Use of short-chain fatty acids as an anti-dandruff agent

Abstract

It provides an effective activator for inhibiting the growth of Malassezia yeast without affecting other bacteria that make up the majority of the skin microbiome. [Solution] The present invention relates to the field of cosmetics, and more particularly to cosmetic uses of a prepared culture medium obtained from at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, or from at least one microorganism capable of producing such short-chain fatty acids, as an anti-dandruff agent, for preventing and / or treating desquamation disorders of the skin associated with excessive proliferation of Malassezia yeasts on the skin, and for maintaining and / or restoring the skin's ecoflora to a normal level, particularly by preventing excessive colonization of the skin by Malassezia and / or by mediating the proliferation of Cuticobacterium acnes.

Description

[Technical Field]

[0001] The present invention relates to the cosmetic field, and particularly to a cosmetic use of at least one short-chain fatty acid selected from the group consisting of propionic acid, butyric acid, valeric acid, non-metal salts thereof, esters thereof and mixtures thereof, or a conditioned culture medium obtained from at least one microorganism capable of producing such a short-chain fatty acid, as an anti-dandruff agent for preventing and / or treating desquamating skin disorders associated with excessive growth of Malassezia yeasts on the skin, and for maintaining and / or restoring skin ecoflora to normal levels, particularly by preventing excessive colonization of skin by Malassezia yeasts and / or by mediating the growth of Cutibacterium acnes. [Background Art]

[0002] Desquamating skin disorders such as dandruff or seborrheic dermatitis affect up to 50% of the world's population. This disorder affects both men and women and is perceived as having very negative psychosocial impacts. The appearance of dandruff is unpleasant both aesthetically and because it causes discomfort, particularly tingling or pruritus, so many people facing this problem desire to eliminate dandruff efficiently and permanently.

[0003] These disorders correspond to visible excessive skin desquamation resulting from excessively rapid proliferation of epithelial cells and their abnormal maturation. This phenomenon can be caused in particular by excessively harsh skin or hair treatments, severe climatic conditions, stress, diet, fatigue and pollution. Dandruff and seborrheic dermatitis conditions usually result from an impairment of the skin microflora, more particularly excessive colonization by fungi belonging to the yeast family of the genus Malassezia, particularly the Malassezia restricta species, and a lower abundance of Cutibacterium acnes compared to healthy scalp.

[0004] Many treatments have been developed with the primary objective of eradicating Malassezia yeast from the skin. Therefore, the activity of currently used activators, such as zinc pyrithione, piroctone olamine, or selendisulfide, is primarily based on their fungicidal properties. However, it is well known that many of these conventional agents exert their antimicrobial effects against at least one other type of bacteria and are therefore not selective for Malassezia yeast, particularly Malassezia restricta, and thus can kill or impair beneficial skin microflora.

[0005] Therefore, finding a novel activator that has selective growth inhibitory activity against Malassezia yeasts (known to be the cause of desquamative disorders of the skin), particularly Malassezia restricta, is a major challenge in maintaining and / or restoring a healthy skin microflora, and thus meets consumer demand. [Prior art documents] [Non-patent literature]

[0006] [Non-Patent Document 1] International Cosmetic Ingredient Dictionary and Handbook [Overview of the project] [Problems that the invention aims to solve]

[0007] The object of the present invention is to provide an active agent that is effective in inhibiting the growth of Malassezia yeasts (which are the cause of desquamative disorders of the skin), particularly Malassezia restricta species, without its antibacterial effect extending to other bacteria that make up the majority of the skin microbiome, particularly Staphylococcus epidermidis and / or Staphylococcus capitis and / or Cutibacterium acnes.

[0008] Another object of the present invention is to propose an activator that can maintain and / or repair the skin's ecoflora at a normal level, particularly by preventing excessive colonization of the skin by Malassezia yeasts and / or by mediating the proliferation of Cutibacterium acnes.

[0009] The applicant has surprisingly discovered that cosmetic applications of at least one short-chain fatty acid having a chain length containing five or fewer carbon atoms, selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters, and mixtures thereof, enable effective treatment of dandruff and / or seborrheic dermatitis conditions associated with the proliferation of Malassezia yeasts, without having antimicrobial effects against Staphylococcus epidermidis and Cutibacterium acnes, in contrast to medium-chain fatty acids having a chain length containing more than six carbon atoms, such as caproic acid, caprylic acid, ethyl caproate, glyceryl monocaprylate, and propylene glycol monocaprylate, as shown in Example 3 of this application, and metal salts of short-chain fatty acids such as zinc propionate.

[0010] In addition to their selectivity for Malassezia, the aforementioned short-chain fatty acids mediate the growth of Cutibacterium acnes, a skin commensal bacterium that is reduced in dandruff and seborrheic dermatitis conditions.

[0011] Therefore, these effects contribute to restoring the balance of the skin's ecoflora. [Means for solving the problem]

[0012] Therefore, the subject of the present invention is the cosmetic use of a prepared culture medium as an anti-dandruff agent, obtained from i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters and mixtures thereof, or ii) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters and mixtures thereof, obtained from at least one microorganism capable of producing one or more short-chain fatty acids.

[0013] Another subject of the present invention is a cosmetic application of a prepared culture medium obtained from i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, and comprising at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, for cosmetic applications to prevent and / or treat desquamative disorders of the skin, such as dandruff and / or seborrheic dermatitis, associated with the proliferation of Malassezia yeast, more particularly Malassezia restricta species.

[0014] The present invention also relates to cosmetic applications of a prepared culture medium containing at least one short-chain fatty acid selected from i) propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and comprising at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, for the purpose of maintaining and / or repairing the skin's ecoflora at a normal level, particularly by preventing excessive colonization on the skin by Malassezia yeast, more particularly by Malassezia restricta species, and / or by mediating the proliferation of Cutibacterium acnes.

[0015] Another subject of the present invention is a cosmetic method intended to prevent and / or treat a desquamating disorder of the skin, such as dandruff and / or seborrheic dermatitis, associated with the proliferation of Malassezia yeast, more particularly Malassezia restricta species, comprising the step of applying a cosmetic composition to the hair and / or skin, comprising the steps of: i) applying to the hair and / or skin a cosmetic composition containing an effective amount of a prepared culture medium obtained from at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and containing at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof.

[0016] Another subject of the present invention is a cosmetic method intended to maintain and / or restore the skin's ecoflora to normal levels by preventing excessive colonization on the skin, particularly by Malassezia yeasts, more particularly by Malassezia restricta species, and / or by mediating the proliferation of Cutibacterium acnes, the method comprising the step of applying a cosmetic composition to the hair and / or skin, comprising: i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, or ii) a prepared culture medium obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and containing at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof.

[0017] definition As used herein, the terms “treat” or “treatment” refer to any action aimed at improving the well-being or health of an individual. Therefore, the term encompasses, but is not limited to, cosmetic treatments, reducing, alleviating, or suppressing the symptoms of dandruff or seborrheic dermatitis.

[0018] For the purposes of the present invention, the term "non-metallic salt" refers to a salt that does not contain metal ions such as zinc ions, aluminum ions, copper ions, iron ions, and mixtures thereof.

[0019] For the purposes of the present invention, the term "skin" means the skin of the whole body including the scalp, preferably the scalp and facial skin, for example, the skin of the forehead, nose, cheeks, chin, chest and neck.

[0020] As used herein, the term "skin ecoflora" means the microflora naturally present on healthy skin, particularly skin commensal microorganisms such as Staphylococcus epidermidis, and / or Staphylococcus capitis, and / or Cutibacterium acnes.

[0021] For the purposes of the present invention, the term "prevent" means reducing the risk of manifestation of a phenomenon, particularly dandruff and seborrheic dermatitis in the context of the present invention.

[0022] For the purposes of the present invention, the term "effective amount" means an amount sufficient to obtain the expected effect.

[0023] As used herein, the term "cosmetic composition" means a composition suitable for application to the skin, particularly a composition comprising a physiologically acceptable medium.

[0024] The term "physiologically acceptable medium" means a medium that is suitable for topical administration of a composition, that is, compatible with the skin of the face, body and scalp.

[0025] For the purposes of the present invention, the term "short-chain fatty acid" means a carboxylic acid having an aliphatic chain containing 3 to 5 carbon atoms, preferably a carboxylic acid having an aliphatic chain containing 3 carbon atoms. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] [Figure 1]This graph shows the effect of SCFA on the proliferation of Malassezia restrictor. [Figure 2] This graph shows the effect of SCFA on the proliferation of Cutibacterium acnes. [Figure 3] This graph shows the effect of SCFA on the proliferation of Staphylococcus epidermidis. [Figure 4] This graph shows the effect of fatty acids (50 mM) on the proliferation of Malassezia restrictor, quantified using fluorescent staining. [Figure 5] This graph shows the effects of fatty acid esters and metal salts (50 mM) on the proliferation of Malassezia restrictor, quantified by measuring ATP. [Figure 6] This graph shows the effects of fatty acids, esters, and metal salts on the proliferation of Cutibacterium acnes. [Figure 7] This graph shows the effects of fatty acids, esters, and metal salts on the growth of Staphylococcus epidermidis. [Figure 8] This graph shows the effect of SCFA on the growth of Cutibacterium acnes when it is cultured in co-culture with Staphylococcus epidermidis under aerobic conditions. [Modes for carrying out the invention]

[0027] Short-chain fatty acids (SCFAs) As already described, the subject of the present invention is the cosmetic use of a modified culture medium as an anti-dandruff agent, obtained from i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters and mixtures thereof, or ii) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters and mixtures thereof, obtained from at least one microorganism capable of producing one or more short-chain fatty acids.

[0028] The present invention also relates to cosmetic applications of a prepared culture medium containing i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and comprising at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, for cosmetic applications to prevent and / or treat desquamative disorders of the skin, such as dandruff and / or seborrheic dermatitis, associated with the proliferation of Malassezia yeast, more particularly Malassezia restricta species.

[0029] The present invention also relates to cosmetic applications of a prepared culture medium containing at least one short-chain fatty acid selected from i) propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and comprising at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, for maintaining and / or repairing the skin's ecoflora at normal levels, particularly by preventing excessive colonization on the skin by Malassezia yeast, more particularly by Malassezia restricta, and / or by mediating the proliferation of Cuticobacterium acnes.

[0030] Nonmetallic salts of short-chain fatty acids according to the present invention are particularly preferred, and these salts can be any safe and effective nonmetallic salt of such acids. For example, certain preferred salts may include calcium salts, sodium salts, magnesium salts, and potassium salts, with sodium salts being the most preferred.

[0031] As a further example, amino acid salts may be used. For instance, carnitine or lysine salts of short-chain fatty acids according to the present invention may be used. Those skilled in the art will recognize that various other amino acids may also be used.

[0032] As examples of esters of propionic acid, butyric acid, or valeric acid, any safe and effective ester of such acids may be used. For example, if SCFA is an ester of propionic acid, the components may be expressed as follows:

[0033] [ka]

[0034] In the formula, R1 = CH3, and R2 is the ester chain or ester of propionic acid.

[0035] For example, the ester chain of a selected acid may be linear or branched, but typically contains about eight or fewer carbon atoms. More preferably, this ester chain contains 1 to about five carbon atoms and may also be linear (e.g., n-propyl) or branched (e.g., iso-propyl). Highly preferred ester chains include those forming methyl esters (i.e., R2 is -CH3), ethyl esters, n-propyl esters, iso-propyl esters, n-butyl esters, iso-butyl esters, and mixtures thereof. Examples of propionic acid esters that may be used herein include methyl propionate, ethyl propionate, n-propyl propionate, iso-propyl propionate, n-butyl propionate, and iso-butyl propionate. Esters of propionic acid, butyric acid, or valeric acid may also be selected.

[0036] Examples of short-chain fatty acids include sodium propionate (reference number P1880), sodium butyrate (reference number 303410), and valeric acid (reference number 75054), all sold by Sigma.

[0037] In certain embodiments, short-chain fatty acids are obtained from at least one microorganism capable of producing one or more short-chain fatty acids. The at least one microorganism capable of producing one or more short-chain fatty acids can be selected from the group consisting of Lactobacillus species, Bifidobacterium species, Ruminococcus species, Rosebria species, Akkermansia muciniphila, Faecalibacterium species, Eubacterium rectale, and Cutibacterium acnes, preferably Lactobacillus species, Bifidobacterium species, and Cutibacterium acnes, more preferably Cutibacterium acnes, for example, the Cutibacterium acnes strain ATCC6919.

[0038] In another embodiment, the short-chain fatty acids according to the present invention are contained in a prepared culture medium (or supernatant) obtained from at least one microorganism capable of producing one or more short-chain fatty acids.

[0039] The at least one microorganism capable of producing one or more short-chain fatty acids can be selected from the group consisting of Lactobacillus species, Bifidobacterium species, Ruminococcus species, Rosebria species, Akkermansia muciniphila, Faecalibacterium species, Eubacterium rectore, and Cutibacterium acnes, preferably Lactobacillus species, Bifidobacterium species, and Cutibacterium acnes, more preferably Cutibacterium acnes, for example, the Cutibacterium acnes ATCC6919 strain.

[0040] "Culture supernatant," also called "prepared culture medium," is typically obtained by culturing concentrated microorganisms in a medium suitable for the survival and / or growth of microorganisms, and then separating the medium and microorganisms so as to collect the medium that has been in contact with the microorganisms. Preferably, the culturing is carried out over a period of time and under conditions that are likely to cause the microorganisms to release into the medium an activator having the desired anti-dandruff properties, particularly short-chain fatty acids according to the present invention.

[0041] An environment suitable for the survival and / or growth of microorganisms is any nutrient medium suitable for the survival and / or cultivation of microorganisms. This medium typically contains sufficient amounts of carbon and nitrogen sources, such as amino acids, sugars, proteins, fatty acids, phosphates, sulfates, minerals, growth factors, and vitamins.

[0042] For the purposes of this application, the terms "prepared culture medium" and "culture supernatant" are used without distinction to specify the entire culture supernatant obtained after culturing the microorganism in question, or any fraction or subcompound of the supernatant obtained by dialysis, preparative separation, phase separation, filtration chromatography, affinity chromatography, precipitation, concentration, lyophilization, etc.

[0043] In the context of the present invention, a prepared culture medium obtained from at least one microorganism capable of producing one or more short-chain fatty acids according to the present invention is prepared by the following steps: i) A step of culturing at least one microorganism capable of producing one or more short-chain fatty acids, preferably a microorganism of the species Cutibacterium acnes, for example, Cutibacterium acnes ATCC6919, ii) A step of separating the culture supernatant from the biomass, particularly by centrifugation, iii) A step of collecting the culture supernatant, iv) A step to stabilize the culture supernatant, if necessary, for example by filtration and / or autoclaving. It is obtained by a method that includes the following.

[0044] As used herein, the term “biomass” refers to the cells of Cutibacterium acnes obtained after carrying out step i).

[0045] Preferably, filtration is performed using a syringe filter with a pore size between 0.2 μm and 0.45 μm.

[0046] The short-chain fatty acid or modified culture medium according to the present invention is used in an amount of 0.01% to 5% by mass relative to the total mass of the composition, preferably in an amount of 0.3% to 1.0% by mass relative to the total mass of the composition.

[0047] Another subject of the present invention is a cosmetic method intended to prevent and / or treat a desquamating disorder of the skin, such as dandruff and / or seborrheic dermatitis, associated with the proliferation of Malassezia yeast, more particularly Malassezia restricta species, comprising the step of applying a cosmetic composition to the hair and / or skin, comprising the steps of: i) applying to the hair and / or skin a cosmetic composition containing an effective amount of a prepared culture medium obtained from at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and containing at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof.

[0048] Another subject of the present invention is a cosmetic method intended to maintain and / or restore the skin's ecoflora to normal levels by preventing excessive colonization on the skin, particularly by Malassezia yeasts, more particularly by Malassezia restricta species, and / or by mediating the proliferation of Cutibacterium acnes, the method comprising the step of applying a cosmetic composition to the hair and / or skin, comprising: i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, or ii) a modified culture medium obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and containing at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof.

[0049] Preferably, the cosmetic composition includes a cosmetically acceptable culture medium, in other words, a keratinous substance, particularly a culture medium suitable for topical application to the skin.

[0050] Preferably, the pH of the cosmetic composition is between 6 and 8, particularly between 6.5 and 7.5, and especially a neutral pH of 7.0.

[0051] Advantageously, the short-chain fatty acid or modified culture medium according to the present invention is present in an amount of 0.01% to 5% by mass relative to the total mass of the composition, preferably in an amount of 0.3% to 1% by mass relative to the total mass of the composition.

[0052] In a first preferred embodiment, the cosmetic composition is a composition for the scalp, which may be a composition that is rinsed off or left on. The composition is preferably in the form of a shampoo, cream, mousse (aerosol or non-aerosol), paste, gel, emulsion, lotion, or even a stick. Preferably, the composition for the hair is a shampoo, gel, or lotion.

[0053] In a second preferred embodiment, the cosmetic composition is a composition for the skin, which may be fluid to varying degrees and may have the appearance of a white or colored cream, ointment, emulsion, lotion, serum, paste, or foam. The composition may be applied to the skin in aerosol form as needed. The composition may also be in solid form, such as a stick or cosmetic compact. In particular, the cosmetic composition may be in the form of an aftershave gel or lotion, a body hygiene composition, such as a shower gel, a solid composition, such as a soap or cleansing bar, or a composition for caring for or cleansing the skin.

[0054] The cosmetic composition preferably comprises water and / or one or more water-miscible organic solvents, the solvent of which may be selected from linear or branched C1-C6 monoalcohols, e.g., ethanol, isopropanol, tert-butanol, or n-butanol; polyols, e.g., glycerol, propylene glycol, hexylene glycol (or 2-methyl-2,4-pentanediol), and polyethylene glycol; polyol ethers, e.g., dipropylene glycol monomethyl ether; and mixtures thereof.

[0055] Preferably, the cosmetic composition contains water in an amount ranging from 30 to 98% by mass, particularly 40 to 95% by mass, and more preferably 50 to 90% by mass, based on the total mass of the composition.

[0056] Preferably, the cosmetic composition contains an amount of organic solvent ranging from 0.05 to 60% by mass, preferably 0.5 to 50% by mass, and more preferably 1 to 40% by mass, based on the total mass of the cosmetic composition.

[0057] The cosmetic compositions according to the present invention may also contain at least one conventional cosmetic ingredient selected from, in particular, plant, mineral, animal, or synthetic oils; liquid fatty alcohols; liquid fatty esters; solid fatty substances, especially waxes, solid fatty esters, and solid alcohols; anionic, cationic, amphoteric, and nonionic surfactants; anionic, nonionic, amphoteric, and cationic polymers; anti-dandruff agents other than short-chain fatty acids or modified culture media according to the present invention; antioxidants; agents for combating hair loss; silicones; fragrances; polymers or nonpolymer thickeners, especially associative polymers; preservatives as needed; chelating agents; and colorants. Naturally, the compositions may contain some of the cosmetic ingredients listed above. Those skilled in the art will carefully select the components and their amounts to constitute the composition so that the advantageous properties of the composition according to the present invention are not adversely affected or substantially affected by the intended additions.

[0058] Following the application of the short-chain fatty acid or the prepared culture medium or composition to the hair and / or skin, a rinsing step, for example with water, may or may not be performed.

[0059] Throughout this description, including the claims, the phrase "comprising a" should be understood as synonymous with "comprising at least one," unless otherwise specified.

[0060] Furthermore, the phrase "at least one" should be understood as synonymous with "one or more" unless the opposite is specified.

[0061] The expressions "greater than," "between... and...," and "within the range of..." should be understood to include both limits unless the opposite is specified.

[0062] The following embodiments and figures are presented as examples of the present invention, without limiting the scope of the invention.

[0063] Compounds are listed by their chemical name or CTFA name (International Cosmetic Ingredient Dictionary and Handbook), as appropriate.

[0064] The present invention will be illustrated in more detail in the following embodiments. [Examples]

[0065] (Example 1) Evaluation of selective inhibition of Malassezia restrictor proliferation by sodium propionate, sodium butyrate, and sodium valerate (according to the present invention). A) Materials and methods Organisms and propagation conditions M. restricta ATCC MYA-4611, S. epidermidis ATCC12228, and C. acnes ATCC6919 were purchased from ATCC. M. restricta was routinely cultured in modified Dixon (MD) medium (pH 6) consisting of 36 g of malt extract (Sigma 70167), 20 g of dessicated oxbile (Sigma 70168), 6 g of Bacto® peptone (BD 211677), 1% v / v Tween 40 (Sigma P1504), 0.2% v / v oleic acid (Fluka 75096), and 0.2% v / v glycerol (Promega H5433) in 1 liter of dH2O. C. acnes and S. epidermidis were cultured in modified Brain Heart Infusion (MBHI) medium (pH 7) containing 37 g of BHI substrate (Accumedia 7116B), 0.4% (v / v) Tween-40 (Sigma P1504), 0.2% (v / v) oleic acid (Fluka 75096), and 0.2% (v / v) glycerol (Promega H5433) in 1 liter of dH2O. M. restricta and S. epidermidis were routinely grown under aerobic conditions with shaking at 200 rpm, while C. acnes was grown under anaerobic conditions. All organisms were grown at 33°C. The culture medium was supplemented with sodium acetate (CH3COOH, Sigma reference number 32319), sodium propionate (CH3CH2COOH, Sigma reference number P1880), sodium butyrate (CH3(CH2)2COOH, Sigma reference number 303410), and valeric acid (CH3(CH2)3COOH, Fluka reference number 75054) where necessary. For valeric acid, the pH of the modified cell medium was neutralized with NaOH (sodium hydroxide, Sigma 283060).

[0066] Quantification of total cell count using SYTO9 M. restrictor cells were collected from 1 ml of culture by centrifugation at 10,000 × g for 5 minutes at room temperature. The cells were washed, pelletized, and resuspended in 0.9% NaCl solution. The same volume of the cell suspension and SYTO 9 working stock (3 μl of SYTO 9 component from the Live / Dead BacLight bacterial viability kit, ThermoFisher Scientific, L7012, diluted in 1 ml of 0.9% NaCl solution) were thoroughly mixed and incubated in a dark room for 10 minutes. Fluorescence intensity units were measured using a Tecan microplate reader with excitation / emission wavelengths of 485 / 530 nm. Standards for the correlation between fluorescence units and concentration were prepared using a fluorimeter and hemocytometer under a light microscope. Total cell counts were calculated using these standard curves.

[0067] Growth assay M. restrictor cells, stored as a glycerol stock (30% glycerol in MD medium), were revived by seeding onto MD agar plates and incubated at 33°C for 2-3 days. A pre-culture was prepared from the microbial flora of the cells grown on the agar plates. Cells were scraped from 1 / 4 of the plate, suspended in 70 ml of MD medium in a 250 ml baffled Erlenmeyer flask, homogenized, and incubated at 33°C at 200 rpm for 24 hours. For the experiment, 10 7 Cells / ml were inoculated into fresh MD medium with or without short-chain fatty acids. Growth was monitored for up to 24 or 96 hours by measuring cell density using SYTO 9. C. acnes and S. epidermidis were each inoculated into OD medium. 600 Bacterial cells were inoculated into MBHI medium at starting cell densities of 0.05 and 0.25. Bacterial growth was measured at various time points by seeding serial dilution cultures onto Brainheart infusion agar plates, and colonies were counted at 72 hours for C. acnes and at 24 hours for S. epidermidis.

[0068] Quantification of Colony-Forming Units (CFU) Bacterial growth was quantified by seeding serial dilution cultures onto Brainheart infusion agar plates at various time points, and colonies were counted at 72 hours for C. acnes and at 24 hours for Staphylococcus species.

[0069] B) Result Sodium propionate, sodium butyrate, and sodium valerate completely inhibit the growth of Malassezia restrictor at a concentration of 30 mM. Acetates showed no effect at the same concentration (Figure 1).

[0070] Sodium propionate, sodium butyrate, and sodium valerate did not affect the growth of C. acnes and S. epidermidis at concentrations (30 mM) at which complete inhibition of Malassezia restrictor was observed (Figures 2 and 3).

[0071] Conclusion: Sodium propionate, sodium butyrate, and sodium valerate selectively inhibit Malassezia restrictor and do not affect the growth of other major skin symbiotic microorganisms such as C. acnes and S. epidermidis.

[0072] (Example 2) Evaluation of the promotion of Cutibacterium acnes growth by sodium propionate (according to the present invention). A) Materials and methods Organisms and propagation conditions S. epidermidis ATCC12228 and C. acnes ATCC6919 were purchased from ATCC. C. acnes and S. epidermidis were cultured in modified Brain Heart Infusion (MBHI) medium (pH 7) containing 37 g of BHI substrate (Accumedia 7116B), 0.4% (v / v) Tween-40 (Sigma P1504), 0.2% (v / v) oleic acid (Fluka 75096), and 0.2% (v / v) glycerol (Promega H5433) in 1 liter of dH2O. S. epidermidis was routinely grown under aerobic conditions with shaking at 200 rpm, and C. acnes was grown under anaerobic conditions. All organisms were grown at 33°C. The culture medium was supplemented with sodium propionate (CH3CH2COOH, Sigma reference number P1880) wherever necessary.

[0073] Growth assay C. acnes and S. epidermidis, respectively, OD 600 Bacterial cells were inoculated into MBHI medium at starting cell densities of 0.05 and 0.25. Bacterial growth was measured at various time points by seeding serial dilution cultures onto Brainheart infusion agar plates, and colonies were counted at 72 hours for C. acnes and at 24 hours for S. epidermidis.

[0074] Co-culture assay For co-culture, two exponentially growing cell species of the same volume are cultured in a 24-well glass-bottom plate at a rate of 10 per ml. 5 The cells were mixed at individual cell densities. The growth of various organisms in the co-culture was quantified by planting the culture suspension on agar plates. For the selective quantification of each bacterium in the co-culture, C. acnes was selectively counted by growing the agar plates under aerobic conditions to promote the selective growth of S. epidermidis, or by growing them anaerobicly using furazolidone, which kills only S. epidermidis.

[0075] Quantification of Colony-Forming Units (CFU) Bacterial growth was quantified by seeding serial dilution cultures onto Brainheart infusion agar plates at various time points, and colonies were counted at 72 hours for C. acnes and at 24 hours for Staphylococcus species.

[0076] B) Result S. epidermidis was cultured for 24 hours until a biofilm formed to maximize its growth, and then C. acnes cells were added to oxygen-saturated fresh medium with or without propionate. Growth of C. acnes was minimal unless the propionate was metabolized by the S. epidermidis layer, which can create conditions for C. acnes growth.

[0077] The proliferation of C. acnes and S. epidermidis was monitored at 24, 48, and 72 hours, and it was found that the proliferation of C. acnes increased in the presence of propionate (Figure 8). The significantly higher proliferation of C. acnes under aerobic conditions in the presence of propionate demonstrates that propionate can modulate the community dynamics of the skin microbiome by promoting the proliferation of C. acnes in the presence of Staphylococcus species.

[0078] (Example 3) Evaluation of selective inhibition of Malassezia restrictor proliferation by sodium propionate, sodium butyrate, and sodium valerate (according to the present invention) compared with medium-chain fatty acids: caproic acid, caprylic acid, ethyl caproate, glyceryl monocaprylate, propylene glycol monocaprylate (other than those specified in the present invention) and metal salts of short-chain fatty acids: zinc propionate (other than those specified in the present invention). A) Materials and methods Organisms and propagation conditions M. restricta ATCC MYA-4611, S. epidermidis ATCC12228, and C. acnes ATCC6919 were purchased from ATCC. M. restricta was routinely cultured in modified Dixon (MD) medium (pH 6) consisting of 36 g of malt extract (Sigma 70167), 20 g of dried oxbile (Sigma 70168), 6 g of Bacto® peptone (BD 211677), 1% (v / v) Tween 40 (Sigma P1504), 0.2% (v / v) oleic acid (Fluka 75096), and 0.2% (v / v) glycerol (Promega H5433) in 1 liter of dH2O. C. acnes and S. epidermidis were cultured in modified Brain Heart Infusion (MBHI) medium (pH 7) containing 37 g of BHI substrate (Accumedia 7116B), 0.4% (v / v) Tween-40 (Sigma P1504), 0.2% (v / v) oleic acid (Fluka 75096), and 0.2% (v / v) glycerol (Promega H5433) in 1 liter of dH2O. M. restricta and S. epidermidis were routinely grown under aerobic conditions with shaking at 200 rpm, while C. acnes was grown under anaerobic conditions. All organisms were grown at 33°C. The culture medium was supplemented with 50 mM sodium propionate (CH3CH2COOH, Sigma reference number P1880), sodium butyrate (CH3(CH2)2COOH, Sigma reference number 303410), and valeric acid (CH3(CH2)3COOH, Fluka reference number 75054) where necessary. For valeric acid, the pH of the modified cell medium was neutralized with NaOH (sodium hydroxide, Sigma 283060).

[0079] Quantification of total cell number of M. restrictor using SYTO9 Cells of *M. restricta* obtained from 1 ml of culture were collected by centrifugation at 10,000 × g for 5 minutes at room temperature. The cells were washed, pelletized, and resuspended in 0.9% NaCl solution. The same volume of the cell suspension and SYTO 9 working stock (3 ul of SYTO 9 component from the Live / Dead BacLight bacterial viability kit, ThermoFisher Scientific, L7012, diluted in 1 ml of 0.9% NaCl solution) were thoroughly mixed and incubated in a dark room for 10 minutes. Fluorescence intensity units were measured using a Tecan microplate reader with excitation / emission wavelengths of 485 / 530 nm. Standards for the correlation between fluorescence units and concentration were prepared using a fluorometer and hemocytometer under a light microscope. Total cell counts were calculated using these standard curves.

[0080] Growth assay M. restrictor cells, stored as a glycerol stock (30% glycerol in MD medium), were revived by seeding onto MD agar plates and incubated at 33°C for 2-3 days. A pre-culture was prepared from the microbial flora of the cells grown on the agar plates. Cells were scraped from 1 / 4 of the plate, suspended in 70 ml of MD medium in a 250 ml baffled Erlenmeyer flask, homogenized, and incubated at 33°C at 200 rpm for 24 hours. For the experiment, 10 7 Cells / ml were inoculated into fresh MD medium with or without 50 mM short-chain fatty acids. Growth was monitored for up to 24 or 96 hours by measuring cell density using SYTO 9 or by estimating ATP concentration (see below). C. acnes and S. epidermidis were each inoculated into OD medium. 600 Bacterial cells were inoculated into MBHI medium at starting cell densities of 0.05 and 0.25. Bacterial growth was measured at various time points by seeding serial dilution cultures onto Brainheart infusion agar plates, and colonies were counted at 72 hours for C. acnes and at 24 hours for S. epidermidis.

[0081] Estimation of ATP Cellular ATP was measured using the BacTiterGlo microbial cell viability kit (Promega) according to the manufacturer's protocol. In other words, luciferase activity, which utilizes ATP as a substrate, was estimated as a direct measurement of ATP produced by the cells.

[0082] B) Result Due to various characteristics of different compounds that interfered with assay interpretation, the growth of M. restrictor was estimated using two different methods: - For all fatty acids and caproic acid esters, the total number of cells after 72 hours of incubation was quantified by fluorescence measurement of cells stained with the fluorescent dye SYTO9 (see Methods). The starting cell density of M. restricta was 1 × 10⁻⁶. 7 The cell count was 100 cells / ml (Figure 4). - For caprylic acid esters and zinc propionate, cell viability was quantified by estimating the total amount of ATP in the cell lysate prepared from cells grown for 24 hours (because these compounds made the culture medium opaque). The ATP measurements were expressed as relative luminescence units (RLU) of the luminescence released by the enzyme luciferase in the presence of ATP (Figure 5).

[0083] The growth of C. acnes was quantified by estimating the colony-forming units (CFU) after growing them in a culture medium containing fatty acids and their esters for 72 hours (Figure 6).

[0084] The growth of S. epidermidis was quantified by estimating the colony-forming units (CFU) after growing them for 24 hours in a culture medium containing fatty acids and their esters (Figure 7). The results are presented in Tables 1 and 2 below.

[0085] [Table 1]

[0086] [Table 2]

[0087] Conclusion: Sodium propionate, sodium butyrate, and sodium valerate (compounds according to the present invention) selectively inhibit M. restricta, in contrast to caproic acid, ethyl caproate, caprylic acid, glyceryl monocaprylate, propylene glycol monocaprylate, and zinc propionate (compounds other than those according to the present invention), which have been found to inhibit at least one skin symbiont, and do not affect other major skin symbiont microorganisms such as C. acnes and S. epidermidis.

[0088] (Example 3) Facial cream Prepare the following composition.

[0089] [Table 3]

[0090] The composition is applied to the skin in a seborrheic dermatitis state.

Claims

1. i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters and mixtures thereof, or ii) a prepared culture medium obtained from at least one microorganism capable of producing one or more short-chain fatty acids, containing at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters and mixtures thereof, for cosmetic use as an anti-dandruff agent.

2. i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, or ii) a prepared culture medium obtained from at least one microorganism capable of producing one or more short-chain fatty acids, comprising at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof, for cosmetic use in preventing and / or treating desquamative disorders of the skin associated with the proliferation of Malassezia yeasts, such as dandruff and / or seborrheic dermatitis.

3. i) at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters and mixtures thereof, or ii) a modified culture medium obtained from at least one microorganism capable of producing one or more short-chain fatty acids, comprising at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, its nonmetallic salts, its esters and mixtures thereof, for cosmetic applications, particularly by preventing excessive colonization of the skin by Malassezia yeasts and / or by mediating the proliferation of Cutibacterium acnes.

4. The cosmetic use according to any one of claims 1 to 3, wherein the short-chain fatty acid is selected from sodium propionate, sodium butyrate, sodium valerate, and mixtures thereof.

5. The cosmetic use according to any one of claims 1 to 4, wherein the short-chain fatty acid is obtained from at least one microorganism of the Cutibacterium acnes species, preferably from at least one microorganism of the Cutibacterium acnes strain ATCC6919.

6. The prepared culture medium is prepared by the following steps: i) A step of culturing at least one microorganism capable of producing one or more short-chain fatty acids, preferably a microorganism of the species Cutibacterium acnes, for example, Cutibacterium acnes ATCC6919, ii) A step of separating the culture supernatant from the biomass, particularly by centrifugation, iii) A step of collecting the culture supernatant, iv) A step to stabilize the culture supernatant, if necessary, for example by filtration and / or autoclaving. A cosmetic use according to any one of claims 1 to 3, obtained by a method comprising:

7. A cosmetic method intended to prevent and / or treat desquamative disorders of the skin associated with the proliferation of Malassezia yeasts, such as dandruff and / or seborrheic dermatitis, comprising the step of applying a cosmetic composition to the hair and / or skin, comprising the steps of: i) applying to the hair and / or skin a cosmetic composition containing an effective amount of a prepared culture medium obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and containing at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts, esters and mixtures thereof.

8. A cosmetic method intended to maintain and / or restore the skin's ecoflora to a normal level, particularly by preventing excessive colonization on the skin by Malassezia yeasts and / or by mediating the proliferation of Cutibacterium acnes, comprising the step of applying a cosmetic composition to the hair and / or skin, comprising the steps of: i) applying to the hair and / or skin a cosmetic composition containing an effective amount of a prepared culture medium obtained from at least one microorganism capable of producing one or more short-chain fatty acids, and containing at least one short-chain fatty acid selected from propionic acid, butyric acid, valeric acid, nonmetallic salts thereof, esters thereof and mixtures thereof.

9. The beauty method according to any one of claims 7 or 8, wherein the short-chain fatty acid or the prepared culture medium is present in an amount of 0.01% to 5% by mass, preferably 0.3% to 1% by mass, relative to the total mass of the composition.

10. The beauty method according to any one of claims 7 to 9, wherein the cosmetic composition is in the form of a shampoo, cream, mousse, paste, gel, emulsion, lotion, or even a stick.

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