Cosmetic treatment based on pycnidione

Abstract

Treatments are intended for use on the skin and its appendages to improve their overall condition and / or sensory comfort. They may be selected from treatments targeting signs of chronological or premature aging, moisturizing treatments, treatments for dull and / or tired skin, treatments for sensory discomfort of the skin, treatments for oily and / or acne-prone skin, and / or hair treatments. Pycnidione may be provided in the form of an extract of biomass capable of producing pycnidione as a secondary metabolite in vitro culture.

Description

Technical Field

[0001] This invention relates to non-therapeutic cosmetic treatments of the skin and its appendages that can be performed locally.

[0002] This invention is intended for the cosmetics, dermatology, and hygiene and personal care product industries. These industries deal with humans and animals and prepare and implement active ingredients and final formulations. These industries are constantly seeking new treatments to provide improvements in the aesthetic condition of the skin and / or its appendages or to enhance their sensory comfort. Background Technology

[0003] Typically, cosmetic treatments involve preventing or addressing unsightly defects or discomforts caused by the chronological or premature aging of the skin and / or its appendages, such as surface unevenness (wrinkles, fine lines, roughness, etc.), color unevenness (spots, redness, etc.) and skin tone luster (loss of luster, brightness), sagging (appearance of double chin, drooping eyelids, etc.), or sensory discomfort (tingling, tightness, etc.).

[0004] It may also include treatments that do not necessarily involve mature skin or appendages, such as in the case of preventative treatments, or treatments designed for all skin types, such as moisturizing treatments, improving skin tone radiance, or sensory discomfort that may be caused by factors other than aging, such as pollution or unhealthy lifestyle.

[0005] In the context of cosmetic treatments, it will always be a matter of providing a curative treatment for diseased skin. Invention Overview

[0007] The object of this invention is to provide novel cosmetic treatments for the skin (including the scalp and lips) and / or its appendages (including, in particular, hair, body hair, eyelashes and eyebrows, fingernails and toenails), which are advantageously suited for implementation within the logic of “natural” products and sustainability using one or more active compounds obtained through biotechnological pathways.

[0008] Therefore, this invention proposes the use of pycnidione for non-therapeutic cosmetic treatments of the skin (including the scalp and lips) and its appendages to improve their overall condition and / or sensory comfort.

[0009] Pycnidione has been described as having antimalarial, anticancer, and more recently antibacterial activity, particularly through its action on Malassezia bacteria for dandruff control (WO2010 / 061185).

[0010] The test results are given below in the instruction manual, demonstrating the advantageous use of pycnidione for non-therapeutic cosmetic treatments of the skin and / or its appendages, particularly for the treatment of signs of chronological aging or premature aging, moisturizing treatments, treatment of dull and / or tired skin (including dark circles and eye bags), treatment of sensory discomfort of the skin, treatment of oily and / or acne-prone skin (treatment of pre-acne conditions of the skin), and / or treatment of brittle and / or split ends.

[0011] More specifically and advantageously, the treatment according to the invention is applicable to:

[0012] - Maintain or improve the mechanical and viscoelastic properties of the skin (including the scalp);

[0013] - Enhance the dermal-epidermal junction (DEJ);

[0014] - Combat the harmful effects of glycation (anti-glycation treatment);

[0015] - Combats the harmful effects of free radicals (antioxidant or anti-free radical treatment, photoprotection);

[0016] - Prevents or reduces rough skin, fine lines, and wrinkles;

[0017] - Prevent or reduce sagging skin (double chin, droopy eyelids);

[0018] - To prevent or reduce pigmentation defects; and / or

[0019] - Stimulates the production of hyaluronic acid.

[0020] Those skilled in the art may also consider other cosmetic activities or benefits.

[0021] pycnidione has a bi-cycloheptatrienolone-like chemical structure, corresponding to the following formula (I) in its natural form (stereochemically described in the scientific publication "Total synthesis and computer investigations of sesquiterpene tropolones reduce stereochemical inconsistencies and resolve an ambiguous biosynthetic relationship", C. Bemis et al., J. Am. Chem. Soc. 2021, 143, 6006-6017):

[0022] (I)

[0023] According to the present invention, the term "pycnidione" also includes dehydroxypycnidione (in which the hydroxyl group of the 10-carbon ring is replaced by hydrogen) and eupenifeldin (a stereoisomer of pycnidione). Pycnidione is preferred.

[0024] According to the present invention, the term "pycnidione" includes pycnidione in a suitable, physiologically acceptable salt form.

[0025] Pycnidione can be produced through chemical synthesis, or it can be derived from natural sources and obtained through in vitro bio-fermentation.

[0026] The following examples of microorganisms (fungi) can be used to produce pycnidione as a secondary metabolite through in vitro biofermentation culture: *Echinochloa glutinis* (… Neosetophoma cerealis (Gastrodia elata) Coniothyrium cereal ), Shield Scale ( Cornichthyrium cerealis ), Neosetophoma samararum Synonyms, strain RKDO834), species of the genus *Neoechocarpus* ( Neosetophoma sp.) (MSX50044 strain), Theissenia rogersii (Strains 9203120, 92031201, OS-F69284 (ATCC 74390)), species of the genus *Stemona* ( Food sp.) (XZ068 strain (CGMCC 10481), Penicillium brevis ( Eupenicillium brefeldianum ) (ATCC 74184 strain), Kionochaeta pughii (BCC 3878 strain) Kionochaeta ramifera (BCC 7585 strain) Gloeotinia sp. (FKI-3416 strain), Aspergillus hencocki ( Aspergillus hancockii ), Entodesmium sp. or Phomopsis tersa (FS441 strain).

[0027] Such strains can be obtained from collections or banks. For example, for Echinochloa oryzae, it can be obtained from the Central Bureau for Fungal Biodiversity (CBS) under the number CBS 963.68.

[0028] Preferably, according to the invention, pycnidione is provided in the form of an extract of biomass from an in vitro culture of microorganisms capable of producing pycnidione as a secondary metabolite. This extract can be purified to increase the pycnidione content. Preferably, the extract is produced by a strain of *Neocetus griseus*. According to specific examples, the in vitro produced extract contains a group of cycloheptatrienolones as secondary metabolites, wherein at least 80% by weight, preferably at least 90% by weight, corresponds to pycnidione, and the remainder consists of epolone A and / or epolone B (which are cycloheptatrienolones, such as pycnidione).

[0029] When the in vitro production route via bio-fermentation is chosen, pycnidione is produced in biomass under suitable and optimal culture conditions for the microorganisms.

[0030] Cultures can be carried out on any suitable synthetic or natural medium, provided it contains appropriate carbon, nitrogen, and inorganic salts. If necessary, the medium can be supplemented with vitamins and other nutrients as appropriate. Examples of common carbon sources include (but are not limited to) sugars such as glucose, maltose, fructose, sucrose, and starch; alcohols such as glycerol and mannitol; amino acids such as glycine, alanine, and asparagine; and oils and fats such as soybean oil and olive oil. Examples of nitrogen sources include organic nitrogenous compounds such as soybean meal, corn steep liquor, beef extract, peptone, yeast extract, amino acid mixtures, and fish meal; and inorganic nitrogen compounds such as ammonium salts and nitrates. Micronutrients in the form of inorganic salts, such as calcium carbonate, sodium chloride, potassium chloride, magnesium sulfate, copper sulfate, manganese chloride, zinc sulfate, cobalt chloride, and various phosphates, can also be used.

[0031] The microorganism can be cultured at a suitable temperature that allows for its growth and efficient production of pycnidione. Preferred culture temperatures are 10°C to 32°C, and more preferably 20°C to 25°C. The initial pH at the start of culture is preferably about 6 to 8, and the culture duration is from one day to several weeks.

[0032] When a sufficient amount of pycnidione is produced, preferably the maximum possible amount, the culture can be stopped.

[0033] Cultivation can be carried out in a bioreactor in either the solid or liquid phase with appropriate stirring.

[0034] Appropriate processing of the biomass (once separated from the supernatant) allows for the recovery of extracts containing pycnidione. Any separation method typically performed in in vitro cultures to extract metabolites from biomass can be used to recover the extract, such as extraction with a suitable solvent in which the metabolite can be dissolved.

[0035] More preferably, according to the invention, in order to extract pycnidione from biomass, extraction is carried out using a solvent, and more preferably a physiologically acceptable solvent, particularly a glycolated solvent, such as pentanediol or butylene glycol or a mixture thereof. If necessary, a clarification step is also performed at the end of the processing to remove any remaining cell debris.

[0036] The extract containing a suitable amount of pycnidione can be advantageously used as is for the implementation of the present invention. If desired, the obtained extract can also be purified to increase the pycnidione content until purified pycnidione is obtained at a high purity level (e.g., with a pycnidione content greater than 80%). Purification of pycnidione can be carried out by chromatography or any other suitable method known to those skilled in the art.

[0037] If the biomass extraction solvent is not a physiologically acceptable solvent, and therefore if the extract cannot be used as is, it is properly treated to remove all trace amounts of non-physiologically acceptable solvents, and then the residue of the obtained extract is contained in a physiologically acceptable medium so that it can be used in the cosmetic treatment according to the invention at an appropriate dose of pycnidione.

[0038] According to the present invention, "physiologically acceptable" means that the composition is suitable for topical or transdermal use, in contact with the mucous membranes, nails, scalp, hair, and skin of mammals, and more particularly humans, and that the composition can be ingested or injected into the skin without risks of toxicity, incompatibility, instability, or allergic reactions. This "physiologically acceptable medium" forms a substance conventionally referred to as an excipient of the composition.

[0039] Preferably, according to the present invention, the physiologically acceptable medium may be an alcohol medium, a glycol medium, an aqueous medium, a water-glycol medium, or a water-alcohol medium, or formed from a water-in-oil emulsion, a water-in-oil emulsion, or a microemulsion. Preferably, according to the present invention, the physiologically acceptable medium is a glycol medium.

[0040] The “effective” amount of pycnidione used in the treatment according to the invention depends on the intended use of the composition containing it, particularly the face, hands, body, hair / scalp, and also on various factors such as the person’s age, condition, or degree of impairment. An effective amount means a non-toxic amount sufficient to achieve the desired effect.

[0041] As a guideline, in a composition intended for use by a consumer in accordance with the invention, the amount of pycnidione to be present in an effective quantity in the composition may range from 0.000001% (0.001 ppm) to 10% (100,000 ppm), preferably 0.000001% (0.01 ppm) to 1% (10,000 ppm), more preferably 0.00001% (0.1 ppm) to 0.1% (1000 ppm), more preferably 0.0001% (1 ppm) to 0.1% (100 ppm), and even more preferably 0.001% (10 ppm) to 0.05% (50 ppm) relative to its total weight, depending on the intended use of the composition and the desired effect (whether more or less pronounced) and the recommended number of applications.

[0042] Unless otherwise stated, all percentages and ratios used in this application are based on the weight of the total composition and all measurements were performed at 25°C.

[0043] In addition, for guidance purposes, the European Cosmetics Directive has specified a standard application rate of 2.72 mg / cm² / day / person for facial cosmetic treatments and 0.5 mg / cm² / day / person for body lotion-type compositions.

[0044] Depending on other features, the cosmetic treatment according to the invention can be combined with one or more other treatments for the skin or hair, such as phototherapy, thermotherapy, or aromatherapy.

[0045] According to the present invention, a multi-compartment device or kit may be proposed for carrying out the methods described above, and the multi-compartment device or kit may contain (e.g., and not limited to) a pycnidione-containing composition in a first compartment and an additional active ingredient acting together in a second compartment, wherein the compositions contained in the first and second compartments are herein considered to be combined compositions for simultaneous, separate, or gradual use over time, particularly in one of the processes defined above.

[0046] According to other advantageous features, pycnidione can be used in combination with one or more additional active ingredients at effective concentrations according to the present invention to advantageously provide synergistic effects or for activity enhancement.

[0047] Other active ingredients can be selected from, for example, agents for skin brightening, anti-redness, anti-blemish, and slimming; soothing active ingredients for treating sensitive and reactive skin; agents that filter radiation (especially UVA, UVB, IR, and visible light, including blue light); humectants for moisturizing, exfoliating, smoothing, toning, anti-aging, anti-wrinkle and fine line reduction, improving mechanical and elastic properties, skin radiance, detoxifying active ingredients, anti-hair growth or conversely promoting hair growth, anti-dandruff, acting on the skin barrier, anti-acne, acting on sebum secretion, mattifying, unifying, anti-inflammatory, antioxidant, anti-free radical, anti-glycation, agents for treating the eye contour (anti-dark circles and for treating eye bags), agents for promoting blood circulation, peptides, vitamins, ceramides, hyaluronic acid, etc. These active ingredients can be obtained through synthetic routes or derived from plant materials, such as plant extracts or products from plant cell culture, fermentation, bacterial culture, or enzyme culture.

[0048] The International Cosmetic Ingredient Dictionary & Handbook, published by the Cosmetic, Toiletry, and Fragrance Association, Inc. (CTFA) of Washington, DC, describes a variety of non-limiting cosmetic and pharmaceutical ingredients commonly used in the skincare industry, suitable for use as additional ingredients in compositions according to the invention, provided they are physically and chemically compatible with the other ingredients of the composition, and especially with the active agents of the invention. Furthermore, the properties 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 derived from biotechnological methods.

[0049] Other skin, scalp, or hair care active agents that can be used in combination with the compositions according to the invention can be found in the sales literature of Sederma, Crodarom, Alban Muller, and Croda, and on the websites www.croda.com and www.crodabeauty.com.

[0050] As examples, the following commercial active agents can also be mentioned: betaine, glycerin, Actimoist Bio 2™ (Active Organics), AquaCacteen™ (Mibelle AG Cosmetics), Aquaphyline™ (Silab), AquaregulK™ (Solabia), Carciline™ (Greentech), Codiavelane™ (BiotechMarine), Dermaflux™ (Arch Chemicals, Inc), Hydra'Flow™ (Sochibo), Hydromoist L™ (Symrise), RenovHyal™ (Soliance), Seamoss™ (Biotech Marine), Argireline™ (trade name for acetyl hexapeptide-3 from Lipotec), Gatuline Expression™ (Gattefossé) extract of Acmella oleracea, Boswellia serrata extract (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™, Revitat™, Resistem™, Chronodyn™, Kombuchka™, Chromocare™, Calmosensine™, Glycokin factor S™, Biobustyl™, Idealift™, Ceramide 2™, Ceramide A2™, Ceramide HO3™, Legance™, Intenslim™, Prodizia™, Beautifeye™, Pacifeel TM , Zingerslim™, Meiritage™, Sebuless™, Apiscalp™, Rubistem™, Citystem TMNeonyca TM ,NG Shea Butter Unsaponifiables TM Majestem TM Hydrogenesis TM Poretect TM Crystallide TM Amberstem TM , Synchrolife™, Sylverfree™, Feminage™, Ameyezing™, BB-Biont™, Revitalide™, Mel[o]stem TM Luceane TM Or a mixture thereof.

[0051] Among plant extracts (in the form of conventional extracts or prepared by in vitro methods) that can be used as additional active agents, the following extracts may be specifically mentioned: ivy (e.g., climbing ivy (English ivy)). Ivy spiral ))), Bupleurum chinense ( Chinese Bupleurum ), Altai Bupleurum ( Bupleurum falcatum Arnica (Yangju) Mountain arnica L.)), rosemary (rosemary ( Rosemary officinalis N.)), Calendula (Calendula ( Marigold officinalis )), Sage (European Sage ( Sage officinalis L.)), ginseng (ginseng ( Panax ginseng )),ginkgo( Ginkgo biloba St. John's wort (St ... Hypericum perforated )) 、 Butcher's Tree (Butcher's Tree) Ruscus aculeatus L.)), Spiraea (Zootopia 'Zootopia') Filipendula ulmaria L.)), cat's whiskers (cat's whiskers ( Orthosiphon stamincus Benth.), artichoke (Artichoke ( Cynara scolymus ), seaweed (fucus vesiculosus) Vesicular fly ), birch (white birch ( Birch white ), green tea, cola nuts (white cola ( Clean filter Horse chestnut, bamboo, centella asiatica ()), horse chestnut, bamboo, centella asiatica () Centella asiatica ), photinia, algae (wrack), willow, pilosella, horse chestnut extract, atractylodes extract, golden chamomile ( Chrysanthemum indica ) extract, apricot ( Armenian ) plants, Atractylodis flat-leaved , Synonyms pharbitidis, genus *Pharbitis* Flanders ), Coleus (such as Coleus pubescens) C. forskohlii ), Coleus ( C. blumei ), hairy calyx sheath stamen ( C. esquirolii ), Five-colored Su ( C. scutellarioides ), Yellow-sheathed stamen flower ( C. xanthan ) and hairy throat coleopterus ( C. bearded Such as extracts from the roots of Coleus forsythia, extracts from Prunus armeniaca, and extracts from the genus Tripterygium (…). Guioa ), Drynaria fortunei ( Davallia ), Terminalia genus ( Terminalia ), genus *Gnaphalium* ( Barringtonia ), genus *Hemp* ( Trembling ), Antioquia genus *Horntree* ( Cecropia ), genus Caprifolia ( Argan ), Dioscorea ( Dioscorea Such as yam ( Dioscorea opposita ) or Mexican tortoise-shell dragon ( Dioscorea mexicana ), toothpick celery ( Ammi's hometown Extracts of ) and Siegesbeckia ( Siegesbeckia Especially Siegesbeckia orientalis ( Siegesbeckia orientalis ) 、 From Ericaceae ( Ericaceae Plant extracts (especially European blueberry (dwarf blueberry)) Vaccinium angustifolium )), Bearberry ( Bearberry Extracts of aloe vera ( ) Aloe vera Plants containing sterols (especially phytosterols) Manjistha (Madder genus) Blonde Plants, especially madder ( Rubia cordifolia (extracts) Guggal (Myrrh) Commiphora ) plants, especially myrrh (mukur myrrh) Commiphora mukul Extracts of ), Kola extract, chamomile, red clover, kava pepper ( Peppermint (From Sederma's Kava Kava), False Purslane ( Bacopa monnieri (Bacocalmine™, Sederma) and sea whip, licorice ( Glycyrrhiza glabra ), mulberry, Melaleuca genus ( Melaleuca (Tea tree), Extremely forked Laria ( Larrea spread apart ), broken rice strips ( Red rhododendron ), slender Euglena ( Euglena slender ) 、 Heavenly Vine ( Cut leeches), Jungle sorghum ( Chaparral sorghum Sunflower, Green Flower Ennan Tomato ( Enantia chlorantha ), Buttongrass genus ( Sperm cell )of Mitracarpus , Buchu barosma , frothy flowers ( Lawsonia unarmed L.), maidenhair fern ( Adiantium hair-veneris L.), Celandine ( Greater celandine ), loofah ( Cylindrical loofah ), Japanese mandarin (Wenzhou mandarin ( Citrus reticulata white var. unshiu)), tea Camellia sinensis ), White Grass ( Cylindrical Imperata ), Yellow Flower Sea Poppy ( Yellow Glaucium Mediterranean cypress ( Cypress evergreen ), Kowloon Ring Polygonum multiflorum ), Lovely Hemsleya, Elderberry ( Black elderberry ), cotton beans ( Phaseolus lunatus ), genus *Lysimachia* ( Centaury Giant kelp Macrocystis pyrifera ), creeping clock flower ( Turnera diffusa ), Anemarrhena asphodeloides ( Anemarrhena asphodel ), hairy purslane ( Hairy purslane ), Hops ( Hops hops Small-bean coffee Arabic coffee Paraguayan holly ( Ilex paraguariensis ), Heart-leaved Ground Flower ( Globularia heart-shaped ), acid wood ( Oxydendron arboreum ), Albizia ( Albizzia julibrissin ), red ginger ( Ginger root Smith), Astragalus ( Astragalus membranaceus Atractylodes macrocephala ( ) Atractylodes macrocephala ), longleaf plantain ( Plantago lanceolata ), Alpine Edelweiss ( Alpine Leontopodium (or eldelweiss), four o'clock flower ( Wonderful jalapa ), dry celery ( Sweet celery ), European summer solstice ( Common horehound ), large-leaved Buddleja ( Buddleja davidii Franch), American mint ( Monarda twin ),Lavender( Lavandula angustifolia Orchid.

[0052] The compositions of the present invention may comprise peptides, including but not limited to dipeptides, tripeptides, tetrapeptides, pentapeptides, and hexapeptides and their derivatives. According to a particular embodiment, the concentration of one or more additional peptides in the composition ranges from 1 x 10⁻⁶. -7 weight% to 20% by weight, preferably 1x10 -6 From % to 10% by weight, preferably 1x10 -5 From 5% to 5% by weight.

[0053] In the context of this invention, the term "peptide" refers to peptides containing ten or fewer amino acids, their derivatives, isomers, and complexes with other substances such as metal ions (e.g., copper, zinc, manganese, magnesium, etc.). The term "peptide" refers to both natural peptides and (bio)synthetic peptides. It also refers to compositions containing peptides that are found in nature and / or are commercially available.

[0054] Non-limiting examples of dipeptides used in the context of this invention include carnosine (β-AH), YR, VW, NF, DF, KT, KC, CK, KP, KK, TT, PA, PM, or PP.

[0055] Non-limiting examples of tripeptides are RKR, HGG, GHK, GGH, GHG, GKH, KPK, KFK, KavaK, KβAK, KabuK, KacaK, KPK, KMOK, KMO2K (MO2 is sulfoxide methionine), PPL, PPR, SPR, QPA, LPA, or SPA.

[0056] Non-limiting examples of tetrapeptides are KTFK (SEQ ID NO: 1), GQPR (SEQ ID NO: 2), RSRK (SEQ ID NO: 3), KTAK (SEQ ID NO: 4), KAYK (SEQ ID NO: 5), KFYK (SEQ ID NO: 6), TKPR (SEQ ID NO: 7), AVPG (SEQ ID NO: 8), VPGA (SEQ ID NO: 9), LKLE (SEQ ID NO: 10), ELED (SEQ ID NO: 11), or LLAN (SEQ ID NO: 12).

[0057] Non-limiting examples of pentapeptides are KTTKS (SEQ ID NO: 13) and KTSKS (SEQ ID NO: 14).

[0058] Non-limiting examples of hexapeptides are GKTTKS (SEQ ID NO: 15) or VGVAPG (SEQ ID NO: 16).

[0059] Other peptides used in the context of this invention may be selected from (this list is not limiting): lipophilic derivatives of peptides, preferably palmitoyl (Pal) derivatives or myristoyl (Myr) derivatives, and metal complexes as described above (e.g., copper complexes of tripeptides HGG or GHK).

[0060] Preferred dipeptides include, for example, N-palmitoyl-β-Ala-His, Pal-KT, and Pal-RT (Sederma). Preferred tripeptide derivatives include, for example, copper derivatives of HGG (Lamin™, Sigma), Pal-GHK, Lipospondin (N-transoleoyl-KFK) and its conservatively substituted analogs, N-acetyl-RKR-NH2 (peptide CK+), Pal-KavaK, Pal-KβAlaK, Pal-KabuK, Pal-KacaK, Pal-KMO2K, N-Biot-GHK (Sederma), and its derivatives.

[0061] Here we can also mention the general formula X–Pro described in WO2015181688. –Pro –Xaa–Y is an anti-aging tripeptide, where Xaa is selected from Leu, Arg, Lys, Ala, Ser, and Asp, and at the N-terminus, X is selected from H, -CO-R. 1 and SO2-R 1 Furthermore, at the C-terminus, Y is selected from OH, OR1, NH2, NHR1, or NR1R2, and R1 and R2 are independently selected from alkyl, aryl, aralkyl, alkylaryl, alkoxy, and aryloxy groups, which can be straight-chain, branched, cyclic, polycyclic, unsaturated, hydroxylated, carbonylated, phosphorylated, and / or sulfided. These groups may have heteroatoms in their main chain, particularly O, S, and / or N, and Pro Corresponding to proline, its analogues or derivatives; including, for example, Myr-PPL-OH and Myr-PPR-OH.

[0062] Here we can further refer to the general formula X–(Xaa1)n–Pro disclosed in WO2014 / 080376. –Xaa2–Y is a staining-promoting and / or pro-MEC dipeptide and tripeptide, where n = 0, 1, or 2; Xaa1 is a hydrophobic amino acid selected from Ala, Val, Met, Leu, Iso, Phe, Pro, and its analogues and derivatives; or a polar amino acid selected from Ser, Thr, Tyr, Asp, Glu, and its analogues and derivatives; and when n = 2, the two amino acids Xaa1 are the same or different; Xaa2 is a hydrophobic amino acid selected from Ala, Val, Met, Leu, Iso, Phe, and its analogues and derivatives. Biological, or basic amino acids selected from Arg, Lys, His, and their analogues and derivatives; at the N-terminus, X is selected from H, -CO-R1, and -SO2-R1; at the C-terminus, Y is selected from OH, OR1, NH2, NHR1, or NR1R2; R1 and R2 are independently selected from alkyl, aryl, aralkyl, alkylaryl, alkoxy, and aryloxy groups, which can be straight-chain, branched, cyclic, polycyclic, saturated, unsaturated, hydroxylated, carbonylated, phosphorylated, and / or sulfided, said groups having or not having O, S, and / or N heteroatoms in their skeleton, and Pro Corresponding to proline, its analogues or derivatives; including, 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.

[0063] Suitable tetrapeptide derivatives for use as other peptides according to the invention include, but are not limited to, Ela-KTAK (SEQ ID NO: 17), Ela-KAYK (SEQ ID NO: 18), Ela-KFYK (SEQ ID NO: 19), Pal-GQPR (SEQ ID NO: 20) or Pal-KTFK (SEQ ID NO: 21).

[0064] Suitable pentapeptide derivatives for use as other peptides herein include, but are not limited to, Pal-KTTKS (SEQ ID NO: 22), Pal-KTSKS (SEQ ID NO: 23), and Pal-YGGFXaa (SEQ ID NO: 24), wherein Xaa is Leu or Pro.

[0065] Suitable hexapeptide derivatives for use as other peptides herein include, but are not limited to, Pal-HLDIIXaa (SEQ ID NO: 25) (where Xaa is Trp, Phe, Tyr, Tic, 7-hydroxy-Tic or Tpi, or a mixture thereof), Pal-GKTTKS (SEQ ID NO: 26), and Pal-VGVAPG (SEQ ID NO: 27).

[0066] Preferred compositions that are commercially available and sold by Sederma:

[0067] - Tripeptides or derivatives include Biopeptide-CL™, Maxi Lip™, or Procapil™ containing GHK;

[0068] - Tetrapeptides or derivatives include RIGIN™ and Eyeliss™ containing Pal-GQPR (SEQ ID NO: 20) and excipients.

[0069] - Pentapeptides or derivatives, such as Matrixyl™ containing Pal-KTTKS (SEQ ID NO: 22).

[0070] You can also mention:

[0071] - A mixture of Pal-GHK and Pal-GQPR (SEQ ID NO: 20) (Matrixyl™ 3000), and

[0072] - A mixture of Pal-GHK and Pal-VGVAPG (SEQ ID NO: 27) (Biobustyl™)

[0073] The following peptides and other active ingredients may be mentioned:

[0074] - Vialox™, Syn-ake™ (β-Ala-Pro-Dab-NH-Bzl), or Syn-Coll™ (Pal-Lys-Val-Lys-OH) sold by Pentapharm;

[0075] - 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), and Trylagen, marketed by Lipotec. TM(INCI name = Pseudomonas aeruginosa fermentation product extract, hydrolyzed wheat protein, hydrolyzed soybean protein, tripeptide-10 citrulline (reaction product of citrulline and tripeptide-10 (a synthetic peptide composed of aspartic acid, isoleucine and lysine)), tripeptide-1), Eyeseryl™ (Ac-β-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 (a synthetic peptide composed of aspartic acid, isoleucine and lysine);

[0076] - Collaxyl™ (Gly-Pro-Gln-Gly-Pro-Gln (SEQ ID NO:32)) or Quintescine™ (Cys-Gly) sold by Vincience;

[0077] - Cytokinol™ LS (casein hydrolysate) sold by Les Laboratoires Serobiologiques / Cognis;

[0078] - Kollaren™ (Gly-His-Lys), IP2000™ (Pal-Val-Tyr-Val), or Meliprene™ (INCI name = monofluoroheptapeptide-1: a reaction product of acetic acid and a synthetic peptide containing arginine, glycine, glutamic acid, histidine, leucine, p-fluorophenylalanine, and tryptophan) sold by l'institut Européen de Biologie Cellulaire;

[0079] - Neutrazen™ (Pal-His-D-Phe-Arg-NH2) sold by Innovations; or

[0080] - BONT-L-Peptide™, Timp-Peptide™, or ECMModuline™ sold by lnfinitec Activos.

[0081] It is also conceivable to combine the present invention with one or more cyclic peptides, particularly the cyclic peptides extracted from flaxseed oil described in the applicant’s patent application WO2019 / 149450.

[0082] More specifically, pycnidione may be combined with at least one of the following: compounds selected from vitamins B, C, E, F, D and A, particularly compounds such as nicotinamide or tocopherol, retinoid compounds such as retinol, hyaluronic acid, hexamidine, alpha-lipoic acid, resveratrol, or DHEA, peptides, ceramides, and chemical or physical UV or IR filters, which are conventional active ingredients used in topical cosmetic compositions.

[0083] The present invention also provides the use of pycnidione in preparing compositions for cosmetic treatments as described above.

[0084] The compositions according to the invention can be applied to the face, body, neckline, scalp, hair, eyelashes, and body hair in any form or medium known to those skilled in the art, particularly as a solution, dispersion, emulsion, paste, or powder alone or as a premix, or as a medium alone or as a premix in combination, incorporated into or adsorbed in a carrier such as macrocapsules, microcapsules, or nanocapsules, macrospheres, microspheres, or nanospheres, liposomes, oleosomes, or chylomicrons, macroparticles, microparticles, or nanoparticles, or macrosponges, microsponges, or nanosponges, microemulsions, or nanoemulsions, or adsorbed on organic polymer powders, talc, bentonite, spores, or exoskeletons and other inorganic or organic carriers.

[0085] In cosmetics, it can be specifically applied in the range of skin care for the face, body, hair and body hair, as well as in the range of makeup treatments (especially eyelashes and eyebrows).

[0086] For example, the composition may be in the form of an emulsion, cream, lotion, milk, solid form, foam, gel, deodorant, antiperspirant, shampoo, conditioner, hair mask, face mask, shower gel, etc.

[0087] Galen formulations can be used in various product ranges for personal care and / or beautification products, including skin care, cleansing, makeup, cleansing, sun protection, artificial tanning, pre-shave, during-shave or post-shave, moisturizers, humectants, emollients, conditioning agents, exfoliants, astringents, depilatory or antiperspirant, deodorants, etc.

[0088] The composition can be incorporated into nonwoven or woven materials having natural or synthetic fibers, wool, or any material intended to come into contact with the skin and which can be used in clothing, including tights and socks, shorts, day or night underwear, tissues, handkerchiefs, or fabrics, to exert its cosmetic effects and achieve continuous local delivery (cosmetic textiles) through contact with the skin / textiles.

[0089] In a specific case of the cosmetic hair treatment according to the present invention, the composition may contain at least one surfactant, which may be selected from anionic, nonionic, amphoteric and / or cationic surfactants.

[0090] Suitable anionic surfactants include alkyl sulfates, alkyl ether sulfates, α-olefin sulfonates, sulfosuccinates, hydroxyethyl sulfonates, acylamides, acylglutamates, alkyl ether carboxylates, and alkyl phosphates. The alkyl group preferably comprises 6 to 30, more preferably 8 to 20, particularly 10 to 14, and especially 12 carbon atoms. Alkyl ether sulfates and / or alkyl sulfates are preferred, particularly alkali metals, such as their sodium and / or ammonium salts. Lauryl ether sulfates and / or lauryl sulfates are particularly preferred anionic surfactants.

[0091] Suitable nonionic surfactants include fatty alcohols or amide ethoxylates, alkanolamides and alkoxylated alkanolamides, monoglyceride ethoxylates, sorbitol ester ethoxylates, alkyl polyglycosides, ethylene glycol monoesters, ethylene glycol diesters and mixtures thereof.

[0092] Suitable amphoteric surfactants include alkyliminodipropionates, alkyl amphoteric glycinates, alkyl amphoteric propions, alkyl amphoteric acetates (mono- and di-), N-alkyl β-aminopropionic acid, alkyl polyaminocarboxylates, phosphorylated imidazolines, and mixtures thereof.

[0093] Suitable cationic surfactants include alkyl quaternary ammonium salts, benzyl quaternary ammonium salts, quaternary esters, ethoxylated quaternary ammonium salts, alkylamines, and mixtures thereof. The alkyl groups preferably comprise 6 to 30, more preferably 8 to 22, and particularly 10 to 20 carbon atoms.

[0094] Cationic surfactants can also be polyquaternium (or polyquat) materials. Polyquaterniums include acrylamide-based polymers and / or dimethylallyl ammonium chloride-based polymers, such as polyquaternium 6, polyquaternium 7, etc. Guar gum polymeric quaterniums, such as guar hydroxypropyltrimethylammonium chloride, can be used. Cellulose polymeric quaterniums, such as polyquaternium 10, and starch polymeric quaterniums can also be used.

[0095] The hair care composition may also contain at least one secondary surfactant. If present, the secondary surfactant may be selected from nonionic, amphoteric, betaine, and / or cationic surfactants.

[0096] Suitable betaines include alkyl betaines, alkylamido betaines, alkyl sulfono betaines, alkylamido sulfono betaines, and mixtures thereof. Alkylamido betaines are preferred. The alkyl group preferably contains 6 to 30, more preferably 8 to 20, and particularly 10 to 14 carbon atoms.

[0097] The hair composition may also include lubricants such as talc, magnesium stearate and mineral oil, wetting agents, emulsifiers and suspending agents, preservatives such as methylparaben and propylparaben, sweeteners and fragrances.

[0098] It may include one or more other standard ingredients or carriers commonly used in hair care products, including shine enhancers, moisturizers, herbal additives, hair strengthening agents, vitamin additives, colorants, hair thickeners, fixatives and styling agents, UV absorbers, silicone oils, essential oils and fragrances, thickeners or viscosity improvers, detergents, stabilizers, emollients, chelating agents or sequestering agents, preservatives, disinfectants, antioxidants / free radical scavengers, antistatic agents, conditioning agents, detangling ingredients, emulsifiers or dispersants, stimulants, softeners, solvents, carriers, etc.

[0099] It may include additional hair care active ingredients to support or complement its activity, including the following active ingredients sold by Sederma:

[0100] Active ingredients that act on the scalp:

[0101] - Apiscalp TM Celery seed extract, obtained by titration with ligusticum lactone and supercritical CO2 extraction, functions to relieve dandruff and reduce scalp itching. It also moisturizes and reduces sebum production.

[0102] Hairspa TM The combination of lactitol and xylitol in glycerin soothes and moisturizes the scalp. It helps combat scalp discomfort—dryness, itching, dandruff, and irritation—by rebalancing the skin's microbiome.

[0103] Active ingredients that affect hair quality or color:

[0104] FruitBio TM This is a complex of alpha-hydroxy acids obtained through lactic acid fermentation with the addition of malic acid and citric acid, combined with green tea extract. The active ingredient smooths the hair cuticles, and the green tea softens them, resulting in smoother and softer hair.

[0105] Heliogenol TMA water-diol extract of sunflower seed meal titrated with polyphenols. It possesses strong anti-free radical activity. It protects and repairs natural and color-treated hair from damage caused by shampoo and UV radiation.

[0106] - Ceramide A2 TM Ceramide 2 is a chemical analogue of ceramide 2 (a natural molecule in hair). Its function is to restructure, smooth, and strengthen hair, restoring the volume, shine, and vitality of damaged hair.

[0107] - Sylverfree TM A hydrated glycerol solution of palmitoylprolylproline (Pal-PP). It helps combat graying hair.

[0108] Active ingredients that promote hair growth:

[0109] - Capigen TM It is composed of high taurine, synthetic taurine analogs, natural amino acids, peptide-rich bacterial filtrate, and marine-derived sulfopolysaccharide extract. It is a hair growth stimulant and helps prevent hair loss.

[0110] - Procapil TM This product combines vitamin-rich matrikine (Biotinyl-GHK) with apigenin (a natural flavonoid) and naturally occurring oleanolic acid. It strengthens and rejuvenates hair follicles to slow hair loss. It promotes hair anchoring by strengthening metabolism and hair follicle structure. Oleanolic acid inhibits 5α-reductase, apigenin improves microcirculation, and biotinyl-GHK stimulates cell metabolism.

[0111] - Kelisoft TM Chelidonin is a highly purified plant molecule used as an excipient. Its function is to reduce hair growth.

[0112] - SPHINGO HAIR TM The series of hair care ceramides, especially Sphingo Hair™ Drypure™, are bio-fermented ceramide NPs that replenish the skin with essential longer-chain ceramides and promote a healthy scalp and hair barrier structure.

[0113] The hair composition may also contain an anti-dandruff active agent selected from one of the following: nystatin, copper micaceous, tolnaftate, candidacloprid, haloperidol, clopidogrel, clotrimazole, undecanoic acid, propionic acid, caprylic acid, benzoic acid, salicylic acid, griseofulvin, amphotericin B, ketoconazole, miconazole, pembrolizumab, haromycin, cymoxanil, schizomycin, bifonazole, butoconazole, econazole, fenteconazole, isoconazole, omeconazole, oxiconazole, sertaconazole, thioconazole, tiaconazole, abaconazole, fluconazole, isaconazole, itraconazole, posaconazole, rivanazole, tebuconazole. The following are included: azole, voriconazole, abafenidine, amorolfen, butenafine, naftifine, terbinafine, anidifine, caspofungin, micafungin, iclopiroxolamine, flucytosine, crystal violet, pyroxone ethanolamine salt, zinc pyrithione, selenium sulfide, coal tar, and tea tree oil, preferably selected from ketoconazole, zinc pyrithione (ZPT), pyroxone ethanolamine salt, octopirox, salicylic acid, selenium sulfide, coal tar, azelaic acid, grippazole, salicylic acid, undecanoic acid, and mixtures thereof.

[0114] Therefore, the present invention covers a method for cosmetic, non-therapeutic topical treatment for improving the appearance and overall condition of skin and its appendages, as well as their sensory comfort, comprising applying an effective amount of pycnidione topically to the skin of a subject in need, the treatment being as defined in the examples given above and below.

[0115] Detailed Explanation

[0116] The invention will be better understood from the following detailed description of exemplary embodiments.

[0117] 1- Preparation Example of Extract Containing Pycnidione

[0118] The microorganism used: *Echinochloa oryzae* strain available from the Culture Collection of Fungi (CBS).

[0119] Pre-inoculation is performed using cryopreserved spores or mycelia in a specific pre-inoculation medium (containing gelling agents, sugars, and nitrogen in the form of peptone or yeast extract). Several pre-inoculation steps are performed to increase biomass.

[0120] Once the pre-inoculum has grown sufficiently, the resulting biomass is used to inoculate the bioreactor with a rice- and micronutrient-based culture medium. In the bioreactor (also known as the generator), the biomass expands, producing pycnidione.

[0121] Pycnidione yields are highest after 7 days of incubation in the generator.

[0122] Then, solid / liquid separation techniques such as centrifugation, centrifugal decanting, or filtration via a filter press are used to harvest the biomass. The supernatant is discarded.

[0123] The harvested biomass was mixed with a solvent (designed to form a physiologically acceptable excipient, consisting of a mixture of pentylene glycol (PTG) and butylene glycol (BG) (20 / 80 w / w)) and stirred at 60°C for 1 hour to extract pycnidione into the excipient. Following extraction, cell debris was removed by clarification using known solid / liquid separation techniques, as mentioned above.

[0124] Analysis of the dry extract determined a final concentration of 1200 ppm of total cycloheptatrienolone in the extract, including approximately 1100 ppm of pycnidione (the target active molecule according to the invention) and the remaining cycloheptatrienolone (including, in particular, Epolone A and Epolone B).

[0125] 2- Examples of preparation of active ingredients for the treatment according to the present invention

[0126] The PTG / BG extract (containing physiologically acceptable excipients) prepared in point 1 below constitutes the active ingredient, which may be used in cosmetic formulations in a defined weight percentage, depending on the desired effect (whether more or less pronounced), as explained in the description above. Various cosmetic formulations using this cosmetic ingredient are further described in point 4.

[0127] 3- In vitro efficacy testing

[0128] Test with the dried extract of the biomass prepared in step 1 above, diluted in DMSO, or with pure pycnidione (approximately 82%) also diluted in DMSO. The ppm values ​​indicated in the table below correspond to the ppm values ​​of pycnidione molecules.

[0129] All statistical analyses were performed using Student's t-test to determine the significance of the results in paired comparisons (controls versus treated).

[0130] 3.1- Stimulation of hyaluronic acid synthesis

[0131] Hyaluronic acid is an essential component of the skin. It is present in the dermis and epidermis, and due to its unusual three-dimensional structure, it can capture thousands of times its weight in water and thus occupy a very large volume relative to its molecular weight. The intracellular spaces of the extracellular matrix are filled with water-swollen hyaluronic acid, which gives the skin its excellent viscoelastic properties.

[0132] Therefore, by stimulating hyaluronic acid synthesis, hydration is maintained or increased, resulting in more elastic and supple skin with a smoother texture and a more comfortable feel, without stinging or tightness.

[0133] plan:

[0134] Human keratinocytes (HK) were cultured to a subconfluence and then contacted with the active ingredient according to the invention. Following this contact, hyaluronic acid synthesis was assessed using an ELISA assay. Cell count estimation using the Hoechst method allowed for normalization of the results.

[0135] result

[0136] Changes in hyaluronic acid production by keratinocytes compared to the control group.

[0137] [Table 1]

[0138]

[0139] The results showed that pycnidione significantly increased hyaluronic acid synthesis in keratinocytes.

[0140] 3.2- Enhance the dermal-epidermal junction (DEJ)

[0141] DEJ ensures cohesion between the epidermis and dermis. During skin aging (whether normal or accelerated by external factors), there is a decrease in the synthesis of DEJ components, particularly laminin and collagen VII, leading to disorder, with effects on skin elasticity and resilience, and loss of vitality.

[0142] plan

[0143] KH cells were cultured to subconfluence and then contacted (or not contacted, for control) with the active ingredient according to the invention. Following contact, collagen VII and laminin synthesis were assessed using an ELISA kit. Cell counts were estimated using the Hoechst method to normalize the results.

[0144] result

[0145] Compared to the control, changes in the production of laminin and collagen VII by keratinocytes:

[0146] [Table 2]

[0147]

[0148] The results showed that pycnidione significantly increased the production of laminin and collagen VII in keratinocytes.

[0149] 3.3- Anti-glycation:

[0150] The human body needs sugar to produce energy. However, the body never fully utilizes the sugar it consumes for this beneficial purpose. The remaining portion of the absorbed sugar reacts non-enzymatically with the amino groups of proteins, nucleic acids, or lipids to produce advanced glycation end products (AGEs), called glycotoxins, or AGEs, which accumulate in tissues over time. Glycation alters the function, enzymes, and structural properties of molecules, affecting the normal functioning of cells and the body.

[0151] In the skin, AGEs are present not only in the dermis but also in the epidermis, all the way down to the stratum corneum. Tissues are particularly affected when proteins such as collagen, elastin, fibronectin, and laminin are present for extended periods. This leads to changes in the mechanical and elastic properties of the extracellular matrix (ECM) in the dermis, making it less supple, more rigid, and also more relaxed and less reactive.

[0152] In addition, mitochondrial proteins in skin cells are also glycated, leading to reduced efficiency in the production of adenosine triphosphate (ATP) and impaired energy production.

[0153] Therefore, AGEs represent a major source of cellular dysfunction, and their accumulation often alters the properties and performance of tissues and cells: reduced flexibility, reduced mobility, reduced responsiveness, and reduced energy production. Glycation leaves its mark on the face, for example, with dark circles under the eyes, puffy eye bags, dull skin tone, and a tired appearance, as well as less responsive, tired, and sagging skin. Studies have also shown that the amount of AGEs is related to hair quality. Decreased protein content and increased AGEs lead to poorer hair quality with reduced breakage resistance.

[0154] Therefore, glycation manifests visually on the skin as the appearance of wrinkles and fine lines, dullness, sagging, lack of luster, tired skin, and skin lacking tone and smoothness, and on the hair as poorer quality hair, split ends, and brittleness.

[0155] plan

[0156] Studies of non-enzymatic glycation were conducted between a model protein (serum albumin) serving as the target and edible reducing sugars derived from fruit. In the presence or absence of the product according to the invention, the protein irreversibly and gradually undergoes glycation (binding to sugar). This change is monitored by fluorescence.

[0157] result

[0158] Compared to the control, the changes produced by AGE:

[0159] [Table 3]

[0160]

[0161] The results showed that pycnidione has strong anti-glycation potential.

[0162] 3.4 - Reduced synthesis of pro-inflammatory molecules:

[0163] Skin subjected to sustained stress (exposure to UV rays, smog, pollutants, etc.) triggers direct or indirect inflammatory responses. Even low-intensity, uncontrolled or persistent inflammatory responses lead to the production of cytokines designed to attract or stimulate other cells, triggering a cascade of reactions. The resulting pro-inflammatory microenvironment alters skin homeostasis and gradually leads to changes or even damage to cellular and tissue biomolecules. It also disrupts the integrity of the skin barrier. Therefore, the inflammatory mediators IL-6 and IL-8 are known to induce premature aging through microinflammation. Furthermore, sensitive and irritated skin is characterized by abnormally high secretion of these cytokines. By reducing the production of these cytokines after stress, the present invention demonstrates a preventative anti-inflammatory effect. Cells are better protected. From a cosmetic perspective, the treatment according to the invention allows for the prevention of discomfort, such as tightness, and unsightly conditions, such as microredness of the skin, caused by external aggressors (such as pollution, radiation, or cold).

[0164] plan

[0165] Normal human keratinocytes (NHK) were seeded and exposed to different concentrations of the test active ingredient, either in contact or without contact, for 48 hours, and then irradiated with UVB. Unirradiated plates were processed in parallel. The amounts of synthesized IL-6 and IL-8 in the culture supernatant were measured using ELISA. At the end of culture, Hoechst staining was used on the fixed cell lawn to estimate cell viability (cell number estimation).

[0166] result

[0167] Changes in IL-6 and IL-8 synthesis after keratinocyte stress compared to the control:

[0168] [Table 4]

[0169]

[0170] The results showed that pycnidione induced a decrease in the synthesis of IL-6 and IL-8 after stress (UVB conditions).

[0171] 3.5- Decreased intracellular production of reactive oxygen species (ROS):

[0172] It is now recognized that the accumulation of oxidative free radicals is essential as the origin of the very biochemical disorders that cause aging. Whether generated by UV rays (via complex cascade reactions), by various pollutants that interact with the skin over a long period, or by natural aging, these free radicals will predictably and over short periods cause skin inflammation, and throughout the body and over longer periods cause skin aging. Under so-called normal conditions, in young, healthy skin relatively protected from external aggressors, there is a balance between the production of oxidative free radicals in the body and the antioxidant defenses responsible for neutralizing them. These defense systems consist of enzymes (SOD, catalase, glutathione peroxidase) or classic antioxidants (vitamin C, vitamin E, glutathione, carotene, etc.). When an imbalance occurs between production and neutralization, there is an excess of oxidative free radicals, causing oxidative stress. This leads to damage to DNA, proteins, and cellular lipids, which undergo oxidation, causing changes in their structure and function. This leads to signs of premature aging, damage to the dermis and epidermis, accompanied by the loss of their mechanical and viscoelastic properties, thinning of the skin, sagging skin, fine lines, wrinkles, spots, etc.

[0173] plan

[0174] Normal human fibroblasts (NHF) were seeded and exposed to the product for 24 hours. After removing the active ingredient, the cells were washed and loaded with the DCFH-DA probe, which became fluorescent after being metabolized by cellular enzymes in the presence of ROS. This fluorescence was further enhanced when a stressor (H2O2) was applied to the cells. The active ingredient reduces both basal fluorescence and induced fluorescence (ROS-induced fluorescence).

[0175] Cell viability was assessed in parallel using Hoechst staining (DNA intercalation reagent).

[0176] result

[0177] Compared with the control, the changes in ROS synthesis by fibroblasts under conditions of oxidative stress and without:

[0178] [Table 5]

[0179]

[0180] Pycnidione significantly reduced the generation of reactive oxygen species (ROS) after H2O2 stress and significantly reduced the generation under basal conditions (non-stress conditions), without dose-dependent effects.

[0181] Pycnidione helps combat the formation of intracellular ROS (known for its role in skin aging).

[0182] 3.6- Anti-lipid peroxidation:

[0183] Lipid peroxidation is the peroxidation of the lipid membranes surrounding cells and mitochondria, leading to their weakening and amplifying damage through the autocatalytic generation of reactive oxygen species (free radicals).

[0184] plan

[0185] Lipid peroxidation is assessed using a test that monitors the occurrence of peroxidation of lipid membranes in liposome form after oxidative UVA radiation (measuring the amount of conjugated dienes formed). The product is then used in this test after irradiation.

[0186] result

[0187] Changes in lipid peroxidation compared to the control:

[0188] [Table 6]

[0189]

[0190] The results demonstrate the strong potential of pycnidione to combat lipid peroxidation and its consequent detrimental effects on cell membranes and their vitality.

[0191] 3.7- Reduces the growth of Propionibacterium acnes (… C. acnes Proliferation of )

[0192] The skin microbiome, a highly complex ecosystem composed of a variety of living microorganisms (bacteria, yeasts, viruses, and parasites), has multiple functions: defense, skin barrier, and immune system regulation. Maintaining its balance is crucial by preventing certain substances from damaging the skin due to overgrowth. For example, *Propionibacterium acnes* (… Cutibacterium acnes Acne bacteria are a case in point. Although they are part of the normal skin microbiome, if they multiply too rapidly, they promote the proliferation and migration of keratinocytes, participate in the formation of free radical substances such as superoxide anions, and trigger cascade reactions that lead to the production of pro-inflammatory molecules, thereby contributing to the development of acne.

[0193] By slowing or inhibiting the growth of Propionibacterium acnes, the epidermis is protected from acne. This effect is of particular concern in cosmetic treatments for so-called oily and acne-prone skin. This type of skin typically corresponds to oily skin during puberty due to excessive sebum production caused by hormonal reasons. By preventing or limiting the proliferation of Propionibacterium acnes, the formation of waste in the hair sheath within the hair follicle is prevented. This is important because bacteria in the hair sheath grow anaerobically by feeding on the sebum and producing waste (including dead cells), leading to blockage of the hair follicle sheath (blackheads), inflammation, and eventually acne papules, the treatment of which is a dermatological problem. Therefore, reducing the amount of Propionibacterium acnes helps prevent the development of a pre-acne state.

[0194] plan

[0195] Under anaerobic conditions at 37°C, suspensions of Propionibacterium acnes at equivalent densities were cultured in a suitable culture medium with or without the test product (test case). After incubation for 48 to 72 hours, bacterial population growth was measured by reading the optical density (OD) at 600 nm.

[0196] result

[0197] Changes in the proliferation of Propionibacterium acnes compared to the control:

[0198] [Table 8]

[0199]

[0200] The results showed that pycnidione significantly slowed the proliferation of Propionibacterium acnes bacteria that cause the pre-acne state.

[0201] 4- Examples of cosmetic formulations

[0202] Various cosmetic formulations are described below. Additional active ingredients may be added to suitable phases based on their hydrophobic or hydrophilic properties, where appropriate, these additional active ingredients support and / or complement the activity of the active ingredients according to the invention. These ingredients may belong to any category, depending on their function, application site (body, face, neck, chest, hands, hair, eyelashes, eyebrows, body hair, etc.), desired end effect, and target consumer, such as antioxidants, moisturizers, nourishing agents, protective agents, smoothing agents, reshaping agents, plumping agents, agents that act on skin tone for shine, anti-spot agents, anti-dark circle agents, anti-glycation agents, slimming agents, soothing agents, muscle relaxants, anti-redness agents, anti-stretch mark agents, etc. They are mentioned above in the description.

[0203] According to the invention: the ingredients prepared in paragraph 2) above (an in vitro prepared extract containing approximately 1100 ppm of pycnidione in a mixture of butylene glycol and pentanediol).

[0204] Recommended effective amount of ingredient: for example, 1% to 15%, preferably 1% to 10%, more preferably about 1% to 5%, which corresponds to about 10 ppm to 60 ppm of pycnidione, expressed as 3% (about 35 ppm) in the examples of the following formulations.

[0205] 4.1 - Lightweight cream formula suitable for face

[0206] [Table 9]

[0207]

[0208] preparation:

[0209] Weigh part A. Weigh and mix part B. While stirring, slowly add part B to part A and mix thoroughly for 30 minutes. Heat part A+B in a water bath to 75°C. Weigh part C and heat it in a water bath to 75°C. While stirring vigorously, add part C to part A+B. Add part D to part A+B+C until approximately 45°C. While stirring, add part E to the previous part. If applicable, add part F to the previous part and mix thoroughly.

[0210] Obtain a light, non-sticky cream that can be used, for example, as a face cream for relaxing massage applications.

[0211] 4.2 - Cream formulations suitable for day creams

[0212] [Table 10]

[0213]

[0214]

[0215] preparation:

[0216] Weigh part A and mix thoroughly with regular stirring. Weigh part B and mix. Pour part B into part A with regular stirring for one hour. Heat part A+B in a water bath to 75°C. Weigh part C and heat in a water bath to 75°C. Add part C to part A+B and stir rapidly. Weigh part D and add to the previous part at approximately 45°C, mixing thoroughly. Weigh part E and add to the previous part, mixing thoroughly. If necessary, weigh part F and add to the previous part, mixing thoroughly.

[0217] This product is a light, opaque white cream with a soft, supple texture suitable for daytime skincare. The active ingredient Crystallide® supports and helps repair the skin, resulting in a radiant complexion.

[0218] 4.3 - Cream formula suitable for oily, acne-prone skin

[0219] [Table 11]

[0220]

[0221] preparation:

[0222] Weigh and mix portion A. Weigh and mix portion B. Add portion B to portion A while stirring for 1 hour. Heat portion A+B in a water bath to 75°C. Weigh portion C and heat it to 75°C in a water bath. Add portion C to portion A+B while stirring vigorously. Weigh and mix portion D. Add portion D to the previous portions and mix thoroughly. Add portion E and mix thoroughly. If applicable, add portion F and mix thoroughly.

[0223] Achieving an opaque, grayish-white, viscous cream, suitable for oily skin due to its dry feel and quick absorption. In conjunction with the ingredient Fruitbio... TM In combination, a gentle chemical exfoliation treatment is added to further smooth the skin's texture.

[0224] 4.4 - Cream formula suitable for hands

[0225] [Table 12]

[0226]

[0227] preparation:

[0228] Weigh portion A and mix under normal stirring. Weigh portion B and mix. Pour portion B into portion A under normal stirring for one hour. Activate portion A+B by vigorous stirring for 1 minute. Add portion C to portion A+B and stir thoroughly. Heat portion A+B+C to 75°C in a water bath. Weigh portion D and heat to 75°C in a water bath. Weigh portion E and add it thoroughly to portion D to form an emulsion. Add portion D+E to the previous portion and mix thoroughly with rapid stirring. Weigh and mix portion F. Add portion F to the previous portion at approximately 45°C and mix thoroughly. Weigh portion G and add it to the previous portion and mix thoroughly. Weigh portion H and add it to the previous portion and mix thoroughly.

[0229] Due to the ingredients according to the invention, and the additional combination with ceramides (a natural lipid component of the skin), a cream perfectly suited for treating dry hands is obtained.

[0230] 4.5 - Cream formula suitable for face and neck.

[0231] [Table 13]

[0232]

[0233]

[0234] preparation:

[0235] Weigh part A. Weigh and mix part B. While stirring, slowly add part B to part A and mix thoroughly for 30 minutes. Heat part A+B to 75°C in a water bath. Weigh part C and heat to 75°C in a water bath. Add part C to part A+B while stirring vigorously. Add part D to part A+B+C while stirring. If necessary, add part E to the previous part while stirring. Mix thoroughly during the cooling phase.

[0236] A pale yellow cream is obtained, ideal for the face and neck, and is quickly absorbed by the skin.

[0237] 4.6 - Cream formula for day creams suitable for the neckline

[0238] [Table 14]

[0239]

[0240]

[0241] preparation:

[0242] Weigh portion A and mix under normal stirring. Weigh portion B and mix. Pour portion B into portion A under normal stirring for one hour. Activate portion A+B by vigorous stirring for 1 minute. Weigh portion C. Add portion C to portion A+B and stir thoroughly. Heat portion A+B+C to 75°C in a water bath. Weigh portion D and heat to 75°C in a water bath. Weigh portion E and add it thoroughly to portion D to form an emulsion. Add portion D+E to the previous portion and mix thoroughly with rapid stirring. Weigh portion F and add it to the previous portion and mix thoroughly. Weigh portion G and add it to the previous portion and mix thoroughly.

[0243] Obtain an opaque cream lotion with a soft and lightweight texture that is quickly absorbed by the skin and can be applied to large areas such as the chest and neck.

[0244] 4.7 - Cream Emulsion Formulation

[0245] [Table 15]

[0246]

[0247] preparation:

[0248] Weigh part A and mix thoroughly under normal stirring. Weigh part B and mix. Add part C to part B and mix thoroughly. Weigh part D and add it to B+C, mix thoroughly. Weigh part E and mix thoroughly. Adjust the pH of B+C+D to pH + / - 5.40 using part E, mix thoroughly. Slowly add part B+C+D+E to part A, stirring rapidly.

[0249] A white, opaque fluid emulsion is obtained, forming an ultra-concentrated emulsion with a moisturizing cream feel comparable to a W / O emulsion.

[0250] 4.8- Hydrogel Formulation

[0251] [Table 16]

[0252]

[0253] preparation:

[0254] Heat portion A to 80°C in a water bath with stirring for 1 hour. Weigh portion B and mix thoroughly. Slowly add portion B to portion A in a water bath at 80°C with stirring. Allow portion A+B to heat to 80°C in a water bath with stirring for 1 hour. Add portion C to portion A+B in a water bath at 80°C. Add portion D to portion A+B+C and mix thoroughly. If applicable, add portion E and mix thoroughly. Pour immediately into molds while still warm.

[0255] This hydrogel provides an immediate refreshing and soothing sensation, while one or more active ingredients work deeply to soothe and protect the skin for a long time. The addition of Crodateric CAB30™, a gentle surfactant, allows for gentle cleansing of the skin with water at the end of use.

[0256] 4.9 - Concentrated Essence Formula

[0257] [Table 17]

[0258]

[0259] preparation:

[0260] Weigh part A and mix thoroughly under normal stirring. Weigh part B and mix. Pour part B into part A and continue stirring under normal stirring for one hour. Weigh part C and add it to part A+B under rapid stirring. Weigh part D and mix thoroughly. Add part D to the previous mixture and mix thoroughly. Weigh and add part E to the previous mixture and mix thoroughly. If applicable, weigh and add part F to the previous mixture and mix thoroughly.

[0261] A highly concentrated essence is obtained by forming an emulsion that resembles a gel and has the feel of a moisturizing cream. In combination, according to the ingredients of the present invention and Synchrolife... TM The ingredients provide complete regenerating nighttime care.

[0262] 4.10 - Paste Formulation

[0263] [Table 18]

[0264]

[0265] preparation:

[0266] Weigh part A and heat it to approximately 65°C. Weigh part B. While stirring, add it to part A and mix thoroughly. Weigh part C and add it to part A+B while stirring. Weigh part C and add it to the previous part at 50°C. Weigh and add part D. Mix thoroughly. If applicable, weigh and add part E and mix thoroughly. Pour immediately into a container.

[0267] Obtain an opaque beige paste, which can be used as a concealer and a screen to protect against urban pollution.

[0268] 4.11 - Cream formula for daytime care around the eyes

[0269] [Table 19]

[0270]

[0271] preparation:

[0272] Weigh part A and mix it while stirring. Weigh part B. Add part B to part A while stirring normally. Weigh part C, if applicable. Add part C to part A+B and mix thoroughly. Weigh part D. Weigh part D and mix it thoroughly while stirring normally. Add part A+B+(C) to part D while stirring rapidly and mix thoroughly.

[0273] Obtain an opaque, off-white, viscous cream, perfectly suited for eye contour care, firming and brightening the area. Feminage TM The active ingredients enhance the treatment by acting on sagging skin.

[0274] 4.12 - Spray Formulation

[0275] [Table 20]

[0276]

[0277] preparation:

[0278] Weigh part A. Weigh and mix part B. Pour part B into part A while stirring as usual. Weigh and add part C to part A+B, and mix thoroughly. Weigh and add part D to the previous part, and mix thoroughly. Weigh and mix part E. Add part E to the previous part, and mix thoroughly. If applicable, weigh and add part F to the previous part, and mix thoroughly.

[0279] A clear, pale yellow liquid is obtained, which can be used as a spray with a light texture. BB-Biont TM The added ingredients also help regulate the microbiome and smooth acne scars. The formula is especially suitable for application to treat skin covered by camouflage.

[0280] 4.13 - Emulsion formulations suitable for the body

[0281] [Table 21]

[0282]

[0283] preparation:

[0284] Weigh part A and mix thoroughly. Heat part A to 75°C in a water bath. Weigh part B and mix thoroughly. Add part B to part A while rapidly stirring at 75°C in a water bath for one hour. Combine parts A and B while stirring vigorously for 1 minute. Weigh and heat part C to 75°C in a water bath. Add part C to part A and B while stirring vigorously. Weigh part D and mix thoroughly. Adjust the pH to 5.80 + / - 0.10 with part D at approximately 45°C and mix thoroughly. Add part E to the previous part and mix thoroughly. If applicable, add part F to the previous part and mix thoroughly.

[0285] Obtain an opaque, fluid cream with a refreshing and silky texture. It can be used as a post-hair removal emulsion, for example, with the anti-hair regrowth active ingredient Kelisoft. TM combination.

[0286] 4.14 - Solid Rod Formulation

[0287] [Table 22]

[0288]

[0289] preparation:

[0290] Weigh portion A and heat it to 75°C in a water bath. Weigh portion B and add it to portion A. Weigh portion C and mix them. Heat portion C to 75°C in a water bath. While stirring rapidly, add portion C to portion A+B. Weigh portion D and mix them. While stirring rapidly, add portion D to the previous portion at approximately 50°C. Immediately pour the mixture into a mold at approximately 45°C to form a rod, which solidifies at room temperature.

[0291] A solid, opaque, light yellow stick is obtained. It can be used for quick and direct contact care. The combination of the ingredients according to the invention with the PoreTect™ ingredient allows for the integration of moisturizing, anti-aging, and specific care actions for enlarged pores. In daytime care, the Solaveyil™ CT-300 ingredient also protects against potential damage from UV rays.

[0292] 4.15 - A moisturizing cream formula with a whipped texture.

[0293] [Table 23]

[0294]

[0295]

[0296] preparation:

[0297] Weigh part A and mix thoroughly. Weigh part B and mix thoroughly. Add part B to part A and stir rapidly for one hour. Heat part A+B in a water bath to 75°C. Weigh and heat part C in a water bath to 75°C. Add part C to part A+B and stir vigorously. Weigh part D and mix thoroughly. Add part D to the previous part at approximately 45°C and stir thoroughly. Add part E to the previous part and stir thoroughly. If applicable, add part F to the previous part and stir thoroughly.

[0298] After a 24-hour interval, the emulsion is stirred with a U-shaped paddle for one hour.

[0299] The desired result is a pale yellow, opaque, viscous cream with a whipped mousse-like texture. The cream is moisturizing but leaves the skin feeling comfortable and non-greasy.

[0300] 4.16 - Roll-on lip formula

[0301] [Table 24]

[0302]

[0303]

[0304] preparation:

[0305] Weigh portion A and heat to approximately 75°C. While stirring, add portion B and mix thoroughly for 30 minutes. Weigh portion C and heat to approximately 75°C. While stirring at high speed, add portion C to portion A+B. Weigh portion D and mix thoroughly. When the temperature is below 45°C, adjust the pH to 6.00-6.20 with portion D and mix thoroughly. Weigh portion E and add it to the previous portions, mixing thoroughly. Weigh portion F and add it to the previous portions, mixing thoroughly.

[0306] Gain a deep red fluid emulsion, compatible with roll-on applications, that provides a repairing, moisturizing, and protective cosmetic treatment thanks to its active ingredients, and gives lips a plump and glossy aesthetic look.

[0307] 4.17 - Shampoo Formula

[0308] [Table 25]

[0309]

[0310] preparation:

[0311] Weigh part A. Mix thoroughly. While stirring, add the components of part B one by one to part A+B. Mix thoroughly. Weigh part C and mix thoroughly. Add part C to part A+B while stirring normally. Add part D to the previous part while stirring normally. Add part E to the previous part while stirring normally.

[0312] Achieve a clear, pale yellow fluid gel that forms a shampoo to gently cleanse and treat hair and scalp.

[0313] 4.18 - Leave-on sprays for hair applications

[0314] [Table 26]

[0315]

[0316] preparation:

[0317] Weigh and mix portion A. Weigh and mix portion B thoroughly. Weigh and mix portion C thoroughly. Mix portion B into portion C. Pour portion B+C into portion A using a regular spiral mixer. Add portion D to the previous portions using a regular spiral mixer.

[0318] Obtain an opaque, off-white fluid emulsion with a texture suitable for application by spraying, which can be gently deposited on hair and scalp.

[0319] 4.19 - Hair mask formulations for hair applications

[0320] [Table 27]

[0321]

[0322]

[0323] preparation:

[0324] Weigh part A and heat it to 85°C in a water bath. Weigh part B and heat it to 90°C in a water bath. Weigh part C and mix thoroughly. Pour part C into part A with normal stirring. Disperse part B into part A + C with vigorous stirring and mix thoroughly. If necessary, pour part D into the previous part at approximately 40°C while stirring. If necessary, add part E and mix thoroughly. If necessary, add part F to the previous part and mix thoroughly. Add part G and mix thoroughly. Add part H and mix thoroughly.

[0325] A yellow, opaque, viscous emulsion is obtained, forming a thick cream suitable for polydimethylsiloxane-free hair and scalp masks, which may include a range of active ingredients such as those according to the invention for complete cosmetic treatment.

[0326] 4.20 - Serum formula for hair application

[0327] [Table 28]

[0328]

[0329] preparation:

[0330] Sprinkle carbomer in water and allow it to swell for 30 minutes. Weigh part B. While stirring, add part B to part A. Weigh and mix part C. Neutralize part C to part A + B. Homogenize thoroughly while stirring. Weigh and mix part D. While stirring, pour part D into part A + B + C. Add part E and mix thoroughly. Add part F and mix thoroughly.

[0331] A colorless fluid gel is obtained, forming a serum with a light, non-greasy texture.

Claims

1. Pycnidione is used for non-therapeutic cosmetic treatments of the skin and its appendages to improve their overall condition and / or sensory comfort.

2. Use according to claim 1, characterized in that, The pycnidione is provided as an extract of biomass from microbial in vitro cultures capable of producing pycnidine as a secondary metabolite, the extract being optionally purified to increase the pycnidione content.

3. The use according to claim 2, characterized in that, The extract is derived from the biomass of in vitro cultures of a strain of Pythium irpicalis Coniothyrium cereals ).

4. Use according to any of the preceding claims, characterized in that The pycnidione is added to a physiologically acceptable medium.

5. Use according to claim 4, characterized in that, The physiologically acceptable medium corresponds to the biomass extraction solvent.

6. Use according to any one of the preceding claims, characterized in that, The processing is local.

7. Use according to any of the preceding claims, characterized in that, The processing is selected from: - Treatment for signs of chronological aging or premature aging - Moisturizing treatment, - Treatment for dull and / or tired skin - Treatment for sensory discomfort of the skin - Treatments for oily and / or acne-prone skin, and / or - Hair treatment.

8. Use according to any of the preceding claims, characterized in that The treatment is suitable for maintaining or improving the mechanical and viscoelastic properties of the skin, including the scalp.

9. Use according to any one of the preceding claims, characterized in that, The treatment is suitable for preventing or reducing roughness, fine lines, and wrinkles on the skin.

10. The use according to any one of the preceding claims, characterized in that, The treatment is suitable for preventing or reducing skin sagging.

11. The use according to any one of the preceding claims, characterized in that, The treatment is intended to stimulate the production of hyaluronic acid.

12. The use according to any one of the preceding claims, characterized in that, The treatment is applicable to enhancing the dermal-epidermal junction (DEJ).

13. The use according to any one of the preceding claims, characterized in that, The treatment is applicable to combating the harmful effects of glycation.

14. The use according to any one of the preceding claims, characterized in that, The treatment is applicable to combating the harmful effects of free radicals.

15. The use according to any one of the preceding claims, characterized in that, The hair treatment is applicable to treating brittle or split ends, and / or preventing and / or treating pigmentation defects.

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