KR20190066264A - Method for Preparing Aged Opuntia Ficusindica Fruit Powder Using Magma Seawater and a Composition for Moisturizing Skin Using an Extract of the Powder - Google Patents

Abstract

Disclosed are a method of producing Opuntia ficus indica powder aged with lava seawater, a skin-moisturizing composition using extracts of the powder, and a composition for atopic dermatitis. The method comprises: a step of soaking Opuntia ficus indica powder in lava seawater; and a step of aging the Opuntia ficus indica powder for 7 days or longer after soaking.

Description

TECHNICAL FIELD [0001] The present invention relates to a method for preparing a lily water-aged biannual seed powder and a composition for skin moisturizing using the extract of the powder,

The present invention relates to a method for producing a lily-of-the-sea water-soluble perennial plant powder and a composition for skin moisturizing using the powdery extract.

The skin that forms the outermost layer of the body is continuously affected by other artificial factors such as temperature change, moisture, dust, various microorganisms, etc., and force majeure stress, and the skin that protects the body from such external harmful factors It is a big institution. The skin is referred to as the epidermis, the dermis, and the subcutaneous fat tissue. The epidermis is composed of epithelial tissue. The keratinocyte, which is a main constituent cell, forms a stratum corneum by continuously changing new cells through different stages of differentiation, and the stratum corneum is located at the outermost part of the skin It maintains the homeostasis of the human body by acting as a barrier to protect against microbial invasion, external stimuli such as tension, and moisture loss.

The dermis is connective tissue below the epidermis and it is composed of extracellular matrix composed of collagenous fiber and elastic fibers which occupies more than 90% of the skin, And to maintain the tension. This extracellular matrix is mainly made up of fibroblasts, and is composed of fibrous proteins such as collagen and elastin, and has elasticity. In addition, between the collagen fiber and the elastic fiber, it is filled with a ground substance such as hyaluronic acid, which is involved in nutrition and moisture balance in the connective tissue while supporting other tissues around the substrate.

As a fiber component, collagen is the most important protein that maintains skin moisture and maintains skin flexibility and elasticity. It gives strength and tension to skin, protects skin from external stimuli, occupies 90% of dermal layer, (J Am Acad Dermatol, 17 (4): 610-613, 2001), which accounts for 3-4% of the dermal layer and is an important factor in the elasticity of the skin as well as collagen. Most of the collagen is Type I collagen, which is synthesized by fibroblasts and degraded by collagenases such as MMP-1. Examples of the polysaccharide include high molecular substances such as hyaluronic acid, mucopolysaccharide, and proteoclycan, which have strong water retention capability.

When skin aging is exposed to aging or ultraviolet rays, keratinocytes and fibroblasts produce reactive oxygen species, and reactive oxygen species oxidize DNA, cell membranes, proteins, fats, and nucleic acids in cells to cause skin aging (Bhawan et 1992; Brenneisen et al., 1997; Scharfferrs-Kochaneketal, 1997). In particular, more than a certain amount of UVB activates protease by a large amount of reactive oxygen species in the skin, collapses collagen and elastin, and damages the connective tissue of the dermis and collapses the skin barrier to cause oil and moisture loss and skin aging. In addition, UVB-induced reactive oxygen species induce oxidative stress, promote the secretion of various cytokines, activate the inflammatory reaction, and cause various skin diseases and skin cancer.

Hyaluronic acid, known as extracellular matrix component, is known to possess moisture. It is known that hyaluronic acid in the skin decreases with the progress of aging (Kim, Jin-Hwa, PhD thesis, Chungbuk National University, 2014).

The skin barrier consists of the stratum corneum at the outermost part of the skin. In order to do this, the keratinocytes are differentiated from the basal layer, and ultimately, the keratin and the lipid material that surrounds it are produced. The skin barrier maintains moisture retention by simultaneously retaining moisture loss and moisture retention (Kim et al, Kor. al, Ph.D. dissertation, University of Konkuk, Seoul, Korea, 2013)

Many researches are currently under way to inhibit skin aging and many functional cosmetics are on the market. However, some substances have limited use due to low stability or skin irritation resulting in safety problems. Accordingly, the search for a substance having excellent physiological activity and safety has been continued.

Meanwhile, in the cosmetics industry, many products using natural products have been developed to reduce skin irritation caused by various chemical substances. In addition to low adverse effects on skin, natural materials have recently become increasingly appreciated as a cosmetic raw material as consumers' response to cosmetics using natural materials has increased.

However, the compositions using the natural extracts extracted by the conventional methods have not only a small effect of desired whitening, wrinkle reduction, antioxidation, etc., but also have difficulty in maintaining and controlling the activity of these extracts.

Lava seawater, which is distributed only in the eastern part of Jeju Island (Gucheong-eup, Seongsan-eup, and Seokseon-myeon), is clean and has a constant water temperature throughout the year and contains a large amount of various minerals not found in ordinary salt groundwater or seawater. Table 1 shows the comparison of minerals content in the samples.

From Table 1, lava seawater is similar to general seawater in items such as salt, magnesium, calcium, and bromine, but there is a clear difference in trace element items such as germanium, vanadium, and selenium.

The present invention has been completed by confirming that the content of active ingredient is increased when natural materials are aged using lava sea water and that the aged extract enhances the skin physiological activities such as anti-inflammatory activity and hyaluronic acid production promoting activity.

It is an object of the present invention to provide a method for producing aged biannual seed powder using lava sea water.

Another object of the present invention is to provide a composition for skin moisturizing using the lava seaweed agar-agar powder.

Other and further objects of the present invention will be described below.

The inventors of the present invention found that the flavor content of Quercetin, Campferol, and the like was higher than that of the perennial powder extract in which the extract was not aged when aged (left as it is) for 7 days to 14 days after immersing the white perennial powder into lava sea water (NO production inhibitory activity), as well as excellent skin barrier strengthening activity and hyaluronic acid production promoting activity. In addition, the inhibitory activity of TARC (Thymus & activation-regulated chemokine) production in HaCaT cells, a human keratinocyte cell line treated with INF-γ and TNF-α, was remarkably superior to that of the non-aged Celophaceae powder extract. The TARC is a typical CC chemokine that acts on Th2 cells together with MDC (macrophage-derived chemokine) and acts on CCR4 (CC chemokine receptor 4) of Th2 cells to induce migration and invasion of Th2 cells as inflammatory lesions Serum concentrations of TARC and MDC have been reported to be significantly increased in patients with atopic dermatitis (J Clin Invest. 113 (5): 651-7, 2004; Curr Allergy Asthma Rep. 5 (4): 284-90, 2005) J Allergy Clin Immunol., 113 (2): 334-340) reported that the serum levels of MDC and TARC decreased when ciclosporin A or corticosteroids were used in the treatment of atopic dermatitis (J Dermatol Sci., 34: 201-208, 2004). Also, in vitro experiments showed that HaCaT cells, which are human keratinocyte cell lines, are highly expressed in MDC and TARC when treated with INF-? Or TNF-? The inhibiting substance is atopic skin (Int Immunol., 14 (7): 767-773, 2002).

In view of the above, the present invention relates to a method for producing a lily water seaweed perennial plant powder in one aspect, and in another aspect it relates to a skin moisturizing composition, an anti-inflammatory composition or an atopic dermatitis improvement ≪ / RTI >

The method for producing the lava seaweed aged perennial plant powder according to the present invention comprises a step of immersing the perennial plant powder in lava seawater and a step of aging (leaving) for 7 days or more after immersion.

In the method of the present invention, it is preferable that the perennial plant is a fruit part when referring to the following examples. Opuntia ficusindica is a perennial plant belonging to the cactus and Opuntia. It originated in the tropics. It originated in Korea for ornamental use on Jeju Island. It is now designated as Jeju Island Local Monument No. 35, And is widely distributed in the south coast region.

In addition, in the method of the present invention, the lava sea water may be a source of purified water from which raw water and suspended substances are removed, a salt and minerals-containing lava sea water generated as a byproduct during desalination by reverse osmosis or electrodialysis, It may be lava water generated as a byproduct of desalination by the osmotic pressure method. Reverse osmosis (RO) is a method in which ionic substances such as salts dissolved in water are used as a membrane, and only pure water is passed through. Therefore, lava water generated as a by-product contains salt and ionic minerals .

In this specification, "lava sea water" is defined in Korean Patent No. 0853244 (Application No.: 10-2008-0027861), entitled " Method for producing mineral composition and mineral composition obtained by the method & It is synonymous with the "saltwater underground in the eastern part of Jeju Island" as described. Specifically, "eastern part of Jeju Island" means Jeju-eup, Seongsan-eup and Seokseon-myeon in Jeju-do in administrative district, and "saltwater groundwater" means groundwater containing salinity of a certain amount or more. Specifically, According to the criteria for classification of groundwater in Jeju Island based on the salinity concentration used in the study of the hydrological geology in the eastern part of Cheju Island (Ⅱ), 2003, Kyungwon University, considered as part of this specification), low salt groundwater (electric conductivity of 1,700 ㎲ / cm (17,350 / / cm) and saltwater groundwater (electrical conductivity of 17,350 / / cm or more), but preferably the electrical conductivity is 17,350 / / cm or more or the salt concentration is 30 ‰ ) Or more (usually the saline concentration of the seawater is 32 to 35 ‰).

In the method of the present invention, there is no particular limitation on the temperature at the time of aging, but when referring to the following examples, it is preferable that the temperature is lower than 15 ° C, particularly 4 ° C to 15 ° C.

In the method of the present invention, the aging period is preferably 7 days or more and 14 days or less with reference to the following examples.

In the composition for skin moisturizing, antiinflammatory composition or composition for improving atopic dermatitis of the present invention (hereinafter collectively referred to as "composition of the present invention") of the present invention is an extract of the above-mentioned lava seaweed aged perennial powder, The powder is dissolved in water, a lower alcohol having 1 to 4 carbon atoms (methanol, ethanol, butanol, etc.), methylene chloride, ethylene, acetone, hexane, ether, chloroform, ethyl acetate, butyl acetate, N, An extract obtained by leaching using dimethylsulfoxide (DMSO), 1,3-butylene glycol, propylene glycol or a mixed solvent thereof, an extract obtained by using a supercritical extraction solvent such as carbon dioxide, pentane, or an extract thereof Fraction, and the extraction method may be selected from the group consisting of cold treatment, reflux, warming, ultrasonic irradiation, It can be applied to any method, such as extraction threshold. In the case of the fractionated extract, a fraction obtained by suspending the extract in a specific solvent and mixing and leaving with a solvent having a different polarity, the crude extract is adsorbed on a column packed with silica gel, and then a hydrophobic solvent, a hydrophilic solvent, Quot; means fractions obtained as a mobile phase. Also, the meaning of the extract includes a concentrated liquid extract or a solid extract in which the extraction solvent is removed by a method such as freeze drying, vacuum drying, hot air drying, spray drying and the like. Preferably an extract obtained by using water, ethanol or a mixed solvent thereof as an extraction solvent.

In the present specification, the term " active ingredient "alone means an ingredient which exhibits the desired activity or which can exhibit activity together with a carrier which is not itself active.

The composition of the present invention may be contained in any amount (effective amount) as long as it can exhibit a moisturizing activity, an anti-inflammatory activity, an atopic dermatitis-improving activity, etc., which are intended to be treated or improved depending on the use, formulation, , A typical effective amount will be determined within the range of from 0.001% to 15% by weight based on the total weight of the composition. Herein, the term "effective amount" as used herein refers to an amount of an effective amount of a dermatologic / pharmacological agent, such as a moisturizing agent, an antiinflammatory effect, etc., when the composition of the present invention is administered to a mammal, preferably a human, Quot; refers to the amount of the active ingredient contained in the composition of the present invention, which can exhibit the effect of the present invention. Such effective amounts can be determined experimentally within the ordinary skill of those skilled in the art.

The composition of the present invention can be used not only for the effective ingredient but also for the enhancement or reinforcement of the skin moisturizing effect or the addition of the similar activity such as skin irritation inhibiting activity, skin protecting activity (skin whitening, skin damaging by ultraviolet rays, May further comprise any compound or natural extract known to have safety in the art and known to have a corresponding activity in order to facilitate the taking, ingestion, and ease of use. These compounds or extracts include the national pharmacopoeia (Korean pharmacopoeia), the national health functional food circulation in Korea (in Korea, the health functional food standard and specification), the functional cosmetics circulation in each country (in Korea, Compounds or extracts listed in the official documents such as "Functional Cosmetics Standards and Test Methods"), compounds or extracts approved by the respective countries in accordance with the laws of the respective countries (in Korea, "Pharmaceutical Affairs Law") regulating the manufacture and sale of pharmaceuticals In accordance with the laws of each country that regulate the manufacture and sale of compounds or extracts and functional cosmetics that are recognized as functioning in accordance with the laws of the respective countries ("Act on Health Functional Foods" in Korea) that regulate the manufacture and sale of functional foods &Quot;).≪ / RTI > Examples of skin whitening ingredients include arbutin, niacinamide, ascorbyl glucoside, alpha-bisabolol and oil soluble licorice (Glycyrrhiza) extract. Examples of the skin wrinkle improving agent include retinol, retinyl palmitate , Adenosine, polyethoxylated amide, and the like. Examples of the ultraviolet protecting component include drometrizol, drometrizol trisiloxane, dipalayyltriolate, Dimethicyldecibenzalmalonate, diethylhexylbutadimido Triazone, and pine bark extract, phosphatidylserine, finger root extract powder, and complex extracts of red ginseng and mountain juice. Examples of the moisturizing component include AP collagen enzyme digesting peptide, collactive collagen peptide, N-acetyl glucosamine, konjac potato extract, dandelion extract, rice bran extract, corn germ extract, low molecular weight collagen peptide, ground extract powder, phosphatidylserine, hyaluronic acid Examples of functional ingredients for improving 'irritable skin condition' include L. sakei Probio 65, gamma linolenic acid-containing oil, L. plantarum CJLP133, and probiotic ATP. Such compounds or natural extracts may be included in the compositions of the present invention in combination with one or more of their active ingredients.

In a specific embodiment, the composition of the present invention can be identified as a food composition. When the composition of the present invention is identified as a food composition, the anti-inflammatory use can be understood as the use of inflammatory skin irritation mitigation, and the atopic dermatitis improving activity can also be understood as a skin hypersensitive reaction inhibiting activity.

The food composition of the present invention can be produced in any form and can be used in various forms such as beverages such as tea, juice, carbonated drink, ionic drink, processed oil such as milk and yogurt, gum, rice cake, Korean confectionery, A food, a health food, a food, a tablet, a capsule, a ring, a granule, a liquid, a powder, a slice, a paste, a syrup, a gel, a jelly and a bar.

In addition, the food composition of the present invention may be classified into any product category as long as it meets the laws and regulations on the time of manufacture and distribution in the legal and functional category. For example, it is a health functional food according to the 「Health Functional Food Act」 in Korea, or a food functional food according to the Korean Food Sanitation Law (Food Standards and Specifications) , Special-purpose food, and the like.

The food composition of the present invention may contain food additives in addition to the active ingredients thereof. Food additives are generally understood to be substances that are added to foods and mixed or infiltrated into food in the manufacture, processing or preservation of food, and their safety must be ensured since they are ingested daily with food and for long periods of time. In food additives according to the laws of the respective countries ("Food Sanitation Act" in Korea) regulating the manufacture and distribution of food, food additives with safety are specified in terms of ingredient or function. In the Food Additives Code of Korea (Food Additives Standards and Standards), the food additives are classified into chemical compounds, natural additives and mixed preparations in terms of ingredients. Such food additives are classified into sweeteners, flavors Preservatives, emulsifiers, acidulants, and thickeners.

The sweetener is used for imparting a sweet taste suitable for foods, and both natural and synthetic sweeteners can be used in the composition of the present invention. Preferably, natural sweeteners are used. Examples of natural sweeteners include sugar sweeteners such as corn syrup solids, honey, sucrose, fructose, lactose and maltose.

Flavors may be used to enhance taste or flavor, both natural and synthetic. Preferably, a natural one is used. When using natural ones, the purpose of nutritional fortification can be performed in addition to the flavor. Examples of natural flavoring agents include those obtained from apples, lemons, citrus fruits, grapes, strawberries, peaches, and the like, or those obtained from green tea leaves, Asiatica, Daegu, Cinnamon, Chrysanthemum leaves and Jasmine. Also, those obtained from ginseng (red ginseng), bamboo shoots, aloe vera, banks and the like can be used. The natural flavoring agent may be a liquid concentrate or a solid form of extract. Synthetic flavors may be used depending on the case, and synthetic flavors such as esters, alcohols, aldehydes, terpenes and the like may be used.

As the preservative, calcium sorbate, sodium sorbate, potassium sorbate, calcium benzoate, sodium benzoate, potassium benzoate and EDTA (ethylenediaminetetraacetic acid) can be used. As the emulsifier, acacia gum, carboxymethyl cellulose, Pectin and the like. As the acidulant, math, malic acid, fumaric acid, adipic acid, phosphoric acid, gluconic acid, tartaric acid, ascorbic acid, acetic acid, phosphoric acid and the like can be used. The acidulant may be added so that the food composition has a proper acidity for the purpose of inhibiting the growth of microorganisms other than the purpose of enhancing the taste.

Examples of the thickening agent include suspending agents, sedimentation agents, gel-forming agents, bulking agents and the like.

The food composition of the present invention may contain physiologically active substances or minerals which are known in the art and which are stable as a food additive in addition to the above-mentioned food additives in order to supplement and supplement functional and nutritional properties.

Examples of such physiologically active substances include catechins contained in green tea and the like, vitamins such as vitamin B1, vitamin C, vitamin E and vitamin B12, tocopherol, dibenzoyl thiamine, etc. Examples of minerals include calcium preparations such as calcium citrate, magnesium stearate , Iron preparations such as iron citrate, chromium chloride, potassium iodide, selenium, germanium, vanadium, zinc and the like.

The food composition of the present invention may contain an appropriate amount of the above-mentioned food additives according to the product type so as to achieve the purpose of addition thereof.

With regard to other food additives that may be included in the food composition of the present invention, reference may be made to the Food Code of the respective countries or the Food Additives Code.

In another specific embodiment, the composition of the present invention can be identified as a pharmaceutical composition.

The pharmaceutical composition of the present invention may be prepared into oral formulations or parenteral formulations according to the route of administration by conventional methods known in the art, including pharmaceutically acceptable carriers in addition to the active ingredient. Where the route of administration may be any suitable route including local routes, oral routes, intravenous routes, intramuscular routes, and direct absorption through mucosal tissues, and combinations of two or more routes may be used. An example of a combination of two or more routes is a combination of two or more formulations of the drug according to the route of administration, for example, one drug is administered intravenously and another drug is administered via a local route.

Pharmaceutically acceptable carriers are well known in the art depending on the route of administration and formulation, and specific reference may be made to the pharmacopoeia of each country, including the "Korean Pharmacopoeia ".

When the pharmaceutical composition of the present invention is prepared into an oral formulation, it may be formulated into powder, granules, tablets, pills, sugar tablets, capsules, solutions, gels, syrups, suspensions, wafers And the like. Examples of suitable carriers include starches such as lactose, glucose, sucrose, dextrose, sorbitol, mannitol and xylitol, corn starch, potato starch and wheat starch, cellulose, methylcellulose, ethylcellulose, sodium carboxymethylcellulose, Hydroxypropylmethylcellulose and the like; polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, magnesium stearate, mineral oil, malt, gelatin, talc, polyol, vegetable oil, ethanol Serol, and the like. In case of formulation, suitable binders, lubricants, disintegrants, coloring agents, diluents and the like may be included as needed. Examples of suitable binders include starch, magnesium aluminum silicate, starch pellets, gelatin, methyl cellulose, sodium carboxymethyl cellulose, polyvinyl pyrrolidone, glucose, corn sweetener, sodium alginate, polyethylene glycol, wax and the like. Examples of the disintegrating agent include starch, methylcellulose, magnesium stearate, magnesium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, silica, talcum, stearic acid, Agar, bentonite, xanthan gum, starch, alginic acid or its sodium salt, and the like. Examples of the diluent include lactose, dextrose, sucrose, mannitol, sorbitol, cellulose, glycine and the like.

When the pharmaceutical composition of the present invention is prepared into a parenteral dosage form, it may be formulated in the form of an injection, transdermal drug delivery, nasal aspirate and suppository together with a suitable carrier according to methods known in the art. As the carrier suitable for injection preparation, aqueous isotonic solutions or suspensions may be used. Specifically, PBS (phosphate buffered saline) containing triethanolamine, sterile water for injection, and isotonic solution such as 5% dextrose may be used . When formulated with a transdermal preparation, it can be formulated in the form of ointments, creams, lotions, gels, external liquids, pastes, liniments, and air-lozenges. Nasal inhalers may be formulated in the form of aerosol sprays using suitable propellants such as dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide, etc., and when formulated as a suppository, witepsol, tween 61, polyethylene glycols, cacao butter, laurin, polyoxyethylene sorbitan fatty acid esters, polyoxyethylene stearates, and sorbitan fatty acid esters.

The formulation of pharmaceutical compositions is well known in the art and can be found, for example, in Remington ' s Pharmaceutical Sciences (19th ed., 1995). This document is considered part of this specification.

The preferred dosage of the pharmaceutical composition of the present invention is 0.001 mg / kg to 10 g / kg per day, preferably 0.001 mg / kg to 1 g / day, depending on the patient's condition, body weight, sex, age, / kg < / RTI > The administration can be carried out once or several times a day. Such dosages should in no way be construed as limiting the scope of the invention.

In another specific embodiment, the composition of the present invention can be identified as a cosmetic composition. When the composition of the present invention is identified as a cosmetic composition, anti-inflammatory use can be understood as an application for the relief of inflammatory skin irritation, and the atopic dermatitis improving activity can also be understood as a skin hypersensitive reaction inhibiting activity.

Even when the composition of the present invention is identified as a cosmetic composition, the cosmetic composition may be classified into any product category according to the use of the cosmetic composition. Specifically, the cosmetic composition may contain functional cosmetic products having applications such as improvement of skin troubles and improvement of atopic dermatitis, General cosmetics, and the like. Specific examples of the product form include a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing oil, powder foundation, emulsion foundation, wax Foundation, spray, and the like. In a specific product form, it may be a form of flexible lotion, nutritional lotion, nutritional cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder.

The cosmetic composition of the present invention may contain, in addition to its active ingredient, conventional additives such as stabilizers, solubilizing agents, surfactants, vitamins, colorants and antioxidants, and carriers commonly used in cosmetic compositions.

When the formulation of the present invention is a paste, a cream or a gel, an animal oil, vegetable oil, wax, paraffin, starch, tracant, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide may be used as a carrier component .

When the formulation of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component. In the case of a spray, in particular, / Propane or dimethyl ether.

When the formulation of the present invention is a solution or emulsion, a solvent, a solubilizing agent or an emulsifying agent is used as a carrier component. Specifically, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, 1,3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol, fatty acid esters of sorbitan, and the like.

When the formulation of the present invention is a suspension, a carrier, such as water, a liquid diluent such as ethanol or propylene glycol, a suspension such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester, polyoxyethylene sorbitan ester, Cellulose, aluminum metahydroxide, bentonite, agar, etc. may be used.

When the formulation of the present invention is an interfacial active agent-containing cleansing, the carrier component is selected from aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid amide Ether sulfates, alkylamidobetaines, aliphatic alcohols, fatty acid glycerides, fatty acid diethanolamides, vegetable oils, lanolin derivatives or ethoxylated glycerol fatty acid esters.

The cosmetic composition of the present invention can be produced according to a method for producing a cosmetic composition which is usually carried out in the art, except that it contains the effective ingredient showing skin wrinkle improving activity.

According to the present invention described above, it is possible to provide a method capable of producing a lava seaweed-ripened perennial plant having various skin physiological activities such as an anti-inflammatory activity and an activity of promoting the production of hyaluronic acid, have.

Lava seaweed aged biannual powder extract can be used in cosmetics and the like because of its functionalities such as skin moisturizing, atopic dermatitis improvement, inflammatory dermatitis improvement, and inflammatory skin irritation mitigation.

Figure 1 shows the results of HPLC analysis of the standard material.
FIG. 2 shows the results of HPLC analysis of the fruit extract of Baekjangju before and after aging.
Figure 3 shows the results of cytotoxicity experiments.
Fig. 4 shows experimental results of inhibition of nitrogen monoxide (NO) production.
Fig. 5 shows experimental results of inhibition of TARC production.
FIG. 6 shows the result of the experiment for promoting the formation of the CE Formation.
Fig. 7 shows the results of stimulating PPARa expression.
Fig. 8 shows experimental results of accelerating hyaluronic acid (HA) production.

Hereinafter, the present invention will be described with reference to Examples and Experimental Examples. However, the scope of the present invention is not limited to these examples and experimental examples.

<Examples> Production of fruit extract of Baekseongcho aged in lava sea water

Example 1 Production of Lycopersicum Seaweed,

The fruit was cut in half and then dried in a hot air dryer at 55 ° C for 2 days and pulverized to prepare a powder of white perennial. Then, 10 times as much weight of lava water (concentrated water of raw water, salt product and ionic minerals as a byproduct produced in the production of desalted lava water by the reverse osmosis method) was added, and the temperature was maintained at 4 to 15 ° C Dried powder of Baekryeomgi was aged for 7 to 14 days.

The mixture of the perennial seed powder and the lava sea water was filtered with a 120 mesh nylon filter to obtain aged white perilla seed powder, 10 times the weight of purified water was added to the perennial seed powder, and the mixture was filtered with a 120 mesh nylon filter. And the remaining salinity was removed from the aged Baekjangcho powder. The salt - free white acorn powder was dried in a hot air dryer at 55 ℃ for 1 day and then pulverized using a dry grinder.

Example 2 Preparation of Lycopersicum Seedlings Fruit Leaf Extract

70% ethanol was added 10 times by weight of the aged white birch seed powder obtained in Example 1, and the mixture was ultrasonically extracted for 3 hours and filtered through a filter paper (1 μm pore size). The filtrate was concentrated under reduced pressure and lyophilized, The fruit extract was prepared.

&Lt; Comparative Example > Preparation of fruit extract of common white perennial plant

The fruit extract of Baekjunseok was prepared in the same manner as in Example 2, except that the non-aged, ordinary white perennial powder was extracted.

<Experimental Example 1> Comparison of components of fruit juice extract of Leuconostoc sp.

The experimental conditions for the analysis of the content of active ingredients are as follows.

Samples of aging periodicity were collected from high-performance liquid chromatography (Waters Alliance 2695 HPLC), which was used for the analysis.

The column used here was Mightysil RP 18GP 5 μm (4.6 × 250 mm), the mobile phase was 100% acetonitrile, water (1% phosphoric acid) and absorbance 200-400 nm Waters 2996 PAD). At this time, the flow rate of the mobile phase was maintained at 1.0 mL per minute.

The sample was diluted with methanol to a concentration of 30 mg / mL, filtered through a 0.45 μm polytetrafluoroethylene (PTFE) filter, and injected into a high performance liquid chromatography. The conditions of the high performance liquid chromatography were as follows.

1. Reference material

Quercetin: 12.5, 25, 50, 100 ppm (Sigma products)

3-O-methyl quercetin: 12.5, 25, 50, 100 ppm (Sigma product)

Camphor roll: 12.5, 25, 50, 100 ppm (product of sigma)

Isolametine: 12.5, 25, 50, 100 ppm (Sigma product)

2. Analysis of flavonoids

To perform flavonoid analysis, Park et al. J Korean Soc Food Sci Nutr, Dentification of Flavonoids from Extracts of Opuntia ficus-indica var. Analysis was carried out according to the method of Saboten and Content Determination of Marker Components Using HPLC-PDA, 2017, 46 (2), 210219 and the results are shown in [Table 2] and [Figure 1] to [Figure 2] Respectively.

As a result of HPLC analysis, the content of flavonoid was increased significantly in the water extract of Baekjangcho, which was aged in lava sea water, and the content of flavonoid was greatly increased at 7 days of fermentation.

<Experimental Example 2> Skin-related activity test of lycopodium agar

1. Experimental Method

1.1 Cell culture

Human Keratinocyte HaCaT and Mouse macrophage cell Raw 264.7 were inoculated on the bottom of the culture dish and DMEM (Dulbecco's Modified Eagle's Medium) medium containing 10% FBS (fetal bovine serum) was added and incubated at 37 ° C with 5% &Lt; / RTI &gt; incubator.

1.2 Cell toxicity experiments

96 of Human Keratinocyte the HaCaT the ^{well plate 1.5 × 10 4 cells /} well, Mouse macrophage cell of Raw 264.7 to 6 × 10 ⁴ cells / well busy then increments, 37 ℃ so that the cells attach well to the plate for 24 hours, 5 They were cultured in% CO ₂ cell incubator.

 After 24 hours, HaCaT and Raw264.7 were transformed into starvation using DMEM medium (serum-free medium) containing no FBS, and then cultured for the next day. Cells were treated with WST-1 solution (DoGen, EZ-3000), and absorbance was measured at 450 nm with an ELISA reader (Thermo, 51119300).

1.3 Determination of NO (Nitric oxide)

Macrophage is known to play an important role in the inflammatory response by producing nitric oxide (NO) and various inflammatory cytokines by chemical stimulation (Ito T., et. al., Curr Drug Traget Inflamm Allergy, 2 (3): 257-265, 2003). NO is known to react with superoxide to form peroxynitrite, which acts as a strong oxidizing agent and damages cells to cause an inflammatory reaction (Gupta SC et al., Exp Biol Med. , 236: 658-671, 2011; Riehemann et al., FEBS Lett., 442: 89-94, 1999; Stamler et al., Science, 258: 1898-1902, 1992).

Raw 267.7 cells were seeded at 6 × 10 ⁴ cells / well in a 96-well plate, and cultured in the cell culture conditions for 24 hours. Raw264.7 was transformed into starvation using FBS-free medium (serum-free medium), and the next day, LPS (㎍ / mL) was treated with a certain concentration of sample. The cell culture medium was reacted with Griess reagent 1: 1, and the absorbance was measured at 540 nm with an ELISA reader. The NO concentration was determined using a standard curve of sodium nitrite and the results were expressed as a percentage of the untreated (control) group.

1.4 Quantification of TARC

In vitro studies have shown that MDC (macrophage-derived chemokine) and TARC (Thymus & activation-regulated chemokine) are highly expressed when human keratinocyte HaCaT cells are treated with INF-γ or TNF-α. (Int Immunol., 14 (7): 767-773, 2002), which has been proposed to be used as a therapeutic agent for atopic dermatitis.

HaCaT was dispensed in a 96-well plate at 1.5 × 10 ⁴ cells / well, and then cultured in the cell culture conditions for 24 hours. HaCaT was transformed into starvation using a medium containing no FBS (serum-free medium). The next day, IFN-γ and TNF-α were mixed and used as a stimulant. After the experiment using the TARC ELISA kit (R & D System, DY364), the absorbance was measured at 450 nm with an ELISA reader. The results are expressed as a percentage of the sample-treated group (control group).

1.5 CE (Cornified envelope) assay

The keratinocytes form a cornified envelope through the differentiation process, which combines with lipid components to form a physical skin barrier to protect the body from the external environment. The CE assay was performed to confirm skin barrier enhancement efficacy by using calcium as a positive control, which is a typical factor that maintains homeostasis by controlling proliferation and differentiation of keratinocytes.

HaCaT was dispensed into 96-well plates at 2.5 × 10 ⁵ cells / well, and cultured in the cell culture conditions for 24 hours. HaCaT was cultured in a DMEM medium without calcium, treated with a certain concentration of the sample, and cultured for 7 days. At this time, the medium was changed every 2 days. After 7 days of incubation, cells were washed with PBS and treated with 2% sodium dodecyl sulphate (SDS) and 20 mM dithiothreitol (DTT). After boiling at 100 ° C for 5 minutes, the absorbance was measured at 340 nm with an ELISA reader.

1.6 Weatern blot

HaCaT was dispensed into 96-well plates at 2.5 × 10 ⁵ cells / well, and cultured in the cell culture conditions for 24 hours. HaCaT was cultured in medium containing no calcium and FBS, and treated with a certain concentration of sample.

The cells were washed with PBS to recover the cells, and the cells were lysed using RIPA buffer, followed by centrifugation at 15000 rpm for 30 minutes to obtain a cell lysate. A certain amount of protein obtained from protein quantification and 4 × sample buffer were mixed and heated at 100 ° C. for 5 minutes. The proteins were separated by electrophoresis on Bis-Tris Plus gel gel (Invitrogen, NW04122BOX), and the separated proteins were transferred to a nitrocellulose membrane (Invitrogen, IB23002).

Membranes were blocked with blocking buffer (5% fat-free dried milk powder in TBST) at 4 ° C for 16 hours. Human PPAR-α (Santa Cruz) was diluted in blocking buffer and allowed to react at room temperature for 3 hours. Secondary antibody bound to peroxidase was reacted at room temperature for 1 hour. Were analyzed using an enhanced chemiluminescence instrument.

1.7 Quantification of HA (Hyaluronic acid)

HaCaT was dispensed in a 96-well plate at 1.5 × 10 ⁴ cells / well, and then cultured in the cell culture conditions for 24 hours. HaCaT was transformed into starvation using FBS-free medium (serum-free medium) and then cultured for the next day at a constant concentration. After the experiment using Hyaluronic acid (R & D System, DY3614) ELISA kit, absorbance was measured at 450 nm with an ELISA reader. The results are expressed as a percentage of the sample-treated group (control group).

1.8 Quantification of protein

Protein quantification was performed by measuring the amount of protein contained in the cells. The medium was recovered, washed with PBS, and then treated with 1N NaOH to dissolve the cells. Lowry assay, one of the protein assay methods, was used and the DC Protein Assay (500-0116) product of Bio-Rad was used.

  Reagent A: Reagent S = 50: 1, reacted with Reagent B, and allowed to react at room temperature for 20 minutes. Absorbance was measured at 750 nm using an ELISA reader to confirm the amount of protein. The amount of protein was determined using a standard curve obtained from bovine serum albumin.

2. Experimental results

2.1 Cytotoxicity

Each extract was dissolved in dimethyl sulfoxide (DMSO), and the cytotoxicity test was performed on HaCaT and Raw 264.7 cells at a maximum concentration of 800 μg / ml. The results are shown in [Table 3] and [Figure 3] below. Based on the cytotoxicity test results, the efficacy test was carried out by treating the sample at a concentration not cytotoxic.

2.2 NO determination

NO measurement results are shown in [Table 4] and [Figure 4] below. NO production was increased by 57.52% in the LPS treated group compared to the untreated group, and it was confirmed that the extract of Baekseokcho aged with Lava seawater had better inhibitory effect on NO production than the Baekseobacho fruit extract not aged. At this time, the aged extract showed 29.41% higher inhibitory amount of NO production than 400mg / mL of aged fruit extract, which was not aged.

2.3 Quantification of TARC

TARC quantification results are shown in [Table 5] and [Figure 5] below. TARC production was increased by 173.00% in the TNF-α and IFN-γ treatment groups compared to the untreated control group, and that the extracts of Baekjangcho extract aged with lava seawater showed better inhibitory effect on TARC formation I could. At this time, the fermented extract showed an inhibitory amount of TARC production of 400 ㎍ / mL higher by 119.12% than that before fermentation.

2.4 CE Quantification

CE determination results are shown in [Table 6] and [Figure 6] below. The extracts of non-fermented non-fermented fruits showed that the differentiation promoting effect of keratinocytes was promoted, whereas the extracts of fermented Leuconostoc spp. Promoted the differentiation of keratinocytes in a concentration-dependent manner. The skin barrier enhancement effect could be indirectly confirmed. At this time, the fermented extract showed 27.15% higher CE production at 400 ㎍ / mL compared to before fermentation.

2.5 PPAR quantitation

The results of PPAR expression are shown in [Table 7] and [Figure 7] below. It was confirmed that the expression level of PPARα was increased at 400 μg / ml concentration of the fruit extract of Baekryeomgi aged with lava sea water. At this time, the ripening extract showed 62.5% higher PPAR expression level at 400 μg / mL compared to before aging.

2.6 HA quantification

The HA quantification results are shown in [Table 8] and [Figure 8] below. On the other hand, the fruit extract of Baekjangcho, which was aged with Lava seawater, showed the effect of stimulating HA production in a dose - dependent manner. At this time, the extract of fruit aged 100 years old showed 240.63% higher increase of HA production at 400 ㎍ / mL compared to that before aging.

Claims (12)

A step of immersing the perennial plant powder in lava seawater, and a step of aging by standing for at least 7 days after immersion
&Lt; / RTI &gt;
The method according to claim 1,
Wherein the perennial plant powder is a perennial plant fruit powder.
The method according to claim 1,
Wherein said lava seawater is lava water containing raw water, refined raw water from which suspended substances are removed, or salt and minerals generated as a byproduct of desalination of raw lava water
The method according to claim 1,
Wherein the aging is carried out at a temperature ranging from 4 캜 to 15 캜.
A skin-moisturizing composition comprising, as an active ingredient, an extract from a lagoon-hydrothermal leucocyte obtained by a method according to any one of claims 1 to 4.
6. The method of claim 5,
Wherein the extract is water, ethanol or a mixed solvent thereof.
6. The method of claim 5,
Wherein the composition is a cosmetic composition.
6. The method of claim 5,
Wherein the composition is a food composition.
A composition for improving atopic dermatitis comprising, as an active ingredient, an extract from a lagoon-hydrothermal leucocyte obtained by a method according to any one of claims 1 to 4.
10. The method of claim 9,
Wherein the extract is water, ethanol or a mixed solvent thereof.
10. The method of claim 9,
Wherein the composition is a cosmetic composition.
10. The method of claim 9,
Wherein the composition is a food composition.

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