A highly efficient endogenous anti-aging supramolecular composition, its preparation method and application

Abstract

This invention belongs to the field of cosmetics, specifically relating to a highly efficient endogenous anti-aging supramolecular composition, its preparation method, and its application. This invention discovers that the synergistic effect of tetrapeptide-1 and PDRN can enhance the free radical scavenging capacity of mitochondria, while simultaneously increasing the expression level of mitochondrial mtDNA, reducing mitochondrial oxidative stress damage, and maintaining youthful mitochondrial function. Specifically, this invention obtains a composition with uniform and stable particle size and better transdermal properties through a combination of microfluidic and microfluidic processing. Adding this composition to skincare products and applying it to the skin can stabilize the mitochondrial function of skin cells, maintain normal physiological activities, and thus delay skin aging caused by endogenous factors.

Description

Technical Field

[0001] This invention belongs to the field of cosmetics, specifically relating to a highly efficient endogenous anti-aging supramolecular composition, its preparation method, and its application. Background Technology

[0002] The information disclosed in this background section is intended only to enhance understanding of the overall background of the invention and is not necessarily to be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art.

[0003] Most people begin to show signs of aging from the age of 25, such as fine lines, sagging skin, and rough, dull skin. This is because the skin's self-repair ability begins to weaken. As we age, we can no longer rely solely on our own skin repair system to maintain the skin's dynamic balance; the rate of skin cell aging far exceeds the rate of cell regeneration. Therefore, to truly combat aging, we need to pay more attention to skin aging caused by endogenous cellular aging.

[0004] Mitochondria are essential for all physiological activities of skin cells. As energy factories, mitochondria synthesize ATP for skin cells and maintain a dynamic balance in the number of free radicals, ensuring the normal functioning of our skin. Normal mitochondria ensure the efficient operation of the skin's self-repair system, maintaining youthful skin. However, with age and the influence of the external environment, mitochondria undergo aging, resulting in decreased energy supply capacity and reduced efficiency in scavenging free radicals. This weakens the flow of ATP within skin cells, causing the rate of skin cell aging to exceed the rate of cell regeneration, leading to wrinkles, fine lines, and sagging skin.

[0005] The inventors discovered during their research that traditional anti-aging product development often prioritizes using active ingredients to inhibit the growth of free radicals that cause skin cell aging, promote stratum corneum metabolism, reduce fine lines, and stimulate collagen production. These active ingredients are the anti-aging targets that traditional anti-aging products focus on. However, to achieve better anti-aging effects, it is necessary to consider whether the active ingredients have the ability to fundamentally regulate mitochondrial function, reduce mitochondrial damage, and maintain skin cell vitality. Therefore, researchers urgently need to design and develop a product that can perform endogenous anti-aging within skin cells to achieve better endogenous anti-aging effects. Summary of the Invention

[0006] To address the shortcomings of existing technologies, the present invention aims to provide a highly efficient endogenous anti-aging supramolecular composition, its preparation method, and its applications. This invention uses fish egg lipid extract and caprylic / capric triglycerides as the external phase components to encapsulate the internal phase components PDRN and tetrapeptide-1. The synergistic effect between these components not only enhances the skin's deep antioxidant capacity and mitochondrial free radical scavenging ability but also increases mitochondrial mtDNA expression, reduces mitochondrial oxidative stress damage, maintains youthful mitochondrial function, and delays the endogenous aging process of the skin.

[0007] To achieve the above objectives, the present invention is implemented through the following technical solution:

[0008] In a first aspect, the present invention provides a highly efficient endogenous anti-aging supramolecular composition comprising an external phase component and an internal phase component; the external phase component is composed of fish egg lipid extract and caprylic / capric triglyceride; the internal phase component is composed of an aqueous solution of tetrapeptide-1 and PDRN.

[0009] Preferably, the mass ratio of the external phase component to the internal phase component is 3 to 5:1.

[0010] Preferably, the mass percentage concentration of the tetrapeptide-1 and PDRN in the aqueous solution is 0.5-2%.

[0011] Preferably, the molecular weight of the PDRN is 0.5-1.5 million.

[0012] Preferably, the mass ratio of the fish roe lipid extract to caprylic / capric triglyceride is 1:5 to 10; more preferably, the mass ratio of the fish roe lipid extract to caprylic / capric triglyceride is 1:7 to 9.

[0013] More preferably, the fish egg lipid extract is fish egg from salmon, sturgeon or flounder; the specific extraction method is as follows: the fish egg is mixed with purified water, ultrasonically crushed, treated with a high-pressure homogenizer, and the bottom product is collected after centrifugation, which is the fish egg lipid extract.

[0014] More preferably, the high-pressure homogenization has a pressure of 5-60 MPa, a flow rate of 20-45 L / h, and a homogenization frequency of 1-3 times.

[0015] A second aspect of the present invention provides a method for preparing the above-mentioned highly efficient endogenous anti-aging supramolecular composition, comprising the following steps:

[0016] S1. Tetrapeptide-1 and PDRN are dissolved in water, and then the pH of the solution is adjusted to obtain the internal phase component;

[0017] S2, a mixture of fish egg lipid extract and caprylic / capric triglyceride was prepared to obtain the external phase component;

[0018] S3. Using microfluidics, the external phase component and the internal phase component are brought into contact to form a composition;

[0019] S4. The composition is subjected to high-pressure microfluidic treatment to obtain a supramolecular composition.

[0020] Preferably, in step S1, the pH value is 4.7 to 7.2.

[0021] Preferably, in step S3, the flow rate of the external phase component in the microfluidic technology is 3-8 mL / h, and the flow rate of the internal phase component is 0.5-1 mL / h.

[0022] Preferably, in step S4, the pressure is set to 15,000-20,000 psi and the number of treatments is 3 to 5.

[0023] A third aspect of the present invention provides a composition comprising the highly efficient endogenous anti-aging supramolecular composition described in the first aspect and a functional peptide.

[0024] Preferably, the functional polypeptide is selected from one or more of palmitoyl polypeptides, acetyl polypeptides, arginine / lysine polypeptides, decapeptide-4, hexapeptide-9, nonapeptide-1, and dipeptide diaminobutyryl benzylamide diacetate.

[0025] Preferably, the composition further comprises a humectant, an emulsifier, a preservative, and water; more preferably, the humectant is selected from one or more of glycerin, propylene glycol, butylene glycol, hexanediol, hyaluronic acid, betaine, and trehalose; the emulsifier is selected from one or more of glyceryl stearate, hydrogenated lecithin, polysorbate-80, polysorbate-20, sorbitan olive oil ester, polyglycerol-6 stearate, and polyglycerol-6 behenate; and the preservative is selected from one or more of methylparaben, phenoxyethanol, p-hydroxyacetophenone, sodium benzoate, and potassium sorbate.

[0026] Preferably, the amount of the highly effective endogenous anti-aging supramolecular composition added to the composition is 0.001-3% by mass; the amount of the functional peptide added to the composition is 0.0001-1% by mass; the amount of the moisturizer added to the composition is 0.1-10% by mass; the amount of the preservative added to the composition is 0.001-2% by mass; and the amount of the emulsifier added to the composition is 0.001-5% by mass.

[0027] A fourth aspect of the present invention provides the use of the highly efficient endogenous anti-aging supramolecular composition described in the first aspect or the composition described in the third aspect in the preparation of skin care products.

[0028] Preferably, the skin care product is a serum, lotion, toner, cream, or mask.

[0029] Preferably, the skin care product has the effects of anti-photoaging, anti-oxidation, whitening, wrinkle removal and anti-aging.

[0030] The beneficial effects achieved by one or more technical solutions of the present invention are as follows:

[0031] (1) The present invention provides a highly efficient endogenous anti-aging supramolecular composition that combines PDRN with tetrapeptide-1 to synergistically delay endogenous aging of cells, improve the skin’s own antioxidant capacity, improve the free radical scavenging capacity of mitochondria, and at the same time increase the expression level of mitochondrial mtDNA, reduce mitochondrial oxidative stress damage, maintain the youthful state of mitochondrial function, and delay the endogenous aging process of skin.

[0032] (2) This invention innovatively combines fluid chemistry and supramolecular chemistry, combining external phase components and internal phase components into a composition in a microfluidic channel. After being treated with microjet, the resulting supramolecular composition has good quality stability and uniform particle size.

[0033] (3) The skin care products containing highly efficient endogenous anti-aging compositions prepared by the present invention can better maintain the youthful state of skin cells, enhance the skin's self-repair ability, and alleviate the aging of skin cells from the endogenous perspective, and have a more obvious and longer-lasting anti-aging effect compared with other skin care products. Attached Figure Description

[0034] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an improper limitation of the invention.

[0035] Figure 1 The sample prepared in Example 1 of this invention;

[0036] Figure 2 This is a particle size distribution diagram of the sample prepared in Example 1 of the present invention;

[0037] Figure 3 This is a particle size variation diagram of the samples prepared in Example 1 and Comparative Examples 3-5 of the present invention;

[0038] Figure 4 This is a graph showing the changes in cell ROS content according to the present invention.

[0039] Figure 5 This is a graph showing the changes in mtDNA expression levels in cells according to the present invention.

[0040] Figure 6 This is a diagram showing the changes in mitochondrial density in cells according to the present invention. Detailed Implementation

[0041] It should be noted that the following detailed descriptions are exemplary and intended to provide further illustration of the invention. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0042] A first typical embodiment of the present invention provides a highly efficient endogenous anti-aging supramolecular composition, the supramolecular composition comprising an external phase component and an internal phase component; the external phase component is composed of fish egg lipid extract and caprylic / capric triglyceride; the internal phase component is composed of an aqueous solution of tetrapeptide-1 and PDRN.

[0043] In one or more embodiments of this implementation, the mass ratio of the external phase component to the internal phase component is 3 to 5:1.

[0044] In one or more embodiments of this implementation, the mass percentage concentration of the tetrapeptide-1 and PDRN in the aqueous solution is 0.5-2%.

[0045] In one or more embodiments of this implementation, the molecular weight of the PDRN is 0.5-1.5 million.

[0046] In one or more embodiments of this implementation, the mass ratio of the fish egg lipid extract to caprylic / capric triglyceride is 1:5 to 10.

[0047] In one or more embodiments of this implementation, the mass ratio of the fish egg lipid extract to caprylic / capric triglyceride is 1:7-9.

[0048] In one or more embodiments of this implementation, the fish egg lipid extract is the fish egg of salmon, sturgeon or flounder; the specific extraction method is as follows: the fish egg is mixed with purified water, ultrasonically crushed, treated by a high-pressure homogenizer, and the bottom product is collected after centrifugation, which is the fish egg lipid extract.

[0049] In one or more embodiments of this implementation, the pressure of the high-pressure homogenization is 5-60 MPa, the flow rate is 20-45 L / h, and the number of homogenization cycles is 1-3.

[0050] A second typical embodiment of the present invention provides a method for preparing the above-mentioned highly efficient endogenous anti-aging supramolecular composition, comprising the following steps:

[0051] S1. Tetrapeptide-1 and PDRN are dissolved in water, and then the pH of the solution is adjusted to obtain the internal phase component;

[0052] S2, a mixture of fish egg lipid extract and caprylic / capric triglyceride was prepared to obtain the external phase component;

[0053] S3. Using microfluidics, the external phase component and the internal phase component are brought into contact to form a composition;

[0054] S4. The composition is subjected to high-pressure microfluidic treatment to obtain a supramolecular composition.

[0055] In one or more embodiments of this implementation, in step S1, the pH value is 4.7 to 7.2.

[0056] In one or more embodiments of this implementation, in step S3, the flow rate of the external phase component in the microfluidic technology is 3-8 mL / h, and the flow rate of the internal phase component is 0.5-1 mL / h.

[0057] In one or more embodiments of this implementation, in step S4, the pressure is set to 15000-20000 psi and the number of treatments is 3 to 5.

[0058] A third typical embodiment of the present invention provides a composition comprising the highly efficient endogenous anti-aging supramolecular composition described in the first aspect and a functional peptide.

[0059] In one or more embodiments of this implementation, the functional polypeptide is selected from one or more of palmitoyl polypeptides, acetyl polypeptides, arginine / lysine polypeptides, decapeptide-4, hexapeptide-9, nonapeptide-1, and dipeptide diaminobutyryl benzylamide diacetate.

[0060] In one or more embodiments of this implementation, the composition further comprises a humectant, an emulsifier, a preservative, and water; more preferably, the humectant is selected from one or more of glycerin, propylene glycol, butylene glycol, hexanediol, sodium hyaluronate, betaine, and trehalose; the emulsifier is selected from one or more of glyceryl stearate, hydrogenated lecithin, polysorbate-80, polysorbate-20, sorbitan olive oil ester, polyglycerol-6 stearate, and polyglycerol-6 behenate; and the preservative is selected from one or more of methylparaben, phenoxyethanol, p-hydroxyacetophenone, sodium benzoate, and potassium sorbate.

[0061] In one or more embodiments of this implementation, the amount of the highly efficient endogenous anti-aging supramolecular composition added to the composition is 0.001-3% by mass; the amount of the functional peptide added to the composition is 0.0001-1% by mass; the amount of the moisturizer added to the composition is 0.1-10% by mass; the amount of the preservative added to the composition is 0.001-2% by mass; and the amount of the emulsifier added to the composition is 0.001-5% by mass.

[0062] The fourth typical embodiment of the present invention provides the application of the highly efficient endogenous anti-aging supramolecular composition described in the first aspect or the composition described in the third aspect in the preparation of skin care products.

[0063] In one or more embodiments of this implementation, the skin care product is a serum, lotion, toner, cream, or mask.

[0064] In one or more embodiments of this implementation, the skin care product has the effects of anti-photoaging, anti-oxidation, whitening, wrinkle removal and anti-aging.

[0065] PDRN, a component with highly effective anti-inflammatory, repairing, and anti-aging properties, is widely used in various cosmetics. Tetrapeptide-1 is a polypeptide that can stimulate fibroblast proliferation and promote the synthesis of extracellular matrix proteins such as collagen. The inventors discovered that when PDRN and tetrapeptide-1 are used in combination at a certain ratio, in addition to improving fine lines and enhancing skin elasticity, they can also significantly improve problems such as fine lines, dullness, and roughness caused by mitochondrial damage or dysfunction, thus playing a synergistic role in delaying endogenous aging. The synergistic effect of the two components can enhance the skin's own antioxidant capacity, improve the mitochondrial free radical scavenging ability, increase the expression level of mitochondrial mtDNA, reduce mitochondrial oxidative stress damage, maintain youthful mitochondrial function, and delay the endogenous aging process of the skin.

[0066] Meanwhile, to enhance the transdermal absorption of PDRN and tetrapeptide-1 and better maintain mitochondrial function, this invention uses fish egg lipid extract and caprylic / capric triglycerides as the exogenous components to encapsulate PDRN and tetrapeptide-1. Fish egg lipid extract, as a naturally derived lipid component, has higher compatibility with human skin and can better carry the endogenous components into the deeper layers of the skin.

[0067] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0068] Example 1: This example provides a highly efficient endogenous anti-aging supramolecular composition and its preparation method.

[0069] In this embodiment, 100g of a highly efficient endogenous anti-aging supramolecular composition was prepared, wherein the mass ratio of the external phase component to the internal phase component was 4:1, the mass ratio of fish egg lipid extract to caprylic / capric triglyceride was 1:7, and the mass percentage concentration of tetrapeptide-1 and PDRN in aqueous solution was 0.5%.

[0070] The specific components and their contents are as follows:

[0071] The total amount of external phase components is 80g, including 10g of fish egg lipid extract and 70g of caprylic / capric triglycerides;

[0072] The total amount of internal phase components is 20g.

[0073] The specific preparation method includes the following steps:

[0074] (1) Weigh 0.1g of tetrapeptide-1 and PDRN respectively, dissolve them in water, add water to make a total weight of 20g, adjust the pH to 6, and obtain the internal phase component;

[0075] (2) Mix 10g of fish egg lipid extract with 70g of caprylic / capric triglyceride to obtain the external phase component;

[0076] (3) Inject the external phase component and the internal phase component into the microfluidic channel, wherein the flow rate of the external phase component is 5 mL / h and the flow rate of the internal phase component is 0.5 mL / h;

[0077] (4) The composition from step (3) is subjected to microfluidic treatment at a pressure of 18000 psi for 3 times. Application Example 1: This application example provides a method for preparing a skincare water containing a highly effective endogenous anti-aging supramolecular composition.

[0078] The specific preparation method includes the following steps:

[0079] (1) Heat phase C and phase E to 35°C beforehand, stir until completely dissolved, and keep warm for later use;

[0080] (2) Heat each component of phase D to 35°C, sonicate until completely dissolved, and keep warm for later use;

[0081] (3) Heat each component of phase A to 35°C, stir until completely dissolved, add phase B and keep warm for later use;

[0082] (4) Add the liquid from step (3) to the liquid from step (2), stir for 5-8 minutes, and shear for 1-2 minutes;

[0083] (5) Add the pre-dissolved C phase and E phase, stir evenly and remove from the pot.

[0084] The specific components of the skin care water are shown in Table 1:

[0085] Table 1. Skincare Water Components Containing Highly Effective Endogenous Anti-aging Supramolecular Compositions

[0086]

[0087]

[0088] Comparative Example 1:

[0089] The difference from Example 1 is that tetrapeptide-1 is not added in step (1), while the other components and preparation methods are the same. Comparative Example 2:

[0090] The difference from Example 1 is that PDRN is not added in step (1), while the other components and preparation methods are the same. Comparative Example 3:

[0091] The difference from Example 1 is that in step (3), the external phase component and the internal phase component are directly mixed and stirred evenly, while the other components and preparation methods are the same.

[0092] Comparative Example 4:

[0093] The difference from Example 1 is that the microfluidic treatment method is not used in step (4), that is, step (4) is not included in the preparation method, while the other components and preparation methods are the same.

[0094] Comparative Example 5:

[0095] The difference from Example 1 is that in step (3), the external phase component and the internal phase component are directly mixed and then subjected to microfluidic treatment.

[0096] Comparative Example 6:

[0097] The difference from Application Example 1 is that the other components and their amounts remain unchanged, the E phase component is changed to the composition prepared in Comparative Example 3, and the preparation method remains unchanged.

[0098] Comparative Example 7:

[0099] The difference from Application Example 1 is that no E-phase component is added, while the other components, amounts added, and preparation methods remain unchanged.

[0100] Experimental Example 1:

[0101] In this experiment, a laser particle size analyzer was used to detect the particle size and polydispersity index (PDI) of Example 1 and Comparative Examples 3-5 of the present invention. The results are shown in Table 2. The appearance and particle size distribution of Example 1 are shown in the figure. Figure 1-2 As shown.

[0102] Table 2. Particle size and PDI test results of Example 1 and Comparative Examples 3-5

[0103]

[0104] Meanwhile, stability tests were conducted on Example 1 and Comparative Examples 3-5 of the present invention. The four groups of samples were placed at 45°C, and the particle size changes were measured on the day of preparation, one month, and three months later. The results are shown in Table 3 and [Table data would be inserted here]. Figure 3 As shown.

[0105] Table 3 Results of particle size variation

[0106]

[0107] As can be seen from the results in Tables 2 and 3, compared with Example 1, Comparative Example 3 could not obtain a transparent liquid under the same component addition amount. The particle size and PDI value of the system were larger than those of other systems. After 3 months of storage, the particle size changed from 310 nm to 786 nm, with the most significant change in particle size. This indicates that the composition system obtained by simple physical mixing has uneven particle size and poor stability. Compared with Comparative Examples 4 and 5, Example 1 had the smallest particle size and more uniform distribution under the same component content. After 3 months of storage, the particle size change was very small. This indicates that the combined treatment of microfluidics and microjets is beneficial to obtaining supramolecular compositions with good stability, small particle size, and uniform distribution.

[0108] Experimental Example 2:

[0109] Reactive oxygen species (ROS) in cells reflect the state of mitochondria to some extent. When mitochondrial function is normal, ROS levels are in dynamic equilibrium and do not fluctuate significantly. When mitochondrial function is impaired, intracellular ROS levels surge, causing cell damage and accelerating aging.

[0110] Experimental Procedure: Based on literature and related reports, human fibroblasts were selected to investigate the effects of Example 1 and Comparative Examples 1-5 on the content of reactive oxygen species (ROS) in cells. The compositions obtained in Example 1 and Comparative Examples 1-5 were diluted to 10% (g / g, mass fraction) using serum-free DMEM medium. After centrifugation, the supernatant was filtered through a 0.22 μm filter membrane to obtain 6 sets of sample stock solutions.

[0111] In this experiment, 21st generation fibroblasts were used as the young cell control, and 54th generation cells were used as the senescent cell control. The 54th generation cells were treated with 2% sample stock solution as the sample group. Each group was divided into three replicates. After culturing for 24 hours, DCFH-DA was added and the cells were cultured for another 30 minutes. After excitation at 488 nm using a multi-functional microplate reader, the fluorescence intensity of each well in the culture plate was detected at a wavelength of 525 nm. The changes in ROS content in each group are shown in Table 4. Figure 4 As shown.

[0112] Table 4 Results of changes in cellular ROS content

[0113]

[0114]

[0115] Test Results: As shown in Table 4, the ROS content in the senescent group cells was significantly higher than that in the young group cells (approximately 1.85 times), indicating that cell state is related to ROS content. Normal mitochondrial function is beneficial for maintaining cell youthfulness and delaying aging. After treating senescent cells with Examples 1, 1, and 2, the ROS content decreased in all cases. Among them, the ROS content of senescent cells treated with Example 1 showed the most significant decrease, decreasing by 0.55%, indicating that tetrapeptide-1 and PDRN have a synergistic effect in reducing intracellular ROS content. The combined use of these two components can significantly improve the antioxidant capacity of senescent cells, maintain normal mitochondrial function, and play a highly efficient endogenous anti-aging role.

[0116] Meanwhile, compared with the senescent cell groups treated with Comparative Examples 3 to 5, the senescent cells treated with Example 1 showed the most significant decrease in ROS content, which was reduced by 0.55. This indicates that the composition obtained by the present invention using the combined treatment method of microfluidics and microjet has the strongest antioxidant capacity of senescent cells and the best endogenous anti-aging effect.

[0117] Experimental Example 3:

[0118] When mitochondria themselves are impaired, they are susceptible to free radicals, leading to oxidative stress, which damages cellular mtDNA and reduces its expression level. This invention uses PCR-fluorescent probe method to investigate the effects of Example 1 and Comparative Examples 1-5 on cellular mtDNA expression levels.

[0119] Experimental procedure: The compositions obtained in Example 1 and Comparative Examples 1-5 were diluted to 10% (g / g, mass fraction) using serum-free DMEM medium. After centrifugation, the supernatant was filtered through a 0.22 μm filter membrane to obtain 6 sets of sample mother liquors.

[0120] In this experiment, 21st generation fibroblasts were used as the young cell control, and 54th generation cells were used as the senescent cell control. The 54th generation cells were treated with 2% sample stock solution as the sample group. After culturing for 24 hours, the supernatant was collected by centrifugation. Subsequent incubation procedures were performed according to the mtDNA nucleic acid detection kit instructions (Shanghai Bohu Biotechnology Co., Ltd.). After incubation, stop solution was added for 15 minutes, and the OD value at each well was measured at 450 nm. The changes in mtDNA expression levels in each group are shown in Table 5. Figure 5 As shown.

[0121] Table 5 Results of changes in cellular mtDNA expression levels

[0122] aging group 1 / Young Group 4 3 Example 1 3.2 2.2 Comparative Example 1 2.3 1.3 Comparative Example 2 1.9 0.9 Comparative Example 3 2.21 1.21 Comparative Example 4 2.05 1.05 Comparative Example 5 2.09 1.09

[0123] Test Results: As shown in Table 5, the mtDNA expression level in the young group cells was significantly higher than that in the senescent group cells (approximately 4 times), indicating that mtDNA expression level is related to cell state, and timely enhancement of mtDNA expression level in damaged cells is beneficial for delaying aging. After treating senescent cells with Examples 1, 1, and 2, the mtDNA expression level of cells was increased, but the mtDNA expression level of senescent cells treated with Example 1 showed the most significant increase, increasing by 2.2, indicating that tetrapeptide-1 and PDRN have a synergistic effect on enhancing cellular mtDNA expression level. The combined use of the two components can significantly increase cellular mtDNA expression level, which is beneficial for repairing cells damaged by oxidative stress, maintaining normal mitochondrial function, and playing a highly efficient endogenous anti-aging role.

[0124] Meanwhile, compared with the senescent cell groups treated with Comparative Examples 3-5, the expression level of mtDNA in senescent cells treated with Example 1 was significantly increased by 2.2, indicating that the composition obtained by the present invention using the combined treatment method of microfluidics and microjets has the strongest ability to improve the expression level of mtDNA in senescent cells and has the best endogenous anti-aging effect.

[0125] Experiment Example 4:

[0126] In addition to examining two indirect indicators of mitochondrial function in Experimental Examples 2 and 3, this invention also uses a cell permeation probe to directly measure the effect of Example 1 and Comparative Examples 1-5 on mitochondrial density.

[0127] Experimental procedure: The compositions obtained in Example 1 and Comparative Examples 1-5 were diluted to 10% (g / g, mass fraction) using serum-free DMEM medium. After centrifugation, the supernatant was filtered through a 0.22 μm filter membrane to obtain 6 sets of sample mother liquors.

[0128] In this experiment, 21st generation fibroblasts were used as the young cell control, and 54th generation cells were used as the senescent cell control. 54th generation cells treated with 2% sample stock solution were used as the sample group. The selected fibroblasts were seeded at a density of 3 × 10⁴ cells / well in 96-well culture plates and cultured for 24 h. Sample was then added, and the cells were cultured for another 24 h. PBS solution and Mitotracker Green probe were added to each well, and the plates were incubated at 37°C for 30 min. Cells were washed with PBS solution, excited at 490 nm using a SpectraMAX molecular detection device, and the fluorescence intensity of each well was detected at a wavelength of 516 nm. The relative mitochondrial density changes of each group are shown in Table 6. Figure 6 As shown.

[0129] Table 6 Results of changes in relative mitochondrial density in cells

[0130] aging group 100 / Young Group 146 / Example 1 134 34 Comparative Example 1 112 12 Comparative Example 2 123 23 Comparative Example 3 119 19 Comparative Example 4 116 16 Comparative Example 5 113 13

[0131] Test Results: As shown in Table 6, the mitochondrial density of the young group cells was significantly higher than that of the senescent group cells (approximately 1.46 times). This indicates that as cells age, mitochondrial density decreases, leading to insufficient energy supply and an inability to maintain normal cellular physiological activities. Treatment of senescent cells with Examples 1, 1, and 2 all increased mitochondrial density, but the increase was most significant in the senescent cells treated with Example 1, with a 34% increase. This demonstrates that tetrapeptide-1 and PDRN have a synergistic effect in increasing mitochondrial density. The combined use of these two components can significantly improve mitochondrial density, which is beneficial for mitochondrial activation, maintaining normal cellular physiological activities, mitigating the loss of mitochondrial numbers caused by aging, and thus playing a highly effective endogenous anti-aging role.

[0132] Compared with the senescent cell groups treated with Comparative Examples 3-5, the mitochondrial density of the senescent cells treated with Example 1 was significantly increased by 34%, indicating that the composition obtained by the combined microfluidic and microjet processing method of the present invention is more conducive to mitochondrial activation and has the best endogenous anti-aging effect.

[0133] Experimental Example 5:

[0134] This experiment selected Application Example 1 and Comparative Examples 6-7 to conduct a human anti-aging and firming efficacy test.

[0135] Experimental Procedure: This anti-wrinkle test was divided into 4 groups, each with 10 volunteers aged 35-55 years with loose facial skin and fine lines or wrinkles. Before the test, the same tester used a Visioline VL650 wrinkle analyzer to measure the area of ​​wrinkles on the volunteers' faces before the test, which was recorded as S0. Then, the tester applied 0.5g of the test sample evenly to the volunteers' faces twice a day (once in the morning and once in the evening). The control group applied an equal amount of purified water. The area of ​​wrinkles on the faces was recorded after 0, 7, 14, 21, and 28 days of continuous use, which was recorded as St. The reduction in wrinkle area was calculated using the following formula:

[0136] Reduction in skin wrinkle area (%) = (St-S0) / S0*100%

[0137] The test results are shown in Table 7 based on the above test plan.

[0138] Table 7 Results of the reduction in skin wrinkles

[0139]

[0140] Analysis of the data in Table 7 shows that the wrinkle removal effect is more obvious after using the skin care water containing the composition of Example 1. The area of ​​skin wrinkles can be reduced by 50.5% after 28 days. In contrast, the area of ​​skin wrinkles reduced by 20.9% after 28 days when using the skin care water containing the composition of Comparative Example 7. This indicates that adding the composition prepared in this invention to skin care products can significantly improve the wrinkle removal and anti-aging effects of skin care products. This is because the composition of this invention can reduce mitochondrial oxidative stress damage, maintain the youthful state of mitochondrial function, and delay the endogenous aging process of the skin at the cellular level, thus having a better effect.

[0141] Compared with the wrinkle-reducing and anti-aging effects of using the skin care water containing the composition of Example 1, the skin care water containing the composition of Comparative Example 6 can also reduce wrinkles, but the reduction in wrinkle area after 28 days is only 29.1%. Meanwhile, the efficiency of reducing the number of wrinkles after applying the skin care water containing Example 1 is very small at 14, 21, and 28 days, at 10.4%, 9.8%, and 6.9%, respectively, while the efficiency after applying the skin care water containing Comparative Example 6 is 5.8%, 4.1%, and 0.5%, respectively. This indicates that even with the same components added, the composition prepared by this invention has a better, more efficient, and longer-lasting wrinkle-reducing and anti-aging effect. This further verifies the effectiveness of the microfluidic and microfluidic preparation method used in this invention. The low-particle-size, highly stable, and highly efficient endogenous anti-aging composition is more conducive to exerting its efficient and long-lasting wrinkle-reducing and anti-aging effects.

[0142] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A highly efficient endogenous anti-aging supramolecular composition, characterized in that, The supramolecular composition is made of an external phase component and an internal phase component; the external phase component consists of fish egg lipid extract and caprylic / capric triglyceride; the internal phase component consists of an aqueous solution of tetrapeptide-1 and PDRN. The mass ratio of the external phase component to the internal phase component is 3~5:1; the mass percentage concentration of both the tetrapeptide-1 and PDRN in the aqueous solution is 0.5-2%. The raw material for the fish roe lipid extract is the roe of salmon, sturgeon, or flounder; the specific extraction method is as follows: the fish roe is mixed with purified water, ultrasonically crushed, treated by a high-pressure homogenizer, and the bottom product is collected after centrifugation, which is the fish roe lipid extract. The molecular weight of the PDRN is 0.5-1.5 million; the mass ratio of the fish egg lipid extract to caprylic / capric triglyceride is 1:5-10; The preparation method of the highly efficient endogenous anti-aging supramolecular composition includes the following steps: S1. Tetrapeptide-1 and PDRN are dissolved in water, and the pH of the solution is adjusted to obtain the internal phase component; the pH value is 4.7~7.

2. S2, a mixture of fish egg lipid extract and caprylic / capric triglyceride was prepared to obtain the external phase component; S3. The external phase component and the internal phase component are brought into contact with each other using microfluidic technology to form a composition; in the microfluidic technology, the flow rate of the external phase component is 3~8 mL / h, and the flow rate of the internal phase component is 0.5~1 mL / h. S4. The composition is subjected to high-pressure microfluidic treatment to obtain a supramolecular composition; in the high-pressure microfluidic treatment, the pressure is set to 15000-20000 psi, and the treatment is performed 3 to 5 times.

2. The supramolecular composition according to claim 1, characterized in that, The mass ratio of the fish egg lipid extract to caprylic / capric triglyceride is 1:7~9.

3. The supramolecular composition according to claim 1, characterized in that, The high-pressure homogenizer operates at a pressure of 5-60 MPa, a flow rate of 20-45 L / h, and undergoes 1-3 homogenization cycles.

4. A method for preparing the highly efficient endogenous anti-aging supramolecular composition according to claim 1, characterized in that, Includes the following steps: S1. Tetrapeptide-1 and PDRN are dissolved in water, and the pH of the solution is adjusted to obtain the internal phase component; the pH value is 4.7~7.

2. S2, a mixture of fish egg lipid extract and caprylic / capric triglyceride was prepared to obtain the external phase component; S3. The external phase component and the internal phase component are brought into contact with each other using microfluidic technology to form a composition; in the microfluidic technology, the flow rate of the external phase component is 3~8 mL / h, and the flow rate of the internal phase component is 0.5~1 mL / h. S4. The composition is subjected to high-pressure microfluidic treatment to obtain a supramolecular composition; in the high-pressure microfluidic treatment, the pressure is set to 15000-20000 psi, and the treatment is performed 3 to 5 times.

5. A composition, characterized in that, The composition comprises the highly efficient endogenous anti-aging supramolecular composition and functional peptides as described in any one of claims 1 to 3.

6. The composition according to claim 5, characterized in that, The functional polypeptide is selected from one or more of palmitoyl polypeptides, acetyl polypeptides, arginine / lysine polypeptides, decapeptide-4, hexapeptide-9, nonapeptide-1, and dipeptide diaminobutyryl benzylamide diacetate.

7. The composition according to claim 5, characterized in that, The composition also includes a humectant, an emulsifier, a preservative, and water.

8. The composition according to claim 7, characterized in that, The moisturizer is selected from one or more of glycerin, propylene glycol, butylene glycol, hexanediol, sodium hyaluronate, betaine, and trehalose; the emulsifier is selected from one or more of glyceryl stearate, hydrogenated lecithin, polysorbate-80, polysorbate-20, sorbitan olive oil ester, polyglycerol-6 stearate, and polyglycerol-6 behenate; the preservative is selected from one or more of methylparaben, phenoxyethanol, p-hydroxyacetophenone, sodium benzoate, and potassium sorbate.

9. The composition according to any one of claims 5 to 8, characterized in that, The composition contains 0.001-3% by mass of a highly effective endogenous anti-aging supramolecular composition; 0.0001-1% by mass of a functional peptide; 0.1-10% by mass of a moisturizer; 0.001-2% by mass of a preservative; and 0.001-5% by mass of an emulsifier.

10. The use of the highly effective endogenous anti-aging supramolecular composition according to any one of claims 1 to 3 or the composition according to claims 5 to 9 in the preparation of skin care products; The skincare products mentioned are serums, lotions, toners, creams, or masks; The skincare product has wrinkle-reducing and anti-aging effects.

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