Anti-aging eggshell membrane peptide, solid-phase synthesis method and application thereof, anti-aging eggshell membrane peptide composition and preparation method thereof
By screening anti-aging peptides from eggshell membranes and preparing peptides with the amino acid sequence LGPVGYPKLR using solid-phase synthesis, the application challenges of eggshell membrane peptides in anti-aging products have been solved, realizing the industrialization and improved efficacy of anti-aging products.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- 四川牧舟科技有限公司
- Filing Date
- 2026-03-10
- Publication Date
- 2026-06-05
AI Technical Summary
There is limited research in the current technology on which polypeptide components in eggshell membrane peptides have good anti-aging effects, making it difficult to realize their large-scale application in anti-aging products.
By screening out peptide components with anti-aging effects from eggshell membranes, a peptide with the amino acid sequence LGPVGYPKLR was synthesized using solid-phase synthesis, including steps such as amino acid fixation, linkage, cleavage, precipitation and purification, to prepare an anti-aging eggshell membrane peptide composition, which was then applied to anti-aging products.
The industrialization of eggshell membrane peptides for anti-aging has been achieved, significantly improving the effectiveness of anti-aging products, including enhancing skin hydration, improving skin structure, and enhancing antioxidant capacity.
Smart Images
Figure CN122145564A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of eggshell membrane peptides, and more specifically, to an anti-aging eggshell membrane peptide, a solid-phase synthesis method thereof and its application, an anti-aging eggshell membrane peptide composition thereof and its preparation method. Background Technology
[0002] The eggshell membrane is a thin film located between the eggshell and the egg white, mainly composed of proteins and containing small amounts of polysaccharides, lipids, and minerals. Studies have identified nearly 500 proteins in the eggshell membrane, which endow it with anti-inflammatory, antibacterial, and antioxidant biological activities. There is an urgent need to identify the active ingredients in the eggshell membrane and their corresponding effects in order to mass-produce and apply them to products with corresponding benefits. Currently, there is limited research on which polypeptide components in the eggshell membrane have good anti-aging effects. Summary of the Invention
[0003] The purpose of this invention is to provide an anti-aging eggshell membrane peptide, a solid-phase synthesis method and its application, an anti-aging eggshell membrane peptide composition and its preparation method, and to propose a new polypeptide with anti-aging effect in eggshell membrane hydrolysate.
[0004] The embodiments of the present invention are achieved through the following technical solutions:
[0005] An anti-aging eggshell membrane peptide composition comprising: a polypeptide with an amino acid sequence as shown in SEQ ID NO.1.
[0006] The amino acid sequence is LGPVGYPKLR, which stands for Leu–Gly–Pro–Val–Gly–Tyr–Pro–Lys–Leu–Arg.
[0007] A solid-phase synthesis method for the anti-aging eggshell membrane peptide includes the following steps: using the amino acid sequence: Leu–Gly–Pro–Val–Gly–Tyr–Pro–Lys–Leu–Arg as a standard, Arg is fixed as the first amino acid on a solid-phase support, and the remaining amino acids are sequentially linked. Finally, the product peptide is obtained through a subsequent process. The subsequent process includes: a cleavage process, a precipitation process, and a purification process.
[0008] Preferably, the specific steps for immobilizing amino acids on a solid support include: after the amino acid protected by FMOC is activated by HATU or HBTU, it is linked to the previous amino acid or solid support by a condensing agent, and then after elution protection and washing, the next amino acid is linked; the condensing agent includes N,N-diisopropylethylamine.
[0009] The preceding amino acid refers to the amino acid that has completed the previous condensation.
[0010] Preferably, the deprotection process uses a 15%-25% piperidine / N,N-dimethylformamide solution, and the deprotection time is 25-35 min; the amount of piperidine / N,N-dimethylformamide solution used is 10-20 mL / g, based on the weight of the solid support.
[0011] Preferably, the subsequent process includes: adding the polypeptide resin obtained after amino acid synthesis to a pyrolysis agent, reacting at 25-35℃ for 2-3 hours, then filtering, adding the precipitate to diethyl ether at 0-4℃, letting it stand for 12-18 hours, centrifuging and filtering to obtain the precipitate, drying the precipitate to obtain the crude product; the crude product is then purified by reversed-phase high-performance liquid chromatography to obtain the product polypeptide.
[0012] The pyrolysis agent comprises, by volume, 2-3 parts triisopropylsilyl, 90-95 parts trifluoroacetic acid and 2-3 parts water; the amount of the pyrolysis agent used is 8-12 ml / g based on the weight of the polypeptide resin.
[0013] A method for preparing an anti-aging eggshell membrane peptide composition includes the following steps:
[0014] S100. Add eggshell membrane powder to pure water at a ratio of 1:(20-40) and adjust the pH to 3.5-4.5. Then add pepsin and hydrolyze for 2-5 hours at a temperature of 35-38°C to obtain the first mixture.
[0015] S200, after adjusting the pH of the first mixture to 8-9, add alkaline protease, and hydrolyze for 6-10 hours at a temperature of 36-38℃ to obtain the second mixture;
[0016] S300 and the second mixture are sequentially passed through an enzyme inactivation process and a centrifugation process to obtain a supernatant;
[0017] S400, the supernatant is passed through an ultrafiltration process to obtain an anti-aging eggshell membrane peptide composition;
[0018] The anti-aging eggshell membrane peptide composition includes: the anti-aging eggshell membrane peptide.
[0019] Preferably, the ultrafiltration process uses a 1kDa ultrafiltration membrane; the enzyme inactivation process is carried out at a temperature of 85-95℃ for 8-12 minutes; and the centrifugation process is carried out at a speed of 5000-6000 ×g for 10-20 minutes.
[0020] Preferably, the amount of pepsin in S100 is 3000-6000 U / g; and the amount of alkaline protease in S300 is 5000-10000 U / g.
[0021] An anti-aging eggshell membrane peptide composition prepared by the aforementioned method.
[0022] An application of the aforementioned anti-aging eggshell membrane peptide in the preparation of anti-aging products.
[0023] The present invention has at least the following beneficial effects:
[0024] This invention screens eggshell membrane peptides with good anti-aging effects from eggshell membrane hydrolysis products and synthesizes polypeptides with corresponding amino acid sequences using solid-phase synthesis. This makes it easy to industrialize eggshell membrane peptides for anti-aging purposes, so that eggshell membrane peptides can be used in anti-aging products. Attached Figure Description
[0025] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present invention and should not be regarded as a limitation on the scope. For those skilled in the art, other related drawings can be obtained based on these drawings without creative effort.
[0026] Figure 1 This is the mass spectrum of the polypeptide;
[0027] Figure 2 This is a graph showing the test results for skin moisture content.
[0028] Figure 3 Microscopic images of HE-stained sections;
[0029] Figure 4 Image showing the results of mouse epidermal thickness measurement;
[0030] Figure 5 Image showing the results of dermal thickness measurement in mice;
[0031] Figure 6 Microscopic images of Masson-stained sections;
[0032] Figure 7 The graph shows the results of MMP-1 expression level measurement.
[0033] Figure 8 The graph shows the results of MMP-3 expression level measurement.
[0034] Figure 9 The image shows the test results of the endogenous component Hyp in mouse skin;
[0035] Figure 10 The graph shows the test results for type I collagen content;
[0036] Figure 11 The graph shows the test results for type III collagen content;
[0037] Figure 12The graph shows the test results of relative gene expression levels of MMP-1.
[0038] Figure 13 The graph shows the test results of relative gene expression levels of MMP-3;
[0039] Figure 14 The graph shows the results of the mRNA gene expression test for type I collagen precursor;
[0040] Figure 15 The graph shows the results of the mRNA gene expression test for type III collagen precursor. Detailed Implementation
[0041] To make the objectives, methods, and advantages of the embodiments of the present invention clearer, the methods in the embodiments of the present invention will be clearly and completely described. Obviously, the described embodiments are some embodiments of the present invention, but not all embodiments.
[0042] Example 1: An anti-aging eggshell membrane peptide composition comprising: a polypeptide with an amino acid sequence as shown in SEQ ID NO.1.
[0043] The amino acid sequence is LGPVGYPKLR, which stands for Leu–Gly–Pro–Val–Gly–Tyr–Pro–Lys–Leu–Arg.
[0044] Anti-aging eggshell membrane peptide compositions refer to peptides that, in addition to those containing amino acid sequences as shown in SEQ ID NO.1, may also contain other peptides.
[0045] Example 2: A method for preparing an anti-aging eggshell membrane peptide composition, comprising the following steps:
[0046] S100. Add eggshell membrane powder to pure water at a ratio of 1:(20-40) and adjust the pH to 3.5-4.5. Then add pepsin and hydrolyze for 2-5 hours at a temperature of 35-38°C to obtain the first mixture.
[0047] S200, after adjusting the pH of the first mixture to 8-9, add alkaline protease, and hydrolyze for 6-10 hours at a temperature of 36-38℃ to obtain the second mixture;
[0048] S300 and the second mixture are sequentially passed through an enzyme inactivation process and a centrifugation process to obtain a supernatant;
[0049] S400 and the supernatant are passed through an ultrafiltration process to obtain an anti-aging eggshell membrane peptide composition.
[0050] The ultrafiltration process uses a 1kDa ultrafiltration membrane; the enzyme inactivation process is carried out at a temperature of 85-95℃ for 8-12 minutes; the centrifugation process is carried out at a speed of 5000-6000 ×g for 10-20 minutes.
[0051] The amount of pepsin used in S100 is 3000-6000 U / g; the amount of alkaline protease used in S300 is 5000-10000 U / g.
[0052] Example 3: After screening the polypeptide with the best anti-aging effect from the anti-aging eggshell membrane peptide composition obtained in step S400, the product polypeptide with the corresponding sequence is obtained by solid-phase synthesis according to the amino acid sequence of the polypeptide; the anti-aging eggshell membrane peptide composition includes: product polypeptide.
[0053] In practice, the screening process also involves solid-phase synthesis. After identifying the amino acid sequence of the polypeptide in the anti-aging eggshell membrane peptide composition by mass spectrometry, the polypeptide with the corresponding amino acid sequence is synthesized by solid-phase synthesis. Then, anti-aging tests are conducted on each polypeptide to obtain polypeptides with good anti-aging effects. Based on their corresponding amino acid sequences, solid-phase synthesis can be used for industrial production.
[0054] Example 4: A method for synthesizing the anti-aging eggshell membrane peptide composition, comprising: a solid-phase synthesis method of a polypeptide with an amino acid sequence as shown in SEQ ID NO.1;
[0055] The solid-phase synthesis method includes the following steps: using the amino acid sequence: Leu–Gly–Pro–Val–Gly–Tyr–Pro–Lys–Leu–Arg as the standard, Arg is fixed as the first amino acid on a solid-phase support, and the remaining amino acids are sequentially linked. Finally, the product peptide is obtained through a subsequent process. The subsequent process includes: a cutting process, a precipitation process, and a purification process.
[0056] The specific steps for immobilizing amino acids on a solid support include: after the amino acid protected by FMOC is activated by HATU or HBTU, it is linked to the previous amino acid or solid support by a condensing agent, and then the next amino acid is linked after elution protection and washing; the condensing agent includes N,N-diisopropylethylamine.
[0057] The deprotection process uses a 15%-25% piperidine / N,N-dimethylformamide solution, and the deprotection time is 25-35 min. Based on the weight of the solid support, the amount of piperidine / N,N-dimethylformamide solution used is 10-20 mL / g.
[0058] The subsequent process includes: adding the polypeptide resin obtained after amino acid synthesis to a pyrolysis agent, reacting at 25-35℃ for 2-3 hours, then filtering, adding the precipitate to diethyl ether at 0-4℃, letting it stand for 12-18 hours, centrifuging and filtering to obtain the precipitate, drying the precipitate to obtain the crude product; the crude product is then purified by reversed-phase high-performance liquid chromatography to obtain the product polypeptide.
[0059] The pyrolysis agent comprises, by volume, 2-3 parts triisopropylsilyl, 90-95 parts trifluoroacetic acid and 2-3 parts water; the amount of the pyrolysis agent used is 8-12 ml / g based on the weight of the polypeptide resin.
[0060] Example 5: Application of the aforementioned anti-aging eggshell membrane peptide composition in the preparation of anti-aging products.
[0061] In practice, anti-aging products may include creams, lotions, masks, and injections, etc., and this invention does not impose any specific limitations.
[0062] Effect verification experiment:
[0063] Solid-phase synthesis of peptides was performed using the amino acid sequence: Leu–Gly–Pro–Val–Gly–Tyr–Pro–Lys–Leu–Arg as a standard. The specific steps are as follows:
[0064] Add 1g of solid support: Rink Amide MBHA amino resin to the peptide synthesis tube, and swell with 15mL of dichloromethane for 30min. After removing the solvent, remove the Fmoc protecting agent with a 20% piperidine / N,N-dimethylformamide solution for 20min. The amount of piperidine / N,N-dimethylformamide solution used is 15mL / g based on the weight of the solid support. After the reaction, dry the solid support with nitrogen and wash with N,N-dimethylformamide. Then begin the amino acid condensation process:
[0065] (1) Add 4.0 eq FMOC-protected Arg, 80. eq N,N-diisopropylethylamine and 4.0 eq HBTU, react at 25 °C for 2 h, add 20% piperidine / N,N-dimethylformamide solution, continue reaction for 30 min, and finally wash 4 times with N,N-dimethylformamide. The amount of piperidine / N,N-dimethylformamide solution used is 15 mL / g, based on the weight of the solid support.
[0066] (2) Refer to step (1) and sequentially connect the remaining amino acids to obtain polypeptide resin.
[0067] (3) Add polypeptide resin and cleavage agent to the reaction tube, react at 30°C for 2.5 h, then filter, add ether at 0°C to the precipitate, let stand for 16 h, centrifuge and filter to obtain the precipitate, and dry the precipitate to obtain the crude product.
[0068] The pyrolysis agent comprises, by volume, 2.5 parts triisopropylsilyl, 95 parts trifluoroacetic acid and 2.5 parts water; the amount of the pyrolysis agent used is 8-12 ml / g based on the weight of the polypeptide resin.
[0069] (4) The crude product was purified by reversed-phase high-performance liquid chromatography to obtain the product polypeptide, as follows:
[0070] A 250 mm × 4.6 mm standard column was used. 20 mg of crude product was dissolved in mobile phase A. Mobile phase A was an aqueous solution containing 0.1% TFA, and mobile phase B was an acetonitrile solution containing 0.1% TFA. Gradient elution was performed, from 10% B to 60% B within 30 min, at a flow rate of 1 mL / min.
[0071] Mass spectrometry identification, such as Figure 1 As shown.
[0072] In vitro antioxidant capacity of the product peptides was determined: the total antioxidant capacity, DPPH free radical scavenging capacity, and ABTS free radical scavenging capacity of the peptides were measured. The total antioxidant capacity kit, DPPH free radical scavenging capacity kit, and ABTS free radical scavenging capacity kit were used for the determination. The experimental results are shown in Table 1.
[0073] Table 1
[0074]
[0075] Mouse smear test: Sixty female mice (20±2 g) were selected and acclimatized for 3-7 days. After ear tagging, they were randomly divided into 6 groups (blank group, model group, positive group, low-dose product peptide group, medium-dose product peptide group, and high-dose product peptide group). The normal group was injected with an equal volume of physiological saline, while the other five groups received continuous subcutaneous injections of D-galactose (D-GAL) into the neck and back to establish a mouse aging model. The model group underwent UVB irradiation combined with subcutaneous D-GAL injection, and the smear matrix was epidermal growth factor gel. The positive group underwent UVB irradiation combined with subcutaneous D-GAL injection, and the smear matrix was mixed with vitamin E. The eggshell membrane peptide group underwent UVB irradiation combined with subcutaneous D-GAL injection, and the smear matrix was mixed with product peptide. Four hours after modeling, the matrix, positive control drug, and eggshell membrane peptide were applied to the backs of mice according to their groups. 100 μL per mouse was applied evenly to the bare skin on the back of the mouse once daily for three weeks. The UVB irradiation dose was one MED in the first week, increasing by 0.5 MED each subsequent week. After the three-week experiment, the mice were euthanized, and the skin tissue from their backs was collected for subsequent testing. The treatment methods for each group in the application test are shown in Table 2.
[0076] Table 2
[0077]
[0078] After feeding, the mice underwent hair removal on their backs. Blood was collected from the eyeballs, and skin, liver, kidneys, and muscle from the back of the neck were harvested and weighed. After standing the blood for 15 minutes, the serum was separated by centrifugation at 3500 rpm for 10 minutes and stored at -80°C for analysis. 0.1 g of the same sample from the skin, liver, kidneys, and muscle was added to 0.9 mL of physiological saline, homogenized, centrifuged at 4000 rpm for 10 minutes, and the supernatant was separated. BCA protein quantification was performed according to the manufacturer's instructions, and the samples were stored at -80°C for analysis.
[0079] Skin moisture content determination: Skin from the back of mice was collected, cleaned after hair removal, and accurately weighed for wet weight. The skin was then placed in an oven and dried at 80℃ for 12 hours, and its dry weight was measured. The difference between the wet and dry weights, and the ratio of the wet weight to the dry weight, represents the skin moisture percentage. The test results are shown in [Figure number missing]. Figure 2 .
[0080] HE and Masson staining: Skin tissue was fixed with 4% paraformaldehyde for more than 48 hours, rinsed with running water for 30 minutes, and then the tissue was trimmed into blocks and placed in a pathological embedding plastic basket for dehydration (75% ethanol for 6 hours, 85% ethanol for 10 hours, 95% ethanol for 4 hours, anhydrous ethanol for 2 hours, anhydrous ethanol for 2 hours), cleared (xylene for 20 minutes, xylene for 15 minutes), immersed in paraffin for 3 hours, and then embedded in paraffin.
[0081] The tissue was cut into 5µm thick sections, flattened in warm water, and placed on glass slides. The sections were then baked at 60℃ for 2 hours. After dewaxing with xylene, the sections were washed with running water for 20 minutes and then stained.
[0082] HE staining: stain with hematoxylin for 30 min, wash with running water for 20 min, differentiate with hydrochloric acid and alcohol, stain with eosin for 5 min, finally dehydrate with graded alcohol, clear with xylene and mount with resin.
[0083] Masson staining: Hematoxylin staining for 2-5 min, wash with water, differentiate with hydrochloric acid and alcohol, blue with ammonia, and rinse with running water; 1% phosphomolybdic acid staining for 1-2 min, observe under a microscope, and stop when muscle fibers appear red and collagen fibers appear light red; stain directly in aniline blue staining solution for 2-5 min; wash quickly with water, dry in a 60℃ oven, clear with xylene, and seal with neutral resin.
[0084] HE-stained sections were photographed under a 40× objective lens and a 10× eyepiece, and under a 4× objective lens and a 10× eyepiece. The thickness of the epidermis and dermis in the images was measured using ImageProPlus 6.0. See the microscopic images below. Figure 3 As shown, the measurement results are as follows. Figures 4-5 As shown, Figure 4This is a diagram showing the results of mouse epidermal thickness measurements. Figure 5 This is a graph showing the results of dermal thickness measurement in mice.
[0085] After Masson staining, collagen fibers appear blue, and muscle fibers appear red. After capturing images using a microscopic imaging system, ImageProPlus 6.0 measures the area of collagen fibers in the images. See the microscopic images below. Figure 6 As shown in Table 3, the area measurement results are shown in Table 3.
[0086] Table 3
[0087]
[0088] Skin and serum antioxidant index detection: The levels of CAT, ROS, GSH-Px, T-AOC, SOD and MDA in the serum of mice in each group were detected according to the kit instructions. The test results are shown in Tables 4, 5, 6 and 7.
[0089] Table 4: Serum antioxidant indicators of each group
[0090]
[0091] Table 5: Serum antioxidant enzyme activities in each group
[0092]
[0093] Table 6: Skin antioxidant enzyme indices for each group
[0094]
[0095] Table 7: Skin antioxidant enzyme activity in each group
[0096]
[0097] ELISA was used to detect the expression levels of skin collagen degradation markers: Approximately 0.1 g of back skin was added to 0.9 mL of pre-cooled PBS and thoroughly homogenized in a homogenizer to obtain a 10% skin tissue homogenate. The homogenate was centrifuged at 4000 r / min and 4℃ for 10 min. The supernatant was collected, and the expression levels of mouse skin collagen degradation markers MMP-1 and MMP-3 were determined according to the kit instructions. The results are shown below. Figures 7-8 .
[0098] ELISA was used to detect the expression levels of endogenous skin components: Approximately 0.1 g of back skin was added to 0.9 mL of pre-cooled PBS and thoroughly homogenized in a homogenizer to obtain a 10% skin tissue homogenate. The homogenate was centrifuged at 4000 r / min and 4℃ for 10 min. The supernatant was collected, and the contents of endogenous mouse skin components Hyp, type I collagen, and type III collagen were determined according to the kit instructions. The test results are shown below. Figures 9-11 .
[0099] Total RNA extraction and real-time quantitative PCR analysis: Total RNA was extracted from skin using the Trizol method. RNA purity and concentration were determined. The reverse transcriptase kit was strictly followed according to the instructions to reverse transcribe RNA into cDNA. The cDNA, primers, and SYBR Green Master Mix were mixed thoroughly, and real-time quantitative PCR was used for amplification, with fluorescence intensity detected. The amplification program was as follows: 95℃ for 5 min, 1 cycle; 95℃ for 20 s; 60℃ for 30 s, 72℃ for 20 s, 45 cycles; terminal extension at 72℃ for 2 min. Results are shown in [Figure number missing]. Figures 12-15 .
[0100] The above are merely preferred embodiments of the present invention and are 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. An anti-aging eggshell membrane peptide, characterized in that, The amino acid sequence is shown in SEQ ID NO.
1.
2. A solid-phase synthesis method for the anti-aging eggshell membrane peptide according to claim 1, characterized in that, The process includes the following steps: using the amino acid sequence: Leu–Gly–Pro–Val–Gly–Tyr–Pro–Lys–Leu–Arg as the standard, Arg is immobilized on a solid-phase support as the first amino acid, and the remaining amino acids are sequentially linked. Finally, the product peptide is obtained through a subsequent process. The subsequent process includes: a cleavage process, a precipitation process, and a purification process.
3. The solid-phase synthesis method according to claim 2, characterized in that, The specific steps for immobilizing amino acids on a solid support include: after the amino acid protected by FMOC is activated by HATU or HBTU, it is linked to the previous amino acid or solid support by a condensing agent, and then the next amino acid is linked after elution protection and washing; the condensing agent includes N,N-diisopropylethylamine.
4. The solid-phase synthesis method according to claim 3, characterized in that, The deprotection process uses a 15%-25% piperidine / N,N-dimethylformamide solution, and the deprotection time is 25-35 min. Based on the weight of the solid support, the amount of piperidine / N,N-dimethylformamide solution used is 10-20 mL / g.
5. The solid-phase synthesis method according to claim 2, characterized in that, The subsequent process includes: adding the polypeptide resin obtained after amino acid synthesis to a pyrolysis agent, reacting at 25-35℃ for 2-3 hours, then filtering, adding the precipitate to diethyl ether at 0-4℃, letting it stand for 12-18 hours, centrifuging and filtering to obtain the precipitate, drying the precipitate to obtain the crude product; the crude product is then purified by reversed-phase high-performance liquid chromatography to obtain the product polypeptide. The pyrolysis agent comprises, by volume, 2-3 parts triisopropylsilyl, 90-95 parts trifluoroacetic acid and 2-3 parts water; the amount of the pyrolysis agent used is 8-12 ml / g based on the weight of the polypeptide resin.
6. A method for preparing an anti-aging eggshell membrane peptide composition, characterized in that, Includes the following steps: S100. Add eggshell membrane powder to pure water at a ratio of 1:(20-40) and adjust the pH to 3.5-4.
5. Then add pepsin and hydrolyze for 2-5 hours at a temperature of 35-38°C to obtain the first mixture. S200, after adjusting the pH of the first mixture to 8-9, add alkaline protease, and hydrolyze for 6-10 hours at a temperature of 36-38℃ to obtain the second mixture; S300 and the second mixture are sequentially passed through an enzyme inactivation process and a centrifugation process to obtain a supernatant; S400, the supernatant is passed through an ultrafiltration process to obtain an anti-aging eggshell membrane peptide composition; The anti-aging eggshell membrane peptide composition comprises: the anti-aging eggshell membrane peptide according to claim 1.
7. The preparation method according to claim 6, characterized in that, The ultrafiltration process uses a 1kDa ultrafiltration membrane; the enzyme inactivation process is carried out at a temperature of 85-95℃ for 8-12 minutes; the centrifugation process is carried out at a speed of 5000-6000 ×g for 10-20 minutes.
8. The preparation method according to claim 6, characterized in that, The amount of pepsin used in S100 is 3000-6000 U / g; the amount of alkaline protease used in S200 is 5000-10000 U / g.
9. An anti-aging eggshell membrane peptide composition prepared by the method according to any one of claims 6-8.
10. The application of the anti-aging eggshell membrane peptide of claim 1 in the preparation of anti-aging products.