Anti-wrinkle firming composition as well as preparation method and application thereof

Abstract

The invention relates to an anti-wrinkle and firming composition as well as a preparation method and application thereof. The anti-wrinkle and firming composition comprises the following components in parts by mass: 0.01-10 parts of composite polypeptide, 0.01-5 parts of recombinant collagen and 0.01-20 parts of amino acid or a derivative thereof, the composite polypeptide comprises acetyl peptide, palmitoyl peptide and carnosine. The brand-new anti-wrinkle and firming composition is developed through multi-dimensional formula design, acetyl peptide, palmitoyl peptide, carnosine and recombinant collagen are compounded, amino acid or amino acid derivatives are supplemented, the mutual synergistic effect is achieved, and the anti-wrinkle and firming purposes are achieved.

Description

Technical Field

[0001] The present invention belongs to the technical field of cosmetics, and relates to an anti-wrinkle and firming composition, a preparation method and application thereof, and specifically relates to an anti-wrinkle and firming composition, a preparation method thereof and application thereof in the preparation of cosmetics. Background Art

[0002] Anti-aging is an eternal theme. With the increase of age, the skin thickness becomes thinner, collagen is lost, and elastic fibers are broken, resulting in wrinkles, sagging, and gradual aging of the skin. People are often keen on using anti-aging cosmetics to stimulate the regeneration of collagen and elastin in the skin to achieve the effect of repairing the dermis and achieving the purpose of anti-aging. In response to the loss of collagen, collagen is added to cosmetics to fight against aging and wrinkles, but large-molecule collagen is difficult to be absorbed through the skin and has immunogenicity, which brings allergy risks, and the effect is not obvious. In recent years, with the development of synthetic biology and the recognition of the efficacy of some active peptides, people have begun to compete to use two or more peptides in cosmetics for compounding. By adding functional peptides, it can directly stimulate the absorption, metabolism, circulation, growth and reproduction of human skin to achieve anti-aging and wrinkle effects. This has become a new research direction and product orientation for beauty and skin care.

[0003] CN116236411A uses recombinant type III humanized collagen polypeptide, which retains a certain activity of collagen and is easier to be absorbed through the skin. It is supplemented with arginine to achieve the function of anti-wrinkle and firming the skin. CN115417843A discloses a compound extracted from Eryngium littoralis, which achieves the purpose of anti-aging by adding palmitoyl tripeptide-5 and palmitoyl tetrapeptide-7 to promote the production of skin collagen. CN116172913A further comprehensively considers endogenous and exogenous anti-aging and moisturizing. Its component, the filtrate of the fermentation product of bifid yeast, provides skin moisturizing, while the combination of acetyl hexapeptide-1, acetyl hexapeptide-7 and acetyl hexapeptide-8 achieves the effect of internal and external anti-aging.

[0004] At present, there are several problems in the application of active peptides in cosmetics: first, most of the active ingredients can only be absorbed through the epidermis and are difficult to enter the dermis to be absorbed; second, there is a lack of research on the synergistic effect of several peptides, which leads to overlapping functions and waste, and even mutual interference; third, cosmetics are affected by oxygen content, microorganisms, temperature, etc., which destroy the stability of peptides, and conventionally added preservatives, surfactants, etc. are also difficult to ensure the stability of peptides, and long-term use increases the burden on the skin, easily accelerates pigmentation, and even changes the structure of the peptides, thereby losing their activity. Therefore, creating a stable and preservative-free system can make the various peptides work together to maximize their effectiveness. Summary of the invention

[0005] In view of the deficiencies in the prior art, the object of the present invention is to provide an anti-wrinkle and firming composition, a preparation method and application thereof, and specifically to provide an anti-wrinkle and firming composition, a preparation method thereof and application thereof in the preparation of cosmetics.

[0006] In order to achieve the purpose of the invention, the present invention adopts the following technical solutions:

[0007] In a first aspect, the present invention provides an anti-wrinkle and firming composition, which comprises, by mass, 0.01-10 parts of a composite polypeptide, 0.01-5 parts of recombinant collagen, and 0.01-20 parts of amino acids or their derivatives; the composite polypeptide comprises acetyl peptide, palmitoyl peptide and carnosine.

[0008] The present invention has developed a new anti-wrinkle and firming composition through a multi-dimensional formula design, which compounds acetyl peptide, palmitoyl peptide, carnosine and recombinant collagen, and supplements them with amino acids or amino acid derivatives to exert their synergistic effects to achieve the purpose of anti-wrinkle and firming.

[0009] Among them, acetyl peptides can reduce muscle shrinkage and relieve expression lines by inhibiting the transmission of nerve signals; palmitoyl peptides can simulate the mechanism of the body's own transforming growth factor to produce collagen, effectively making up for the lack of collagen in the skin; collagen in the dermis will undergo a Maillard reaction with sugar to produce glycation end products, causing the skin to lose elasticity and turn yellow. The glycation reaction can be inhibited by adding carnosine. The structure of carnosine is similar to the glycosylation site of protein. By sacrificing itself to undergo a glycation reaction with sugar, the level of protein glycation can be reduced; recombinant collagen and amino acids or their derivatives can give the skin elasticity and firmness, smooth fine lines, delay skin aging, promote cell regeneration, and brighten the skin tone. The addition of arginine can not only play a synergistic role with recombinant collagen in giving skin elasticity, but also solve the characteristic that palmitoyl peptides need to be solubilized by adding emulsifiers, avoiding the risk of irritation caused by the addition of emulsifiers, and better play the role of palmitoyl peptides.

[0010] The mass fractions of the composite polypeptide can be 0.01 parts, 0.05 parts, 0.1 parts, 0.5 parts, 1 parts, 2 parts, 3 parts, 4 parts, 5 parts, 6 parts, 7 parts, 8 parts, 9 parts, 10 parts, etc.

[0011] The mass fraction of the recombinant collagen can be 0.01 parts, 0.05 parts, 0.1 parts, 0.5 parts, 1 parts, 2 parts, 3 parts, 4 parts, 5 parts, etc.

[0012] The mass fraction of the amino acid or its derivative can be 0.01 parts, 0.05 parts, 0.1 parts, 0.5 parts, 1 parts, 3 parts, 5 parts, 7 parts, 9 parts, 10 parts, 13 parts, 15 parts, 17 parts, 20 parts, etc.

[0013] Other specific point values ​​within the above numerical ranges can be selected and will not be described in detail here.

[0014] Preferably, the mass ratio of the acetyl peptide, palmitoyl peptide and carnosine is (0.05-5):(0.05-5):(0.05-20).

[0015] The specific point values ​​in the (0.05-5) can be selected from 0.05, 0.1, 0.5, 1, 2, 3, 4, 5, etc.

[0016] The specific point values ​​in the mentioned (0.05-20) can be selected as 0.05, 0.1, 0.2, 0.3, 0.4, 0.5, 1, 2, 4, 6, 8, 10, 12, 14, 16, 18, 20, etc.

[0017] Other specific point values ​​within the above numerical ranges can be selected and will not be described in detail here.

[0018] Preferably, the acetyl peptide includes any one of acetyl dipeptide-1 cetyl ester, acetyl tripeptide-1, acetyl tripeptide-5, acetyl tetrapeptide-2, acetyl tetrapeptide-3, acetyl tetrapeptide-5, acetyl tetrapeptide-9, acetyl hexapeptide-1, acetyl hexapeptide-3, acetyl hexapeptide-7, acetyl hexapeptide-8, acetyl heptapeptide-4 or acetyl octapeptide-3, or a combination of at least two thereof.

[0019] More preferably, acetyl hexapeptide-1 and / or acetyl tetrapeptide-9 are used. Compared with other types of acetyl peptides, acetyl hexapeptide-1 and / or acetyl tetrapeptide-9 have better effects in combination with palmitoyl peptide, carnosine and recombinant collagen.

[0020] The amino acid sequence of acetyl hexapeptide-1 is: Ac-Arg-Trp--His-Ala-Leu-Phe-NH2. It inhibits the synthesis of SNARE receptors and the release of catecholamine and acetylcholine neurotransmitters (Ach), thereby locally blocking the transmission of muscle contraction signals by nerves, regulating the depolarization state of the postsynaptic membrane, and reducing the postsynaptic membrane Ca 2+ The inflow and cross-linking of myosin relax the facial muscles, smoothing out dynamic lines, static lines and fine lines. Acetyl tetrapeptide-9 strengthens the skin's resistance to traction, stimulates the synthesis of type I collagen and basement membrane polysaccharide, and improves the thickness and firmness of the skin.

[0021] Preferably, the palmitoyl peptide includes any one of palmitoyl oligopeptide, palmitoyl dipeptide-7, palmitoyl tripeptide-1, palmitoyl tripeptide-5, palmitoyl tripeptide-8, palmitoyl tetrapeptide-3, palmitoyl tetrapeptide-7, palmitoyl pentapeptide-3, palmitoyl pentapeptide-4 or palmitoyl hexapeptide-12, or a combination of at least two thereof.

[0022] Palmitoyl tripeptide-5 is further preferred. Palmitoyl tripeptide-5 has a better effect in combination with acetyl peptide, carnosine and recombinant collagen than other types of palmitoyl peptides.

[0023] Palmitoyl tripeptide-5 can mimic our own polypeptide signals and amplify the signals, which can cause the sequence KRFK to bind to the inactive transforming growth factor β (TGFβ), leading to the release of active TGFβ, which can significantly reduce the average wrinkle parameters by 12%, and act on more ECM components to produce a greater and more uniform repair effect, thereby promoting the proliferation of collagen, elastic fibers and hyaluronic acid, and increasing the water content of the skin. In addition, palmitoyl tripeptide-5 can inhibit tyrosinase activity and inhibit melanin production. Arginine can provide immune function, regulate acid-base moisture, and balance oil.

[0024] Preferably, the recombinant collagen comprises recombinant type III collagen and / or recombinant type XVII collagen.

[0025] Recombinant type III collagen is further preferred. Compared with other types of recombinant collagen, recombinant type III collagen has a better effect in combination with acetyl peptide, palmitoyl peptide and carnosine.

[0026] The structure of recombinant human type III collagen is a single-chain structure. The basic repeating unit is GERGAPGFRPAGPNGIPGEKGPAGERGAP, which is a type III peptide of human collagen. The terminal sequence is GPPGPCCGGG, which is a type II peptide of human collagen. It has a unique 164.88° triple helix flexible curvature. This structure can significantly enhance the adhesion of collagen to cells. It can promote the expression of TβR II in the body and activate the TGF-β / Smads signaling pathway. TβRⅡ can phosphorylate TβRⅠ, and then phosphorylate Smad protein to mediate the signal pathway to the cell nucleus. Phosphorylation of Smad2 and Smad3 proteins is a key step in the Smad pathway, and their activity can be inhibited by Smad6 and Smad7; while collagen peptides can reduce the expression of Smad6 and Smad7. It can protect and promote fibroblast proliferation, relieve oxidative stress, and regulate related signaling pathways, thereby improving skin aging.

[0027] Preferably, the amino acid or its derivative comprises any one or a combination of at least two of lysine or its derivative, arginine or its derivative, and histidine or its derivative.

[0028] Arginine or its derivatives are further preferred. Compared with other types of amino acids or their derivatives, arginine or its derivatives have better effects in combination with recombinant collagen.

[0029] Arginine can directly reach the dermal fibroblasts, has strong permeability and biological activity, can make the skin more elastic and smooth, and can be used to repair and prevent skin damage.

[0030] Preferably, the anti-wrinkle and firming composition further comprises an energy-enabling substance; the energy-enabling substance comprises any one of glucose, nicotinamide riboside, ATP, fructose, galactose, sorbitol, mannose, sucrose or plant polysaccharides, or a combination of at least two thereof.

[0031] In view of the technical problem that most of the active ingredients in existing cosmetics can only be absorbed through the epidermis and are difficult to enter the dermis to be absorbed, the present invention also creatively adds an enabling substance, which can stimulate ATP synthesis in skin cells, activate cells, promote transdermal absorption of other active ingredients, optimize skin metabolism, help repair cell damage, further promote skin elasticity and alleviate or prevent fine lines and wrinkles. In addition, the enabling substance added by the present invention can also meet the freeze-drying needs of the product, minimize the damage to the cell structure and characteristics in the freeze-dried product, prevent the biological characteristics of the product from changing, protect the activity and efficacy of the polypeptide to the greatest extent, and ensure the efficacy stability of the product.

[0032] Preferably, the energy-enhancing substance comprises a combination of sucrose, nicotinamide riboside and glucose.

[0033] When the enabling substance is selected in the form of a combination of sucrose, nicotinamide riboside and glucose, the above effects are more significant.

[0034] Preferably, the anti-wrinkle and firming composition comprises, by weight, 0.01-10 parts of composite polypeptide, 0.01-5 parts of recombinant collagen, 0.01-20 parts of amino acids or their derivatives, and 0.01-10 parts of energy-enhancing substances.

[0035] Preferably, the anti-wrinkle and firming composition further comprises a radiation protectant; the radiation protectant comprises any one of niacinamide, carnosine, sodium DNA, citric acid, glutathione or vitamin C, or a combination of at least two thereof.

[0036] Considering that the existing cosmetic skin is susceptible to the influence of oxygen content, microorganisms, temperature, etc., which destroys the stability of the polypeptide, and the conventionally added preservatives and surfactants are also difficult to ensure the stability of the polypeptide, and long-term use increases the burden on the skin, easily accelerates pigmentation, and even changes the structure of the polypeptide to lose its activity, the present invention adopts sterilization filtration supplemented by irradiation sterilization, abandons the potential hazards of preservatives and cosolvents, does not contain preservatives, and makes the product safe to use, good biocompatibility, and no toxic side effects. At the same time, in order to meet the needs of this sterilization method, a specific type of radiation protectant is added to prevent the active ingredients such as polypeptides from being inactivated during the irradiation process.

[0037] Preferably, the radiation protectant comprises a combination of niacinamide, glutathione and vitamin C.

[0038] When the radiation protector is selected in the form of a combination of nicotinamide, glutathione and vitamin C, its effect in protecting the effective ingredients such as polypeptides to stably take effect is more significant.

[0039] Preferably, the anti-wrinkle and firming composition further comprises a moisturizer and / or a pH adjuster.

[0040] Preferably, the moisturizing agent comprises hyaluronic acid and / or sodium hyaluronate.

[0041] Hyaluronic acid is a member of the glycosaminoglycan (also known as mucopolysaccharide) family. Like other glycosaminoglycans, hyaluronic acid is also a high molecular weight linear polysaccharide composed of repeated connections of disaccharide units composed of aminohexose and hexuronic acid. There are also a large number of negatively charged carboxyl groups on the hyaluronic acid skeleton, which play a role in ion exchange and can regulate the concentration of cations around cells. In addition, hyaluronic acid also acts as a signal transduction molecule, and participates in cell signal transduction and regulates various cell activities, including cell proliferation, migration, differentiation, adhesion, etc., by binding to various protein receptors on the extracellular matrix and cell membrane, thereby regulating the physiological functions of the body. For example, hyaluronic acid can promote the aggregation of white blood cells at the site of inflammation by binding to the CD44 receptor, thereby promoting the body's immune and anti-inflammatory effects.

[0042] Preferably, the pH adjuster includes any one of citric acid, sodium hydroxide, disodium hydrogen phosphate or sodium dihydrogen phosphate, or a combination of at least two thereof.

[0043] Preferably, the pH value of the anti-wrinkle and firming composition is 3.0-8.0, for example, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, more preferably 5.0-7.0.

[0044] In a second aspect, according to the method for preparing the anti-wrinkle and firming composition of the first aspect, the method comprises: physically mixing the raw materials for preparation evenly.

[0045] Preferably, the preparation method specifically comprises:

[0046] The enabling substance and the moisturizing agent are mixed and dissolved in water, and then mixed with the composite polypeptide, recombinant collagen, amino acid or its derivatives, and then mixed with the radiation protective agent, filtered for sterilization, freeze-dried to obtain freeze-dried powder, and finally the freeze-dried powder is sterilized by irradiation.

[0047] Preferably, the freeze-drying process parameters are: pre-freezing at -60 to -40°C for 2-4h; sublimation drying temperature is -15 to -5°C, then vacuuming, vacuum degree is 10-30Pa, maintained for 24-48h; analytical drying temperature is 20-35°C, vacuum degree is 1-10Pa, maintained for 16-20h.

[0048] Preferably, the irradiation sterilization is carried out at 2-8°C, and the irradiation dose is 5-15 kGy.

[0049] In a third aspect, the present invention provides use of the anti-wrinkle and firming composition according to the first aspect in the preparation of cosmetics.

[0050] Preferably, the dosage form of the cosmetic includes microspheres, microcapsules, liquids, emulsions, creams, gels or patches.

[0051] Compared with the prior art, the present invention has the following beneficial effects:

[0052] The present invention has developed a new anti-wrinkle and firming composition through a multi-dimensional formula design, which compounds acetyl peptide, palmitoyl peptide, carnosine and recombinant collagen, and supplements them with amino acids or amino acid derivatives to exert their synergistic effects to achieve the purpose of anti-wrinkle and firming.

[0053] Among them, acetyl peptides can reduce muscle shrinkage and relieve expression lines by inhibiting the transmission of nerve signals; palmitoyl peptides can simulate the mechanism of the body's own transforming growth factor to produce collagen, effectively making up for the lack of collagen in the skin; collagen in the dermis will undergo a Maillard reaction with sugar to produce glycation end products, causing the skin to lose elasticity and turn yellow. The glycation reaction can be inhibited by adding carnosine. The structure of carnosine is similar to the glycosylation site of protein. By sacrificing itself to undergo a glycation reaction with sugar, the level of protein glycation can be reduced; recombinant collagen and amino acids or their derivatives can give the skin elasticity and firmness, smooth fine lines, delay skin aging, promote cell regeneration, and brighten the skin tone. The addition of arginine can not only play a synergistic role with recombinant collagen in giving skin elasticity, but also solve the characteristic that palmitoyl peptides need to be solubilized by adding emulsifiers, avoiding the risk of irritation caused by the addition of emulsifiers, and better play the role of palmitoyl peptides. DETAILED DESCRIPTION

[0054] In order to further explain the technical means and effects adopted by the present invention, the technical solution of the present invention is further described below in conjunction with the preferred embodiments of the present invention, but the present invention is not limited to the scope of the embodiments.

[0055] The high molecular weight sodium hyaluronate involved in the following embodiments or comparative examples is the product model HA-T purchased from Bloomage Biotechnology Co., Ltd.; the low molecular weight sodium hyaluronate is the product model HA-TLM purchased from Bloomage Biotechnology Co., Ltd.; the hydrolyzed sodium hyaluronate is the product model Nano HA purchased from Bloomage Biotechnology Co., Ltd.; the recombinant human type III collagen is the product model BSS purchased from Jiangsu Jiangshan Juyuan Biotechnology Co., Ltd.; the recombinant type XVII collagen is the product model ReCol3-TTA03-02SC purchased from Jiangsu Chuangjian Medical Technology Co., Ltd.

[0056] Example 1

[0057] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is as follows (based on the total mass of the product as 100%):

[0058] (1) Weigh 0.1% high molecular weight sodium hyaluronate, 0.2% low molecular weight sodium hyaluronate, 0.3% hydrolyzed sodium hyaluronate, and 5% sucrose into purified water and stir to dissolve completely;

[0059] (2) Add 0.1% palmitoyl tripeptide-5 and 1% arginine, and after complete dissolution, add 0.1% acetyl hexapeptide-1, 0.1% acetyl tetrapeptide-9, 0.2% recombinant human type III collagen, and 0.3% carnosine, stir at a speed of 500 r / min, and completely dissolve for 30 minutes;

[0060] (3) Add 0.05% niacinamide, 0.025% glutathione, and 0.002% vitamin C in sequence and keep stirring until completely dissolved;

[0061] (4) 0.22 μm sterile filtration;

[0062] (5) The solution obtained in the above step was pre-frozen at -40°C for 4 hours; the sublimation drying temperature was -10°C, and then vacuumed to a vacuum degree of 20 Pa for 24 hours; the analytical drying temperature was 25°C, the vacuum degree was 5 Pa, and the temperature was maintained for 16 hours to obtain a composite polypeptide freeze-dried powder;

[0063] (6) Sterilize by irradiation at 4°C with a dose of 10 kGy.

[0064] Example 2

[0065] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is as follows (based on the total mass of the product as 100%):

[0066] (1) Weigh 0.1% high molecular weight sodium hyaluronate, 0.1% low molecular weight sodium hyaluronate, 0.4% hydrolyzed sodium hyaluronate, and 5% sucrose, add to purified water, and stir to dissolve completely;

[0067] (2) Add 0.1% palmitoyl tripeptide-5 and 1% arginine, and after complete dissolution, add 0.05% acetyl hexapeptide-1, 0.05% acetyl tetrapeptide-9, 0.2% recombinant human type III collagen, and 0.25% carnosine, stir at a speed of 500 r / min, and completely dissolve for 30 minutes;

[0068] (3) Add 0.1% niacinamide, 0.025% glutathione, and 0.002% vitamin C in sequence and keep stirring until completely dissolved;

[0069] (4) 0.22 μm sterile filtration;

[0070] (5) The solution obtained in the above step was pre-frozen at -40°C for 4 hours; the sublimation drying temperature was -10°C, and then vacuumed to a vacuum degree of 20 Pa for 24 hours; the analytical drying temperature was 25°C, the vacuum degree was 5 Pa, and the temperature was maintained for 16 hours to obtain a composite polypeptide freeze-dried powder;

[0071] (6) Sterilize by irradiation at 4°C with a dose of 10 kGy.

[0072] Example 3

[0073] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of Embodiment 1 only in that acetyl hexapeptide-1 and acetyl tetrapeptide-9 in step (2) are replaced by acetyl tripeptide-5 and acetyl heptapeptide-4, respectively, the added amounts remain unchanged, and other conditions remain unchanged.

[0074] Example 4

[0075] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of Embodiment 1 only in that palmitoyl tripeptide-5 in step (2) is replaced by palmitoyl tetrapeptide-3, the added amount remains unchanged, and other conditions remain unchanged.

[0076] Example 5

[0077] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of embodiment 1 only in that the recombinant type III collagen in step (2) is replaced with recombinant type XVII collagen, the added amount remains unchanged, and other conditions remain unchanged.

[0078] Example 6

[0079] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of embodiment 1 only in that arginine in step (2) is replaced by lysine, the added amount remains unchanged, and other conditions remain unchanged.

[0080] Example 7

[0081] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of Example 1 only in that the sucrose in step (1) is replaced with an equal amount of nicotinamide riboside, and other conditions remain unchanged.

[0082] Example 8

[0083] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of Embodiment 1 only in that the sucrose in step (1) is replaced by an equal amount of glucose, and other conditions remain unchanged.

[0084] Example 9

[0085] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of Embodiment 1 only in that the sucrose in step (1) is replaced by a combination of sucrose, nicotinamide riboside and glucose in an equal mass ratio, the total amount added remains unchanged, and other conditions remain unchanged.

[0086] Example 10

[0087] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of Embodiment 1 only in that step (3) is removed, i.e., no radiation protection agent is added, and other conditions remain unchanged.

[0088] Embodiment 11

[0089] This embodiment provides an anti-wrinkle and firming composition, and the preparation method thereof is different from that of Embodiment 1 only in that sucrose is not added in step (1), and other conditions remain unchanged.

[0090] Comparative Example 1

[0091] This comparative example provides an anti-wrinkle and firming composition, the preparation method of which is different from that of Example 1 only in that acetyl hexapeptide-1 and acetyl tetrapeptide-9 are not contained, and their reduced amounts are added proportionally to the mass of palmitoyl tripeptide-5, carnosine and recombinant type III collagen, and other conditions remain unchanged.

[0092] Comparative Example 2

[0093] This comparative example provides an anti-wrinkle and firming composition, the preparation method of which is different from that of Example 1 only in that palmitoyl tripeptide-5 is not contained, and its reduced amount is added proportionally to the mass of acetyl hexapeptide-1 and acetyl tetrapeptide-9, carnosine and recombinant type III collagen, and other conditions remain unchanged.

[0094] Comparative Example 3

[0095] This comparative example provides an anti-wrinkle and firming composition, the preparation method of which is different from that of Example 1 only in that: carnosine is not contained, and its reduced amount is added proportionally to the mass of acetyl hexapeptide-1 and acetyl tetrapeptide-9, palmitoyl tripeptide-5 and recombinant type III collagen, and other conditions remain unchanged.

[0096] Comparative Example 4

[0097] This comparative example provides an anti-wrinkle and firming composition, the preparation method of which is different from that of Example 1 only in that: no recombinant type III collagen is contained, and its reduced amount is added proportionally to the mass of acetyl hexapeptide-1 and acetyl tetrapeptide-9, palmitoyl tripeptide-5 and carnosine, and other conditions remain unchanged.

[0098] Comparative Example 5

[0099] This comparative example provides an anti-wrinkle and firming composition, the preparation method of which is different from that of Example 1 only in that arginine is not contained, and the reduced amount thereof is all added to the mass of recombinant type III collagen, and other conditions remain unchanged.

[0100] Test Example 1

[0101] Fibroblast proliferation assay:

[0102] Specific operation: Prepare L929 mouse fibroblast suspension at 1×10 4 / mL was inoculated in a culture plate, and the supernatant was removed after culturing for 24 hours; the experimental group was added with 50mg / ml of the lyophilized powder prepared in Example 1-11 and Comparative Example 1-5, and dissolved with fresh cell culture medium; the blank control group was added with an equal amount of fresh cell culture medium; the culture was continued for 72 hours, and the absorbance (A) value was measured at a wavelength of 490nm on an ELISA reader to calculate the relative cell proliferation rate. The sample without adding active ingredients was used as the blank control group, and its cell survival rate was 100%. The results are shown in Table 1:

[0103] Table 1

[0104]

[0105]

[0106] As shown in Table 1, compared with the comparative example product, the composition product of the present invention has excellent efficacy in promoting fibroblast proliferation, and there is a significant synergistic effect between acetyl peptide, palmitoyl peptide, carnosine and recombinant collagen, and arginine can promote the efficacy of recombinant collagen. At the same time, the selection of enabling substances, the selection of acetyl peptide, the selection of palmitoyl peptide, and the selection of recombinant collagen will affect the performance of the above-mentioned efficacy to a certain extent.

[0107] Test Example 2

[0108] Collagen synthesis test:

[0109] Specific operation: Human dermal fibroblasts HSF (SV40) are treated with trypsin / EDTA at a concentration of 0.25%, and the amount used in a T25 culture flask is 1 ml. Observe under an inverted microscope until most of the cells become round and are in a suspended state, add about 3 times the volume of trypsin-containing DMEM medium to terminate digestion, collect into a centrifuge tube, and centrifuge at 800r / min for 6min. After centrifugation, discard the supernatant, add a certain volume of cell culture medium to the centrifuge tube, mix the cells with an elbow pipette, and count them with a cell counter or a hemocytometer. After diluting with cell culture medium to the inoculation density, inoculate into a 96-well plate with a liquid volume of 200μL per well. After inoculation, place in a CO2 incubator and culture for 24h.

[0110] The culture medium in the 96-well plate was discarded, and the test medium was added to the test wells. The experimental groups were added with lyophilized powders prepared in Examples 1-11 and Comparative Examples 1-5 at a concentration of 10%, and dissolved with cell culture medium; the positive control group was TGF-β1 cytokine, and the blank control group was added with an equal amount of fresh cell culture medium, 200 μL per well. After the administration, the 96-well plate was placed in a CO2 incubator for 24 hours. After the incubation and culture, 200 μL of cell culture supernatant was collected from each well in a 1.5 mL sterile centrifuge tube and placed in a -80°C ultra-low temperature refrigerator for freezing.

[0111] ELISA test: Perform the test according to the instructions of the human type I collagen enzyme-linked immunosorbent assay kit.

[0112] The concentration of the standard is taken as the vertical axis, OD 450 The value is the horizontal axis, and a standard curve is made according to the OD of the sample. 450 , find out the corresponding concentration.

[0113] Calculation formula for upregulation rate of type I collagen: Upregulation rate (%) = (T / C-1) × 100%

[0114] Among them, T is the average content of type I collagen in each experimental group, and C is the average content of type I collagen in the blank control.

[0115] The results are shown in Table 2:

[0116] Table 2

[0117]

[0118]

[0119] As shown in Table 2, compared with the comparative example product, the composition product of the present invention has excellent effect of promoting collagen production, and there is a significant synergistic effect between acetyl peptide, palmitoyl peptide, carnosine and recombinant collagen, and arginine can promote the effect of recombinant collagen. At the same time, the selection of enabling substances, the selection of acetyl peptide, the selection of palmitoyl peptide, and the selection of recombinant collagen will affect the performance of the above effects to a certain extent.

[0120] Test Example 3

[0121] Elastase inhibition test:

[0122] Elastase can specifically hydrolyze elastase substrates. The absorbance of the hydrolysis product can be measured by spectrophotometry to calculate the inhibition rate of elastase activity. The in vitro elastase activity inhibition experiment can be used to evaluate the inhibitory effect of the sample on elastase activity, thereby evaluating the potential anti-aging and firming effect of the sample.

[0123] Specific operation: Take a 50% sample solution in the experimental group, add Tris-HCl buffer and elastase solution, incubate at room temperature (25°C) for 5 minutes, add elastase substrate solution, react at room temperature (25°C) for 10 minutes, and immediately measure the absorbance at 410nm using an enzyme reader. Vitamin C (VC, 100mg / mL) is used as a positive control, and 3 parallel samples are set for each experimental group.

[0124] The results are shown in Table 3:

[0125] Table 3

[0126]

[0127]

[0128] As shown in Table 3, compared with the comparative example product, the composition product of the present invention has excellent efficacy in inhibiting elastase, and there is a significant synergistic effect between acetyl peptide, palmitoyl peptide, carnosine and recombinant collagen, and arginine can promote the efficacy of recombinant collagen. At the same time, the selection of enabling substances, the selection of acetyl peptide, the selection of palmitoyl peptide, and the selection of recombinant collagen will affect the performance of the above-mentioned efficacy to a certain extent.

[0129] Test Example 4

[0130] Antioxidant efficacy test:

[0131] Solution preparation: First, dissolve 1 mg of DPPH in about 20 mL of anhydrous ethanol, ultrasonicate for 5 minutes, and shake thoroughly to make the upper and lower parts uniform. Take 1 mL of the DPPH solution and measure the A value at 517 nm to make A=1.2-1.3; then prepare each sample solution: dilute each group of samples with anhydrous ethanol to prepare a series of sample solutions with a concentration of 50% (V / V).

[0132] Specific method: The sample addition system of each group of experiments is shown in the table below. After the reaction solution is shaken well, it is placed in a 37°C water bath for 15 minutes and then taken out. The absorbance value of the solution is measured with a UV-visible spectrophotometer.

[0133] Reagents C1 / mL C2 / mL C3 / mL DPPH solution 2 2 0 Sample solution 0 2 2 Anhydrous ethanol 2 0 2

[0134] The calculation formula of DPPH free radical scavenging rate is shown as follows:

[0135] DPPH free radical scavenging rate = 1-(A2-A3) / A1×100%

[0136] Where A1, A2, and A3 are the absorbance values ​​of C1, C2, and C3, respectively.

[0137] The results are shown in Table 4:

[0138] Table 4

[0139]

[0140]

[0141] It can be seen from the results in Table 4 that, compared with the comparative example product, the composition product of the present invention has excellent antioxidant effect, and the selection of the enabling substance and the selection of the polypeptide type will affect the performance of the above-mentioned effect to a certain extent.

[0142] The applicant declares that the present invention uses the above-mentioned embodiments to illustrate an anti-wrinkle and firming composition and its preparation method and application, but the present invention is not limited to the above-mentioned embodiments, that is, it does not mean that the present invention must rely on the above-mentioned embodiments to be implemented. Those skilled in the art should understand that any improvement of the present invention, equivalent replacement of various raw materials of the product of the present invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of the present invention.

[0143] The preferred embodiments of the present invention are described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept of the present invention, a variety of simple modifications can be made to the technical solution of the present invention, and these simple modifications all belong to the protection scope of the present invention.

[0144] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not further describe various possible combinations.

Claims

1. An anti-wrinkle and firming composition, characterized in that: The anti-wrinkle and firming composition comprises, by weight, 0.01-10 parts of a composite polypeptide, 0.01-5 parts of recombinant collagen, and 0.01-20 parts of amino acids or their derivatives; the composite polypeptide comprises acetyl peptide, palmitoyl peptide and carnosine.

2. The anti-wrinkle and firming composition according to claim 1, characterized in that: The mass ratio of the acetyl peptide, palmitoyl peptide and carnosine is (0.05-5):(0.05-5):(0.05-20); Preferably, the acetyl peptide comprises any one of acetyl dipeptide-1 cetyl ester, acetyl tripeptide-1, acetyl tripeptide-5, acetyl tetrapeptide-2, acetyl tetrapeptide-3, acetyl tetrapeptide-5, acetyl tetrapeptide-9, acetyl hexapeptide-1, acetyl hexapeptide-3, acetyl hexapeptide-7, acetyl hexapeptide-8, acetyl heptapeptide-4 or acetyl octapeptide-3, or a combination of at least two thereof; acetyl hexapeptide-1 and / or acetyl tetrapeptide-9 are further preferred; Preferably, the palmitoyl peptide includes any one of palmitoyl oligopeptide, palmitoyl dipeptide-7, palmitoyl tripeptide-1, palmitoyl tripeptide-5, palmitoyl tripeptide-8, palmitoyl tetrapeptide-3, palmitoyl tetrapeptide-7, palmitoyl pentapeptide-3, palmitoyl pentapeptide-4 or palmitoyl hexapeptide-12, or a combination of at least two thereof; palmitoyl tripeptide-5 is further preferred.

3. The anti-wrinkle and firming composition according to claim 1 or 2, characterized in that: The recombinant collagen includes recombinant type III collagen and / or recombinant type XVII collagen; further preferably, recombinant type III collagen; Preferably, the amino acid or its derivatives include any one of lysine or its derivatives, arginine or its derivatives, histidine or its derivatives, or a combination of at least two thereof; arginine or its derivatives are further preferred.

4. The anti-wrinkle and firming composition according to any one of claims 1 to 3, characterized in that: The anti-wrinkle and firming composition further comprises an energy-enhancing substance; the energy-enhancing substance comprises any one or a combination of at least two of glucose, nicotinamide riboside, ATP, fructose, galactose, sorbitol, mannose, sucrose or plant polysaccharides; Preferably, the energy-enhancing substance comprises a combination of sucrose, nicotinamide riboside and glucose; Preferably, the anti-wrinkle and firming composition comprises, by weight, 0.01-10 parts of composite polypeptide, 0.01-5 parts of recombinant collagen, 0.01-20 parts of amino acids or their derivatives, and 0.01-10 parts of energy-enhancing substances.

5. The anti-wrinkle and firming composition according to any one of claims 1 to 4, characterized in that: The anti-wrinkle and firming composition further comprises a radiation protectant; the radiation protectant comprises any one of niacinamide, carnosine, sodium DNA, citric acid, glutathione or vitamin C, or a combination of at least two thereof; Preferably, the radiation protectant comprises a combination of niacinamide, glutathione and vitamin C.

6. The anti-wrinkle and firming composition according to any one of claims 1 to 5, characterized in that: The anti-wrinkle and firming composition further comprises a moisturizer and / or a pH adjuster; Preferably, the moisturizing agent comprises hyaluronic acid and / or sodium hyaluronate; Preferably, the pH adjuster comprises any one of citric acid, sodium hydroxide, disodium hydrogen phosphate or sodium dihydrogen phosphate, or a combination of at least two thereof; Preferably, the pH value of the anti-wrinkle and firming composition is 3.0-8.0, more preferably 5.0-7.

0.

7. The method for preparing the anti-wrinkle and firming composition according to any one of claims 1 to 6, characterized in that: The preparation method comprises: physically mixing the raw materials for preparation evenly.

8. The method for preparing the anti-wrinkle and firming composition according to claim 7, characterized in that: The preparation method specifically comprises: The enabling substance and the moisturizing agent are mixed and dissolved in water, and then mixed with the composite polypeptide, recombinant collagen, amino acid or its derivatives, and then mixed with the radiation protective agent, filtered for sterilization, freeze-dried to obtain freeze-dried powder, and finally the freeze-dried powder is sterilized by irradiation.

9. The method for preparing the anti-wrinkle and firming composition according to claim 8, characterized in that: The freeze-drying process parameters are: pre-freezing at -60 to -40°C for 2-4 hours; sublimation drying temperature is -15 to -5°C, then vacuuming, vacuum degree is 10-30Pa, and maintained for 24-48 hours; analytical drying temperature is 20-35°C, vacuum degree is 1-10Pa, and maintained for 16-20 hours; Preferably, the irradiation sterilization is carried out at 2-8°C, and the irradiation dose is 5-15 kGy.

10. Use of the anti-wrinkle and firming composition according to any one of claims 1 to 6 in the preparation of cosmetics; Preferably, the dosage form of the cosmetic includes microspheres, microcapsules, liquids, emulsions, creams, gels or patches.

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