Anti-aging and repairing compositions containing PDRN, their lipid nanoparticles and applications

Abstract

This application provides an anti-aging and repairing composition containing PDRN, its lipid nanoparticles, and its applications. The composition provided in this application includes polydeoxyribonucleotides, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5. This application combines polydeoxyribonucleotides, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5, exhibiting synergistic effects in promoting cell proliferation and inhibiting intracellular ROS levels, resulting in significant antioxidant, anti-inflammatory, anti-aging, soothing, and repairing effects. This application prepares the PDRN-containing composition into lipid nanoparticles, improving the stability of the PDRN-containing composition and increasing its permeability to the skin surface, thereby enhancing its bioavailability, promoting cell survival and uptake, adenosine A2A receptor expression, collagen synthesis, angiogenesis, and inhibiting anti-inflammatory factors, achieving significant anti-aging effects.

Description

Technical Field

[0001] This invention relates to the field of daily chemical technology, and in particular to an anti-aging and repairing composition containing PDRN, its lipid nanoparticles, and its applications. Background Technology

[0002] Polydeoxyribonucleotide (PDRN) is a polymer of DNA fragments extracted and purified from sperm cells of specific fish (usually salmon). Its molecular weight typically ranges from 50 to 1500 kDa, and it exhibits high biocompatibility. The mechanisms of action of PDRN mainly involve activating A2A receptors, promoting cell proliferation and differentiation, regulating inflammatory responses, and promoting angiogenesis. When used as an active ingredient in cosmetics, it can effectively alleviate wrinkles caused by skin aging and help improve pigmentation.

[0003] Further enhancing the role of PDRN in promoting cell proliferation and differentiation, regulating inflammatory responses, and promoting angiogenesis is currently a hot research topic. Summary of the Invention

[0004] In view of this, the technical problem to be solved by this application is to provide an anti-aging and repair composition containing PDRN, its lipid nanoparticles and applications. The components in the composition provided by this application have a synergistic effect in promoting cell proliferation and inhibiting intracellular ROS levels, and have significant antioxidant, anti-inflammatory, anti-aging, soothing and repair effects.

[0005] This application provides a composition containing PDRN, comprising polydeoxyribonucleotide, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5.

[0006] This application combines polydeoxyribonucleotides, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5, which have a synergistic effect in promoting cell proliferation and inhibiting intracellular ROS levels, and have significant antioxidant, anti-inflammatory, anti-aging, soothing and repairing effects.

[0007] Polydeoxyribonucleotide (PDRN) is a DNA fragment polymer extracted and purified from sperm cells of specific fish (usually salmon). It has functions such as activating A2A receptors, promoting cell proliferation and differentiation, regulating inflammatory responses, and promoting angiogenesis. This application does not impose any special restrictions on the source of the PDRN; it can be purchased commercially.

[0008] The peptide sequence of microcurrent tetrapeptide-1 is LEAP, which can improve mitochondrial function and increase ATP production. This application does not impose any special restrictions on the source of the microcurrent tetrapeptide-1; it can be purchased commercially, for example, under the trade name Uplevity.TM e-Liftpeptide solution products.

[0009] Palmitoyl tripeptide-5 can activate skin tissue growth factor-β (TGF-β), significantly increasing the production of type I, II, and IV collagen, enhancing skin elasticity, and reducing wrinkles and sagging. This application does not impose any special restrictions on the source of the palmitoyl tripeptide-5; it can be purchased commercially, for example, from a product marketed as SYN@-COLL.

[0010] In some specific implementations, the mass ratio of the polydeoxyribonucleotide, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 is 0.1~5:0.000005~0.0005:0.0001~1. In some specific implementations, the mass ratio of the polydeoxyribonucleotide, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 is 0.5~4.5:0.00001~0.0003:0.0005~0.5. In some specific implementations, the mass ratio of the polydeoxyribonucleotide, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 is 1~4:0.00005~0.0001:0.001~0.1. In some specific implementations, the mass ratio of the polydeoxyribonucleotide, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 is 4:0.0001:0.04.

[0011] This application also provides a PDRN-containing lipid nanoparticle, comprising the composition described in the above technical solution and a lipid nanoparticle raw material, wherein the lipid nanoparticle raw material comprises at least two of phospholipids, emulsifiers, polyols, and water.

[0012] PDRN has drawbacks such as large molecular weight, strong polarity, difficulty in penetrating the skin barrier and accumulating at the skin's anti-aging target sites, and easy degradation by nucleases on the skin surface. Microcurrent tetrapeptide-1 and palmitoyl tripeptide-5 also have the problems of instability and poor penetration. This application prepares the PDRN-containing composition into lipid nanoparticles to improve the stability of the PDRN-containing composition and increase its permeability on the skin surface, thereby improving its bioavailability, promoting cell survival and uptake, expression of adenosine A2A receptor, collagen synthesis, angiogenesis, and inhibition of anti-inflammatory factors, achieving significant anti-aging effects.

[0013] In some specific implementations, the lipid nanoparticles include:

[0014] 0.1wt%~5wt% polydeoxyribonucleotides;

[0015] 0.000005wt%~0.0005wt% of microcurrent tetrapeptide-1;

[0016] 0.0001wt%~1wt% palmitoyl tripeptide-5;

[0017] 0.1wt%~3wt% phospholipids;

[0018] 0.1wt%~3wt% of non-phospholipid functional lipids;

[0019] 10wt%~40wt% emulsifier;

[0020] 10wt%~50wt% of alcohols;

[0021] The remaining water.

[0022] In some specific implementations, the lipid nanoparticles include:

[0023] 0.5wt%~4.5wt% polydeoxyribonucleotides;

[0024] 0.00001wt%~0.0003wt% of microcurrent tetrapeptide-1;

[0025] 0.0005wt%~0.5wt% palmitoyl tripeptide-5;

[0026] 0.1wt%~3wt% phospholipids;

[0027] 0.1wt%~3wt% of non-phospholipid functional lipids;

[0028] 10wt%~40wt% emulsifier;

[0029] 10wt%~50wt% of alcohols;

[0030] The remaining water.

[0031] In some specific implementations, the phospholipids include, but are not limited to, lecithin (e.g., soybean lecithin, hydrogenated lecithin, egg yolk lecithin, hydrogenated soybean lecithin, hydrogenated egg yolk lecithin, etc.), distearate phosphatidylcholine (DSPC), natural sphingomyelin (SM), phosphatidylglycerol (PG), phosphatidylinositol (PI), and phosphatidyltryptophan, which may be one or more of these.

[0032] In some specific implementations, the non-phospholipid functional lipids include, but are not limited to, stearamide, oleoyl fatty amine derivatives, polyethylene glycol stearate N-hydroxysuccinimide ester, N-(methylpolyoxyethyleneoxycarbonyl)-1,2-distearate-sn-glycerol-3-phosphate ethanolamine sodium salt, and may be one or more of these.

[0033] In some specific implementations, the emulsifier includes, but is not limited to, polyoxyethylene castor oil emulsifiers, polyoxyethylene hydrogenated castor oil emulsifiers, polyglycerol emulsifiers, poloxamer, cocoyl glucoside, polyglycerol-10 laurate, glyceryl citrate stearate, sodium bis(lauramide glutamine) lysine, and triglyceride emulsifiers (e.g., caprylic / capric triglyceride, palmitic triglyceride, 1,3-dioleoyl-2-palmitoyl triglyceride, caprylic / capric / succinic triglyceride, docosane). The ingredients may be one or more of the following: triglycerides, hydrooleic acid / linoleic acid triglycerides, caprylic / capric acid / linoleic acid triglycerides, etc.), lauryl ether-23, PPG-26-butanol ether-26, tridecyl alcohol ether-12, polyglycerol esters, polysorbate-80 (Tween-80), polysorbate-60 (Tween-60), polysorbate-20 (Tween-20), PEG-20 hydrogenated castor oil, PEG-40 hydrogenated castor oil, and PEG-60 hydrogenated castor oil.

[0034] In some specific implementations, the alcohol compound can be a monohydric alcohol, a dihydric alcohol, or a polyhydric alcohol, including but not limited to glycerol, 1,2-propanediol, 1,2-butanediol, 1,3-butanediol, 1,4-butanediol, 2,3-butanediol, 1,3-propanediol, 1,2-pentanediol, ethoxydiethylene glycol, 1,2-hexanediol, dipropylene glycol, isopropanol, methylpropanediol, polyethylene glycol-200, PPG-10 sorbitol, and octyldodecyl alcohol, etc., and can be one or more of them.

[0035] This application also provides a method for preparing the lipid nanoparticles described in the above technical solution, comprising the following steps:

[0036] (1) Mix polydeoxyribonucleotides, emulsifiers, water and a first mass fraction of alcohol to obtain mixture 1;

[0037] Mixture 2 is prepared by mixing phospholipids, non-phospholipid functional lipids, and a second mass fraction of alcohol compounds.

[0038] Mixture 3 was prepared by mixing microcurrent tetrapeptide-1, palmitoyl tripeptide-5, emulsifier and a third part by weight of alcohol compound;

[0039] (2) Mix mixture 3 with mixture 1, and then mix it with mixture 2 to obtain mixture 4;

[0040] (3) The mixture 4 was nano-sized to obtain lipid nanoparticles.

[0041] This application first mixes polydeoxyribonucleotides, emulsifiers, water, and a first mass part of an alcohol compound under an inert gas, such as nitrogen, to obtain mixture 1. Then, phospholipids, non-phospholipid functional lipids, and a second mass part of the alcohol compound are mixed to obtain mixture 2. Next, microcurrent tetrapeptide-1, palmitoyl tripeptide-5, an emulsifier, and a third mass part of the alcohol compound are mixed to obtain mixture 3. After obtaining mixtures 1, 2, and 3, mixture 3 is first mixed with mixture 1, and then with mixture 2 to obtain mixture 4. Finally, mixture 4 is nano-sized to obtain lipid nanoparticles.

[0042] In some specific implementations, the mixing in the above steps is carried out at 10℃~30℃.

[0043] In some specific implementations, the nano-sizing process specifically refers to:

[0044] Mixture 4 was homogenized under nitrogen protection, pressure of 500-1200 bar, and homogenization 1-5 times.

[0045] In some specific implementations, after nano-processing, the material is discharged at 20°C under nitrogen protection.

[0046] The method provided in this application enables phospholipids to encapsulate active ingredients, resulting in lipid nanoparticles with a particle size of 100nm~300nm and a PDI of 0.1~0.6, which are relatively stable.

[0047] This application also provides a composition containing lipid nanoparticles, comprising the lipid nanoparticles described in the above technical solution and a fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract.

[0048] Bifidobacterium / Lactobacillus / Soybean Seed Extract Ferment Filtrate (Huanfuyuan), National Cosmetic Product Registration Number 20230032, registered Chinese name is Bifidobacterium / Lactobacillus / Soybean Seed Extract Ferment Filtrate, INCI name is Bifidobacterium / Lactobacillus / Soybean Seed Extract Ferment Filtrate, it can be purchased from Yixian e-commerce platform, or it can be prepared by the following steps: soaking black beans with water in a certain proportion, heating and water extraction to obtain soybean seed extract, sterilizing to obtain fermentation base, then transferring to a fermentation tank, adding a bacterial strain composition (containing Bifidobacterium and Lactobacillus) for fermentation, and obtaining fermentation filtrate after sterilization and filtration.

[0049] This application combines lipid nanoparticles with fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract, enabling the active ingredients to efficiently penetrate the skin barrier and quickly reach the deep dermis layer of the skin. This effectively stimulates fibroblast renewal, thereby promoting the synthesis of collagen and elastin, enhancing skin elasticity, and promoting skin renewal; as well as inhibiting the production of inflammatory factors, enhancing the expression of adenosine A2A receptors, and delaying skin aging.

[0050] In some specific implementations, the mass ratio of the lipid nanoparticles to the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract is 1~10:1. In some specific implementations, the mass ratio of the lipid nanoparticles to the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract is 2~15:1.

[0051] This application also provides a method for preparing a composition containing lipid nanoparticles, comprising the following steps:

[0052] After mixing the liposomes with the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract, the mixture was discharged at 20°C under nitrogen protection.

[0053] In some specific implementations, the mixing is carried out at 10~30℃. In some specific implementations, the mixing can be achieved by stirring, shearing, or other methods to achieve uniform mixing.

[0054] The PDRN-containing composition, lipid nanoparticles, and / or lipid nanoparticle-containing composition described in the above-mentioned technical solutions provided in this application have anti-aging, anti-inflammatory, soothing, and repairing effects, and can be used in products with corresponding effects.

[0055] This application also provides a product comprising the PDRN-containing composition described in the above technical solution, the lipid nanoparticles described in the above technical solution, and / or the lipid nanoparticle-containing composition described in the above technical solution.

[0056] In some embodiments of the present invention, the above-mentioned products include, but are not limited to, personal care products, pharmaceuticals, etc. Those skilled in the art will understand that, in addition to the compositions described in the above technical solutions, other excipients are also included, such as pharmaceutical excipients or excipients for personal care products.

[0057] In some embodiments of the present invention, the personal care products include: basic care products and / or makeup products.

[0058] In some embodiments of the present invention, the cosmetic products include, but are not limited to:

[0059] (1) Base makeup:

[0060] Foundation / cream: Used to even out skin tone and cover blemishes;

[0061] BB cream / CC cream: A lightweight base makeup product that combines skincare and makeup application;

[0062] Concealer / pen: Used to cover up specific areas such as pimples and dark circles;

[0063] Loose powder / setting powder: Sets makeup and reduces facial shine.

[0064] (2) Eye makeup:

[0065] Eyeshadow: Adds color and dimension to the eyes;

[0066] Eyeliner pencil / liquid / gel: Used to draw lines on the eyes, making them look more vibrant;

[0067] Mascara: Lengthens and thickens eyelashes, enhancing the depth of the eyes;

[0068] Eyebrow pencil / powder / gel: Fill in gaps in eyebrows and create the ideal eyebrow shape.

[0069] (3) Cheek makeup:

[0070] Blush: Adds natural color to the cheeks and improves complexion;

[0071] Contouring powder / stick: Use shading techniques to make facial contours more three-dimensional;

[0072] (4) Lip makeup:

[0073] Lipstick / lip gloss / lip tint: to change or emphasize the color of the lips;

[0074] Lip liner: Defines the clear outline of the lips and prevents lipstick from bleeding.

[0075] (5) Multifunctional cosmetics:

[0076] Highlighter sticks / liquids / powders: Highlight high points on the face (such as the bridge of the nose and cheekbones) to create a luminous effect.

[0077] In addition, there are products designed specifically for special occasions, such as waterproof and sweatproof eyeliners and long-lasting, non-fading lipsticks.

[0078] In some embodiments of the present invention, the basic care products include, but are not limited to:

[0079] Facial cleanser: Gently removes dirt, oil, and makeup residue from the face;

[0080] Cleansing oil / cleansing water / cleansing balm: specially designed to thoroughly remove makeup, especially waterproof cosmetics;

[0081] Toner / lotion: Used after cleansing, it can further cleanse the skin's surface of residue, while replenishing the skin's moisture, restoring the skin's pH balance, and laying a good foundation for the absorption of subsequent skin care products;

[0082] Serum: Contains a high concentration of active ingredients, providing deep nourishment and repair for specific skin problems (such as anti-aging, moisturizing, whitening, etc.);

[0083] Eye cream: Specially designed for the sensitive area around the eyes, it has the effects of reducing fine lines and dark circles and firming the skin around the eyes. The texture is usually light and easily absorbed.

[0084] Day / night lotion or cream: It has functions such as protection, repair and nourishment, and promotes cell regeneration. It can provide the skin with the necessary moisture and lock in the replenished moisture.

[0085] Sunscreen: Used for ultraviolet protection and prevention of photoaging, etc.

[0086] Face masks: provide extra nourishment and care for the skin, such as hydration, pore cleansing, or brightening of the complexion.

[0087] In some specific implementations, the personal care product includes 0.1wt% to 10wt% of the PDRN-containing composition described in the above technical solution, the lipid nanoparticles described in the above technical solution, and / or the lipid nanoparticle-containing composition described in the above technical solution, preferably including 0.5wt% to 9.5wt%, more preferably including 1wt% to 9wt%.

[0088] In some specific implementations, the personal care product can be an essence water, a typical formula of which includes: component A, component B, active ingredients, and water. By total mass (100%), component A contains: 5% dipropylene glycol, 10% glycerin, 15% propylene glycol, and 0.15% carbomer 941; component B contains: 0.15% triethanolamine, and 0.5% 1,2-hexanediol / caprylyl hydroxamic acid / ethylhexylglycerin / propylene glycol; the active ingredients include 0.1wt%~10wt% of the PDRN-containing composition described in the above technical solution, the lipid nanoparticles described in the above technical solution, and / or the composition containing lipid nanoparticles described in the above technical solution; the balance being water.

[0089] This application combines polydeoxyribonucleotides, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5, which have a synergistic effect in promoting cell proliferation and inhibiting intracellular ROS levels, and have significant antioxidant, anti-inflammatory, anti-aging, soothing and repairing effects.

[0090] Furthermore, this application prepares lipid nanoparticles by combining polydeoxyribonucleotides, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 with phospholipids, non-phospholipid functional lipids, emulsifiers, alcohols, and water. These nanoparticles are readily soluble in water, easy to use, and exhibit good stability and water dispersibility, increasing the solubility of the active ingredients while also reducing their irritation. This allows the active ingredients to reach sufficient concentrations in the product to exert their corresponding functional effects. The lipid nanoparticles provided by this application effectively improve the solubility of each active ingredient while providing a stable storage space, preventing unnecessary degradation or inactivation of the active ingredients before storage and use, and thus facilitating the increase of the concentration of active substances during use.

[0091] This application combines the lipid nanoparticles described in the above technical solution with the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract, which enables the active ingredients to efficiently penetrate the skin barrier and quickly reach the deep dermis layer of the skin. This effectively stimulates fibroblast renewal, thereby promoting the synthesis of collagen and elastin, enhancing skin elasticity, and promoting skin renewal; as well as inhibiting the generation of cellular inflammatory factors, enhancing the expression of adenosine A2A receptors, and delaying skin aging.

[0092] Experimental results show that the lipid nanoparticles and their compositions provided in this application have good skin permeability and retention properties after use. They can be enriched at high concentrations in target tissues, remain for a long time, and be effectively taken up by target cells, thereby improving the bioavailability of active ingredients, enhancing anti-aging effects, and prolonging the duration of action. Attached Figure Description

[0093] Figure 1 The electrophoresis results provided in the embodiments of the present invention. Detailed Implementation

[0094] This invention provides a PDRN-containing composition, its lipid nanoparticles, and its applications. Those skilled in the art can refer to the content of this document and appropriately modify the process parameters to achieve the desired results. The methods and applications of this invention have been described through preferred embodiments. Those skilled in the art can obviously make modifications or appropriate changes and combinations to the methods and applications described herein without departing from the content, spirit, and scope of this invention to realize and apply the technology of this invention.

[0095] The terms “including,” “having,” or “containing,” including the use of their grammatical synonyms, should generally be understood as open-ended and non-restrictive, for example, not excluding other unstated elements or steps, unless otherwise specifically stated or understood from the context.

[0096] It should be understood that the order of the steps or the order in which certain actions are performed is not important as long as the invention remains operational. Furthermore, two or more steps or actions can be performed simultaneously.

[0097] The use of any and all instances or exemplary language such as “e.g.” or “including” in this document is merely intended to better illustrate the invention and is not intended to limit the scope of the invention unless the claims are made. No language in this specification should be construed as indicating that any unclaimed element is essential to the practice of the invention.

[0098] Furthermore, the numerical ranges and parameters used to define the present invention are approximate values, and the relevant values ​​in the specific embodiments have been presented as precisely as possible. However, any value inevitably contains standard deviations due to individual test methods. Therefore, unless explicitly stated otherwise, it should be understood that all ranges, quantities, values, and percentages used in this disclosure are modified with the word "approximately". Here, "approximately" generally means that the actual value is within plus or minus 10%, 5%, 1%, or 0.5% of a specific value or range.

[0099] This application provides an anti-aging and repair composition containing PDRN, comprising polydeoxyribonucleotide, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5.

[0100] This application combines polydeoxyribonucleotides, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5, which have a synergistic effect in promoting cell proliferation and inhibiting intracellular ROS levels, and have significant antioxidant, anti-inflammatory, anti-aging, soothing and repairing effects.

[0101] This application also provides a PDRN-containing lipid nanoparticle, comprising the composition described in the above technical solution and a lipid nanoparticle raw material, wherein the lipid nanoparticle raw material comprises at least two of phospholipids, non-phospholipid functional lipids, emulsifiers, alcohol compounds, and water.

[0102] This application prepares lipid nanoparticles by combining polydeoxyribonucleotides, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 with phospholipids, non-phospholipid functional lipids, emulsifiers, alcohols, and water. These nanoparticles are readily soluble in water, easy to use, and exhibit good stability and water dispersibility, increasing the solubility of the active ingredients while also reducing their irritation. This allows the active ingredients to reach sufficient concentrations in the product to exert their corresponding functional effects. The lipid nanoparticles provided by this application effectively improve the solubility of each active ingredient while providing a stable storage space, preventing unnecessary degradation or inactivation of the active ingredients before storage and use, and thus facilitating higher concentrations of active substances during use.

[0103] This application also provides a method for preparing the lipid nanoparticles described in the above technical solution, comprising the following steps:

[0104] (1) Mix polydeoxyribonucleotides, emulsifiers, water and a first mass fraction of alcohol to obtain mixture 1;

[0105] Mixture 2 is prepared by mixing phospholipids, non-phospholipid functional lipids, and a second mass fraction of alcohol compounds.

[0106] Mixture 3 was prepared by mixing microcurrent tetrapeptide-1, palmitoyl tripeptide-5, emulsifier and a third part by weight of alcohol compound;

[0107] (2) Mix mixture 3 with mixture 1, and then mix it with mixture 2 to obtain mixture 4;

[0108] (3) The mixture 4 was nano-sized to obtain lipid nanoparticles.

[0109] The method provided in this application enables phospholipids to encapsulate active ingredients, resulting in lipid nanoparticles with a particle size of 100nm~300nm and a PDI of 0.1~0.6, which are relatively stable.

[0110] This application also provides a composition containing lipid nanoparticles, comprising the lipid nanoparticles described in the above technical solution and a fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract.

[0111] This application combines lipid nanoparticles with fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract, enabling the active ingredients to efficiently penetrate the skin barrier and quickly reach the deep dermis layer of the skin. This effectively stimulates fibroblast renewal, thereby promoting the synthesis of collagen and elastin, enhancing skin elasticity, and promoting skin renewal; as well as inhibiting the production of inflammatory factors, enhancing the expression of adenosine A2A receptors, and delaying skin aging.

[0112] The present application will be further described below with reference to the embodiments.

[0113] Example 1, Comparative Examples 1-6

[0114] According to the formula shown in Table 1, the components are mixed to obtain the composition.

[0115] Table 1. Formulations of Example 1 and Comparative Examples

[0116]

[0117] Example 2

[0118] (1) 4% PDRN, 20% methyl propylene glycol, 12% 1,3-butanediol, 5% lauryl ether-23, 20% cocoyl glucoside, 10% Tween-80, and the remainder water were stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0119] (2) 2% soybean lecithin, 0.5% phosphatidylglycerol, 10% 1,2-pentanediol and 3% isopropanol were stirred at 30°C to form a homogeneous and clear liquid 2.

[0120] (3) Microcurrent tetrapeptide-1 0.0001%, palmitoyl tripeptide-5 0.04%, polyglycerol-10 laurate 5%, and 1,3-propanediol 5% were stirred at 20°C to form a homogeneous and clear liquid 3.

[0121] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0122] (5) After homogenizing liquid 4 three times with a high-pressure homogenizer under nitrogen protection and 800 bar conditions, PDRN lipid nanoparticles were obtained.

[0123] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0124] Example 3

[0125] (1) 4% PDRN, 20% methyl propylene glycol, 12% 1,3-butanediol, 5% lauryl ether-23, 20% cocoyl glucoside, 10% Tween-80, and the remainder water were stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0126] (2) 2% hydrogenated lecithin, 0.5% polyethylene glycol N-hydroxysuccinimide stearate, 10% 1,2-pentanediol and 3% isopropanol were stirred at 30°C to form a homogeneous and clear liquid 2.

[0127] (3) Microcurrent tetrapeptide-1 0.0001%, palmitoyl tripeptide-5 0.04%, polyglycerol-10 laurate 5%, and 1,3-propanediol 5% were stirred at 20°C to form a homogeneous and clear liquid 3.

[0128] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0129] (5) After homogenizing liquid 4 three times with a high-pressure homogenizer under nitrogen protection and 800 bar conditions, PDRN lipid nanoparticles were obtained.

[0130] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0131] Example 4

[0132] (1) 4% PDRN, 20% methyl propylene glycol, 12% 1,3-butanediol, 5% lauryl ether-23, 20% cocoyl glucoside, 10% Tween-80, and the remainder water were stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0133] (2) 1% phosphatidylinositol, 1% hydrogenated lecithin, 0.5% polyethylene glycol N-hydroxysuccinimide stearate, 10% 1,2-pentanediol, and 3% isopropanol were stirred at 30°C to form a homogeneous and clear liquid 2.

[0134] (3) Microcurrent tetrapeptide-1 0.0001%, palmitoyl tripeptide-5 0.04%, polyglycerol-10 laurate 5%, and 1,3-propanediol 5% were stirred at 20°C to form a homogeneous and clear liquid 3.

[0135] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0136] (5) After homogenizing liquid 4 three times with a high-pressure homogenizer under nitrogen protection and 800 bar conditions, PDRN lipid nanoparticles were obtained.

[0137] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0138] Example 5

[0139] (1) 4% PDRN, 20% methyl propylene glycol, 12% 1,3-butanediol, 5% lauryl ether-23, 20% cocoyl glucoside, 10% Tween-80, and the remainder water were stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0140] (2) 1% of oleoyl fatty amine derivative, 1% stearamide, 0.5% natural sphingomyelin, 10% 1,2-pentanediol, and 3% isopropanol were stirred at 30°C to form a homogeneous and clear liquid 2.

[0141] (3) Microcurrent tetrapeptide-1 0.0001%, palmitoyl tripeptide-5 0.04%, polyglycerol-10 laurate 5%, and 1,3-propanediol 5% were stirred at 20°C to form a homogeneous and clear liquid 3.

[0142] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0143] (5) After homogenizing liquid 4 three times with a high-pressure homogenizer under nitrogen protection and 800 bar conditions, PDRN lipid nanoparticles were obtained.

[0144] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0145] Example 6

[0146] (1) 4% PDRN, 20% methyl propylene glycol, 12% 1,3-butanediol, 5% lauryl ether-23, 20% cocoyl glucoside, 10% Tween-80, and the remainder water were stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0147] (2) Egg yolk lecithin 1%, natural sphingomyelin 1%, phosphatidylglycerol 0.5%, 1,2-pentanediol 10%, isopropanol 3%, are stirred at 30°C to form a homogeneous and clear liquid 2.

[0148] (3) Microcurrent tetrapeptide-1 0.0001%, palmitoyl tripeptide-5 0.04%, polyglycerol-10 laurate 5%, and 1,3-propanediol 5% were stirred at 20°C to form a homogeneous and clear liquid 3.

[0149] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0150] (5) After homogenizing liquid 4 three times with a high-pressure homogenizer under nitrogen protection and 800 bar conditions, PDRN lipid nanoparticles were obtained.

[0151] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0152] Example 7

[0153] (1) 4% PDRN, 20% methyl propylene glycol, 12% 1,3-butanediol, 5% lauryl ether-23, 20% cocoyl glucoside, 10% Tween-80, and the remainder water were stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0154] (2) 1% phosphatidyltryptophan, 0.5% natural sphingomyelin, 0.5% phosphatidylglycerol, 0.5% N-(methylpolyoxyethyleneoxycarbonyl)-1,2-distearate-sn-glycerol-3-phosphate ethanolamine sodium salt, 10% 1,2-pentanediol, and 3% isopropanol were stirred at 30°C to form a homogeneous and clear liquid 2.

[0155] (3) Microcurrent tetrapeptide-1 0.0001%, palmitoyl tripeptide-5 0.04%, polyglycerol-10 laurate 5%, and 1,3-propanediol 5% were stirred at 20°C to form a homogeneous and clear liquid 3.

[0156] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0157] (5) After homogenizing liquid 4 three times with a high-pressure homogenizer under nitrogen protection and 800 bar conditions, PDRN lipid nanoparticles were obtained.

[0158] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0159] Example 8

[0160] (1) 2.5% PDRN, 10% octyl dodecanol, 8% ethoxydiethylene glycol, 10% caprylic / capric / succinic triglyceride, 15% sodium di(lauramide glutamine) lysine, and the remainder water are stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0161] (2) 1.5% soybean lecithin, 0.5% phosphatidylglycerol and 10% isopropanol were stirred at 30°C to form a homogeneous and clear liquid 2.

[0162] (3) Microcurrent tetrapeptide-1 0.00008%, palmitoyl tripeptide-5 0.02%, glyceryl citrate stearate 3%, PPG-26-butanol polyether-26 5%, and glycerol 5% are stirred at 20°C to form a homogeneous and clear liquid 3.

[0163] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0164] (5) After homogenizing liquid 4 once with a high-pressure homogenizer under nitrogen protection and 1000 bar, PDRN lipid nanoparticles are obtained.

[0165] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0166] Example 9

[0167] (1) PDRN 1%, 1,2-pentanediol 10%, PPG-10 sorbitol 5%, tridecyl alcohol polyether-12 5%, PEG-20 hydrogenated castor oil 10%, lauryl alcohol polyether-23 5%, and the balance water are stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0168] (2) Soybean lecithin 0.5%, phosphatidylglycerol 0.5%, 1,2-hexanediol 3%, dipropylene glycol 2% were stirred at 30°C to form a homogeneous and clear liquid 2.

[0169] (3) Microcurrent tetrapeptide-1 0.00005%, palmitoyl tripeptide-5 0.01%, sodium bis(lauramide glutamine) lysine 1%, and 1,2-propanediol 6% were stirred at 20°C to form a homogeneous and clear liquid 3.

[0170] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0171] (5) After homogenizing liquid 4 four times with a high-pressure homogenizer under nitrogen protection and 600 bar conditions, PDRN lipid nanoparticles were obtained.

[0172] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0173] Example 10

[0174] (1) 0.1% PDRN, 5% glycerol, 2,3-butanediol, 5% lauryl ether-23, 3% poloxamer, and the balance water were stirred at 20°C under nitrogen protection to form a homogeneous and clear liquid 1.

[0175] (2) Soybean lecithin 0.3%, phosphatidylglycerol 0.2%, 1,2-pentanediol 1%, isopropanol 2% were stirred at 30°C to form a homogeneous and clear liquid 2.

[0176] (3) Microcurrent tetrapeptide-1 0.00001%, palmitoyl tripeptide-5 0.001%, polyglycerol-10 laurate 3%, and 1,2-propanediol 3% are stirred at 20°C to form a homogeneous and clear liquid 3.

[0177] (4) Pour liquid 3 into liquid 1 and mix well, then pour it into liquid 2 and stir to form a homogeneous liquid 4.

[0178] (5) After homogenizing liquid 4 five times with a high-pressure homogenizer under nitrogen protection and 500 bar conditions, PDRN lipid nanoparticles were obtained.

[0179] In this embodiment, the percentages of each component are all by mass, and their sum is 100%.

[0180] Example 11

[0181] 20 parts of the PDRN lipid nanoparticles prepared in Example 2 were mixed with 1 part of the fermentation product filtrate of Bifidobacterium / Lactobacillus / Soybean Seed Extract (Huanfuyuan) to obtain the PDRN lipid nanoparticle composition.

[0182] The mass ratio of PDRN liposomes to Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) in this PDRN liposome composition is 20:1.

[0183] Example 12

[0184] 15 parts of the PDRN lipid nanoparticles prepared in Example 2 were mixed with 1 part of the fermentation product filtrate of Bifidobacterium / Lactobacillus / Soybean Seed Extract (Huanfuyuan) to obtain the PDRN lipid nanoparticle composition.

[0185] The mass ratio of PDRN liposomes to Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) in this PDRN liposome composition is 15:1.

[0186] Example 13

[0187] Ten parts of the PDRN liposomes prepared in Example 2 were mixed with one part of the fermentation product filtrate of Bifidobacterium / Lactobacillus / Soybean Seed Extract (Huanfuyuan) to obtain the PDRN liposome composition.

[0188] In this PDRN liposome composition, the mass ratio of PDRN liposomes to Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) is 10:1.

[0189] Example 14

[0190] Five parts of the PDRN liposomes prepared in Example 2 were mixed with one part of the filtrate of Bifidobacterium / Lactobacillus / Soybean Seed Extract Fermentation Product (Huanfuyuan) to obtain the PDRN liposome composition.

[0191] The PDRN liposome composition contains PDRN liposomes in a mass ratio of 5:1 to the fermentation product filtrate of Bifidobacterium / Lactobacillus / Soybean Seed Extract.

[0192] Example 15

[0193] Two parts of the PDRN liposomes prepared in Example 2 were mixed with one part of the fermentation product filtrate of Bifidobacterium / Lactobacillus / Soybean Seed Extract (Huanfuyuan) to obtain the PDRN liposome composition.

[0194] The PDRN liposome composition contains PDRN liposomes in a mass ratio of 2:1 to Bifidobacterium / Lactobacillus / Soybean Seed Extract Fermentation Product Filtrate (Huanfuyuan).

[0195] Example 16

[0196] One part of the PDRN lipid nanoparticles prepared in Example 2 was mixed with one part of the fermentation product filtrate of Bifidobacterium / Lactobacillus / Soybean Seed Extract (Huanfuyuan) to obtain the PDRN lipid nanoparticle composition.

[0197] In this PDRN liposome composition, the mass ratio of PDRN liposomes to Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) is 1:1.

[0198] Example 17

[0199] This embodiment provides an essence for testing efficacy samples. Its raw materials include component A, component B, and water. By total mass, component A contains: 5% dipropylene glycol, 10% glycerin, 15% propylene glycol, and 0.15% carbomer 941; component B contains: 0.15% triethanolamine, 0.5% 1,2-hexanediol / capryloyl hydroxamic acid / ethylhexylglycerin / propylene glycol; and the balance is water.

[0200] The preparation method of the serum includes the following steps:

[0201] (1) Add component A to water and stir at 35~45℃ and 400~500rpm until a clear, uniform liquid without agglomeration is obtained to obtain phase A.

[0202] (2) Add triethanolamine and 1,2-hexanediol / octanoyl hydroxamic acid / ethylhexylglycerin / propylene glycol from component B to phase A while stirring at 400~5000 rpm, and then stir slowly at 200~300 rpm for 15 min until the liquid is homogeneous to obtain blank essence.

[0203] The blank essence is a transparent liquid with a pH between 5 and 7.

[0204] Test Example 1: Cell Proliferation Experiment

[0205] Test samples: compositions prepared in Example 1 and Comparative Examples 1-6, PDRN lipid nanoparticles prepared in Examples 2-4 and Examples 8-10, and PDRN lipid nanoparticle compositions prepared in Examples 11-16.

[0206] HDF cells in the logarithmic growth phase were subjected to 8 × 10⁻⁶ cells. 3The cells were seeded at a density of 100 μL per well in 96-well plates and incubated at 5% CO2 and 37°C for 24 h. Add 100 μL of each well containing: PDRN nanoparticles prepared in Examples 2-4 and 8-10 (PDRN nanoparticle concentration of 500 μg / mL); PDRN nanoparticle compositions prepared in Examples 11-16 (PDRN nanoparticle concentration of 500 μg / mL, and Huanfuyuan concentrations of 25 μg / mL, 33.3 μg / mL, 50 μg / mL, 100 μg / mL, 250 μg / mL, and 500 μg / mL, respectively); the composition prepared in Example 1 (consistent with the active ingredient concentration in PDRN liposomes at 500 μg / mL, i.e., PDRN, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 concentrations of 20 μg / mL, 0.0005 μg / mL, and 0.2 μg / mL, respectively); and compositions prepared in Comparative Examples 1-6 (Comparative Example 1 active ingredient concentration, PDRN concentration of 20 μg / mL). Comparative Example 2: Active ingredient concentration, microcurrent tetrapeptide-1 concentration was 0.0005 μg / mL; Comparative Example 3: Active ingredient concentration, palmitoyl tripeptide-5 concentration was 0.2 μg / mL; Comparative Example 4: Active ingredient concentration, PDRN and microcurrent tetrapeptide-1 concentrations were 20 μg / mL and 0.0005 μg / mL, respectively; Comparative Example 5: Active ingredient concentration, PDRN and palmitoyl tripeptide-5 concentrations were 20 μg / mL and 0.2 μg / mL, respectively; Comparative Example 6: Active ingredient concentration, microcurrent tetrapeptide-1 and palmitoyl tripeptide-5 concentrations were 0.0005 μg / mL and 0.2 μg / mL, respectively. All samples were cultured in DMEM complete medium. A blank control group was added with 100 μL of DMEM complete medium. Each group had three replicates. Cells were cultured for another 48 h, and cell proliferation rate was measured using the CCK-8 assay. The results are shown in Table 2, which presents the cell proliferation experiment results provided in the examples and comparative examples of this application.

[0207] Table 2. Cell proliferation experiment results provided in the embodiments and comparative examples of this application.

[0208]

[0209] Note: Compared with the control group, a p < 0.05, aa p < 0.01; compared with Example 1, b p < 0.05, bb p < 0.01; compared with Example 2, c p < 0.05, cc p < 0.01.

[0210] Table 2 shows that, compared with the blank control group, the cell viability of the compositions prepared in Example 1 and Comparative Examples 1-6, the PDRN nanoparticles prepared in Examples 2-4 and 8-10, and the PDRN nanoparticle compositions prepared in Examples 11-16 were all improved. Compared with Example 1, the cell viability of Comparative Examples 1-6 was significantly reduced, indicating that PDRN, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 have a triple synergistic effect in the free state, and the effect is better than that of single or two-component combinations. The synergistic effect of PDRN depends on the complete three-component system, and its effect is limited in the free two-component combination. Compared with the blank control group, the cell viability of Comparative Examples 1-6 increased by 5.32%, 3.81%, 2.54%, 10.98%, 9.36%, and 5.62%, respectively, and the cell viability of Example 1 increased by 20.36%. Using Comparative Example 1 as agent A and Comparative Example 6 as agent B, the q-value of Example 1 was calculated using the Jin Zhengjun method.

[0211] q=E A+B / (E A +E B -E A *E B )

[0212] The q value of 1.9 further indicates that the combination of PDRN, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 has a synergistic effect.

[0213] Compared with Example 1, the cell survival rate of Example 2 was significantly improved, indicating that the cell survival effect of the active ingredient after being encapsulated by the nanocarrier was significantly enhanced compared with the free active ingredient.

[0214] Compared with Example 2, the cell survival rate of PDRN lipid nanoparticles prepared in Examples 3 and 4 was significantly reduced. Example 2 showed better cell proliferation promotion effect, indicating that specific phospholipid combinations may optimize the biocompatibility or cell affinity of nanocarriers, thereby more effectively promoting cell survival and cell proliferation.

[0215] Compared with Example 2, the PDRN lipid nanoparticles and Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) prepared in Examples 11-16 can promote cell proliferation. In particular, when the mass ratio of PDRN lipid nanoparticles to Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) is (2-15):1, the cell survival rate is significantly improved. This indicates that PDRN lipid nanoparticles and Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) may enhance the effect of PDRN lipid nanoparticles by synergistically promoting the repair of the cell microenvironment and by optimizing the local skin environment through fermentation products, thereby improving the bioavailability of PDRN and peptides.

[0216] Test Example 2: ROS Fluorescence Intensity Detection

[0217] Test samples: compositions prepared in Examples 1 and Comparative Examples 1-6, PDRN lipid nanoparticles prepared in Examples 2-4 and 8-10, and PDRN lipid nanoparticle compositions prepared in Examples 11-16.

[0218] HDF cells were loaded at 4 × 10 4 Inoculate 500 μL per well into a 24-well plate at a density of 1000 cells / well. After 24 h of incubation, discard the supernatant. The model group was given 1 mL of DMEM medium containing 0.6 mmol / L H2O2. Other sample groups were given 1 mL of DMEM medium containing 0.6 mmol / L H2O2 and PDRN liposomes from Examples 2-4 and 8-10 (PDRN liposome concentration of 500 μg / mL), PDRN liposome compositions from Examples 11-16 (PDRN liposome concentration of 500 μg / mL, with revitalizing concentrations of 25 μg / mL, 33.3 μg / mL, 50 μg / mL, 100 μg / mL, 250 μg / mL, and 500 μg / mL, respectively), the composition prepared in Example 1 (consistent with the active ingredient concentration in 500 μg / mL PDRN liposomes, with PDRN, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 concentrations of 20 μg / mL, 0.0005 μg / mL, and 0.2 μg / mL, respectively), and the compositions prepared in Comparative Examples 1-6 (comparative Example 1 active ingredient concentration...). The concentrations of PDRN and microcurrent tetrapeptide-1 were as follows: 20 μg / mL for Comparative Example 2; 0.0005 μg / mL for Comparative Example 3; 0.2 μg / mL for Comparative Example 4; 20 μg / mL for PDRN and 0.0005 μg / mL for microcurrent tetrapeptide-1; 20 μg / mL for PDRN and 0.2 μg / mL for palmitoyl tripeptide-5; and 0.0005 μg / mL for microcurrent tetrapeptide-1 and 0.2 μg / mL for palmitoyl tripeptide-5. For Comparative Example 6, 0.0005 μg / mL for microcurrent tetrapeptide-1 and 0.2 μg / mL for palmitoyl tripeptide-5 were added to DMEM medium. A blank control group without H2O2 was also included. After culturing for 24 h, 1 mL of serum-free DMEM containing 20 µM DCFH-DA was added, and incubation continued for another 20 h. The cells were washed three times with PBS, and the fluorescence intensity was observed under a fluorescence microscope. After collecting the cells, the fluorescence intensity was detected by flow cytometry. The results are shown in Table 3. Table 3 shows the ROS fluorescence intensity detection results of the examples and comparative examples of this application.

[0219] Table 3. ROS fluorescence intensity detection results of the embodiments and comparative examples of this application.

[0220]

[0221] Note: Compared with the model group, a p < 0.05, aa p < 0.01; compared with Example 1, bb p < 0.01; compared with Example 2, cc p < 0.01.

[0222] As shown in Table 3, compared with the model group, the compositions prepared in Example 1 and Comparative Examples 1-6, the PDRN lipid nanoparticles prepared in Examples 2-4 and 8-10, and the PDRN lipid nanoparticle compositions prepared in Examples 11-16 can all significantly inhibit intracellular ROS levels.

[0223] Compared with Example 1, the intracellular ROS levels of Comparative Examples 1-6 were significantly increased, indicating that PDRN, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 have a triple synergistic effect in the free state, and the effect is better than that of single or two-component combinations. The synergistic effect of PDRN depends on the complete three-component system, and its effect in the free combination of two components is limited.

[0224] Compared to the model group, the ROS inhibition rates of the samples prepared in Comparative Examples 1-6 were 17.05%, 10.59%, 5.72%, 26.04%, 23.36%, and 18.13%, respectively. The ROS inhibition rate of the sample prepared in Example 1 was 44.18%. Using Comparative Example 1 as agent A and Comparative Example 6 as agent B, the q-value of Example 1 was calculated using the Jin Zhengjun method.

[0225] q=E A+B / (E A +E B -E A *E B )

[0226] The q value of 1.4 further indicates that the combination of PDRN, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 has a synergistic effect.

[0227] Compared with Example 1, the intracellular ROS levels of the PDRN lipid nanoparticles prepared in Examples 2-4 were significantly reduced, indicating that the active ingredients, after being encapsulated by nanocarriers, significantly enhanced their ROS inhibition and cellular antioxidant effects compared with the free active ingredients.

[0228] Compared with Example 2, the intracellular ROS level of the PDRN lipid nanoparticles prepared in Examples 3 and 4 was significantly increased, and the cellular antioxidant effect of Example 2 was better. This indicates that the specific phospholipid combination may optimize the biocompatibility or cell affinity of the nanocarrier, thereby more effectively inhibiting ROS and promoting the cellular antioxidant effect.

[0229] Compared with Example 2, the PDRN liposomes and Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) compositions prepared in Examples 11-16 can further inhibit ROS. In particular, when the mass ratio of PDRN liposomes to Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) is (2-15):1, the ROS inhibition effect is significant. This further demonstrates that PDRN liposomes and Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) can further support their synergistic effect through the dual pathway of "reducing oxidative damage + promoting repair".

[0230] Test Example 3

[0231] 1. Stability Test 1

[0232] The PDRN lipid nanoparticles from Examples 2-10 were placed at room temperature for 3 months, and then the particle size, PDI and appearance were observed to see if there were any changes; the results are shown in Table 4, which shows the stability test results of the lipid nanoparticles prepared in the examples of this application.

[0233] Table 4. Stability test results of lipid nanoparticles prepared in the embodiments of this application.

[0234]

[0235] Table 4 shows that the average particle size of the PDRN lipid nanoparticles is 100~300nm, and the PDI is 0.1~0.6. By comparing the effects of different phospholipid combinations on the stability of the PDRN lipid nanoparticles in each example, the results show that at room temperature, the particle size and PDI of Examples 6 and 7 are larger than those of Examples 2~5 and Examples 8~10. The PDRN lipid nanoparticles exhibit stratification or chromatography, indicating that the two phospholipid combinations of egg yolk lecithin, natural sphingomyelin, phosphatidylglycerol and phosphatidyltryptophan, natural sphingomyelin, phosphatidylglycerol, and N-(methylpolyoxyethyleneoxycarbonyl)-1,2-distearate-sn-glycerol-3-phosphate ethanolamine sodium salt have an impact on the stability of the carrier.

[0236] 2. Stability Test 2

[0237] After the above experiments, the PDRN lipid nanoparticles prepared in Examples 2-5 and 8-10, which have good room temperature stability, were placed in a sealed container and placed at -20℃, 4℃, and 45℃ for 3 months respectively. The appearance of the PDRN lipid nanoparticles was observed to see whether layering or precipitation occurred. The results are shown in Table 5. Table 5 shows the stability test results of the lipid nanoparticles prepared in the examples of this application.

[0238] Table 5. Stability test results of lipid nanoparticles prepared in the embodiments of this application 2

[0239]

[0240]

[0241] As shown in Table 5, the PDRN nanoparticles prepared in Example 5 exhibited stratification under high temperature conditions in the second month. The PDRN nanoparticles prepared in Examples 2-4 and 8-10 did not show stratification or precipitation after being placed at 4℃ and 45℃ for 3 months, respectively. This indicates that the PDRN nanoparticles prepared in Examples 2-4 and 8-10 have good stability. The phospholipid combination used to prepare the PDRN nanoparticles in Example 5 affects the stability of the carrier at high temperatures.

[0242] Test Example 4: Evaluation of Irritation of Chicken Embryo Allantoic Membrane

[0243] Test samples: PDRN lipid nanoparticles prepared in Examples 2-4 and 8-10, PDRN lipid nanoparticle compositions prepared in Examples 11-14, and the composition of Example 1.

[0244] Test method: Dilute the test sample 10 times, take 0.2 mL, drop it onto the surface of the chorioallantoic membrane, observe the changes in CAM vessels within 5 min and record the initial time of CAM vessel congestion, hemorrhage and coagulation, and calculate the stimulus score IS.

[0245] The stimulus score (IS) is calculated using the following formula:

[0246] IS=[(301-secH)×5+(301-secL)×7+(301-secC)×9] / 300

[0247] In the above formula, secH represents the initial time of congestion (s); secL represents the initial time of bleeding (s); and secC represents the initial time of clotting (s).

[0248] Calculate the mean of repeated trials and classify the irritation of the test substance according to the magnitude of the mean. The mean values ​​of 0~0.9, 1.0~4.9, 5.0~8.9 and 9~21.0 are respectively classified as no irritation, mild irritation, moderate irritation and severe irritation.

[0249] Test results showed that after the PDRN liposomes prepared in Examples 2-4 and 8-10 diluted 10 times, the PDRN liposome compositions prepared in Examples 11-14, and the free composition (Example 1) were in contact with the chicken embryo allantoic membrane for 300 s, there was no capillary bleeding, no vascular dissolution, and no coagulation. The reaction scores were 0.07, 0.06, 0.08, 0.07, 0.06, 0.08, 0.09, 0.08, 0.11, 0.09, and 0.08, respectively.

[0250] Test Example 5 Patch Test

[0251] Fifty subjects were selected, and 10% of the PDRN liposomes prepared in Examples 2-4, 8-10, and PDRN liposome compositions prepared in Examples 11-14, as well as a blank control, were applied to the flexor side of the subjects' forearms for 24 hours. After removing the applicator, the skin reaction was observed after a 30-minute interval until the indentation disappeared. The skin reaction was observed again 24 hours and 48 hours after removing the applicator.

[0252] The results showed that none of the 50 subjects developed light red spots, erythema, edematous erythema, significant redness and swelling, infiltration or papules and papules or vesicles, indicating that the PDRN liposomes prepared in Examples 2-4 and 8-10 and the PDRN liposome compositions prepared in Examples 11-14 are non-irritating to human skin.

[0253] Test Example 6: Observation of Skin Penetration Behavior using Laser Confocal Microscopy

[0254] Transdermal experiments on isolated porcine skin were conducted using the vertical Franz diffusion cell method. The skin was fixed between the receiving and supply chambers. RhoB (0.1%) was loaded into the aqueous phase as a water-soluble dye, and the corresponding RhoB compositions were prepared according to the methods described in Examples 2, 8-16, and the literature. RhoB (0.1%) was also added to the mixture in Example 1 to prepare the corresponding free RhoB compositions. 0.5 g of each sample was placed in the supply chamber, and PBS was used as the receiving solution for diffusion at 32°C with stirring. After 2 h and 4 h, residual sample on the skin was gently wiped away, and the skin from the target area was removed. The skin was rinsed again, thoroughly cleaned, and dried. The samples were frozen and sectioned, and the sections were observed using a laser confocal microscope. The results are shown in Table 6, which presents the experimental results of skin permeation and cellular uptake behavior in the embodiments of this application.

[0255] Table 6. Experimental results of skin penetration and cellular uptake behavior in the embodiments of this application.

[0256]

[0257] Note: Compared with Example 1, aa p < 0.01; compared with Example 2, b p < 0.05, bb p < 0.01.

[0258] As shown in Table 6, compared with Example 1, Examples 2 and 8-10 showed significantly better skin penetration, indicating that the skin penetration of active ingredients was significantly enhanced after being encapsulated by nanocarriers compared with that of free active ingredients.

[0259] Compared to Example 2, at 4 hours, the combination of PDRN nanoparticles from Examples 12-15 with Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) showed significant skin penetration, indicating that the Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) has microecological regulation to assist penetration, enhancing the delivery efficiency of active ingredients through physical penetration. Compared to Example 2, the combination of PDRN nanoparticles from Example 16 with Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) showed decreased skin penetration. This indicates that a mass ratio of (2-15):1 of PDRN nanoparticles to Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) is more effective. At a reasonable low concentration, it may mainly play a role in enhancing penetration, while excessive amounts (such as 1:1) may lead to decreased nanoparticle stability. Organic acids or enzymes in high-concentration fermentation products may interfere with the structure of lipid nanoparticles, thus reducing the delivery efficiency of PDRN nanoparticles.

[0260] Test Example 7: Flow Cytometry Detection of Cell Uptake Behavior

[0261] Test samples: RhoB (addition amount of 0.1%) was loaded into the aqueous phase as a water-soluble dye, and the corresponding RhoB compositions were prepared according to the methods described in Examples 2 and 8-16; RhoB (addition amount of 0.1%) was added to Example 1 to prepare the corresponding free RhoB compositions.

[0262] HDF cells were spaced at 3.0 × 10⁶ cells per well. 5 Cells were seeded at a density of 1000 g / mL in 6-well plates and incubated for 24 h to allow cell adhesion. The old culture medium was discarded, and each well was then filled with DMEM complete medium diluted 1000-fold with the RhoB compositions prepared in Examples 2, 8-16, and the free RhoB composition prepared in Example 1 (RhoB concentration 1 μg / mL). Untreated cells served as a negative control. After culturing for 2 h and 4 h, the cells were washed with cold PBS, trypsinized, centrifuged, and the cell pellet was collected and resuspended in 0.5 mL of cold PBS. Intracellular fluorescence intensity was detected by flow cytometry. The results are shown in Table 7, which presents the flow cytometry results of cell uptake behavior in this application.

[0263] Table 7. Experimental results of cell uptake behavior detected by flow cytometry in this application embodiment.

[0264]

[0265] Note: Compared with Example 1, aa p < 0.01; compared with Example 2, b p < 0.05, bb p < 0.01.

[0266] As shown in Table 7, compared with Example 1, Examples 2 and 8-10 showed significantly better cell uptake, indicating that the cell uptake of active ingredients was significantly enhanced after being encapsulated by nanocarriers compared with that of free active ingredients.

[0267] Compared to Example 2, the PDRN nanoparticles combined with the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract (Huanfuyuan) in Examples 12-14 showed significant cellular uptake effects, indicating that the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract (Huanfuyuan) has microecological regulation to assist in penetration and synergistically enhance cellular uptake, improving the delivery efficiency of active ingredients through physical penetration enhancement and bioactivity synergy. Compared to Example 2, there was no significant difference in cellular uptake effects between the PDRN nanoparticles combined with the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract (Huanfuyuan) in Examples 11 and 16. This indicates that a mass ratio of PDRN nanoparticles to fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract (Huanfuyuan) of (2-15):1 is more effective, possibly because the biosurfactant components in the fermentation filtrate (such as fermentation-produced peptides or polysaccharides) slightly modify the surface of the nanoparticles, forming a more stable "invisible" hydrophilic layer (similar to a PEGylation effect), reducing macrophage clearance and enhancing the specific uptake by target cells (such as fibroblasts). When the ratio of regenerator to liposome is too high (1:1), excessive fermentation components may over-encapsulate lipid nanoparticles, leading to surface charge neutralization or reversal (such as negatively charged fermentation products covering positively charged lipids), reducing electrostatic adsorption to the cell membrane. When the ratio of regenerator to liposome is too low (20:1), the absorption-promoting effect decreases. Secondly, due to steric hindrance, it may hinder the binding of lipid nanoparticles to cell membrane receptors (such as CD36 or SR-B1-mediated endocytosis).

[0268] Test Example 8: Detection of Anti-aging Factors

[0269] Test samples: PDRN lipid nanoparticles prepared in Examples 2, 8-10, PDRN lipid nanoparticle compositions prepared in Examples 11-16, and the free composition of Example 1.

[0270] HDF cells were loaded at 4 × 10 4Cells were seeded at a density of 500 μL per well into 24-well cell culture plates and cultured at 37°C with 5% CO2 for 24 h. 1 mL of DMEM complete medium containing PDRN nanoparticles prepared in Example 2 (PDRN nanoparticle concentration of 500 μg / mL), PDRN nanoparticles prepared in Examples 8-10 (PDRN nanoparticle concentration of 500 μg / mL), PDRN nanoparticle compositions prepared in Examples 11-16 (PDRN nanoparticle concentration of 500 μg / mL, with the same concentration as in Test Examples 1 and 2), and the free composition of Example 1 (active substance test concentration as in Test Examples 1 and 2) was added to each well. The blank control group received only 100 µL of DMEM complete medium. Each group had 3 replicates. The plates were incubated in a CO2 incubator for 24 h. The supernatant was collected, and the Col I and Col III contents were tested using an ELISA kit. The results are shown in Table 8, which presents the Col I and Col III contents results from the examples in this application.

[0271] Table 8. Results of Col I and Col III content in the embodiments of this application.

[0272]

[0273] Note: Compared with the model group, a p < 0.05, aa p < 0.01; compared with Example 1, b p < 0.05, bb p < 0.01; compared with Example 2, cc p < 0.01

[0274] As shown in Table 8, compared with the model group, Examples 2, 8-16, and 1 all promoted the secretion of Col I and Col III by cells. Compared with Example 1, Examples 2 and 8 significantly increased the content of Col I and Col III, indicating that the active ingredients, after being encapsulated, have a more significant effect on promoting collagen synthesis and anti-aging, which is significantly better than the same dose of free ingredients. Compared with Example 2, the PDRN liposomes prepared in Examples 12-15 and the filtrate of Bifidobacterium / Lactobacillus / soybean seed extract fermentation product (Huanfuyuan) significantly increased the content of COLⅠ and COLⅢ. The PDRN liposome compositions prepared in Examples 11 and 16 showed no significant difference in the synthesis of COLⅠ and COLⅢ. This may be because the mass ratio of PDRN liposomes to Bifidobacterium / Lactobacillus / soybean seed extract fermentation product (Huanfuyuan) (2-15):1 is more conducive to maintaining the integrity of the liposomes and ensuring the continuous release of PDRN to the target cells. The surface-active components in the fermentation filtrate (such as polysaccharides or peptides produced by fermentation) may disrupt the lipid bilayer structure at a high ratio (e.g., 1:1), leading to premature release or degradation of PDRN. On the other hand, if the ratio of fermentation filtrate is too low (e.g., 20:1), the absorption-promoting effect will decrease, and PDRN cannot be continuously released to the target cells. This study demonstrates that PDRN liposomes and Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) may synergistically promote collagen synthesis and achieve significant anti-aging effects when used in a specific ratio.

[0275] Test Example 9: Effects on Inflammatory Factors

[0276] Test samples: PDRN lipid nanoparticles prepared in Examples 2 and 8-10, PDRN lipid nanoparticle compositions prepared in Examples 11-16, and the free composition of Example 1.

[0277] HDF cells were loaded at 1.0 × 10⁻⁶ 5 The cells were seeded at a density of 1 mL / mL into 24-well cell culture plates. After overnight incubation, TNF-α / INF-γ (final concentration of 10 ng / mL) and 1 mL of sample were added for co-incubation. DMEM complete medium containing the following test samples were then added: PDRN nanoparticles prepared in Examples 2 and 8-10 (PDRN nanoparticle concentration of 500 μg / mL), PDRN nanoparticle compositions prepared in Examples 11-16 (PDRN nanoparticle concentration of 500 μg / mL, Huanfuyuan test concentration same as in Examples 1 and 2), and the free composition of Example 1 (test concentration same as in Example 8). Five replicates were used for each group. After 24 h, the cell supernatant was collected, and the levels of IL-6 and TNF-α secreted by HDF cells were measured using an ELISA kit. The results are shown in Table 9, which presents the IL-6 and TNF-α content results of the examples in this application.

[0278] Table 9. Results of IL-6 and TNF-α content in the embodiments of this application.

[0279]

[0280] Note: Compared with the model group, a p < 0.05, aa p < 0.01; compared with Example 1, b p < 0.05, bb p < 0.01; compared with Example 2, cc p < 0.01

[0281] As shown in Table 9, compared with the model group, Examples 2, 8-10, 11-16 and Example 1 all inhibited the secretion of IL-6 and TNF-α by cells. Compared with Example 1, Examples 2 and 8 significantly inhibited the content of IL-6 and TNF-α, indicating that the anti-inflammatory effect of the active ingredient after encapsulation is more significant and is significantly better than that of the same dose of free ingredient. Compared with Example 2, the PDRN liposomes prepared in Examples 12-15 and the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract (Huanfuyuan) significantly inhibited the secretion of IL-6 and TNF-α. The PDRN liposomes prepared in Examples 11 and 16 and the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract (Huanfuyuan) had no significant effect on inhibiting the secretion of IL-6 and TNF-α. This may be because the fermentation product of Bifidobacterium / Lactobacillus / soybean seed extract may contain probiotic metabolites (such as short-chain fatty acids and peptides), which have dual immunomodulatory effects. At a suitable low concentration ratio, it may slightly activate TLR2 / 4 or PPAR-γ, promote the secretion of anti-inflammatory cytokines (such as IL-10), indirectly inhibit IL-6 / TNF-α, and synergistically enhance the anti-inflammatory signal of A2A receptor with PDRN. High concentrations may overstimulate TLR or NLRP3 inflammasomes, and organic acids in the fermentation products (such as lactic acid) may lower the pH, affecting PDRN stability or cellular uptake. Conversely, excessively low concentrations lead to decreased absorption and prevent the sustained release of active ingredients to target cells, all resulting in reduced anti-inflammatory effects. This indicates that PDRN liposomes and the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract (Huanfuyuan) at a specific ratio may synergistically inhibit the secretion of inflammatory factors, achieving a significant anti-inflammatory effect.

[0282] Test Example 10: Expression of Adenosine A2A Receptor (qRT-PCR)

[0283] Test samples: PDRN lipid nanoparticles prepared in Examples 2, 8-10, PDRN lipid nanoparticle compositions prepared in Examples 11-16, and the free composition of Example 1.

[0284] HDF cells were loaded at 3 × 10 5 Cells were seeded at a density of 500 μL / well into 6-well cell culture plates and cultured at 37°C with 5% CO2 for 24 h. 2 mL of DMEM complete medium containing PDRN nanoparticles prepared in Examples 2 and 8-10 (PDRN nanoparticle concentration 500 μg / mL), PDRN nanoparticle compositions prepared in Examples 11-16 (PDRN nanoparticle concentration 500 μg / mL, with the same concentration as in Test Examples 1 and 2), and the free composition of Example 1 (active ingredient concentration same as in Test Examples 1 and 2) was added to each well. The blank control group received only 100 µL of DMEM complete medium. The plates were incubated in a CO2 incubator for 24 h. Cells were collected, and total RNA was extracted using an RNA extraction kit. RNA concentration and purity were analyzed using a microplate reader. Complementary DNA synthesis was performed using an M-MLV reverse transcriptase kit. Quantitative real-time PCR was performed using a real-time PCR instrument according to the operating procedures of the one-step quantitative RT-PCR MasterMix kit. Primer sequences and product sizes are shown in Table 10. Using ACTIN as an internal reference, the relative expression of A2A mRNA was quantified using the 2-ΔΔCt method based on the Δcycle threshold (Ct). The control group result was set to 1, and the ratio of the control group to ACTIN was used as the relative value of the corresponding protein mRNA expression level. Each sample was tested in triplicate, and the results were averaged. The results are shown in Table 11, which presents the expression results of adenosine A2A receptor (qRT-PCR).

[0285] Table 10 Primer sequences and product sizes

[0286]

[0287] Table 11. Expression of adenosine A2A receptor (qRT-PCR) results.

[0288]

[0289] Note: Compared with the blank control group, a p < 0.05, aa p < 0.01; compared with Example 1, bb p < 0.01; compared with Example 2, cc p < 0.01.

[0290] As shown in Table 11, compared with the blank control group, Examples 2, 8-10, 11-16 and 1 all showed the expression effect of adenosine A2A receptor;

[0291] Compared to Example 1, Examples 2 and 8 showed significantly enhanced adenosine A2A receptor expression, indicating that the expression of adenosine A2A receptor in the encapsulated PDRN nanoparticles was more significant and significantly superior to that of the same dose of free component. Compared to Example 2, the PDRN nanoparticle compositions prepared in Examples 12-15 showed significantly increased adenosine A2A receptor expression, possibly because the fermentation filtrate at low concentrations synergistically enhanced receptor expression rather than inhibited it; maintaining nanoparticle stability ensured effective PDRN delivery. Compared to Example 2, the PDRN nanoparticle composition prepared in Example 16 showed no significant difference in adenosine A2A receptor expression, possibly because a high proportion of fermentation filtrate led to reduced adenosine production or receptor downregulation due to metabolic interference, pH changes, or carrier destruction; while a low proportion of fermentation filtrate resulted in decreased absorption promotion, preventing the continuous release of the active ingredient to target cells, leading to reduced adenosine production or receptor downregulation. This indicates that the combined use of PDRN liposomes and Bifidobacterium / Lactobacillus / Soybean Seed Extract Fermentation Product Filtrate (Huanfuyuan) is more effective than PDRN liposomes alone, and that PDRN liposomes and Bifidobacterium / Lactobacillus / Soybean Seed Extract Fermentation Product Filtrate (Huanfuyuan) maintain the PDRN-dominated adenosine-A2A receptor signaling pathway at a specific ratio.

[0292] Test Example 11: Nuclease Degradation Experiment

[0293] Test samples: PDRN lipid nanoparticles prepared in Example 2, PDRN lipid nanoparticle composition prepared in Example 12, and free composition of Example 1.

[0294] Take 20 μL each of the PDRN lipid nanoparticles prepared in Example 2, the composition prepared in Example 12, and the free composition from Example 1, and place them in enzyme-free EP tubes. Add an equal volume of DNase I solution (enzyme concentration of 1 U) to each reaction system. Place the reaction system in a constant temperature water bath, set the reaction temperature to 37°C according to the kit instructions, and maintain the same reaction time to ensure experimental consistency. After the reaction is complete, terminate the reaction by adding an appropriate amount of EDTA and chelate the free DNase I. The system is then heated at 65°C for 10 min to inactivate the DNase I enzyme. Then, add 2.5 μL of 1× loading buffer and 7.5 μL of each sample to the sample plate, mix well, and use a 10 μL micropipette to add the samples to the sample slots of the gel plate. Immediately after sample addition, electrophoresis is performed on the gel plate at a voltage of 190V. Electrophoresis is stopped when the bromophenol blue moves to about 1 cm from the bottom edge of the gel plate. After electrophoresis, the gel was removed and observed under a UV lamp. The presence of PDRN was indicated by fluorescent bands. Images were then taken and saved using a gel imaging system. The results are shown below. Figure 1 , Figure 1 The electrophoresis results provided in the embodiments of the present invention.

[0295] like Figure 1 As shown, the PDRN fragment sizes in the PDRN nanoparticles prepared in Example 2, the composition prepared in Example 12, and the free composition in Example 1 are in the range of 50-2000 bp, consistent with literature reports. After nuclease degradation, the bands in the free composition of Example 1 are concentrated below 100 bp, indicating that they have been degraded into small molecular fragments; the PDRN nanoparticles prepared in Example 2 are degraded to a certain extent, but some large molecular fragments are retained, and the effect is better than that of the free composition of Example 1, indicating that the carrier encapsulation can inhibit the complete degradation of PDRN raw materials by nucleases; while the composition prepared in Example 12 is degraded, but more large molecular fragments are retained, and the effect is better than that of the PDRN nanoparticles prepared in Example 2, indicating that the PDRN nanoparticles prepared in Example 2, when used in combination with the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract (Huanfuyuan), have a synergistic protective effect on PDRN.

[0296] Test Example 12: Human Efficacy Evaluation

[0297] Test Methods: 180 healthy women (aged 35-55) were selected and divided into 18 groups of 10 each. Each group used the products obtained in Examples 2, 12, and 1, respectively. Participants used the samples twice daily (morning and evening) for 56 days. Participants were not allowed to use other products during the study period. Results were evaluated after 56 days of sample use. Before the trial, skin wrinkles and skin elasticity were measured. After using the serum containing 5% of each sample (5% sample + 95% blank serum prepared in Example 17) for 7, 14, 28, and 56 days, skin wrinkles and skin elasticity were measured again. Facial wrinkles were measured using the Shanghai Fuhuan Vplus® Intelligent Skin Analysis System; skin elasticity was measured using a Cutometer® dual MPA580 skin elasticity meter, and the average value for each group of 30 participants was calculated.

[0298] Wrinkle change rate (%) = (average wrinkle value of each group after using the product in the example / comparative example - average wrinkle value of each group before use) / average wrinkle value of each group before use * 100%. The wrinkle improvement rate is calculated by the formula and then the positive value is taken. The results are shown in Table 12.

[0299] Table 12 Wrinkle improvement rate after different periods of product use in the examples

[0300]

[0301] As shown in Table 12, both the products of Examples 2 and 12 exhibit significant wrinkle-reducing effects after use, with better results observed over longer usage. The wrinkle-reducing and anti-aging properties of Examples 2 and 12 are significantly superior to those of Example 1, indicating that the synergistic effect of the three components—PDRN, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5—encapsulated in the nanocarrier of this invention is crucial for maximizing the wrinkle-reducing and anti-aging effects of the active ingredients. Compared to the free component (Example 1), the wrinkle-reducing and anti-aging performance of the encapsulated PDRN nanoparticles is significantly better than that of the same dosage of the free component. Comparing Example 2 and Example 12, the wrinkle-reducing effect of the PDRN nanoparticle composition (PDRN nanoparticles combined with Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan)) is significantly better than that of PDRN nanoparticles alone, demonstrating that the combined use of PDRN nanoparticles and Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate (Huanfuyuan) has a synergistic effect, achieving better anti-aging efficacy.

[0302] The above are merely preferred embodiments of the present invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A composition containing PDRN, characterized in that, It is composed of polydeoxyribonucleotides, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5; The mass ratio of the polydeoxyribonucleotide, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 is 0.5~4.5:0.00001~0.0003:0.0005~0.5; The peptide sequence of microcurrent tetrapeptide-1 is LEAP.

2. The composition according to claim 1, characterized in that, The mass ratio of polydeoxyribonucleotide, microcurrent tetrapeptide-1, and palmitoyl tripeptide-5 is 1~4:0.00005~0.0001:0.001~0.

1.

3. A lipid nanoparticle containing PDRN, characterized in that, The composition includes the composition according to any one of claims 1 to 2 and the lipid nanoparticle raw material, wherein the lipid nanoparticle raw material includes at least two of phospholipids, non-phospholipid functional lipids, emulsifiers, alcohols, and water.

4. The lipid nanoparticles according to claim 3, characterized in that, include: 0.5wt%~4.5wt% polydeoxyribonucleotides; 0.00001wt%~0.0003wt% of microcurrent tetrapeptide-1; 0.0005wt%~0.5wt% palmitoyl tripeptide-5; 0.1wt%~3wt% phospholipids; 0.1wt%~3wt% of non-phospholipid functional lipids; 10wt%~40wt% emulsifier; 10wt%~50wt% of alcohols; The remaining water.

5. The lipid nanoparticles according to claim 4, characterized in that, The phospholipid is one or more of the following: lecithin, distearate phosphatidylcholine, natural sphingomyelin, phosphatidylglycerol, phosphatidylinositol, and phosphatidyltryptophan; The non-phospholipid functional lipids are one or more of the following: stearamide, oleoyl fatty amine derivatives, polyethylene glycol stearate N-hydroxysuccinimide ester, and sodium salt of N-(methylpolyoxyethyleneoxycarbonyl)-1,2-distearate-sn-glycerol-3-phosphate ethanolamine. The emulsifier is one or more selected from polyoxyethylene castor oil emulsifier, polyoxyethylene hydrogenated castor oil emulsifier, polyglycerol emulsifier, poloxamer, cocoyl glucoside, polyglycerol-10 laurate, glyceryl citrate stearate, sodium bis(lauramide glutamine) lysine, triglyceride emulsifier, lauryl ether-23, PPG-26-butanol ether-26, tridecyl alcohol ether-12, polyglycerol ester, polysorbate-80, polysorbate-60, polysorbate-20, PEG-20 hydrogenated castor oil, PEG-40 hydrogenated castor oil, and PEG-60 hydrogenated castor oil. The alcohol compounds are one or more selected from glycerol, 1,2-propanediol, 1,2-butanediol, 1,3-butanediol, 1,4-butanediol, 2,3-butanediol, 1,3-propanediol, 1,2-pentanediol, ethoxydiethylene glycol, 1,2-hexanediol, dipropylene glycol, isopropanol, methylpropanediol, polyethylene glycol-200, PPG-10 sorbitol, and octyldodecyl alcohol.

6. The method for preparing lipid nanoparticles according to any one of claims 3 to 5, characterized in that, Includes the following steps: (1) Mix polydeoxyribonucleotides, emulsifiers, water and a first mass fraction of alcohol to obtain mixture 1; Mixture 2 is prepared by mixing phospholipids, non-phospholipid functional lipids, and a second mass fraction of alcohol compounds. Mixture 3 was prepared by mixing microcurrent tetrapeptide-1, palmitoyl tripeptide-5, emulsifier and a third part by weight of alcohol compound; (2) Mix mixture 3 with mixture 1, and then mix it with mixture 2 to obtain mixture 4; (3) The mixture 4 was nano-sized to obtain lipid nanoparticles.

7. A composition containing lipid nanoparticles, characterized in that, Includes the lipid nanoparticles and Bifidobacterium / Lactobacillus / soybean seed extract fermentation product filtrate as described in any one of claims 3 to 5; The mass ratio of the lipid nanoparticles to the fermentation product filtrate of Bifidobacterium / Lactobacillus / soybean seed extract is 1~20:

1.

8. A personal care product, characterized in that, include: The composition according to any one of claims 1 to 2, the lipid nanoparticles according to any one of claims 3 to 5, and / or the composition containing lipid nanoparticles according to claim 7.

9. The use of the composition according to any one of claims 1 to 2, the lipid nanoparticles according to any one of claims 3 to 5, and / or the composition containing lipid nanoparticles according to claim 7 in the preparation of anti-aging, anti-inflammatory, soothing, and repairing products.

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