A composition for relieving skin desquamation and its application

Abstract

The present invention discloses a composition for relieving skin desquamation and its application, belonging to the field of daily chemicals. The composition includes any one of the following (1)-(4): (1) shikonin; (2) shikonin and eriodictyol; (3) shikonin and lauroside H; (4) shikonin, eriodictyol and lauroside H. By weight, the composition includes 1-125 parts of shikonin, 0-30 parts of eriodictyol and 0-30 parts of lauroside H. Through experimental verification, it is found that the complex composed of shikonin, eriodictyol and lauroside H has the effects of inhibiting the excessive proliferation of skin keratinocytes and relieving skin inflammation by inducing ferroptosis. Therefore, by adding eriodictyol and lauroside H to shikonin, the activity of shikonin can be improved, while its toxicity can be reduced, and the effect of reducing toxicity and increasing efficacy can be enhanced, thereby improving the effect of shikonin in relieving skin desquamation.

Description

Technical Field

[0001] The present invention relates to the field of daily chemical products, and particularly relates to a composition for relieving skin desquamation and its application. Background Art

[0002] The skin is the largest organ of the human body and plays a protective and barrier function. Skin diseases not only pose a major problem to human health but also affect normal social interactions. As a barrier between the human body and the environment, factors causing skin diseases include, in addition to genetic factors, stress, friction, temperature changes, light, radiation, environmental pollution, mosquito bites, food and drug reactions, unbalanced nutritional intake, and adverse skin reactions caused by other diseases. Skin desquamation is one of the most common skin diseases. Healthy skin desquamation is a phenomenon in the normal metabolic process of the skin, where dead stratum corneum cells shed from the skin surface. Some studies have observed white blood cells on the scalp surface of patients with desquamative scalp diseases through microscopy, indicating the presence of inflammation. Subsequent clinical studies have also proven that abnormal skin desquamation is often accompanied by subclinical inflammatory changes in skin cells. The causes of inflammation may be Malassezia infection, autoimmune diseases, ultraviolet radiation, or the use of poor-quality cosmetics, etc. Inflammatory reactions lead to disordered adhesion of keratinocytes and subsequent desquamation. Currently, the main treatment methods for dandruff are focused on: 1) anti-Malassezia; 2) promoting exfoliation of the stratum corneum; 3) inhibiting the growth of scalp cells. Therefore, based on these treatment methods, seeking drug components that can effectively relieve dandruff is extremely important for solving the problem of skin desquamation. Summary of the Invention

[0003] The purpose of the present invention is to provide a composition for relieving skin desquamation and its application to solve the problems existing in the above prior art. By combining shikonin, eriocitrin, and lauroside H in different combinations, it is found that when eriocitrin and lauroside H are added to shikonin, it can reduce keratinocyte hyperplasia, alleviate skin inflammation, reduce inflammatory cell infiltration from the root cause, thereby reducing the occurrence of skin desquamation, and effectively enhancing the effect of shikonin in relieving skin desquamation.

[0004] To achieve the above purpose, the present invention provides the following solutions:

[0005] The present invention provides a composition for relieving skin desquamation, which comprises any one of the following (1)-(4):

[0006] (1) Shikonin;

[0007] (2) Shikonin and eriocitrin;

[0008] (3) Shikonin and lauroside H;

[0009] (4) Shikonin, eriocitrin, and lauroside H.

[0010] Preferably, by weight, the composition comprises 1-125 parts of shikonin, 0-30 parts of eriodictyol, and 0-30 parts of osmanthuside H.

[0011] Preferably, by weight, the composition comprises 1-125 parts of shikonin, 10-30 parts of eriodictyol, and 10-30 parts of osmanthuside H.

[0012] Preferably, by weight, the composition comprises 25 parts of shikonin, 10 parts of eriodictyol, and 10 parts of osmanthuside H.

[0013] In the above technical solution, shikonin, with the English name of shikonin, CAS No.: 517-89-5, molecular weight of 288.2994, chemical name: (+)-5,8-dihydroxy-2-(1-hydroxy-4-methyl-3-pentenyl)-1,4-naphthoquinone, and the molecular formula is C 16 H 16 O5.

[0014] Eriodictyol, with the English name of (+ / -)-Eriodictyol, CAS No.: 552-58-9, molecular weight of 288.252, chemical name: 2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-2,3-dihydrochromen-4-one, and the molecular formula is C 15 H 12 O6.

[0015] Osmanthuside H, namely osmanthuside H, CAS No.: 149155-70-4, molecular weight of 432.4, chemical name: (2R,3S,4S,5R,6R)-2-[[(2R,3R,4R)-3,4-dihydroxy-4-(hydroxymethyl)oxolan-2-yl]oxymethyl]-6-[2-(4-hydroxyphenyl)ethoxy]oxane-3,4,5-triol, and the molecular formula is C 19 H 28 O 11 .

[0016] The present invention also provides a product for relieving skin desquamation, and the product comprises the above composition and a pharmaceutically acceptable carrier.

[0017] Preferably, the carrier comprises at least one of an excipient, an emulsifier, and a surfactant.

[0018] More preferably, the excipient is selected from at least one of calcium carbonate, lactose, calcium phosphate and sodium phosphate. The emulsifier is selected from at least one of egg yolk, gum arabic, tragacanth, gelatin, magnesium hydroxide, aluminum hydroxide, silicon dioxide, calcium hydroxide, zinc hydroxide, magnesium stearate, calcium hydroxide, zinc hydroxide, magnesium stearate, MC, gum arabic, cetyl alcohol, stearic acid, etc. The surfactant is selected from at least one of sodium dodecyl sulfate (SDS), stearate, oleate, span (sorbitan fatty acid esters), tween (polysorbates), poloxamer (Pluronic F68), sucrose stearate, polyoxyethylene fatty alcohol ethers (benzethonium), polyoxyethylene fatty acid esters (myrj), etc.

[0019] The present invention also provides the application of the composition in the preparation of a product for relieving skin desquamation. The composition improves skin desquamation by inhibiting the excessive proliferation of epidermal cells and relieving skin inflammation.

[0020] Preferably, the product includes drugs, beauty products and personal care products.

[0021] The present invention discloses the following technical effects:

[0022] The present invention provides a composition for relieving skin desquamation. Through experimental verification, it is found that 0.25 mg / L shikonin has the effect of inhibiting the proliferation of HaCaT cells (P<0.01) and the activity of inducing ferroptosis of cells; on this basis, adding eriodictyol and aucubin can relieve the activity of skin cell over-proliferation leading to skin desquamation, and further reduce the toxicity of shikonin and improve the drug efficacy, playing the role of reducing toxicity and enhancing efficacy. Therefore, by compounding shikonin, eriodictyol and aucubin, the present invention can improve the effect of shikonin in relieving skin desquamation, reduce inflammatory cell infiltration from the root, thereby reducing skin inflammation and reducing the occurrence of skin desquamation. Description of the Drawings

[0023] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for use in the embodiments. Obviously, the drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can be obtained based on these drawings.

[0024] Figure 1 It is the cytotoxicity experiment of HaCaT cells; A: The cytotoxicity of shikonin is clarified; B: The cytotoxicity of shikonin to HaCaT cells is relieved by the combination of eriodictyol or (and) aucubin.

[0025] Figure 2Back skin pictures of mice in each group (A), psoriasis area and severity index (PASI score) (B), and comparison of inflammatory factor concentrations in serum (C-D);

[0026] Figure 3 Activity of different treatments in alleviating skin hyperplasia in psoriasis model mice; A: Staining results; B: Statistical results of epidermal thickness data; "*" indicates p < 0.0001 compared with the normal group; "ns" indicates no statistical significance; "#" indicates compared with the imiquimod group, "#" for p < 0.01, "#" for p < 0.0001;

[0027] Figure 4 Expression of inflammatory factors and ferroptosis-related proteins in the back skin tissues of mice in different treatment groups; A: Results of Western blot experiments for the expression of inflammatory factors; B: Results of Western blot experiments for the expression of ferroptosis-related proteins; C: Statistical results of TNF expression; D: Statistical results of NCOA4 expression; E: Statistical results of GPX4 expression; "*" indicates compared with the normal group, "*" for p < 0.05, "**" for p < 0.01; "#" indicates compared with the imiquimod group, "#" for p < 0.05, "##" for p < 0.01, "" for p < 0.001, "#" for p < 0.0001. Detailed implementation manners

[0028] The various exemplary implementation manners of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, characteristics, and implementation schemes of the present invention.

[0029] It should be understood that the terms used in the present invention are only for describing specific implementation manners and are not used to limit the present invention. Additionally, for the numerical ranges in the present invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Each intermediate value within any stated value or stated range, as well as each smaller range between any other stated value or intermediate value within the stated range, is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded from the range.

[0030] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the present invention pertains. Although the present invention only describes preferred methods and materials, any methods and materials similar or equivalent to those described herein can also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials related to the documents. In case of conflict with any incorporated document, the content of this specification shall prevail.

[0031] Without departing from the scope or spirit of the present invention, various modifications and variations can be made to the specific embodiments of the present invention description, which are obvious to those skilled in the art. Other embodiments obtained from the description of the present invention are obvious to those skilled in the art. The description and examples of the present application are merely exemplary.

[0032] Regarding the terms "comprising", "including", "having", "containing", etc. used herein, they are all open-ended terms, meaning including but not limited to.

[0033] Unless otherwise specified, the "parts" described in the present invention are all by mass.

[0034] Example 1 The combination of eriocitrin, osmanthuside H and shikonin reduced the cytotoxicity of shikonin used alone

[0035] HaCaT cells are immortalized human keratinocytes and are a commonly used cell line for studying skin growth, repair, and regeneration. In this example, this cell line was used as the experimental material to compare the activities of shikonin complex and shikonin in inhibiting cell proliferation and cytotoxicity.

[0036] 1. Experimental materials

[0037] HaCaT cells were purchased from the American Type Culture Collection (ATCC).

[0038] Coconut oil and alcohol were mixed at a volume ratio of 7:3 for dissolving the following active compounds:

[0039] 1 mg / L, 5 mg / L, 25 mg / L, and 125 mg / L shikonin;

[0040] 10 mg / L, 20 mg / L, and 30 mg / L osmanthuside H;

[0041] 10 mg / L, 20 mg / L, and 30 mg / L eriocitrin.

[0042] 2. Experimental methods

[0043] Freshly cultured HaCaT cells were collected by centrifugation and seeded in 24-well plates at a concentration of 2×10 5 cfu / mL, and 1 mL of DMEM / F12 medium containing 10% fetal bovine serum was added to each well. 10 μL of shikonin solutions with concentrations of 1 mg / L, 5 mg / L, 25 mg / L, and 125 mg / L were added respectively to make the final concentrations of shikonin 0.01 mg / L, 0.05 mg / L, 0.25 mg / L, and 1.25 mg / L, and the cell viability was detected by the MTT method.

[0044] In addition, 24-well plates were seeded with fresh HaCaT cells, 10 μL of shikonin at a concentration of 25 mg / L was added, and then 10 μL of eriocitrin at concentrations of 10 mg / L, 20 mg / L, and 30 mg / L, or phyllodulcin H at concentrations of 10 mg / L, 20 mg / L, and 30 mg / L, or a mixture of 10 mg / L eriocitrin and 10 mg / L phyllodulcin H was added respectively. The cells were treated for 16 h, and cell viability was detected by the MTT method. The specific method was as follows: 20 μL of MTT solution (prepared with PBS at 5 mg / mL, pH = 7.4) was added to each well. Incubation was continued for 3 h, and then the culture was terminated. The culture supernatant in the wells was carefully aspirated. For suspension cells, centrifugation was required before aspirating the culture supernatant in the wells. 150 μL of DMSO was added to each well, and the mixture was shaken for 10 min to fully dissolve the crystals. The absorbance of each well was measured at a wavelength of 490 nm on an enzyme-linked immunosorbent assay monitor, and the results were recorded.

[0045] 3. Results and Analysis

[0046] The results are as Figure 1 shown. The cytotoxicity of shikonin on HaCaT cells was studied, and the results showed that shikonin had significant cytotoxicity on HaCaT cells when the final concentration exceeded 0.25 mg / L, and this toxicity increased with the increase in concentration ( Figure 1 A). The combination of eriocitrin or phyllodulcin H with shikonin could not relieve the cytotoxicity of shikonin. However, a mixture of 0.10 mg / L eriocitrin and 0.10 mg / L phyllodulcin H relieved the cytotoxicity of shikonin to a certain extent ( Figure 1 B).

[0047] Example 2 The combination of eriocitrin, phyllodulcin H and shikonin enhanced their active effects on desquamating mice

[0048] 1. Experimental Method

[0049] (1) Male BALB / c mice at 7 weeks of age were adaptively fed for 1 week. The mice were shaved with scissors and then depilated with depilatory cream. Imiquimod was applied to the back skin of the mice for modeling. The modeled mice were divided into five groups, namely the imiquimod group (62.5 mg imiquimod cream), the shikonin group, the combination group (a mixture of 0.10 mg / L eriocitrin and 0.10 mg / L phyllodulcin H), and the positive drug group (10 mg / kg tacrolimus cream). 0.1 mL of shikonin or shikonin combination at a concentration of 1 mg / mL was given, and local treatment was performed on the skin of the modeled mice once every 2 days. On the 5th day of treatment, the back of the mice was photographed with a digital camera at a distance of 20 cm.

[0050] (2) To evaluate the degree of skin desquamation and inflammation, on the 5th day of implementation, mouse skin tissue samples were taken. The samples were fixed in 4% paraformaldehyde for 24 h, and then dehydrated in 30% sucrose prepared with PBS for 1 day. Paraffin embedding was performed, and longitudinal sections with a thickness of 30 μm were obtained by sectioning, and then stained with HE. The tissue was incubated with 4% paraformaldehyde (PFA) solution for 5 min and washed with tap water for 5 min. The tissue was stained with hematoxylin for 3 min and rinsed with tap water for 5 min. Next, the tissue was incubated with 70% ethanol containing 1% HCl for 5 s and incubated with eosin solution for 30 s. The slides were successively placed in 95%, 100% ethanol and xylene for 3 s each. The tissue was covered with a thin coverslip.

[0051] (3) To comparatively evaluate the therapeutic activities of shikonin and the compound on psoriasis, on the 5th day of implementation, mouse serum was taken, and the contents of inflammatory factors IL-6 and TNF-α in the serum of mice in each group were detected using a specific Elisa kit.

[0052] (4) To comparatively evaluate the therapeutic activities of shikonin and the compound on psoriasis, on the 5th day of treatment, mouse skin tissue samples were taken. The cells were lysed by grinding after being frozen in liquid nitrogen and adding cell lysate with the same volume as the skin, and the cell lysate was collected into a 1.5 mL centrifuge tube. The mixture was centrifuged at 12000 rpm at low temperature for 15 min, and the supernatant was aspirated. The protein concentration was measured using a BCA protein concentration kit, and then 2× loading buffer was added and mixed well. The protein samples were fully denatured by heating in a metal bath, and then centrifuged at 12000 rpm. After 5 min, they were ready for use. The proteins in the samples were separated by 10% SDS-PAGE gel electrophoresis and transferred to an acetate cellulose membrane. After the transfer was completed, it was first blocked with 5% skim milk powder at room temperature, and then incubated overnight at 4 °C with specific antibodies of NCOA4, GPX4, and TNF-α prepared with 3% skim milk powder. After washing three times with 1×TBST solution, it was incubated with the detection antibody prepared with 5% skim milk powder at room temperature in the dark for 2 h. Finally, after washing three times with 1×TBST solution, the chromogenic solution was added and the membrane was scanned with an Odyssey fluorescence imaging system. The images were analyzed using Odessey V3.0 software and Image J software.

[0053] 2. Results and analysis:

[0054] BALB / c mice are recognized as the most suitable mice for psoriasis models, so 8-week-old male BALB / c mice were selected for the experiment. Imiquimod (IMQ) is a toll-like receptor 7 / 8 ligand activator, which is expressed by monocytes, macrophages, and plasmacytoid dendritic cells. Therefore, the production of pro-inflammatory cytokines and chemokines increases abnormally, accompanied by profound Th1 and Th17 responses. These events lead to the direct influx of immune cells into the IMQ application site, thereby inducing inflamed squamous skin lesions. In this example, through animal experiments, the effects of the shikonin group and the compound group on treating desquamation were compared. Through photographic observation and psoriasis area and severity index (PASI score) evaluation, it was found that the compound group was slightly inferior to the positive drug group (tacrolimus treatment) in relieving squamous skin lesions and reducing the psoriasis skin area, and was superior to the shikonin group( Figure 2 ). Although shikonin treatment could reduce the concentrations of two inflammatory factors, TNF-α and IL-6, in the serum, it was not statistically significant. In contrast, the inhibitory effect of the compound treatment on the concentrations of the above two inflammatory factors, although not as good as that of the positive drug, was significant( Figure 2 ).

[0055] The degree of excessive proliferation of mouse epidermal cells, and the integrity of the epidermal layer and basal layer after compound treatment were better than those of the shikonin group( Figure 3 ). At the same time, the compound treatment had significant or extremely significant effects on the expressions of the inflammatory factor TNF-α, the ferroptosis-related proteins NCOA4 and GPX4 in mouse skin, and this activity was significantly better than those of shikonin and the positive drug( Figure 4 ). It shows that shikonin and the compound in the present invention can inhibit the excessive proliferation of skin cells leading to desquamation by promoting ferroptosis, and at the same time also show the activity of inhibiting inflammation.

[0056] The above-described embodiments are only descriptions of the preferred embodiments of the present invention, and do not limit the scope of the present invention. Without departing from the design spirit of the present invention, various deformations and improvements made by those of ordinary skill in the art to the technical solutions of the present invention shall fall within the protection scope determined by the claims of the present invention.

Claims

1. A composition for relieving skin desquamation, characterized in that, By weight, the composition comprises 25 parts of shikonin, 10 parts of eriocitrin, and 10 parts of lauroside H.

2. A product for relieving skin peeling, characterized in that, The product comprises the composition according to claim 1 and a pharmaceutically acceptable carrier.

3. The product for relieving skin desquamation according to claim 2, wherein The carrier comprises at least one of excipients, emulsifiers, and surfactants.

4. Use of the composition according to claim 1 in the preparation of a product for relieving skin desquamation, characterized in that, The composition improves skin desquamation by inhibiting excessive proliferation of epidermal cells and alleviating skin inflammation.

5. The application according to claim 4, wherein The product comprises drugs, beauty products, and personal care products.

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