Cosmetic composition for improving hair density

Abstract

The present invention relates to a cosmetic composition for improving hair density comprising active ingredients of a lipid extract of Rhodobryum gigantum, a hydrophilic extract of Saxifraga oppositifolia, a lipid extract of Triticum vulgare and a hydrophilic extract of Quillaja saponaria, in combination with at least five biomimetic peptides. The present invention also relates to a method for the cosmetic care of the hair and scalp, as well as a dermatological composition for use as a medicament for treating alopecia.

Description

Technical Field

[0001] The technical field of this invention relates to cosmetic and dermatological compositions for improving hair density, the compositions comprising effective amounts of at least four plant extracts—derived from Coleus forskolhii, Saxifraga rotundifolia, Triticum vulgare, and Quillajasaponaria—and active ingredients combined with at least five biomimetic peptides. Background Technology

[0002] There are approximately 100,000 hair shafts on the human scalp. Each hair grows at a different pace, with an average lifespan of about 3.5 years and a growth rate of 1 mm per month. Hair has three regions: the medulla (the innermost layer, formed by clear cells, which varies depending on hair type), the cortex (the middle layer, which provides strength to the hair shaft and is composed of keratin), and the keratin layer (the outermost layer). The hair follicle can be divided into the lower, middle (isthmus), and upper (infundibulum) segments. The lower segment includes the bulb and the area above the bulb. The middle segment includes the area extending from the attachment point of the arrector pili muscle to the opening of the sebaceous gland duct. The upper segment includes the area between the opening of the sebaceous gland duct and the follicle opening [Park, AM; Khan, S.; Rawnsley, J. Hair biology: Growth and pigmentation. Facial Plast. Surg. Clin. N. Am. 2018, 26, 415–424.].

[0003] Hair color, density, curvature, and diameter are all affected by hair aging, and these factors collectively determine the appearance and plasticity of hair [Trüeb, RM Aging of hair. J. Cosmet. Dermatol. 2005, 4, 60-72.]. Many factors influence hair health and the hair growth cycle. Pollution (such as heavy metals—thallium, mercury, arsenic), as well as PM2.5 particles from petroleum compound combustion, toxins (botulinum toxin, podostroma cornu-damae), medications, genetics, stress, smoking, menopause, lifestyle, and diet are some of the major factors related to hair health. The combination of ultraviolet radiation and pollutants (such as particulate matter from petroleum combustion) may exacerbate the biochemical and clinical effects of hair and skin tissue aging and may trigger alopecia areata and other types of hair loss.

[0004] Alopecia areata (AA) has a history dating back to around 1500 BC, but scientific research on hair loss has seen rapid development in publications in recent decades [Broadley, D.; McElwee, KJ A “hair-raising” history of alopecia areata. Exp. Dermatol. 2020, 29, 208–222.]. Although hair is not biologically important to humans, hair loss has significant social, psychological, and emotional impacts on anyone affected. Therefore, treating hair loss and / or improving hair and scalp health is a social problem that needs to be addressed.

[0005] Existing technologies describe numerous cosmetic and dermatological compositions for stimulating hair growth. The use of many plant extracts, even peptides, or combinations of both, has been proposed to prevent hair loss and / or stimulate hair regrowth.

[0006] Surprisingly, the inventors discovered that a novel complex composed of four plant extracts [Coleus forskohlii, Saxifraga rotundifolia, Triticum vulgare, and Quillaja saponaria] and five synthetic peptides [EGF, IGF-1, acidic FGF, basic FGF, and VEGF] exhibits strong stimulating activity for hair regeneration and can improve scalp quality.

[0007] This specific combination of ingredients has never been described before in the field of hair care. Summary of the Invention

[0008] This invention relates to a cosmetic composition for improving hair density, comprising an effective amount of at least four plant extracts combined with at least five synthetic biomimetic peptides as active ingredients. The four plant extracts are selected from lipid extracts of the roots of *Coleus forsythia suspensa*, hydrophilic extracts of *Saxifraga stolonifera*, lipid extracts of common wheat, and hydrophilic extracts of *Gnaphalium affine* wood. The five synthetic biomimetic peptides are selected from EGF, IGF-1, acidic FGF, basic FGF, and VEGF, as well as physiologically acceptable excipients.

[0009] The present invention also relates to a method for cosmetic treatment of hair and scalp, the method comprising: applying about 0.5 to 1.5 g of an effective amount of such composition to hair and scalp twice daily for about 1 to 9 minutes, particularly 3 to 7 minutes, and optionally rinsing with water or shampoo.

[0010] The present invention also relates to a dermatological composition for use as a medicament for the prevention and treatment of hair loss, comprising an effective amount of at least four plant extracts combined with at least five synthetic biomimetic peptides as active ingredients. The four plant extracts are selected from a lipid extract of the root of Coleus forsythia, a hydrophilic extract of Saxifraga stolonifera, a lipid extract of common wheat, and a hydrophilic extract of the wood of Leymus chinensis. The five synthetic biomimetic peptides are selected from EGF, IGF-1, acidic FGF, basic FGF, and VEGF, as well as dermatologically acceptable excipients.

[0011] Finally, this invention relates to a method for preparing the cosmetic and dermatological compositions of the present invention, the method comprising: The lipid extract from the root of Coleus fasciatus was mixed with the lipid extract from common wheat. The mixture was added to an aqueous phase containing a hydrophilic extract of Saxifraga stolonifera, a hydrophilic extract of Gleditsia sinensis wood, and the at least five synthetic biomimetic peptides. Stir and cool to room temperature.

[0012] definition In this invention, the terms are defined as follows: "Improving hair density" refers to improving scalp quality and stimulating hair regeneration. This improvement in hair density is applicable to hair loss or baldness, as well as to fine and sparse hair, and to maintaining the results of hair transplantation.

[0013] "Synthetic biomimetic peptides" refer to peptides involved in cell repair processes, such as EGF, IGF-1, acidic FGF, basic FGF, and VEGF. These peptides are reproduced through synthetic or biosynthetic methods and, in turn, mimic these various cell repair processes—hence the name biomimetic peptides. Synthetic reproduction is a chemical synthesis method, while biosynthetic reproduction is a biotechnological synthesis method (by inserting its encoding gene into bacteria).

[0014] "Coleus Forskohlii Root Extract" refers to an extract obtained, for example, by supercritical CO2 extraction.

[0015] Coleus fortis, or Indian Coleus fortis, is a tropical perennial plant with recognized lipid metabolism benefits. It has been used in Ayurvedic medicine for centuries and grows in Southeast Asia: India, Thailand, Nepal, Pakistan, and Sri Lanka. In traditional Indian medicine, the root extract of *Coleus forsythia* is used as a tonic due to its components, including chameccydine, alpha-hydroxydemethylcryptojaponol, alpha-cedrene, oleanolic acid, forskolin G, forskolin J, 1,6-diacetyl-9-deoxyforskolin, forskolin A, forskolin H, 6-acetyl-1-deoxyforskolin, betulinic acid, and beta-sitosterol. Other related therapeutic properties include anthelmintic effects and efficacy in treating skin infections and rashes.

[0016] "Hydrophilic extract of Saxifraga rotundifolia (INCI name: Saxifraga rotundifolia extract)" refers to an extract obtained, for example, using an ultrasonic extractor from the REUS brand, in the presence of water and glycerin.

[0017] *Saxifraga stolonifera*, a perennial plant in the Saxifragaceae family, can grow to 20-50 cm tall and has round leaves with large serrated edges. Its flowers are small, white, star-shaped, with five petals adorned with small spots, transitioning from yellow at the center to red at the petal tips. It prefers cool environments and grows in shady spots or along streams. In tall mountain grass communities, it often hides in the shade of larger plants. Its hydrophilic extract, composed of polyphenols and glycoside flavonoids, imparts anti-inflammatory activity.

[0018] "Lipid extracts of common wheat (INCI name: Glycosphingolipids (and) Glycolipids)" refers to extracts obtained, for example, by supercritical CO2 extraction.

[0019] Lipid extracts from common wheat belong to the "anti-inflammatory" class and are primarily used to treat vaginal yeast infections, pressure sores (bedsores), skin injuries, and burns. This common wheat extract contains vitamin E, which can be used as a skin conditioner. It also aids in the healing process and skin regeneration. It contains antioxidants, which are used to treat skin scars. When used for yeast infections, it effectively reduces the severity of symptoms such as vaginal dryness, pain, burning sensation, itching, and discharge.

[0020] "Hydrophilic extract of soapberry wood (INCI name: Quillaja Saponaria Wood Extract)" refers to an extract obtained, for example, using an ultrasonic extractor from the REUS brand in the presence of deionized water.

[0021] The soapberry tree (Quillaja saponaria), also known as the Chilean soapberry, has had its hydrophilic wood extract used in its native region for a long time to treat various chest and skin ailments. It is considered particularly effective as an expectorant, helping the body clear mucus by promoting the production of thin mucus. The soapberry tree has also been used to treat dandruff and skin ulcers as it is a mild cleanser. Recent research suggests that the tree may be effective in reducing or even preventing rotavirus infections in children. Topical uses of the soapberry tree include treating dandruff, skin ulcers, and other epidermal problems. Extracts from the plant have anti-inflammatory properties. It is also believed to soothe itchy skin and scalp irritation.

[0022] "EGF peptide," or epidermal growth factor, is a polypeptide composed of 53 amino acids. A chemically synthesized biomimetic EGF peptide is called sh-Oligopeptide-1.

[0023] "IGF-1 peptide," or insulin-like growth factor 1 (IGF1), also known as somatomedin C, is a protein produced by the liver, muscles, and other tissues in response to growth hormone (GH) stimulation. IGF-1 promotes bone growth and muscle mass increase. A chemically synthesized biomimetic IGF-1 peptide is named sh-Oligopeptide-2.

[0024] "Acidic FGF peptide," or acidic fibroblast growth factor, is marketed, for example, under the brand name SIGMA on the priority date. A chemically synthesized biomimetic acidic FGF peptide is named sh-Polypeptide-1.

[0025] "Basic FGF peptide," or Basic Fibroblast Growth Factor, is marketed, for example, under the brand name SIGMA on the priority date. A chemically synthesized biomimetic FGF Acid peptide is named sh-Polypeptide-9.

[0026] "VEDF peptide," or Vascular Endothelium Growth Factor, was marketed, for example, under the brand name SIGMA on the priority date. A chemically synthesized biomimetic VEDF peptide was named sh-Polypeptide-11.

[0027] "Active ingredient" refers to a substance that exhibits the desired activity and / or therapeutic effect in a cosmetic composition or dermatological medicine, and is usually present in a very small proportion relative to the excipients.

[0028] "Physiologically acceptable" and "dermatologically acceptable" means that it is suitable for topical use, use in contact with human skin, or use via other routes of administration (such as oral or injectable administration) without the risk of toxicity, incompatibility, instability, or allergic reactions. Attached Figure Description

[0029] Other features, advantages, and details of the invention will be better understood by reading the following description in conjunction with the accompanying drawings, wherein: [ Figure 1 The results of the labeling were shown, which also revealed the expression of hyaluronic acid in the papillary dermis and epidermis of the scalp; [ Figure 2 The image shows the labeling results of PINK1 in hair follicle cells, and the resulting expression levels. [ Figure 3 The image shows the labeling results of Nrf2 and the resulting representations. Detailed Implementation

[0030] This invention relates to a cosmetic composition for improving hair density, comprising an effective amount of at least four plant extracts combined with at least five synthetic biomimetic peptides as active ingredients. The four plant extracts are selected from lipid extracts of the roots of *Coleus forsythia suspensa*, hydrophilic extracts of *Saxifraga stolonifera*, lipid extracts of common wheat, and hydrophilic extracts of *Gnaphalium affine* wood. The five synthetic biomimetic peptides are selected from EGF, IGF-1, acidic FGF, basic FGF, and VEGF, as well as physiologically acceptable excipients.

[0031] Surprisingly, the inventors discovered that by combining four plant extracts—a lipid extract selected from the root of *Coleus fortis*, a hydrophilic extract from *Saxifraga stolonifera*, a lipid extract from common wheat, and a hydrophilic extract from the wood of *Gnaphalium affine*—with five synthetic biomimetic peptides—selected from EGF, IGF-1, acidic FGF, basic FGF, and VEGF—the cosmetic composition of the present invention achieves the following: It can increase the expression of hyaluronic acid in the dermis of the scalp nipple, thus it has moisturizing and repairing properties for skin tissue and participates in the cell proliferation of skin tissue cells; It can increase the expression of PINK1 in hair follicle cells, thus it is involved in maintaining cell homeostasis; It can reduce the expression of Nrf2 in the epidermis, thus possessing anti-free radical properties; It can increase the concentration of ATP in hair follicle cells, thus promoting proliferation and inhibiting terminal differentiation of keratinocytes; In addition, it helps in the repair of the scalp skin barrier.

[0032] According to a particular embodiment, the cosmetic composition of the present invention may optionally further comprise at least one additional plant extract different from the four extracts described above, and / or at least one additional biomimetic peptide different from the five peptides described above.

[0033] According to a particular embodiment, in the cosmetic composition according to the invention, each of the at least four plant extracts is present at a weight of 0.01% to 5.5% relative to the total weight of the composition, and each of the at least five peptides is present at a weight of 0.005% to 1.5% relative to the total weight of the composition.

[0034] According to a particular embodiment, in the cosmetic composition according to the invention, each of the at least four plant extracts is present at a weight of 0.05% to 5% relative to the total weight of the composition, and each of the at least five peptides is present at a weight of 0.01% to 1% relative to the total weight of the composition, for example, the plant extracts are present at a weight of 0.2%, 0.5%, 0.7%, and 0.9%, and the biomimetic peptides are present at a weight of 0.02%, 0.04%, 0.07%, and 0.08%.

[0035] According to one embodiment, in the cosmetic composition according to the invention, the biomimetic peptides are chemically synthesized biomimetic peptides sh-oligopeptide-1, sh-oligopeptide-2, sh-polypeptide-1, sh-polypeptide-9, and sh-polypeptide-11.

[0036] According to one particular embodiment, in the cosmetic composition according to the invention, the active ingredient comprises four plant extracts combined with five biomimetic peptides (preferably biomimetic peptides or chemically synthesized peptides).

[0037] The cosmetic compositions according to the invention may optionally further include any additives commonly used in the intended application area, as well as auxiliaries required for their formulation, such as solvents, thickeners, diluents, antioxidants, colorants, sunscreens, self-tanning agents, pigments, fillers, preservatives, fragrances, deodorants, cosmetic or pharmaceutical active ingredients, essential oils, vitamins, essential fatty acids, surfactants, film-forming polymers, etc.

[0038] The INCI Dictionary and Handbook (International Nomenclature of Cosmetic Ingredients (13th Edition, 2010), published by the Personal Care Products Council, Inc., Washington, D.C., describes a variety of (but not limited to) cosmetic and pharmaceutical ingredients commonly used in the skincare industry that are suitable as additional ingredients in the compositions of this invention.

[0039] When present in a cosmetic composition according to the invention, the total amount of additional compounds, in addition to the active ingredient of the invention, is advantageously from 0.01% to 60% relative to the total weight of the composition. It should be understood that those skilled in the art should carefully select these one or more possible additional compounds so that the addition of these possible compounds does not, or substantially does not, alter the inherently relevant advantageous properties of the cosmetic composition according to the invention.

[0040] The present invention also relates to a method for hair and scalp beauty care, characterized in that the method comprises applying about 0.5 to 1.5 g of an effective amount of the composition according to the invention to the hair and scalp twice daily for about 1 to 9 minutes, particularly 3 to 7 minutes, and optionally rinsing with water or shampoo.

[0041] According to one embodiment of the cosmetic care method of the present invention, the scalp is treated with light emitted by light-emitting diodes (LEDs) at wavelengths of 670 nm and 880 nm for a duration between 10 and 60 minutes, particularly between 15 and 45 minutes, typically between 25 and 35 minutes. The treatment can be applied from once a week to once daily, typically 2 to 3 times a week.

[0042] The present invention also relates to a dermatological composition for use as a medicament for the prevention and treatment of hair loss, comprising an effective amount of at least four plant extracts combined with at least five synthetic biomimetic peptides as active ingredients. The four plant extracts are selected from a lipid extract of the root of Coleus forsythia, a hydrophilic extract of Saxifraga stolonifera, a lipid extract of common wheat, and a hydrophilic extract of the wood of Leymus chinensis. The five synthetic biomimetic peptides are selected from EGF, IGF-1, acidic FGF, basic FGF, and VEGF, as well as dermatologically acceptable excipients.

[0043] According to one embodiment, in the dermatological composition according to the invention, each of the at least four plant extracts is present at a weight of 3% to 5.5% relative to the total weight of the composition, and each of the at least five peptides is present at a weight of 0.6% to 1.5% relative to the total weight of the composition.

[0044] According to one embodiment, in the dermatological composition according to the invention, the biomimetic peptides are chemically synthesized biomimetic peptides sh-oligopeptide-1, sh-oligopeptide-2, sh-polypeptide-1, sh-polypeptide-9, and sh-polypeptide-11.

[0045] According to one particular embodiment, in the dermatological composition according to the invention, the active ingredient comprises four plant extracts combined with five biomimetic peptides (preferably chemically synthesized biomimetic peptides).

[0046] The dermatological compositions according to the invention may optionally further include any additives commonly used in the intended application area, as well as excipients required for their formulation, as previously mentioned for cosmetic compositions.

[0047] The present invention also relates to a method for preparing the cosmetic and dermatological compositions as described above, the method comprising: - The lipid extract of the root of Coleus fasciatus is mixed with the lipid extract of common wheat, for example, by stirring, and the mixing is carried out at a temperature above room temperature, for example, above 70°C, or even above 75°C, or even still above 80°C. - The mixture is added to an aqueous phase containing a hydrophilic extract of Saxifraga rotundifolia, a hydrophilic extract of Sapindus mukorossi wood, and at least five synthetic biomimetic peptides. This addition is, for example, carried out with stirring, or added to an aqueous phase heated to a temperature above room temperature (e.g., above 70°C, or even above 75°C, or even above 80°C, for example, the same temperature as the mixture). - Stir and cool to room temperature. Stirring may continue for at least 30 minutes, for example, between 45 and 90 minutes. During this stage, preservatives may be added to the composition. Cooling may be forced or natural.

[0048] The compositions according to the invention may be particularly present in the form of creams, oil-in-water emulsions or water-in-oil emulsions or multiple emulsions, serums, solutions, suspensions, gels, lotions, lotions, sticks or even powders, and are suitable for application to the skin and / or hair.

[0049] The compositions according to the invention are particularly composed of a hair care composition, especially shampoos, conditioners, conditioning lotions, styling creams or gels, hair rebuilding lotions, hair masks, etc. The cosmetic compositions according to the invention can be used specifically for application involving rinseable or non-rinseable treatments, or in the form of a shampoo. They can also be taken as hair dyes or mascaras applied with a brush or comb, particularly for application to eyelashes, eyebrows, or hair.

[0050] According to one embodiment of the present invention, the cosmetic or dermatological composition according to the present invention is in the form of gel, lotion, foam, conditioner, cream, spray or serum.

[0051] Example The invention will be better understood by reading the following embodiments which are described in a non-limiting manner.

[0052] Example 1: Testing of different biological activities of the composition To demonstrate the efficacy of the composition of the present invention, the inventors analyzed the bioactivity of the composition in vitro on human skin explants with hair follicles through histological, biochemical, and genomic studies, and found that the composition according to the present invention induces the following effects in human skin and hair follicles: - Increase hyaluronic acid synthesis; - Added PINK1 expression; - Reduce Nrf2 expression; - Increase ATP synthesis; - Regeneration of the scalp skin barrier; - Differentiation of keratinocytes.

[0053] Histological and biochemical studies further include analyzing the effects of the composition according to the invention on the expression of hyaluronic acid, PINK1 and Nrf2 and their ATP concentrations in cells derived from human skin explants.

[0054] Therefore, the effects of the following products were tested: Product P1 contains 5% by weight of Coleus forsythia lipid extract, 5% by weight of Saxifraga rotundifolia hydrophilic extract, 5% by weight of common wheat lipid extract, 5% by weight of Sapindus mukorossi hydrophilic extract, and 1% by weight of five biosynthetic peptides [EGF, IGF-1, acidic FGF, basic FGF, and VEGF]. This mixture is replenished with 79% by weight of deionized water and sonicated for 10 minutes at room temperature (BRANSON ultrasound probe and equipment).

[0055] Product P2 contains 1% by weight of Coleus forsythia lipid extract, 1% by weight of Saxifraga rotundifolia hydrophilic extract, 1% by weight of common wheat lipid extract, 1% by weight of Sophora japonica extract, and 0.1% by weight of five biosynthetic peptides [EGF, IGF-1, acidic FGF, basic FGF, and VEGF]. This mixture is replenished with 95.9% by weight of deionized water and sonicated at room temperature for 10 minutes.

[0056] Product E consists of 100% distilled water by mass.

[0057] Human skin explants obtained from scalp reshaping surgery were prepared and divided into four batches: - Batch T0, corresponding to the plastic surgery control group, contains 3 explants: - Batch T, corresponding to the control group, contains 6 explants; - Batch E, corresponding to the batch treated with product E, contains 6 explants. - Batch P1 corresponds to the batch treated with product P1 and contains 6 explants; - Batch P2 corresponds to the batch treated with product P2 and contains 6 explants.

[0058] Each skin explant was placed in a culture medium at 37°C in a humid atmosphere rich in 5% CO2, and was in survival mode.

[0059] At T = 0 days (D0), T = 1 day (D1), T = 4 days (D4), T = 6 days (D6), and T = 7 days (D7), 5 μL of product E, P1, and P2 were applied topically to the explants of batches E, P1, and P2, respectively.

[0060] At D0, three explants from batch T0 were collected and cut into three parts. One part was fixed in buffered formalin solution, the second part was frozen at -80°C for histological analysis, and the third part was frozen at -80°C for biochemical analysis.

[0061] At T = 5 days (D5) and T = 8 days (D8), three explants from batches T, E, P1 and P2 were collected and treated in the same manner as the explants from batch T0.

[0062] After soaking in buffered formalin for 24 hours, the samples were dehydrated, embedded in paraffin, and prepared into paraffin blocks. 5 μm thick sections were prepared and mounted on histological glass slides.

[0063] After staining with Masson's trichrome staining method and Goldner's variation method, the overall morphology of the dermis and epidermis was analyzed on paraffin sections and evaluated by microscopic examination.

[0064] Hyaluronic acid (HABP) immunolabeling was performed on samples from batches T0, T at days 5 (TJ5) and 8 (TJ8), E at days 5 (EJ5) and 8 (ED8), P1 at days 5 (P1J5) and 8 (P1J8), and P2 at days 5 (P2J5) and 8 (P2J8) to observe the expression of hyaluronic acid in the dermis and epidermis of the scalp nipple. Hyaluronic acid was labeled with HABP (hyaluronic acid binding protein) on formalin-fixed paraffin sections. HABP was diluted 1:800 in 0.3% PBS-BSA and labeled for 1 hour at room temperature using a biotin / streptavidin amplification system, followed by VIP staining. The immunolabeling was then evaluated by microscopic observation.

[0065] PINK1 expression in hair follicle cells was observed by immunolabeling of samples from T0, T on days 5 (TJ5) and 8 (TJ8), E on days 5 (EJ5) and 8 (EJ8), P1 at J5 (P1J5) and J8 (P1J8), and P2 at J5 (P2J5) and J8 (P2J8). PINK1 labeling was performed on paraffin sections using a monoclonal anti-PINK1 antibody diluted 1:100 in 0.3% PBS20 BSA 0.3%, incubated overnight at 4°C, amplified using a biotin / streptavidin system, and developed via VIP staining. Immunolabeling was then evaluated by microscopic observation.

[0066] Nrf2 expression in cells was observed by immunolabeling of batches T0, T on day 5 (TJ5) and day 8 (TJ8), E on day 5 (EJ5) and day 8 (EJ8), P1 on J5 (P1J5) and J8 (P1J8), and P2 on J5 (P2J5) and J8 (P2J8). Nrf2 labeling was performed on paraffin sections using a monoclonal anti-Nrf2 antibody diluted 1:400 in 0.3% PBS-BSA-Tween 20 0.05%, incubated at room temperature for one hour, amplified using a biotin / streptavidin system, and developed with VIP staining. The immunolabeling was then evaluated by microscopic observation.

[0067] ATP was measured at T0, T on day 5 (TJ5) and day 8 (TJ8), E on day 5 (EJ5) and day 8 (EJ8), P1 on day 5 (P1J5) and day 8 (P1J8), and P2 on day 5 (P2J5) and day 8 (P2J8). Frozen skin explants were first ground and then sonicated to disrupt the cells. After centrifugation at 4000 rpm at 18°C, the protein content in the supernatant was determined using the Bradford method. ATP in the explants was quantified using an ATP assay kit and a fluorometer.

[0068] exist Figures 1 to 3 In this context, TF corresponds to very weak labeling, F to weak labeling, M to moderate labeling, AN to fairly obvious labeling, N to obvious labeling, TH to very obvious labeling, and Fo to strong labeling.

[0069] Figure 1 The labeling results of hyaluronic acid in the dermis and epidermis of the scalp nipple are shown, along with the resulting expression.

[0070] Hyaluronic acid is a non-sulfated glycosaminoglycan composed of repeating units of D-glucuronic acid and DN-acetylglucosamine. It is a major component of the skin and significantly participates in various cellular processes, including hydration, skin tissue repair, and cell proliferation. For example, under chronic ultraviolet radiation, hyaluronic acid synthesis decreases, while hyaluronidase activity remains unchanged. Therefore, significant degradation of hyaluronic acid can be observed. HABP (hyaluronic acid-binding protein) is a protein that binds hyaluronic acid to other components of the dermal extracellular matrix, including proteoglycans. It is used in dermatological studies to visualize hyaluronic acid within skin tissue.

[0071] from Figure 1 As can be seen, the hyaluronic acid level in the epidermis is low in batch T0, low to medium in batches TJ8, EJ8, P1J8 and P2J8, and medium in batches TJ5, EJ5, P1J5 and P2J5.

[0072] Therefore, neither product P1 nor product P2 caused any change in hyaluronic acid expression compared to the T control group and product E, whether on day 5 or day 8.

[0073] Hyaluronic acid markers in the dermis of the nipple were low to moderate in batches TJ8, EJ8, P1J8, and P2J8; moderate in batches TJ5, EJ5, and P2J5; and quite pronounced in batches T0 and P1J5.

[0074] Therefore, in the dermis of the nipple, product P1 caused a moderate increase in hyaluronic acid expression compared with control group T and product E.

[0075] Therefore, the composition according to the invention is effective in increasing hyaluronic acid expression in the dermal layer of the scalp nipple. Thus, it possesses moisturizing and skin tissue repair properties and participates in the proliferation of skin tissue cells.

[0076] Figure 2 The results of PINK1 labeling in hair follicle cells and the resulting expression are shown.

[0077] PINK1 (P-TEN-Induced Putative Kinase 1) is a protein involved in mitophagy, the selective autophagy of damaged and nonfunctional mitochondria. It specifically accumulates on the outer membrane surface of damaged mitochondria whose electrochemical potential is disturbed. In skin tissue and hair follicle cells, mitophagy is stimulated by oxidative stress, which is associated with the accumulation of reactive oxygen species (ROS), helping to clear damaged mitochondria and maintain cellular homeostasis. PINK1 is also associated with anti-aging cellular mechanisms, particularly at the skin level.

[0078] PINK1 markers in hair follicle cells were low in batch T0, low to moderate in batches TJ8 and EJ8, moderate in batches TJ5 and EJ5, and moderate to quite prominent in batches P1J5, P2J5, P1J8, and P2J8.

[0079] Therefore, both products P1 and P2 caused an increase in PINK1 expression compared to the T control group and product E, whether on day 5 or day 8.

[0080] Therefore, the composition according to the present invention increases PINK1 expression at the cellular level. Thus, it participates in maintaining the cellular homeostasis of hair follicle cells and possesses anti-aging properties.

[0081] Figure 3 The labeling results of Nrf2 in cells are shown, along with the resulting expression.

[0082] Nrf2 is a transcription factor that plays a crucial role in oxidative stress response and is constitutively expressed in a variety of cells and tissues. Under oxidative stress (including UV exposure), Nrf2 is phosphorylated, prompting its translocation from the cytoplasm to the nucleus. Once translocated to the nucleus, Nrf2 can bind to specific regions of DNA known as HAREs (Human Antioxidant Response Elements). Nrf2 plays a key role in protecting hair follicle keratinocytes from damage caused by UV radiation and PM2.5 particulate matter pollution.

[0083] The Nrf2 markers in the epidermis were low to moderate in batches TJ8, EJ8, P1J8, and P2J8; moderate in batch P1J5; moderate to fairly prominent in batches TJ5, EJ5, and P2J5; and fairly prominent to prominent in batch T0.

[0084] Therefore, compared with the control group T and product E, product P1 caused a decrease in Nrf2 expression.

[0085] Therefore, the compositions of the present invention reduce Nrf2 expression at the cellular level. This reduction in Nrf2 expression leads to its translocation from the cytoplasm to the nucleus, where it activates genes responsible for cellular protection, particularly the synthesis of antioxidant enzymes. Thus, the compositions of the present invention possess free radical scavenging and antioxidant properties.

[0086] The following table shows the ATP measurement results for batches T0, TJ5, EJ5, P1J5, P2J5, TJ8, EJ8, P1J8, and P2J8.

[0087] [Table 1] ATP (adenosine triphosphate) is a nucleotide known to stimulate and participate in numerous intracellular processes. Extracellular ATP and its metabolite adenosine can exert a variety of effects on almost all human skin cell types, including scalp cells, and the skin is a potentially rich source of ATP. Sources of ATP in the skin include, for example, ATP released during mechanical injury due to tissue hypoxia and cell lysis, and ATP released by keratinocytes in response to nucleotides or cytokines. ATP promotes keratinocyte proliferation and inhibits terminal differentiation of keratinocytes through P2Y2 receptors located in the basal layer of the epidermis. ATP also plays an important role in the tissue skin's response to mechanical injury, both by regulating the inflammatory process of Langerhans cells and by directly stimulating skin regeneration and healing processes. Furthermore, ATP is a crucial energy molecule for intracellular processes and plays a major role in skin regeneration. With age, ATP levels in the dermis decline, leading to energy deficiency that affects skin appearance. Therefore, ATP is effective in providing anti-aging protection and also possesses moisturizing and soothing properties.

[0088] Therefore, it was observed that products P1 and P2 both caused an increase in ATP concentration compared with the control group T and product E.

[0089] The compositions according to the invention increase the ATP concentration in epidermal cells. Increased ATP production at the skin level helps improve skin's metabolic function. Therefore, the compositions according to the invention promote the proliferation of follicular keratinocytes and inhibit their terminal differentiation. The compositions according to the invention also help repair the skin barrier of the scalp.

[0090] Compositions containing 0.1% to 1% by weight of *Coleus fordii* lipid extract, 0.1% to 1% by weight of *Saxifraga rotundifolia* hydrophilic extract, 0.1% to 1% by weight of common wheat lipid extract, 0.1% to 1% by weight of *Sapindus mukorossi* extract, and 0.01% to 0.09% by weight of five biosynthetic peptides [EGF, IGF-1, acidic FGF, basic FGF, and VEGF]. These mixtures were replenished with deionized water and sonicated at room temperature for 10 minutes. These compositions were found to have the same properties as products P1 and P2 described above.

[0091] The following description relates to examples of cosmetic compositions according to the present invention.

[0092] Example 2: Scalp Serum [Table 2] Example 3: Hair Mask [Table 3] Example 4: Hair Spray [Table 4] Example 5: CBD Hair Conditioning Toner [Table 5]

Claims

1. A cosmetic composition for improving hair density, comprising an effective amount of an active ingredient combining at least four plant extracts with at least five synthetic biomimetic peptides, wherein the four plant extracts are selected from a lipid extract of the root of *Coleus forsythia suspensa*, a hydrophilic extract of *Saxifraga stolonifera*, a lipid extract of common wheat, and a hydrophilic extract of *Gnaphalium affine* wood, and the five synthetic biomimetic peptides are selected from EGF, IGF-1, acidic FGF, basic FGF, and VEGF, as well as physiologically acceptable excipients.

2. The composition according to claim 1, characterized in that, The at least four plant extracts are each present in an amount of 0.01% to 5.5% of the total weight of the composition, and the at least five peptides are each present in an amount of 0.005% to 1.5% of the total weight of the composition.

3. The composition according to claim 1 or 2, characterized in that, The at least four plant extracts are each present in an amount of 0.05% to 5% relative to the total weight of the composition, and the at least five peptides are each present in an amount of 0.01% to 1% relative to the total weight of the composition.

4. The composition according to any one of claims 1 to 3, characterized in that, The biomimetic peptides are chemically synthesized peptides sh-oligopeptide-1, sh-oligopeptide-2, sh-polypeptide-1, sh-polypeptide-9, and sh-polypeptide-11.

5. The composition according to any one of claims 1 to 4, characterized in that, The active ingredient consists of four plant extracts combined with five biomimetic peptides (preferably chemically synthesized biomimetic peptides).

6. A method for hair and scalp beauty care, characterized in that, The method comprises applying about 0.5 g to 1.5 g of an effective amount of the composition according to any one of claims 1 to 5 to the hair and scalp twice daily for about 1 to 9 minutes, particularly 3 to 7 minutes, and optionally rinsing with water or shampoo.

7. The method according to claim 6, characterized in that, Light treatment with wavelengths of 670 nm and 880 nm is applied to the scalp for a duration of 10 to 60 minutes, particularly between 15 and 45 minutes, typically between 25 and 35 minutes.

8. A dermatological composition for the prevention and treatment of hair loss, comprising an effective amount of an active ingredient consisting of at least four plant extracts combined with at least five synthetic biomimetic peptides, wherein the four plant extracts are selected from a lipid extract of the root of *Coleus forsythia suspensa*, a hydrophilic extract of *Saxifraga stolonifera*, a lipid extract of common wheat, and a hydrophilic extract of *Gnaphalium affine* wood, and the five synthetic biomimetic peptides are selected from EGF, IGF-1, acidic FGF, basic FGF, and VEGF, as well as dermatologically acceptable excipients.

9. A method for preparing cosmetic and dermatological compositions according to claims 1 and 8, the method comprising: - Mix the lipid extract from the root of Coleus fasciatus with the lipid extract from common wheat; - Add the mixture to an aqueous phase containing a hydrophilic extract of Saxifraga stolonifera, a hydrophilic extract of Sapindus mukorossi wood, and the at least five synthetic biomimetic peptides; - Stir and cool to room temperature.

10. The composition according to claim 1 or claim 8, characterized in that, It comes in the form of gel, lotion, foam, conditioner, cream, spray, or serum.

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