Cosmetic composition, preparation method thereof and composition

Abstract

The invention provides a beautifying composition, a preparation method thereof and a composition. The cosmetic composition provided by the invention comprises 0.01 wt%-0.5 wt% of a cyclic hexapeptide compound with a structure as shown in a formula (I); 0.1 wt% to 5 wt% of a thickening agent; 1 wt% to 10 wt% of a polyol; 1 wt% to 10 wt% of an emulsifier; 1 wt% to 10 wt% of an oily raw material; 0.1 wt% to 5 wt% of a preservative; and the balance of water. The cyclic hexapeptide compound with the structure shown in the formula (I) has high stability and bioavailability, can remarkably promote secretion of collagen, especially I-type collagen, III-type collagen and IV-type collagen, has the effects of tightening, moisturizing and the like, and is used as an active component of the beautifying composition, so that the beautifying composition has good moisturizing and tightening effects, and the beautifying composition is free of toxic and side effects. The method can be applied to the fields of cosmetics and beauty and health care.

Description

Technical Field

[0001] The present invention relates to the technical field of daily chemical industry, and in particular to a cosmetic composition, a preparation method thereof and the composition. Background Art

[0002] Collagen is the main component of animal connective tissue and the most abundant and widely distributed functional protein in mammals, accounting for 25% to 30% of the total protein. It plays a vital role in maintaining the elasticity, firmness and moisture content of the skin. As we age, collagen in the human body will gradually be lost, causing the skin to show signs of sagging, so supplementing or stimulating the production of collagen becomes the key to firming.

[0003] Retinol is an effective firming ingredient, but it is photosensitive and irritating, so it is not suitable for people with sensitive skin. Special attention should be paid to sun protection when using it, and improper use may cause adverse reactions such as skin allergies. Peptide ingredients are relatively mild and have high safety. They are suitable for people of all skin types, especially Asians, whose skin has a thin stratum corneum and is easily sensitive. Peptide ingredients are more in line with the skin firming needs of Asians. Summary of the invention

[0004] In view of this, the technical problem to be solved by the present invention is to provide a cosmetic composition, a preparation method thereof and the composition. The cosmetic composition provided by the present application uses a cyclohexapeptide compound as a main ingredient and has good moisturizing and firming effects.

[0005] The present invention provides a cosmetic composition, comprising: 0.01 wt% to 0.5 wt% of a cyclohexapeptide compound; 0.1 wt% to 5 wt% of a thickener; 1 wt% to 10 wt% of a polyol; 1 wt% to 10 wt% of an emulsifier; 1 wt% to 10 wt% of an oily raw material; 0.1 wt% to 5 wt% of a preservative; and the balance of water;

[0006] The cyclohexapeptide compound has the structure of formula (I):

[0007]

[0008] In formula (I), x and y are independently selected from integers of 1 to 5;

[0009] R 1 , R 2 , R 3 and R 4 Independently selected from H, hydroxy, halogen or C1-C6 alkoxy.

[0010] In some specific implementations, x and y are independently 1, 2, 3, 4, or 5, preferably 1 or 2.

[0011] In some specific implementations, R 1 , R 2 , R 3 and R 4 At least one of them is hydroxyl, halogen or C1-C6 alkoxy. Among them, halogen is selected from F, Cl, Br or I, preferably F. C1-C6 alkoxy is preferably methoxy, ethoxy, propoxy, and more preferably methoxy. In some specific implementations, R 1 , R 2 , R 3 and R 4 At least one of them is a hydroxyl group.

[0012] In some specific implementations, R 4 is hydroxyl or H. In some specific implementations, R 4 is hydroxyl or H, and R 2 is H, hydroxyl, halogen or C1-C6 alkoxy. In some specific implementations, R 4 is hydroxyl or H, R 2 is H, hydroxyl, halogen or C1-C6 alkoxy, and R 1 is H or hydroxyl; R 3 For H.

[0013] In some specific implementations, the cyclohexapeptide compound has a structure of formula I-1, I-2, I-3, I-4, I-5, I-6, I-7, I-8, I-9, I-10, I-11, I-12, I-13, I-14, I-15, I-16, I-17, I-18, I-19, I-20, I-21, I-22, I-23 or I-24:

[0014]

[0015]

[0016] The present application has no particular limitation on the source of the cyclohexapeptide compound, which can be prepared according to the following method:

[0017] a) using solid phase synthesis to obtain a linear hexapeptide, wherein the amino acid sequence of the linear hexapeptide is: the amino C2-C6 alkyl acid-4 position is replaced by R 1 The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 Replaced Pro;

[0018] Or: 4 bits are R1 The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 Substituted Pro-amino C2-C6 alkyl acids;

[0019] Or: 4 bits are R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 1 Replaced Pro;

[0020] b) cyclizing the linear hexapeptide to obtain a cyclic hexapeptide compound.

[0021] The present application firstly adopts solid phase synthesis to prepare a linear hexapeptide, and then cyclizes and cuts it to obtain a cyclic hexapeptide compound. Since cyclization is required, the present application has no special restrictions on the starting amino acid when preparing the linear hexapeptide. For example, the amino acid sequence of the linear hexapeptide can be: the amino acid C2 to C6 alkyl acid -4 position is replaced by R 1 The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 Pro replaced; or: 4 digits replaced by R 1 The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 Substituted Pro-amino C2-C6 alkyl acid; or: R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 1 The following is a linear hexapeptide with amino C2-C6 alkyl acid-4 position replaced by R 1 The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 The replaced Pro is used as an example for explanation.

[0022] In some specific implementations, the linear hexapeptide is prepared according to the following method:

[0023] a) Sequentially connect 4 R 4 Pro replaced, 4th position replaced by R 3 Substituted Pro, amino C2-C6 alkyl acid, R 2 Pro replaced, 4th position replaced by R 1 Substituted Pro and amino C2-C6 alkyl acid to obtain a resin containing a peptide chain;

[0024] b) Removal of the resin to obtain a linear hexapeptide.

[0025] The present invention adopts a solid phase synthesis method to sequentially connect 4 R 4 Pro replaced, 4th position replaced by R 3 Substituted Pro, amino C2-C6 alkyl acid, R 2 Pro replaced, 4th position replaced by R 1 Substituted Pro and amino C2~C6 alkyl acid to obtain a resin containing a peptide chain.

[0026] The present invention uses solid phase synthetic resin as the starting point for synthesis, firstly reacts with 4-position R 4 The substituted proline raw material reacts and the 4-position R is attached to the resin 4 Specifically, the present invention comprises a solid phase synthesis resin, a proline substituted by R 4 The substituted proline raw material, the reaction solvent and the base are mixed for the first reaction, and then mixed with the alcohol raw material for the second reaction. After deprotection, the 4-position R 4 Replaced Pro-Resin.

[0027] In some specific implementations, the solid phase synthetic resin includes but is not limited to Wang-resin, CTC-resin, etc., preferably CTC-resin. 4 The substituted proline raw material is preferably Fmoc-Pro (4-R 4 )-OH. The reaction solvent includes but is not limited to dichloromethane (DCM), N,N-dimethylformamide (DMF), dichloromethane (DCM), dimethyl sulfoxide (DMSO), etc., preferably DCM. The base includes but is not limited to N,N-diisopropylethylamine (DIEA), triethylamine (TEA), pyridine (Py), etc., preferably DIEA. In some specific implementations, the solid phase synthesis resin, the 4-position R 4 The molar ratio of the substituted proline raw material to the base is 1-10:5-20:0.005-0.1, preferably 5-8:10-15:0.01-0.05. In some specific implementations, the temperature of the first reaction is preferably 10°C-30°C, preferably 15°C-25°C, and the time is preferably 2h-5h, more preferably 2.5h-4.5h.

[0028] After the first reaction is completed, an alcohol raw material is directly added thereto to carry out a second reaction. In some specific implementations, the time of the second reaction is preferably 1 min to 10 min, more preferably 3 min to 8 min. In some specific implementations, the alcohol raw material includes but is not limited to methanol, ethanol, isopropanol, etc., preferably methanol. After the second reaction is completed, the reaction product is filtered, and the obtained resin is washed twice with a solvent such as DCM, washed twice with methanol, and then washed twice with DMF, and then deprotected. The present application has no special restrictions on the deprotection method. The deprotection method can be selected according to the type of protecting group. For example, when the solid phase synthetic resin is CTC-resin and the 4-position is replaced by R 4 The substituted proline raw material is preferably Fmoc-Pro (4-R 4 )-OH, preferably piperidine is used for deprotection, specifically comprising: mixing the obtained resin with a piperidine solution for reaction. In some specific implementations, the piperidine solution is preferably a DMF solution of piperidine, wherein the mass concentration of piperidine is preferably 10% to 30%, more preferably 15% to 25%. The reaction temperature is preferably 10°C to 30°C, preferably 15°C to 25°C, and the reaction time is preferably 20min to 50min, more preferably 25min to 45min. After the reaction is completed, the obtained product is washed with a reaction solvent such as DMF, preferably washed 6 times, and the resin is drained.

[0029] Get 4 bits by R 4 After the Pro-resin is substituted, the 4-position is connected to R 3 Pro replaced. This application first replaces the 4th position with R 3 The substituted Pro raw material, condensation agent and solvent are mixed for reaction, and the obtained reaction solution is mixed with the 4-position R 4 The substituted Pro-resin was mixed and reacted, and after deprotection, the 4-position R 3 The Pro-4 position was replaced by R 4 In some specific implementations, the 4 position is replaced by R 3 The substituted Pro raw material is Fmoc-Pro (4-R 3 )-OH; the condensing agent is a mixture of HOBt / DIC; and the solvent is DMF. Specifically, the present invention firstly 3 )-OH and HOBt are mixed and dissolved with DMF, and then mixed with DIC for reaction. In some specific implementations, the Fmoc-Pro (4-R 3The molar ratio of )-OH, HOBt and DIC is preferably 5-30:5-30:5-30, more preferably 8-25:8-25:8-25. In some specific implementations, the reaction is carried out under stirring, the reaction temperature is preferably 2°C to 8°C, and the reaction time is preferably 10min to 20min. After the reaction is completed, the obtained reaction product is mixed with the 4-positioned R 4 The substituted Pro-resin is mixed for reaction, the reaction temperature is preferably 10°C to 30°C, preferably 15°C to 25°C, the time is preferably 1h to 3h, more preferably 1.5h to 2.5h. After the reaction is completed, the reaction product is filtered, and the obtained resin is washed with a reaction solvent such as DMF, preferably three times, and then deprotected. The present application has no special restrictions on the deprotection method, and the deprotection method can be selected according to the type of protecting group. For example, when the proline raw material is R- 3 When Pro is substituted, piperidine is preferably used for deprotection, specifically comprising: mixing the obtained resin with a piperidine solution for reaction. In some specific implementations, the piperidine solution is preferably a DMF solution of piperidine, wherein the mass concentration of piperidine is preferably 10% to 30%, more preferably 15% to 25%. The reaction temperature is preferably 10°C to 30°C, preferably 15°C to 25°C, and the time is preferably 20min to 40min, more preferably 25min to 35min. After the reaction is completed, the obtained product is washed with a reaction solvent such as DMF, preferably washed 6 times, and the resin is drained.

[0030] Get 4 bits by R 3 The Pro-4 position was replaced by R 4 After the Pro-resin is substituted, an amino C2-C6 alkyl acid is connected thereto. In the present application, the amino C2-C6 alkyl acid raw material, a condensing agent and a solvent are mixed for reaction, and then the obtained reaction solution is mixed with R 3 The Pro-4 position was replaced by R 4 The substituted Pro-resin is mixed and reacted, and after deprotection, amino C2-C6 alkyl acid-R 3 The Pro-4 position was replaced by R 4Substituted Pro-resin. In some specific implementations, the amino C2-C6 alkyl acid raw material is Fmoc-amino C2-C6 alkyl acid, such as aminoacetic acid (glycine, Gly) or β-aminopropionic acid (β-Ala); the condensing agent is a mixture of HOBt / DIC; the solvent is DMF. Specifically, the present application first mixes and dissolves the amino C2-C6 alkyl acid raw material and HOBt with DMF, and then mixes with DIC for reaction. In some specific implementations, the molar ratio of the amino C2-C6 alkyl acid raw material, HOBt and DIC is preferably 5-30:5-30:5-30, and more preferably 8-25:8-25:8-25. In some specific implementations, the reaction is carried out under stirring, the reaction temperature is preferably 2°C to 8°C, and the reaction time is preferably 10min to 20min. After the reaction is completed, the obtained reaction product is mixed with R at the 4 position. 3 The Pro-4 position was replaced by R 4 The substituted Pro-resin is mixed for reaction, and the reaction temperature is preferably 10°C to 30°C, preferably 15°C to 25°C, and the time is preferably 1h to 3h, and more preferably 1.5h to 2.5h. After the reaction is completed, the reaction product is filtered, and the obtained resin is washed with a reaction solvent such as DMF, preferably washed three times, and then deprotected. The present application has no special restrictions on the deprotection method, and the deprotection method can be selected according to the type of protecting group. For example, when the amino C2 to C6 alkyl acid raw material is Fmoc-amino C2 to C6 alkyl acid, piperidine is preferably used for deprotection, specifically including: mixing the obtained resin with a piperidine solution for reaction. In some specific implementations, the piperidine solution is preferably a DMF solution of piperidine, wherein the mass concentration of piperidine is preferably 10% to 30%, and more preferably 15% to 25%. The reaction temperature is preferably 10°C to 30°C, preferably 15°C to 25°C, and the time is preferably 20min to 40min, and more preferably 25min to 40min. After the reaction is completed, the obtained product is washed with a reaction solvent such as DMF, preferably 6 times, and the resin is drained to obtain amino C2-C6 alkyl acid-R 3 The Pro-4 position was replaced by R 4 Replaced Pro-Resin.

[0031] Obtain amino C2~C6 alkyl acid-R 3 The Pro-4 position was replaced by R 4 The Pro-resin substituted by R 2 Pro replaced, 4th position replaced by R 1 Substituted Pro and amino C2-C6 alkyl acid to obtain amino C2-C6 alkyl acid -4 position R 1The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 The substituted Pro-CTC resin is not described in detail in this application.

[0032] The amino C2-C6 alkyl acid -4 position is R 1 The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 After replacing the Pro-CTC resin, remove the resin thereon. In this application, trifluoroacetic acid is preferably used to remove the resin. The specific steps are as follows:

[0033] The amino C2-C6 alkyl acid -4 position is replaced by R 1 The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 The substituted Pro-CTC resin was mixed with trifluoroacetic acid / DCM solution and reacted, and then filtered to remove the resin.

[0034] In some specific implementations, the mass concentration of trifluoroacetic acid in the trifluoroacetic acid / DCM is 0.5% to 2%, preferably 1% to 1.5%. In some specific implementations, the reaction temperature is 20°C to 40°C, preferably 25°C to 35°C, and the reaction time is 20min to 40min, preferably 25min to 35min. In some specific implementations, the reaction is preferably carried out under stirring.

[0035] After removing the resin, the sequence is amino C2~C6 alkyl acid -4 position is replaced by R 1 The Pro-4 position was replaced by R 2 The substituted Pro-amino C2-C6 alkyl acid-4 position is replaced by R 3 The Pro-4 position was replaced by R 4 A linear hexapeptide of substituted Pro, cyclizing the linear hexapeptide to obtain a cyclic hexapeptide compound. Specifically, the present application reacts the linear hexapeptide under the action of a condensing agent to cyclize the hexapeptide. In some specific implementations, the condensing agent is a composition of DIC, HOBt and DIEA, and the molar ratio of DIC, HOBt and DIEA is 1 to 2:1 to 2:1 to 2, preferably 1:1:1. The reaction temperature is 20°C to 40°C, preferably 25°C to 35°C, and the reaction time is 10h to 20h, preferably 12h to 16h.

[0036] As mentioned above, R 1 , R 2 , R 3 and R 4 are independently selected from H, hydroxyl, halogen or C1-C6 alkoxy, when R 1 , R 2 , R 3 and R 4 When R is independently selected from H, the corresponding amino acid is proline (Pro), and there is no need to protect the 4-position substituent; 1 , R 2 , R 3 and R 4 When R is independently selected from halogen or C1-C6 alkoxy, there is no need to protect the 4-substituent, and the corresponding cyclohexapeptide compound can be obtained after the cyclization reaction is completed. 1 , R 2 , R 3 and R 4 When independently selected from hydroxyl, the corresponding amino acid is hydroxyproline (Hyp), which needs to protect the hydroxyl group of the 4-position substituent. The present application has no special restrictions on the protecting group, and is preferably a tert-butyl protecting group. When the 4-position substituent is protected by a protecting group, it is preferably deprotected after the cyclization reaction is completed. The present application preferably uses trifluoroacetic acid for deprotection, and the specific steps are as follows:

[0037] The cyclohexapeptide compound containing a protecting group is mixed with a mixture of trifluoroacetic acid / triisopropylsilane / water, reacted, filtered, and the obtained filtrate is crystallized in a solvent to obtain a crude peptide.

[0038] In some specific implementations, the volume ratio of trifluoroacetic acid / triisopropylsilane / water is 80-90:5-10:5-10, more preferably 90:5:5. In some specific implementations, the reaction temperature is 0-5°C, and the reaction time is 1h-3h, preferably 1.5h-2.5h. In some specific implementations, the solvent includes but is not limited to tert-butyl methyl ether (MTBE), ether, isopropyl ether, etc., preferably MTBE. In some specific implementations, the crystallization temperature is preferably 2°C-8. The crystallized solid is centrifuged and vacuum dried to obtain a crude peptide.

[0039] After the cyclohexapeptide compound is obtained by cyclization reaction or the crude peptide is obtained by deprotection and crystallization, it is preferably purified by high performance liquid chromatography (HPLC), and the cyclohexapeptide compound represented by formula (I) can be obtained after freeze-drying. In some specific implementations, the HPLC column is a reverse phase C18 column.

[0040] The cyclohexapeptide compound provided in the present application has higher stability and bioavailability, can significantly promote the secretion of collagen, especially type I collagen, type III collagen and type IV collagen, has the effects of firming and moisturizing, and can be used as an active ingredient in a cosmetic composition.

[0041] In the cosmetic composition provided in the present application, the concentration of the cyclohexapeptide compound is 0.01 wt% to 0.5 wt%, preferably 0.03 wt% to 0.3 wt%, and more preferably 0.05 wt% to 0.2 wt%.

[0042] The cosmetic composition provided in the present application includes a thickener, and the concentration of the thickener is 0.1wt% to 5wt%, preferably 0.5wt% to 4wt%, and more preferably 0.8wt% to 3wt%. In some specific implementations, the thickener is preferably an ion-resistant thickener, such as acrylic acid (ester) / C10-30 alkyl acrylate crosspolymer, polyacrylate crosspolymer-6, etc., which can be one or more thereof. When the thickener is a mixture, the present application has no special restrictions on the ratio between the specific substances.

[0043] The cosmetic composition provided in the present application includes a polyol, which is used to promote the penetration of active ingredients, and the concentration of the polyol is 1wt% to 10wt%, preferably 2wt% to 8wt%, and more preferably 3wt% to 7wt%. In some specific implementations, the polyol includes but is not limited to glycerol, 1,3-propylene glycol, dipropylene glycol, 1,2-hexanediol and 1,2-pentanediol, etc., and can be one or more of them. When the polyol is a mixture, the present application has no special restrictions on the ratio between the specific substances.

[0044] The cosmetic composition provided in the present application includes an emulsifier, and the concentration of the emulsifier is 1wt% to 10wt%, preferably 2wt% to 8wt%, and more preferably 3wt% to 7wt%. In some specific implementations, the emulsifier is selected from at least one emulsifier with an HLB value of 8 to 15, such as cetearyl alcohol, polyglyceryl-10 stearate, cetearyl glucoside, etc., and can be one or more thereof. When the emulsifier is a mixture, the present application has no special restrictions on the ratio between the specific substances.

[0045] The cosmetic composition provided in the present application includes an oily raw material, and the concentration of the oily raw material is 1wt% to 10wt%, preferably 2wt% to 8wt%, and more preferably 3wt% to 7wt%. In some specific implementations, the oily raw material includes but is not limited to caprylic / capric glycerides, squalane, avocado tree fat, hydrogenated olive oil, isononyl isononanoate, white meadowfoam seed oil, etc., preferably a mixture of caprylic / capric glycerides, squalane and avocado tree fat. The mass ratio of caprylic / capric glycerides, squalane and avocado tree fat is 2 to 4:2 to 4:1 to 2, preferably 2:2:1.

[0046] The cosmetic composition provided in the present application includes a preservative, and the concentration of the preservative is 0.1wt% to 5wt%, preferably 0.2wt% to 3wt%, and more preferably 0.3wt% to 2wt%. In some specific implementations, the preservative includes but is not limited to phenoxyethanol, p-hydroxyacetophenone and 1,2-hexanediol, etc., and can be one or more of them. When the preservative is a mixture, the present application has no special restrictions on the ratio between the specific substances.

[0047] In order to improve the efficacy of the cosmetic composition, in some specific implementations, it also includes 0.1wt% to 0.5wt% of Tremella polysaccharide and / or 0.05wt% to 0.5wt% of sodium hyaluronate. Tremella polysaccharide is a polysaccharide extracted from Tremella fruiting body, which has the functions of moisturizing, anti-oxidation and improving skin quality. It is used in combination with the above-mentioned cyclohexapeptide compound to make the obtained cosmetic composition have good moisturizing and firming effects. In some specific implementations, the concentration of the Tremella polysaccharide is preferably 0.15wt% to 0.45wt%, more preferably 0.2wt% to 0.4wt%. Sodium hyaluronate has multiple effects such as moisturizing, lubricating, repairing skin, improving skin elasticity, inhibiting bacterial growth, etc., and is used in combination with the above-mentioned cyclohexapeptide compound, or with the above-mentioned cyclohexapeptide compound and Tremella polysaccharide, so that the obtained cosmetic composition has good moisturizing, firming and moisturizing effects. In some specific implementations, the concentration of the sodium hyaluronate is preferably 0.08wt% to 0.45wt%, more preferably 0.1wt% to 0.4wt%. In some specific implementations, the molecular weight of the sodium hyaluronate is 100 KDa to 1000 KDa, preferably 150 KDa to 900 KDa, and more preferably 150 KDa to 900 KDa.

[0048] The present application also provides a method for preparing the cosmetic composition described in the above technical solution, comprising the following steps:

[0049] Dissolving the cyclohexapeptide compound and the preservative respectively to obtain a cyclohexapeptide compound solution and a preservative solution;

[0050] After the polyol, thickener and water are uniformly mixed, an aqueous phase solution is obtained;

[0051] After the emulsifier and the oily raw material are uniformly mixed, an oil phase solution is obtained;

[0052] The oil phase solution is added to the water phase solution and then homogenized to obtain an emulsion;

[0053] The emulsion, the cyclohexapeptide compound solution and the preservative solution are uniformly mixed to obtain a cosmetic composition.

[0054] The present application firstly dissolves the cyclohexapeptide compound and the preservative in water to obtain the cyclohexapeptide compound and the preservative solution respectively, mixes the polyol, the thickener and the water to obtain the aqueous phase solution, and mixes the emulsifier and the oily raw material to obtain the oil phase solution.

[0055] In some specific implementations, the present application preferably dissolves the cyclohexapeptide compound and the preservative in water at 20°C to 65°C. In some specific implementations, the present application preferably mixes the polyol, thickener and water at 50°C to 85°C to obtain an aqueous phase solution, and keeps it warm at 50°C to 85°C. In some specific implementations, the present application preferably mixes the emulsifier and the oily raw material at 50°C to 85°C to obtain an oil phase solution, and keeps it warm at 50°C to 85°C. When the cosmetic composition also includes Tremella polysaccharide and / or sodium hyaluronate, Tremella polysaccharide and / or sodium hyaluronate are uniformly mixed with polyol, thickener and water.

[0056] After obtaining the oil phase solution and the water phase solution respectively, the oil phase solution is added to the water phase solution for homogenization, and an oil-in-water emulsion is formed after uniform emulsification. The present application has no special restrictions on the homogenization method, and it can be stirred evenly. In some specific implementation methods, the homogenization time is 5min to 25min.

[0057] The present application provides a composition, including the cyclohexapeptide compound, Tremella polysaccharide and hyaluronic acid described in the above technical solution in a mass ratio of 0.01-0.5:0.1-0.5:0.05-0.5. Among them, the cyclohexapeptide compound can significantly promote the secretion of collagen, especially type I collagen, type III collagen and type IV collagen, and has the effects of firming and moisturizing; Tremella polysaccharide is a polysaccharide extracted from Tremella fruiting body, which has the functions of moisturizing, anti-oxidation, firming and improving skin quality, and sodium hyaluronate has multiple effects such as moisturizing, lubricating, repairing skin, improving skin elasticity, and inhibiting bacterial growth. The above-mentioned cyclohexapeptide compound, Tremella polysaccharide and sodium hyaluronate are used in combination, so that the obtained beauty composition has good moisturizing and firming effects.

[0058] The cosmetic composition provided by the present invention comprises: 0.01wt% to 0.5wt% of a cyclohexapeptide compound having a structure of formula (I); 0.1wt% to 5wt% of a thickener; 1wt% to 10wt% of a polyol; 1wt% to 10wt% of an emulsifier; 1wt% to 10wt% of an oily raw material; 0.1wt% to 5wt% of a preservative; and the remainder of water. Among them, the cyclohexapeptide compound having a structure of formula (I) has high stability and bioavailability, can significantly promote the secretion of collagen, especially type I collagen, type III collagen and type IV collagen, has the effects of firming and moisturizing, etc., and as an active ingredient of the cosmetic composition, the cosmetic composition has good moisturizing and firming effects, and can be applied to the field of cosmetics and the field of beauty and health care. BRIEF DESCRIPTION OF THE DRAWINGS

[0059] Figure 1 The results of the effect of the cyclohexapeptide prepared in Example 1 on human keratinocytes;

[0060] Figure 2 The expression level of type I collagen in the cyclic hexapeptide experimental group prepared in Example 1;

[0061] Figure 3 The expression level of type III collagen in the cyclic hexapeptide experimental group prepared in Example 1;

[0062] Figure 4 The expression level of type IV collagen in the cyclohexapeptide experimental group prepared in Example 1;

[0063] Figure 5 This is a statistical chart of the water content increase rate of the cyclic collagen hexapeptide essence lotion;

[0064] Figure 6 This is a statistical chart of the elastase inhibition rate of the essence milk provided in the examples of the present application. DETAILED DESCRIPTION

[0065] The present invention provides a cosmetic composition, a preparation method thereof and a composition. A person skilled in the art can refer to the content of this invention and appropriately improve the process parameters to achieve the above. It should be particularly noted that all similar substitutions and modifications are obvious to a person skilled in the art and are considered to be included in the present invention. The method and application of the present invention have been described through preferred embodiments, and relevant persons can obviously modify or appropriately change and combine the method and application of this invention without departing from the content, spirit and scope of the present invention to achieve and apply the technology of the present invention.

[0066] The present invention will be further described below in conjunction with embodiments:

[0067] Example 1

[0068] A method for preparing a cyclohexapeptide compound 1, comprising:

[0069] Step S11: CTC resin (6.0 g, 6 mmol) was placed in a 100 mL solid phase synthesis reactor, and amino acid Fmoc-Hyp(tBu)-OH (4.9 g, 12 mmol) was added, and 60 mL of dichloromethane (DCM) was added, and then N,N-diisopropylethylamine (DIEA) (4.0 mL) was added, and the reaction was carried out at 25°C for 3 h, and 6.0 mL of methanol was added, and the reaction was carried out for 5 min. The resin was filtered, and the resin was washed twice with 60 mL of dichloromethane (DCM), 60 mL of methanol, and 60 mL of DMF. Then, 60 mL of 20% piperidine (Pip) / DMF solution was added to the solid phase synthesis reactor, and the reaction was stirred for 30 min. The deprotection solution was removed by suction filtration, and then the resin was washed 6 times with 60 mL of DMF solution, and dried for use.

[0070] Step S12: Take Fmoc-Pro-OH (5.1 g, 15 mmol) and HOBt (2.03 g, 15 mmol) in a 100 mL beaker, cool to 2°C to 8°C, add 25 mL of DMF solution and DIC (2.3 mL, 15 mmol) and let it react for 10 min to 20 min. Add the solution in the 100 mL beaker into a 100 mL solid phase synthesis reactor and stir to react for 1.5 h. After the reaction is completed, wash the resin three times with 60 mL of DMF solution each time. After washing, add 60 mL of 20% Pip / DMF solution and stir to react for 30 min. Filter to remove the deprotection solution, then wash 6 times with 60 mL of DMF solution and drain for use.

[0071] Step S13: Take Fmoc-Gly-OH (4.5 g, 15 mmol) and HOBt (2.03 g, 15 mmol) in a 100 mL beaker, cool to 2°C to 8°C, add 25 mL of DMF solution and DIC (2.3 mL, 15 mmol) and let it react for 10 min to 20 min. Add the solution in the 100 mL beaker into a 100 mL solid phase synthesis reactor and stir to react for 1.5 h. After the reaction is completed, wash the resin three times with 60 mL of DMF solution each time. After washing, add 60 mL of 20% Pip / DMF solution and stir to react for 30 min. Filter to remove the deprotection solution, then wash 6 times with 60 mL of DMF solution and drain for use.

[0072] Step S14: Take Fmoc-Hyp(tBu)-OH (6.1 g, 15 mmol) and HOBt (2.03 g, 15 mmol) in a 100 mL beaker, cool to 2°C to 8°C, add 25 mL of DMF solution and DIC (2.3 mL, 15 mmol) and let it react for 10 min to 20 min. Add the solution in the 100 mL beaker into a 100 mL solid phase synthesis reactor and stir to react for 1.5 h. After the reaction is completed, wash the resin three times with 60 mL of DMF solution each time. After washing, add 60 mL of 20% Pip / DMF solution and stir to react for 30 min. Filter to remove the deprotection solution, then wash 6 times with 60 mL of DMF solution and drain for standby use.

[0073] Step S15: Take Fmoc-Pro-OH (5.1 g, 15 mmol) and HOBt (2.03 g, 15 mmol) in a 100 mL beaker, cool to 2°C to 8°C, add 25 mL of DMF solution and DIC (2.3 mL, 15 mmol) and let it react for 10 min to 20 min. Add the solution in the 100 mL beaker into a 100 mL solid phase synthesis reactor and stir to react for 1.5 h. After the reaction is completed, wash the resin three times with 60 mL of DMF solution each time. After washing, add 60 mL of 20% Pip / DMF solution and stir to react for 30 min. Filter to remove the deprotection solution, then wash 6 times with 60 mL of DMF solution and drain for use.

[0074] Step S16: Take Fmoc-Gly-OH (4.5g, 15mmol), HOBt (2.03g, 15mmol) in a 100mL beaker, cool to 2°C to 8°C, add 25mL of DMF solution, DIC (2.3mL, 15mmol) and let stand for 10min to 20min, and add the solution in the 100mL beaker to a 100mL solid phase synthesis reactor, stir and react for 1.5h, the reaction is complete, the resin is washed three times with DMF solution, 60mL each time, after washing, add 60mL of 20% Pip / DMF solution, stir and react for 30min, filter to remove the deprotection solution, and then wash 6 times with 60mL of DMF solution, 2 times with 60mL of methanol, 2 times with 60mL of DCM solution, and 2 times with 60mL of methanol. Vacuum drying to obtain the peptide resin of H-Gly-Pro-Hyp(tBu)-Gly-Pro-Hyp(tBu)-CTC resin.

[0075] Step S2: Add 200 mL of 1% trifluoroacetic acid (TFA) / DCM solution to the peptide resin of H-Gly-Pro-Hyp(tBu)-Gly-Pro-Hyp(tBu)-CTC resin, stir and react at 30°C for 30 minutes, filter, remove the resin, and obtain a filtrate. The filtrate is drained to obtain the fully protected peptide H-Gly-Pro-Hyp(tBu)-Gly-Pro-Hyp(tBu)-OH.

[0076] Step S3: Dissolve the fully protected polypeptide H-Gly-Pro-Hyp(tBu)-Gly-Pro-Hyp(tBu)-OH in 1.0 L of dichloromethane (DCM), add DIC (1.5 mL, 10 mmol), HOBt (1.4 g, 10 mmol), and DIEA (1.7 mL, 10 mmol), and stir the reaction at 30°C for 12 to 16 hours to form Cyclo(Gly-Pro-Hyp(tBu)-Gly-Pro-Hyp(tBu)).

[0077] Step S4: The Cyclo(Gly-Pro-Hyp(tBu)-Gly-Pro-Hyp(tBu)) solid was cut with TFA / TIS / H2O=90 / 5 / 5 (50 mL) for 2.5 hours, the cutting solution was added to 500 mL of tert-butyl methyl ether (2°C~8°C) solution, a white solid was precipitated, and the mixture was centrifuged to obtain a white solid crude peptide. The white solid crude peptide was dried under vacuum to obtain a crude peptide powder Cyclo(Gly-Pro-Hyp-Gly-Pro-Hyp).

[0078] Step S5: The crude peptide powder Cyclo (Gly-Pro-Hyp-Gly-Pro-Hyp) was purified by reverse phase C18 preparative chromatography and freeze-dried to obtain cyclohexapeptide compound 1: Cyclo (Gly-Pro-Hyp-Gly-Pro-Hyp), i.e., the cyclohexapeptide compound shown in formula I-1. Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.64-–4.60(m,2H),4.40-–4. 35(m,2H),4.32(ddd,J=7.0,5.3,0.7Hz,2H),3.99(dd,J=15.8,6.0Hz,2H),3 .88(dd,J=15.8,6.0Hz,2H),3.71-–3.59(m,6H),3.48(d,J=5.7Hz,2H),3.45 -–3.38(m,2H),2.10-–2.03(m,2H),2.03-–1.93(m,4H),1.93-–1.86(m,6H).C24 H 34 N6O8,[M+H] + 535.25.

[0079]

[0080] Example 2

[0081] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH to obtain cyclohexapeptide compound 2 shown in formula I-2: Cyclo (b-Ala-Pro-Hyp-Gly-Pro-Hyp). Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ7.38(t,J=5.6Hz,1H),6.82(t,J=5.9Hz,1H),4.64–-4.57(m,2H),4.41–-4.34(m,2H), 4.31(dtd,J=6.5,5.6,0.9Hz,2H),3.99(dd,J=15.7,6.0Hz,1H),3.88(dd,J=15.7,6.0Hz,1H),3.70-–3.59(m, 5H),3.55(dt,J=12.3,4.2Hz,1H),3.47(dd,J=15.5,5.8Hz,3H),3.44–3.37(m,3H),2.56(dt,J=14.8,6.1Hz,1 H),2.48(dt,J=14.8,6.1Hz,1H),2.17--2.09(m,2H),2.09--1.92(m,5H),1.89(dtt,J=6.5,5.0,3.4Hz,5H).C 25 H 36 N6O8,[M+H] + 549.26.

[0082]

[0083] Example 3

[0084] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH to obtain cyclohexapeptide compound 3 shown in formula I-3: Cyclo(Gly-Hyp-Hyp-Gly-Pro-Hyp). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.65-–4.60(m,1H),4.57(ddd,J=7 .7,5.9,0.7Hz,1H),4.45-–4.35(m,3H),4.32(ddt,J=7.1,5.3,0.9Hz,2H),3.97(d dd,J=17.8,15.8,6.0Hz,2H),3.88(dd,J=15.9,5.9Hz,2H),3.67-–3.59(m,7H),3. 48(d,J=5.7Hz,3H),3.45--3.38(m,1H),2.13--1.93(m,7H),1.93--1.85(m,3H).C 24 H 34 N6O9,[M+H] + 551.25.

[0085]

[0086] Example 4

[0087] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-F)-OH to obtain cyclohexapeptide compound 4 shown in formula I-4: Cyclo(Gly-Pro-Pro(4-F)-Gly-Pro-Hyp). Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.91–4.77(m,1H),4.66–-4.59(m,2H),4.51(ddd ,J=6.6,4.8,0.8Hz,1H),4.41--4.34(m,1H),4.34--4.28(m,1H),3.99(dd,J=15.8,6.0Hz,2H), 3.88(dd,J=15.8,6.0Hz,2H),3.80--3.71(m,1H),3.71--3.68(m,1H),3.68--3.59(m,4H),3.48 (d,J=5.7Hz,1H),3.45–3.38(m,2H),2.47–2.29(m,2H),2.10–1.95(m,5H),1.95–1.86(m,5H).C 24 H 33 FN6O7,[M+H] + 537.25.

[0088]

[0089] Example 5

[0090] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-OMe)-OH to obtain cyclohexapeptide compound 5 shown in formula I-5: Cyclo(Gly-Pro-Pro(4-OMe)-Gly-Pro-Hyp). Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.65–4.59(m,2H),4.41–4.34(m,2H),4.32 (ddd,J=7.0,5.3,0.7Hz,1H),4.26–4.20(m,1H),3.99(dd,J=15.8,6.0Hz,2H),3.88(dd,J =15.8,6.0Hz,2H),3.75–3.60(m,6H),3.48(d,J=5.7Hz,1H),3.45–3.38(m,2H),3.19(d,J =1.5Hz,3H),2.25(ddd,J=12.4,5.8,4.9Hz,1H),2.10--1.95(m,5H),1.95--1.86(m,6H).C 25 H 36 N6O8,[M+H] + 549.27.

[0091]

[0092] Example 6

[0093] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro-OH to obtain cyclohexapeptide compound 6 shown in formula I-6: Cyclo(Gly-Pro-Pro-Gly-Pro-Hyp). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ6.82(t,J=5.9Hz,1H),6.73(t,J=6.0Hz,1H),4.61(dt,J=6 .1,4.3Hz,2H),4.47(dd,J=5.4,3.6Hz,1H),4.41--4.35(m,1H),4.34--4.30(m,1H ),3.99(dd,J=15.8,6.0Hz,2H),3.88(dd,J=15.8,6.0Hz,2H),3.69-–3.56(m,5H), 3.54--3.47(m,2H),3.45--3.38(m,2H),2.10--1.95(m,6H),1.95--1.82(m,8H).C 24 H 34 N6O7,[M+H] + 519.25.

[0094]

[0095] Example 7

[0096] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro-OH, and Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro-OH, and step S4 is not required, to obtain cyclohexapeptide compound 7 shown in formula I-7: Cyclo(Gly-Pro-Pro-Gly-Pro-Pro). Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ6.73(t,J=6.0Hz,2H),4.64–-4.58(m,2H),4.47(dd,J=5.4,3.7Hz,2H),3.99(dd,J=15.7,6.0Hz,2H),3.88(dd,J=15.7,6.0Hz, 2H),3.68–-3.56(m,4H),3.56––3.48(m,2H),3.45––3.38(m,2H),2.12–-2. 02(m,4H),1.98(dddd,J=11.5,7.8,5.5,3.8Hz,2H),1.94-–1.82(m,10H).C 24 H 34 N6O6,[M+H] + 503.26.

[0097]

[0098] Example 8

[0099] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Pro-OH in step S12 is replaced by Fmoc-Hyp(tBu)-OH, and Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH to obtain cyclohexapeptide compound 8 shown in formula I-8: Cyclo(Gly-Hyp-Hyp-Gly-Hyp-Hyp). Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.57(ddd,J=7.7,5.9,0.7Hz,2H),4.44–-4.35(m,2H),4.32(ddd,J=7.1,5.4,0.8Hz,2H),3.95(dd ,J=15.8,6.0Hz,2H),3.88(dd,J=15.7,6.0Hz,2H),3.70-–3.59(m,8H),3 .48(d,J=5.7Hz,4H),2.10(dt,J=12.2,6.0Hz,2H),2.06-–1.93(m,8H).C 24 H 34 N6O 10 ,[M+H] + 567.24.

[0100]

[0101] Example 9

[0102] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH, to obtain cyclohexapeptide compound 9 shown in formula I-9: Cyclo(b-Ala-Hyp-Hyp-Gly-Pro-Hyp). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.38(t,J=5.6Hz,1H),6.82(t,J=6.0Hz,1H),4.66–-4.59(m,1H),4.51(d dd,J=7.9,6.0,0.8Hz,1H),4.46-–4.35(m,3H),4.31(dddd,J=7.3,6.5,5.4,0.7Hz,2H),3.99(dd ,J=15.8,6.0Hz,1H),3.88(dd,J=15.7,6.0Hz,1H),3.71-–3.58(m,7H),3.52-–3.35(m,6H),2.58 (dt,J=14.8,6.1Hz,1H),2.49(dt,J=14.8,6.0Hz,1H),2.14--1.94(m,7H),1.94--1.86(m,3H).C 25 H 36 N6O9,[M+H] + 565.26.

[0103]

[0104] Example 10

[0105] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Pro-OH in step S12 is replaced by Fmoc-Hyp(tBu)-OH, Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH to obtain cyclohexapeptide compound 10 shown in formula I-10: Cyclo(b-Ala-Hyp-Hyp-Gly-Hyp-Hyp). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.38(t,J=5.6Hz,1H),6.82(t,J=6.0Hz,1H),4.57(ddd,J=7.7,5.9,0.8Hz,1H),4.51( dddd,J=7.9,6.0,0.8Hz,1H),4.45–-4.35(m,4H),4.31(dddd,J=7.3,6.4,5.4,0.7Hz,2H),3.95(dd,J=15.8,6. 0Hz,1H),3.88(dd,J=15.7,6.0Hz,1H),3.71--3.59(m,8H),3.49(t,J=5.5Hz,4H),3.47--3.36(m,2H),2.58(d t,J=14.8,6.1Hz,1H),2.49(dt,J=14.8,6.0Hz,1H),2.10(dtd,J=12.2,6.1,2.3Hz,2H),2.06-–1.93(m,6H).C 25 H 36 N6O 10 ,[M+H] + 581.26.

[0106]

[0107] Embodiment 11

[0108] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-F)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH to obtain cyclohexapeptide compound 11 shown in formula I-11: Cyclo(b-Ala-Pro-Pro(4-F)-Gly-Pro-Hyp). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.38(t,J=5.6Hz,1H),6.82(t,J=5.9Hz,1H),4.91-–4.76(m,1H),4.64–-4.58(m ,2H),4.51(ddd,J=6.6,4.8,0.8Hz,1H),4.41–-4.35(m,1H),4.33-–4.28(m,1H),3.99(dd,J=15.7,6.0Hz ,1H),3.88(dd,J=15.7,6.0Hz,1H),3.80--3.67(m,2H),3.67--3.59(m,3H),3.55(dt,J=12.3,4.2Hz,1H ),3.51--3.36(m,5H),2.61--2.45(m,2H),2.45--2.30(m,2H),2.17--2.03(m,3H),2.03--1.85(m,7H).C 25 H 35 FN6O7,[M+H] + 551.26.

[0109]

[0110] Example 12

[0111] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-OMe)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH to obtain cyclohexapeptide compound 12 shown in formula I-12: Cyclo(b-Ala-Pro-Pro(4-OMe)-Gly-Pro-Hyp). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.38(t,J=5.6Hz,1H),6.82(t,J=6.0Hz,1H),4.64-–4.57(m,2H),4.37(dddd,J=7.7,7.0,4 .2,1.3Hz,2H),4.31(ddd,J=7.1,5.5,0.8Hz,1H),4.26--4.20(m,1H),3.99(dd,J=15.8,6.0Hz,1H),3.88(dd,J=15. 8,6.0Hz,1H),3.75–3.53(m,6H),3.51–-3.36(m,5H),3.19(d,J=1.5Hz,3H),2.56(dt,J=14.8,6.1Hz,1H),2.48(dt, J=14.8,6.1Hz,1H),2.25(ddd,J=12.4,5.8,4.9Hz,1H),2.16-–2.09(m,2H),2.09–1.93(m,4H),1.93–1.86(m,5H).C 26 H 38 N6O8,[M+H] + 563.28.

[0112]

[0113] Example 13

[0114] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-F)-OH, Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-F)-OH, Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH, and step S4 is not required, to obtain cyclohexapeptide compound 13 shown in formula I-13: Cyclo(b-Ala-Pro-Pro(4-F)-Gly-Pro-Pro(4-F)). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.38(t,J=5.6Hz,1H),6.82(t,J=5.9Hz,1H),4.93–-4.76(m,2H),4.61(dd d,J=10.5,6.1,4.5Hz,2H),4.54–-4.46(m,2H),3.99(dd,J=15.7,6.0Hz,1H),3.88(dd,J=15.8,6.0 Hz,1H),3.80–3.67(m,4H),3.67–3.60(m,1H),3.55(dt,J=12.3,4.2Hz,1H),3.51-–3.36(m,4H),2. 59-–2.45(m,2H),2.45-–2.30(m,5H),2.17–-2.09(m,2H),2.09-–2.02(m,1H),1.94–1.86(m,5H).C 25 H 34 F2N6O6,[M+H] + 553.26.

[0115]

[0116] Embodiment 14

[0117] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-OMe)-OH, Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-OMe)-OH, Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH, and step S4 is not required, to obtain cyclohexapeptide compound 14 shown in formula I-14: Cyclo(b-Ala-Pro-Pro(4-OMe)-Gly-Pro-Pro(4-OMe)). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.38(t,J=5.6Hz,1H),6.82(t,J=6.0Hz,1H),4.64–4.58(m,2H),4.35(dddd,J=7.5,6.9 ,4.9,0.8Hz,2H),4.23(tddd,J=6.7,4.1,1.6,1.0Hz,2H),3.99(dd,J=15.8,6.0Hz,1H),3.88(dd,J=15.8,6.0H z,1H),3.72(ddd,J=12.4,4.2,0.7Hz,2H),3.69--3.60(m,3H),3.59--3.52(m,1H),3.51--3.36(m,4H),3.19(d ,J=1.5Hz,6H),2.59–2.45(m,2H),2.25(ddd,J=12.3,5.9,5.0Hz,2H),2.16-–2.01(m,5H),1.95–-1.85(m,5H).C 27 H 40 N6O8,[M+H] + 577.30.

[0118]

[0119] Embodiment 15

[0120] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that Fmoc-Gly-OH in step S13 is replaced by Fmoc-b-Ala-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH to obtain cyclohexapeptide compound 15 shown in formula I-15: Cyclo (b-Ala-Pro-Hyp-b-Ala-Pro-Hyp). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.34(t,J=5.6Hz,2H),4.60(dd,J=5.9,4.6Hz,2H),4.37(ddddd, J=7.3,6.3,5.4,3.2,0.8Hz,2H),4.31(ddd,J=7.1,5.5,0.8Hz,2H),3.70–-3.59(m,4H),3 .59-–3.53(m,2H),3.51-–3.44(m,4H),3.44–3.36(m,4H),2.56(dt,J=14.8,6.1Hz,2H), 2.48(dt,J=14.8,6.0Hz,2H),2.16–2.08(m,4H),2.07–-1.94(m,4H),1.92–1.85(m,4H).C 26 H 38 N6O8,[M+H] + 563.28.

[0121]

[0122] Example 16

[0123] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Pro-OH in step S12 is replaced by Fmoc-Hyp(tBu)-OH, Fmoc-Gly-OH in step S13 is replaced by Fmoc-b-Ala-OH, Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH to obtain cyclohexapeptide compound 16 shown in formula I-16: Cyclo

[0124] (b-Ala-Hyp-Hyp-b-Ala-Hyp-Hyp). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.34(t,J=5.6Hz,2H),4.51(ddd,J=7.9,6.0,0.8Hz,2H),4.45–4. 34(m,4H),4.31(ddd,J=7.1,5.5,0.8Hz,2H),3.67(ddd,J=12.4,5.1,2.5Hz,4H),3.62(dd d,J=12.4,4.4,2.2Hz,4H),3.49(t,J=5.5Hz,4H),3.47–3.36(m,4H),2.58(dt,J=14.8,6. 1Hz,2H),2.49(dt,J=15.0,6.1Hz,2H),2.11(dt,J=12.4,6.2Hz,2H),2.05–1.94(m,6H).C 26 H 38 N6O 10 ,[M+H] + 595.27.

[0125]

[0126] Embodiment 17

[0127] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-F)-OH, Fmoc-Gly-OH in step S13 is replaced by Fmoc-b-Ala-OH, Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-F)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH, and step S4 is not required, thereby obtaining cyclohexapeptide compound 17 shown in formula I-17: Cyclo(b-Ala-Pro-Pro(4-F)-b-Ala-Pro-Pro(4-F)). Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ7.34(t,J=5.6Hz,2H),4.91–-4.76(m,2H),4.60(dd,J=5.9,4.6Hz,2H),4.49(ddd,J=6.4,5.0,0.8Hz,2H),3.78–3.72(m, 2H),3.72–3.65(m,2H),3.58–3.52(m,2H),3.51–-3.36(m,6H),2.60-–2 .45(m,4H),2.45–2.30(m,4H),2.18-–2.07(m,4H),1.92-–1.86(m,4H).C26 H 36 F2N6O6,[M+H] + 567.27.

[0128]

[0129] Embodiment 18:

[0130] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-OMe)-OH, Fmoc-Gly-OH in step S13 is replaced by Fmoc-b-Ala-OH, Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-OMe)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH, and step S4 is not required, to obtain cyclohexapeptide compound 18 shown in formula I-18: Cyclo(b-Ala-Pro-Pro(4-OMe)-b-Ala-Pro-Pro(4-OMe)). Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ7.34(t,J=5.6Hz,2H),4.60(dd,J=5.9,4.6Hz,2H),4.34(ddd,J=7.0,4.9,0.8Hz,2 H),4.26--4.20(m,2H),3.72(dd,J=12.2,4.2Hz,2H),3.66(dd,J=12.6,2.7Hz,2H),3.58--3.53(m,2H),3. 51-–3.36(m,6H),3.19(d,J=1.5Hz,6H),2.56(dt,J=14.8,6.1Hz,2H),2.48(dt,J=14.8,6.0Hz,2H),2.25( ddd,J=12.5,5.8,4.9Hz,2H),2.16–-2.09(m,4H),2.04(ddd,J=12.4,6.8,4.0Hz,2H),1.92–1.86(m,4H).C 28 H 42 N6O8,[M+H] + 591.31.

[0131]

[0132] Embodiment 19:

[0133] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-F)-OH, and Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-F)-OH, and step S4 is not required, to obtain cyclohexapeptide compound 19 shown in formula I-19: Cyclo(Gly-Pro-Pro(4-F)-Gly-Pro-Pro(4-F)). Its NMR data are as follows: 1 H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.91-–4.76(m,2H),4.65–4.59(m,2H),4.51(ddd,J=6.6,4.9,0.8Hz,2H),3.99(dd,J=15.8,6 .0Hz,2H),3.88(dd,J=15.8,6.0Hz,2H),3.80–3.60(m,6H),3.46–-3.37(m,2H),2.46-–2.30(m,4H),2.10–-2.01(m,2H),1.94–1.85(m,6H).C 24 H 32 F2N6O6,[M+H] + 539.24.

[0134]

[0135] Embodiment 20

[0136] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-OMe)-OH, and Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-OMe)-OH, and step S4 is not required, to obtain cyclohexapeptide compound 20 shown in formula I-20: Cyclo(Gly-Pro-Pro(4-OMe)-Gly-Pro-Pro(4-OMe)). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.65-–4.58(m,2H),4.36(ddd,J=6.8,5.0,0.8Hz,2 H),4.23(dddt,J=4.9,4.1,3.3,1.5Hz,2H),3.99(dd,J=15.8,6.0Hz,2H),3.88(dd,J=15.8,6.0Hz ,2H),3.72(dd,J=12.5,4.2Hz,2H),3.69--3.65(m,2H),3.65--3.61(m,2H),3.44--3.38(m,2H),3 .19(d,J=1.5Hz,6H),2.25(ddd,J=12.5,5.8,5.0Hz,2H),2.10--2.00(m,4H),1.95--1.85(m,6H).C 26 H 38 N6O8,[M+H] + 563.28.

[0137]

[0138] Embodiment 21

[0139] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-F)-OH, Fmoc-Pro-OH in step S12 is replaced by Fmoc-Hyp(tBu)-OH, Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-F)-OH, and Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH to prepare cyclohexapeptide compound 21 shown in formula I-21: Cyclo(Gly-Hyp-Pro(4-F)-Gly-Hyp-Pro(4-F)). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.91-–4.77(m,2H),4.57(ddd,J=7.7,5.9,0.7Hz ,2H),4.51(ddd,J=6.6,4.9,0.8Hz,2H),4.44-–4.38(m,2H),3.95(dd,J=15.8,6.0Hz,2H),3.88 (dd,J=15.7,6.0Hz,2H),3.78–3.71(m,2H),3.71–3.67(m,2H),3.67-–3.61(m,4H),3.48(d,J=5 .7Hz,2H),2.46–2.29(m,4H),2.10(dt,J=12.2,6.0Hz,2H),2.01(ddd,J=12.2,7.8,4.5Hz,2H).C 24 H 32 F2N6O8,[M+H] + 571.23.

[0140]

[0141] Embodiment 22

[0142] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-OMe)-OH, Fmoc-Pro-OH in step S12 is replaced by Fmoc-Hyp(tBu)-OH, Fmoc-Hyp(tBu)-OH in step S14 is replaced by Fmoc-Pro(4-OMe)-OH, and Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH to prepare cyclohexapeptide compound 22 shown in formula I-22: Cyclo(Gly-Hyp-Pro(4-OMe)-Gly-Hyp-Pro(4-OMe)). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ6.82(t,J=6.0Hz,2H),4.57(ddd,J=7.7,5.9,0.7Hz,2H),4.44-–4.38(m,2H),4 .35(ddd,J=7.0,5.0,0.8Hz,2H),4.23(dtt,J=5.8,2.6,1.6Hz,2H),3.95(dd,J=15.8,6.0Hz,2H),3.88( dd,J=15.7,6.0Hz,2H),3.72(dd,J=12.2,4.2Hz,2H),3.69–-3.61(m,6H),3.48(d,J=5.7Hz,2H),3.19( d,J=1.5Hz,6H),2.25(ddd,J=12.5,5.8,4.9Hz,2H),2.10(dt,J=12.2,6.0Hz,2H),2.07-–1.97(m,4H).C 26 H 38 N6O 10 ,[M+H] + 595.27.

[0143]

[0144] Embodiment 23

[0145] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-F)-OH, Fmoc-Pro-OH in step S12 is replaced by Fmoc-Hyp(tBu)-OH, Fmoc-Gly-OH in step S13 is replaced by Fmoc-b-Ala-OH, and Fmoc-Hyp( In step S14, tBu)-OH is replaced by Fmoc-Pro(4-F)-OH, Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH to obtain cyclohexapeptide compound 23 shown in formula I-23: Cyclo(b-Ala-Hyp-Pro(4-F)-b-Ala-Hyp-Pro(4-F)). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.34(t,J=5.6Hz,2H),4.91--4.76(m,2H),4.55--4.46(m,4H),4.45 -–4.38(m,2H),3.78–-3.71(m,2H),3.71-–3.65(m,4H),3.62(dd,J=12.4,4.2Hz,2H),3.49( d,J=5.7Hz,2H),3.48--3.36(m,4H),2.58(dt,J=14.8,6.1Hz,2H),2.49(dt,J=15.0,6.1Hz, 2H),2.45–-2.30(m,4H),2.11(dt,J=12.3,6.2Hz,2H),1.99(ddd,J=12.2,7.7,4.4Hz,2H).C 26 H 36 F2N6O8,[M+H] + 599.27.

[0146]

[0147] Embodiment 24

[0148] The raw materials and preparation method used in this example are basically the same as those in Example 1, except that: Fmoc-Hyp(tBu)-OH in step S11 is replaced by Fmoc-Pro(4-OMe)-OH, Fmoc-Pro-OH in step S12 is replaced by Fmoc-Hyp(tBu)-OH, Fmoc-Gly-OH in step S13 is replaced by Fmoc-b-Ala-OH, and Fmoc-Hyp(tB u)-OH is replaced by Fmoc-Pro(4-OMe)-OH, Fmoc-Pro-OH in step S15 is replaced by Fmoc-Hyp(tBu)-OH, and Fmoc-Gly-OH in step S16 is replaced by Fmoc-b-Ala-OH to obtain cyclohexapeptide compound 24 shown in formula I-24: Cyclo(b-Ala-Hyp-Pro(4-OMe)-b-Ala-Hyp-Pro(4-OMe)). Its NMR data are as follows: 1H NMR(500MHz,MeOH-d4)δ7.34(t,J=5.6Hz,2H),4.51(ddd,J=7.9,6.1,0.8Hz,2H),4.44–-4.38(m,2H) ,4.34(ddd,J=7.0,5.0,0.8Hz,2H),4.23(dtt,J=5.8,2.6,1.6Hz,2H),3.74-–3.59(m,8H),3.49(d,J= 5.7Hz,2H),3.48--3.36(m,4H),3.19(d,J=1.5Hz,6H),2.58(dt,J=14.8,6.1Hz,2H),2.49(dt,J=15. 0,6.1Hz,2H),2.25(ddd,J=12.5,5.8,4.9Hz,2H),2.11(dt,J=12.4,6.2Hz,2H),2.07-–1.95(m,4H).C 28 H 42 N6O 10 ,[M+H] + 623.31.

[0149]

[0150] Test Example 1 Cytotoxicity Detection

[0151] Human keratinocytes (HaCaT) were cultured at 2×10 4 The cells were cultured in a 96-well cell culture plate at a density of 1 / 96well and placed in a 37°C, 5% CO2 incubator. After 48 hours, the culture medium was discarded, and the cells were washed once with 1×PBS. The cells were treated with bioactive substances and the cyclohexapeptide compounds prepared in Examples 1 to 7 at different concentrations were added. After 24 hours of treatment, the culture medium was removed, 100 μL of culture medium containing 10% cck-8 solution was added, and the cells were placed in an incubator for 1 hour. The absorbance at 450 nm was detected with an enzyme marker. At the same time, the solvent without the sample was used as a control group, and the cell survival rate was calculated according to the following formula:

[0152] Cell survival rate % = (absorbance of experimental group / absorbance of control group) × 100%.

[0153] Results Figure 1 and Table 1, Figure 1 Table 1 shows the effect of the cyclohexapeptide prepared in Example 1 on human keratinocytes, and Table 2 shows the effect of the cyclohexapeptide prepared in Examples 1 to 7 on keratinocytes. Figure 1 As can be seen from Table 1, the cyclohexapeptides prepared in Examples 1 to 7 have no significant cytotoxicity to human keratinocytes.

[0154] Table 1 Effect of cyclohexapeptides prepared in Examples 1 to 7 on keratinocytes

[0155] Relative cell viability (%) 1000 μg / mL 100 μg / mL 10 μg / mL 1 μg / mL Example 1 85.59 96.84 95.73 99.35 Example 2 84.32 95.44 95.21 98.45 Example 3 83.44 94.22 92.46 98.74 Example 4 81.36 92.82 94.54 96.26 Example 5 80.68 95.48 93.86 98.52 Example 6 82.31 93.54 95.12 94.88 Example 7 82.45 93.62 93.54 96.84

[0156] Test Example 2 Firming Effect

[0157] Detection reagents: Human type I collagen α1 (COL1α1) enzyme-linked immunosorbent assay kit (catalog number: E-EL-H0869, Elabscience, China); human type III collagen (COL3) enzyme-linked immunosorbent assay kit (catalog number: E-EL-H6049, Elabscience, China); human type IV collagen (COL4) enzyme-linked immunosorbent assay kit (catalog number: E-EL-H0178, Elabscience, China).

[0158] Test cells: fibroblasts (HFF-1), obtained from the Cell Bank of Type Culture Collection Committee of the Chinese Academy of Sciences (Cell Bank of the Chinese Academy of Sciences).

[0159] Detection method: Fibroblasts (HFF-1) were cultured at 5×10 4 / 6wel was cultured in a 6-well cell culture plate and placed in a 37°C, 5% CO2 incubator. After 48 hours, the culture medium was discarded, washed once with 1×PBS, and 500μg / mL of the cyclohexapeptide prepared in Examples 1 to 7 was added. The volume of each well was 2mL. A normal control group and an experimental group were set, with three biological replicates for each group. Incubate in a cell culture incubator for 48 hours. Then, the cell supernatant was aspirated, centrifuged at 1000g, 4°C for 20 minutes, the supernatant was collected, and the precipitate was discarded. Follow the instructions of the collagen ELISA kit for subsequent operations.

[0160] Results Figure 2 , Figure 3 , Figure 4 , Table 2, Table 3 and Table 4, Figure 2 The expression level of type I collagen in the cyclohexapeptide experimental group prepared in Example 1, Figure 3 The expression level of type III collagen in the cyclohexapeptide experimental group prepared in Example 1, Figure 4 Table 2 is the expression of type I collagen in the cyclohexapeptide experimental group prepared in Example 1, Table 3 is the expression of type III collagen in the cyclohexapeptide experimental group prepared in Examples 1 to 7, and Table 4 is the expression of type IV collagen in the cyclohexapeptide experimental group prepared in Examples 1 to 7. It can be seen that after the treatment with the cyclohexapeptide prepared in Examples 1 to 7, the expression of type I, type III and type IV collagen increased significantly, which shows that the cyclohexapeptide has an excellent firming effect.

[0161] Table 2 Expression of type I collagen in the cyclic hexapeptide experimental group prepared in Examples 1 to 7

[0162] Group Type I collagen concentration (ng / mL) P-value Blank control 6.07±0.1 / Example 1 6.82±0.11 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 2 6.37±0.02 <![CDATA[ ＊ P<0.5]]> Example 3 6.44±0.06 <![CDATA[ ＊ P<0.5]]> Example 4 6.37±0.05 <![CDATA[ ＊ P<0.5]]> Example 5 6.38±0.11 <![CDATA[ ＊ P<0.5]]> Example 6 6.41±0.1 <![CDATA[ ＊ P<0.5]]> Example 7 6.38±0.08 <![CDATA[ ＊ P<0.5]]>

[0163] Table 3 Expression of type III collagen in the cyclic hexapeptide experimental group prepared in Examples 1 to 7

[0164] Group Type III collagen concentration (ng / mL) P-value Blank control 0.86±0.18 / Example 1 2.19±0.05 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 2 1.83±0.05 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 3 1.58±0.2 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 4 1.68±0.03 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 5 1.78±0.03 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 6 1.68±0.03 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 7 1.82±0.02 <![CDATA[ ＊＊＊＊ P<0.0001]]>

[0165] Table 4 Expression of type IV collagen in the cyclohexapeptide experimental group prepared in Examples 1 to 7

[0166] Group Type IV collagen concentration (ng / mL) P-value Blank control 6.97±0.23 / Example 1 15.15±0.36 <![CDATA[ ＊＊＊＊ P<0.0001 <!-- 22 -->]]> Example 2 13.64±0.32 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 3 12.26±0.41 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 4 11.86±0.48 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 5 13.41±0.15 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 6 12.09±0.63 <![CDATA[ ＊＊＊＊ P<0.0001]]> Example 7 11.88±0.32 <![CDATA[ ＊＊＊＊ P<0.0001]]>

[0167] Embodiments 25 to 28

[0168] According to the formula shown in Table 5, the essence milk was prepared as follows:

[0169] (1) After dissolving and dispersing cyclocollagen hexapeptide, Tremella polysaccharide, sodium hyaluronate, polyol, thickener and water at 80° C., the mixture is kept warm at 80° C., wherein the polyol is selected from glycerol, 1,3-propylene glycol and 1,2-hexanediol in a mass ratio of 4:5:1, and the thickener is selected from acrylic acid (ester) / C10-30 alkyl acrylate cross-linked polymer.

[0170] (2) After the emulsifier and the oil are stirred, dissolved and dispersed at 80° C., the temperature is maintained at 80° C., wherein the emulsifier is selected from cetearyl alcohol and polyglyceryl-10 stearate in a mass ratio of 1:2, and the oil is selected from caprylic / capric glycerides, squalane, and avocado butter in a mass ratio of 2:2:1.

[0171] (3) The solution obtained in step (2) was mixed with the solution obtained in step (1), homogenized for 20 minutes, emulsified evenly, and cooled.

[0172] (4) The solution obtained in step (3) is cooled to below 40° C., phenoxyethanol as a preservative is added, and stirred evenly to obtain an essence lotion.

[0173] Table 5 Formula of the essence milk prepared in Examples 25 to 28

[0174]

[0175] Test Example 3 Moisturizing Efficacy Test

[0176] Twenty healthy volunteers aged 20 to 50 years without allergic diseases were selected for the experiment at room temperature (25±2)℃ and humidity 40% to 60%. Before the experiment, the volunteers were required to be quiet for at least 30 minutes in this environment. After the environment reached equilibrium, three 3cm×3cm experimental areas were drawn on the left forearm and three 3cm×3cm experimental areas were drawn on the right forearm. The interval between each area was at least 1 to 2cm. The samples were smeared on both arms. The blank group samples, the samples of Example 25 and the samples of Example 26 were smeared on the left forearm, and the samples of Example 27 and the samples of Example 28 were smeared on the right forearm. The concentration was (2.0±0.1) mg / cm 2 The dose was applied to the test area. After the subjects used the essence lotion 0min, 30min, 2h, 4h, and 8h, the technicians used the skin moisture tester probe to test the moisture content of the test area, and the average value of the three measurements was taken as the test value.

[0177] The moisture rise rate is calculated according to the following formula:

[0178] Moisture increase rate = (moisture content after use - moisture content before use) / moisture content before use × 100%.

[0179] The results are shown in Tables 6 and Figure 5 Table 6 is a statistical table of the water content increase rate of the cyclocollagen hexapeptide essence milk. Figure 5 This is a statistical chart of the moisture increase rate of the cyclic collagen hexapeptide essence lotion.

[0180] Table 6 Statistical table of moisture increase rate of cyclic collagen hexapeptide essence milk

[0181] Example 30min(%) 2h(％) 4h(％) 8h(％) Blank Group 171.86 145.76 108.47 64.75 Embodiment 25 177.74 165.11 153.16 90.36 Embodiment 26 170.26 162.09 150.98 96.73 Embodiment 27 189.96 180.27 174.39 126.99 Embodiment 28 175.83 164.76 150.33 88.59

[0182] Compared with the experimental groups, the blank group had a P<0.05.

[0183] Moisturizing is the basis and key to healthy skin care, as shown in Table 6 and Figure 5 It can be seen that the cyclic collagen hexapeptide essence lotion can effectively moisturize and has a sustained moisturizing effect.

[0184] Test Example 4 Firming Effect Test

[0185] (1) Test principle

[0186] The firming effect biochemical method mainly characterizes the firming effect of the active substance by evaluating its inhibition rate on elastase. Elastase is the most important enzyme in the matrix metalloproteinase family, which is mainly synthesized and secreted by fibroblasts and can degrade elastin in the skin, leading to skin sagging.

[0187] (2) Instruments, equipment and reagents

[0188] 2.1 Instruments and equipment:

[0189] Analytical balance;

[0190] Full wavelength multifunctional microplate reader.

[0191] 2.2 Reagents and consumables:

[0192] Tris(hydroxymethyl)aminomethane (AR);

[0193] Concentrated hydrochloric acid (GR);

[0194] Elastase (30 U / mg);

[0195] EGCG (epigallocatechin gallate);

[0196] Substrate: Suc-AAPF-pNA (N-succinyl-propionyl-propionyl-prolyl-phenylalanine p-nitroanilide);

[0197] 2.3 Test substances

[0198] Sample: Dilute with pure water to a sample concentration of 1%;

[0199] Negative control: pure water;

[0200] Positive control: EGCG aqueous solution with a mass concentration of 0.1%.

[0201] (3) Test methods and results

[0202] A total of six groups of experiments were set up, including a positive control group, a negative control group, a blank group, Example 25, Example 26 and Example 27, with three parallels in each group. The experiment was carried out in a 96-well plate (356ul / well). First, 100ul of buffer, 30ul of sample (sample concentration is 1%) and 10ul of substrate were added and mixed, and then 30ul of enzyme solution (30U / mg) was added. The well plate was placed in a 37°C shaking water tank and the shaking mode was turned on. The shaking frequency was 200rpm; the amplitude was 25mm. After the solution inside reacted completely, the absorbance value of each group was measured at a wavelength of 410nm. Among them, the buffer solution was a buffer solution composed of 6.057g of tris(hydroxymethyl)aminomethane and 4.2ml of concentrated hydrochloric acid, diluted to 100ml with ultrapure water).

[0203] The elastase inhibition rate was calculated according to the following formula:

[0204]

[0205] Where A is the absorbance of the reaction solution containing the sample and the enzyme;

[0206] B is the absorbance of the reaction solution containing the sample but not the enzyme;

[0207] C is the absorbance of the reaction solution containing enzyme but not sample;

[0208] D is the absorbance of the reaction solution without sample and enzyme.

[0209] The results are shown in Tables 7 and Figure 6 Table 7 is a statistical table of the elastase inhibition rate of the essence milk provided in the examples of the present application. Figure 6 This is a statistical chart of the elastase inhibition rate of the essence milk provided in the examples of the present application.

[0210] Table 7 Statistical table of elastase inhibition rate of the essence milk provided in the examples of this application

[0211] Group Elastase inhibition rate % p-value Positive control group 72.255±0.505 ＜0.05 Negative control group 0±0.481 / Blank Group 50.322±0.503 ＜0.05 Embodiment 29 62.377±0.500 ＜0.05 Embodiment 30 68.101±0.504 ＜0.05 Embodiment 31 79.866±0.502 ＜0.05

[0212] The test results show that, under the premise of ensuring the effectiveness of the test reaction system (the elastase inhibition rate of the positive control is >50%), when the samples are the blank group, Example 29, Example 30, and Example 31, the elastase inhibition rates are 50.322%, 62.377%, 68.101%, and 79.866%, respectively, which are significantly different from those of the positive control (P<0.05), indicating that the samples have good firming effects.

[0213] The above are only preferred embodiments of the present invention. It should be pointed out that, for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A cosmetic composition comprising: 0.01 wt% to 0.5 wt% of a cyclohexapeptide compound; 0.1 wt% to 5 wt% of a thickener; 1wt% to 10wt% of polyol; 1wt% to 10wt% of emulsifier; 1wt% to 10wt% of oily raw material; 0.1 wt% to 5 wt% of a preservative; and the balance of water; The cyclohexapeptide compound has the structure of formula (I): In formula (I), x and y are independently selected from integers of 1 to 5; R 1 , R 2 , R 3 and R 4 Independently selected from H, hydroxy, halogen or C1-C6 alkoxy.

2. The cosmetic composition according to claim 1, characterized in that The cyclohexapeptide compound has a structure of formula I-1, formula I-2, formula I-3, formula I-4, formula I-5, formula I-6, formula I-7, formula I-8, formula I-9, formula I-10, formula I-11, formula I-12, formula I-13, formula I-14, formula I-15, formula I-16, formula I-17, formula I-18, formula I-19, formula I-20, formula I-21, formula I-22, formula I-23 or formula I-24:

3. The cosmetic composition according to claim 1 or 2, characterized in that: The invention also comprises 0.1wt% to 0.5wt% of Tremella polysaccharide and / or 0.05wt% to 0.5wt% of sodium hyaluronate.

4. The cosmetic composition according to claim 3, characterized in that The polyol is selected from at least one of glycerol, 1,3-propylene glycol, dipropylene glycol, 1,2-hexanediol and 1,2-pentanediol; The emulsifier is selected from at least one emulsifier having an HLB value of 8 to 15; The oily raw material is selected from a mixture of caprylic / capric glycerides, squalane and avocado butter; The preservative is selected from at least one of phenoxyethanol, p-hydroxyacetophenone and 1,2-hexanediol; The thickener is selected from ion-resistant thickeners.

5. The cosmetic composition according to claim 4, characterized in that The emulsifier is selected from one or more of cetearyl alcohol, polyglyceryl-10 stearate and cetearyl glucoside.

6. The cosmetic composition according to claim 4, characterized in that In the oily raw material, the mass ratio of caprylic / capric glycerides, squalane and avocado butter is 2-4:2-4:1-2.

7. The cosmetic composition according to claim 4, characterized in that The molecular weight of the sodium hyaluronate is 100KDa to 1000KDa.

8. The method for preparing the cosmetic composition according to any one of claims 1 to 7, comprising the following steps: Dissolving the cyclohexapeptide compound and the preservative respectively to obtain a cyclohexapeptide compound solution and a preservative solution; After the polyol, thickener and water are uniformly mixed, an aqueous phase solution is obtained; After the emulsifier and the oily raw material are uniformly mixed, an oil phase solution is obtained; The oil phase solution is added to the water phase solution and then homogenized to obtain an emulsion; The emulsion, the cyclohexapeptide compound solution and the preservative solution are uniformly mixed to obtain a cosmetic composition.

9. A composition comprising a cyclohexapeptide compound, Tremella polysaccharide and hyaluronic acid in a mass ratio of 0.01-0.5:0.1-0.5:0.05-0.5; The cyclohexapeptide compound has the structure of formula (I): In formula (I), x and y are independently selected from integers of 1 to 5; R 1 , R 2 , R 3 and R 4 Independently selected from H, hydroxy, halogen or C1-C6 alkoxy.

10. The composition according to claim 9, characterized in that The cyclohexapeptide compound has a structure of formula I-1, formula I-2, formula I-3, formula I-4, formula I-5, formula I-6, formula I-7, formula I-8, formula I-9, formula I-10, formula I-11, formula I-12, formula I-13, formula I-14, formula I-15, formula I-16, formula I-17, formula I-18, formula I-19, formula I-20, formula I-21, formula I-22, formula I-23 or formula I-24:

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