KR102472093B1 - Composition for improving skin - Google Patents

Abstract

The present invention relates to a composition for improving skin. The honeysuckle extract according to the present invention promotes the proliferation of skin stem cells and exhibits the effect of maintaining stem cell properties, so it can be used in the manufacture of cosmetics for promoting stem cell activity and medicine, cosmetics, or food for skin regeneration.

Description

Composition for improving skin {Composition for improving skin}

The present invention relates to a composition for skin improvement that exhibits skin regeneration and stem cell activity promoting effects.

Embryologically, it is known that all components of human skin are derived from ectoderm or mesoderm. Epidermis, hair follicles, sebaceous and sweat glands originate from the ectoderm, and melanocytes, nerves and special sensory receptors originate from the neuroectoderm. Depending on the developmental stage, stem cells in the embryo repeatedly differentiate to become cells with characteristics suitable for each tissue function, and even after becoming an adult, a certain number of stem cells remain in tissues. Stem cells mainly exist in two places in the skin. . The first is in the hair follicle. This area is known to play an important role in regeneration of the epidermis as a cell before cell differentiation occurs, and also plays an important role in regeneration and growth of hair. The second is the basal layer of epidermis. It has been revealed that the stem cells located here play an important role in maintaining skin health by managing not only the epidermis but also the fibroblasts in the dermal layer. Stem cells here are widely used in skin stem cell research because they have the advantage of being relatively large and easy to obtain. The skin is continuously regenerated, and stem cells present in the epithelium of the skin, ie, epidermal stem cells of the skin, are involved in epithelial recovery after injury [Non-Patent Document 1]. The epithelial stem cells of the skin are also referred to as skin stem cells, and when the skin stem cells are activated, skin wounds such as trauma can be treated or wound healing can be promoted, and wrinkle relief of the skin can also be expected. have. Integrin β1 and integrin α6 are used as indicators of skin stem cells, and it has been reported that the maintenance of skin stem cells necessary for epithelial morphogenesis and differentiation is regulated by p63 protein.

Skin stem cells play an important role in maintaining skin health and physiological and biochemical homeostasis. Stem cells present in the skin also malfunction due to the effects of aging, and as a result, various problems occur as the homeostasis of the skin is broken. Therefore, it will be possible to improve various phenomena of skin aging through the activity of these stem cells [Non-Patent Document 2].

In addition, there is an urgent need to develop a component that is safe for the body, stable in active ingredients, and above all, has a skin regeneration activity that is superior to existing substances with a skin regeneration effect.

On the other hand, honeysuckle (Lonicera japonica Thunb.) is a semi-evergreen vine that grows wild in the foothills of mountains and banks in the vicinity of houses and mountains at an altitude of 50 to 600 m in Japan, China, and Korea. ), and Indongdeung (忍冬藤) refers to the stem and branch parts.

Honeysuckle is known to be effective in antibacterial application, anti-inflammatory analgesic action, and anticancer action, and there are records in oriental medicine that it is useful for various inflammatory diseases and antipyretic.

Active ingredients of honeysuckle include flavonoids such as apigenin, loniserin, and luteolin, as well as alkaloids, luterunin, inositol, and saponin.

Despite the many human body effects of honeysuckle, there is no research on skin stem cells of honeysuckle. Accordingly, the inventors of the present invention, while researching a candidate capable of promoting the proliferation of stem cells in natural products, discovered an excellent stem cell proliferation effect such as honeysuckle, and completed the present invention.

1. Epidermal Stem Cells of the Skin, Cedric Blanpain, Elaine Fuchs, Annual Review of Cell and Developmental Biology, November 2006, Vol. 22, Pages 339-373 2. Human skin stem cells and the aging process, Catherine Niemann, Stem Cell Aging and Regenerative Medicine, November 2008, Pages 986-997

Accordingly, the inventors of the present invention completed the present invention by confirming the effect of promoting stem cell activity since the honeysuckle extract promotes the proliferation of skin stem cells in the skin to promote skin regeneration and maintain stem cell properties.

Accordingly, an object of the present invention is to provide a composition for skin regeneration comprising a honeysuckle extract as an active ingredient.

In addition, another object of the present invention is to provide a composition for promoting stem cell activity comprising a honeysuckle extract as an active ingredient.

As a means for solving the above problems, the present invention provides a composition for skin regeneration comprising a honeysuckle extract as an active ingredient for manufacturing medicine, cosmetics, or food.

As another means for solving the above problems, the present invention provides a composition for promoting stem cell activity comprising a honeysuckle extract as an active ingredient for preparing a cosmetic formulation.

The honeysuckle extract according to the present invention promotes the proliferation of skin stem cells and exhibits an effect of promoting skin regeneration, so it can be used in medicine, cosmetics, or food production.

In addition, the honeysuckle extract promotes the proliferation of skin stem cells and exhibits the effect of maintaining stem cell properties, so it can be used in the preparation of cosmetic formulations for promoting stem cell activity.

Hereinafter, the configuration of the present invention will be described in detail.

In order for skin regeneration ingredients to exert excellent effects when applied to the skin, they must be highly active at low concentrations, have excellent ability to penetrate and be absorbed through the skin, and must be volatile enough to remain for a sufficient time to exhibit skin regeneration effects. It is preferable that the active ingredient is stably maintained in the composition or on the skin, is easy to manufacture into medicines, cosmetics, or foods, and is safe for the skin. However, among known components, components satisfying all of the above characteristics are not common. For example, some skin regeneration ingredients have excellent skin regeneration activity even at low concentrations in in vitro experiments, but their ability to penetrate and absorb through the skin is poor, making it difficult to apply them to actual skin. Other active ingredients are difficult to formulate as medicines, cosmetics, or foods due to their low hydrophilicity. In addition, some skin regeneration ingredients are decomposed or transformed into other compounds when exposed to heat, light, or oxygen, so that the effect disappears before application to the skin.

As can be seen in the following examples, since the honeysuckle extract exhibits excellent skin stem cell proliferation promotion and stem cell maintenance effects at low concentrations, it can be used as an active ingredient in pharmaceutical, cosmetic, or food compositions for skin regeneration.

Accordingly, the present invention provides a composition for skin regeneration comprising a honeysuckle extract as an active ingredient.

The honeysuckle extract may be extracted by a method known in the art, and the method is not particularly limited. Alternatively, commercially available extracts may be used.

Preferably, the honeysuckle extract may use an extract obtained by extracting honeysuckle with water and/or an organic solvent, and the organic solvent may be a polar organic solvent, a nonpolar organic solvent, or a mixture thereof. The polar organic solvent may be a lower alcohol having 1 to 5 carbon atoms, ethyl acetate or acetone, and the non-polar organic solvent may be ether, chloroform, benzene, hexane or dichloromethane. For example, the lower alcohol having 1 to 5 carbon atoms may be methanol, ethanol, propanol, butanol or isopropanol.

In one embodiment, the Indongdeung extract may include a fraction obtained by fractionating the primary extract extracted using the aforementioned extraction solvent using an extraction solvent having a different polarity. For example, the Indongdeung extract may be a fraction obtained by extracting with an alcohol having 1 to 5 carbon atoms and then fractionating again with a solvent having a different polarity such as ether, benzene, or hexane. At the time of the fractionation, two or more solvents may be used, and each solvent extract may be prepared by sequentially using or mixing according to the polarity of the solvent.

In the present invention, the solvent may be removed by filtering, concentrating, or drying the extract prepared above or the fraction obtained by performing the fractionation process, and all of the filtration, concentration, and drying may be performed. Specifically, the filtration may use filter paper or a vacuum filter, the concentration may be concentrated under reduced pressure using a vacuum concentrator, etc., and the drying may be performed by lyophilization.

In addition, silica gel column chromatography, thin layer chromatography, or high performance liquid chromatography are used for the prepared extract or the fraction obtained by performing the fractionation process. Further purified fractions can be obtained by purification using various chromatographies.

The composition of the present invention can be used for the preparation of pharmaceutical formulations.

The pharmaceutical formulation may be in the form of a solution, suspension or emulsion in an oily or aqueous medium, or may be in the form of an extract, powder, granule, tablet or capsule.

In addition, the composition may further contain one or more active ingredients exhibiting the same or similar functions. For example, a known skin regeneration component may be included. When an additional skin regeneration component is included, the skin regeneration effect of the composition of the present invention can be further enhanced. When adding the above components, skin safety according to combined use, ease of formulation, and stability of active ingredients may be considered. In one embodiment of the present invention, the composition is a skin regeneration component known in the art, retinoic acid, TGF, animal placenta-derived protein, betulinic acid and chlorella extract, as an antioxidant component known in the art, tocopherol, selenium , vitamin C and phenolic compounds, whitening ingredients known in the art, substances that inhibit tyrosinase enzyme activity such as kojic acid and arbutin, hydroquinone, vitamin-C (L- Ascorbic acid) and derivatives thereof and various plant extracts may further include one or more components selected from the group consisting of. Additional components may be included in an amount of 0.0001% to 10% by weight based on the total weight of the composition, and the content range may be adjusted according to requirements such as skin safety and ease of formulation of the honeysuckle extract.

In addition, the composition of the present invention may further include a pharmaceutically acceptable carrier.

Pharmaceutically acceptable carriers may contain various ingredients such as buffer, sterile water for injection, normal saline or phosphate buffered saline, sucrose, histidine, salts and polysorbates.

The composition of the present invention can be administered orally or parenterally, and can be administered in the form of general pharmaceutical formulations, for example, in various oral and parenteral formulations for clinical administration, and when formulated, commonly used fillers and extenders , It may be prepared using diluents or excipients such as binders, wetting agents, disintegrants, and surfactants.

Solid preparations for oral administration include tablets, pills, powders, granules, capsules, etc., and these solid preparations contain at least one excipient, for example, starch, calcium carbonate, water, etc., in the pharmaceutical composition of the present invention. It may be prepared by mixing sucrose or lactose, gelatin, and the like.

In addition to simple excipients, lubricants such as magnesium styrate and talc are also used. Liquid preparations for oral use include suspensions, solutions for oral use, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweeteners, aromatics, and preservatives may be included.

Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried formulations, and suppositories. Propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate may be used as non-aqueous solvents and suspending agents. As a base for the suppository, witepsol, macrogol, tween 61, cacao butter, laurin paper, glycerogeratin and the like may be used.

In the present invention, the 'skin regeneration effect' refers to the recovery of skin tissue against damage caused by external and internal skin causes as the activity of skin stem cells is promoted. At this time, the damage caused by the external cause may include ultraviolet rays, external contaminants, wounds or trauma, and the like, and the damage caused by the internal cause may include stress.

In the present invention, 'effective amount' means the amount of the extract that promotes the regeneration of damaged skin. When the composition of the present invention contains an effective amount of the honeysuckle extract, it can provide a desirable skin regeneration effect. An effective amount of the honeysuckle extract included in the composition of the present invention will vary depending on the type of commercialization of the composition, how the compound is applied to the skin, and how long the compound stays on the skin. For example, when the composition is commercialized as a pharmaceutical formulation, the honeysuckle extract may be included in a higher concentration than when the composition is commercialized as a cosmetic product that is routinely applied to the skin. Therefore, the daily dose is 0.1 to 100 mg/kg, preferably 30 to 80 mg/kg, more preferably 50 to 60 mg/kg, based on the amount of the honeysuckle extract, 1 to 6 per day. Can be administered twice.

The composition of the present invention can be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy, and biological response modifiers.

The pharmaceutical formulation of the present invention may include a formulation for external application to the skin.

When the honeysuckle extract is used as an external skin preparation, in addition, a fatty substance, an organic solvent, a solubilizing agent, a thickening agent and a gelling agent, a softening agent, an antioxidant, a suspending agent, a stabilizer, a foaming agent, a fragrance, a surfactant, Water, ionic or nonionic emulsifiers, fillers, sequestering agents and chelating agents, preservatives, vitamins, blocking agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic actives, lipid vesicles or external preparations for skin It may contain adjuvants commonly used in the field of dermatology, such as any other ingredients used. In addition, the components may be introduced in an amount generally used in the field of skin science.

When the honeysuckle extract is provided as a formulation for external application to the skin, it may have formulations such as, but not limited to, ointments, patches, gels, creams, or sprays.

In addition, the composition for skin regeneration of the present invention can be used for preparing cosmetic formulations.

The cosmetic formulation may be in the form of a general emulsified formulation and a solubilized formulation. For example, lotion such as softening lotion or nourishing lotion, emulsion such as facial lotion, body lotion, etc., cream such as nourishing cream, moisture cream, eye cream, essence, cosmetic ointment, spray, gel, pack, sunscreen, makeup base, liquid It may have formulations such as foundation, powder, cleansing cream, cleansing lotion, makeup remover such as cleansing oil, cleanser such as cleansing foam, soap, body wash, and the like.

In addition to the honeysuckle extract, the cosmetics include a fatty substance, an organic solvent, a solubilizing agent, a thickening agent and a gelling agent, a softening agent, an antioxidant, a suspending agent, a stabilizer, a foaming agent, a fragrance, a surfactant, water, and an ion. type or nonionic emulsifiers, fillers, sequestering and chelating agents, preservatives, vitamins, blocking agents, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic actives, lipid vesicles or any commonly used in cosmetics It may contain adjuvants commonly used in the field of cosmetology, such as other ingredients.

The cosmetic formulation may contain the honeysuckle extract at a relatively high concentration in the case of a wash-off type cosmetic such as a make-up remover or cleanser in which the active ingredient stays on the skin for a short period of time. On the other hand, in the case of leave-on type cosmetics such as lotion, lotion, cream, essence, etc., in which active ingredients stay on the skin for a long time, the honeysuckle extract is contained at a lower concentration than wash-off type cosmetics. It will be fine even if you do. Although not limited thereto, in one embodiment of the present invention, the composition may include the honeysuckle extract in an amount of 0.0001% to 10% by weight (preferably 0.0001% to 1% by weight) based on the total weight of the composition. have. When the composition of the present invention contains less than 0.0001% by weight of the honeysuckle extract, a sufficient skin regeneration effect cannot be expected, and when it contains more than 10% by weight, unwanted reactions such as allergies occur or problems with skin safety occur. There may be, so this is to prevent it.

In addition, the composition for skin regeneration of the present invention can be used for preparing food formulations.

The food formulation refers to a food prepared by adding the honeysuckle extract to food materials such as beverages, teas, spices, chewing gum, and confectionery, or encapsulated, powdered, or suspended.

Since the food formulation can be consumed on a daily basis, it is very useful because a high skin regeneration effect can be expected.

When using the honeysuckle extract as a food additive, the honeysuckle extract may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be appropriately determined depending on the purpose of use (prevention, health or therapeutic treatment). In general, when preparing food or beverage, the composition of the present invention is added in an amount of 15 parts by weight or less, preferably 10 parts by weight or less, based on the raw material. However, in the case of long-term intake for the purpose of health and hygiene or health control, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount above the above range. .

There is no particular limitation on the type of food. Examples of foods to which the substance can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, There are alcoholic beverages and vitamin complexes, and includes all health foods in a conventional sense.

When the food formulation is a beverage, it may contain various flavoring agents or natural carbohydrates as additional ingredients, as in conventional beverages. The aforementioned natural carbohydrates include monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As the sweetener, natural sweeteners such as thaumatin and stevia extract, or synthetic sweeteners such as saccharin and aspartame may be used. The proportion of the natural carbohydrate is generally about 0.01 to 0.04 g, preferably about 0.02 to 0.03 g per 100 mL of the composition of the present invention.

In addition to the above, food formulations include various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonated beverages It may contain a carbonation agent used for Other food formulations may contain fruit flesh for the production of natural fruit juice, fruit juice beverages and vegetable beverages. These components may be used independently or in combination. The ratio of these additives is not very important, but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the composition of the present invention.

The present invention also provides a composition for promoting stem cell activity comprising the honeysuckle extract as an active ingredient.

In the present invention, the term 'stem cell' is a cell that can divide itself and has the ability to differentiate into a very diverse specific cell type. The type of these stem cells is not particularly limited, and in one embodiment, the stem cells may be skin stem cells. The 'skin stem cells' refer to stem cells capable of differentiating into cells constituting the skin (epidermis, dermis, and subcutaneous fat layer). Cells forming the skin include keratinocytes present in the epidermis, melanocytes present in the epidermis, and fibroblasts present in the dermis (mainly responsible for the biosynthesis of collagen and elastin).

The type of skin stem cells is not particularly limited. Skin stem cells used in the present invention can be used regardless of where they originate. For example, skin stem cells can be obtained from known sources of skin stem cells, such as hair follicles or the basal layer of the epidermis, and the animal to be collected may be a mammal. In one embodiment, a mammal can include, but is not limited to, a human, mouse, rat, guinea pig, rabbit, monkey, pig, horse, cow, sheep, antelope, dog, or cat. Preferably the mammal may be a human. Such a method for obtaining skin stem cells from a skin stem cell source is well known in the art.

In the present invention, the 'stem cell activity promoting effect' refers to the effect of promoting stem cell proliferation and/or maintaining the specific indicator molecule of stem cells and stem cell nature, which is the property of self division.

In addition, the composition of the present invention can be used for the preparation of cosmetic formulations. These cosmetic formulations are the same as those previously described in the description of cosmetic formulations containing honeysuckle extract.

Hereinafter, the present invention will be described in detail by examples. However, the following examples are only to illustrate the present invention, and the content of the present invention is not limited to the following examples.

manufacturing example One: artificial light Preparation of extract

The honeysuckle was washed, powdered, and cooled in 10-fold ethanol (w/v) for 5 days, and then the extracted stock solution was filtered to obtain a concentrated extract. A mixed solution of purified water and ethanol was added thereto to dissolve, and then filtered to prepare a final extract.

manufacturing example 2: serum free in the medium condition of skin stem cells culture

Human epidermal stem cells purchased from Cellntec were placed in a 96-well plate (5×10 ³ cells/well), and BPE (Bovine pituitary extract) similar to fetal bovine serum was added. CNT-57 medium (Cellntec) was used for 24 hours in 5% CO ₂ and cultured at 37°C. Thereafter, the culture medium was removed through the suction tube, and then the medium components were removed using a PBS solution (GibcoBRL), and the CNT-57 medium without BPE was treated with 5 μg/mL of honeysuckle extract, and 5% CO ₂ And incubated for 72 hours at 37 ℃ conditions.

Experimental example One: CCK -8 Through the evaluation method skin stem cells Proliferative effect

CCK-8 (Cell counting kit-8) evaluation was performed on the skin stem cells cultured in Preparation Example 2. CCK-8 evaluation is an analysis method that indirectly indicates the density of living cells by measuring the absorbance of formazan produced by dehydrogenase in the intracellular electron transport chain decomposing tetrazolium salt.

The cells were treated with a CCK-8 solution (1/10 of the volume of the medium) at 37° C. for 2 hours, and then the absorbance at 450 nm was measured spectrophotometrically. For the proliferation promoting effect of skin stem cells of Indongdeung extract, the proliferation rate (%) was calculated according to Equation 1 below compared to the control group (CNT-57 medium without the extract) from the measured absorbance value, and the value is shown in Table 1 below. showed up

[Equation 1]

Proliferation rate (%) = (absorbance of sample treatment group / absorbance of control group) × 100

Skin stem cell proliferation promotion effect (number of repetitions = 3) Additive sample density Cell proliferation rate (%) compared to the control group honeysuckle extract 5 μg/mL 127% control group - 100%

As shown in Table 1, the skin stem cells in the medium conditions treated with the honeysuckle extract showed excellent proliferation promoting effects.

Experimental example 2: of skin stem cells stem cell nature ( stemness ) check the retention effect

In order to confirm the effect of maintaining stem cell nature of the skin stem cells cultured in Preparation Example 2, the expression level of p63 was evaluated. As an intracellular transcription factor, p63 is well known as a maintenance marker for skin stem cells. Cellular RNA was isolated to synthesize cDNA, and real-time PCR was performed using Taqman staining solution to measure the level of p63 expression. The concentration of RNA in this experiment was normalized to S16 ribosomal RNA.

The experimental results are shown in Table 2 below. In Table 2 below, the value of p63 increase fold means the fold compared to the control group (CNT-57 medium supplemented with BPE).

Stemness maintenance effect (repeat number = 3) Additive sample density p63 increase fold (fold) honeysuckle extract 5 μg/mL 2.17 control group - 1.0

As shown in Table 2, the medium condition treated with the honeysuckle extract promotes the expression of p63 in skin stem cells, resulting in an excellent stem cell maintenance effect.

formulation example 1: Preparation of pharmaceutical preparations

1. Preparation of tablets

Indongdeung Extract 0.2mg

Corn Starch 100mg

lactose 100mg

Magnesium Stearate 2mg

After mixing the above ingredients, tablets were prepared by tableting according to a conventional tablet manufacturing method.

formulation example 2: Manufacturing of cosmetics

1. Manufacture of nutritional cream

As shown in the following composition, a nutritious cream containing the honeysuckle extract as an active ingredient was prepared according to a conventional method.

Indongdeung extract 0.2% by weight

Beta-1,3-glucan 5.0% by weight

10.0% by weight of beeswax

Polysorbate 60 1.5% by weight

PEG 60 hydrogenated castor oil 2.0% by weight

Sorbitan sesquioleate 0.5% by weight

10.0% by weight of liquid paraffin

Squalane 5.0% by weight

Caprylic/capric triglyceride 5.0% by weight

Glycerin 5.0% by weight

Butylene glycol 3.0% by weight

Propylene glycol 3.0% by weight

Triethanolamine 0.2% by weight

Preservative 0.05% by weight

Pigment 0.05% by weight

Perfume 0.05% by weight

Purified water to 100% by weight

formulation example 3: Preparation of topical skin preparation

1. Preparation of ointment

As shown in the following composition, an ointment containing the honeysuckle extract as an active ingredient was prepared according to a conventional method.

Indongdeung extract 0.5% by weight

Beta-1,3-glucan 10.0% by weight

10.0% by weight of beeswax

Polysorbate 60 5.0% by weight

PEG 60 hydrogenated castor oil 2.0% by weight

Sorbitan sesquioleate 0.5% by weight

Vaseline 5.0% by weight

10.0% by weight of liquid paraffin

Squalane 5.0% by weight

Shea butter 3.0% by weight

Caprylic/capric triglyceride 5.0% by weight

Glycerin 10.0% by weight

Propylene glycol 10.2% by weight

Triethanolamine 0.2% by weight

Preservative 0.05% by weight

Pigment 0.05% by weight

Perfume 0.05% by weight

Purified water to 100% by weight

Claims (7)

A composition for skin regeneration comprising a honeysuckle extract as an active ingredient.
According to claim 1,
Indongdeung extract is a skin regeneration composition obtained by extracting indongdeung with at least one selected from the group consisting of water and organic solvents.
According to claim 2,
The organic solvent is a polar solvent including a lower alcohol having 1 to 5 carbon atoms, ethyl acetate, or acetone; non-polar solvents including ether, chloroform, benzene, hexane or dichlorohexane; Or a composition for skin regeneration that is a mixed solvent thereof.
According to claim 1,
The composition is a composition for skin regeneration, for the preparation of medicines, cosmetics or foods for skin regeneration.
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