METHOD FOR APPLYING A NATURAL EXTRACT TO THE SKIN TO ACHIEVE BIOSYNTHESIS OF METABOLITES OF INTEREST AT THE LEVEL OF THE SKIN MICROBIOTA

Abstract

The invention relates to a cosmetic and / or dermo-cosmetic application method for the skin, comprising the application of a natural extract to the skin surface to induce biosynthesis in the skin microbiota. This biosynthesis induces the production of one or more metabolites of interest by the skin microbiota. The metabolite(s) of interest, associated with the applied natural extract, induce a beneficial effect on the skin. FIGURE: 3

Description

Title of the invention: METHOD FOR APPLYING A NATURAL EXTRACT TO THE SKIN TO PRODUCE A BIOSYNTHESIS OF METABOLITES OF INTEREST AT THE LEVEL OF THE SKIN MICROBIOTA Technical field of the invention

[0001] The field of the invention is that of cosmetology and dermo-cosmetology. More specifically, the invention relates to a method for applying a natural extract to the surface of the skin to carry out biosynthesis of metabolites of interest at the level of the cutaneous microbiota, alone or in association with the skin cells, the association of the active extract and the metabolites produced in situ inducing a beneficial effect for the skin. Technical background

[0002] Human skin is an organ that constitutes an interface with our external environment. It is composed of numerous molecules originating from host cells, the microbiota and external molecules, deliberately or not applied to the skin. All the metabolites, which are the end products of cellular processes, constitute the metabolome. The metabolome is thus composed of a large number of small metabolites, from primary metabolites, such as sugars, amino acids, fatty acids, or lipids, to secondary metabolites, such as polyphenols, alkaloids, drugs, or toxins.

[0003] In particular, the quantity and heterogeneity of skin metabolites are influenced by the functioning of the skin microbiota, as well as by the lifestyle and the environment surrounding and affecting this skin microbiota: diet, consumption of medication and / or tobacco, the composition of the outside air, or even the composition of clothing are all factors that can interfere with the composition of these metabolites produced by the skin microbiota. Cosmetic ingredients applied to the skin, and therefore to the skin microbiota, are also part of these factors.

[0004] The skilled person knows how to detect and quantify cutaneous metabolites produced by the skin in response to an external aggression, such as contact with pesticide ingredients. Thus, for example, in the medical field, we are able to identify biomarkers indicating diseases, such as fibrosis, psoriasis, Parkinson's disease, or even certain cancers, and in particular by using a metabolomic approach.

[0005] However, until now, the impact of natural active ingredients applied to the skin has never been exploited or evaluated, in particular at the level of the skin microbiota, and a fortiori for a cosmetic or dermo-cosmetic application. Summary of the invention

[0006] The subject of the present invention is therefore a method of cosmetic and / or dermo-cosmetic application to the skin comprising the application of a natural extract to the surface of the skin to carry out biosynthesis at the level of the cutaneous microbiota, this biosynthesis inducing the production of one or more metabolites of interest by the cutaneous microbiota, this or these metabolites of interest associated with the natural extract applied, inducing a beneficial effect for the skin.

[0007] Such a method thus advantageously makes it possible to analyze the functioning of the skin microbiota, and has made it possible to determine that the application of a natural active cosmetic or dermo-cosmetic ingredient to the skin allows the bacteria of the microbiota to produce another active natural ingredient, distinct from the applied ingredient, in order to optimally meet a cosmetic need at the skin level. The bacteria of the skin microbiota recognize the applied active ingredient, they are in a way "activated" by this ingredient, which has the function of sending a signal to the bacteria. In response to this signal, the bacteria of the skin microbiota recognize the needs of the skin and consequently produce, in situ, at least one natural active metabolite, this metabolite being beneficial for the skin.The skin microbiota is therefore a target for cosmetic and dermo-cosmetic ingredients: it allows the implementation of biosynthesis, which corresponds to the production at the level of the skin microbiota of an active natural ingredient in response to the topical application of a distinct active natural ingredient. The presence at the level of the skin of these two active natural ingredients, one applied and the other produced, makes it possible to produce a cosmetic effect of better quality / more effective than in the presence of only one of these ingredients. The active natural ingredient acts directly on the skin microbiota but also on the skin, because it allows the microbiota to produce a metabolite of interest to generate a directly measurable cosmetic effect on the skin.

[0008] For the purposes of the present invention, skin means not only human skin, but also the scalp and lips. The method according to the invention can also produce cosmetic and dermo-cosmetic effects on the appendages, for example on the nails and hair.

[0009] For the purposes of the present invention, the term metabolite means any compound which can be produced by a skin microbiota and which can have a beneficial effect on the skin (both qualitatively and in terms of appearance), for example a peptide or a vitamin.

[0010] Such a process also makes it possible to obtain a cosmetic or dermo-cosmetic effect. beneficial for the skin in a natural, economical and environmentally friendly way: the active metabolite(s) are produced in situ by the skin microbiota, so there is no need to apply synthetic ingredients to the skin, which are expensive, potentially irritating to the skin and harmful to the environment. In addition, it is the combination of the natural ingredient applied initially and the metabolite produced naturally by the skin microbiota that will produce the beneficial effect for the skin: there is therefore no need to apply the initial ingredient to the skin in high proportions, which would exceed cosmetically acceptable doses.

[0011] Such a method is furthermore advantageously non-invasive and easy to implement.

[0012] Advantageously, the biosynthesis implemented in this method according to the invention is analyzed and quantified according to the following steps:

[0013] - A) a qualitative analysis to identify the metabolite of interest produced by the mi cutaneous microbiota in response to topical application of the natural extract;

[0014] - B) an assay of the production of the metabolite of interest identified in step A by the skin microbiota, comprising the collection of at least one sample of skin microbiota; then bringing it into contact with the natural extract and quantifying the metabolite of interest in the skin microbiota sample.

[0015] Such a method according to the invention therefore makes it possible to reliably characterize the metabolites of interest which are biosynthesized by the cutaneous microbiota in response to the topical application of an active natural extract, because in a first step, this method is based on the identification of the metabolite(s), and in a second step, it confirms this identification by being capable of quantitatively dosing this or these identified metabolites.

[0016] According to one embodiment, the qualitative analysis A) for identifying the metabolite of interest is a metabolomic analysis.

[0017] Metabolomics makes it possible to effectively evaluate the effectiveness of active cosmetic ingredients on the skin. It allows the skin metabolome to be characterized as broadly as possible. Metabolomic studies are currently carried out by combining liquid chromatography, mass spectrometry and / or nuclear magnetic resonance. It is therefore a simple method to implement, which implements analytical techniques well known to those skilled in the art.

[0018] It is understood that the identification and quantification of metabolites of interest at the level of the skin microbiota are not limited to a study by metabolomics, and that any other analysis technique allowing such identification / quantification to be carried out can be used.

[0019] Advantageously, the natural extract is a plant extract or an extract of a product derived from a bacterium.

[0020] The method according to the invention thus applies to numerous cosmetic or dermo-cosmetic extracts, comprising active ingredients from very varied sources.

[0021] According to one embodiment, the metabolite of interest produced by the skin microbiota is a small organic molecule chosen from vitamins, carotenoids, amino acids, peptides, glycopeptides, lipids, glycolipids, nucleic acids, metabolites of phenol type such as flavonoids, or of nitrogen type such as glycosinolates, or of terpene type such as monoterpenes, diterpenes and triterpenes, indolamines such as serotonin and catecholamines such as dopamine.

[0022] According to a preferred embodiment, the metabolite of interest produced by the skin microbiota is a vitamin and / or an intermediate in the production of a vitamin, such a vitamin preferably belonging to the group of B vitamins.

[0023] With age, the human body finds it increasingly difficult to absorb and metabolize vitamins. In particular, B vitamins are important for combating cognitive decline, osteoporosis and for protecting the cardiovascular system. The method according to the invention therefore allows the biosynthesis of metabolites of interest to the human body, which are particularly sought after by users of cosmetic and dermo-cosmetic products.

[0024] According to a preferred embodiment, this vitamin is chosen from vitamin B3 and vitamin B5.

[0025] Vitamin B3 advantageously exhibits anti-inflammatory, antioxidant, anti-microbial, lightening, photoprotective, and skin barrier protective effects.

[0026] Vitamin B5, for its part, is a cosmetic active ingredient that produces beneficial effects in terms of skin soothing, dermal remodeling and barrier function restoration. It has moisturizing, soothing and regenerating properties. It is also effective in combating hair loss. In addition, since vitamin B5 is unstable, it is generally not used as such in known cosmetic formulations, but in its stabilized form, which corresponds to panthenol. The concentrations of this stabilized ingredient derived from vitamin B5 are therefore very often high in these formulations, beyond cosmetically acceptable doses, which can lead to skin irritation.The invention, on the contrary, allows the in situ production of vitamin B5 as such, which then allows it to be used in physiologically and cosmetically acceptable concentrations in cosmetic formulations, that is to say in substantially lower concentrations compared to known and currently marketed formulations.

[0027] According to a first advantageous embodiment, the plant extract is an extract of a terrestrial plant, this terrestrial plant being obtained by harvesting, by cultivation or by biotechnology such as dedifferentiated plant cells.

[0028] According to a second advantageous embodiment, the plant extract is an extract of a marine plant, this marine plant being chosen from macroalgae and microalgae. Preferably, the marine plant extract is an extract of the macroalga Codium tomentosum, this extract being fermented in the presence of the bacterium Exiguobacterium marinum in seawater and stimulating the production of vitamin B5 by the skin microbiota.

[0029] According to a third advantageous embodiment, the extract of product derived from a bacterium is an extract of a product derived from a marine bacterium, this marine bacterium being chosen from cyanobacteria and bacteria producing polyhydroxyalcanoates and exopolysaccharides.

[0030] According to a fourth advantageous embodiment, the extract of product derived from a marine bacterium is an extract of polyhydroxyalkanoate produced by the bacterium CNCM 1-5658 and stimulates the production of an intermediate for the production of vitamin B3 by the skin microbiota.

[0031] According to a preferred embodiment, this intermediate for the production of vitamin B3 is the metabolite kynurenine.

[0032] The extract used in the process according to the invention can be of various natures, for example an aqueous, hydroglycolic, glycolic, oily extract. It can be obtained by various types of extraction, such as supercritical CO2 extraction or even cold extraction. It can also be fermented in the presence of bacteria, like the extract of the macroalga Codium tomentosum, fermented in the presence of the bacterium Exiguobacterium marinum.

[0033] According to one embodiment, the metabolite(s) of interest associated with the applied natural extract exhibit at least one of the following effects: hydration, and / or healing, and / or an anti-blemish effect, and / or a soothing effect, and / or a lightening effect, and / or an effect on the complexion, and / or an effect on the appearance of the skin, and / or an anti-wrinkle effect.

[0034] The method according to the invention thus makes it possible to understand how certain cosmetic effects are obtained at the skin level with active cosmetic ingredients, and what are the mechanisms of action at the skin level which allow such effects to be obtained.

[0035] Other advantages and particularities of the present invention will result from the description which follows, given by way of non-limiting example and made with reference to the figures and examples. Brief description of the figures

[0036] [Fig-1] - [Fig.l] is a diagram representing the concentration of vitamin B5 (in ppb) versus time (in hours) for three pre-cultured skin microbiota samples including different concentrations of fermented natural extract of the macroalga Codium tomentosum;

[0037] [Fig.2] - [Fig.2] is a diagram representing the concentration of vitamin B5 (in ppb) as a function of time (in hours) for a sample of wild skin microbiota comprising the natural fermented extract of the macroalga Codium tomentosum at 5% by weight;

[0038] [Fig.3] - [Fig.3] is a histogram representing the prosta-glandin-E2 concentration (in pg / mL) of different skin samples having been stressed with phorbol 12-myristate 13-acetate and comprising the natural fermented extract of the macroalga Codium tomentosum and / or synthetic vitamin B5, to analyze the anti-inflammatory properties of the effect of the algae extract;

[0039] [Fig.4] - [Fig.4] is a histogram representing the relative expression of the FLG gene for different skin samples comprising the natural fermented extract of the macroalga Codium tomentosum and / or synthetic vitamin B5, to analyze the effect of the algae extract on cell differentiation;

[0040] [Fig.5] - [Fig.5] is a histogram representing the relative expression of the IVL gene for different skin samples comprising the natural fermented extract of the macroalga Codium tomentosum and / or synthetic vitamin B5, to also analyze the effect of the algae extract on cell differentiation. EXAMPLES

[0041] EXAMPLE 1: application of a natural fermented extract of the macroalgae Codium tomentosum

[0042] Preparation of the extract

[0043] A macroalga Codium tomentosum was brought into contact with the bacterium Exiguobacterium marinum, which was isolated from the surface of the alga Codium tomentosum. This bacterium has properties of halotolerance (adaptability to environments with high salt concentrations), resistance to strong variations in pH and temperature, resistance to UV, and includes high levels of heavy metals, such as arsenic: it can therefore advantageously adapt to extreme environmental conditions. The macroalga / bacteria mixture was fermented in seawater. The culture medium was then filtered, and the retentate was mixed with pentylene glycol in a content of 10%, relative to the total weight of the mixture.

[0044] Non-targeted study on the skin

[0045] The natural fermented extract of the macroalgae Codium tomentosum described above was applied to the cheek of several volunteers in a concentration of 5% by weight, on a skin surface of 9 cm2, twice a day, for 28 days. A placebo was applied to the other cheek of the volunteers under the same conditions. After 28 days, skin samples from both cheeks of the volunteers were taken by swabs, on a skin surface of 2 cm2. These samples were then frozen at -80°C.

[0046] These samples were then thawed and prepared for chromatographic and spectrometric analysis. For this, they were rinsed several times with a 50 / 50 water / ethanol mixture by volume, with stirring, at a temperature of 4°C. After a centrifugation step, the supernatant of the centrifuged sample was recovered and analyzed by ultra-performance liquid chromatography coupled with a high-resolution mass spectrometer (UHPLC-MS / MS).

[0047] This study identified the production and inhibition of various metabolites in the skin by the application of the fermented natural extract of the macroalga Codium tomentosum. Among these metabolites, it was determined in particular that vitamin B5 was produced in response to the application of this algae extract, and that this production was doubled compared to the topical application of the placebo.

[0048] A targeted study was then conducted on the application of the fermented natural extract of the macroalga Codium tomentosum to the skin, in order to confirm that vitamin B5 is indeed produced in response to the topical application of this extract, and if necessary, to quantify it.

[0049] Targeted study on the skin

[0050] A composition comprising 5% by weight of the fermented natural extract of the macroalga Codium tomentosum was applied to the cheek of five volunteers, over a skin area of ​​9 cm2, twice a day, for 28 days. A placebo was applied to the other cheek of the volunteers under the same conditions. At the end of the 28 days, skin samples from both cheeks of the volunteers were taken by swabs, over a skin area of ​​4 cm2. These swabs were then rinsed with a 50 / 50 water / ethanol mixture by volume, and frozen at -80°C.

[0051] These samples were then thawed and prepared for chromatographic and spectrometric analysis. For this, the swabs were subjected to ultrasound in 1 mL of the 50 / 50 water / ethanol mixture by volume, for 20 min. After a centrifugation step, the supernatant of the centrifuged sample was recovered and analyzed by high-performance liquid chromatography coupled with a high-resolution mass spectrometer (HPLC-MS / MS).

[0052] This study confirmed that vitamin B5 is indeed produced in situ by the skin in response to the application of the natural fermented extract of the macroalgae Codium tomentosum. This study also made it possible to quantify vitamin B5.

[0053] Targeted studies on the skin microbiota were then carried out, in order to verify that the production of vitamin B5 is indeed done specifically by the bacteria of the skin microbiota.

[0054] Targeted study on pre-cultured skin microbiota

[0055] A swab was rinsed with a solution comprising 9 g / L of sodium chloride (NaCl) and 0.1% of polyoxyethylene sorbitan monolaurate (Tween20®). A sample of skin microbiota was taken by contact of this rinsed swab with the skin of the cheek of a volunteer. This swab was then placed in contact with a culture medium comprising water, 7 g / L of NaCl, 1 g / L of yeast extract, 1 g / L of wheat peptone and 1 g / L of sucrose, for a period of 24 hours, at a temperature of 35°C and with stirring, to allow the growth of a consortium of skin microbiota microorganisms. This culture was then placed in contact with sterile demineralized water for some samples, and with sterile demineralized water comprising either 2% by weight or 5% by weight of the natural fermented extract of the macroalga Codium tomentosum for other samples.These mixtures were then incubated at a temperature of 35°C with shaking for 144 hours. The growth of the skin microorganisms was assessed by measuring the optical density (OD) at a wavelength of 750 nm. Samples of these cultures were taken at different times and frozen at -20°C to monitor changes in vitamin B5 content over time.

[0056] These samples were then prepared for chromatographic and spectrometric analysis. For this, they were thawed and filtered at 0.2 pm, then analyzed by ultra-performance liquid chromatography coupled with a high-resolution mass spectrometer (UPLC-MS).

[0057] [Fig.l] illustrates the results of this study focused on pre-cultured skin microbiota. It corresponds to a diagram which represents the evolution of the concentration of vitamin B5, in parts per billion (usually designated by the acronym ppb, meaning "part per billion" in English), as a function of time, in hours, for a sample of pre-cultured microbiota which has not been put into contact with the fermented natural extract of the macroalga Codium tomentosum (curve with points), and for samples which have been put into contact with this algal extract in a concentration of 2% by weight (curve with squares) and 5% (curve with triangles) by weight.

[0058] Thus, it is clearly observable that the samples of skin microbiota in contact with the fermented natural extract of the macroalga Codium tomentosum produce vitamin B5 in greater quantity compared to the sample which was not put in contact with this extract. This production is observed over the entire duration of the study, that is to say from the time the microbiota is put in contact with the algae extract, until the end of the observation.

[0059] In addition, the sample brought into contact with the algae extract at a concentration of 5% by weight produce more vitamin B5 than the sample brought into contact with this extract at a concentration of 2% by weight: after 50 hours, the sample in the presence of 5% by weight of extract comprises approximately 930 ppb of vitamin B5, while the sample in the presence of 2% by weight of extract comprises approximately 900 ppb of vitamin B5; and after 150 hours, the sample in the presence of 5% by weight of extract comprises approximately 1000 ppb of vitamin B5, while the sample in the presence of 2% by weight of extract comprises approximately 950 ppb of vitamin B5. The production of vitamin B5 is therefore dependent on the concentration of the algae extract applied to the skin.

[0060] In addition, the production of vitamin B5 by the sample in the presence of 5% by weight of extract increases over time, with an increase of 42% after 144 hours. This result therefore tends to demonstrate that the natural fermented extract of the macroalga Codium tomentosum, when applied to the skin, acts continuously over time, allowing the biosynthesis of vitamin B5 by the skin microbiota also continuously over time.

[0061] Targeted study on the wild microbiota of the skin

[0062] A sample of skin microbiota was taken from the cheek skin of five volunteers under the same conditions as for the previous study on pre-cultured skin microbiota. These samples, which do not include skin cells, were not cultured, but directly placed in contact with sterile demineralized water comprising 5% by weight of the fermented natural extract of the macroalga Codium tomentosum. These mixtures were then incubated at a temperature of 35°C, with stirring, for a period of 72 hours. The growth of the skin microorganisms was evaluated by measuring the optical density (OD) at a wavelength of 750 nm. Samples of these cultures were taken at different times and frozen at -20°C, in order to monitor the evolution of the vitamin B5 content over time.

[0063] These samples were then prepared for chromatographic and spectrometric analysis under the same conditions as for the previous study on pre-cultured skin microbiota.

[0064] [Fig.2] illustrates the results of this study focused on the wild skin microbiota. Like [Fig.l] relating to the pre-cultured microbiota, this [Fig.2] corresponds to a diagram which represents the evolution of the concentration of vitamin B5, in ppb, as a function of time, in hours, for a sample of wild microbiota having been put in contact with the natural fermented extract of macroalga C odium tomentosum in a concentration of 5% by weight.

[0065] Thus, it is clearly observable that the sample of wild skin microbiota in contact with the algae extract also produces vitamin B5. In particular, a production peak at approximately 300 ppb can be observed after approximately 30 hours.

[0066] Therefore, these two studies targeted on the cutaneous microbiota (pre-cultured and wild) have therefore made it possible to demonstrate initially that the microbiota is capable of developing in the presence of the natural fermented extract of the macroalga Codium to-mentosum.

[0067] They also made it possible to demonstrate that vitamin B5, identified as being produced in situ by the skin in response to the topical application of Codium tomentosum extract, is specifically produced by the bacteria of the skin microbiota, since these studies relate to microbiota samples not comprising skin cells. Biosynthesis of this metabolite was therefore indeed implemented at the level of the skin microbiota, which recognized the active extract applied to the skin, and which responded by producing vitamin B5, necessary for the skin to meet certain needs.

[0068] In vitro study on the anti-inflammatory effects of the natural fermented extract of the macroalga Codium tomentosum.

[0069] Normal human dermal fibroblasts (usually referred to by the acronym NHDF) were obtained in the plastic surgery department of a local hospital following "abdominoplasty" type surgical procedures. These NHDFs are in particular derived from a 38-year-old woman. For the invention, they were used at passage 4, to reproduce normal skin. These NHDFs were seeded in a basal culture medium, usually referred to as "Dulbecco's Modified Eagle Medium" in English (known by the acronym DMEM), comprising 10% heat-inactivated fetal calf serum (usually referred to by the acronym FCS), for a period of 3 days, in a 24-multiwell plate. Following this culture period, the cells were either left untreated (control sample) or treated (test sample) with the natural fermented extract of the macroalga Codium tomentosum at a concentration of 1% by weight for a period of 16 hours.The samples were then mixed with phorbol 12-myristate 13-acetate (commonly referred to as PMA) and incubated for 24 hours. After this incubation period, the culture media were removed and frozen at -80°C.

[0070] Prostaglandin-E2 (usually referred to by the acronym PGE2) was measured in these samples. Indeed, this fatty acid plays an important role in the immune and inflammatory phenomena of the human body. Its study therefore makes it possible to analyze the anti-inflammatory effects of Codium tomentosum extract on the skin. PGE2 measurements were carried out using an ELISA assay kit, following the procedures recommended by the manufacturer (Enzo Lifescience, ref ADL901-001). The optical density was measured at a wavelength of 450 nm, with a correction at 540 nm using a spectrophotometer plate.

[0071] [Fig.3] illustrates the results of this study on the inflammatory effects of the extract of macroalga Codium tomentosum. It corresponds to a histogram which represents the concentration of PGE2, in pg / mL, for different samples: a sample of unstressed NHDF which has not been put in contact with the algae extract (black column of the histogram); a sample of NHDF which has been stressed with PMA at 0.1 qg / mL (column with vertical lines); two samples of control NHDF (flat column and column with dots); a sample of NHDF which has been stressed with PMA and put in contact with the algae extract at 1% by weight (squared column); a sample of NHDF which has been stressed with PMA and put in contact with synthetic vitamin B5 at 10 qg / mL (0.001%) (column with wavy lines); and a sample of NHDF stressed with PMA and contacted with 1% by weight seaweed extract and 10 μg / mL (0.001%) synthetic vitamin B5 (column with horizontal lines).

[0072] Thus, it is clearly observable that vitamin B5 acts in reducing skin inflammation: the samples comprising vitamin B5, whether synthetic or produced in situ in response to the presence of the Codium tomentosum extract, all have a lower PGE2 concentration than the samples stressed by PMA and not treated with the extract or with synthetic vitamin B5. Indeed, the PGE2 concentration is approximately 38,000 pg / mL for the sample only stressed by PMA, while this concentration is approximately 32,000 pg / mL for the sample stressed by PMA and put in contact with the 1% by weight algae extract, approximately 27,000 pg / mL for the sample stressed by PMA and put in contact with synthetic vitamin B5, and approximately 22,000 pg / mL for the sample stressed by PMA and put in contact with the 1% by weight algae extract and synthetic vitamin B5.

[0073] Furthermore, it can be observed that the sample having been put in contact with synthetic vitamin B5 and the algae extract has a significantly lower concentration of PGE2 compared to the samples having been treated either with the algae extract alone or with synthetic vitamin B5 alone: ​​this result clearly demonstrates that the application of the algae extract associated with the vitamin B5 bio-synthesized by the skin microbiota generates a synergy which produces an improved cosmetic effect, here anti-inflammatory, on the skin.

[0074] In vitro study on the effects of cell differentiation and proliferation of the natural fermented extract of the macroalga Codium tomentosum.

[0075] Normal human epidermal keratinocytes (usually referred to by the acronym NHEK) were obtained in the plastic surgery department of a local hospital following "abdominoplasty" type surgical procedures. These NHEK are in particular derived from a 27-year-old woman. For the invention, they were used in passage 3. They were seeded in EpiLife™ medium (Gibco™) for a duration of 4 days, in a 24-multiwell plate. Following this culture period, the cells were either untreated (control sample) or treated with the natural fermented extract of the macroalga Codium tomentosum at a concentration of 0.5%, by weight for a period of 72 hours.

[0076] After extraction and quantification of RNA, polymerase chain reactions (usually referred to by the acronym PCR, meaning “Polymerase Chain Reaction” in English) were carried out.

[0077] Figures 4 and 5 illustrate the results of this study on the effects of the extract of the macroalga Codium tomentosum on the expression of genes coding for cell differentiation and for cell proliferation markers.

[0078] [Fig.4] is a histogram that represents the relative expression of the FLG gene, which codes for the cell differentiation process, for different samples: an untreated NHEK sample (black column of the histogram); an NHEK sample that has been contacted with 0.5% by weight seaweed extract (checkered column); an NHEK sample that has been contacted with 1 μg / mL synthetic vitamin B5 (column with wavy lines); and an NHEK sample that has been contacted with 0.5% by weight seaweed extract and 1 μg / mL synthetic vitamin B5 (column with horizontal lines).

[0079] [Fig.5] corresponds to a similar histogram relating to the IVL gene, which also codes for the process of cellular differentiation.

[0080] Thus, it is clearly observable that vitamin B5 acts in the stimulation of cell differentiation in the skin: the samples comprising vitamin B5, whether synthetic or produced in situ in response to the presence of the fermented natural extract of the macroalga Codium tomentosum, almost all stimulate the expression of the FLG and IVL genes. Concerning the FLG gene, a respective increase in its expression of 201% is observed for the sample in contact with the algae extract alone, of 61% for the sample in contact with the synthetic vitamin B5 alone, and of 300% for the sample in contact with the algae extract and the synthetic vitamin B5, compared to the sample that has not been treated.Concerning the IVL gene, a respective increase in its expression of 836% was observed for the sample in contact with the algae extract alone, and of 1045% for the sample in contact with the algae extract and synthetic vitamin B5, compared to the sample that had not been treated.

[0081] Furthermore, it can be observed that the sample having been put in contact with synthetic vitamin B5 and algae extract stimulates the expression of the FLG and IVL genes to a significantly greater extent compared to the samples having been treated either with algae extract alone or with synthetic vitamin B5 alone: ​​this result clearly demonstrates that the application of the algae extract associated with the vitamin B5 biosynthesized by the skin microbiota generates a synergy which produces a cosmetic effect, here stimulation of cell differentiation, improved on the skin.

[0082] All of these studies relating to the topical application of the fermented natural extract of the macroalga Codium tomentosum have made it possible to demonstrate that it advantageously induces the biosynthesis of vitamin B5 specifically at the level of the cutaneous microbiota, and that the association of this active natural extract with the natural vitamin B5 produced in situ makes it possible to produce an improved cosmetic effect on the skin, compared to the presence on the skin of the extract alone, or of the metabolite alone.

[0083] EXAMPLE 2: application of a natural extract of poly(3-hydroxyoctanoate) (PHO) from the marine bacterium CNCM-I 5658

[0084] A poly(3-hydroxyoctanoate) (usually designated by the acronym PHO) was obtained by fermentation of the bacterium CNCM 1-5658 in a suitable culture medium. After isolation and purification, it was diluted in glycerol triheptanoate (usually designated by the acronym THG), at a content of 5% by weight, to obtain a natural extract of PHO.

[0085] The bacterial strain CNCM 1-5658, living in the waters bordering the department of Finistère (France), was deposited according to the Budapest Treaty, at the National Collection of Cultures of Microorganisms (usually designated by the acronym CNCM), by the company Polymaris Biotechnology.

[0086] This PHO has already been used in the field of cosmetics. Thus, FR3121353A1 proposes makeup compositions, such as foundation, incorporating this polyhydroxyalkanoate for its film-forming properties. FR31213525A1 also discloses its use for improving the skin's barrier function, for moisturizing the skin, or for its antibacterial effect on the skin.

[0087] Such an extract was then applied to the cheek of several volunteers, over an area of ​​9 cm2 of skin, twice a day, for 28 days. A placebo was applied to the other cheek of the volunteers under the same conditions.

[0088] A metabolomics-based analysis identified that the application of such a PHO extract leads to the biosynthesis of kynurenine, which is an intermediate in the production of vitamin B3. This production is increased twofold compared to the topical application of the placebo.

Claims

Claims

1. Method of cosmetic and / or dermo-cosmetic application to the skin comprising the application of a natural extract to the surface of the skin to induce biosynthesis at the level of the skin microbiota, said biosynthesis inducing the production of one or more metabolites of interest by the skin microbiota, alone or in association with skin cells, said metabolite(s) of interest associated with said applied natural extract, inducing a beneficial effect for the skin.

2. Application method according to claim 1, wherein said biosynthesis is analyzed and quantified according to the following steps: - A) a qualitative analysis to identify said metabolite of interest produced by the skin microbiota in response to the topical application of said natural extract; - B) an assay of the production of said metabolite of interest identified in step A by the skin microbiota, comprising the collection of at least one sample of skin microbiota; then bringing it into contact with said natural extract and the quantification of said metabolite of interest in said sample of skin microbiota.

3. Application method according to claim 2, in which the qualitative analysis A) for identifying said metabolite of interest is a metabolomic analysis.

4. Application method according to any one of claims 1 to 3, wherein said natural extract is a plant extract or an extract of a product derived from a bacterium.

5. Method according to any one of claims 1 to 4, according to which said metabolite of interest produced by the skin microbiota is a small organic molecule chosen from vitamins, carotenoids, amino acids, peptides, glycopeptides, lipids, glycolipids, nucleic acids, metabolites of phenol type such as flavonoids, or of nitrogen type such as glycosinolates, or of terpene type such as monoterpenes, diterpenes and triterpenes, indolamines such as serotonin and catecholamines such as dopamine.

6. Method according to claim 5, according to which said metabolite of interest produced by the skin microbiota is a vitamin and / or an intermediate in the production of a vitamin, said vitamin preferably belonging to the group of B vitamins.

7. A method according to claim 6, wherein said vitamin is selected from vitamin B3 and vitamin B5.

8. A method according to any one of claims 1 to 7, wherein said plant extract is an extract of a terrestrial plant, said terrestrial plant being obtained by harvesting, cultivation or biotechnology such as dedifferentiated plant cells.

9. A method according to any one of claims 1 to 7, wherein said plant extract is an extract of a marine plant, said marine plant being selected from macroalgae and microalgae.

10. The method of claim 9, wherein said marine plant extract is an extract of the macroalga Codium tomentosum, said extract being fermented in the presence of the bacterium Exiguobacterium marinum in seawater and stimulating the production of vitamin B5 by the skin microbiota.

11. The method of any one of claims 1 to 7, wherein said extract of product from a bacterium is an extract of a product from a marine bacterium, said marine bacterium being selected from cyanobacteria and bacteria producing polyhydroxyalkanoates and exopolysaccharides.

12. The method of claim 11, wherein said marine bacterium product extract is a polyhydroxyalkanoate extract produced by the CNCM-I5658 bacterium and stimulates the production of a vitamin B3 production intermediate by the skin microbiota.

13. The method of claim 12, wherein said vitamin B 3 production intermediate is the metabolite kynurenine.

14. Method according to any one of claims 1 to 13, according to which said metabolite(s) of interest associated with said applied natural extract exhibit at least one of the following effects: hydration, and / or healing, and / or an anti-blemish effect, and / or a soothing effect, and / or a lightening effect, and / or an effect on the complexion, and / or an effect on the appearance of the skin, and / or an anti-wrinkle effect.

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