US20200054549A1 - Hydrolysate of pichia minuta and cosmetic use thereof for controlling hair loss and stimulating regrowth - Google Patents

Hydrolysate of pichia minuta and cosmetic use thereof for controlling hair loss and stimulating regrowth Download PDF

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US20200054549A1
US20200054549A1 US16/476,428 US201816476428A US2020054549A1 US 20200054549 A1 US20200054549 A1 US 20200054549A1 US 201816476428 A US201816476428 A US 201816476428A US 2020054549 A1 US2020054549 A1 US 2020054549A1
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active ingredient
hydrolysate
hair
cosmetic
pichia minuta
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Jean Paufique
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Limousine D'application Biologique Silab Ste
Societe Industrielle Limousine dApplication Biologique SA SILAB
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
    • A61K8/99—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from microorganisms other than algae or fungi, e.g. protozoa or bacteria
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00—Cosmetics or similar toiletry preparations
    • A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
    • A61K8/64—Proteins; Peptides; Derivatives or degradation products thereof
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q7/00—Preparations for affecting hair growth

Definitions

  • This invention relates to a particular cosmetic active ingredient obtained by yeast hydrolysis and its use for fighting hair loss and promoting regrowth.
  • hair is a symbol of beauty and strength whose condition has a significant impact on one's self-esteem and image.
  • the acceleration of hair loss scientifically known as alopecia, is perceived as a sign of early aging and can dramatically affect the quality of life of those affected.
  • Alopecia is a common disorder with its androgenetic form being the most common. It affects up to 30% of men under 30 and more than 50% of men over age 50.
  • This progressive disorder which is not a disease (Prie B. et al., “Oxidative stress in androgenetic alopecia”, Journal of Medicine and Life Vol. 9, Issue 1, January-March 2016, pp. 79-83), is characterized by the appearance of finer, shorter and non-pigmented hair which turns to vellus hair. It is caused by two main factors, genetic and hormonal, which cause hypersensitivity to a hormone: dihydrotestosterone (DHT). The latter is responsible for all biological changes associated with this disorder.
  • DHT dihydrotestosterone
  • DHT dihydrotestosterone
  • DHT dihydrotestosterone
  • AGA androgenetic alopecia
  • Retention of thick hair is the result of equilibrium between hair loss and regrowth.
  • Losing hair is a natural phenomenon, with an individual losing between 50 and 125 every day. Thanks to a process of regeneration, a constant renewal thereof is ensured: the so-called life cycle of the hair or hair growth cycle. It is divided into three phases:
  • the hair or hair follicle consists of two main elements.
  • the dermal papilla is the driving force of its growth and lies at the base of the follicle. It is composed of fibroblasts able to emit growth signals. These are then captured by the cells of the hair matrix. This structure that surrounds the papilla is home to keratinocytes. In response to these signals, they will proliferate to trigger the formation of a new hair stalk.
  • Hair growth is an energy-consuming process that requires functional mitochondria in the cells, the morphological state of these dynamic organelles conditioning energy production.
  • the invention proposes a natural cosmetic active ingredient, derived from yeast, able to act on three mechanisms for regulation of the regrowth of hair.
  • hydrolysates of yeast have already been used in cosmetics, such as hydrolysates of Saccharomyces cerevisiae, Pichia anomala or Torulaspora delbrueckii.
  • the invention relates to a hydrolysate obtained from a different yeast, Pichia minuta, which is very different from already known yeasts used in cosmetics and which exhibits its own characteristics.
  • the invention relates to a cosmetic active ingredient including a hydrolysate of Pichia minuta, in particular a hydrolysate of Pichia minuta at least comprising peptides.
  • such an active ingredient is able to both stimulate mitochondrial dynamics for hair growth and correct expression defects affecting the “signal” and epigenetic molecules associated with androgenetic alopecia.
  • the cosmetic active ingredient of the invention thus makes it possible to reactivate the dermal papilla and stimulate growth of hair follicles.
  • the invention also relates to the non-therapeutic cosmetic use of such an active ingredient when applied to hair and/or scalp, for fighting hair loss and enable regrowth.
  • the invention also relates to cosmetic compositions, containing at least 0.05% by weight, of the active ingredient of the invention, as well as a non-therapeutic cosmetic method for fighting hair loss and activating its regrowth by application of such compositions to the hair or scalp.
  • FIG. 1 represents the chromatographic profile (HPLC/RI) of the hydrolysate peptides of example 1,
  • FIG. 2 represents the aminogram of the peptides of the active ingredient of the invention of example 1,
  • FIG. 3 represents the chromatogram of the proteins of the three products presented in the Comparative Test 1, obtained using the same process starting for 3 different yeasts: Pichia minuta, Pichia heedii and Saccharomyces cerevisiae,
  • FIG. 4A represents an agar culture medium on which a biomass of Pichia minuta was inoculated and incubated for 48 h at 30° C.
  • FIG. 4B represents an agar culture medium on which a hydrolysate of Pichia minuta was inoculated and incubated for 48 h at 30° C.
  • FIG. 5A represents an enlarged photo of the scalp before application of a composition of the invention
  • FIG. 5B shows a zoomed in picture of a scalp after 8 months of application of a composition of the invention (point II.1)
  • FIG. 6A represents a photo of a scalp before application of a composition of the invention.
  • FIG. 6B shows a picture of a scalp after 8 months of application of a composition of the invention (point II.1)
  • the term “cosmetic active agent” or “cosmetic active ingredient” means at least one molecule, preferentially a set of molecules having an effect on the cells of the hair and/or the scalp, in particular on the fibroblasts of the dermal papilla and the cells of the hair matrix.
  • protein fraction or “peptide compounds” of the hydrolysate of Pichia minuta means all proteins and peptides present in the hydrolysate of Pichia minuta.
  • hydrolysate of Pichia minuta means any active ingredient derived from Pichia minuta yeast, obtained by a process comprising at least one step of enzymatic or chemical hydrolysis of Pichia minuta.
  • hydrolysate of Pichia minuta excludes molecules produced solely by fermentation of Pichia minuta.
  • Pichia minuta within the meaning of the invention, means any yeast of the Saccharomycetaceae family, of the Ogataea genus and Pichia minuta species.
  • the Pichia minuta yeast has been registered in several yeast collections, under numbers ATCC 26176, CBS 6511, MUCL 27758, MUCL 29976, DSM-70275, NRRL Y-411, NRRL Y-7953, NRRL Y-10948, Y-172, Y-2081, Y-2516, NCYC-499, 3622T, 9442, 3615T, NBRC 1473, NBRC 0975T, NBRC 10402, NBRC 10746. It is also known by other names: Candida methanolovescens, Ogataea minuta, Ogataea nonfermentans. Pichia minuta can be isolated from the azalea flower, Rhododendron indicum.
  • atomization carrier is understood to mean a neutral adjuvant added to a solution in order to produce a powder during spray drying.
  • the invention therefore relates to a cosmetic active ingredient including a hydrolysate of Pichia minuta.
  • the hydrolysate of Pichia minuta at least contains peptides.
  • the hydrolysate of Pichia minuta contains peptides which have a molecular weight of less than 3500 Da, preferably between 243 and 3500 Da. These peptides play an important role in the effectiveness of the active ingredient of the invention.
  • the peptides which have a molecular weight of less than 3500 Da represent at least 50% by weight of the protein fraction of the hydrolysate, still more preferably at least 80%.
  • the protein fraction represents at least 40% by weight of the hydrolysate dry matter, preferably between 40 and 90%, more preferably between 50 and 70%.
  • the peptide compound content is preferentially determined by the LOWRY method (Lowry et al., Protein measurement with the folin reagent, J. Biol. Chem., 193, 265, 1951).
  • amino acids which make up more than 5%, constituting the peptides of the hydrolysate are preferably: alanine, arginine, aspartic acid, glutamic acid, leucine, lysine, serine, threonine, valine.
  • the aminogram of the peptides which make up the hydrolysate of the invention can be obtained by ion exchange chromatography after acid hydrolysis of the sample.
  • the hydrolysate also preferably comprises sugars and/or minerals.
  • the sugars represent at least 5% by weight of hydrolysate dry matter, in particular between 5 and 25%, more preferably between 10 and 20%.
  • the sugar content of the hydrolysate can be determined by the DUBOIS method (Dubois M et al., Analytical Chemistry, 28, 3, 350-356, 1956).
  • the minerals present in the hydrolysate are preferably calcium, potassium, sodium, chlorine and phosphorus.
  • the analysis of the minerals constituting the ashes of the hydrolysate was carried out by optical emission spectrometry (ICP/OES) and the measurement of chloride ions by titration with silver nitrate.
  • the ash content is between 5 and 35% by weight of hydrolysate dry matter, more preferably between 20 and 30%.
  • the crude ash content can be determined by weighing the residues resulting from the incineration of the samples of the active ingredient of the invention at 550° C., in an electric muffle furnace.
  • the hydrolysate of the invention can be obtained by chemical (acid or base) or enzymatic hydrolysis.
  • an enzymatic hydrolysate is used.
  • the active ingredient of the invention can be in solid or liquid form.
  • the active ingredient of the invention is preferably the hydrolysate as described above. It is in the form of a clear liquid, with a weak odor and a slightly orangish yellow color.
  • the active ingredient of the invention is preferably made up of the hydrolysate of Pichia minuta as previously described and of a carrier chosen from maltodextrin, gum arabic, soy lecithin or isomalt.
  • the hydrolysate represents between 25 and 75% of the weight of the active ingredient, preferably between 40 and 60%.
  • the carrier generally comprising mainly of sugars.
  • the atomization carrier is maltodextrin, in an amount between 25 and 75%, preferably between 40 and 60%:
  • the peptides represent at least 12%, in particular between 12 and 53% by weight of dry matter of the active ingredient, preferably between 20 and 42%,
  • the sugars represent between 32 and 80% by weight of dry matter of the active ingredient, preferably between 46 and 68%,
  • the ashes represent between 5 and 23% by weight of dry matter of the active ingredient, preferably between 8 and 18%.
  • the hydrolysate constituting the active ingredient of the invention can be obtained by any process comprising at least one step of hydrolysis of Pichia minuta, in particular a step for hydrolysis of the proteins.
  • it is obtained by a process involving a step of enzymatic hydrolysis, in particular of enzymatic hydrolysis of the proteins, that is to say a hydrolysis carried out enzymatically by means of proteolytic enzymes.
  • proteolytic enzymes may be, for example, proteases of plant origin or derived from microorganisms.
  • the biomass of Pichia minuta Prior to the process for obtaining the hydrolysate as such, the biomass of Pichia minuta should be produced. This step is carried out according to the culture method of the yeasts in a medium appropriate for their development, in a manner conventional to those skilled in the art.
  • the active ingredient is obtained by the implementation of the following steps:
  • the hydrolysate obtained at this stage may be further concentrated and/or purified in order to select the fractions of low molecular weight, preferably less than 3500 kDa, or even possibly less than 2000 kDa, by successive ultrafiltration steps through varying porosity filters, retaining the filtrates at each stage and/or by means of a chromatographic type method, for example to specifically enrich the hydrolysate in these molecules.
  • the hydrolysate can then be dried and combined with a carrier to be in solid form.
  • This phase can be achieved by implementing the following steps:
  • This presentation of the peptide hydrolysate corresponds to a highly concentrated form of active molecules, in particular, peptides.
  • the active ingredient of the invention is particularly effective against hair loss and to activate its regrowth.
  • the growth of hair follicles is a process requiring a high energy input.
  • mitochondria that ensure energy production.
  • these organelles are able to adapt their morphology and, ultimately, their activity: mitochondrial dynamics This is based on two phenomena: fission and fusion.
  • Fission is defined by the separation of mitochondria which are then smaller and produce less energy.
  • fusion corresponds to the grouping of mitochondria in the form of a network. This relies notably on the intervention of mitofusin 1 (MFN1), a membrane protein involved in the fusion of the outer mitochondrial membranes. This process allows them to intensify their metabolism and produce more energy in the form of ATP.
  • MFN1 mitofusin 1
  • Mitochondrial dynamics is involved in the biology of hair follicles.
  • the morphology of mitochondria evolves towards a fused form in dermal papilla cells. This phenomenon is accompanied by an increase in mitochondrial enzymatic activity and the production of ATP. These mitochondrial changes provide the energy required to pass or maintain the growth phase of the hair cycle (Mifude et al. “PDGF-AA-induced filamentous mitochondria benefit dermal papilla cells in cellular migration”. International Journal of Cosmetic Science, 37, 266-271 (2015)).
  • the active ingredient of the invention is able to stimulate the synthesis of mitofusin 1 in the fibroblasts of the dermal papilla and thus to promote the growth of hair follicles.
  • the “signal” molecules present in the microenvironment of the dermal papilla are essential for hair growth. Three of them are deregulated in androgenetic alopecia:
  • DKK1 Dihydrotestosterone-Inducible Dickkopf 1 from Balding's Dermal Papilla Cells Causes Apoptosis in Follicular Keratinocytes” Journal of Investigative Dermatology, 128, 262-269 (2008)) and the expression of DKK1 (Fawzi M M T & al. “Assessment of tissue levels of dickkopf-1 in androgenetic alopecia and alopecia areata.” Journal of Cosmetic Dermatology, 15, 10-15 (2015)). These deregulations are also observed following treatment with DHT and may even be accentuated following the activation of the androgen signaling pathway (Yang et al. “Androgen Receptor Accelerates Premature Senescence of Human Dermal Papilla Cells in Association with DNA Damage”. Plos One, 8, 1-10 (2013)).
  • the active ingredient of the invention is able to reduce the secretion of IL-6 and the expression of DKK1 and P16 in the fibroblasts of the dermal papilla. It thus makes it possible to correct the abnormal expression of the “signal” molecules involved in androgenetic alopecia.
  • epigenetics also plays a major role in the biology of hair.
  • Epigenetics studies the mechanisms that can regulate the expression of genes in response to certain environmental stresses, without modifying the DNA pattern. All actors involved are grouped under the name of epigenome. Among them, miRNAs hold an important place. These small RNA molecules are able to inactivate genes on the basis of the environment by modifying the stability of their target messenger RNAs. It has recently been demonstrated that miRNAs regulate various processes such as proliferation and differentiation of dermal papilla fibroblasts and keratinocytes of the hair matrix. In addition, they regulate apoptosis during the catagen phase (Andl et al. “MicroRNAs (miRNAs) in the control of HF development and cycling: the next frontiers in hair research” Experimental dermatology, 24, 821-826 (2015)).
  • the fibroblasts of the dermal papilla have a deregulated miRNA expression profile.
  • These modifications include overexpression of miR-3663-3p and of let-7a-3p (Lee et al. “Analysis of the microRNA expression of normal human dermal papilla cells treated with 5 ⁇ -dihydrotestosterone” Molecular Medicine Reports, 12, 1205-1212 (2015)).
  • Some are involved in the Wnt/ ⁇ -catenin signaling pathway, an essential pathway for hair growth. It is activated following the binding of a ligand of the Wnt family to its specific receptor and causes the accumulation of ⁇ -catenin.
  • the active ingredient of the invention is able to limit the expression of miRNA 3663-3p and of let-7a-3p in fibroblasts of the dermal papilla.
  • it attenuates epigenetic deregulations associated with androgenetic alopecia.
  • the dermal papilla is the control center for hair growth.
  • the fibroblasts of the papilla emit signals to the keratinocytes of the hair matrix in order to regulate the development of the hair. In a normal context, it takes place in a cycle having three phases:
  • the active ingredient of the invention is able to stimulate the synthesis of versican in fibroblasts of the dermal papilla and thus preserve the activation of the dermal papilla. In addition, it improves the growth of hair follicles and helps to maintain a significant synthesis of versican and Ki-67. Thus, the active ingredient of the invention makes it possible to reduce the lack of growth associated with androgenetic alopecia by keeping hair follicles in the growth phase.
  • the invention thus relates to the use of an active ingredient of the invention by means of application to the hair and/or scalp, for fighting hair loss and activate its regrowth, especially in people with androgenetic alopecia. It acts on molecular abnormalities associated with androgenetic alopecia by correcting deregulation affecting the signal molecules and epigenetics and by stimulating mitochondrial dynamics It is thus effective in normalizing two essential parameters for hair development, namely: activation of the dermal papilla and stimulation of the growth of hair follicles.
  • compositions comprising a cosmetically acceptable medium.
  • compositions in different galenic forms, suitable for application on the scalp and hair.
  • compositions can, in particular, be in the form of oil-in-water emulsions, water-in-oil emulsions, multiple emulsions (Water/Oil/Water or Oil/Water/Oil), which can possibly be micro emulsions or nanoemulsions, or in the form of solutions, suspensions, hydrodispersions, aqueous gels or powders. They can be more or less fluid and have the appearance of lotion, shampoo, cream or mousse.
  • compositions comprising at least 0.05% of a hydrolysate of Pichia minuta of the invention, preferably between 0.5 and 10%.
  • compositions comprise, on top of the active ingredient, a physiologically acceptable and preferably cosmetically acceptable medium, i.e. which does not cause unacceptable discomfort for the user, such as redness, pulling or tingling.
  • compositions of the invention may contain as adjuvant at least one compound chosen from:
  • compositions are especially intended to be used for anti-hair loss and hair regrowth processes.
  • the invention thus also relates to a non-therapeutic method, namely a non-therapeutic cosmetic method for fighting hair loss and activating its regrowth, which includes applying a composition which includes an active ingredient of the invention to the hair and/or scalp, preferably at least once a day for at least one month.
  • a composition which includes an active ingredient of the invention to the hair and/or scalp, preferably at least once a day for at least one month.
  • the composition is a composition of the invention.
  • the active ingredient according to Example 1 is obtained by implementing the following steps:
  • the hydrolysate obtained is in the form of a clear liquid, of a slightly orangish yellow color, with a weak odor.
  • the peptides were characterized in order to determine their molecular weights and to quantify the various protein fractions by size exclusion FPLC chromatography.
  • the hydrolysate is analyzed under the following conditions:
  • the different fractions collected from the hydrolysate are quantified by spectrophotometric assay according to the LOWRY method (Lowry et al., Protein measurement with the folie reagent, J. Biol. Chem., 193, 265, 1951).
  • More than 93% of peptides in the main active ingredient of Example 1 of the invention have molar masses of between 243 and 3500 Da.
  • the amount of protein greater than 3500 Da represents less than 5% of the total protein.
  • the ash distribution was determined after incineration at 550° C.
  • the analysis of the minerals was carried out by optical emission spectrometry (ICP/OES) and the determination of chloride ions by titration with silver nitrate.
  • Example 2 The active ingredient of Example 1 is combined with maltodextrin to constitute the active principle of Example 2.
  • the active ingredient is in the form of a powder having a particle size of less than 500 ⁇ m, a weak odor, a slightly orangish yellow color.
  • maltodextrin includes only non-active sugars
  • the peptide composition, the aminogram and the mineral composition of the active ingredient are identical to those of Example 1.
  • composition of the invention in serum form includes:
  • the pH of the composition is 5. It is in the form of an opalescent liquid, without odor.
  • the serum of Example 3 has the following characteristics: sprayable lotion, fresh and silky application, slippery effect, immediate penetration, soft and dry finish, smoothing effect on the hair.
  • composition of the invention in the form of a mask for the hair is constituted by:
  • the pH of the composition is 4.5.
  • the mask of Example 3 has the following characteristics: firm grip, easy application, film-forming effect, easy rinsing, soft and dry finish.
  • composition of the invention in the form of treatment for the hair is constituted by:
  • the pH of the composition is 6.1.
  • Example 5 The treatment of Example 5 has the following characteristics: Gentle grip, soft and even application on the skin and hair, cool and moisturizing effect, easy rinsing, smooth and soft finish.
  • composition of the invention in the form of treatment for the hair is constituted by:
  • the pH of the composition is 5.
  • the washing base of Example 6 has the following characteristics: fluid flow, soft touch, extremely strong foaming power, good water dispersion with creation of a soft, white and compact foam, easy rinse, very slightly film-forming soft finish, moisturizing effect.
  • composition of the invention in the form of a lotion for hair includes:
  • the objective of this study is to evaluate the effect of an active ingredient of the invention on the mitochondrial dynamics, an element necessary for the growth of hair follicles.
  • Mitofusin 1 is a major player in mitochondrial fusion in human fibroblasts of the dermal papilla. When hair growth is encouraged, mitochondrial dynamics are modified. This results in an increase in the proportion of large fused mitochondria compared to small fissioned mitochondria in dermal papilla fibroblasts. This state is favorable for energy production.
  • the human fibroblasts of the dermal papilla are inoculated into culture medium and incubated at 37° C.
  • the cells are treated: the fibroblasts are treated with medium either containing or not the active ingredient at 0.02% and 0.04% (final V/V).
  • the objective of this study is to evaluate the effect of an active ingredient of the invention on the regulation of “signal” molecules described in androgenetic alopecia.
  • the human fibroblasts of the dermal papilla are inoculated into culture medium and incubated at 37° C.
  • the fibroblasts are treated with a solution of DHT containing or not the active ingredient at 0.02% and 0.04% (final V/V).
  • the IL-6 assay is performed using an ELISA assay kit.
  • the RNAs were reverse-transcribed and the resulting complementary DNAs were analyzed by the quantitative PCR technique.
  • DKK1 and P16 mRNAs were also studied.
  • mRNAs of Ribosomal Protein S27 (RPS27), Hypoxanthine Phosphoribosyltransferase 1 (HPRT1), beta-Glucuronidase (GUSB) were analyzed as internal reference controls for standardization.
  • DHT significantly increases IL-6 secretion and expression of DKK1 and P16 by human fibroblasts of the dermal papilla, thus mimicking the expression modification of “signal” molecules described in androgenetic alopecia.
  • the active ingredient of the invention makes it possible to significantly limit the secretion of IL-6 by 97% and the expressions of DKK1 and P16 by 91% and 71%, respectively.
  • the objective of this study is to evaluate the effect of an active ingredient of the invention on the epigenetic modifications associated with androgenetic alopecia.
  • DHT dihydrotestosterone
  • the human fibroblasts of the dermal papilla are inoculated in culture medium and incubated at 37° C. in an atmosphere containing 5% CO 2 .
  • the fibroblasts are treated with a solution of DHT which does or does not contain the active ingredient of the invention at 0.02% and 0.04% (final V/V).
  • RNAs were reverse-transcribed and the resulting complementary DNAs were analyzed by quantitative PCR technique.
  • microRNAs 3663-3p and let-7a-3p has been studied.
  • small nucleolar RNA C/D box 68 (SNORD68) was analyzed as an internal reference control for standardization.
  • Quantification of fluorescence incorporation is measured continuously using a thermocycler.
  • the analysis of the Ct (relative quantification) is performed using software.
  • DHT induces expression of 3663-3p and let-7a-3p miRNAs in human dermal papilla fibroblasts, mimicking the epigenetic changes associated with androgenetic alopecia.
  • an active ingredient of the invention makes it possible to limit the epigenetic modifications associated with androgenetic alopecia.
  • the objective of this study is to evaluate the effect of an active ingredient of the invention on the activation of the dermal papilla.
  • the versican synthesis was measured. It is a proteoglycan that is specific to the growth phase.
  • the evaluation was performed in a 3D model of fibroblasts in the form of spheroids, treated with a solution of DHT. This model mimics the inactivation of the dermal papilla, a factor contributing to the lack of growth associated with androgenetic alopecia.
  • the human fibroblasts of the dermal papilla are inoculated into culture medium and incubated at 37° C.
  • the spheroids formed are treated with a solution of DHT which does or does not contain the active ingredient of the invention at 0.04% (final V/V).
  • Immunohistological labeling of versican is performed. Visualization is performed on a confocal microscope coupled to an image analysis system.
  • Versican synthesis is proportional to the intensity of fluorescence (green color) present on the spheroids.
  • the nuclei of the cells appear to be blue in color.
  • DHT induces a decrease in the synthesis of versican, in a model of human fibroblasts of dermal papilla in the form of spheroids, mimicking an inactivation of the dermal papilla induced during androgenetic alopecia.
  • the active ingredient of the invention stimulates the synthesis of versican in this model.
  • it helps maintain the activity of the dermal papilla and its ability to induce the growth of hair follicles.
  • the objective of this study is to evaluate the ability of an active ingredient of the invention to maintain hair follicles in a growth phase (anagen phase).
  • the evaluation of the elongation is carried out by measuring the size of hair follicles after photographs.
  • the syntheses of versican and Ki-67 are studied by immunohistological labeling. The operating protocol is described hereafter.
  • the hair follicles are obtained from scalp fragments and incubated in culture medium at 37° C.
  • the follicles are photographed and measured on a microscope coupled to an image analysis system.
  • the follicles selected are then treated with a solution of DHT which does or does not contain the active ingredient of the invention at 0.02% and 0.04% (final V/V).
  • the follicles are photographed and measured on a microscope coupled to an image analysis system.
  • the elongation of each follicle is equal to its growth between day 0 and day 5.
  • the follicles are treated with a solution of DHT containing or not the active ingredient at 0.02% and 0.04% (final V/V).
  • the follicles are recovered and included in Tissue-Tek® and then frozen. Sections (4 ⁇ m) are then made using a cryostat Immunohistological labeling of versican and Ki-67 is performed. Visualization is performed on a confocal microscope coupled to an image analysis system.
  • Versican synthesis is proportional to the intensity of fluorescence (green color) present on sections of hair follicles.
  • Ki67 The synthesis of Ki67 is proportional to the intensity of the fluorescence (yellow color) present on sections of hair follicles.
  • each follicle On day 7, the follicles are again photographed and measured. The elongation of each follicle is equal to its growth between day 0 and day 7.
  • DHT significantly reduces the growth of hair follicles at day 5 and day 7 by ⁇ 31% and ⁇ 37%, respectively. As a result, DHT mimics the lack of hair growth associated with androgenetic alopecia.
  • an active ingredient of the invention makes it possible to significantly increase the growth of hair follicles on day 5 and day 7 by respectively +31% and +37%.
  • DHT significantly reduces the synthesis of versican and Ki-67. As a result, DHT mimics the metabolic alterations associated with androgenetic alopecia.
  • an active ingredient of the invention makes it possible to maintain a significant synthesis of versicans, and the proliferative power of keratinocytes.
  • the use of the invention makes it possible to limit the reduction in growth associated with androgenetic alopecia and keeps hair follicles in an anagen phase.
  • the volunteers participating in the in vivo tests were selected according to several criteria. These are men under 55 years of age, with mild to moderate alopecia and a minimum of 20% hair in the telogen phase.
  • the objective of this study was to evaluate, in vivo, the anti-hair loss effect of an active ingredient of the invention (Example 2) formulated at 0.3% in a lotion (Example 7), in volunteers with mild to moderate alopecia.
  • the anti-hair loss effect was studied by the phototrichogram method from measurements taken at time 0, 3, 6 and 8 months after the start of treatment. The following parameters were analyzed:
  • an active ingredient of the invention formulated at 0.3% increases the capillary density already after 3 months of application (+21.3%). This effect is prolonged and intensified after 6 and 8 months of treatment (respectively +22.1 and +24.9%).
  • the active ingredient of the invention shows an action on capillary growth, already after 3 months of application, which intensifies during treatment.
  • the active ingredient of the invention formulated at 0.3% in a lotion significantly reduces the proportion of hair in the telogen phase after 3 months of application ( ⁇ 14.0%). This effect intensifies after 6 and 8 months of treatment (respectively ⁇ 21.9% and ⁇ 26.1%).
  • FIGS. 5A / 6 A at day 0 before application
  • 5 B/ 6 B after 8 months of application
  • the total hair gain on the scalp between day 0 and 8 months for this illustration is 74,160 hairs.
  • an active ingredient of the invention makes it possible to strengthen and redensify the hair.
  • the objective of this study was to evaluate, in vivo, the ability of an active ingredient of the invention (Example 2) formulated at 0.3% in a lotion (Example 7), to curb the hair loss of volunteers with mild to moderate alopecia.
  • the magnitude of hair loss was studied using the wash test method performed before starting and 3, 6 and 8 months after the start of treatment.
  • composition of the invention slows hair loss by reducing the number of hairs recovered after shampooing by 21.2%.
  • the invention stabilizes hair loss by limiting the number of hairs lost by 34.0%.
  • hydrolysates do not have the same analytical characteristics, especially as regards the protein content, which are very different for the three products obtained from the same process, but from different yeasts.
  • the size of the proteins is also different, as can be seen in the chromatogram of the proteins of the three products, as shown in FIG. 3 .
  • the aim of this test is to compare the effectiveness of a cosmetic active ingredient, a hydrolysate of Pichia anomala described in patent FR3016521, with that of the active ingredient of the invention in order to show that a cosmetic active ingredient derived from a yeast of the genus Pichia, does not necessarily work.
  • Protein size is also similar, since both hydrolysates contain 59% protein, more than 87% of which have a molecular weight of less than 5 kDa.
  • the aim of this test is to compare the effectiveness of a hydrolysate of Torulaspora delbrueckii, described in patent FR2979541, with that of the active ingredient of the invention of Example 1, in order to show that a cosmetic active ingredient derived from a yeast having a similar analytical characteristic, does not necessarily work.
  • Protein size is also similar, since both hydrolysates contain more than 93% having a molecular weight of less than 5 kDa.
  • the objective of this test is to demonstrate that a hydrolysate of Pichia minuta does not have the same analytical characteristics and is therefore different from a Pichia minuta biomass, as it exists in nature.
  • the hydrolysate does not contain live yeasts whereas the biomass contains them.
  • FIG. 4B No growth is not observed ( FIG. 4B ) on the agar with the hydrolysate of example 1, whereas on the other hand ( FIG. 4A ) a growth of white, round, shiny, oily colonies with a diameter of less than 0.5 mm is observed on agar with Pichia minuta biomass.
  • Biomass is the natural raw material on which the enzymatic hydrolysis process is applied to obtain the active ingredient of the invention, hydrolysate of Pichia minuta.

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US16/476,428 2017-01-09 2018-01-09 Hydrolysate of pichia minuta and cosmetic use thereof for controlling hair loss and stimulating regrowth Abandoned US20200054549A1 (en)

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FR1770024A FR3061656B1 (fr) 2017-01-09 2017-01-09 Hydrolysat de pichia minuta et utilisation cosmetique pour lutter contre la chute des cheveux et favoriser leur repousse
PCT/EP2018/050462 WO2018127604A1 (fr) 2017-01-09 2018-01-09 Hydrolysat de pichia minuta et utilisation cosmetique pour lutter contre la chute des cheveux et favoriser leur repousse

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CN115252485A (zh) * 2022-08-08 2022-11-01 澳宝化妆品（惠州）有限公司 毕赤酵母提取物的制备方法及含有其的头皮护理组合物
US11604559B2 (en) 2007-09-04 2023-03-14 Apple Inc. Editing interface
US11650713B2 (en) 2005-12-30 2023-05-16 Apple Inc. Portable electronic device with interface reconfiguration mode
US11675476B2 (en) 2019-05-05 2023-06-13 Apple Inc. User interfaces for widgets
US11733656B2 (en) 2016-06-11 2023-08-22 Apple Inc. Configuring context-specific user interfaces
US11736602B2 (en) 2006-09-06 2023-08-22 Apple Inc. Portable multifunction device, method, and graphical user interface for configuring and displaying widgets
US11809700B2 (en) 2010-04-07 2023-11-07 Apple Inc. Device, method, and graphical user interface for managing folders with multiple pages
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FR3138309A1 (fr) 2022-07-27 2024-02-02 L'oreal Composition cosmétique comprenant du 2,4-diaminopyrimidine-3-N-oxyde, de la piroctone olamine, de la caféine, et au moins 0,5% en poids de composé hydroxylé particulier
FR3138314A1 (fr) 2022-07-27 2024-02-02 L'oreal Composition cosmétique comprenant du 2,4-diaminopyrimidine-3-N-oxyde, de la piroctone olamine, de la caféine, et un extrait d’écorce d’Eperua falcata

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US11650713B2 (en) 2005-12-30 2023-05-16 Apple Inc. Portable electronic device with interface reconfiguration mode
US11736602B2 (en) 2006-09-06 2023-08-22 Apple Inc. Portable multifunction device, method, and graphical user interface for configuring and displaying widgets
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EP3565640B1 (fr) 2020-12-02
EP3565640A1 (fr) 2019-11-13
ES2859608T3 (es) 2021-10-04
US20230240976A1 (en) 2023-08-03

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