Hyaluronic acid modified tetrapeptide as well as preparation method and application thereof

Abstract

The invention discloses hyaluronic acid modified tetrapeptide as well as a preparation method and application thereof, and relates to the field of cosmetic peptides. The hyaluronic acid modified cosmetic peptide has a structural formula as shown in a formula (I): M-C (I), wherein M represents sodium hyaluronate, the structure of the sodium hyaluronate is shown in the formula (II), and in the formula (II), y is a natural number larger than or equal to 1; c represents a cosmetic peptide, the cosmetic peptide comprises a polypeptide with cosmetic and / or skin care efficacy or a derivative thereof, and the polypeptide comprises a dipeptide, a tripeptide, a tetrapeptide, a pentapeptide, a hexapeptide, a heptapeptide, an octapeptide or a nonapeptide; wherein M-terminal glucuronic acid is in open-loop linkage with amino in the C structure. The hyaluronic acid modified cosmetic peptide provided by the invention not only can maintain the property of sodium hyaluronate, but also can have the functionality of polypeptide, and the stability of the prepared novel compound is obviously improved, so that the hyaluronic acid modified cosmetic peptide has a wide application prospect in the field of cosmetics / skin care products.

Description

Technical Field

[0001] The present invention belongs to the field of cosmetic peptides, and in particular relates to a hyaluronic acid-modified tetrapeptide, a preparation method and uses thereof. Background Art

[0002] Hyaluronic acid is a macromolecular polysaccharide first isolated from the bovine vitreous humor. Its aqueous solution is transparent and glassy, ​​hence its name, hyaluronic acid. Hyaluronic acid is a natural biological substance found widely in tissues such as the vitreous humor, skin, umbilical cord, and synovial fluid. Early hyaluronic acid raw materials were primarily isolated and extracted from rooster combs. Due to cost and raw material limitations, this process was not widely adopted. Currently, hyaluronic acid for cosmetic and pharmaceutical applications is primarily produced through microbial fermentation. This process is not restricted by animal raw materials, offers low costs, is easily scalable, and produces high product purity, making it easier to control molecular weight. Hyaluronic acid is a major component of the extracellular matrix and intercellular matrix, acting as a filler between cells and playing a vital role in the morphology, structure, and function of the skin. Due to its moisturizing, repairing, and nourishing properties, its good skin compatibility, and its safety, hyaluronic acid is increasingly used in cosmetics. Due to its biodegradability, biocompatibility, chemical modifiability, and in vivo targeting, hyaluronic acid has attracted considerable attention in the field of protein and peptide modification. The present invention adopts hyaluronic acid-modified cosmetic peptides and further studies the synthesis method and application thereof. Summary of the Invention

[0003] The present invention aims to provide a hyaluronic acid-modified cosmetic peptide, a preparation method, and its use. The hyaluronic acid-modified cosmetic peptide can maintain the properties of sodium hyaluronate while containing the functionality of a polypeptide. In addition, the stability of the prepared novel compound is significantly improved, and the peptide has broad application prospects in the field of cosmetics / skin care products.

[0004] The technical solutions adopted by the present invention to achieve the above-mentioned purpose are: A hyaluronic acid-modified beauty peptide, the structural formula of the hyaluronic acid-modified beauty peptide is shown in formula (I): MC (I); Wherein, M represents sodium hyaluronate, and its structure is shown in formula (II): (II), where y is a natural number ≥ 1; C represents a beauty peptide, which includes a polypeptide or a derivative thereof having beauty and / or skin care effects, and includes a dipeptide, tripeptide, tetrapeptide, pentapeptide, hexapeptide, heptapeptide, octapeptide, nonapeptide or other polypeptides; The amino group in the M-terminal glucuronic acid linkage C structure is linked. The application adopts cosmetic polypeptide to modify hyaluronic acid, adopts substitution reaction to combine sodium hyaluronate and polypeptide through a covalent bond to form a new chemical substance, the new chemical substance can maintain the properties of sodium hyaluronate and contain the functionality of polypeptide; and the compound has multiple effects, and the stability effect of the compound is obviously enhanced compared with the original effect. For example, the compound has more excellent moisturizing performance; the anti-wrinkle firming performance, whitening performance, antioxidant capacity and anti-sugar performance of part of the compound are obviously improved, and the soothing capacity is also effectively improved. The application generates hyaluronic acid modified cosmetic peptide by coupling hyaluronic acid and cosmetic polypeptide through a covalent bond, directly modifies through chemical modification, forms a new chemical structure, can fundamentally endow the compound with more excellent characteristics, and the preparation method is simple; meanwhile, the application also provides the application of the compound in cosmetics and cosmetic products, significantly enhances the use effect of functional cosmetics, such as moisturizing, anti-aging firming, whitening, soothing, antioxidant and anti-sugar effects, and further can improve the use satisfaction of users.

[0005] Preferably, y is selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12; more preferably, y = 1 or 2 or 3.

[0006] Specifically, the compound represented by formula (I) includes the structure represented by formula (III) as follows: (III); wherein, The above n is a natural number; The above R is the remaining part of the cosmetic peptide structure except the reactive group amino group; The above cosmetic peptide includes a polypeptide or derivative thereof having a cosmetic and / or skin care effect, and the above polypeptide includes a dipeptide, a tripeptide, a tetrapeptide, a pentapeptide, a hexapeptide, a heptapeptide, an octapeptide, a nonapeptide or other polypeptide.

[0007] Preferably, n = 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23 or 24. More preferably, n = 0 or 1 or 2 or 3.

[0008] Specifically, the dipeptide or derivative thereof includes one of dipeptide-2 and carnosine; preferably carnosine.

[0009] Specifically, the tripeptide or derivative thereof includes one of dermasebtide, tripeptide-1, tripeptide-5, tripeptide-8, tripeptide-38 and palmitoyl tripeptide-38; preferably tripeptide-1 or tripeptide-8.

[0010] Specifically, the tetrapeptide or its derivative includes one of tetrapeptide-5, tetrapeptide-7, tetrapeptide-9, tetrapeptide-11, tetrapeptide-30 and tetrapeptide-15, or includes a tetrapeptide having an amino acid sequence of H-Asp-Val-Lys-Tyr-OH; preferably tetrapeptide-7 or tetrapeptide-15.

[0011] Specifically, the pentapeptide or its derivative includes one of pentapeptide-4 and myristyl pentapeptide-4; preferably pentapeptide-4.

[0012] Specifically, the hexapeptide or its derivative includes one of hexapeptide-1, hexapeptide-8, hexapeptide-9, hexapeptide-11 and hexapeptide-38, or includes one of the hexapeptides having an amino acid sequence of H-Arg-Arg-Gln-Met-Glu-Glu-NH2, H-Arg-Arg-Gln-D-Met-Glu-Glu-NH2, H-Trp-Phe-Arg-Leu-Ala-His-NH2 and H-Trp-Phe-Arg-D-Leu-Ala-His-NH2.

[0013] Preferably, the hexapeptide or its derivative includes hexapeptide-1, hexapeptide-8, hexapeptide-9, and hexapeptide-11; more preferably, the amino acid sequence is one of the hexapeptides H-Arg-Arg-Gln-Met-Glu-Glu-NH2, H-Arg-Arg-Gln-D-Met-Glu-Glu-NH2, H-Trp-Phe-Arg-Leu-Ala-His-NH2 and H-Trp-Phe-Arg-D-Leu-Ala-His-NH2.

[0014] Specifically, the heptapeptide or its derivatives include a heptapeptide having an amino acid sequence of H-Glu-Glu-Met-Gln-Arg-Arg-Ala-OH or H-Glu-Glu-Met-Gln-Arg-Arg-Ala-NH2.

[0015] Specifically, the octapeptide or its derivative includes an octapeptide having an amino acid sequence of H-Glu-Glu-Met-Gln-Arg-Arg-Ala-Asp-NH2.

[0016] Specifically, the nonapeptide or its derivative includes nonapeptide-1.

[0017] Specifically, the compound represented by the above (I) includes one or more of H-Glu-Glu-Met-Gln-Arg-Arg-NH-hyaluronic acid (sodium), hyaluronic acid (sodium)-NH-Phe-Val-Ala-Pro-Phe-Pro-OH, H-His-D-Phe-Arg-NH-hyaluronic acid (sodium), H-His-Ala-Leu-Arg-Phe-Trp-NH-hyaluronic acid (sodium), H-Trp-Phe-Arg-D-Leu-Ala-His-NH-hyaluronic acid (sodium), H-Trp-Phe-Arg-Leu-Ala-His-NH-hyaluronic acid (sodium), H-Arg-Arg-Gln-D-Met-Glu-Glu-NH-hyaluronic acid (sodium), or H-Arg-Arg-Gln-Met-Glu-Glu-NH-hyaluronic acid (sodium).

[0018] Further, the compound represented by the above (I) can also include one or more of hyaluronic acid (sodium)-HN-Gly-Gln-Pro-Arg-OH, hyaluronic acid (sodium)-HN-Lys-Thr-Thr-Lys-Ser-OH, H-Lys-Leu-Ala-Lys-Lys-NH-hyaluronic acid (sodium), H-Gly-Pro-Gln-Gly-Pro-Gln-NH-hyaluronic acid (sodium), H-Met-Pro-{D-Phe}-Arg-{D-Trp}-Phe-Lys-Pro-Val-NH-hyaluronic acid (sodium), H-Tyr-Pro-Phe-Phe-NH-hyaluronic acid (sodium), hyaluronic acid (sodium)-NH-Glu-Glu-Met-Gln-Arg-Arg-Ala-OH, or hyaluronic acid (sodium)-NH-Asp-Val-Lys-Tyr-OH.

[0019] It should be noted that the hyaluronic acid modified cosmetic peptide represented by the above can represent a case where there is only one reactive site between the hyaluronic acid (sodium) and the cosmetic peptide in the structure, and can also represent a case where there are multiple reactive sites. Further, it should be noted that the bonding portion includes a reaction between an active amino group in the cosmetic polypeptide structure and an active hydroxyl site capable of further polymerization in the end group ring of the hyaluronic acid (sodium) structure.

[0020] More preferably, the compound represented by the above formula (I) includes one or more of the following: A2; A4; A6; B2; B4; B6; D2; D4; D6; E2; E4; E6; F2; F4; F6; G2; G4; G6。 The application further discloses a preparation method of the hyaluronic acid modified cosmetic peptide.

[0021] Further, the synthesis route of the hyaluronic acid modified cosmetic peptide is shown as follows: ; wherein n is a natural number; Y is Na or H; Peptide=R; R is the remaining part of the cosmetic peptide structure except the reactive amino group; X includes H, sodium or potassium; The cosmetic peptide includes a polypeptide or derivative thereof having a cosmetic and / or skin care effect, and the polypeptide includes a dipeptide, tripeptide, tetrapeptide, pentapeptide, hexapeptide, heptapeptide, octapeptide, nonapeptide or other polypeptide.

[0022] It should be noted that the final form of the hyaluronic acid modified cosmetic peptide provided by the application can be the hyaluronic acid modified cosmetic peptide, or a hydrochloride, sulfate, phosphate, acetate, sodium salt, potassium salt, trifluoroacetate, maleate or fumarate, etc.

[0023] The preparation method of the hyaluronic acid modified cosmetic peptide includes that sodium hyaluronate and a cosmetic peptide are subjected to ring-opening reaction to prepare the hyaluronic acid modified cosmetic peptide.

[0024] Specifically, the preparation method of the hyaluronic acid modified cosmetic peptide includes the following steps: The cosmetic peptide is added into an organic solvent, stirred and dissolved, then TFA is added, stirred for 0.5-1 h, then DIEA is added, then sodium hyaluronate is weighed and added; stirring is carried out at 40-50℃ overnight, sample detection is carried out by LC-MS, the reaction is basically complete, acetic acid is added, temperature control is carried out by a water bath pot at 30-45℃, rearrangement reaction is carried out for 2-4 h, sample detection is carried out by LC-MS, the reaction is basically complete; then reverse chromatography purification is carried out, and the hyaluronic acid modified cosmetic peptide is prepared.

[0025] Specifically, the above-mentioned organic solvent is selected from single solvents or mixed solvents of DMF, N,N-dimethylacetamide, DMSO, acetonitrile, azomethylnitrile pyrrolidone (NMP), methanol, ethanol, acetone, THF, dichloromethane, ethyl acetate, dioxane and water; and DMSO is preferred.

[0026] Specifically, the mass-volume ratio of the above-mentioned cosmetic peptide to DMSO is 1g:8-25mL; and 1g:10mL is preferred.

[0027] Specifically, the mass-volume ratio of the above-mentioned cosmetic peptide to TFA is 1g:0-1.2mL.

[0028] Specifically, the mass ratio of the above-mentioned cosmetic peptide to DIEA is 1:0.5-5.5; and 1:0.8-2.5 is preferred.

[0029] Specifically, the molar ratio of the above-mentioned cosmetic peptide to hyaluronic acid is 1:2-8; 1:3-5 is preferred; and 1:4 is more preferred.

[0030] Specifically, the mass-volume ratio of the above-mentioned cosmetic peptide to acetic acid is 1g:4-10mL; 1g:5-9mL is preferred; and 1g:8mL is more preferred.

[0031] Another purpose of the present application is to disclose the use of the above-mentioned hyaluronic acid modified cosmetic peptide in the preparation of cosmetics and / or skin care products.

[0032] The present application further discloses the use of the above-mentioned hyaluronic acid modified cosmetic peptide in enhancing the moisturizing performance of cosmetics and / or skin care products.

[0033] The present application further discloses the use of the above-mentioned hyaluronic acid modified cosmetic peptide in enhancing the soothing performance of cosmetics and / or skin care products.

[0034] The present application further discloses the use of the above-mentioned hyaluronic acid modified cosmetic peptide in enhancing the anti-wrinkle and firming performance of cosmetics and / or skin care products.

[0035] The present application further discloses the use of the above-mentioned hyaluronic acid modified cosmetic peptide in enhancing the whitening performance of cosmetics and / or skin care products.

[0036] The application also discloses use of the hyaluronic acid modified cosmetic peptide in enhancing anti-glycation performance of the cosmetic and / or skin care product.

[0037] The application also discloses use of the hyaluronic acid modified cosmetic peptide in enhancing anti-oxidation performance of the cosmetic and / or skin care product.

[0038] A cosmetic product comprising the hyaluronic acid modified cosmetic peptide.

[0039] A skin care product comprising the hyaluronic acid modified cosmetic peptide.

[0040] The application has the following advantages: The application modifies hyaluronic acid by using a cosmetic polypeptide, forms a new compound through a Maillard reaction, the compound can maintain the properties of sodium hyaluronate and contain the functionality of the polypeptide, and the stability of the compound is obviously enhanced. For example, the compound has more excellent moisturizing performance, the anti-wrinkle firming performance, whitening performance, anti-oxidation ability and anti-glycation performance of some compounds are obviously improved, and the soothing ability is also effectively improved. The hyaluronic acid modified cosmetic peptide provided by the application is directly modified by chemical modification, forms a new chemical structure, can fundamentally endow the compound with more excellent characteristics, and the preparation method is simple. The application also provides application of the hyaluronic acid modified cosmetic peptide in cosmetic products and cosmetic products, significantly enhances the use effect of functional cosmetic products, such as moisturizing, anti-aging firming, whitening, soothing, anti-oxidation and anti-glycation, and thus can improve the use satisfaction of users.

[0041] Therefore, the application provides a hyaluronic acid modified cosmetic peptide, a preparation method and application thereof, the hyaluronic acid modified cosmetic peptide can maintain the properties of sodium hyaluronate and contain the functionality of the polypeptide, and the stability of the prepared new compound is obviously improved, and the hyaluronic acid modified cosmetic peptide has a wide application prospect in the field of cosmetic products / skin care products. BRIEF DESCRIPTION OF DRAWINGS

[0042] Figure 1 Mass spectrum test results of H-Gly-Gln-Pro-Arg-OH prepared for Example 1; Figure 2 Liquid chromatogram test results of H-Gly-Gln-Pro-Arg-OH prepared for Example 1; Figure 3 Mass spectrum test results of the hyaluronic acid modified cosmetic peptide prepared for Example 1; Figure 4 Liquid chromatogram test results of the hyaluronic acid modified cosmetic peptide prepared for Example 1; Figure 5Mass Spectrometry test results for H-Lys-Thr-Thr-Lys-Ser-OH prepared for Example 2; Figure 6 Liquid Chromatography test results for H-Lys-Thr-Thr-Lys-Ser-OH prepared for Example 2; Figure 7 Liquid Chromatography test results for hyaluronic acid modified cosmetic peptide prepared for Example 2; Figure 8 Mass Spectrometry test results for sample at 5.801 min in Liquid Chromatography of hyaluronic acid modified cosmetic peptide prepared for Example 2; Figure 9 Mass Spectrometry test results for sample at 6.102 min in Liquid Chromatography of hyaluronic acid modified cosmetic peptide prepared for Example 2; Figure 10 Mass Spectrometry test results for sample at 6.558 min in Liquid Chromatography of hyaluronic acid modified cosmetic peptide prepared for Example 2; Figure 11 Mass Spectrometry test results for H-Lys-Leu-Ala-Lys-Lys-NH2 prepared for Example 3; Figure 12 LC-MS test results for H-Lys-Leu-Ala-Lys-Lys-NH2 prepared for Example 3; Figure 13 Mass Spectrometry test results for hyaluronic acid modified cosmetic peptide prepared for Example 3; Figure 14 Liquid Chromatography test results for hyaluronic acid modified cosmetic peptide prepared for Example 3; Figure 15 Mass Spectrometry test results for H-Glu-Glu-Met-Gln-Arg-Arg-NH2 prepared for Example 4; Figure 16 LC-MS test results for H-Glu-Glu-Met-Gln-Arg-Arg-NH2 prepared for Example 4; Figure 17 Mass Spectrometry test results for hyaluronic acid modified cosmetic peptide D2 prepared for Example 4; Figure 18 Liquid Chromatography test results for hyaluronic acid modified cosmetic peptide D2 prepared for Example 4; Figure 19 Mass Spectrometry test results for hyaluronic acid modified cosmetic peptide D4 prepared for Example 4; Figure 20 Liquid Chromatography test results for hyaluronic acid modified cosmetic peptide D4 prepared for Example 4; Figure 21 Mass spectrometry test results for hyaluronic acid modified cosmetic peptide D6 prepared for Example 4; Figure 22 Liquid chromatography test results for hyaluronic acid modified cosmetic peptide D6 prepared for Example 4; Figure 23 Mass spectrometry test results for hyaluronic acid modified cosmetic peptide prepared for Example 5; Figure 24 Liquid chromatography test results for hyaluronic acid modified cosmetic peptide prepared for Example 5; Figure 25 Mass spectrometry test results for H-Gly-Pro-Gln-Gly-Pro-Gln-NH2 prepared for Example 6; Figure 26 Liquid chromatography test results for H-Gly-Pro-Gln-Gly-Pro-Gln-NH2 prepared for Example 6; Figure 27 Mass spectrometry test results for hyaluronic acid modified cosmetic peptide prepared for Example 6; Figure 28 Liquid chromatography test results for hyaluronic acid modified cosmetic peptide prepared for Example 6; Figure 29 Mass spectrometry test results for H-Met-Pro-{D-Phe}-Arg-{D-Trp}-Phe-Lys-Pro-Val-NH2 prepared for Example 7; Figure 30 Liquid chromatography test results for H-Met-Pro-{D-Phe}-Arg-{D-Trp}-Phe-Lys-Pro-Val-NH2 prepared for Example 7; Figure 31 Mass spectrometry test results for hyaluronic acid modified cosmetic peptide prepared for Example 7; Figure 32 Liquid chromatography test results for hyaluronic acid modified cosmetic peptide prepared for Example 7; Figure 33 Mass spectrometry test results for H-Phe-Val-Ala-Pro-Phe-Pro-OH prepared for Example 8; Figure 34 LC-MS test results for H-Phe-Val-Ala-Pro-Phe-Pro-OH prepared for Example 8; Figure 35 Mass spectrometry test results for hyaluronic acid modified cosmetic peptide H2 prepared for Example 8; Figure 36LC-MS test result of hyaluronic acid modified cosmetic peptide H2 prepared for Example 8; Figure 37 MS test result of hyaluronic acid modified cosmetic peptide H4 prepared for Example 8; Figure 38 LC-MS test result of hyaluronic acid modified cosmetic peptide H4 prepared for Example 8; Figure 39 MS test result of hyaluronic acid modified cosmetic peptide H6 prepared for Example 8; Figure 40 LC-MS test result of hyaluronic acid modified cosmetic peptide H6 prepared for Example 8; Figure 41 MS test result of H-His-D-Phe-Arg-NH2 prepared for Example 9; Figure 42 LC-MS test result of H-His-D-Phe-Arg-NH2 prepared for Example 9; Figure 43 MS test result of hyaluronic acid modified cosmetic peptide L2 prepared for Example 9; Figure 44 LC-MS test result of hyaluronic acid modified cosmetic peptide L2 prepared for Example 9; Figure 45 MS test result of hyaluronic acid modified cosmetic peptide L4 prepared for Example 9; Figure 46 LC-MS test result of hyaluronic acid modified cosmetic peptide L4 prepared for Example 9; Figure 47 MS test result of hyaluronic acid modified cosmetic peptide L6 prepared for Example 9; Figure 48 LC-MS test result of hyaluronic acid modified cosmetic peptide L6 prepared for Example 9; Figure 49 MS test result of H-Tyr-Pro-Phe-Phe-NH2 prepared for Example 10; Figure 50 LC-MS test result of H-Tyr-Pro-Phe-Phe-NH2 prepared for Example 10; Figure 51 MS test result of hyaluronic acid modified cosmetic peptide prepared for Example 10; Figure 52 LC-MS test result of hyaluronic acid modified cosmetic peptide prepared for Example 10; Figure 53Mass spectrometry test results of the hyaluronic acid-modified cosmetic peptide part products (reaction products of sodium hyaluronate and cosmetic peptide with y = 1, y = 2, and y = 3) prepared for Example 11; Figure 54 High performance liquid chromatography test results of the hyaluronic acid-modified cosmetic peptide part products (reaction products of sodium hyaluronate and cosmetic peptide with y = 1, y = 2, and y = 3) prepared for Example 11; Figure 55 Mass spectrometry test results of the hyaluronic acid-modified cosmetic peptide part products (reaction products of sodium hyaluronate and cosmetic peptide with y = 2 and y = 3) prepared for Example 11; Figure 56 High performance liquid chromatography test results of the hyaluronic acid-modified cosmetic peptide part products (reaction products of sodium hyaluronate and cosmetic peptide with y = 2 and y = 3) prepared for Example 11; Figure 57 Mass spectrometry test results of H-Glu-Glu-Met-Gln-Arg-Arg-Ala-OH prepared for Example 12; Figure 58 High performance liquid chromatography test results of H-Glu-Glu-Met-Gln-Arg-Arg-Ala-OH prepared for Example 12; Figure 59 High performance liquid chromatography test results of the hyaluronic acid-modified cosmetic peptide prepared for Example 12; Figure 60 Mass spectrometry test results of the sample at 11.577 min in the high performance liquid chromatography of the hyaluronic acid-modified cosmetic peptide prepared for Example 12; Figure 61 Mass spectrometry test results of the sample at 12.223 min in the high performance liquid chromatography of the hyaluronic acid-modified cosmetic peptide prepared for Example 12; Figure 62 Mass spectrometry test results of H-Asp-Val-Lys-Tyr-OH prepared for Example 13; Figure 63 High performance liquid chromatography test results of H-Asp-Val-Lys-Tyr-OH prepared for Example 13; Figure 64 High performance liquid chromatography test results of the hyaluronic acid-modified cosmetic peptide prepared for Example 13; Figure 65 Mass spectrometry test results of the sample at 8.226 min in the high performance liquid chromatography of the hyaluronic acid-modified cosmetic peptide prepared for Example 13; Figure 66Mass spectrum test result of sample at 12.711 min in the liquid chromatogram of hyaluronic acid modified cosmetic peptide prepared for Example 13; Figure 67 Mass spectrum test result of H-His-Ala-Leu-Arg-Phe-Trp-NH2 prepared for Example 14; Figure 68 Liquid chromatogram test result of H-His-Ala-Leu-Arg-Phe-Trp-NH2 prepared for Example 14; Figure 69 Mass spectrum test result of hyaluronic acid modified cosmetic peptide A2 prepared for Example 14; Figure 70 Liquid chromatogram test result of hyaluronic acid modified cosmetic peptide A2 prepared for Example 14; Figure 71 Mass spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 72 Liquid chromatogram test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 73-1 Partial nuclear magnetic hydrogen spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 73-2 Partial nuclear magnetic hydrogen spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 73-3 Partial nuclear magnetic hydrogen spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 73-4 Partial nuclear magnetic hydrogen spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 73-5 Partial nuclear magnetic hydrogen spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 74-1 Partial nuclear magnetic carbon spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 74-2 Partial nuclear magnetic carbon spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 74-3 Partial nuclear magnetic carbon spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 74-4 Partial nuclear magnetic carbon spectrum test result of hyaluronic acid modified cosmetic peptide A4 prepared for Example 14; Figure 74-5Test results of the partial nuclear magnetic carbon spectrum of hyaluronic acid modified cosmetic peptide A4 prepared in Example 14; Figure 75 Test results of the two-dimensional cosy spectrum of hyaluronic acid modified cosmetic peptide A4 prepared in Example 14; Figure 76 Test results of the mass spectrum of hyaluronic acid modified cosmetic peptide A6 prepared in Example 14; Figure 77 Test results of the liquid chromatogram of hyaluronic acid modified cosmetic peptide A6 prepared in Example 14; Figure 78 Test results of the mass spectrum of H-Trp-Phe-Arg-D-Leu-Ala-His-NH2 prepared in Example 27; Figure 79 Test results of the liquid chromatogram of H-Trp-Phe-Arg-D-Leu-Ala-His-NH2 prepared in Example 27; Figure 80 Test results of the liquid chromatogram of hyaluronic acid modified cosmetic peptide E2 prepared in Example 27; Figure 81 Test results of the mass spectrum of hyaluronic acid modified cosmetic peptide E2 prepared in Example 27; Figure 82 Test results of the mass spectrum of hyaluronic acid modified cosmetic peptide E4 prepared in Example 27; Figure 83 Test results of the liquid chromatogram of hyaluronic acid modified cosmetic peptide E4 prepared in Example 27; Figure 84 Test results of the mass spectrum of hyaluronic acid modified cosmetic peptide E6 prepared in Example 27; Figure 85 Test results of the liquid chromatogram of hyaluronic acid modified cosmetic peptide E6 prepared in Example 27; Figure 86 Test results of the mass spectrum of H-Trp-Phe-Arg-Leu-Ala-His-NH2 prepared in Example 28; Figure 87 Test results of the liquid chromatogram of H-Trp-Phe-Arg-Leu-Ala-His-NH2 prepared in Example 28; Figure 88 Test results of the mass spectrum of hyaluronic acid modified cosmetic peptide B2 prepared in Example 28; Figure 89 Test results of the liquid chromatogram of hyaluronic acid modified cosmetic peptide B2 prepared in Example 28; Figure 90Liquid chromatogram test results for hyaluronan-modified cosmetic peptide B4 prepared for Example 28; Figure 91 Liquid chromatogram test results for hyaluronan-modified cosmetic peptide B4 prepared for Example 28; Figure 92 Mass spectrometry test results for hyaluronan-modified cosmetic peptide B6 prepared for Example 28; Figure 93 Liquid chromatogram test results for hyaluronan-modified cosmetic peptide B6 prepared for Example 28; Figure 94 Mass spectrometry test results for H-Arg-Arg-Gln-D-Met-Glu-Glu-NH2 prepared for Example 29; Figure 95 Liquid chromatogram test results for H-Arg-Arg-Gln-D-Met-Glu-Glu-NH2 prepared for Example 29; Figure 96 Liquid chromatogram test results for hyaluronan-modified cosmetic peptide G2 prepared for Example 29; Figure 97 Liquid chromatogram test results for hyaluronan-modified cosmetic peptide G2 prepared for Example 29; Figure 98 Liquid chromatogram test results for hyaluronan-modified cosmetic peptide G4 prepared for Example 29; Figure 99 Mass spectrometry test results for hyaluronan-modified cosmetic peptide G4 prepared for Example 29; Figure 100 Mass spectrometry test results for hyaluronan-modified cosmetic peptide G6 prepared for Example 29; Figure 101 Liquid chromatogram test results for hyaluronan-modified cosmetic peptide G6 prepared for Example 29; Figure 102 Mass spectrometry test results for H-Arg-Arg-Gln-Met-Glu-Glu-NH2 prepared for Example 30; Figure 103 Liquid chromatogram test results for H-Arg-Arg-Gln-Met-Glu-Glu-NH2 prepared for Example 30; Figure 104 Liquid chromatogram test results for hyaluronan-modified cosmetic peptide F2 prepared for Example 30; Figure 105 Liquid chromatogram test results for hyaluronan-modified cosmetic peptide F2 prepared for Example 30; Figure 106Test results of the liquid chromatogram of hyaluronic acid modified cosmetic peptide F4 prepared for example 30; Figure 107 Test results of the mass spectrum of hyaluronic acid modified cosmetic peptide F4 prepared for example 30; Figure 108 Test results of the mass spectrum of hyaluronic acid modified cosmetic peptide F6 prepared for example 30; Figure 109 Test results of the liquid chromatogram of hyaluronic acid modified cosmetic peptide F6 prepared for example 30. DETAILED DESCRIPTION

[0043] In order to make the purpose, technical solutions and advantages of the present application clearer and more explicit, the technical solutions of the present application are further described in detail below in combination with specific embodiments: It should be noted that the hyaluronic acid used in examples 1-30 of the present application is hydrolyzed sodium hyaluronate, which is purchased from Shandong Bolijia Biological Technology Co., Ltd. It is a mixture, and the main components include disaccharide sodium hyaluronate, tetrasaccharide sodium hyaluronate, and hexasaccharide sodium hyaluronate, with a mass ratio of 1:5:3.

[0044] Example 1: A method for synthesizing H-Gly-Gln-Pro-Arg-OH, comprising: 3.13 mmol of Wang resin was placed in a 125 mL solid-phase synthesis reactor, 7.5 mmol of amino acid Fmoc-Arg(Pbf)-OH was added, 15 mL of dichloromethane (DCM) was added, then 2.01 mL of pyridine and 1.78 mL of DBU were added, and the reaction was carried out at 25°C for 3 h. Filtration was performed, and the resin was washed with a DMF solution 3 times, each time with 15 mL. 15 mL of capping liquid (the capping liquid contains Ac2O, DMF, and DIEA, with a mass ratio of Ac2O:DMF:DIEA of 10:84:6) was added, and the reaction was carried out for 15 min. Filtration was performed, and the resin was washed with dichloromethane 2 times, each time with 15 mL, washed with methanol 2 times, each time with 15 mL, and washed with DMF 2 times, each time with 15 mL. 15 mL of 20% Pip / DMF (v / v) solution was added, and the reaction was carried out with stirring for 30 min. Filtration was performed, the deprotection liquid was removed, and then the resin was washed with DMF 6 times, each time with 15 mL, and was dried by suction.

[0045] Take 5 mmol Fmoc-Pro-OH.H2O, 5 mmol HOBt in 50 mL beaker, cooling to 5 ℃, add 5 mL DMF, 0.8 mL DIC standing reaction 15 min, and the solution in 50 mL beaker is added to the above-mentioned 125 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 15 mL each time; after washing, the next step reaction is carried out; 20% Pip / DMF (v / v) solution 15 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, 15 mL each time, suction dry for use.

[0046] Take 7.5 mmol Fmoc-Gln(Trt)-OH, 7.5 mmol HOBt in 50 mL beaker, cooling to 5 ℃, add 5 mL DMF, 1.2 mL DIC standing reaction 15 min, and the solution in 50 mL beaker is added to the above-mentioned 125 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 15 mL each time; after washing, the next step reaction is carried out; 20% Pip / DMF (v / v) solution 15 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 15 mL 6 times, suction dry for use.

[0047] Take 7.5 mmol Fmoc-Gly-OH, 7.5 mmol HOBt in 50 mL beaker, cooling to 5 ℃, add 5 mL DMF, 1.2 mL DIC standing reaction 15 min, and the solution in 50 mL beaker is added to the above-mentioned 125 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 15 mL each time; after washing, the next step reaction is carried out; 20% Pip / DMF (v / v) solution 15 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, 15 mL each time; methanol washing 2 times, 15 mL each time; DCM solution washing 2 times, 15 mL each time; methanol washing 2 times, 15 mL each time; vacuum drying, to obtain the peptide resin of H-AA1-AA2-AA3-AA4-Wang-resin, wherein AA1 is Gly; AA2 is Gln(Trt); AA3 is Pro; AA4 is Arg(Pbf). The above-mentioned peptide resin is cleaved with TFA / Tis / H2O (the volume ratio of TFA, Tis and H2O is 90:5:5), the amount is 30 mL, the time is 2.5 h, the cleavage solution is added to 300 mL ether (5 ℃) solution, the white solid is precipitated, centrifuged, to obtain H-Gly-Gln-Pro-Arg-OH, its mass spectrum ( Figure 1 ) and high performance liquid chromatography ( Figure 2As shown in Figure 1-2

[0048] A preparation method of hyaluronic acid modified cosmetic peptide, comprising the following steps: Take 0.46g H-Gly-Gln-Pro-Arg-OH.2TFA, add 5mL DMSO, stir and dissolve, add 0.785g DIEA, and the tetrapeptide is precipitated. Take 2.1g hyaluronic acid, control the temperature of the water bath at 45℃, stir for 15min, and the reaction solution is clear. Continue to control the temperature at 45℃ for overnight reaction. Take sample for LC-MS detection, and the reaction is basically complete. Add 4mL acetic acid, control the temperature of the water bath at 35℃, and carry out rearrangement reaction for 3h. Take sample for LC-MS detection, and the reaction is basically complete. Then carry out reverse chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5g of the crude product, add 100mL H2O for dilution; Filler: 21.2*250mm, 10-120, C18; flow rate: 10mL / min; wavelength: 220nm; Mobile phase: A: 1%HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium for 10min, flow rate: 10mL / min; Loading: flow rate: 10mL / min; Elution: 0-10%B for 60min; Column cleaning: clean with 80% ACN to baseline equilibrium; Collect the qualified product, and hyaluronic acid modified cosmetic peptide is prepared. The mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 3 and Figure 4

[0049] Example 2: A synthesis method of H-Lys-Thr-Thr-Lys-Ser-OH, comprising: Put 8.75mmol CTC resin into a 250mL solid phase synthesis reactor, add 17.5mmol amino acid Fmoc-Ser(tBu)-OH, add dichloromethane 75mL, add 7.0mL DIEA, and react at 25℃ for 3h. Add 10mL of methanol and react for 5min. Filter, wash the resin with dichloromethane twice, 75mL each time; wash with methanol twice, 75mL each time; wash with DMF twice, 75mL each time; add 20% Pip / DMF (v / v) solution 75mL, stir for 30min, filter, remove the deprotection solution, and then wash with DMF solution for 6 times, 75mL each time, and dry by suction.

[0050] ​​Take 21 mmol Fmoc-Lys(Boc)-OH, 21 mmol HOBt in a 100 mL beaker, cool to 5°C, add 50 mL of DMF solution, 3.2 mL of DIC, and let the reaction stand for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir the reaction for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 75 mL each time; after the washing is complete, the next step is carried out; 20% Pip / DMF (v / v) solution 75 mL is added, the reaction is stirred for 30 min, suction filtration is performed, the deprotection solution is removed, and then the resin is washed with DMF solution 6 times, 65 mL each time, and suction dried for use.

[0051] Take 21 mmol Fmoc-Lys(Boc)-OH, 21 mmol HOBt in a 100 mL beaker, cool to 5°C, add 50 mL of DMF solution, 3.2 mL of DIC, and let the reaction stand for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir the reaction for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 75 mL each time; after the washing is complete, the next step is carried out; 20% Pip / DMF (v / v) solution 75 mL is added, the reaction is stirred for 30 min, suction filtration is performed, the deprotection solution is removed, and then the resin is washed with DMF solution 6 times, 65 mL each time, and suction dried for use.

[0052] Take 21 mmol Fmoc-Lys(Boc)-OH, 21 mmol HOBt in a 100 mL beaker, cool to 5°C, add 50 mL of DMF solution, 3.2 mL of DIC, and let the reaction stand for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir the reaction for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 75 mL each time; after the washing is complete, the next step is carried out; 20% Pip / DMF (v / v) solution 75 mL is added, the reaction is stirred for 30 min, suction filtration is performed, the deprotection solution is removed, and then the resin is washed with DMF solution 6 times, 65 mL each time, and suction dried for use.

[0053] Take 21 mmol Fmoc-Lys(Boc)-OH, 21 mmol HOBt in a 100 mL beaker, cool to 2-8℃, add 50 mL DMF solution, 3.2 mL DIC, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, the reaction is complete; the resin is washed with DMF solution for 3 times, 75 mL each time; after the washing is completed, the next step reaction is carried out; 20% Pip / DMF (v / v) solution 75 mL is added, stirred for reaction for 30 min, suction filtered, the deprotection solution is removed, then washed with DMF solution for 6 times, 65 mL each time, then washed with methanol for 2 times, 75 mL each time, DCM solution for 2 times, 75 mL each time, methanol for 2 times, 75 mL each time, vacuum dried to obtain H-Lys(Boc)-Thr(tBu)-Thr(tBu)-Lys(Boc)-Ser(tBu)-CTC-resin; Take 4.14 g of the above-mentioned peptide resin, add 40 mL of the cutting solution TFA / Tis / H2O (the volume ratio of TFA, Tis and H2O is 90:5:5), stir for reaction for 2.5 h at 30℃, filter, remove the resin to obtain the filtrate; the filtrate is dried to obtain H-Lys-Thr-Thr-Lys-Ser-OH, and the mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 5 and Figure 6 .

[0054] The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Lys-Thr-Thr-Lys-Ser-OH.3TFA, add 5 mL of DMSO to stir and dissolve, add 82 μL of TFA, stir for 0.5 h, then add 0.641 g of DIEA, take 1.71 g of hyaluronic acid, control the temperature of the water bath to 45℃, stir for reaction overnight, take a sample to detect LC-MS, and the reaction is basically complete; then add 4 mL of acetic acid, control the temperature of the water bath to 35℃, and carry out rearrangement reaction for 3 h, take a sample to detect LC-MS, and the reaction is basically complete; then carry out reverse chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=100:0, equilibrate for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-10% B for 60 min; Column washing: 80% ACN wash to baseline equilibrium; The collected product was hyaluronic acid modified cosmetic peptide, and its high performance liquid chromatography characterization was as shown in Figure 7 The samples collected at 5.801 min, 6.102 min and 6.558 min were subjected to mass spectrometry characterization, and the results were as shown in Figure 8 、 Figure 9 、 Figure 10

[0055] Example 3: A method for synthesizing H-Lys-Leu-Ala-Lys-Lys-NH2, comprising: Put 10 mmol AM resin into a 250 mL solid-phase synthesis reactor, add 70 mL of 20% Pip / DMF (v / v) solution, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution for 6 times, 70 mL each time, and dry for standby.

[0056] Take 20 mmol Fmoc-Linker and 20 mmol HOBt in a 100 mL beaker, cool to 5°C, add 35 mL of DMF solution and 3.1 mL of DIC, stand for reaction for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete; the resin is washed with DMF solution for 3 times, 70 mL each time; after washing is completed, the next step reaction is carried out; add 70 mL of 20% Pip / DMF (v / v) solution, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution for 6 times, 70 mL each time, and dry for standby.

[0057] Take 20 mmol Fmoc-Lys(Boc)-OH and 20 mmol HOBt in a 100 mL beaker, cool to 5°C, add 35 mL of DMF solution and 3.1 mL of DIC, stand for reaction for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete; the resin is washed with DMF solution for 3 times, 70 mL each time; after washing is completed, the next step reaction is carried out; add 70 mL of 20% Pip / DMF (v / v) solution, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution for 6 times, 70 mL each time, and dry for standby.

[0058] ​Take 20 mmol Fmoc-Lys(Boc)-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5°C, add DMF solution 35 mL, 3.1 mL DIC, stand for 15 min, and add the solution in 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for 1.5 h, the reaction is complete; resin is washed with DMF solution 3 times, 70 mL each time; after washing, the next step is carried out; add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 70 mL each time, and dry for use.

[0059] Take 20 mmol Fmoc-Ala-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5°C, add DMF solution 35 mL, 3.1 mL DIC, stand for 15 min, and add the solution in 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for 1.5 h, the reaction is complete; resin is washed with DMF solution 3 times, 70 mL each time; after washing, the next step is carried out; add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 70 mL each time, and dry for use.

[0060] Take 20 mmol Fmoc-Leu-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5°C, add DMF solution 35 mL, 3.1 mL DIC, stand for 15 min, and add the solution in 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for 1.5 h, the reaction is complete; resin is washed with DMF solution 3 times, 70 mL each time; after washing, the next step is carried out; add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0061] Take 20 mmol Fmoc-Lys(Boc)-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5℃, add DMF solution 35 mL, 3.1 mL DIC stand by reaction 15 min, and add the solution in 100 mL beaker to the above 250 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 70 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 70 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, suction dry for use. Then add methanol wash 2 times, 70 mL each time, DCM solution wash 2 times, 70 mL each time, methanol wash 2 times, 70 mL each time. Vacuum drying, get H-Lys(Boc)-Leu-Ala-Lys(Boc)-Lys(Boc)-Linker-AM resin peptide resin, In the above peptide resin, add 100 mL cutting fluid TFA / Tis / H2O (TFA, Tis and H2O in the volume ratio of 90:5:5), cutting 2.5 hours, add the cutting fluid to 1000 mL ether (5℃) solution, precipitate white solid, centrifugation, vacuum drying, prepared H-Lys-Leu-Ala-Lys-Lys-NH2, its mass spectrum and LC-MS characterization results are shown in Figure 11 and Figure 12

[0062] The preparation method of hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g H-Lys-Leu-Ala-Lys-Lys-NH2.4TFA, add 10 mL DMSO stirring to dissolve, add 107 μL TFA, stirring at room temperature for 0.5 h, then add 0.557 g DIEA, add 1.49 g hyaluronic acid, water bath temperature control 45℃, stirring reaction overnight, sample detection LC-MS, the reaction is basically complete, add 4 mL acetic acid, water bath temperature control 35℃, rearrangement reaction 3 h, sample detection LC-MS, the reaction is basically complete; then carry out reverse phase chromatography purification, purification conditions: Dissolution: take 0.5 g of crude product and dilute with 100 mL H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; ​Elution: 0-10% B for 60 min; Column washing: 80% ACN wash to baseline equilibration; The product was collected and the hyaluronic acid modified cosmetic peptide was prepared, and the mass spectrum and high performance liquid chromatography characterization results thereof were as shown in Figure 13 and Figure 14 .

[0063] Example 4: A method for synthesizing H-Glu-Glu-Met-Gln-Arg-Arg-NH2, comprising: 5 mmol AM resin was placed in a 100 mL solid phase synthesis reactor, 30 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, filtered, the deprotection solution was removed, and then washed with DMF solution for 6 times, 30 mL each time, and dried for standby; 10 mmol Fmoc-Linker, 10 mmol HOBt were taken in a 100 mL beaker, cooled to 5°C, 15 mL of DMF solution, 1.5 mL of DIC were added, and the solution in the 100 mL beaker was added to the above-mentioned 100 mL solid phase synthesis reactor, stirred for 1.5 h, and the reaction was completed; the resin was washed with DMF solution for 3 times, 20 mL each time; after washing was completed, the next step reaction was carried out; 30 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, filtered, the deprotection solution was removed, and then washed with DMF solution for 6 times, 30 mL each time, and dried for standby.

[0064] 10 mmol Fmoc-Arg(Pbf)-OH, 10 mmol HOBt were taken in a 100 mL beaker, cooled to 5°C, 15 mL of DMF solution, 1.5 mL of DIC were added, and the solution in the 100 mL beaker was added to the above-mentioned 100 mL solid phase synthesis reactor, stirred for 1.5 h, and the reaction was completed; the resin was washed with DMF solution for 3 times, 20 mL each time. After washing was completed, the next step reaction was carried out; 30 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, filtered, the deprotection solution was removed, and then washed with DMF solution for 6 times, 30 mL each time, and dried for standby.

[0065] Take 10 mmol Fmoc-Arg(Pbf)-OH, 10 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 15 mL, 1.5 mL DIC standing reaction 15 min, and the solution in 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; resin is washed with DMF solution 3 times, 20 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 30 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for use.

[0066] Take 10 mmol Fmoc-Gln(Trt)-OH, 10 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 15 mL, 1.5 mL DIC standing reaction 15 min, and the solution in 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; resin is washed with DMF solution 3 times, 20 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 30 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for use.

[0067] Take 10 mmol Fmoc-Met-OH, 10 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 15 mL, 1.5 mL DIC standing reaction 15 min, and the solution in 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; resin is washed with DMF solution 3 times, 20 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 40 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, suction dry for use.

[0068] Take 10 mmol Fmoc-Glu(otBu)-OH, 10 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 15 mL, 1.5 mL DIC standing reaction 15 min, and the solution in 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; resin is washed with DMF solution 3 times, 20 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 40 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, suction dry for use.

[0069] Take 15mmol Fmoc-Glu(otBu)-OH, 1mmol HOBt in 100mL beaker, cool to 5℃, add DMF solution 15mL, 2.3mL stand reaction 15min, and add 100mL beaker solution to the above 100mL solid phase synthesis reactor, stir reaction 1.5h, reaction is complete; the resin is washed with DMF solution 3 times, 30mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 40mL, stir reaction 30min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40mL each time, and dry by suction; then wash with methanol 2 times, 40mL each time, DCM solution 2 times, 40mL each time, methanol 2 times, 40mL each time, vacuum drying, to obtain H-Glu(otBu)-Glu(otBu)-Met-Gln(Trt)-Arg(Pbf)-Arg(Pbf)-Linker-AM resin peptide resin; The above peptide resin is cut with 100mL TFA / anisole / phenol / H2O / EDT (TFA, anisole, phenol, H2O and EDT are in a mass ratio of 87.5:5:2.5:2.5:2.5) for 2.5h, and the cutting solution is added to 1000mL ether (5℃) solution to precipitate white solid, centrifugation, vacuum drying, to obtain H-Glu-Glu-Met-Gln-Arg-Arg-NH2, and its mass spectrum and high performance liquid chromatography characterization results are shown in Figure 15 and Figure 16 .

[0070] A synthesis route of a hyaluronic acid modified cosmetic peptide is: ; Wherein, -COOX is -COONa; The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 1g H-Glu-Glu-Met-Gln-Arg-Arg-NH2.3TFA, add 10mL DMSO and stir to dissolve, add 889mg DIEA, then add 3.67g hyaluronic acid, control the temperature of water bath to 45℃, stir to react overnight, take sample for LC-MS detection, and the reaction is basically complete; add 8mL acetic acid, control the temperature of water bath to 35℃, and carry out rearrangement reaction for 3h, take sample for LC-MS detection, and the reaction is basically complete; then carry out reverse phase chromatography purification, and the purification conditions are: Dissolution: take 1g of crude product and dilute with 200mL of H2O; Filler: 50DAC, C18; flow rate: 60mL / min; wavelength: 220nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 100:0, equilibrium 10 min, flow rate: 60 mL / min; Loading: flow rate: 60 mL / min; Elution: 0-20% B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; Collecting qualified products, three kinds of hyaluronic acid modified cosmetic peptides with the following structures are obtained: D2, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 17 and Figure 18 ; D4, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 19 and Figure 20 ; D6, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 21 and Figure 22 .

[0071] Example 5: The preparation method of hyaluronic acid modified cosmetic peptides comprises the following steps: Take 0.5 g H-Gly-His-Lys-OH.2AcOH, add 10 mL DMSO and stir for 5 min to dissolve, add 286 μL TFA, stir for 1 min, then the solution becomes clear, stir for another 0.5 h, then add 3 g hyaluronic acid, stir to dissolve, then add 1.11 g DIEA, stir for 1 min, then control the temperature of the water bath at 45°C, stir overnight, take a sample for LC-MS detection, and the reaction is basically complete, add 4 mL acetic acid, control the temperature of the water bath at 35°C, and perform rearrangement reaction for 3 h, take a sample for LC-MS detection, and the reaction is basically complete; then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 100:0, equilibrium 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-20% B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; The collected product was modified with hyaluronic acid to obtain a cosmetic peptide, and the mass spectrum and high performance liquid chromatography test results thereof are shown in Figure 23 and Figure 24 .

[0072] Example 6: A method for synthesizing H-Gly-Pro-Gln-Gly-Pro-Gln-OH, comprising: 10 mmol AM resin was placed in a 100 mL solid phase synthesis reactor, 30 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, filtered, the deprotection solution was removed, and then washed with 6 times of DMF solution, 30 mL each time, and dried for standby.

[0073] 20 mmol Fmoc-Linker, 20 mmol HOBt was taken in a 100 mL beaker, cooled to 5°C, 15 mL of DMF solution, 3.1 mL of DIC was added, and the solution in the 100 mL beaker was added to the above-mentioned 100 mL solid phase synthesis reactor, stirred for 1.5 h, the reaction was completed; the resin was washed with 3 times of DMF solution, 20 mL each time; after washing, the next reaction was carried out; 30 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, filtered, the deprotection solution was removed, and then washed with 6 times of DMF solution, 30 mL each time, and dried for standby.

[0074] 20 mmol Fmoc-Glu-OtBu, 20 mmol HOBt was taken in a 100 mL beaker, cooled to 5°C, 15 mL of DMF solution, 4.6 mL of DIC was added, and the solution in the 100 mL beaker was added to the 100 mL solid phase synthesis reactor, stirred for 1.5 h, the reaction was completed; the resin was washed with 3 times of DMF solution, 20 mL each time; after washing, the next reaction was carried out; 30 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, filtered, the deprotection solution was removed, and then washed with 6 times of DMF solution, 30 mL each time, and dried for standby.

[0075] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 15 mL, 4.6 mL DIC standing reaction 15 min, and the solution in 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, reaction is completed; resin is washed with DMF solution 3 times, 20 mL each time; after washing is completed, the next step reaction is carried out; 20%Pip / DMF (v / v) solution 30 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for standby.

[0076] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 15 mL, 4.6 mL DIC standing reaction 15 min, and the solution in 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, reaction is completed; resin is washed with DMF solution 3 times, 20 mL each time; after washing is completed, the next step reaction is carried out; 20%Pip / DMF (v / v) solution 30 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for standby.

[0077] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 15 mL, 4.6 mL DIC standing reaction 15 min, and the solution in 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, reaction is completed; resin is washed with DMF solution 3 times, 20 mL each time; after washing is completed, the next step reaction is carried out; 20%Pip / DMF (v / v) solution 30 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for standby.

[0078] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 15 mL, 4.6 mL DIC standing reaction 15 min, and the solution in 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, reaction is completed; resin is washed with DMF solution 3 times, 20 mL each time; after washing is completed, the next step reaction is carried out; 20%Pip / DMF (v / v) solution 30 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for standby.

[0079] Take 20 mmol Fmoc-Gly-OH, 20 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 4.6 mL DIC stand by reaction 15 min, and add the solution in the 100 mL beaker to the above 100 mL solid phase synthesis reactor, stir for 1.5 h, the reaction is complete; the resin is washed with DMF solution 3 times, 30 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry by suction. Then wash with methanol 2 times, 40 mL each time, DCM solution 2 times, 40 mL each time, and methanol 2 times, 40 mL each time. Vacuum drying, get H-BB1-BB2-BB3-BB4-BB5-BB6-Linker-AM resin, wherein AA1 is Gly; AA2 is Pro; AA3 is Gln; AA4 is Gly; AA5 is Pro; and AA6 is Gln.

[0080] Take 8.3 g of the above peptide resin and cut it with 50 mL of cutting solution (TFA, Tis and H2O in a volume ratio of 90:5:5) for 2.5 h, add the cutting solution to 500 mL of ether (5°C) solution, precipitate white solid, centrifuge, vacuum drying, prepared H-Gly-Pro-Gln-Gly-Pro-Gln-OH, its mass spectrum and high performance liquid chromatography characterization results are shown in Figure 25 and Figure 26 .

[0081] The preparation method of hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Gly-Pro-Gln-Gly-Pro-Gln-OH. TFA, add 5 mL of DMSO and stir to dissolve, add 0.834 g of DIEA, then add 2.23 g of hyaluronic acid, control the temperature of water bath to 45°C, stir overnight, take sample for LC-MS detection, the reaction is basically complete, add 4 mL of acetic acid, control the temperature of water bath to 35°C, and carry out rearrangement reaction for 3 h, take sample for LC-MS detection, the reaction is basically complete; then carry out reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=100:0, equilibrate for 10 min, flow rate: 10 mL / min; Load: flow rate: 10 mL / min; Elution: 0-20% B 60 min; Column washing: 80% ACN wash to baseline equilibration; Collect the product, and prepare the hyaluronic acid modified cosmetic peptide, and the mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 27 and Figure 28 .

[0082] Example 7: A method for synthesizing H-Met-Pro-{D-Phe}-Arg-{D-Trp}-Phe-Lys-Pro-Val-NH2, comprising: Place 5 mmol AM resin in a 100 mL solid-phase synthesis reactor, add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for use.

[0083] Take 10 mmol Fmoc-Linker, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 1.5 mL DIC, and let stand for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. Wash the resin with DMF solution 3 times, 20 mL each time; after washing is complete, proceed to the next reaction; add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for use.

[0084] Take 15 mmol Fmoc-Val-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 2.3 mL DIC, and let stand for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. Wash the resin with DMF solution 3 times, 20 mL each time; after washing is complete, proceed to the next reaction; add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for use.

[0085] Take 15 mmol Fmoc-Pro-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 2.3 mL DIC standing reaction 15 min, and the solution in the 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; resin with DMF solution washing 3 times, each 20 mL; after washing, the next step reaction; add 20% Pip / DMF (v / v) solution 30 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, each 30 mL, suction dry for use.

[0086] Take 15 mmol Fmoc-Pro-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 2.3 mL DIC standing reaction 15 min, and the solution in the 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; resin with DMF solution washing 3 times, each 20 mL; after washing, the next step reaction; add 20% Pip / DMF (v / v) solution 30 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, each 30 mL, suction dry for use.

[0087] Take 15 mmol Fmoc-Pro-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 2.3 mL DIC standing reaction 15 min, and the solution in the 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; resin with DMF solution washing 3 times, each 20 mL; after washing, the next step reaction; add 20% Pip / DMF (v / v) solution 40 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, each 40 mL, suction dry for use.

[0088] Take 15 mmol Fmoc-Pro-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 2.3 mL DIC standing reaction 15 min, and the solution in the 100 mL beaker is added to the above 100 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; resin with DMF solution washing 3 times, each 20 mL; after washing, the next step reaction; add 20% Pip / DMF (v / v) solution 40 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, each 40 mL, suction dry for use.

[0089] Take 15 mmol Fmoc-Arg(Pbf)-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 2.3 mL DIC, and let the reaction stand for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir the reaction for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 30 mL each time. After washing is complete, the next step is carried out; add 20% Pip / DMF (v / v) solution 40 mL, stir the reaction for 30 min, filter, remove the deprotection solution, and then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0090] Take 10 mmol Fmoc-D-Phe-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 1.5 mL DIC, and let the reaction stand for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir the reaction for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 30 mL each time; after washing is complete, the next step is carried out; add 20% Pip / DMF (v / v) solution 40 mL, stir the reaction for 30 min, filter, remove the deprotection solution, and then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0091] Take 15 mmol Fmoc-Pro-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 2.3 mL DIC, and let the reaction stand for 15 min, and then add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir the reaction for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 30 mL each time; after washing is complete, the next step is carried out; add 20% Pip / DMF (v / v) solution 40 mL, stir the reaction for 30 min, filter, remove the deprotection solution, and then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0092] Take 15 mmol Fmoc-Met-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, 2.3 mL DIC stand by reaction 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, the reaction is complete; the resin is washed with DMF solution 3 times, 30 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry by suction. Then wash with methanol 2 times, 40 mL each time, DCM solution 2 times, 40 mL each time, and methanol 2 times, 40 mL each time; vacuum drying, to obtain the peptide resin of H-Met-Pro-{D-Phe}-Arg-{D-Trp}-Phe-Lys-Pro-Val-Linker-AM resin.

[0093] Take 5.7 g of the above-mentioned peptide resin and cut it with 50 mL of cutting solution TFA / anisyl sulfide / EDT / phenol / H2O (the mass ratio of TFA:anisyl sulfide:EDT:phenol:H2O is 87.5:5:2.5:2.5:2.5) for 2.5 h, add the cutting solution to a 500 mL solution of ethyl ether (5°C), precipitate white solid, centrifuge, and vacuum dry to obtain H-Met-Pro-{D-Phe}-Arg-{D-Trp}-Phe-Lys-Pro-Val-NH2, whose mass spectrum and high performance liquid chromatogram are shown in Figure 29 and Figure 30 .

[0094] The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Met-Pro-{D-Phe}-Arg-{D-Trp}-Phe-Lys-Pro-Val-NH2.3TFA, add 5 mL of DMSO to stir and dissolve, add 0.375 g of DIEA, and then add 1 g of hyaluronic acid, control the temperature of the water bath to 45°C, and stir to react overnight; take a sample to detect LC-MS, and the reaction is basically complete; then add 4 mL of acetic acid, control the temperature of the water bath to 35°C, and perform rearrangement reaction for 3 h; take a sample to detect LC-MS, and the reaction is basically complete; then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibration: A:B = 95:5, equilibration 10 min, flow rate: 10 mL / min; Load: flow rate: 10 mL / min; Elution: 15-35%B 60 min; Column cleaning: 80%ACN clean to baseline equilibration; Collecting the qualified product, the hyaluronic acid modified cosmetic peptide is prepared, and the mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 31 and Figure 32 .

[0095] Example 8: A method for synthesizing H-Phe-Val-Ala-Pro-Phe-Pro-OH, comprising: 12.5 mmol of CTC resin was placed in a 125 mL solid-phase synthesis reactor, 25 mmol of amino acid Fmoc-Pro-OH was added, 100 mL of dichloromethane was added, 10.9 mL of DIEA was added, and the reaction was carried out at 25°C for 3 h, 15 mL of methanol was added, and the reaction was carried out for 5 min; filtration, the resin was washed with dichloromethane twice, 100 mL each time, methanol was washed twice, 100 mL each time, and DMF was washed twice, 100 mL each time; 100 mL of 20% Pip / DMF (v / v) solution was added, and the reaction was stirred for 30 min, then the deprotection solution was removed by suction filtration, and then the resin was washed with 20 mL of DMF solution 6 times, and then suction dried for use.

[0096] 20 mmol of Fmoc-Phe-OH and 20 mmol of HOBt were taken in a 50 mL beaker, cooled to 5°C, 30 mL of DMF solution, 3.1 mL of DIC was added, and the reaction was carried out for 15 min, and then the solution in a 100 mL beaker was added to the above-mentioned 250 mL solid-phase synthesis reactor, and the reaction was carried out for 1.5 h, and then the reaction was completed; the resin was washed with 100 mL of DMF solution 3 times; after the washing was completed, the next step reaction was carried out; 100 mL of 20% Pip / DMF (v / v) solution was added, and the reaction was stirred for 30 min, then the deprotection solution was removed by suction filtration, and then the resin was washed with 20 mL of DMF solution 6 times, and then suction dried for use.

[0097] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in 50 mL beaker, cooling to 5℃, add DMF solution 30 mL, 3.1 mL DIC, stand for reaction 15 min, and the solution in 100 mL beaker is added to the above 250 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 100 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 100 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 20 mL each time, suction dry for use.

[0098] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in 50 mL beaker, cooling to 5℃, add DMF solution 30 mL, 3.1 mL DIC, stand for reaction 15 min, and the solution in 100 mL beaker is added to the above 250 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 100 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 100 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 20 mL each time, suction dry for use.

[0099] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in 50 mL beaker, cooling to 5℃, add DMF solution 30 mL, 3.1 mL DIC, stand for reaction 15 min, and the solution in 100 mL beaker is added to the above 250 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 100 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 100 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 20 mL each time, suction dry for use.

[0100] Take 20 mmol Fmoc-Phe-OH, 20 mmol HOBt in a 50 mL beaker, cool to 5℃, add DMF solution 30 mL, 3.1 mL DIC, stand for reaction 15 min, and add the solution in a 100 mL beaker to the above 250 mL solid phase synthesis reactor, stir for 1.5 h, the reaction is complete; the resin is washed with DMF solution 3 times, 100 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 100 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 20 mL each time, then wash with methanol 2 times, 100 mL each time, DCM solution 2 times, 100 mL each time, methanol 2 times, 100 mL each time, vacuum drying, get H-Phe-Val-Ala-Pro-Phe-Pro-CTC-resin; 2 g of the above peptide resin is cut with 20 mL of cutting solution TFA / DCM (TFA and DCM in a volume ratio of 2:98) at 30℃ for 1 hour, and the cutting solution is dried to obtain H-Phe-Val-Ala-Pro-Phe-Pro-OH, and the mass spectrum and LC-MS characterization results are as shown in Figure 33 and Figure 34 The synthesis route of a hyaluronic acid modified cosmetic peptide is as follows: ; Among them, -COOX is -COONa; The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.56 g of H-Phe-Val-Ala-Pro-Phe-Pro-OH.TFA, add 6 mL of DMSO and stir to dissolve, add 0.75 g of DIEA, then add 2.2 g of hyaluronic acid, control the temperature of the water bath to 45℃, stir for overnight, take sample for LC-MS detection, and the reaction is basically complete, add 4 mL of acetic acid, control the temperature of the water bath to 35℃, and carry out rearrangement reaction for 3 h, take sample for LC-MS detection, and the reaction is basically complete; then carry out reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=95:5, equilibration for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 12-32% B for 60 min; Column washing: 80% ACN wash to baseline equilibrium; Collecting the product, three kinds of hyaluronic acid modified cosmetic peptides with different structures were obtained, respectively: Hyaluronic acid modified cosmetic peptide H2, in which n = 0, that is, y = 1 in the structure of the raw material hyaluronic acid, and the product prepared by the cosmetic peptide, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 35 and Figure 36 ; Hyaluronic acid modified cosmetic peptide H4, in which n = 1, that is, y = 2 in the structure of the raw material hyaluronic acid, and the product prepared by the cosmetic peptide, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 37 and Figure 38 ; Hyaluronic acid modified cosmetic peptide H6, in which n = 2, that is, y = 3 in the structure of the raw material hyaluronic acid, and the product prepared by the cosmetic peptide, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 39 and Figure 40 .

[0101] Example 9: A method for synthesizing H-His-D-Phe-Arg-NH2, comprising: Put 10 mmol AM resin into a 250 mL solid phase synthesis reactor, add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 70 mL each time, and dry for use.

[0102] Take 20 mmol Fmoc-Linker, 20 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 35 mL, 3.1 mL DIC, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid phase synthesis reactor, stir for 1.5 h, the reaction is complete; the resin is washed with DMF solution 3 times, 70 mL each time; after washing, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 70 mL each time, and dry for use.

[0103] Take 20 mmol Fmoc-Arg(Pbf)-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 35 mL, 3.1 mL DIC, standing reaction 15 min, and the solution in 100 mL beaker is added to the above 250 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 70 mL each time; after washing is completed, the next step reaction is carried out; 20% Pip / DMF (v / v) solution 70 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, 70 mL each time, suction dry for standby.

[0104] Take 20 mmol Fmoc-D-Phe-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 35 mL, 3.1 mL DIC, standing reaction 15 min, and the solution in 100 mL beaker is added to the above 250 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 70 mL each time; after washing is completed, the next step reaction is carried out; 20% Pip / DMF (v / v) solution 70 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, 70 mL each time, suction dry for standby.

[0105] Take 20 mmol Fmoc-His(Trt)-OH, 20 mmol HOBt in 100 mL beaker, cooling to 5 ℃, add DMF solution 35 mL, 20 mmol DIC, standing reaction 15 min, and the solution in 100 mL beaker is added to the above 250 mL solid phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed; the resin is washed with DMF solution 3 times, 70 mL each time; after washing is completed, the next step reaction is carried out; 20% Pip / DMF (v / v) solution 70 mL is added, stirring reaction 30 min, suction filtration, remove the deprotection solution, then washed with DMF solution 6 times, 40 mL each time, suction dry for standby. Then methanol is added to wash 2 times, 70 mL each time, DCM solution is washed 2 times, 70 mL each time, methanol is washed 2 times, 70 mL each time. Vacuum drying, H-His(Trt)-D-Phe-Arg(Pbf)-Linker-AM resin peptide resin is obtained; The above peptide resin is cleaved with TFA / Tis / H2O (the volume ratio of TFA, Tis and H2O is 90:5:5), the amount is 100 mL, the time is 2.5 h, the cleavage solution is added to 1000 mL ether (5 ℃) solution, white solid is precipitated, centrifuged, vacuum dried, H-His-D-Phe-Arg-NH2 is prepared, and the mass spectrum and LC-MS characterization results are as followsFigure 41 and Figure 42 A synthetic route of the hyaluronic acid modified cosmetic peptide is shown in the following formula: ; wherein n = 2, -COOX is -COONa; A preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: 0.5 g of H-His-D-Phe-Arg-NH2.3TFA was weighed, stirred and dissolved in 5 mL of DMSO, 661 mg of DIEA was added, and then 1.94 g of hyaluronic acid was added. The water bath was controlled at 45°C, and the stirring reaction was carried out overnight. The sample was detected by LC-MS, and the reaction was basically complete. 4 mL of acetic acid was added, the water bath was controlled at 35°C, and the rearrangement reaction was carried out for 3 h. The sample was detected by LC-MS, and the reaction was basically complete. Then reverse phase chromatography purification was carried out, and the purification conditions were as follows: Dissolution: 0.5 g of the crude product was diluted with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 100:0, equilibrium for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-20% B for 60 min; Column cleaning: 80% ACN was used to clean to the baseline equilibrium; The qualified product was collected, and three kinds of hyaluronic acid modified cosmetic peptides were obtained, which were respectively: The hyaluronic acid modified cosmetic peptide L2, wherein n = 0, that is, y = 1 in the structure of the raw material hyaluronic acid, and the product prepared by the cosmetic peptide, the mass spectrum and the high performance liquid chromatography characterization results are shown in Figure 43 and Figure 44 ; The hyaluronic acid modified cosmetic peptide L4, wherein n = 1, that is, y = 2 in the structure of the raw material hyaluronic acid, and the product prepared by the cosmetic peptide, the mass spectrum and the high performance liquid chromatography characterization results are shown in Figure 45 and Figure 46 ; The hyaluronic acid modified cosmetic peptide L6, wherein n = 2, that is, y = 3 in the structure of the raw material hyaluronic acid, and the product prepared by the cosmetic peptide, the mass spectrum and the high performance liquid chromatography characterization results are shown in Figure 47 and Figure 48 .

[0106] Example 10: A synthesis method of H-Tyr-Pro-Phe-Phe-NH2, comprising: Take 10 mmol AM resin, place it in a 250 mL solid-phase synthesis reactor, add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 70 mL each time, and dry for use.

[0107] Take 20 mmol Fmoc-Linker, 20 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 35 mL, DIC 3.1 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 70 mL each time; after washing is complete, the next step is carried out; add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 70 mL each time, and dry for use.

[0108] Take 20 mmol Fmoc-Linker, 20 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 35 mL, DIC 3.1 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 70 mL each time; after washing is complete, the next step is carried out; add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 70 mL each time, and dry for use.

[0109] Take 20 mmol Fmoc-Linker, 20 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 35 mL, DIC 3.1 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete; the resin is washed with DMF solution 3 times, 70 mL each time; after washing is complete, the next step is carried out; add 20% Pip / DMF (v / v) solution 70 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 70 mL each time, and dry for use.

[0110] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 35 mL, DIC 3.1 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 70 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 70 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 70 mL each time, and suction dry for standby.

[0111] Take 20 mmol Fmoc-Pro-OH, 20 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 35 mL, DIC 3.1 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 70 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 70 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 70 mL each time, and suction dry for standby. The above-mentioned peptide resin is cleaved with TFA / Tis / H2O (the volume ratio of TFA, Tis and H2O is 90:5:5), the amount is 100 mL, and the time is 2.5 h. The cleavage solution is added to a 1000 mL ether (5℃) solution, white solid is precipitated, centrifuged, and vacuum dried to obtain H-Tyr-Pro-Phe-Phe-NH2, and the mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 49 and Figure 50

[0112] The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Tyr-Pro-Phe-Phe-NH2.TFA, add 5 mL of DMSO to stir and dissolve, add 0.771 g of DIEA, and then add 2.27 g of hyaluronic acid. Stir for reaction overnight in a water bath controlled at 45℃. Take a sample for LC-MS detection, and the reaction is basically complete. Add 4 mL of acetic acid, and perform rearrangement reaction in a water bath controlled at 35℃ for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then, reverse phase chromatography purification is carried out, and the purification conditions are as follows:​ Dissolution: take 0.2 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 95:5, equilibrium for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 12-32% B for 60 min; Column cleaning: clean to baseline with 80% ACN; Collect the qualified product to prepare the hyaluronic acid modified cosmetic peptide, and the mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 51 and Figure 52 .

[0113] Example 11: The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-β-Ala-His-OH.TFA, stir and dissolve in 10 mL of DMSO, add 0.5 g of TFA, stir for 10 min, then add 2.5 g of DIEA, then add 6.85 g of hyaluronic acid, control the temperature in a water bath at 45°C, stir and react overnight, take a sample for LC-MS detection, and the reaction is basically complete, add 4 mL of acetic acid, control the temperature in a water bath at 35°C, and perform rearrangement reaction for 3 h, take a sample for LC-MS detection, and the reaction is basically complete; then perform reverse chromatography purification, and the purification conditions are: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 100:0, equilibrium for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-10% B for 60 min; Column cleaning: clean to baseline with 80% ACN; Collect the qualified product to prepare the hyaluronic acid modified cosmetic peptide, and the mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 53 and Figure 54The peak position of high performance liquid chromatography is 6.460 min, and the molecular weight of the product is 872.3 / 582.0. The mass spectrum and high performance liquid chromatography characterization results of the product of the cosmetic peptide and one six-sugar sodium hyaluronate and one four-sugar sodium hyaluronate are shown in Figure 55 and Figure 56 The peak position in the high performance liquid chromatography is 7.202 min, and the molecular weight of the product is 708.4 / 1061.8 / 1415.8.

[0114] Example 12: A method for synthesizing H-Glu-Glu-Met-Gln-Arg-Arg-Ala-OH, comprising: 7.5 mmol Wang resin was placed in a 125 mL solid phase synthesis reactor, 15 mmol of amino acid Fmoc-Ala-OH was added, 50 mL of dichloromethane was added, 4.8 mL of pyridine, 4.2 mL of DBU was added, and the reaction was carried out at 25°C for 3 h. Filtration was performed, and the resin was washed with DMF solution 3 times, each time 50 mL. 50 mL of capping liquid Ac2O / DMF / DIEA (volume ratio of Ac2O, DMF and DIEA is 10:84:6) was added and reacted for 15 min. Filtration was performed, and the resin was washed with dichloromethane 2 times, each time 50 mL, methanol 2 times, each time 50 mL, and DMF 2 times, each time 50 mL. 50 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, and then filtered to remove the deprotection liquid. Then the resin was washed with DMF solution 6 times, each time 50 mL, and dried for use.

[0115] 15 mmol of Fmoc-Arg(Pbf)-OH and 15 mmol of HOBt were taken in a 50 mL beaker, cooled to 5°C, and 20 mL of DMF solution, 2.32 mL of DIC was added. The solution in the 50 mL beaker was added to the above-mentioned 125 mL solid phase synthesis reactor, and the reaction was carried out for 1.5 h. The resin was washed with DMF solution 3 times, each time 15 mL. After washing, the next step was carried out. 50 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, and then filtered to remove the deprotection liquid. Then the resin was washed with DMF solution 6 times, each time 50 mL, and dried for use.

[0116] Take 15 mmol Fmoc-Arg(Pbf)-OH, 15 mmol HOBt in a 50 mL beaker, cool to 5°C, add DMF solution 20 mL, DIC 2.32 mL, stand for reaction for 15 min, and add the solution in the 50 mL beaker to the above-mentioned 125 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 15 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 50 mL, stir for 30 min, filter, remove the deprotection solution, and then wash with DMF solution 6 times, 50 mL each time, and dry for use.

[0117] Take 15 mmol Fmoc-Arg(Pbf)-OH, 15 mmol HOBt in a 50 mL beaker, cool to 5°C, add DMF solution 20 mL, DIC 2.32 mL, stand for reaction for 15 min, and add the solution in the 50 mL beaker to the above-mentioned 125 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 15 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 50 mL, stir for 30 min, filter, remove the deprotection solution, and then wash with DMF solution 6 times, 50 mL each time, and dry for use.

[0118] Take 15 mmol Fmoc-Arg(Pbf)-OH, 15 mmol HOBt in a 50 mL beaker, cool to 5°C, add DMF solution 20 mL, DIC 2.32 mL, stand for reaction for 15 min, and add the solution in the 50 mL beaker to the above-mentioned 125 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 15 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 50 mL, stir for 30 min, filter, remove the deprotection solution, and then wash with DMF solution 6 times, 50 mL each time, and dry for use.

[0119] Take 15 mmol Fmoc-Arg(Pbf)-OH, 15 mmol HOBt in a 50 mL beaker, cool to 5°C, add DMF solution 20 mL, DIC 2.32 mL, stand for reaction for 15 min, and add the solution in the 50 mL beaker to the above-mentioned 125 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 15 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 50 mL, stir for 30 min, filter, remove the deprotection solution, and then wash with DMF solution 6 times, 50 mL each time, and dry for use.

[0120] Take 15 mmol Fmoc-Glu(OtBu)-OH, 15 mmol HOBt in a 50 mL beaker, cool to 5℃, add DMF solution 20 mL, DIC 2.32 mL, stand for 15 min, and add the solution in the 50 mL beaker to the above-mentioned 125 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 15 mL each time. After washing, the next step is carried out; add 20% Pip / DMF (v / v) solution 50 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 50 mL each time, methanol 2 times, 50 mL each time, DCM solution 2 times, 50 mL each time, and methanol 2 times, 50 mL each time. Vacuum drying, get H-AA1-AA2-AA3-AA4-AA5-AA6-AA7-Wang-resin, wherein AA1 is Glu(OtBu); AA2 is Glu(OtBu); AA3 is Met; AA4 is Gln(Trt); AA5 is Arg(Pbf); AA6 is Arg(Pbf); AA7 is Ala.

[0121] Take 4.22 g of the above-mentioned peptide resin and cut it with 40 mL of TFA / anisole / phenol / H2O / EDT (TFA, anisole, phenol, H2O and EDT in a mass ratio of 87.5:5:2.5:2.5:2.5) for 2.5 h, add the cutting solution to 400 mL of ethyl ether (5℃) solution, precipitate white solid, centrifuge, and vacuum dry to obtain H-Glu-Glu-Met-Gln-Arg-Arg-Ala-OH, whose mass spectrum and high performance liquid chromatography characterization results are shown in Figure 57 and Figure 58 .

[0122] The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Glu-Glu-Met-Gln-Arg-Arg-Ala-OH.3TFA, add 5 mL of DMSO and stir to dissolve, then add 0.46 g of DIEA, and then add 1.24 g of hyaluronic acid, control the temperature of the water bath to 45℃, and stir to react overnight. Take a sample for LC-MS detection, and the reaction is basically complete. Add 4 mL of acetic acid, control the temperature of the water bath to 35℃, and carry out rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then carry out reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 100:0, equilibrium 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-10% B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; Collecting the qualified product, hyaluronic acid modified cosmetic peptide is prepared, and the high performance liquid chromatography test result is as shown in Figure 59 The sample at 11.577 min is the product of the combination of cosmetic peptide and one tetrasaccharide sodium hyaluronate, and the mass spectrum characterization result is as shown in Figure 60 The sample at 12.223 min is the product of the combination of cosmetic peptide and one hexasaccharide sodium hyaluronate, and the mass spectrum characterization result is as shown in Figure 61

[0123] Example 13: A synthesis method of H-Asp-Val-Lys-Tyr-OH, comprising: 6.25 mmol of CTC resin was placed in a 250 mL solid phase synthesis reactor, 12.5 mmol of amino acid Fmoc-Tyr(tBu)-OH was added, 120 mL of dichloromethane was added, 8.7 mL of DIEA was added, and the reaction was carried out at 25°C for 3 h, 12.5 mL of methanol was added, and the reaction was carried out for 5 min. Filtration, the resin was washed with dichloromethane twice, 150 mL each time, methanol was washed twice, 150 mL each time, and DMF was washed twice, 390 mL each time. 65 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, filtered, and the deprotection solution was removed, then washed with DMF solution for 6 times, 120 mL each time, and dried for standby.

[0124] 15 mmol of Fmoc-Lys(Boc)-OH and 15 mmol of HOBt were taken in a 100 mL beaker, cooled to 5°C, 14 mL of DMF solution, 1.89 mL of DIC was added, and the reaction was carried out for 15 min. The solution in the 100 mL beaker was added to the above-mentioned 250 mL solid phase synthesis reactor, and the reaction was carried out for 1.5 h. The resin was washed with DMF solution for 3 times, 120 mL each time. After washing, the next step reaction was carried out. 65 mL of 20% Pip / DMF (v / v) solution was added, stirred for 30 min, filtered, and the deprotection solution was removed, then washed with DMF solution for 6 times, 120 mL each time, and dried for standby.

[0125] ​Take 15 mmol Fmoc-Val-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 10 mL, DIC 2.32 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 120 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 65 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 120 mL each time, and suction dry for standby.

[0126] Take 15 mmol Fmoc-Val-OH, 15 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 10 mL, DIC 2.32 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 250 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 120 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 65 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 120 mL each time. Then wash with methanol 2 times, 125 mL each time, DCM solution 2 times, 125 mL each time, methanol 2 times, 125 mL each time, vacuum dry to obtain H-Asp(OtBu)-Val-Lys(Boc)-Tyr(tBu)-CTC-resin; Cut 4.89 g of the above-mentioned peptide resin with TFA / Tis / H2O (the volume ratio of TFA, Tis and H2O is 90:5:5), the amount is 40 mL, stir for reaction for 2.5 h at 30℃, filter, remove the resin to obtain the filtrate. The filtrate is pulled dry to obtain the crude peptide H-Asp-Val-Lys-Tyr-OH, and the mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 62 and Figure 63 .

[0127] The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Asp-Val-Lys-Tyr-OH.2TFA, add 7 mL of DMSO to stir and dissolve, then add 0.773 g of DIEA, and then add 2.1 g of hyaluronic acid. Control the temperature of the water bath to 45℃, stir for 15 min to dissolve, and then stir for reaction overnight. Take a sample to detect LC-MS, and the reaction is basically complete. Then add 4 mL of acetic acid, control the temperature of the water bath to 35℃, and carry out rearrangement reaction for 3 h. Take a sample to detect LC-MS, and the reaction is basically complete. Then carry out reverse phase chromatography purification, and the purification conditions are: Dissolution: 0.5 g of the crude product was diluted with 100 mL of H2O; Filler: 21.2*250mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-20% B for 60 min; Column cleaning: 80% ACN was used to clean to the baseline equilibrium; The product was collected, and hyaluronic acid modified cosmetic peptides were prepared, and the high performance liquid chromatography test results are shown in Figure 64 The sample at 8.226 min is the product of cosmetic peptides combined with two six-sugar sodium hyaluronate, and the mass spectrometry characterization results are shown in Figure 65 The sample at 12.711 min is the product of cosmetic peptides combined with one six-sugar sodium hyaluronate, one disaccharide sodium hyaluronate, or cosmetic peptides combined with two four-sugar sodium hyaluronate, and the mass spectrometry characterization results are shown in Figure 66 .

[0128] Example 14: A method for synthesizing H-His-Ala-Leu-Arg-Phe-Trp-NH2, comprising: 30 mmol AM resin was placed in a 500 mL solid-phase synthesis reactor, 20% Pip / DMF (v / v) solution 200 mL was added, stirred for 30 min, filtered, and the deprotection solution was removed, then washed with DMF solution 6 times, 200 mL each time, and dried for use.

[0129] 10 mmol Fmoc-Linker, 60 mmol HOBt were taken in a 100 mL beaker, cooled to 5°C, 60 mL of DMF solution, 9.3 mL of DIC were added, and the solution in the 100 mL beaker was added to the above-mentioned 500 mL solid-phase synthesis reactor, stirred for 1.5 h, and the reaction was completed. The resin was washed with DMF solution 3 times, 100 mL each time. After washing, the next reaction was carried out; 20% Pip / DMF (v / v) solution 200 mL was added, stirred for 30 min, filtered, and the deprotection solution was removed, then washed with DMF solution 6 times, 200 mL each time, and dried for use.

[0130] Take 60 mmol Fmoc-Trp(Boc)-OH, 60 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 60 mL, DIC 9.3 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 500 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 100 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 200 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 200 mL each time, and suction dry for standby.

[0131] Take 60 mmol Fmoc-Phe-OH, 60 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 60 mL, DIC 9.3 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 500 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 100 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 200 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 200 mL each time, and suction dry for standby.

[0132] Take 60 mmol Fmoc-Arg(Pbf)-OH, 60 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 60 mL, DIC 9.3 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 500 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 100 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 200 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 200 mL each time, and suction dry for standby.

[0133] Take 60 mmol Fmoc-Leu-OH, 60 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 60 mL, DIC 9.3 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 500 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 100 mL each time. After the washing is complete, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 200 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 200 mL each time, and suction dry for standby.

[0134] Take 60 mmol Fmoc-Leu-OH, 60 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 60 mL, DIC 9.3 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 500 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 100 mL each time. After the washing is complete, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 200 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 200 mL each time, and suction dry for standby.

[0135] Take 60 mmol Fmoc-Leu-OH, 60 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 60 mL, DIC 9.3 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 500 mL solid-phase synthesis reactor, stir for reaction for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 100 mL each time. After the washing is complete, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 200 mL, stir for reaction for 30 min, suction filter, remove the deprotection solution, and then wash with DMF solution 6 times, 200 mL each time, and suction dry for standby.

[0136] The peptide resin was cleaved with 690 mL TFA / anisole / phenol / H2O / EDT (the mass ratio of TFA, anisole, phenol, H2O and EDT is 87.5:5:2.5:2.5:2.5) for 2.5 h, and the cleavage solution was added to 7000 mL of ethyl ether (5°C) solution to precipitate white solid, centrifuged, vacuum dried to obtain H-His-Ala-Leu-Arg-Phe-Trp-NH2, and the mass spectrum and high performance liquid chromatography characterization results are shown in Figure 67 and Figure 68 .

[0137] A synthetic route of a hyaluronic acid modified cosmetic peptide is as follows: ; wherein -COOX is -COONa; A preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: 1 g of H-His-Ala-Leu-Arg-Phe-Trp-NH2.TFA was weighed, stirred and dissolved in 10 mL of DMSO, 1.28 g of DIEA was added, stirred for 5 min, then 3.75 g of hyaluronic acid was added, and the water bath was controlled at 45°C, and the stirring reaction was carried out overnight. Sample detection LC-MS, the reaction was basically complete, 8 mL of acetic acid was added, the water bath was controlled at 35°C, and the rearrangement reaction was carried out for 3 h. Sample detection LC-MS, the reaction was basically complete; then reverse phase chromatography purification was carried out, and the purification conditions were as follows: Dissolution: 0.2 g of crude product was diluted with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=95:5, equilibrium for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 12-32% B for 60 min; Column cleaning: 80% ACN was used to clean to the baseline equilibrium; The qualified product was collected to obtain three kinds of hyaluronic acid modified cosmetic peptides, as shown below: A2, and the mass spectrum and high performance liquid chromatography characterization results are shown in Figure 69 and Figure 70 . A4, and the mass spectrum and high performance liquid chromatography characterization results are shown in Figure 71 and Figure 72 , the nuclear magnetic hydrogen spectrum is shown in Figure 73-1 to Figure 73-5 , and the carbon spectrum is shown inFigure 74-1 to 74-5 The two-dimensional cosy spectrum thereof is shown in Figure 1 1. Figure 75 The mass spectrum and high performance liquid chromatography characterization results of A6 are shown in Figures 1 1 and 12. Figure 76 Figure 77

[0138] Example 15: A method for preparing a hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-β-Ala-Pro-Dab-NH-Bzl.TFA, add 5 mL of DMSO to stir and dissolve, add 704.32 mg of DIEA, stir for 5 min, then add 2064.08 mg of hyaluronic acid, control the temperature of the water bath at 45°C, stir and react overnight, take a sample for LC-MS detection, and the reaction is basically complete. Add 3 mL of acetic acid, control the temperature of the water bath at 35°C, and perform rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.2 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 95:5, equilibrate for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 12-32% B for 60 min; Column cleaning: clean with 80% ACN to the baseline equilibrium; Collect the qualified product to obtain the hyaluronic acid modified cosmetic peptide.

[0139] Example 16: A method for preparing a hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Dab-Val-Dab-OH.2TFA, add 7 mL of DMSO to stir and dissolve, add 833.2 mg of DIEA, stir for 5 min, then add 2441.79 mg of hyaluronic acid, control the temperature of the water bath at 45°C, and stir and react overnight. Take a sample for LC-MS detection, and the reaction is basically complete. Add 3 mL of acetic acid, control the temperature of the water bath at 35°C, and perform rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; ​​​Filler: 21.2*250mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-20% B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; Collect the qualified product, and prepare the hyaluronic acid modified cosmetic peptide.

[0140] Example 17: The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Lys-Val-Lys-OH.2TFA, add 7 mL of DMSO, stir and dissolve, then add 708.03 mg of DIEA, stir for 5 min, then add 2074.97 mg of hyaluronic acid, control the temperature in a water bath at 45°C, and stir overnight. Take a sample for LC-MS detection, and the reaction is basically complete. Add 3 mL of acetic acid, control the temperature in a water bath at 35°C, and perform rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-20% B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; Collect the qualified product, and prepare the hyaluronic acid modified cosmetic peptide.

[0141] Example 18: The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5g H-Gln-Asp-Val-His-OH.TFA, add 5mL DMSO to stir and dissolve, add 531.55mg DIEA, stir for 5min, then add 1557.76mg hyaluronic acid, water bath temperature control 45℃, stir overnight, sample detection LC-MS, basically complete reaction, add 3mL acetic acid, water bath temperature control 35℃, rearrangement reaction for 3h, sample detection LC-MS, basically complete reaction; then reverse phase chromatography purification, purification conditions: Dissolution: take 0.2g of crude product and dilute with 100mL H2O; Filler: 21.2*250mm, 10-120, C18; flow rate: 10mL / min; wavelength: 220nm; Mobile phase: A: 1%HAc; B: ACN; Equilibrium: A:B=95:5, equilibrium 10min, flow rate: 10mL / min; Loading: flow rate: 10mL / min; Elution: 12-32%B 60min; Column cleaning: 80%ACN cleaning to baseline equilibrium; Collect the qualified product, and prepare the hyaluronic acid modified cosmetic peptide.

[0142] Example 19: The method for preparing the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5g H-Pro-Pro-Tyr-Leu-OH.TFA, add 5mL DMSO to stir and dissolve, add 541.25mg DIEA, stir for 5min, then add 1586.20mg hyaluronic acid, water bath temperature control 45℃, stir overnight, sample detection LC-MS, basically complete reaction, add 3mL acetic acid, water bath temperature control 35℃, rearrangement reaction for 3h, sample detection LC-MS, basically complete reaction; then reverse phase chromatography purification, purification conditions: Dissolution: take 0.2g of crude product and dilute with 100mL H2O; Filler: 21.2*250mm, 10-120, C18; flow rate: 10mL / min; wavelength: 220nm; Mobile phase: A: 1%HAc; B: ACN; Equilibrium: A:B=95:5, equilibrium 10min, flow rate: 10mL / min; Loading: flow rate: 10mL / min; Elution: 12-32%B 60min; Column cleaning: 80%ACN cleaning to baseline equilibrium; The collected product is a hyaluronic acid modified cosmetic peptide.

[0143] Example 20: A method for preparing a hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Pro-Lys-Glu-Lys-OH.2TFA, add 7 mL of DMSO and stir to dissolve, then add 528.27 mg of DIEA, stir for 5 min, then add 1548.14 mg of hyaluronic acid, control the temperature in a water bath at 45°C, and stir overnight. Take a sample for LC-MS detection, and the reaction is basically complete. Add 3 mL of acetic acid, control the temperature in a water bath at 35°C, and perform rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-20% B for 60 min; Column cleaning: clean with 80% ACN to baseline equilibrium; The collected product is a hyaluronic acid modified cosmetic peptide.

[0144] Example 21: A method for preparing a hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Val-Trp-OH.2TFA, add 7 mL of DMSO and stir to dissolve, then add 871.74 mg of DIEA, stir for 5 min, then add 2554.72 mg of hyaluronic acid, control the temperature in a water bath at 45°C, and stir overnight. Take a sample for LC-MS detection, and the reaction is basically complete. Add 3 mL of acetic acid, control the temperature in a water bath at 35°C, and perform rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium for 10 min, flow rate: 10 mL / min; Load: flow rate: 10 mL / min; Elution: 0-20% B 60 min; Column washing: 80% ACN wash to baseline equilibration; Collect the qualified product, and prepare the hyaluronic acid modified cosmetic peptide.

[0145] Example 22: The method for preparing the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Ser-Val-Val-Val-Arg-Thr-NH2.TFA, stir and dissolve in 5 mL of DMSO, add 404.41 mg of DIEA, stir for 5 min, then add 1176.38 mg of hyaluronic acid, control the temperature of the water bath at 45°C, and stir overnight. Take a sample for LC-MS detection, and the reaction is basically complete. Add 3 mL of acetic acid, control the temperature of the water bath at 35°C, and perform rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.2 g of the crude product, dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibration: A:B = 95:5, equilibration for 10 min, flow rate: 10 mL / min; Load: flow rate: 10 mL / min; Elution: 12-32% B 60 min; Column washing: 80% ACN wash to baseline equilibration; Collect the qualified product, and prepare the hyaluronic acid modified cosmetic peptide.

[0146] Example 23: The method for preparing the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of Pal-Lys-Met(O)2-Lys-OH.2TFA, stir and dissolve in 7 mL of DMSO, add 391.22 mg of DIEA, stir for 5 min, then add 1146.50 mg of hyaluronic acid, control the temperature of the water bath at 45°C, and stir overnight. Take a sample for LC-MS detection, and the reaction is basically complete. Add 3 mL of acetic acid, control the temperature of the water bath at 35°C, and perform rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product, dilute with 100 mL of H2O; Filler: 21.2*250mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 100:0, equilibrium 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-20% B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; Collect the qualified product, and prepare the hyaluronic acid modified cosmetic peptide.

[0147] Example 24: The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5 g of H-Lys-Met(O)2-Lys-OH.2TFA, add 7 mL of DMSO, stir and dissolve, then add 604.37 mg of DIEA, stir for 5 min, then add 1771.19 mg of hyaluronic acid, control the temperature in a water bath at 45°C, and stir overnight. Take a sample for LC-MS detection, and the reaction is basically complete. Add 3 mL of acetic acid, control the temperature in a water bath at 35°C, and perform rearrangement reaction for 3 h. Take a sample for LC-MS detection, and the reaction is basically complete. Then perform reverse phase chromatography purification, and the purification conditions are as follows: Dissolution: take 0.5 g of the crude product and dilute with 100 mL of H2O; Filler: 21.2*250mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B = 100:0, equilibrium 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 0-20% B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; Collect the qualified product, and prepare the hyaluronic acid modified cosmetic peptide.

[0148] Example 25: The preparation method of the hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5g H-Val-Gly-Val-Ala-Pro-Gly-OH.2TFA, add 7mL DMSO to stir and dissolve, then add 530.41mg DIEA, stir for 5min, then add 1554.41mg hyaluronic acid, control the temperature at 45°C in water bath, then stir overnight, take sample to detect LC-MS, basically complete reaction, add 3mL acetic acid, control the temperature at 35°C in water bath, carry out rearrangement reaction for 3h, take sample to detect LC-MS, basically complete reaction; then carry out reverse phase chromatography purification, purification conditions: Dissolution: take 0.5g crude product and dilute with 100mL H2O; Filler: 21.2*250mm, 10-120, C18; flow rate: 10mL / min; wavelength: 220nm; Mobile phase: A: 1%HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium for 10min, flow rate: 10mL / min; Loading: flow rate: 10mL / min; Elution: 0-20%B 60min; Column cleaning: clean with 80%ACN to baseline equilibrium; Collect the qualified product to prepare hyaluronic acid modified cosmetic peptide.

[0149] Example 26: The method for preparing hyaluronic acid modified cosmetic peptide comprises the following steps: Take 0.5g H-bataAla-His-Ser-His-OH.2TFA, add 7mL DMSO to stir and dissolve, then add 587.07mg DIEA, stir for 5min, then add 1720.46mg hyaluronic acid, control the temperature at 45°C in water bath, then stir overnight, take sample to detect LC-MS, basically complete reaction, add 3mL acetic acid, control the temperature at 35°C in water bath, carry out rearrangement reaction for 3h, take sample to detect LC-MS, basically complete reaction; then carry out reverse phase chromatography purification, purification conditions: Dissolution: take 0.5g crude product and dilute with 100mL H2O; Filler: 21.2*250mm, 10-120, C18; flow rate: 10mL / min; wavelength: 220nm; Mobile phase: A: 1%HAc; B: ACN; Equilibrium: A:B=100:0, equilibrium for 10min, flow rate: 10mL / min; Loading: flow rate: 10mL / min; Elution: 0-20%B 60min; Column washing: 80% ACN wash to baseline equilibrium; The collected product is hyaluronic acid modified cosmetic peptide.

[0150] Example 27: A method for synthesizing H-Trp-Phe-Arg-D-Leu-Ala-His-NH2, comprising: Put 5 mmol AM resin in a 100 mL solid-phase synthesis reactor, add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for standby.

[0151] Take 10 mmol Fmoc-Linker, 10 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. Wash the resin with DMF solution 3 times, 20 mL each time. After washing is complete, proceed to the next reaction; add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for standby.

[0152] Take 10 mmol Fmoc-His(Trt)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. Wash the resin with DMF solution 3 times, 20 mL each time. After washing is complete, proceed to the next reaction. Add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for standby.

[0153] Take 10 mmol Fmoc-Ala-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for use.

[0154] Take 10 mmol Fmoc-D-Leu-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for use.

[0155] Take 10 mmol Fmoc-Arg(Pbf)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0156] Take 10 mmol Fmoc-Phe-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use 15 mmol of Fmoc-Trp(Boc)-OH and 1 mmol of HOBt were placed in a 100 mL beaker and cooled to 5°C. 15 mL of DMF solution and 2.3 mL of DIC were added and allowed to react for 15 minutes. The solution in the 100 mL beaker was then added to the 100 mL solid-phase synthesis reactor described above and stirred for 1.5 hours until the reaction was complete. The resin was washed three times with 30 mL of DMF solution. After washing, the resin was proceeded to the next step. 40 mL of a 20% Pip / DMF (v / v) solution was added and stirred for 30 minutes. The deprotection solution was removed by filtration and then washed six times with 40 mL of DMF solution. The resin was then dried and set aside. The resin was then washed twice with 40 mL of methanol, twice with 40 mL of DCM solution, and twice with 40 mL of methanol. Drying under vacuum gave a peptide resin of H-AA1-AA2-AA3-AA4-AA5-AA6-Linker-AM resin, wherein AA1 is Trp(Boc); AA2 is Phe; AA3 is Arg; AA4 is D-Leu; AA5 is Ala; and AA6 is His(Trt).

[0157] The peptide resin was cleaved with 110 mL of TFA / thioanisole / phenol / H2O / EDT (the mass ratio of TFA, thioanisole, phenol, H2O and EDT was 87.5:5:2.5:2.5:2.5) for 2.5 h. The cleavage solution was added to 1000 mL of ether (5°C) to precipitate a white solid, which was centrifuged and dried in vacuo to obtain H-Trp-Phe-Arg-D-Leu-Ala-His-NH2. Its mass spectrometry and HPLC characterization results are shown in Figure 2. Figure 78 and Figure 79 shown.

[0158] A synthetic route for a hyaluronic acid-modified beauty peptide is: ; Wherein, -COOX is -COONa; The preparation method of hyaluronic acid-modified cosmetic peptide comprises the following steps: Weigh 1g of H-Trp-Phe-Arg-D-Leu-Ala-His-NH2.TFA, add 10mL of DMSO and stir to dissolve, add 1.28g of DIEA, stir for 5 minutes, then add 3.75g of hyaluronic acid, control the temperature in a water bath at 45°C, stir and react overnight, take a sample and detect by LC-MS, if the reaction is basically complete, add 8mL of acetic acid, keep the temperature constant for rearrangement reaction for 2h, take a sample and detect by LC-MS, if the reaction is basically complete; then perform reverse chromatography purification, purification conditions: Dissolution: dilute 0.2 g of crude product with 100 mL of H2O; Packing: 21.2*250mm, 10-120, C18; Flow rate: 10mL / min; Wavelength: 220nm; Mobile phase: A: 1% HAc; B: ACN; Balance: A:B=95:5, balance 10 min, flow rate: 10 mL / min; Sample loading: flow rate: 10 mL / min; Elution: 12-32%B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; Qualified products were collected to obtain three structures of hyaluronic acid-modified beauty peptides, as shown below: E2, its mass spectrometry and HPLC characterization results are shown in Figure 80 and Figure 81 As shown; E4, its mass spectrometry and HPLC characterization results are shown in Figure 82 and Figure 83 As shown; E6, its mass spectrometry and HPLC characterization results are shown in Figure 84 and Figure 85 shown.

[0159] Example 28: A method for synthesizing H-Trp-Phe-Arg-Leu-Ala-His-NH2 comprises: Place 5 mmol AM resin in a 100 mL solid-phase synthesis reactor, add 30 mL of 20% Pip / DMF (v / v) solution, stir and react for 30 min, filter to remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and drain to set aside.

[0160] Place 10 mmol of Fmoc-Linker and 10 mmol of HOBt in a 100 mL beaker, cool to 5°C, add 15 mL of DMF solution and 1.5 mL of DIC, and allow to react for 15 minutes. The solution in the 100 mL beaker was then added to the 100 mL solid-phase synthesis reactor described above and stirred for 1.5 hours until the reaction was complete. The resin was washed three times with 20 mL of DMF solution. After washing, proceed to the next step: add 30 mL of 20% Pip / DMF (v / v) solution, stir for 30 minutes, filter to remove the deprotection solution, and then wash six times with 30 mL of DMF solution before draining.

[0161] Take 10 mmol Fmoc-His(Trt)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction 15 min, and the solution in the 100 mL beaker is added to the above-mentioned 100 mL solid-phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for use.

[0162] Take 10 mmol Fmoc-His(Trt)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction 15 min, and the solution in the 100 mL beaker is added to the above-mentioned 100 mL solid-phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for use.

[0163] Take 10 mmol Fmoc-His(Trt)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction 15 min, and the solution in the 100 mL beaker is added to the above-mentioned 100 mL solid-phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for use.

[0164] Take 10 mmol Fmoc-His(Trt)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction 15 min, and the solution in the 100 mL beaker is added to the above-mentioned 100 mL solid-phase synthesis reactor, stirring reaction 1.5 h, the reaction is completed. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stirring reaction 30 min, suction filtration, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, suction dry for use.

[0165] Take 10 mmol Fmoc-Phe-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5 ℃, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use Take 15 mmol Fmoc-Trp(Boc)-OH, 1 mmol HOBt in a 100 mL beaker, cool to 5 ℃, add DMF solution 15 mL, DIC 2.3 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 30 mL each time. After washing, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use. Then wash with methanol 2 times, 40 mL each time, DCM solution 2 times, 40 mL each time, and methanol 2 times, 40 mL each time. Vacuum drying to obtain a peptide resin of H-AA1-AA2-AA3-AA4-AA5-AA6-Linker-AM resin, wherein AA1 is Trp(Boc); AA2 is Phe; AA3 is Arg; AA4 is D-Leu; AA5 is Ala; and AA6 is His(Trt).

[0166] The above-mentioned peptide resin is cut with 110 mL TFA / anisole / phenol / H2O / EDT (TFA, anisole, phenol, H2O and EDT have a mass ratio of 87.5:5:2.5:2.5:2.5) for 2.5 h, and the cutting solution is added to a 1000 mL ether (5 ℃) solution to precipitate white solids, centrifuged, and vacuum dried to obtain H-Trp-Phe-Arg-Leu-Ala-His-NH2, the mass spectrum and high performance liquid chromatography characterization results of which are shown in Figure 86 and Figure 87 .

[0167] A synthesis route of a hyaluronic acid modified cosmetic peptide is: ; wherein -COOX is -COONa; A method for preparing hyaluronic acid modified cosmetic peptides, comprising the following steps: Weigh 1 g of H-Trp-Phe-Arg-Leu-Ala-His-NH2.TFA, stir and dissolve in 10 mL of DMSO, add 1.28 g of DIEA, stir for 5 min, then add 3.75 g of hyaluronic acid, control the temperature at 45°C in water bath, stir overnight, take sample for LC-MS detection, the reaction is basically complete, add 8 mL of acetic acid, constant temperature rearrangement reaction for 2 h, take sample for LC-MS detection, the reaction is basically complete; then perform reverse phase chromatography purification, the purification conditions are as follows: Dissolution: take 0.2 g of crude product and dilute with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=95:5, equilibrium for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 12-32% B for 60 min; Column cleaning: clean with 80% ACN to baseline equilibrium; Collect the qualified product to obtain three kinds of hyaluronic acid modified cosmetic peptides, as shown below: B2, the mass spectrum and high performance liquid chromatography characterization results thereof are as shown in Figure 88 and Figure 89 B4, the mass spectrum and high performance liquid chromatography characterization results thereof are as shown in Figure 90 and Figure 91 B6, the mass spectrum and high performance liquid chromatography characterization results thereof are as shown in Figure 92 and Figure 93 .

[0168] Example 29: A method for synthesizing H-Arg-Arg-Gln-D-Met-Glu-Glu-NH2, comprising: Put 5 mmol of AM resin into a 100 mL solid phase synthesis reactor, add 30 mL of 20% Pip / DMF (v / v) solution, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for use.

[0169] ​​Take 10 mmol Fmoc-Linker, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min reaction, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out; add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for use.

[0170] Take 10 mmol Fmoc-Glu(otBu)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min reaction, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use; Take 10 mmol Fmoc-Glu(otBu)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min reaction, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use; Take 10 mmol Fmoc-D-Met-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min reaction, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0171] Take 10 mmol Fmoc-Gln(Trt)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry under suction.

[0172] Take 10 mmol Fmoc-Gln(Trt)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry under suction.

[0173] Take 10 mmol Fmoc-Gln(Trt)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry under suction.

[0174] The peptide resin was cleaved with 110 mL TFA / anisole / phenol / H2O / EDT (mass ratio of TFA, anisole, phenol, H2O and EDT: 87.5:5:2.5:2.5:2.5) for 2.5 h, and the cleavage solution was added to 1000 mL of ethyl ether (5°C) solution to precipitate white solid, centrifuged, vacuum dried to obtain H-Arg-Arg-Gln-D-Met-Glu-Glu-NH2, and the mass spectrum and high performance liquid chromatography characterization results are shown in Figure 94 and Figure 95 .

[0175] A synthetic route of a hyaluronic acid modified cosmetic peptide is as follows: ; wherein -COOX is -COONa; A preparation method of a hyaluronic acid modified cosmetic peptide comprises the following steps: 1 g of H-Arg-Arg-Gln-D-Met-Glu-Glu-NH2.TFA was weighed, stirred and dissolved in 10 mL of DMSO, 1.25 g of DIEA was added, stirred for 5 min, then 3.67 g of hyaluronic acid was added, and the water bath was controlled at 45°C, and the stirring reaction was carried out overnight. Sample detection LC-MS, the reaction was basically complete, 8 mL of acetic acid was added, and the rearrangement reaction was carried out at constant temperature for 2 h. Sample detection LC-MS, the reaction was basically complete. Then reverse phase chromatography purification was carried out, and the purification conditions were as follows: Dissolution: 0.2 g of crude product was diluted with 100 mL of H2O; Filler: 21.2*250 mm, 10-120, C18; flow rate: 10 mL / min; wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=95:5, equilibrium for 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 12-32% B for 60 min; Column cleaning: cleaned with 80% ACN to baseline equilibrium; The qualified product was collected to obtain three kinds of hyaluronic acid modified cosmetic peptides, as shown below: G2, and the mass spectrum and high performance liquid chromatography characterization results are shown in Figure 96 and Figure 97 . G4, and the mass spectrum and high performance liquid chromatography characterization results are shown in Figure 98 and Figure 99 . G6, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 100 and Figure 101 .

[0176] Example 30: A synthesis method of H-Arg-Arg-Gln-Met-Glu-Glu-NH2, comprising: Put 5 mmol AM resin into a 100 mL solid phase synthesis reactor, add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for standby.

[0177] Take 10 mmol Fmoc-Linker, 10 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is completed, the next step reaction is carried out; add 20% Pip / DMF (v / v) solution 30 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 30 mL each time, and dry for standby.

[0178] Take 10 mmol Fmoc-Glu(otBu)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for standby Take 10 mmol Fmoc-Glu(otBu)-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5℃, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction for 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is completed, the next step reaction is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for standby Take 10 mmol Fmoc-Met-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0179] Take 10 mmol Fmoc-Met-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0180] Take 10 mmol Fmoc-Met-OH, 10 mmol HOBt in a 100 mL beaker, cool to 5°C, add DMF solution 15 mL, DIC 1.5 mL, stand for reaction 15 min, and add the solution in the 100 mL beaker to the above-mentioned 100 mL solid-phase synthesis reactor, stir for 1.5 h, and the reaction is complete. The resin is washed with DMF solution 3 times, 20 mL each time. After washing is complete, the next step is carried out. Add 20% Pip / DMF (v / v) solution 40 mL, stir for 30 min, filter, remove the deprotection solution, then wash with DMF solution 6 times, 40 mL each time, and dry for use.

[0181] Place 15 mmol of Fmoc-Arg(Pbf)-OH and 15 mmol of HOBt in a 100 mL beaker, cool to 5°C, add 20 mL of DMF solution and 2.3 mL of DIC, and let the mixture react for 15 minutes. The mixture was then added to the 100 mL solid-phase synthesis reactor described above and stirred for 1.5 hours until the reaction was complete. The resin was washed three times with 20 mL of DMF solution. After washing, the resin was moved to the next step. 40 mL of a 20% Pip / DMF (v / v) solution was added and stirred for 30 minutes. The deprotection solution was removed by filtration, and the resin was then washed six times with 40 mL of DMF solution and dried. The resin was then washed twice with 40 mL of methanol, twice with 40 mL of DCM solution, and twice with 40 mL of methanol. Drying under vacuum gave a peptide resin of H-AA1-AA2-AA3-AA4-AA5-AA6-Linker-AM resin, wherein AA1 is Arg(Pbf); AA2 is Arg(Pbf); AA3 is Gln(Trt); AA4 is Met; AA5 is Glu(otBu); and AA6 is Glu(otBu).

[0182] The peptide resin was cleaved with 110 mL of TFA / thioanisole / phenol / H2O / EDT (the mass ratio of TFA, thioanisole, phenol, H2O and EDT was 87.5:5:2.5:2.5:2.5) for 2.5 h. The cleavage solution was added to 1000 mL of ether (5°C) to precipitate a white solid, which was centrifuged and dried in vacuo to obtain H-Arg-Arg-Gln-Met-Glu-Glu-NH2. Its mass spectrometry and HPLC characterization results are shown in Figure 2. Figure 102 and Figure 103 shown.

[0183] A synthetic route for a hyaluronic acid-modified beauty peptide is: ; Wherein, -COOX is -COONa; The preparation method of hyaluronic acid-modified cosmetic peptide comprises the following steps: Weigh 1g of H-Arg-Arg-Gln-Met-Glu-Glu-NH2.TFA, add 10mL of DMSO and stir to dissolve, add 1.25g of DIEA, stir for 5 minutes, then add 3.67g of hyaluronic acid, control the temperature in a water bath at 45°C, stir and react overnight, take a sample and detect by LC-MS, if the reaction is basically complete, add 8mL of acetic acid, keep the temperature constant for rearrangement reaction for 2h, take a sample and detect by LC-MS, if the reaction is basically complete, then perform reverse phase chromatography purification, purification conditions: Dissolution: dilute 0.2 g of crude product with 100 mL of H2O; Filler: 21.2*250mm, 10-120, C18; Flow rate: 10 mL / min; Wavelength: 220 nm; Mobile phase: A: 1% HAc; B: ACN; Equilibrium: A:B=95:5, equilibrium 10 min, flow rate: 10 mL / min; Loading: flow rate: 10 mL / min; Elution: 12-32% B 60 min; Column cleaning: 80% ACN cleaning to baseline equilibrium; Collecting qualified products, three kinds of hyaluronic acid modified cosmetic peptides with the following structures are obtained: F2, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 104 and Figure 105 ; F4, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 106 and Figure 107 ; F6, its mass spectrum and high performance liquid chromatography characterization results are as shown in Figure 108 and Figure 109 .

[0184] Example 31: A preparation method of a hyaluronic acid modified cosmetic peptide, comprising the following steps: Take 0.56 g of H-Phe-Val-Ala-Pro-Phe-Pro-OH.TFA (prepared in Example 8), add 6 mL of DMSO to stir and dissolve, add 0.75 g of DIEA, then add 2.2 g of hyaluronic acid (y=1), control the temperature of the water bath at 45°C, stir and react overnight, take a sample for LC-MS detection, and the reaction is basically complete. Add 4 mL of acetic acid, control the temperature of the water bath at 35°C, and carry out rearrangement reaction for 3 h, take a sample for LC-MS detection, and the reaction is basically complete. Then carry out reverse phase chromatography purification to obtain a hyaluronic acid modified cosmetic peptide.

[0185] Example 32: A preparation method of a hyaluronic acid modified cosmetic peptide, comprising the following steps: Take 0.56g H-Phe-Val-Ala-Pro-Phe-Pro-OH.TFA (prepared in Example 8), add 6mL DMSO stirring to dissolve, add 0.75g DIEA, then add 2.2g hyaluronic acid (y=2), water bath temperature control 45℃, stirring overnight, sample detection LC-MS, the reaction is substantially complete, add 4mL acetic acid, water bath temperature control 35℃, rearrangement reaction for 3h, sample detection LC-MS, the reaction is substantially complete; then reverse phase chromatography purification to obtain hyaluronic acid modified cosmetic peptide.

[0186] Example 33: A method for preparing a hyaluronic acid modified cosmetic peptide, comprising the following steps: Take 0.56g H-Phe-Val-Ala-Pro-Phe-Pro-OH.TFA (prepared in Example 8), add 6mL DMSO stirring to dissolve, add 0.75g DIEA, then add 2.2g hyaluronic acid (y=2), water bath temperature control 45℃, stirring overnight, sample detection LC-MS, the reaction is substantially complete, add 4mL acetic acid, water bath temperature control 35℃, rearrangement reaction for 3h, sample detection LC-MS, the reaction is substantially complete; then reverse phase chromatography purification to obtain hyaluronic acid modified cosmetic peptide.

[0187] Test Example 1: 1. Moisturizing performance test AQP3 content test as follows: (1) Cell inoculation: inoculate cells into 24-well plates and incubate in an incubator (37℃, 5% CO2) overnight; (2) Liquid preparation: prepare working solutions of test substances according to the experimental design (as shown in Table 1); Table 1 AQP3 experimental design table

[0188] (3) Add test substances: after incubation in an incubator (37℃, 5% CO2) for 24h, add test substances according to the table, and continue to incubate for 24h; (4) Sample collection: discard the supernatant and rinse the cells with PBS for 3 times; (5) Immunofluorescence staining: a. Add methanol to fix the cells, rinse with PBS for 3 times, and add 1mL BSA to each well for blocking for 1h; b. Discard the blocking solution, add primary antibody to each well, and place in a 4℃ refrigerator overnight. Discard the primary antibody and rinse with PBS for 3 times; c. Add secondary antibody to each well and act for 2h; discard the secondary antibody and rinse with PBS for 3 times; d. Add DAPI to each well for nuclear staining, act for 10 min, discard DAPI, rinse with PBS for 3 times, and then take pictures by using a fluorescence microscope; (6) Result analysis: the AQP3 fluorescence intensity was quantitatively analyzed by using Image Pro Plus software.

[0189] The HA content was tested as follows: (1) Cell inoculation: inoculate cells into a 24-well plate, and incubate in an incubator (37℃, 5% CO2) overnight; (2) Liquid preparation: prepare the test substance working solution according to the experimental design (Table 2); Table 2 HA experimental design table

[0190] (3) Add test substance: after 24h of incubation in an incubator (37℃, 5% CO2), add the test substance according to the table, and continue to incubate for 24h; (4) Sample collection: collect the supernatant, and determine the HA content by using an ELISA kit.

[0191] Result analysis: Table 3-1 AQP3 test results

[0192] From the data analysis in Table 3-1, it can be seen that the hyaluronic acid modified cosmetic peptides prepared in the embodiments of the present application have excellent moisturizing performance and can effectively improve the content of AQP3. Specifically, the hyaluronic acid modified cosmetic peptides D4 and D6 prepared in Example 4 of the present application have excellent moisturizing efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL, and the hyaluronic acid modified cosmetic peptides of the three structures (D2, D4, D6) mixed at a certain mass ratio also have excellent moisturizing efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. The hyaluronic acid modified cosmetic peptides of the three structures (E2, E4, E6) prepared in Example 27 of the present application mixed at a certain mass ratio have excellent moisturizing efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the hyaluronic acid modified cosmetic peptides of the three structures (B2, B4, B6) prepared in Example 28 of the present application mixed at a certain mass ratio have excellent moisturizing efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. The hyaluronic acid modified cosmetic peptides of the three structures (G2, G4, G6) prepared in Example 29 of the present application mixed at a certain mass ratio have excellent moisturizing efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. The hyaluronic acid modified cosmetic peptides of the three structures (F2, F4, F6) prepared in Example 30 of the present application mixed at a certain mass ratio have excellent moisturizing efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. The hyaluronic acid modified cosmetic peptides A4 and A6 prepared in Example 14 of the present application have excellent moisturizing efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL, and the hyaluronic acid modified cosmetic peptides of the three structures (A2, A4, A6) mixed at a certain mass ratio also have excellent moisturizing efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL.

[0193] Table 3-2 AQP3 test results

[0194] From the data analysis in Table 3-2, it can be seen that the hyaluronic acid modified cosmetic peptides prepared in the embodiments of the present application exhibit excellent promotion effect on AQP3 and better content improvement effect. Specifically, the hyaluronic acid modified cosmetic peptides prepared in embodiments 1-2, 5-7 and 10-13 of the present application have excellent moisturizing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. Moreover, the hyaluronic acid modified cosmetic peptides of three structures (D2, D4, D6) prepared in embodiment 4 of the present application, after being mixed at a certain mass ratio, have excellent moisturizing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the hyaluronic acid modified cosmetic peptides of three structures (H2, H4, H6) prepared in embodiment 8 of the present application, after being mixed at a certain mass ratio, have excellent moisturizing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the hyaluronic acid modified cosmetic peptides of three structures (L2, L4, L6) prepared in embodiment 9 of the present application, after being mixed at a certain mass ratio, have excellent moisturizing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL.

[0195] Table 4 HA test results

[0196] From the data analysis in Table 4, it can be seen that the hyaluronic acid modified cosmetic peptides prepared in embodiments 2 and 8 of the present application exhibit excellent promotion effect on HA and better content improvement effect. Specifically, the hyaluronic acid modified cosmetic peptides prepared in embodiment 2 of the present application have excellent moisturizing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. Moreover, the hyaluronic acid modified cosmetic peptides of three structures (H2, H4, H6) prepared in embodiment 8 of the present application, after being mixed at a certain mass ratio, have excellent moisturizing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL.

[0197] 2. Anti-wrinkle firming performance determination The type I collagen and MMP-1 content tests are as follows: (1) Cell inoculation: inoculate cells into a 24-well plate and incubate in an incubator (37°C, 5% CO2) overnight; (2) Liquid preparation: prepare the test substance working solution according to the experimental design (Table 5); Table 5 Experimental design table

[0198] (3) UVA radiation: after 24 hours of culture, the negative control group, the positive control group and the sample group received a total dose of 9 J / cm 2 of UVA radiation, while the blank control group was placed in the same environment (UVA radiation dose of 0 J / cm 2 ); (4) adding the test substance: according to the experimental design, after irradiation, the test substance was added to each group, 1 mL of cell culture solution was added to each well of the blank control group and the negative control group; 1 mL of cell culture solution containing vitamin C and vitamin E was added to each well of the positive control group; 1 mL of culture solution containing the corresponding concentration of the test substance was added to each well of the sample group; after the addition of the test substance was completed, the 24-well plate was placed in the incubator (37°C, 5% CO2) for 24 hours of culture; (5) collecting the supernatant for determination of the contents of collagen type I and MMP-1; (6) result analysis: t-test statistical analysis was used for comparison between groups, and statistical analysis was two-tailed.

[0199] Table 6 MMP-1 test results

[0200] Table 7-1 Collagen type I test results

[0201] From the data analysis in Table 6 and Table 7-1, it can be seen that the hyaluronic acid modified cosmetic peptides prepared in the embodiments of the present application exhibit excellent inhibitory effect on MMP-1 and excellent promoting effect on Collagen I. Specifically, the hyaluronic acid modified cosmetic peptide D4 and the hyaluronic acid modified cosmetic peptide D6 prepared in Example 4 of the present application have excellent anti-wrinkle firming efficacy at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL, and the hyaluronic acid modified cosmetic peptides of the three structures (D2, D4, D6) mixed at a certain mass ratio also have excellent anti-wrinkle firming efficacy at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. The hyaluronic acid modified cosmetic peptides of the three structures (E2, E4, E6) mixed at a certain mass ratio in Example 27 of the present application have excellent anti-wrinkle firming efficacy at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the hyaluronic acid modified cosmetic peptides of the three structures (B2, B4, B6) mixed at a certain mass ratio in Example 28 of the present application have excellent anti-wrinkle firming efficacy at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. The hyaluronic acid modified cosmetic peptides of the three structures (G2, G4, G6) mixed at a certain mass ratio in Example 29 of the present application have excellent anti-wrinkle firming efficacy at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. The hyaluronic acid modified cosmetic peptides of the three structures (F2, F4, F6) mixed at a certain mass ratio in Example 30 of the present application have excellent anti-wrinkle firming efficacy at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. The hyaluronic acid modified cosmetic peptide A4 and the hyaluronic acid modified cosmetic peptide A6 prepared in Example 14 of the present application have excellent anti-wrinkle firming efficacy at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL, and the hyaluronic acid modified cosmetic peptides of the three structures (A2, A4, A6) mixed at a certain mass ratio also have excellent anti-wrinkle firming efficacy at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL.

[0202] Table 7-2 Collagen I test results

[0203] From the data analysis in Table 7-2, it can be seen that the hyaluronic acid modified cosmetic peptides prepared in Examples 1-2, 5-7 and 10-13 of the present application have excellent anti-wrinkle firming efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL. Furthermore, the three structures (D2, D4, D6) of hyaluronic acid modified cosmetic peptides prepared in Example 4 of the present application, mixed in a certain mass ratio, have excellent anti-wrinkle firming efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the three structures (H2, H4, H6) of hyaluronic acid modified cosmetic peptides prepared in Example 8 of the present application, mixed in a certain mass ratio, have excellent anti-wrinkle firming efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the three structures (L2, L4, L6) of hyaluronic acid modified cosmetic peptides prepared in Example 9 of the present application, mixed in a certain mass ratio, have excellent anti-wrinkle firming efficacy at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL.

[0204] 3. Soothing performance test IL-6 content test (1) Cell inoculation: inoculate cells into a 24-well plate and incubate overnight in an incubator (37℃, 5% CO2); (2) Liquid preparation: prepare the working solution of the test substance according to the experimental design (as shown in Table 8); Table 8. IL-6 synthesis experimental design table

[0205] (3) Add test substance: according to the experimental grouping, when the cell plating rate in the 24-well plate reaches 40%~60%, add the test substance in groups, with 3 replicate holes in each group, and place the 24-well plate in an incubator (37℃, 5% CO2) for incubation for 24 h; (4) Detection: after 24 hours of culture, collect the supernatant and determine the IL-6 content by using an ELISA kit.

[0206] Table 9. Soothing performance test results

[0207] From the data analysis in Table 9, it can be seen that the hyaluronic acid modified cosmetic peptides prepared in Example 2 of the present application have soothing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the hyaluronic acid modified cosmetic peptides prepared in Example 5 of the present application have soothing effect at a concentration of 0.063 mg / mL; the hyaluronic acid modified cosmetic peptides prepared in Example 6 of the present application have soothing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the hyaluronic acid modified cosmetic peptides prepared in Example 7 of the present application have soothing effect at a concentration of 0.125 mg / mL; the three structures of hyaluronic acid modified cosmetic peptides prepared in Example 9 of the present application are mixed at a certain mass ratio, and have soothing effect at concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; the hyaluronic acid modified cosmetic peptides prepared in Example 10 of the present application have soothing effect at a concentration of 0.063 mg / mL; the hyaluronic acid modified cosmetic peptides prepared in Example 13 of the present application have soothing effect at a concentration of 0.125 mg / mL; the hyaluronic acid modified cosmetic peptides prepared in Example 11 of the present application have soothing effect at concentrations of 0.063 mg / mL and 0.125 mg / mL; the hyaluronic acid modified cosmetic peptides prepared in Example 12 of the present application have soothing effect at concentrations of 0.063 mg / mL and 0.25 mg / mL.

[0208] 4. Whitening performance test The melanin content test is as follows: Table 10 Melanin content experimental design table

[0209] Logarithmic growth phase cells were collected and inoculated into a 24-well plate, and after 24 h of culture in an incubator (37℃, 5% CO2), according to the cytotoxicity results, the tested substances were added according to the table, with untreated cells as a blank control, and 3 parallel groups were set for each group.

[0210] After drug addition, the culture was continued in an incubator (37℃, 5% CO2) for 24 h, the supernatant was discarded, 0.5 mL of 1M NaOH containing 10% DMSO was added, and incubation was carried out at 80℃ for 1 h. 1M NaOH containing 10% DMSO was used as a solvent control, and the absorbance value was read under an enzyme marker and the relative inhibition rate of cell melanin was calculated.

[0211]

[0212] Table 11 Whitening performance test results

[0213] From the data analysis in Table 11, it can be seen that the hyaluronic acid modified cosmetic peptide prepared in Example 7 of the present application has excellent whitening effect at the concentrations of 0.063 mg / mL, 0.125 mg / mL and 0.25 mg / mL; and the whitening effect at the concentrations of 0.125 mg / mL and 0.25 mg / mL is equivalent to or higher than that of the positive control group.

[0214] 5. Anti-glycation performance test A mixed solution containing bovine serum albumin and glucose was prepared using PBS, filtered through a 0.22 μm filter membrane, and used as 2x glycation reaction solution. Each group of reaction systems was prepared according to the table.

[0215] Table 12. AGEs clearance test reaction system table

[0216] After mixing evenly, incubate at 55°C for 4d. Use PBS instead of the sample as the negative control, use aminoguanidine hydrochloride (100 mg / mL) as the positive control, and use PBS instead of the glycosylation reaction solution as the control system. After the reaction is completed, cool the incubated solution to room temperature, centrifuge at 2000 r / min for 5 min, take the supernatant, filter it through a 0.22 μm filter membrane, and then take 200 μL of the reaction solution and add it to a 96-well plate. Use a fluorescence plate reader to detect under the conditions of an excitation wavelength of 320 nm and an emission wavelength of 460 nm, and calculate the AGEs inhibition rate according to the following formula.

[0217]

[0218] In the formula: A - the fluorescence intensity of the glycation system to which the test substance is added; B - the fluorescence intensity of the PBS solution to which the test substance is added; C - the fluorescence intensity of the glycation system without the addition of the test substance; D - the fluorescence intensity of the PBS solution without the addition of the test substance.

[0219] Table 13. Anti-glycation performance test results

[0220] From the data analysis in Table 13, it can be seen that the hyaluronic acid modified cosmetic peptide prepared in Example 11 of the present application has a clearance effect on advanced glycosylation end products at the concentrations of 0.0625 mg / mL and 0.25 mg / mL.

[0221] 6. Antioxidant performance test The ROS content test is as follows: (1) Cell inoculation: inoculate cells into a 24-well plate and incubate in an incubator (37°C, 5% CO2) overnight; (2) Liquid preparation: according to the experimental design (Table 13), the working solution of the test substance was prepared.

[0222] Table 14 ROS experimental design table

[0223] (3) Add test substance: after 24h of incubation in an incubator (37℃, 5% CO2), add the test substance according to the table, and continue to incubate for 24h; (4) Modeling: PBS was washed twice, and untreated cells were used as a blank control. The rest of the groups received UVB stimulation according to the conditions in the table, and the VC+VE group was used as a positive control; (5) ROS content detection: the DCFH-DA probe mother liquor was diluted with serum-free culture medium. 500μL of diluted DCFH-DA probe was added to each well, and incubated in a 37℃ cell incubator; after 30 min, the cells were washed 3 times with serum-free DMEM culture medium to thoroughly remove the probe that did not enter the cells; using a fluorescence microscope, observe and take pictures under the condition of excitation wavelength 488 nm.

[0224] (6) Result analysis: the ROS fluorescence intensity was quantitatively analyzed using Image Pro Plus software.

[0225] Table 15 Antioxidant performance test results

[0226] From the data analysis in Table 15, it can be seen that the hyaluronic acid modified cosmetic peptide prepared in Example 11 has better antioxidant capacity at a concentration of 0.125mg / mL and 0.25mg / mL, and its antioxidant capacity level at a concentration of 0.25mg / mL is comparable to that of the positive control.

[0227] The conventional techniques in the above examples are prior art known to those skilled in the art, and therefore will not be described in detail here.

[0228] The above is only a specific embodiment of the present application, but the protection scope of the present application is not limited thereto. Any person skilled in the art can easily think of changes or replacements within the technical scope disclosed by the present application, which should be covered within the protection scope of the present application. Therefore, the protection scope of the present application should be subject to the protection scope of the claims.

Claims

1. A hyaluronic acid-modified beauty peptide, wherein the structural formula of the hyaluronic acid-modified beauty peptide is shown in formula (I): MC (I); in, M represents sodium hyaluronate, and its structure is shown in formula (II): (II), where y is a natural number ≥ 1; C represents a beauty peptide, which includes a polypeptide or a derivative thereof having beauty and / or skin care effects, and the polypeptide includes a tetrapeptide; Among them, the M terminal glucuronic acid links the amino group in the C structure.

2. The hyaluronic acid-modified cosmetic peptide according to claim 1, characterized in that: The compound represented by formula (I) includes the structure represented by formula (III): (III); in, Said n is a natural number; The R is the remaining part of the cosmetic peptide structure after removing the reactive amino group; The cosmetic peptides include polypeptides or derivatives thereof having cosmetic and / or skin care effects, and the polypeptides include tetrapeptides.

3. The hyaluronic acid-modified cosmetic peptide according to claim 1 or 2, characterized in that: The tetrapeptide or its derivative includes one of tetrapeptide-5, tetrapeptide-7, tetrapeptide-9, tetrapeptide-11, tetrapeptide-30 and tetrapeptide-15, or includes a tetrapeptide with an amino acid sequence of H-Asp-Val-Lys-Tyr-OH.

4. The method for preparing the hyaluronic acid-modified cosmetic peptide according to claim 1 or 2, comprising: Sodium hyaluronate is used to react with beauty peptides to produce hyaluronic acid-modified beauty peptides.

5. The method for preparing the hyaluronic acid-modified cosmetic peptide according to claim 4, characterized in that: The molar ratio of the cosmetic peptide to sodium hyaluronate is 1:2-8.

6. Use of the hyaluronic acid-modified beauty peptide obtained by the preparation method according to claim 4 in the preparation of cosmetics and / or skin care products.

7. Use of the hyaluronic acid-modified beauty peptide obtained by the preparation method of claim 4 in enhancing the moisturizing, firming, anti-wrinkle or anti-aging properties of cosmetics and / or skin care products.

8. Use of the hyaluronic acid-modified beauty peptide obtained by the preparation method according to claim 4 in enhancing the soothing, antioxidant or whitening properties of cosmetics and / or skin care products.

9. A cosmetic comprising the hyaluronic acid-modified beauty peptide according to claim 1.

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