeans, is then subjected to precipitation of the proteins by varying the ionic strength in neutral or acidic medium, which makes it possible to eliminate the soluble carbohydrate components, the lipids and the nucleic acids. pH 3.5. The supernatant is removed and the precipitate is then washed with a solvent such as, for example, ethanol or methanol and the solvent is evaporated by drying in vacuo.

  At this stage, about 50 grams of light yellow powder of crude protein extract are obtained containing: Proteins: 75% Carbohydrates: 20% Lipids: 5% The precipitate rich in proteins is dissolved in water or other solvent. The crude protein extract is then subjected to a series of gentle and selective hydrolyses consisting of chemical and enzymatic hydrolyses in the presence of 0.5% PVPP (Polyclar V) and cysteine endopeptidases (papain, ficin). After reaction, the hydrolyzate is filtered on a plate and then on a sterilizing cartridge (0.21am). A light-colored hydrolyzate titrant of 15 to 30 g / l of dry extract is then obtained, which is then diluted so that the concentration of peptides determined by the method of Lowry is between 0.1 and 5 g. / 1 and preferably between 0.5 and 2 -12 g / l. The physicochemical analysis of the vegetable hydrolyzate, which constitutes the active ingredient, shows that its pH is between 4 and 7, and preferably between 5 and 6, the dry extract is between 1 to 8 g / l and preferred way between 2 and 5 g / 1, its content of peptide compounds is between 0.1 and 5 g / 1 and preferably between 0.5 to 2 g / 1 and its sugar content between 0.5 to 2, 5 g / 1.

  Example 2 Preparation of Active Ingredient from Soybean (Glycine max L.) A variant of the protocol of Example 1 consists of carrying out the same sequence of controlled and selective enzymatic hydrolyses but in the presence of 0.5% of PVPP. A light-colored hydrolyzate of 15 to 30 g / l of dry extract is then obtained after sterilizing filtration. The solution is then ultrafiltered on a Millipore Helicon filtration cartridge (cut off 1 kDa). The high molecular weights contained in the retentate are removed and the filtrate is preserved. The concentration of compounds of peptidic nature is determined by the method of Lowry, between 0.1 and 5 g / l and preferably between 0.5 and 2 g / l. The physicochemical analysis of the vegetable hydrolyzate, which constitutes the active ingredient, shows that its pH is between 4 and 7, and preferably between 5 and 6, the dry extract is between 1 to 8 g / l and preferred way between 2 and 5 g / l, its content of peptide compounds is between 0.1 and 5g / 1 and preferably between 0.5 to 2 g / 1 and its sugar content between 0.5 to 2, 5 g / 1.

  Another variant consists in carrying out a purification of the active principle, obtained according to Example 1 or 2, by ion exchange chromatography, on a TSK gel (TosoHaas) column with a Ph7 phosphate buffer.

  EXAMPLE 3 Demonstration of the Activating Effect of the Active Principle According to Example 1 on the Enzymatic Activity of Aconitase The aim of this study is to determine the influence of the active ingredient according to Example 1 on the enzymatic activity of aconitase. For this, the total enzymatic activity (cytoplasmic and mitochondrial) and the mitochondrial enzymatic activity of aconitase were measured. Protocol: Normal human fibroblasts are treated with a 1% solution, of the active ingredient according to Example 1, for 48 hours, then subjected to irradiation with UVB at 100 mJ / cm 2. After irradiation, the cells are treated under the same conditions for a further 24 hours. Negative controls are carried out under the same conditions but in the absence of active principle according to Example 1. To evaluate the enzymatic activity of aconitase, the fibroblasts are then harvested and centrifuged in PBS buffer. Part of the cells is then treated to perform the assay of the total enzyme activity, another part is used to isolate the mitochondrial fraction according to the following protocol: The cells are lysed using a Dounce type homogenizer and the homogenate is centrifuged cold at 10,000 g for 10 minutes. The pellet, rich in mitochondria, is washed with TES buffer (10 mM Tris, 1 mM EGTA, 0.25 M sucrose) and resuspended in 0.2 mM trisodium citrate buffer before being used for enzymatic assay. The determination of the enzymatic activity of aconitase is carried out using the OXIS Bioxytech Aconitase-340 assay Kit (Oxys International), according to the following principle: the aconitase causes the isomerization of citrate to isocitrate. The isocitrate is then converted to alphaketoglutarate by the enzyme isocitrate dehydrogenase. this second reaction is accompanied by the transformation of NADP + into NADPH. It is the formation of NADPH which is quantified by spectrophotometry at 340 nm. Under the chosen experimental conditions, the rate of formation of NADPH is proportional to the amount of aconitase. Analysis of the appearance kinetics of NADPH makes it possible to calculate the aconitase concentration of the sample (in mU / ml). Results: Assays for the total enzymatic activity of aconitase show an increase of more than 55% in activity in cells treated with the active ingredient according to Example 1 compared to untreated control cells. When the cells are irradiated with UVB, on the contrary, a decrease of more than 30% of this activity is observed. The treatment, before and after the UVB irradiation, with the active principle according to Example 1, makes it possible to limit the decrease observed in the untreated control cells. Assays of enzymatic activity of mitochondrial aconitase show comparable results. Conclusions: The active ingredient according to Example 1 strongly stimulates the enzymatic activity of cytoplasmic and mitochondrial aconitases of cutaneous cells. The action of the active ingredient according to Example 1 protects cells and mitochondria against UVB stress. EXAMPLE 4 Evaluation of the Protective Effect of the Active Principle According to Example 1 on Cutaneous Cells Subject to Ultraviolet Radiation (UVB) The aim of this study is to determine the protective effect of the active ingredient according to US Pat. Example 1 vis-à-vis dermal fibroblasts subjected to stress by UVB radiation. For this, cell viability tests were performed by the MTT technique. Protocol: Human dermal fibroblasts are treated with a solution containing 1% of the active principle according to Example 1, for 24 hours, irradiated with UVB (from 25 to 100 mJ / cm 2) and then cultured for a further 24 hours in the presence of the same concentration of active principle according to Example 1. Untreated and non-irradiated controls and untreated but irradiated controls are carried out under the same conditions. At the end of the experiment, the cells are incubated in a solution containing 0.1 mg / ml of MTT (344.5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide). This compound is absorbed by living cells and then metabolized by the mitochondrial enzymes into a blue violet compound, formazan, which will be assayed spectrophotometrically at 540 nm. The optical density (OD) is then directly proportional to the mitochondrial enzymatic activity as well as to the number of living cells. Results: The evaluation of the cell viability by the MTT technique shows, on the one hand, that the active ingredient according to Example 1, independently of any UVB radiation, increases the cell viability and, on the other hand, that the irradiations by UVB have had a dose-dependent cytotoxic effect on human fibroblasts, but this cytotoxic effect is not manifested in the presence of 1% of active ingredient according to Example 1. Conclusions: The active ingredient according to Example 1 increases cell viability and on the other hand, effectively protects the skin cells against the cytotoxic effects of UVB radiation.

  EXAMPLE 5 Evaluation of the Protective Effect of the Active Principle According to Example 1 on Cutaneous Cells Under Oxidative Stress The aim of this study is to determine the protective effect of the active ingredient according to Example 1 with respect to dermal fibroblast screw subjected to oxidative stress caused by hydrogen peroxide (H2O2) at 4 mM or 5 mM. For this, cell viability tests were performed by the MTT technique. Protocol: The human dermal fibroblasts are treated with a 1% solution of the active principle according to Example 1, for 24 hours, subjected to oxidative stress caused by 4 mM or 5 mM H2O2 for 30 minutes. minutes and then cultivated another 24 hours in the presence of the same concentration of active principle according to Example 1. Controls not treated with the active ingredient or with H2O2 and treatment controls with the active principle alone are made in the same conditions. At the end of the experiment, the cells are incubated in a solution containing 0.1 mg / ml of MTT (344.5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide), according to the protocol described in Example 4. Results: The evaluation of cell viability by the MTT technique shows, on the one hand, that the active ingredient according to Example 1, independently of oxidative stress, increases cell viability, and on the other hand, that oxidative stress caused by 4 mM or 5 mM H2O2 for 30 minutes had a dose-dependent cytotoxic effect on human fibroblasts, but that this cytotoxic effect does not occur in the presence of 1% of active ingredient according to Example 1.

  Conclusions: The active ingredient according to Example 1 increases cell viability and effectively protects the skin cells against the cytotoxic effects of oxidative stress.

  EXAMPLE 6 Evaluation of the cytoprotective effect of the active principle according to Example 1 on cutaneous cells subjected to ultraviolet radiation (UVB) A study of the cytoprotective effect of the active principle according to Example 1 was carried out by evaluating the damage caused at the DNA level. This study was carried out using the comet test (or Single Cell Gel Electrophoresis), a short and sensitive microelectrophoretic technique that allows the visualization and quantification of DNA breaks on individual cells. Protocol: Primary human fibroblasts are seeded in boxes of diameters 100. When the cells reached 70% confluence, they are treated with a 1% solution of active principle according to Example 1 for 24 hours. The cells are then subjected to UVB irradiation at 100 mJ / cm 2 and then cultured in the presence of the active ingredient for a further 24 hours. Boxes not treated with the active ingredient serve as control. A cell suspension at 100,000 cells / ml is then performed, 500 l are taken, included in an agarose solution and cast between slide and coverslip. The cells are lysed cold. The DNA is then denatured with an alkaline buffer and then subjected to short electrophoresis (250 mA for 30 minutes) and evidenced by propidium iodide staining. The slides are then observed under a fluorescence microscope. An altered cell sees its DNA stretch toward the anode in proportion to the number of breaks in the DNA and form a comet. The highly degraded DNA is in the tail of the comet. An intact cell remains round and its DNA is then compacted at the head of the comet. The evaluation of DNA breaks is carried out using an image analysis software which makes it possible to determine the percentage of degradation of the DNA, by the quantitative evaluation of the Tail Moment, a parameter that defines itself. as the product of the length of the comet by the percentage of DNA in its distal part. Results: The results obtained show that the cells subjected to UVB irradiation, in the absence of the active principle according to Example 1 (UVB control), have the most important Tail Moment, that is to say the condition where DNA is the most damaged. In contrast, the cells treated with the active ingredient according to Example 1, have a Tail Moment, that is to say, damage to the DNA, decreased by 25%. Conclusion: The active ingredient according to Example 1 plays an important role in protecting the DNA of cutaneous cells subjected to irradiation with UVB.

  EXAMPLE 7 Preparation of compositions 1 - Sun protection cream: Trade names INCI names% by mass PHASE A Demineralized water Aqua (Water) qsp Pemulen TRI Acrylates / C10-30 Alkyl Acrylal 0.40 Crosspolymer Glycerine Glycerin 3.00 Nipastat Sodium Sodium Methylparaben ( and) Sodium 0.15 Ethylparaben (and) Sodium Butyl paraben (and) Sodium Propylparaben (and) Sodium Isobutylparaben PHASE B Parsol MCX Ethylhexyl Methoxycinnamate 7.50 Eusolex 4360 Benzophenone-3 3.00 Parsol 1789 Butyl Methoxydibenzoylmethane 2.00 -17- Trade Names INCI Names% by mass Myritol 318 Caprylic / Capric Triglyceride 4.00 Emulgade SEV Hydrogenated Palm Glycerides (and) 5.00 Ceteareth-20 (and) Ceteareth-12 (and) Cetearyl Alcohol Propylparaben Propylparaben 0.15 Nacol 16-98 Cetyl Alcohol 1.00 PHASE C TEA Triethanolamine 0.20 PHASE D Active principle according to Example 1 3 Fragrance Perfume (Fragrance) qsp Colorant qsp The constituents of phase A and phase B are heated separately between 70 ° C. and 75 ° C. Phase B is emulsified in phase A with stirring. Phase C is added at 45 ° C., increasing stirring. Phase D is then added when the temperature is below 40 C. Cooling is continued to 25 C with vigorous stirring. 2 ûLait après-soleil: Trade names INCI names% by mass PHASE A Montanov LC 14-22 Alcohols (and) C 12-20 3.00 Alkyl Glucoside Waglinol 2559 Cetearyl Isononanoate 4.00 Tegosoft TN C12-15 Alkyl Benzoate 3.00 Oil of Apricot Kernels Prunus Armeniaca (Apricot) Kernel 2.00 Oil Avocado Oil Persea Gratissima (Avocado) Oil 1.00 Abil 350 Dimethicone 1.00 PHASE B Demineralized Water Aqua (Water) qsp -18- Trade Names INCI Names % Mass PHASE C Simulgel EG Sodium Acrylate / Acryloyldimethyl 0.4 Taurate Copolymer (and) Isohexadecane (and) Polysorbate 80 Copolymer (and) Polysorbate 80 PHASE D Phenonip Phenoxyethanol (and) 0.30 Methylparaben (and) Ethylparaben (and) Butylparaben (and) and) Propylparaben (and) Isobutylparaben Ethylparaben and Propylparaben and Buthylparaben Germall 115 Imidazolidinyl Urea 0.20 PHASE E Active principle according to Example 1 0.1 Prepare phase A with stirring. Incorporate the xanthan gum gradually, with deflocculating stirring. Phases C and D will be incorporated once the gel is complete. Phase E, prepared before perfect dissolution of the DHA, will be added later.

  Adjust the pH if necessary to 4 - 4.5. Color and perfume.

  3 - Anti-Aging Cream: Trade Names INCI Names% Massic Phase A Montanov 68 Cetearyl Alcohol (and) Cetearyl Glucoside 6.00 Squalane Squalane 3.00 Cetiol SB 45 Butyrospermum Parkii (Shea Butter) 2.00 Waglinol 250 Cetearyl Ethylhexanoate 3.00 -19- Trade Names INCI Names% by mass Amerchol L- 101 Ore Oil (and) Lanolin Alcohol 2.00 Abil 350 Dimethicone 1.50 BHT BHT 0.01 Coenzyme Q10 Ubiquinone 0.10 Phase B Avocado Oil Persea Gratissima (Avocado ) Oil 1.25 Phenonip Phenoxyethanol (and) Methylparaben (and) 0.75 Ethylparaben (and) Butylparaben (and) Propylparaben (and) Isobutylparaben Phase C Demineralized Water Aqua (Water) qsp Butylene Glycol Butylene Glycol 2.00 Glucam El0 Methyl Gluceth Allantoin Allantoin 0.15 Carbopol Ultrez 10 Carbomer 0.20 Phase D TEA I Triethanolamine 0.18 Phase E Active ingredient according to Example 1 0.5 GP4G Water (and) Artemia Extract 1.50 Collaxyl Water ( and) Butylene Glycol (and) Hexapeptide- 3.00 9 Phase F Perfume Fragrance (Fragrance) qsp Col Preparing and melting phase A at 65-70C. Heat phase C at 65-70C. Phase B is added to phase A just before emulsifying A in B. At about 45C, the carbomer is neutralized by addition of phase D. Phase E is then added with gentle stirring and cooling is continued to 25 ° C. Phase F is then added if desired.

  4 ûShield Day Cream: Trade Names INCI Names% by mass Phase A Emulium Delta Cetyl alcohol (and) Glyceryl Stearate (and) 4.00 PEG-75 Stearate (and) Ceteth-20 (and) Steareth-20 Lanette O Cetearyl Alcohol 1 , 50 DC 200 Fluid / 100cms Dimethicone 1.00 DUB 810C Coco Caprylate / Caprate 1.00 DPPG Propylene Glycol Dipelargonate 3.00 DUB DPHCC Dipentaerythrityl Hexacaprylate / Hexacaprate 1.50 Cegesoft PS6 Vegetable Oil 1.00 Vitamin E Tocopherol 0.30 Phenonip Phenoxyethanol (and) Methylparaben (and) 0.70 Ethylparaben (and) Butylparaben (and) Propylparaben (and) Isobutylparaben Phase B Demineralized Aqua qs 100 Glycerine Glycerin 2.00 Carbopol EDT 2020 Acrylates / C 10-3OAlkyl Acrylate 0.15 Crosspolymer Keltrol BT Xanthan Gum 0.30 Phase C -21- Sodium Hydroxide (Sodium Hydroxide Sodium 0.30 10%) Phase D Demineralized Water Aqua 5.00 Stay-C 50 Sodium Ascorbyl Phosphate 0.50 Phase E Butylene Glycol Butylene Glycol 2.00 Dekaben CP Chlorphenesin 0.20 Phase F GP4G Water (and) Artemia Extract F 1.00 Active ingredient according to Example 1 Prepare phase A and heat to 75 ° C with stirring. Prepare phase B by dispersing the carbopol, then the xanthan gum with stirring. Let rest. Heat to 75 C. At temperature, emulsify A in B with rotor-stator stirring. Neutralize with phase C with rapid stirring. After cooling to 40 ° C., add phase D and then phase E. Cooling is continued with gentle stirring and phase F added.

Claims (14)

  1. Use of an effective amount of at least one active principle, derived from the hydrolysis of soybean (Glycine max L.), in a cosmetic composition or for the preparation of a pharmaceutical composition, characterized in that the active ingredient, or the composition containing it, is intended to activate aconitase and protect mitochondria.   2. Use according to claim 1, characterized in that the active principle is of peptide nature and comes from the hydrolysis of soy proteins (Glycine max L.).   3. Use according to any one of the preceding claims, characterized in that the active ingredient contains at least 0.1 and 5 g / 1, and preferably at least 0.5 to 2 g / 1 of peptide-like compounds. . 15   4. Use according to any one of the preceding claims, characterized in that the active ingredient is used in an amount representing from 0.0001% to 20% of the total weight of the composition, and preferably in an amount representing 0.05 % to 5% of the total weight of the composition. 20   5. Use according to any one of the preceding claims, characterized in that the active ingredient is first solubilized in one or more cosmetically or pharmaceutically acceptable solvents, such as water, glycerol, ethanol, propylene glycol, butylene glycol, dipropylene glycol, ethoxylated or propoxylated diglycols, cyclic polyols, petrolatum, a vegetable oil or any mixture of these solvents.   6. Use according to any one of the preceding claims, characterized in that the composition is in a form suitable for topical application comprising a cosmetically or dermatologically acceptable medium.   7. Use according to any one of the preceding claims, characterized in that the composition additionally contains at least one antioxidant active principle and / or at least one active principle stimulating the synthesis of the extracellular matrix, and / or at least one active principle stimulating the energetic cellular metabolism.   8. Use of an effective amount of active ingredient, as defined in any one of claims 1 to 3, in a cosmetic composition or for the preparation of a pharmaceutical composition for protecting the skin and the integuments against all types of external aggression.   9. Use of an effective amount of active ingredient, as defined in any one of claims 1 to 3, in a cosmetic composition or for the preparation of a pharmaceutical composition for protecting mitochondria.   10. Use of an effective amount of active ingredient, as defined in any one of claims 1 to 3, in a cosmetic composition or for the preparation of a pharmaceutical composition for preventing or treating damage to the skin. the skin and to the integuments by UV radiation.   11. Use of an effective amount of an active ingredient as defined in any one of claims 1 to 3 in a cosmetic composition or for the preparation of a pharmaceutical composition for preventing or treating damage to the skin. skin and skin appendages by free radicals.   12. Use of an effective amount of active ingredient, as defined in any one of claims 1 to 3, in a cosmetic composition or for the preparation of a pharmaceutical composition for preventing or treating cutaneous signs of the skin. aging and / or photo-aging.   13. Use of an effective amount of active ingredient, as defined in any one of claims 1 to 3, for preparing pharmaceutical compositions for preventing or combating mitochondrial dysfunction.   14. A cosmetic treatment method for stimulating the defenses and protecting the skin and the integuments of the external aggressions, characterized in that it is applied topically on the skin or integuments to treat a composition as defined according to US Pat. any of claims 1 to 7.