Small molecule compounds with sulfonic or sulfinic lactone structure, compositions and uses thereof

Abstract

This application discloses a small molecule compound, composition, and application having a sulfonic acid or sulfinic acid lactone structure. This application provides a class of small molecule compounds with a sulfonic acid or sulfinic acid lactone structure, which have been verified to have extremely low skin irritation, excellent permeability, and high stability. They can not only significantly scavenge reactive oxygen species and reduce oxidative stress, but also alleviate skin inflammation, exhibiting excellent ability to protect and repair skin cells, and can be used as a new raw material for cosmetics.

Description

[0001] Cross-reference declaration

[0002] This patent application claims priority to Chinese Patent Application No. 2023114494701, filed on November 1, 2023, entitled “Small molecule compounds, compositions and applications thereof having a sulfonic acid or sulfinic acid lactone structure”, the entire contents of which are incorporated herein by reference. Technical Field

[0003] This invention relates to the field of biochemistry, and particularly to small molecule compounds, compositions, and their applications having sulfonic acid or sulfinic acid lactone structures. Background Technology

[0004] Protecting and repairing damaged skin is typically a crucial function of skincare and cosmetic products. Factors contributing to skin damage include hormone levels, excessive sun exposure, smog, and improper diet. Inflammation and damage to the skin can easily lead to roughness, pigmentation, reduced collagen, and decreased antioxidant capacity, ultimately accelerating skin aging in the long run. Studies have shown that quenching reactive oxygen species (ROS) and eliminating harmful free radicals can alleviate skin inflammation and repair damaged skin. While traditional antioxidants in existing cosmetics, such as vitamin C, vitamin E, niacinamide, and arbutin, possess some ROS scavenging ability, they still suffer from serious problems such as irritation, weak stability, and poor skin permeability.

[0005] Therefore, developing new cosmetic raw materials that are highly permeable, have low irritation, can quench ROS, eliminate harmful oxygen free radicals, and alleviate skin inflammation is of great value. Summary of the Invention

[0006] The purpose of this invention is to provide a small molecule compound having a sulfonic acid or sulfinic acid lactone structure.

[0007] Another object of the present invention is to provide a composition.

[0008] Another object of the present invention is to provide a method for relieving skin inflammation.

[0009] Another object of the present invention is to provide a method for reducing reactive oxygen species in skin cells.

[0010] Another object of the present invention is to provide uses of the above-described compounds or compositions.

[0011] To address the aforementioned technical problems, the first aspect of this invention provides a compound having the structure shown in general formula I, its salt, stereoisomer, or solvate.

[0012]

[0013] Where X represents oxygen or none;

[0014] R 1 and R 2 Each is independently hydrogen, hydroxyl, C 1-6 Alkyl, C 1-6 Alkoxy, amino, -NC(O)R 1-1 -NR a R b -CH2C(O)OR 1-2 , phenyl, -OC(O)R 1-3 -OCH2OC(O)R 1-4 Halogen, cyano, nitro, -C(O)R 1-5 or -C(O)OR 1-6 ;

[0015] R 1-1 R 1-2 R 1-3 R 1-4 R 1-5 and R 1-6 Each independently is hydrogen, C 1-6 Alkyl, halogen-substituted C 1-6 Alkyl or -NR a R b ;

[0016] R a and R b Independently hydrogen, C 1-6 alkyl or halogen-substituted C 1-6 alkyl.

[0017] In some preferred embodiments, X is oxygen.

[0018] In some preferred solutions, R 1 and R 2 Each is independently hydrogen, hydroxyl, C 1-6 Alkyl, C 1-6 Alkoxy, amino, -NC(O)R 1-1 -NR a R b -CH2C(O)OR 1-2 , phenyl, -OC(O)R 1-3 or -OCH2OC(O)R 1-4 .

[0019] In some preferred solutions, R 1 and R 2 Each independently consists of hydroxyl and C. 1-6 Alkoxy,

[0020] In some preferred embodiments, the C 1-6 Alkyl group is C 1-4 Alkyl; more preferably, the C 1-4 The alkyl group is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, or tert-butyl.

[0021] In some preferred embodiments, the C 1-6 The alkoxy group is C 1-4 Alkyl group; more preferably, C 1-4 The alkoxy group can be methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, or tert-butoxy.

[0022] In some preferred embodiments, the halogen is fluorine, chlorine, bromine, or iodine.

[0023] In some preferred embodiments, the halogen-substituted C 1-6 Alkyl groups are halogen-substituted C 1-4 Alkyl; more preferably halogen-substituted methyl, halogen-substituted ethyl, halogen-substituted n-propyl, halogen-substituted isopropyl, halogen-substituted n-butyl, halogen-substituted isobutyl or halogen-substituted tert-butyl.

[0024] In some preferred solutions, R 1 The compounds are hydroxyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy. More preferably, R 1 Hydroxyl, methoxy

[0025] In some preferred solutions, R 2 The compounds are hydroxyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy. More preferably, R 2 Hydroxyl, methoxy

[0026] In some preferred solutions, R 1-3 It can be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, or More preferably, R 1-3 It is isopropyl, tert-butyl or

[0027] In some preferred solutions, R 1-4 It can be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, or More preferably, R 1-4 It is tert-butyl.

[0028] In some preferred embodiments, the compound is selected from any of the following:

[0029]

[0030]

[0031]

[0032]

[0033] In some preferred embodiments, the compound is not

[0034] A second aspect of the present invention provides a composition comprising a salt, stereoisomer, or solvate of the compound described in the first aspect of the present invention; and a cosmetically acceptable excipient.

[0035] A third aspect of the invention provides the use of the compounds described in the first aspect of the invention, their salts, stereoisomers or solvates, or the compositions described in the second aspect of the invention, in the preparation of cosmetics.

[0036] A fourth aspect of the present invention provides a method for relieving skin inflammation, the method comprising the steps of: administering to a test subject a compound, salt thereof, stereoisomer thereof, or solvate according to the first aspect of the present invention; or administering to a test subject a composition according to the second aspect of the present invention.

[0037] A fifth aspect of the present invention provides a method for reducing reactive oxygen species in skin cells, the method comprising the steps of: administering to a test subject a compound, salt thereof, stereoisomer thereof, or solvate according to the first aspect of the present invention; or administering to a test subject a composition according to the second aspect of the present invention.

[0038] Compared with the prior art, the present invention has at least the following advantages:

[0039] This invention provides a class of small molecule compounds with sulfonic acid or sulfinic acid lactone structures, which have been verified to have extremely low skin irritation, excellent permeability and high stability. They can not only significantly scavenge reactive oxygen species and reduce oxidative stress, but also relieve skin inflammation. They have a good ability to protect and repair skin cells and can be used as new raw materials for cosmetics.

[0040] It should be understood that, within the scope of this invention, the above-described technical features of this invention and the technical features specifically described below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be described in detail here. Attached Figure Description

[0041] One or more embodiments are illustrated by way of example with reference to the accompanying drawings, and these illustrative descriptions do not constitute a limitation on the embodiments.

[0042] Figure 1 This is a behavioral trajectory diagram of a representative zebrafish selected from each group according to an embodiment of the present invention;

[0043] Figure 2 This is a statistical chart of the total distance traveled by each group of zebrafish within 20 minutes according to an embodiment of the present invention;

[0044] Figure 3 This is a statistical chart of the average movement speed of each group of zebrafish over 20 minutes according to an embodiment of the present invention;

[0045] Figure 4 This is a representative staining image of inflammatory cells at the nerve tumulus of each group of zebrafish according to an embodiment of the present invention;

[0046] Figure 5 This is a statistical diagram of the number of inflammatory cells at the neurotumor of each group of zebrafish according to an embodiment of the present invention;

[0047] Figure 6 This describes the effect of I-8 on wound repair in zebrafish according to embodiments of the present invention;

[0048] Figure 7 This is a statistical analysis of the caudal fin area of ​​each group of zebrafish according to an embodiment of the present invention;

[0049] Figure 8 This is a diagram illustrating the effect of I-8 in embodiments of the present invention on the number of inflammatory cells at the wound site of zebrafish;

[0050] Figure 9 This is a statistical chart of the number of inflammatory cells at each group of wounds according to an embodiment of the present invention.

[0051] Figure 10 is According to the embodiments of the present invention, the ROS content in zebrafish is shown in the white line area, which is the zebrafish yolk sac (ROS quantitative area), and the value in parentheses is the I-8 concentration, in μg / mL.

[0052] Figure 11 is Statistical analysis of ROS content in zebrafish according to embodiments of the present invention;

[0053] Figure 12 is According to the embodiments of the present invention, the zebrafish β-galactosidase activity image shows that the tissue appears blue-green after β-galactosidase staining. Comparing the tissues pointed to by the arrows on the dorsal fin, the more arrows there are, the deeper the blue-green color, and the higher the β-galactosidase activity.

[0054] Figure 13 isStatistics on β-galactosidase activity in zebrafish according to embodiments of the present invention;

[0055] Figure 14 is Statistical analysis of the expression of antioxidant and anti-aging related genes in zebrafish according to embodiments of the present invention;

[0056] Figure 15 is According to the embodiments of the present invention, compound I-8 can downregulate TNF-α in HaCaT;

[0057] Figure 16 is According to embodiments of the present invention, compound I-8 can downregulate IL-8 in HaCaT;

[0058] Figure 17 is According to embodiments of the present invention, compound I-8 can downregulate IL-1β of HaCaT;

[0059] Figure 18 is According to the embodiments of the present invention, compound I-8 can downregulate the EP2 of HaCaT;

[0060] Figure 19 is According to the embodiments of the present invention, compound I-8 can downregulate the TRPV1 of HaCaT;

[0061] Figure 20 is Compound I-8 scavenges ROS according to embodiments of the present invention;

[0062] Figure 21 is According to embodiments of the present invention, compound I-8 increases the secretion of type I collagen;

[0063] Figure 22 is According to embodiments of the present invention, compound I-8 upregulates COL3A1 gene expression;

[0064] Figure 23 is According to embodiments of the present invention, compound I-8 upregulates COL4A1 gene expression;

[0065] Figure 24 is According to embodiments of the present invention, compound I-8 upregulates COL7A1 gene expression;

[0066] Figure 25 is According to embodiments of the present invention, compound I-8 upregulates LAMA5 gene expression. Detailed Implementation

[0067] Traditional cosmetic antioxidants suffer from poor stability and require stringent application conditions; for example, they decompose in daylight and can only be applied at night. They also have poor skin permeability, often requiring high concentrations and large doses to be effective, and often showing no effect on some individuals. Furthermore, these antioxidants are highly irritating, requiring strict dosage control for those with sensitive skin, otherwise they can easily trigger severe allergic and inflammatory reactions. Through extensive and in-depth research, the inventors have developed a class of small-molecule polyphenolic compounds with sulfonic acid or sulfinic acid lactone structures. These compounds exhibit low irritation, good permeability, and high stability, and can significantly quench reactive oxygen species (ROS), scavenge harmful free radicals, eliminate oxidative stress, alleviate skin inflammation, and soothe and repair damaged skin cells. These compounds, along with their salts, stereoisomers, or solvates, can serve as novel cosmetic raw materials with the potential to replace existing antioxidants.

[0068] compound

[0069] This invention relates to a class of small molecule compounds having a sulfonic acid or sulfinic acid lactone structure, the structure of which is shown in general formula I below:

[0070]

[0071] In the above general formula I, X is oxygen or absent. Based on the beneficial effects of further improving the stability and permeability of the compound, X is oxygen.

[0072] R 1 and R 2

[0073] R 1 The quantity is 0-4, for example, 0, 1, 2, 3, or 4. As R 1 It can be hydrogen, hydroxyl, C 1-6 Alkyl, C 1-6 Alkoxy, amino, -NC(O)R 1-1 -NR a R b -CH2C(O)OR 1-2 , phenyl, -OC(O)R 1-3 -OCH2OC(O)R 1-4 Halogen, cyano, nitro, -C(O)R 1-5 or -COO R 1-6 Furthermore, R 1-1 R 1-2 R 1-3 R 1-4 R 1-5 and R 1-6 Each independently is hydrogen, C 1-6 Alkyl, halogen-substituted C 1-6 Alkyl or -NR a Rb ;R a and R b Independently hydrogen, C 1-6 alkyl or halogen-substituted C 1-6 Alkyl group. In a preferred embodiment, R 1 For hydrogen, hydroxyl, C 1-6 Alkyl, C 1-6 Alkoxy, amino, -NC(O)R 1-1 -NR a R b -CH2C(O)OR 1-2 , phenyl, -OC(O)R 1-3 or -OCH2OC(O)R 1-4 .

[0074] Similarly, R 2 The quantity is 0-4, for example, 0, 1, 2, 3, or 4. As R 2 It can be hydrogen, hydroxyl, C 1-6 Alkyl, C 1-6 Alkoxy, amino, -NC(O)R 1-1 -NR a R b -CH2C(O)OR 1-2 , phenyl, -OC(O)R 1-3 -OCH2OC(O)R 1-4 Halogen, cyano, nitro, -C(O)R 1-5 or -COO R 1-6 Furthermore, R 1-1 R 1-2 R 1-3 R 1-4 R 1-5 and R 1-6 Each independently is hydrogen, C 1-6 Alkyl, halogen-substituted C 1-6 Alkyl or -NR a R b ;R a and R b Independently hydrogen, C 1-6 alkyl or halogen-substituted C 1-6 Alkyl group. In a preferred embodiment, R 2 For hydrogen, hydroxyl, C 1-6 Alkyl, C 1-6 Alkoxy, amino, -NC(O)R 1-1 -NR a R b -CH2C(O)OR 1-2 , phenyl, -OC(O)R 1-3or -OCH2OC(O)R 1-4 .

[0075] In a more preferred embodiment of the present invention, the above-mentioned R 1-3 It can be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, or More preferably, R 1-3 It is isopropyl, tert-butyl or R 1-4 It can be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, or More preferably, R 1-4 It is tert-butyl.

[0076] R 1 and R 2 The quantities can be equal or unequal. In the preferred embodiment, R 1 and R 2 The quantities are equal.

[0077] R 1 and R 2 The substitution position is not limited, but in the preferred embodiment, R 1 and R 2 The positions are 4' and 4' of the biphenyl ring, respectively (as shown in I-8 of specific embodiments of the present invention). As used in the invention, the position numbering rule for the biphenyl ring is as follows: Furthermore, in the sulfonic acid or sulfinic acid lactone structure, the O atom is covalently bonded to the carbon atom on the 2' side, the S atom is covalently bonded to the carbon atom on the 2' side, and the oxygen atom and sulfur atom are covalently bonded to each other to form the lactone structure.

[0078] Based on the excellent effects of promoting the scavenging of reactive oxygen species and alleviating the effects of the compound, in a more preferred embodiment, R 1 and R 2 At least one of them has the ability to supply electrons to the parent ring, and preferably all of them have the ability to supply electrons to the target. For example, R 1 and R 2 Each independently consists of hydroxyl and C. 1-6 Alkoxy, The electron-donating group is shown. More preferably, R 1 and R 2 The compounds are hydroxyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy. In a specific embodiment of the present invention, R 1 Hydroxyl, methoxy In a specific embodiment of the present invention, R 2 Hydroxyl, methoxy

[0079] In several specific embodiments of the present invention, the specific structures of the compounds are shown in the table below, wherein compounds I-1 to I-12, as well as I-17 and I-18, have better permeability than I-13 to I-16.

[0080] surface

[0081]

[0082]

[0083]

[0084]

[0085] Composition

[0086] This invention also relates to compositions containing the compounds of this invention, their salts, stereoisomers, or solvates. The compositions of this invention can be cosmetic compositions or topical formulations.

[0087] As used in this invention, the term "cosmetics" refers to a product applied to the surface of the human body (such as the epidermis, hair, lips, etc.) by smearing, spraying or other similar methods to cleanse, care for, beautify or eliminate unpleasant odors, and the product has a soothing effect on the application site.

[0088] When used as a cosmetic composition, the composition of the compounds, salts, stereoisomers or solvates of the present invention serves as a cosmetic agent in the cosmetic composition. In addition, the composition also includes a cosmetically acceptable medium for diluting or dispersing the cosmetic agent or for use as a carrier thereon, so as to promote its distribution when the composition is applied to the skin.

[0089] These media can be aqueous, anhydrous, or emulsions. Oily carriers, in the presence of water and emulsifiers, form emulsion systems that act as carriers. Preferably, the composition is aqueous or an emulsion, especially water-in-oil or oil-in-water emulsions, with oil-in-water emulsions being more preferred. Cosmetic compositions are typically, but not limited to, liquids, creams, or emulsions. In a preferred embodiment, water is used as a carrier to form the cosmetic composition. In a preferred embodiment, the cosmetic composition further includes a carrier other than water, such as oil, fat, wax, oily matrix (e.g., coconut oil, palm oil, olive oil, castor oil, mink oil, snake oil, silicone oil and its derivatives, tallow, lanolin and its derivatives, carnauba wax, cetacean, beeswax, liquid paraffin, petrolatum, microcrystalline wax, squalane, fatty acids, fatty alcohols and esters, etc.), powdery matrix (e.g., talc, kaolin, zinc dioxide, titanium dioxide, bentonite, magnesium stearate, zinc stearate, calcium carbonate, magnesium carbonate, calcium hydrogen phosphate, etc.), and solvent matrix (e.g., alcohols, small molecule ketones, ethers, small molecule esters).

[0090] In the cosmetic compositions of the present invention, cosmetically acceptable excipients may also be added. The term "cosmetically acceptable excipient" refers to substances that contribute to the shaping, stabilization, or imparting color, fragrance, and other properties to a cosmetic. Examples include preservatives (such as benzoic acid and its derivatives, chlorobutanol, xylene, salicylic acid and its derivatives, sorbic acid and its derivatives, imidazolidinyl urea, phenylethanol, etc.), antioxidants (such as butylated hydroxyanisole, tert-butylated hydroxyanisole, vitamin E, propyl gallate, etc.), moisturizers (such as glycerin, propylene glycol, sorbitol, polyethylene glycol, lactic acid, sodium lactate, sodium pyrrolidone carboxylate, hyaluronic acid, hydrolyzed collagen, chitin and its derivatives, glucose esters), sunscreens (titanium dioxide, zinc oxide, aminobenzoates and their derivatives, salicylic acid and its derivatives, benzophenone, etc.), and surfactants (lecithin, soapberry). Glycosides, alkyl glycosides, etc.), antioxidants, colorants (such as organic synthetic pigments such as azo and anthraquinone pigments, or inorganic pigments such as zinc oxide, titanium dioxide, ferric oxide, ferrous hydroxide, chromium oxide, and iron oxide, or natural pigments such as capsicum red, sorghum red, and anthocyanins), fragrances (natural or blended fragrances such as ambergris, castoreum, musk, civet, rose oil, peppermint oil, spearmint oil, lavender oil, and fennel oil), water-soluble polymers, chelating agents (such as disodium EDTA and its derivatives), and film-forming agents (such as polyvinyl acetate, polyacrylate emulsions, hyaluronic acid, and polyglycerol-2 isostearate).

[0091] In the cosmetic composition of the present invention, there is no limitation on the amount of each raw material and excipient added.

[0092] Use of the compound or composition

[0093] The present invention also relates to the use of the above-described compounds, their salts, stereoisomers or solvates, or compositions containing the above-described compounds, their salts, stereoisomers or solvates, for: (i) preparing cosmetics; (ii) relieving skin inflammation; (iii) reducing reactive oxygen species in skin cells; and / or (iv) repairing damaged skin.

[0094] As used herein, the term "skin" includes the skin on the face, neck, chest, back, arms, armpits, buttocks, hands, legs, and scalp. As used herein, a cosmetic beneficial agent refers to a component that (a) improves facial or body characteristics such as skin characteristics upon topical application, (b) is beneficial to facial or body characteristics such as skin characteristics, or (c) both (a) and (b). In a preferred embodiment, the compound or cosmetic composition is used for topical application.

[0095] Methods to relieve skin inflammation and / or reduce reactive oxygen species in skin cells and / or repair damaged skin

[0096] The present invention also relates to methods for (a) alleviating skin inflammation and / or (b) reducing reactive oxygen species in skin cells and / or (c) repairing damaged skin, comprising the steps of administering the above-described compound, its salt, stereoisomer, or solvate to a test subject; or, administering the above-described composition to a test subject. The administration method is non-enteric; preferably topical, such as by smearing or applying topically, in appropriate amounts to the skin, mucous membranes, etc.

[0097] the term

[0098] As used herein, the term "alkyl" refers to a linear or branched saturated monovalent hydrocarbon group, wherein the alkyl group may optionally be substituted with one or more substituents. In certain embodiments, the alkyl group is having a carbon number of 1 to 20 (C0). 1-20 ), 1 to 15 (C 1-15 ), 1 to 12 (C 1-12 ), 1 to 10 (C 1-10 ) or 1 to 6 (C 1-6 A linear saturated monovalent hydrocarbon group with 3 to 20 carbon atoms, or a hydrocarbon group with 3 to 20 carbon atoms. 3-20 ), 3 to 15 (C 3-15 ), 3 to 12 (C 3-12 ), 3 to 10 (C 3-10 ) or 3 to 6 (C 3-6 A branched, saturated monovalent hydrocarbon group with 1 carbon atom. The linear C used here... 1-6 and C with branches 3-6 Alkyl groups are also called "lower alkyl groups". Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl (including all isomers), n-propyl, isopropyl, butyl (including all isomers), n-butyl, isobutyl, tert-butyl, pentyl (including all isomers), and hexyl (including all isomers). For example, C 1-6 Alkyl refers to a linear saturated monovalent hydrocarbon group having 1 to 6 carbon atoms or a branched saturated monovalent hydrocarbon group having 3 to 6 carbon atoms. In one embodiment, the alkyl group is an optionally substituted alkyl group as described elsewhere herein. In some embodiments, C 1-6 Alkyl group is C 1-4 Alkyl, C 1-4 The alkyl group is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, or tert-butyl. In other embodiments, C 1-6 One or more hydrogen atoms in the alkyl group are substituted with halogens. In other embodiments, the halogen-substituted C... 1-6 Alkyl groups are halogen-substituted C 1-4 Alkyl group. In other embodiments, halogen-substituted C 1-4The alkyl group is a halogen-substituted methyl, a halogen-substituted ethyl, a halogen-substituted n-propyl, a halogen-substituted isopropyl, a halogen-substituted n-butyl, a halogen-substituted isobutyl, or a halogen-substituted tert-butyl.

[0099] As used herein, the term "alkoxy group" refers to a stable straight-chain or branched, or cyclic, hydrocarbon group, or a combination thereof, consisting of the indicated number of carbon atoms and one or more (one to three in one embodiment) O atoms. Examples of alkoxy groups include, but are not limited to, -O-CH3, -O-CH2-CH3, -O-CH2-CH2-CH3, -O-CH-(CH3)2, and -O-CH2-CH2-O-CH3. In one embodiment, the alkoxy group is an optionally substituted alkoxy group as described elsewhere herein. In some embodiments, the alkoxy group is C 1-6 Alkyl group. In some embodiments, the alkoxy group is C60. 1-4 Alkyl group. In some embodiments, the alkoxy group is methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, or tert-butoxy. In other embodiments, C 1-6 One or more hydrogen atoms in the alkoxy group are substituted with halogens. In other embodiments, the halogen-substituted C... 1-6 The alkoxy group is a halogen-substituted methoxy, a halogen-substituted ethoxy, a halogen-substituted n-propoxy, a halogen-substituted isopropoxy, a halogen-substituted n-butoxy, a halogen-substituted isobutoxy, or a halogen-substituted tert-butoxy.

[0100] As used in this invention, the term "halogen" refers to fluorine, chlorine, bromine, or iodine.

[0101] As used in this invention, "(O)" represents the =O structure, such as -NH-C(O)- being an amide group, -OC(O)- being an acyloxy group, and -C(O)- being a carbonyl group.

[0102] As used herein, the term "hydrogen" includes protons (¹H), deuterium (²H), tritium (³H), and / or mixtures thereof. In the compounds described herein, one or more hydrogen-occupied sites may be enriched with deuterium and / or tritium. Such isotopically enriched analogs can be prepared from suitable isotopically labeled starting materials available from commercial sources or by known literature procedures.

[0103] As used in this article, the term "hydroxyl" refers to -OH.

[0104] As used in this article, the term "amino" refers to -NH2.

[0105] As used in this article, the term "cyano" refers to -CN.

[0106] As used in this article, the term "nitro" refers to -NO2.

[0107] The term “substitution” refers to a portion having a substituent that replaces a hydrogen atom or one or more non-hydrogen atoms on the molecule.

[0108] As used herein, the term "solvent compound" refers to a compound formed by the interaction of a solvent with a compound provided in this invention or a salt thereof.

[0109] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the present invention is further described below in conjunction with specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments that do not specify specific conditions are generally performed under conventional conditions or as recommended by the manufacturer. Unless otherwise stated, percentages and parts are weight percentages and parts by weight. Unless otherwise specified, the experimental materials and reagents used in the following embodiments are commercially available.

[0110] Unless otherwise specified, the technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains. It should be noted that the terms used herein are for the purpose of describing particular embodiments only and are not intended to limit the exemplary embodiments of this application.

[0111] General Synthesis Step 1

[0112]

[0113] General Synthesis Step Two

[0114]

[0115] General Synthesis Step 3

[0116]

[0117] General Synthesis Step Four

[0118]

[0119] Example 1: Synthesis and Characterization of Compound I-8

[0120]

[0121] Synthesis of intermediate 22-Iodo-5-methoxyphenol (0.73 g, 2.92 mmol, 1.2 eq) and DMAP (0.03 g, 0.24 mmol, 0.1 eq) were dissolved in DCM (5.00 mL) and pyridine (10.00 mL). The mixture was cooled to 0 °C and stirred until homogeneous. Compound 1 (0.50 g, 2.43 mmol, 1 eq) was then dissolved in DCM (5.00 mL). This solution was then slowly added dropwise to the reaction system while maintaining the temperature at 0 °C. After the addition was complete, the system was allowed to warm naturally to room temperature, and the reaction was allowed to proceed overnight. LC-MS showed that the starting material had reacted completely. Stirring was stopped, and the mixture was extracted with water and DCM. The organic phase was separated, concentrated, and purified by silica gel column chromatography (PE:EA = 10:1) to obtain intermediate 2 (0.96 g, 2.23 mmol, 94.61% yield, 93.42% purity) as a yellow oil. LC-MS: [M+1] + =421.

[0122] Synthesis of intermediate 3 (i.e., I-17) Compound 2 (0.2 g, 0.48 mmol, 1 eq), Pd(Pivate)2 (0.015 g, 0.048 mmol, 0.1 eq), TBAB (0.18 g, 0.57 mmol, 1.2 eq), and potassium acetate (0.14 g, 1.43 mmol, 3 eq) were dissolved in DMAc (2.00 mL). The mixture was fully purged with nitrogen, heated to 60 °C, and stirred overnight under nitrogen protection. The reaction was monitored by LC-MS until the reaction was complete. The mixture was then filtered, extracted with EA, concentrated, and purified by silica gel column chromatography (PE:EA = 1:1) to give intermediate 3 (0.12 g, 0.42 mmol, 88.49% yield, 93.32% purity) as a white solid. 1 H NMR(400MHz,Chloroform-d)δ7.76(d,J=2.8Hz,1H),7.74(d,J=2.7Hz,1H),7.43(d,J=2.7Hz,1H),7.28– 7.23(m,1H),6.93(dd,J=8.8,2.6Hz,1H),6.84(d,J=2.6Hz,1H),3.91(s,3H),3.87(s,3H).LC-MS: [M+1] + =293.

[0123] Synthesis of compound I-8Compound 3 (1.00 g, 3.42 mmol, 1 eq) was dissolved in DCM (20.00 mL), cooled to 0 °C, and BBr3 (1.0 M in DCM, 13.68 mmol, 13.68 mL, 4 eq) was slowly added dropwise under nitrogen protection, with the temperature controlled at 0 °C. After the addition was complete, the system was allowed to warm naturally to room temperature and reacted overnight. The reaction was monitored by LC-MS until it ended. The reaction was quenched with ice water, extracted with EA and DCM, concentrated, and purified by silica gel column chromatography (PE:EA = 1:1) to obtain compound I-8 (0.25 g, 0.95 mmol, 27.78% yield, 100% purity) as a pale yellow solid. 1H NMR (400MHz, DMSO-d6) δ7.96(d,J=8.5Hz,1H),7.92(d,J=8.8Hz,1H),7.27–7.22(m,2H),6.87(dd,J=8.6,2.4Hz,1H),6.78(d,J=2.4Hz,1H).LC-MS: [M-1] - =263.

[0124] Example 2: Synthesis and Characterization of Compound I-11

[0125]

[0126] In Example 1, compound 3' was also generated during the reaction from compound 2 to compound 3. Compound 3' was then subjected to the same demethylation step to give compound I-11. LC-MS: [M-1] - =263.

[0127] Example 3: Synthesis and Characterization of Compound I-13

[0128]

[0129] Compound I-8 (130 mg, 0.49 mmol), DMAP (6 mg, 0.05 mmol), and DIEA (254 mg, 1.97 mmol) were dissolved in THF (5 mL) and cooled to 0 °C. Then, dimethylcarbamoyl chloride (212 mg, 1.97 mmol) was slowly added dropwise to the reaction system while maintaining the temperature at 0 °C. After the addition was complete, the system was allowed to warm naturally to room temperature and reacted overnight. The mixture was then concentrated and purified by silica gel column chromatography (DCM:EA = 100:5) to obtain I-13 (90 mg, yield 45.01%) as a white solid. 1H NMR (400MHz, DMSO-d6) δ8.33(d,J=8.8Hz,1H),8.30(d,J=8.8Hz,1H),7.92(d,J=2.4Hz,1H),7.74(dd,J=8.7,2.5Hz,1H) ,7.45(d,J=2.3Hz,1H),7.34(dd,J=8.7,2.4Hz,1H),3.08(d,J=6.2Hz,6H),2.94(d,J=2.8Hz,6H).LCMS:m / z=407.1(M+H + (ESI).

[0130] Example 4: Synthesis and characterization of compounds I-14 and I-15

[0131] Using the synthesis method described in Example 3, and again using I-8 as the starting material, replacing dimethylcarbamoyl chloride with isobutyryl chloride or tervapotranol chloride yields I-14 and I-15, both white solids. I-14: LCMS: m / z = 405 (M+H) + (ESI).I-15: 1 HNMR (400MHz, DMSO-d6) δ8.24(d,J=8.8Hz,1H),8.07(dd,J=7.7,1.3Hz,1H),7.83(t,J=8.0Hz,1H),7.75(dd,J=8 .3,1.3Hz,1H),7.54(d,J=2.4Hz,1H),7.38(dd,J=8.8,2.4Hz,1H),1.35(s,9H),1.32(s,9H).LCMS:m / z=433(M+H + (ESI).

[0132] Example 5: Synthesis and Characterization of Compound I-1

[0133] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-1 was prepared: LCMS: m / z = 247 (MH). + (ESI).

[0134] Example 6: Synthesis and Characterization of Compound I-2

[0135] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-2 was prepared: LCMS: m / z = 295 (MH). + (ESI).

[0136] Example 7: Synthesis and Characterization of Compound I-3

[0137] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-3 was prepared: LCMS: m / z = 247 (MH). + (ESI).

[0138] Example 8: Synthesis and Characterization of Compound I-4

[0139] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-1 was prepared: LCMS: m / z = 311 (MH). + (ESI).

[0140] Example 9: Synthesis and Characterization of Compound I-5

[0141] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-5 was prepared: LCMS: m / z = 279 (MH). + (ESI).

[0142] Example 10: Synthesis and Characterization of Compound I-6

[0143] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-6 was prepared: LCMS: m / z = 295 (MH). + (ESI).

[0144] Example 11: Synthesis and Characterization of Compound I-7

[0145] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-7 was prepared: LCMS: m / z = 295 (MH). + (ESI).

[0146] Example 12: Synthesis and Characterization of Compound I-9

[0147] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-9 was prepared: LCMS: m / z = 263 (MH). + (ESI).

[0148] Example 13: Synthesis and Characterization of Compound I-10

[0149] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-10 was prepared: LCMS: m / z = 279 (MH). + (ESI).

[0150] Example 14: Synthesis and Characterization of Compound I-12

[0151] Following general synthetic steps one and two, and employing the synthetic method described in Example 1, the compound was synthesized... Using [a specific ingredient], compound I-12 was prepared: LCMS: m / z = 247 (MH). + (ESI).

[0152] Example 15: Synthesis and Characterization of Compound I-16

[0153] According to general synthetic step four, with compound I-3 and Using [M+H] as a raw material, compound I-16 was prepared: LCMS: m / z = 493 (M+H) + (ESI).

[0154] Example 16: Synthesis and Characterization of Compound I-18

[0155] According to the general synthetic step one, with compound Using raw materials, intermediate 4 was prepared:

[0156]

[0157] Intermediate 4 was deprotected by hydrogenation to obtain compound I-18: LCMS: m / z = 293 (MH) + (ESI).

[0158] Example 17: Synthesis and Characterization of Compound I-19

[0159] According to the general synthetic step one, with compound Compound I-19 was prepared from the raw material: 1HNMR (400MHz, DMSO-d6) δ10.95(s,1H),10.67(s,1H),8.58(d,J=8.9Hz,1H),7.32–7.29(m,1H),7.28–7. 26(m,1H),7.26–7.23(m,1H),6.98(dd,J=8.4,1.2Hz,1H),6.90(dd,J=8.1,1.2Hz,1H).LCMS:m / z=263(MH + (ESI).

[0160] Example 18: Synthesis and Characterization of Compound I-20

[0161] According to the general synthetic step one, with compound Compound I-20 was prepared from the raw material: 1 H-NMR(400MHz,DMSO-d6)δ10.77(s,1H),10.46(s,1H),10.22(s,1H),8.45–8.43(d,1H),7.2 5-7.24(d,1H),7.21–7.18(dd,1H),6.45–6.44(d,1H),6.28-6.27(d,1H).LCMS: m / z=279(MH + (ESI).

[0162] Example 19: Synthesis and Characterization of Compound I-21

[0163] According to the general synthetic step one, with compound Using raw materials, intermediate 5 was prepared:

[0164]

[0165] Intermediate 5 was deprotected by hydrogenation to obtain compound I-21: 1 H-NMR (400MHz, DMSO-d6) δ10.48(s,1H),8.07(d,J=8.7Hz,1H),7.99(d,J=8.7Hz,1H),7.48–7 .41(m,2H),6.89(dd,J=8.7,2.4Hz,1H),6.81(d,J=2.4Hz,1H),3.90(s,3H).LCMS:m / z=277(MH + (ESI).

[0166] Example 20: Synthesis and Characterization of Compound I-22

[0167] According to the general synthetic step one, with compound Using raw materials, intermediate 6 was prepared:

[0168]

[0169] Intermediate 6 was deprotected by hydrogenation to obtain compound I-22: 1 H-NMR (400MHz, DMSO-d6) δ11.04(s,1H),8.67(d,J=9.0Hz,1H),7.49(d,J=2.8Hz,1H),7.44(dd,J=9.1,2.9Hz,1H ),7.30(t,J=8.2Hz,1H),7.01(dd,J=8.3,1.2Hz,1H),6.92(dd,J=8.1,1.1Hz,1H),3.91(s,3H).LCMS:m / z=277(MH + (ESI).

[0170] Example 21: Synthesis and Characterization of Compound I-23

[0171] According to the general synthetic step one, with compound Using [a specific ingredient], compound I-23 was prepared: LCMS: m / z = 277 (MH). + (ESI).

[0172] Example 22: Synthesis and Characterization of Compound I-24

[0173] According to the general synthetic step one, with compound Using [a specific ingredient], compound I-24 was prepared: LCMS: m / z = 281 (MH). + (ESI).

[0174] Example 23: Synthesis and Characterization of Compound I-25

[0175] According to the general synthetic step one, with compound Using [a specific ingredient], compound I-25 was prepared: LCMS: m / z = 297 (MH). + (ESI).

[0176] Example 24: Synthesis and Characterization of Compound I-26

[0177] According to the general synthetic step one, with compound Using [a specific ingredient], compound I-26 was prepared: LCMS: m / z = 277 (MH). + (ESI).

[0178] Example 25: Synthesis and Characterization of Compound I-27

[0179] According to the general synthetic step one, with compound Using [a specific ingredient], compound I-27 was prepared: LCMS: m / z = 281 (MH). + (ESI).

[0180] Example 26: Synthesis and Characterization of Compound I-28

[0181] According to the general synthetic step one, with compound Using [a specific ingredient], compound I-28 was prepared: LCMS: m / z = 297 (MH). + (ESI).

[0182] Test Example 1: Zebrafish model to evaluate the soothing and anti-inflammatory effects of compounds.

[0183] Laboratory animals: Healthy wild-caught AB strain zebrafish that have reached 3 dpf (days post-fertilization).

[0184] Evaluation of the efficacy in relieving stinging: Healthy AB-strain zebrafish were randomly divided into four groups: blank control, model group, positive control group, and sample group. The blank control group zebrafish were treated with embryo culture water; the model group zebrafish were treated with a modeling agent (100 μM MSDS, sodium dodecyl sulfate); the positive control zebrafish were treated with both the modeling agent and a positive control drug (200 μM asiaticoside solution); the sample group zebrafish were treated with both the modeling agent and different concentrations of the test sample (high, medium, and low concentrations). After continuous treatment for a certain period, the zebrafish's behavioral trajectories were recorded using a behavior analyzer, and the changes in their total distance and speed were statistically analyzed. The results are presented in... Figures 2-4 In this study, SDS caused stinging pain in zebrafish, prompting them to move more vigorously. Therefore, compared with the blank control group, the total distance and average speed of movement in the model group were significantly increased. Figure 2 In the behavioral trajectory diagram, the black lines represent the zebrafish's movement trajectory inside the hole. The more numerous and chaotic the lines, the more vigorous the zebrafish's movement.

[0185] Evaluation of soothing and anti-inflammatory efficacy: As described above, the modeling agent was 40 μM copper sulfate, the positive control agent was 200 μM asiaticoside, and the I-8 concentrations were 66.6, 22.2, and 7.4 μg / mL. After treating zebrafish as described above, inflammatory cells in their bodies were stained, the number of inflammatory cells at the neurothalamus was counted, and statistical analysis was performed. The results are presented in... Figures 5-6 middle.

[0186] Conclusion: I-8 exhibits certain analgesic effects at effective concentrations of 22.2, 66.6, and 200 μg / mL. It also exhibits certain anti-inflammatory effects at effective concentrations of 7.4, 22.2, and 66.6 μg / mL.

[0187] Test Example 2: Evaluating the ability of compounds to penetrate the skin using the Franz diffusion cell

[0188] Experimental materials: skin from the back of a one-month-old Bama miniature pig, 0.8–1.0 mm thick; 0.1 mg / mL solution of the test compound, in 50 mM Sodium Phosphate Buffer (PB) containing 20% ​​ethanol.

[0189] Experimental procedure:

[0190] 1. Pipette 400 μL of the test compound solution into the feed cell, repeat three times.

[0191] 2. Add approximately 7600 μL of 50 mM PB buffer solution to the appropriate receiving cell.

[0192] 3. The test compound was permeated at 32℃ for 2 h and 6 h.

[0193] 4. At 2h and 6h time points, the skin was washed with 50mM PB buffer solution containing 20% ​​ethanol, the skin from the infiltrated area was removed and homogenized, and acetonitrile solution containing internal standard was added.

[0194] 5. Centrifuge, take 100 μL of the supernatant and add it to 100 μL of purified water. Mix well and then perform quantitative analysis by LC-MS / MS.

[0195] The quantitative detection results of the compounds in the examples penetrating the skin are shown in Table 2 below:

[0196] Table 2

[0197]

[0198] Compound I-8 has good skin penetration ability, and can reach a penetration amount of 10ug / g within 6 hours.

[0199] Test Example 3: Evaluation of Compound Repair Efficacy Using a Zebrafish Model

[0200] Laboratory animals: Healthy wild-caught AB strain zebrafish that have reached 3 dpf (days post-fertilization).

[0201] Evaluation of wound repair efficacy: Healthy AB strain zebrafish with a developmental age of 3dpf (3 days post-floor) were used as experimental animals and randomly divided into a blank control group, a model group, a positive control group, and a sample group. The blank control group zebrafish were treated with culture water; the model group zebrafish were treated with culture water after caudal fin removal; the positive control and sample groups zebrafish were treated with 600 μg / mL hyaluronic acid and different concentrations of I-8 (1, 3, 9 μg / mL), respectively, after caudal fin removal. All groups of larvae were placed in a constant temperature incubator at 28.5℃ and kept in the dark for 48 hours, with the medium changed every 24 hours. The caudal fin area of ​​each group of zebrafish was calculated using Image-Pro Plus software, and the data were statistically analyzed using GraphPad software. The results are presented in... Figures 7-8 middle.

[0202] Evaluation of anti-inflammatory efficacy: As described above, the fry in each group were placed in a constant temperature incubator at 28.5℃ and continuously protected from light for 24 hours. Inflammatory cells in the fish were labeled with neutral red, and the number of inflammatory cells at the tail docking site was counted. GraphPad software was used for statistical analysis of the data. The results are presented in… Figures 9-10 .

[0203] Conclusion: Figures 7-8 As shown, compared with the blank control group, although the caudal fin of zebrafish in the model group regenerated, its area was significantly reduced. When the zebrafish tail-amputation model was treated with I-8 at concentrations of 1, 3, and 9 μg / mL, the caudal fin area of ​​the zebrafish increased significantly compared with the model group. These results suggest that when the concentration of I-8 is 1, 3, or 9 μg / mL, it can further promote the regeneration of caudal fin tissue and has a certain wound-repairing effect. Figures 9-10 The results showed that, compared with the blank control group, the number of inflammatory cells at the tail amputation site of zebrafish in the model group was significantly increased. When the zebrafish tail amputation model was treated with I-8 at concentrations of 1, 3, and 9 μg / mL, the number of inflammatory cells at the wound site was significantly reduced compared with the model group. These results suggest that I-8 has a certain anti-inflammatory effect at concentrations of 1, 3, and 9 μg / mL.

[0204] Test Example 4: Kinetic solubility in aqueous buffer compared to urolithin A

[0205] Experimental Method: 30 μL of the 10 mM DMSO stock solution of the test compound was placed in a solubility plate (two replicates). 970 μL of aqueous buffer solution was then added to the plate. The plate was sealed with a sealing membrane and shaken at 1100 rpm for 2 hours at 25°C. The sample from the solubility plate was transferred to a filter plate and filtered. 10 μL of the filtrate was added to 980 μL of methanol along with 10 μL of DMSO. The solution was then diluted 10-fold with methanol:water (1:1). The resulting sample was analyzed by LC-MS / MS to obtain the Area (filtered). Separately, 10 mM of the DMSO stock solution of the test compound was diluted to 300 μM with DMSO. 10 μL of this 300 μM DMSO solution and 10 μL of aqueous buffer solution were added to 980 μL of methanol. The solution was then diluted 10-fold with methanol:water (1:1). The resulting sample was analyzed by LC-MS / MS to obtain the Area (std). Calculate the kinetic solubility using the following formula:

[0206]

[0207] Where DF is the dilution factor.

[0208] Urolithin A 98.76 I-8 >300*

[0209] *When the actual solubility is greater than 300 μM, an accurate kinetic solubility value cannot be measured.

[0210] Conclusion: Compared with the comparative compound urolithin A, I-8 has better solubility, overcoming the problem that urolithin A has poor water solubility and is difficult to develop into topical formulations.

[0211] Test Example 5: Evaluating the antioxidant efficacy of compounds using a zebrafish model

[0212] Laboratory animals: Healthy wild-caught AB strain zebrafish at 48 hpf (huors post-fertilization).

[0213] Evaluation of antioxidant efficacy: Wild AB strain zebrafish with a growth rate of 48 hpf were used as experimental animals. A zebrafish oxidative aging model was constructed using chemical mutagenesis with 4 μM menadione as the modeling agent. Fucoxanthin was used as a positive control compound. After treating the zebrafish oxidative aging model with different concentrations of I-8 for 22 h, changes in the levels of reactive oxygen species (ROS), β-galactosidase activity, and the expression of three antioxidant and anti-aging related genes (Cu / Zn-sod, Mn-sod, and ampk) were detected and statistically analyzed.

[0214] During the experiment, the modeling agent, positive compound, and I-8 were dispersed together in the zebrafish culture water. The larvae in each group were placed in a constant temperature incubator at 28.5℃ and continuously protected from light for 22 hours. Cell ROX was used. TM Deep Red fluorescent probes and β-galactosidase staining kits were used to specifically label ROS and β-galactosidase in fish, respectively, and the staining intensity was quantified using Image-Pro Plus software. After extracting RNA from whole fish, the expression changes of three genes—Cu / Zn-sod, Mn-sod, and ampk—were detected at the mRNA level using quantitative real-time PCR (RT-qPCR). The data were statistically analyzed using GraphPad software. The results are shown in Figures 10–14.

[0215] Conclusion: As shown in Figures 10-13, compared with the blank control group, after continuous treatment with menadione for 22 hours, the ROS content and β-galactosidase activity in zebrafish were significantly increased, and these increases were statistically significant. Menadione is an oxidant that can generate unstable semiquinones through the intracellular reductase system, leading to the production of large amounts of ROS and accelerating aging. β-galactosidase is a hydrolytic enzyme in lysosomes, and its increased activity is a significant characteristic of cellular aging. When the zebrafish oxidative aging model was treated with I-8 at a concentration of 22.2 μg / mL, compared with the model group, both the ROS content and β-galactosidase activity were significantly reduced. These results suggest that I-8 at a concentration of 22.2 μg / mL has certain antioxidant effects.

[0216] like Figure 14 The results showed that when zebrafish oxidative aging models were treated with I-8 at concentrations of 2.5, 7.5, and 22.2 μg / mL, the expression of both Cu / Zn-sod and Mn-sod genes was upregulated significantly compared to the model group. When the zebrafish oxidative aging model was treated with I-8 at a concentration of 2.5 μg / mL, the expression of the ampk gene was upregulated significantly compared to the model group.

[0217] The results suggest that when the concentration of I-8 is 2.5, 7.5 and 22.2 μg / mL, the abnormal expression of the Cu / Zn-sod and Mn-sod genes induced by menadione can be reversed; and when the concentration is 2.5 μg / mL, the abnormal expression of the ampk gene induced by menadione can be reversed.

[0218] Inflammation is a physiological response that protects the body from various forms of harm, such as physical injury, pathogens, exposure to toxic chemicals, and ultraviolet radiation. Early inflammation is primarily characterized by capillary dilation, increased permeability, and edema. Various inflammatory mediators play crucial roles in the development of acute and chronic inflammation, including interleukin-1α (IL-1β), interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α), and prostaglandin E2 (PGE2). PGE2 is the most abundant prostaglandin produced in the body, and its production begins with arachidonic acid. Arachidonic acid is first converted to prostaglandin H2 (PGH2) by cyclooxygenase (COX), and then further catalyzed by prostaglandin E synthase to generate PGE2. Ultimately, PGE2 exerts its predominantly inflammatory biological functions by acting on four prostaglandin E (EP) receptors, EP1-4. All EP2 receptors are coupled to Gs proteins and primarily signal via the cAMP-PKA-CREB pathway triggered by adenylate cyclase. Studies have also demonstrated that selective small-molecule antagonists targeting EP2 receptors can be developed to alleviate downstream pathological processes mediated by EP2 receptors, thus enabling the development of next-generation anti-inflammatory therapies.

[0219] Furthermore, the TRPV1 receptor is a nociceptor that can be activated by various factors, including chemical substances (capsaicin), noxious thermal stimuli, and acidification. TRPV1 receptors are widely distributed in class C sensory nerve afferent fibers and keratinocytes. Once activated, TRPV1 receptors release monovalent and divalent cations (primarily Ca2+). 2+ TRPV1 enters cells, triggers action potentials, and transmits signals to the higher central nervous system, producing a burning sensation. Therefore, by acting on the test substance, blocking or inhibiting the expression of the TRPV1 receptor protein can help relieve the burning sensation and achieve a soothing effect. Thus, the expression level of the TRPV1 gene in keratinocytes after sample treatment can be detected as a preliminary indicator to determine whether the sample has a soothing effect.

[0220] In the following test examples 6-1 and 6-2, immortalized keratinocytes (HaCaT) and UVB were used to establish a model. After the test compound was applied to HaCaT, the expression levels of TNF-α, IL-8, IL-1β and EP2 were detected. In the test example 6-2, immortalized keratinocytes (HaCaT) and capsaicin were used to establish a model. After the test compound was applied to HaCaT, the expression level of TRPV1 was detected. This was done to evaluate the soothing effect of the test substance from multiple dimensions.

[0221] Test Example 6-1: Detection of TNF-α, IL-8, IL-1β and EP2 gene expression in HaCaT after UVB radiation

[0222] 1) Cell seeding: HaCaT cells were seeded at 6×10⁵ cells / well in 6-well plates and cultured in an incubator (37℃, 5% CO₂) for 12 h. Normal control group (0 mJ / cm² + solvent control), model control group (80 mJ / cm² + solvent control), low concentration group (80 mJ / cm² + low concentration compound), medium concentration group (80 mJ / cm² + medium concentration compound), and high concentration group (80 mJ / cm² + high concentration compound) were set up.

[0223] 2) UVB modeling: Before UVB irradiation, HaCaT cells were washed three times with D'Hanks. 1 mL of D'Hanks was added to the wells to submerge the cells. The control group was wrapped in tin foil and placed in the dark. For the experimental groups, UVB modeling (80 mJ / cm²) was performed on the groups requiring irradiation.

[0224] 3) Drug administration: Each experimental group was added to DMEM medium containing different concentrations and compounds and cultured for another 24 hours;

[0225] 4) After the culture was completed, the cells were collected, total RNA was extracted from each experimental group, cDNA was synthesized, and the gene expression of β-actin and the target gene was detected by q-PCR.

[0226] 5) Using β-actin as an internal reference for gene expression, calculate the relative RNA expression level of the target gene.

[0227] relative RNA expression level = 2 △△C(t)

[0228]

[0229] △C(t)=C(t) 目的基因 -C(t) β-actin

[0230] Test Example 6-2: Detection of TRPV1 gene expression in HaCaT after capsaicin stimulation

[0231] 1) Cell seeding: HaCaT cells were seeded at 6×10⁵ cells / well in 6-well plates and cultured in an incubator (37℃, 5% CO₂) for 12 h. Normal control group (0 μM capsaicin + solvent control), model control group (0 μM capsaicin + solvent control), low concentration group (15 μM capsaicin + low concentration compound), medium concentration group (15 μM capsaicin + medium concentration compound), and high concentration group (15 μM capsaicin + high concentration compound) were set up.

[0232] 2) Induction and Drug Administration: Discard the culture medium in the 6-well plate and proceed with the drug administration. Add 1 mL of culture medium containing the test substance and capsaicin stock solution to each well according to the drug groups described above. After drug administration, place the 24-well plate in an incubator (37℃, 5% CO2) and incubate for 24 h ± 2 h.

[0233] 3) After incubation, D-Hanks gently rinsed the cells once or twice. Fresh culture medium was added to the normal control group, and fresh culture medium containing the corresponding concentration of the compound was added to the sample group. The cells were cultured at 37°C and 5% CO2 for 24 hours.

[0234] 4) After the culture was completed, the cells were collected, total RNA was extracted from each experimental group, cDNA was synthesized, and the gene expression of β-actin and the target gene was detected by q-PCR.

[0235] 5) Using β-actin as an internal reference for gene expression, calculate the relative RNA expression level of the target gene.

[0236] relative RNA expression level = 2 △△C(t)

[0237]

[0238] △C(t)=C(t) 目的基因 -C(t) β-actin

[0239] like Figure 15~ Figure 19 The results showed that compound I-8 could downregulate the expression levels of TNF-α, IL-8, IL-1β, EP2, and TRPV1 in a dose-dependent manner, suggesting that compound I-8 has a soothing effect.

[0240] Test Example 7: Evaluation of the antioxidant efficacy of compounds using HaCaT human immortalized keratinocytes

[0241] Oxidative reactions such as respiration and metabolism in organisms produce reactive free radicals. Under normal bodily conditions, free radicals have a stable scavenging system, maintaining a low concentration of free radicals in the body. Oxidative stress (OS) refers to an imbalance between oxidation and antioxidation in the body due to endogenous and / or exogenous stimuli, resulting in excessive free radical production. Excessive free radicals produce a series of negative effects in the body, including oxidative damage to biomolecules and further causing cell death and tissue damage. In the body, almost all ROS, except for trace amounts utilized by the body, should be promptly eliminated. UVB (280-319nm) radiation damage is the most important factor causing photoaging of the skin, primarily damaging keratinocytes (HaCaT), specifically manifested as the accumulation of photoproducts, increased ROS, and exacerbated oxidative damage. Over long periods of evolution, organisms have developed a complete antioxidant system, maintaining a dynamic balance between the production and elimination of free radicals. This is an adaptive mechanism of organisms, and its main components include antioxidant enzymes, antioxidants, and proteins that separate transition metals, all of which can specifically limit oxidative damage to the body. Therefore, the antioxidant efficacy of a test compound can be evaluated by detecting the ROS scavenging rate in keratinocytes after UVB radiation. Example 7 below uses HaCaT immortalized human keratinocytes to evaluate the antioxidant efficacy of a compound.

[0242] Test Example 7: Detection of ROS content in HaCaT after UVB radiation

[0243] 1) Cell seeding: HaCaT was seeded at a rate of 2 × 10⁻⁶. 4 100 cells / well were seeded into 96-well plates and incubated in an incubator (37℃, 5% CO2) for 12 h. A control group (120 mJ / cm²) was also included. 2 +solvent control), low concentration group (120mJ / cm³) 2 +low concentration compounds), medium concentration group (120mJ / cm) 2 +medium concentration compounds), high concentration group (120mJ / cm) 2 (+high concentration compounds);

[0244] 2) UVB modeling: Before UVB irradiation, HaCaT cells were washed three times with D'Hanks, and 50 μL of D'Hanks was added to the wells to submerge the cells. The control group was wrapped in tin foil and placed in the dark. UVB modeling was performed on the groups requiring irradiation according to the experimental groups; 3) Drug administration: Each experimental group was added to DMEM medium containing different concentrations of the compound and cultured for another 24 h.

[0245] 4) ROS fluorescent probe transfer: Dilute DCFH-DA to a final concentration of 10 μmol / L using PBS at a dilution ratio of 1:1000. Discard the culture medium in all wells except for the bare cell wells. After washing three times with PBS, add 200 μL of DCFH-DA working solution to each well and incubate in a CO2 incubator for 30 min. After incubation, wash the cells in each well three times with PBS.

[0246] 5) Fluorescence analysis. Place the 96-well plate to be tested on the detection stage of the fluorescence microplate reader, set the incident light wavelength to 525nm and the excitation light wavelength to 488nm, and read the values ​​for analysis.

[0247] Experimental results: such as Figure 20 The results show that compound I-8 can scavenge ROS in a dose-dependent manner, suggesting that compound I-8 has antioxidant effects.

[0248] As we age, human skin gradually atrophies (thinns), becomes fragile, develops poor pigmentation, and experiences delayed wound healing. Skin fragility is partly attributed to changes in hemidesmosomes and the downregulation of various collagen proteins (Collagen I, III, IV, VII) or laminins (such as LN-5) at the dermal-epidermal junction. LN-5 has been proven to be a component of anchoring fibers in the basement membrane of the skin, cornea, conjunctiva, and other tissues. LN-5 participates in cell-cell interactions through integrins and proteoglycans, playing a crucial role in cell adhesion, growth, migration, and differentiation. The appearance of wrinkles is closely related to the normal synthesis and expression of collagen and laminins. Therefore, increasing collagen levels and LN-5 can help resist wrinkle formation and play an important role in the skin aging process. Therefore, the firming and anti-wrinkle efficacy of the test compounds was evaluated by detecting the Collagen I protein level in epidermal cells (fibroblasts) using ELISA and the transcriptional levels of epidermal cells (keratinocytes) III, IV, VII, and LN-5 using Real-Time PCR. Test Examples 8-1 and 8-2 below used HaCaT immortalized human keratinocytes and HSF human skin fibroblasts to evaluate the anti-wrinkle efficacy of the compounds.

[0249] Test Example 8-1: Detection of Type I Collagen Expression in HSF Human Skin Fibroblasts

[0250] 1) Cell seeding: Seed cells into 96-well plates at a density of 2 × 10⁶ cells / well. 4 Cells / well (37℃, 5% CO2), cultured for 24 hours.

[0251] 2) Drug administration: Discard the culture medium in the 96-well plate and proceed with the drug administration procedure. Add 200 μL of culture medium containing the compound to the sample group and 200 μL of cell culture medium without the compound to the control group. After drug administration, place the 96-well plate in an incubator (37℃, 5% CO2) for 24 h ± 2 h.

[0252] 3) Type I collagen detection: After incubation, cell supernatant was collected and the content of type I collagen was determined using a human type I collagen enzyme-linked immunosorbent assay kit.

[0253] Test Example 8-2: Detection of Collagen III, IV, VII and LN-5 gene expression levels in HaCaT immortalized human keratinocytes

[0254] 1) Cell seeding: Epidermal cells were seeded at a concentration of 1×10⁻⁶. 6 Cells / well were seeded into 6-well plates and cultured in an incubator (37℃, 5% CO2) for 12 h. Control group (0% compound), low concentration group (low concentration compound), medium concentration group (medium concentration compound) and high concentration group (high concentration compound) were set up respectively.

[0255] 2) Drug administration: For each experimental group, DMEM medium containing different concentrations and compounds was added and cultured for another 24 hours;

[0256] 3) Cell collection: After culture, cell samples were collected for subsequent Collagen III, IV, VII and LN-5 expression detection.

[0257] like Figure 21~ Figure 25 The results showed that compound I-8 could increase the secretion of type I collagen, upregulate the expression of COL3A1, COL4A1, COL7A1 and LAMA5 genes in a dose-dependent manner, suggesting that compound I-8 has anti-wrinkle effects.

[0258] Test Example 9: Detection and calculation of the ROS scavenging rate of various compounds in HaCaT cells after UVB radiation

[0259] Using the experimental method in Test Example 7, the ROS clearance rate of each compound on HaCaT cells after UVB irradiation was calculated according to the following formula, and the results are listed in Table 1.

[0260]

[0261] Where S: fluorescence intensity

[0262] Table 1.

[0263] I-1(0.0025％) 5 I-14(0.0025％) 29 I-2(0.0025％) 41 I-15(0.0025％) 27 I-3(0.0025％) 6 I-16(0.0025％) 38 I-4(0.0025％) 37 I-17(0.0025％) 2 I-5(0.0025％) 31 I-18(0.0025％) 21 I-6(0.0025％) 28 I-19(0.0025％) 19 I-7(0.0025％) 40 I-20(0.0025％) 37 I-8(0.00015625％) 12 I-21(0.0025％) 8 I-8(0.000625％) 22 I-22(0.0025％) 9 I-8(0.0025％) 26 I-23(0.0025％) 34 I-9(0.0025％) 11 I-24(0.0025％) 31 I-10(0.0025％) 15 I-25(0.0025％) 29 I-11(0.0025％) 12 I-26(0.0025％) 36 I-12(0.0025％) 33 I-27(0.0025％) 35 I-13(0.0025％) 35 I-28(0.0025％) 30

[0264] Those skilled in the art will understand that the above embodiments are specific examples of implementing the present invention, and in practical applications, various changes in form and detail may be made without departing from the spirit and scope of the present invention.

Claims

1. A compound having the structure shown in general formula I, or a salt thereof, in, X is oxygen or none; R 1 is hydroxy, -NR a R b , -CH2C(O)OR 1-2 , -OC(O)R 1-3 , -OCH2OC(O) R 1-4 , halogen, cyano or -COOR 1-6 , R 1 is 1-4, said halogen is fluorine, chlorine or bromine; R 2 Hydroxyl group, C 1-6 Alkoxy, -NR a R b -CH2C(O)OR 1-2 -OC(O)R 1-3 -OCH2OC(O) R 1-4 Or cyano, R 2 The quantity is 1-4; R 1-2 R 1-3 R 1-4 and R 1-6 Each independently is hydrogen, C 1-6 Alkyl or -NR a R b ; R a and R b Independently hydrogen or C 1-6 alkyl.

2. The compound or its salt according to claim 1, characterized in that, X represents oxygen.

3. The compound or its salt according to claim 1, characterized in that, The C 1-6 The alkyl group is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, or tert-butyl; The C 1-6 The alkoxy group can be methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, or tert-butoxy.

4. The compound or its salt according to claim 1, characterized in that, R 1 Hydroxyl group, -NR a R b -CH2C(O)OR 1-2 -OC(O)R 1-3 or -OCH2OC(O) R 1-4 ;R 2 Hydroxyl group, C 1-6 Alkoxy, -NR a R b -CH2C(O)OR 1-2 -OC(O)R 1-3 -OCH2OC(O) R 1-4 .

5. The compound or its salt according to claim 1, characterized in that, R 1 hydroxyl group or ;R 2 Hydroxyl group, C 1-6 Alkoxy, or ; Among them, R 1-3 and R 1-4 Each of the following is independently methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, or .

6. The compound or its salt according to claim 1, characterized in that, R 1 hydroxyl group , , or ;R 2 The compounds are hydroxyl, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, tert-butoxy. , , or .

7. The compound or its salt according to claim 1, characterized in that, The compound is selected from any one of the following: 。 8. A composition, characterized in that, The composition comprises the compound or a salt thereof as described in any one of claims 1-7; And cosmetic-acceptable excipients.

9. Use of the compound or salt thereof as described in any one of claims 1-7, or the composition as described in claim 8, in the preparation of cosmetics.

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