A blue copper peptide composition with instant cooling and penetration-enhancing effects, its preparation method and application.

Abstract

This invention relates to the field of cosmetics, specifically to a copper peptide composition with instant cooling and penetration-enhancing effects, its preparation method, and its application. The composition comprises the active ingredient copper peptide and a cooling and penetration-enhancing system composed of at least two of the following: menthol alkylformyl ethylamine, menthone glyceryl ketal, and methyl diisopropylpropionamide. The active ingredient may further include hexapeptide-11, cyanobacterial extract, and yeast fermentation products. The preparation method includes three core steps: providing an aqueous phase, adding the active ingredient, and adding the cooling and penetration-enhancing system and other components. This invention, through the synergistic combination of specific cooling agents, achieves highly efficient penetration enhancement of copper peptide while significantly reducing the dosage of each individual ingredient. It provides a rapid-onset, long-lasting, and gentle layered cooling experience, avoiding the skin irritation problems commonly found with traditional penetration enhancers or high-concentration single cooling agents, and possesses excellent transdermal efficiency, active ingredient stability, and user comfort.

Description

Technical Field

[0001] This invention relates to the field of cosmetic technology, and in particular to a blue copper peptide composition with instant cooling and penetration-enhancing effects, its preparation method, and its application. Background Technology

[0002] Damaged skin barrier, sensitive skin, and aging are core issues that urgently need to be addressed in the current skincare industry. Blue copper peptide (GHK-Cu) is an endogenous complex composed of a tripeptide (glycine, histidine, lysine) bound to copper ions, named for the blue color of its aqueous solution. Current research has found that blue copper peptide can promote the synthesis of collagen, elastin, and hyaluronic acid by fibroblasts, repair the skin barrier, accelerate wound healing, and synergistically scavenge free radicals, provide antioxidant protection, reduce inflammatory factors, alleviate inflammation, improve skin elasticity, and mitigate photoaging. It is widely used in high-end anti-wrinkle and repair skincare products and in the field of medical wound repair. Its gentle and hypoallergenic properties make it suitable for sensitive skin, but it should be avoided when using with high-concentration acids or strong oxidants. After opening, it should be stored away from light.

[0003] Copper peptides, as a highly effective repair and anti-aging active ingredient, have broad application prospects. However, this ingredient faces two major technical bottlenecks in practical applications: First, due to its large molecular weight and strong hydrophilicity, its transdermal absorption efficiency in traditional formulations is extremely low, resulting in insufficient bioavailability and difficulty in fully realizing its efficacy; Second, chemical penetration enhancers (such as azone) added to promote absorption or potent single cooling agents (such as menthol) added to improve skin feel are often highly irritating to the skin, especially unsuitable for skin with damaged barriers or sensitive skin, and may even affect the chemical stability of copper peptides themselves.

[0004] While existing technologies offer various complex formulas aimed at achieving multi-effect skincare, they often fail to fundamentally resolve the contradiction between the transdermal efficiency and gentleness of the aforementioned active ingredients. For example, Chinese invention patent CN120661418A discloses an anti-aging and moisturizing essence with a complex composition, including multiple active ingredients such as copper peptide, hexapeptide, and niacinamide, as well as plant extracts and fermentation products. This technical solution focuses on achieving multi-faceted skincare—"anti-aging, whitening, moisturizing, and repairing"—through a combination of multiple ingredients. However, it does not disclose any penetration-enhancing technology for the specific active ingredient copper peptide, and its formula does not contain any effective cooling penetration-enhancing system, failing to provide an immediate comfortable skin feel. Furthermore, it may have shortcomings in terms of the stability of active ingredients and targeted delivery efficiency.

[0005] To improve the transdermal absorption of copper peptides, existing technologies have proposed several solutions. For example, Chinese invention patent CN114983856B discloses a copper peptide solution with a penetration-enhancing system and its preparation method. This technology uses an ionic liquid or an ionic liquid combined with 4-hydroxyethylpiperazine ethanesulfonic acid (HEPES) as a penetration-enhancing system. The ionic liquid encapsulates the copper peptides, while HEPES loosens the stratum corneum to achieve the purpose of enhancing penetration. Although this technical solution has improved the transdermal absorption of copper peptides to a certain extent, it still has the following limitations: (1) The preparation process of its core penetration-enhancing component, the ionic liquid, is relatively complex. It requires specific organic bases and organic acids or polyols to react and synthesize under heating conditions (40-80℃) for a long time (10-12 hours), which increases the production energy consumption and the difficulty of cost control; (2) This solution mainly focuses on solving the transdermal problem and does not involve or optimize the immediate skin feel when using the product, which cannot meet consumers' needs for a comfortable experience of skin care products.

[0006] Therefore, there is an urgent need in this field for a new technical solution that can overcome the shortcomings of existing technologies, namely: while ensuring the high transdermal efficiency and high stability of copper peptides, it can provide excellent immediate skin comfort, and the preparation process is simple and easy to industrialize, so as to meet the market's urgent demand for high-performance, low-irritation, and high-experience skin care products. Summary of the Invention

[0007] To address the aforementioned technical problems, the present invention aims to provide a copper peptide composition with instant cooling and penetration-enhancing effects, its preparation method, and its application, thereby overcoming the shortcomings of the prior art and providing a copper peptide composition and its application that combines highly efficient penetration enhancement, excellent soothing and repair capabilities, and a layered, comfortable cooling sensation.

[0008] To achieve the above-mentioned technical effects, the present invention adopts the following technical solution:

[0009] In a first aspect, the present invention provides a blue copper peptide composition having an instant cooling sensation and a penetration-enhancing effect, comprising an active ingredient and a cooling and penetration-enhancing system, wherein the active ingredient comprises at least blue copper peptide, and the cooling and penetration-enhancing system is composed of at least two of menthol alkyl ethylamine, menthone glycerol ketal, and methyl diisopropyl propionamide.

[0010] Preferably, the cooling and penetration-enhancing system is composed of menthol carbamoyl ethylamine, menthone glycerol ketal, and methyl diisopropyl propionamide in a mass ratio of (0.2-0.5):(0.5-1.0):(0.8-1.5).

[0011] Preferably, the content of the cooling and penetration-enhancing system in the composition is 1.0-3.0% by weight percentage.

[0012] Preferably, the content of the blue copper peptide is 0.4-2% by weight.

[0013] Preferably, the active ingredient further comprises yeast fermentation product, which is selected from at least one of galactosomal yeast fermentation product filtrate and yeast polypeptides.

[0014] Preferably, the active ingredients also include hexapeptide-11 and cyanobacterial extract.

[0015] More preferably, the active ingredients include copper peptide, galactosomal yeast fermentation product filtrate, yeast polypeptides, hexapeptide-11, and cyanobacterial extract.

[0016] Most preferably, the mass ratio of the active ingredients, including copper peptide, hexapeptide-11 and cyanobacterial extract, is (1.2-1.5):(0.3-0.6):(1-1.2).

[0017] Furthermore, the composition also includes a moisturizing and repairing component, which includes at least one or more combinations of trehalose, erythritol, dipropylene glycol, and butylene glycol.

[0018] Preferably, the moisturizing and repairing component is present in the composition at a weight percentage of 1-35% wt, more preferably 5-20% wt.

[0019] Secondly, the present invention also provides the application of the aforementioned composition of blue copper peptides having an instant cooling sensation and penetration-enhancing effect in skin care products.

[0020] Furthermore, the skincare product in question is a leave-on skincare product.

[0021] Thirdly, the present invention further provides a method for preparing the aforementioned blue copper peptide composition with instant cooling and penetration-enhancing effects, comprising the following steps:

[0022] S1: Provide the aqueous phase: Mix at least a portion of water and optional moisturizing and repairing ingredients, heat to 40-50°C and stir until completely dissolved to form a homogeneous aqueous phase;

[0023] S2: Add active ingredients: Cool the aqueous phase obtained in step S1 to 30-35℃, add copper peptide, hexapeptide-11, blue algae extract and optional yeast fermentation product, and stir until completely dissolved.

[0024] S3: Add cooling and penetration-enhancing system and other components: Add cooling and penetration-enhancing system and other remaining components to the system obtained in step S2, and continue stirring until the system is clear and transparent. Add the remaining water in the formula to make up the volume, and the composition is obtained.

[0025] Preferably, in S1, the portion of water accounts for 10-90% of the total water volume of the formula.

[0026] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0027] Compared with existing technologies, this invention has the following significant advantages: This invention creatively constructs a penetration-enhancing system with a specific ternary cooling agent as its core. Through the complementarity and synergy of the components in terms of action mechanism, onset time, and duration, a synergistic effect of "1+1+1>3" is achieved. In vitro transdermal experiments confirm that this system's penetration-enhancing efficiency for copper peptides is significantly better than that of single cooling agents, binary compound systems, and the traditional chemical penetration enhancer azone. Accelerated stability tests show that this system can maintain the chemical stability of copper peptides very well, preventing their dissociation and inactivation, and its effect is far superior to the control group. Human sensory evaluation results further demonstrate that this composition can provide a distinct and comfortable cooling sensation, with rapid onset and long duration, while reducing unpleasant sensations such as stinging and burning to extremely low levels. Its mildness is comparable to the blank matrix, successfully resolving the contradiction between high-efficiency penetration enhancement and skin irritation, making it particularly suitable for sensitive skin and those with damaged skin barriers.

[0028] In summary, the core innovation of this invention lies in constructing a synergistic delivery system centered on a "triple cooling agent compound system." This system, through the complementary and synergistic effects of menthol alkyl ethylamine, menthone glycerol ketal, and methyl diisopropyl propionamide in terms of their mechanisms of action, onset time, and duration of action, achieves highly efficient penetration enhancement of copper peptides while significantly reducing the dosage of each individual ingredient. It provides a layered cooling experience that is "rapidly effective and long-lastingly gentle," fundamentally avoiding skin irritation that may be caused by single potent cooling agents or traditional penetration enhancers. Detailed Implementation

[0029] The following embodiments are only used to illustrate the technical solutions of the present invention more clearly, and are therefore only examples and should not be used to limit the scope of protection of the present invention.

[0030] Those skilled in the art will understand that the present invention can be practiced even without certain specific details. In some other embodiments, methods, means, apparatus, and steps well known to those skilled in the art have not been described in detail in order to highlight the spirit of the invention. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art. Unless otherwise specified, all units used in this specification are International Standard Units (SI), and all numerical values ​​and ranges appearing in this invention should be understood to include systematic errors unavoidable in industrial production.

[0031] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods; the materials, reagents, or instruments used, unless otherwise specified by the manufacturer, are all commercially available reagents and materials; the conditions not specified in the examples are all carried out under conventional conditions or conditions recommended by the manufacturer. Furthermore, this invention does not limit the source of the raw materials used; unless otherwise specified, the raw materials used in this invention are all commercially available products in this technical field. Unless otherwise specified, the "ratios" referred to in the following examples are ratios of mass parts. It should be specifically noted that the butanediol used in this invention is 1,3-butanediol. Blue-green algae extract: Shaanxi Haoyang Biotechnology Co., Ltd., E18. Galactosomalidomic fermentation product filtrate: Huzhou Purui Cosmetic Technology Co., Ltd. Yeast polypeptides: Guangzhou Qiancui Biotechnology Co., Ltd.

[0032] Example 1

[0033] The purpose of this embodiment is to provide a blue copper peptide composition with instant cooling and penetration-enhancing effects, and its preparation method. Specifically, its formulation is as follows:

[0034] The formulation (based on a yield of 1000g) comprises: copper peptide: 15.0g, hexapeptide-11: 3.0g, blue algae extract: 12.0g, galactosomal yeast fermentation product filtrate: 10.0g, yeast polypeptides: 0.0005g, menthol alkyl ethylamine (WS-3): 5.0g, menthone glycerol ketal: 5.0g, methyl diisopropyl propionamide: 10.0g, erythritol: 50.0g, trehalose: 50.0g, dipropylene glycol: 15.0g, butylene glycol: 80.0g, ethylhexylglycerin: 0.5g, 1,2-hexanediol: 40.0g, and water to a total of 1000.0g.

[0035] The composition is prepared as follows:

[0036] S1: Provide the aqueous phase: Add measured amounts of water, erythritol, trehalose, dipropylene glycol, and 1,3-butanediol to a mixing pot, heat to 45°C and stir until all components are completely dissolved to form a homogeneous aqueous phase.

[0037] S2: Add active ingredients: Cool the aqueous phase obtained in step S1 to 35°C, add copper peptide, galactosomal yeast fermentation product filtrate and yeast polypeptides, and stir until completely dissolved.

[0038] S3: Add cooling and penetration-enhancing system and other components: Menthol alkylformyl ethylamine, menthone glycerol ketal and methyl diisopropyl propionamide are added sequentially to the system obtained in step S2, followed by ethylhexylglycerin and 1,2-hexanediol. The mixture is stirred until it is clear and transparent to obtain the composition, which is referred to as composition A.

[0039] Example 2

[0040] The purpose of this embodiment is to provide a blue copper peptide composition with instant cooling and penetration-enhancing effects, and its preparation method. Specifically, its formulation differs from that of the previous embodiment. The formulation used in this embodiment is as follows:

[0041] The formula, based on the preparation of 1000g, includes: copper peptide: 18.0g, hexapeptide-11: 5.0g, blue algae extract: 10.0g, galactosomal yeast fermentation product filtrate: 15.0g, yeast polypeptides: 0.0008g, menthol carbamoyl ethylamine: 3.0g, menthone glycerol ketal: 5.0g, methyl diisopropyl propionamide: 15.0g, erythritol: 60.0g, trehalose: 40.0g, dipropylene glycol: 25.0g, butylene glycol: 100.0g, ethylhexylglycerin: 0.8g, 1,2-hexanediol: 60.0g, and water to 1000.0g.

[0042] The composition is prepared as follows:

[0043] S1: Provide the aqueous phase: Add appropriate amounts of water, erythritol, trehalose, dipropylene glycol and butylene glycol to the preparation pot, heat to 48°C and stir until all components are completely dissolved to form a homogeneous aqueous phase.

[0044] S2: Add active ingredients: Cool the aqueous phase obtained in step S1 to 33°C, add copper peptide, galactosomal yeast fermentation product filtrate and yeast polypeptides, and stir until completely dissolved.

[0045] S3: Add cooling and penetration-enhancing system and other components: Menthol alkyl ethylamine, menthone glycerol ketal and methyl diisopropyl propionamide are added sequentially to the system obtained in step S2, followed by ethylhexylglycerin and 1,2-hexanediol. The mixture is stirred until it is clear and transparent to obtain the composition, which is referred to as composition B.

[0046] Example 3

[0047] The purpose of this embodiment is to provide a blue copper peptide composition with instant cooling and penetration-enhancing effects, and its preparation method. Specifically, its formulation differs from that of the previous embodiment. The formulation used in this embodiment is as follows:

[0048] The formula, based on the preparation of 1000g, includes: copper peptide: 12.0g, hexapeptide-11: 6.0g, blue algae extract: 11.0g, galactosomal yeast fermentation product filtrate: 5.0g, yeast polypeptides: 0.0001g, menthol carbamoyl ethylamine: 8.0g, menthone glycerol ketal: 2.0g, methyl diisopropyl propionamide: 3.0g, erythritol: 40.0g, trehalose: 60.0g, dipropylene glycol: 30.0g, butylene glycol: 70.0g, ethylhexylglycerin: 0.1g, 1,2-hexanediol: 30.0g, and water to 1000.0g.

[0049] The composition is prepared as follows:

[0050] S1: Provide the aqueous phase: Add appropriate amounts of water, erythritol, trehalose, dipropylene glycol and 1,3-butanediol to the preparation pot, heat to 42°C and stir until all components are completely dissolved to form a homogeneous aqueous phase.

[0051] S2: Add active ingredients: Cool the aqueous phase obtained in step S1 to 30°C, add copper peptide, galactosomal yeast fermentation product filtrate and yeast polypeptides, and stir until completely dissolved.

[0052] S3: Add cooling and penetration-enhancing system and other components: Menthol alkylformyl ethylamine, menthone glycerol ketal and methyl diisopropylpropionamide are added sequentially to the system obtained in step S2, followed by ethylhexylglycerin and 1,2-hexanediol. The mixture is stirred until it is clear and transparent to obtain the composition, which is referred to as composition C.

[0053] Experimental Example 1

[0054] The purpose of this experiment is to observe the synergistic effect of the cooling and penetration-enhancing system of the present invention (menthol alkyl ethylamine, menthone glycerol ketal, and methyl diisopropyl propionamide) on the penetration enhancement of copper peptides, and to compare it with single / dual cooling agents and traditional formulations, as detailed below:

[0055] 1.1 Experimental and Control Group Design

[0056] Except for the "cooling and penetration-enhancing system", all other components of all groups are completely identical (refer to the basic formula of Example 1, but only prepare 100mL of system), only the type or composition of the penetration-enhancing system is changed to ensure that the variable is unique.

[0057] The group settings for this embodiment are shown in Table 1:

[0058] Table 1. Experimental and control group settings

[0059] Group number Group type Composition of the penetration enhancement system The specific composition of the cooling and penetration-enhancing system (parts by mass, total mass 2.0 g, consistent with the total concentration of the ternary system in composition A) EG experimental group The ternary system of this invention (same as the composition) Menthyl alkyl ethylamine 0.5g + Menthone glycerol ketal 0.5g + Methyl diisopropyl propionamide 1.0g CG-1 Blank control No penetration enhancer none CG-2 Single control 1 Mono-mentholmethaneformylethylamine Menthyl methylformamide 2.0g CG-3 Single control 2 Mono-menthone glycerol acetal Menthone glycerol ketal 2.0g CG-4 Single control 3 Mono-methyldiisopropylpropionamide Methyl diisopropyl propionamide 2.0g CG-5 Binary comparison 1 Menthyl alkyl ethylamine + Menthone glycerol ketal Menthyl methylformamide 1.0g + Menthone glycerol ketal 1.0g CG-6 Binary control 2 Menthyl carbamoyl ethylamine + methyl diisopropyl propionamide Menthyl benzoyl ethylamine 0.67g + methyl diisopropyl propionamide 1.33g CG-7 Binary control 3 Menthone glycerol ketal + methyl diisopropyl propionamide Menthone glycerol acetal 0.67g + methyl diisopropyl propionamide 1.33g CG-8 Traditional permeation control Azone Azone 2.0g

[0060] 1.2 Experimental Methods

[0061] Skin model preparation: Take fresh detached pig skin (back skin, remove subcutaneous fat), rinse it 3 times with physiological saline, cut it into skin slices with a thickness of 0.8±0.1mm, and store it in 0.9% physiological saline (containing 0.1% sodium azide) at 4℃. Use it within 24 hours.

[0062] Franz diffusion cell parameter settings: A vertical Franz diffusion cell was used, with an effective diffusion area of ​​1.77 cm², a receiving chamber volume of 7 mL, and a receiving solution of pH 7.4 phosphate buffered saline (PBS, containing 0.02% Tween-80 to maintain the solubility of the blue copper peptide); the magnetic stirring speed was 300 rpm, and the temperature was controlled by a constant temperature water bath at 37 ± 0.5℃.

[0063] Sample loading and sampling: The pigskin was fixed between the supply and receiving chambers of the diffusion cell. The receiving chamber was filled with receiving liquid and air bubbles were removed. 0.2 mL of sample was added to the supply chamber, ensuring a consistent amount per unit area. After sealing, timing was started. Sampling was performed after 24 hours: 0.5 mL of receiving liquid was aspirated, and the sample was filtered through a 0.22 μm organic phase filter membrane. The concentration of blue copper peptide was determined by HPLC.

[0064] Based on the experimental results, the cumulative permeation rate (Qn, μg / cm²) and steady-state transdermal rate (Jss, μg / cm² / h) were calculated.

[0065] The experimental results are shown in Table 2:

[0066] Table 2. In vitro transdermal permeation parameters of copper peptide in each experimental group and control group (n=3, Mean±SD)

[0067] Group describe 24-hour cumulative permeability Qn (μg / cm²) Steady-state transdermal rate Jss (μg / cm² / h) Qn improvement factor relative to CG-1 EG Tri-element cooling and penetration-enhancing system 28.56±2.23 1.42±0.12 4.94 times CG-1 No penetration enhancer 5.78±0.69 0.29±0.03 / CG-2 Single Menthylcarbamate 16.32±1.45 0.82±0.07 2.82 times CG-3 Menthone glycerol ketal 10.15±0.98 0.51±0.05 1.76 times CG-4 Monomethyldiisopropylpropionamide 12.48±1.13 0.62±0.06 2.16 times CG-5 Dimeric (menthol alkylformyl ethylamine + menthone glycerol ketal) 20.75±1.81 1.03±0.09 3.59 times CG-6 Dimeric (mentholmaneformylethylamine + methyldiisopropylpropionamide) 22.14±1.76 1.11±0.08 3.83 times CG-7 Dimeric (menthol glycerol acetal + methyl diisopropyl propionamide) 18.92±1.63 0.95±0.07 3.27 times CG-8 Traditional penetration enhancer (azone) 23.35±1.92 1.17±0.10 4.04 times

[0068] The above experimental results show that:

[0069] The 24-hour Qn (28.56 μg / cm²) of the experimental group EG (ternary system) was 4.94 times that of the blank control CG-1 (5.78 μg / cm²), and significantly higher than all single cooling agent groups (CG-2~CG-4: 10.15~16.32 μg / cm²). This indicates that the combination of the three cooling agents is not a simple "concentration additive," but rather significantly enhances the transdermal ability of copper peptides through synergistic effects. The speculated mechanism may be: menthol ethylamine rapidly loosens the intercellular spaces of the stratum corneum, menthone glycerol ketal prolongs the hydration time of the stratum corneum, and methyl diisopropyl propionamide increases the solubility of copper peptides in the stratum corneum. The three work synergistically from three dimensions—"channel opening-duration effect-component dissolution"—overcoming the limitations of single cooling agents.

[0070] Meanwhile, the experimental results above show that the Qn (18.92~22.14 μg / cm²) of all binary system groups (CG-5~CG-7) is lower than that of EG (28.56 μg / cm²). Among them, the best performing binary group CG-6 (menthol ethylcarbamate + methyl diisopropyl propionamide) has a Qn of only 77.5% of EG, and Jss is only 78.1% of EG. This indicates that the absence of any cooling agent (such as the absence of methyl diisopropyl propionamide in CG-5 and the absence of menthol ethylcarbamate in CG-7) will lead to the breakage of the penetration-promoting chain: for example, due to the absence of methyl diisopropyl propionamide, the solubility of copper peptide in the stratum corneum of CG-5 is insufficient, and it cannot make full use of the opportunity of "opening of the stratum corneum gaps" to penetrate; due to the absence of menthol ethylcarbamate, the stratum corneum loosening efficiency of CG-7 is low, and the subsequent penetration-promoting effect cannot be effectively exerted.

[0071] The traditional chemical penetration enhancer azone group (CG-8) has a Qn (23.35 μg / cm²) that is 21.7% lower than that of EG. Furthermore, azone, as a synthetic penetration enhancer, has the drawback of being "highly irritating to sensitive skin". In contrast, the ternary system of this invention is based on a combination of natural and mild cooling agents, which achieves higher penetration efficiency while avoiding the irritation problem of traditional penetration enhancers, making it more suitable for the use of sensitive skin and skin with damaged skin barrier.

[0072] In summary, the ternary cooling and penetration-enhancing system of sample A of this invention (menthol alkylformyl ethylamine, menthone glyceryl ketal, and methyl diisopropyl propionamide) is significantly superior to no penetration enhancer, single cooling agent, binary cooling agent, and traditional penetration enhancer in terms of the penetration-enhancing effect of blue copper peptides. It is the optimal solution to achieve "highly efficient penetration enhancement + gentle skin feel".

[0073] Experimental Example 2

[0074] The purpose of this experiment is to observe the effect of the cooling and penetration-enhancing system of the present invention (menthol alkylformyl ethylamine, menthone glycerol ketal, and methyl diisopropyl propionamide) on the chemical stability of copper peptides, and to compare it with single / dual cooling agents and traditional formulations, as detailed below:

[0075] 2.1 Experimental and Control Group Design

[0076] All group formulations are completely based on Experiment Example 1, and the specific group composition and numbering of the penetration-enhancing system are also consistent with the design of Experiment Example 1.

[0077] 2.2 Experimental Methods

[0078] Prepare samples according to the above group formulations (refer to the basic formulation of Example 1, the basic formulations are the same, only the composition of the permeation-enhancing system is different), prepare 3 parallel samples for each group, and package them in brown light-proof vials (to avoid light affecting the stability of blue copper peptides), seal them and keep them for later use.

[0079] Accelerated stability test conditions: All samples were placed in a constant temperature and humidity accelerated stability test chamber. The conditions were set as follows: temperature 40±2℃, relative humidity 75±5% (refer to the standard for accelerated stability testing of cosmetics), and stored away from light. Samples were taken and tested at the end of 0 months (initial state), 1 month, 2 months and 3 months respectively, and 3 parallel samples were taken for testing each time.

[0080] The testing indicators include:

[0081] (1) Residual content of blue copper peptide

[0082] The HPLC method was used for determination, and the chromatographic conditions were the same as those in Experiment 1: C18 column (250 mm × 4.6 mm, 5 μm), mobile phase methanol-0.1% phosphoric acid aqueous solution (30:70, v / v), flow rate 1.0 mL / min, detection wavelength 220 nm, column temperature 30 ℃, and injection volume 20 μL.

[0083] Content residual rate (%) = (measured concentration of blue copper peptide at the sampling time point / measured concentration at 0 months) × 100%.

[0084] (2) Solution appearance and clarity

[0085] According to the appearance inspection method in the "Cosmetic Safety Technical Specifications", visually observe the color, clarity and presence of sediment / turbidity of the sample against a white background, and record the characteristics.

[0086] The experimental results are shown in Table 3:

[0087] Table 3. Accelerated stability test results of blue copper peptides in each experimental group (n=3, Mean±SD)

[0088]

[0089] The above experimental results show that:

[0090] After 3 months of accelerated testing, the residual content of copper peptides in the EG group reached 97.8%, only 1.7% higher than that of CG-1 (96.1%) without any penetration enhancer, and both solutions remained clear and transparent throughout (no turbidity, no precipitation). This result indicates that when the ternary cooling and penetration-enhancing system of the present invention (menthol alkylformyl ethylamine + menthone glycerol ketal + methyl diisopropyl propionamide) is combined with copper peptides, no additional chemical interactions are introduced. It neither destroys the copper-peptide coordination structure of copper peptides nor causes a decrease in the physical stability of the system.

[0091] Meanwhile, the experimental results showed that the residual rates of all single cooling agent groups (CG-2~CG-4) were significantly lower than those of EG after 3 months, and the stability differences were obvious. Among them, CG-4 (monomethyl diisopropyl propionamide) had the worst stability, with a residual rate of only 78.5% after 3 months, which was 19.3 percentage points lower than EG. Moreover, the residual rate had dropped to 84.8% after 2 months, and there was obvious turbidity in 3 months. This indicates that when methyl diisopropyl propionamide is used alone and in large quantities, its molecular structure may compete weakly with the copper ions of blue copper peptide, accelerating the dissociation of the active ingredient. The above experimental results prove that single cooling agents not only have limited penetration-enhancing effects, but also pose a "risk of stability degradation". The ternary system, through the synergistic combination of the three components, can offset the negative impact of a single component on the stability of blue copper peptide and significantly improve the stability of the system. In the binary combination, "menthol ethylamine + methyl diisopropyl propionamide (CG-6)" generally performed better than the single-component cooling agent group, but was still significantly lower than the EG group. Among them, CG-6 (menthol ethylamine 0.67g + methyl diisopropyl propionamide 1.33g) was the best performing binary combination, with a residue rate of 88.3% after 3 months, but still 9.5 percentage points lower than EG, and slight turbidity appeared after 2 months. The CG-7 group had the worst stability, with a residue rate of only 84.2% after 3 months. This result proves that binary cooling agents can only partially alleviate the stability problem of single components, and only the ternary system can achieve the optimal balance between "penetration efficiency" and "stability".

[0092] The above experimental results demonstrate that the ternary cooling and penetration-enhancing system of this invention is not only the optimal solution for improving the transdermal efficiency of copper peptides, but also a key technology for protecting the chemical stability of copper peptides. Through the synergistic effect of the three cooling agents, it avoids the stability defects of single / binary cooling agents and overcomes the irritation and degradation risks of traditional penetration enhancers and comparative solutions. This provides irreplaceable technical support for the development of "highly effective + stable + gentle" skincare products for sensitive skin and skin with damaged barriers.

[0093] Experimental Example 3

[0094] This experimental case aims to quantify and verify, through subjective human evaluation, the layered cooling sensation provided by the ternary cooling and penetration-enhancing system (EG) of this invention in actual use, characterized by "rapid onset and long-lasting gentleness," and to evaluate its advantages over single cooling agents, binary compound systems, and traditional penetration enhancers in terms of skin irritation, as detailed below:

[0095] 3.1 Experimental and Control Group Design

[0096] Except for the "cooling and penetration-enhancing system", all other components of all groups were prepared in accordance with the basic formula of Example 1 to ensure that the variables were unique; the group numbers and the composition of the penetration-enhancing system were completely consistent with those of Experiment 1, and will not be repeated here.

[0097] 3.2 Test Methods

[0098] Baseline criteria: Participants: Ninety healthy volunteers (half male and half female, aged 20-48 years) with no history of skin diseases were recruited and underwent acclimatization preparation as required before the test. They were randomly divided into 9 groups of 10 each and underwent bilateral facial testing.

[0099] Environment: Constant temperature and humidity room (temperature 25±1℃, relative humidity 50±5%). Subjects sat quietly for 30 minutes before the test to adapt to the environment.

[0100] 3.3 Sample preparation and application methods

[0101] Each sample was prepared as a uniform, transparent liquid and dispensed into colorless blind tubes of uniform size (only the group number was marked; both the subjects and the raters were double-blind). Three independent test areas of 2.5cm × 2.5cm were selected on the left and right sides of the face and the forehead, and 0.5mL of sample was accurately taken from each area. The sample was then evenly spread in the same direction with a sterile cotton swab and left to stand for 1 minute for natural absorption.

[0102] Subjects self-rated the results, and the scoring was conducted at three time points: 0.5 min (onset of action), 5 min, and 60 min after application. The scoring indicators and standards are shown in Table 4.

[0103] Table 4 Definition of Cooling Sensation Index

[0104] index Definitions and Classification Standards Cooling intensity 0 = No cooling sensation; 1~3 = Slight (perceived only at fingertips); 4~6 = Mild (comfortable and non-irritating); 7~9 = Strong (distinctly cooling); 10 = Stinging cooling sensation stinging sensation 0 = None; 1~2 = Slight (negligible); ≥3 = Obvious (requires attention); ≥5 = Strong (discomfort) burning sensation 0 = None; 1~2 = Slight; ≥3 = Obvious; ≥5 = Strong lingering numbness Evaluation at 60 minutes: 0 = none; 1-2 = slight; ≥3 = noticeable (skin dullness).

[0105] 3.4 Test Results

[0106] Table 5 shows the intensity scores of cooling sensation at each time point (only the cooling sensation group is included; CG-1 and CG-8 have no cooling sensation), and Table 6 shows the peak scores of unpleasant sensations.

[0107] Table 5. Average cooling intensity scores (Mean±SD, 0~10 points) at different time points for each group

[0108]

[0109] Table 6. Peak scores of adverse feelings in each group (Mean±SD, 0~10 points)

[0110] Evaluation indicators EG (Ternary) CG-1 (Blank) CG-2 (Single) CG-3 (Single 2) CG-4 (Single 3) CG-5 (binary 1) CG-6 (binary 2) CG-7 (binary 3) CG-8 (Azone) Maximum stinging sensation 0.7 ± 0.3 0.0 ± 0.0 3.8 ± 1.0 1.2 ± 0.4 2.5 ± 0.8 2.1 ± 0.7 2.3 ± 0.8 1.5 ± 0.5 2.8 ± 0.9 Maximum burning sensation 0.4 ± 0.2 0.0 ± 0.0 3.2 ± 0.9 0.8 ± 0.3 1.8 ± 0.6 1.5 ± 0.5 1.7 ± 0.6 1.0 ± 0.4 2.2 ± 0.7 60 minutes of numbness 1.1 ± 0.4 0.0 ± 0.0 2.8 ± 0.7 0.5 ± 0.2 1.2 ± 0.5 1.3 ± 0.5 1.5 ± 0.6 0.8 ± 0.3 0.0 ± 0.0 Statistical differences - p>0.05 p<0.01 p<0.05 p<0.01 p<0.01 p<0.01 p<0.05 p<0.01

[0111] 3.5 Results Analysis and Conclusions

[0112] As shown in Table 5, the cooling sensation curve of the experimental group (EG) exhibited an ideal "rapid onset - smooth transition - long-lasting maintenance" characteristic. At 0.5 minutes, its cooling intensity (6.3) was lower than that of CG-2 (7.5), but it avoided an excessively strong instantaneous impact; at 5 minutes, the cooling intensity of EG rose to its peak (6.7) and remained mild, while CG-2 had significantly decreased (5.1); most importantly, at 60 minutes, EG could still maintain a perceptible cooling sensation (2.4), which was significantly better than (p<0.01) all control groups, demonstrating the perfect complementarity of the three cooling agents in terms of duration and achieving a layered cooling experience.

[0113] Table 6 clearly shows that EG scored extremely low on key irritation indicators such as stinging and burning sensations, with no statistically significant difference from the control group (CG-1) (p>0.05), but significantly lower than all other experimental groups (p<0.01 or p<0.05). This indicates that by reducing the dosage of a single ingredient through triple compounding technology, this invention effectively avoids stimulation caused by excessive activation of TRPM8 receptors from the source. Although EG provides a lasting cooling sensation, its numbness after 60 minutes (1.1) is also at a very low level, far lower than that of the high-concentration single cooling agent CG-2 (2.8), demonstrating good skin comfort.

[0114] The above human sensory evaluation experiments, using objective scale data, confirm that the ternary cooling and penetration-enhancing system (EG) of this invention can provide a layered cooling experience that is fast-acting, moderately strong, and significantly longer-lasting than the control group. Simultaneously, this system greatly reduces the stinging and burning sensations commonly experienced with single or binary cooling agents. Its mildness is comparable to the blank matrix, achieving a high degree of unity between powerful efficacy and extreme gentleness. This demonstrates the success of this invention in achieving the technical effect of "rapid onset, long-lasting effect, and low irritation" through the compounding of specific cooling agents, resolving the contradiction between strong penetration enhancement and skin irritation.

[0115] The above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit it. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the spirit and scope of the present invention, and all such modifications and substitutions should be covered within the scope of the claims of the present invention. Technical aspects, shapes, and structures not described in detail in this invention are all well-known technologies.

Claims

1. Use of a combination of menthanecarboxylic acid ethylamide, menthone glycerol ketal and methyl diisopropyl propionamide in a weight ratio of 1 : 1 : 2 in a synergistically enhanced blue copper peptide in-vitro transdermal penetration enhancing composition, characterized in that, The composition comprises: Blue Copper Peptide: 15.0 g, Hexapeptide-11: 3.0 g, Cyanobacterial Extract: 12.0 g, Fermented filtrate of Galactomyces: 10.0 g, Yeast Polypeptides: 0.0005 g, Menthanecarboxylic Acid Ethylamide: 5.0 g, Menthone Glycerin Acetal: 5.0 g, Methyl Diisopropyl Propionamide: 10.0 g, Erythritol: 50.0 g, Fuculose: 50.0 g, Dipropylene Glycol: 15.0 g, Butylene Glycol: 80.0 g, Ethylhexylglycerin: 0.5 g, 1,2-Hexanediol: 40.0 g, and water to 1000.0 g.

2. Use of a combination of menthanecarboxylic acid ethylamide, menthone glycerol ketal and methyldiisopropylpropionamide in a weight ratio of 1 : 1 : 2 in synergistically enhancing the stability of ceruloplasmin peptide in a composition, characterized in that, The composition comprises: Blue Copper Peptide: 15.0 g, Hexapeptide-11: 3.0 g, Cyanobacterial Extract: 12.0 g, Fermented filtrate of Galactomyces: 10.0 g, Yeast Polypeptides: 0.0005 g, Menthanecarboxylic Acid Ethylamide: 5.0 g, Menthone Glycerin Acetal: 5.0 g, Methyl Diisopropyl Propionamide: 10.0 g, Erythritol: 50.0 g, Fuculose: 50.0 g, Dipropylene Glycol: 15.0 g, Butylene Glycol: 80.0 g, Ethylhexylglycerin: 0.5 g, 1,2-Hexanediol: 40.0 g, and water to 1000.0 g.

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