A multilayer encapsulated nano-lyophilized formulation, its preparation method, and its skincare application.

Abstract

This invention discloses a multilayer encapsulated lyophilized nano-formulation, its preparation method, and its skincare application, belonging to the field of encapsulation technology. The process includes the following steps: mixing an oil-soluble component and a vegetable oil, vortexing to obtain an oil phase; mixing whey protein isolate powder, deionized water, and tea polyphenols, stirring until homogeneous, adjusting the pH, continuing stirring, adding a water-soluble component, mixing, and stirring to obtain an aqueous phase; mixing the aqueous and oil phases, ultrasonically dispersing, adding a sodium alginate aqueous solution, adjusting the pH, ultrasonically dispersing, then adding a carboxymethyl chitosan aqueous solution, continuing stirring, then adding a calcium chloride solution, continuing stirring, rotary evaporation to remove water, and freeze-drying to obtain the multilayer encapsulated lyophilized nano-formulation. This method achieves efficient encapsulation of both oil-soluble and water-soluble natural active ingredients while avoiding the reduction in activity caused by environmental damage to the natural active ingredients.

Description

Technical Field

[0001] This invention belongs to the field of encapsulation technology, specifically relating to a multilayer encapsulated nano-lyophilized formulation, its preparation method, and its skincare application. Background Technology

[0002] Most natural active ingredients are structurally sensitive and easily affected by light, temperature, oxygen, and other components in cosmetic formulations, leading to oxidative degradation and loss of activity. This not only reduces product efficacy but may also cause skin irritation due to ingredient deterioration. To address these issues, encapsulation technology has become a key method in the cosmetics industry to improve the stability and bioavailability of active ingredients. Existing encapsulation technologies, such as monolayer liposome encapsulation and microcapsule encapsulation, while able to isolate active ingredients from external environmental damage to a certain extent, still have significant drawbacks: monolayer encapsulation structures have poor compatibility with components of different polarities, making it difficult to simultaneously achieve efficient encapsulation of both oil-soluble and water-soluble components; some encapsulation carriers also have poor biocompatibility, potentially affecting the product's feel and safety. Therefore, developing multilayer encapsulated lyophilized nanoparticle formulations that combine multi-component compatibility, high stability, and efficient delivery capabilities has become a research hotspot in the skincare field, and is of great significance for promoting the efficient application of natural active ingredients in cosmetics and enhancing the core competitiveness of skincare products. Summary of the Invention

[0003] The purpose of this invention is to provide a multilayer encapsulated nano-lyophilized formulation, its preparation method, and its skincare application, which can achieve efficient encapsulation of both oil-soluble and water-soluble natural active ingredients while avoiding the problem of reduced activity of natural active ingredients due to damage from the external environment.

[0004] The objective of this invention can be achieved through the following technical solutions: A method for preparing a multilayer encapsulated nano-lyophilized formulation includes the following steps: S1. Take the oil-soluble components and vegetable oil, mix them, and vortex until homogeneous to obtain the oil phase; S2. Take whey protein isolate, deionized water and tea polyphenols, mix them, stir well, adjust the pH to 8-10, continue stirring, add water-soluble ingredients and mix, stir to obtain the aqueous phase; S3. Take the aqueous phase and oil phase, mix them, disperse them by ultrasonication, add sodium alginate aqueous solution, adjust the pH to 4-6, disperse by ultrasonication, then add carboxymethyl chitosan aqueous solution, continue stirring, then add calcium chloride solution, continue stirring, remove water by rotary evaporation, and freeze dry to obtain the multilayer embedded nano-lyophilized formulation.

[0005] As a preferred embodiment of the present invention, in step S1, the vegetable oil is selected from at least one of sunflower oil, soybean oil, olive oil, flaxseed oil and palm oil.

[0006] As a preferred embodiment of the present invention, in step S1, the mass ratio of the oil-soluble component to the vegetable oil is 2-3:18-26.

[0007] As a preferred embodiment of the present invention, in step S2, the mass ratio of the whey protein isolate, deionized water, tea polyphenols, and water-soluble components is 1-2:30-40:8-12:4-6.

[0008] As a preferred embodiment of the present invention, in step S3, the volume ratio of the aqueous phase, oil phase, sodium alginate aqueous solution, carboxymethyl chitosan aqueous solution, and calcium chloride solution is 7-9:1-3:10:0.2-0.3:0.01-0.02.

[0009] As a preferred technical solution of the present invention, in step S3, the freeze drying refers to drying at -45 to -35°C and a vacuum degree of 100-120 Pa for 4-5 hours.

[0010] A multilayer encapsulated nano-lyophilized formulation prepared using the above-described preparation method.

[0011] The application of a multi-layer encapsulated nano-lyophilized formulation in skincare, wherein the application refers to adding the multi-layer encapsulated nano-lyophilized formulation to a cosmetic formulation, wherein the multi-layer encapsulated nano-lyophilized formulation comprises 5%-25% by mass in the cosmetic formulation.

[0012] As a preferred embodiment of the present invention, the oil-soluble components include Terminalia chebula extract and Rosa canina fruit oil; the mass ratio of Terminalia chebula extract and Rosa canina fruit oil is 3-5:6-8.

[0013] As a preferred embodiment of the present invention, the water-soluble components include extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sibiricum*, and extracts of *Cladosporium cladosporium*; the mass ratio of the extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sibiricum*, and extracts of *Cladosporium cladosporium* is 4-6:5-9:1-3.

[0014] The following is a summary of some of the raw materials used in this application: Terminalia arjuna extract, purchased from Shanghai Zhenzhun Biotechnology Co., Ltd. Rosa Canina Fruit Oil, purchased from Beijing Lingbao Technology Co., Ltd. FICUS PUMILA HYPANTHIUM EXTRACT, purchased from Xi'an Huilin Biotechnology Co., Ltd. Eryngium maritimum callus culture filtrate, purchased from Guangzhou Gaosheng Chemical Co., Ltd. Cladospora novae-caledoniae extract, purchased from Zhuhai Haixin Biotechnology Co., Ltd. Tea polyphenols, CAMELLIA SINENSIS POLYPHENOLS, are isolated from green tea.

[0015] The beneficial effects of this invention are: This invention utilizes the covalent bonding of plant oil components, whey protein isolate, and tea polyphenols to form an amphiphilic structure that possesses both lipophilic and hydrophilic groups, enabling it to act as a molecular emulsifier and achieve uniform dispersion of the oil and aqueous phases. Simultaneously, the molecular network formed by the covalent composite layer, combined with the subsequent multilayer encapsulation of sodium alginate, carboxymethyl chitosan, and calcium ions, forms a protective barrier, isolating the encapsulated natural active ingredients from direct damage by the external environment. The strong covalent bonds make the composite layer encapsulation structure more stable, further protecting the activity of the encapsulated material.

[0016] Furthermore, the extracts of Terminalia chebula, Rosa canina fruit oil, Ficus hirta syconium extract, Erycibe obtusifolia callus culture filtrate, and Cladosporium cladosporium extract are combined for skincare to synergistically improve skin aging. Through the targeted complementarity and synergistic mechanism of each skincare ingredient, excellent wrinkle reduction and anti-aging effects are ultimately achieved. Detailed Implementation

[0017] To further illustrate the technical means and effects of the present invention in achieving its intended purpose, the following detailed description of the specific implementation methods, structures, features, and effects of the present invention, in conjunction with embodiments, is provided below.

[0018] Example 1 A method for preparing a multilayer encapsulated nano-lyophilized formulation includes the following steps: S1. Take the oil-soluble components and olive oil, mix them, and vortex until homogeneous to obtain the oil phase; S2. Mix whey protein isolate, deionized water and tea polyphenols, stir well, adjust pH to 8, continue stirring for 10 hours, add water-soluble components and mix, stir for 20 minutes to obtain the aqueous phase; S3. Take the aqueous phase and oil phase, mix them, and ultrasonically disperse for 4 min. Add sodium alginate aqueous solution, adjust the pH to 4, ultrasonically disperse for 30 s, then add carboxymethyl chitosan aqueous solution, continue stirring for 10 min, then add calcium chloride solution, continue stirring for 5 min, rotary evaporate to remove water, and freeze dry to obtain the multilayer embedded nano-freeze-dried formulation. In step S1, the oil-soluble components include Terminalia chebula extract and Rosa canina fruit oil; the mass ratio of Terminalia chebula extract to Rosa canina fruit oil is 3:6. In step S1, the mass ratio of the oil-soluble component to the vegetable oil is 2:18; In step S2, the water-soluble components include extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium*; the mass ratio of the extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium* is 4:5:1. In step S2, the mass ratio of whey protein isolate, deionized water, tea polyphenols, and water-soluble components is 1:30:8:4. In step S3, the volume ratio of the aqueous phase, oil phase, sodium alginate aqueous solution, carboxymethyl chitosan aqueous solution, and calcium chloride solution is 7:3:10:0.2:0.01; In step S3, the mass fraction of the sodium alginate aqueous solution is 1%; In step S3, the mass fraction of the carboxymethyl chitosan aqueous solution is 1%. In step S3, the mass fraction of the calcium chloride solution is 0.13%. In step S3, freeze drying refers to drying at -45°C and a vacuum of 100 Pa for 4 hours.

[0019] Example 2 A method for preparing a multilayer encapsulated nano-lyophilized formulation includes the following steps: S1. Take the oil-soluble components and olive oil, mix them, and vortex until homogeneous to obtain the oil phase; S2. Take whey protein isolate, deionized water and tea polyphenols, mix them, stir evenly, adjust the pH to 9, continue stirring for 11 hours, add water-soluble components and mix, stir for 25 minutes to obtain the aqueous phase; S3. Take the aqueous phase and oil phase, mix them, and ultrasonically disperse for 5 min. Add sodium alginate aqueous solution, adjust the pH to 5, ultrasonically disperse for 40 s, then add carboxymethyl chitosan aqueous solution, continue stirring for 15 min, then add calcium chloride solution, continue stirring for 8 min, remove water by rotary evaporation, and freeze dry to obtain the multilayer embedded nano-freeze-dried formulation. In step S1, the oil-soluble components include Terminalia chebula extract and Rosa canina fruit oil; the mass ratio of Terminalia chebula extract to Rosa canina fruit oil is 4:7. In step S1, the mass ratio of the oil-soluble component to the vegetable oil is 2.5:22; In step S2, the water-soluble components include extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium*; the mass ratio of the extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium* is 5:7:2. In step S2, the mass ratio of whey protein isolate, deionized water, tea polyphenols, and water-soluble components is 1.5:35:10:5. In step S3, the volume ratio of the aqueous phase, oil phase, sodium alginate aqueous solution, carboxymethyl chitosan aqueous solution, and calcium chloride solution is 8:2:10:0.25:0.015. In step S3, the mass fraction of the sodium alginate aqueous solution is 1%; In step S3, the mass fraction of the carboxymethyl chitosan aqueous solution is 1%. In step S3, the mass fraction of the calcium chloride solution is 0.13%. In step S3, freeze drying refers to drying at -40°C and a vacuum of 110 Pa for 4.5 hours.

[0020] Example 3 A method for preparing a multilayer encapsulated nano-lyophilized formulation includes the following steps: S1. Take the oil-soluble components and olive oil, mix them, and vortex until homogeneous to obtain the oil phase; S2. Take whey protein isolate, deionized water and tea polyphenols, mix them, stir evenly, adjust the pH to 10, continue stirring for 12 hours, add water-soluble components and mix, stir for 30 minutes to obtain the aqueous phase; S3. Take the aqueous phase and oil phase, mix them, and ultrasonically disperse for 6 min. Add sodium alginate aqueous solution, adjust the pH to 6, ultrasonically disperse for 50 s, then add carboxymethyl chitosan aqueous solution, continue stirring for 20 min, then add calcium chloride solution, continue stirring for 10 min, remove water by rotary evaporation, and freeze dry to obtain the multilayer embedded nano-freeze-dried formulation. In step S1, the oil-soluble components include Terminalia chebula extract and Rosa canina fruit oil; the mass ratio of Terminalia chebula extract to Rosa canina fruit oil is 5:8. In step S1, the mass ratio of the oil-soluble component to the vegetable oil is 3:26; In step S2, the water-soluble components include extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium*; the mass ratio of the extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium* is 6:9:3. In step S2, the mass ratio of whey protein isolate, deionized water, tea polyphenols, and water-soluble components is 2:40:12:6. In step S3, the volume ratio of the aqueous phase, oil phase, sodium alginate aqueous solution, carboxymethyl chitosan aqueous solution, and calcium chloride solution is 9:1:10:0.3:0.02. In step S3, the mass fraction of the sodium alginate aqueous solution is 1%; In step S3, the mass fraction of the carboxymethyl chitosan aqueous solution is 1%. In step S3, the mass fraction of the calcium chloride solution is 0.13%. In step S3, freeze drying refers to drying at -35°C and a vacuum of 120 Pa for 5 hours.

[0021] Comparative Example 1 A method for preparing a multilayer encapsulated nano-lyophilized formulation includes the following steps: S1. Take the oil-soluble components and olive oil, mix them, and vortex until homogeneous to obtain the oil phase; S2. Take deionized water and water-soluble components, mix them, and stir for 25 minutes to obtain the aqueous phase; S3. Take the aqueous phase and oil phase, mix them, and ultrasonically disperse for 5 min. Add sodium alginate aqueous solution, adjust the pH to 5, ultrasonically disperse for 40 s, then add carboxymethyl chitosan aqueous solution, continue stirring for 15 min, then add calcium chloride solution, continue stirring for 8 min, remove water by rotary evaporation, and freeze dry to obtain the multilayer embedded nano-freeze-dried formulation. In step S1, the oil-soluble components include Terminalia chebula extract and Rosa canina fruit oil; the mass ratio of Terminalia chebula extract to Rosa canina fruit oil is 4:7. In step S1, the mass ratio of the oil-soluble component to the vegetable oil is 2.5:22; In step S2, the water-soluble components include extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium*; the mass ratio of the extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium* is 5:7:2. In step S2, the mass ratio of deionized water to water-soluble components is 46.5:5; In step S3, the volume ratio of the aqueous phase, oil phase, sodium alginate aqueous solution, carboxymethyl chitosan aqueous solution, and calcium chloride solution is 8:2:10:0.25:0.015. In step S3, the mass fraction of the sodium alginate aqueous solution is 1%; In step S3, the mass fraction of the carboxymethyl chitosan aqueous solution is 1%. In step S3, the mass fraction of the calcium chloride solution is 0.13%. In step S3, freeze drying refers to drying at -40°C and a vacuum of 110 Pa for 4.5 hours.

[0022] Comparative Example 2 A method for preparing a multilayer encapsulated nano-lyophilized formulation includes the following steps: S1. Take the oil-soluble components and olive oil, mix them, and vortex until homogeneous to obtain the oil phase; S2. Take whey protein isolate, deionized water and tea polyphenols, mix them, stir evenly, adjust the pH to 9, continue stirring for 11 hours, add water-soluble components and mix, stir for 25 minutes to obtain the aqueous phase; S3. Take the aqueous phase and oil phase, mix them, ultrasonically disperse for 5 min, add sodium alginate aqueous solution, adjust the pH to 5, ultrasonically disperse for 40 s, then add carboxymethyl chitosan aqueous solution, continue stirring for 15 min, remove water by rotary evaporation, freeze dry to obtain the multilayer embedded nano-lyophilized formulation. In step S1, the oil-soluble components include Terminalia chebula extract and Rosa canina fruit oil; the mass ratio of Terminalia chebula extract to Rosa canina fruit oil is 4:7. In step S1, the mass ratio of the oil-soluble component to the vegetable oil is 2.5:22; In step S2, the water-soluble components include extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium*; the mass ratio of the extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium sambac*, and extracts of *Cladosporium cladosporium* is 5:7:2. In step S2, the mass ratio of whey protein isolate, deionized water, tea polyphenols, and water-soluble components is 1.5:35:10:5. In step S3, the volume ratio of the aqueous phase, oil phase, sodium alginate aqueous solution, carboxymethyl chitosan aqueous solution, and calcium chloride solution is 8:2:10:0.25:0.015. In step S3, the mass fraction of the sodium alginate aqueous solution is 1%; In step S3, the mass fraction of the carboxymethyl chitosan aqueous solution is 1%. In step S3, freeze drying refers to drying at -40°C and a vacuum of 110 Pa for 4.5 hours.

[0023] Comparative Example 3 The only difference from Example 3 is that the components do not contain the extract of Terminalia chebula, and the reduced amount is allocated to the Rosa canis fruit oil in proportion to the number of parts, while the other components and their amounts remain unchanged.

[0024] Comparative Example 4 The only difference from Example 3 is that the components do not contain Rosa canis fruit oil, and the reduced amount is allocated to the Terminalia chebula extract in proportion to the number of parts, while the other components and their amounts remain unchanged.

[0025] Comparative Example 5 The only difference from Example 3 is that the components do not contain the extract of creeping fig syconium inflorescence. The reduced amount is allocated to the filtrate of callus culture of Erycibe obtusifolia and the extract of Cladosporium cladosporium, while the other components and amounts remain unchanged.

[0026] Comparative Example 6 The only difference from Example 3 is that the components do not contain the callus culture filtrate of *Erycibe obtusifolia*. The reduced amount is allocated to the extracts of *Ficus hirta* and *Cladosporium cladoceratum* in proportion to the number of parts, while the other components and their amounts remain unchanged.

[0027] Comparative Example 7 The only difference from Example 3 is that the components do not contain the extract of Cladosporium cladoceratum. The reduced amount is allocated to the extract of Syringa vulgaris and the filtrate of Erycibe obtusifolia callus culture in proportion to the number of parts. The other components and their amounts remain unchanged.

[0028] Performance testing An essence emulsion is prepared from the following components in weight percentage: 0.5% glyceryl stearate, 0.1% tocopheryl acetate, 1% polydimethylsiloxane, 2% isononyl isononanoate, 0.2% cetearyl alcohol, 0.02% ubiquinone, 0.5% xanthan gum, 0.1% polydimethylsiloxane / vinyl polydimethylsiloxane crosspolymer, 5% glycerin, 5% 1,3-butanediol, 1% nano-lyophilized formulations obtained from Examples 1-3 and Comparative Examples 1-7 respectively, and the balance being deionized water.

[0029] 1) Subjects: 100 volunteers aged 48-65 with wrinkles and healthy skin were randomly divided into 10 groups of 10 each.

[0030] 2) Use of the product: Use the samples in Examples 1-3 and Comparative Examples 1-7 once in the morning and once in the evening for 30 days.

[0031] 3) Usage: After cleansing your face in the morning and evening, take an appropriate amount of sample and apply it evenly to your face. Gently massage until absorbed. Each group of volunteers used the samples prepared in Examples 1-3 and Comparative Examples 1-7, respectively.

[0032] 4) Anti-aging test: Specifically, before the trial began, the participants' skin wrinkles and skin elasticity were measured. After using the serum product for 30 days, the participants' skin wrinkles and skin elasticity were measured again. Skin wrinkles were measured using a skin image analysis system (skinvisiometersv600), and skin elasticity was measured using a cutometer dualMPa580.

[0033] Facial wrinkle improvement rate (%) = (wrinkle parameter value after using serum - wrinkle parameter value before using serum) / wrinkle parameter value before using serum. The wrinkle improvement rate is calculated by formulating the rate of change and then taking the positive value. Facial elasticity improvement rate (%) = (elasticity value after using serum - elasticity value before using serum) / elasticity value before using serum. The elasticity improvement rate is calculated by formulating the rate of change and taking a positive value. The samples prepared in Examples 1-3 and Comparative Examples 1-7 of this invention were tested for anti-aging, and the results are shown in Table 1.

[0034] Table 1 As shown in Table 1, the multi-layer encapsulated nano-lyophilized formulations prepared in Examples 1-3 of this application have better anti-aging effects compared with the comparative samples.

[0035] This product utilizes a blend of extracts from Terminalia chebula, Rosa canina fruit oil, Ficus hirta syconium inflorescence extract, Erycibe obtusifolia callus culture filtrate, and Cladosporium cladosporium extract for skincare. It synergistically improves skin aging through the targeted complementarity and mechanistic synergy of the various skincare ingredients. Addressing the core aspects of skin aging (oxidative damage, loss of supporting structures, decreased cell vitality, and loosening of the epidermal-dermal junction), it forms a four-dimensional closed loop of protection, repair, regeneration, and consolidation, ultimately achieving excellent wrinkle reduction and anti-aging effects. Its synergistic principle can be seen from the complementary and enhanced effects between the ingredients in the following four aspects: 1) Utilizing the synergistic effects of extracts from Terminalia chebula, Cladosporium cladodes, and Ficus hirta to provide antioxidant stress protection, blocking damage at the source of aging, scavenging free radicals, and reducing collagen degradation and cell aging, these three ingredients work together to block damage through dual antioxidant and collagen protection. Flavonoids and tannins from *Terminalia catappa* extract, phycobiliproteins and polysaccharides from *Cladosporium cladosporum*, and polyphenols and flavonoids from *Ficus pumila* are all potent antioxidants that can target and eliminate different types of ROS (such as superoxide anions and hydroxyl radicals). Their action sites are complementary (cell membrane, cytoplasm, mitochondria), resulting in higher elimination efficiency than single antioxidants. *Cladosporium cladosporum* inhibits free radical production at its source; polyphenols from *Ficus pumila* inhibit lipid peroxidation, protecting unsaturated fatty acids in skin cell membranes from oxidative damage; and extracts from *Terminalia catappa* and *Ficus pumila* also inhibit matrix metalloproteinases (MMPs) activity, preventing existing collagen from being oxidized and damaged, thus laying the foundation for subsequent repair.

[0036] 2) The synergistic effect of extracts from Terminalia chebula, syconium syringae, and callus culture filtrate from Erycibe obtusifolia can repair the supporting structure of the dermis, fill the core of wrinkles, rebuild the collagen and elastic fiber network, and restore the skin's support.

[0037] The extract of Terminalia chebula directly promotes collagen synthesis by fibroblasts by activating the TGF-β / Smad signaling pathway; the polysaccharides and flavonoids of Ficus pumila extract synergistically promote fibroblast proliferation and increase the total secretion of collagen and elastin; Terminalia chebula and Ficus pumila continue to exert their MMP inhibitory effects, preventing the decomposition of newly synthesized collagen and improving repair efficiency; the callus filtrate of Erycibe obtusifolia is rich in growth factor-like active ingredients, which can mimic the effects of epidermal growth factor and fibroblast growth factor, promote the cross-linking and network reconstruction of elastic fibers, and improve the support of the dermis in both quantity and quality, allowing the skin to regain firmness and elasticity from the inside.

[0038] 3) The combination of Rosa canis fruit oil, Erycibe spp. callus filtrate, and Cladosporium cladosporium extract promotes the activation and renewal of epidermal cells, improves skin metabolism and aging, accelerates epidermal cell turnover, and makes the skin smooth and vibrant.

[0039] The active ingredients (such as adenosine and amino acids) in the callus tissue of *Erycibe obtusifolia* can activate the AMPK energy metabolism pathway and increase cellular ATP production; the polysaccharides of *Cladosporium cladosporum* provide nutrients to cells and further enhance cell vitality; the vitamin A (a precursor to retinol) in *Rosa canis* fruit oil can be converted into retinol, activating the proliferation and differentiation of keratinocytes, accelerating the shedding of dead skin cells, promoting the generation of new cells, and making the skin smoother; the unsaturated fatty acids in *Rosa canis* fruit oil can replenish epidermal lipids and repair the skin barrier; the polysaccharides of *Cladosporium cladosporum* can form a moisturizing film on the skin surface, working synergistically with lipids to lock in moisture, allowing epidermal cells to proliferate and differentiate better in a hydrated environment.

[0040] 4) The combination of Rosa canis fruit oil, Cremastrae syconium inflorescence extract, and Erycibe obtusifolia callus filtrate enhances the anti-aging effect, strengthens the connection between the epidermis and dermis, and improves the overall skin structure and elasticity.

[0041] The vitamin C in Rosa canis fruit oil is an essential coenzyme for collagen synthesis, which can promote the synthesis of collagen and laminin; the polysaccharides in fig extract can bind to adhesion proteins, enhance the adhesion between epidermal basal cells and dermis, and make the skin structure tighter; the active ingredients in callus tissue of Erycibe obtusifolia can regulate the expression of DEJ-related genes, repair damaged DEJ structure, and form a complete support system.

[0042] The five ingredients are not simply added together, but form a synergistic effect across multiple targets and the entire process: at the antioxidant level, they reduce collagen damage, easing the burden on the repair level; at the dermal repair level, they provide core support, giving the skin after epidermal renewal an elastic foundation; at the DEJ reconstruction level, they enable the epidermis and dermis to work together, enhancing the overall effect. By working synergistically across four dimensions—blocking damage, repairing and supporting, activating metabolism, and consolidating structure—it covers the core mechanisms of skin aging, achieving a comprehensive effect of wrinkle reduction, anti-aging, and improved elasticity. Moreover, the various ingredients complement each other without antagonism, synergistically amplifying the efficacy of individual ingredients.

[0043] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make some modifications or alterations to the above-disclosed technical content to create equivalent embodiments without departing from the scope of the present invention. Any simple modifications, equivalent changes and alterations made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention shall still fall within the scope of the present invention.

Claims

1. A method for preparing a multilayer encapsulated nano-lyophilized formulation, characterized in that, Includes the following steps: S1. Take the oil-soluble component and vegetable oil, mix them, and vortex until homogeneous to obtain the oil phase; the mass ratio of the oil-soluble component to the vegetable oil is 2-3:18-26. S2. Take whey protein isolate, deionized water and tea polyphenols, mix them, stir evenly, adjust the pH to 8-10, continue stirring, add water-soluble ingredients and mix, stir to obtain the aqueous phase; the mass ratio of whey protein isolate, deionized water, tea polyphenols and water-soluble ingredients is 1-2:30-40:8-12:4-6. S3. Mix the aqueous phase and oil phase, disperse ultrasonically, add sodium alginate aqueous solution, adjust the pH to 4-6, disperse ultrasonically, then add carboxymethyl chitosan aqueous solution, continue stirring, then add calcium chloride solution, continue stirring, remove water by rotary evaporation, and freeze-dry to obtain the multilayer embedded nano-lyophilized formulation; the volume ratio of the aqueous phase, oil phase, sodium alginate aqueous solution, carboxymethyl chitosan aqueous solution, and calcium chloride solution is 7-9:1-3:10:0.2-0.3:0.01-0.02; The oil-soluble components include Terminalia chebula extract and Rosa canina fruit oil; the mass ratio of Terminalia chebula extract to Rosa canina fruit oil is 3-5:6-8; The water-soluble components include extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium spp.*, and extracts of *Cladosporium spp.*; the mass ratio of the extracts of *Ficus pumila* syconium, filtrate of callus culture from *Eryngium spp.*, and extracts of *Cladosporium spp.* is 4-6:5-9:1-3.

2. The method for preparing a multilayer encapsulated nano-lyophilized formulation according to claim 1, characterized in that, In step S1, the vegetable oil is selected from at least one of sunflower oil, soybean oil, olive oil, flaxseed oil, and palm oil.

3. The method for preparing a multilayer encapsulated nano-lyophilized formulation according to claim 1, characterized in that, In step S3, the freeze drying is carried out at -45 to -35°C and a vacuum of 100-120 Pa for 4-5 hours.

4. A multilayer encapsulated nano-lyophilized formulation prepared by the preparation method according to any one of claims 1-3.

5. The application of a multilayer encapsulated nano-lyophilized formulation prepared by the method according to any one of claims 1-3 in skin care, characterized in that, The application involves adding the multi-layer encapsulated nano-lyophilized formulation to a cosmetic formulation, wherein the multi-layer encapsulated nano-lyophilized formulation constitutes 5%-25% of the cosmetic formulation by mass.

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