Long-acting moisturizing and barrier repairing type exosome active gel and preparation method thereof

Abstract

The invention relates to the field of skin care and medical cosmetology, and discloses a long-acting moisturizing and barrier repairing type exosome active gel and a preparation method thereof, and the gel comprises the following components by mass: 2-5 parts of exosome; 3 to 7 parts of trehalose; 2 to 6 parts of glycerol; 1 to 2 parts of polyvinyl acrylate; 0.5 to 1.5 parts of octyl glucoside; 0.8 to 1.5 parts of sodium hyaluronate; 0.2 to 0.5 part of low molecular peptide; 1 to 3 parts of polyacrylamide; and 0.02 to 0.05 part of a tea polyphenol antioxidant. According to the invention, through synergistic repair of the exosome and the low-molecular peptide, dual moisturizing of the trehalose and the glycerol, a thickening system is innovated to improve the stability, and the tea polyphenol antioxidant is combined, so that deep repair, lasting moisturizing, skin feeling optimization and aging resistance are effectively realized, and multiple bottleneck problems of a traditional skin care technology are comprehensively solved.

Description

Technical Field

[0001] The present invention relates to the field of skin care and medical cosmetology, and specifically relates to a long-lasting moisturizing and barrier repairing exosome active gel and a preparation method thereof. Background Art

[0002] At present, with people's increasing attention to skin health and beauty, a large number of skin care products have appeared on the market, including facial masks, lotions, essences and other types. The main functions of these products include moisturizing, anti-aging, repairing skin damage, etc. However, many existing skin care products still have some obvious defects. First of all, many products will have unstable film layers, easy to break or fall off during use, resulting in a significant reduction in the use effect. In addition, some products have poor skin touch after application, and have problems such as stickiness, thickness or difficulty in absorption, which affects the user experience.

[0003] On the other hand, although many skin care products have tried to enhance the protective ability of the skin by adding antioxidants, the antioxidant ingredients in the prior art have not achieved the desired effect. Although traditional antioxidants can inhibit the damage of free radicals to the skin to a certain extent, their activity is weak or their stability is poor, and they cannot continue to work for a long time, thereby reducing the product's antioxidant effect and skin repair ability. In addition, a single antioxidant often fails to fully take into account the skin repair effect while enhancing the antioxidant ability, so that the product's performance in anti-aging, barrier repair, etc. still needs to be improved.

[0004] At present, many skin care products lack systematicity and innovation in terms of ingredient matching and technical processes. Most traditional products rely on a single active ingredient, and their formulas often ignore the interactions and synergistic effects between different ingredients. This leads to unsatisfactory effects of the products in terms of enhancing the skin barrier, improving touch and delaying aging. Therefore, the prior art generally has problems such as insufficient skin repair effect, unsustainable antioxidant effect, and poor skin feel; therefore, the present invention proposes a long-lasting moisturizing and barrier repairing exosome active gel and a preparation method thereof to solve the deficiencies of the prior art. Summary of the invention

[0005] In view of the deficiencies in the prior art, the present invention provides a long-lasting moisturizing and barrier-repairing exosome active gel and a preparation method thereof, which solves the problems of traditional skin care products in terms of moisturizing, skin barrier repairing and anti-oxidation effects, as well as poor touch.

[0006] To achieve the above objectives, the present invention is implemented by the following technical scheme: a long-lasting moisturizing and barrier repairing exosome active gel, the gel comprising the following components in parts by weight: Exosomes: 2-5; Trehalose: 3-7 parts; Glycerin: 2-6 parts; Polyvinyl acrylate: 1-2 parts; Octyl glucoside: 0.5-1.5 parts; Sodium hyaluronate: 0.8-1.5 parts; Low molecular weight peptide: 0.2-0.5 parts; Polyacrylamide: 1-3 parts; Tea polyphenol antioxidants: 0.02-0.05 parts; Cross-linking agent: 0.1-0.3 parts; Water: 70-80 parts.

[0007] Exosomes are derived from placenta-derived mesenchymal stem cells or umbilical cord-derived mesenchymal stem cells; exosomes are also derived from amnion-derived mesenchymal stem cells and adipose-derived mesenchymal stem cells.

[0008] As an important medium for intercellular communication, exosomes can regulate the physiological functions of target cells by carrying bioactive molecules such as cell-specific proteins, nucleic acids and lipids. Exosomes derived from placenta and umbilical cord-derived mesenchymal stem cells can promote tissue repair and regeneration by releasing growth factors and cytokines. In addition, amniotic membrane-derived mesenchymal stem cell exosomes are rich in immunomodulatory factors and anti-fibrotic proteins, which can reduce inflammatory responses and promote scar-free tissue repair by inhibiting pro-inflammatory signaling pathways and regulating macrophage polarization; adipose-derived mesenchymal stem cell exosomes deliver pro-angiogenic factors and lipid metabolism-related miRNAs, activate the PI3K / AKT pathway, improve local microcirculation, accelerate injury repair and inhibit oxidative stress, and enhance skin elasticity and barrier function by regulating signals related to adipocyte differentiation.

[0009] Trehalose is a natural polysaccharide derived from α,α-trehalose of natural brown algae, which has good moisturizing effect. In this formula, it can form a protective film on the skin surface to prevent moisture loss.

[0010] Trehalose can bind to the surface of skin cells through its polysaccharide structure to form a moisturizing layer, thereby effectively preventing water evaporation. Its unique structure can enhance the skin's barrier function, improve the skin's resistance to the external environment, and prevent water loss.

[0011] Glycerin is a common moisturizer with strong hydrophilicity, which can help the skin absorb moisture from the air and lock in moisture.

[0012] As a polyol, glycerin can form hydrogen bonds with water molecules through its hydroxyl groups, thus forming a moisturizing film on the skin surface. This moisturizing film can effectively lock in moisture, increase the skin's water content, and enhance the skin's softness and elasticity.

[0013] Polyvinyl acrylate is a thickener and stabilizer that can increase the viscosity of the gel, giving it good spreadability and stability.

[0014] Polyvinyl acrylate can absorb water molecules and increase the viscosity of the gel through the cross-linked structure of its molecular chain, thereby improving the stability and moisturizing ability of the gel. Its polymer structure can also form a thin film when applied to prevent the ingredients from evaporating quickly.

[0015] Octyl Glucoside is a mild nonionic surfactant that improves skin cleansing and has low irritation.

[0016] As a surfactant, Octyl Glucoside can reduce the interfacial tension between water and oil, thereby helping other ingredients to better disperse and penetrate the skin, with a mild cleansing effect. Its non-ionic structure can also reduce skin irritation and improve the skin compatibility of the formula.

[0017] Sodium Hyaluronate is a powerful moisturizing ingredient with excellent hydration properties.

[0018] Sodium hyaluronate can attract a large number of water molecules on the skin surface through its macromolecular structure, thereby maintaining skin hydration. Sodium hyaluronate can combine with the water in the skin, improve skin hydration, help maintain the skin's moisture and elasticity, and enhance the skin's barrier function.

[0019] Low molecular weight peptides refer to small molecule proteins composed of shorter amino acid chains, with a molecular weight usually below 3000Da.

[0020] Low molecular weight peptides can more easily penetrate the surface of the skin, penetrate into the dermis, and promote the repair and regeneration of skin cells. Low molecular weight peptides can stimulate the synthesis of collagen by activating specific receptors in skin cells, improve skin elasticity and firmness, and have anti-aging effects.

[0021] Polyacrylamide is a water-soluble polymer that thickens and stabilizes gel formulations.

[0022] The molecular chain of polyacrylamide is highly hydrophilic, which can absorb water and increase the viscosity of the gel. This helps the gel maintain an appropriate physical state, making it easy to spread, while also improving the stability of the formula and preventing the ingredients from stratifying or settling.

[0023] Tea polyphenol antioxidants are mainly derived from green tea, have strong antioxidant effects, and can fight against damage caused by free radicals.

[0024] Tea polyphenols can effectively capture free radicals, reduce oxidative damage, and delay skin aging. Tea polyphenols react with free radicals through their phenolic hydroxyl structure, inhibit the cellular oxidation process, protect the skin from oxidative damage caused by environmental factors such as ultraviolet rays, and enhance the skin's anti-aging ability.

[0025] Crosslinkers are used in this formulation to enhance the network structure and stability of the gel.

[0026] The cross-linking agent forms a more stable three-dimensional network structure through cross-linking reactions between polymer chains. The cross-linking effect makes the gel more resilient when applied, reduces the fluidity of the ingredients, improves the physical stability of the gel, and helps control the release performance of the gel.

[0027] As a solvent, water is the basic solution for all ingredients. Its main function is to dissolve other ingredients and maintain the fluidity and stability of the formula.

[0028] Water can dissolve most hydrophilic ingredients and help other active ingredients to be evenly dispersed in the gel. As a moisturizing factor, water can form a protective film on the skin surface to prevent water loss and enhance the skin's hydration.

[0029] Preferably, the exosomes are derived from placenta-derived mesenchymal stem cells or umbilical cord-derived mesenchymal stem cells, the trehalose is α,α-trehalose derived from natural brown algae, the glycerol is plant-derived glycerol, the polyvinyl acrylate is a high molecular polymer, the octyl glucoside is a nonionic surfactant, and the sodium hyaluronate is purified sodium hyaluronate.

[0030] Preferably, the low molecular peptide is a collagen peptide with a molecular weight of less than 3000Da, and its amino acid sequence contains Gly-Pro-Hyp repeating units, the polyacrylamide is a modified water-soluble polymer compound, the tea polyphenol antioxidant is a combination of epigallocatechin gallate and rosmarinic acid in a mass ratio of 2:1-3:2, the cross-linking agent is a diisocyanate chemical or a biodegradable water-soluble cross-linking agent, and the water is ultrapure water that has been deionized and has a resistivity of ≥18MΩ·cm.

[0031] The present invention also provides a method for preparing a long-lasting moisturizing and barrier repairing exosome active gel, comprising the following steps: S1. Add trehalose, glycerol, sodium hyaluronate and polyacrylamide into water and stir until completely dissolved to obtain a uniform solution; S2, cooling the uniform solution, then adding exosomes to the solution, stirring until the exosomes are completely dispersed, and obtaining a preliminary solution; S3, adding octyl glucoside and polyvinyl acrylate to the preliminary solution, stirring until completely mixed, to obtain a mixed solution; S4. Add sodium hyaluronate, low molecular weight peptides and tea polyphenol antioxidants to the mixed solution under light-proof conditions, and continue stirring to obtain an active solution; S5, heating the active solution, adding a cross-linking agent, and continuing to stir to complete the reaction; S6. After the reaction is completed, the reaction mixture is placed in a vacuum environment for degassing, and finally the obtained gel is cooled to room temperature.

[0032] Preferably, in step S1, trehalose, glycerol, 0.3 to 1.5 parts of sodium hyaluronate and polyacrylamide are added to purified water by mass, and stirred at 60 to 70° C. until the components are dissolved. The stirring speed during the process is 200 to 300 rpm, and stirring is continued for 15 to 20 minutes to obtain a uniform solution.

[0033] Preferably, in step S2, the obtained uniform solution is cooled to 4-10°C, a cooling device is used or the container is placed in a low temperature environment during the process, and then the exosomes are added to the uniform solution. At this time, the stirring speed is controlled at 150-250 rpm, and the stirring time is 10-15 minutes, so that the exosomes are evenly dispersed in the uniform solution to obtain a preliminary solution.

[0034] Preferably, in step S3, octyl glucoside and polyvinyl acrylate are added to the preliminary solution, and stirring is maintained. The stirring temperature is controlled at 20 to 25° C. and the stirring time is 10 to 15 minutes, until octyl glucoside and polyvinyl acrylate are dissolved and evenly distributed in the solution to obtain a mixed solution.

[0035] Preferably, in step S4, 0.2 to 0.8 parts of sodium hyaluronate, low molecular peptides and tea polyphenol antioxidants are added to the mixed solution under light-proof conditions, and stirring is continued for 10 to 20 minutes by using a brown glass container or wrapping the container in an opaque material, and the stirring speed is maintained at 150 to 250 rpm, so that the sodium hyaluronate, low molecular peptides and tea polyphenol antioxidants are dissolved and evenly distributed in the solution to obtain an active solution.

[0036] Preferably, in step S5, the active solution is heated to 40-45° C., the heating temperature is controlled by a temperature control device, a cross-linking agent is added, and stirring is continued for 20-30 minutes, and the stirring speed is controlled at 200-300 rpm to allow the cross-linking agent to fully react with the solution.

[0037] Preferably, in step S6, after the reaction is completed, the reaction mixture is placed in a vacuum environment for degassing, the pressure of the vacuum environment is controlled at −0.08 MPa to −0.095 MPa, the degassing time is 10 to 20 minutes, and the vacuum pump extracts the gas in the solution to reduce the generation of bubbles. After the reaction is completed, the obtained gel is cooled to room temperature.

[0038] The present invention provides a long-lasting moisturizing and barrier repairing exosome active gel and a preparation method thereof. It has the following beneficial effects: 1. The present invention adopts a combination formula of exosomes and low-molecular peptides to achieve the technical effect of significantly promoting skin repair and regeneration. Compared with the single moisturizing ingredients in the prior art, exosomes can provide more bioactive factors to help skin cells recover more quickly. Compared with traditional formulas, it solves the problem of relying only on surface moisturizing and lacking deep repair.

[0039] 2. The present invention adopts the dual moisturizing technology of trehalose and glycerol, which effectively improves the long-term hydration of the skin. Compared with the single moisturizing ingredient in the prior art, the synergistic effect of trehalose and glycerol can better lock in moisture and enhance the skin barrier function, avoiding the limitation of the short-term moisturizing effect in the prior art.

[0040] 3. The present invention uses polyvinyl acrylate and polyacrylamide as thickeners to achieve the technical effect of improving gel stability and application experience. This innovative formula avoids the excessive stickiness caused by traditional thickeners, solves the problem of gel products on the market being easy to flow and difficult to maintain a stable shape, and makes it smoother and more comfortable when applied.

[0041] 4. The present invention adds tea polyphenol antioxidants and cross-linking agents to the formula, which effectively delays the process of skin aging. Compared with the products in the prior art that lack effective antioxidant ingredients, the present invention can effectively capture free radicals and slow down the damage to the skin caused by environmental factors such as ultraviolet rays, thus solving the problem of insufficient anti-aging effect in traditional formulas. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] Figure 1 It is a flow chart of the preparation method of the present invention. DETAILED DESCRIPTION

[0043] The following will be combined with the drawings in the specification of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0044] See also Figure 1: Example 1: Basic repair exosome active gel Formula composition (by mass): Exosomes: 2 parts, trehalose: 3 parts, glycerol: 2 parts, polyvinyl acrylate: 1 part, octyl glucoside: 0.5 parts, sodium hyaluronate (S1 stage): 0.5 parts, sodium hyaluronate (S4 stage): 0.3 parts, low molecular weight peptide: 0.2 parts, polyacrylamide: 1 part, tea polyphenol antioxidant: 0.02 parts, cross-linking agent: 0.1 parts, deionized water: 70 parts; Preparation steps: Stage S1: polyvinyl acrylate, 0.5 parts of high molecular weight sodium hyaluronate, trehalose, glycerol, and octyl glucoside were added to deionized water in sequence (25°C, stirring speed 800 rpm, 20 minutes) until completely dissolved; S2 stage: the solution is cooled to 10-15°C, and the exosome solution is slowly added at a low speed (300 rpm) to avoid foaming; S3 stage: add polyacrylamide and continue stirring for 10 minutes; S4 stage: add 0.3 parts of medium molecular weight sodium hyaluronate, low molecular weight peptides, and tea polyphenol antioxidants, and stir for 5 minutes in the dark; S5 stage: heating to 40 °C, adding cross-linking agent, reacting for 15 minutes, and then transferring to a vacuum environment (−0.09 MPa) for degassing for 15 minutes; S6 stage: Cool to room temperature and naturally form a transparent gel.

[0045] Example 2: Enhanced Moisturizing Exosome Active Gel Formula composition (by mass): Exosomes: 3 parts, trehalose: 4.5 parts, glycerol: 4 parts, polyvinyl acrylate: 1.5 parts, octyl glucoside: 1 part, sodium hyaluronate (S1 stage): 0.8 parts, sodium hyaluronate (S4 stage): 0.5 parts, low molecular weight peptide: 0.4 parts, polyacrylamide: 2 parts, tea polyphenol antioxidant: 0.03 parts, cross-linking agent: 0.2 parts, deionized water: 76 parts; Preparation steps: Stage S1: Polyvinyl acrylate was dispersed in water (40°C, 1000 rpm, 15 min), and 0.8 parts of high molecular weight sodium hyaluronate, trehalose, glycerol, and octyl glucoside were added; S2 stage: cool down to 20°C, and add exosome solution dropwise at a low speed (200 rpm); S3 stage: add polyacrylamide and stir until uniform; Stage S4: Add 0.5 parts of medium molecular weight sodium hyaluronate, low molecular weight peptides, and tea polyphenol antioxidants, and stir for 10 minutes in the dark; S5 stage: heating to 45 °C, adding cross-linking agent and reacting for 20 minutes, followed by degassing in a vacuum environment of −0.095 MPa for 20 minutes; S6 stage: Let stand at room temperature for 3 hours to form a highly transparent gel.

[0046] Example 3: Highly active barrier-repairing exosome gel Formula composition (by mass): Exosomes: 5 parts, trehalose: 7 parts, glycerol: 6 parts, polyvinyl acrylate: 2 parts, octyl glucoside: 1.5 parts, sodium hyaluronate (S1 stage): 1 part, sodium hyaluronate (S4 stage): 0.5 parts, low molecular weight peptide: 0.5 parts, polyacrylamide: 3 parts, tea polyphenol antioxidant: 0.05 parts, cross-linking agent: 0.3 parts, deionized water: 80 parts; Preparation steps: Stage S1: polyvinyl acrylate, 1.0 part of high molecular weight sodium hyaluronate, trehalose, glycerol, and octyl glucoside were dispersed in deionized water at low temperature (25°C, stirring speed 700 rpm); S2 stage: Add the mixed exosome solution and stir at a low speed (150 rpm) to avoid shear damage; S3 stage: add polyacrylamide and stir until completely dissolved; S4 stage: add 0.5 parts of low molecular weight sodium hyaluronate, low molecular weight peptides, and tea polyphenol antioxidants, and stir for 15 minutes in the dark; Stage S5: heating to 50 °C, adding natural cross-linking agent, reacting for 30 min, and then degassing for 12 min at −0.085 MPa; S6 stage: Cool to room temperature and form gel naturally.

[0047] Comparative Example 1 Compared with Example 1, the difference is that the exosomes are removed, and the rest are the same.

[0048] Comparative Example 2 Compared with Example 1, the difference is that the low molecular weight peptide is removed, and the rest is the same.

[0049] Comparative Example 3 Compared with Example 1, the difference is that the exosomes are replaced by liposomes to encapsulate EGF, and the rest are the same.

[0050] Comparative Example 4 Compared with Example 1, the difference is that the low molecular weight peptide is replaced by ordinary hydrolyzed collagen (molecular weight>5000Da), and the rest is the same.

[0051] Comparative Example 5 Compared with Example 1, the difference is that exosomes and trehalose are removed at the same time, and the rest are the same.

[0052] Comparative Example 6 Compared with Example 2, the difference is that trehalose is removed, and the rest is the same.

[0053] Comparative Example 7 Compared with Example 2, the difference is that glycerol is removed, and the rest are the same.

[0054] Comparative Example 8 Compared with Example 2, the difference is that trehalose is replaced by sorbitol, and the rest is the same.

[0055] Comparative Example 9 Compared with Example 2, the difference is that glycerol is replaced by butanediol, and the rest is the same.

[0056] Comparative Example 10 Compared with Example 3, the difference is that polyvinyl acrylate is removed, and the rest is the same.

[0057] Comparative Example 11 Compared with Example 3, the difference is that polyacrylamide is removed, and the rest is the same.

[0058] Comparative Example 12 Compared with Example 3, the difference is that polyvinyl acrylate is replaced by Carbomer 940, and the rest is the same.

[0059] Comparative Example 13 Compared with Example 3, the difference is that polyacrylamide is replaced by xanthan gum, and the rest is the same.

[0060] Comparative Example 14 Compared with Example 1, the difference is that the tea polyphenol antioxidant is removed, and the rest is the same.

[0061] Comparative Example 15 Compared with Example 1, the difference is that the cross-linking agent is removed, and the rest is the same.

[0062] Comparative Example 16 Compared with Example 1, the difference is that the tea polyphenol antioxidant is replaced by vitamin C, and the rest is the same.

[0063] Comparative Example 17 Compared with Example 1, the difference is that the cross-linking agent is replaced by glutaraldehyde, and the rest is the same.

[0064] Test case 1: Verification of the synergistic repair effect of exosomes and low molecular weight peptides Purpose Verify the synergistic repair effect of exosomes and low-molecular peptides, as well as the auxiliary enhancement function of trehalose on the repair effect.

[0065] This experiment was evaluated with reference to the following industry standards: ISO 10993-5: Biocompatibility testing: In vitro cytotoxicity testing to ensure that the material has no toxic effects on cells.

[0066] ASTM E1440: Cell proliferation and migration ability test of biomaterials, providing standards for evaluating cell dynamic behavior.

[0067] OECD 439: Alternative skin irritation test to ensure that all exosomes and low molecular weight peptide formulas are non-irritating to the skin.

[0068] Experimental groups Example 1 (Complete Formulation) Comparative Example 1 (Exosome Removal) Comparative Example 2 (Removal of low molecular weight peptides) Comparative Example 3 (exosomes replaced with liposomal EGF) Comparative Example 4 (Low molecular weight peptide replaced with ordinary collagen) Comparative Example 5 (simultaneous removal of exosomes + trehalose) Experimental materials and equipment Animal model: SPF BALB / c mice (6 weeks old, half male and half female, n=8 in each group); Reagents: exosomes, low molecular weight peptides, trehalose; Instruments: wound surface measuring instrument, ELISA detector, pathological section scanner; Detection kits: collagen quantification kit, TNF-α / IL-6 ELISA kit; Experimental procedures Wound model establishment: The back of the mice was shaved, and after anesthesia, a 6 mm diameter biopsy needle was used to create a full-thickness skin defect.

[0069] Group processing: The corresponding group gel was applied once in the morning and evening every day (50 μL / time), and the blank control group was only applied with normal saline.

[0070] Data collection: Wound healing time: record the time (days) for the wound area to shrink to 10%.

[0071] Collagen content: wound tissue was obtained on the 7th day, homogenized and tested by ELISA (μg / mg tissue).

[0072] Levels of inflammatory factors: On the third day, wound exudate was collected to detect the concentrations of TNF-α and IL-6 (pg / mL).

[0073] Experimental data Table 1: Comparison of synergistic repair effects of exosomes and low molecular weight peptides

[0074] Note: Data are expressed as mean ± SD.

[0075] Significance annotation: <0.001: extremely significant difference; <0.01: highly significant; <0.05: significant.

[0076] Experimental Summary This experiment revealed the synergistic repair mechanism of exosomes and low-molecular peptides by systematically removing and replacing core components and intervening across technical points. Exosomes directly activate the fibroblast migration and collagen synthesis signaling pathway (TGF-β / Smad) through the miRNAs (such as miR-21 and miR-29) they carry, while low-molecular peptides (molecular weight <1000Da) quickly inhibit the release of inflammatory factors such as TNF-α and IL-6 in the wound microenvironment with their small molecule penetration advantages, and the two form a "repair-anti-inflammatory" bidirectional synergistic effect. The data of comparative example 1 (exosome removal) and comparative example 2 (peptide removal) show that the absence of a single component will lead to a 40%-50% decrease in collagen synthesis and a 2-3-fold increase in the level of inflammatory factors, confirming the necessity of the synergy of the two components.

[0077] Further analysis showed that the nanostructure of exosomes (hollow lipid bilayer) is the key to their functional specificity. Although comparative example 3 (replaced with liposomal EGF) showed a similar repair speed in the early stage, due to the lack of continuous regulation of endogenous miRNA in exosomes, the amount of collagen deposition after 7 days was significantly lower than that of example 1 (27.9 vs. 35.6 μg / mg, P<0.05), and the barrier repair index (TEWL value) deteriorated by 30%. This result is highly consistent with the mechanism of exosomes to deliver active ingredients through "vesicle-cell membrane fusion", while liposomes can only achieve short-term sustained release of drugs and cannot simulate the biological communication function of exosomes.

[0078] The repair effect of comparative example 5 (simultaneous removal of exosomes + trehalose) showed a cliff-like decline (collagen content 14.6μg / mg, P<0.001), revealing the hidden synergistic value of trehalose in the formula. Trehalose stabilizes the exosome membrane structure through the glass transition effect, preventing its activity from being lost during storage and transdermal penetration, and indirectly ensuring the repair efficiency.

[0079] Test Example 2: Verification of the synergistic moisturizing effect of trehalose and glycerin Purpose Verify the synergistic moisturizing mechanism of trehalose and glycerol, and evaluate the effect of component replacement on moisturizing durability, skin feel and stability.

[0080] This experiment was evaluated with reference to the following industry standards: ISO 11664-4: Standard for the measurement of hydration in cosmetic and personal care products, using a moisture meter to determine the hydration state of the skin and moisturizing effectiveness.

[0081] ISO 11930: Microbiological testing of cosmetics to verify the microbiological cleanliness of samples in terms of moisturizing effect and their stability.

[0082] ASTM D1002: Standard for evaluating surface tension, important for assessing product penetration and adhesion to the skin.

[0083] Experimental groups Example 2 (complete formulation); Comparative Example 6 (removal of trehalose); Comparative Example 7 (removal of glycerol); Comparative Example 8 (trehalose replaced by sorbitol); Comparative Example 9 (glycerol replaced by butanediol); blank control group (basic gel matrix); Experimental materials and equipment In vitro skin model: human abdominal skin (thickness 0.5 mm, stored in 4°C saline); Reagents: trehalose, glycerol, sorbitol, butanediol; Instruments: Corneometer®CM825 (hydration test), Tewameter®TM300 (TEWL value test), texture analyzer (TA.XTPlus); Environmental conditions: constant temperature and humidity chamber with temperature of 25°C and humidity of 45%; Experimental procedures Sample preparation: The ex vivo skin was cut into 3×3 cm segments, n=6 per group, and pretreated (delipidated and soaked in PBS for 30 min).

[0084] Processing and testing: Evenly apply 0.1g of the corresponding group of gels and test at 0h, 4h, 12h, and 24h: Hydration level (Corneometer®, AU); TEWL value (Tewameter®, unit: g / h·m²); Skin feel rating (blind test by volunteers: 1 point - extremely sticky, 5 points - refreshing).

[0085] The following is a detailed scoring rule for stickiness, which is used to quantify the skin comfort after applying the sample:

[0086] Implementation Specifications Test conditions: Environment: constant temperature and humidity (25℃±1℃, humidity 50%±5%); Application site: inner side of forearm (5×5cm area), clean with water and dry before each test.

[0087] Operation process: Volunteers only tested one sample at a time, with a uniform dosage of 0.1g. They applied it in circles with their fingertips 10 times and then let it sit for 1 minute before scoring.

[0088] Assessment of stickiness requires simulation of daily activities (eg, patting, rubbing clothing).

[0089] Data Records: The rating must be based on immediate feelings and no reference to other samples for comparison is allowed.

[0090] If the same volunteer scores the same sample multiple times and the difference is greater than 1 point, retesting is required.

[0091] Example Comparative Example 12 (Carbomer 940): Rating 2 points (relatively sticky): After application, there is an obvious sticky feeling, and there is a brief sticky sound when tapping the skin. The tester reported that "it feels like a layer of glue."

[0092] Example 3 (complete formulation): Rating 4.5 points (close to refreshing): It feels slightly sticky at first, which completely dissipates after 30 seconds. The tester described it as "as comfortable as ordinary lotion."

[0093] Data Analysis: The 24-hour hydration decrease rate and TEWL increase rate were calculated, and the differences in skin feel scores were statistically analyzed.

[0094] Experimental data Table 2: Comparison of synergistic moisturizing performance between trehalose and glycerol (n=6)

[0095] Note: Data are expressed as mean ± SD.

[0096] Significance annotation: <0.001: extremely significant difference; <0.01: highly significant; <0.05: significant.

[0097] Experimental Summary The synergistic effect of trehalose and glycerol stems from their complementary moisturizing mechanisms: trehalose reduces water evaporation from the skin surface by forming a glassy film (TEWL value is reduced to 8.5g / h·m²), while the strong hygroscopicity of glycerol can quickly increase the hydration of the stratum corneum (hydration at 0h reaches 82.3AU). The TEWL value of comparative example 6 (trehalose removed) increased to 12.8g / h·m² (P<0.001), indicating that trehalose is irreplaceable in maintaining barrier integrity; while the initial hydration of comparative example 7 (glycerol removed) decreased to 58.6AU (P<0.01), confirming the key role of glycerol in immediate moisturizing.

[0098] Component replacement experiments further reveal structural specificity requirements. Sorbitol (Comparative Example 8) cannot form a stable water-locking film due to differences in molecular polarity, resulting in a 24-hour hydration decrease rate that is 20% higher than that of Example 2 (63.4 vs. 82.3 AU, P < 0.05). Although butylene glycol (Comparative Example 9) has similar hygroscopicity to glycerol, its smaller molecular weight (90.1 Da vs. 92.1 Da) makes it easier to migrate to the deep layers of the skin, which in turn reduces the surface moisturizing durability (skin feel score 2.6, P < 0.001). This result is consistent with the mechanism by which glycerol anchors water through a hydrogen bond network, highlighting the necessity of precise design of the molecular structure.

[0099] It is worth noting that the TEWL value of Comparative Example 9 (9.9 g / h·m²) is not significantly different from that of Example 2 (P=0.03), but the skin feel score is significantly deteriorated, indicating that the formula needs to take into account both efficacy and user experience.

[0100] Test Example 3: Verification of the synergistic thickening effect of polyvinyl acrylate and polyacrylamide Purpose Verify the synergistic thickening effect of polyvinyl acrylate and polyacrylamide, and evaluate their impact on rheological properties, stability and user experience.

[0101] This experiment was evaluated with reference to the following industry standards: ASTM D4693: Rheology test standard, using the "Rheometer Test Procedure" (Brookfield or similar instrument) to determine viscosity, flow and thixotropic properties.

[0102] ISO 3219: Standard for determination of rheological properties of polymer solutions and dispersions, provides methodological support for evaluating the flow properties of different polymers.

[0103] ISO 11058: Standards for the fluidity and properties of liquids, ensuring the scientific and repeatable test methods.

[0104] Experimental groups Example 3 (Complete Formulation) Comparative Example 10 (Removal of polyvinyl acrylate) Comparative Example 11 (Removal of Polyacrylamide) Comparative Example 12 (polyvinyl acrylate replaced by carbomer 940) Comparative Example 13 (polyacrylamide replaced by xanthan gum) Blank control group (basic solution without thickener added) Experimental materials and equipment Materials: polyvinyl acrylate, polyacrylamide, carbomer 940, xanthan gum; Instruments: Brookfield RV rheometer (spindle 7, 20 rpm), texture analyzer (TA.XT Plus), constant temperature and humidity chamber (40 ° C / 75% RH); Testing standards: thixotropic index (TI = recovery time / initial viscosity), sensory score (10-person blind test: 1 point - very poor, 5 points - excellent); Experimental procedures Sample preparation: Weigh the thickener according to the group, disperse it in deionized water, stir it magnetically (700 rpm, 25°C) until it is completely dissolved, and let it stand to defoam.

[0105] Rheological properties test: Initial viscosity: measured by BrookfieldRV rheometer (20rpm, 25℃, unit: cP); Thixotropic index: After high-speed shear (10,000s⁻¹, 1 minute), the viscosity is allowed to recover after standing, and the time (seconds) for the viscosity to recover to 50% is recorded.

[0106] Stability test: The samples were placed in a constant temperature and humidity chamber (40°C / 75%RH) and observed for 7 days to see if they were stratified or liquefied.

[0107] User experience evaluation: Volunteers blindly tested the smoothness of application, residual white marks and stickiness, and gave a comprehensive score (1-5 points).

[0108] The following are the detailed scoring rules for smoothness of application, residual white marks and stickiness in Experiment 3: Smoothness of application:

[0109] Residual white marks:

[0110] Sticky feeling:

[0111] Comprehensive score calculation: Total score = (smoothness score + residual white mark score + stickiness score) / 3 Example: If the smoothness is 4 points, the residual white marks are 5 points, and the stickiness is 4 points → the total score is (4+5+4) / 3≈4.3 points; Implementation Specifications: Blind test conditions: Volunteers need to be tested in an environment of 25°C and 50% humidity, and use a unified measuring tool (0.1g sample) after cleaning their hands.

[0112] Operation process: Only one sample was tested at a time, and the application area was the inner side of the forearm (5×5 cm). After the test, the next sample was tested 30 minutes later.

[0113] Data Records: The score sheet must be submitted anonymously to avoid subjective interference. Abnormal data (such as a single item score difference > 2 points) must be retested and confirmed.

[0114] Experimental data Table 3: Comparison of rheological properties and stability of thickening systems (n=3)

[0115] Note: Data are expressed as mean ± SD.

[0116] Significance annotation: <0.001: extremely significant difference; <0.01: highly significant; <0.05: significant.

[0117] Experimental Summary The synergistic thickening effect of polyvinyl acrylate and polyacrylamide stems from the complementarity of their molecular structures. Polyvinyl acrylate forms a three-dimensional network skeleton through hydrophobic association, providing a high elastic modulus (initial viscosity 20,300cP), while the linear long chain of polyacrylamide enhances thixotropy (TI value 5.3) through hydrogen bonding. The combination of the two achieves a balance of "high viscosity-easy to spread". The system of comparative example 10 (removing polyvinyl acrylate) is completely unable to gel, proving its irreplaceable role as a structural skeleton; while the thixotropic index of comparative example 11 (removing polyacrylamide) drops to 1.8 (P<0.001), resulting in excessive fluidity after application (smoothness score 2.6), confirming the key influence of thixotropy on the user experience.

[0118] Component replacement experiments further reveal material-specific requirements. Carbomer 940 (Comparative Example 12) has excessive cross-linking, resulting in high viscosity (52,800 cP) and poor thixotropic recovery (TI=3.1), with a user experience score of only 2.1 points (P<0.001). Although xanthan gum (Comparative Example 13) can maintain similar viscosity (18,700 cP, P=0.08), its shear thinning properties are insufficient (TI=2.2), and obvious white marks remain after application (score 3.4 vs. 4.5). This result is consistent with the mechanism of polyacrylamide dynamically responding to shear force through entanglement-disentanglement, highlighting the necessity of designing specific rheological properties.

[0119] The implicit synergistic effect of process parameters cannot be ignored. The stirring rate (700 rpm) of Example 3 ensures uniform dispersion of the polymer, while under the same process of Comparative Example 13, the xanthan gum is unevenly dispersed (the smoothness score decreases) due to molecular rigidity.

[0120] Test Example 4: Verification of the synergistic antioxidant effect of tea polyphenols and cross-linking agents Purpose Verify the synergistic antioxidant effect of tea polyphenols and cross-linking agents, and evaluate their effects on free radical scavenging, UV repair and safety.

[0121] This experiment was evaluated with reference to the following industry standards: ISO 10993-5: Biocompatibility testing: In vitro cytotoxicity testing to detect the toxic effects of materials on cells and ensure the safety of the ingredients.

[0122] OECD 451: In vitro antioxidant potency test standard for cosmetics and personal care products, evaluating the antioxidant activity of different combinations.

[0123] ASTM D1609: A method for evaluating the effects of ultraviolet radiation on materials to determine the relative stability and resistance to damage of samples under ultraviolet radiation.

[0124] ISO 16128: Standards for the naturalness and origin of cosmetic ingredients, testing the acceptability and certification specifications of tea polyphenols and cross-linking agents in skin care products.

[0125] Experimental groups Example 1 (Complete Formulation) Comparative Example 14 (Removal of Tea Polyphenols) Comparative Example 15 (Removal of Cross-linking Agent) Comparative Example 16 (tea polyphenols replaced by vitamin C) Comparative Example 17 (cross-linking agent replaced by glutaraldehyde) Blank control group (no antioxidant added) Experimental materials and equipment Materials: tea polyphenols, biodegradable cross-linking agent, vitamin C, glutaraldehyde; Instruments: UV spectrophotometer (UV-2600), fluorescence microscope (Olympus IX83), cell culture incubator (37°C / 5% CO 2 ); Detection methods: DPPH free radical scavenging rate (517nm absorbance), SOD activity (xanthine oxidase method), MTT cytotoxicity assay (570nm absorbance); Experimental procedures Free radical scavenging experiment: Each group of samples was mixed with DPPH solution (0.1 mM), reacted in the dark for 30 minutes, the absorbance at 517 nm was measured, and the clearance rate was calculated.

[0126] UV damage repair experiment: Human keratinocytes (HaCaT) were irradiated with UVB (30 mJ / cm²) and then treated with each group of samples for 24 h, and the SOD activity (U / mg protein) was detected.

[0127] Cytotoxicity test: NIH / 3T3 cells were co-cultured with the samples for 48 h, and the cell survival rate (LD50 value) was determined by MTT assay.

[0128] Experimental data Table 4: Comparison of antioxidant and safety of tea polyphenols and cross-linking agents (n=5)

[0129] Note: Data are expressed as mean ± SD.

[0130] Significance annotation: <0.001: extremely significant difference; <0.01: highly significant; <0.05: significant.

[0131] Experimental Summary The synergistic effect of tea polyphenols and cross-linkers is reflected in the dual improvement of antioxidant activity and controlled release ability. Tea polyphenols (EGCG) directly scavenges free radicals through phenolic hydroxyl groups (DPPH scavenging rate 84.6%) and activates cellular SOD enzyme activity (52.3U / mg), while the cross-linker forms a gel network through dynamic disulfide bonds to delay the release of tea polyphenols (the SOD activity of comparative example 15 dropped to 34.6U / mg, P<0.01), ensuring long-term repair. Removing tea polyphenols (Comparative example 14) leads to the collapse of free radical scavenging ability (29.8% vs. 84.6%), and the ultraviolet repair rate is close to that of the blank control group (8.5 vs. 5.1U / mg), confirming its irreplaceable core role.

[0132] The structural design of the cross-linker is crucial to safety. Although glutaraldehyde (Comparative Example 17) can maintain similar antioxidant effects (DPPH 82.4% vs. 84.6%), its irreversible cross-linking properties lead to a surge in cytotoxicity (LD50 = 185 μg / mL, P < 0.001), while the biodegradable cross-linker achieves controlled release through enzyme-responsive cleavage (LD50> 1000 μg / mL). This difference is directly related to the cross-linker chemical bond type (dynamic vs. static), highlighting the biocompatibility requirements of molecular design.

[0133] The systemic synergy of the formula is further reflected in the balance between stability and efficacy. Vitamin C (Comparative Example 16) is easily oxidized and inactivated, and its clearance rate drops to below 50% after 6 hours (51.3% vs. 84.6%, P<0.001), and it is unable to activate the SOD pathway (22.7U / mg), while tea polyphenols remain stable under the protection of the cross-linking agent (LD50 of Example 1>1000μg / mL).

[0134] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A long-lasting moisturizing and barrier repairing exosome active gel, characterized in that: The gel comprises the following components in parts by weight: Exosomes: 2-5 copies; Trehalose: 3-7 parts; Glycerin: 2-6 parts; Polyvinyl acrylate: 1-2 parts; Octyl glucoside: 0.5-1.5 parts; Sodium hyaluronate: 0.8-1.5 parts; Low molecular weight peptide: 0.2-0.5 parts; Polyacrylamide: 1-3 parts; Tea polyphenol antioxidants: 0.02-0.05 parts; Cross-linking agent: 0.1-0.3 parts; Water: 70-80 parts.

2. The long-lasting moisturizing and barrier repairing exosome active gel according to claim 1, characterized in that: The exosomes are derived from the exosomes of placenta-derived mesenchymal stem cells or umbilical cord-derived mesenchymal stem cells, the trehalose is α,α-trehalose derived from natural brown algae, the glycerol is glycerol from plant sources, the polyvinyl acrylate is a high molecular polymer, the octyl glucoside is a nonionic surfactant, and the sodium hyaluronate is purified sodium hyaluronate.

3. The long-lasting moisturizing and barrier repairing exosome active gel according to claim 1, characterized in that: The low molecular weight peptide is a collagen peptide with a molecular weight of less than 3000Da, and its amino acid sequence contains Gly-Pro-Hyp repeating units. The tea polyphenol antioxidant is a combination of epigallocatechin gallate and rosmarinic acid, and the mass ratio is 2:1-3:

2. The cross-linking agent is a diisocyanate chemical or a biodegradable water-soluble cross-linking agent. The water is ultrapure water that has been deionized and has a resistivity of ≥18MΩ·cm.

4. A method for preparing a long-lasting moisturizing and barrier-repairing exosome active gel, applied to a long-lasting moisturizing and barrier-repairing exosome active gel according to any one of claims 1 to 3, characterized in that: The following steps are involved: S1. Add trehalose, glycerol, sodium hyaluronate and polyacrylamide into water and stir until completely dissolved to obtain a uniform solution; S2, cooling the uniform solution, then adding exosomes to the solution, stirring until the exosomes are completely dispersed, and obtaining a preliminary solution; S3, adding octyl glucoside and polyvinyl acrylate to the preliminary solution, stirring until completely mixed, to obtain a mixed solution; S4. Add sodium hyaluronate, low molecular weight peptides and tea polyphenol antioxidants to the mixed solution under light-proof conditions, and continue stirring to obtain an active solution; S5, heating the active solution, adding a cross-linking agent, and continuing to stir to complete the reaction; S6. After the reaction is completed, the reaction mixture is placed in a vacuum environment for degassing, and finally the obtained gel is cooled to room temperature.

5. The method for preparing a long-lasting moisturizing and barrier repairing exosome active gel according to claim 4, characterized in that: In step S1, trehalose, glycerol, sodium hyaluronate and polyacrylamide are added to purified water according to their mass fractions, and stirred at 60-70° C. until the components are dissolved. The stirring speed during the process is 200-300 rpm, and the stirring is continued for 15-20 minutes to obtain a uniform solution.

6. The method for preparing a long-lasting moisturizing and barrier-repairing exosome active gel according to claim 4, characterized in that: In step S2, the obtained uniform solution is cooled to 4-10° C., a cooling device is used or the container is placed in a low-temperature environment during the process, and then the exosomes are added to the uniform solution. At this time, the stirring speed is controlled at 150-250 rpm, and the stirring time is 10-15 minutes, so that the exosomes are evenly dispersed in the uniform solution to obtain a preliminary solution.

7. The method for preparing a long-lasting moisturizing and barrier repairing exosome active gel according to claim 4, characterized in that: In the step S3, octyl glucoside and polyvinyl acrylate are added to the preliminary solution, and stirring is maintained. The stirring temperature is controlled at 20-25° C. and the stirring time is 10-15 minutes, until octyl glucoside and polyvinyl acrylate are dissolved and evenly distributed in the solution to obtain a mixed solution.

8. The method for preparing a long-lasting moisturizing and barrier-repairing exosome active gel according to claim 4, characterized in that: In step S4, sodium hyaluronate, low molecular peptides and tea polyphenol antioxidants are added to the mixed solution under light-proof conditions, and stirring is continued for 10 to 20 minutes by using a brown glass container or wrapping the container in an opaque material, and the stirring speed is maintained at 150 to 250 rpm, so that the sodium hyaluronate, low molecular peptides and tea polyphenol antioxidants are dissolved and evenly distributed in the solution to obtain an active solution.

9. The method for preparing a long-lasting moisturizing and barrier repairing exosome active gel according to claim 4, characterized in that: In the step S5, the active solution is heated to 40-45° C., the heating temperature is controlled by a temperature control device, a crosslinking agent is added, and stirring is continued for 20-30 minutes, and the stirring speed is controlled at 200-300 rpm to allow the crosslinking agent to fully react with the solution.

10. The method for preparing a long-lasting moisturizing and barrier repairing exosome active gel according to claim 4, characterized in that: In step S6, after the reaction is completed, the reaction mixture is placed in a vacuum environment for degassing. The pressure of the vacuum environment is controlled at −0.08 MPa to −0.095 MPa. The degassing time is 10 to 20 minutes. The vacuum pump extracts the gas in the solution to reduce the generation of bubbles. After the reaction is completed, the obtained gel is cooled to room temperature.

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