Application of Ectoine or derivative thereof as 11 beta-HSD1 expression inhibitor

Abstract

The invention discloses an application of ectoin or a derivative thereof as an 11beta-HSD1 expression inhibitor, so that conversion of cordinone to cortisol can be weakened, and skin problems caused by imbalance of cortisol can be prevented and / or relieved.

Description

Technical Field

[0001] The present application relates to the technical field of personal care products, and particularly to the use of ectoine or its derivatives as inhibitors of 11β-HSD1 expression. Background Art

[0002] The skin is the largest organ of the human body and the "boundary" between the body and the environment. Different from other tissues and organs, the skin is continuously exposed to various stress events and can independently "perceive", "think", "judge" and respond. When the human body senses negative emotions such as tension, anxiety, and isolation, the hypothalamic-pituitary-adrenal (HPA) axis in the human body is activated, and then other physiological systems in the body are quickly activated to perform certain physiological functions to resist stress.

[0003] Cortisol, also known as hydrocortisone, is a glucocorticoid secreted by the adrenal gland in response to the activation of the HPA axis and is regarded as a "stress hormone". It plays an important regulatory role in carbohydrate, lipid, and protein metabolism, regulates the immune response, and affects changes in mood, behavior, neuroendocrine function, body temperature, and pain perception. In addition, most of the physiological and pharmacological effects of glucocorticoids are mediated by specific receptors, which participate in the transmission of information contained in the molecule and act as transcription factors.

[0004] Normally, after the human body is stressed, cortisol is rapidly secreted within 60 minutes to increase blood sugar and heart rate to cope with stress, and generally returns to normal levels gradually after several hours. However, if the body is exposed to stress for a long time and the HPA axis is continuously activated, a series of destructive consequences will occur. For example, cortisol increases blood sugar levels through hepatic gluconeogenesis and at the same time transmits hunger signals to the brain, resulting in the accumulation of a large amount of unused glucose as visceral (especially abdominal and facial) fat because the expression levels of cortisol receptors are high in the abdomen and face. Cortisol acts on the dermal papilla cells of hair follicles, causing the hair follicle cycle to stay in the resting phase, resulting in stress-induced hair loss. For the skin, the cortisol released suddenly under stress stimulates blood vessel dilation and sebum secretion of the skin, leading to acne or eczema; the soaring adrenaline and cortisol brought about by anger directly affect the regeneration of collagen in the dermis of the skin, making expression lines permanently imprinted on the skin surface; cortisol inhibits the activity of immune cells in the skin, causing damage to the innate immune barrier and making the skin sensitive and fragile. In addition, if too much cortisol is produced or glucocorticoids at supra-physiological doses are used for a long time due to systemic diseases such as autoimmune diseases, it can lead to an increase in cortisol, and symptoms such as increased capillary fragility, skin ecchymosis / petechiae, inhibition of wound healing, inhibition of cell-mediated immune response, and skin ulcers may occur.

[0005] In addition to psychological stress, external environmental stressors such as ultraviolet and blue light radiation, environmental pollutants, and smoking can also lead to the activation of the HPA axis. Social factors such as high-intensity and high-pressure work, exams, staying up late, insomnia, and anxiety can also have the same effect. It should be noted that the definition of "stress" is anything that can increase HPA axis activity, as the stress experienced by an individual can vary greatly from person to person. Research has shown that cortisol is the main glucocorticoid in humans, and the presence of a hydroxyl group at position C11 of cortisol affects its biological activity. Due to the oxidation of this group, biologically active cortisol (hydrocortisone) is converted to inactive cortisone by 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2). The pattern is as follows Figure 13 shown Figure 13 from Reference 1. (Among them, Reference 1: Terao M, Katayama I. Local cortisol / corticosterone activation in skin physiology and pathology. J Dermatol Sci. 2016 Oct;84(1):11-16. doi: 10.1016 / j.jdermsci.2016.06.014. Epub 2016 Jun 29. PMID: 27431412.)

[0006] The enzyme plays an important role in the peripheral mechanism of cortisol production. 11β-HSD1 is expressed in the liver, adipose tissue, brain, blood vessels, gonads, and skin, and catalyzes the conversion of cortisone to cortisol (hydrocortisone). The conversion is as Figure 14 shown Figure 14 from Reference 2. (Among them, Reference 2: Kupczyk D, Bilski R, Kozakiewicz M, Studzińska R, K, Kosmalski T, Pedrycz-Wieczorska A, M. 11β-HSD as a New Target in Pharmacotherapy of Metabolic Diseases. Int J Mol Sci. 2022 Aug 11;23(16):8984. doi: 10.3390 / ijms23168984. PMID: 36012251; PMCID: PMC9409048.)

[0007] Its function is to increase the concentration of active - form glucocorticoids in peripheral tissues. In the skin, 11βHSD1 is distributed in keratinocytes, fibroblasts, and sebaceous gland cells, etc., and can activate cortisol locally in the skin, causing harm such as hyaluronic acid decomposition, epidermal barrier disruption, and loss of dermal collagen fibers. Further research shows that with the increase of age, the expression levels of 11β - HSD1 in skin cells such as keratinocytes and fibroblasts also gradually increase, further increasing the local cortisol level in the skin. Conversely, the function of 11β - HSD2 is to convert cortisol into inactive cortisone, and it appears in tissues related to mineralocorticoid action, namely the kidney, large intestine, placenta, and salivary gland. Its effect causes cortisol inactivation and prevents the activation of mineralocorticoid receptors.

[0008] The human skin is mainly composed of the epidermis, dermis, and subcutaneous tissue. As the body's first - line barrier, it can resist exogenous chemical and physical invasions, and participate in internal metabolism to maintain homeostasis.

[0009] The skin barrier consists of many structures and components, including stratum corneum - related structures, intercellular lipids, natural moisturizing factors, and dermo - epidermal junction (DEJ) structures, etc., constructing a skin barrier model that conforms to the "brick - wall structure". The outermost physical barrier comes from the lipids secreted to the skin surface after the disintegration of mature sebum cells. These lipids interact with water in the environment to form a water - lipid film with a sealing effect on the skin, keeping the skin surface moist. There is extensive cross - linking between cell membranes within the stratum corneum, forming a cornified envelope mainly composed of loricrin and involucrin, etc., which together with the stratum corneum form the "brick" structure of the skin barrier. The main components outside keratinocytes are composed of hydrophobic free ceramides, fatty acids, and cholesterol, forming the mortar in the "brick - wall structure". Natural moisturizing factors such as hyaluronic acid (HA), amino acids, pyrrolidone carboxylic acid, and uric acid, which are widely present in the epidermis - dermis, can bind a large number of water molecules and retain moisture deep in the skin. In addition, the dermo - epidermal junction (DEJ), also known as the basement membrane zone, is also an indispensable structure for the integrity of the skin barrier. They bulge upward in a papillary shape and are embedded between epidermal projections. This meshing structure, on the one hand, facilitates the dermo - epidermal connection, maintains skin firmness and prevents the formation of skin wrinkles. On the other hand, the basement membrane is a complex grid structure formed by laminin, claudin, type IV collagen, etc. buried in a gel matrix rich in viscous polysaccharides, which can increase the dermo - epidermal contact area, facilitate dermal - epidermal substance exchange, and play an osmotic and barrier function. The dermis of the skin is mainly composed of a dense extracellular matrix (ECM) rich in collagen, which provides mechanical and structural support, mainly composed of collagen, glycosaminoglycans, and elastin, and undertakes the functions of skin firmness and support.

[0010] Under various stress events, the excessively accumulated circulating cortisone in the skin is converted into biologically active cortisol by 11β-HSD1 expressed in keratinocytes, fibroblasts or sebaceous gland cells. As previously mentioned, cortisol stimulates the excessive secretion of skin oil, clogs pores, and is susceptible to bacteria, which may cause acne, folliculitis, skin laxity and other hazards. The integrity of the dermo-epidermal tight junction plays an indispensable role in preventing the formation of skin wrinkles and maintaining the epidermal barrier function. Studies have shown that long-term use of glucocorticoids may cause adverse reactions such as thinning of the dermis-epidermis, flattening of the dermo-epidermal junction, and collagen breakdown. During stress-induced skin aging, the extracellular matrix of skin cells undergoes anti-synthetic damage, mainly caused by the inhibition of collagen and hyaluronic acid production. The accumulation of stress disrupts skin homeostasis, easily triggers inflammation and an obvious appearance of fatigue, manifested as collagen reduction and skin aging, disruption of dermo-epidermal tight junction integrity and epidermal barrier dysfunction, and decline in skin moisturizing and oil control ability and imbalance of water-oil balance.

[0011] The imbalance of cortisol levels brings various hazards. In particular, stress has an adverse impact on people of all ages and is widely regarded as an inevitable part of daily life. Only by actively establishing a feedback mechanism can the body return to a normal physiological state, thereby eliminating the consequences brought about by cortisol and restoring homeostasis. However, as the main glucocorticoid in the human body, cortisol undertakes a series of important physiological functions. Therefore, how to locally weaken the skin problems caused by the excessive conversion of cortisone to cortisol in the skin remains an urgent problem to be solved. Summary of the Invention

[0012] After in-depth research, the inventors of the present application found that ectoine or its derivatives can inhibit the expression of 11β-HSD1, thereby weakening the conversion of cortisone to cortisol, preventing and / or alleviating skin problems caused by cortisol imbalance, and thus completing the present application.

[0013] The specific technical solutions of the present application are as follows:

[0014] Use of ectoine or its derivatives as an inhibitor of 11β-HSD1 expression.

[0015] Furthermore, the ectoine or its derivatives are used to reduce the production of cortisol.

[0016] Furthermore, the ectoine or its derivatives are used to reduce the production of cortisol in the skin.

[0017] Use of ectoine or its derivatives as an inhibitor of 11β-HSD1 expression in preventing and / or alleviating skin problems caused by cortisol imbalance.

[0018] Use of ectoine or its derivatives as an 11β-HSD1 expression inhibitor in preventing and / or alleviating stress-induced skin problems.

[0019] Furthermore, the skin problems include one or more of skin aging, damaged skin barrier, decreased skin moisturizing ability, decreased oil control ability, acne, skin inflammation, appearance of fatigued skin, skin ulcers, skin ecchymosis / petechiae, increased capillary fragility, and difficult healing of skin wounds.

[0020] Furthermore, the derivatives of ectoine are selected from one or more of methyl ectoine, hydroxyectoine, sodium ectoinate, potassium ectoinate, and ammonium ectoinate.

[0021] Advantages of the Invention

[0022] This application discovers that ectoine or its derivatives can act as an 11β-HSD1 expression inhibitor, thereby being used to inhibit the production of, for example, cortisol.

[0023] Furthermore, ectoine or its derivatives can prevent and / or alleviate skin problems caused by cortisol imbalance.

[0024] Furthermore, ectoine or its derivatives can prevent and / or alleviate stress-induced skin problems. Brief Description of the Drawings

[0025] The drawings are used to better understand this application and do not unduly limit this application. Among them:

[0026] Figure 1A and Figure 1B is a schematic diagram of the results of Example 1;

[0027] Figure 2A and Figure 2B is a schematic diagram of the results of Example 2;

[0028] Figure 3 is a schematic diagram of the results of Example 3;

[0029] Figure 4A and Figure 4B is a schematic diagram of the results of Example 4;

[0030] Figure 5A 、 Figure 5B 、 Figure 5C 、 Figure 5D is a schematic diagram of the results of Example 5;

[0031] Figure 6A and Figure 6B is a schematic diagram of the results of Example 6;

[0032] Figure 7A andFigure 7B Schematic diagram of the results of Example 7;

[0033] Figure 8A and Figure 8B Schematic diagram of the results of Example 8;

[0034] Figure 9A and Figure 9B Schematic diagram of the results of Example 9;

[0035] Figure 10A and Figure 10B Schematic diagram of the results of Example 10;

[0036] Figure 11A and Figure 11B Schematic diagram of the results of Comparative Example 1;

[0037] Figure 12A and Figure 12B Schematic diagram of the results of Comparative Example 2;

[0038] Figure 13 Schematic diagram of Document 1;

[0039] Figure 14 Schematic diagram of Document 2. Detailed implementation manners

[0040] The following is an illustration of exemplary embodiments of the present application, including various details of the embodiments of the present application to facilitate understanding. It should be considered merely exemplary. Therefore, those of ordinary skill in the art should recognize that various changes and modifications can be made to the embodiments described herein without departing from the scope and spirit of the present application. Similarly, for the sake of clarity and conciseness, descriptions of well-known functions and structures are omitted below.

[0041] In the present application, the term "expression inhibitor" refers to any reagent that can reduce or down-regulate the expression of a corresponding gene at the level of a certain protein. Those skilled in the art can understand that the reduction or down-regulation can be measured by conventional techniques in the art. An 11β-HSD1 expression inhibitor refers to a reagent that can reduce or down-regulate the expression of 11β-HSD1.

[0042] In the present application, the term "cortisol" refers to the naturally occurring glucocorticoid (also known as hydrocortisone) produced by the zona fasciculata of the adrenal gland.

[0043] The present application provides the use of ectoine or its derivatives as an inhibitor of 11β-HSD1 expression. Through research, it is found in the present application that ectoine or its derivatives can inhibit the expression of 11β-HSD1 and can be used as an inhibitor of 11β-HSD1 expression, and thus can be applied to personal care products for preventing and / or alleviating skin problems caused by cortisol imbalance. The personal care products can be oral preparations or topical preparations.

[0044] In some embodiments of the present application, as an inhibitor of 11β-HSD1 expression, ectoine or its derivatives can reduce the production of cortisol, for example, the production of cortisol in the skin, thereby preventing and / or alleviating skin problems caused by cortisol imbalance.

[0045] For example, when cortisol increases, it can lead to skin water-oil imbalance and damage to the skin barrier function caused by increased sebum secretion on the skin surface, reduction of natural moisturizing factors such as hyaluronic acid, and decreased expression of epidermal structural proteins (such as involucrin and loricrin); further damage to the skin barrier and skin aging caused by reduction of proteins in the dermo-epidermal junction structure (such as claudin and laminin) and decomposition of the dermal extracellular matrix (such as hyaluronic acid and collagen); and skin problems such as skin inflammation, appearance of tired skin, skin ulcers, skin ecchymosis / petechiae, increased capillary fragility, and difficult healing of skin wounds.

[0046] The present application also provides the use of ectoine or its derivatives as an inhibitor of 11β-HSD1 expression in preventing and / or alleviating skin problems caused by stress.

[0047] In the present application, the stress refers to factors that can activate the HPA axis and cause the release and secretion of glucocorticoids by the adrenal gland.

[0048] In some embodiments of the present application, the skin problems caused by cortisol imbalance or stress include one or more of skin aging, damaged skin barrier, decreased skin moisturizing ability, decreased oil control ability, acne, skin inflammation, appearance of tired skin, skin ulcers, skin ecchymosis / petechiae, increased capillary fragility, and difficult healing of skin wounds.

[0049] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of skin aging and damaged skin barrier.

[0050] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of skin aging and decreased skin moisturizing ability.

[0051] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of skin aging and decreased oil control ability.

[0052] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of damaged skin barrier and decreased skin moisturizing ability.

[0053] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of damaged skin barrier and decreased oil control ability.

[0054] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of decreased skin moisturizing ability and decreased oil control ability.

[0055] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of skin aging, damaged skin barrier and decreased skin moisturizing ability.

[0056] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of skin aging, damaged skin barrier and decreased oil control ability.

[0057] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of damaged skin barrier, decreased skin moisturizing ability and decreased oil control ability.

[0058] In some embodiments of the present application, the ectoine or its derivatives can solve the problems of skin aging, damaged skin barrier, decreased skin moisturizing ability and decreased oil control ability.

[0059] In some embodiments of the present application, the ectoine or its derivatives can also solve one, two, three, four, five, six or seven problems among acne, skin inflammation, appearance of tired skin, skin ulcer, skin ecchymosis / petechia, increased capillary fragility, and difficult healing of skin wounds. In some embodiments of the present application, the ectoine or its derivatives can solve one, two, three, four, five, six or seven problems among decreased skin collagen, decreased skin hyaluronic acid, decreased skin tight junction protein, decreased skin laminin, decreased skin corneodesmosin, decreased skin involucrin, and increased sebum secretion of sebaceous glands.

[0060] In one embodiment of the present application, the skin collagen is type I collagen. Preferably, the skin collagen is type I collagen in dermal fibroblasts.

[0061] In one embodiment of the present application, the skin collagen is the collagen of dermal fibroblasts.

[0062] In one embodiment of the present application, the skin hyaluronic acid is the hyaluronic acid of dermal fibroblasts.

[0063] In one embodiment of the present application, the skin hyaluronic acid is the hyaluronic acid in the epidermis and dermis.

[0064] In one embodiment of the present application, the skin tight junction protein is the tight junction protein of epidermal keratinocytes.

[0065] In one embodiment of the present application, the skin laminin is the laminin of epidermal keratinocytes.

[0066] In one embodiment of the present application, the skin loricrin is the loricrin of epidermal keratinocytes.

[0067] In one embodiment of the present application, the skin involucrin is the involucrin of epidermal keratinocytes.

[0068] In one embodiment of the present application, the sebum of the sebaceous gland is derived from the sebum secretion of dermal sebaceous gland cells.

[0069] In some embodiments of the present application, ectoine or its derivatives can act on the epidermis, dermis and subcutaneous tissue of the skin.

[0070] Those skilled in the art can understand that the present application may aim to achieve the above applications in the field of skin care and beauty.

[0071] The present application provides the use of ectoine or its derivatives in the preparation of an 11β-HSD1 expression inhibitor.

[0072] The present application provides the use of ectoine or its derivatives for non-therapeutic purposes in reducing the production of cortisol.

[0073] The present application provides the use of ectoine or its derivatives for non-therapeutic purposes in reducing the production of cortisol in the skin.

[0074] The present application provides the use of ectoine or its derivatives for non-therapeutic purposes in preventing and / or alleviating skin problems caused by cortisol imbalance.

[0075] The present application provides the use of ectoine or its derivatives for non-therapeutic purposes in preventing and / or alleviating skin problems caused by stress.

[0076] For the description of skin problems, see the above text.

[0077] In some embodiments of the present application, the inhibitor is a topical preparation.

[0078] In some embodiments of the present application, the derivatives of ectoine are selected from one or more of hydroxyectoine, methyl ectoine, sodium ectoinate, potassium ectoinate and ammonium ectoinate.

[0079] Example

[0080] This application generally and / or specifically describes the materials and test methods used in the experiments. In the following examples, unless otherwise specified, % represents wt%, that is, weight percentage. Reagents or instruments without indicating the manufacturer can be obtained as conventional reagent products through commercial purchase.

[0081] Example 1 Effect of ectoine on the expression of 11β-HSD1

[0082] Experimental materials: Ectoine, DMEM medium, solar simulator.

[0083] Experimental sample solution 1: Dilute the ectoine powder with DMEM medium to prepare a standby solution with 0.02 wt% ectoine.

[0084] Method: Seed the human immortalized keratinocyte cell line in a 24-well plate with cell slides at a density of 30,000 cells per well, and culture in an incubator for 72 h until the cell confluence reaches 70 - 80%. The cells are divided into three groups: untreated group, experimental group, and UVB irradiation group.

[0085] Untreated group: Aspirate the above cell medium, replace it with PBS solution, then replace the liquid in the cell wells with DMEM medium and continue culturing for 24 h.

[0086] Experimental group: Aspirate the above cell medium, replace it with PBS solution, then irradiate the well plate under the solar simulator with an irradiation dose of UVB = 5 mJ. After irradiation, replace the liquid in the cell wells with experimental sample solution 1 and continue culturing for 24 h.

[0087] UVB irradiation group: Aspirate the above cell medium, irradiate the well plate under the solar simulator with an irradiation dose of UVB = 5 mJ. After irradiation, replace the liquid in the cell wells with DMEM medium and continue culturing for 24 h.

[0088] Respectively collect the cell slides, fix the cells with ice methanol for 15 min, add anti-HSD11B1 antibody and incubate overnight at 4°C. Discard the supernatant, then add Goat-anti-mouse IgG secondary antibody and incubate at room temperature for 1.5 h. Use DAPI mounting medium to mount the slides. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then use Image J software to quantify the fluorescence intensity. Use GraphPad prism6 software to plot the graph and test the significance of the expression level of 11-β-hydroxysteroid dehydrogenase 1 between samples. *p < 0.05 indicates statistical difference. The results are as Figure 1A and Figure 1B shown.

[0089] As Figure 1A and Figure 1B shown, after UVB irradiation, the expression of 11β-HSD1 in HaCaT cells was significantly up-regulated. However, after treatment with ectoin, its expression level decreased, indicating that ectoin can significantly inhibit the expression of 11β-HSD1 on keratinocytes after UVB irradiation.

[0090] Example 2 Determination of Collagen Content

[0091] Experimental materials: Cortisone, ectoin (ectoin, purchased from Bloomage Biotechnology Co., Ltd., the same below), DMEM medium.

[0092] Experimental sample solution 2: Weigh a certain mass of cortisone and ectoin powders and dilute them with DMEM medium to prepare an experimental sample solution 2 containing both cortisone and ectoin. In experimental sample solution 2, the concentration of cortisone is 100 μM and the concentration of ectoin is 0.02 wt%.

[0093] Cortisone solution: Weigh a certain mass of cortisone powder and dilute it with DMEM medium to prepare a cortisone solution. In the cortisone solution, the concentration of cortisone is 100 μM.

[0094] Control group: Diluted DMEM medium.

[0095] Method: Seed human fibroblasts at a density of 12,000 cells / well in a 24-well plate equipped with cell slides and culture in an incubator for 24 h. Discard the supernatant, and add the cortisone solution, experimental sample solution 2 and the control group to the wells respectively, and incubate in the incubator for 72 h. Collect the cell slides, fix the cells with ice-cold methanol for 15 min, add anti-CollagenⅠ antibody and incubate overnight at 4 °C. Discard the supernatant, then add Goat-anti-rabbit IgG secondary antibody and incubate at room temperature for 1.5 h, and mount with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, as Figure 2A described, where Collagen I represents type I collagen, Nuclear represents the cell nucleus, and Merge represents the merged picture of the two.

[0096] Use Image J software to quantify the fluorescence intensity of type I collagen, use GraphPad prism6 software to plot graphs, and test the significance of the collagen expression levels among samples. *p < 0.05 indicates statistical difference. Take the fluorescence intensity of the wells in the untreated group as N 0 , and set it as 100%, as a reference for the collagen production content under normal conditions. The fluorescence intensity of the wells treated with cortisone (i.e., treated with the cortisone solution) is N, and the fluorescence intensity of the wells treated with experimental sample solution 2 is Nt, N / N0 is the relative fluorescence intensity of the cortisone treatment group, Nt / N 0 is the relative fluorescence intensity of the wells of experimental sample solution 2, as Figure 2B shown.

[0097] Among them,

[0098] "Untreated" means without any treatment with cortisone, ectoin or others;

[0099] "Cortisone" means treated with cortisone solution;

[0100] "Cortisone + Ectoin" means treated with experimental sample solution 2, the same below.

[0101] After adding cortisone to the cells, 11β-HSD1 in the cells will convert cortisone into cortisol, from Figure 2A , 2B It can be seen that the amount of collagen production after treatment with cortisone solution is significantly reduced compared with the untreated group, indicating that cortisone is converted into cortisol and then affects the expression of type I collagen, that is, stress will reduce the production of type I collagen in dermal fibroblasts; Experimental sample solution 2 added ectoin, and compared with treatment with only cortisone solution, the production amount of type I collagen was significantly increased, indicating that ectoin can improve the reduction of collagen under stress, that is, ectoin has a certain anti-stress and anti-aging activity.

[0102] Example 3

[0103] Experimental materials: Cortisone, Ectoin, DMEM medium.

[0104] Experimental sample solution 3: Weigh a certain mass of cortisone and ectoin powders and dilute them with DMEM medium to prepare experimental sample solution 3 containing both cortisone and ectoin. In experimental sample solution 3, the concentration of cortisone is 100 μM and the concentration of ectoin is 0.02 wt%.

[0105] Cortisone solution: Weigh a certain mass of cortisone and dilute it with DMEM medium to prepare cortisone solution. In the cortisone solution, the concentration of cortisone is 100 μM.

[0106] Method: After co-culturing human dermal fibroblasts with experimental sample solution 3 and cortisone solution for 1 day respectively, use a total RNA extraction kit to extract total cellular RNA, reverse transcribe it into cDNA, and use primers for RT-PCR experiments to detect the mRNA expression level of COL1A1 (type I collagen α1 chain gene). Use Graphpad prism6 software to plot graphs and test the significance of mRNA expression levels between samples.

[0107] AsFigure 3 As shown, only cortisone treatment caused a significant decrease in the mRNA of the cellular α1-1 type collagen gene; experimental sample solution 3 added ectoin, and reversed the expression of α1-1 type collagen mRNA compared to treatment with only cortisone solution, indicating that ectoin can significantly promote the de novo generation of collagen in dermal fibroblasts after cortisone treatment and has certain anti-stress anti-aging activity.

[0108] Example 4 Determination of hyaluronic acid content

[0109] Experimental materials: Cortisone, ectoin, DMEM medium.

[0110] Experimental sample solution 4: Weigh a certain mass of cortisone and ectoin powders and dilute them with DMEM medium to prepare experimental sample solution 4 containing both cortisone and ectoin. In experimental sample solution 4, cortisone is 100 μM and ectoin is 0.02 wt%.

[0111] Cortisone solution: Weigh a certain mass of cortisone and dilute it with DMEM medium to prepare a cortisone solution. In the cortisone solution, the cortisone concentration is 100 μM.

[0112] Control group: Diluted DMEM medium.

[0113] Method: Seed human fibroblasts at a density of 12,000 cells / well in a 24-well plate containing cell slides and culture in an incubator for 24 h. Discard the supernatant, add the cortisone solution, experimental sample solution 4, and the control group to the wells and incubate in the incubator for 72 h. Collect the cell slides, fix the cells with ice-cold methanol for 15 min, block them using an Avidin / Biotin blocking solution kit, add the B-HABP antibody and incubate at room temperature for 2 h. Discard the supernatant, then add the Steptavidin antibody and incubate at room temperature for 15 min, and mount the slides using a DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then use Image J software to quantify the fluorescence intensity. Use GraphPad prism6 software to plot the graph and test the significance of the hyaluronic acid expression levels between samples. *p < 0.05 indicates a statistically significant difference. Take the fluorescence intensity of the wells in the untreated group as N 0 , and set it as 100%, as a reference for the hyaluronic acid production content under normal conditions. The fluorescence intensity of the wells treated with cortisone (i.e., treated with the cortisone solution) is N, and the fluorescence intensity of the wells treated with experimental sample solution 4 is Nt. N / N 0 is the relative fluorescence intensity of the cortisone treatment group, and Nt / N 0 is the relative fluorescence intensity of the wells of experimental sample solution 4. The results are as shown in Figure 4A and Figure 4B shown.

[0114] As Figure 4A and Figure 4B shown, only triamcinolone treatment caused a significant loss of hyaluronic acid in fibroblasts; Experimental sample solution 4 increased ectoin, and restored the production of hyaluronic acid compared with treatment with only triamcinolone solution, indicating that ectoin can significantly promote the production of hyaluronic acid in dermal fibroblasts after triamcinolone treatment, and plays an important role in promoting the skin moisturizing ability under stress.

[0115] Example 5 Determination of hyaluronic acid synthase 2 content

[0116] Experimental materials: Triamcinolone, Ectoin, DMEM medium.

[0117] Experimental sample solution 5: Weigh a certain mass of triamcinolone and ectoin powders and dilute them with DMEM medium to prepare experimental sample solution 5 containing both triamcinolone and ectoin. In experimental sample solution 5, triamcinolone is 100 μM and ectoin is 0.02 wt%.

[0118] Triamcinolone solution: Weigh a certain mass of triamcinolone and dilute it with DMEM medium to prepare triamcinolone solution. In the triamcinolone solution, the concentration of triamcinolone is 100 μM.

[0119] Control group: Diluted DMEM medium.

[0120] Method: Human fibroblasts (FB) and human immortalized keratinocyte cell line (HaCaT) were seeded on 24-well plates with cell slides at a density of 12,000 / well and 40,000 / well respectively, and cultured in an incubator for 24 h. Discard the supernatant, add triamcinolone solution, experimental sample solution 5 and control group (i.e., untreated group) to the wells, and incubate in an incubator for 72 h. Collect the cell slides, fix the cells with ice-cold methanol for 15 min, add anti-HAS2 antibody and incubate overnight at 4 °C. Discard the supernatant, then add Goat-anti-mouse IgG secondary antibody and incubate at room temperature for 1.5 h, and mount with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then use Image J software to quantify the fluorescence intensity. Use GraphPad prism6 software to plot the graph and test the significance of the expression level of hyaluronic acid synthase 2 between samples. *p < 0.05 indicates statistical difference. Take the fluorescence intensity of the wells in the untreated group as N 0 , and set it as 100%, as a reference for the production content of hyaluronic acid synthase 2 under normal conditions. The fluorescence intensity of the wells treated with triamcinolone (i.e., treated with triamcinolone solution) is N, and the fluorescence intensity of the wells treated with experimental sample solution 5 is Nt. N / N 0 is the relative fluorescence intensity of the triamcinolone treatment group, Nt / N 0It is the relative fluorescence intensity of the wells of experimental sample solution 5.

[0121] As Figure 5A , Figure 5B , Figure 5C , Figure 5D shown, only treatment with dexamethasone caused a significant loss of hyaluronan synthase 2 in cells; experimental sample solution 5 increased ectoine, and restored the production of hyaluronan synthase 2 compared to treatment with only dexamethasone solution, indicating that ectoine can significantly promote the production of hyaluronan synthase 2 in epidermal keratinocytes and dermal fibroblasts after dexamethasone treatment, thereby promoting the de novo production of hyaluronic acid in the epidermis and dermis, which is beneficial to the protection of skin moisturizing ability under pressure.

[0122] Example 6 Determination of tight junction protein content

[0123] Experimental materials: Dexamethasone, Ectoine, DMEM medium.

[0124] Experimental sample solution 6: Weigh a certain mass of dexamethasone and ectoine powders and dilute them with DMEM medium to prepare experimental sample solution 6 containing both dexamethasone and ectoine. In experimental sample solution 6, dexamethasone is 100 μM and ectoine is 0.02 wt%.

[0125] Dexamethasone solution: Weigh a certain mass of dexamethasone and dilute it with DMEM medium to prepare a dexamethasone solution. In the dexamethasone solution, the concentration of dexamethasone is 100 μM.

[0126] Control group: Diluted DMEM medium.

[0127] Method: Seed human immortalized keratinocyte cell line at a density of 40,000 cells / well in a 24-well plate equipped with cell slides and culture in an incubator for 24 h. Discard the supernatant, add the dexamethasone solution, experimental sample solution 6 and the control group to the well plate and incubate in the incubator for 72 h. Collect the cell slides, fix the cells with ice-cold methanol for 15 min, add anti-claudin-1 antibody and incubate overnight at 4 °C. Discard the supernatant, then add Goat-anti-rabbit IgG secondary antibody and incubate at room temperature for 1.5 h, and mount the slides with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then use Image J software to quantify the fluorescence intensity. Use GraphPad prism6 software to plot the graph and test the significance of the tight junction protein expression levels between samples. *p < 0.05 indicates a statistically significant difference. Take the fluorescence intensity of the wells of the untreated group as N 0, and set it to 100% as a reference for the content of tight junction protein generated under normal conditions. The fluorescence intensity of the well treated with dexamethasone (i.e., treated with dexamethasone solution) is N, and the fluorescence intensity of the well treated with experimental sample solution 6 is Nt. N / N 0 is the relative fluorescence intensity of the dexamethasone treatment group, and Nt / N 0 is the relative fluorescence intensity of the well of experimental sample solution 6.

[0128] As Figure 6A and Figure 6B shown, only dexamethasone treatment caused a significant loss of tight junction protein in cells; Experimental sample solution 6 increased ectoin, and restored the production of tight junction protein compared with treatment with only dexamethasone solution, indicating that ectoin can significantly promote the production of tight junction protein in epidermal keratinocytes after dexamethasone treatment, and plays an important role in maintaining the skin barrier function under stress.

[0129] Example 7 Determination of laminin content

[0130] Experimental materials: Dexamethasone, ectoin, DMEM medium.

[0131] Experimental sample solution 7: Weigh a certain mass of dexamethasone and ectoin powder and dilute them with DMEM medium to prepare experimental sample solution 7 containing both dexamethasone and ectoin. In experimental sample solution 7, dexamethasone is 100 μM and ectoin is 0.02 wt%.

[0132] Dexamethasone solution: Weigh a certain mass of 100 mM dexamethasone and dilute it with DMEM medium to prepare a dexamethasone solution. In the dexamethasone solution, the concentration of dexamethasone is 100 μM.

[0133] Control group: Diluted DMEM medium.

[0134] Method: Seed the human immortalized keratinocyte cell line at a density of 30,000 cells per well in a 24-well plate containing cell slides, and culture it in an incubator for 24 h. Discard the supernatant, add the dexamethasone solution, experimental sample solution 7 and the control group (i.e., the untreated group) to the well plate, and incubate it in an incubator for 72 h. Collect the cell slides, fix the cells with ice-cold methanol for 15 min, add anti-laminin-5 antibody and incubate overnight at 4 °C. Discard the supernatant, then add Goat-anti-mouse IgG secondary antibody and incubate at room temperature for 1.5 h, and mount the slides with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then use Image J software to quantify the fluorescence intensity. Use GraphPad prism6 software to plot the graph and test the significance of the laminin expression level between samples. *p < 0.05 indicates a statistically significant difference. Take the fluorescence intensity of the well of the untreated group as N0 and set it to 100% as a reference for the production content of laminin 5 under normal conditions. The fluorescence intensity of the wells treated with dexamethasone (i.e., treated with dexamethasone solution) is N, and the fluorescence intensity of the wells treated with experimental sample solution 7 is Nt. N / N 0 is the relative fluorescence intensity of the dexamethasone treatment group, and Nt / N 0 is the relative fluorescence intensity of the wells of experimental sample solution 7.

[0135] As Figure 7A and Figure 7B shown, only dexamethasone treatment caused a significant loss of laminin in the cell layer; experimental sample solution 7 increased ectoine and restored the production of laminin compared with treatment with only dexamethasone solution, indicating that ectoine can significantly promote the production of laminin in epidermal keratinocytes after dexamethasone treatment and plays an important role in maintaining the skin barrier function under stress.

[0136] Example 8 Determination of Loricrin Content

[0137] Experimental materials: Dexamethasone, Ectoine, DMEM medium.

[0138] Experimental sample solution 8: Weigh a certain mass of dexamethasone and ectoine powders and dilute them with DMEM medium to prepare experimental sample solution 8 containing both dexamethasone and ectoine. In experimental sample solution 8, dexamethasone is 100 μM and ectoine is 0.02 wt%.

[0139] Dexamethasone solution: Weigh a certain mass of dexamethasone and dilute it with DMEM medium to prepare a dexamethasone solution. In the dexamethasone solution, the concentration of dexamethasone is 100 μM.

[0140] Control group: Diluted DMEM medium.

[0141] Method: Seed the human immortalized keratinocyte cell line at a density of 30,000 cells / well in a 24-well plate equipped with cell slides and culture in an incubator for 24 h. Discard the supernatant, add the dexamethasone solution, experimental sample solution 8 and the control group to the well plate and incubate in an incubator for 72 h. Collect the cell slides, fix the cells with ice-cold methanol for 15 min, add anti-loricrin antibody and incubate overnight at 4 °C. Discard the supernatant, then add Goat-anti-rabbit IgG secondary antibody and incubate at room temperature for 1.5 h, and mount the slides with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then use Image J software to quantify the fluorescence intensity. Use GraphPad prism6 software to plot graphs and test the significance of the loricrin expression levels between samples. *p < 0.05 indicates statistical difference. The fluorescence intensity of the wells of the untreated group is N 0, and set to 100%, as a reference for the production content of loricrin under normal conditions. The fluorescence intensity of the well treated with cortisone (i.e., treated with cortisone solution) is N, and the fluorescence intensity of the well treated with experimental sample solution 8 is Nt. N / N 0 is the relative fluorescence intensity of the cortisone treatment group, and Nt / N 0 is the relative fluorescence intensity of the well of experimental sample solution 8.

[0142] As Figure 8A and Figure 8B shown, only cortisone treatment caused a significant loss of loricrin in cells; experimental sample solution 8 increased ectoine, and restored the production of loricrin compared with treatment with only cortisone solution, indicating that ectoine can significantly promote the production of loricrin in epidermal keratinocytes after cortisone treatment, and plays an important role in maintaining the skin barrier function under stress.

[0143] Determination of involucrin content in Example 9

[0144] Experimental materials: Cortisone, Ectoine, DMEM medium.

[0145] Experimental sample solution 9: Weigh a certain mass of cortisone and ectoine powders and dilute them with DMEM medium to prepare experimental sample solution 9 containing both cortisone and ectoine. In experimental sample solution 9, cortisone is 100 μM and ectoine is 0.02 wt%.

[0146] Cortisone solution: Weigh a certain mass of cortisone and dilute it with DMEM medium to prepare cortisone solution. In the cortisone solution, the concentration of cortisone is 100 μM.

[0147] Control group: Diluted DMEM medium.

[0148] Method: Seed the human immortalized keratinocyte cell line at a density of 30,000 cells per well in a 24-well plate containing cell slides, and culture in an incubator for 24 h. Discard the supernatant, add cortisone solution, experimental sample solution 9 and the control group to the well plate, and incubate in an incubator for 72 h. Collect the cell slides, fix the cells with ice-cold methanol for 15 min, add anti-involucrin antibody and incubate overnight at 4 °C. Discard the supernatant, then add Goat-anti-rabbit IgG secondary antibody and incubate at room temperature for 1.5 h, and mount the slides with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then use Image J software to quantify the fluorescence intensity. Use GraphPad prism6 software to plot the graph and test the significance of the involucrin expression level between samples. *p < 0.05 indicates a statistical difference. Take the fluorescence intensity of the well of the untreated group as N 0, and set to 100%, as a reference for the content of endopin generated under normal conditions. The fluorescence intensity of the wells treated with prednisolone (i.e., treated with prednisolone solution) is N, and the fluorescence intensity of the wells treated with experimental sample solution 9 is Nt. N / N 0 is the relative fluorescence intensity of the prednisolone treatment group, and Nt / N 0 is the relative fluorescence intensity of the wells of experimental sample solution 9.

[0149] As Figure 9A and Figure 9B shown, only treatment with prednisolone caused a significant loss of endopin in cells; experimental sample solution 9 increased ectoin, and restored the production of endopin compared with treatment only with prednisolone solution, indicating that ectoin can significantly promote the production of endopin in epidermal keratinocytes after prednisolone treatment, and plays an important role in maintaining the skin barrier function under stress.

[0150] Example 10 Determination of oil content

[0151] Experimental materials: Prednisolone, Ectoin, DMEM medium.

[0152] Experimental sample solution 10: Weigh a certain mass of prednisolone and ectoin powders and dilute them with DMEM medium to prepare experimental sample solution 10 containing both prednisolone and ectoin. In experimental sample solution 10, prednisolone is 100 μM and ectoin is 0.02 wt%.

[0153] Prednisolone solution: Weigh a certain mass of prednisolone and dilute it with DMEM medium to prepare a prednisolone solution. In the prednisolone solution, the concentration of prednisolone is 100 μM.

[0154] Control group: Diluted DMEM medium.

[0155] Method: Seed human sebaceous gland cells (SZ95) at a density of 40,000 cells per well in a 24-well plate equipped with cell slides, and culture them in an incubator for 24 h. Discard the supernatant, add the prednisolone solution, experimental sample solution 10 and the control group to the well plate, and incubate in an incubator for 72 h. Collect the cell slides, add 1 μg / mL Nile red and incubate at 37 °C for 30 min, and seal the slides with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then use Image J software to quantify the fluorescence intensity. Use GraphPad prism6 software to plot graphs and test the significance of the oil expression levels between samples. *p < 0.05 indicates a statistically significant difference. Take the fluorescence intensity of the wells of the untreated group as N 0and set to 100%, as a reference for the oil production content under normal conditions. The fluorescence intensity of the wells treated with prednisolone (i.e., treated with prednisolone solution) is N, and the fluorescence intensity of the wells treated with the experimental sample solution 10 is Nt. N / N 0 is the relative fluorescence intensity of the prednisolone treatment group, and Nt / N 0 is the relative fluorescence intensity of the wells of the experimental sample solution 10.

[0156] As Figure 10A and Figure 10B shown, only prednisolone treatment significantly increased cell oil secretion; the experimental sample solution 10 increased ectoine and reduced oil production compared with treatment with only prednisolone solution, indicating that ectoine can significantly inhibit the oil secretion of dermal sebaceous gland cells after prednisolone treatment and plays an important role in maintaining skin oil regulation under stress.

[0157] Effect of ergothioneine on the expression of 11β-HSD1 in Comparative Example 1

[0158] Experimental materials: ergothioneine, DMEM medium, solar simulator.

[0159] Experimental sample solution 11: Dilute ergothioneine powder with DMEM medium to prepare a 5 μM stock solution for use.

[0160] Method: Seed the human immortalized keratinocyte cell line at a density of 30,000 cells per well in a 24-well plate containing cell slides, and culture in an incubator for 72 h until the cell confluence reaches 70-80%. These cells are used to prepare the untreated group, experimental group, and UVB irradiation group respectively.

[0161] Untreated group: Aspirate the above cell culture medium, replace it with PBS solution, and then replace the liquid in the cell wells with DMEM medium and continue to culture for 24 h.

[0162] Experimental group: Aspirate the above cell culture medium, replace it with PBS solution, then irradiate the well plate under the solar simulator with an irradiation dose of UVB = 5 mJ. After irradiation, replace the liquid in the cell wells with the experimental sample solution and continue to culture for 24 h.

[0163] UVB irradiation group: Aspirate the above cell culture medium, irradiate the well plate under the solar simulator with an irradiation dose of UVB = 5 mJ. After irradiation, replace the liquid in the cell wells with DMEM medium and continue to culture for 24 h.

[0164] Collect cell slides separately, fix the cells with ice-cold methanol for 15 min, add anti-HSD11B1 antibody and incubate overnight at 4°C. Discard the supernatant, then add Goat-anti-mouse IgG secondary antibody and incubate for 1.5 h at room temperature. Seal the slides with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope, and then quantify the fluorescence intensity using Image J software. Use GraphPad prism6 software to plot the graph and test the significance of the expression level of 11-β-hydroxysteroid dehydrogenase 1 between samples. *p<0.05 indicates statistical difference.

[0165] As Figure 11A and Figure 11B shown, the expression of 11β-HSD1 in HaCaT cells was significantly up-regulated after UVB irradiation. After treatment with ergothioneine, its expression did not change significantly, indicating that ergothioneine could not inhibit the expression of 11β-HSD1 on keratinocytes after UVB irradiation.

[0166] Comparative Example 2 Determination of Collagen Content

[0167] Experimental materials: Cortisone, ergothioneine, DMEM medium.

[0168] Experimental sample solution 12: Weigh a certain mass of cortisone and ergothioneine powders and dilute them with DMEM medium to prepare an experimental sample solution 12 containing both cortisone and ergothioneine. In experimental sample solution 12, the concentration of cortisone is 100 μM and the concentration of ergothioneine is 5 μM.

[0169] Cortisone solution: Weigh a certain mass of cortisone and dilute it with DMEM medium to prepare a cortisone solution. In the cortisone solution, the concentration of cortisone is 100 μM.

[0170] Control group: Diluted DMEM medium.

[0171] Method: Seed human fibroblasts at a density of 12,000 cells / well in a 24-well plate containing cell slides and culture in an incubator for 24 h. Discard the supernatant, and add the cortisone solution, experimental sample solution 12 and the control group to the wells respectively, and incubate in an incubator for 72 h. Collect the cell slides, fix the cells with ice-cold methanol for 15 min, add anti-CollagenⅠ antibody and incubate overnight at 4°C. Discard the supernatant, then add Goat-anti-rabbit IgG secondary antibody and incubate for 1.5 h at room temperature. Seal the slides with DAPI mounting medium. Observe and take fluorescence pictures under a Leica inverted fluorescence microscope. As Figure 10A described, where Collagen I represents type I collagen, Nuclear represents the cell nucleus, and Merge represents the merged picture of the two.

[0172] The fluorescence intensity of type I collagen was quantified using Image J software, graphs were plotted using GraphPad prism6 software, and the significance of the collagen expression levels between samples was examined. *p < 0.05 indicates a statistical difference. The fluorescence intensity of the wells in the untreated group was taken as N 0 , and set as 100%, as a reference for the collagen production content under normal conditions. The fluorescence intensity of the wells treated with cortisone (i.e., treated with cortisone solution) was N, and the fluorescence intensity of the wells treated with experimental sample solution 12 was Nt. N / N 0 was the relative fluorescence intensity of the cortisone-treated group, and Nt / N 0 was the relative fluorescence intensity of the wells of experimental sample solution 12. The results were as Figure 12B shown.

[0173] It can be seen that the amount of collagen produced after treatment with cortisone solution was significantly reduced compared to the untreated group, indicating that stress can reduce the production of type I collagen in dermal fibroblasts; experimental sample solution 12 increased ergothioneine, and there was no statistical difference in the amount of type I collagen produced compared to treatment with cortisone solution alone, indicating that ergothioneine cannot improve the reduction of collagen under stress.

[0174] From the above experiments, it can be seen that ectoine has the effect of inhibiting the expression of 11β-HSD1, and thus can regulate various skin problems caused by cortisol imbalance, such as skin aging caused by cortisol imbalance.

[0175] To further confirm the differences between skin problems caused by cortisol imbalance and those caused by other factors, the applicant further tested the effect of ergothioneine with anti-aging effects on cortisol expression. The test results showed that ergothioneine did not have the effect of inhibiting the expression of 11β-HSD1 and could not solve the skin aging caused by cortisol.

[0176] The possible reason may be as follows: After adding cortisone to the cells, 11β-HSD1 in the cells will convert cortisone into cortisol, resulting in a decrease in the expression levels of collagen, hyaluronic acid, tight junction proteins, laminin, loricrin, and involucrin in the cells, an increase in the oil secretion of the cells, and further causing a series of skin problems such as a decrease in skin collagen, a decrease in skin hyaluronic acid, a decrease in skin tight junction proteins, a decrease in skin laminin, a decrease in skin loricrin, a decrease in skin involucrin, and an increase in the oil secretion of sebaceous glands, triggering symptoms such as skin aging, a decline in skin moisturization, a breakdown of the skin barrier, and a decline in oil control ability; Since ectoine or its derivatives can inhibit the expression of 11β-HSD1 in the cells, therefore, it can alleviate the series of skin problems caused by cortisol. However, substances such as ergothioneine cannot inhibit the expression of 11β-HSD1 in the cells, so they cannot solve the series of skin problems caused by cortisol.

[0177] Although the embodiments of the present application have been described in combination above, the present application is not limited to the above specific embodiments and application fields. The above specific embodiments are merely illustrative and guiding, rather than restrictive. Those of ordinary skill in the art can also make many forms under the inspiration of this specification and without departing from the scope protected by the claims of the present application, and these all fall within the scope of protection of the present application.

Claims

1. Use of ectoine or its derivatives as an inhibitor of 11β-HSD1 expression.

2. The use according to claim 1, wherein the ectoine or its derivatives are used to reduce cortisol production.

3. The use according to claim 1 or 2, wherein the ectoine or its derivatives are used to reduce cortisol production in the skin.

4. Use of ectoine or its derivatives as an inhibitor of 11β-HSD1 expression in the prevention and / or alleviation of skin problems caused by cortisol imbalance.

5. Use of ectoine or its derivatives as an inhibitor of 11β-HSD1 expression in the prevention and / or alleviation of skin problems caused by stress.

6. The use according to claim 4 or 5, wherein the skin problems include one or more of skin aging, impaired skin barrier, decreased skin moisturizing ability, decreased oil control ability, acne, skin inflammation, appearance of tired skin, skin ulcers, skin ecchymosis / petechiae, increased capillary fragility, and difficult healing of skin wounds.

7. The use according to any one of claims 1 to 6, wherein the derivatives of ectoine are selected from one or more of methyl ectoine, hydroxyectoine, sodium ectoinate, potassium ectoinate, and ammonium ectoinate.

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