External healing-promoting skin-care preparation for simulating dynamic regeneration of sebum and preparation method of external healing-promoting skin-care preparation

Abstract

The invention relates to the technical field of cosmetics, and discloses a bionic sebum dynamic regeneration external healing-promoting skin care preparation and a preparation method thereof, and the preparation comprises a bionic triglyceride substrate, stabilized lipase and a targeted synergistic red ginseng extract. The method comprises the following steps: catalyzing hydrolysis of a bionic triglyceride substrate on the surface of skin in situ by using stabilized lipase to continuously generate free fatty acid; the generated free fatty acid not only can repair the skin barrier for a long time, but also can be used as an endogenous penetration enhancer to synergistically improve the transdermal absorption of active ingredients in the target synergistic red ginseng extract and enhance the anti-inflammatory and healing-promoting effects of the target synergistic red ginseng extract. The preparation method adopts the steps of high-temperature emulsification and low-temperature active component addition, so that the activity of the thermosensitive component and the product stability are ensured. By constructing a skin in-situ biological reaction system, the technical problems that a traditional preparation is short in repairing effect and low in active matter absorption efficiency are solved.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of cosmetic technology, in particular to a bionic sebum dynamic regeneration external use healing-promoting skin care preparation and a preparation method thereof. BACKGROUND

[0002] Skin is the first line of defense of the human body, and the outermost stratum corneum is the core structure for realizing the barrier function. A healthy stratum corneum can effectively prevent water loss from the body and resist external physical, chemical and biological damage. When the skin barrier function is damaged, it will lead to increased transdermal water loss, dry and sensitive skin, and easily cause inflammation and a series of problems. Therefore, repairing the damaged skin barrier is an important direction in the field of skin care.

[0003] At present, the external use preparation for repairing the skin barrier usually directly adds ingredients simulating the intercellular lipids of the stratum corneum cells, such as ceramides, cholesterol and free fatty acids, to the product, in order to fill the structural defects of the barrier by external supplementation. Although this method can have an immediate occlusive and moisturizing effect, its effect is often temporary. Once the product is removed from the skin surface, the externally supplemented lipids decrease, and the skin barrier returns to the damaged state, making it difficult to achieve long-term self-repair.

[0004] In addition, in order to enhance the efficacy of the preparation, some active ingredients such as plant extracts are often added. However, the dense structure of the stratum corneum also constitutes a major obstacle to the transdermal absorption of these active ingredients, resulting in low bioavailability and limited actual effect. In order to solve this problem, the prior art often uses the addition of chemical penetration enhancers (such as alcohols, azone, etc.) to improve the penetration of active substances. However, such chemical penetration enhancers may irritate the already damaged skin, or further interfere with the normal structure of the barrier, and their application has certain limitations, especially for sensitive or damaged skin. Therefore, how to achieve efficient transdermal delivery of active ingredients without introducing additional irritation, while achieving long-term barrier repair, is a technical problem to be solved in the field. SUMMARY

[0005] The technical problem solved by the present application is that the existing external use skin care preparations usually rely on physical occlusion or one-time supplementation of lipids for repairing the skin barrier, and the effect is short-acting. At the same time, the transdermal delivery efficiency of active substances is limited, resulting in low bioavailability of active substances at the target site and difficulty in fully exerting their physiological efficacy.

[0006] To solve the above problems, the present application provides the following technical solutions: In a first aspect, the present application provides a bionic sebum dynamic regeneration external use healing-promoting skin care preparation, which comprises the following components by weight percentage: Biomimetic triglyceride substrate: 3.0%–5.0%; Stabilized lipase: 0.1%–0.4%; Targeted synergistic red ginseng extract: 0.3%–0.8%; Acceptable carrier for cosmetics: excess.

[0007] By employing the above technical solution, this invention constructs a bioreaction system in the formulation that can be initiated in situ on the skin surface. This system utilizes a stabilized lipase and a biomimetic triglyceride substrate within the formulation. During product storage, the components remain stable in a low-water-activity matrix. When the formulation is applied to the skin surface, the skin's temperature and trace amounts of moisture activate the lipase, initiating the reaction. The specific working mechanism of this reaction system includes: 1. In-situ enzymatic hydrolysis: The activated and stabilized lipase catalyzes the hydrolysis of the biomimetic triglyceride substrate, continuously and slowly generating free fatty acids and glycerol.

[0008] 2. Long-term repair of the skin barrier: Free fatty acids generated in situ are key components of the intercellular lipids of the stratum corneum. This dynamic generation mechanism can continuously replenish the lipids needed by the skin barrier, achieving long-term repair and maintenance of the damaged barrier. Its mode of action differs from simple physical closure or one-time addition of end products.

[0009] 3. Endogenous Permeability Enhancement: The free fatty acids (such as oleic acid) generated during the reaction act as endogenous permeability enhancers, reversibly altering the ordered structure of the stratum corneum lipid bilayer and increasing its fluidity. This reduces the resistance to transdermal absorption of active ingredients in targeted synergistic red ginseng extract, improving its permeation efficiency. Compared to a one-time addition of permeability enhancers, this dynamic generation mechanism allows the permeation-enhancing microenvironment to be maintained for a longer period, thus achieving a sustained permeation effect.

[0010] 4. Enhanced bioavailability and efficacy: The above mechanism enhances the transdermal penetration of active ingredients in targeted synergistic red ginseng extract, enabling them to reach higher effective concentrations at the site of action, thereby enhancing their biological effects, such as exhibiting stronger anti-inflammatory activity.

[0011] This invention achieves skin barrier repair, active ingredient penetration enhancement, and efficacy enhancement simultaneously through a unified enzymatic reaction system. The synergistic effect of each step improves the overall performance of the formulation.

[0012] Preferably, the biomimetic triglyceride substrate is composed of high-oleic sunflower seed oil and palmitic triglyceride, wherein the mass ratio of high-oleic sunflower seed oil to palmitic triglyceride is (70-90):(10-30).

[0013] By employing the above technical solution, the selected raw material ratio ensures that the composition of the free fatty acids generated after enzymatic hydrolysis closely resembles the fatty acid profile of healthy human sebum. This component design enables the generated lipids to have good affinity with the skin, thereby effectively participating in the reconstruction of the skin barrier.

[0014] Preferably, the preparation method of the targeted synergistic red ginseng extract includes: enzymatic conversion treatment of crude red ginseng extract with β-glucosidase to enrich ginsenoside Rg3.

[0015] By employing the above-mentioned technical solution, through a pre-conversion step, the original ginsenosides, which are abundant but have weak activity and large molecular weight in red ginseng, are converted into rare ginsenoside Rg3, which has a smaller molecular weight, stronger lipid solubility, and higher biological activity. This step enhances the efficacy of the core active ingredient, allowing it to further enhance the final physiological effects when combined with the subsequent delivery system.

[0016] Preferably, the stabilized lipase is a triacylglycerol lipase derived from Aspergillus oryzae, which is stabilized using maltodextrin as a carrier.

[0017] By employing the above technical solution and using stabilized lipase, the lipase maintains stable activity throughout the shelf life of the formulation, preventing premature reaction with the substrate. Simultaneously, the maltodextrin carrier dissolves rapidly upon contact with skin moisture, allowing the lipase to be released and activated, ensuring immediate initiation and controllability of the reaction system.

[0018] Preferably, the cosmetically acceptable carrier comprises an oil phase component, an aqueous phase component, and an emulsifier, and the formulation is an oil-in-water emulsion.

[0019] By adopting the above technical solution, this dosage form can stably disperse oil-soluble biomimetic triglyceride substrates and water-soluble enzymes, red ginseng extracts and other components in a homogeneous system, ensuring that the components can effectively contact and act during application.

[0020] Secondly, the present invention provides a method for preparing the aforementioned biomimetic sebum dynamic regeneration topical skin care preparation, the method comprising the following steps: (a) The oil phase component and the aqueous phase component are emulsified under heating conditions to form an emulsion; (b) Cool the emulsion obtained in step (a) to below 40°C; (c) At a temperature below 40°C, add the biomimetic triglyceride substrate, stabilized lipase and targeted synergistic red ginseng extract to the emulsion and mix well to obtain the final product.

[0021] By employing the above technical solution, the preparation method utilizes a process of high-temperature emulsification followed by low-temperature addition of active components. Step (a) is carried out at a relatively high temperature to ensure that all components in the oil phase are completely melted and form a stable emulsion matrix. Step (c) is performed after the system has been sufficiently cooled. Because bioactive components such as stabilized lipases are heat-sensitive, their structures undergo irreversible changes and become inactive at high temperatures. Placing the addition of these active components at a temperature below 40°C effectively maintains their bioactivity and structural integrity, enabling the final product to achieve the predetermined in-situ enzymatic reaction.

[0022] Preferably, the method further includes a pretreatment step for preparing a targeted synergistic red ginseng extract: take crude red ginseng extract, add β-glucosidase and react at pH 5.0 and 55℃ for 36 hours, then purify and dry to obtain the targeted synergistic red ginseng extract.

[0023] By adopting the above technical solution, the active ingredients are processed at the initial stage of the overall preparation process. This pretreatment step results in a higher concentration and efficacy of active ingredients in the red ginseng extract ultimately added to the emulsion system, providing a foundation for achieving the overall efficacy of the formulation.

[0024] Preferably, in step (a), the emulsification temperature is 75°C to 85°C.

[0025] By adopting the above technical solution, this temperature range can ensure that high-melting-point components such as cetearyl alcohol in the oil phase are completely melted, reducing the overall viscosity of the oil phase and facilitating the formation of fine and uniformly distributed microdroplets under homogeneous shearing, thereby obtaining an emulsion matrix with good physical stability.

[0026] Preferably, in step (c), the cooling temperature is 38°C to 40°C; and the addition of the component is carried out under vacuum conditions of -0.06 MPa to -0.08 MPa.

[0027] By adopting the above technical solution, the temperature for adding active ingredients is controlled at 38℃~40℃, which effectively maintains enzyme activity and ensures the paste has suitable fluidity for uniform mixing. Simultaneously, adding and stirring under vacuum conditions removes air trapped in the system, preventing air bubbles from forming in the finished formulation. Furthermore, this operation reduces the oxygen content in the system, protecting easily oxidized lipases and active ingredients in red ginseng extract, thus improving the product's storage stability.

[0028] Preferably, step (c) is implemented by first adding and mixing a biomimetic triglyceride substrate, and then adding a stabilized lipase and a targeted synergistic red ginseng extract.

[0029] By adopting the above technical solution, this order of addition facilitates the uniform dispersion of each component in the system. The biomimetic triglyceride substrate is an oil-soluble component; adding it first to the oil-in-water emulsion helps it disperse uniformly in the oil phase. Adding stabilizing lipase and red ginseng extract further ensures a uniform distribution of the enzyme, substrate, and active ingredient within the system, preventing instability or uneven reactions that might result from localized accumulation, and ensuring consistent in-situ reaction conditions when the product is applied to the skin.

[0030] In summary, the present invention has at least one of the following beneficial technical effects: 1. This invention constructs a system capable of in-situ reaction on the skin surface by introducing stabilized lipase and biomimetic triglyceride substrates into the formulation. This system responds to the skin's microenvironment, continuously hydrolyzing the substrate to generate free fatty acids, thereby dynamically replenishing the core lipid components required for the skin barrier. Compared to traditional methods of applying lipids only once, this invention provides long-lasting barrier repair and maintenance effects, and can more persistently improve the skin's water retention capacity and resistance to external stimuli.

[0031] 2. This invention combines an in-situ enzymatic reaction system with a targeted, synergistic red ginseng extract, resulting in a synergistic effect. The free fatty acids (such as oleic acid) generated by enzymatic hydrolysis act as endogenous penetration enhancers, improving the transdermal absorption efficiency of active ingredients (such as ginsenoside Rg3) in the targeted, synergistic red ginseng extract. This enhances the bioavailability of the active ingredients at the target site on the skin, allowing their anti-inflammatory and other physiological activities to be more fully realized.

[0032] 3. The preparation method employed in this invention, particularly the addition of heat-sensitive active ingredients such as stabilized lipases under low-temperature conditions, ensures the activity and stability of these components during product storage, preventing premature reactions in the system. This guarantees the product's shelf-life stability and achieves controlled release of the active ingredients only upon application to the skin. The reliable process design ensures the reproducibility of product efficacy. Detailed Implementation

[0033] To further clarify the purpose, technical solution, and advantages of this invention, the following detailed description is provided in conjunction with specific embodiments. It should be noted that the following embodiments are merely preferred examples of this invention and are not intended to limit the scope of the invention. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of this invention should be included within the protection scope of this invention.

[0034] The main raw materials and reagents used in the following examples and comparative examples have the following sources and specifications. Unless otherwise specified, all reagents are commercially available cosmetic grade, analytical grade or higher products.

[0035] High oleic sunflower seed oil, with oleic acid (C18:1) content accounting for more than 80% of the total fatty acid content, CAS No.: 8001-21-6.

[0036] Polydimethylsiloxane, with a kinematic viscosity of 250–450 mm. 2 / s (25℃), CAS No.: 63148-62-9.

[0037] β-glucosidase, derived from Aspergillus niger, EC number: 3.2.1.21, with an enzyme activity specification of 10000 U / g.

[0038] The stabilized lipase, a triacylglycerol lipase, is derived from Aspergillus oryzae (EC No. 3.1.1.3). This enzyme was stabilized using maltodextrin as a carrier and then freeze-dried, achieving an enzyme activity of 20,000 U / g.

[0039] Preparation Example 1: Preparation of Biomimetic Triglyceride Substrate Preparation Example 1-1: This preparation example provides a method for preparing biomimetic triglyceride substrate A, including the following steps: Weigh 800g of high-oleic sunflower seed oil and 200g of palmitic acid triglyceride (mass ratio 80:20), and put them into a reaction vessel equipped with a stirrer; turn on the stirrer, heat to 50-60℃, and keep stirring until the palmitic acid triglyceride is completely dissolved and the system is a clear and homogeneous liquid; slowly cool to room temperature to obtain the final product.

[0040] Preparation Examples 1-2: This preparation example provides a method for preparing biomimetic triglyceride substrate B, including the following steps: Weigh 900g of high-oleic sunflower seed oil and 100g of palmitic acid triglyceride (mass ratio 90:10), and put them into a reaction vessel equipped with a stirrer; turn on the stirrer, heat to 50-60℃, and keep stirring until the palmitic acid triglyceride is completely dissolved and the system is a clear and homogeneous liquid; slowly cool to room temperature to obtain the final product.

[0041] Preparation Examples 1-3: This preparation example provides a method for preparing biomimetic triglyceride substrate C, including the following steps: Weigh 700g of high oleic sunflower seed oil and 300g of palmitic acid triglyceride (mass ratio 70:30), and put them into a reaction vessel equipped with a stirrer; turn on the stirrer, heat to 50-60℃, and keep stirring until the palmitic acid triglyceride is completely dissolved and the system is a clear and homogeneous liquid; slowly cool to room temperature to obtain the final product.

[0042] Preparation Example 2: Preparation of Targeted Synergistic Red Ginseng Extract This preparation example provides a method for preparing a targeted synergistic red ginseng extract, including the following steps: (1) Extraction: Take 1 kg of dried red ginseng root powder, add 10 L of 70% (v / v) ethanol solution, and reflux extract twice at 80℃ for 2 hours each time. Combine the two extracts, filter, and concentrate the filtrate under reduced pressure at 60℃ and -0.08 MPa until there is no alcohol odor to obtain crude red ginseng extract; (2) Enzymatic conversion: The crude extract of red ginseng obtained in step (1) was dissolved in pure water to prepare a solution with a concentration of 15 mg / mL. The pH was adjusted to 5.0 using citrate-phosphate buffer. 2000 U of β-glucosidase was added per gram of extract, and the mixture was reacted in a constant temperature water bath at 55°C for 36 hours. (3) Termination and purification: Heat the reaction solution from step (2) to 100°C and maintain for 15 minutes to completely inactivate β-glucosidase. After cooling, load the reaction solution onto a pretreated AB-8 macroporous adsorption resin column. First, elute with 5 column volumes of deionized water to remove sugars and polar impurities; then elute with 8 column volumes of 45% (v / v) ethanol solution and collect this eluent. (4) Drying: The eluent collected in step (3) is concentrated under reduced pressure at 60℃ and -0.08MPa, and then freeze-dried to obtain a brownish-yellow powder of targeted synergistic red ginseng extract.

[0043] Example: Preparation of skin care formulations Example 1: This embodiment provides a method for preparing a biomimetic sebum dynamic regeneration topical skin care preparation, including the following steps: (1) Preparation of Phase A (oil phase): Weigh the following oil phase components by weight percentage: mineral oil 8.0%, lanolin 5.0%, cetearyl alcohol 4.0%, glyceryl stearate 2.0%, potassium lauryl phosphate 1.0%, and silicone oil 1.5%. Put the above components into the main emulsifying pot, heat to 80°C, and stir at 40-60 RPM until completely melted and mixed evenly. Keep warm for later use.

[0044] (2) Preparation of Phase B (Aqueous Phase): Weigh the following aqueous phase components by weight percentage: glycerol 10.0%, 1,2-hexanediol 2.0%, citric acid and sodium citrate (total) 0.3%, hydroxyacetophenone 0.5%, phenoxyethanol 0.8%, and the remainder is deionized water. Add the above components to the aqueous phase pot, heat to 80°C, stir until completely dissolved, and keep warm for later use.

[0045] (3) Emulsification: Turn on the high-speed homogenizer of the main emulsification pot and set the speed to 3500 RPM. Slowly pump the B phase from step (2) into the A phase from step (1) and homogenize for 5 minutes to form a homogeneous emulsion.

[0046] (4) Cooling: Stop homogenization, turn on the jacket cooling water, and cool the emulsion by stirring at a slow speed of 20-40 RPM until the temperature of the liquid drops to 40℃.

[0047] (5) Addition of active ingredients: Add 4.0% of the biomimetic triglyceride substrate A obtained in Preparation Example 1-1 to the paste in step (4) and stir slowly for 10 minutes. Then, under vacuum (-0.07 MPa) conditions, add 0.2% of the stabilized lipase and 0.5% of the targeted synergistic red ginseng extract obtained in Preparation Example 2 to the pot and continue stirring at 25 RPM for 20 minutes.

[0048] (6) Finished product: Finally, add 0.1% flavoring and stir well. Stop stirring, let stand to remove bubbles, and then discharge the product.

[0049] Example 2: This embodiment provides a method for preparing a biomimetic sebum dynamic regeneration topical skin care preparation, including the following steps: (1) Preparation of Phase A (oil phase): Weigh the following oil phase components by weight percentage: mineral oil 6.0%, lanolin 4.0%, cetearyl alcohol 3.0%, glyceryl stearate 2.5%, potassium lauryl phosphate 1.2%, silicone oil 2.0%. Put the above components into the main emulsifying pot, heat to 75°C, and stir at 40-60 RPM until completely melted and mixed evenly. Keep warm for later use.

[0050] (2) Preparation of Phase B (Aqueous Phase): Weigh the following aqueous phase components by weight percentage: glycerol 12.0%, 1,2-hexanediol 3.0%, citric acid and sodium citrate (total) 0.4%, hydroxyacetophenone 0.8%, phenoxyethanol 1.0%, and the remainder is deionized water. Add the above components to the aqueous phase pot, heat to 75°C, stir until completely dissolved, and keep warm for later use.

[0051] (3) Emulsification: Turn on the high-speed homogenizer of the main emulsification pot and set the speed to 2500 RPM. Slowly pump the B phase from step (2) into the A phase from step (1) and homogenize for 7 minutes to form a homogeneous emulsion.

[0052] (4) Cooling: Stop homogenization, turn on the jacket cooling water, and cool the emulsion by stirring at a slow speed of 20-40 RPM until the temperature of the liquid drops to 38°C.

[0053] (5) Addition of active ingredients: Add 5.0% of the biomimetic triglyceride substrate B obtained in Preparation Example 1-2 to the paste in step (4) and stir slowly for 10 minutes. Then, under vacuum (-0.08 MPa) conditions, add 0.4% of the stabilized lipase and 0.8% of the targeted synergistic red ginseng extract obtained in Preparation Example 2 to the pot and continue stirring at 30 RPM for 15 minutes.

[0054] (6) Finished product: Finally, add 0.1% flavoring and stir well. Stop stirring, let stand to remove bubbles, and then discharge the product.

[0055] Example 3: This embodiment provides a method for preparing a biomimetic sebum dynamic regeneration topical skin care preparation, including the following steps: (1) Preparation of Phase A (oil phase): Weigh the following oil phase components by weight percentage: mineral oil 10.0%, lanolin 6.0%, cetearyl alcohol 5.0%, glyceryl stearate 1.5%, potassium lauryl phosphate 0.8%, and silicone oil 1.0%. Put the above components into the main emulsifying pot, heat to 85°C, and stir at 40-60 RPM until completely melted and mixed evenly. Keep warm for later use.

[0056] (2) Preparation of Phase B (Aqueous Phase): Weigh the following aqueous phase components by weight percentage: glycerol 8.0%, 1,2-hexanediol 1.5%, citric acid and sodium citrate (total) 0.2%, hydroxyacetophenone 0.5%, phenoxyethanol 0.7%, and the remainder is deionized water. Add the above components to the aqueous phase pot, heat to 85°C, stir until completely dissolved, and keep warm for later use.

[0057] (3) Emulsification: Turn on the high-speed homogenizer of the main emulsification pot and set the speed to 4500 RPM. Slowly pump the B phase from step (2) into the A phase from step (1) and homogenize for 3 minutes to form a homogeneous emulsion.

[0058] (4) Cooling: Stop homogenization, turn on the jacket cooling water, and cool the emulsion by stirring at a slow speed of 20-40 RPM until the temperature of the liquid drops to 39°C.

[0059] (5) Addition of active ingredients: Add 3.0% of the biomimetic triglyceride substrate C obtained in Preparation Examples 1-3 to the paste in step (4) and stir slowly for 10 minutes. Then, under vacuum (-0.06 MPa) conditions, add 0.1% of the stabilized lipase and 0.3% of the targeted synergistic red ginseng extract obtained in Preparation Example 2 to the pot and continue stirring at 15 RPM for 25 minutes.

[0060] (6) Finished product: Finally, add 0.1% flavoring and stir well. Stop stirring, let stand to remove bubbles, and then discharge the product.

[0061] Comparative Example 1: Compared with Example 1, the difference is that no stabilizing lipase is added, and its mass is made up by deionized water. The other components and preparation methods are the same.

[0062] Comparative Example 2: Compared with Example 1, the difference is that biomimetic triglyceride substrate A is not added, and its mass is made up by deionized water. The other components and preparation methods are the same.

[0063] Comparative Example 3: Compared with Example 1, the difference is that no targeted synergistic red ginseng extract is added, and its mass is made up by deionized water. The other components and preparation methods are the same.

[0064] Comparative Example 4: Compared with Example 1, the difference is that 0.5% of the targeted synergistic red ginseng extract was replaced with an equal mass of ordinary red ginseng extract, while the other components and preparation methods are the same. Ordinary red ginseng extract refers to the product of the crude red ginseng extract after extraction in step (1) of Preparation Example 2, without enzymatic conversion in step (2), and after drying.

[0065] Comparative Example 5: Compared with Example 1, the difference is that: no stabilizing lipase and biomimetic triglyceride substrate A are added; instead, oleic acid and glycerol are added in the same molar amount as theoretically generated after complete hydrolysis of 4.0% biomimetic triglyceride substrate A in Example 1, and the amount of deionized water is adjusted accordingly to make up the total mass. The other components and preparation methods are the same.

[0066] Test Example 1: The experimental steps are as follows: (1) 1.0g of the paste from Example 1 was evenly spread on a porous ceramic carrier that had been pre-wetted with pH 5.5 buffer solution and placed in a constant temperature and humidity chamber at 32°C and 80% relative humidity for incubation.

[0067] (2) Samples were taken at 0h, 2h, 4h, 8h and 12h after incubation.

[0068] (3) Use a surface pH meter to measure the pH value of the sample surface.

[0069] (4) Extract the sample with a hexane-isopropanol mixed solvent to separate the organic phase and the aqueous phase. After methylation treatment, the organic phase was quantitatively analyzed by gas chromatography-mass spectrometry (GC-MS). The aqueous phase was quantitatively analyzed by high performance liquid chromatography (HPLC) to determine the glycerol content.

[0070] The experimental results are shown in Table 1.

[0071] Table 1. Product formation and pH change data of the sample from Example 1 in a simulated skin environment: The data in Table 1 show that the enzymatic reaction in the formulation of Example 1 can be initiated under simulated skin temperature, humidity, and acidity conditions. At 0 h, the levels of free fatty acids and glycerol in the system are at baseline, representing the background values ​​of the raw materials.

[0072] With prolonged incubation, the chemical composition of the system changed. The content of free fatty acids increased from 14.2 μg / g to 1059.7 μg / g, and the content of glycerol increased from 11.5 μg / g to 107.4 μg / g. Simultaneously, the pH value of the sample surface decreased from 6.38 to 5.51. The simultaneous and continuous formation of both free fatty acids and glycerol, and their changing trend, are consistent with the reaction mechanism of triglyceride hydrolysis catalyzed by lipase.

[0073] This result confirms from the perspective of changes in chemical composition that, under application conditions, the formulation's built-in enzymes and substrates can constitute a functional reaction system. This system continuously generates free fatty acids and glycerol by catalyzing the hydrolysis of triglycerides in situ on the skin surface. This process not only replenishes the lipid components on the skin surface but also adjusts the local pH value towards the slightly acidic range of normal skin through the accumulation of products (free fatty acids).

[0074] Test Example 2: The experimental steps are as follows: (1) Take the skin from the back of a pig outside the body, remove the subcutaneous fat and connective tissue, and fix it between the donor chamber and the receiver chamber of the Franz diffusion pool with the stratum corneum facing upward.

[0075] (2) A skin barrier damage model was constructed by applying a 5% sodium dodecyl sulfate (SLS) aqueous solution to the skin surface for 2 hours. After treatment, the skin surface was washed with phosphate buffer, and the initial TEWL value (0h) was measured and recorded using a transdermal water loss tester.

[0076] (3) Samples from Examples 1-3, Comparative Examples 1-5, and a blank control group (without any sample applied) were respectively treated with 2 mg / cm³. 2 Apply the appropriate amount evenly to the surface of the damaged skin.

[0077] (4) All Franz diffusion cells were placed in a constant temperature environment of 32°C. The TEWL value of each group of skin was measured using a transdermal moisture loss tester at 1, 4, 8, 12 and 24 hours after the sample was applied.

[0078] The experimental results are shown in Table 2.

[0079] Table 2. Transdermal water loss (TEWL) values ​​(g / h·m) for each test group at different time points. 2 ): This test case evaluates the ability of each formulation to repair the damaged skin barrier and its long-lasting effect by measuring changes in transdermal water loss (TEWL).

[0080] Table 2 shows that all samples reduced the TEWL value increased by SLS treatment in the initial stage of application (1h–4h), which is due to the occlusive physical film formed by the cream matrix itself. However, the effects of different formulations diverged over time.

[0081] The TEWL values ​​of groups 1-3 in Examples 1-3 continuously decreased or remained at a low level throughout the 24-hour testing period. At 24 hours, the measured values ​​were (8.8-9.5 g / h·m). 2 The levels were significantly lower than all comparative and blank control groups, indicating that all three formulations possess sustained barrier repair capabilities and that the effects remain stable within the claimed parameter range.

[0082] Compared to Comparative Example 1 and Comparative Example 1 (without enzyme) and Comparative Example 2 (without substrate), the TEWL values ​​of the latter two began to rebound after 4 hours. This indicates that without any key component of the enzymatic reaction system, the formulation can only provide a temporary physical blocking effect and cannot achieve long-term maintenance of barrier function. The long-lasting effect of Example 1 stems from the fact that its enzymatic reaction system continuously generates lipids such as free fatty acids during the test, supplementing the chemical structure of the barrier.

[0083] Comparative Example 1 and Comparative Example 5 (direct addition of product) showed similar results in the initial stage of testing. However, after 8 hours, the TEWL value of Comparative Example 5 began to rise, while that of Example 1 continued to decline. This indicates that the effect of adding the reaction product in one step diminishes over time; while the in-situ dynamic generation mechanism proposed in this invention can maintain the barrier function for a longer period.

[0084] Comparing Example 1 with Comparative Example 3 (without extract) and Comparative Example 4 (using common extract), there was no significant difference in TEWL values ​​among the groups. This indicates that in this test, the physical repair of the skin barrier is mainly contributed by enzymatically generated lipids, and the direct effect of red ginseng extract on this indicator is not obvious; its synergistic effect is manifested in other aspects.

[0085] In summary, the TEWL test data verified the core technical effect of the present invention: through the mechanism of in-situ and dynamic lipid generation via enzymatic reaction, it can provide a longer-lasting skin barrier repair effect than simple physical sealing or direct addition of products.

[0086] Test Example 3: The experimental steps are as follows: (1) Take skin from the back of a pig in vitro, remove subcutaneous fat and connective tissue, and fix it between the donor and receiver chambers of the Franz diffusion tank. The receiver chamber is filled with phosphate buffer containing 2% (v / v) ethanol at pH 7.4 and kept in a 32°C water bath for circulation.

[0087] (2) The samples from Example 1, Comparative Example 1, Comparative Example 4, and Comparative Example 5 were subjected to a concentration of 10 mg / cm³. 2 Apply the appropriate amount evenly to the stratum corneum of the skin.

[0088] (3) At 2, 4, 8, 12 and 24 hours after the sample is applied, all the receiving liquid is taken out from the receiving pool as the test sample, and an equal amount of fresh receiving liquid is immediately added to maintain the sedimentation conditions.

[0089] (4) The test samples at each time point were analyzed using high performance liquid chromatography (HPLC). Ginsenoside Rg3 was used as the standard, and its concentration was quantitatively detected by external standard method. The cumulative permeation per unit area was calculated based on the concentration at each time point and the volume of the receiving cell.

[0090] The experimental results are shown in Table 3.

[0091] Table 3. Cumulative permeation of ginsenoside Rg3 in each test group (μg / cm³) 2 ): This test case evaluates the effect of the formulation on promoting the penetration of ginsenoside Rg3 through an in vitro transdermal permeation experiment.

[0092] Table 3 shows that the cumulative penetration of ginsenoside Rg3 in Example 1 group was higher than that in other comparative groups at all test time points.

[0093] Compared with Comparative Example 1 (without enzyme), the cumulative penetration amount of the former over 24 hours was more than ten times that of the latter, indicating that the enzymatic reaction is the key to the penetration of active ingredients. The mechanism is that the free fatty acids generated in situ act as penetration promoters, reversibly disturbing the orderly arrangement of lipids in the stratum corneum, creating penetration channels for ginsenoside Rg3 molecules.

[0094] Comparing Example 1 and Comparative Example 5 (direct addition of product), the effects of both were similar in the initial stage of penetration. However, as time progressed, the penetration rate of Example 1 remained at a high level, while the rate of Comparative Example 5 slowed down. This indicates that the effect of a one-time addition of a penetration enhancer will decay, while the in-situ dynamic generation mechanism can continuously maintain a highly efficient penetration driving force.

[0095] Compared with Example 1 and Comparative Example 4 (using ordinary red ginseng extract), under the same penetration-enhancing system, the cumulative penetration of Example 1 was 1.75 times that of Comparative Example 4. This is because Example 1 used a targeted synergistic red ginseng extract with a higher content of ginsenoside Rg3, indicating that the combination of a high concentration of active ingredient source and an efficient delivery system can achieve a high penetration efficiency.

[0096] In summary, by generating a permeation enhancer through in-situ hydrolysis with lipase and combining it with the core active ingredient enriched through biotransformation, the transdermal permeation efficiency of this active ingredient can be synergistically improved.

[0097] Test Example 4: The experimental steps are as follows: (1) Mouse macrophages (RAW 264.7) were 1×10 5 The cells / mL were seeded in 96-well plates and incubated at 37°C with 5% CO2 for 24 hours.

[0098] (2) Experimental groups: normal group, model group, transdermal solution group of Example 1, transdermal solution group of Comparative Example 1, and transdermal solution group of Comparative Example 4.

[0099] (3) Cell treatment: Discard the old culture medium. Except for the normal group, all other groups were given culture medium containing 1 μg / mL lipopolysaccharide (LPS). Subsequently, the three experimental groups were given an equal volume of the 24-hour transdermal solution collected in Test Example 3 (the receiving cell solution of Example 1, Comparative Example 1, and Comparative Example 4), while the normal group and the model group were given an equal volume of blank culture medium. The cells were cultured for another 24 hours.

[0100] (4) NO content detection: Collect cell culture supernatant from each well. Follow the instructions of the Griess kit. After reacting the supernatant with the Griess reagent, measure the absorbance at 540 nm using a microplate reader. Calculate the concentration of nitric oxide (NO) in the supernatant of each group using the sodium nitrite standard curve.

[0101] The experimental results are shown in Table 4.

[0102] Table 4. Effects of each test group on LPS-induced NO production in RAW 264.7 cells: This test case aims to correlate the results of in vitro transdermal penetration with biological effects to verify the effect of the present invention on improving the bioavailability of active ingredients.

[0103] Table 4 shows that the model group cells produced a large amount of NO under LPS stimulation, indicating that the inflammation model was successfully constructed. In Example 1, the transdermal solution group suppressed the NO concentration to 16.3 μM, which was superior to the inhibitory effect of the comparative example 1 transdermal solution group (42.8 μM) and the comparative example 4 transdermal solution group (30.5 μM).

[0104] This difference in biological activity directly corresponds to the cumulative permeation data of ginsenoside Rg3 in Test Example 3. Comparative Example 1, due to its extremely low Rg3 permeation, exhibited the weakest anti-inflammatory activity in its transdermal solution, confirming that the enzymatic delivery system is fundamental to the active ingredient's efficacy. Comparative Example 4, with its lower Rg3 concentration at the source of the active ingredient, showed weaker transdermal permeation and final anti-inflammatory activity compared to Example 1, indicating that the optimization of the active ingredient itself, combined with the delivery system, has a synergistic effect on the final efficacy.

[0105] The test results correlated the improved penetration efficiency with the final biological activity, demonstrating that the technical solution of the present invention can achieve a concentration at the target site sufficient to produce the expected physiological effect by increasing the penetration amount of the active ingredient.

[0106] The results of test examples 1 to 4 above verify the technical solution of the present invention and its beneficial effects from different dimensions.

[0107] The results of Test Example 1 confirmed the reaction mechanism of the present invention. Under simulated skin conditions, the enzymatic system in the formulation was activated, hydrolyzing the biomimetic triglyceride substrate into free fatty acids and glycerol, while simultaneously adjusting the environmental pH, demonstrating the feasibility of dynamically generating the product.

[0108] Transdermal water loss (TEWL) data from Test Example 2 show that, compared to the control group lacking enzymatic reaction components or with added end products, the embodiment based on this mechanism maintains skin barrier function for a longer period. This indicates that in-situ continuous lipid replenishment is more effective in repairing the skin barrier than simple physical occlusion or ingredient mixing.

[0109] The results of Test Example 3 explain the source of the synergistic effect. The free fatty acids generated by the enzymatic reaction increased the transdermal permeability of ginsenoside Rg3. This effect was superior to systems without enzymatic reactions or with statically added products, indicating that a dynamically generated permeation-enhancing environment can provide a sustained permeation effect. The test also showed that the total permeation was correspondingly increased when using biotransformed and enriched red ginseng extract.

[0110] Test Example 4 verified the enhanced biological activity resulting from high penetration efficiency. The results showed that the anti-inflammatory activity of the transdermal solution was positively correlated with the penetration amount of ginsenoside Rg3. The transdermal solution of Example 1 exhibited the strongest NO inhibition ability, demonstrating that the present invention improves the bioavailability and biological activity at the site of action by increasing the penetration amount of the active ingredient.

[0111] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. An external healing-promoting skin care preparation for dynamic regeneration of sebaceous glands, characterized by comprising, The formulation comprises the following components by weight percentage: Biomimetic triglyceride substrate: 3.0%~5.0%; Stabilized lipase: 0.1%~0.4%; Targeting and synergistic red ginseng extract: 0.3%~0.8%; Cosmetically acceptable carrier: balance.

2. The biogenic sebum dynamic regeneration, over-the-counter, pro-healing skin care formulation as claimed in claim 1, wherein, The biomimetic triglyceride substrate is composed of high-oleic sunflower oil and palmitic acid triglyceride, wherein the mass ratio of high-oleic sunflower oil to palmitic acid triglyceride is (70~90):(10~30).

3. The biogenic sebum dynamic regeneration, over-the-counter, pro-healing skin care formulation as claimed in claim 1, wherein, The preparation method of the targeting and synergistic red ginseng extract comprises: performing enzymatic conversion treatment on the red ginseng crude extract by β-glucosidase to enrich ginsenoside Rg3.

4. The biogenic sebum dynamic regeneration, over-the-counter, pro-healing skin care formulation as claimed in claim 1, wherein, The stabilized lipase is a triacylglycerol lipase derived from Aspergillus oryzae, which is stabilized by maltodextrin as a carrier.

5. The biogenic sebum dynamic regeneration, over-the-counter, pro-healing skin care formulation as claimed in claim 1, wherein, The cosmetically acceptable carrier comprises oil phase components, aqueous phase components and emulsifiers, and the formulation is an oil-in-water emulsion.

6. A method of preparing the external use healing-promoting skin care formulation for dynamic sebum regeneration of the biomimetic skin according to any one of claims 1 to 5, characterized in that, The method comprises the following steps: (a) emulsifying the oil phase components and the aqueous phase components under heating conditions to form an emulsion; (b) cooling the emulsion obtained in step (a) to below 40℃; (c) adding the biomimetic triglyceride substrate, the stabilized lipase and the targeting and synergistic red ginseng extract to the emulsion below 40℃ and mixing uniformly to obtain the product.

7. The preparation method according to claim 6, characterized in that, The method further comprises a pretreatment step of preparing the targeting and synergistic red ginseng extract: Take red ginseng crude extract, add β-glucosidase, and react at pH 5.0 and 55℃ for 36 hours, and then purify and dry to obtain the targeting and synergistic red ginseng extract.

8. The preparation method according to claim 6, characterized in that, In step (a), the temperature of the emulsification is 75℃~85℃.

9. The preparation method according to claim 6, characterized in that, In step (c), the temperature of the cooling is 38℃~40℃; and the operation of adding components is performed under vacuum conditions of-0.06 MPa~-0.08 MPa.

10. The method of claim 6, wherein, The specific implementation of step (c) is: First, add the biomimetic triglyceride substrate and mix, and then add the stabilized lipase and the targeting and synergistic red ginseng extract.

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