TWI788019B - Use of chamaemelum nobile extract for manufacturing skincare composition - Google Patents

Abstract

A use of Chamaemelum nobileextract for manufacturing a skincare composition. The Chamaemelum nobileextract is extracted with a solvent containing water. The composition is used for anti-inflammatory, increasing the activity of superoxide dismutase, strengthening the skin barrier, reducing the formation of skin erythema, and/or improving skin moisture.

Description

Use of fruity chrysanthemum extract for preparing skin conditioning composition

This case is about the use of an extract of Chamaemelum nobile , especially the use of the extract of Chamaemelum nobile for preparing a skin conditioning composition.

The skin will age normally with age. In modern life, people are under various pressures, such as abnormal work and rest, staying up late, lack of sleep, day and night upside down, unbalanced diet and other physiological and psychological pressures that cause skin redness and the main reason of uneven skin tone, people can only cover up skin blemishes with makeup, however, the use of makeup aggravates the damage of the skin, resulting in decreased skin elasticity, insufficient water retention, roughness, dullness, enlarged pores, wrinkles, The skin has blemishes and/or an overall dull, sallow, and other aging appearance. In addition, these stresses cause metabolic disorders in the skin, making it impossible for the skin to decompose and discharge metabolic waste in time, and a large amount of waste accumulates in the cells. Accelerates the rate of skin aging, leaving the overall appearance looking listless.

In addition, people with sensitive skin usually have a thinner epidermis, which is not only prone to peeling and desquamation, but also has more obvious subcutaneous microvessels, which are often affected by the external environment and substances, such as sunlight, food, fragrance, pigment, and synthetic fibers. Stimulates the capillaries and triggers an inflammatory response.

In order to solve the above problems, those skilled in the art urgently need to develop a composition for conditioning the skin, so as to benefit a large group of people in need.

In view of this, the object of the present invention is to provide an extract of Chamaemelum nobile for preparing a composition for conditioning the skin. The extract of Chamaemelum nobile has anti-inflammation and increases the activity of superoxide dismutase , Strengthen the skin barrier, reduce skin erythema and/or increase skin moisture. The fruity chrysanthemum extract of the present invention improves skin defense, reduces oxidative stress to form free radicals, reduces skin inflammation, prevents capillaries from being stimulated to produce inflammatory reactions, calms and soothes skin, reduces skin redness, and improves skin stability. Sexuality and avoid uneven skin tone.

In addition, the fruity chrysanthemum extract of the present invention also strengthens the skin barrier to prevent water loss, keeps the skin moisturized, and provides the best protection for the skin.

The present invention provides a use of chamaemelum nobile extract for preparing a skin conditioning composition, wherein the chamaemelum nobile extract is extracted with a solvent containing water.

In some embodiments, an extract of Chamaemelum nobile is used for preparing a skin conditioning composition, wherein the chamaemelum nobile extract is extracted with an aqueous solvent.

In some embodiments, the composition is used for anti-inflammation and/or increasing the activity of superoxide dismutase.

In some embodiments, the composition works by reducing reactive oxygen species and/or reducing the expression of interleukin-8 (IL-8) gene.

In some embodiments, the composition is used to strengthen the skin barrier.

In some embodiments, the composition is used to reduce erythema production in the skin.

In some embodiments, the composition is used to increase skin hydration.

In some embodiments, the concentration of the chrysanthemum extract is at least 1% by weight.

In some embodiments, the composition further comprises a dermatologically acceptable carrier.

In some embodiments, the composition is in the form of powder, granule, liquid, jelly or paste.

In some embodiments, the composition is further prepared into a skin care product composition, a health food composition, or a cosmetic composition.

In summary, the chrysanthemum extract of any embodiment can be used to prepare a composition for conditioning the skin. The chrysanthemum extract has the functions of anti-inflammation, improving the activity of superoxide dismutase, strengthening the skin barrier, reducing skin Erythema formation and/or increase skin hydration. The fruity chrysanthemum extract of the present invention improves skin defense, reduces oxidative stress to form free radicals, reduces skin inflammation, prevents capillaries from being stimulated to produce inflammatory reactions, calms and soothes skin, reduces skin redness, and improves skin stability. Sexuality and avoid uneven skin tone. In addition, the fruity chrysanthemum extract of the present invention also strengthens the skin barrier to prevent water loss, keeps the skin moisturized, and provides the best protection for the skin.

Some specific implementation aspects of this case will be described below. Without departing from the spirit of this case, this case can still be practiced in many different forms, and the scope of protection should not be limited to the conditions specifically stated in the specification.

In this case, Excel software was used for statistical analysis. Data are expressed as mean ± standard deviation (SD), and differences among groups were analyzed by Student's t-test.

The values used in this article are approximate values, and all experimental data are expressed within the range of plus or minus 10%, and the best is within the range of plus or minus 5%.

As used herein, the term "extract" refers to a product prepared by extraction. The extract may be presented as a solution in a solvent, or the extract may be presented as a concentrate or essence free or substantially free of solvent.

The scientific name of fruity chrysanthemum is Chamaemelum nobile , also known as Roman chamomile, which grows in Europe, America and Asia. Fruity Chrysanthemum is a perennial herb of the genus Fruity Chrysanthemum in the family Asteraceae. It has a strong fragrance and can reach a height of 30 cm. The stems are erect, the leaves are alternate, sessile, pinnatifid two to three times, and the last lobes are very large. Narrow, capitulum solitary on the top of stems and long branches, heteromorphic flowers; involucral bracts with broad membranous margins, imbricate. Receptacles conical, ligulate flowers female, white, tubular flowers bisexual, yellow. Achenes without crown hairs. Fruity chrysanthemum promotes digestion, antiemetic, antispasmodic, and mildly sedative. Fructus chamomile can also be used to treat chapped skin, sore gums, rashes, inflammation and to promote wound healing.

As used herein, "fruit chrysanthemum" generally refers to the flower of a plant, wherein the flower can include raw, dried or otherwise physically processed to facilitate handling, which can further include whole, chopped, diced, milled, ground Flowers that have been otherwise processed to affect the size and physical integrity of the original material.

In some embodiments, firstly, the flowers of Chrysanthemum chrysanthemum are picked manually or by machine and rinsed with clear water for later use. The dry material of chrysanthemum chrysanthemum is extracted with water, alcohol, or a mixture of alcohol and water. The extraction solvent is preferably Water is mixed at a weight ratio of 15-75:1-5, homogenized and then extracted in a solvent at 50-100°C for 0.5-3 hours. After cooling to room temperature after extraction, the crude extract was filtered through a 400 mesh filter to obtain a filtrate. Finally, the filtrate is concentrated under reduced pressure at 45-70° C., and finally, the filtrate is spray-dried by a spray dryer, and the powder obtained is the chrysanthemum chrysanthemum extract of the present invention.

In some embodiments, the juice extracted from chrysanthemum chrysanthemum (sometimes referred to as chrysanthemum chrysanthemum extract) reduces melanin production, improves skin hydration, reduces skin wrinkles, calms and soothes skin, and reduces skin discoloration. red area. Therefore, the chrysanthemum extract can be used to prepare skin conditioning compositions for anti-inflammation, increasing the activity of superoxide dismutase, strengthening the skin barrier, reducing skin erythema and/or improving skin moisture.

In some embodiments, the composition containing the fruit extract can be a pharmaceutical composition, a skin care product composition, a food composition, or a health food composition.

The pharmaceutical composition can be formulated into a dosage form suitable for enterally, parenterally or topically administered by techniques well known to those skilled in the art. For example, it may be an injection (for example, a sterile aqueous solution or dispersion), a sterile powder, an external preparation, or the like.

The pharmaceutical composition may further contain a pharmaceutically acceptable carrier (pharmaceutically acceptable carrier) which is widely used in pharmaceutical manufacturing technology. For example, a pharmaceutically acceptable carrier may contain one or more of the following: emulsifier, suspending agent, decomposer, disintegrating agent, dispersing agent ), binder (binding agent), excipient (excipient), stabilizer (stabilizing agent), chelating agent (chelating agent), diluent (diluent), gelling agent (gelling agent), preservative (preservative), Wetting agents, lubricants, absorption delaying agents, liposomes, and the like. The selection and quantity of these carriers can be selected by those skilled in the art depending on the circumstances.

In some embodiments, the pharmaceutically acceptable carrier may comprise one of the following solvents: water, normal saline, phosphate buffered saline (PBS), aqueous solution containing alcohol (aqueous solution containing alcohol) and any other suitable solvent.

In some embodiments, the pharmaceutical composition may be administered by any of the following parenteral routes: subcutaneous injection, intraepidermal injection, intradermal injection ) and intralesional injection.

In some embodiments, the pharmaceutical composition may be formulated as an external preparation suitable for topical application to the skin using techniques well known to those skilled in the art. For example, it may be any of the following, but not limited to: emulsion (emulsion), gel (gel), ointment (ointment), cream (cream), patch (patch), liniment ( liniment), powder (powder), aerosol (aerosol), spray (spray), emulsion (lotion), serum (serum), paste (paste), foam (foam), drop (drop), suspension ( suspension), salve, and bandage.

In some embodiments, topical formulations are prepared by mixing the pharmaceutical composition with a base well known to those skilled in the art.

In some embodiments, the substrate may contain one or more of the following additives: water, alcohols, glycols, hydrocarbons (such as petroleum jelly and white Vaseline (white petrolatum,)], waxes (such as paraffin and yellow wax), preservatives (preserving agents), antioxidants (antioxidants), surfactants (surfactants), absorption enhancers ( absorption enhancers), stabilizing agents, gelling agents (such as carbopol® 974P (carbopol® 974P), microcrystalline cellulose, and carboxymethylcellulose], Active agents, humectants, odor absorbers, fragrances, pH adjusting agents, chelating agents, emulsifiers, occlusive agents (occlusive agents), emollients, thickeners, solubilizing agents, penetration enhancers, anti-irritants, colorants and propellants (propellants) etc. The selection and amounts of these additives are within the professionalism and routine skill of those skilled in the art.

In some embodiments, the skin care product may contain an acceptable adjuvant that is widely used in the manufacturing technology of skin care products. For example, the acceptable adjuvant may comprise one or more of the following adjuvants: solvents, gelling agents, active agents, preservatives, antioxidants, screening agents, chelating agents, surfactants, coloring agents agent), thickening agent, filler, fragrance and odor absorber. The selection and quantity of these reagents can be appropriately adjusted according to actual needs.

In some embodiments, the skin care product can be manufactured into a form suitable for skincare or makeup using techniques well known to those skilled in the art, which can be any of the following, but not limited thereto : Aqueous solution, water-alcohol solution or oily solution, oil-in-water type, water-in-oil type type) or compound emulsion, gel, ointment, cream, mask, patch, pack, liniment, powder, aerosol, spray, lotion, whey, paste, foam, Dispersion, drops, mousse, sunblock, sunscreen essence, sunscreen cream, sunscreen spray, sunscreen essence, sunscreen cream, sunscreen spray, tonic water, foundation ), makeup remover products (makeup remover products), soap (soap) and other body cleansing products (body cleansing products), etc.

In one embodiment of the present invention, cosmetic compositions and personal care products are provided comprising a coloring composition capable of exhibiting color change, such as a pigment grind comprising pigment particles in a carrier and styrene-maleic anhydride copolymer. The cosmetic composition may be in the form of any cosmetic or personal care product that provides color to human skin. For example, the cosmetic composition can be, but is not limited to, lipstick, lip gloss, moisturizing lipstick, nail polish, foundation, face powder, primer, concealer, blush, eye shadow, eyeliner, mascara, or bronzer. The personal care product may be in any suitable form for imparting color to human skin. For example, personal care products may include day creams or lotions, night creams or lotions, sunscreen lotions, creams or oils and other SPF products, moisturizers, salves, ointments, gels, body milks, artificial browning compositions, hair removal etc.

The cosmetic compositions and personal care products of the present invention are applied to the human dermal system, including skin, lips, nails, hair and other keratinous surfaces. The term "keratinous surface" as used herein refers to the portion of the human skin system comprising keratin, including but not limited to mammalian, preferably human, skin, lips, hair (including scalp, eyelashes, eyebrows, facial and body hair) and Nails (toenails, fingernails, nail epithelium, etc.).

Cosmetic or personal care products capable of exhibiting discoloration are applied to any area of the skin and preferably the face, neck, hands, feet or other areas of the body, such as the walls, legs and back.

In some embodiments, skin care products may also be used in combination with one or more of the following known active external use agents: whitening agents (such as tretinoin, catechin catechin, kojic acid, arbutin, and vitamin C], humectants, bactericides, ultraviolet absorbers, plant extracts (such as aloe extract) , skin nutrients, anesthetics, anti-acne agents, antipruritics, analgesics, antidermatitis agents, anti-keratosis antihyperkeratolytic agents, anti-dry skin agents, antipsoriatic agents, antiaging agents, antiwrinkle agents, antiseborrheic agents, Wound-healing agents, corticosteroids, and hormones. The selection and amount of these topical agents are within the professionalism and routine skill of those skilled in the art.

In some embodiments, the pharmaceutical composition can be used as a food additive, which can be added during the preparation of raw materials or during the production of food by conventional methods, and can be formulated with any edible material Prepared as a food product intended for consumption by humans and non-human animals.

In some embodiments, the types of food may be but not limited to: beverages, fermented foods, bakery products, health foods and dietary supplements. The food includes but is not limited to beverages, jellies, dairy products, meal replacement packs, etc.

Unless otherwise specified, the experimental procedures in the following examples are carried out at room temperature (25±5° C.) and normal pressure (1 atm).

[Example 1]: the preparation method of fruity chrysanthemum extract

In this embodiment, at first, the flowers of Chrysanthemum chrysanthemum are picked manually or by machine and rinsed with clear water for later use. The dry material of chrysanthemum chrysanthemum is extracted with water, alcohol, or a mixture of alcohol and water. The extraction solvent is preferably Mix water, dried chrysanthemum and extraction solvent at a weight ratio of 1:15, and extract in the solvent at 85°C for 1 hour after homogenization. After cooling to room temperature after extraction, the crude extract was filtered through a 400 mesh filter to obtain a filtrate. Finally, the filtrate is concentrated under reduced pressure at 60°C±5°C, and finally, the filtrate is spray-dried by a spray dryer, and the powder obtained is the chrysanthemum chrysanthemum extract of the present invention.

[Example 2]: detection of total flavonoid content of fruity chrysanthemum extract

The analytical method for detecting the content of total flavonoids is mainly based on the method described by Zhishen Jia et al., (1999), Food Chemistry, 64: 555-559 and modified. Take 200 μL of an appropriately diluted sample and add 1000 μL of H ₂ O. After mixing evenly, add 200 μL of 5% sodium citrate (Sodium nitrite), let it stand for 6 minutes, then add 200 μL of 10% aluminum nitrate (Aluminum nitrate), and let it stand for another 6 minutes. Finally, 2 mL of 4% sodium hydroxide (Sodium hydroxide) and 1.4 mL of H ₂ O were added, mixed uniformly, analyzed at 500 nm, and a standard curve was drawn with Rutin as a standard. In this embodiment, the total flavonoid content of the chrysanthemum extract of embodiment 1 is at least 3.0 mg/mL.

[Example 3]: Cell experiment - fruity chrysanthemum extract reduces active oxygen species

Herein, the degree of green fluorescence in human dermal fibroblasts CCD-966sk cells was observed with a fluorescent microscope in the dark, so as to determine the changes in the content of reactive oxygen species after being treated with the extract of chrysanthemum chrysanthemum.

Materials and Instruments 1. Cell line: Human skin fibroblast CCD-966sk (Biological Resource Conservation and Research Center (BCRC), No. 60153). 2. Medium: basal medium containing 10 vol% FBS (fetal bovine serum, purchased from Gibco). Among them, the basal medium is Eagle's minimum essential medium (Eagle's minimum essential medium (MEM), purchased from Gibco, product number 15188-319) with additional components added to make it contain 1 mM sodium pyruvate (sodium pyruvate, purchased from Gibco), 1.5 g/L sodium bicarbonate (sodium bicarbonate, purchased from Sigma) was prepared. 3. Phosphate buffered saline solution (PBS solution): purchased from Gibco, product number 10437-028. 4. DCFH-DA solution: Dissolve dichlorodihydrofluorescein diacetate (2,7-dichloro-dihydro-fluorescein diacetate, DCFH-DA; product number SI-D6883, purchased from Sigma) in dimethyl Dimethyl sulfoxide (DMSO, purchased from Sigma, product number SI-D6883-50MG) was prepared into a 5 mg/ml DCFH-DA solution. 5. Hydrogen peroxide (H ₂ O ₂ ): purchased from Sigma-Aldrich, product model 95299-1L. 6. Trypsin (Trypsin-EDTA): 10X Trypsin-EDTA (purchased from Gibco) was diluted 10 times with 1X PBS solution. 7. Inverted microscope equipped with a camera system (ZEISS, Cat. Vert.A1) 8. Chrysanthemum chrysanthemum extract: The chrysanthemum chrysanthemum extract used in this experiment was prepared in the same manner as in Example 1.

Experimental procedure

The experiment will be divided into the experimental group, the control group (the group that did not add the extract of chrysanthemum chrysanthemum, and the group that was not treated with hydrogen peroxide), and the control group (the group that did not add the extract of chrysanthemum chrysanthemum, but was treated with hydrogen peroxide) The experiment was carried out in groups, and each group carried out three repeated experiments: 1. CCD-966sk cells were inoculated in a 6-well culture plate containing 2ml of medium in each well at a rate of 1×10 ⁵ per well. 2. Place the culture plate in 5% CO ₂ at 37°C and incubate for 24 hours. 3. Remove medium. 4. Add 2 mL of assay medium to each well of the culture plate and incubate at 37°C for 1 hour. The experimental medium of the experimental group was 2 mL of cell culture medium supplemented with 5 μL of the fruity chrysanthemum extract obtained in Example 1 (that is, the volume percentage of the fruity chrysanthemum extract to the cell culture medium was 0.125%). The experimental medium of the control group was simple 2 mL cell culture medium (that is, no fruit extract). The experimental medium of the control group was a simple 2 mL cell culture medium (that is, no fruit extract). 5. Add DCFH-DA solution so that the concentration in the culture medium is 10 μg/ml, and react at 37°C for 40 minutes. 6. After DCFH-DA treatment, add H ₂ O ₂ to the experimental medium of the experimental group and the experimental medium of the control group respectively, and react at 37°C for 1 hour. Specifically, 35% wt hydrogen peroxide was first diluted to 100 mM (add 10 μL of hydrogen peroxide to 990 μL of double distilled water), and then 20 μL of 100 mM hydrogen peroxide was added to a 2 mL cell culture dish. 7. After the reaction, wash the cells twice with 1 ml PBS solution per well. 8. Stain with Hoechst 33342 (diluted 1:20000) for 5 minutes in the dark. 9. Use a fluorescent microscope to observe each group of cells in the dark, and take pictures to record the degree of green fluorescence in the cells. The relevant diagram is shown in Figure 1.

Experimental results

As shown in Figure 1, the ROS content of the control group was 1.0. From the results of the blank control group and the control group, it can be seen that after hydrogen peroxide treatment, the proportion of cells with high ROS expression (high fluorescence expression) increased significantly to 13.9 times, showing that Hydrogen peroxide does lead to intracellular production of reactive oxygen species with subsequent damage to fibroblasts. On the other hand, according to the results of the control group and the experimental group, it can be seen that when the cells were treated with the fruit extract, the proportion of cells with high ROS expression was significantly reduced to 1.4 times compared with the control group. It is shown that the extract of chrysanthemum chrysanthemum can effectively reduce the production or accumulation of active oxygen species in cells, and can be used as a scavenger of active oxygen species. In other words, the fruit extract can reduce the oxidative damage to cells caused by strong oxidizing substances such as hydrogen peroxide by reducing the content of active oxygen species in cells.

[Example 4]: Cell experiment - fruity chrysanthemum extract enhances the activity of superoxide dismutase

Human dermal fibroblasts were purchased from Taiwan Bioresource Collection and Research Center (BCRC), number BCRC 60153. The cells were cultured in a medium supplemented with 10% fetal bovine serum (FBS) (GIBCO Company, No. 10438-026, U.S.), 0.1 mM non-essential amino acids, 1.5 g/L sodium bicarbonate (Sigma Company, No. S5761, the United States), 1mM sodium pyruvate (GIBCO company, No. 11360-070, the United States) minimum essential medium (Minimum essential medium, MEM) (Eagle) (mixed with Earle's Balanced Salt Solution (Earle's Balanced Salt Solution, Earle's BSS)) (GIBCO Corporation, No. 41500-034, USA).

Inoculate 2×105 human dermal fibroblasts into each well of a 6-well culture plate containing 2 mL of the above medium, and then culture overnight at 37°C. After removing the medium, the cells were divided into 2 groups, including 1 control group and 1 experimental group. Add 0.5mg/mL chrysanthemum extract and 1mM H2O2 to the cells of the experimental group and act at 37°C for 6 hours, while the cells of the control group are not treated. Next, each group of cells was cultured at 37° C. for 24 hours. Afterwards, the medium was removed and washed once with 1X PBS (Gibco), and then trypsin was added to treat the cells for 3 minutes, followed by medium to stop trypsin activity and transferred to a 1.5 mL centrifuge tube. Next, centrifuge at 300xg for 5 minutes, then wash with PBS, and then centrifuge at 300xg for 5 minutes. Afterwards, 30 μL of 1X cell extraction buffer was added, and then the cell suspension was reacted on ice for 30 minutes, vortexed every 10 minutes to mix. Next, the broken cell suspension was centrifuged at 10,000 xg for 10 minutes at 4°C to remove insoluble material, and then 25 μL of 1X SOD buffer was added to the bottom of the active control well. After that, add 25 μL of prepared SOD standards (S1~S7, see Table 1).

Table 1 : 1X SOD solution S1 248 2 μL of SOD standard S2 100 100 S1 S3 150 100 S2 S4 100 100 S3 S5 100 100 S4 S6 150 100 S5 S7 100 100 S6

Afterwards, 25 μL of the sample was added to the bottom of the appropriate well, followed by the addition of 150 μL of Master Mix to each well. Next, 25 μL of 1X Xanthine solution was added to all wells using a multichannel pipet to initiate the reaction, and the plate was immediately transferred to a microtiter plate reader. ), and read the absorbance at 450 nm every minute for 10 minutes at room temperature.

Statistically significant differences between groups were determined by Student's t-test. The results of this example are shown in FIG. 2 .

Fig. 2 is a data graph showing the efficacy of the extract of chrysanthemum chrysanthemum of the present invention in enhancing the SOD activity of dermal fibroblasts. As can be seen from Figure 2, compared with the control group, the SOD activity of the experimental group was significantly improved (compared with the control group, the SOD activity of the experimental group increased by about 117.5%). The results of this example show that the extract of Chrysanthemum chrysanthemum of the present invention has the effect of enhancing the SOD activity of dermal fibroblasts.

[Example 5]: Efficacy Evaluation of Anti-Skin Inflammation

Here, the human CXCL8/IL8 ELISA analysis kit, ultraviolet radiation chamber and enzyme immunoassay analyzer (ELISA reader) were used to measure the anti-inflammatory effect in the cells of primary human skin keratinocytes HPEK-50 treated with chrysanthemum extract. factor changes.

Materials and Instruments 1. Cell line: Human primary epidermal keratinocytes (Human primary epidermal keratinocytes) HPEKp (CELLnTEC (Switzerland), HPEK-50). 2. Culture medium: serum-free keratinocyte culture medium (keratinocyte-SFM) (Gibco Company, USA, No. #10724-011). 3. Ultraviolet irradiation chamber (Vilber Corporation). 4. Human IL8 ELISA analysis kit: purchased from R&D systems, model D8000CD8000C, this kit contains the following reagents: (1) capture antibody (2) detection antibody (3) streptavidin Avidin-HRP (Streptavidin-HRP) (4) Washing buffer: phosphate buffered saline (PBS solution) (5) Substrate solution (tetramethyl-benzidine, TMB) (6) Termination Solution (stop solution) 5. Enzyme immunoassay analyzer (ELISA reader), purchased from BioTek Company. 6. Substrate solution (R&D systems). 7. H ₂ SO ₄ , purchased from Sigma-Aldrich Company. 8. Cell culture box, purchased from Astec Company. 9. Shaker, purchased from GenePure. 10. Fruity chrysanthemum extract: The fruity chrysanthemum extract used in this experiment was obtained as described in Example 1 above.

Detection purpose

Interleukin 8 (interleukin-8, IL-8) is known as a cytokine, which has the function of promoting inflammatory response. When an inflammatory response occurs, the inflamed tissue will release specific cytokines to attract cells with specific functions IL-8 plays an important role in the chemotaxis of neutrophils to specific tissues.

UVB in ultraviolet light has high energy, which can penetrate into the epidermis of the skin and cause sunburn, and cause skin inflammation and redness.

Experimental procedure

The experiment will be divided into three groups: 1 control group, 1 control group (UVB irradiation), and 1 experimental group (UVB irradiation and adding chrysanthemum chrysanthemum extract), and each group will carry out three repeated experiments: 1. Human primary skin keratinocytes were inoculated at 5×10 ⁴ per well and cultured in a 24-well culture plate containing 0.5ml medium per well. 2. Place the culture plate in the cell culture incubator, culture at 37°C for 24 hours, then remove the culture medium. 3. Using the ultraviolet irradiation room, with the intensity of 300mJ/ ^cm2 , the control group and the experimental group were induced to inflammatory response, and at the same time, 0.125 mg/mL fruit extract was added to the cells of the experimental group respectively, as for the control group The cells were left without any treatment (no UVB exposure, and no chrysanthemum extract). 4. After each group of cells was cultured at 37°C for 24 hours, 120 μL of cell culture supernatant was taken as a sample, and IL8 in these samples was analyzed using the human IL8 ELISA analysis kit. 5. First, dilute the capture antibody with PBS, apply the diluted capture antibody to the bottom of a 96-well microplate at an amount of 100 μL per well, and react overnight at 4°C. 6. Next, wash the 96-well microplate three times with 200 μL of washing buffer (containing 0.05% Tween 20 (Tween 20), prepared in PBS). 7. Block with 300 μL of blocking buffer (Block buffer) (1% BSA prepared in PBS), and then act at 37°C for 3 hours. 8. Afterwards, wash the 96-well microplate 3 times with 200 μL of washing buffer. 9. Add 100 μL of samples and standards (configured in diluent (0.1% BSA in PBS)), and then bind to the capture antibody at 37°C for 2 hours. 10. Next, wash the 96-well microplate 3 times with 200 μL of washing buffer. 11. Then add 100 μL of detection antibody, and then detect the capture antibody at 37°C for 2 hours. 12. Afterwards, wash the 96-well microplate 3 times with 200 μL of washing buffer. 13. Add 100 μL of streptavidin-HRP (Streptavidin-HRP), then react at room temperature for 20 minutes. 14. After that, wash the 96-well microplate 3 times with 200 μL of washing buffer. 15. Then add 100 μL of substrate solution (R&D systems) for 20 minutes at room temperature. 16. Next, add 50 μL of stop solution (stop solution) to stop the reaction, and finally measure its absorbance at 450 nm with an enzyme immunoassay analyzer. Statistical analysis was performed with Excel software.

Statistically significant differences between groups were determined by Student's t-test. The results of this example are shown in FIG. 3 .

Fig. 3 is a data graph of the anti-inflammatory effect of the chrysanthemum extract of the present invention on skin. It can be seen from Figure 3 that compared with the control group, the IL-8 production in the UVB group was significantly increased to about 115.2%, which indicates that UVB can cause an inflammatory response to human primary skin keratinocytes. Compared with the UVB group, the production of IL-8 in the experimental group decreased by 105.9%. The results of this example show that the extract of the chrysanthemum chrysanthemum of the present invention has the effect of anti-inflammation of the skin, and can increase the protective power of the skin.

In order to confirm the effect of the fruit extract on the human body, relevant human experiments are also carried out as follows.

[Example 6]: human body experiment-skin moisturizing degree detection

Samples used: facial mask containing the essence of chrysanthemum extract (made by soaking the mask cloth in the essence solution containing chrysanthemum extract as described below to make it fully absorbed) and placebo mask (made with the same The mask cloth of the model is soaked in the aforementioned essence, but the chrysanthemum extract in it is replaced with water of the same weight), and then it is fully absorbed. In this example, each facial mask contains 25mL of the corresponding essence.

Essence ingredients containing fruity chrysanthemum extract: 0.2wt% sanxian gum, 0.6wt% xinxinone, 5wt% 1,3-butanediol, 0.1wt% triethanolamine, 0.6wt% hexanediol, fruity Chrysanthemum extract 0.5wt%, the rest is water. Wherein, the chrysanthemum chrysanthemum extract is obtained by the preparation method in Example 1.

Number of subjects: 7 male and female subjects aged 20-55.

Experimental method: After the subject cleans the whole face, according to different test items, use the corresponding instrument and measurement method to record the values of the left and right half of the face before using the mask, or take a photo before using it. Then, for the instant test, the fruity chrysanthemum extract mask and the placebo mask were applied to the right half of the face and the left half of the face of the subject respectively, and they were removed after 15 minutes, and then the left half was massaged with the fingertips. Half face and right half face. After the mask is removed for about 5 minutes, use the corresponding instrument and measurement method to measure or take a photo, and then compare it with the value of your original left half face or right half face (when you want to test before and after use) , the temperature and humidity of the test area where the subject is located are consistent to reduce the impact of external temperature and humidity on the skin). For the long-acting test, the test subjects were tested before and after continuous use for 4 weeks by the aforementioned method.

The skin moisture content detection probe Corneometer ^® CM825 (C+K Multi Probe Adapter System, Germany) purchased from Courage+Khazaka electronic in Germany was used to detect the facial skin of the same subject before and after using the mask. The detection probe is based on the principle of capacitance to measure. When the water content changes, the capacitance value of the skin also changes, so the water content of the skin surface can be analyzed by measuring the skin capacitance value.

See Figure 4A. The figure shows the instant (15 minutes) moisture content of the skin of the subject before and after using the mask containing the chrysanthemum chrysanthemum extract, and the moisture content before use is taken as 100%. Among them, the water content of the subjects' skin increased to about 119.5% after use, and the extract of chrysanthemum chrysanthemum can immediately provide good moisturizing effect, improve the moisturizing degree of the skin, and make the skin no longer rough due to dryness.

See Figure 4B. The figure is the long-term (4 weeks) skin moisture content of the subject before and after using the mask containing the extract of the fruit chrysanthemum in this case, and the moisture content before the subject's use is 100%. Among them, the water content of the subjects' skin increased to about 110.7% after use, and the fruity chrysanthemum extract can provide good moisturizing effect for a long time, improve the moisturizing degree of the skin, and make the skin no longer rough due to dryness.

[Example 7]: Human experiment - skin redness detection

Using the VISIA full-face skin quality detector sold by Canfield in the United States to take pictures, use the multiple spectral imaging technology to analyze the value of the skin redness area on the face of the same subject in [Example 6] before and after using the mask. The red area (dark color) of the skin is used to detect the inflammation and sensitivity of the face. The lower the value, the lower the sensitivity of inflammation.

Please refer to Figure 5A, which is the instant (15 minutes) reddened area of the subject's skin before and after using the mask containing the fruity chrysanthemum extract of this case. The reddened area of the skin of the subject before use is 100%. Among them, the reddened area of the subjects after using it was reduced to 89.4%. After using the mask containing the fruity chrysanthemum extract of this case, the reddened area on the skin was significantly reduced immediately, which has a good skin calming effect and prevents skin inflammation and sensitivity.

Please refer to Figure 5B, which is the long-term (4 weeks) skin redness area of the subject before and after using the mask containing the fruity chrysanthemum extract of this case, and the redness area of the skin before the subject's use is 100%. Among them, the reddened area of the subjects after use was reduced to 91.0%. After long-term use of the mask containing the fruity chrysanthemum extract of this case, the reddened area on the skin is significantly reduced, and it has a good skin calming effect, preventing skin inflammation and sensitivity.

[Example 8]: Human experiment - detection of transdermal water loss

Use the skin water loss detection probe TM 300 (C+K Multi Probe Adapter System, Germany) purchased from Courage+Khazaka electronic company in Germany to test the same subject in [Example 6] who used the chrysanthemum extract essence. Facial skin was tested before and after the mask or placebo. The detection probe uses a cylindrical cavity with open ends to form a relatively stable test environment on the skin surface. By measuring the water vapor pressure gradient at two different points, the amount of water evaporated through the epidermis is calculated to measure the skin surface moisture. Churn situation.

It can be seen from Figure 6 that, compared with before using the facial mask of fruity chrysanthemum extract essence, after 4 weeks of continuous use of the facial mask of fruity chrysanthemum extract essence, the water loss rate through the epidermis was significantly improved by about 94.2%. Based on this, it can be seen that the external application of the fruit extract on the skin can indeed reduce the transepidermal water loss rate, thus improving the skin's moisturizing ability, and has the potential to improve skin diseases related to abnormal TEWL values.

In summary, the chrysanthemum extract of any embodiment can be used to prepare a composition for conditioning the skin. The chrysanthemum extract has the functions of anti-inflammation, improving the activity of superoxide dismutase, strengthening the skin barrier, reducing skin Erythema formation and/or increase skin hydration. The fruity chrysanthemum extract of the present invention improves skin defense, reduces oxidative stress to form free radicals, reduces skin inflammation, prevents capillaries from being stimulated to produce inflammatory reactions, calms and soothes skin, reduces skin redness, and improves skin stability. Sexuality and avoid uneven skin tone. In addition, the fruity chrysanthemum extract of the present invention also strengthens the skin barrier to prevent water loss, keeps the skin moisturized, and provides the best protection for the skin.

none.

Figure 1 is a comparison chart of the results of inhibiting the generation of active oxygen species in the control group, the control group and the experimental group. ### p-value < 0.001 compared to control group, ***p-value < 0.001 compared to control group. Fig. 2 is a comparison chart of the expression level of the interleukin-8 (IL-8) gene induced by the resistance to ultraviolet rays in the control group, the control group and the experimental group. ### p-value < 0.001 compared to control group, ***p-value < 0.001 compared to control group. Figure 3 is a comparison chart of the results of the wrinkled area of the skin before and after the subject uses the mask containing the essence of the experimental group and the image taken by the detector, taking the wrinkled area of the subject before use as 100%. Figures 4A and 4B are the results comparison charts of the instant (15 minutes) and long-term (4 weeks) results of the skin moisturizing degree of the subject using the mask containing the essence of the placebo and 1% fruity chrysanthemum extract, respectively. Before use, the skin moisture level of the subject was 100%. Figures 5A and 5B are the results comparison charts of the instant (15 minutes) and long-term (4 weeks) results of the erythema generation index of the facial mask containing placebo and 1% fruity chrysanthemum extract, respectively. The erythema production index of the subjects before use was 100%. Figure 6 is a long-term (4 weeks) comparison chart of the results of the transdermal water loss of the mask containing the essence of the placebo and 1% fruity chrysanthemum extract, based on the erythema formation of the previous subjects The index is 100%.

Claims (7)

A use of Chamaemelum nobile extract for preparing a composition for strengthening the skin barrier, wherein the Chamaemelum nobile extract is obtained by extracting Chamaemelum nobile flowers at 85°C for 1 hour at 85°C. The chrysanthemum flower and the solvent are mixed at a weight ratio of 1:15 for extraction. A use of Chamaemelum nobile extract for preparing a composition for reducing skin erythema, wherein the Chamaemelum nobile extract is obtained by extracting Chamaemelum nobile flowers at 85°C for 1 hour at 85°C. Fruity chrysanthemum flowers are mixed with the solvent at a weight ratio of 1:15 for extraction. A use of Chamaemelum nobile extract for preparing a composition for improving skin moisture retention, wherein the Chamaemelum nobile extract is obtained by extracting Chamaemelum nobile flowers at 85°C for 1 hour at 85°C. Fruity chrysanthemum flowers are mixed with the solvent at a weight ratio of 1:15 for extraction. The use as described in any one of claims 1 to 3, wherein the concentration of the chrysanthemum extract is at least 0.5wt%. The use according to any one of claims 1 to 3, wherein the composition further comprises a skin-acceptable carrier. The use as described in any one of claims 1 to 3, wherein the composition is in the form of powder, granule, liquid, jelly or paste. The use as described in any one of claims 1 to 3, wherein the composition is further prepared into a skin care composition, a health food composition, or a cosmetic composition.

Images