Scalp anti-aging composition as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of daily chemical products, and particularly relates to a scalp anti-aging composition and a preparation method and application thereof. The scalp anti-aging composition provided by the invention is prepared from the following components in parts by mass: 20 to 40 parts of nicotinamide adenine dinucleotide, 20 to 40 parts of medicinal phellinus igniarius fermentation product filtrate, 0.8 to 3.0 parts of hydrolyzed sodium hyaluronate, 2 to 8 parts of collagen, 12 to 40 parts of mannitol and 8 to 25 parts of trehalose. The scalp anti-aging composition provided by the invention can promote dermal papilla cell proliferation, inhibit inflammatory factor TNF-alpha gene expression, inhibit fibroblast matrix metalloproteinase-1 gene expression, and can go deep into a cellular level to realize a scalp anti-aging effect. The anti-aging composition for scalp is used as a core component, the prepared essence can relieve red and swollen scalp, nourish scalp and the like, and a very good anti-aging effect for scalp is achieved.

Description

Technical Field

[0001] This invention belongs to the field of daily chemical products technology, specifically relating to a scalp anti-aging composition, its preparation method, and its application. Background Technology

[0002] With the improvement of living standards and the deepening of health and beauty concepts, people's attention to skin care has extended from the face to the scalp. As an important part of human skin, the scalp shares the same structure as facial skin and plays a special role in supporting hair and regulating the microenvironment of hair follicles. At the same time, it is more susceptible to external stimuli and physiological metabolic effects. In recent years, problems related to scalp aging, such as thinning hair, scalp sensitivity, and dry, brittle hair, have become increasingly prominent, becoming a common need affecting people of different ages. This has driven scalp anti-aging to become a research hotspot in the beauty and health fields.

[0003] Scalp aging is not a single-dimensional physiological change, but a complex process involving multiple aspects such as cellular function decline, inflammatory response imbalance, and extracellular matrix degradation. For example, dermal papilla cells, as the "signaling center" for hair follicle growth, directly determine the growth cycle and activity of hair follicles through their proliferative capacity. A decrease in the number or functional decline of dermal papilla cells can lead to hair follicle atrophy, resulting in thinning and hair loss. Abnormal fibroblast function can cause the dermal layer of the scalp to loosen and lose its water retention capacity, accompanied by the uncontrolled expression of inflammatory factors, further exacerbating damage to the scalp barrier.

[0004] Currently, most scalp anti-aging products on the market focus on a single function, such as moisturizing products with hydrolyzed sodium hyaluronate as the core, soothing products with plant extracts as the core, or structural repair products that only add a single collagen ingredient. They lack the ability to systematically regulate scalp aging across multiple targets and pathways. For example, while some products can improve scalp dryness by supplementing with sodium hyaluronate, they cannot intervene in the proliferation disorders of hair papilla cells or the expression of inflammatory factors; and traditional collagen products are difficult to be effectively absorbed by the scalp due to their molecular structure. The limitations of existing technology make the development of a scalp anti-aging composition with synergistic ingredients, precise action, and comprehensive effects an urgent task to meet market demands and overcome technological bottlenecks. Summary of the Invention

[0005] To address the technical shortcomings of existing scalp anti-aging products, such as limited functionality, narrow target areas, low penetration of active ingredients, and poor synergy, this invention aims to provide a scalp anti-aging composition, its preparation method, and its applications. The scalp anti-aging composition provided by this invention can promote the proliferation of dermal papilla cells, inhibit the expression of the inflammatory factor TNF-α gene, and inhibit the expression of the matrix metalloproteinase-1 gene in fibroblasts, achieving anti-aging effects at the cellular level. Using this scalp anti-aging composition as a core component, the resulting serum can soothe scalp redness and swelling, nourish the scalp, and achieve excellent anti-aging effects.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: The first objective of this invention is to provide a scalp anti-aging composition comprising, by weight: 20-40 parts nicotinamide adenine dinucleotide, 20-40 parts medicinal fumonis fermentation product filtrate, 0.8-3.0 parts hydrolyzed sodium hyaluronate, 2-8 parts collagen, 12-40 parts mannitol and 8-25 parts trehalose.

[0007] Preferably, the medicinal Fermentation Product Filtrate is prepared by fermentation with Lactobacillus fermentation; the collagen is recombinant type III collagen.

[0008] Preferably, the method for preparing the fermentation product filtrate of the medicinal Fertilizer includes: S1. Inoculate Lactobacillus fermentum colonies into MRS medium and culture at 33-40℃ and 80-120 rpm for 40-54 h to obtain a bacterial suspension. S2. Inoculate the bacterial suspension into MRS medium at an inoculation rate of 3-7% (v / v) and culture for 40-54 hours at 33-40℃ and 80-120 rpm to obtain the seed culture. S3. Inoculate the seed culture into a fermentation medium containing medicinal Fertilizer powder at an inoculation rate of 1-3% (v / v), and ferment at 33-40℃, 80-120 rpm, and sterile air ventilation rate of 0.2-0.6 vvm for 30-45 h to obtain crude fermentation broth. S4. Sterilize the crude fermentation broth at 80~90℃ for 20~40 minutes, filter, and obtain the fermentation product filtrate of medicinal Fertilizer.

[0009] Preferably, the fermentation medium containing medicinal fumonis in step S3 includes medicinal fumonis powder, carbon source, nitrogen source, inorganic salts and water; the amount of medicinal fumonis added is 10~40g / L; The nitrogen source includes at least two of peptone, beef extract, and yeast powder, and the amount of nitrogen source added is 15~35g / L; The carbon source includes at least one of glucose, sucrose, and fructose syrup, and the amount of nitrogen source added is 15~30g / L; The inorganic salt includes at least three of the following: magnesium sulfate, manganese sulfate, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, diammonium hydrogen phosphate, and ammonium dihydrogen phosphate. The amount of the inorganic salt added is 2-8 g / L.

[0010] A second objective of this invention is to provide a method for preparing the above-mentioned scalp anti-aging composition, comprising the following steps: (1) Prepare the fermentation product filtrate of medicinal fumonis through fermentation with Lactobacillus fermentum, and set it aside for later use; (2) Mix the fermentation product filtrate of medicinal fumonis from step (1), hydrolyzed sodium hyaluronate, collagen, mannitol and trehalose, and homogenize under high pressure at 100-150 MPa for 3-6 times to obtain mixture A1; (3) Add nicotinamide adenine dinucleotide to mixture A1 from step (2) and stir at 500-1000 rpm for 20-40 min to obtain the scalp anti-aging composition.

[0011] Another object of the present invention is to provide an application of the above-mentioned anti-aging scalp composition in the preparation of hair products, wherein the hair products are selected from at least one of scalp care essence, scalp care water, scalp nourishing cream, scalp massage cream, hair essence, hair oil, shampoo, shampoo bar, shampoo powder, hair mask, conditioner, hair spray, hair gel, and hair lotion.

[0012] Preferably, the amount of the composition added to the hair product is 0.1 to 5 wt%.

[0013] Another object of the present invention is to provide a scalp anti-aging serum, comprising, by weight percentage: 0.1-5% of the above-mentioned scalp anti-aging composition, 3-10% glycerin, 3-10% 1,3-butanediol, 0.005-0.02% sodium hyaluronate (small molecule), 0.5-2.0% erythritol, 0.1-0.4% Active Moist 24 polyamino acid polysaccharide, 0.05-0.2% Lactobacillus fermentation product, 0.3-1.0% phenoxyethanol, 0.3-1.0% 1,2-hexanediol, 0.05-0.2% Microcare Emollient EHG, 0.4-1.2% DN-5, 0.05-0.2% citric acid, and the balance being water.

[0014] Preferably, the small molecule sodium hyaluronate was purchased from Baihong Huajia Biotechnology (Shandong) Group Co., Ltd., model Polyglycin® microsphere recombinant collagen; the Active Moist 24 multi-amino acid polysaccharide was purchased from Guangzhou Fuyulong Biotechnology Co., Ltd.; the Lactobacillus fermentation product was purchased from Guangzhou Runheng Biotechnology Co., Ltd.; the MicrocareEmollient EHG was purchased from Tor Specialty Chemicals (Zhenjiang) Co., Ltd.; and the DN-5 was purchased from Sanei Chemical Co., Ltd., Japan.

[0015] Another object of the present invention is to provide a method for preparing the above-mentioned anti-aging scalp serum, comprising the following steps: S1. Weigh out water, glycerin and 1,3-butanediol according to the formula, stir them evenly and heat them to 80~85℃, keep them at this temperature and stir to obtain mixture B1. S2. After the mixture B1 from step S1 has cooled to 40~45℃, add the scalp anti-aging composition, small molecule sodium hyaluronate, erythritol, Active Moist 24 polyamino acid polysaccharide, lactobacillus fermentation product, phenoxyethanol, 1,2-hexanediol and Microcare Emollient EHG to it according to the formula amount, and stir evenly to obtain mixture B2. S3. Add the formula amount DN-5 to the mixture B2 in step S2, mix evenly, and obtain mixture B3; S4. Add the prescribed amount of citric acid to mixture B3 from step S3, stir well, and obtain the scalp anti-aging essence.

[0016] The design principles and concepts of this invention: The scalp anti-aging composition of the present invention is composed of nicotinamide adenine dinucleotide, medicinal filtrate of fermentation product of Phytophoresis flocculation, hydrolyzed sodium hyaluronate, collagen, mannitol and trehalose. These components are key ingredients of the scalp anti-aging composition of the present invention, and they work synergistically to achieve the anti-aging effect of the scalp.

[0017] First, nicotinamide adenine dinucleotide (NADP) also plays a crucial role in this invention. As a core substrate of the mitochondrial respiratory chain, it participates in over 90% of ATP synthesis. Hair follicles, as the "factories" for hair growth, depend on a sufficient energy supply to maintain cell division, keratin synthesis, and cell cycle transitions (anagen-catagen-telogen phase). NADP can provide hair follicle cells with a large amount of energy, maintaining follicle vitality. Furthermore, it provides energy for hair matrix cells to synthesize keratin, revitalizing them and increasing the amount of keratin synthesized, thus transforming hair from "fine, weak downy hair" to "thick, strong hair strands." NADP can also improve the scalp microenvironment by inhibiting inflammatory pathways and scavenging free radicals, alleviating chronic scalp inflammation and thus slowing scalp aging.

[0018] Secondly, the medicinal Fermentation Product Filtrate of *Polyporus medica* is produced through fermentation by the edible fungus *Lactobacillus fermentum*. Its metabolites are rich in low-molecular-weight active components, which have a positive effect on scalp anti-aging. During preparation, the *Polyporus medica* fermentation product filtrate is used as a liquid medium and initially mixed with hydrolyzed sodium hyaluronate, collagen, mannitol, and trehalose. Subsequently, the mixture undergoes high-pressure homogenization. This process activates the weak interactions between the molecules, allowing them to spontaneously aggregate into structurally complete and stable microspheres, using mannitol and trehalose as carriers, in conjunction with the active molecules of collagen and hydrolyzed sodium hyaluronate. This microsphere structure optimizes the skin penetration of active substances such as collagen and hydrolyzed sodium hyaluronate, solving the problem of insufficient permeability of traditional collagen-based ingredients, allowing the active components to more easily reach the deep tissues of the scalp. Simultaneously, the microsphere structure also enables the orderly and sustained release of active substances, preventing rapid loss of active ingredients and prolonging their duration of action on the scalp, thus continuously exerting anti-aging effects.

[0019] Ultimately, the scalp anti-aging composition of the present invention, through the synergistic effect of the above-mentioned multiple components, can effectively promote the proliferation of dermal papilla cells, inhibit the expression of the inflammatory factor TNF-α gene in dermal papilla cells, and inhibit the expression of the MMP-1 gene in scalp fibroblasts, thereby exhibiting a significant scalp anti-aging effect.

[0020] Compared with the prior art, the present invention has the following beneficial effects: The scalp anti-aging composition provided by this invention scientifically combines nicotinamide adenine dinucleotide, medicinal fumonis fermentation product filtrate, hydrolyzed sodium hyaluronate, collagen and other components to form a synergistic system, achieving synergistic enhancement of multi-dimensional anti-aging effects. It can precisely target key anti-aging points of the scalp, with a clear mechanism of action, and slows down scalp aging from the root.

[0021] The preparation process of the scalp anti-aging composition of this invention is mild, without the involvement of high-temperature or highly toxic reagents, thus maximizing the preservation of the activity of each component and balancing safety and efficacy. Simultaneously, high-pressure homogenization activates the weak interactions between the molecules of each component, causing the composite units to spontaneously aggregate into structurally complete and stable microspheres. This solves the problems of insufficient permeability and easy loss of activity in traditional collagen-based ingredients, improving transdermal efficiency and the longevity of efficacy. The preparation process of this invention is simple, low-cost, and easily industrialized.

[0022] The scalp anti-aging composition provided by this invention has mild ingredients and a wide range of applications. It can be used to prepare various scalp anti-aging products such as scalp care essences, shampoos, and hair masks, and has broad market application prospects. Detailed Implementation

[0023] Experimental methods in the following embodiments of the present invention, unless otherwise specified, are generally performed under conventional conditions or as recommended by the manufacturer. All commonly used chemical reagents used in the embodiments are commercially available products.

[0024] Unless otherwise defined, all technical and scientific terms used in this invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The terminology used in this specification is for the purpose of describing particular embodiments only and is not intended to limit the invention.

[0025] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments. It should be understood that these descriptions are merely exemplary and not intended to limit the scope of the invention. Furthermore, descriptions of well-known structures and technologies are omitted in the following description to avoid unnecessarily obscuring the concept of the invention.

[0026] The following embodiments further describe the present invention, but these embodiments are not intended to limit the scope of protection of the present invention.

[0027] 1. Raw materials and reagents The hydrolyzed sodium hyaluronate is sourced from Baihong Huajia Biotechnology (Shandong) Group Co., Ltd., and is modeled as Polyglycin® Microsphere Recombinant Collagen. The recombinant type III collagen was sourced from Baihong Huajia Biotechnology (Shandong) Group Co., Ltd. The small molecule sodium hyaluronate is sourced from Bloomage Biotechnology Co., Ltd. The Active Moist 24 polyamino acid polysaccharide is sourced from Guangzhou Fuyulong Biotechnology Co., Ltd. The lactobacillus fermentation product was obtained from Guangzhou Runheng Biotechnology Co., Ltd. The Microcare Emollient EHG is sourced from Tor Specialty Chemicals (Zhenjiang) Co., Ltd. The DN-5 is sourced from Sanei Chemical Co., Ltd. of Japan; The fermenting Lactobacillus is BNCC194390, which is sourced from Beina Chuanglian Biotechnology Co., Ltd. The Lactobacillus plantarum is ATCC 8014, which is derived from Ningbo Mingzhou Biotechnology Co., Ltd. The Lactobacillus rhamnosus mentioned is BNCC 187896, which originated from Beina Chuanglian Biotechnology Co., Ltd.

[0028] Examples 1-10 Preparation of the scalp anti-aging composition of the present invention The anti-aging scalp composition of the present invention, by weight, is shown in Table 1 below: Table 1. Raw material formulation of the scalp anti-aging composition of the present invention

[0029] The preparation method of the scalp anti-aging composition in Examples 1-8 includes the following steps: (1) Prepare the fermentation product filtrate of medicinal fumonis through fermentation with Lactobacillus fermentum, and set it aside for later use; (2) The fermentation product filtrate of medicinal fumonis from step (1), hydrolyzed sodium hyaluronate, collagen, mannitol and trehalose were mixed and homogenized at 120 MPa for 4 times to obtain mixture A1. (3) Add nicotinamide adenine dinucleotide to mixture A1 from step (1) and stir at 800 rpm for 30 min to obtain the scalp anti-aging composition.

[0030] The preparation of medicinal Fermentation Product Filtrate from Lactobacillus fermentation includes the following steps: S1. Inoculate Lactobacillus fermentum colonies into MRS medium and culture at 37℃ and 100rpm for 48h to obtain a bacterial suspension. S2. Inoculate the bacterial suspension into MRS medium at an inoculum of 5% (v / v) and culture for 48 hours at 37℃ and 100 rpm to obtain the seed culture. S3. Inoculate the seed culture at a rate of 2% (v / v) into a fermentation medium containing 20 g / L of medicinal fumonis powder, and ferment at 37°C, 100 rpm and sterile air flow rate of 0.4 vvm for 36 h to obtain crude fermentation broth. S4. Sterilize the crude fermentation broth at 85℃ for 30 minutes, and filter it through a diatom filter to obtain the fermentation product filtrate of medicinal fumonis.

[0031] The fermentation medium containing medicinal Ferula melitica has the following formula: 20 g / L medicinal Ferula melitica powder, 12 g / L peptone, 10 g / L beef extract powder, 20 g / L glucose, 0.2 g / L magnesium sulfate, 0.1 g / L manganese sulfate, 3 g / L potassium dihydrogen phosphate, and the balance being water.

[0032] Example 9: A method for preparing a scalp anti-aging composition, comprising the following steps: (1) Prepare the fermentation product filtrate of medicinal fumonis through fermentation with Lactobacillus fermentum, and set it aside for later use; (2) The fermentation product filtrate of medicinal fumonis from step (1), hydrolyzed sodium hyaluronate, collagen, mannitol and trehalose were mixed and homogenized under high pressure at 100 MPa 6 times to obtain mixture A1; (3) Add nicotinamide adenine dinucleotide to mixture A1 from step (1) and stir at 500 rpm for 40 min to obtain the scalp anti-aging composition.

[0033] The preparation of medicinal Fermentation Product Filtrate from Lactobacillus fermentation includes the following steps: S1. Inoculate Lactobacillus fermentation colonies into MRS medium and culture at 33℃ and 120rpm for 54h to obtain a bacterial suspension. S2. Inoculate the bacterial suspension into MRS medium at an inoculum of 7% (v / v) and culture for 40 hours at 33℃ and 120 rpm to obtain the seed culture. S3. Inoculate the seed culture at a rate of 1% (v / v) into a fermentation medium containing 15 g / L of medicinal fumonis powder, and ferment at 33°C, 120 rpm, and sterile air ventilation rate of 0.4 vvm for 36 h to obtain crude fermentation broth. S4. Sterilize the crude fermentation broth at 85℃ for 30 minutes, and filter it through a diatom filter to obtain the fermentation product filtrate of medicinal fumonis.

[0034] The fermentation medium containing medicinal fumonis is formulated as follows: 15 g / L medicinal fumonis powder, 5 g / L yeast powder, 10 g / L beef extract powder, 30 g / L sucrose, 0.3 g / L manganese sulfate, 1 g / L potassium dihydrogen phosphate, 0.5 g / L diammonium hydrogen phosphate, and the balance being water.

[0035] Example 10: A method for preparing a scalp anti-aging composition, comprising the following steps: (1) Prepare the fermentation product filtrate of medicinal fumonis through fermentation with Lactobacillus fermentum, and set it aside for later use; (2) The fermentation product filtrate of medicinal fumonis from step (1), hydrolyzed sodium hyaluronate, collagen, mannitol and trehalose were mixed and homogenized three times under high pressure at 150 MPa to obtain mixture A1. (3) Add nicotinamide adenine dinucleotide to mixture A1 from step (1) and stir at 1000 rpm for 20 min to obtain the scalp anti-aging composition.

[0036] The preparation of medicinal Fermentation Product Filtrate from Lactobacillus fermentation includes the following steps: S1. Inoculate Lactobacillus fermentation colonies into MRS medium and culture at 40℃ and 80rpm for 40h to obtain a bacterial suspension. S2. Inoculate the bacterial suspension into MRS medium at an inoculation rate of 7% (v / v) and culture for 54 hours at 40℃ and 80 rpm to obtain the seed culture. S3. Inoculate the seed culture at a rate of 3% (v / v) into a fermentation medium containing 35 g / L of medicinal fumonis powder, and ferment at 40°C, 80 rpm and sterile air flow rate of 0.6 vvm for 30 h to obtain crude fermentation broth. S4. Sterilize the crude fermentation broth at 90℃ for 20 minutes, and filter it through a diatom filter to obtain the fermentation product filtrate of medicinal fumonis.

[0037] The fermentation medium containing medicinal fumonis is formulated as follows: 35 g / L medicinal fumonis powder, 10 g / L peptone, 8 g / L beef extract powder, 12 g / L yeast powder, 15 g / L fructose syrup, 0.2 g / L magnesium sulfate, 0.04 g / L manganese sulfate, 2 g / L dipotassium hydrogen phosphate, 0.8 g / L ammonium dihydrogen phosphate, and the balance being water.

[0038] Comparative Example 1 Compared with Example 1, the difference is that the medicinal Fermentation Product Filtrate, Mannitol, and Trehalose are removed, and only the basic matrix components in the formula are retained. Otherwise, it is the same as Example 1.

[0039] Comparative Example 2 Compared with Example 1, the difference is that the filtrate of the medicinal Fermentation Product of Polyporus medicata is removed, while the other components and amounts remain unchanged, and everything else is the same as in Example 1.

[0040] Comparative Example 3 Compared with Example 1, the difference is that mannitol is removed, while the other components and their amounts remain unchanged, and everything else is the same as in Example 1.

[0041] Comparative Example 4 Compared with Example 1, the difference is that trehalose is removed, while the other ingredients and amounts remain unchanged, and everything else is the same as in Example 1.

[0042] Comparative Example 5 Compared with Example 1, the difference is that the amount of medicinal Fermentation Product Filtrate added is 15 parts, and the rest is the same as in Example 1.

[0043] Comparative Example 6 Compared with Example 1, the difference is that the amount of medicinal Fermentation Product Filtrate added is 45 parts, and the rest is the same as in Example 1.

[0044] Comparative Example 7 Compared with Example 1, the difference is that the amount of mannitol added is 8 parts, and the rest is the same as in Example 1.

[0045] Comparative Example 8 Compared with Example 1, the difference is that the amount of mannitol added is 50 parts, and the rest is the same as in Example 1.

[0046] Comparative Example 9 Compared with Example 1, the difference is that the amount of trehalose added is 4 parts, and the rest is the same as in Example 1.

[0047] Comparative Example 10 Compared with Example 1, the difference is that the amount of trehalose added is 30 parts, and everything else is the same as in Example 1.

[0048] Comparative Example 11 Compared with Example 1, the difference is that mannitol is replaced with an equal mass of glycerol, otherwise the same as Example 1.

[0049] Comparative Example 12 Compared with Example 1, the difference is that trehalose is replaced with an equal mass of maltose, otherwise the same as Example 1.

[0050] Comparative Example 13 Compared with Example 1, the difference is that the filtrate of the medicinal Fermentation Product of Polyporus flocculation was replaced with that prepared by fermentation of Lactobacillus plantarum (ATCC 8014), otherwise it is the same as Example 1.

[0051] The preparation method of the fermentation product filtrate of medicinal fibroblasts prepared by fermentation of *Lactobacillus plantarum* (ATCC 8014) is the same as the preparation method of the fermentation product filtrate of medicinal fibroblasts prepared by fermentation of *Lactobacillus plantarum* in Example 1.

[0052] Comparative Example 14 Compared with Example 1, the difference is that the filtrate of the medicinal Fermentation Product of Polyporus flocculation was replaced with that prepared by fermentation of Lactobacillus rhamnosus (BNCC 187896), otherwise it is the same as Example 1.

[0053] The preparation method of the fermentation product filtrate of medicinal fibroblasts prepared by fermentation of Lactobacillus rhamnosus (BNCC 187896) is the same as the preparation method of the fermentation product filtrate of medicinal fibroblasts prepared by fermentation of Lactobacillus rhamnosus in Example 1.

[0054] Experiment 1 Functional Verification Experiment (1) Detection experiment of dermal papilla cell proliferation and viability Test Principle: Hair follicle stem cells are the source of hair follicle regeneration and the hair cycle; their functional decline is one of the root causes of scalp aging and hair loss. Dermal dermal papilla cells are the "brain" that directs hair follicle growth. Detecting cell proliferation activity is the primary step in evaluating whether a test substance can revitalize basal hair follicle cells and promote hair growth.

[0055] Test samples: The scalp anti-aging compositions of Examples 1-10 and Comparative Examples 1-14 were diluted with DMEM medium to a mass concentration of 0.5%.

[0056] The dermal dermal papilla cells used in the experiment were obtained from Suzhou Haixing Biotechnology Co., Ltd. Testing instruments and reagents: the microplate reader was a Bio-Rad iMark absorbance microplate reader, model 1681130; the CCK-8 reagent kit was obtained from Beyotime Biotechnology Co., Ltd.

[0057] Test Procedure: Dermal papilla cells in the logarithmic growth phase were seeded into 96-well cell culture plates at a density of 100 μL per well, with 10,000 cells per well. The cells were incubated at 37°C with 5% CO2 for 24 h. After incubation, the original culture medium was discarded, and the cells were divided into two groups: the test group received 100 μL of test sample per well, and the negative control group received 100 μL of DMEM medium per well. After incubation at 37°C with 5% CO2 for 24 h, the sample solution was discarded, and 100 μL of 10% CCK-8 solution was added to each well under light-protected conditions for 2 h. The absorbance was measured at 410 nm using a microplate reader. The relative cell viability (Viability) of each group was calculated, with the negative control group's cell viability set at 100%.

[0058]

[0059] OD TA -OD Blank This refers to the actual absorbance of formazan produced by the test group cells after background subtraction.

[0060] OD NC -OD Blank This refers to the actual absorbance of formazan produced by the negative control group cells after background subtraction.

[0061] OD Blank The absorbance values ​​are for blank wells containing only DMEM medium and CCK-8 solution.

[0062] The test results are shown in Table 2 below.

[0063] (2) Inhibition of inflammatory factor TNF-α experiment Test Principle: Scalp microinflammation is a key factor accelerating aging and leading to hair follicle atrophy. This experiment establishes an inflammatory cell model (stimulating dermal papilla cells with lipopolysaccharide LPS) to detect the inhibitory effect of the test substance on the expression of the key inflammatory factor TNF-α, thereby evaluating its potential to soothe scalp inflammation.

[0064] Test samples: The scalp anti-aging compositions of Examples 1-10 and Comparative Examples 1-14 were diluted with DMEM medium to a mass concentration of 1%; the dermal papilla cells used in the experiment were obtained from Suzhou Haixing Biotechnology Co., Ltd.; the lipopolysaccharide (LPS) was obtained from Sigma-Aldrich, USA; the real-time fluorescence PCR instrument was the CFX Opus 96 real-time fluorescence quantitative PCR system.

[0065] Test procedure: Dermal dermal papilla cells in the logarithmic growth phase were seeded into 6-well cell culture plates, 2 × 10⁶ cells per well. 5Cells were cultured in DMEM medium (hereinafter referred to as culture medium) at 37℃ for 24 h, and the original culture medium was discarded after the culture. 1 mL of culture medium was added to the negative control group, and 1 mL of 5 μg / mL LPS solution was added to the model control group and the test sample group. Cells were incubated at 37℃ for 18 h to induce cell modeling, and the culture medium was discarded afterward. 1 mL of culture medium was added to the negative control group and the model control group, and 1 mL of the corresponding sample solution was added to the test sample group. Cells were cultured for another 24 h. After the culture, the expression level of TNF-α was determined by quantitative real-time PCR performed by Sangon Biotech (Shanghai) Co., Ltd.

[0066] The test results are shown in Table 2 below.

[0067] (3) Test for inhibiting MMP-1 expression in scalp fibroblasts Test Principle: MMP-1 (matrix metalloproteinase-1) is a key enzyme in the degradation of type I, II, and III collagen. Its overexpression damages the extracellular matrix (ECM), leading to skin / scalp aging and hair follicle atrophy. Cells in the scalp and hair follicles are one of the main sources of MMP-1. Scalp fibroblasts are the main cells in the dermis of the scalp, responsible for synthesizing and secreting collagen. External factors such as ultraviolet radiation and pollution can induce scalp fibroblasts to express MMP-1, leading to scalp laxity and wrinkles.

[0068] Test samples: The scalp anti-aging compositions of Examples 1-10 and Comparative Examples 1-14 were diluted with DMEM medium to a mass concentration of 1%; the scalp fibroblasts used in the experiment were obtained from Suzhou Haixing Biotechnology Co., Ltd.

[0069] The ultraviolet irradiator was sourced from Shanghai Precision Instruments Co., Ltd., model ZDZ-1; the real-time fluorescence PCR instrument was the CFX Opus 96 real-time fluorescence PCR system.

[0070] Test procedure: Scalp fibroblasts in the logarithmic growth phase were seeded into 6-well cell culture plates, 2 × 10⁶ cells per well. 5 Cells were added to DMEM medium (hereinafter referred to as culture medium) and cultured in a 37℃, 5% CO2 incubator for 24 h. After culture, the original culture medium was discarded. 1 mL of culture medium was added to the negative control group and the model control group, and 1 mL of test sample solution was added to the test sample group. After culturing for another 24 h, the cells were analyzed using 30 mJ / cm²... 2Scalp fibroblasts were irradiated with UVB for 1 hour (the negative control group did not receive UVB irradiation). After irradiation, the corresponding culture medium or sample solution was added, and the cells were cultured at 37°C in a 5% CO2 incubator for 24 hours. After culture, the expression level of the collagen-related gene MMP-1 was detected by qRT-PCR technology at Sangon Biotech (Shanghai) Co., Ltd. Higher MMP-1 mRNA expression levels indicate more MMP-1, making collagen and elastin fibers in the skin more susceptible to damage, and increasing the likelihood of wrinkles and aging on the scalp.

[0071] The test results are shown in Table 2 below.

[0072] (4) Experimental results and analysis are shown in the table below: Table 2. Results of Functional Verification Experiments for the Invention

[0073] As shown in Table 2 above, compared with Example 1, Comparative Example 1 did not add the fermentation product filtrate of *Fomitopsis medicata*, mannitol, and trehalose (only the basic matrix was retained); Comparative Example 2 did not add the fermentation product filtrate of *Fomitopsis medicata*; Comparative Example 3 did not add mannitol; Comparative Example 4 did not add trehalose; Comparative Examples 5-6 adjusted the amount of fermentation product filtrate of *Fomitopsis medicata*; Comparative Examples 7-8 adjusted the amount of mannitol; Comparative Examples 9-10 adjusted the amount of trehalose; Comparative Example 11 replaced mannitol with an equal mass of glycerol; Comparative Example 12 replaced trehalose with an equal mass of maltose; and Comparative Examples 13-14 used *Lactobacillus plantarum* (ATCC 8014) and *Lactobacillus rhamnosus* (BNCC 187896) to prepare the fermentation product filtrate of *Fomitopsis medicata* (fermentation method was the same as in Example 1). The comparative examples 1-14 all showed significantly worse anti-aging indicators than Examples 1-10 due to the absence of one or more core components, deviations from the optimal range of core component dosage, or substitution of core components / fermentation strains. This indicates that the medicinal Fermentation Product Filtrate, mannitol, and trehalose of the present invention are the core key components for achieving excellent anti-aging effects in the scalp anti-aging composition, and all three must be controlled within a specific optimal dosage range. Furthermore, the medicinal Fermentation Product Filtrate must be prepared by exclusive fermentation with Lactobacillus fermentum, and mannitol and trehalose cannot be replaced by conventional ingredients such as glycerol and maltose. These three components, along with nicotinamide adenine dinucleotide, hydrolyzed sodium hyaluronate, and collagen, form an irreplaceable synergistic system; deviation from any factor will disrupt the synergistic effect. Secondly, the experimental results above show that the scalp anti-aging compositions of Examples 1-10 of the present invention have higher cell activity values ​​and lower relative levels of TNF-α mRNA and MMP-1 mRNA. This means that the scalp anti-aging compositions of the present invention can better promote the proliferation of dermal papilla cells, reduce the production of the inflammatory factor TNF-α in dermal papilla cells, and reduce the production of matrix metalloproteinases in scalp fibroblasts. This indicates that the scalp anti-aging compositions of the present invention have superior scalp anti-aging effects.

[0074] In summary, the scalp anti-aging composition of the present invention achieves optimal anti-aging efficacy through the specific ratio and synergistic effect of nicotinamide adenine dinucleotide, the fermentation product filtrate of medicinal fumonis prepared by fermentation of Lactobacillus fermentum, hydrolyzed sodium hyaluronate, collagen, mannitol, and trehalose. It can effectively promote the proliferation of dermal papilla cells and significantly inhibit the expression of inflammatory factors TNF-α and matrix metalloproteinase MMP-1. However, the absence of core components, the replacement of key components, or the exceeding of reasonable ratios will lead to a decrease in anti-aging efficacy.

[0075] Application Examples 1-3: Preparation of the Anti-aging Scalp Essence of the Present Invention The formula for preparing the scalp anti-aging essence of this invention is shown in Table 3 below: Table 3. Formula of the anti-aging scalp serum of the present invention

[0076] The preparation method of the scalp anti-aging serum in Application Example 1 includes the following steps: S1. Weigh out water, glycerin and 1,3-butanediol according to the formula, stir them evenly and heat them to 80°C. Keep stirring and maintaining the temperature to obtain mixture B1. S2. After the mixture B1 from step S1 has cooled to 40°C, add the scalp anti-aging composition, small molecule sodium hyaluronate, erythritol, Active Moist 24 polyamino acid polysaccharide, lactobacillus fermentation product, phenoxyethanol, 1,2-hexanediol and Microcare Emollient EHG to it according to the formula amount, and stir evenly to obtain mixture B2. S3. Add the formula amount DN-5 to the mixture B2 in step S2, mix evenly, and obtain mixture B3; S4. Add the prescribed amount of citric acid to mixture B3 from step S3, stir well, and obtain the scalp anti-aging essence.

[0077] The preparation method of the scalp anti-aging serum in Application Example 2 includes the following steps: S1. Weigh out water, glycerin and 1,3-butanediol according to the formula, stir them evenly and heat them to 83°C. Maintain this temperature while stirring to obtain mixture B1. S2. After the mixture B1 from step S1 has cooled to 43°C, add the scalp anti-aging composition, small molecule sodium hyaluronate, erythritol, Active Moist 24 polyamino acid polysaccharide, lactobacillus fermentation product, phenoxyethanol, 1,2-hexanediol and Microcare Emollient EHG to it according to the formula amount, and stir evenly to obtain mixture B2. S3. Add the formula amount DN-5 to the mixture B2 in step S2, mix evenly, and obtain mixture B3; S4. Add the prescribed amount of citric acid to mixture B3 from step S3, stir well, and obtain the scalp anti-aging essence.

[0078] The preparation method of the scalp anti-aging serum in Application Example 3 includes the following steps: S1. Weigh out water, glycerin and 1,3-butanediol according to the formula, stir them evenly and heat them to 85°C. Keep stirring and maintaining the temperature to obtain mixture B1. S2. After the mixture B1 from step S1 has cooled to 45°C, add the scalp anti-aging composition, small molecule sodium hyaluronate, erythritol, Active Moist 24 polyamino acid polysaccharide, lactobacillus fermentation product, phenoxyethanol, 1,2-hexanediol and Microcare Emollient EHG to it according to the formula amount, and stir evenly to obtain mixture B2. S3. Add the formula amount DN-5 to the mixture B2 in step S2, mix evenly, and obtain mixture B3; S4. Add the prescribed amount of citric acid to mixture B3 from step S3, stir well, and obtain the scalp anti-aging essence.

[0079] Experiment 2: Evaluation of Human Use Test samples: The serums from Application Examples 1-3, and the base serum (prepared in a similar manner to Application Example 2, except that the scalp anti-aging composition in Application Example 2 is replaced with an equal percentage of deionized water by mass). Test Procedure: Forty volunteers aged 20-50 years with symptoms of itchy, dry, and red scalp were selected (one test sample was used for every 10 volunteers). Each volunteer applied 2mL of the serum to their scalp twice daily, once in the morning and once in the evening, for two consecutive weeks. After using the sample, volunteers evaluated its user experience and efficacy. The rating scale was 0 for no effect and 10 for excellent effect. Ratings could be any value between 0 and 10. The test result was the average of the 10 volunteers.

[0080] The test results are shown in Table 4 below.

[0081] Table 4 Test results of human efficacy

[0082] As shown in Table 4 above, compared with the base essence, the essence of the scalp anti-aging composition of the present invention in Examples 1-3 is more effective in relieving scalp itching, redness, and sensitivity. At the same time, volunteers who used the essence of the application examples of the present invention reported that their scalps were fully nourished, and the originally dry and rough scalp condition was significantly improved, presenting a moist and delicate texture. The overall scalp firmness and luster were improved, further confirming that the scalp anti-aging composition of the present invention achieves the dual effects of scalp anti-aging and nourishing repair by inhibiting the overexpression of MMP-1, protecting scalp collagen fibers, and soothing inflammatory responses.

[0083] In summary, this invention addresses the shortcomings of existing scalp anti-aging products, such as limited functionality, narrow target range, low penetration of active ingredients, and poor synergy. It provides a scientifically formulated scalp anti-aging composition with a clear mechanism of action, along with its preparation method and applications. This composition uses nicotinamide adenine dinucleotide and the fermentation product filtrate of medicinal *Polyporus thuringiensis* prepared by *Lactobacillus fermentation* as its core components, combined with hydrolyzed sodium hyaluronate, recombinant type III collagen, mannitol, and trehalose. Through the synergistic effect of these components, it achieves multi-dimensional scalp anti-aging effects—efficiently promoting the proliferation of dermal papilla cells to provide impetus for hair follicle growth; significantly inhibiting the expression of the inflammatory factor TNF-α gene to soothe scalp inflammation; and effectively inhibiting the expression of the matrix metalloproteinase-1 gene in fibroblasts to protect scalp collagen fibers and delay scalp structural aging. In terms of preparation process, this invention promotes the formation of microsphere molecular aggregates by high pressure homogenization technology, which not only solves the problem of insufficient permeability of traditional collagen components, but also achieves orderly sustained release of active substances, prolongs the duration of efficacy, and the process is mild, cost controllable, and easy to industrialize and promote.

[0084] Secondly, this composition has a wide range of applications and can be added to various hair care products such as scalp care essences, shampoos, and hair masks. Only 0.1-5 wt% is needed to achieve excellent results. Therefore, this invention, through synergistic innovation of ingredients and process optimization, breaks through the bottlenecks of existing technologies. The scalp anti-aging composition provided is safe, effective, and practical, offering a new technical solution for the research and development of daily anti-aging scalp chemical products, and has broad market application prospects and industrialization value.

[0085] The above embodiments are merely illustrative of the principles and effects of the present invention and are not intended to limit the invention. Any person skilled in the art can modify or alter the above embodiments without departing from the spirit and scope of the present invention. Therefore, all equivalent modifications or alterations made by those skilled in the art without departing from the spirit and technical concept disclosed in the present invention should still be covered by the claims of the present invention.

Claims

1. A scalp anti-aging composition characterized in that, By mass parts, comprising: 20~40 parts of nicotinamide adenine dinucleotide, 20~40 parts of medicinal Fomes fomentarius fermentation product filtrate, 0.8~3.0 parts of hydrolyzed sodium hyaluronate, 2~8 parts of collagen, 12~40 parts of mannitol and 8~25 parts of trehalose.

2. The scalp anti-aging composition according to claim 1, characterized by, The medicinal Fomes fomentarius fermentation product filtrate is prepared by fermentation of Lactobacillus fermentum; and the collagen is recombinant type III collagen.

3. The scalp anti-aging composition according to claim 1, characterized by, The preparation method of the medicinal Fomes fomentarius fermentation product filtrate comprises: S1, inoculating Lactobacillus fermentum colonies into MRS culture medium, expanding culture at 33~40℃ and 80~120rpm for 40~54h to obtain a bacterial suspension; S2, inoculating the bacterial suspension into MRS culture medium at an inoculation amount of 3~7% (v / v), secondary culture at 33~40℃ and 80~120rpm for 40~54h to obtain a seed liquid; S3, inoculating the seed liquid into a fermentation culture medium containing medicinal Fomes fomentarius powder at an inoculation amount of 1~3% (v / v), fermentation culture at 33~40℃, 80~120rpm and a sterile air ventilation amount of 0.2~0.6vvm for 30~45h to obtain a crude fermentation liquid; S4, sterilizing the crude fermentation liquid at 80~90℃ for 20~40min, filtering to obtain the medicinal Fomes fomentarius fermentation product filtrate.

4. The scalp anti-aging composition according to claim 1, characterized by, The fermentation culture medium containing medicinal Fomes fomentarius in step S3 comprises medicinal Fomes fomentarius powder, a carbon source, a nitrogen source, inorganic salts and water; and the addition amount of the medicinal Fomes fomentarius is 10~40g / L; The nitrogen source comprises at least two of peptone, beef infusion powder and yeast powder, and the addition amount of the nitrogen source is 15~35g / L; The carbon source comprises at least one of glucose, sucrose and fructose glucose syrup, and the addition amount of the nitrogen source is 15~30g / L; The inorganic salts comprise at least three of magnesium sulfate, manganese sulfate, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, diammonium hydrogen phosphate and ammonium dihydrogen phosphate, and the addition amount of the inorganic salts is 2~8g / L.

5. A method of preparing a composition for anti-aging of the scalp according to any one of claims 1 to 4, characterized in that, The method comprises the following steps: (1) preparing the medicinal Fomes fomentarius fermentation product filtrate by fermentation of Lactobacillus fermentum for standby; (2) mixing the medicinal Fomes fomentarius fermentation product filtrate, hydrolyzed sodium hyaluronate, collagen, mannitol and trehalose in step (1), high-pressure homogenization at 100~150MPa for 3~6 times to obtain a mixture A1; (3) adding nicotinamide adenine dinucleotide to the mixture A1 in step (2), stirring at 500~1000rpm for 20~40min to obtain a scalp anti-aging composition.

6. Use of a scalp anti-aging composition according to any one of claims 1 to 4 for the preparation of a hair product, characterized in that, The product for hair use is selected from at least one of scalp care essence, scalp care water, scalp nourishing cream, scalp massage cream, hair essence, hair essential oil, shampoo, hair soap, hair powder, hair mask, hair conditioner, hair spray, hair jelly and hair emulsion.

7. Use according to claim 6, characterized in that, The addition amount of the composition in the product for hair use is 0.1~5wt%.

8. A scalp anti-aging serum, characterized by, By mass percentage, it comprises: 0.1~5% of the scalp anti-aging composition according to any one of claims 1~4, 3~10% of glycerin, 3~10% of 1,3-butanediol, 0.005~0.02% of small molecule sodium hyaluronate, 0.5~2.0% of erythritol, 0.1~0.4% of Active Moist 24 polyamino acid polysaccharide, 0.05~0.2% of lactobacillus fermentation product, 0.3~1.0% of phenoxyethanol, 0.3~1.0% of 1,2-hexanediol, 0.05~0.2% of Microcare Emollient EHG, 0.4~1.2% of DN-5, 0.05~0.2% of citric acid and the balance of water.

9. The scalp anti-aging serum of claim 8, wherein, The small molecule sodium hyaluronate is purchased from Bihonghua Biological Technology (Shandong) Group Co., Ltd., model Polyglyte®Microspheric Recombinant Collagen; the Active Moist 24 polyamino acid polysaccharide is purchased from Guangzhou Fuyulong Biological Technology Co., Ltd.; the lactobacillus fermentation product is purchased from Guangzhou Runheng Biological Technology Co., Ltd.; the Microcare Emollient EHG is purchased from Tor Special Chemicals (Zhenjiang) Co., Ltd.; and the DN-5 is purchased from Yamaguchi Chemical Industry Co., Ltd.

10. A method of preparing the scalp anti-aging serum according to claim 8 or 9, characterized in that, The method comprises the following steps: S1, water, glycerin and 1,3-butanediol are weighed according to the formula amount, stirred uniformly, heated to 80~85℃, and kept at this temperature for stirring and preservation, to obtain a mixture B1; S2, after the mixture B1 in step S1 is cooled to 40~45℃, the scalp anti-aging composition, small molecule sodium hyaluronate, erythritol, Active Moist 24 polyamino acid polysaccharide, lactobacillus fermentation product, phenoxyethanol, 1,2-hexanediol and Microcare Emollient EHG are added thereto according to the formula amount, stirred uniformly, to obtain a mixture B2; S3, DN-5 is added to the mixture B2 in step S2 according to the formula amount, mixed uniformly, to obtain a mixture B3; S4, citric acid is added to the mixture B3 in step S3 according to the formula amount, stirred uniformly, to obtain the scalp anti-aging essence.

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