Mild Anti-aging composition containing recombinant collagen and use thereof

Abstract

Provided are a mild anti-aging composition containing a recombinant collagen and use thereof. The mild anti-aging composition comprises vitamin A alcohol, glycolic acid, the recombinant collagen, schizophyllan, and chondroitin sulfate. The combination of vitamin A alcohol, glycolic acid, and the recombinant collagen possesses a synergistic anti-aging effect. Also, considering the irritation of the vitamin A alcohol and the glycolic acid to the skin, the combination of the recombinant collagen, schizophyllan, and chondroitin sulfate is used as the anti-irritation component. The three components have a significant synergistic effect in reducing the irritation to the skin. The composition features an excellent anti-aging effect and excellent mildness, and has good application prospects in the preparation of cosmetics.

Description

A mild anti-aging composition containing recombinant collagen and its application Technical Field

[0001] The present application belongs to the technical field of cosmetics and relates to a mild anti-aging composition containing recombinant collagen and its application. Background Art

[0002] Aging, also known as aging, is an inevitable stage in the life process, with skin aging being the most obvious manifestation. Aging skin not only manifests as wrinkles, sagging, and hyperpigmentation, but also leads to reduced epidermal metabolism, a weakened ability to repair damaged skin, and a diminished barrier function. Numerous ingredients offer anti-aging benefits, but few are both highly effective and gentle. For example, hydroxy acids can improve the appearance of photoaged or naturally aged skin, but they can be somewhat irritating, causing varying degrees of redness or stinging.

[0003] With the booming cosmetics market, consumers are not only demanding anti-aging and whitening benefits, but are also increasingly demanding mildness and non-irritation in skincare products. China's cosmetics market is the world's largest emerging market, and the proliferation of skincare and beauty products is overwhelming. Beyond learning how to use quality cosmetics, consumers are also increasingly concerned with their effectiveness and safety.

[0004] Factors influencing skin irritation include internal factors such as ethnicity, age, and skin health, as well as external factors such as climate and irritating cosmetic ingredients. Skin irritation can cause damage to the skin barrier, neurological dysfunction, and inflammation. For those with sensitive skin, skin irritation can lead to dryness, tightness, and redness. Even mild irritation can cause itching, scaling, burning, and stinging. In severe cases, negative emotions or mental symptoms can develop.

[0005] Therefore, it is very meaningful to develop a product that can achieve both high anti-aging effects and mild and non-irritating effects.

[0006] Summary of the Invention

[0007] The following is a summary of the subject matter described in detail herein. This summary is not intended to limit the scope of the claims.

[0008] In view of the deficiencies of the prior art, the purpose of the present application is to provide a mild anti-aging composition containing recombinant collagen and its application.

[0009] To achieve this goal, this application adopts the following technical solutions:

[0010] In a first aspect, the present application provides a mild anti-aging composition containing recombinant collagen, wherein the mild anti-aging composition comprises retinol, glycolic acid, recombinant collagen, schizophyllan and chondroitin sulfate.

[0011] Retinol is a fat-soluble vitamin composed of 3,7-dimethoxyvitamin A derivatives and is an active form of vitamin A. Retinol can promote the synthesis of collagen, promote skin metabolism, increase skin elasticity, and prevent and reduce wrinkles.

[0012] Glycolic acid penetrates deep into the skin to exfoliate, gently dissolving dead skin cells and improving skin texture, color, and pore size. It also boosts collagen and elastin levels in the skin to help fight wrinkles.

[0013] This application creatively combines two ingredients, retinol and glycolic acid, and cooperates with recombinant collagen to jointly exert anti-aging effects. The three have a potential mutually reinforcing effect in anti-aging, and there is a synergistic effect between retinol and glycolic acid, as well as between retinol and recombinant collagen. At the same time, considering that retinol and glycolic acid have certain irritation to the skin, this application uses a combination of recombinant collagen, schizophyllan and chondroitin sulfate as an anti-irritation component. The three have a significant synergistic effect in reducing skin irritation. This composition can well balance excellent anti-aging efficacy and excellent mildness, and its application in the preparation of cosmetics has prospects.

[0014] In one embodiment, the mild anti-aging composition comprises, by mass, 0.1-0.5 parts of retinol, 0.5-2 parts of glycolic acid, 0.01-1 parts of recombinant collagen, 0.1-0.5 parts of schizophyllan, and 0.1-3 parts of chondroitin sulfate.

[0015] Based on the potential interactions between the above ingredients, when the above five ingredients are combined in a specific mass ratio, they have better effects in reducing skin irritation and improving skin anti-aging properties.

[0016] The mass fraction of the retinol can be selected as 0.1 part, 0.15 part, 0.2 part, 0.25 part, 0.3 part, 0.35 part, 0.4 part, 0.45 part, 0.5 part, etc.; the mass fraction of the glycolic acid can be selected as 0.5 part, 0.6 part, 0.7 part, 0.8 part, 1 part, 1.2 parts, 1.5 parts, 1.8 parts, 2 parts, etc.; the mass fraction of the recombinant collagen can be selected as 0.01 part, 0.03 part, 0.05 part, 0.08 part, 0.1 part, 0.2 part, 0.4 The mass fraction of the carboxymethyl schizophyllan can be selected as 0.1 part, 0.15 part, 0.2 part, 0.25 part, 0.3 part, 0.35 part, 0.4 part, 0.45 part, 0.5 part, etc.; the mass fraction of the chondroitin sulfate can be selected as 0.1 part, 0.2 part, 0.3 part, 0.5 part, 0.8 part, 1 part, 1.5 parts, 2 parts, 2.5 parts, 3 parts, etc.; other specific point values ​​within the above numerical ranges can be selected, and they will not be repeated here.

[0017] In one embodiment, the nucleotide sequence encoding the recombinant collagen is the sequence shown in SEQ ID NO: 1.

[0018] Compared with recombinant collagen from other sources, the present application also found that the recombinant collagen from the above-mentioned specific source is more effective in supplementing skin collagen, anti-oxidation, anti-aging, and reducing skin irritation.

[0019] In a second aspect, the present application provides use of the mild anti-aging composition containing recombinant collagen according to the first aspect in the preparation of cosmetics.

[0020] In one embodiment, the mass percentage of the mild anti-aging composition containing recombinant collagen in the cosmetic is 0.1-10%, for example, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 7%, 8%, 9%, 10%, etc. Other specific point values ​​within this numerical range can be selected and will not be described here one by one.

[0021] In a third aspect, the present application provides an essence milk with anti-aging effects. The raw materials for preparing the essence milk with anti-aging effects include, in terms of mass percentage, 0.1-10% of the mild anti-aging composition containing recombinant collagen described in the first aspect, 1-5% of an emulsifier, 0.1-2% of a thickener, 5-20% of a polyol, and 50-95% of water.

[0022] The mass percentage of the mild anti-aging composition can be 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 7%, 8%, 9%, 10%, etc.; the mass percentage of the emulsifier can be 1%, 2%, 3%, 4%, 5%, etc.; the mass percentage of the thickener can be 0.1%, 0.5%, 0.7%, 0.8%, 1%, 1.2%, 1.4%, 1.5%, 1.6%, 1.8%, 2%, etc.; the mass percentage of the polyol can be 5%, 6%, 8%, 10%, 12%, 14%, 15%, 17%, 18%, 19%, 20%, etc.; the mass percentage of water can be 50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, etc.; other specific point values ​​within the above numerical ranges can be selected, and they will not be repeated here.

[0023] In one embodiment, the emulsifier includes any one of potassium cetyl phosphate, polysorbate 60, methyl glucose sesquistearate, sorbitan stearate, sorbitan polyether-30 tetraoleate, polysorbate 20, cetearyl olivate / sorbitan olivate or cetyl PEG / PPG-10 / 1 polydimethylsiloxane, or a combination of at least two thereof.

[0024] In one embodiment, the thickener includes any one or a combination of at least two of sodium polyacrylate, sodium polyacrylate grafted starch, ammonium acryloyldimethyltaurate / VP copolymer, or xanthan gum.

[0025] In one embodiment, the polyol includes any one or a combination of at least two of glycerol, 1,3-propylene glycol, panthenol, 1,2-hexanediol, or butanediol.

[0026] In one embodiment, the raw materials for preparing the essence milk with anti-aging efficacy further include, in terms of mass percentage, any one or a combination of at least two of the following: 1-10% plant-based conditioner, 0.1-2% pH adjuster, 0.01-0.5% chelating agent, 0.05-0.5% antioxidant, and 0.1-0.5% fragrance.

[0027] The mass percentage of the plant-derived conditioning agent can be selected as 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, etc.; the mass percentage of the pH adjusting agent can be selected as 0.1%, 0.3%, 0.5%, 0.8%, 1%, 1.2%, 1.4%, 1.6%, 1.8%, 2%, etc.; the mass percentage of the chelating agent can be selected as 0.01%, 0.03%, 0.05%, 0.08%, 0.1%, 0. 15%, 0.2%, 0.3%, 0.4%, 0.5%, etc.; the mass percentage of the antioxidant can be selected as 0.05%, 0.08%, 0.1%, 0.15%, 0.2%, 0.3%, 0.4%, 0.5%, etc.; the mass percentage of the fragrance can be selected as 0.1%, 0.15%, 0.2%, 0.3%, 0.4%, 0.5%, etc.; other specific point values ​​within the above numerical ranges can be selected, and they will not be repeated here.

[0028] In one embodiment, the plant-derived conditioning agent is selected from any one or a combination of at least two of perilla leaf extract, milk thistle extract, and shiitake mushroom extract; and can further be selected from a combination of perilla leaf extract, milk thistle extract, and shiitake mushroom extract.

[0029] When the above composition is used to prepare cosmetics, further adding perilla leaf extract, milk thistle extract and shiitake mushroom extract ingredients can further enhance the soothing and anti-irritation effects of the product.

[0030] In a fourth aspect, the present application provides a method for preparing the essence milk with anti-aging efficacy according to the third aspect, the preparation method comprising:

[0031] Mix water and part of the polyol in an aqueous phase pot, heat to 60-80°C, keep warm for 10-40 minutes, cool to 25-40°C, and then mix with retinol, glycolic acid, recombinant collagen, and chondroitin sulfate to obtain an aqueous phase mixture;

[0032] The emulsifier, thickener, remaining polyol, schizophyllan, and remaining raw materials are mixed in an emulsifier, homogenized at 30-50° C. for 5-15 minutes, then mixed with the aqueous phase mixture, and homogenized at 25-40° C. for 5-30 minutes to obtain the essence milk with anti-aging effect.

[0033] The specific point value in the range of 60-80°C can be selected as 60°C, 65°C, 70°C, 75°C, 80°C, etc.; the specific point value in the range of 10-40min can be selected as 10min, 15min, 20min, 25min, 30min, 35min, 40min, etc.; the specific point value in the range of 25-40°C can be selected as 25°C, 30°C, 35°C, 40°C, etc.; the specific point value in the range of 30-50°C can be selected as 30°C, 32°C, 33°C, 35°C, etc.; the specific point value in the range of 5-15min can be selected as 5min, 6min, 7min, 8min, 10min, 13min, 15min, etc.; the specific point value in the range of 5-30min can be selected as 5min, 6min, 7min, 8min, 10min, 13min, 15min, 20min, 25min, 30min, etc.; other specific point values ​​within the above numerical ranges are all selectable and will not be repeated here.

[0034] Compared with the prior art, this application has the following beneficial effects:

[0035] This application creatively combines two ingredients, retinol and glycolic acid, and cooperates with recombinant collagen to jointly exert anti-aging effects. The three have a potential mutually reinforcing effect in anti-aging, and there is a synergistic effect between retinol and glycolic acid, as well as between retinol and recombinant collagen. At the same time, considering that retinol and glycolic acid have certain irritation to the skin, this application uses a combination of recombinant collagen, schizophyllan and chondroitin sulfate as an anti-irritation component. The three have a significant synergistic effect in reducing skin irritation. This composition can well balance excellent anti-aging efficacy and excellent mildness, and its application in the preparation of cosmetics has prospects.

[0036] Still other aspects will become apparent upon reading and understanding the detailed description. DETAILED DESCRIPTION

[0037] In order to further illustrate the technical means and effects adopted by this application, the technical solution of this application is further explained below in combination with the preferred embodiments of this application, but this application is not limited to the scope of the embodiments.

[0038] The mass fractions of each component in the composition products involved in the following content are calculated based on the actual active ingredient content in the commercially available raw materials.

[0039] The schizophyllum polysaccharide involved in the following content is derived from the product model TZCSW02905 of Shaanxi Tianzhicheng Biotechnology Co., Ltd.; chondroitin sulfate is derived from the products of Guangzhou Huayu Biotechnology Co., Ltd.; perilla leaf extract is derived from the products of Lanzhou Waterless Biotechnology Co., Ltd.; milk thistle extract is derived from the products of Lanzhou Waterless Biotechnology Co., Ltd.; shiitake mushroom extract is derived from the products of Lanzhou Waterless Biotechnology Co., Ltd.; sodium polyacrylate is derived from the products of Guangzhou Tianjia Biotechnology Co., Ltd.; xanthan gum is derived from the products of Shanghai Puzhen Biological Co., Ltd.

[0040] The recombinant collagen involved in the following content is the product prepared in the preparation example. The pET-28a(+) vector in the preparation example was purchased from Maclean; BL21(DE3) chemically competent cells were purchased from Merck with product number CMC0014-4X40UL.

[0041] Preparation Example

[0042] This preparation example provides a recombinant collagen freeze-dried powder, and the preparation method thereof is carried out according to the method disclosed in Preparation Example 1 in ZL 202310123476.3, as follows:

[0043] (1) Construction of recombinant plasmid: According to the nucleotide sequence shown in SEQ ID NO: 1, the pET-28a(+) vector suitable for the expression vector Escherichia coli host was selected and sent to a sequencing company for synthesis to obtain the recombinant collagen expression vector pET-28a-1.

[0044] (2) Construction of recombinant fermentation strain: The pET-28a-1 expression vector was transformed into Escherichia coli (BL21 (DE3)) competent cells by chemical transformation. Positive clones were screened in the transformed plate colonies by agarose gel electrophoresis. The screened positive clones were transferred to 2 mL LB medium and cultured at 37°C, 200 rpm for 12 h. The cultured bacterial solution was diluted 1-fold with 50% glycerol by volume, distributed into cryovials, and frozen at -80°C to obtain a recombinant collagen fermentation strain.

[0045] (3) Fermentation culture of recombinant fermentation strain: 10 μL of recombinant collagen fermentation liquid was spread on a solid LB plate, cultured in a 37°C incubator for 12 h, and then a single clone was selected and inoculated into 50 mL of LB liquid culture medium containing 50 μg / mL kanamycin. The culture was placed in a shaker at 37°C, 200 rpm for 14 h, and then inoculated into 200 mL of LB liquid culture medium containing 50 μg / mL kanamycin at a ratio of 1:100. The culture was cultured at 37°C until the OD value of the bacterial liquid was 0.6, and then IPTG was added to make the final concentration reach 1 mM. The culture was continued for 4 h, and then the bacteria were collected by centrifugation at 4°C, 4000 rpm for 15 min.

[0046] (4) Purification of recombinant collagen: The cells were ultrasonically disrupted and centrifuged, and the precipitate was washed with Triton X-100 washing solution, and the supernatant was removed by centrifugation again. 6 M guanidine hydrochloride was added to the precipitate, and the pH was adjusted to 8. After the precipitate was fully dissolved, the solution was passed through nickel affinity chromatography resin and eluted in sequence using 20 mM, 50 mM, 100 mM, 200 mM, and 500 mM concentration gradient imidazole solutions. The eluate was concentrated using a protein concentration column, and the concentrate was collected and dialyzed overnight (dialysis bag, RC membrane, 14 KD, flat width 44 mm (Sangong F600112)). The protein concentration was then detected using a BCA kit, and 0.5 mL of a 5% volume concentration mannitol aqueous solution was added to every 5 mg of protein and mixed with the mannitol aqueous solution. The solution was then pre-frozen at -40°C for 10 min using a freeze dryer, and finally freeze-dried at -35°C to obtain recombinant collagen freeze-dried powder.

[0047] Example 1

[0048] This embodiment provides a mild anti-aging composition, which is composed of the following components in parts by mass: 0.2 parts of retinol, 0.7 parts of glycolic acid, 0.05 parts of recombinant collagen, 0.2 parts of schizophyllan, and 0.3 parts of chondroitin sulfate.

[0049] Example 2

[0050] This embodiment provides a mild anti-aging composition, which is composed of the following components in parts by mass: 0.1 part of retinol, 1 part of glycolic acid, 0.01 part of recombinant collagen, 0.5 part of schizophyllan, and 1 part of chondroitin sulfate.

[0051] Example 3

[0052] This embodiment provides a mild anti-aging composition, which is composed of the following components in parts by mass: 0.4 parts of retinol, 0.5 parts of glycolic acid, 0.05 parts of recombinant collagen, 0.1 parts of schizophyllan, and 0.4 parts of chondroitin sulfate.

[0053] Comparative Example 1

[0054] This comparative example provides a composition, which differs from Example 1 only in that it does not contain retinol, the mass fraction of glycolic acid is 0.9 parts, and the other components and contents remain unchanged.

[0055] Comparative Example 2

[0056] This comparative example provides a composition, which differs from Example 1 only in that it does not contain glycolic acid, the mass fraction of vitamin A is 0.9 parts, and the other components and contents remain unchanged.

[0057] Comparative Example 3

[0058] This comparative example provides a composition, which differs from Example 1 only in that it does not contain recombinant collagen, and its mass fraction is distributed to the mass of glycolic acid, while other components and contents remain unchanged.

[0059] Comparative Example 4

[0060] This comparative example provides a composition, which differs from Example 1 only in that it does not contain Schizophyllan, the mass fraction of chondroitin sulfate is 0.5 parts, and the other components and contents remain unchanged.

[0061] Comparative Example 5

[0062] This comparative example provides a composition, which differs from Example 1 only in that it does not contain chondroitin sulfate, the mass fraction of Schizophyllan is 0.5 parts, and the other components and contents remain unchanged.

[0063] Comparative Example 6

[0064] This comparative example provides a composition, which differs from Example 1 only in that it does not contain recombinant collagen, and its mass fraction is distributed to the mass of Schizophyllan, while other components and contents remain unchanged.

[0065] Application Example 1

[0066] This application example provides an essence lotion, the formula of which includes the following ingredients:

[0067] Its preparation method is:

[0068] (1) Mixing water and glycerin in an aqueous phase pot, heating to 70° C., keeping warm for 20 minutes, cooling to 35° C., and then mixing with the vitamin A alcohol, glycolic acid, recombinant collagen, and chondroitin sulfate in the composition to obtain an aqueous phase mixture;

[0069] (2) The emulsifier, thickener, remaining polyol, and Schizophyllan were mixed in an emulsifier, homogenized at 45°C for 10 minutes, and then mixed with the aqueous phase mixture, homogenized at 35°C for 20 minutes, and the pH value was adjusted to obtain an essence milk.

[0070] Application Example 2-3

[0071] This application example provides two essence lotions, the formula of which differs from that of Application Example 1 only in that the mild anti-aging composition of Example 1 is replaced with an equal amount of the anti-aging composition prepared in Examples 2-3, and other components and contents remain unchanged.

[0072] Its preparation method is:

[0073] (1) Mixing water and glycerin in an aqueous phase pot, heating to 65° C., keeping warm for 30 minutes, cooling to 30° C., and then mixing with the vitamin A alcohol, glycolic acid, recombinant collagen, and chondroitin sulfate in the composition to obtain an aqueous phase mixture;

[0074] (2) The emulsifier, thickener, remaining polyol, and Schizophyllan were mixed in an emulsifier, homogenized at 40°C for 15 minutes, and then mixed with the aqueous phase mixture, homogenized at 30°C for 25 minutes, and the pH value was adjusted to obtain an essence milk.

[0075] Comparative Application Examples 1-6

[0076] Comparative Application Examples 1-6 provide six serums. Their formulations differ from those of Application Example 1 only in that the mild anti-aging composition of Example 1 is replaced with an equal amount of the compositions prepared in Comparative Examples 1-6. Other components and amounts remain unchanged. Their preparation methods follow those of Application Example 1.

[0077] Application Example 4

[0078] This application example provides an essence lotion, the formula of which includes the following ingredients:

[0079] Its preparation method is:

[0080] (1) Mixing water and glycerin in an aqueous phase pot, heating to 70° C., keeping warm for 20 minutes, cooling to 35° C., and then mixing with the vitamin A alcohol, glycolic acid, recombinant collagen, and chondroitin sulfate in the composition to obtain an aqueous phase mixture;

[0081] (2) The emulsifier, thickener, remaining polyol, and Schizophyllan were mixed in an emulsifier, homogenized at 45°C for 10 minutes, and then mixed with the aqueous phase mixture and the plant extract, homogenized at 35°C for 20 minutes, and the pH value was adjusted to obtain an essence milk.

[0082] Application Example 5

[0083] This application example provides an essence lotion, the formula of which differs from that of Application Example 4 only in that the perilla leaf extract is missing, and its mass is proportionally distributed to the mass of the milk thistle extract and the shiitake mushroom extract, while other combinations and contents remain unchanged.

[0084] Application Example 6

[0085] This application example provides an essence lotion, the formula of which differs from that of Application Example 4 only in that milk thistle extract is missing, and its mass is proportionally distributed to the mass of perilla leaf extract and shiitake mushroom extract, while other combinations and contents remain unchanged.

[0086] Application Example 7

[0087] This application example provides an essence milk, the formula of which differs from that of application example 4 only in that shiitake mushroom extract is missing, and its mass is proportionally distributed to the mass of milk thistle extract and perilla leaf extract, while other combinations and contents remain unchanged.

[0088] Test Example 1

[0089] Chicken embryo chorioallantoic membrane vascular test:

[0090] (1) Experimental conditions

[0091] Chicken embryo: SPF grade Bailaihang chicken.

[0092] Culture conditions: incubator temperature 37.5±0.5℃, relative humidity 55%-70%.

[0093] Reagents and controls: Positive control (PC1): 1.0% SDS solution;

[0094] Positive control (PC2): 0.1 mol / L NaOH solution;

[0095] Negative control (NC): 0.9% NaCl solution;

[0096] Test sample (TA): The products prepared in Examples 1-3 and Comparative Examples 1-6 were diluted to 5% with deionized water.

[0097] (2) Experimental methods

[0098] CAM preparation: Purchase 0-day-old chicken embryos. During the incubation process, turn them over once a day. When the embryos are 5 days old, check their developmental status. On the 6th day, peel off the eggshell to expose the white egg membrane; carefully remove the inner membrane with tweezers to ensure that the vascular membrane is not damaged.

[0099] Formal test: For each group of six chick embryos, the test substance was ground into a powder. After gentle squeezing into a 0.3 mL volume in a graduated container (e.g., a microcentrifuge tube), the powder was applied directly to the CAM, ensuring that at least 50% of the CAM surface was covered by the test substance. After 3 minutes of application, the test substance on the CAM membrane was gently rinsed with normal saline. This rinse may quickly mask any slight bleeding changes on the CAM membrane. Therefore, the results should be observed after 30 seconds of rinsing. The extent of each toxic effect, including bleeding, coagulation, and vascular lysis, was observed to the nearest second, and the extent of the reaction was recorded. The same procedure was used for the positive and negative controls.

[0100] Data analysis: The severity of each reaction was recorded, graded and scored. Bleeding was divided into no bleeding (0 point), mild bleeding (1 point), moderate bleeding (2 points), and severe bleeding (3 points) according to the severity. Coagulation was divided into no coagulation (0 point), mild coagulation (1 point), moderate coagulation (2 points), and severe coagulation (3 points) according to the severity. Vascular melting was divided into no vascular melting (0 point), mild vascular melting (1 point), moderate vascular melting (2 points), and severe vascular melting (3 points) according to the severity.

[0101] Calculate the endpoint score (ES) according to the following formula, rounded to two decimal places: Score per embryo = Sum of the degree of bleeding, coagulation, and vascular lysis observed in each embryo; ES = Mathematical sum of the scores of all six embryos. Classify the eye irritation of the test substance according to the table below based on the ES value.

[0102] The endpoint scores ES values ​​of the tested samples are shown in Table 1.

[0103] Table 1

[0104] It can be seen from the data results in Table 1 that the irritation classification of the composition involved in the present application is non-irritating. Compared with the results of Comparative Examples 4-6, recombinant collagen, schizophyllan and chondroitin sulfate can effectively reduce the irritation of the composition, and there is a significant synergistic effect between schizophyllan and chondroitin sulfate, as well as between them and recombinant collagen.

[0105] Test Example 2

[0106] In vitro hyaluronidase inhibition assay:

[0107] Hyaluronic acid is a component within the tissue matrix that restricts the diffusion of water and other extracellular substances. After being hydrolyzed by hyaluronidase, the intercellular space becomes non-sticky, causing cell degranulation and the exudation of newly synthesized mediators. This exerts biological effects and leads to the occurrence of immediate allergic reactions. Hyaluronidase is a participant in allergic reactions and is strongly correlated with allergies. The anti-allergic effect of a product can be evaluated by measuring its inhibitory activity.

[0108] The in vitro anti-allergic efficacy of Examples 1-3 and Comparative Examples 1-6 was investigated using the hyaluronidase inhibition method. The products obtained in Examples 1-3 and Comparative Examples 1-6 were diluted with deionized water to prepare solutions with a concentration of 5% as sample solutions.

[0109] 0.1 mL of CaCl2 solution (0.25 mmol / L) and 0.5 mL of hyaluronidase solution (100 U / mL) were reacted in a 37°C water bath for 20 min. 0.5 mL of the extract was added and the mixture was kept warm for 20 min. 0.5 mL of sodium hyaluronate solution (0.5 g / L) was then added and the mixture was allowed to react for 30 min, after which it was removed and cooled for 5 min. 0.1 mL of sodium hydroxide solution (0.4 mol / L) and 0.5 mL of acetylacetone solution (3.5 mL of acetylacetone dissolved in 50 mL of 1.0 mol / L sodium carbonate solution) were added and the mixture was boiled in a water bath for 15 min, then immediately transferred to an ice water bath for 5 min. 1 mL of Ehrlich reagent (0.8 g of p-dimethylaminobenzaldehyde dissolved in 15 mL of concentrated hydrochloric acid and 15 mL of anhydrous ethanol) was added dropwise and diluted with 3 mL of anhydrous ethanol. The mixture was allowed to stand at 25°C for 20 min for color development. The absorbance was measured at 540 nm, and dipotassium glycyrrhizate was used as a control.

[0110] The hyaluronidase inhibition rate was calculated according to the following formula:

[0111] Hyaluronidase inhibition rate = {[(A-B)-(C-D)] / (AB)}×100%

[0112] In the formula: A is the absorbance of the control solution (acetic acid buffer solution is used instead of the sample solution); B is the absorbance of the control blank solution (acetic acid buffer solution is used instead of the sample solution and enzyme solution); C is the absorbance of the sample solution; D is the absorbance of the sample blank solution (acetic acid buffer solution is used instead of the enzyme solution).

[0113] The inhibition rate of each group of samples on hyaluronidase was investigated, and the results are shown in Table 2:

[0114] Table 2

[0115] As shown in Table 2, the composition of the present application has a certain degree of inhibitory effect on hyaluronidase, with an inhibition rate of over 38%, indicating that the composition can alleviate or inhibit allergic reactions. Furthermore, the recombinant collagen, schizophyllan, and chondroitin sulfate in the composition have a significant synergistic effect in achieving the above-mentioned effects.

[0116] Test Example 3

[0117] Skin elasticity evaluation test:

[0118] (1) Subjects: A total of 140 volunteers aged 18-50 years (male:female ratio 1:4) without hypersensitivity were randomly divided into 14 groups, with 2 males and 8 females in each group.

[0119] Preparation before the test: The subject should mark the measurement area on the inner side of the forearms of both hands. The experimental area should be at least 3cm×3cm, and the interval between each test area should be at least 1cm.

[0120] Test samples: emulsions prepared in Application Examples 1-7 and Comparative Application Examples 1-6.

[0121] (2) Test process: 13 groups of test samples were placed at 2 ml / cm 2 A single application of the test sample was performed (the blank control group was pure water). Using latex gloves, the sample was evenly applied to the test area and gently patted until absorbed. Use once daily for 28 days. On the 14th day, skin elasticity was measured. After thoroughly cleansing the face with a standard cleanser and gently drying it with a paper towel, the skin elasticity tester probe was placed vertically on the test area in a room with a room temperature of 22°C ± 2°C and a relative humidity of 40 ± 2%. The test was started after 15 minutes in a room with a room temperature of 22°C ± 2°C and a relative humidity of 40 ± 2%. The probe was then removed after the test cycle was completed. Three points were measured on each test area. The results are shown in Table 3. On the 28th day, the anti-wrinkle effect was measured. After thoroughly cleansing the face with a standard cleanser and gently drying it with a paper towel, the skin elasticity tester probe was placed vertically on the test area in a room with a room temperature of 22°C ± 2°C and a relative humidity of 40 ± 2%. The VISIA facial image analyzer was used to photograph the subject's face. The analysis area should be minimized from interference from reflective areas and hair. The subjects' faces (including the front, left side, and right side) were photographed, and the absolute scores obtained were statistically analyzed to record the wrinkle detection data of the subjects' faces. The test results are shown in Table 3.

[0122] Table 3

[0123] As can be seen from the data results in Table 3, the composition involved in the present application has a good effect of restoring skin elasticity, among which glycolic acid and retinol promote each other in promoting skin elasticity, and work together with recombinant collagen to achieve the best effect.

[0124] Test Example 4

[0125] Sensory evaluation test:

[0126] Test Example 3: The same group of volunteers rated the anti-aging and mildness of the product after use, with the highest score being 5 and the lowest being 1, with a score range of 1-5. The evaluation results are shown in Table 4 (average score):

[0127] Table 4

[0128] As can be seen from Table 4, the composition involved in the present application and the cosmetics prepared therefrom have both excellent anti-aging properties and mildness. The plant extracts added to the cosmetics can further enhance the mildness of the composition, and the perilla leaf extract, milk thistle extract, and shiitake mushroom extract have a certain synergistic effect in improving the mildness of the product.

[0129] The applicant declares that while the above-mentioned embodiments are used to illustrate the technical solutions of this application, this application is not limited to these embodiments, and does not imply that this application must rely on these embodiments in order to be implemented. Persons skilled in the art should understand that any improvements to this application, equivalent replacements for the raw materials of the products of this application, addition of auxiliary ingredients, and selection of specific methods, etc., fall within the scope of protection and disclosure of this application.

[0130] The preferred embodiments of the present application are described in detail above. However, the present application is not limited to the specific details of the above embodiments. Within the technical concept of the present application, various simple modifications can be made to the technical solution of the present application, and these simple modifications all fall within the scope of protection of the present application.

[0131] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any appropriate manner unless there is any contradiction. In order to avoid unnecessary repetition, this application will not further describe various possible combinations.

Claims

1. A mild anti-aging composition containing recombinant collagen, comprising retinol, glycolic acid, recombinant collagen, schizophyllan and chondroitin sulfate.

2. The mild anti-aging composition containing recombinant collagen according to claim 1, wherein: The mild anti-aging composition comprises, by weight, 0.1-0.5 parts of retinol, 0.5-2 parts of glycolic acid, 0.01-1 parts of recombinant collagen, 0.1-0.5 parts of schizophyllan and 0.1-3 parts of chondroitin sulfate.

3. The mild anti-aging composition containing recombinant collagen according to claim 1 or 2, wherein: The nucleotide sequence encoding the recombinant collagen is the sequence shown in SEQ ID NO:

1.

4. Use of the mild anti-aging composition containing recombinant collagen according to any one of claims 1 to 3 in the preparation of cosmetics.

5. The use according to claim 4, wherein: The mass percentage of the mild anti-aging composition containing recombinant collagen in the cosmetics is 0.1-10%.

6. An essence milk with anti-aging effect, wherein the raw materials for its preparation include, by mass percentage: A mild anti-aging composition containing 0.1-10% recombinant collagen as claimed in any one of claims 1 to 3, 1-5% emulsifier, 0.1-2% thickener, 5-20% polyol and 50-95% water.

7. The essence milk with anti-aging effect according to claim 6, wherein: The emulsifier includes any one or a combination of at least two of potassium cetyl phosphate, polysorbate-60, methyl glucose sesquistearate, sorbitan stearate, sorbitan polyether-30 tetraoleate, polysorbate-20, cetearyl olivate / sorbitan olivate or cetyl PEG / PPG-10 / 1 polydimethylsiloxane; Optionally, the thickener includes any one or a combination of at least two of sodium polyacrylate, sodium polyacrylate grafted starch, ammonium acryloyldimethyltaurate / VP copolymer or xanthan gum; Optionally, the polyol includes any one of glycerol, 1,3-propylene glycol, panthenol, 1,2-hexanediol or butanediol, or a combination of at least two thereof.

8. The essence milk with anti-aging effect according to claim 6, wherein: The raw materials for preparing the essence milk with anti-aging effect also include, by mass percentage, any one or a combination of at least two of: 1-10% plant-derived conditioner, 0.1-2% pH adjuster, 0.01-0.5% chelating agent, 0.05-0.5% antioxidant, and 0.1-0.5% fragrance.

9. The essence milk with anti-aging effect according to claim 8, wherein: The plant-derived conditioning agent is selected from any one of perilla leaf extract, milk thistle extract, and shiitake mushroom extract, or a combination of at least two of them; further, a combination of perilla leaf extract, milk thistle extract, and shiitake mushroom extract can be selected.

10. The method for preparing the essence milk with anti-aging effect according to any one of claims 6 to 9, comprising: Mix water and part of the polyol in a water phase pot, heat to 60-80°C, keep warm for 10-40 minutes, cool to 25-40°C, and then mix with retinol, glycolic acid, recombinant collagen, and chondroitin sulfate to obtain a water phase mixture; The emulsifier, thickener, remaining polyol, Schizophyllan polysaccharide, and remaining raw materials are mixed in an emulsifying pot, homogenized at 30-50° C. for 5-15 minutes, and then mixed with the aqueous phase mixture, and homogenized at 25-40° C. for 5-30 minutes to obtain the essence milk with anti-aging effect.

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