A polypeptide complex composition and its use in skin lesion repair

Abstract

The application discloses a kind of polypeptide complex composition and its application in skin damage repair, belong to cosmetic technical field, specifically related to a kind of polypeptide complex composition, including carnosine and rutin, and the mass ratio of carnosine and rutin is 1-10:1-10.Carnosine and rutin are compounded in specific ratio, and under suitable ratio compounding, it has good effect to skin oxidative damage repair, and the compounding of carnosine and rutin has synergistic effect, in the compounding of carnosine and rutin, auxiliary materials can also be added, and the auxiliary materials include 6-hydroxy nicotinic acid and / or coffee acyl tartaric acid and / or penetration enhancer.The application is a kind of polypeptide complex composition and its application in skin damage repair, which has good anti-glycosylation effect, has protective effect on cell oxidative damage, has high expression of type I collagen (Col1A1), and can up-regulate antioxidant genes.

Description

Technical Field

[0001] This invention belongs to the field of cosmetic technology, specifically relating to a polypeptide compound composition and its application in skin damage repair. Background Technology

[0002] Oxidative stress is a core cause of skin aging and functional damage. Excessive production of reactive oxygen species (ROS) can trigger lipid peroxidation, cell damage, and metabolic disorders, which in turn can lead to problems such as dull skin, wrinkles, and decreased elasticity. At the same time, oxidative stress can accelerate the accumulation of advanced glycation end products (AGEs), leading to cross-linking and hardening of collagen fibers. This, combined with ultraviolet (UV)-induced photodamage (acute sunburn, chronic photoaging), results in multiple skin hazards of "oxidation-glycation-photodamage".

[0003] Reactive oxygen species (ROS) are a major threat to skin health, including hydroxyl radicals (・OH), superoxide anions (O2⁻), and hydrogen peroxide (H2O2). Excessive generation of these substances disrupts the skin's oxidation-antioxidant balance. Oxidative stress not only directly damages the structure and function of keratinocytes and fibroblasts but also activates inflammatory pathways, accelerates collagen degradation and glycation, and induces skin aging. Combined with ultraviolet radiation, this further exacerbates DNA damage and collagen metabolism imbalance, creating a vicious cycle of photodamage compounded by oxidative damage. Developing compound ingredients with high-efficiency antioxidant properties and multiple protective effects is a core requirement in the functional skincare field. Summary of the Invention

[0004] The purpose of this invention is to provide a polypeptide compound composition with good anti-glycation effect, protective effect against cellular oxidative damage, high expression level of type I collagen (Col1A1), and the ability to upregulate antioxidant genes, and its application in skin damage repair.

[0005] The technical solution adopted by the present invention to achieve the above objectives is as follows: A polypeptide compound composition comprising carnosine and rutin, wherein the mass ratio of carnosine to rutin is 1-10:1-10.

[0006] Carnosine, β-alanyl-L-histidine, is widely found in mammalian tissues and possesses multiple biological functions. It can directly scavenge ROS such as hydroxyl radicals and hydrogen peroxide, and chelate Fe... 2+ Cu 2+ It uses transition metal ions to inhibit lipid peroxidation; reduces the formation of advanced glycation end products (AGEs) and prevents collagen fiber cross-linking and hardening; and downregulates the expression of pro-inflammatory factors such as IL-6 and TNF-α by inhibiting the NF-κB pathway. Preclinical studies have shown that it can improve skin oxidative dullness, but its activation of endogenous antioxidant pathways is insufficient when used alone.

[0007] Rutin is a natural flavonoid compound whose core advantage lies in its antioxidant function. Firstly, its polyhydroxy structure (three phenolic hydroxyl groups) acts as a hydrogen donor, effectively scavenging various ROS induced by UV radiation or metabolic processes and inhibiting lipid peroxidation. Secondly, by upregulating the nuclear transcriptional activity of Nrf2 (nuclear factor E2-related factor 2), it promotes the expression of antioxidant enzymes such as SOD (superoxide dismutase), GPx (glutathione peroxidase), and CAT (catalase), thereby enhancing the skin's own antioxidant capacity. In addition, rutin also has certain collagen-protective and anti-inflammatory effects, but its intervention effect on glycation is limited.

[0008] This invention discovers that peptide complex compositions comprising carnosine and rutin, when safe, effective, and in appropriate proportions, exhibit a synergistic effect of "core antioxidant - auxiliary anti-glycation - synergistic anti-photodamage." Under appropriate proportions, the effect of the peptide complex composition at the same dosage is superior to that of the same dosage of carnosine or rutin. When carnosine and rutin are mixed, the effect of the peptide complex composition at the same dosage should be between that of carnosine and rutin alone. However, the effect of the peptide complex composition at the same dosage is far superior to that of carnosine or rutin alone, indicating that the combined use of carnosine or rutin has a good effect. However, if the ratio of carnosine to rutin is inappropriate, there is almost no synergistic effect.

[0009] Preferably, the polypeptide compound composition further includes a penetration enhancer.

[0010] Preferably, the mass ratio of carnosine to rutin is 3:1; or, the mass ratio of carnosine to rutin is 1:1; or, the mass ratio of carnosine to rutin is 1:3; or, the mass ratio of carnosine to rutin is 2.5:1; or, the mass ratio of carnosine to rutin is 3.5:1.

[0011] Preferably, the penetration enhancer includes at least one of azone, NMP, and Span 80.

[0012] Preferably, the polypeptide compound composition further includes 6-hydroxynicotinic acid, and the mass ratio of carnosine to 6-hydroxynicotinic acid is 1-10:0.01-0.2. In this invention, by further adding 6-hydroxynicotinic acid to the compound of carnosine and rutin, the anti-glycation effect and the protective effect against cell damage can be improved.

[0013] More preferably, the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.01-0.2.

[0014] Preferably, the polypeptide compound composition further contains caffeoyl tartaric acid, and the mass ratio of carnosine to caffeoyl tartaric acid is 1-10:0.01-0.1. In this invention, by further adding caffeoyl tartaric acid to the compound of carnosine and rutin, the anti-glycation effect and the protective effect against cell damage can be improved.

[0015] More preferably, the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.01-0.1.

[0016] Preferably, the polypeptide compound composition also contains 6-hydroxynicotinic acid and caffeoyl tartaric acid. When using carnosine and rutin, the present invention can also add 6-hydroxynicotinic acid and caffeoyl tartaric acid together. The amount of 6-hydroxynicotinic acid and caffeoyl tartaric acid used needs to meet a certain range. Under the appropriate amount of combined use, it has a better anti-glycation effect and a protective effect against cell damage.

[0017] This invention discloses a cosmetic product comprising the above-mentioned polypeptide compound composition.

[0018] Preferably, the cosmetic also includes water or PBS.

[0019] Preferably, the concentration of carnosine is 0.05-0.5 mg / mL.

[0020] Preferably, the cosmetic comprises a polypeptide complex composition and water. The polypeptide complex composition comprises carnosine and rutin, wherein the mass ratio of carnosine to rutin is 1-10:1-10, and the mass ratio of carnosine to water is 0.00005-0.0005:1.

[0021] More preferably, cosmetics also contain penetration enhancers.

[0022] More preferably, the penetration enhancer includes azone, and the mass ratio of carnosine and azone is 1-10:0.1-1.

[0023] More preferably, the penetration enhancer includes NMP, and the mass ratio of carnosine and NMP is 1-10:0.1-1.

[0024] More preferably, the penetration enhancer includes Span 80, and the mass ratio of carnosine to Span 80 is 1-10:0.1-1.

[0025] This invention discloses the use of the above-mentioned polypeptide complex composition in the preparation of cosmetics and / or skin repair products and / or topical medicines.

[0026] This invention utilizes a combination of carnosine and rutin to prepare a polypeptide compound composition. Carnosine and rutin are combined in a specific ratio, and under appropriate ratios, they exhibit good effects on repairing oxidative damage to the skin. The combination of carnosine and rutin has a synergistic effect. Excipients, including 6-hydroxynicotinic acid and / or caffeoyl tartaric acid and / or penetration enhancers, can also be added to the carnosine and rutin compound composition, resulting in the following beneficial effects: the polypeptide compound composition has good anti-glycation effects, protects against cellular oxidative damage, and exhibits high expression levels of type I collagen (Col1A1), which can upregulate antioxidant genes. Therefore, this invention relates to a polypeptide compound composition with good anti-glycation effects, protective effects against cellular oxidative damage, high expression levels of type I collagen (Col1A1), and the ability to upregulate antioxidant genes, and its application in skin damage repair. Attached Figure Description

[0027] Figure 1 This is a graph showing the inhibition rate of AGEs.

[0028] Figure 2 This is a graph showing cytotoxicity.

[0029] Figure 3 The figure shows the results of the protective effect against oxidative damage to cells.

[0030] Figure 4 This is a graph showing the intracellular MDA content.

[0031] Figure 5 This is a graph of intracellular SOD activity.

[0032] Figure 6 Immunofluorescence image of COL1A1 in a skin model.

[0033] Figure 7 This is a graph showing relative gene expression levels.

[0034] Figure 8 This is a graph showing the results of the anti-glycation test.

[0035] Figure 9 The figure shows the results of the cell oxidative damage protection test. Detailed Implementation

[0036] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0037] The concepts involved in this application will first be described with reference to the accompanying drawings. It should be noted that the following descriptions of various concepts are only for the purpose of making the content of this application easier to understand and do not constitute a limitation on the scope of protection of this application; furthermore, the embodiments and features in the embodiments of this application can be combined with each other unless otherwise specified. This application will now be described in detail with reference to the accompanying drawings and embodiments.

[0038] Research basis: In this study, the safe concentrations of carnosine and rutin were tested using fibroblasts. Based on the results of the cytotoxicity experiment, the safe concentration range of carnosine is 0.05-0.5 mg / mL, at which the fibroblast survival rate is ≥80%; the safe concentration range of rutin is 0.05-0.5 mg / mL, at which the fibroblast survival rate is ≥80%.

[0039] The purity of carnosine and rutin was ≥98%. Carnosine and rutin were prepared into a stock solution using deionized water as a diluent. When preparing the experimental concentration, the stock solution was sterilized by filtration through a 0.22μm filter membrane and stored in a light-proof, sealed container.

[0040] 1. Anti-glycation test 1) Sample preparation: Take a 96-well microplate and add 20 mg / mL BSA solution and 1 wt% MGO solution to each well at a mass ratio of BSA to MGO of 1:1. Adjust the volume as needed to ensure that the system is suitable for the well volume of the microplate.

[0041] 2) Group processing: Add positive control aminoguanidine acid (AG) solution, test sample solution (0.12 mg / mL) or negative control DMSO to the corresponding wells respectively, and shake to mix for 1-2 min.

[0042] 3) Initial fluorescence detection: Using a fluorescence microplate reader, the initial fluorescence value of each well (RFU blank control, i.e., the fluorescence value at 0h) was detected under the conditions of excitation wavelength 370nm and emission wavelength 440nm.

[0043] 4) Incubation at constant temperature: Place the ELISA plate in a constant temperature shaker, set the temperature to 45℃ and keep it in the dark, and incubate for 16 hours.

[0044] 5) Post-incubation treatment: After incubation, place the microplate in a centrifuge and centrifuge at 1000 rpm for 5 minutes.

[0045] 6) Terminal fluorescence detection: The terminal fluorescence value of each well was detected again using a fluorescence microplate reader at the same excitation / emission wavelength (370nm / 440nm) (the fluorescence value of the negative control well in the RFU control group and the fluorescence value of the test sample / positive control well in the RFU experimental group).

[0046] Formula for calculating AGEs inhibition rate R% = 1 - (RFU test group - RFU blank control / RFU negative control - RFU blank control) × 100%.

[0047] Explanation: RFU blank control = fluorescence values ​​of BSA and MGO at 0h; RFU control group = fluorescence value of negative control (DMSO) after 16h incubation; RFU experimental group = fluorescence value of test sample / positive control after 16h incubation.

[0048] AGEs inhibition rate results are as follows Figure 1 As shown, in the experiment of inhibiting AGEs formation, the compound of carnosine and rutin showed a significant anti-glycation effect compared with the ginseng drug aminoguanidine (AG); the anti-glycation inhibition rate of ginseng AG at the same concentration (0.12 mg / mL) was about 63.4%, and the anti-glycation inhibition rate of the carnosine:rutin (3:1) compound at the same concentration (0.12 mg / mL) was about 61.7%.

[0049] 2. Cytotoxicity test 1) Cell treatment This study used HaCaT cells, at a ratio of 1×10⁶ cells per well. 4 Cells were seeded at a density of 0.2 mL per well in 96-well plates. The culture medium consisted of 90% DMEM, 10% FBS, and 1% penicillin-streptomycin (100X). The plates were then incubated at 37°C in a 5% CO2 incubator for 24 h. A blank control group, a negative control group, and experimental groups were set up. The blank control group contained no sample treatment and no cell-containing complete culture medium. The negative control group contained no sample treatment and cell-containing complete culture medium. The experimental groups included rutin, carnosine, and mixtures of rutin and carnosine in different mass ratios of 3:1, 1:1, and 1:3. The rutin group contained rutin at three concentrations: 0.05, 0.1, and 0.2 mg / mL. The carnosine group contained carnosine at three concentrations: 0.05, 0.1, and 0.2 mg / mL. In the rutin and carnosine mixtures with different mass ratios, the concentrations of rutin and carnosine were 0.09 mg / mL and 0.03 mg / mL in the 3:1 mixture, 0.06 mg / mL and 0.06 mg / mL in the 1:1 mixture, and 0.03 mg / mL and 0.09 mg / mL in the 1:3 mixture.

[0050] 2) CCK-8 detection After cell culture, 20 μL of CCK-8 solution was added to each well, and the cells were incubated for another 1 h. The absorbance was measured at 450 nm using a microplate reader, and the cell viability was calculated.

[0051] Cytotoxicity test results as follows Figure 2 As shown, the survival rate of keratinocytes in the negative control was approximately 100%, representing the baseline of normal cell viability. The survival rates of keratinocytes in the rutin monotherapy group, the carnosine monotherapy group, and the combined treatment groups of carnosine-rutin (3:1), carnosine-rutin (1:1), and carnosine-rutin (1:3) were all maintained in the range of 90% to 100%, with no significant difference from the level of the negative control.

[0052] In summary, under the conditions of this experiment, neither rutin nor carnosine, whether used alone or in different proportions, produced significant cytotoxicity to keratinocytes, and cell viability was not significantly inhibited.

[0053] 3. Protective test against cellular oxidative damage 1) Cell viability The experimental group was divided into a negative control group, a model group, and a sample group, with three replicates in each group. The control group received cell-containing culture medium without drug treatment; the model group received H2O2; and the sample group received the test solution + H2O2. HaCaT cells were cultured at 1×10⁻⁶ cells / day. 4 Cells were seeded at a density of 100 cells / well in 96-well plates and cultured for 24 h. The cells were then treated with the prepared sample solution and cultured for another 24 h. After washing twice with PBS, 500 μM H2O2 was added and the cells were cultured for 4 h. After washing twice with PBS, 20 μL of CCK-8 solution was added to every 200 μL of culture medium. After culturing for 2 h, the OD value of each well was measured at 450 nm using a microplate reader. The negative control group received no sample treatment and no H2O2 treatment. The model group received H2O2 treatment. The sample groups included carnosine, rutin, and mixtures of carnosine and rutin in different mass ratios. The carnosine concentration in the carnosine group was 0.12 mg / mL, and the rutin concentration in the rutin group was 0.12 mg / mL. The mixtures of carnosine and rutin in different mass ratios were 3:1, 1:1, and 1:3. In the 3:1 mixture, the concentrations of rutin and carnosine were 0.09 mg / mL and 0.03 mg / mL, respectively; in the 1:1 mixture, the concentrations were 0.06 mg / mL and 0.06 mg / mL, respectively; and in the 1:3 mixture, the concentrations were 0.03 mg / mL and 0.09 mg / mL, respectively. The negative control group was treated with PBS, and PBS was used as the diluent in the experiment.

[0054] Cell viability = [(OD model - OD blank) / (OD control - OD blank)] × 100%.

[0055] The results of the cell viability test are as follows: Figure 3As shown in the figure, a cell oxidative damage model was established using 500 μM H2O2 to investigate the protective effects of rutin, carnosine, and their different ratios (3:1, 1:1, 1:3) against H2O2-induced oxidative damage in HaCaT cells. The results in the figure show that, compared with the H2O2 model group, the carnosine-rutin (3:1) combination group showed a highly significant protective effect against cell oxidative damage (p<0.01), with a cell survival rate of 80.17%. The carnosine monotherapy group and the carnosine-rutin (1:1) combination group also showed significant protective effects against cell oxidative damage (p<0.05), with the carnosine monotherapy group achieving a cell survival rate of 71.73% and the carnosine-rutin (1:1) combination group achieving a cell survival rate of 75.17%. This indicates that among the treatments of rutin and carnosine alone and in different ratios, a total of five treatment methods showed significant protective effects against H2O2-induced oxidative damage in HaCaT cells, with the 3:1 ratio showing the best protective effect.

[0056] 2) Tests of SOD activity and MDA content in HaCaT cells damaged by H2O2 SOD and MDA activity assay: Cells were used at a concentration of 2×10⁻⁶ 5 Cells were seeded at a density of 100 cells / well in 6-well plates and cultured for 24 h. Different groups of samples were treated for 24 h, then treated with H2O2 for 4 h. After washing twice with PBS, cells were collected by adding cell lysis buffer, and the content of SOD, MDA, etc. in the cell supernatant was detected according to the kit instructions.

[0057] MDA content test results are as follows Figure 4 As shown, compared with the model group, the intracellular MDA content was significantly reduced in the carnosine monotherapy group, the carnosine-rutin (3:1) combination group, and the carnosine-rutin (1:3) combination group (P<0.01). The intracellular MDA content was also significantly reduced in the rutin monotherapy group and the carnosine-rutin (1:1) combination group (P<0.05).

[0058] SOD activity test results are as follows: Figure 5 As shown, compared with the H2O2-induced oxidative damage model group, the SOD activity in HaCaT cells treated with the carnosine-rutin (3:1), carnosine-rutin (1:1), and carnosine-rutin (1:3) combination groups was significantly increased (P<0.01). This indicates that rutin and carnosine, used alone or in combination at different ratios, have antioxidant capabilities that enhance SOD activity and reduce MDA content in HaCaT cells, with the 3:1 ratio showing a particularly significant effect on enhancing cellular antioxidant activity.

[0059] 4. 3D dermal skin model testing 1) Model construction: A 3D dermal skin model was constructed using keratinocytes and dermal fibroblasts. The cells were cultured in the corresponding complete culture medium for 1 week and then subjected to UV irradiation to construct a photodamage model.

[0060] 2) Grouping Treatment: A negative control group, a positive control group, and a compound composition treatment group were set up. The compound composition was applied to the model surface and incubated for 48 hours. The negative control group was treated with PBS, and the positive control group was treated with TGF-β. The concentration of TGF-β in the positive control group was 10 ng / mL. In the compound composition treatment group, the mass ratio of carnosine to rutin was 3:1, and the concentrations of rutin and carnosine were 0.09 mg / mL and 0.03 mg / mL, respectively. PBS was used as the diluent in the experiment.

[0061] 3) Detection indicators: Collagen expression: Immunofluorescence staining was used to detect the localization and fluorescence intensity of type I collagen (COL1A1), and ImageJ software was used for quantitative analysis.

[0062] Immunofluorescence: Sample preparation and fixation: The 3D skin model (6 mm in diameter) after UV irradiation was peeled off, immersed in 4% paraformaldehyde solution and fixed at room temperature for 2-4 hours, and rinsed 3 times with PBS (5 minutes each time).

[0063] Permeability and Blocking: Transfer to 0.3% Triton X-100 solution and incubate at room temperature for 30 minutes to 1 hour, or incubate at room temperature for 1 hour with 5% BSA (or 10% normal goat serum). For 3D models, 0.1% Tween-20 can be added to enhance the blocking effect.

[0064] Antibody incubation: Incubate primary antibody (anti-COL1A1 antibody diluted 1:200) at 4°C overnight (or at 37°C for 2 hours), rinse with PBS, add fluorescently labeled secondary antibody (diluted 1:500), and incubate at room temperature in the dark for 1 hour.

[0065] Nuclear staining and mounting: Incubate with DAPI solution (1:1000 dilution) at room temperature in the dark for 5 minutes, fix with anti-quenching mounting medium, dry in the dark for 24 hours, and store at 4°C.

[0066] Imaging and Analysis: Laser confocal microscope images were taken, and ImageJ software was used to set a threshold to exclude the background. The average fluorescence intensity of the target area was calculated. At least three fields of view were analyzed for each group, and the results were expressed as "mean ± standard deviation".

[0067] The results of the localization and expression levels of type I collagen (Col1A1) are as follows: Figure 6As shown, the merging refers to the blue staining of cell nuclei and the green staining of type I collagen. After UVA irradiation, the expression level of type I collagen (Col1A1) in the 3D dermal skin model decreased, while the expression level of type I collagen (Col1A1) significantly increased after treatment with the rutin and carnosine combination. These results indicate that the rutin and carnosine combination significantly promotes the function of type I collagen. The rutin and carnosine combination significantly increases the content of type I collagen.

[0068] 5. Quantitative Genetic Detection Study (QGP) 1) Cell culture refers to cell culture in section 2(1).

[0069] 2) According to 6×10 per hole 4 Cells were seeded at a density of 0.5 mL in 24-well plates. The plates were incubated for 24 h. The experiment included a negative control group, a carnosine group, a rutin group, and mixed groups with different mass ratios of carnosine and rutin, each group containing three replicates. The negative control group was treated with PBS. The carnosine group contained 0.12 mg / mL of carnosine, and the rutin group contained 0.12 mg / mL of rutin. In the mixed groups with different mass ratios of carnosine and rutin (3:1, 1:1, and 1:3), the concentrations of rutin and carnosine were 0.09 mg / mL and 0.03 mg / mL, respectively; in the 1:1 group, the concentrations were 0.06 mg / mL and 0.06 mg / mL, respectively; and in the 1:3 group, the concentrations were 0.03 mg / mL and 0.09 mg / mL, respectively. PBS was used as the diluent in the experiment.

[0070] 3) After 24 h of culture, the supernatant was removed, and 0.2 mL of cell lysis buffer (Lysis Mixture) was added to the wells containing cells to lyse the cells. The lysed cells were stored at -80°C.

[0071] 4) Place the cell lysate and cell lysis buffer at room temperature in advance, and then incubate at 37°C for 30 min.

[0072] 5) Reagent processing: Thaw and mix the probe set and blocking reagent, briefly centrifuge the probe set and collect the bottom precipitate; store proteinase K on ice; remove the capture beads and store them in the dark.

[0073] 6) Prepare the working bead mix according to the instructions. Add 20 μL of the working bead mix to each well of the hybridization plate. At the same time, add 80 μL of cell lysates and diluent lysis mixture.

[0074] 7) Seal the hybridization plate and incubate at 54±1℃ and 600 rpm for 18-22 h.

[0075] 8) After the pre-amplifier solution, amplifier solution, and label probe solution have been brought to room temperature, they should be incubated at 37°C for 30 min for later use; the SAPE diluent should be kept at room temperature for later use.

[0076] 9) Prepare 1X Wash Buffer.

[0077] 10) Centrifuge the hybridization plate, remove the seal, transfer the liquid up and down 5 times, and then transfer the liquid to the magnetic separation plate.

[0078] 11) Wash the plate: Place the magnetic separation plate into the washing machine and then invert it to remove the solution.

[0079] 12) Perform the reaction of the pre-amplifier solution, amplifier solution, and label probe solution.

[0080] 13) After incubation, wash the plate and repeat the washing steps.

[0081] 14) Combine SAPE (Bind SAPE) to carry out the SAPE reaction.

[0082] 15) Wash the plate and repeat the washing steps.

[0083] 16) For analysis, add 130 μL of SAPE wash buffer to each well, shake, and then perform detection and analysis.

[0084] Quantitative gene detection results as follows Figure 7 As shown, compared with the negative control, both rutin and carnosine monotherapy upregulated antioxidant-related genes (rutin targets NFE2L2, SOD2, and GSR, while carnosine targets NFE2L2 and CAT) and extracellular matrix-related genes COL1A1 and AQP3, while downregulating pro-inflammatory genes TIMP2 (rutin) and IL6 (carnosine), respectively. Rutin is more focused on activating the antioxidant enzyme system, while carnosine is more focused on regulating matrix homeostasis and anti-glycation function.

[0085] The regulatory effect of the combination treatment with carnosine and rutin was significantly better than that of the single drug group, demonstrating a synergistic effect: the combination of carnosine and rutin at a mass ratio of 1:3 significantly upregulated the antioxidant genes CAT and GSR (p<0.05) and the matrix-related gene COL1A1 (p<0.01), while downregulating TIMP2 (p<0.01) and IL6 (p<0.05). Carnosine and rutin achieved balanced regulation at a mass ratio of 1:1, significantly upregulating antioxidant genes SOD2 (p<0.05) and CAT (p<0.01), matrix-related genes COL1A1 (p<0.01) and ELN (p<0.05), and AQP3 and anti-glycation-related genes (p<0.05), while downregulating IL6 (p<0.05). The optimal regulatory intensity was achieved when carnosine and rutin were in a mass ratio of 3:1, which significantly upregulated antioxidant genes SOD2, GSR, and CAT (p<0.01), matrix-related genes COL1A1, ELN, and AQP3, and anti-glycation-related genes, while significantly downregulating TIMP2 and IL6.

[0086] In summary, the combination of carnosine and rutin achieves multi-dimensional regulation of "antioxidation-promoting repair-suppressing inflammation" through synergistic effects. Among them, the complementary effect of carnosine and rutin in the regulation of antioxidation, collagen synthesis and anti-glycation is the most prominent when the mass ratio of carnosine to rutin is 3:1, and the regulatory efficiency is comprehensively better than that of single drugs and other combination ratios.

[0087] Example 1: A polypeptide compound composition The polypeptide compound composition includes carnosine and rutin, with a mass ratio of carnosine to rutin of 3:1.

[0088] Example 2: A polypeptide compound composition The polypeptide compound composition includes carnosine and rutin, with a mass ratio of carnosine to rutin of 1:1.

[0089] Example 3: A polypeptide compound composition The polypeptide compound composition includes carnosine and rutin, with a mass ratio of carnosine to rutin of 1:3.

[0090] Comparative Example 1: A polypeptide complex composition The polypeptide compound composition includes carnosine and rutin, with a mass ratio of carnosine to rutin of 1:0.01.

[0091] Comparative Example 2: A polypeptide complex composition A polypeptide compound composition comprising carnosine and rutin, wherein the mass ratio of carnosine to rutin is 1:100.

[0092] Example 4: A polypeptide compound composition The polypeptide compound composition includes carnosine, rutin and 6-hydroxynicotinic acid, wherein the mass ratio of carnosine to rutin is 3:1 and the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.1.

[0093] Example 5: A polypeptide compound composition The polypeptide compound composition includes carnosine, rutin and 6-hydroxynicotinic acid, wherein the mass ratio of carnosine to rutin is 3:1 and the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.02.

[0094] Example 6: A polypeptide compound composition The polypeptide complex composition includes carnosine, rutin, and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1, and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.09.

[0095] Example 7: A polypeptide compound composition The polypeptide complex composition includes carnosine, rutin and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1 and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.03.

[0096] Example 8: A polypeptide compound composition The polypeptide complex composition includes carnosine, rutin, 6-hydroxynicotinic acid and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1, the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.1, and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.09.

[0097] Example 9: A polypeptide compound composition The polypeptide complex composition includes carnosine, rutin, 6-hydroxynicotinic acid and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1, the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.1, and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.03.

[0098] Example 10: A polypeptide compound composition The polypeptide complex composition includes carnosine, rutin, 6-hydroxynicotinic acid and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1, the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.02, and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.09.

[0099] Example 11: A polypeptide compound composition The polypeptide complex composition includes carnosine, rutin, 6-hydroxynicotinic acid and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1, the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.03, and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.03.

[0100] Comparative Example 3: A polypeptide complex composition The polypeptide complex composition includes carnosine, rutin, 6-hydroxynicotinic acid and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1, the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.008, and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.09.

[0101] Comparative Example 4: A polypeptide complex composition The polypeptide complex composition includes carnosine, rutin, 6-hydroxynicotinic acid and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1, the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.1, and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.009.

[0102] Comparative Example 5: A polypeptide complex composition The polypeptide complex composition includes carnosine, rutin, 6-hydroxynicotinic acid and caffeoyl tartaric acid, wherein the mass ratio of carnosine to rutin is 3:1, the mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.008, and the mass ratio of carnosine to caffeoyl tartaric acid is 3:0.009.

[0103] Example 12: A cosmetic Cosmetics, including peptide complex compositions and water. The peptide complex composition includes carnosine and rutin in a mass ratio of 3:1, and carnosine and water in a mass ratio of 0.00009:1.

[0104] Example 13: A cosmetic Cosmetics, including peptide complex compositions, azone, and water. The peptide complex composition includes carnosine and rutin in a mass ratio of 3:1, carnosine and azone in a mass ratio of 10:1, and carnosine and water in a mass ratio of 0.00009:1.

[0105] Example 14: A cosmetic Cosmetics, including a peptide complex composition, NMP, and water. The peptide complex composition includes carnosine and rutin in a mass ratio of 3:1, carnosine and NMP in a mass ratio of 10:1, and carnosine and water in a mass ratio of 0.00009:1.

[0106] Example 15: A cosmetic Cosmetic products, including a peptide complex composition, Span 80, and water. The peptide complex composition includes carnosine and rutin in a mass ratio of 3:1, carnosine and Span 80 in a mass ratio of 10:1, and carnosine and water in a mass ratio of 0.00009:1.

[0107] Experimental example: 1. Anti-glycation test The present invention tested the polypeptide complex compositions prepared in Examples 1-11 and Comparative Examples 1-5 according to the above-described anti-glycation test method. Deionized water was used as a diluent to prepare a stock solution, which was then prepared to a concentration of carnosine and rutin of 0.1 mg / mL. The solution was sterilized by filtration through a 0.22 μm filter membrane. The test results are as follows: Figure 8 As shown, S1 is Example 1, S4 is Example 4, S5 is Example 5, S6 is Example 6, S7 is Example 7, S8 is Example 8, S9 is Example 9, S10 is Example 10, S11 is Example 11, D1 is Comparative Example 1, D2 is Comparative Example 2, D3 is Comparative Example 3, D4 is Comparative Example 4, and D5 is Comparative Example 5. In this invention, the concentrations of carnosine and rutin are controlled to be 0.1 mg / mL. When carnosine and rutin are used in combination, they need to meet a certain ratio. Within a suitable ratio range, the combined use of carnosine and rutin is superior to the use of carnosine or rutin alone. However, if the proportion of one of the carnosine and rutin in the combined use is too low, there is no synergistic effect. Furthermore, in the research section of this invention, the concentrations of carnosine and rutin are controlled to be 0.1 mg / mL. The concentration of 0.12 mg / mL also demonstrates that carnosine and rutin, under certain ratios, have excellent synergistic effects. In preparing polypeptide complex compositions using carnosine and rutin, this invention can also add 6-hydroxynicotinic acid. At effective concentrations and ratios of carnosine and rutin, the use of 6-hydroxynicotinic acid can improve the AGEs inhibition rate and anti-glycation effect of the polypeptide complex composition. Furthermore, at effective concentrations and ratios of carnosine and rutin, this invention can also add caffeoyl tartaric acid. The resulting polypeptide complex composition also exhibits better anti-glycation effects. Further, 6-hydroxynicotinic acid and caffeoyl tartaric acid can be added together. At effective concentrations and ratios of carnosine and rutin, the combined use of 6-hydroxynicotinic acid and caffeoyl tartaric acid is superior to the use of either 6-hydroxynicotinic acid or caffeoyl tartaric acid alone.

[0108] 2. Tests for protection against cellular oxidative damage The present invention tested the polypeptide complex compositions prepared in Examples 1-11 and Comparative Examples 1-5 according to the above-described test method for protection against cellular oxidative damage. Deionized water was used as a diluent to prepare a stock solution, which was then prepared to a concentration of carnosine and rutin of 0.1 mg / mL. The solution was sterilized by filtration through a 0.22 μm filter membrane. The test results are as follows: Figure 9 As shown, S1 is Example 1, S4 is Example 4, S5 is Example 5, S6 is Example 6, S7 is Example 7, S8 is Example 8, S9 is Example 9, S10 is Example 10, S11 is Example 11, D1 is Comparative Example 1, D2 is Comparative Example 2, D3 is Comparative Example 3, D4 is Comparative Example 4, and D5 is Comparative Example 5. In this invention, the concentrations of carnosine and rutin are controlled to be 0.1 mg / mL. When carnosine and rutin are used in combination, they need to meet a certain ratio. Within a suitable ratio range, the combined use of carnosine and rutin is better than the use of carnosine or rutin alone. However, if the proportion of one of the carnosine and rutin in the combined use is too small, there will be no synergistic effect. In the research part of this invention, the concentrations of carnosine and rutin are controlled to be 0.1 mg / mL. The concentration of 12 mg / mL also demonstrates that carnosine and rutin, when combined in a certain ratio, exhibit excellent synergistic effects. In preparing polypeptide complex compositions using carnosine and rutin, this invention can further incorporate 6-hydroxynicotinic acid. At effective concentrations and ratios of carnosine and rutin, the use of 6-hydroxynicotinic acid can enhance the protective effect of the polypeptide complex composition against cellular oxidative damage. Furthermore, at effective concentrations and ratios of carnosine and rutin, this invention can also incorporate caffeoyl tartaric acid. The resulting polypeptide complex composition also exhibits better protective effects against cellular oxidative damage. Additionally, 6-hydroxynicotinic acid and caffeoyl tartaric acid can be added together. At effective concentrations and ratios of carnosine and rutin, the combined use of 6-hydroxynicotinic acid and caffeoyl tartaric acid is superior to the single use of either 6-hydroxynicotinic acid or caffeoyl tartaric acid.

[0109] The embodiments and / or implementation methods described above are merely preferred embodiments and / or implementation methods for implementing the technology of the present invention, and are not intended to limit the implementation methods of the technology of the present invention in any way. Any person skilled in the art can make some modifications or alterations to other equivalent embodiments without departing from the scope of the technical means disclosed in the content of the present invention, but they should still be regarded as the technology or embodiments that are substantially the same as the present invention.

[0110] This document uses specific examples to illustrate the principles and implementation methods of this application. The descriptions of the above embodiments are only for the purpose of helping to understand the methods and core ideas of this application. The above descriptions are only preferred embodiments of this application. It should be noted that due to the limitations of written expression, while there are objectively infinite specific structures, those skilled in the art can make several improvements, modifications, or changes without departing from the principles of this application, and can also combine the above technical features in an appropriate manner. These improvements, modifications, changes, or combinations, or the direct application of the inventive concept and technical solution to other situations without modification, should all be considered within the scope of protection of this application.

Claims

1. A polypeptide compound composition comprising carnosine and rutin, wherein the mass ratio of carnosine to rutin is 1-10:1-10; The polypeptide compound composition further includes 6-hydroxynicotinic acid, and the mass ratio of carnosine to 6-hydroxynicotinic acid is 1-10:0.01-0.2; The polypeptide compound composition also contains caffeoyl tartaric acid, and the mass ratio of carnosine to caffeoyl tartaric acid is 1-10:0.01-0.

1.

2. The polypeptide compound composition according to claim 1, characterized in that, The polypeptide compound composition also includes a penetration enhancer.

3. The polypeptide compound composition according to claim 1, characterized in that, The mass ratio of carnosine to rutin is 3:1; or, the mass ratio of carnosine to rutin is 1:1; or, the mass ratio of carnosine to rutin is 1:3; or, the mass ratio of carnosine to rutin is 2.5:1; or, the mass ratio of carnosine to rutin is 3.5:

1.

4. The polypeptide compound composition according to claim 2, characterized in that, The penetration enhancer includes at least one of azone, NMP, and Span 80.

5. The polypeptide compound composition according to claim 1, characterized in that, The mass ratio of carnosine to 6-hydroxynicotinic acid is 3:0.01-0.

2.

6. A cosmetic product comprising the polypeptide complex composition according to any one of claims 1-5.

7. The cosmetic product according to claim 6, characterized in that, The cosmetics also include water.

8. The cosmetic product according to claim 6, characterized in that, The concentration of carnosine is 0.05-0.5 mg / mL.

9. Use of the polypeptide complex composition of claim 1 in the preparation of cosmetics and / or skin repair products and / or topical medicines.

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