Hyaluronic acid derivatives, methods for preparing them, and their use

Abstract

The present application discloses hyaluronic acid derivatives, specifically hyaluronic acid esters synthesized from hyaluronic acid or its salt and salicylic acid esters. The hyaluronic acid esters disclosed in the present application are characterized by good water solubility, low irritation, and mild properties, optimizing the stratum corneum. Furthermore, the water-soluble esters inhibit the penetration of salicylic acid into the stratum corneum and extend the duration of salicylic acid's action in the stratum corneum. The unique chemical structure of the salicylic acid hyaluronic acid esters of the present application immobilizes acidic functional groups, reducing the irritation of salicylic acid. The increased hyaluronic acid polysaccharide content effectively delays the penetration of salicylic acid into the deeper layers of the skin, prolonging its action time in the epidermis, targeting the stratum corneum and contributing to the long-term regulation of keratinocytes. The improved solubility of salicylic acid itself in the aqueous phase facilitates its use in formulations.

Description

Technical Field

[0001] The present application relates to the field of biotechnology, in particular to a hyaluronic acid derivative, a preparation method therefor and use thereof.

Background Art

[0002] Hyaluronic acid (HA) is a mucopolysaccharide formed by alternately linked disaccharide units of glucuronic acid and N-acetylglucosamine. Due to its unique molecular structure and physical and chemical properties, it has been effectively used in cosmetics, medical aesthetics, ophthalmic surgery, and arthritis treatment, among other fields.

[0003] Hyaluronic acid is widely distributed in nature. In the human body, more than 50% of hyaluronic acid is present in the skin, lungs and intestines. Furthermore, it is also present in interstitial tissues such as synovial fluid, cartilage, umbilical cord, and vascular wall. In early research, the main source of hyaluronic acid was the umbilical cord. Currently, hyaluronic acid products can be extracted from animal tissues such as chicken combs, vitreous body, brain cartilage and synovial fluid, and can also be obtained through fermentation of bacteria such as Streptococcus and Pseudomonas aeruginosa. The fermentation method for producing hyaluronic acid is gradually replacing the tissue extraction method because of its low cost, abundant raw materials, ease of large-scale production, and the high molecular weight of the obtained hyaluronic acid. In recent years, various hyaluronic acid derivatives have been developed by modifying hyaluronic acid in various ways, and in addition to retaining the functions of hyaluronic acid itself, the hyaluronic acid derivatives also have improved stability, biocompatibility, water solubility, and other properties.

[0004] Salicylic acid (simply referred to as SA), also known as o-hydroxybenzoic acid, is a white crystalline powder that can exfoliate the stratum corneum, remove excessively thickened layers, and promote metabolism. However, because SA is a small-molecule acid, it is highly permeable, dissolving the stratum corneum and destroying the sebum film while easily penetrating the dermis and subcutaneous tissue. As a result, it does not remain in the stratum corneum for a sufficient amount of time, failing to exert its characteristic exfoliating effect and instead irritating the microvessels and nerves distributed deep within the skin. For these reasons, its application is limited. [Overview of the project]

[0005] In view of the above issues, this application provides hyaluronic acid derivatives and methods for preparing them. Specifically, this application adopts the following technical proposal.

[0006] 1. A salicylic acid hyaluronic acid ester whose structural formula is represented by formula (I),

[0007] [ka]

[0008] In the formula, R1 is H or

[0009] [ka]

[0010] Hyaluronic acid ester, where R2 is H or a metal ion, x1≧0, x2≧0, y≧1. 2. The hyaluronic acid ester according to item 1, wherein the degree of substitution of salicylic acid in hyaluronic acid or a salt thereof is 15% to 60%, preferably 25% to 50%. 3. The hyaluronic acid ester according to any one of items 1 to 3, wherein the molecular weight of the hyaluronic acid ester is 500 kDa or less, preferably 5 kDa to 300 kDa, and more preferably 5 kDa to 100 kDa. 4. A method for preparing a salicylic acid hyaluronic acid ester, wherein the hyaluronic acid ester is an ester synthesized from hyaluronic acid or a salt thereof and a salicylic acid ester. 5. The method for preparing the hyaluronic acid ester described in item 4, wherein the hyaluronic acid ester is obtained by reacting a quaternary ammonium salt of hyaluronic acid with a salicylic acid ester in the presence of an inorganic base. 6. The method for preparing a hyaluronic acid ester according to item 4 or 5, wherein the molar ratio of the quaternary ammonium hyaluronic acid salt to the salicylic acid ester is 1:(1~10). 7. The preparation method according to any one of items 4 to 6, wherein the molar ratio of the inorganic base to the quaternary ammonium hyaluronic acid salt is 1:(2 to 10). 8. The preparation method according to any one of items 4 to 7, wherein the inorganic base comprises one or more of sodium hydroxide, potassium hydroxide, ammonium hydroxide, barium hydroxide, sodium carbonate, sodium bicarbonate, potassium carbonate, or potassium bicarbonate. 9. The preparation method according to any one of items 4 to 8, wherein the reaction temperature is 50 to 100°C, preferably 60 to 80°C. 10. The inorganic base has been treated, and the treatment is A preparation method according to any one of claims 4 to 9, comprising mixing and dissolving an inorganic base and a tetrabutylammonium salt, and then freeze-drying the mixture. 11. The preparation method according to any one of items 4 to 10, wherein, in the treatment of the inorganic base, the molar ratio of the inorganic base to the tetrabutylammonium salt is 1:(0.1~1). 12. Use in skincare products of hyaluronic acid esters described in any one of items 1 to 3, or hyaluronic acid esters prepared by the preparation method described in any one of items 4 to 11. 13. Use of hyaluronic acid esters described in any one of items 1 to 6, or hyaluronic acid esters prepared by the preparation method described in any one of items 7 to 15, in the preparation of exfoliating and / or hypoallergenic topical agents. 14. A composition comprising a hyaluronic acid ester described in any one of items 1 to 3, or a hyaluronic acid ester prepared by the preparation method described in any one of items 4 to 11. [Effects of the Invention]

[0011] 1. The salicylic acid hyaluronic acid ester of this application has good water solubility, low irritancy, and mild properties, and is characterized by its ability to optimize the stratum corneum. Furthermore, the water-soluble ester inhibits the penetration of salicylic acid into the stratum corneum and extends the duration of action of salicylic acid in the stratum corneum. 2. The unique chemical structure of the salicylic acid hyaluronic acid ester of this application can fix the acidic functional group and reduce the irritation of salicylic acid. The increase in hyaluronic acid polysaccharides effectively delays the penetration of salicylic acid into the deeper layers of the skin, extending the duration of action in the epidermis, targeting the stratum corneum, aiding in the long-term regulation of keratinocytes, and improving the aqueous solubility of salicylic acid itself, making it easier to use in formulations. 3. In the salicylic acid hyaluronic acid ester preparation method of this application, the catalytic effect of the inorganic base in the reaction solvent is improved by mixed freeze-drying treatment with an inorganic base, the reaction temperature is lowered, and the reaction efficiency is improved. Compared to untreated inorganic bases, the catalytic effect is much higher and higher than that of organic bases. Furthermore, the preparation process of this application is easy to operate, suitable for industrial production, and has broad potential in the cosmetic raw materials market. [Brief explanation of the drawing]

[0012] The attached drawings are provided for the purpose of better understanding this application and do not constitute any unwarranted limitation thereto. [Figure 1] This is the 1H NMR spectrum of the salicylic acid hyaluronic acid ester prepared in Example 18. [Figure 2] This is the full-wavelength ultraviolet scanning spectrum of the salicylic acid hyaluronic acid ester prepared in Example 18. Details of the invention

[0013] Exemplary embodiments of the present application will be described below. These embodiments contain various details of the embodiments of the present application for facilitating understanding, and they should be considered as merely illustrative. Therefore, those skilled in the art will understand that various changes and modifications to the embodiments described herein can be made without departing from the scope and spirit of the present application. In addition, in the following description, descriptions of well-known functions and structures are omitted for clarity and conciseness.

[0014] The present application provides a hyaluronic acid derivative, specifically provides a hyaluronic acid ester which is an ester synthesized from hyaluronic acid or a salt thereof and a salicylic acid ester.

[0015] The hyaluronic acid obtained in the present application refers to a biopolymer material composed of linearly linked repeating units of N-acetyl-D-glucosamine and D-glucuronic acid. Said hyaluronic acid or a salt thereof includes hyaluronic acid per se, a salt thereof, or a combination thereof. Examples of hyaluronic acid salts include, but are not limited to, inorganic salts such as sodium hyaluronate, potassium hyaluronate, calcium hyaluronate, magnesium hyaluronate, zinc hyaluronate, gold hyaluronate, and cobalt hyaluronate, and organic salts such as quaternary ammonium hyaluronate. In the present application, hyaluronic acid per se or a salt thereof may be used alone, or two or more kinds of hyaluronic acid or salts thereof may be used in combination.

[0016] The hyaluronic acid or a salt thereof described in the present application is not limited. In a preferred embodiment, the hyaluronic acid salt is a water-soluble salt of hyaluronic acid, more preferably a quaternary ammonium salt of hyaluronic acid.

[0017] The salicylic acid ester described in the present application is preferably an alkyl ester or an alkenyl ester, wherein the alkyl or alkenyl group has 1 to 6 carbon atoms, including but not limited to pentyl salicylate, ethyl salicylate, cis-3-hexenyl salicylate, hexyl salicylate, isobutyl salicylate, and methyl salicylate.

[0018] In a preferred embodiment, the alkyl salicylate is methyl salicylate, that is, methyl o-hydroxybenzoate, also known as methyl salicylate, which is an organic compound with the chemical formula C8H8O3, is a colorless to pale yellow transparent liquid, and has a strong wintergreen oil odor.

[0019] The present application also provides hyaluronic acid salicylate having the structural formula shown in formula (I),

[0020]

Chemical Formula

[0021] wherein R1 is H or

[0022]

Chemical Formula

[0023] , R2 is H or a metal ion, x1≧0, x2≧0, y≧1, and x1, x2 and y are each independent repeating units.

[0024] The term "grafting" refers to a reaction in which appropriate branched chains or functional side groups are chemically bonded to polymer chains, and the resulting product is called a graft copolymer. Graft rate generally refers to graft efficiency, and is given by: Graft rate = [Amount of graft monomer or copolymer branched chain grafted onto the graft copolymer / (Total amount of grafted monomer or grafted polymer initially added)] × 100%. In this application, the concept of degree of substitution is used to express graft efficiency. The term "degree of substitution" refers to the amount of substance that substitutes for the active hydroxyl group of each D-glucose unit of cellulose. In this application, the degree of substitution specifically refers to: [Amount of graft monomer or copolymer branched chain grafted onto the graft copolymer / (Total amount of grafted monomer or grafted polymer initially added)] × 100%.

[0025] The degree of substitution of salicylic acid hyaluronic acid ester in this application is 15% to 60%, and may be, for example, 16%, 17%, 18%, 19%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 51%, 52%, 53%, 54%, 55%, 56%, 57%, 58%, or 59%. Preferably, the degree of substitution is 25% to 50%. The molecular weight of the salicylic acid hyaluronic acid ester of this application is 500 kDa or less, and may be, for example, 1 kDa, 2 kDa, 3 kDa, 4 kDa, 5 kDa, 6 kDa, 7 kDa, 8 kDa, 9 kDa, 10 kDa, 20 kDa, 30 kDa, 40 kDa, 50 kDa, 60 kDa, 70 kDa, 80 kDa, 90 kDa, 100 kDa, 200 kDa, 300 kDa, 400 kDa, or 500 kDa. Preferably, it is 5 kDa to 300 kDa, and more preferably 5 kDa to 100 kDa.

[0026] This application further provides a method for preparing a hyaluronic acid ester described herein, the method comprising using an inorganic base as a catalyst to perform a graft reaction with a salicylic acid ester on a quaternary ammonium hyaluronic acid salt, that is, preparing the hyaluronic acid ester by covalently grafting methyl salicylate onto a quaternary ammonium hyaluronic acid salt.

[0027] The aforementioned quaternary ammonium hyaluronic acid salt refers to a hyaluronic acid graft polymer formed by linking a positively charged quaternary ammonium group to the active group of hyaluronic acid. By binding with a quaternary ammonium base, hyaluronic acid enhances its own excellent moisturizing properties, improves its adhesion to the surface of skin and hair, and enhances the moisturizing, lubricating, nourishing, and restorative effects of hyaluronic acid. Patent documents such as CN101316864 (Shiseido, Japan), CN101715457A (Kewpie, Japan), CN107739417A (Yangzhou Zhongfu Biotechnology Co., Ltd.), and CN107556402A (Chongqing University of Technology and Commerce) disclose methods for preparing quaternary ammonium hyaluronic acid salts. The quaternary ammonium hyaluronic acid salt used in the preparation method of this application may be a quaternary ammonium hyaluronic acid salt prepared by any of the prior art methods, including the above-mentioned patent document, or any of the quaternary ammonium hyaluronic acid salts that are commonly available for purchase. In a preferred embodiment, this application uses a quaternary ammonium hyaluronic acid salt prepared by the method disclosed in patent document CN107459590B.

[0028] The type of inorganic base is not limited, and any inorganic base in the art that can accelerate the reaction in the preparation method of this application may be used. In this application, the inorganic base is a catalyst in the reaction process, but the term catalyst is not limited to inorganic bases. Those skilled in the art will understand that catalysis refers to a process that changes the rate of a chemical reaction. Any substance that can play such a role in change can be called a catalyst and can be used in the reaction process of this application. In some preferred embodiments, the inorganic base may include one or more of sodium hydroxide, potassium hydroxide, ammonium hydroxide, barium hydroxide, sodium carbonate, sodium bicarbonate, potassium carbonate, or potassium bicarbonate. For example, the inorganic base may be sodium hydroxide. The inorganic base may be potassium hydroxide. The inorganic base may be ammonium hydroxide. The inorganic base may be barium hydroxide. The inorganic base may be sodium carbonate. The inorganic base may be potassium carbonate. The inorganic base may be potassium bicarbonate.

[0029] In the preparation method of this application, the specific method involves first dissolving the quaternary ammonium hyaluronic acid salt in an organic solvent, then adding methyl salicylate to the organic solvent containing the dissolved quaternary ammonium hyaluronic acid salt, then adding an inorganic base to form a reaction solution, stirring while heating, allowing the reaction to proceed for a certain period of time, then adding a salt solution and continuing to stir, followed by alcohol precipitation, alcohol washing, dehydration, suction filtration, and drying to obtain the hyaluronic acid ester.

[0030] In the above preparation method, the organic solvent used to dissolve the quaternary ammonium hyaluronic acid salt is not limited. In preferred embodiments, the organic solvent is DMSO (dimethyl sulfoxide) or DMF (dimethylformamide).

[0031] The heating can be any method in the art, such as water bath heating, oil bath heating, air bath heating, sand bath heating, or electric heating. The heating temperature, i.e., the reaction temperature between the quaternary ammonium hyaluronic acid salt and methyl salicylate in the preparation method of this application, is not limited as long as the two can undergo a covalent bond reaction. In a preferred embodiment, the reaction temperature is 50 to 100°C, and may be, for example, 55°C, 60°C, 61°C, 62°C, 63°C, 64°C, 65°C, 66°C, 67°C, 68°C, 69°C, 70°C, 71°C, 72°C, 73°C, 74°C, 75°C, 76°C, 77°C, 78°C, 79°C, 80°C, 85°C, 90°C, or 95°C. In a preferred embodiment, the reaction temperature is 60 to 80°C.

[0032] The stirring can be carried out in any manner in the art, as long as the reactants are uniformly mixed and uniformly heated.

[0033] The aforementioned "reacting for a certain period of time" refers to the time required for the quaternary ammonium hyaluronic acid salt and methyl salicylate to react completely, and is not limited as long as the two react completely.

[0034] The salt solution is not limited, but can be, for example, a sodium chloride solution, a potassium chloride solution, or a calcium chloride solution. In a preferred embodiment, during the reaction, the salt solution is an NaCl solution, the concentration of the salt solution is 2-5%, and its volume is more than half the volume of the reaction solution.

[0035] In the above preparation method, a salt solution is added, and stirring is continued for a certain period of time, for example 10 to 50 minutes, to promote precipitation and complete ion displacement. After that, an alcohol solution can be used for precipitation and washing. The concentration of the alcohol solution is not limited, but is preferably 60 to 90%, and the number of washes is not limited, but is preferably 1 to 10 times. In the preferred preparation method, after washing with alcohol, the product is dehydrated once with ethanol, then filtered by suction, and vacuum dried to obtain the final product.

[0036] In the above preparation method, the molar ratio of the quaternary ammonium hyaluronic acid salt to methyl salicylate is not limited. In preferred embodiments, the molar ratio of the quaternary ammonium hyaluronic acid salt to methyl salicylate is 1:(1~10), and may be, for example, 1:1.5, 1:2, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5, 1:5, 1:5.5, 1:6, 1:6.5, 1:7, 1:7.5, 1:8, 1:8.5, 1:9, or 1:9.5.

[0037] In the above preparation method, the molar ratio of the inorganic base to the quaternary ammonium hyaluronic acid salt is not limited. In preferred embodiments, the molar ratio of the inorganic base to the quaternary ammonium hyaluronic acid salt is 1:(2~10), and may be, for example, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5, 1:5, 1:5.5, 1:6, 1:6.5, 1:7, 1:7.5, 1:8, 1:8.5, 1:9, or 1:9.5.

[0038] In a preferred embodiment, the inorganic base is a treated inorganic base.

[0039] One method of the aforementioned treatment is to mix and dissolve an inorganic base and a tetrabutylammonium salt, and then freeze-dry them.

[0040] The mixing can be carried out by any conventional method in the art, and in the dissolution, the inorganic base and the tetrabutylammonium salt are mixed and dissolved in water.

[0041] The type of tetrabutylammonium salt is not limited, but may include, for example, tetrabutylammonium fluoride, tetrabutylammonium chloride, tetrabutylammonium bromide, or tetrabutylammonium iodide.

[0042] The aforementioned freeze-drying is a drying method that involves freezing a water-containing substance to solidify it, and then using the property of water to sublimate under low temperature and low pressure conditions to dehydrate and dry it at low temperatures. The freeze-drying process of this application can be carried out by any method in the art, for example, by pre-freezing the substance in a refrigerator, freeze-dryer, or liquid nitrogen, and then freeze-drying it.

[0043] In the above processing steps, the molar ratio of the inorganic base to the tetrabutylammonium salt is not limited. In preferred embodiments, the molar ratio of the inorganic base to the tetrabutylammonium salt is 1:0.1 to 1:1, and may be, for example, 1:0.2, 1:0.25, 1:0.3, 1:0.35, 1:0.4, 1:0.45, 1:0.5, 1:0.55, 1:0.6, 1:0.65, 1:0.7, 1:0.75, 1:0.8, 1:0.85, 1:0.9, or 1:0.95.

[0044] This application further provides for the use of any of the hyaluronic acid esters described above, or hyaluronic acid esters prepared using the preparation methods provided herein, in skin protection products or skin care products.

[0045] The aforementioned skin protection or skincare products include, but are not limited to, water, lotions, ointments, creams, essences, sunscreens, oils, makeup removers, solutions, gels, shower gels, shampoos, facial cleansers, shampoo creams, shaving creams, shaving foams, shaving waters, moisturizing lotions, moisturizing sprays, moisturizing masks, toners, lotions, softeners, exfoliating gels, acne gels, facial creams, body creams, hand creams, foot creams, repair essences, moisturizing lotions, moisturizing creams, repair lotions, moisturizing lipsticks, lip glosses, lip glazes, and hair removal creams.

[0046] In a preferred embodiment, the skincare product is a skincare product having an exfoliating function.

[0047] The hyaluronic acid ester provided in this application is a novel hyaluronic acid derivative that is completely soluble in water and has good water solubility. It can be prepared as a cosmetic ingredient and can also be combined with various cosmetic ingredients to prepare cosmetics. Furthermore, since the molecular weight of the hyaluronic acid ester can be selected according to various needs, it offers further possibilities to the cosmetic ingredient market.

[0048] The hyaluronic acid ester of this application possesses the properties of both hyaluronic acid and salicylic acid, and its unique chemical structure allows for the fixation of acidic functional groups, thereby reducing the irritation of salicylic acid. The increased hyaluronic acid polysaccharide effectively delays the penetration of salicylic acid into the deeper layers of the skin, extending its duration of action in the epidermis. It targets the stratum corneum, exhibits low irritation, a mild nature, and optimizes the stratum corneum. Human skin studies have demonstrated that the hyaluronic acid ester of this application exhibits lower irritation and a better exfoliation rate compared to compositions of salicylic acid and hyaluronic acid ester.

[0049] This application provides a method for preparing the salicylic acid hyaluronic acid ester and optimizes the type and amount of raw materials in the preparation method. By mixing and freeze-drying an inorganic base, the catalytic effect of the inorganic base in the reaction solvent is improved, the reaction temperature is lowered, and the reaction efficiency is improved. By comparison, the catalytic effect is much higher than that of an untreated inorganic base and higher than that of an organic base. At the same time, by optimizing the preparation conditions, it is also possible to prepare hyaluronic acid esters with a higher degree of substitution. [Examples]

[0050] Example 1 1) Treatment of inorganic base: 5 mmol of potassium bicarbonate and 2.5 mmol of tetrabutylammonium chloride were mixed and dissolved in 10 ml of purified water. After complete dissolution, the mixture was pre-frozen in liquid nitrogen and freeze-dried to obtain the treated inorganic base. 2) Preparation of quaternary ammonium hyaluronic acid salt: Hyaluronic acid with a molecular weight of 100 kDa was selected, and quaternary ammonium hyaluronic acid salt was prepared by the method disclosed in Chinese Patent CN107459590B. 3) Preparation of water-soluble hyaluronic acid ester: 10 mmol of quaternary ammonium hyaluronic acid salt prepared in step 2) was dissolved in DMSO (dimethyl sulfoxide), and after complete dissolution, 50 mmol of methyl salicylate was added. Add all of the inorganic base prepared in Step 1), stir at 80°C, and allow to react for 4 hours. Half of the total volume of 2% NaCl solution was added, and the mixture was stirred for 20 minutes. Then, ethanol was added to allow precipitation, and the mixture was allowed to stand. The supernatant was taken and washed with 80% ethanol. This process was repeated five times, and then the mixture was dehydrated once with ethanol, filtered by suction, and vacuum dried to obtain a water-soluble hyaluronic acid ester powder.

[0051] The degree of substitution in the product was detected by quantitative NMR analysis, which compares the intensities of different absorption peaks. In quantitative analysis, the integral of the absorption peak intensity of the H atom is proportional to its molar concentration, and this is converted to the degree of substitution. The degree of substitution was 15%.

[0052] Examples 2-27 Except for using the types and amounts of raw materials shown in Table 2, water-soluble hyaluronic acid powder was prepared according to the method of Example 1, in the same manner as in Example 1, unless otherwise specified.

[0053] The structural formula of the water-soluble hyaluronic acid ester prepared in Example 18 is as follows.

[0054] [ka]

[0055] 1 The 1H NMR spectrum is shown in Figure 1.

[0056] [Table 1]

[0057] 1 The H spectrum shows 16 hydrogen groups, and the integral ratios from low-field hydrogen to high-field hydrogen are 1:1:1:1:2:2:4:2:2:2:2:2:2:2:2:2:2:2:6, which is consistent with the structure of this product. Of these, the hydrogen at δ8.015 is a double-lined peak group, has 1 proton, and is attributed to the hydrogen at position 7''. The hydrogen at δ7.600 is a triple-lined peak group, has 1 proton, and is attributed to the hydrogen at position 5''. The hydrogen at δ7.053 is a multi-lined peak group, has 1 proton, and is attributed to the hydrogen at position 6''. The hydrogen at δ7.032 is a multi-lined peak group, has 1 proton, and is attributed to the hydrogen at position 4''. Chemical shifts in the range of δ7 to δ8.5 are generally caused by the benzene ring structure. The results above indicate that this product contains a benzene ring structure and has a 1:1 relationship with hyaluronic acid molecules.

[0058] The ultraviolet absorption spectrum of the benzene ring: 203 nm (ε=8700) is called the E2 band. Between 230 and 270 nm, there is a series of relatively weak absorption bands called the fine structure, centered at 254 nm (ε=204) and called the B band. When groups such as -OCH3, -CHO, -COOH, and -NO2 are introduced, both the E2 and B bands generally undergo redshift. The full-wavelength ultraviolet scanning spectrum of this product is shown in Figure 2. The peak in the range of 230 nm to 240 nm is the E2 band of the benzene ring, and the peak in the range of 300 nm to 310 nm is the B band of the benzene ring. The above results indicate that this product contains carbonyl and substituted benzene ring structures.

[0059] [Table 2-1]

[0060] [Table 2-2]

[0061] [Table 2-3]

[0062] The degree of substitution of the prepared hyaluronic acid ester was detected by nuclear magnetic resonance spectroscopy and is shown in Table 3.

[0063] [Table 3-1]

[0064] [Table 3-2]

[0065] Test example Test Example 1: Cytotoxicity Test 1.1 Sample: Salicylic acid hyaluronic acid ester obtained in Example 18. 1.2 Experimental Methods and Procedures: 1.2.1 Plating HaCaT cells in the logarithmic growth phase were collected and placed in a 96-well plate in a 1 × 10⁶ layer. 5 Cells were inoculated at a density of 100 μL per well. The culture system consisted of DMEM basal medium supplemented with 10% fetal bovine serum. The inoculated cells were placed in a carbon dioxide incubator and cultured at 37°C and 5% CO2 for 24 hours. 1.2.2 Preparation of Sample Solution The samples were prepared as 2.0% solutions using serum-free medium and filtered and sterilized through a 0.22 μm filter membrane. The final concentrations were 0.05%, 0.1%, 0.2%, and 0.5%, and were prepared each time they were used. 1.2.3 Administration After 24 hours of normal incubation, the old culture medium was discarded, 100 μL of sample was added to the experimental group, and an equal volume of serum-free medium was added to the normal control group, creating six parallel wells at each level. 1.3 Detection After 24 hours of incubation, the relative cell growth rate was detected using CCK-8. The culture medium was discarded, 100 μL of CCK-8 diluted 10-fold in serum-free medium was added to each well, and the cells were placed in a cell culture incubator and incubated for 2 hours. Absorbance was measured at a wavelength of 450 nm using a plate reader. Relative growth rate (RGR) is the ratio of the absorbance of the experimental group to the absorbance of the normal control group. According to the requirements of GB / T16886.5-2017, if the RGR is less than 70%, the sample is considered cytotoxic. 1.4 Results and Analysis: A relative proliferation rate of 70% or higher is considered non-cytotoxic. Salicylic acid hyaluronic acid ester is non-cytotoxic at concentrations of 0.5% or less.

[0066] [Table 4]

[0067] Test Example 2: Irritation Test 1.1 Sample: Sample 1: The salicylic acid hyaluronic acid ester obtained in Example 18 was dissolved in water at a mass concentration of 5%. Sample 2: A composition of hyaluronic acid and salicylic acid was dissolved in water. Here, the molecular weight of hyaluronic acid was 100 kDa, the mass concentration of hyaluronic acid was 4.35%, and the mass concentration of salicylic acid was 0.7%. 1.2 Experimental Methods and Procedures: We recruited healthy volunteers to participate in this project voluntarily. Exclusion criteria: (1) Persons with severe systemic disease, immunodeficiency or autoimmune disease, skin disease or medical history (such as severe freckles, atopic dermatitis, psoriasis, eczema, severe acne, etc.). (2) Anyone who has received dermatological treatment or cosmetic surgery, or who has used hormones or immunosuppressants within the past month. (3) Individuals with an allergic constitution, allergic dermatitis, or active allergic disease. (4) Pregnant, breastfeeding, or menopausal women. (5) Any other person who has a skin condition that may affect the trial and is deemed unsuitable to participate in this project based on clinical evaluation. Number of valid subjects: 30. The subjects washed their faces with facial cleanser, dried them with a tissue, and waited 15 minutes. 20 μL of each of the two samples was dropped onto a 7 mm diameter circular filter paper and randomly attached to the nasolabial folds on both sides of the subjects' faces. At 0, 1, 5, and 10 minutes, the subjects evaluated the level of discomfort (itchiness, tingling, and burning) at the test sites. The sum of the sensory scores at each time point was the total sensory score. Scoring Criteria: 0 points: No tingling, itching, or burning sensation. 1 point: Mild tingling, itching, or burning sensation (slightly felt). 2 points: Moderate tingling, itching, or burning sensation (strong but tolerable). 3 points: Obvious tingling, itching, or burning sensation (relatively strong, unbearable). 1.3 Statistical methods: A paired t-test was used for statistical analysis. Test level α = 0.05 If the test mean is in the beneficial direction and the test result is p<0.05, it means the test is effective. 1.4 Results and Analysis: The overall skin irritation score of samples containing 5% salicylic acid hyaluronic acid was low within 10 minutes of application, demonstrating that salicylic acid hyaluronic acid is less irritating.

[0068] [Table 5]

[0069] Test Example 3: Exfoliation ability against keratinocytes - Stratum corneum desquamation assay 1.1 Sample: Sample 1: The salicylic acid hyaluronic acid ester obtained in Example 18 was dissolved in water at a mass concentration of 5%. Sample 2: A composition of hyaluronic acid and salicylic acid was dissolved in water. Here, the molecular weight of hyaluronic acid was 100 kDa, the mass concentration of hyaluronic acid was 4.35%, and the mass concentration of salicylic acid was 0.7%. 1.2 Experimental Methods and Procedures: Twenty-five healthy subjects were recruited. After wiping the flexor side of their forearms with a dry tissue, a 3cm x 3cm test area was marked at the same location on both the left and right forearm flexor sides. Sample application: 20 μL of each of the two samples was applied to the test areas on both arms, massaged for 30 seconds, waited for 1 minute, then rinsed with warm water for 10 seconds, and the area was patted dry with a tissue. 1.3 Method for measuring desquamation: Fifteen minutes after absorbing moisture, the release film was lightly pressed onto the test area to collect exfoliated material, and the percentage of keratin exfoliation was measured using a Visioscope PC 35. 1.4 Results and Analysis: A 5% concentration of salicylic acid hyaluronic acid ester has strong exfoliating properties.

[0070] [Table 6]

[0071] While embodiments of this application have been described above, this application is not limited to the specific embodiments or fields of application described herein. The specific embodiments described above are merely illustrative and guideline, and not limiting. A person skilled in the art can create many forms in accordance with the guidelines herein without departing from the scope of protection of the claims of this application, all of which are protected by this application.

Claims

1. The salicylic acid hyaluronic acid ester whose structural formula is shown in formula (I), 【Chemistry 1】 In the formula, R １ is H or 【Chemistry 2】 And R ２ is H or a metal ion, and x １ ≥ 0, x ２ Hyaluronic acid esters such that ≥ 0 and y ≥ 1.

2. The hyaluronic acid ester according to claim 1, characterized in that the degree of substitution of salicylic acid in the hyaluronic acid or its salt is 10% to 60%.

3. The hyaluronic acid ester according to claim 1 or 2, characterized in that the molecular weight of the hyaluronic acid ester is 500 kDa or less.

4. A method for preparing salicylic acid hyaluronic acid ester, characterized in that the method for preparing the hyaluronic acid ester includes a step of reacting hyaluronic acid or a salt thereof with a salicylic acid ester.

5. The method for preparing a hyaluronic acid ester according to claim 4, characterized in that the hyaluronic acid ester is obtained by reacting a quaternary ammonium salt of hyaluronic acid with a salicylic acid ester in the presence of an inorganic base.

6. The preparation method according to claim 5, characterized in that the molar ratio of the quaternary ammonium hyaluronic acid salt to the salicylic acid ester is 1:(1 to 10).

7. The preparation method according to claim 6, characterized in that the molar ratio of the inorganic base to the quaternary ammonium hyaluronic acid salt is 1:(2 to 10).

8. The preparation method according to claim 5, characterized in that the inorganic base comprises one or more of sodium hydroxide, potassium hydroxide, ammonium hydroxide, barium hydroxide, sodium carbonate, sodium bicarbonate, potassium carbonate, or potassium bicarbonate.

9. The preparation method according to claim 4, characterized in that the reaction temperature is 50 to 100°C.

10. The inorganic base is treated, and the treatment is, The preparation method according to claim 5, characterized by comprising mixing and dissolving an inorganic base and a tetrabutylammonium salt, and then freeze-drying the mixture.

11. The preparation method according to claim 10, characterized in that, in the treatment of the inorganic base, the molar ratio of the inorganic base to the tetrabutylammonium salt is 1:(0.1 to 1).

12. Use of a hyaluronic acid ester according to claim 1 or 2, or a hyaluronic acid ester prepared by the preparation method according to claim 4 or 5, in a skincare product.

13. Use of a hyaluronic acid ester according to claim 1 or 2, or a hyaluronic acid ester prepared by the preparation method according to claim 7 or 5, in the preparation of an exfoliating and / or low-irritation topical agent.

14. A composition comprising the hyaluronic acid ester according to claim 1 or 2.

15. The hyaluronic acid ester according to claim 1, characterized in that the degree of substitution of salicylic acid in the hyaluronic acid or its salt is 10% to 40%.

16. The hyaluronic acid ester according to claim 1 or 2, characterized in that the molecular weight of the hyaluronic acid ester is 5 kDa to 100 kDa.

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