Exopolysaccharide and oligosaccharides in particular for cosmetics

Abstract

The present invention relates to an exopolysaccharide, to oligosaccharides obtainable by cleavage of said exopolysaccharide, to a microorganism for their manufacture, to a process obtaining them, to compositions containing them, and to uses, particularly cosmetic uses. Oligosaccharides are particularly adapted to improve skin radiance, relief, firmness and hydration and to protect the skin from aging and / or pollution effects.

Description

EXOPOLYSACCHARIDE AND OLIGOSACCHARIDES IN PARTICULAR FOR COSMETICS

[0001] The present invention relates to an exopolysaccharide, to oligosaccharides obtainable by cleavage of said exopolysaccharide, to a microorganism for their manufacture, to a process for obtaining them, to compositions containing them, and to uses, particularly cosmetic uses.

[0002] The fields of application of the present invention are mainly the chemical industries, for example the food, cosmetics, pharmaceuticals, veterinary, agricultural, environmental or paints and coatings industries.

[0003] The present invention is applicable more particularly to the fields of the cosmetic and dermo-pharmaceutical industries which manufacture and / or use products intended for the treatment of the skin (including the scalp), the mucous membranes and the appendages (such as body hair, eyelashes, eyebrows, nails, hair) of mammals, animals or humans, for improving the appearance and / or the general condition thereof.

[0004] Polysaccharides consist of a longer or shorter concatenation of monosaccharides and / or of monosaccharide acids linked by glycosidic bonds. They may be linear or branched. They are present in natural state in plants, mammals or microorganisms, their roles being in particular the stockage of energy, the structure or support of tissues, the regulation of tissue viscoelasticity, the water retention, the protection against bacterial or protease attacks, and / or the blood coagulation.

[0005] The family of the polysaccharides contains a particular class consisting of the exopolysaccharides which are characterized in that they are produced by microorganisms living in ecosystems under extreme conditions and in that they are secreted on the outside of the cells.

[0006] One of the advantages of exopolysaccharides for industry is that the microorganisms which produce them can be cultured in high volume under controlled conditions. This facilitates the production of exopolysaccharides on a large scale relative to polysaccharides, which often require complex extractions from natural sources such as plants or algae.

[0007] A number of exopolysaccharides have been described for the field of cosmetics and / or dermatology.

[0008] Patent application US2009 / 069213 describes a number of exopolysaccharides obtained from strains of microalgae from thePorphyridiumfamily. An exopolysaccharide obtained from the strainPorphyridiumcruentumis presented,composed of three monosaccharides, xylose, glucose and galactose, and of the glucuronic monosaccharide acid, and recommended for its antiwrinkle and moisturizing properties.

[0009] Another example is given in patent application WO2012 / 072245, which describes an exopolysaccharide isolated from a strain ofPseudoalteromonas sp. deposited under number I-4150, which is a specifically non-hydrothermal strain. This exopolysaccharide is composed of five monosaccharides, mannose, glucose, galactose, rhamnose and N-acetylglucosamine, and of two monosaccharide glucuronic and galacturonic acids. It is recommended as an alternative to hyaluronic acid, again for antiwrinkle and moisturizing effects.

[0010] A further example is given in patent application WO2015 / 117985, which describes an exopolysaccharide obtained from a culture ofPseudomonas sp. deposited under number CECT8437. This exopolysaccharide is composed of three monosaccharides, glucose, galactose and fucose, and of a uronic monosaccharide acid. It is recommended for its emulsifying activity in the cosmetics and food fields.

[0011] Furthermore, exopolysaccharides may advantageously be used as a source of saccharide monomers or oligosaccharides by virtue of appropriate cutting. For the skilled person, oligosaccharides are small polysaccharides often characterized in that they comprise fewer than 25 monosaccharides and / or monosaccharide acids. The advantage of the oligosaccharides is that they more readily penetrate the skin barrier and exhibit a better stability in formulations, owing to their reduced size.

[0012] Accordingly, the abovementioned patent application WO2015 / 117985, for example, describes the hydrolysis of exopolysaccharide to oligosaccharides and monosaccharides which can be used in cosmetic or pharmaceutical products. No particular activity is presented.

[0013] Another patent application, WO98 / 50013, describes the use of oligosaccharides, without specifying their provenance. The oligosaccharides described possess between two and six oside residues, including a galactose residue in terminal position. They are recommended in cosmetic compositions for increasing the synthesis and / or reducing the degradation of proteoglycans in the connective tissue, allowing the skin to be moisturized, and for fighting against the effects of free radicals.

[0014] However, there is still a high commercial demand for new active ingredients to respond to the current problem of sustainability, particularly to offer products obtained from biotechnological processes that preserve natural resources.

[0015] A particular aim of the present invention is to meet this demand.

[0016] For this purpose, the invention, according to afirstaspect, provides an exopolysaccharide characterized in that it comprises at least one monosaccharide unit that is of N-acetylquinovosamine (NAcQ).

[0017] This new exopolysaccharide advantageously constitutes a source of new oligosaccharides, owing to the presence of N-acetylquinovosamine, which exhibit original cosmetic properties, as shown by the results of tests set out below in the description.

[0018] Furthermore, this new exopolysaccharide may be used advantageously in the technical fields mentioned above, in the native or partially cleaved state. For example, it may be used as a rheological or film-forming agent in formulations. In the field of cosmetics, it may be used for improving the general condition of the skin and appendages, by forming a protective barrier to prevent water loss and to create a sensation of softness and comfort. It may also have antiaging propertiesviaa tightening effect.

[0019] According to other features, the molecular weight of the exopolysaccharide is greater than or equal to 260 000 Da, more particularly greater than or equal to 500 000 Da.

[0020] According to yet other particular features of the exopolysaccharide according to the invention:

[0021] - it further comprises at least one monosaccharide acid unit selected from glucuronic acid (Glu) and galacturonic acid (Gal), and preferably comprises these two monosaccharide acid units; and / or

[0022] - it further comprises at least one monosaccharide unit that is of rhamnose (Rham); and / or

[0023] - it comprises between 15% and 55% of N-acetylquinovosamine, preferably substantially 25%; and / or

[0024] - it comprises between 15% and 55% of glucuronic acid, preferably substantially 25%; and / or

[0025] - it comprises between 15% and 55% of galacturonic acid, preferably substantially 25%; and / or

[0026] - it comprises between 15% and 55% of rhamnose, preferably substantially 25%; and / or

[0027] - it comprises a concatenation of the following three saccharide units: glucuronic acid, galacturonic acid and N-acetylquinovosamine (Glu-Gal-NAcQ); and / or

[0028] - it further comprises rhamnose, with the following concatenation: glucuronic acid, galacturonic acid, N-acetylquinovosamine and rhamnose (Glu-Gal-NAcQ-Rham); and / or

[0029] - it consists of the repetition of the concatenation glucuronic acid, galacturonic acid, N-acetylquinovosamine, rhamnose (Glu-Gal-NAcQ-Rham), each saccharide unit being present substantially in equimolar amount in the exopolysaccharide.

[0030] All of the percentages expressed in the paragraph above are by weight relative to the total weight of the exopolysaccharide.

[0031] Further, the exopolysaccharide may be derivatized on the carboxylic acid functions when they are present; the exopolysaccharide particularly may be derivatized such that at least one carboxylic acid end of the glucuronic acid or of the galacturonic acid is esterified or acylated, for example.

[0032] According to yet other features, the exopolysaccharide is obtained from the bacterial strainPseudoalteromonas sp., deposited by the Applicant at the Collection Nationale de Culture de Microorganismes (CNCM) at Paris under number I-5893 according to the Treaty of Budapest. This bacterial strain was isolated at the surface of a coral in candelabra shape of the octocoral type belonging to the genusPleuxaura, collected in the warm waters of the Bimini Islands in the Bahamas. The unsporulated bacterium is stick-shaped and grows in colonies in salt-rich environments on account of its marine origin.

[0033] As mentioned above, the exopolysaccharide according to the invention may be used as a source of oligosaccharides.

[0034] Therefore, according to asecondaspect, the present invention also provides an oligosaccharide obtainable by cleavage from the exopolysaccharide according to the invention and composed of at least one monosaccharide unit that is of N-acetylquinovosamine.

[0035] The oligosaccharide according to the invention may also be manufactured synthetically by a chemical or enzymatic way, starting from monosaccharides.

[0036] Advantageously, oligosaccharides according to the invention are presented in the detailed description as possessing particularly interesting and original cosmetic activities as well as conventional moisturizing and antiwrinkle properties. In particular, the results of tests show that the oligosaccharides help to fight against cellular hypoxia for a very rapid cosmetic benefit on the radiance of the complexion.

[0037] According to particular features, the oligosaccharide according to the invention:

[0038] - has a degree of polymerization of between 2 and 12, preferably a degree of polymerization of 3, 4, 7 or 11; and / or

[0039] - comprises, further to the N-acetylquinovosamine, at least one monosaccharide acid unit selected from glucuronic acid and galacturonic acid, and preferably it comprises these two monosaccharide acid units; and / or

[0040] - comprises the concatenation glucuronic acid, galacturonic acid and N-acetylquinovosamine (Glu-Gal-NAcQ); and / or

[0041] - further comprises rhamnose, with the following concatenation: glucuronic acid, galacturonic acid, N-acetylquinovosamine, rhamnose (Glu-Gal-NAcQ-Rham).

[0042] According to other features, since the average molecular weight of a saccharide unit of the exopolysaccharide according to the invention is approximatively 200 Da (corresponding to the average of the mass of one unit of N-acetylquinovosamine, of glucuronic acid, of galacturonic acid and of rhamnose), the oligosaccharides according to the invention preferably have a molecular weight of less than or equal to 5000 Da, corresponding to a degree of polymerization of 25, more preferably a molecular weight of less than or equal to 3000 Da, corresponding to a degree of polymerization of 15, more preferentially less than or equal to 2200 Da, corresponding to a degree of polymerization of 11, more preferentially less than or equal to 1400 Da, corresponding to a degree of polymerization of 7, more preferentially less than or equal to 800 Da for a degree of polymerization of less than or equal to 4, more preferentially less than or equal to 600 Da for a polymerization degree of less than or equal to 3, and more preferably less than or equal to 400 Da for a polymerization degree of less than or equal to 2.

[0043] The preferred oligosaccharides according to the invention are:

[0044] - a trisaccharide with a degree of polymerization of 3, comprising one N-acetylquinovosamine unit, one glucuronic acid unit and one galacturonic acid unit and having a molecular weight of 557 Da, consisting more particularly of the concatenation Glu-Gal-NAcQ; and / or

[0045] - a heptasaccharide with a degree of polymerization of 7, comprising two N-acetylquinovosamine units, two glucuronic acid units, two galacturonic acid units and one rhamnose unit and having a molecular weight of 1242 Da, consisting more particularly of the concatenation Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ; and / or

[0046] - a hendecasaccharide with a degree of polymerization of 11, comprising three N-acetylquinovosamine units, three glucuronic acid units, three galacturonic acid units and two rhamnose units and having a molecular weight of 1927 Da, consisting more particularly of the concatenation Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ.

[0047] Further, the oligosaccharide according to the invention may be derivatized on the carboxylic acid functions when they are present; the oligosaccharide particularly may be derivatized such that at least one carboxylic acid end of the glucuronic acid or of the galacturonic acid is esterified or acylated, for example.

[0048] As presented above, the exopolysaccharide according to the invention and consequently the resultant oligosaccharides may be obtained by a fermentation process, particularly starting from the strain I-5893 deposited at the CNCM.

[0049] According to athirdaspect, the present invention therefore provides a bacterial strainPseudoalteromonas sp., deposited by the Applicant at the Collection Nationale de Culture de Microorganismes (CNCM) at Paris under number I-5893.

[0050] According to afourthaspect, the present invention provides a process for cultured manufacture of this strain for producing the exopolysaccharide according to the first aspect and oligosaccharides according to the second aspect, said process comprising the following steps:

[0051] - a first step of culturing a bacterial strain of the genusPseudoalteromonas sp. I-5893 deposited at the CNCM in a culture medium adapted to the production of the bacterium excreting the exopolysaccharide in the culture medium; and

[0052] -a second step of recovering the exopolysaccharide separated from the culture medium.

[0053] Advantageously, in the first step, the exopolysaccharide is excreted by the bacterial strain in the culture medium at the same time as the amount of bacterial strain is increased.

[0054] The culture medium is liquid and may contain macroelements, microelements, yeast extracts, vitamins, and sugars. It is selected to promote the growth and the production of the exopolysaccharide according to the invention.

[0055] Mineral salts may be used as microelements for the culture. For example and without limitation, salts providing the ions Na+, K+, NH4+, Ca²+, Mg²+, PO43-, SO42-, Cl-and / or CO32-, or trace elements such as Cu, Mn, Fe and / or Zn, or a mixture thereof. These salts are selected to promote the growth and the production of the exopolysaccharide.

[0056] Similarly, sugars may be used as a carbon source to feed the bacterial strain, such as, without limitation thereto, galactose, glucose, mannose, amygdaline, cellobiose, maltose, starch, glycogen, lactose or a mixture thereof. The addition of sugars also increases the production of the exopolysaccharide.

[0057] According to other features, in the second step, the separation of the exopolysaccharide from the culture medium takes place in two stages.

[0058] In a first stage, the exopolysaccharide present in the liquid culture medium is separated from the biomass, which is solid, by a process enabling separation of a liquid phase from a solid. For example, without limitation thereto, the separation may be carried out by centrifugation, ultrafiltration, tangential filtration or press filtration. The biomass present in the solid phase may be removed or retained to be added, after extraction, to the exopolysaccharide obtained at the end of the process, with the aim of enriching it with proteins and with sugars, which are the compounds present in the biomass.

[0059] In a second stage, the exopolysaccharide is purified by isolating it from the liquid culture medium containing other molecules, which may be salts, proteins, etc. For example, these molecules can be precipitated by virtue of a decrease in the temperature, referred to as cold precipitation, or by means of a water-miscible solvent in which the exopolysaccharide is insoluble - for example, methanol, isopropanol, ethanol or a mixture thereof - or else a tangential filtration may be carried out.

[0060] Preferably, the exopolysaccharide is isolated from the culture medium by virtue of a tangential filtration. More particularly, a filter membrane with a lower cut-off than the exopolysaccharide is used. More particularly, the filter membrane has a cut-off of lower than 150 000 Da.

[0061] In an additional step, the exopolysaccharide obtained by the process above may be cleaved to obtain at least one oligosaccharide according to the invention. This cleavage may be carried out by chemical or enzymatic hydrolysis.

[0062] Chemical hydrolysis in acid medium may be carried out by a strong acid, such as hydrochloric acid, nitric acid, sulfuric acid or trifluoroacetic acid.

[0063] Enzymatic hydrolysis may be carried out by a specific hydrolase enzyme.

[0064] For these different types of hydrolysis, the concentration of the solvent or enzyme, the reaction time, and the temperature are defined as a function of the size of the oligosaccharides it is desired to obtain. For example, the higher the acid concentration or the greater the reaction time, the greater the extent to which small oligosaccharides will be obtained.

[0065] Acid hydrolysis is the preferred method and the solvent preferably used is hydrochloric acid at a concentration of 0.2 M.

[0066] During the hydrolysis, precipitates may appear in the liquid medium containing the oligosaccharides. These precipitates may consist of salts, proteins, cell debris, etc., and attempts are made to remove these in order to purify the liquid phase. To accomplish this, any process enabling separation of a liquid phase from a solid is employed. For example, the separation may be carried out by centrifugation, ultrafiltration, tangential filtration or press filtration.

[0067] In the exopolysaccharide according to the invention, the chemical bonds around the rhamnose are cleaved, being the most fragile bonds. Since the exopolysaccharide has the repeating concatenation glucuronic acid, galacturonic acid, N-acetylquinovosamine and rhamnose, the following oligosaccharides which may particularly be obtained predominantly are:

[0068] - trisaccharides having the concatenation glucuronic acid, galacturonic acid and N-acetylquinovosamine (Glu-Gal-NAcQ);

[0069] -other oligosaccharides, having a concatenation of the trisaccharides with a rhamnose between each trisaccharide, such as particularly:

[0070] - heptasaccharides corresponding to two trisaccharides Glu-Gal-NAcQ connected by a rhamnose (Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ);

[0071] - hendecasaccharides corresponding to three trisaccharides Glu-Gal-NAcQ each connected by a rhamnose (Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ).

[0072] According to afifthaspect,the present invention provides a mixture of oligosaccharides obtainable by the process of the fourth aspect of the invention, preferably composed mainly of trisaccharides and / or heptasaccharides according to the invention, i.e., more than 50%, more particularly more than 80%. The percentages are expressed by weight relative to the total weight of the mixture of oligosaccharides.

[0073] Each oligosaccharide according to the invention may be isolated from the mixture of oligosaccharides obtained by the process according to the invention by an appropriate technique - for example, by flash chromatography on silica.

[0074] The exopolysaccharide and the oligosaccharides according to the invention, particularly obtained by the process described above, may be used in cosmetic or dermatological compositions as active ingredients.

[0075] Therefore, according to asixthaspect, the present invention provides a cosmetic or dermatological composition comprising the exopolysaccharide according to the invention as active agent and a physiologically acceptable medium.

[0076] Therefore likewise, according to aseventhaspect, the present invention provides a cosmetic or dermatological composition comprising at least one oligosaccharide according to the invention as active ingredient according to the invention and a physiologically acceptable medium.

[0077] The term "physiologically acceptable medium" means according to the present invention, without limitation, an aqueous or hydro-alcoholic solution, a water-in-oil emulsion, an oil-in-water emulsion, a micro-emulsion, an aqueous gel, an anhydrous gel, a serum, a dispersion of vesicles, or a powder.

[0078] The term "physiologically acceptable" means that the compositions are suitable for topical or transdermal use, in contact with mucous membranes, nails, scalp, hair, bodily hairs and skin of mammals and more particularly humans, compositions that may be ingested or injected into the skin, without risk of toxicity, incompatibility, instability, allergic response, and the like.

[0079] Preferably, the physiologically acceptable medium may be an aqueous, hydroglycolic or hydroalcoholic solution, or a water-in-oil emulsion, an oil-in-water emulsion, or a microemulsion, or a mixture thereof. Preferably, it is a mixture of water, glycol, and diol.

[0080] According to other advantageous features, the exopolysaccharide and / or the at least one oligosaccharide may be associated with one or more other active ingredients at effective concentrations that can act synergistically or additionally for reinforcing and achieving the desired effects described for the invention, such as the following agents: filtering radiations, in particular UVA, UVB, IR or generated by blue light, hydrating, moisturizing, humectant, calming, muscle relaxant, slimming, restructuring, firming, replumping, lifting, smoothing, acting on blood microcirculation, inflammation, free radicals, anti-aging, anti-fine lines and wrinkles, lightening, acting on complexion, anti-glycation, anti-carbonylation, pro- pigmenting, acting on stratum corneum, on dermal -epidermal junction, on HSP protein production, on firmness, elasticity and tone of skin, on hair growth or anti-regrowth (including eyelashes and eyebrows), on eye contours (dark circles and under eye bags), promoting blood circulation, regenerating, acting favorably or unfavourably on the skin microbiota and its appendages, promoting hair growth, promoting the elimination of toxins, etc. These active ingredients may be obtained from plant materials, such as plant extracts or products of plant culture or fermentation production methods.

[0081] More specifically, the exopolysaccharide and / or the at least one oligosaccharide according to the invention may be combined with at least one compound chosen among B3 vitamin compounds, compounds as niacinamide or tocopherol, retinoids compounds as retinol, in particular in encapsulated form, hexamidine, α-lipoic acid, resveratrol or DHEA, hyaluronic acid, ceramides or ceramide analogues, obtained synthetically or by biotechnological way, peptides, in particular N-acétyl-Tyr-Arg-O-hexadécyl, VW, Pal-PP, Biot-GHK, Ela-KFK, Pal-KPK, Pal-KHG, Myr-SPA, Pal-VGVAPG (SEQ ID NO : 1), Pal-KTTKS (SEQ ID NO: 2), Pal-GHK, Pal KMO2K, Pal-GQPR (SEQ ID NO : 3), Pal K(Ac)HG, Pal K(P)HG and Pal KTSKS (SEQ ID NO : 4), Pal KTFK (SEQ ID NO : 5), Pal YGGFL (SEQ ID NO : 6), cyclolinopeptides, the plant extract, in particularMarrubium vulgareextract,Leontopodium Alpinumextract obtained by in vitro cellular culture,Mirabilisjalapaextract, or mixture thereof, asRosmarinus officinalisand Pal-GQPR (SEQ ID NO : 3), which are known active ingredients that may be used in topical cosmetic or dermopharmaceutical compositions.

[0082] According to aneighthaspect, the present invention provides the use of the exopolysaccharide and / or of at least one oligosaccharide and / or of one of the compositions containing it according to the various aspects of the invention in the industrial fields mentioned above, namely – mainly – the chemical industries - for example, the food, cosmetics, pharmaceutical, veterinary, agricultural, environmental or paints and coatings industries.

[0083] More particularly, the present invention provides the use of at least one oligosaccharide and / or of the composition containing it according to the various aspects of the invention for a non-therapeutic cosmetic treatment of the skin and its appendages. Preferentially according to the invention, the treatment is topical.

[0084] Preferentially, the treatment is carried out with a mixture of oligosaccharides according to the fifth aspect of the invention.

[0085] According to the invention, “topical treatment” or “topical use” refers to an application which is intended to act at the site where it is applied: skin, mucous membrane and / or appendages.

[0086] More particularly, the cosmetic treatment provided by the present invention aims to fight against the unsightly skin signs that result from the loss of vitality of the cells and from the loss of or reduction in certain essential skin functions.

[0087] The loss of radiance of complexion, of firmness, of moisturization and / or the appearance of roughness of the skin form part of these aesthetical signs. They are due to normal chronological aging of skin, and / or to premature aging involving extrinsic factors such as, for example, aggressive external exposures, particularly atmospheric pollutants or forms of radiation. Over time and / or in the presence of these extrinsic factors, the vital functions of the cells of the skin and certain functions on the scale of the skin as well will deteriorate.

[0088] According to particular features, the cosmetic treatment implementing at least one oligosaccharide according to the invention is adapted to:

[0089] - improve the radiance of the complexion, also called brightness;

[0090] - improve the relief of the skin;

[0091] - improve the firmness of the skin; and / or

[0092] - improve the moisturization of the skin.

[0093] The tests carried outin vitroand presented below on in the detailed description show that the oligosaccharide-based treatment according to the invention makes it possible advantageously:

[0094] - to act at the level of the skin cells: by countering oxidation and cellular hypoxia, by stimulating the production of energy, the consumption of oxygen, and cell respiration; and

[0095] - to act on the scale of the skin: by reinforcing the barrier function, the moisturizing function, the function of the dermal-epidermal junction (DEJ) and the microbiotic function.

[0096] More specifically at the level of theskincells:

[0097] The tests show that the treatment according to the invention confersa beneficial antioxidant effectby decreasing the synthesis of the reactive oxygen species (ROSs)viaan increase in the synthesis of the antioxidant proteins encoded by the genes Nrf2, NQO-1 and HMOX-1.

[0098] Under an oxidizing stress, the protein encoded by the gene Nrf2 enters the nucleus of the cell and induces the activity of certain genes such as NQO-1 and HMOX-1.

[0099] NQO-1 is a gene coding for an antioxidant protein which serves to maintain a balance in the intracellular oxidative state and also to detoxify aromatic-type pollutants. This protein is synthesized when the ROSs increase in the cell.

[0100] HMOX-1 is the gene coding for the protein heme oxygenase (HO-1), which is induced quickly after exposure to free radicals. HMOX-1 exhibits a protective role against oxidative stress, owing essentially to its capacity to degrade heme into carbon monoxide and biliverdin, a precursor of bilirubin, which is a powerful antioxidant.

[0101] By virtue of this antioxidant action, skin aging is limited, particularly via preservation of the dermal extracellular matrix (ECM). The oxidation acts negatively on the synthesis of molecules, such as the fibroblasts which are responsible for the synthesis of collagen, elastin and other components of the ECM which allow the structure to be maintained, avoiding wrinkles and fine lines, allow firmness and elasticity to be maintained in the skin.

[0102] The tests show that the treatment according to the invention is active oncellular hypoxia:

[0103] - the mitochondrial potential, which creates energy in the cells which need it in order to manufacture proteins, operate enzymes, detoxify the cells from accumulated pollutants, for moisturizing the skin, etc., is increased;

[0104] - the consumption of oxygen also creates energy. In fact, oxygen is consumed to enable removal of CO2and to produce energy;

[0105] - the manufacture of adenosine triphosphate (ATP), which activates cell respiration by stimulating the mechanisms of detoxification and by increasing the production of the cells, is increased.

[0106] Greater energy and oxygenation of the cells result in detoxification and regeneration of the cells, improving the quality of the epidermis and reinforcing the skin barrier for a better radiance of the complexion.

[0107] More specifically on the scale of the skin:

[0108] Theinvitrotests show that the treatment according to the invention is active onthe reinforcement of theskinbarrierby acting on the quality and the thickness of thestratum corneum.

[0109] Thestratum corneum,or horny layer, is the outermost layer of the epidermis and constitutes a physical and functional barrier, its role being in one direction to prevent penetration of harmful external agents, such as allergens, pathogens and pollutants, and in the other direction to prevent dehydration of the skin.

[0110] Thestratum corneumis composed of corneocytes, lipids, lamellar bodies and natural moisturizing factors (NMFs - substances capable of fixing water in thestratum corneum) which maintain the degree of moisturization of the skin at an optimal level.

[0111] The transepidermal water loss (TEWL) characterizes the evaporation of the water contained in the skin: the greater the damage to the skin barrier, the more easily the water evaporates.

[0112] Improved differentiation of the keratinocytes leads to reinforcement of the skin barrier and, consequently, contributes to better protection of the epidermis and better moisturization.

[0113] The keratinocytes are viable cells of the epidermis with a lipid contour which migrate from the basal layer to the most superficial layer, differentiating themselves into corneocytes, which are small flat bricks without a nucleus. Proteins such as involucrin, filaggrin and claudin-1 participate in the formation and the migration of the keratinocytes into corneocytes. Furthermore, corneocytes are strongly linked to one another by the corneodesmosomes, in which the principal protein is corneodesmosin, and are “cemented” by organized layers of complex extracellular lipids: cholesterol, ceramides and neutral lipids.

[0114] Involucrin is a protein synthesized in the cytoplasm of the keratinocytes of thestratum granulosum. By bonding to the protein loricrin, involucrin contributes to the formation of a cellular envelope which protects the corneocytes of the skin.

[0115] The protein claudin-1 is a constituent of the tight junctions of thestratum granulosum, which form a selective barrier controlling the paracellular transport of molecules and of inflammatory cells. These tight junctions are also essential for limiting water loss.

[0116] The tests show an action on these essential proteins: involucrin, corneodesmosin and claudin-1.

[0117] The tests also show that the treatment according to the invention is active onthe moisturizing function of the skin, by also stimulating the synthesis of aquaporin-3 (AQP-3), which is a known marker for moisturization and which is present in the epidermis, and also the synthesis of filaggrin, which plays a pivotal role for maintaining the barrier function and which has degradation products that are essential to the regulation of pH and the skin moisturization. Advantageously, therefore, thanks to the treatment according to the invention, not only is dehydration of the skin prevented, with a reinforced skin barrier, but also the moisturization function itself is stimulated, by increasing the synthesis of one of the macromolecules that are responsible for moisturization in the epidermis. A better- moisturized skin is plumper, more supple and smoother (less rough).

[0118] Thanks to the treatment according to the invention, the skin is better protected and better moisturized, and therefore more radiant.

[0119] The treatment according to the invention acts also onthe function of theDEJ, by ensuring effective anchoring of the keratinocytes to the basal layer and better communication between the cells, enabling polarization of the keratinocytes. Synthesis of the main components of the DEJ (collagen IV, collagen VII and laminins) decreases with aging, and have therefore significant repercussions on the firmness of the skin.

[0120] The tests show that the treatment according to the invention is active on the markers of the DEJ, namely, collagen IV and collagen VII for improving the firmness of the skin. These molecules are also involved in reinforcement of the skin barrier.

[0121] In addition, the treatment according to the invention acts on theskinmicrobioticfunction.

[0122] This function is ensured mainly by the presence of the strainStaphylococcus epidermidisat the surface of the skin. This is a germ which colonizes the skin and is known to have beneficial effects on skin immunity. Furthermore, it has been demonstrated thatStaphylococcus epidermidiswas favorable for the homeostasis of the barrier function, owing to its capacity to cut sphingomyelin 1) into phosphocholine, which acts as a nutrient substrate for the germ and allows it to colonize the cutaneous space, and 2) into type-2 ceramide, which promotes the quality of the skin barrier.

[0123] The colonization of the skin surface byStaphylococcus epidermidisreinforces the effects of production of ceramides which are known to improve the skin barrier, prevent dehydration and smooth the skin which appear more radiant.

[0124] The tests show that the treatment according to the invention is active on the growth ofStaphylococcus epidermidisand on the production of ceramides by the skin and byStaphylococcus epidermidis.

[0125] All of these effects, advantageously and synergistically, can fight against a dull complexion (loss of radiance), a loss of firmness and a lack of moisturization of the skin, countering aesthetical signs on the skin.

[0126] Advantageously, the treatment according to the invention is particularly adapted to fight against pollution, particularly atmospheric pollution.

[0127] Results of specific tests on cellular hypoxia and detoxification of the cells are given below in the detailed description, performed in the presence of a solution of urban atmospheric microparticles (UMP) to simulate atmospheric pollutants.

[0128] In vivotests have been carried out confirming the cosmetic benefits according to invention, particularly a test on dull complexion (or radiance), a test on microrelief demonstrating a smoothing effect, a hydration test and a firmness test.

[0129] The present invention thus provides a non-therapeutic cosmetic treatment method for beautifying or improving the appearance and general condition of the skin and / or its appendages and for treating imperfections, in a subject in need thereof, comprising the application of an effective amount of the oligosaccharide and or a mixture of oligosaccharide according to the invention or of a composition comprising it, in a physiologically acceptable excipient.

[0130] The "effective" amount according to the invention, i.e. its dosage in the composition, depends on the intended use of the composition. It depends on various factors, such as the age and condition of the patient and the severity of the disorder. An effective amount means a non-toxic amount sufficient to obtain the desired effect.

[0131] In a cosmetic composition according to the invention containing at least one oligosaccharide, to be present in an effective amount, it is generally present in an amount ranging from 0.000001% and 15% based on the total weight of the composition, preferably ranging from 0.00001% and 10%, depending on the destination of the composition and the more or less pronounced desired effect. More preferably, the effective amount is between 0.0001% and 5% based on the total weight of the composition.

[0132] According to other features, the cosmetic treatment method according to the invention may be combined with one or more other treatment methods targeting the skin such as luminotherapy, heat or aromatherapy treatments.

[0133] According to the invention, devices with several compartments or kits may be proposed to apply the method described above which may include for example and non-restrictively, a first compartment containing a composition comprising the oligosaccharide of the invention, and in a second compartment an additional active ingredient, the compositions contained in the said first and second compartments in this case being considered to be a combination composition for simultaneous, separate or stepwise use in time, particularly in one of the treatment methods recited above.

[0134] For indication, for a cosmetic face treatment, the European standard dosage of a cream is 2.72 mg / cm² / day / person and for a cosmetic body treatment the European standard dosage of a lotion is 0.5 mg / cm² / day / person.

[0135] The present invention also relates to the use of the exopolysaccharide and / or the oligosaccharide and / or the oligosaccharide mixture according to the invention, for the manufacture of compositions for a cosmetic treatment, as described above.

[0136] The composition for the use according to the invention may be provided in any galenic form (examples are given below) defined according to the composition destination and application site.

[0137] A composition according to the invention may be applied to the face, body, neckline, scalp, hair, eyelashes, body hair, in any form or vehicle known to those skilled in the art, in particular in the form of a solution, dispersion, emulsion, paste or powder, individually or as a premix or vehicle individually or as a premix in a bound form, incorporated or adsorbed in vectors such as macro-, micro-, or nanocapsules, macro-, micro- or , nanospheres, liposomes, oleosomes or chylomicrons, macro-, micro-, or nanoparticles or macro-, micro or nanosponges, micro- or nanoemulsions, or adsorbed on organic polymer powders, talcs, bentonites, spores or exines, and other inorganic or organic supports.

[0138] In cosmetics, applications may be proposed particularly in the ranges of skin care for the face, body, hair and body hair and ranges of make-up treatments, in particular eyelashes and eyebrows.

[0139] For example, the galenic form of the composition may be a lotion, a cream, a butter, a milk, a solid form, a foam, a gel, a deodorant, an antiperspirant, a shampoo, a conditioner, a hair mask, a face mask, a shower gel, etc.

[0140] The galenical formulations may enter in different product ranges for personal care and / or beauty products including skin care, cleaning, makeup, cleansing, sunscreen, artificial tanning, pre-shave, shaving or aftershave, moisturizer, humectant, emollient, conditioning, exfoliating, astringent, depilatories or antiperspirant, deodorant, etc.

[0141] The composition may be incorporated onto a non-woven or woven material, with natural or synthetic fibres, wool, or any material intended to come into contact with skin and that may be used in clothing, including tights and socks, shorty, day or night underwear, tissues, handkerchiefs or fabric to exert its cosmetic effect via the contact skin / textile and enable continuous topical delivery (cosmetic-textiles).

[0142] According to characteristics of particular interest, the cosmetic treatment implementing oligosaccharides according to the invention is adapted to:

[0143] - improve the radiance of the complexion;

[0144] - improve the relief of the skin;

[0145] - improve the firmness of the skin; and / or

[0146] - improve the moisturization of the skin.

[0147] More particularly, the cosmetic treatment may be a treatment which protects the skin:

[0148] - from aging; and / or

[0149] - from pollution, especially atmospheric pollution.

[0150] More particularly, the treatment:

[0151] - improves the quality of the skin by reinforcing the skin barrier, for a skin which will be firmer, more supple, smoother and more moisturized and therefore for better radiance of the complexion; and / or

[0152] - fight against cellular hypoxia, for a very rapid cosmetic benefit on the radiance of the complexion.

[0153] These cosmetic effects may be envisaged according to the invention separately or in combination, offering advantageously a use combining beautifying and sensory effects.DETAILED DESCRIPTION

[0154] A greater appreciation of the present invention will be obtained in the light of the detailed description of working examples, studies and figures which are described below.

[0155] [Figure 1]: the figure shows a graph of deformation of the skin under the effect of a mechanical deformation as a function of time.

[0156] 1.Example of manufacture of theexopolysaccharidefrom the strainI-5893

[0157] First step:Production of the exopolysaccharide.

[0158] The strainPseudoalteromonasspnumber I-5893 is inoculated in a bioreactor containing a culture medium for a time sufficient to allow the amount of strain to multiply.

[0159] The culture medium is subsequently transferred to a larger bioreactor and fresh culture medium may be added to the reactor to increase production of the strain. Incubation is carried out for a time sufficient to obtain an even greater amount of thePseudoalteromonas sp. Strain. This step is optionally repeated.

[0160] The culture medium used comprises:from 0.1% to 0.5% of yeast extract;from 0.1% to 0.5% of KH2PO4;from 0.1% to 0.5% of Na2HPO4;from 0.1% to 0.5% of NH4Cl;from 0.001% to 0.01% of KCl;from 0.001% to 0.01% of iron citrate;at least 0.05% of antifoam; andfrom 1% to 5% of sea salts.

[0161] Glucose, a carbon source, at between 1% and 5% is also added to the culture medium to increase the production of the exopolysaccharide.

[0162] It is optionally possible during the incubation to add fresh culture medium in the bioreactor and / or to add minerals and / or to add sugars during the incubation.

[0163] The culture under these conditions is left for a time sufficient for the strain to excrete the exopolysaccharide according to the invention into the culture medium and to obtain the desired amount thereof.

[0164] Preferably, the incubation is carried out between 25°C and 35°C, 30°C for example, the pH between 6.5 and 8 and the dissolved oxygen pressure (pO2) between 10% and 50% by increased agitation. When the bacteria are in exponential phase, the pO2is regulated to stimulate the excretion of the exopolysaccharide into the medium.

[0165] During the incubation, the bacterial strain excretes the exopolysaccharide into the culture medium.

[0166] Second step:Recovery of the exopolysaccharide.

[0167] When the incubation period is over, the culture medium (liquid) containing the exopolysaccharide is separated from the biomass (solid). The liquid phase is separated from the solid phase using a continuous centrifugation at a velocity of 9000 g.

[0168] The biomass contained in the solid phase may be either removed or retained to be added, after extraction, to the exopolysaccharide obtained at the end of the process according to the invention or added to the mixture of oligosaccharides.

[0169] When the liquid phase containing the exopolysaccharide has been recovered, the exopolysaccharide according to the invention is isolated from the other molecules contained in the liquid phase. A tangential filtration is used with a 100 000 Da membrane.

[0170] The tangential filtration allows the molecules of the liquid to be separated by their size: the molecules having a size less than 100 000 Da pass through the membrane and are removed. The exopolysaccharide has a size greater than 260 000 Da; it does not pass through the membrane and therefore can be recovered.

[0171] Accordingly, the exopolysaccharide, washed from undesirable molecules and concentrated, is obtained.

[0172] Optionally, the exopolysaccharide obtained may subsequently undergo various types of treatment: dilution, preservation, purification, fractionation by precipitation or by chromatography, freeze drying, zeodration or spray drying.Characterization of the exopolysaccharide

[0173] During the fermentation, the secretion of the exopolysaccharide may be monitored by size exclusion chromatography (SEC) in aqueous phase with detection by an evaporative light scattering detector (ELSD).

[0174] The exopolysaccharide obtained was characterized by nuclear magnetic resonance (NMR) spectroscopy and direct infusion into a high-resolution (HR) Orbitrap mass spectrometer.

[0175] Cross-analysis of the spectra determined the repeating sequence composing the exopolysaccharide: Glu-Gal-NAcQ-Rham.

[0176] 2.Example of manufacture of the mixture of oligosaccharides from theexopolysaccharide

[0177] The exopolysaccharide according to the invention obtained by the process described above in section 1. may be cleaved to give oligosaccharides.

[0178] The cleavage is carried out by an acid hydrolysis step. The exopolysaccharide is mixed with 0.2 M concentrated hydrochloric acid solution with a pH <1. The mixture is heated at a temperature of between 85°C and 95°C, 90°C for example, for a time of between 2 hours and 4h30.

[0179] The hydrolysis is terminated by using an NaOH solution to raise the pH of the mixture, to pH 3 for example.

[0180] During the hydrolysis step, precipitates appear in the liquid medium containing the oligosaccharides. A clarification step is therefore carried out. They may be removed by centrifugation, ultrafiltration, tangential filtration or press filtration.

[0181] A mixture of oligosaccharides is obtained which comprises mainly trisaccharides and / or heptasaccharides according to the invention.

[0182] Optionally, each oligosaccharide according to the invention may be separated from the rest of the mixture of oligosaccharides by flash chromatography on silica.

[0183] Characterization of theobtainedmixture of oligosaccharides

[0184] The oligosaccharides present in the mixture have a degree of polymerization of greater than 2 and less than 12 and correspond to assemblies of the saccharide units of the exopolysaccharides, such as N-acetylquinovosamine, galacturonic acid and glucuronic acid and where appropriate rhamnose.

[0185] The mixture of oligosaccharides comprises approximatively 85% of oligosaccharides having the sequence Glu-Gal-NAcQ, mainly once the sequence, corresponding to trisaccharides, and / or twice the sequence, corresponding to heptasaccharides.

[0186] The major sequence Glu-Gal-NAcQ present in the mixture of oligosaccharides was characterized by NMR spectrometry and by ORBITRAP ultrahigh-performance liquid chromatography-high-resolution mass spectrometry.

[0187] Cross-analysis of the spectra determined:

[0188] - that the structure of the oligosaccharides is a concatenation of the units of the exopolysaccharide: glucuronic acid, galacturonic acid, N-acetylquinovosamine and where appropriate rhamnose.

[0189] - that the majority oligosaccharides in the mixture of oligosaccharides are:

[0190] - trisaccharides with the following concatenation: Glu-Gal-NAcQ; and / or

[0191] - heptasaccharides with the following concatenation: Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ;

[0192] - that the mixture of oligosaccharides comprises other oligosaccharides in minor amounts, such as:

[0193] - tetrasaccharides with the following concatenation: Glu-Gal-NAcQ-Rham;

[0194] - hendecasaccharides with the following concatenation: Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ.

[0195] 3.Example of manufacture of an active ingredient, particularly for cosmetics, containing the mixture of oligosaccharides

[0196] The mixture of oligosaccharides according to the invention obtained in section 2. above is dissolved in a physiologically acceptable medium comprising water, humectants and preservation boosters such as pentylene glycol and propanediol, to give a concentration of oligosaccharides of between 4000 ppm and 7000 ppm - for example, of 5000 ppm.

[0197] 4.Example of compositionscomprisingthe active ingredient

[0198] Various cosmetic formulations described below comprising the active ingredient according to the invention as described above in section 3.

[0199] Additional active ingredients, where appropriate to support and / or complement the activity, may be added in the appropriate phase according to their hydrophobic, hydrophilic, or amphiphilic nature. These ingredients may be of any category, for example ingredients of land-based, benthic, marine, aerial, vegetable, or microbial origin such as bacteria, archaea, or fungi.

[0200] These formulations may contain additional active ingredients, where appropriate to support and / or complement the activity of the active ingredient according to the invention. These ingredients may be of any category depending on their function(s), the place of application (body, face, neck, bust, hands, scalp, hair, bodily hairs, etc.), the desired end effect and the targeted consumer, for example anti-wrinkles, moisturizing, concealer, firming, anti-glycation, volumizing, soothing, myorelaxant, anti-redness, detoxifying, etc.

[0201] In the context of the present invention, the following additional active ingredients marketed by Sederma are particularly advantageous:To reinforce the activity of the active ingredient according to the invention such as BB-BIONTTMor skin radiance and smoothing, CITYSTEM™, PACIFEEL™ and / or SYNCHROLIFETMfor their anti-pollution activity, AQUALANCE™ fot its moisturizing action, CERAMIDE 2™, CERAMIDE A2™, CERAMIDE HO3™ and / or one or more products of DS-CERAMIDE™ range for their skin barrier reinforcement activity;in addition to the activity of the active ingredient according to the invention such as MEL[O]STEM™ for its white and brown anti-spots activity, CALMOSENSINETMand / or PACIFEEL™ for their skin soothing activity, MAJESTEMTM DRYPURETMfor its anti-ageing activity and one or more products of MATRIXYL™ range, particularly for their activity on the dermal EMC.

[0202] The International Cosmetic Ingredient Dictionary & Handbook published by the Cosmetic, Toiletry, and Fragrance Association, Inc. (CTFA), Washington, D.C., describes a wide variety, without limitation, of cosmetic and pharmaceutical ingredients commonly used in the skincare industry, which are suitable for use as additional ingredients in compositions according to the present invention, as long as they are physically and chemically compatible with the other ingredients of the composition and especially with the active agents of the present invention. Moreover, the nature of these additional ingredients must not impair the benefits of the active agents of the invention. These additional ingredients may be synthetic or natural, such as plant extracts, or be derived from a biofermentation method.

[0203] Other skincare active agents that are particularly useful in combination with the composition according to the invention can be found in the sales literature of Sederma, Crodarom and Alban Muller and on the website www.croda.fr.

[0204] Mention may also be made, by way of example, of the following commercial active agents: betaine, glycerol, Actimoist Bio 2™ (Active Organics), AquaCacteen™ (Mibelle AG Cosmetics), Aquaphyline™ (Silab), AquaregulK™ (Solabia), Carciline™ (Greentech), Codiavelane™ (Biotech Marine), Dermaflux™ (Arch Chemicals, Inc), Hydra'Flow™ (Sochibo), Hydromoist L™ (Symrise), RenovHyal™ (Soliance), Seamoss™ (Biotech Marine), Argireline™ (trade name of acetyl hexapeptide-3 from Lipotec), spilanthol or an extract ofAcmella oleraceaknown as Gatuline Expression™ (Gattefossé), an extract ofBoswellia serrataknown as Boswellin™, Deepaline PVB™ (SEPPIC), Syn-AKE™ (Pentapharm), Ameliox™, Bioxilift™ (Silab), PhytoCellTec™Argan (Mibelle), Papilactyl D™ (Silab), Preventhelia™ (Lipotec), or one or more of the following active ingredients sold by Sederma: Subliskin™, Venuceane™, Moist 24™, Vegesome Moist 24™, Essenskin™, Juvinity™, Revidrat™, Resistem™, Chronodyn™, Kombuchka™, Chromocare™, Calmosensine™, Glycokin factor S™, Biobustyl™, Idealift™, Ceramide 2™, Ceramide A2™, Ceramide HO3™, Legance™, Intenslim™, Prodizia™, Beautifeye™, PacifeelTM, Zingerslim™, Meiritage™, Sebuless™, Apiscalp™, Rubistem™, CitystemTM, NeonycaTM, NG Shea Butter UnsaponifiablesTM, MajestemTM, HydronesisTM, PoretectTM, CrystalideTM, AmberstemTM, Synchrolife™, Sylverfree™, Feminage™, Ameyezing™, BB-Biont™, Revitalide™, Mel[o]stemTM, or mixtures thereof.

[0205] Among the plant extracts (in the form of conventional extracts or prepared by anin vitromethod) that may be used as additional active agents, mention may also be made, in particular, of extracts of ivy, for example climbing ivy (Hedera helix),Bupleurum chinensis,Bupleurumfalcatum, arnica (ArnicamontanaL.), rosemary (Rosmarinus officinalisN.), calendula (Calendula officinalis), sage (Salvia officinalisL.), ginseng (Panax ginseng),Ginkgo biloba, St. John's wort (Hyperycumperforatum), butcher's broom (RuscusaculeatusL.), meadowsweet (FilipendulaulmariaL.), orthosiphon (OrthosiphonstamincusBenth.), artichoke (Cynara scolymus), seaweed (Fucus vesiculosus), birch (Betula alba), green tea, kola nut (Cola nitida), horse chestnut, bamboo,Centellaasiatica, heather, wrack, willow, pilosella, escin extracts, cangzhu extracts,Chrysanthellumindicumextracts, plants of theArmeniaceagenus,Atractylodisplaticodon,Sinnomenum, pharbitidis,Flemingia, coleus such asC.forskohlii,C.blumei,C.esquirolii,C.scutellaroides,C.xanthantusandC. barbatus, such asColeus barbatusroot extract, extracts of horehound,Guioa,Davallia,Terminalia,Barringtonia,Trema,Antirobia,Cecropia,Argania,Dioscoreaesuch asDioscoreaoppositaorDioscoreamexicana, extracts ofAmmivisnaga,Siegesbeckia, in particularSiegesbeckiaorientalis, plant extracts from theEricaceaefamily, in particular extracts of bilberry (Vaccinium angustifolium),Arctostaphylos uvaursi,Aloe vera, sterol-containing plants (particularly phytosterols),Manjistha(extract of plants of theRubiagenus, in particularRubia cordifolia),Guggal(extract of plants of the genusCommiphora, in particularCommiphoramukul), an extract of kola, chamomile, red clover,Pipermethysticum(Kava Kava from Sederma),Bacopamonieri(Bacocalmine™, Sederma) and sea whip,Glycyrrhiza glabra, mulberry,Melaleuca(tea tree),Larreadivaricata,Rabdosiarubescens,Euglenagracilis,Fibraurearecisahirudinea,Chaparral sorghum, sunflower,Enantiachlorantha,Mitracarpeof theSpermacoceagenus,Buchubarosma,LawsoniainermisL.,Adiantiumcapillus-venerisL.,Chelidonium majus,Luffa cylindrica, Japanese mandarin (Citrus reticulatablancovar. unshiu),Camelia sinensis,Imperata cylindrica,Glauciumflavum,Cupressussempervirens,Polygonatummultiflorum, lovely hemsleya,Sambucus nigra,Phaseolus lunatus,Centaurium,Macrocystispyrifera,Turneradiffusa,Anemarrhenaasphodeloides,Portulacapilosa,Humuluslupulus,Coffea arabica,Ilexparaguariensis,Globularia cordifolia,Oxydendronarboreum,Albizziajulibrissin,Zingiber zerumbetsmith,Astragalusmembranaceus,Atractylodesmacrocephalae,Plantago lanceolata,Leontopodium alpinum(or eldelweiss),Mirabilisjalapa,Apium graveolens,Marrubium vulgare,Buddleja davidiiFranch,Monarda didyma, deLavandula angustifoliaor orchids.

[0206] The compositions of the present invention may include peptides, including, without limitation, di-, tri-, tetra-, penta-and hexapeptides and their derivatives. According to a particular embodiment, the concentration of the additional peptide(s), in the composition, ranges from 1x10-7% and 20%, preferably from 1x10-6% and 10%, preferably between 1x10-5% and 5% by weight.

[0207] In the context of the present invention, the term “peptide” refers here to peptides containing 10 amino acids or less, their derivatives, isomers and complexes with other species such as a metal ion e.g., copper, zinc, manganese, magnesium, and others). The term "peptides" refers to both natural peptides and (bio)synthetic peptides. It also refers to compositions that contain peptides and which are found in nature, and / or are commercially available.

[0208] Non-limiting examples of dipeptides for use in the context of the present invention comprises to Carnosine (β-AH), YR, VW, NF, DF, KT, KC, CK, KP, KK, TT, PA, PM or PP.

[0209] Non-limiting examples of tripeptides are RKR, HGG, GHK, GGH, GHG, GKH, KPK, KFK, KavaK, KbêtaAK, KabuK, KacaK, KPK, KMOK, KMO2K (MO2being a di-oxygenated sulfoxide methionine), PPL, PPR, SPR, QPA, LPA or SPA.

[0210] Non-limiting examples of tetrapeptides are KTFK (SEQ ID NO : 7), GQPR (SEQ ID NO : 8), RSRK (SEQ ID NO : 9), KTAK (SEQ ID NO : 10), KAYK (SEQ ID NO :11), KFYK (SEQ ID NO : 12), TKPR (SEQ ID NO : 13), AVPG (SEQ ID NO : 14), VPGA (SEQ ID NO : 15), LKLE (SEQ ID NO : 16), ELED (SEQ ID NO : 17) or LLAN (SEQ ID NO : 18).

[0211] Non-limiting examples of pentapeptide are KTTKS (SEQ ID NO: 19) and KTSKS (SEQ ID NO :20).

[0212] Non-limiting examples of hexapeptides are GKTTKS (SEQ ID NO: 21) or VGVAPG (SEQ ID NO: 22).

[0213] Other peptides for use in the context of the present invention may be selected, this list being not limitative, from: lipophilic derivatives of peptides, preferably palmitoyl (Pal) derivatives or myristoyl (Myr), and metal complexes as aforementioned (e.g., copper complex of the tripeptide HGG or GHK).

[0214] Preferred dipeptides include for example N-Palmitoyl-β-Ala-His, Pal-KT, Pal-RT (Sederma). Preferred tripeptide derivatives include for example the copper derivative of HGG (Lamin™, Sigma), Pal-GHK, Lipospondin (N-Elaidoyl-KFK) and its analogs of conservative substitution, N-Acetyl-RKR-NH2(Peptide CK+), Pal-KavaK, Pal-KβAlaK, Pal-KabuK, Pal-KacaK, Pal-KMO2K, N-Biot-GHK (Sederma) and derivatives thereof.

[0215] Mention may also be made here of the anti-aging tripeptides of general formula X–Pro*–Pro*–Xaa–Y described in WO2015181688 Xaa selected from Leu, Arg, Lys, Ala, Ser, and Asp, at the N-terminus , X chosen from H, -CO-R1and -SO2-R1and at the C-terminal end Y chosen from OH, OR1, NH2, NHR1or NR1R2, R1and R2being, independently of one another, chosen from a alkyl, aryl, aralkyl, alkylaryl, alkoxy and aryloxy group, which may be linear, branched, cyclic, polycyclic, unsaturated, hydroxylated, carbonylated, phosphorylated and / or sulfurized, said group possibly possessing in its backbone a heteroatom particularly O, S and / or or N, and Pro* corresponding to Proline, an analogue or derivative thereof; comprising, for example, Myr-PPL-OH and Myr-PPR-OH.

[0216] Here can further be cited also the propigmenting and / or pro-MEC dipeptides and tripeptides of general Formula X–(Xaa1)n–Pro*–Xaa2–Y disclosed in WO2014 / 080376, with n=0, 1 or 2, Xaa1an hydrophobic aminoacid selected from Ala, Val, Met, Leu, Iso, Phe, Pro, and analogues and derivatives thereof; or a polar aminoacid selected from Ser, Thr, Tyr, Asp, Glu and analogues and derivatives thereof; and when n=2 the two aminoacids Xaa1being the same or different; Xaa2being an hydrophobic aminoacid selected from Ala, Val, Met, Leu, Iso, Phe, and analogues and derivatives thereof, or a basic aminoacid selected from Arg, Lys, His, and analogues and derivatives thereof; at the N terminal end X being selected from H, -CO-R1and -SO2-R1; at the C terminal end Y being selected from OH, OR1, NH2, NHR1or NR1R2; R1and R2being, independently from each other, selected from an alkyl, aryl, aralkyl, alkylaryl, alkoxy et aryloxy group, that can be linear, branched, cyclic polycyclic, saturated, unsaturated, hydroxylated, carbonylated, phosphorylated and / or sulfured, said group having or not an O, S and / or N heteroatom in its skeleton and Pro* corresponding to a Proline, analogue or derivative thereof; comprising for example the following peptides Pal-SPR-OH, Pal-PPR-OH, Pal-QPA-OH, Pal-LPA-OH, Myr-SPA-OH, Pal-PM-OH, Pal-PA-OH and Pal-PP-OH.

[0217] Suitable tetrapeptides derivatives for use as additional peptides according to the present invention include but are not limited to Ela-KTAK (SEQ ID NO: 23), Ela-KAYK (SEQ ID NO: 24), Ela-KFYK (SEQ ID NO : 25), Pal-GQPR (SEQ ID NO : 3) or Pal-KTFK (SEQ ID NO : 5).

[0218] Suitable pentapeptides derivatives for use as additional peptides herein include, but are not limited to, Pal-KTTKS (SEQ ID NO: 2), Pal-KTSKS (SEQ ID NO: 4), Pal-YGGFXaa (SEQ ID NO: 6) with Xaa being Leu or Pro.

[0219] Suitable hexapeptides derivatives for use as additional peptides herein include, but are not limited to, Pal-HLDIIXaa (SEQ ID NO: 26) with Xaa being Trp, Phe, Tyr, Tic, 7-hydroxy-Tic or Tpi, or mixtures thereof, Pal-GKTTKS (SEQ ID NO: 27), Pal-VGVAPG (SEQ ID NO: 1).

[0220] The preferred compositions available commercially and sold by Sederma:

[0221] - tripeptides or a derivative include Biopeptide-CL™, Maxi Lip™, or Procapil™ containing GHK;

[0222] - tetrapeptides or a derivative include RIGIN™, Eyeliss™ containing Pal-GQPR (SEQ ID NO: 3) and an excipient,

[0223] - pentapeptide or a derivative as Matrixyl™ containing Pal-KTTKS (SEQ ID NO: 2).

[0224] Can also be mentioned:

[0225] - the mixture of Pal-GHK and Pal-GQPR (SEQ ID NO: 3) (Matrixyl™ 3000), and

[0226] - the mixture of Pal-GHK and Pal-VGVAPG (SEQ ID NO: 1) (Biobustyl™)

[0227] The following marketed peptides can be mentioned as well as additional active ingredients:

[0228] - Vialox™, Syn-ake™ (β -Ala-Pro-Dab-NH-Bzl) or Syn-Coll™ (Pal-Lys-Val-Lys-OH) marketed by Pentapharm;

[0229] - Argireline™ (Ac-Glu-Glu-Met-Gln-Arg-Arg-NH2(INCI name = Acetyl hexapeptide-3)(SEQ ID NO: 28), Leuphasyl™ (Tyr-D-Ala-Gly-Phe-Leu) (SEQ ID NO: 29), Aldenine™ (Gly-His-Lys), TrylagenTM(INCI name = Pseudoalteromonas Ferment Extract, Hydrolyzed Wheat Protein, Hydrolyzed Soy Protein, Tripeptide-10 Citrulline (reaction product of Citrulline and Tripeptide-10 (synthetic peptide constituted of aspartic acid, isoleucine and lysine)), Tripeptide-1), Eyeseryl™ (Ac-bêta-Ala-His-Ser-His)(SEQ ID NO: 30), Serilesine™ (Ser-Ile-Lys-Val-Ala-Val) (SEQ ID NO: 31) or Decorinyl™ (INCI name: Tripeptide-10 Citrulline = reaction product of Citrulline and Tripeptide-10 (synthetic peptide constituted of aspartic acid, isoleucine and lysine) marketed by Lipotec

[0230] - Collaxyl™ (Gly-Pro-Gln-Gly-Pro-Gln (SEQ ID NO: 32)) or Quintescine™ (Cys-Gly) marketed by Vincience;

[0231] - Cytokinol™LS (casein hydrolysate) marketed by Les Laboratoires Serobiologiques / Cognis;

[0232] - Kollaren™ (Gly-His-Lys), IP2000™ (Pal-Val-Tyr-Val) or Meliprene™ (INCI name = Monofluoroheptapeptide-1: reaction product of acetic acid and a synthetic peptide comprising arginine, glycine, glutamic acid, histidine, norleucine, p-fluorophenylalanine and tryptophan) marketed by l’lnstitut Européen de Biologie Cellulaire;

[0233] - Neutrazen™ (Pal-His-D-Phe-Arg-NH2) marketed by Innovations; or

[0234] - BONT-L-Peptide™, Timp-Peptide™ or ECM Moduline™ marketed by lnfinitec Activos.

[0235] It is also possible to envisage combining the invention with one or more cyclic peptides, in particular those extracted from linseed oil described in the Applicant's patent application WO2019 / 149450.Serum form

[0236] Starting materials%INCI namePhase AH2OqsAqua / waterSodium benzoateqsSodium benzoatePotassium SorbateqsPotassium sorbatePhase BButylene Glycol5.50Butylene GlycolXanthan Gum0.25Xanthan GumArlasolve DMI PC2.50Dimethyl IsosorbidePhase CH2O0.40Aqua / waterLactic acid0.04Lactic acidPhase DIngredient according to the invention3.0Protocol :

[0237] Weigh and mix A. Weigh and mix B. Add B to A while stirring, mix. Weigh and mix C. Add C to B+A while stirring, mix. Add D to B+A+C, mix.Milky mist form

[0238] Starting materials%INCI namePhase AH2OqsAqua / waterPotassium sorbate0.10Potassium sorbatePhase BGlycerin5.00GlycerinMicrocare® Emollient DCP1.50Pentylene Glycol (and) Caprylyl Glycol (and) Decylene GlycolAmigum0.20Sclerotium GumPhase CSP Span™ 20 MBAL1.00Sorbitan LauratePhase DIngredient according to the invention3.0Protocol :

[0239] Weigh and mix A. Weigh and mix B. Add B to A while stirring, mix. Activate A+B by mixing them strongly. Weigh and mix C. Add C to B+A while stirring, mix. Add D to B+A+C, homogenize.5. Results of the in vitro tests

[0240] The mixture of oligosaccharides obtained in section 3. above was used for thein vitrotests carried out below, denoted as “active ingredient according to the invention”.

[0241] 5.1. Protectionagainst loss of radiance and the appearance of signs of skin aging5.1.1 Resistance to oxidation5.1.1.1 Effect on reactive oxygen speciesProtocol

[0242] Human keratinocytes (HK) are cultivated to confluence in their culture medium. The cells are subsequently contacted with the ingredient according to the invention for 28 hours and then receive a fluorescent probe intended to mark the intracellular production of ROSs. After incorporation, the cells become sensitive to oxidizing agents. Subsequently, the cells do or do not receive an agent intended to create ROSs (oxidizing stress). Reading the fluorescence (ex: 490 nm / em: 520 nm) allows the amount of intracellular ROSs to be estimated. The number of cells is estimated using the Hoechst method (DNA staining) to weight the data obtained.Results

[0243] Variation of the production of ROSs in HK. Effect of 2% of the active ingredient according to the invention relative to the control (n=6):

[0244] Variation (%); significanceControl caseReference2% of the active ingredient according to the invention-39.6%; p<0.01

[0245] The results show that 2% of the active ingredient according to the invention significantly reduces the production of ROSs, by -39.6% (p<0.01).5.1.1.2. Effect on the protein Nrf2Protocol

[0246] HK are cultivated to confluence in their culture medium. The cells are subsequently contacted with the ingredient according to the invention for 2 hours or 24 hours. Subsequently, immuno-cytochemical labeling of the protein Nrf2 took place and passage thereof from the cytoplasm to the nucleus was monitored by image analysis.Results

[0247] Variation of the translocation of Nrf2 as a function of time in a culture of HK. Effect of 3% of the active ingredient according to the invention relative to the control case (n=4):

[0248] Control case: contact with 0.1% DMSOContact with 3% of the active ingredient according to the inventionVariation (%); significanceAfter 2 hours of contactReference 1+41%; p<0.01After 24 hours of contactReference 2+36%; p<0.05

[0249] The results show that 3% of the active ingredient according to the invention significantly increases the translocation of the protein Nrf2 by +41% (p<0.01) at 2 hours of contact and by +36% (p<0.05) after 24 hours of contact.5.1.1.3. NQO-1Protocol

[0250] HK are cultivated to confluence in their culture medium. The cells are subsequently contacted with the ingredient according to the invention for 6 hours. The HK carpets are then rinsed, and the cells are ground for extraction of their mRNA. These mRNA are then converted into small DNA sequences, which are analyzed after having been applied to DNA chips and amplified by a method closely related to qRT-PCR. The variations in mRNA due to the ingredient according to the invention are compared with the control case.Results

[0251] The results show that 3% of the active ingredient according to the invention significantly increases the production of the protein NQO-1 (+67%,p<0.01).5.1.1.4. HMOX-1Protocol

[0252] In a first phase, HK are cultivated to confluence in their culture medium. The cells are subsequently contacted with the ingredient according to the invention for 54 hours. The cells are rinsed, dried, and stored frozen. The amount of HMOX-1 is measured by an ELISA method, after extraction.

[0253] The amount of total proteins was evaluated after extraction, by the bicinchoninic acid method, to weight the data obtained.

[0254] In a second phase, the cells are contacted with 500 µM Trolox in place of the ingredient according to the invention. Trolox is a hydrophilic close analogue of vitamin E which is known for its antioxidant activity and is used to limit the damage associated with oxidizing stresses.Results

[0255] Variation of the stimulation of production of HMOX-1. Effect of 3% of the active ingredient according to the invention at 54 hours relative to the control case (n=4):

[0256] Variation (%); significanceControl case 1Reference 13% of the active ingredient according to the invention+65%; p<0.01Control case 2Reference 2500 µM Trolox+24%; p<0.01

[0257] The results show that the stimulation of production of HMOX-1 is increased significantly in both cases, but that 3% of the active ingredient according to the invention stimulates the production of HMOX-1 more (+65%; p<0.01vsits control) relative to 500 µM Trolox (+24%; p<0.01vsits control).5.1.2. Resistance to cellular hypoxia

[0258] 5.1.2.1.Mitochondrial potentialΔΨ,promotiveforceProtocol

[0259] HK are cultivated to confluence in their culture medium. The cells are subsequently contacted with the ingredient according to the invention for 28 hours and then are labeled with a fluorescent probe.

[0260] Reading of these probes by fluorescence is used to measure the two forms and to establish the monomer / polymer ratio (520 nm / 590 nm). A high value of this ratio is less favorable to the survival of the cell than a low value.

[0261] The number of cells is estimated using the Hoechst method (DNA staining) to weight the data obtained.Results

[0262] Variation of the mitochondrial potential ΔΨ in HK. Effect of 3% of the active ingredient according to the invention relative to the control case (n=6):

[0263] Variation (%); significanceControl caseReference3% of the active ingredient according to the invention+33.2%; p<0.01

[0264] The results show that 3% of the active ingredient according to the invention significantly increases the mitochondrial potential ΔΨ by +33.2% (p<0.01).

[0265] 5.1.2.2. Protectionofrespiration, consumptionof oxygenProtocol

[0266] HK are cultivated to confluence in their culture medium. The cells are subsequently contacted with the ingredient according to the invention for 24 hours. The HK are subsequently detached from their support and placed in equivalent amounts into the wells of a plate having sensors which measure the consumption of oxygen by these cells as a function of time.Results

[0267] Variation of the oxygen consumption (oxygen pressure per minute: pO2) in HK. Effect of 3% of the active ingredient according to the invention relative to the control case (n=6):

[0268] Variation (%); significanceControl caseReference3% of the active ingredient according to the invention+23.6%; p<0.05

[0269] The results show that 3% of the active ingredient according to the invention acts favorably on the respiration of the cells, stimulating it by +23.6% (p<0.05).5.1.2.3. Synthesis of ATPProtocol

[0270] HK are cultivated to confluence in their culture medium. The cells are subsequently contacted with the ingredient according to the invention for 26 hours. The intracellular ATP is extracted and assayed by bioluminescence, the signal obtained being proportional to the amount of ATP in the cells.

[0271] The number of cells is estimated using a propidium iodide fluorescent probe to weight the data obtained.Results

[0272] Variation of the synthesis of ATP in HK. Effect of 3% of the active ingredient according to the invention relative to the control case (n=6):

[0273] Variation (%); significanceControl caseReference3% of the active ingredient according to the invention+84%; p<0.01

[0274] The results show that 3% of the active ingredient according to the invention significantly increases the synthesis of ATP, by +84% (p<0.01) dose-dependently relative to the control case.5.1.3. Reinforcing the barrier function5.1.3.1. Thickness of the stratum corneumProtocol

[0275] Equivalent skins are prepared and contacted with the ingredient according to the invention for 2 days. The skins are subsequently labeled with hematoxylin and eosin to visualize the macrostructure of the various layers of the skin. This labeling is monitored by image analysis.Results

[0276] Variation of the thickness of thestratum corneum. Effect of 3% of the active ingredient according to the invention relative to the control case (n=6):

[0277] Variation (%); significanceControl caseReference3% of the active ingredient according to the invention+43.4%; p<0.01

[0278] The results show that 3% of the active ingredient according to the invention significantly increases the thickness of thestratum corneum, by +43.4% (p<0.01).5.1.3.2. Involucrin and corneodesmosinProtocol

[0279] Equivalent skins are prepared and contacted with the ingredient according to the invention for 2 days. The skins are subsequently labeled using antibodies directed against involucrin and corneodesmosin. This labeling is monitored by image analysis (N = 57 to 60 photos per case).

[0280] The number of cells is estimated using the Hoechst method (DNA staining) to weight the data obtained.Results

[0281] Variation of the synthesis of corneodesmosin and involucrin. Effect of 3% of the active ingredient according to the invention relative to the control case (n=5):

[0282] InvolucrinVariation (%); significanceCorneodesmosinVariation (%); significanceControl caseReference 1Reference 23% of the active ingredient according to the invention+42.0%; p<0.01+46.0%; p<0.01

[0283] The results show that 3% of the active ingredient according to the invention significantly increases the production of involucrin (+42%; p<0.01) and of corneodesmosin (+46%; p<0.01). The quality of the barrier function is therefore improved.5.1.4. Reinforcing the moisturization functionProtocol

[0284] Skin explants are prepared and the ingredient according to the invention is placed on their surfaces for 3 days. Subsequently, the explants are sectioned into thin 7 µm slices and labeled using antibodies directed against moisturization markers: filaggrin and aquaporin-3. This labeling is monitored by image analysis (N = 41 to 45 photos per case).Results

[0285] Variation of the synthesis of filaggrin and aquaporin-3 in skin explants. Effect of 3% of the active ingredient according to the invention relative to the control case (n=3):

[0286] Variation (%); significanceAquaporin-3+11.0%; p<0.01Filaggrin+10.0%; p<0.01

[0287] The results show that 3% of the active ingredient according to the invention significantly increases the two moisturization markers.

[0288] 5.1.5.Reinforcing the function of the dermal-epidermal junction(DEJ)Protocol

[0289] The same protocol as for filaggrin and aquaporin-3 was used for claudin-1, collagen IV and collagen VII. The labeling is monitored by image analysis (N = 41 to 45 photos per case).Results

[0290] Variation of the synthesis of claudin-1, collagen IV and collagen VII in skin explants. Effect of 3% of the active ingredient according to the invention relative to the control case (n=3):

[0291] Variation (%); significanceCollagen IV+26.0%; p<0.01Collagen VII+27.0%; p<0.01Claudin-1+7.0%; p<0.01

[0292] The results show that 3% of the active ingredient according to the invention significantly increases the markers of the DEJ (collagen IV and collagen VII) and the marker of the tight junctions (claudin-1).5.1.6. Reinforcing the microbiotic function5.1.6.1. Growth of Staphylococcus epidermidisProtocol

[0293] The ingredient according to the invention is contacted with a culture ofStaphylococcus epidermidis(ATCC 14990) at 4.105CFU* / mL, the whole at 37°C in a culture medium allowing the growth of this bacterium. The growth kinetics are monitored by measurements of optical density (OD) at 600 nm.Results

[0294] Growth kinetics ofS. epidermidisover time. Effect of 3% of the active ingredient according to the invention relative to the control case (n=3):

[0295] Variation between T=0 and 12 hours (%); significanceVariation between T=0 and 16 hours (%); significanceControl caseReference 1Reference 23% of the active ingredient according to the invention+1120%; p<0.01+1497%; p<0.01

[0296] The results show that 3% of the active ingredient according to the invention significantly increases growth ofStaphylococcus epidermidis.5.1.6.2. Synthesis of ceramides by the skinProtocol

[0297] Keratinocytes in culture are contacted with the ingredient according to the invention for 24 hours. At the end of this contact, the cells are detached from the culture medium. The quantification of the ceramides is measured by an ELISA method. The number of cells is standardized to harmonize the results.Results

[0298] Variation of the production of ceramides by the skin. Effect of 3% of the active ingredient according to the invention relative to the control case (n=4):

[0299] Variation between T=0 and 24 hours (%); significanceControl caseReference3% of the active ingredient according to the invention+22%; p<0.05

[0300] The results show that 3% of the active ingredient according to the invention significantly increases the synthesis of ceramides by the skin.

[0301] 5.1.6.3.Synthesis of ceramides byStaphylococcus epidermidisProtocol

[0302] Staphylococcus epidermidisis contacted with the ingredient according to the invention for 24 hours. The RNA is extracted and the expression of thesphgene of sphingomyelinase is quantified by RT-qPCR.Results

[0303] Variation of the expression of thesphgene of sphingomyelinase. Effect of 3% of the active ingredient according to the invention relative to the control case (n=4):

[0304] Variation between T=0 and 24 hours (%); significanceControl caseReference3% of the active ingredient according to the inventionx3.41; p<0.01

[0305] The results show that 3% of the active ingredient according to the invention significantly increases the expression of thesphgene of sphingomyelinase, the sphingomyelinase ofStaphylococcus epidermidisbeing capable of converting sphingomyelin into ceramide and phosphocholine.5.1.6.4. Conclusion

[0306] All thein vitroresults presented above show that the mixture of oligosaccharides according to the invention protects theskinmicrobioticfunctionwith an increase in the growth ofStaphylococcus epidermidisand on the production of ceramides by the skin and byStaphylococcus epidermidis.5.2. Protection against pollution

[0307] The tests were carried out in the presence of atmospheric UMP to simulate the effect of pollution.5.2.1. Resistance to oxidation5.2.1.1. Action toward reactive oxygen speciesProtocol

[0308] The effects of atmospheric pollutants on the formation of ROSs were evaluated. The protocol used is the same as in section 5.1.1.1. and a solution of UMP is contacted with the cultivated cells and labeled as before.Results

[0309] Variation of the production of ROSs in HK (n=6) with effect of atmospheric pollutants. Effect of 2% of the active ingredient according to the invention relative to the control with UMP (n=6):

[0310] Variation (%); significanceControl case without UMPReference 1Control case with UMP+26.9%; p<0.01Reference 22% of the active ingredient according to the invention, with UMP-41.9%; p<0.01

[0311] The results show that pollutant microparticles increase the production of ROSs by +26.9% (p<0.01) and that 2% of the active ingredient according to the invention can forestall the effects of the pollution by significantly decreasing the production of oxidative radical species by -41.9% (p<0.01).5.2.1.2. Protein Nrf2Protocol

[0312] The effects of atmospheric pollutants on the synthesis of the protein Nrf2 were evaluated. The protocol used is the same as in section 5.1.1.2. and a solution of UMP or benzo[a]pyrene is contacted with the cultivated cells and labeled as before.Results

[0313] Variation of the translocation of Nrf2 as a function of time in a culture of HK. Effect of 3% of the active ingredient according to the invention, the atmospheric UMP solution and benzo[a]pyrene (B[a]P):

[0314] Control: contact with 0.1% DMSOContact with UMP;Contact with B[a]P;Contact with 3% of the active ingredient according to the inventionVariation (%); significanceVariation (%); significanceVariation (%); significanceAfter 2 hours of contactReference 1+136%; p<0.01+58%; p<0.05+41%; p<0.01After 24 hours of contactReference 2+401%; p<0.01+199%; p<0.01+36%; p<0.05

[0315] The results show that the translocation of the protein Nrf2 from the cytoplasm to the nucleus is increased significantly in the three cases, but that there are large differences in both time and intensity.

[0316] Regarding the atmospheric UMP, the results show a large increase from 2 hours of contact (+136%; p<0.01 relative to the control case) and a very large increase in the translocation after 24 hours of contact (+401%; p<0.01).

[0317] Regarding the benzo[a]pyrene, one of the most well-known atmospheric pollutants and carried by UMP, the results show a more moderate increase in the translocation, of +58% (p<0.05) after 2 hours of contact, and an exponential increase after 24 hours of contact (+199%; p<0.01).

[0318] Regarding the contact with the 3% of the active ingredient according to the invention, the results show an increase in the translocation of the protein Nrf2 of +41% (p<0.01) from 2 hours of contact and of +36% (p<0.05) after 24 hours of contact.

[0319] These results remain moderate relative to the other two cases (UMP and B[a]P), but significant relative to the control case. This underlines the controlled “powering up” of the detoxification system of the cells for the purpose of countering pollutants.5.2.2. Resistance to cellular hypoxia

[0320] 5.2.2.1.Mitochondrial potentialΔΨ,promotive forceProtocol

[0321] The effects of atmospheric pollutants on the mitochondrial potential ΔΨ were evaluated. The protocol used is the same as in section 5.1.2.1. and a solution of atmospheric UMP is contacted with the cultivated cells and labeled as before.Results

[0322] Variation of the mitochondrial potential ΔΨ in HK. Effect of 3% of the active ingredient according to the invention relative to the control case with or without UMP (n=6):

[0323] Variation (%); significanceControl case without UMPReference 1Control case with UMP-72.0%; p<0.01Reference 23% of the active ingredient according to the invention, with UMP+49.6%; p<0.01

[0324] The results show that the pollutant microparticles reduce the mitochondrial potential ΔΨ by -72% (p<0.01) and that 3% of the active ingredient according to the invention can forestall the effects of the pollution by significantly increasing the mitochondrial potential ΔΨ by +49.6% (p<0.01).

[0325] 5.2.2.2. Protectionofrespiration, consumption ofoxygenProtocol

[0326] The effects of atmospheric pollutants on the consumption of oxygen by the HK were evaluated. The protocol used is the same as in section 5.1.2.2. and a solution of atmospheric UMP is contacted with the cultivated cells as before.Results

[0327] Variation of the consumption of oxygen (oxygen pressure per minute: pO2) in HK. Effect of 3% of the active ingredient according to the invention relative to the control case with or without UMP (n=6):

[0328] Variation (%); significanceControl case without UMPReference 1Control case with UMP-29.8%; p<0.01Reference 23% of the active ingredient according to the invention, with UMP+24.2%; p<0.01

[0329] The results show that the pollutant microparticles reduce the consumption of oxygen by the cells by -29.8% (p<0.01) and that 3% of the active ingredient according to the invention can forestall the effects of the pollution by significantly increasing the cellular respiration by +24.2% (p<0.01).5.2.2.3. Protection of ATP productionProtocol

[0330] The effects of atmospheric pollutants on the synthesis of ATP by the HK were evaluated. The protocol used was the same as in section 5.1.2.3. and a solution of atmospheric UMP was contacted with the cultivated cells as before.Results

[0331] Variation of the synthesis of ATP in HK. Effect of 3% of the mixture of oligosaccharides relative to the control case with or without UMP (n=6):

[0332] Variation (%); significanceControl case without UMPReference 1Control case with UMP-21%; p<0.01Reference 23% of the active ingredient according to the invention, with UMP+74%; p<0.01

[0333] The results show that the pollutant microparticles reduce the production of ATP by -21% (p<0.01) and that 3% of the active ingredient according to the invention can forestall the effects of the pollution by significantly increasing the synthesis of ATP by +74% (p<0.01).5.2.2.4. Conclusion

[0334] All thein vitroresults presented above show that the mixture of oligosaccharides according to the invention protects the cells against oxidation and hypoxia, with the aim of detoxifying the cells from the effects of micropollutants.5.2.3. Reinforcing the skin barrier5.2.3.1. Thickness of the stratum corneumProtocol

[0335] The effects of atmospheric pollutants on the thickness of thestratum corneumare evaluated. The protocol used is the same as in section 5.1.3.1. and a solution of atmospheric UMP is contacted with equivalent skins.Results

[0336] Variation of the thickness of thestratum corneum. Effect of 3% of the active ingredient according to the invention (n=4):

[0337] Variation (%); significanceControl case without UMPReference 1Control case with UMP-61.3%; p<0.01Reference 23% of the active ingredient according to the invention, with UMP+136.4%; p<0.01

[0338] The results show that the pollutant microparticles reduce the thickness of thestratum corneumby -61.3% (p<0.01) and that 3% of the active ingredient according to the invention can forestall the effects of the pollution by significantly increasing the thickness of thestratum corneumby +136.4% (p<0.01). This demonstrates the positive effect of the ingredient according to the invention against the harmful effects of pollution.

[0339] 5.3. Comparative results

[0340] The monosaccharides and monosaccharide acids in the mixture of oligosaccharides, namely N-acetylquinovosamine, galacturonic acid, glucuronic acid and rhamnose, were tested alone and as a mixture to enable comparison with the active ingredient according to the invention. The amounts tested are:

[0341] - 50 ppm of rhamnose;

[0342] - 50 ppm of N-acetylquinovosamine;

[0343] - 50 ppm of galacturonic acid;

[0344] - 50 ppm of glucuronic acid;

[0345] - in the mixture of the 3 saccharides: 50 ppm of N-acetylquinovosamine, 50 ppm of galacturonic acid, and 50 ppm of glucuronic acid;

[0346] - in the mixture of the 4 saccharides: 50 ppm of N-acetylquinovosamine, 50 ppm of rhamnose, 50 ppm of galacturonic acid and 50 ppm of glucuronic acid.5.3.1. Synthesis of ATPProtocol

[0347] The protocol used was the same as in section 5.1.2.3.Results

[0348] Variation of the synthesis of ATP in HK. Comparison of the effects of saccharides alone or as a mixture and of 3% of the mixture of oligosaccharides according to the invention, relative to the control case (n=8):

[0349] Effect on production of ATPControl caseReference50 ppm of rhamnose050 ppm of glucuronic acid050 ppm of galacturonic acid050 ppm of N-acetylquinovosamine0Mixture of the 3 saccharides03% of the active ingredient according to the invention++

[0350] The results show that the monosaccharides, alone or combined, have no effect on the production of ATP, by contrast with the active ingredient according to the invention. It is therefore the oligosaccharides according to the invention, i.e., the monosaccharides bonded to one another, which act on the production of ATP.5.3.2. Kinetics of S. epidermidisProtocol

[0351] The protocol used was the same as in section 5.1.6.Results

[0352] Kinetics of growth ofS. epidermidisover time. Comparison of the effects of the saccharides alone or as a mixture and of 3% of the mixture of oligosaccharides relative to the control case (n= 7):

[0353] Effect on the production ofStaphylococcus epidermidisControl caseReference50 ppm of rhamnose050 ppm of glucuronic acid050 ppm of galacturonic acid050 ppm of N-acetylquinovosamine0Mixture of the 4 saccharides03% of the active ingredient according to the invention++

[0354] The results show that the monosaccharides, alone or combined, have no effect on the production ofStaphylococcus epidermidis, by contrast with the active ingredient according to the invention. It is therefore certainly the oligosaccharides according to the invention, i.e., the monosaccharides bonded to one another, which act on the growth ofS. epidermidis.6. Results of the in vivo testsTest product

[0355] The serum described in table 1 abovevs.a placebo serum, having the same base as the serum without the active ingredient according to the invention.Principle

[0356] The efficacy of the cosmetic treatment according to the invention was evaluated on a total of 181 volunteers in two complementary studies: one on a Caucasian panel and one on a Chinese panel. The panelists had to have a dull complexion and an irregular skin grain.

[0357] The metrological means (measurements) were evaluated by using expert analyses (judgements) and self-evaluations (perceived effects).Protocol

[0358] The study on the Caucasian volunteers was conducted in France on a maximum of 54 volunteers, who were women with an average age of 38 years (20 – 50 years).

[0359] The study on the Chinese volunteers was conducted in China on a maximum of 127 volunteers, men and women, with an average age of 35 years (18 – 50 years).Type of study, duration, applications

[0360] The volunteers did not have knowledge of the type of product (active ingredient according to the invention or placebo).

[0361] The panel of the 54 Caucasian women was divided into two and various measurements and acquisitions were performed before and after the applications, according to table 25 below:

[0362] T0 (D0)T = 28 days (D28)T = 56 days (D56)Evaluation of radiancePhotosPhotos / Evaluation of microreliefFringe projectionFringe projection / Evaluation of moisturizationMoistureMeter-D™MoistureMeter-D™MoistureMeter-D™Evaluation of firmnessCutometer® MPA580 / Cutometer® MPA580Self-evaluation / / Questionnaire

[0363] The panel of 127 Chinese volunteers applied four cloth masks impregnated with the serum according to the invention in 14 days for 15 minutes. The perception of the effect of the serum according to the invention in the mask was evaluated by means of a self-evaluation questionnaire after the first and after the fourth and last applications.Statistics

[0364] For the instrumental parameters, the statistical calculation is made with the aid of a Studentttest or, if necessary, with a non-parametric Wilcoxon or Mann-Whitney test. Unilateral tests were performed on unpaired series.

[0365] For the values in the questionnaire, the statistical calculation is made with the aid of a Khi test². The level of 0.05 is selected as a minimum acceptance point in the statistical significance.6.1. Evaluation of the radiance of the complexion

[0366] The skin radiance index (SRI) is based on a calculation which takes into account four zones of the face, namely the forehead, the nose, the areas under the eyes, and the cheeks, and three colorimetric components, namely L*, which represents the lightness or brightness, H, which represents the hue of a color, and IWA (individual white angle), which is an index developed by the Newtone Technologies company for evaluating the whiteness of the skin, taking into account the parameters L*, a* and b* (CIELAB or CIELCH color space).Protocol

[0367] A Headscan V05TMphotographic bench (Orion Concept, France) is used to acquire photographs of the face in cross-polarized mode to withdraw the natural shine.

[0368] For this evaluation, only 52 Caucasian volunteers took part: 26 applied the serum containing the active ingredient according to the invention, and 26 applied the placebo serum. The results below are an average of the measurements carried out on the various parts of the face.Results

[0369] Variation of the skin radiance index after 28 days of application. Effect of 3% of the active ingredient according to the invention relative to the placebo (N=52):

[0370] Serum according to the inventionPlacebo serumVariation between D0 and D28 (%); significance+3.1%; p<0.05-0.2%; dns*% improvement of the serum according to the inventionvs.the placebo serum; significancevs. placebo+3.3%; p<0.05

[0371] * dns: data not significant

[0372] The results show that the skin radiance index is increased significantly with the serum according to the invention (+3.3%; p<0.05vs. placebo). This shows that the ingredient according to the invention can improve the radiance of the skin.6.2. Evaluation of the microrelief

[0373] A number of parameters are studied to demonstrate the advantage of the active ingredient according to the invention for the skin microrelief.

[0374] The first parameter is called complexity. It represents the ratio between the actual surface and the ideal apparent surface.

[0375] The second parameter is called maximum amplitude. It corresponds to the average height of the highest reliefs. It conveys the changes to the skin.

[0376] The third parameter is called roughness. It corresponds to the average roughness of the facial skin. The greater the roughness, the less smooth the skin.Protocol

[0377] Line shadows (fringes) on the face were projected and the corresponding deformations, which are linked to the relief of the skin, were measured by triangulation. This technology allows the relief to be reconstituted in three dimensions.

[0378] For this evaluation, only 53 Caucasian volunteers were included: 26 applied the serum containing the active ingredient according to the invention, and 27 applied the placebo serum.

[0379] The measurements were carried out on the cheek and surface roughness was analyzed on the basis of the acquisitions to enable the more or less smooth appearance of the skin grain (microrelief, pores, fine micro-lines) to be qualified. The results below are an average of the various measurements carried out.Results

[0380] Variation of the complexity, maximum amplitude and roughness after 28 days of application. Effect of 3% of the active ingredient according to the invention relative to the placebo (N=53):

[0381] % variation between D0 and D28; significancevs. D0% improvement of the serum according to the inventionvs.the placebo serum; significancevs. placeboComplexitySerum according to invention-8%; p<0.01-5.7%; p<0.05Placebo serum-2.3%; dnsMaximum amplitudeSerum according to invention-4.9%; p<0.01-4.1%; p<0.05Placebo serum-0.8%; dnsRoughnessSerum according to invention-3.8%; p<0.01-3.2%; p<0.05Placebo serum-0.6%; dns

[0382] The results show that the serum according to the invention significantly reduces the complexity, maximum amplitude and roughness of the skin after 28 days of application. A decrease in these three parameters conveys a smoothing and a facial skin which is more luminous and more radiant.6.3. Evaluation of the moisturization

[0383] The moisturization of the skin is involved in the formation and maintenance of an effective skin barrier against external aggressions, to counter allergens, to limit water loss, to make the skin stronger, smoother and firmer, and is involved in skin radiance.Protocol

[0384] The MoistureMeter-D™ is an apparatus for measuring the quantity of cutaneous water via the principle of impedancemetry.

[0385] In this study, 6 zones were studied in parallel: forehead, temples, cheekbones, cheeks, jowls, chin. The results below are an average of the various measurements carried out and serve to express the overall moisturization of the face of the volunteers. The measurements were carried out a minimum of 12 hours after the last application.

[0386] For this evaluation, the 54 Caucasian volunteers were included: 27 applied the serum containing the active ingredient according to the invention, and 27 applied the placebo serum. Among the 54 volunteers, 24 have dry skin: 13 applied the serum containing the active ingredient according to the invention, and 11 applied the placebo serum.Results

[0387] Variation of moisturization after 28 days and 56 days of application. Effect of 3% of the active ingredient according to the invention relative to the placebo:

[0388] All skins (N=54) Dry skins (N=24)Serum according to inventionPlacebo serumSerum according to inventionPlacebo serum% variation between D28 and D0; significancevs. D0+8.4%; p<0.010%; dns+13.9%; p<0.01+4.5%; dns% improvement of the serum according to the inventionvs.the placebo serum; significancevs. placebo+8.4%; p<0.01+9.4%; p<0.05% variation between D56 and D0; significancevs. D0+15.0%; p<0.01-0.3%; dns+21.4%; p<0.01+6.6%; p<0.05% improvement of the serum according to the invention vs. the placebo serum; significancevs. placebo+15.3%; p<0.01+14.8%; p<0.01

[0389] The results show that the serum according to the invention significantly increases the moisturization of the skin after 28 days and 56 days of application, relative to the placebo. Moreover, the extraction of the “dry skins” shows that the increased moisturizing over the long term of serum according to the invention also increases significantly.

[0390] Since the measurements were carried out a minimum of 12 hours after the last application of the product on the skin, the results show a genuine beneficial effect of the moisturization of the skin over the long term.6.4. Evaluation of skin firmness

[0391] Skin firmness is one of the most looked-for parameters, as it is associated with youthfulness, the strength of the skin, and its tonicity.Protocol

[0392] The Cutometer® is used to measure the viscoelastic parameters of the skin. This instrument measures the deformation of a zone of skin which is subjected to repeated mechanical suction stresses, and its power of recovery. The instrument supplies graphs of deformation of the skin as a function of time, as shown in [Figure 1]. In this [Figure 1], the parameters Uf and R10 are indicated, respectively representing the extensibility and the “fatigability” of the skin.

[0393] In this study, 3 measurements were performed on the right and left sides of the face. The results for extensibility are an average of the various measurements carried out, and the results for the power of recovery are an average of the last measurements performed on the right and left sides of the face.

[0394] For this evaluation, the 54 Caucasian volunteers were included: 27 applied the serum containing the active ingredient according to the invention, and 27 applied the placebo serum.Results

[0395] Variation of the extensibility (Uf) and of the power of recovery (R10) after 56 days of application (N=54). Effect of the 3% of the active ingredient according to the invention relative to the placebo:

[0396] UfR10Serum according to inventionPlacebo serumSerum according to inventionPlacebo serum% variation between D56 and D0; significancevs. D0-8.0%; p<0.01-2.3%; dns-4.7%; p<0.05-0.2%; dns% improvement of the serum according to the inventionvs.the placebo serum; significancevs. placebo-5.7%; p<0.05-4.5%; p<0.06

[0397] The results show that the serum according to the invention significantly reduces the extensibility and the skin fatigue, thus demonstrating an increase in the firmness of the skin.

[0398] 6.5. Perceptionof the effects of the ingredient according to the invention6.5.1. In the Caucasian panelProtocol

[0399] The opinions of the volunteers were collected by means of a self-evaluation questionnaire after 56 days of application of the serum containing the active ingredient according to the invention or of the placebo serum.Results

[0400] Number of favorable opinions for each statement. Effect of 3% of the active ingredient according to the invention relative to the placebo:

[0401] % Number of favorable opinions for the serum according to invention; significance% Number of favorable opinions for the placebo serum; significance% Number of favorable opinions for the serum according to invention vs.placebo serum; significancevs. placeboMy skin is oxygenated and purified67%; p<0.0546%; dns+11%; p<0.05The product moisturizes immediately and durably83%; p<0.0169%; p<0.005+14%; p<0.08My skin looks prettier89%; p<0.0167%; p<0.005+22%; p<0.05

[0402] The results show that the serum according to the invention has significantly more favorable perceived effects relative to the placebo serum. The panelists find that their skin is more oxygenated, purified and moisturized immediately and durably, and looks prettier.6.5.2. In the Chinese panelProtocol

[0403] The opinions of the volunteers were collected by means of a self-evaluation questionnaire after 56 days of application of a cloth face mask impregnated with serum containing the active ingredient according to the invention or with placebo serum.Results

[0404] Number of favorable opinions for the serum according to the invention:

[0405] % Favorable opinions after 1stapplication% Favorable opinions after 4thapplication% Favorable opinions after 1stvs.4thapplicationMy complexion is less dull78%91%+13%My complexion is brighter82%92%+10%My skin is firmer83%89%+6%My skin is softer90%94%+4%My skin is more moisturized90%95%+5%My skin is fresher87%93%+6%The skin is revitalized / energized87%91%+4%My skin is purified87%91%+4%I look younger77%89%+12%I look less tired89%96%+7%

[0406] The results show that the perception of the quality of the skin is improved after four applications at short intervals of time of a cloth mask impregnated with the serum according to the invention.6.6. Conclusion

[0407] The entirety of thein vivoresults presented above show that the treatment according to the invention advantageously reinforces the radiant appearance of the skin, causes it to appear more luminous and reinvigorated, moisturizes it, smooths the relief, reinforces its firmness, and reduces skin fatigue.

Claims

Exopolysaccharide characterized in that it comprises at least one monosaccharide unit that is of N-acetyl-quinovosamine.Exopolysaccharide according to the claim 1, characterized in that it further comprises at least one monosaccharide acid unit selected from glucuronic acid and galacturonic acid.Exopolysaccharide according to claim 1 or 2, characterized in that it further comprises at least one monosaccharide unit that is of rhamnose.Exopolysaccharide according to anyone of claims 1 to 3, comprising between 15% to 55% of N-acetyl-quinovosamine by weight relative to the total weight of the exopolysaccharide.Exopolysaccharide according to anyone of claims 1 to 4, comprising between 15% to 55% of glucuronic acid and / or between 15% to 55% of galacturonic acid, by weight relative to the total weight of the exopolysaccharide.Exopolysaccharide according to anyone of claims 1 to 5, comprising between 15% to 55% of rhamnose by weight relative to the total weight of the exopolysaccharide.Exopolysaccharide according to anyone of claims 1 to 6, composed of an equimolar mixture of N-acetyl-quinovosamine, glucuronic acid, galacturonic acid and rhamnose.Exopolysaccharide according to anyone of claims 1 to 7, characterized in that it comprises the concatenation of three units comprising N-acetyl-quinovosamine and two monosaccharide acid units selected from glucuronic acid and galacturonic acid.Exopolysaccharide according to the claim 8, characterized in that it further comprises rhamnose.Exopolysaccharide according to anyone of preceding claims, characterized in that it has a molecular weight greater than or equal to 260,000 Da.Exopolysaccharide according to anyone of preceding claims, characterized in that it is obtained from the bacterial strainPseudoalteromonas sp. deposited at the Collection Nationale de Culture de Microorganismes (CNCM) at Paris under number I-5893.Oligosaccharide that can be obtained in particular by cleavage of the exopolysaccharide according to anyone of preceding claims, characterized in that it comprises the N-acetyl-quinovosamine.Oligosaccharide according to claim 12, characterized in that it has a degree of polymerization of between 2 and 12.Oligosaccharide according to claims 12 or 13, characterized in that further comprises at least two monosaccharidic acid units selected from glucuronic acid and galacturonic acid.Oligosaccharide according to anyone of claims 12 to 14, characterized in that it consists of N-acetyl-quinovosamine, glucuronic acid and galacturonic acid.Oligosaccharide according to anyone of claims 12 to 15, characterized in that it comprises the concatenation glucuronic acid, galacturonic acid and N-acetyl-quinovosamine.Process for obtaining the exopolysaccharide according to anyone of claims 1 to 11, comprising the following steps:- A first step of culturing a bacterial strain of the genusPseudoalteromonas sp. deposited at the Collection Nationale de Culture de Microorganismes (CNCM) at Paris under number I-5893 in a culture medium adapted to the production of the bacterium excreting the exopolysaccharide in the culture medium- A second step of recovering the exopolysaccharide separated from the culture medium.Process according to claim 17, characterized in that the exopolysaccharide present in the culture medium is separated from the solid biomass by a process enabling separation of a liquid phase from a solid.Process according to claim 18, characterized in that the exopolysaccharide is separated from the culture medium by a tangential filtration.Process according to anyone of claims 17 to 19, characterized by an additional step of cleaving the exopolysaccharide in at least one oligosaccharide according to anyone of claims 12 to 16.Process according to claim 20, characterized in that the cleavage is carried out by acid hydrolysis.Oligosaccharides mixture that can be obtained by the process according to claim 20 or 21 is composed mainly of trisaccharides and / or heptasaccharides.Oligosaccharides mixtures according to claim 22, characterized in that the trisaccharides have the concatenation: glucuronic acid, galacturonic acid and N-acetyl-quinovosamine (Glu-Gal-NAcQ).Oligosaccharides mixtures according to claim 22, characterized in that the heptasaccharides have the concatenation: glucuronic acid, galacturonic acid, N-acetyl-quinovosamine, rhamnose, glucuronic acid, galacturonic acid and N-acetyl-quinovosamine (Glu-Gal-NAcQ-Rham-Glu-Gal-NAcQ).Bacterial strain of the genusPseudoalteromonas sp. deposited at the Collection Nationale de Culture de Microorganismes (CNCM) at Paris under number I-5893.Cosmetic or dermatological composition comprising an exopolysaccharide according to anyone of claims 1 to 11 and a physiologically acceptable medium.Cosmetic or dermatological composition comprising at least one oligosaccharide according to anyone of claims 12 to 16 or an oligosaccharides mixture according to anyone of claims 22 to 24 and a physiologically acceptable medium.Use of the exopolysaccharide according to anyone of claims 1 to 11 or a composition containing it according to claim 26, in industrial sectors such as the chemical, food, cosmetics, pharmaceutical, veterinary, agricultural, environmental or paint and coatings industries.Use of at least one oligosaccharide according to anyone of claim 12 to 16 or oligosaccharides mixture according to anyone of claims 22 to 24 or a composition containing it according to claim 27, for non-therapeutic cosmetic treatment of the skin and its appendages.Use according to claim 28 or 29, characterized in that the cosmetic treatment is topical.Use according to claim 29 or 30, characterized in that the cosmetic treatment is adapted to:- improve the radiance of the complexion;- improve the relief of the skin;- improve the firmness of the skin; and / or- improve the moisturization of the skin.Use according to anyone of claims 29 to 31, characterized in that the cosmetic treatment is a treatment that protects the skin from ageing and / or pollution, particularly atmospheric pollution.

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