A composition with wrinkle-removing and anti-aging effects and application thereof

Abstract

The application discloses a composition with instant wrinkle-removing and long-term anti-aging effects, which comprises the following components in parts by weight: 10-30 parts of an anti-wrinkle soothing agent, 0.03-1 parts of an extract of Anoectochilus roxburghii, 0.5-3 parts of an extract of Inula japonica var. rotata, and 5-20 parts of a protein generation peptide compound. The composition uses natural plant extracts and the protein generation peptide compound to exert long-term anti-aging and wrinkle-removing effects on the basis of the anti-wrinkle soothing agent, and uses hexapeptide-11 and a marine algae extract to obtain anti-aging effects, so that the double effects of internal and external combination and multiple paths are combined to achieve the purposes of long-term wrinkle-removing, anti-aging and tightening. When the composition is applied to cosmetics, the problems of rough skin texture, skin relaxation, wrinkles and the like can be solved, and the composition is particularly suitable for use in skin with water deficiency, damage and aging phenomena.

Description

Technical Field

[0001] This invention belongs to the field of cosmetic technology, specifically relating to a composition with wrinkle-reducing and anti-aging effects, and also to the application of the composition. Background Technology

[0002] Skin aging manifests comprehensively as dryness, roughness, fine lines, sagging, thickened epidermis, and dullness, influenced by both internal and external factors. Internal factors, namely natural aging, include excess free radicals and mitochondrial DNA damage, which are unavoidable processes. External factors are primarily photoaging. Currently, most anti-aging skincare products on the market focus on eliminating free radicals or stimulating collagen production, but their effects are not significant enough and are prone to rebound.

[0003] Fibronectin plays a vital role in various physiological functions. It promotes cell migration, adhesion, proliferation, coagulation, and repair; activates monocytes and macrophages to eliminate toxins in damaged tissues; and promotes the synthesis of DNA, RNA, and proteins, thereby improving cell shape and function. It also plays a crucial role in cell culture. Fibronectin is a key messenger protein that activates collagen in the extracellular matrix (ECM) and stimulates the production of hyaluronic acid and other proteins, altering the normal arrangement of the ECM and thus alleviating wrinkles. Current cosmetic compositions often utilize the combination of fibronectin and peptides, but most are inclusion complexes and do not consider long-term anti-aging effects. They also lack immediate wrinkle-reducing effects, offering only a single approach to addressing aging or representing a single cosmetic form. Summary of the Invention

[0004] The purpose of this invention is to provide a composition with wrinkle-reducing and anti-aging effects, which has long-lasting wrinkle-reducing, anti-aging, and firming effects.

[0005] Another object of the present invention is to provide the application of the above composition in cosmetic additives.

[0006] The technical solution adopted in this invention is a composition with wrinkle removal and anti-aging effects, comprising the following components by weight: 10-30 parts anti-wrinkle and soothing agent, 0.03-1 part *Cymbidium goeringii* extract, 0.5-3 parts *Inula japonica* extract, and 5-20 parts protein-generating peptide complex.

[0007] The invention is further characterized in that, Preferably, the composition further comprises 30 to 60 parts of hexapeptide.

[0008] Preferably, the composition further comprises 2 to 10 parts of a complex algae extract.

[0009] Preferably, the composition comprises the following components in parts by weight: 15-30 parts of anti-wrinkle and soothing agent, 0.05-1 part of *Cymbidium goeringii* extract, 0.8-3 parts of *Inula japonica* extract, and 10-20 parts of protein-generating peptide complex.

[0010] More preferably, the composition comprises the following components in parts by weight: 15-25 parts of anti-wrinkle and soothing agent, 0.08-1 part of *Cymbidium goeringii* extract, 1-3 parts of *Inula japonica* extract, 10-20 parts of protein-generating peptide complex, and 35-55 parts of hexapeptide.

[0011] More preferably, the composition comprises the following components in parts by weight: 15-20 parts of anti-wrinkle and soothing agent, 0.2-0.8 parts of *Cymbidium goeringii* extract, 1.5-3 parts of *Inula japonica* extract, 15-20 parts of protein-generating peptide complex, 40-55 parts of hexapeptide, and 5-10 parts of complex algae extract.

[0012] The anti-wrinkle and soothing agent is a mixture of water-soluble fibronectin and mussel extract in a mass ratio of 3:0.1. The protein-generating peptide complex is composed of acetyl tetrapeptide-2, tetrapeptide-4, pentapeptide, and time peptide in a mass ratio of 1.5:1:10:2. The compound algae extract is composed of Haematococcus pluvialis extract, Tetranychus iridis extract, and Coralaria spp. extract in a mass ratio of 235:1:40. Another technical solution adopted in this invention is a composition with wrinkle-reducing and anti-aging effects, and its application as a cosmetic additive added to toners, makeup removers, lotions, serums, creams, gels, and masks.

[0013] The beneficial effects of this invention are as follows: Based on an anti-wrinkle and soothing agent, the composition of this invention uses natural plant extracts combined with protein-derived peptide complexes to exert long-term anti-aging and wrinkle-reducing effects. Furthermore, it uses hexapeptide and marine algae extracts to achieve anti-aging effects. This combination of internal and external effects achieves long-lasting wrinkle reduction, anti-aging, and firming through multiple pathways. When applied to cosmetics, it can solve problems such as rough skin texture, loose skin, and wrinkles, and is especially suitable for dehydrated, damaged, and aging skin. Attached Figure Description

[0014] Figure 1 This is a graph showing the changes in skin elasticity characterization value R2 of subjects using Example 12 and Comparative Example 5; Figure 2 This is a before-and-after image of eye wrinkles from a subject using Example 12; Figure 3 This is a before-and-after image of eye wrinkles in a subject using Example 12 after 14 days; Figure 4This is a before-and-after image of eye wrinkles from a subject using Example 12 after 28 days; Figure 5 This is a before-and-after image of eye wrinkles from a subject using a control group of 5. Figure 6 This is a before-and-after image of eye wrinkles from a subject using a control group of 5 after 14 days. Figure 7 This is a before-and-after image of eye wrinkles from a subject using a control group of 5 after 28 days. Figure 8 This is a before-use image of a subject (allergic) using Example 12; Figure 9 This is a result image of a subject (allergic) using Example 12 after 28 days; Figure 10 This is a before-and-after image of a subject (allergic) using the product in a 5:1 ratio. Figure 11 This is a result chart of a 5-fold control group (allergic) using the drug after 28 days. Detailed Implementation

[0015] The present invention will now be described in detail with reference to specific embodiments and accompanying drawings.

[0016] This invention discloses a composition with anti-wrinkle and anti-aging effects that can be added as a cosmetic additive to cosmetics such as toners, makeup removers, lotions, serums, creams, gels, and masks. During production, the selection and preparation processes of other components in the cosmetic can be carried out according to conventional practices in the field.

[0017] The anti-wrinkle and soothing agent in the composition of this invention, as an essential high-molecular-weight glycoprotein, is widely present in tissues and tissue fluids. In the body's life activities, it participates in cell migration, adhesion, proliferation, hemostasis, tissue repair, and embryonic development. It acts as a growth factor, promoting cell proliferation, inducing epidermal cells to pass through granulation tissue, and promoting the reconstruction of the basement membrane and normal keratinization process under the epidermis. Fibronectin also participates in many pathological processes in the body. In skin wound repair and healing, fibronectin can shorten wound healing time and reduce scarring. In skin care, fibronectin (FN) has multiple uses, specifically in wrinkle reduction: wrinkles are mainly caused by the loss of collagen, elastin, and reticular fibers that support the skin. Collagen is the most important component of the extracellular matrix, and together with other extracellular matrix components, it maintains the skin's elasticity, resilience, and luster. As an important signaling molecule, fibronectin induces the expression and secretion of collagen in the extracellular matrix, promotes the secretion of hyaluronic acid and other glycoprotein molecules, and affects the directional alignment of the ECM. Therefore, fibronectin has a good effect on smoothing wrinkles. Mussel extract is rich in collagen and amino acids, which can promote skin elasticity and firmness, reducing the appearance of wrinkles and fine lines. Secondly, mussel extract contains abundant minerals and vitamins, which can nourish and replenish the skin's essential nutrients, improving its nutritional status. Furthermore, mussel extract has antioxidant properties, combating free radical damage and slowing down the skin aging process. Additionally, mussel extract has a soothing and calming effect, relieving sensitivity and inflammation, and promoting skin repair and healing. These two components work synergistically to address wrinkles and other signs of aging while repairing the skin.

[0018] Orchistem™, an active ingredient derived from *Inula japonica*, is a component of the composition of this invention. Provital's in vitro experiments have confirmed that Orchistem™ induces the production of growth factors in adipose-derived stem cells (ADSCs). These secreted growth factors reach dermal fibroblasts, increasing fibroblast proliferation and migration, and activating a positive skin regeneration process. Therefore, Orchistem™ from *Inula japonica* extract can stimulate fibroblast proliferation and activity by activating signaling communication between adipose-derived stem cells and fibroblasts, increasing collagen and elastin synthesis, and restarting the rejuvenating function of fibroblasts. *Inula japonica* extract, by scavenging excess free radicals in cells, prevents and reduces oxidative stress in the skin, significantly improving many skin health-related problems. Both of these components work synergistically, one activating skin regeneration and the other scavenging free radicals, addressing the same skin problem through different mechanisms and pathways.

[0019] The peptide complex in the composition of this invention consists of acetyl tetrapeptide-2, tetrapeptide-4, a pentapeptide, and a time peptide. Acetyl tetrapeptide-2 controls a specific gene that controls elastin production, primarily expressed before birth and within the first year after birth. Its expression decreases significantly with age, so the elastin in an adult body is essentially the protein produced initially. Acetyl tetrapeptide-2 can upregulate the expression of genes related to collagen synthesis and adhesion spots, inducing the synthesis of elastin and type I collagen, thereby increasing skin firmness and rebuilding the epidermal structure. Tetapeptide-4 significantly promotes the production of extracellular matrix proteins such as adhesion proteins and collagen, reducing fine lines and wrinkles, especially wrinkles around the eyes and crow's feet; it also increases the content of hyaluronic acid in the dermis, providing moisturizing effects and can be used as a skin conditioning agent. The pentapeptide is a water-soluble complex of five ingredients obtained through freeze-drying using inclusion technology. It significantly promotes the synthesis of collagen, hyaluronic acid, and adhesion proteins, effectively reducing various wrinkles and improving skin firmness, smoothness, and elasticity. Time peptides promote the production of fibronectin, type I, and type III collagen, and can be used as a skin conditioning agent. They effectively promote collagen and adhesion protein production, increase angiogenesis and antioxidant capacity, and stimulate the production of glucosamines (GAGs), helping the skin restore its natural self-repairing abilities.

[0020] The composite algae extracts in the composition of this invention are Haematococcus pluvialis extract, Iranian algae extract, and coral algae extract. Haematococcus pluvialis extract is rich in astaxanthin, carotene, anthocyanins, and lycopene, possessing antioxidant and anti-aging effects and can be directly applied to the skin. It also exhibits excellent inhibitory effects against *Hypericum melanogaster*, protecting the eyes, skin, and joints. Iranian algae extract helps the skin retain moisture, resist pollution, and improve skin vitality. Coral algae extract has moisturizing and hydrating effects. Rich in vitamins, minerals, and various amino acids, its extract can effectively penetrate the skin, replenishing the skin's required moisture and nutrients, helping to maintain skin hydration and improve dryness and roughness, and possessing antioxidant and anti-inflammatory properties. Coral algae is rich in various polyphenols and antioxidants, which can neutralize free radicals, reduce oxidative stress damage to the skin, and inhibit inflammatory responses, alleviating skin redness and sensitivity. The three algae, in a specific ratio, achieve optimal efficacy, maximizing the overall effect and contributing to the composition's efficacy in the most natural way.

[0021] Example 1 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, consisting of the following components: 0.5g anti-wrinkle and soothing agent, 0.05g *Cymbidium goeringii* extract, 0.05g *Inula japonica* extract, and 3g protein-generating peptide complex.

[0022] Example 2 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, consisting of the following components: 1.0g anti-wrinkle and soothing agent, 0.06g *Cymbidium goeringii* extract, 0.15g *Inula japonica* extract, and 5g protein-generating peptide complex.

[0023] Example 3 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, consisting of the following components: 2.0g anti-wrinkle and soothing agent, 0.07g *Cymbidium goeringii* extract, 0.2g *Inula japonica* extract, and 7g protein-generating peptide complex.

[0024] Example 4 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, consisting of the following components: 3.0g anti-wrinkle and soothing agent, 0.09g *Cymbidium goeringii* extract, 0.3g *Inula japonica* extract, and 9g protein-generating peptide complex.

[0025] Example 5 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, comprising the following components: 0.5g anti-wrinkle and soothing agent, 0.05g *Cymbidium goeringii* extract, 0.05g *Inula japonica* extract, 3.0g protein-generating peptide complex, and 50g hexapeptide.

[0026] Example 6 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, comprising the following components: 0.8g anti-wrinkle and soothing agent, 0.06g *Cymbidium goeringii* extract, 0.1g *Inula japonica* extract, 5.0g protein-generating peptide complex, and 55g hexapeptide.

[0027] Example 7 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, comprising the following components: 2.0g anti-wrinkle and soothing agent, 0.08g *Cymbidium goeringii* extract, 0.25g *Inula japonica* extract, 7.0g protein-generating peptide complex, and 70g hexapeptide.

[0028] Example 8 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, comprising the following components: 3.0g anti-wrinkle and soothing agent, 0.09g *Cymbidium goeringii* extract, 0.3g *Inula japonica* extract, 9.0g protein-generating peptide complex, and 80g hexapeptide.

[0029] Example 9 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, comprising the following components: 0.5g anti-wrinkle and soothing agent, 0.05g *Cymbidium goeringii* extract, 0.05g *Inula japonica* extract, 3.0g protein-generating peptide complex, 50g hexapeptide, and 0.03g complex algae extract.

[0030] Example 10 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, comprising the following components: 0.8g anti-wrinkle and soothing agent, 0.06g *Cymbidium goeringii* extract, 0.1g *Inula japonica* extract, 5.0g protein-generating peptide complex, 55g hexapeptide, and 5.0g complex algae extract.

[0031] Example 11 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, comprising the following components: 2.0g anti-wrinkle and soothing agent, 0.08g *Cymbidium goeringii* extract, 0.25g *Inula japonica* extract, 7.0g protein-generating peptide complex, 60g hexapeptide, and 15.0g complex algae extract.

[0032] Example 12 This embodiment describes a composition with wrinkle-reducing and anti-aging effects, comprising the following components: 3.0g anti-wrinkle and soothing agent, 0.09g *Cymbidium goeringii* extract, 0.3g *Inula japonica* extract, 9.0g protein-generating peptide complex, 65g hexapeptide, and 18.0g complex algae extract.

[0033] The preparation method of the compositions in Examples 1 to 12 above is as follows: weigh each component, stir and mix them evenly at room temperature to obtain the final product.

[0034] The method of using any one of the compositions in Examples 1-12 above involves adding any one of the compositions to a cosmetic product, with the composition accounting for 0.5% to 8% of the total mass of the cosmetic product. This can be done using conventional cosmetic preparation processes and excipients to prepare skincare cosmetics such as toners, makeup removers, lotions, serums, creams, gels, and masks. During production, the selection and preparation processes of other components in the cosmetic product can be carried out according to conventional practices in the field.

[0035] To demonstrate the effectiveness of the present invention, comparative examples of embodiments of the present invention are given below: Comparative Example 1 This comparative composition, which has immediate wrinkle-reducing and long-lasting anti-aging effects, consists of the following components: 1.5g anti-wrinkle and soothing agent, and 0.18g Inula japonica flower extract.

[0036] Comparative Example 2 This comparative composition, which exhibits immediate wrinkle reduction and long-lasting anti-aging effects, comprises the following components: 0.07g of *Cymbidium goeringii* extract and 5.0g of a protein-generating peptide complex. The protein-generating peptide complex consists of acetyl tetrapeptide-2, tetrapeptide-4, a pentapeptide, and a time peptide, in a mass ratio of 1.5:1:10:2.

[0037] Comparative Example 3 This comparative composition, which has immediate wrinkle-reducing and long-lasting anti-aging effects, consists of the following components: 1.5g anti-wrinkle and soothing agent, 0.18g Inula japonica extract, and 55g hexapeptide.

[0038] Comparative Example 4 This comparative composition, which exhibits immediate wrinkle reduction and long-lasting anti-aging effects, comprises the following components: 0.07g of *Cymbidium goeringii* extract, 5g of a protein-generating peptide complex, and 5g of a complex algae extract. The protein-generating peptide complex consists of acetyl tetrapeptide-2, tetrapeptide-4, a pentapeptide, and time peptide in a mass ratio of 1.5:1:10:2; the complex algae extract consists of *Haematococcus pluvialis* extract, *Echinochloa hainanensis* extract, and *Gnaphalium affine* extract in a mass ratio of 235:1:40.

[0039] Comparative Example 5 This comparative composition, which exhibits immediate wrinkle reduction and long-lasting anti-aging effects, comprises the following components: 1.5g anti-wrinkle and soothing agent, 0.18g Inula japonica extract, 55g hexapeptide, and 13g complex algae extract. The complex algae extract consists of Haematococcus pluvialis extract, Tetranychus hainanensis extract, and Coralaria spp. extract in a mass ratio of 235:1:40.

[0040] Comparative Example 6 This comparative composition, exhibiting immediate wrinkle reduction and long-lasting anti-aging effects, comprises the following components: 0.07g of *Cymbidium goeringii* extract, 5g of a protein-generating peptide complex, 55g of hexapeptide, and 13g of a complex algae extract. The protein-generating peptide complex consists of acetyl tetrapeptide-2, tetrapeptide-4, a pentapeptide, and time peptide in a mass ratio of 1.5:1:0.1:0.04. The complex algae extract consists of *Haematococcus pluvialis* extract, *Echinochloa hainanensis* extract, and *Gnaphalium affine* extract in a mass ratio of 235:1:40.

[0041] To better illustrate the advantages of the present invention, the experimental results of the technical solution provided by the present invention are given below: I. Human Safety Testing Methods The human safety testing method refers to the human skin patch test method in Chapter 7, Section 2 of the "Cosmetic Safety Technical Specifications" (2015 edition).

[0042] 1. Test Samples: The compositions of Examples 1-12 and Comparative Examples 1-6 were prepared into 5% aqueous solutions. Water was used as a control group.

[0043] 2. Participant Selection: Volunteers aged 18–60 years who meet the trial requirements will be selected as subjects. The following individuals are ineligible to be subjects: those who have used antihistamines within the past week or immunosuppressants within the past month; those who have applied any anti-inflammatory drugs to the test site within the past two months; subjects with clinically unhealed inflammatory skin diseases; patients with insulin-dependent diabetes mellitus; patients currently receiving treatment for asthma or other chronic respiratory diseases; those who have received chemotherapy for cancer within the past 6 months; patients with immunodeficiency or autoimmune diseases; breastfeeding or pregnant women; those who have undergone bilateral mastectomy and bilateral axillary lymph node dissection; those whose skin defects, such as scars, pigmentation, atrophy, port-wine stains, or other blemishes, affect the determination of test results; those participating in other clinical trials; those with highly sensitive constitutions; non-volunteer participants; and those unable to complete the prescribed content as required by the trial.

[0044] 3. Testing Method: Fifty suitable subjects were selected, including 30 females and 20 males. A qualified spot treatment device with an area not exceeding 50 mm and a depth of approximately 1 mm was used. The test sample was placed in the small chamber of the spot treatment device, with a dosage of approximately 0.020–0.025 g. The spot treatment device containing the test sample was applied to the flexor side of the subject's forearm using hypoallergenic adhesive tape, and the palm was gently pressed to ensure even application to the skin for 24 hours. Skin reactions were observed according to the standards in Table 1 at 30 minutes (after the indentation disappeared), 24 hours, and 48 hours after removing the spot treatment device, and the results were recorded. The results are shown in Table 2.

[0045] Table 1. Grading Criteria for Skin Reactions in Closed Patch Tests

[0046] Table 2 Patch Test Results

[0047] As can be seen from the patch test results in Table 2, the compositions of Examples 1-12 and Comparative Examples 1-6 achieved the expected effects, all showing negative reactions. Regarding the most basic safety requirements for cosmetics, Examples 1-12 and Comparative Examples 1-6 all meet the requirements for human safety, demonstrating feasibility and fulfilling the basic requirements of cosmetics. This also illustrates the safety of the combination of components in the compositions. Further efficacy experiments can be conducted to verify the efficacy of Examples 1-12 and Comparative Examples 1-6.

[0048] II. Antioxidant Efficacy Evaluation Test 1. Evaluation test of ABTS free radical scavenging ability Preparation of ABTS working solution: Mix 7.4 mmol / L ABTS solution with 2.6 mmol / L potassium persulfate solution in equal volumes and place in the dark for 12–16 h. Before use, dilute with 95% ethanol until the absorbance at 734 nm is 0.70 ± 0.02. This solution is the ABTS•+ free radical working solution. Sample group: Mix 0.8 mL of sample solution with 0.8 mL of ABTS•+ free radical working solution. Blank group: Replace the sample with an equal volume of distilled water. After reacting at room temperature in the dark for 10 min, measure the absorbance at 734 nm. The formula for calculating the ABTS free radical scavenging rate is shown in formula (1): ABTS radical scavenging rate (%) = [A0 - A] x / A0]A0×100%(1) In the formula, A0 is blank, A x For the sample.

[0049] 2. Hydroxyl radical scavenging capacity evaluation test Add 3 mL of 2 mmol / L FeSO4 and 3 mL of 1 mmol / L H2O2 to a 25 mL cuvette, shake well, then add 3 mL of 6 mmol / L salicylic acid and shake well. Heat in a water bath at 37℃ for 15 min and then remove and measure the absorbance. Add a certain concentration of sample solution to each, shake well, continue heating in a water bath for 15 min, and then remove and measure the absorbance.

[0050] The formula for calculating the hydroxyl radical scavenging rate is shown in equation (2): Hydroxyl radical scavenging rate (%) = [A0-A] x / A0]A0×100%(2) In the formula, A0, A x The absorbance values ​​are: the absorbance of the reaction system before the sample was added, the absorbance of the system after the sample was used to remove hydroxyl radicals, and the absorbance of the system after the sample solvent was used to remove hydroxyl radicals.

[0051] Table 3 Free radical scavenging rate of compositions with different ratios

[0052] As can be clearly seen from the data in Table 3, all embodiments exhibit higher scavenging rates for ABTS and hydroxyl radicals than all comparative examples. The highest ABTS radical scavenging rate reached 98.6%, and the highest hydroxyl radical scavenging rate reached 86.9%, almost double that of the comparative examples. Among the comparative examples, the highest free radical scavenging rate was 56.8%, and the highest hydroxyl radical scavenging rate was 46.1%. Free radical scavengers or antioxidants can provide electrons and inhibit oxidation. By absorbing free radicals while protecting cells from attack, the purpose of free radical scavenging is achieved, reducing wrinkle formation and thus slowing down the skin aging process. The free radical scavenging rate also demonstrates the excellent antioxidant capacity of this invention in vitro, making it promising for subsequent human trials and related experimental verification.

[0053] III. Human Skin Moisturizing and Anti-aging Effect Test 1. Skin moisture content and skin moisture loss (TEWL) test The test was conducted according to T / ZHCA 003-2018, the test method for the effect of cosmetics on transepidermal moisture loss.

[0054] (1) Test samples: The test samples were prepared by adding Examples 1-12 and Comparative Examples 1-6 to a common emulsion external matrix, wherein the composition accounted for 5% of the total mass of the emulsion.

[0055] (2) Subject selection Number of participants: 220 valid participants, 10 in each group.

[0056] Subject eligibility: Volunteers aged 18–60 years who meet the trial requirements. Subjects with the following conditions will not be selected: those who have used antihistamines within the past week or immunosuppressants within the past month; those who have applied any anti-inflammatory drugs to the test site within the past two months; those with clinically uncured inflammatory skin diseases; patients with insulin-dependent diabetes mellitus; patients currently receiving treatment for asthma or other chronic respiratory diseases; those who have received chemotherapy for cancer within the past six months; those with immunodeficiency or autoimmune diseases; breastfeeding or pregnant women; those who have undergone bilateral mastectomy and bilateral axillary lymph node dissection; those whose skin blemishes such as scars, pigmentation, atrophy, port-wine stains, or other imperfections at the test site affect the determination of trial results; those participating in other clinical trials; those with highly sensitive constitutions; non-volunteer participants; or those unable to complete the prescribed tasks as required by the trial.

[0057] (3) Measurement steps Testing instrument: Tewameter (transdermal moisture loss tester) Measurement Method: Subjects were randomly assigned to either the left or right forearm flexor side as the sample application side and the control side, ensuring statistical balance between the two sides. Before each measurement, the test area was cleaned with water and then patted dry with a lint-free absorbent paper towel. The subject sat quietly in a standardized testing environment for at least 20 minutes, without drinking water or beverages, with the forearm exposed, relaxed, and avoiding touching the test area. The test was performed according to the instructions for the transepidermal water loss (TEWL) tester. When measuring the TEWL value, the instrument probe must be placed in the center of the test area. During measurement, the average of 10 consecutive points on the TEWL value curve was taken [its standard deviation should be less than 0.2 g / (h·m)]. 2 [Measurement values] are used as the initial values; first, the initial values ​​of each region (before using the sample) are measured, and then the TEWL values ​​of the test and control regions are measured after a set time. The set time is 7 days and 8 days. The test for the same subject must be performed using the same instrument by the same tester. If the subject experiences an adverse skin reaction during sample use, the test should be stopped immediately, and the subject should receive appropriate medical treatment. Adverse reactions should be recorded.

[0058] (4) Calculation results Descriptive statistics: Descriptive statistics are performed on the measured values ​​of each test area, including quantity, mean, standard deviation, minimum, median, and maximum.

[0059] Difference analysis: The rate of change of the average TEWL values ​​for both the sample application side and the control side was calculated. This rate of change was then used to statistically analyze the differences between the sample application area and the control area at different measurement time points. If the test data were normally distributed, a t-test was used for statistical analysis; if the test data were not normally distributed, a rank-sum test was used. All statistical methods employed two-tailed tests with a significance level of α = 0.05.

[0060] (5) Result determination Positive result: Compared with Comparative Example 2, the rate of change of TEWL value on the applied side of the sample was significantly different (P < 0.05), indicating that the tested sample has a promoting effect on the recovery of transepidermal water loss in skin with damaged barrier.

[0061] Negative result: Compared with Comparative Example 2, there was no significant difference in the rate of change of TEWL value on the applied side of the sample (P≥0.05), indicating that the tested sample had no promoting effect on the recovery of transepidermal water loss in skin with damaged barrier. The test results are shown in Table 4.

[0062] Table 4. Transdermal water loss (TEWL) values ​​of test samples at different times

[0063] Note: Under the test conditions, the higher the TEWL value, the greater the transepidermal water loss per unit time and per unit cross-sectional area; the lower the TEWL value, the less the transepidermal water loss per unit time and per unit cross-sectional area.

[0064] Statistical analysis of the experimental results in Table 4 showed that both the comparative and example samples exhibited repair effects to varying degrees. However, the rate of change in transepidermal water loss (TEWL) values ​​in all experimental examples was significantly higher than that in the comparative samples. In Example 12, after 7 days of use, the TWL value decreased by 21.88% compared to before use; after 14 days of use, the TWL value decreased by 32.13% compared to before use, demonstrating a highly significant difference. This indicates that the composition already had a repairing effect after 7 days of use. While the comparative samples showed a certain rate of decrease, the degree of decrease was not significantly different. The control group data also showed almost no repairing effect. The repairing effects of all experimental examples were superior to those of the comparative samples, proving that the synergistic effect of various substances in the composition greatly enhanced the repairing effect. The enhanced transepidermal water retention also prevented, to some extent, the causes of wrinkles such as dryness.

[0065] 2. Skin elasticity test: Twenty participants aged 30-60 years with wrinkles at the outer corners of their eyes (wrinkle grades 1-7 on both sides, with the same grade on both sides) were selected, with a 50 / 50 male-to-female ratio. No products (cosmetics, topical medications, or oral health supplements) were used on the test area for 15 days prior to the test. Before the test, participants were required to cleanse their faces and, 2 hours later, sit quietly in a climate-controlled room (22±1℃, 50% relative humidity) for at least 20 minutes, maintaining a relaxed state.

[0066] In the experiment, half-face testing was conducted using a randomized table, with one side serving as the sample and the other as the control. Skin elasticity was tested on both cheeks and corners of the eyes of volunteers, and the average value of three tests was taken as the skin elasticity data (d0) for the left and right corners of the eyes. After the test, volunteers were required to use the product as instructed, applying the sample to one side of the face and the control product to the other side. The product was used continuously for 63 days, and participants were required to return for follow-up on days 7 (d7), 14 (d14), 21 (d21), 28 (d28), 35 (d35), 42 (d42), 49 (d49), 56 (d56), and 69 (d69).

[0067] The skin elasticity index R² is used to represent the skin's ability to recover after being stretched during testing. Younger, more elastic skin has a larger R²; conversely, older, less elastic skin has a smaller R². Figure 1 shows the changes in the skin elasticity index R². The figure shows the trends of R² changes after 69 days of continuous use of the composition of this invention and the control sample. It can be seen that the change in R² after 69 days of use with the control sample was not significant, with an increase of approximately 7.9% after 28 days and only about 20.6% after 63 days. The increase in R² after 69 days of use with the composition of this invention was more significant, with an increase of approximately 25.8% after 28 days and a 45.2% increase after 69 days. This demonstrates that the comparative test results prove that adding the composition of this invention has a significant effect on improving skin elasticity, thus showing that the composition of this invention has a good effect on improving skin elasticity and has good anti-aging effects.

[0068] To more intuitively demonstrate its effects, facial images of one subject from Example 12 and Comparative Example 5 were selected for illustration, such as... Figures 2-7 As shown in the two sets of effect images, it is clear that in the most prominent eye wrinkle area, after using the sample of Example 12 for 14 and 28 days, the wrinkle area of ​​the subjects was significantly reduced compared to before use, and the reduction effect was more pronounced with longer use. This further illustrates that the composition of the present invention, with the synergistic effect of multiple components, can minimize the degree of facial wrinkles and achieve wrinkle removal to a certain extent. Meanwhile, the effect images of using the sample of Comparative Example 5 for 14 and 28 days (…) Figure 5-7 As can be seen, although the wrinkle area at 28 days was not as large or as severe as at 0 days, there were still large areas of wrinkles around the eyes with obvious lines. This indicates that the comparative composition did not have a strong wrinkle-removing function under non-irritating conditions, and there was a clear contrast with the effect of the example.

[0069] Figure 8 , Figure 9 , Figure 10 , Figure 11 The results show the facial redness of volunteers before and after using Example 12 and Comparative Example 5 in an allergic state. Fewer red areas indicate a repairing effect of the composition on allergies, while more red areas or no change indicates no repairing effect. Figure 8 , Figure 9 , Figure 10 , Figure 11 It is obvious that Figure 9 The redness on the face has significantly decreased, and Figure 11 The fact that the redness on the face did not change significantly compared to before use directly demonstrates the repairing effect or the inhibitory effect on allergies of the composition of the present invention.

[0070] In summary, this invention, through the synergistic effect of anti-wrinkle and soothing agents, *Cymbidium goeringii* extract, *Inula japonica* extract, protein-generating peptide complex, hexapeptide, and complex algae extracts, combined with results from human safety tests, water retention tests, and skin elasticity assessments, shows that the compositions of Examples 1-12 have high anti-wrinkle and anti-aging effects. In contrast, the compositions of Comparative Examples 1-6 have less noticeable moisturizing effects or cannot maintain moisture for long. Furthermore, the efficacy of Comparative Examples 1-6 in in vitro antioxidant experiments and human wrinkle-reducing experiments is far lower than that of Examples 1-12, and their effects fall far short of the required intensity. This demonstrates that the synergistic effect of the substances in this invention is necessary and effective, addressing skin aging problems from multiple perspectives and through multiple pathways. While ensuring human safety, it addresses wrinkles and long-term aging from multiple angles, achieving long-lasting anti-aging and effective wrinkle reduction.

Claims

1. A composition having wrinkle-reducing and anti-aging effects, characterized in that, The composition consists of the following components: 3.0g anti-wrinkle and soothing agent, 0.09g *Cymbidium goeringii* extract, 0.3g *Inula japonica* extract, and 9g protein-generating peptide complex; The anti-wrinkle and soothing agent is a mixture of water-soluble fibronectin and mussel extract in a mass ratio of 3:0.1; the protein-generating peptide complex is a mixture of acetyl tetrapeptide-2, tetrapeptide-4, pentapeptide, and time peptide in a mass ratio of 1.5:1:10:2.

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