A fibroblast energy-activated anti-aging nano composition and its preparation method and application

Abstract

The present application relates to the field of medical devices, the field of medical or cosmetic formulations, the field of biopharmaceutical manufacturing technology and the field of preparation manufacturing technology, in particular to a kind of targeted fibroblast energy-activated anti-aging nano composition and its preparation method and application.The nano composition includes: nanoparticle, surfactant, polyol, targeting peptide and water;The nanoparticle includes: active ingredient, encapsulating material, phospholipid and stabilizer;The encapsulating material wraps the active ingredient to form inclusion compound, the phospholipid forms phospholipid bilayer to wrap the inclusion compound, and the stabilizer is embedded in the phospholipid bilayer.The nano composition provided by the present application has excellent appearance, stability of content, particle size and the like, and can efficiently and specifically target fibroblasts and significantly improve the ATP level of fibroblasts, and has excellent energy and anti-aging efficacy of activating fibroblasts.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of medical devices, the field of formulations for medical or cosmetic use, the field of biopharmaceutical manufacturing technology and the field of formulation manufacturing technology, in particular to a fibroblast-targeted anti-aging nano-composition and a preparation method and application thereof. BACKGROUND

[0002] Fibroblasts, as the most predominant cell type in the dermis layer of skin, play a core role in maintaining the structural stability and functional integrity of skin. They are the main producers of extracellular matrix (ECM), responsible for synthesizing key structural proteins such as collagen, elastin, fibronectin, and involved in regulating the structural integrity of the dermal-epidermal junction (DEJ). With age, endogenous and exogenous aging factors together cause the gradual decline of fibroblast function, manifested as decreased synthesis capacity, replicative senescence, and the production of senescence-associated secretory phenotype (SASP), ultimately leading to visible signs of aging such as skin laxity, wrinkle formation, and loss of elasticity. However, most of the anti-aging compositions on the market have poor targeting, unclear efficacy mechanism, etc.

[0003] Currently, the strategies for skin anti-aging mainly include: supplementing exogenous active ingredients (such as vitamin C, retinol, etc.) to stimulate collagen production; applying antioxidants (such as vitamin E, coenzyme Q10, etc.) to resist oxidative stress damage; and promoting skin reconstruction through chemical peeling, laser, etc. However, the above methods have obvious limitations. Most active ingredients have small molecular weight, poor stability, are easy to inactivate during preparation or storage, and are difficult to be effectively absorbed through the skin and targeted to the fibroblasts in the dermis layer. When conventional formulations act on the skin, they are often largely removed or metabolized in the epidermis, and cannot reach effective therapeutic concentrations at the target site. At the same time, non-targeted broad action may cause skin irritation, dryness, sensitivity, and other adverse reactions. Therefore, developing a delivery system that can efficiently penetrate the skin barrier and continuously release effective ingredients is the key to improving anti-aging effects and reducing side effects.

[0004] Nanotechnology provides a new way to solve the above problems. Nanocarriers (such as liposomes, polymeric nanoparticles, nanoemulsions, etc.) can promote the transdermal absorption of active ingredients due to their size advantage, and can achieve cell-targeted delivery through surface modification with targeting peptides. However, in the prior art: phospholipids are prone to oxidation and hydrolytic degradation, and single lecithin carriers are not conducive to the stability of active substances, and nano-compositions specially designed for efficient targeting of fibroblasts in the dermis layer and simultaneously addressing multiple aging mechanisms are still rare.

[0005] Therefore, there is an urgent need to develop an anti-aging composition with good stability and high targeting. SUMMARY

[0006] To address the aforementioned technical problems, the present invention provides the following technical solutions.

[0007] In a first aspect, the present invention provides a nanocomposition.

[0008] A nanocomposition comprising: nanoparticles, surfactant, polyol, targeting peptide, and water;

[0009] The nanoparticles include: active ingredients, encapsulating materials, phospholipids, and stabilizers;

[0010] The encapsulating material encapsulates the active ingredient to form an inclusion complex, the phospholipid forms a phospholipid bilayer that encapsulates the inclusion complex, and the stabilizer is embedded in the phospholipid bilayer;

[0011] The nanoparticles are dispersed in the surfactant, polyol, and water;

[0012] The targeted peptide modifies the surface of the nanoparticles;

[0013] The encapsulating material includes at least one of hydroxypropyl-α-cyclodextrin and hydroxypropyl-β-cyclodextrin;

[0014] The phospholipids include lecithin;

[0015] The stabilizer includes phytol;

[0016] The surfactant includes at least one of oleyl alcohol polyether-20 and sucrose stearate;

[0017] Based on the total weight of the nanocomposition, the content of the active ingredient is 1 wt% to 11 wt%, the content of the encapsulating material is 5 wt% to 20 wt%, the content of the phospholipid is 0.1 wt% to 5 wt%, the content of the stabilizer is 1 wt% to 5 wt%, the content of the surfactant is 0.6 wt% to 21 wt%, the content of the polyol is 5 wt% to 30 wt%, the content of the targeting peptide is 0.001 wt% to 0.01 wt%, and the content of water is 39 wt% to 70 wt%.

[0018] This invention encapsulates the active ingredient in a cyclodextrin-phospholipid system and uses at least one of oleyl alcohol polyether-20 and sucrose stearate as a surfactant. This surfactant not only has a synergistic effect with the cyclodextrin system, but also increases the stability and transdermal absorption rate of lecithin.

[0019] Phytol (CAS No. 7541-49-3) is an unsaturated higher alcohol containing 20 carbon atoms, with the molecular formula C6H2O. 20 H 40O, belongs to the diterpenoid class of compounds. This substance is naturally present in the molecular structure of chlorophyll and has attracted attention in the skincare industry due to its unique biological activity. Known as "phytoretinol," it provides anti-aging effects similar to retinol but is generally gentler. It can activate RXR / PPAR receptors, promoting collagen production; terminate free radical chain reactions; and inhibit inflammatory factors such as IL-1β and NF-κB. In this invention, phytol also acts as a stabilizer, embedding itself in the interstitial spaces of lecithin to form terpene bodies, enhancing the overall stability of the raw material, preventing degradation, and ensuring the active ingredients are not deactivated.

[0020] In some embodiments, based on the total weight of the nanocomposition, the content of the active ingredient is 1.1 wt% to 11 wt%, the content of the encapsulating material is 5 wt% to 20 wt%, the content of the phospholipid is 0.1 wt% to 5 wt%, the content of the stabilizer is 1 wt% to 5 wt%, the content of the surfactant is 0.6 wt% to 21 wt%, the content of the polyol is 5 wt% to 30 wt%, the content of the targeting peptide is 0.001 wt% to 0.01 wt%, and the content of water is 39.694 wt% to 69.594 wt%.

[0021] In some embodiments, based on the total weight of the nanocomposition, the content of the active ingredient is 1.1 wt% to 11 wt%, the content of the encapsulating material is 5 wt% to 20 wt%, the content of the phospholipid is 0.1 wt% to 5 wt%, the content of the stabilizer is 1 wt% to 5 wt%, the content of the surfactant is 0.6 wt% to 21 wt%, the content of the polyol is 5 wt% to 30 wt%, the content of the targeting peptide is 0.001 wt% to 0.01 wt%, and the balance is water.

[0022] In some embodiments, the content of the active ingredient, calculated based on the total weight of the nanocomposition, is 1 wt%, 1.1 wt%, 1.5 wt%, 2 wt%, 2.3 wt%, 2.5 wt%, 3 wt%, 4 wt%, 5 wt%, 6 wt%, 7 wt%, 8 wt%, 9 wt%, 10 wt%, 11 wt%, or any value between any two of these values.

[0023] In some embodiments, the content of the encapsulating material, calculated based on the total weight of the nanocomposition, is 5 wt%, 6 wt%, 7 wt%, 8 wt%, 9 wt%, 10 wt%, 11 wt%, 12 wt%, 13 wt%, 14 wt%, 15 wt%, 16 wt%, 17 wt%, 18 wt%, 19 wt%, 20 wt%, or any value between any two of these values.

[0024] In some embodiments, the phospholipid content, calculated by the total weight of the nanocomposition, is 0.1 wt%, 0.5 wt%, 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%, 4 wt%, 5 wt%, or any value between any two of these values.

[0025] In some embodiments, the stabilizer content is 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%, 4 wt%, 5 wt%, or any value between any two of these values, calculated based on the total weight of the nanocomposition.

[0026] In some embodiments, the surfactant content is 0.6 wt%, 1 wt%, 5 wt%, 6 wt%, 10 wt%, 15 wt%, 20 wt%, 21 wt%, or any value between any two of these, calculated based on the total weight of the nanocomposition.

[0027] In some embodiments, the content of the polyol, calculated by the total weight of the nanocomposition, is 5 wt%, 6 wt%, 10 wt%, 15 wt%, 20 wt%, 25 wt%, 30 wt%, or any value between any two of these values.

[0028] In some embodiments, the content of the targeting peptide, calculated based on the total weight of the nanocomposition, is 0.001 wt%, 0.002 wt%, 0.003 wt%, 0.004 wt%, 0.005 wt%, 0.006 wt%, 0.007 wt%, 0.008 wt%, 0.009 wt%, 0.01 wt%, or any value between any two of these values.

[0029] In some embodiments, the water content, calculated based on the total weight of the nanocomposition, is 39.694 wt%, 40 wt%, 45 wt%, 48.994 wt%, 50 wt%, 52.694 wt%, 54.694 wt%, 55 wt%, 57.69 wt%, 57.694 wt%, 57.699 wt%, 58.094 wt%, 58.694 wt%, 60 wt%, 65 wt%, 68.894 wt%, 69 wt%, or 69.594 wt%. In some embodiments, the weight ratio of oleyl alcohol polyether-20 to sucrose stearate is 5:1 to 20:1. In some embodiments, the weight ratio of oleyl alcohol polyether-20 to sucrose stearate is 5:1, 6:1, 7:1, 8:1, 9:1, 10:1, 11:1, 12:1, 13:1, 14:1, 15:1, 16:1, 17:1, 18:1, 19:1, 20:1, or any value between any two of these values.

[0030] In some embodiments, the polyol includes at least one selected from dipropylene glycol, 1,2-pentanediol, 1,3-butanediol, hexyldecyl alcohol, 1,2-propanediol, 1,3-propanediol, 1,2-hexanediol, and methylpropanediol.

[0031] In some embodiments, the polyol comprises a first polyol and a second polyol, wherein the first polyol comprises at least one selected from dipropylene glycol and methylpropanediol; and the second polyol comprises 1,2-pentanediol. In some embodiments, the polyol is composed of a first polyol and a second polyol, wherein the first polyol is selected from at least one selected from dipropylene glycol and methylpropanediol; and the second polyol is 1,2-pentanediol.

[0032] In some embodiments, the weight ratio of the first polyol to the second polyol is 2:3 to 5:1. In some embodiments, the weight ratio of the first polyol to the second polyol is 2:3, 1:1, 1.5:1, 2:1, 2.5:1, 3:1, 4:1, 5:1, or any value between any two of these values.

[0033] In some embodiments, the weight ratio of dipropylene glycol to methylpropanediol is 0:1 to 6.5:1. In some embodiments, the weight ratio of dipropylene glycol to methylpropanediol is 0:1, 0.5:1, 1:1, 1.5:1, 2:1, 2.5:1, 3:1, 3.5:1, 4:1, 4.5:1, 5:1, 5.5:1, 6:1, 6.5:1, or any value between any two of these values.

[0034] In some embodiments, the active ingredient includes at least one of silymarin and silanetriol.

[0035] Silanetriol is a type of organosilicon compound that possesses the natural bioactive form of silicon in the human body. Its small molecular structure gives it extremely high skin affinity and permeability, allowing it to be directly utilized by the skin. It can also regulate the skin's immune response, effectively relieving external stimuli, and promote the synthesis of collagen and elastin by fibroblasts.

[0036] In some embodiments, the targeting peptide includes acetyl tetrapeptide-3.

[0037] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 2 wt% lecithin, 8 wt% dipropylene glycol, 2 wt% methyl propylene glycol, 2 wt% phytol, 5 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0038] In some embodiments, the nanocomposition comprises the following components in the following proportions: 1 wt% silymarin, 0.1 wt% silanetriol, 5 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 2 wt% lecithin, 8 wt% dipropylene glycol, 2 wt% methyl propylene glycol, 2 wt% phytol, 5 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0039] In some embodiments, the nanocomposition comprises the following components in the following proportions: 10 wt% silymarin, 1 wt% silanetriol, 20 wt% hydroxypropyl-β-cyclodextrin, 0.1 wt% sucrose stearate, 2 wt% lecithin, 8 wt% dipropylene glycol, 2 wt% methyl propylene glycol, 2 wt% phytol, 0.9 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0040] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 2 wt% lecithin, 8 wt% dipropylene glycol, 2 wt% methyl propylene glycol, 1 wt% phytol, 5 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0041] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 2 wt% lecithin, 8 wt% dipropylene glycol, 2 wt% methyl propylene glycol, 5 wt% phytol, 5 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0042] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 0.1 wt% lecithin, 2 wt% dipropylene glycol, 1 wt% methyl propylene glycol, 2 wt% phytol, 5 wt% oleyl alcohol polyether-20, 2 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0043] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 5 wt% lecithin, 15 wt% dipropylene glycol, 10 wt% methyl propylene glycol, 2 wt% phytol, 5 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0044] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 2 wt% lecithin, 2 wt% methyl propylene glycol, 2 wt% phytol, 20 wt% oleyl alcohol polyether-20, 3 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0045] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 0.1 wt% sucrose stearate, 2 wt% lecithin, 13 wt% dipropylene glycol, 2 wt% methyl propylene glycol, 2 wt% phytol, 0.5 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.006 wt% acetyl tetrapeptide-3, with the balance being water.

[0046] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 2 wt% lecithin, 8 wt% dipropylene glycol, 2 wt% methyl propylene glycol, 2 wt% phytol, 5 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.01 wt% acetyl tetrapeptide-3, with the balance being water.

[0047] In some embodiments, the nanocomposition comprises the following components in the following proportions: 2 wt% silymarin, 0.3 wt% silanetriol, 15 wt% hydroxypropyl-β-cyclodextrin, 1 wt% sucrose stearate, 2 wt% lecithin, 8 wt% dipropylene glycol, 2 wt% methyl propylene glycol, 2 wt% phytol, 5 wt% oleyl alcohol polyether-20, 5 wt% 1,2-pentanediol, 0.001 wt% acetyl tetrapeptide-3, with the balance being water.

[0048] In a second aspect, the present invention provides a method for preparing the nanocomposition described in the first aspect.

[0049] A method for preparing the nanocomposition described in the first aspect, characterized by comprising the following steps:

[0050] (1) Preparation of phase A: The active ingredient is mixed with ethanol, heated and stirred to obtain phase A;

[0051] (2) Preparation of phase B: The encapsulating material is dissolved in water, heated and stirred to obtain phase B;

[0052] (3) Preparation of inclusion complex: Under stirring conditions, phase A and phase B are mixed, stirred evenly, and dried to obtain inclusion complex;

[0053] (4) Preparation of phase C: Sucrose stearate, the phospholipid and polyol are mixed to obtain phase C;

[0054] (5) Preparation of mixture 1: Grind the inclusion compound obtained in step (3) and mix it with the C phase obtained in step (4), heat and stir to obtain mixture 1;

[0055] (6) Preparation of mixture 2: The phytol is added to the mixture 1 obtained in step (5), heated and stirred to obtain mixture 2;

[0056] (7) Preparation of phase E: Mix oleyl alcohol polyether-20 with polyol, heat and stir until completely dissolved to obtain phase E;

[0057] (8) Preparation of crude emulsion: The E phase obtained in step (7) is added to the mixture 2 and homogenized and emulsified to obtain crude emulsion;

[0058] (9) Preparation of nanoemulsion: The crude emulsion obtained in step (8) is homogenized under high pressure to obtain nanoemulsion;

[0059] (10) Preparation of the composition: The target peptide is mixed with water to obtain phase F, and then the nanoemulsion obtained in step (9) is mixed with phase F and stirred to obtain the nanocomposition.

[0060] In some embodiments, in step (1), the weight ratio of the active ingredient to ethanol is 1:10 to 11:10. In some embodiments, in step (1), the weight ratio of the active ingredient to ethanol is 1:10, 1.1:10, 1.5:10, 2:10, 2.3:10, 2.5:10, 3:10, 4:10, 5:10, 6:10, 7:10, 8:10, 9:10, 10:10, 11:10, or any value between any two of these values.

[0061] In some embodiments, the heating temperature in step (1) is 60°C to 80°C. In some embodiments, the heating temperature in step (1) is 60°C, 65°C, 70°C, 75°C, 80°C, or any value between any two of these values.

[0062] In some embodiments, the heating time in step (1) is 0.5h to 1h. In some embodiments, the heating time in step (1) is 0.5h, 0.6h, 0.7h, 0.8h, 0.9h, 1h, or any value between any two of these values.

[0063] In some embodiments, in step (2), the weight ratio of water to encapsulating material is 1:1 to 5:1. In some embodiments, in step (2), the weight ratio of water to encapsulating material is 1:1, 2:1, 3:1, 4:1, 5:1, or any value between any two of these values.

[0064] In some embodiments, the heating temperature in step (2) is 55°C to 65°C. In some embodiments, the heating temperature in step (2) is 55°C, 60°C, 65°C, or any value between any two of these values.

[0065] In some embodiments, the heating time in step (2) is 10 min to 30 min. In some embodiments, the heating time in step (2) is 10 min, 15 min, 20 min, 25 min, 30 min, or any value between any two of these values.

[0066] In some embodiments, the drying in step (3) includes vacuum drying.

[0067] In some embodiments, the drying temperature in step (3) is 55°C to 65°C. In some embodiments, the drying temperature in step (3) is 55°C, 60°C, 65°C, or any value between any two of these values.

[0068] In some embodiments, the drying time of step (3) is 18h to 24h. In some embodiments, the drying time of step (3) is 18h, 19h, 20h, 21h, 22h, 23h, 24h, or any value between any two of these values.

[0069] In some embodiments, the polyol in step (4) includes at least one of dipropylene glycol and methyl propylene glycol.

[0070] In some embodiments, the polyol in step (4) is the first polyol in the first aspect.

[0071] In some embodiments, the heating temperature in step (5) is 60°C to 80°C. In some embodiments, the heating temperature in step (5) is 60°C, 65°C, 70°C, 75°C, 80°C, or any value between any two of these values.

[0072] In some embodiments, the heating time in step (5) is 0.5h to 1h. In some embodiments, the heating time in step (5) is 0.5h, 0.6h, 0.7h, 0.8h, 0.9h, 1h, or any value between any two of these values.

[0073] In some embodiments, the heating temperature in step (6) is 50°C to 60°C. In some embodiments, the heating temperature in step (6) is 50°C, 55°C, 60°C, or any value between any two of these values.

[0074] In some embodiments, the heating time in step (6) is 10 min to 30 min. In some embodiments, the heating time in step (6) is 10 min, 15 min, 20 min, 25 min, 30 min, or any value between any two of these values.

[0075] In some embodiments, the polyol in step (7) includes 1,2-pentanediol.

[0076] In some embodiments, the polyol in step (7) is the second polyol in the first aspect.

[0077] In some embodiments, the rotation speed of homogenization emulsification in step (8) is 8000 rpm to 10000 rpm (e.g., 8000 rpm, 8500 rpm, 9000 rpm, 10000 rpm, or any value between any two of these values), and the time is 3 min to 5 min (e.g., 3 min, 3.5 min, 4 min, 4.5 min, 5 min, or any value between any two of these values).

[0078] In some embodiments, the pressure of the high-pressure homogenization in step (9) is 600 bar to 800 bar (such as 600 bar, 650 bar, 700 bar, 750 bar, 800 bar, or any value between any two of these values), and the number of cycles is 3 to 4.

[0079] Thirdly, the present invention provides a formulation product.

[0080] A pharmaceutical product comprising the nanocomposite described in the first aspect or the nanocomposite prepared by the preparation method described in the second aspect;

[0081] In some embodiments, the formulation product includes medical devices or cosmetics.

[0082] In some embodiments, the medical device includes medical dressings.

[0083] In some embodiments, the cosmetic is a functional cosmetic.

[0084] Fourthly, the present invention provides an application of the nanocomposite described in the first aspect or the nanocomposite prepared by the preparation method described in the second aspect.

[0085] The application of the nanocomposite described in the first aspect or the nanocomposite prepared by the method described in the second aspect in the preparation of formulations for anti-aging and / or activating cellular energy metabolism. The core of activating cellular energy metabolism lies in enhancing mitochondrial function and promoting its efficient synthesis of adenosine triphosphate (ATP). By increasing the level of this core energy source, cells are directly infused with vitality, thereby reversing their aging state and effectively promoting the synthesis of youthful substances such as collagen, thus achieving the purpose of anti-aging.

[0086] In some embodiments, the formulation product includes medical devices or cosmetics.

[0087] In some embodiments, the medical device includes medical dressings.

[0088] In some embodiments, the cosmetic is a functional cosmetic.

[0089] Beneficial effects

[0090] Compared with the prior art, the technical solution provided by the present invention has at least one of the following beneficial technical effects:

[0091] (1) The nanocomposition provided by the present invention has excellent physical and chemical stability, good stability of active ingredient silymarin content, good particle size stability, and excellent targeting effect on fibroblasts and anti-aging effect. It can specifically target the core cells of skin anti-aging - fibroblasts, while avoiding non-specific uptake by epidermal keratinocytes, providing a core basis for achieving precise anti-aging; it can also effectively reverse cell energy decline and activate cell function from the root, with excellent unexpected technical effects.

[0092] (2) The nanocomposition provided by the present invention showed no abnormalities in appearance after being placed under normal temperature, high temperature and freezing conditions for three months, and the content retention rate of the active ingredient silymarin was significantly higher than that of the comparative example, proving that the nanocomposition system has the best physical stability and chemical stability.

[0093] (3) The hydroxypropyl-β-cyclodextrin provided by this invention is the key to maintaining the stability of the system. Replacing it with other cyclodextrins or removing it will cause the composition to become cloudy, separate into layers and have a serious decrease in content during storage, resulting in a significant deterioration in stability. Therefore, compared with not adding hydroxypropyl-β-cyclodextrin or replacing it with other cyclodextrins, the use of hydroxypropyl-β-cyclodextrin as the encapsulation material in this invention is more conducive to improving the appearance and stability of the active ingredient content of the obtained nanocomposition, and has unexpected technical effects.

[0094] (4) Compared with not adding lecithin, the addition of lecithin to the composition in this invention is more conducive to improving the appearance and stability of the active ingredient content of the obtained nanocomposition, and has unexpected technical effects.

[0095] (5) Compared with other feed ratios of lecithin, the feed ratio of lecithin of the present invention (calculated based on the total weight of the nanocomposition (excluding ethanol in phase A and water in phase B), with a lecithin content of 0.1 wt% to 5 wt%) is more conducive to improving the appearance and stability of the active ingredient content of the obtained nanocomposition, and has unexpected technical effects.

[0096] (6) Compared with other phospholipids, the lecithin provided by the present invention is more conducive to improving the appearance and stability of the active ingredient content of the obtained nanocomposite, and has unexpected technical effects.

[0097] (7) Compared with other surfactants, using sucrose stearate and oleyl alcohol polyether-20 provided by the present invention as surfactants is more conducive to improving the appearance and stability of the active ingredient content of the obtained nanocomposite, and has unexpected technical effects.

[0098] (8) The specific combination and ratio of hydroxypropyl-β-cyclodextrin, lecithin, sucrose stearate and oleyl alcohol polyether-20 provided by the present invention are crucial for preventing the degradation of active ingredients and maintaining the uniformity and stability of the system. Hydroxypropyl-β-cyclodextrin, lecithin, sucrose stearate and oleyl alcohol polyether-20 work synergistically. The absence or excess of any one of the components or its replacement with other similar components will lead to a sharp decrease in stability.

[0099] (9) The nanocomposition provided by this invention has excellent stability. The preparation method encapsulates milk thistle and silanetriol in a cyclodextrin-phospholipid system, and uses sucrose stearate as a surfactant in the composition. This surfactant has a synergistic effect with the cyclodextrin system, increasing the stability of lecithin. In addition, after the addition of phytol, it is embedded in the interstitial spaces of lecithin, forming terpenoids with lecithin, which enhances the overall stability of the raw materials, prevents degradation of the raw materials, and ensures that the active ingredient silymarin is not deactivated.

[0100] (10) The use of hydroxypropyl-β-cyclodextrin provided in this invention as an encapsulating material is crucial for the formation of stable and uniform nanoparticles; replacing it with other cyclodextrins or removing it will result in a significant increase in particle size, a wider distribution, and a sharp decrease in system stability. It can be seen that the use of hydroxypropyl-β-cyclodextrin as an encapsulating material in this invention has an unexpected technical effect on particle size stability.

[0101] (11) The amount of lecithin added needs to be within the range specified in this invention (0.1 wt%~5 wt%). Removing or adding too much of it is not conducive to forming a uniform and stable nanosystem. It can be seen that the amount of lecithin added (0.1 wt%~5 wt%) provided by this invention has an unexpected technical effect on the stability of particle size.

[0102] (12) The two types of surfactants (sucrose stearate and oleyl alcohol polyether-20) selected in this invention are indispensable for maintaining the stability and uniformity of nanoparticle size. Replacing them with other types of surfactants or removing them will have an adverse effect on the formation of nanoparticles. The two types of surfactants (sucrose stearate and oleyl alcohol polyether-20) selected in this invention have unexpected technical effects on maintaining the stability and uniformity of nanoparticle size.

[0103] (13) The acetyl tetrapeptide-3 provided by the present invention is a key modifying unit for achieving specific targeting. When it is replaced with water (i.e. without the targeting peptide), the composition almost completely loses its targeting ability to HSF cells, proving that this specific peptide is indispensable in guiding nanoparticles to recognize fibroblasts and has unexpected technical effects.

[0104] (14) When acetyl tetrapeptide-3 is replaced with other peptides (such as acetyl hexapeptide-8 or palmitoyl tripeptide-8), its HSF cell targeting efficiency is much lower than that of the present invention, indicating that acetyl tetrapeptide-3 has unexpected advantages in achieving efficient and specific targeting, and cannot be equivalently replaced by other active peptides commonly used in the field, thus having unexpected technical effects.

[0105] (15) The complete cyclodextrin-phospholipid-specific surfactant system provided by this invention is the basis for ensuring effective targeted delivery. When this system is destroyed (e.g., by changing the type of cyclodextrin or surfactant), even if acetyl tetrapeptide-3 is retained, its targeting efficiency will decrease significantly, indicating that there is a synergistic effect between the complete nanocarrier system and the targeting peptide, which together determine the final targeting effect and has unexpected technical effects.

[0106] (16) Acetyl tetrapeptide-3 is a key component for achieving efficient energy activation. Removal of it significantly reduced the ATP-boosting effect of the composition (62.13%), indicating that this specific targeting peptide is crucial for effectively delivering the active ingredient to the site of action, thereby activating cellular energy metabolism, and has unexpected technical effects.

[0107] (17) Compared with other targeting peptides (acetyl hexapeptide-8 or palmitoyl tripeptide-8), the acetyl tetrapeptide-3 provided by this invention has a significantly better effect on ATP enhancement and is superior in synergistically enhancing cellular energy, which cannot be easily replaced by conventional anti-aging peptides in the field. Therefore, the use of the acetyl tetrapeptide-3 provided by this invention has unexpected technical effects.

[0108] (18) The complete nanocarrier system (specific cyclodextrin and surfactant) provided by this invention is the basis for ensuring the energy activation effect. After the system is destroyed, even if the target peptide is retained, its ATP enhancement effect is significantly weakened, indicating that the integrity of the nanocarrier and the precise modification of the target peptide together constitute a synergistic system to achieve significant biological effects, with unexpected technical effects.

[0109] Terminology Explanation

[0110] In the foregoing description of this invention, all figures disclosed herein, whether or not the words “approximately” or “about” are used, are approximate values. Based on the disclosed figures, the value of each figure may vary by less than ±10% or by a difference that is considered reasonable by those skilled in the art, such as ±1%, ±2%, ±3%, ±4%, or ±5%.

[0111] The term “room temperature” refers to ambient temperature, which is between approximately 15°C and approximately 35°C, or approximately 20°C and approximately 30°C, or approximately 25°C.

[0112] The terms “optional,” “optional,” or “optionally” refer to events or situations that may, but are not necessarily, occur as described below.

[0113] The term “and / or” should be understood to mean any one of the options or any combination of two or more of the options.

[0114] The term "wt%" indicates a percentage by mass.

[0115] In the description of this specification, the references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the present invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Moreover, without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification, as well as the features of different embodiments or examples. Detailed Implementation

[0116] To enable those skilled in the art to better understand the technical solutions of the present invention, some non-limiting embodiments are further disclosed below to provide a more detailed description of the present invention.

[0117] All reagents used in this invention can be purchased commercially or prepared by the methods described in this invention.

[0118] Silanetriol: Methylsilanetriol (CH6O3Si), CAS number 2445-53-6.

[0119] Acetyl tetrapeptide-3: Its CAS number is 827306-88-7.

[0120] Acetyl hexapeptide-8: Its CAS number is 616204-22-9.

[0121] Palmitoyl tripeptide-8: Its CAS number is 936544-53-5.

[0122] Dipropylene glycol: its CAS number is 110-98-5.

[0123] Methylpropanediol: Its CAS number is 2163-42-0.

[0124] Silymarin: Composed of silybin (CAS No. 22888-70-6): silybin (CAS No. 33889-69-9): silybinin (CAS No. 29782-68-1) = 4.3:1:0.2 (weight ratio).

[0125] Example 1: Nanocomposition

[0126] Formula: See Table 1.

[0127] Table 1: Formulation of Nanocompositions

[0128]

[0129] Preparation method:

[0130] (1) Preparation of phase A: The active ingredient is mixed with ethanol and heated to 70℃±10℃ and stirred for 0.5h (0.5h~1h) to obtain phase A;

[0131] (2) Preparation of phase B: Dissolve the encapsulating material in water, heat to 60℃±5℃ and stir for 10 min (10 min ~ 30 min) to obtain phase B;

[0132] (3) Preparation of inclusion complex: Mix phase A and phase B under stirring, stir evenly, and vacuum dry at 60℃±5℃ for 18h (18h~24h) to obtain inclusion complex;

[0133] (4) Preparation of phase C: Sucrose stearate, the phospholipid and the first polyol are mixed to obtain phase C;

[0134] (5) Preparation of mixture 1: Grind the inclusion compound obtained in step (3) and mix it with the C phase obtained in step (4), heat it to 70℃±10℃ and stir for 0.5h (0.5h~1h) to obtain mixture 1;

[0135] (6) Preparation of mixture 2: Add the phytol to the mixture 1 obtained in step (5), heat to 55℃±5℃ and stir for 10 min (10 min ~ 30 min) to obtain mixture 2;

[0136] (7) Preparation of phase E: Mix oleyl alcohol polyether-20 with a second polyol, heat to 55℃±5℃ and stir until completely dissolved to obtain phase E;

[0137] (8) Preparation of crude emulsion: Add the E phase obtained in step (7) to the mixture 2 and homogenize and emulsify at high speed of 8000 rpm (or 8000 rpm-10000 rpm) for 5 min (or 3 min-5 min) to obtain crude emulsion;

[0138] (9) Preparation of nanoemulsion: The crude emulsion obtained in step (8) is subjected to high pressure homogenization at 600 bar (600 bar-800 bar) for 3 cycles to obtain nanoemulsion;

[0139] (10) Preparation of the composition: Mix the target peptide with water to obtain phase F, then mix the nanoemulsion obtained in step (9) with phase F and stir for 10 min (10 min ~ 30 min) to obtain the nanocomposition.

[0140] Comparative Examples 1-3: Examination of Encapsulation Materials

[0141] Comparative Example 1: The only difference from Example 1 is that hydroxypropyl-β-cyclodextrin is replaced with sulfobutyl-γ-cyclodextrin, and all other conditions are the same as in Example 1.

[0142] Comparative Example 2: The only difference from Example 1 is that hydroxypropyl-β-cyclodextrin is replaced with methyl-β-cyclodextrin, and all other conditions are the same as in Example 1.

[0143] Comparative Example 3: The only difference from Example 1 is the removal of hydroxypropyl-β-cyclodextrin and the adaptive adjustment of the water in phase F so that the total weight parts of the composition (excluding ethanol in phase A and water in phase B) are 100. Other conditions are the same as in Example 1.

[0144] Comparative Examples 4-7: Phospholipid Investigation

[0145] Comparative Example 4: The only difference from Example 1 is that lecithin was removed and the water in phase F was adjusted to make the total weight parts of the composition (excluding ethanol in phase A and water in phase B) 100. Other conditions were the same as in Example 1.

[0146] Comparative Example 5: The only difference from Example 1 is that the weight of lecithin was modified to 10, and the water in phase F was adjusted to make the total weight of the composition (excluding ethanol in phase A and water in phase B) 100. Other conditions were the same as in Example 1.

[0147] Comparative Example 6: The only difference from Example 1 is that lecithin is replaced with hydrogenated lecithin, and all other conditions are the same as in Example 1.

[0148] Comparative Example 7: The only difference from Example 1 is that lecithin is replaced with soybean lecithin, and all other conditions are the same as in Example 1.

[0149] Comparative Examples 8-9: Surfactant Investigation

[0150] Comparative Example 8: The only difference from Example 1 is that sucrose stearate is replaced with glyceryl stearate, and all other conditions are the same as in Example 1.

[0151] Comparative Example 9: The only difference from Example 1 is the removal of sucrose stearate and the adaptive adjustment of water in phase F so that the total weight parts of the composition (excluding ethanol in phase A and water in phase B) are 100. Other conditions are the same as in Example 1.

[0152] Comparative Examples 10-12: Surfactant Investigation

[0153] Comparative Example 10: The only difference from Example 1 is that oleyl alcohol polyether-20 is replaced with PEG-60 hydrogenated castor oil, and the other conditions are the same as in Example 1.

[0154] Comparative Example 11: The only difference from Example 1 is that oleyl alcohol polyether-20 is replaced with polyglycerol-10 oleate, and the other conditions are the same as in Example 1.

[0155] Comparative Example 12: The only difference from Example 1 is the removal of oleyl alcohol polyether-20 and the adaptive adjustment of the water in phase F so that the total weight parts of the composition (excluding ethanol in phase A and water in phase B) are 100. Other conditions are the same as in Example 1.

[0156] Comparative Examples 13-15: Investigation of Targeting Peptides

[0157] Comparative Example 13: The only difference from Example 1 is that acetyl tetrapeptide-3 is replaced with water, i.e., it does not contain the target peptide. All other conditions are the same as in Example 1.

[0158] Comparative Example 14: The only difference from Example 1 is that acetyl tetrapeptide-3 is replaced with acetyl hexapeptide-8, and all other conditions are the same as in Example 1.

[0159] Comparative Example 15: The only difference from Example 1 is that acetyl tetrapeptide-3 is replaced with palmitoyl tripeptide-8, and all other conditions are the same as in Example 1.

[0160] Examples 2-11: Preparation of Nanocomposites

[0161] Formulas: See Tables 2 to 4.

[0162] Table 2: Formulation of Nanocompositions

[0163]

[0164] Table 3: Formulation of Nanocompositions

[0165]

[0166] Table 4: Formulation of Nanocompositions

[0167]

[0168] Preparation method: Refer to the preparation method in Example 1.

[0169] Experimental Example 1: Stability Study

[0170] The cosmetic compositions obtained in each example and comparative example were placed under normal temperature (25±0.5℃), high temperature (45±0.5℃), and freezing (-15±0.5℃) conditions for three months. After the placement, the silymarin content was tested and the appearance and crystallinity (whether precipitation occurred) were checked. The results are shown in Table 5.

[0171] The content of milk thistle was determined using an LC-20AT high-performance liquid chromatography system equipped with a diode array detector. The chromatographic column was a ShimNex CS C18, 4.6 mm x 250 mm, 5.0 µm; the column temperature was 30 °C; the mobile phase was A% (0.1% formic acid aqueous solution) and B% (methanol); the flow rate was 1.0 mL / min; the injection volume was 10 μL; the peak area at 288 nm was detected; and the content of milk thistle was calculated using the external standard method.

[0172] Table 5: Results of Silymarin Content and Stability After Three Months

[0173]

[0174] Note: No abnormality indicates that compared with day 0, the color remains unchanged and is uniform, transparent, without crystallization, turbidity, or stratification.

[0175] The content is expressed as a percentage of the measured amount to the theoretical amount of material to be fed.

[0176] in conclusion:

[0177] (1) As can be seen from the results of Examples 1-11 in Table 5, the nanocomposition provided by the present invention showed no abnormalities in appearance after being placed under normal temperature, high temperature and freezing conditions for three months, and the content retention rate of the active ingredient silymarin was significantly higher than that of the comparative example, proving that the nanocomposition system has the best physical and chemical stability.

[0178] (2) As can be seen from the results of Example 1 and Comparative Examples 1-3 in Table 5, hydroxypropyl-β-cyclodextrin is the key to maintaining the stability of the system. Replacing it with other cyclodextrins or removing it will cause the composition to become turbid, separate, and have a serious decrease in content during storage, resulting in a significant deterioration in stability. Therefore, compared with not adding hydroxypropyl-β-cyclodextrin or replacing hydroxypropyl-β-cyclodextrin with other cyclodextrins, the present invention uses hydroxypropyl-β-cyclodextrin as an encapsulating material, which is more conducive to improving the appearance and stability of the active ingredient content of the obtained nano-composition, and has unexpected technical effects.

[0179] (3) As can be seen from the results of Example 1 and Comparative Example 4 in Table 5, compared with not adding lecithin, the addition of lecithin to the composition in this invention is more conducive to improving the appearance and stability of the active ingredient content of the obtained nano-composition, and has unexpected technical effects.

[0180] (4) As can be seen from the results of Example 1 and Comparative Example 5 in Table 5, compared with other feed ratios of lecithin, the feed ratio of lecithin of the present invention (calculated based on the total weight of the nanocomposition (excluding ethanol in phase A and water in phase B), with a lecithin content of 0.1 wt% to 5 wt%) is more conducive to improving the appearance and stability of the active ingredient content of the obtained nanocomposition, and has unexpected technical effects.

[0181] (5) As can be seen from the results of Example 1, Comparative Example 6 and Comparative Example 7 in Table 5, compared with other phospholipids, the lecithin provided by the present invention is more conducive to improving the appearance and stability of the active ingredient content of the obtained nanocomposite, and has unexpected technical effects.

[0182] (6) As can be seen from the results of Example 1 and Comparative Examples 8 to 12 in Table 5, compared with other surfactants, using sucrose stearate and oleyl alcohol polyether-20 provided by the present invention as surfactants is more conducive to improving the appearance and stability of the active ingredient content of the obtained nanocomposite, and has unexpected technical effects.

[0183] (7) As can be seen from the results of Example 1 and Comparative Examples 1 to 12 in Table 5, the specific combination and ratio of hydroxypropyl-β-cyclodextrin, lecithin, sucrose stearate and oleyl alcohol polyether-20 are crucial for preventing the degradation of active ingredients and maintaining the uniformity and stability of the system. Hydroxypropyl-β-cyclodextrin, lecithin, sucrose stearate and oleyl alcohol polyether-20 work synergistically. The absence or excess of any one of the components or its replacement with other similar components will lead to a sharp decrease in stability.

[0184] (8) Based on the comparative data, the nanocomposition provided by this invention exhibits excellent stability. Furthermore, the preparation method encapsulates milk thistle and silanetriol within a cyclodextrin-phospholipid system, using sucrose stearate as a surfactant in the composition. This surfactant exhibits a synergistic effect with the cyclodextrin system, increasing the stability of lecithin. In addition, the addition of phytol allows it to intercalate within the lecithin layers, forming terpenoids that enhance the overall stability of the raw materials, prevent degradation, and ensure the inactivation of the active ingredient, silymarin.

[0185] Experimental Example 2: Investigation of Nanoparticle Size

[0186] The particle size of the anti-aging nanocomposites prepared in Examples 1-11 and Comparative Examples 1-15 was measured after 0 days and after being placed under accelerated conditions (45±0.5℃) for three months. The particle size and polydispersity index were determined by a nanoparticle size potentiometer to characterize the anti-aging nanocomposites. The results are shown in Table 6.

[0187] Table 6: Characterization of Particle Size and Polydispersity Index of Anti-aging Nanocompositions

[0188]

[0189] in conclusion:

[0190] Based on the nanoparticle size measurement results of Experiment Example 2, the following conclusions can be drawn:

[0191] (1) As can be seen from the results of Examples 1-11 in Table 6, the anti-aging nanocomposition provided by the present invention has excellent and uniform nanoparticle size (14nm-30nm) and extremely low polydispersity coefficient (0.043-0.083), indicating that the composition system is stable, the particle distribution is uniform, and it has excellent particle size stability.

[0192] (2) As can be seen from the results of Example 1 and Comparative Examples 1-3 in Table 6, the use of hydroxypropyl-β-cyclodextrin provided by the present invention as the encapsulating material is crucial for the formation of stable and uniform nanoparticles; replacing it with other cyclodextrins or removing it will result in a significant increase in particle size, a wider distribution, and a sharp decrease in system stability. Therefore, it can be seen that the use of hydroxypropyl-β-cyclodextrin as the encapsulating material in the present invention has an unexpected technical effect on particle size stability.

[0193] (3) As can be seen from the results of Example 1 and Comparative Examples 4-5 in Table 6, the amount of lecithin added needs to be within the range specified in this invention (0.1 wt%~5 wt%). Removing or adding too much lecithin is not conducive to forming a uniform and stable nanosystem. Therefore, it can be seen that the amount of lecithin added (0.1 wt%~5 wt%) provided by this invention has an unexpected technical effect on particle size stability.

[0194] (4) As can be seen from the results of Example 1 and Comparative Examples 6-10 in Table 6, the two types of surfactants (sucrose stearate and oleyl alcohol polyether-20) selected in this invention are indispensable for maintaining the stability and uniformity of nanoparticle size. Replacing them with other types of surfactants or removing them will have an adverse effect on the formation of nanoparticles. The two types of surfactants (sucrose stearate and oleyl alcohol polyether-20) selected in this invention have unexpected technical effects on maintaining the stability and uniformity of nanoparticle size.

[0195] The anti-aging nanocomposites prepared by Comparative Examples 1, 3, 4, 5, 8, 9, 10, 11, and 12 were unstable and could not be stored for a long time due to stability testing and particle size characterization, and therefore were not further studied.

[0196] Experimental Example 3: Targeted Testing

[0197] Human fibroblasts (HSFs) are crucial cells responsible for synthesizing collagen (such as type I and type III collagen, which maintain skin elasticity) and elastin. The core of aging is the decline in fibroblast function, leading to collagen loss and skin sagging. Supplementing with active ingredients and precisely targeting them is essential to promoting collagen regeneration at its source and achieving effective anti-aging.

[0198] This composition modifies the surface of nanoparticles with specific peptides that can specifically recognize fibroblasts. The experimental examples, through Examples 1-11, Comparative Examples 6, 10, 13, 14, and 15, demonstrate the targeting selectivity of fibroblasts and keratinocytes (HaCaT), further providing core evidence for its precise application in the field of skin anti-aging.

[0199] Test method:

[0200] (1) In this test, silybin, silybinin and silybinin were labeled with CY2, CY5 and AF405 fluorescent groups respectively to detect the targeting effect. After being labeled with CY2, CY5 and AF405 fluorescent groups, silybin, silybinin and silybinin emitted green, red and blue fluorescence respectively. The greater the intensity of fluorescence emitted in the cells, the higher the content of the active composition absorbed by the cells. By comparing the fluorescence intensity, the effect of this nanocomposition on HSF cells was verified.

[0201] Test steps:

[0202] ① Cell seeding: Select HSF cells (Fenghui Biotechnology) / HaCaT (Pronosai) in logarithmic growth phase, collect cells after trypsin digestion, prepare cell suspension, count cells with a cell counter, prepare a cell suspension of 1.2×105 cells / mL with complete culture medium (manufacturer: Thermo Fisher Scientific; brand: Gibco; catalog number 11995), seed into 12-well plates, add 1 mL to each well, and incubate at 37℃ in a 5% CO2 incubator for 24±2 h.

[0203] ② Cell sample loading: Discard the culture medium in the cell culture plate and load the sample (the sample group is loaded with the final concentration of the composition to the detection concentration of 0.25wt%; no composition is added to the negative control group, and the other operations are the same as the sample group). After the sample loading is completed, place the cell culture plate in a CO2 incubator and continue to culture for 24±2 h.

[0204] ③ Photography and Result Analysis: After incubation, photographs were taken under a fluorescence microscope at the same magnification. The fluorescence intensity of different treatment groups was observed to determine the cell uptake efficiency of the sample. The average fluorescence intensity (Mean) = the sum of fluorescence intensity in the region (IntDen) / the area of ​​the region (Area). The results are shown in Table 7.

[0205] Table 7: Comparison of Relative Fluorescence Intensities

[0206]

[0207] in conclusion:

[0208] Based on the target testing results of Experiment 3, the following conclusions can be drawn:

[0209] (1) As can be seen from the results of Examples 1, 2, 3, 7, 10 and 11 in Table 7, the nanocomposition provided by the present invention exhibits high fluorescence intensity on human fibroblasts (HSF) and low fluorescence intensity on human keratinocytes (HaCaT), proving that the composition can specifically target the core cells of skin anti-aging - fibroblasts, and avoid non-specific uptake by epidermal keratinocytes, providing a core basis for achieving precise anti-aging and having unexpected technical effects.

[0210] (2) As can be seen from the results of Example 1 and Comparative Example 13 in Table 7, acetyl tetrapeptide-3 is the key modification unit for achieving specific targeting. After replacing it with water (i.e. without the targeting peptide), the composition almost completely lost its targeting ability to HSF cells, proving that this specific peptide is indispensable in guiding nanoparticles to recognize fibroblasts and has unexpected technical effects.

[0211] (3) As can be seen from the results of Example 1 and Comparative Examples 14 and 15 in Table 7, when acetyl tetrapeptide-3 is replaced with other peptides (such as acetyl hexapeptide-8 or palmitoyl tripeptide-8), its HSF cell targeting efficiency is much lower than that of the present invention. This indicates that acetyl tetrapeptide-3 has unexpected advantages in achieving efficient and specific targeting and cannot be equivalently replaced by other active peptides commonly used in the field, thus having unexpected technical effects.

[0212] (4) As can be seen from the results of Example 1 and Comparative Example 2 in Table 7, the cyclodextrin-phospholipid-specific surfactant system of the composition of the present invention is the basis for ensuring effective targeted delivery. After the system is destroyed, even if acetyl tetrapeptide-3 is retained, its targeting efficiency will decrease significantly, indicating that there is a synergistic effect between the complete nanocarrier system and the targeting peptide, which jointly determines the final targeting effect and has unexpected technical effects.

[0213] Therefore, by modifying acetyl tetrapeptide-3 onto a stable nanocarrier composed of specific excipients, this invention successfully constructed a delivery system that can efficiently and specifically target fibroblasts, laying a solid foundation for the precise anti-aging application of the composition.

[0214] Experimental Example 4: ATP Enhancement Experiment

[0215] Adenosine triphosphate (ATP) is the direct energy source for all intracellular biochemical activities, such as biosynthesis, muscle contraction, and signal transduction. During aging, the decline in mitochondrial function leads to a significant reduction in ATP synthesis efficiency. This weakened energy directly results in insufficient fibroblast production, hindering the efficient synthesis of new collagen and elastin, and impairing effective self-repair. This is one of the core underlying reasons for skin sagging and wrinkles.

[0216] This experiment tested the activation effect of Examples 1-11, Comparative Examples 2, 13, 14, and 15 on the energy metabolism level of human skin fibroblasts, and tested its ability to increase the concentration of intracellular ATP, thereby confirming its ability to reverse cell aging and activate cell function from the perspective of cell energy activation.

[0217] Test method:

[0218] (1) Cell seeding: Select HSF cells (Fenghui Biotechnology) in the logarithmic growth phase. When the plate-laying rate reaches about 60%, seed the cells into 24-well plates and incubate overnight in a CO2 incubator (37℃, 5% CO2).

[0219] (2) Solution preparation: Prepare working solutions of test substances according to test groups.

[0220] (3) Drug administration: According to the test groups, when the cell deposition rate in the 24-well plate reached 30%~50%, the drugs were administered to the groups (concentration of 0.004wt%), with 3 replicates per group. 1 mL of culture medium was added to each well of the blank control group and the negative control group (excluding the cell energy activation composition with anti-aging effects), 1 mL of culture medium containing vitamin E (VE, Sigma) (concentration of 0.05%) was added to each well of the positive control group, and 1 mL of culture medium containing the corresponding test sample was added to each well of the sample group. After drug administration, the 24-well plate was placed in a CO2 incubator (37℃, 5% CO2) and incubated for 24 h.

[0221] (4) UVA irradiation: Except for the blank control group, all other groups were subjected to UVA irradiation at a dose of 30 J / cm². 2 After irradiation, the sample was placed in a CO2 incubator (37℃, 5% CO2) for 24 hours.

[0222] (5) ATP content detection: After incubation, the ATP was washed with PBS phosphate buffer (manufacturer: Solarbio) and detected by fluorescence detection according to the instructions of the ATP detection kit (manufacturer: Baiying Biotechnology).

[0223] (6) Calculation of ATP enhancement rate: ATP enhancement rate (%) = (sample group - negative control group) / negative control group × 100%.

[0224] (7) Results: See Table 8.

[0225] Table 8: ATP Enhancement Rate of Anti-aging Nanocompositions

[0226]

[0227] in conclusion:

[0228] Based on the ATP enhancement rate test results of Experiment Example 4, the following conclusions can be drawn:

[0229] (1) As can be seen from the results of Examples 1-11 in Table 8, the anti-aging nanocomposition provided by the present invention can significantly increase the ATP level in HSF (ATP increase rate 115.89%-142.86%), indicating that the composition can effectively reverse cell energy decline and activate cell function from the root, with unexpected technical effects.

[0230] (2) As can be seen from the results of Example 1 and Comparative Example 13 in Table 8, acetyl tetrapeptide-3 is a key component for achieving efficient energy activation. After its removal, the ATP enhancement effect of the composition was significantly reduced (ATP enhancement rate was 62.13%), indicating that this specific targeting peptide is crucial for effectively delivering the active ingredient to the site of action, thereby activating cellular energy metabolism, and has unexpected technical effects.

[0231] (3) As can be seen from the results of Example 1 and Comparative Examples 14 and 15 in Table 8, when Acetyl tetrapeptide-3 is replaced with other peptides (Acetyl hexapeptide-8 or palmitoyl tripeptide-8), the ATP enhancement effect (ATP enhancement rate of 89.23% and 78.54% respectively) is significantly lower than that of the present invention, which proves that Acetyl tetrapeptide-3 has superiority in synergistic enhancement of cell energy and cannot be simply replaced by conventional anti-aging peptides in the field, and has unexpected technical effects.

[0232] (4) As can be seen from the results of Example 1 and Comparative Example 2 in Table 8, the complete nanocarrier system (specific cyclodextrin and surfactant) is the basis for ensuring the energy activation effect. After the system is destroyed, even if the target peptide is retained, its ATP enhancement effect is significantly weakened, indicating that the integrity of the nanocarrier and the precise modification of the target peptide together constitute a synergistic system to achieve significant biological effects, with unexpected technical effects.

[0233] Experiment 5: Human Testing of Anti-wrinkle Effect

[0234] The wrinkle area of ​​the subjects was measured before using the cosmetic product using the VISIA 7 facial image analyzer (Canfield, USA), and recorded as the initial value. The wrinkle area was measured again after 30 minutes, 14 days, and 28 days of continuous use, and recorded as the termination value. Finally, the wrinkle area change rate was calculated as follows: Wrinkle area change rate (%) = (Initial value - Termination value at the test time point) / Initial value × 100%. The higher the wrinkle area change rate, the better the anti-aging effect of the cosmetic product. The results are shown in Table 9.

[0235] Table 9: Change rate of wrinkle area in experimental cases

[0236]

[0237] Conclusion: According to Table 9, the results of Examples 2 and 3 show that the supramolecular milk thistle anti-aging composition can effectively achieve cosmetic anti-aging effects at 30 min, 14 days and 28 days, and significantly reduce the proportion of wrinkle area.

[0238] The method of this invention has been described through preferred embodiments. Those skilled in the art will readily be able to modify or appropriately alter and combine the methods and applications described herein within the scope, spirit, and context of this invention to implement and apply the technology of this invention. Those skilled in the art can refer to the content herein to appropriately improve process parameters. It should be particularly noted that all similar substitutions and modifications are obvious to those skilled in the art and are considered to be included within the scope of this invention.

Claims

1. A nanocomposition, characterized in that, The nanocomposition comprises: nanoparticles, surfactants, polyols, targeting peptides, and water; The nanoparticles include: active ingredients, encapsulating materials, phospholipids, and stabilizers; The active ingredient is at least one of silymarin and silanetriol; The targeted peptide includes acetyl tetrapeptide-3; The encapsulating material encapsulates the active ingredient to form an inclusion complex, the phospholipid forms a phospholipid bilayer that encapsulates the inclusion complex, and the stabilizer is embedded in the phospholipid bilayer; The nanoparticles are dispersed in the surfactant, polyol, and water; The targeted peptide modifies the surface of the nanoparticles; The encapsulating material is hydroxypropyl-β-cyclodextrin; The phospholipid is lecithin; The stabilizer is phytol; The surfactant is oleyl alcohol polyether-20 and sucrose stearate; The weight ratio of oleyl alcohol polyether-20 to sucrose stearate is 5:1 to 20:1; Based on the total weight of the nanocomposition, the content of the active ingredient is 1.5wt%~5wt%, the content of the encapsulating material is 10wt%~20wt%, the content of the phospholipid is 0.5wt%~4wt%, the content of the stabilizer is 1.5wt%~4wt%, the content of the surfactant is 5wt%~21wt%, the content of the polyol is 5wt%~25wt%, the content of the targeting peptide is 0.001wt%~0.01wt%, and the balance is water.

2. The nanocomposition according to claim 1, wherein the polyol comprises at least one selected from dipropylene glycol, 1,2-pentanediol, 1,3-butanediol, hexyldecyl alcohol, 1,2-propanediol, 1,3-propanediol, 1,2-hexanediol, and methylpropanediol.

3. The nanocomposition according to claim 1, wherein the polyol comprises a first polyol and a second polyol, the first polyol comprising at least one of dipropylene glycol and methyl propylene glycol; the second polyol comprising 1,2-pentanediol, and the weight ratio of the first polyol to the second polyol is 2:3 to 5:

1.

4. A method for preparing the nanocomposition according to any one of claims 1 to 3, characterized in that, Includes the following steps: (1) Preparation of phase A: The active ingredient is mixed with ethanol, heated and stirred to obtain phase A; (2) Preparation of phase B: Dissolve the encapsulating material in water, heat and stir to obtain phase B; in step (2), the weight ratio of water to encapsulating material is 1:1 to 5:1; (3) Preparation of inclusion complex: Under stirring conditions, phase A and phase B are mixed, stirred evenly, and dried to obtain inclusion complex; (4) Preparation of phase C: Sucrose stearate, the phospholipid and polyol are mixed to obtain phase C; the polyol in step (4) includes at least one of dipropylene glycol and methyl propylene glycol; (5) Preparation of mixture 1: Grind the inclusion compound obtained in step (3) and mix it with the C phase obtained in step (4), heat and stir to obtain mixture 1; (6) Preparation of mixture 2: The phytol is added to the mixture 1 obtained in step (5), heated and stirred to obtain mixture 2; (7) Preparation of phase E: Mix oleyl alcohol polyether-20 with a polyol, heat and stir until completely dissolved to obtain phase E; the polyol in step (7) includes 1,2-pentanediol; (8) Preparation of crude emulsion: The E phase obtained in step (7) is added to the mixture 2 and homogenized and emulsified to obtain crude emulsion; (9) Preparation of nanoemulsion: The crude emulsion obtained in step (8) is homogenized under high pressure to obtain nanoemulsion; (10) Preparation of the composition: The target peptide is mixed with water to obtain phase F, and then the nanoemulsion obtained in step (9) is mixed with phase F and stirred to obtain the nanocomposition.

5. According to the preparation method of claim 4, in step (1), the weight ratio of the active ingredient to ethanol is 1:10 to 11:10; The heating temperature in step (1) is 60℃~80℃; The heating temperature in step (2) is 55℃~65℃; The heating temperature in step (5) is 60℃~80℃; The heating temperature in step (6) is 50℃~60℃.

6. A pharmaceutical preparation, characterized in that, The nanocompositions include those described in any one of claims 1 to 3 or those prepared by the methods described in any one of claims 4 to 5.

7. The use of a nanocomposition according to any one of claims 1 to 3 or a nanocomposition prepared by any one of claims 4 to 5 in the preparation of a pharmaceutical product for anti-aging and / or activation of cellular energy metabolism.

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