CN112807417B - Collagen-containing skin-whitening and anti-wrinkle medicine or cosmetic - Google Patents
Collagen-containing skin-whitening and anti-wrinkle medicine or cosmetic Download PDFInfo
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- CN112807417B CN112807417B CN202110377047.XA CN202110377047A CN112807417B CN 112807417 B CN112807417 B CN 112807417B CN 202110377047 A CN202110377047 A CN 202110377047A CN 112807417 B CN112807417 B CN 112807417B
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- collagen
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- fish skin
- adipose
- stem cells
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Images
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Abstract
The invention relates to a collagen-containing medicament or cosmetic for whitening and resisting wrinkles. The invention prepares a novel collagen peptide which has better free radical scavenging capacity. Meanwhile, the polypeptide is used for culturing the adipose-derived stem cells, so that the aging of the adipose-derived stem cells can be effectively prevented, the proliferation of the adipose-derived stem cells can be promoted, the collagen peptide and the adipose-derived stem cells cultured by the polypeptide are used together, the aging can be effectively prevented, the skin can be effectively whitened, and the polypeptide-containing adipose-derived stem cell culture medium has a good application value.
Description
Technical Field
The application relates to the field of medicines or cosmetics, in particular to a collagen-containing medicine or cosmetic for whitening and resisting wrinkles.
Background
ADSCs have become ideal tools in cell therapy and tissue engineering applications due to the advantages of wide sources, convenient material acquisition, small tissue damage, rapid amplification and the like. According to a plurality of researches, the ADSCs can play a more remarkable role in the aspects of in-vivo injury repair, aging resistance and the like; the biological effect is realized partially through the multidirectional differentiation capability, the ADSCs can be differentiated into various cells such as fat, bones, cartilages, muscles, tendons and the like, and aged and defected tissue cells are supplemented and replaced; the other part of the cells has paracrine function, so that the ADSCs can synthesize and secrete active ingredients such as various cytokines, polypeptides, gas molecules and the like, and the vitality and the anti-apoptosis capacity of damaged cells are improved. The ADSCs have remarkable effects in inhibiting skin aging, whitening skin, assisting fat transplantation, promoting hair regeneration and the like in various anti-aging aspects by secreting a series of active ingredients.
A plurality of experiments prove that various cytokines secreted by the ADSCs have the effects of enhancing cell proliferation and migration capacity, promoting angiogenesis, resisting apoptosis and the like, promoting the tissue function to recover to be normal, reducing damage and having a remarkable application effect in the anti-aging field. Researches show that insulin-like growth factors, epidermal growth factors, transforming growth factors, interleukins and tumor necrosis factors secreted by the ADSCs can improve skin fibroblast damage caused by medium-wave ultraviolet rays, and enhance the proliferation of fibroblasts and the capability of synthesizing collagen, wherein the transforming growth factors have the strongest effect of stimulating the fibroblasts in vitro experiments. The ADSCs are injected into a photoaging nude mouse model, the thickness of the dermis layer and the number of fibroblasts at the position where the ADSCs are injected are obviously increased, the collagen synthesis of the fibroblasts is increased, the angiogenesis of the dermis is increased, and the skin aging appearance caused by the irradiation of ultraviolet rays is obviously improved. The ADSCs conditioned medium is applied to the facial skin of a patient to remove wrinkles, and significant improvement in skin texture, increase in dermis thickness and wrinkle reduction are observed 2 months after injection treatment. Patients who are subjected to facial rejuvenation treatment by the combination of the ADSCs injection after the operation are followed, and the skin of the patients who are subjected to the combination treatment is found to be better than that of the patients who are subjected to the simple operation treatment.
Collagen is a common cosmetic raw material, and the application of collagen is rapidly developed in recent years. It can moisten skin, give smooth feeling, and has good conditioning effect on hair. Research shows that the skin care product has excellent effect of resisting various kinds of radiation and can form excellent water maintaining layer to provide skin with required water completely. The collagen can enter into the deep layer of skin, and supply essential nutrients (amino acids) to skin, maintain the stability and integrity of collagen fiber structure, enhance the activity of collagen in skin, improve the living environment of skin cells, promote metabolism of tissue, and achieve the purposes of nourishing, moistening, caring skin and nourishing hair. The similarity of the collagen and the structure of the skin stratum corneum determines the good compatibility, the good affinity and the good permeability of the collagen and the skin, the collagen can permeate into the epidermis layer of the skin and be fully absorbed by the skin, and an extremely thin film layer is formed on the surface of the skin, so that the skin is plump, wrinkles are stretched, the density of the skin is improved, the tension is generated, and the collagen has the anti-wrinkle effect. Tyrosine residues in the collagen compete with tyrosine in the skin and are combined with the active center of tyrosinase, so that the tyrosine in the skin is inhibited from being catalyzed and converted into dopa by the tyrosinase, the formation of melanin in the skin is prevented, and the whitening effect is achieved.
Currently, there are few cases where stem cells are used together with collagen for skin whitening or treatment, and corresponding improvements are also less studied.
Disclosure of Invention
The invention overcomes the defects of the prior art and provides a cosmetic or a medicine with the functions of whitening, water retention and wrinkle resistance.
Specifically, the invention provides a collagen peptide, which is prepared by the following method:
the snakehead skin was washed with tap water and cut into small pieces with a knife. The fish skin is soaked in NaOH solution. The alkali-treated fish skin was washed to neutrality with tap water, then soaked with HCl (v/v) for 1h, and then rinsed to neutrality with tap water. Sterile water was added to the cleaned fish skin and the mixture was incubated at a constant temperature. Centrifuging at 8000r/min for 20min to obtain supernatant as collagen, and freeze drying. Performing enzymolysis on fish skin collagen with compound protease (papain, Guangxi Pombo biological preparation Co., Ltd., enzyme activity of 2900U/mg; trypsin, Shanghai Nanning Biochemical reagent Co., Ltd., enzyme activity of 3600U/mg; alkaline protease, Shandong Long great biological engineering Co., Ltd., enzyme activity of 2000U/mg), performing enzymolysis at pH8.0 and 40 deg.C, heating in boiling water to inactivate enzyme after enzymolysis, cooling to room temperature, centrifuging the enzymolysis liquid, concentrating the supernatant, and freeze-drying to obtain fish skin collagen peptide.
Further, the polypeptide capable of effectively promoting the proliferation of the adipose-derived mesenchymal stem cells and preventing the aging of the stem cells is obtained through early stage identification, separation and screening, and the sequence of the polypeptide is shown as SEQ ID NO:1 is shown.
Further, the present invention provides a pharmaceutical composition or cosmetic composition for delaying aging and whitening skin, the composition comprising fish skin collagen peptide and a peptide prepared from SEQ ID NO: 1.
In an embodiment, the composition or combination includes a pharmaceutically acceptable carrier. In embodiments, the pharmaceutically acceptable carrier comprises an emulsion. In embodiments, the emulsion is an oil-in-water emulsion or a water-in-oil emulsion. In embodiments, the combination or combination comprises or is in the form of a cream (cream), lotion (deposition), suspension or aqueous solution.
In embodiments, the composition is formulated for topical application to the skin (i.e., the composition is a topical composition). In an embodiment, the composition is a pharmaceutical composition.
The composition further comprises a pharmaceutically acceptable carrier.
In embodiments, the pharmaceutical composition is formulated for topical application to the skin. In embodiments, the pharmaceutically acceptable carrier comprises an emulsion. In embodiments, the emulsion is an oil-in-water emulsion or a water-in-oil emulsion. In embodiments, the pharmaceutical composition is in the form of a cream, lotion, suspension or aqueous solution. The composition may be for subcutaneous injection or topical application.
The composition can reduce tyrosinase activity to reduce tyrosinase related protein expression, and finally inhibit melanin synthesis, thereby whitening skin. Moreover, the composition can also result in a skin whitening effect with a reduced melanocyte count.
The invention also relates to a therapeutic or cosmetic treatment method comprising the administration of a pharmaceutical or cosmetic composition as a composition for inhibiting melanin synthesis. In one particular mode, the invention also relates to a therapeutic or cosmetic method comprising administering a pharmaceutical or cosmetic composition comprising said stem cells for the treatment of a pigmentary disorder.
Further, the composition may also be selected from all of the following compounds and functional equivalents thereof:
the further compounds may be chosen in particular from active agents conventionally used in dermatology or cosmetics, such as emollients, moisturizing active agents, keratin synthesis activators, keratin regulators, keratin exfoliants, agents for reestablishing the skin barrier (sebum synthesis activators), peroxisome proliferator-activated receptor (PPAR) agonists, RXR or LXR agonists, sebum regulators, anti-irritants, soothing agents, anti-inflammatory agents, antioxidants and anti-ageing agents, depigmenting or hypodepigmenting agents, colorants, lipolytic or lipogenesis inhibitors or anti-cellulite or reducing agents, inorganic or organic filters and sunscreens, antifungal compounds, antiseptics, antibacterial agents, prebiotics and probiotics, antibiotics, immunomodulators.
More particularly, the agents for healing and/or reestablishing the skin barrier that can be used in the combination are advantageously panthenol (vitamin B5), arabinogalactans, zinc oxide, ceramides, cholesterol, squalane and phospholipids.
The sebum regulators that can be used in the combination are advantageously chosen from 5-alpha-reductase inhibitors. Zinc (and zinc derivatives such as its gluconate, salicylate and pyroglutamate) and spironolactone also have sebum suppressive activity. Other sebum regulators of lipid origin acting on sebum quality, such as linoleic acid, are also of interest.
The anti-inflammatory and/or anti-irritant and/or soothing agent may be arabinogalactan.
The sunscreen active agents which can be used in combination are advantageously UVB and/or UVA filters and sunscreens, for example inorganic and/or organic sunscreens or filters known to the person skilled in the art, whose choice and concentration can be adjusted by the person skilled in the art according to the degree of protection required.
Preservatives that can be used in combination are, for example, those customarily used in cosmetics, molecules having antibacterial activity (pseudo-preservatives) such as octanoic acid derivatives, such as, for example, octanoyl glycine and caprylin; hexanediol, sodium levulinate, and copper and zinc derivatives (gluconate and PCA).
Advantageous effects
The invention prepares a novel collagen peptide which has better free radical scavenging capacity. Meanwhile, the polypeptide is used for culturing the adipose-derived stem cells, so that the aging of the adipose-derived stem cells can be effectively prevented, the proliferation of the adipose-derived stem cells can be promoted, the collagen peptide and the adipose-derived stem cells cultured by the polypeptide are used together, the aging of mice can be effectively prevented, the skin can be whitened, and the polypeptide-containing adipose-derived stem cell whitening composition has a good application value.
Drawings
FIG. 1 is a graph showing the effect of enzyme addition on collagen yield
FIG. 2 is a graph showing the result of the DPPH radical scavenging rate of fish skin collagen polypeptide
FIG. 3 is a graph showing the aging results of adipose-derived mesenchymal stem cells
Detailed Description
To further illustrate the objects, aspects and advantages of the present invention, we shall now describe the invention with reference to the following specific examples, which are only for better illustrating the patent of the present invention and are not intended to limit the scope of the present invention. All other embodiments that can be obtained by a person skilled in the art without making any inventive step based on the examples of the present invention belong to the protection scope of the present invention.
EXAMPLE 1 preparation of collagen peptide
The snakehead skin was washed with tap water and cut into 0.5cm x 0.5cm pieces with a knife. The fish skin was soaked in 1.0mmol/L NaOH solution for 2h (fish skin: solution ═ 1:10, w/v). The alkali-treated fish skin was washed with tap water to neutrality, then soaked with 0.5% HCl (v/v) for 1h (fish skin: solution ═ 1:10, w/v), and then rinsed with tap water to neutrality. Sterile water was added to the cleaned fish skin and the mixture was incubated at a constant temperature. Centrifuging at 8000r/min for 20min to obtain supernatant as collagen, and freeze drying. Performing enzymolysis on fish skin collagen by using compound protease (papain, Guangxi Pombo biological preparation Co., Ltd., enzyme activity of 2900U/mg, trypsin, Shanghai Nanning Biochemical reagent Co., Ltd., enzyme activity of 3600U/mg, alkaline protease, Shandong Long great biological engineering Co., Ltd., enzyme activity of 2000U/mg), setting the enzyme addition amount to five gradients of 0.5%, 1%, 2%, 3%, 4% and 5%, performing enzymolysis at pH8.0 and 40 ℃ for 4h, heating in boiling water for 15min to inactivate enzyme after enzymolysis, cooling to room temperature, centrifuging the enzymolysis liquid at 8000r/min for 20min, taking supernatant, concentrating, and freeze-drying to obtain the fish skin collagen peptide.
The collagen is decomposed into amino acids after enzymolysis, and is oxidized by chloramine T to form oxide containing pyrrole ring, after the oxidation is stopped by perchloric acid, p-dimethylaminobenzaldehyde is used as a color developing agent to generate red compounds, and the colorimetric determination is carried out at the wavelength of 550 nm. The measured hydroxyproline content is calculated to obtain the collagen content. The results are shown in FIG. 1.
As can be seen from fig. 1, the effect of the enzyme addition amount on the collagen yield is that the yield is not increased all the time with the increase of the ratio, and when the enzyme addition amount is 2%, the enzyme is the optimal ratio for the enzyme to exert the activity, and at this time, the enzyme activity is the highest, and a higher addition amount may play a role in blocking the enzymolysis reaction. Therefore, collagen peptide powder was prepared at 2% enzyme addition for subsequent experiments.
EXAMPLE 2 measurement of DPPH radical scavenging ability
The collagen peptide prepared in example 1 was prepared into sample solutions of various concentrations and 0.1mmol/L ethanol DPPH solution. Accurately sucking 0.4mL of diluted samples to be detected with different concentrations into a 5mL centrifuge tube, adding 2.0mL of DPPH ethanol solution, uniformly mixing, standing in a dark place for 30min, continuously shaking, and measuring the light absorption value at 517 nm. DPPH clearance (%) ((a control- (a sample-a sample control))/a control) × 100 in the formula: control A was 0.4mL deionized water +2.0mL LDPPH solution; sample A control was 0.4mL sample +2.0mL ethanol; the sample A is 0.4mL of the diluted sample to be tested +2.0mL of the LDPPH solution. The results are shown in FIG. 2.
The DPPH method is a fast and feasible method to evaluate the radical scavenging activity of natural antioxidants. As can be seen from FIG. 2, the clearance rate of the fish skin collagen polypeptide prepared by the invention to DPPH free radicals is obviously increased along with the increase of the concentration of the sample, and the clearance rate can reach more than 80% under the concentration of 50 mug/mL, so that the fish skin collagen polypeptide has a better effect.
EXAMPLE 3 preparation of adipose-derived Stem cells
Under the aseptic condition, 50mL of fat normal saline mixture is obtained, blood cells are removed by centrifugation and PBS (phosphate buffer solution) cleaning twice, and fat particles with higher purity are obtained. 0.075% collagenase type I is digested in a constant temperature shaker at 37 ℃ for 60min and 1500r/min, centrifuged for 10min, upper undigested adipose tissues and grease are removed, the precipitate is resuspended in a 200-mesh screen for filtration, centrifuged again, the erythrocyte lysate is lysed for 5min, the erythrocyte is washed twice by phosphate buffer solution, the 10% fetal calf serum high-sugar DMEM is resuspended, and the 5 × 10 fetal calf serum high-sugar DMEM is used for resuspending4Per cm2 inoculation to 10cm2In a culture plate. The cells inoculated at the 1 st passage are designated as P0, after 90% fusion of the cells, 0.25% trypsinization is carried out, the cells are inoculated at the 1: 4 passage, and the 3 rd passage cells are taken for experiment.
The hADSCs at passage 3 were trypsinized to form single cell suspensions. 1X 106Anti-human CD34PE, CD73PE, CD90FITC, CD105PE and CD45FITC monoclonal antibodies are added into the cells respectively, and each 1 mu L of the monoclonal antibodies are negative controls PE-IgG1 and FITC-IgG 1. Incubating at 4 ℃ in dark for 30min, and detecting by a flow cytometer. The results are shown in table 1 below:
TABLE 1 results of phenotypic identification of cells
| Phenotypic characterization of cells | Expression rate |
| CD105 | (96.2±2.6)％ |
| CD90 | (95.8±1.9)％ |
| CD34 | (4.2±1.2)％ |
| CD45 | (3.1±0.80)％ |
As can be seen from Table 1, the tested CD105 expression rate (96.2. + -. 2.6)%, CD90 expression rate (95.8. + -. 1.9)%, CD34 expression rate (4.2. + -. 1.2)%, and CD45 expression rate (3.1. + -. 0.80)%. The adipose-derived stem cells isolated by the invention show obvious biological characteristics of adipose-derived stem cells, and express hematopoietic molecules such as CD90 and CD105 with high expression and CD34 and CD45 with low expression.
Example 4 adipose-derived stem cell anti-aging assay
The adipose-derived stem cells isolated in example 3 were digested to prepare a cell suspension, and the cell density was 1X 105individual/mL inoculation, group treatment: subculturing the control group with DMEM medium containing 10% fetal calf serum for 48 h; and (3) an aging group: culturing for 48h by using a DMEM medium containing 8g/L D-galactose and 10% fetal calf serum; aging rescue group: application of D-galactose 8g/L + SEQ ID NO:1, culturing the anti-aging peptide in a DMEM medium containing 1mg/mL plus 10% fetal calf serum for 48 hours; stimulation group: the use of SEQ ID NO:1, culturing the anti-aging peptide in a DMEM medium containing 1mg/mL plus 10% fetal calf serum for 48 hours; and then, operating according to a beta-galactosidase detection kit, removing the cell culture medium by suction, washing with PBS for 1 time, adding a proper amount of SA-beta-gal staining fixing solution, fixing at room temperature for 15min, removing the cell fixing solution by suction, adding PBS for washing for 3 times, removing the PBS by suction, adding a proper amount of cell staining working solution, incubating overnight at 37 ℃, observing that the aged cells are light blue to dark blue under an optical microscope, and expressing the positive rate of the sample by the percentage of the positive sample in the total standard number. The results are shown in FIG. 3.
As can be seen from fig. 3, the SA- β -gal staining result shows that the adipose-derived mesenchymal stem cells can significantly reduce the senescence rate of adipose-derived mesenchymal stem cells under the action of 1mg/mL of polypeptide, and the mesenchymal stem cell culture medium cultured by the polypeptide has fewer blue-green senescent cells, and has significant difference (P <0.01) compared with the control group, which indicates that the adipose-derived mesenchymal stem cells treated by the polypeptide have significant anti-cell senescence effect.
In addition, according to the cell counting result after the culture, the polypeptide can inhibit cell senescence and promote the proliferation of the stem cells, and compared with the control group, the proliferation effect can reach (23.1 +/-2.5)%.
Example 5 Effect of collagen peptides and adipose-derived Stem cells on Guinea pig tyrosinase and skin melanin
Selecting guinea pigs with different colors, wherein the guinea pigs can be used for male and female purposes and have the weight of about 250 g; guinea pigs were randomly divided into 5 groups, i.e., a normal control group (saline injected subcutaneously), a positive control group: (for 100mg/kg of commercial collagen peptide); protein peptide-treated group (subcutaneous injection of 100mg/kg of collagen peptide prepared in example 1), adipose-derived stem cell-treated group (subcutaneous injection of 2 × 10 stem cells prepared in example 3)9pieces/mL/0.1 mL/kg), collagen peptide + adipose-derived stem cell treated group (subcutaneous injection of 50mg/kg of collagen peptide prepared in example 1 +1 x 10 stem cells prepared in example 3)9one/mL/0.1 mL/kg), the frequency of administration for each group was 3 d/time.
Prior to the experiment, the guinea pigs in each group were cut with an electric razor to remove the dark grey skin bristles for use. After continuous administration for 30 days, after the last administration, anaesthetizing each group of guinea pigs by using 20% urethane, separating common carotid artery for blood sampling, centrifuging at 2500r/min for 10min, and taking plasma to measure the content of tyrosinase; the skin of 2cm × 2cm was taken in the area of shaved hair, fixed with 4% formaldehyde, sectioned by conventional paraffin embedding, stained with HE, and stained with melanin by the ferrous sulfate Lillie method, and 50 follicles were observed at will, and the number of follicles containing melanin was counted. Statistical treatment was performed and differences between groups were compared. The results are shown in Table 2.
TABLE 2 Effect of groups on tyrosinase Activity and skin melanin in guinea pigs
As can be seen from table 1, the positive control group, the collagen peptide treated group, the adipose-derived stem cell treated group, and the collagen peptide + adipose-derived stem cell treated group all resulted in a significant decrease in the tyrosinase activity level in guinea pig plasma, and had a significant difference (. about.p <0.05) compared to the contemporary normal control group, and particularly, the treatment with the collagen peptide and the stem cell together resulted in a significant decrease in the serum tyrosinase activity in guinea pig, and showed a significant synergistic effect. In addition, the positive control group, the collagen peptide treatment group, the fat stem cell treatment group and the collagen peptide + fat stem cell treatment group can cause the number of the cells containing the melanin granules in the hair follicle of the guinea pig skin to be obviously reduced, and compared with the contemporary normal control group, the number of the cells is obviously different (P is less than 0.05). Therefore, the collagen peptide and fat stem cell treatment group has a good whitening effect, and the content of melanin particles in the skin of the guinea pig can be reduced.
The invention has been described in detail with reference to specific embodiments and illustrative examples, but the description is not intended to be construed in a limiting sense. Those skilled in the art will appreciate that various equivalent substitutions, modifications or improvements may be made to the technical solution of the present invention and its embodiments without departing from the spirit and scope of the present invention, which fall within the scope of the present invention. The scope of the invention is defined by the appended claims.
Sequence listing
<110> Beijing Vast Mei Biotechnology Ltd
<120> collagen-containing skin-whitening and anti-wrinkle drug or cosmetic
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 45
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
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Ser Phe His His Met Asn Gln Lys Thr Arg Pro Arg Lys Trp Ile Pro
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Ser Pro Ile Ile Thr Arg Gln Val Thr Ala Trp Gly Leu Pro Gln Lys
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His Pro His Lys Gln Ser Arg Gln Trp Val Pro Trp Met
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Claims (6)
1. An anti-aging drug is characterized by comprising collagen peptide and adipose-derived stem cells with anti-aging effect, wherein the collagen peptide is prepared by the following method:
washing snakehead skin with tap water, cutting into small pieces of 0.5cm × 0.5cm with a knife, and soaking the snakehead skin in 1.0mmol/L NaOH solution for 2 h; washing the fish skin subjected to alkali treatment to be neutral by using tap water, soaking the fish skin by using 0.5% of HCl in percentage by volume for 1 hour, and then rinsing the fish skin by using tap water to be neutral; adding sterile water to the cleaned fish skin and incubating the mixture at a constant temperature; centrifuging at 8000r/min for 20min to obtain supernatant as collagen, and freeze drying; with a complex protease: papain, the enzyme activity of 2900U/mg, trypsin, the enzyme activity of 3600U/mg, alkaline protease, the enzyme activity of 2000U/mg for enzymolysis of fish skin collagen, 2 percent of compound protease is added, the fish skin collagen is subjected to enzymolysis for 4 hours at the temperature of pH8.0 and 40 ℃, the boiled water after the enzymolysis is heated for 15min for enzyme deactivation, the temperature is cooled to the room temperature, the enzymolysis liquid is centrifuged for 20min at 8000r/min, and the supernatant is concentrated and freeze-dried to obtain the fish skin collagen peptide;
the adipose-derived stem cells are prepared by digesting the isolated adipose-derived stem cells to prepare a cell suspension, and the cell density is 1 x 105And (3) inoculating the strain per mL, and subculturing the strain by using a DMEM medium containing 10% fetal bovine serum added with 1mg/mL of the polypeptide of SEQ ID NO: 1.
2. The medicament of claim 1, wherein the medicament comprises a pharmaceutically acceptable carrier.
3. An anti-aging and whitening cosmetic is characterized by containing collagen peptide and adipose-derived stem cells with anti-aging effect, wherein the collagen peptide is prepared by the following method:
washing snakehead skin with tap water, cutting into small pieces of 0.5cm × 0.5cm with a knife, and soaking the snakehead skin in 1.0mmol/L NaOH solution for 2 h; washing the fish skin subjected to alkali treatment to be neutral by using tap water, soaking the fish skin by using 0.5% of HCl in percentage by volume for 1 hour, and then rinsing the fish skin by using tap water to be neutral; adding sterile water to the cleaned fish skin and incubating the mixture at a constant temperature; centrifuging at 8000r/min for 20min to obtain supernatant as collagen, and freeze drying; with a complex protease: papain, the enzyme activity of 2900U/mg, trypsin, the enzyme activity of 3600U/mg, alkaline protease, the enzyme activity of 2000U/mg for enzymolysis of fish skin collagen, 2 percent of compound protease is added, the fish skin collagen is subjected to enzymolysis for 4 hours at the temperature of pH8.0 and 40 ℃, the boiled water after the enzymolysis is heated for 15min for enzyme deactivation, the temperature is cooled to the room temperature, the enzymolysis liquid is centrifuged for 20min at 8000r/min, and the supernatant is concentrated and freeze-dried to obtain the fish skin collagen peptide;
the adipose-derived stem cells are prepared by digesting the isolated adipose-derived stem cells to prepare a cell suspension, and the cell density is 1 x 105And (3) inoculating the strain per mL, and subculturing the strain by using a DMEM medium containing 10% fetal bovine serum added with 1mg/mL of the polypeptide of SEQ ID NO: 1.
4. A cosmetic product according to claim 3, characterized in that it comprises a pharmaceutically acceptable carrier.
5. The cosmetic of claim 4, wherein the pharmaceutically acceptable carrier comprises an emulsion.
6. The cosmetic according to claim 5, wherein the emulsion is an oil-in-water emulsion or a water-in-oil emulsion.
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