Use of ectoine or its derivatives in the prevention and / or improvement of gingival damage caused by cigarette smoke and methods for achieving this use.

Abstract

This application discloses the use of ectoine or its derivatives in preventing and / or improving gingival damage caused by cigarette smoke, as well as methods for achieving this use. Ectoine or its derivatives can effectively prevent and / or improve gingival damage caused by cigarette smoke, thus expanding the scope of application of ectoine or its derivatives. Furthermore, ectoine or its derivatives can effectively prevent and / or improve gingival recession and / or gingival inflammation caused by cigarette smoke, and can be used in products such as oral care products.

Description

Technical Field

[0001] This application relates to the field of personal care products technology, and more particularly to the use of ectoine or its derivatives in preventing and / or improving gingival damage caused by cigarette smoke and methods for achieving such use. Background Technology

[0002] The gingiva is a pale pink structure closely attached to the neck of the tooth and adjacent alveolar bone, composed of stratified squamous epithelium and lamina propria. It is part of the oral mucosa, richly vascularized, pale pink, tough, and elastic. Lacking a submucosa, it is directly and tightly connected to the periosteum, thus the gingiva is immobile. Gingival recession occurs when the gingival margin recedes towards the root, exposing the tooth root. After gingival recession, the exposed root surface is prone to caries. Mechanically grinding away the thinner dentin on the root surface can easily lead to wedge-shaped defects or dentin hypersensitivity, and even long-term irritation can cause pulp congestion and degeneration. Receding interdental papillae increase the interproximal space, easily causing food impaction and plaque buildup. Gingival clefts and thickened gingival margins can also hinder plaque removal, leading to more severe inflammation and hyperplasia.

[0003] However, gingival damage caused by cigarette smoke differs from other types of gingival damage. Two main phases have been identified in cigarette smoke: a tar phase and a gas phase, a complex mixture of over 7,000 compounds. Studies have shown that cigarette smoke extract (CSE) has the ability to degrade collagen and can lead to damage to type II alveolar epithelial cells, activation of nuclear factor-κB (NF-κB), and increased secretion of tumor necrosis factor-α (TNF-α). Smoking reduces fibroblast binding and collagen and bone morphogenetic protein production, and increases the levels of tissue-destructive enzymes (MMPs). Therefore, gingival damage caused by cigarette smoke is multifaceted and complex.

[0004] Gum damage caused by cigarette smoke differs from skin damage. In oral cavities, the epithelial cells on the mucosal surface are the first line of defense against harmful environmental stimuli such as cigarette smoke. Direct contact between the gums and other tissues in the oral cavity and cigarette smoke leads to a series of oral diseases. Epidemiological studies have confirmed that smoking is a recognized risk factor for periodontal disease; in patients with periodontitis, smoking is associated with attachment loss and bone resorption. Smoking also negatively impacts the response of periodontal tissues to treatment. Research has found that cigarette smoke affects gingival blood flow, cytokine production, cell morphology, cell migration, proliferation and attachment of connective tissue cells (such as fibroblasts), as well as protein synthesis and secretion. Summary of the Invention

[0005] In order to effectively protect gums from damage caused by cigarette smoke, this application has been completed through research that ectoine or its derivatives can effectively reduce or alleviate gum damage caused by cigarette smoke.

[0006] The specific technical solution of this application is as follows:

[0007] 1. Use of ectoine or its derivatives for the prevention and / or improvement of gingival damage caused by cigarette smoke.

[0008] 2. According to the use described in item 1, the gingival injury includes gingival recession and / or gingival inflammation.

[0009] 3. According to the use described in item 1 or 2, the gingival injury includes gingival fibroblast injury.

[0010] 4. The use according to any one of items 1-3, wherein the ectoine derivative comprises hydroxyectoine, sodium ectoine, or potassium ectoine.

[0011] 5. A method for preventing and / or improving gingival damage caused by cigarette smoke, the method comprising applying ectoine or a derivative thereof into the oral cavity.

[0012] 6. The method according to item 5, wherein the gingival injury includes gingival recession and / or gingival inflammation.

[0013] 7. The method according to item 5 or 6, wherein the gingival injury includes gingival fibroblast injury.

[0014] 8. The method according to any one of items 5-7, wherein the ectoine derivative comprises hydroxyectoine, sodium ectoine, or potassium ectoine.

[0015] Invention Effects

[0016] Ectoine or its derivatives can effectively prevent and / or improve gingival damage caused by cigarette smoke, thus expanding the scope of application of ectoine or its derivatives.

[0017] Furthermore, ectoine or its derivatives can effectively prevent and / or improve gingival recession and / or gingival inflammation caused by cigarette smoke. Therefore, ectoine or its derivatives can be used in oral care products, etc. Attached Figure Description

[0018] The accompanying drawings are provided to better understand this application and do not constitute an undue limitation thereof. Wherein:

[0019] Figure 1 The relative ROS content of different groups in Example 1;

[0020] Figure 2 The relative TNF-α content of different groups in Example 2. Detailed Implementation

[0021] The following description provides exemplary embodiments of this application, including various details to aid understanding, and should be considered merely exemplary. Therefore, those skilled in the art will recognize that various changes and modifications can be made to the embodiments described herein without departing from the scope and spirit of this application. Similarly, for clarity and brevity, descriptions of well-known functions and structures are omitted in the following description.

[0022] This application provides new uses for ectoine or its derivatives, as well as methods for achieving those uses.

[0023] On the one hand, this application provides the use of ectoine or its derivatives for the prevention and / or improvement of gingival damage caused by cigarette smoke.

[0024] The gingiva refers to the pale pink structure that is closely attached to the neck of the tooth and adjacent alveolar bone. It is composed of stratified squamous epithelium and lamina propria and is part of the oral mucosa. The gingiva described in this application conforms to the general definition in the art.

[0025] Ectoine, chemically known as 2-methyl-1,4,5,6-tetrahydropyrimidine-4-carboxylic acid, is an amino acid derivative found in microorganisms and belongs to the cyclic amino acid family. Recent studies have shown that ectoine has effects such as delaying skin aging, protecting against ultraviolet rays, anti-oxidation, and moisturizing, and it is commonly used in skincare products with moisturizing and anti-aging properties.

[0026] The ectoine derivatives used in this application are not limited and can be any kind of ectoine derivative in the art, as long as they can prevent and / or alleviate oral inflammation, including but not limited to ectoine modified with chemical groups or ectoine salts. In one specific embodiment, the ectoine derivative is hydroxyectoine, sodium ectoine, or potassium ectoine.

[0027] Cigarette smoke can cause gum damage. This application, through research, has found that ectoine or its derivatives can effectively prevent and / or improve gum damage caused by cigarette smoke. Therefore, it can be used in products for preventing and / or improving gum damage caused by cigarette smoke, such as personal care products to alleviate gum damage caused by cigarette smoke. These personal care products can be oral or topical formulations. This application does not limit the specific type of formulation; those skilled in the art can choose from existing technologies according to their needs. For example, oral formulations can be powders, granules, capsules, liquid formulations, suspensions, etc., and topical formulations can be patches, sprays, creams, liquid ointments, etc.

[0028] In some embodiments, this application provides the use of ectoine or its derivatives in the preparation of products / personal care products for preventing and / or improving gingival damage caused by cigarette smoke.

[0029] This application does not limit the dosage of ectoine or its derivatives in the product / personal care product. Those skilled in the art can select the dosage based on the conventional usage dosage of ectoine or its derivatives. For example, by weight percentage, it can be 0.01% to 10%. In some embodiments, the concentration of ectoine or its derivatives is 0.01%, 0.05%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5.0%, 6.0%, 7.0%, 8.0%, 9.0%, 10.0%, etc.

[0030] In some embodiments, the personal care product includes mouthwash, toothpaste, powder, lozenges, or oral patches. This application does not impose any limitations on the preparation methods of mouthwash, toothpaste, powder, lozenges, or oral patches, as they can all be prepared using conventional methods in the art.

[0031] In some embodiments, the personal care product further includes one or more of the following: antibacterial agents, anticaries, antiallergens, antistone agents, anti-inflammatory agents, whitening agents, and moisturizers.

[0032] For example, the ectoine or its derivatives may be used in the aforementioned personal care products in conjunction with any one, two, three, or four of the following ingredients: antibacterial agents, anticaries, antiallergens, antistone agents, anti-inflammatory agents, whitening agents, and moisturizers, or in conjunction with all of the aforementioned ingredients.

[0033] In this application, no restrictions are placed on the antibacterial agent, which can be a conventional antibacterial agent in the art, such as stannous chloride, tetrahydrocurcumin, triclosan, etc.

[0034] In this application, no restrictions are placed on the anti-caries agent, which can be any commonly used anti-caries agent in the art, such as calcium phosphate, sodium trimetaphosphate, magnesium glycerophosphate, calcium lactate, etc.

[0035] In this application, no restrictions are placed on the anti-allergy agent, which can be a commonly used anti-allergy agent in the art, such as dipotassium glycyrrhizate, potassium fluoride, potassium chloride, etc.

[0036] In this application, no restrictions are placed on the anti-stone agent, which can be a commonly used anti-stone agent in the art, such as pyrophosphate, tripolyphosphate, citrate, etc.

[0037] In this application, no restrictions are placed on the anti-inflammatory agent, which can be any commonly used anti-inflammatory agent in the art, such as metronidazole, tinidazole, ornidazole, etc.

[0038] In this application, no restrictions are placed on the whitening agent, which can be a whitening agent commonly used in the art, such as peroxide bleaching agent, papain, glucose oxidase, etc.

[0039] In this application, no restrictions are placed on the use of moisturizers, which can be commonly used moisturizers in the art, such as glycerin, propylene glycol, sorbitol, xylitol, hyaluronic acid, etc.

[0040] In some embodiments, the personal care product further includes one or more of a pH adjuster, a thickener, and an osmotic pressure adjuster.

[0041] The pH adjuster can be an acid, alkali, inorganic salt, etc., available in the art; the thickener can be hydroxyethyl cellulose, carboxymethyl cellulose and its salts, xanthan gum, etc.; the osmotic pressure adjuster can be an inorganic salt, etc., available in the art.

[0042] This application also provides for the use of ectoine or its derivatives for non-therapeutic purposes in preventing and / or improving gingival damage caused by cigarette smoke.

[0043] The purpose of preventing and / or improving gingival damage caused by cigarette smoke is non-therapeutic. Non-therapeutic means that it is not for the diagnosis or treatment of disease, but can be for prevention, improvement, relief, reduction, or mitigation, etc.

[0044] This application provides a non-therapeutic method for preventing and / or improving gingival damage caused by cigarette smoke, the method comprising applying ectoine or a derivative thereof to the oral cavity. The application method may be any method permitted in the art, including but not limited to application, sublingual administration, nebulization, oral administration, injection, etc.

[0045] This application does not limit the dosage of ectoine or its derivatives administered into the cavity. Those skilled in the art can select the dosage based on the conventional dosage of ectoine or its derivatives. For example, by weight percentage, it can be 0.01% to 10%. In some embodiments, the concentration of ectoine or its derivatives is 0.01%, 0.05%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.5%, 2.0%, 2.5%, 3.0%, 3.5%, 4.0%, 4.5%, 5.0%, 6.0%, 7.0%, 8.0%, 9.0%, 10.0%, etc.

[0046] The gingival damage described in this application includes, but is not limited to, one or more of the following: gingival bleeding, gingival oxidation, gingival pain, gingival swelling, gingival recession, and gingival inflammation. In one specific embodiment, the gingival damage includes gingival recession and / or gingival inflammation.

[0047] The gingival injury described in this application includes, but is not limited to, one or more of the following: fibroblast injury, lymphocyte injury, plasma cell injury, and macrophage injury in the gingiva. In one specific embodiment, the gingival injury includes gingival fibroblast injury.

[0048] This application provides the use of ectoine or its derivatives in the preparation of products for preventing or inhibiting gingival damage caused by smoking.

[0049] In some embodiments of this application, ectoine or its derivatives are used as the sole active ingredient for the prevention or improvement of gingival damage caused by smoking / cigarette smoke.

[0050] In some embodiments of this application, ectoine or its derivatives are used as the sole active ingredient in the preparation of products / personal care products for the prevention or improvement of gingival damage caused by smoking / cigarette smoke.

[0051] In some embodiments of this application, ectoine or its derivatives are used as the sole active ingredient for the prevention and / or improvement of gingival damage caused by cigarette smoke.

[0052] In some embodiments of this application, ectoine or its derivatives are used as the sole active ingredient for non-therapeutic purposes to prevent and / or improve gingival damage caused by cigarette smoke.

[0053] This application describes the use of ectoine or its derivatives in the preparation of personal care products for the prevention and / or improvement of gingival damage caused by cigarette smoke, or for non-therapeutic purposes in the prevention and / or improvement of gingival damage caused by cigarette smoke. It demonstrates that ectoine or its derivatives can significantly inhibit the increase in reactive oxygen species (ROS) content in gingival fibroblasts after oxidative stimulation by cigarette smoke extracts. Compared to cells not treated with ectoine or its derivatives, the results show a highly significant statistical difference, indicating that ectoine or its derivatives can inhibit gingival oxidative damage induced by cigarette smoke extracts. Simultaneously, it can also inhibit the increase in tumor necrosis factor (TNF) content after stimulation by cigarette smoke extracts, again showing a highly significant statistical difference compared to cells not treated with ectoine or its derivatives. This indicates that ectoine or its derivatives can inhibit the increase in gingival inflammatory factor content induced by cigarette smoke extracts and alleviate gingival inflammation.

[0054] The novel uses of ectoine or its derivatives provided in this application, as well as the methods for achieving these uses, can improve oral health, alleviate oral discomfort caused by smoking, and prevent the formation of oral diseases, such as oral tumors, and have broad application prospects.

[0055] Example

[0056] This application provides a general and / or specific description of the materials and test methods used in the experiments. In the following examples, unless otherwise specified, % represents wt%, i.e., weight percentage. Reagents or instruments used, unless otherwise specified, are all commercially available conventional reagent products.

[0057] Experimental Example 1: Effect of Ectoin on ROS (Reactive Oxygen Species) Generation under CSE Induction

[0058] Weigh out ectoine (hereinafter referred to as Ectoine, purchased from Bloomage Biotechnology) and mix it with HGF-1 cell (gingival fibroblast) complete culture medium (containing 10% FBS and 1% penicillin and streptomycin) to prepare 100 μg / mL and 500 μg / mL Ectoine solutions respectively.

[0059] Preparation of Cigarette Smoke Extract (CSE): A gas collecting bottle was sterilized and filled with HGF-1 cell culture medium. A cigarette of a certain brand was fixed to the long end of the collecting bottle, and the short end of the tube was connected to a vacuum pump. The vacuum pump was turned on and the cigarette was lit, allowing the cigarette smoke to enter the culture medium from the long end of the tube. Cigarette smoke extract was prepared in this manner.

[0060] Test method: Prepare a DCFH-DA solution with a final concentration of 10 μM using phenol red-free medium.

[0061] HGF-1 cells were seeded at a density of 30,000 cells / well in 96-well plates and cultured at 37°C and 5% CO2. After 24 hours of cell attachment, blank, control, and experimental groups containing 100 μg / mL and 200 μg / mL ectoine were established (100 μL / well). After 24 hours of incubation, 200 μL of 5% CSE solution was added to each well (blank and control groups only received culture medium). After 3 hours of incubation, the supernatant was aspirated, and the cells were washed with DPBS. 200 μL of 10 μM DCFH-DA working solution was added, and the cells were incubated for 30 minutes, followed by washing with DPBS. Fluorescence intensity was measured at an excitation wavelength of 485 nm and an emission wavelength of 538 nm.

[0062] Blank group: No ectoine added, no CSE processing performed.

[0063] Control group: No ectoine was added, and CSE treatment was performed.

[0064] Experimental group: Add 100 or 200 μg / mL of ectoine and perform CSE treatment.

[0065] Experimental results are as follows Figure 1 As shown, according to Figure 1 Data analysis, comparing the blank group and the control group, showed that CSE significantly stimulated an increase in ROS content in cells. Simultaneously, observation of the experimental and control groups revealed that 100 μg / mL and 200 μg / mL ectoine inhibited the increase in ROS content after CSE oxidative stimulation, indicating that it can inhibit CSE-induced gingival oxidative damage, and the results were statistically significant.

[0066] The data was analyzed using Graphpad Prism statistical software. Pairwise comparisons were performed using t-tests.

[0067] *P<0.05 indicates a statistically significant difference, **P<0.01 indicates a statistically significant difference, ***P

[0068] <0.001 indicates a highly statistically significant difference.

[0069] Experimental Example 2: Effect of ectoin on TNF-α (tumor necrosis factor) production induced by CSE

[0070] Inflammation and tumors are closely related. Tumor necrosis factor-α (TNF-α) is one of the main inflammatory cytokines involved in inflammation and is highly expressed in various chronic inflammatory diseases and tumor environments. TNF-α can induce the release of other inflammatory factors, amplifying the inflammatory response, and can also directly induce epithelial cell carcinogenesis and promote tumorigenesis by releasing oxygen and nitrogen mediators. This experiment examines the effect of ectoin on TNF-α production induced by CSE to verify the influence of ectoin on CSE-induced gingival inflammation.

[0071] Ectoine (hereinafter referred to as Ectoine, purchased from Bloomage Biotechnology) was weighed and dissolved in HGF-1 complete cell culture medium (containing 10% FBS and 1% penicillin and streptomycin) to prepare 100 μg / mL and 500 μg / mL Ectoine solutions respectively.

[0072] Preparation of Cigarette Smoke Extract (CSE): A gas collecting bottle was sterilized and filled with HGF-1 cell culture medium. A cigarette of a certain brand was fixed to the long end of the collecting bottle, and the short end of the tube was connected to a vacuum pump. The vacuum pump was turned on and the cigarette was lit, allowing the cigarette smoke to enter the culture medium from the long end of the tube. Cigarette smoke extract was prepared in this manner.

[0073] Assay Method: HGF-1 cells were seeded at a density of 30,000 cells / well in 96-well plates and cultured at 37°C and 5% CO2. After 24 hours of cell attachment, blank, control, and experimental groups containing 100 and 500 μg / mL ectoine were established (100 μL / well). After 24 hours of incubation, 200 μL of 5% CSE solution was added to each well (blank and control groups only received culture medium) for 24 hours. The supernatant was collected, and the secretion of TNF-α was detected using an ELISA kit. Simultaneously with the ELISA assay, cell lysis buffer was added to the 96-well plate and centrifuged. The supernatant was used for BCA protein quantification. After homogenization, the relative TNF-α content was expressed as a percentage (%) compared to the control group.

[0074] Blank group: No ectoine added, no CSE processing performed.

[0075] Control group: No ectoine was added, and CSE treatment was performed.

[0076] Experimental group: Add 100 or 500 μg / mL of ectoine and perform CSE treatment.

[0077] Experimental results are as follows Figure 2 As shown, according to Figure 2Data analysis, comparing the blank group and the control group, showed that CSE significantly stimulated an increase in TNF-α levels in cells. Simultaneously, observation of the experimental and control groups revealed that 100 μg / mL and 500 μg / mL of Ectoine inhibited the increase in TNF-α levels after CSE stimulation, indicating that it can suppress CSE-induced gingival inflammation with statistically significant differences.

[0078] The data was analyzed using Graphpad Prism statistical software. Pairwise comparisons were performed using t-tests.

[0079] *P<0.05 indicates a statistically significant difference, **P<0.01 indicates a statistically significant difference, ***P

[0080] <0.001 indicates a highly statistically significant difference.

[0081] Although the embodiments of this application have been described above in conjunction with the specific embodiments described, this application is not limited to the specific embodiments and application fields described above. The specific embodiments described above are merely illustrative and instructive, and not restrictive. Those skilled in the art can make many other forms based on the teachings of this specification and without departing from the scope of protection of the claims of this application, and these are all within the scope of protection of this application.

Claims

1. Use of an ectoine or a derivative thereof for the manufacture of a product for preventing and / or ameliorating gingival inflammatory stimuli caused by smoking / cigarette smoke; wherein the ectoine derivative is selected from the group consisting of an ectoine sodium salt or an ectoine potassium salt.

2. Use according to claim 1, the gingival inflammatory stimuli comprising inflammatory stimuli to gingival fibroblasts.

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