KR102239784B1 - Cosmetic composition comprising mixture of fermented extract of Cirsium japonicum and Moringa oleifera - Google Patents

Abstract

The present invention relates to a mixture of a fermented extract of thistle and moringa, and more particularly, to an antioxidant or anti-inflammatory cosmetic, food, and pharmaceutical composition comprising the mixture of fermented extract of hot spring water as an active ingredient. The mixture of thistle and Moringa hot spring water fermentation extract according to the present invention has low cytotoxicity, excellent electron donating ability and radical scavenging ability, and has the effect of inhibiting the expression of inflammatory factor proteins, fermentation of thistle and Moringa hot spring water of the present invention. The mixture of extracts can be used in various fields in food, cosmetic and pharmaceutical fields.

Description

Cosmetic composition comprising mixture of fermented extract of Cirsium japonicum and Moringa oleifera}

The present invention relates to a mixture of a fermented extract of thistle and moringa, and more particularly, to an antioxidant or anti-inflammatory cosmetic, food, and pharmaceutical composition comprising the mixture of fermented extract of hot spring water as an active ingredient.

Fermentation refers to the phenomenon that when microorganisms use carbohydrates as an energy source, the carbon bonds are broken and alcohol, organic acids, carbon dioxide gas, etc. are generated. Fermented cosmetics were developed with the focus on the substances produced during the fermentation process. In other words, the function of activating cells by enzymes contained in fermented foods that we eat in everyday life was introduced into cosmetics. Fermentation in cosmetics removes fermenting bacteria in the process of fermenting microorganisms and extracts active ingredients in addition to enzymes, thereby doubling the efficacy of only one existing enzyme and allowing the ingredients to be quickly absorbed into the skin.

In general, the spacing between skin cells is 70 to 80 nm, and in order for cosmetic ingredients to penetrate into the skin, the molecular weight of the ingredients must be smaller than the size of the skin cells. In order to solve this problem, the cosmetics industry focused on the development of nanoparticles or liposome-type microcarriers. However, this method requires a high level of technology and has a limitation in that the manufacturing cost is high.

On the other hand, it has been reported that fermented cosmetics can increase skin absorption rate only by fermentation without such advanced technology. In addition, there is an advantage in that various fermentation ingredients can be absorbed into the dermis layer because the molecular particles are decomposed to 80 nm or less due to the action of microorganisms and yeast during the fermentation of raw materials.

Meanwhile, Moringa oleifera is known to have 14 species as perennial plants distributed in tropical and subtropical climate regions. Recently, in various countries around the world, moringa has been used in various ways, such as medicines, cosmetics, water purifiers, animal feeds, and crop growth promoters, depending on the characteristics of each part of moringa. Moringa leaves and seeds, flowers, tree trunks, fruits (pods), roots, and all other parts are used as food ingredients. Moringa leaves have 20 to 25% dry matter compared to other plant leaves. It has been reported that the moisture content is low and the content of various nutrients is relatively high. Moringa leaves are high in protein as well as minerals such as beta-carotene, vitamin C, carotene, calcium, potassium, and iron, so in Africa, where food is absolutely scarce, it is useful as a dietary supplement to cope with malnutrition or as a nutritional food for mothers. It is a plant.

Also thistle ( Cirsium japonicum var. ussuriense ) is a perennial herb belonging to the family Compositae. In Korea, 13 species and 6 varieties and 1 variety are native to the mountains. In oriental medicine, the above-ground part or the underground part (root) is called daegae and is used for medicinal purposes. That is, the above-ground parts such as leaves, stems, flowers and seeds are collected from May to June when the seeds are in bloom during the flowering period, and the roots are collected in autumn and dried. It has been used to treat hepatitis and hypertension. Plants of the genus Cirsium are rich in flavonoid compounds with excellent physiological activity, and have anti-inflammatory, anti-cancer, anti-mutagenic, anti-fungal, neuroprotective and immune enhancing activities. In addition, thistle inhibits lipid peroxidation and promotes alcohol detoxification by increasing the activity of glutathione reductase, thereby promoting liver protection, and thistle leaf extract is known to have antirheumatic effects.

Accordingly, the present inventors completed the present invention by developing a mixture of thistle and Moringa hot spring water fermented extract in order to increase the skin absorption rate of the active ingredient.

Accordingly, an object of the present invention is to provide an antioxidant or anti-inflammatory cosmetic composition comprising a mixture of thistle and Moringa hot spring water fermented extract as an active ingredient.

Another object of the present invention is to provide an antioxidant or anti-inflammatory food composition comprising a mixture of thistle and Moringa hot spring water fermented extract as an active ingredient.

Another object of the present invention is to provide a pharmaceutical composition for preventing or treating diseases caused by oxidative stress or inflammation, comprising a mixture of thistle and Moringa hot spring water fermented extract as an active ingredient.

In order to achieve the above object, the present invention provides an antioxidant or anti-inflammatory cosmetic composition comprising a mixture of thistle and Moringa hot spring water fermented extract as an active ingredient.

In addition, the present invention provides an antioxidant or anti-inflammatory food composition comprising a mixture of thistle and Moringa hot spring water fermented extract as an active ingredient.

In addition, the present invention provides a pharmaceutical composition for preventing or treating diseases caused by oxidative stress or inflammation, comprising a mixture of thistle and Moringa hot spring water fermented extract as an active ingredient.

The mixture of thistle and Moringa hot spring water fermentation extract according to the present invention has low cytotoxicity, excellent electron donating ability and radical scavenging ability, and has the effect of inhibiting the expression of inflammatory factor proteins, fermentation of thistle and Moringa hot spring water of the present invention. The mixture of extracts can be used in various fields in food, cosmetic and pharmaceutical fields.

1 is a diagram showing a method for preparing a mixture of fermented extract of hot spring water of thistle and Moringa according to the present invention.
2 is a diagram showing the result of measuring the electron donating ability of a mixture of fermented extract of hot spring water of Thistle and Moringa according to the present invention.
3 is a diagram showing the result of measuring the ABTS radical scavenging ability of a mixture of fermented extract of hot spring water of thistle and Moringa according to the present invention.
4 is a diagram showing the results of measuring the cytotoxicity of a mixture of fermented extracts of thistle and Moringa hot spring water according to the present invention in Raw264.7 cells, which are macrophages.
5 is a diagram showing the results of measuring the cytotoxicity of a mixture of fermented extracts of thistle and Moringa hot spring water according to the present invention in B16F10, which is melanoma cells.
Figure 6 is a view showing the results of measuring the NO production inhibitory activity of a mixture of thistle and Moringa's hot spring water fermentation extract according to the present invention.
Figure 7 is a diagram showing the results of measuring the expression of the inflammatory factor iNOS protein when Raw264.7 cells are treated with a mixture of fermented extracts of thistle and Moringa's hot spring water according to the present invention.
8 is a diagram showing the results of measuring the expression of COX-2 protein, an inflammatory factor, when Raw264.7 cells were treated with a mixture of fermented extracts of thistle and Moringa's hot spring water according to the present invention.

Hereinafter, the present invention will be described in detail.

According to an aspect of the present invention, the present invention provides an antioxidant or anti-inflammatory cosmetic composition comprising a mixture of thistle and Moringa's hot spring water fermented extract as an active ingredient.

In the present invention, the term "extract" refers to a preparation obtained by squeezing a herbal medicine into an appropriate leachate and evaporating the leachate. It may be a crude product or a purified product.

The thistle and moringa can be used without limitation, such as cultivated or commercially available, and can be washed and used as it is, or can be used after drying, and both dry, shade, hot air drying, and natural drying methods can be used as drying methods. have.

According to a preferred embodiment of the present invention, thistle is a dried stem, leaf or root.

According to another preferred embodiment of the present invention, Moringa is dried Moringa leaves.

In the present invention, the term “hot spring water” refers to groundwater whose water temperature is higher than the annual average temperature of the region or the groundwater temperature of a shallow stratum, and in Korea, the temperature is 25°C or higher, and refers to groundwater that is not harmful to the human body.

In an embodiment of the present invention, the hot spring water is preferably Onyang hot spring water, but is not limited thereto.

In the present invention, the term “fermentation” refers to a phenomenon in which when a microorganism uses a carbohydrate as an energy source, carbon bonds of carbohydrates are cut off to produce alcohol, organic acids, carbon dioxide gas, etc. Examples of the microorganism include Lactobacillus rhamnosus (Lactobacillus rhamnosus). Lactobacillus rhamnosus ), Lactobacillus paracasei , Lactobacillus arizonensis , Saccharomyces cerevisiae ), Saccharomyces ellipsoides , Bacillus subtilis , Bacillus licheniformis and Leuconostoc mesenteroides ) and so on.

In an embodiment of the present invention, the microorganism for fermentation was a mixture of Lactobacillus rhamnosus and Lactobacillus paracasei , but is not limited thereto.

Water or an organic solvent may be used as a suitable solvent for obtaining the fermented extract of thistle and Moringa hot spring water from the fermented product, and any organic solvent may be used as long as it is a pharmaceutically acceptable organic solvent. For example, as the solvent, alone or two or more alcohols having 1 to 4 carbon atoms such as water, methanol, ethanol, propanol, isopropanol, butanol, etc. It can be mixed and used, and it is preferable to use water.

The extraction temperature is preferably 50 to 100 ℃. In addition, as the extraction method, various methods such as hot water extraction, cold sedimentation, reflux cooling extraction, solvent extraction, steam distillation, ultrasonic extraction, elution, compression, etc. may be used, but are not limited thereto, and preferably hot water extraction or reflux. It is to use the cooling extraction method.

The thistle and Moringa hot spring water fermentation extract may be chilled, heated and filtered at room temperature by a conventional method known in the art to obtain a liquid, or additionally, the solvent may be evaporated, spray dried, or freeze-dried. In addition, the thistle and Moringa hot spring water fermentation extract may be prepared in a powder state by additional processes such as distillation under reduced pressure and freeze drying or spray drying. In addition, the extract was obtained as an additionally purified fraction using various chromatography such as silica gel column chromatography, thin layer chromatography, high performance liquid chromatography, etc. I can.

In a specific example of the present invention, hot water was added to each of thistle and moringa to ferment, and the fermented product was extracted with hot water to obtain a fermented extract of each hot spring water. The thistle spring water fermented extract and Moringa spring water fermented extract were mixed to prepare a mixture of thistle and Moringa spring water fermented extract.

In an embodiment of the present invention, the thistle hot spring water fermented extract and Moringa hot spring water fermented extract are preferably mixed in a weight ratio of 1 to 2: 1.

The thistle and Moringa hot spring water fermented extract is preferably contained in an amount of 5 to 1000 μg/ml, more preferably 500 to 1000 μg/ml with respect to the cosmetic composition. If the content is less than 5 μg/ml, the antioxidant or anti-inflammatory effect may be insignificant, and if it exceeds 1000 μg/ml, the increase rate of the effect relative to the amount used may be low, which may be uneconomical.

In addition to the active ingredients, the cosmetic composition of the present invention may further include conventional auxiliary agents and carriers such as antioxidants, stabilizers, solubilizers, vitamins, pigments, fragrances, etc. that are commonly used in cosmetic compositions. For example, the cosmetic composition may further include auxiliary ingredients such as glycerin, butylene glycol, polyoxyethylene hydrogenated castor oil, tocopheryl acetate, citric acid, panthenol, squalane, sodium citrate, and allantoin. .

Since the cosmetic composition of the present invention is basically applied to the skin, it can be prepared in any formulation conventionally prepared with reference to the cosmetic composition in the art. For example, it may be formulated as a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation and spray, etc. It is not limited thereto. In more detail, it may be prepared in the form of a flexible lotion, a nutritional lotion, a nutritional cream, a massage cream, an essence, an eye cream, a cleansing cream, a cleansing foam, a cleansing water, a mask pack, a spray or a powder.

When the formulation of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tracant, cellulose derivatives, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, etc. may be included as carrier components. have.

When the formulation of the present invention is a powder or spray, lactose, talc, silica, aluminum hydroxide, calcium silicate, polyamide powder, etc. may be included as a carrier component. In particular, in the case of a spray, additional chlorofluorohydrocarbon, propane / It may contain a propellant such as butane and dimethyl ether.

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

When the formulation of the present invention is a suspension, as a carrier component, liquid diluents such as water, ethanol, and propylene glycol; Suspending agents such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester, and polyoxyethylene sorbitan ester; Microcrystalline cellulose, aluminum metahydroxide, bentonite, agar, tracant, and the like may be included.

When the formulation of the present invention is a surfactant containing cleansing, as a carrier component, aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, sarcosinate, fatty acid amide Ether sulfate, alkylamidobetaine, fatty alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative, ethoxylated glycerol fatty acid ester, and the like may be included.

According to another aspect of the present invention, the present invention provides an antioxidant or anti-inflammatory food composition comprising a mixture of fermented spring water extract of thistle and Moringa as an active ingredient.

The mixture of thistle and Moringa hot spring water fermented extract of the present invention can be used as a health functional food, food additive or dietary supplement.

When the mixture of thistle and Moringa hot spring water fermented extract is used as a food additive, the mixture may be added as it is or may be used in combination with other foods or food ingredients according to a conventional method.

In addition, the mixing amount of the mixture of thistle and Moringa hot spring water fermented extract may be suitably changed depending on the purpose of use (prevention, health, or therapeutic treatment), as well as 0.01 to 95% by weight based on the total weight of the food composition. It is preferable to be, and more preferably 1 to 80% by weight. When the content is less than 0.01% by weight, the antioxidant or anti-inflammatory effect may be insignificant, and when it exceeds 95% by weight, the increase rate of the effect relative to the amount of use is low, which may be uneconomical.

As a specific example, when preparing food or beverage, the mixture of thistle and Moringa hot spring water fermented extract of the present invention is added in an amount of 15% by weight or less, preferably 10% by weight or less based on the raw material. However, in the case of long-term intake for health and hygiene purposes or for health control purposes, it may be added in an amount below the above range, and since there is no problem in terms of safety, the active ingredient can be used in an amount above the above range. have.

There is no particular limitation on the type of food, but examples of foods to which the mixture of the thistle and Moringa hot spring water fermented extract of the present invention can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, etc. Noodles, gums, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, vitamin complexes, etc., and all health foods in the usual sense are included.

When the food composition of the present invention is prepared as a beverage, it may include additional ingredients such as various flavoring agents or natural carbohydrates, like a conventional beverage. The natural carbohydrates include monosaccharides such as glucose and fructose; Disaccharides such as maltose and sucrose; Natural sweeteners such as dextrin and cyclodextrin; Synthetic sweeteners such as saccharin and aspartame may be used. The natural carbohydrate is contained in an amount of 0.01 to 10% by weight, preferably 0.01 to 0.1% by weight, based on the total weight of the food composition of the present invention.

The food composition of the present invention includes various nutrients, vitamins, electrolytes, flavoring agents, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloidal thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohols, carbonic acid. It may include a carbonation agent used in beverages, and may include flesh for the manufacture of natural fruit juice, fruit juice beverage, and vegetable beverage, but is not limited thereto. These components may be used independently or in combination. The ratio of the additives is not largely limited, but is preferably contained within the range of 0.01 to 0.1% by weight based on the total weight of the food composition of the present invention.

In the case of long-term intake for the purpose of health and hygiene or for the purpose of health control, the food composition of the present invention can be taken for a long time because there is no problem in terms of safety.

According to another aspect of the present invention, the present invention provides a pharmaceutical composition for preventing or treating diseases caused by oxidative stress or inflammation, comprising a mixture of thistle and Moringa's hot spring water fermented extract as an active ingredient.

In the present invention, "disease caused by inflammation" is dermatitis, allergy, atopy, asthma, conjunctivitis, cataract, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, sepsis, gastric ulcer, gastritis, Crohn's disease, hemorrhoids, ankylosing spondylitis, Lupus, fibromyalgia, psoriasis, arthritis, osteoarthritis, rheumatoid arthritis, peri-shoulder joint infection, tendinitis, tendonitis, myositis, hepatitis, cystitis, nephritis, multiple sclerosis, diabetes, skin sclerosis, gout, neurodegenerative diseases, silicosis, atherosclerosis, ischemia, etc. I can.

In the present invention, the "disease caused by oxidative stress" may be heart disease, cancer, diabetes, cerebrovascular disease, arteriosclerosis, etc. among senile diseases, and in particular, oxidative stress is caused by the occurrence of vascular complications and age-related diseases in diabetes. It is recognized as an important factor.

The pharmaceutical composition for preventing or treating diseases caused by oxidative stress or inflammation of the present invention may be formulated and used in various forms according to a conventional method, respectively. For example, it may be formulated in oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, etc., and may be formulated in the form of external preparations, suppositories, and sterile injectable solutions. However, the composition of the present invention may be most preferably provided in the form of an external preparation for skin. Specifically, it may be used in the form of a liquid, ointment, cream, lotion, spray, patch, gel or aerosol.

In addition, a pharmaceutically acceptable carrier, excipient, and diluent may be further included according to each formulation. In addition, it can be formulated and used in the form of external preparations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, and sterile injectable solutions according to a conventional method, preferably creams, gels, patches, sprays, Ointment, oral, lotion, liniment, pasta, or cataplasma formulation may be used. For example, in the case of an external skin preparation that is used locally on the site, it may contain conventional additives, for example, a preservative, a solvent that aids the penetration of medicine, and an emollient in the case of ointments and creams. It may contain conventional carriers. Suitable formulations known in the art are preferably those disclosed in Remington's Pharmaceutical Science, recently, Mack Publishing Company, Easton PA, but are not limited thereto.

Examples of the carrier, excipient and diluent include lactose, dextrose, sucrose, oligosaccharide, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, Microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxy benzoate, propylhydroxy benzoate, talc, magnesium stearate, mineral oil, and the like. When formulating or formulating the pharmaceutical composition, it is prepared using diluents or excipients such as generally used fillers, extenders, binders, wetting agents, disintegrants, and surfactants. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and these solid preparations include at least one excipient in the composition, such as starch, calcium carbonate, and sucrose. , Lactose, gelatin, etc. are mixed and prepared. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral use include suspensions, liquid solutions, emulsions, syrups, etc.In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as humectants, sweeteners, fragrances, preservatives, etc. have. Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized formulations, suppositories, and the like. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate may be used. As a base for suppositories, witepsol, macrogol, tween 61, cacao butter, laurin paper, glycerogelatin, and the like may be used. The above ingredients may be added independently or in combination with the active ingredient, that is, a mixture of thistle and Moringa hot spring water fermented extract.

The term "administration" as used herein means providing a pharmaceutical composition of the present invention to an individual by any suitable method.

The pharmaceutical composition of the present invention is an amount of an active ingredient or pharmaceutical composition that induces a biological or medical response in a tissue system, animal or human, conceived by a researcher, veterinarian, doctor or other clinician, that is, the symptom of the disease or disorder being treated It can be administered in a therapeutically effective amount, which is an amount that induces remission. It is apparent to those skilled in the art that the therapeutically effective dosage and frequency of administration of the pharmaceutical composition of the present invention will vary depending on the desired effect. Therefore, the optimal dosage to be administered can be easily determined by those skilled in the art, and the type of disease, the severity of the disease, the amount of active ingredients and other ingredients contained in the composition, the type of formulation, the patient's age, weight, and general health condition. , Sex and diet, administration time, administration route and secretion rate of the composition, treatment period, it may be adjusted according to various factors, including drugs used simultaneously. The pharmaceutical composition of the present invention can be administered to a subject by various routes. For example, intravenous, intraperitoneal, intramuscular, intraarterial, oral, intracardiac, intramedullary, intrathecal, transdermal, intestinal, subcutaneous, sublingual or topical administration may be performed, but is not limited thereto.

The pharmaceutical composition of the present invention may be administered in an amount of 1 to 10,000 mg/kg/day, preferably 1 to 200 mg/kg/day, and may be administered once a day or divided into several times. have.

Hereinafter, the present invention will be described in more detail through examples. These examples are for illustrative purposes only, and it will be apparent to those of ordinary skill in the art that the scope of the present invention is not construed as being limited by these examples.

Materials and reagents

2,2-diphenyl-1-picryl-hydrazyl (DPPH), a reagent used in the electron donating ability measurement experiment, and potassium persulfate, a reagent for measuring ABTS+ radical scavenging ability, were manufactured by Sigma-Aldrich (St. Louis, MO, USA). ), and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sμlphonic acid) was purchased from Wako Pure Chemical Industries. Ltd. (Japan).

Macrophage Raw 264.7, a cell line used for cell culture and cytotoxicity measurement, was purchased from Korea Cell Line Bank. For cell culture, DMEM (dμlbecco's modified eagle medium) medium, penicillin/streptomycin, trypsin, phosphate buffered saline (PBS), and fetal bovine serum (FBS) were prepared in thermo scientific hyclone (USA). It was purchased and used. The dimethyl sulfoxide (DMSO) used to measure cytotoxicity was purchased from BioShop (Canada) and used, and 3-[4,5-dimethylthiazol]-2-yl]-2,5-diphenyl-tetrazolium bromide ( MTT) was purchased and used from Sigma-Aldrich (St. Louis, MO, USA).

The equipment used in the experiment was a vortexer (Scientific Industries, INC, USA), an autoclave (JS Research Inc, Korea), a freeze dryer (ILShin BioBase Co. Korea), a microplate reader (Tecan, Austria), and a rotary vacuum concentrator ( EYELA, Japan), CO ₂ incubator (Vision Scientific, Korea), microscope (Olympus, Japan), UV/VIS spectrophotometer (Hitachi, Japan), centrifuge (Hanil Science Industrial Co. Korea), Mini-PROTEAN® tetra cell (Bio-Rad, USA), Mini Trans-Blot® Cell (Bio-Rad, USA), microcentrifugation (gyrozen, Korea), digital mixer (Deihan Scientific, Korea) and Davinch-Chemi ^™ Imager CAS-400SM system (Davinch-K Co, Korea) was used.

Example 1. thistle and Moringa Preparation of mixture of fermented extract of hot spring water

A mixture of thistle and Moringa's spring water fermented extract was prepared in the same manner as in FIG. 1.

Specifically, the thistle used for extraction was pulverized into 1 to 3 cm of stems, leaves, and roots, and then pulverized once again to 1,300 mesh. The pulverized thistle was sterilized at 120° C. for 20 minutes using an autoclave. The sterilized thistle and Onyang hot spring water were mixed and homogenized in a ratio of 1:1 (v/v). The Onyang hot spring water was provided by the Asan First Tourist Hotel located in Asan. Activated Lactobacillus rhamnosus and Lactobacillus paracasei strains were inoculated with 10% of the milk thistle and hot spring water mixture, followed by fermentation at 37° C., 120 rpm and anaerobic conditions for 72 hours. Distilled water of 10 times the weight of the sample was added to the fermented product in an extraction flask equipped with a vertical reflux condenser, and then extracted three times at 100° C. for 60 minutes to obtain a thistle hot spring water fermented extract.

The Moringa hot spring water fermented extract was prepared using dried Moringa leaves, and was prepared in the same manner as the thistle hot spring water fermented extract.

A mixture of thistle and Moringa's hot spring water fermented extract (MCF) was prepared by mixing the prepared thistle hot spring water fermented extract and Moringa hot spring water fermented extract at a ratio of 1:1 (w/w).

Example 2. Thistle and Moringa Of a mixture of fermented extracts of hot spring water Electronic donation ability Measure

Electron donating abilities (EDA) was tested by modifying Blois' method. 60 μl of DPPH solution (Sigma, USA), 120 μl of mixture of thistle and Moringa hot spring water fermented extracts (5, 10, 50, 100, 500 and 1000 μg/ml) by concentration, and reacted for 15 minutes, followed by microplate reader Absorbance was measured at 517 nm using. The electron donating ability was expressed as the absorbance reduction rate of the added and non-added sample solutions, and was calculated as shown in Equation 1 below. The result of measuring the electron donating ability of the mixture of thistle and Moringa hot spring water fermented extract is shown in FIG. 2.

[Equation 1]

As shown in FIG. 2, as the concentration of the mixture of thistle and Moringa hot spring water fermented extract increased, the electron donating ability increased, and showed an effect of 77.8% at a concentration of 1,000 μg/ml. Comparing this with the control vitamin C, it can be seen that the electron donating ability of the mixture of thistle and Moringa hot spring water fermented extract was significantly higher than that of vitamin C, which showed 56.8% at the same concentration.

Example 3. Thistle and Moringa Of a mixture of fermented extracts of hot spring water ABTS Radical Erasing ability Measure

The ABTS radical scavenging ability assay (ABTS cation decolorization assay) is a method of measuring antioxidant power using ABTS radicals. Specifically, 7mM 2,2-azino-bis (3-ethyl-benthiazoline-6-sμlfonic acid) and 2.45mM potassium persulfate (potassium persμlfate) were mixed and then reacted at room temperature for 24 hours to generate ^{ABTS +. Thereafter, 100 μl of a sample was added to ABTS + 100} μl diluted with ethanol, and the absorbance was measured at 700 nm. The sample was a mixture (5, 10, 50, 100, 500 and 1000 μg/ml) of thistle and Moringa fermented extracts by concentration, and BHT was used as a control. The ABTS scavenging ability was calculated as in Equation 2 below, The results are shown in FIG. 3.

[Equation 2]

As shown in FIG. 3, the ABTS radical scavenging ability of the mixture of thistle and Moringa hot spring water fermented extract increased in a concentration-dependent manner, and it was confirmed that 95% scavenging ability was exhibited, particularly at a concentration of 1,000 μg/ml. This is a significant result when compared to the control group BHT.

Example 4. Thistle and Moringa Measurement of cytotoxicity of mixtures of fermented extracts from hot spring water

4-1. Cell line and cell culture

In this example, raw 264.7 cells as macrophages and B16F10 cells as melanoma cells were used, and the medium for culturing the cells included 10% FBS and 1% penicillin/streptomycin (penicillin/streptomycin, 100 U/ml). DMEM medium was used. In addition, the cultivation of the Raw 264.7 cells was performed in an incubator at a _{temperature of 37°C and 5% CO 2.}

4-2. MTT analysis

MTT analysis was conducted according to Carmichael's method. ^{1x10 5} in a 96 well plate Raw264.7 cells, cells/well, were dispensed at 180 μl each. 20 μl of a mixture (5, 10, 50, 100, 500, and 1000 μg/ml) of thistle and Moringa hot spring water fermentation extract by concentration was added to the dispensed cells, followed by _{incubation in a 37° C., 5% CO 2} incubator for 24 hours. To this, 40 μl of the MTT solution prepared at a concentration of 2.5 mg/ml was further added and incubated for 4 hours. After removing the culture solution, 100 μl of DMSO was added to each well and reacted at room temperature for 10 minutes, and then absorbance was measured at 540 nm with an ELISA reader. The cytotoxicity measurement was expressed as the absorbance reduction rate of the mixture of thistle and Moringa hot spring water fermented extract and the non-added group, and was calculated as shown in Equation 3 below. The results of MTT analysis of Raw264.7 cells for a mixture of thistle and Moringa hot spring water fermented extract are shown in FIG. 4.

[Equation 3]

As shown in Fig. 4, the experimental group treated with a mixture of thistle and Moringa hot spring water fermented extract at a concentration of 100 μg/ml in Raw264.7 cells confirmed that the cell viability was 90.4% or more.

In addition, the cytotoxicity of a mixture of thistle and Moringa hot spring water fermented extract in melanoma cells B16F10 cells was measured in the same manner as in Raw264.7 cells. The results of measuring cytotoxicity in melanoma cells are shown in FIG. 5.

As shown in FIG. 5, the experimental group treated with a mixture of thistle and Moringa hot spring water fermented extract at a concentration of 100 μg/ml to melanoma cells B16F10 cells was confirmed to have a cell viability of 94.8% or more.

Example 5. Measurement of nitric oxide (NO) production inhibitory activity

The amount of nitric oxide (NO) produced from Raw264.7 cells was measured according to the method of Green et al. Specifically, it was measured in the form of _{NO 2} present in the cell culture solution using the griess reagent. Raw 264.7 cells were dispensed into a 6-well plate at 2×10 ⁵ cells/ml, and then _{cultured in a 37° C. CO 2} incubator for 24 hours. After washing twice with 1X PBS, 20 μg/ml of LPS (Lipopolysaccharide) was treated and incubated for 2 hours. After that, a mixture (5, 10, 50 and 100 μg/ml) of thistle and Moringa hot spring water fermented extract prepared by concentration was treated and cultured for 24 hours. After incubation, the supernatant was added with the same amount of griess reagent and reacted in a 96-well plate for 10 minutes, and the absorbance at 540 nm was measured. The measurement of the NO inhibitory activity was expressed as the absorbance reduction rate of the sample-added group and the non-added group, and was calculated as in Equation 4 below. The results of measuring the NO production inhibitory activity of the mixture of thistle and Moringa hot spring water fermented extract are shown in FIG. 6.

[Equation 4]

As shown in Figure 6, it can be seen that the treatment of a mixture of thistle and Moringa hot spring water fermented extract 5 to 100 μg/ml inhibits NO generation. In particular, it can be seen that the NO concentration was 57.1% in the group treated with 100 μg/ml of the mixture of thistle and Moringa hot spring water fermented extract, which showed excellent NO production inhibitory activity.

Example 6. Western Blotting through Inflammatory factor Protein expression measurement

6-1. iNOS

In order to confirm the expression of the inflammatory factor iNOS protein, Raw264.7 cells were seeded in a 100 mm tissue culture dish and cultured for 24 hours. After removing the medium, the mixture of thistle and Moringa hot spring water fermented extract was treated by concentration (25, 50 and 100 μg/ml), incubated for 24 to 48 hours, and washed twice with PBS. To the washed cells, 100 μl of a solution obtained by adding 1 tab of complete mini to 10 ml of RIPA buffer was added, followed by centrifugation at 4° C. and 13,200 rpm for 20 minutes to obtain a supernatant. The supernatant was quantified using a BCA protein assay kit. 20 μl of the quantified protein was separated by electrophoresis on a 10% SDS gel, and the separated protein was transferred to a PVDF membrane. Blocking buffer (5% skim milk in TBST) was added to the PVDF membrane to perform blocking at room temperature for 1 hour. The primary antibody was diluted and reacted overnight at 4° C., and then washed three times with TBST buffer at 10 minute intervals. The secondary antibody diluted 1:1,000 on the washed PVDF membrane was reacted for 2 hours at room temperature, washed 3 times, and the band was confirmed using a ^{Davinch-Chemi TM Imager CAS-400SM instrument.} Western blotting results of iNOS protein are shown in FIG. 7.

As shown in Figure 7, the mixture of thistle and Moringa hot spring water fermented extract reduced the expression of the inflammatory factor iNOS in a concentration-dependent manner. In particular, it was confirmed that the 50 and 100 μg/ml treatment groups of thistle and Moringa hot spring water fermented extracts significantly inhibited iNOS protein expression compared to the control vitamin C treatment group.

6-2. COX-2

The expression of another inflammatory factor, COX-2 protein, was measured in the same manner as in Example 6-1. The result of measuring the expression of COX-2 protein is shown in FIG. 8.

As shown in Fig. 8, it can be seen that the mixture of thistle and Moringa hot spring water fermented extract reduces the expression of the inflammatory factor COX-2 in a concentration-dependent manner.

Overall, the present inventors confirmed that the mixture of thistle and Moringa hot spring water fermented extract has low cytotoxicity, excellent electron donating ability and radical scavenging ability, and inhibits the expression of inflammatory factor proteins. This means that the mixture of thistle and Moringa hot spring water fermented extract of the present invention has excellent antioxidant and anti-inflammatory effects, and the mixture of thistle and Moringa hot spring water fermented extract of the present invention can be used in various ways in the field of food and cosmetics. have.

Hereinafter, the present invention will be described in more detail through formulation examples. Formulation examples are for illustrative purposes only, and are not to be construed as limiting the scope of the present invention.

Formulation example One. Cosmetic Preparation of the formulation

1-1. Manufacture of flexible cosmetic water

Mixture of thistle and Moringa hot spring water fermentation extract 0.1% by weight, 1,3-butylene glycol 5.2% by weight, oleyl alcohol 1.5% by weight, ethanol 3.2% by weight, polysorbate 20 3.2% by weight, benzophenone-9 2.0% by weight %, 1.0% by weight of carboxylvinyl polymer, 3.5% by weight of glycerin, a trace amount of fragrance, a trace amount of preservative, and the remaining amount of purified water were mixed to prepare a flexible cosmetic water by a conventional method.

1-2. Milk lotion Produce

Mixture of thistle and Moringa spring water fermentation extract 0.1% by weight, glycerin 5.1% by weight, propylene glycol 4.2% by weight, tocopheryl acetate 3.0% by weight, liquid paraffin 4.6% by weight, triethanolamine 1.0% by weight, squalane 3.1% by weight, macadamia nut 2.5% by weight of oil, 1.6% by weight of polysorbate, 1.6% by weight of sorbitan sesquirolate, 0.6% by weight of propylparaben, 1.5% by weight of carboxylvinyl polymer, a trace amount of fragrance, a trace amount of preservative, and the remaining amount of purified water are mixed Milk lotion was prepared by a conventional method.

1-3. Nutrition cream manufacturing

Mixture of thistle and Moringa spring water fermentation extract 0.5% by weight, glycerin 4.0% by weight, petrolatum 3.5% by weight, triethanolamine 2.1% by weight, liquid paraffin 5.3% by weight, squalane 3.0% by weight, beeswax 2.6% by weight, tocopheryl acetate 5.4% by weight %, polysorbate 60 3.2% by weight, carboxylvinyl polymer 1.0% by weight, sorbitan sesquioleate 3.1% by weight, a trace amount of fragrance, a trace amount of preservative and the remaining amount of purified water were mixed to prepare a nutrient cream by a conventional method. .

1-4. Massage cream manufacturing

Mixture of thistle and Moringa hot spring water ferment extract 0.5% by weight, glycerin 4.0% by weight, petrolatum 3.5% by weight, triethanolamine 0.5% by weight, liquid paraffin 24.0% by weight, squalane 3.0% by weight, beeswax 2.1% by weight, tocopheryl acetate 0.1% by weight %, polysorbate 60 2.4% by weight, carboxylvinyl polymer 1.0% by weight, sorbitan sesquioleate 2.3% by weight, a trace amount of fragrance, a trace amount of preservative and the remaining amount of purified water were mixed to prepare a massage cream by a conventional method. .

1-5. For cleaning Body cleanser Produce

Mixture of thistle and Moringa hot spring water fermentation extract 1g, anionic surfactant 18g, nonionic surfactant 5g, glycerin 7g, sodium chloride 3g, natural olive liquid soap 1.5g, fragrance 1g and water 100g for cleaning by a conventional method A body cleanser was prepared.

Above, a specific part of the present invention has been described in detail, and for those of ordinary skill in the art, it is obvious that this specific technique is only a preferred embodiment, and the scope of the present invention is not limited thereby. something to do. Therefore, it will be said that the practical scope of the present invention is defined by the appended claims and their equivalents.

Claims (9)

It contains a mixture of thistle and Moringa's hot spring water fermented extract as an active ingredient,
The mixture is thistle spring water fermented extract and Moringa spring water fermented extract is mixed in a weight ratio of 1 to 2: 1, antioxidant and anti-inflammatory cosmetic composition.
The method of claim 1,
Thistle and Moringa spring water fermentation extract is extracted with one or more solvents selected from water, alcohols having 1 to 4 carbon atoms, and a mixed solvent thereof, an antioxidant and anti-inflammatory cosmetic composition.
The method of claim 1,
The thistle is one or more parts selected from the group consisting of stems, leaves and roots, antioxidant and anti-inflammatory cosmetic composition.
The method of claim 1,
The Moringa is Moringa leaf, an antioxidant and anti-inflammatory cosmetic composition.
The method of claim 1,
The fermentation is by Lactobacillus rhamnosus (Lactobacillus rhamnosus) and Lactobacillus paracasei (Lactobacillus paracasei), an antioxidant and anti-inflammatory cosmetic composition.
It contains a mixture of thistle and Moringa's hot spring water fermented extract as an active ingredient,
The mixture is thistle spring water fermented extract and Moringa spring water fermented extract is mixed in a weight ratio of 1 to 2: 1, antioxidant and anti-inflammatory food composition.
In the pharmaceutical composition for antioxidant and anti-inflammatory,
The composition contains a mixture of thistle and Moringa's hot spring water fermented extract as an active ingredient,
The mixture is a mixture of thistle hot spring water fermented extract and Moringa hot spring water fermented extract in a weight ratio of 1 to 2: 1,
The antioxidant is the prevention or treatment of one or more diseases selected from the group consisting of senile heart disease, cancer, diabetes, cerebrovascular disease and arteriosclerosis,
The anti-inflammatory diseases include dermatitis, allergy, atopy, asthma, conjunctivitis, cataract, periodontitis, rhinitis, otitis media, pharyngitis, tonsillitis, pneumonia, sepsis, gastric ulcer, gastritis, Crohn's disease, hemorrhoids, ankylosing spondylitis, lupus, fibromyalgia, psoriasis, arthritis, Prevention of one or more diseases selected from the group consisting of osteoarthritis, rheumatoid arthritis, peri-shoulder joint inflammation, tendinitis, tendonitis, myositis, hepatitis, cystitis, nephritis, multiple sclerosis, diabetes, scleroderma, gout, neurodegenerative disease, silicosis, atherosclerosis and ischemia Or the therapist,
Pharmaceutical composition for antioxidant and anti-inflammatory.
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