KR20120048208A - A composition for skin protection including skin aging, anti-inflammatory comprising the extracts of lactarius hatsudake as an active ingredients - Google Patents

Abstract

PURPOSE: An anti-inflammatory composition containing Lactarius hatsudake extract or fraction thereof is provided to suppress production or secretion of interferon-gamma and to prevent or treat inflammatory diseases without cytotoxicity. CONSTITUTION: A cosmetic composition for anti-aging, immunity enhancement, or inflammatory diseases prevention or treatment contains 0.01-50 weight parts of Lactarius hatsudake extract or fraction thereof as an active ingredient. The extract is prepared using water, alcohol of C1-C2, or mixture thereof. The fraction is prepared using normal-hexane, ethyl acetate, normal-butanol, and water in order. The cosmetic composition is manufactured in the form of emulsion, cream, essence, body lotion, body gel, body essence, body cleanser, cleansing foam, pack, or cleansing cream.

Description

A composition for skin protection including skin aging, anti-inflammatory comprising the extracts of Lactarius hatsudake as an active ingredients}

The present invention is a mushroom fungus ( Lactarius) hatsudake ) relates to an anti-aging, skin protection and anti-inflammatory composition containing the extract and fractions thereof as an active ingredient.

Inflammatory disease refers to a pathological state of abscesses formed by invasion of bacteria or physical and chemical stimulation by the internal and external environment. The inflammatory diseases include dermatitis, atopy, psoriasis, osteoarthritis, rheumatoid arthritis, gout, ankylosing spondylitis, tendonitis, tendonitis, rheumatic fever, lupus, fibromyalgia, psoriatic arthritis, asthma, atopic dermatitis, Crohn's disease, ulcerative colitis, etc. Acute and chronic inflammatory diseases. Inflammation is a manifestation of normal and protective in vivo defense mechanisms that are localized to tissue damage caused by physical trauma, harmful chemicals, microbial infections or irritants in metabolites in vivo. This inflammation is triggered by chemical mediators such as various cytokines produced from damaged tissues and migrating cells, and these chemical mediators are known to vary according to the type of inflammatory process. In normal cases, the living body restores normal structure and function by neutralizing or eliminating pathogens and regenerating upper and lower tissues through inflammatory reactions. Otherwise, the living body may progress to a disease state such as chronic inflammation. In addition, in case of improperly triggered inflammation by harmless substances such as pollen or autoimmune reactions such as asthma and rheumatoid arthritis, the defense reaction itself damages the tissues, thus preventing or treating inflammatory diseases. In the case of skin, the enzymes used to treat and remove inflammation in most inflammatory reactions destroy and damage surrounding normal tissues, thereby destroying the collagen fibers and elastic fibers of the skin, thereby aging the skin. In almost all clinical diseases, inflammatory reactions can be observed, and some of these inflammatory diseases are bacterial diseases that can be causatively treated with antibiotics, but most of them are due to tissue damage caused by autoimmune reactions. It is known as intractable disease.

The most common inflammatory disease preventing or treating agents for treating such inflammatory diseases are largely divided into steroidal and nonsteroidal inflammatory disease preventing or treating agents. Of these, most synthetic inflammatory disease preventing or treating agents have various side effects in addition to the main action. In many cases it is accompanied by excellent effects and development of a inflammatory disease prevention or treatment agent with fewer side effects is urgently required. In particular, in terms of efficacy and side effects, natural product formulations, which have a long history of clinical experience and excellent evaluation in terms of safety, are considered to be good candidates for the development of preventive and therapeutic agents for inflammatory diseases.

Various biochemical phenomena are involved as a cause of inflammation in living bodies. Macrophage is a multi-functional cell that plays an important role in the inflammatory response by generating various cytokines and NOx by chemical stimulation. In particular, inducible nitric oxide synthase (iNOS) expressed by cytokine stimulation such as interferons-gamma (IFN-γ) in macrophages produces large amounts of nitric oxide (NO) for a long time. This oxidative stress is known to promote NF-κB activity, a transcription factor of the inflammatory response inhibited by IκB. Activated NF-κB is known to promote the expression of genes that induce inflammatory responses, such as iNOS, COX-2, and various cytokines such as IL-1β or TNF-α. It is known to inhibit the inflammatory response (Baeuerle et al., Annu. Rev. Immunol., 12: 141-179, 1994). Inflammation of the skin is caused by UV rays, dryness, contaminants, and the like, and these factors cause collagen fibers and elastic fibers to break down, resulting in dry skin or deepening of wrinkles.

Nitric oxide (NO) is produced from L-arginine by three major nitrogen oxide synthase (NOS) isomers, neuronal NOS (nNOS), endothelial NOS (eNOS), and inducible NOS (iNOS). nNOS and eNOS are regulated by Ca2 + / calmodulin, but iNOS is regulated at the transcription level by inflammatory stimuli such as interleukin, interferon, LPS. Small amounts of NO produced by nNOS or eNOS are responsible for normal physiological functions such as vasodilation, neurotransmission, and cellular destruction of pathogens, whereas NO produced by iNOS in macrophages is associated with various pathophysiology, including inflammation and cancer. It is involved in the process and reacts with superoxide to form peroxynitrite, which acts as a powerful oxidant, damaging cells and activating NF-κB in macrophages activated by inflammatory stimuli. It is known to be involved in chronic diseases such as inflammatory reactions, cancer, and atherosclerosis (Lawrence et al., Nat Med., 7: 1291-1297, 2001; Riehemann et al., FEBS Lett., 442: 89-94, 1999 Stamler et al., Science, 258: 1898-1902, 1992).

Interferon (IFN), on the other hand, refers to cytokines important for the resistance of mammalian hosts to pathogens. The three main functions of interferon are antiviral, antiproliferative, and immunomodulatory functions. The antiviral action of interferon is used to treat viral hepatitis, and the antiproliferative and immunomodulatory actions are used to treat cancer.

In particular, interferon gamma (IFN- [gamma]) is well known as immune or type II IFN and is a homodimer produced rapidly by activated T cells and natural killer (NK) cells. In addition, it plays a crucial role in the treatment of innate and adaptive immunity and tumor therapy against intracellular pathogens. The importance of IFN- [gamma] in the immune system directly begins with the inhibitory effect of viral replication, but most importantly, IFN- [gamma] has immunostimulatory and immunomodulatory effects. In addition to the central role of INF-γ in host defense, aberrant expression of IFN-γ is associated with autoinflammatory and autoimmune diseases. Excessive secretion of IFN-γ is associated with the development of chronic inflammatory and immune diseases. Therefore, the search for a substance that inhibits IFN-γ, an inflammatory cytokine, may prove to be an effective prophylactic or therapeutic substance for inflammatory diseases.

Lactarius belonging to the Russulaceae family of Basidiomycete hatsudake ) is a common edible mushroom widely distributed in Korea, China, Japan, Europe, and North America. It grows or grows on land in red pine forests in summer and autumn. The shape of the shade is 3 ~ 12 ㎝ in diameter, light light brown, and has a dark concentric pattern on the surface. It is round at first and then spreads slowly, and later it becomes a funnel. The meat is crumbly, but its flavor and taste are good. It is a mushroom that has been enjoyed by Japanese people since early autumn, along with early autumn mushrooms. Currently, L. Although the biological activity of antitumor and antiviral effects of hatsudake has been reported, there have been few reports on anti-inflammatory action through IFN-γ inhibition.

Thus, the inventors of the present invention while searching for a novel natural material derived from excellent anti-inflammatory activity, Lactarius hatsudake ) extracts or fractions thereof have the ability to inhibit the production of interferon-gamma (IFN-γ), which is an excellent inflammatory cytokine. The invention was completed.

An object of the present invention is Lactarius hatsudake ) To provide an anti-aging, immune-enhancing and anti-inflammatory composition containing an extract or a fraction thereof as an active ingredient.

In order to achieve the above object, the present invention provides a cosmetic composition for anti-aging, fortifying immunity, or preventing and improving inflammatory diseases containing 0.01 to 50 parts by weight of Lactarius hatsudake extract or fractions thereof as an active ingredient. do.

The present invention also provides a topical skin preparation for the prevention and improvement of an inflammatory disease containing a milk mushroom fungus extract or a fraction thereof as an active ingredient.

In addition, the present invention provides a pharmaceutical composition for the prevention and treatment of inflammatory diseases containing milk mushroom fungus extract or fractions thereof as an active ingredient.

In addition, the present invention provides a health food for the prevention and improvement of inflammatory diseases containing milk mushroom fungus extract or fractions thereof as an active ingredient.

Lactarius milk mushroom of the present invention hatsudake ) extract or fractions thereof inhibits the production of inflammatory cytokines Interferon-gamma and inhibits the cytotoxicity of skin cells due to the lack of cytotoxicity. It can be usefully used as an active ingredient of the composition, it can be usefully used as a composition for the prevention and treatment of inflammatory diseases.

1 is a diagram showing the inhibitory activity of IFN-γ production of NK92 cells treated with the extract of the fungus extract of the present invention or fractions thereof.
Figure 2 is a diagram showing the cytotoxicity of NK92 cells treated with milk mushroom fungus extract of the present invention or fractions thereof.

Hereinafter, the present invention will be described in detail.

The present invention is a mushroom fungus ( Lactarius) hatsudake ) provides an anti-aging, immune-enhancing, or inflammatory disease prevention and a cosmetic composition containing 0.01 to 50 parts by weight of the extract as an active ingredient.

In another aspect, the present invention provides a cosmetic composition for anti-aging, fortifying immunity, or for preventing and improving inflammatory diseases, containing 0.01 to 50 parts by weight of the fractions separated from the milk mushroom fungus extract as an active ingredient.

The inflammatory disease or immune disease is preferably any one selected from the group consisting of edema, dermatitis, allergy, atopy, acne, pruritus, multiple sclerosis, acute inflammatory disease and chronic inflammatory disease.

The cosmetic composition is preferably, but not limited to, improving or preventing wrinkles and aging resulting from inflammatory diseases.

Milk mushroom fungus extract of the present invention is preferably prepared in the following steps, but is not limited thereto;

1) extracting milk mushroom fungus by adding an extraction solvent;

2) filtering the extract of step 1); And

3) preparing the extract by concentrating the filtered extract of step 2) under reduced pressure.

In the production method, the mushroom fungus of step 1) can be used without limitation, such as cultivated, collected or commercially available. The milk mushroom fungus can use the whole plant, it is preferable to use fruiting body, but is not limited thereto.

In the preparation method, the extraction solvent of step 1) may be used any extraction solvent commonly used in the art, water, alcohol or mixtures thereof, preferably C ₁ to C ₂ lower alcohol or It is preferable to use a solvent selected from mixed solvents, and the lower alcohol of C ₁ -C ₂ is more preferably ethanol or methanol, but is not limited thereto. The amount of the extraction solvent is preferably 1 to 20 times the dry weight of the milk mushroom ash ratio, more preferably 10 times, but is not limited thereto. The extraction method may be an extraction method such as hot water extraction, immersion extraction, reflux cooling extraction and ultrasonic extraction, and may be preferably extracted once to five times by the ultrasonic extraction method. When the extraction temperature is preferably 10 ℃ to 100 ℃ and more preferably room temperature, but is not limited thereto. The extraction time is preferably 1 day to 7 days, more preferably 3 days, but is not limited thereto.

In the above production method, the extraction method is a method using an extraction device such as supercritical extraction, subcritical extraction, high temperature extraction, high pressure extraction or ultrasonic extraction method or a method using an adsorption resin containing XAD and HP-20. Conventional extraction methods may be used, and heating is preferably performed at reflux extraction or extraction at room temperature, but is not limited thereto. The extraction number is preferably 1 to 5 times, more preferably 3 times repeated extraction is not limited thereto.

In the production method according to the present invention, the concentration of the obtained extract is preferably a vacuum rotary evaporator of reduced pressure, but is not limited thereto. In addition, the drying is preferably dried under reduced pressure, vacuum drying, boiling drying, spray drying, room temperature drying or lyophilization.

The fraction of the milk mushroom fungus extract is preferably prepared by a manufacturing method comprising the step of preparing an organic solvent fraction by adding an organic solvent to the milk mushroom fungus extract, but is not limited thereto.

In the method for preparing the fraction, the organic solvent may be used any extraction solvent commonly used in the art, and anhydrous or hydrous lower alcohol having 1 to 4 carbon atoms (methanol, ethanol, propanol, butanol, normal-propanol, Iso-propanol or normal-butanol and the like), a mixed solvent of the lower alcohol and water, acetone, ethyl acetate, chloroform, 1,3-butylene glycol, hexane, diethyl ether or butyl acetate, and the like. The organic solvent is preferably normal-hexane, ethyl acetate or normal-butanol, but is not limited thereto.

Normally, hexane, ethyl acetate, butanol, and water are sequentially added to the milk mushroom ash extract in order of low polarity to obtain fractions of the organic solvent layers, and the fractions are fractionated using a separatory funnel. But is not limited thereto. All fractions of each organic solvent layer are available.

The fraction may be obtained by repeating the fractionation process from 1 to 5 times, preferably 3 times, from the extract of the mushroom fungus, preferably concentrated under reduced pressure after the fraction, but is not limited thereto.

In the present invention, the extract of the fruiting bodies of the Lactarius hatsudake with methanol to prepare a methanol extract of the mushroom fungi, suspended in water to add Norman-hexane and fractionate the Norman-hexane layer To obtain a normal-hexane fraction, and then fractionated and ethyl acetate was added to the remaining water layer, and then the ethyl acetate layer was separated to obtain an ethyl acetate fraction, and normal-butanol fraction was added to the remaining water layer. Obtained.

The cosmetic composition is preferably contained in 0.1 to 50 parts by weight of milk mushroom fungus extract or a fraction thereof based on the total weight, but is not limited thereto.

The cosmetic composition may be prepared in one or more formulations selected from the group consisting of emulsion, cream, essence, essence, body lotion, body gel, body essence, body cleaner, cleansing foam, pack and cleansing cream.

The cosmetic composition is, for example, emulsions, suspensions, microemulsions, microcapsules, microgranules or ionic (liposomes) obtained by dispersing an oil phase in a solution, a gel, a solid or dough anhydrous product, an aqueous phase, and a nonionic vesicle dispersant. It may be provided in the form of a cream, skin, lotion, powder, ointment, spray or cone stick. It may also be prepared in the form of a foam or in the form of an aerosol composition further containing a compressed propellant.

In addition, the cosmetic composition is in addition to the extract of milk mushroom fungus of the present invention or fractions thereof, fatty substances, organic solvents, solubilizers, thickening and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents (foaming agents), Fragrances, surfactants, water, ionic or nonionic emulsifiers, fillers, metal ion sequestrants and chelating agents, preservatives, vitamins, blockers, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic active agents, lipid vesicles or It may contain adjuvants conventionally used in the cosmetic field, such as any other ingredient conventionally used in cosmetics.

The inventors of the present invention, the present invention to the NK92 cells (human NK lymphoma), which is interferon-gamma (IFN-γ) -dependent natural killer cell line, in order to determine whether the milk mushroom fungus extract of the present invention or its fraction has anti-inflammatory activity Extracts or fractions thereof were treated at various concentrations to determine the production of IFN-γ, one of the inflammatory cytokines, on the cells. As a result, the production of IFN- [gamma] was reduced in a concentration-dependent manner of the present invention (see FIG. 1). Moreover, as a result of confirming cytotoxicity through WST-1 analysis, it was confirmed that the milk mushroom fungus extract or its fraction is a substance with little cytotoxicity (see FIG. 2).

Therefore, the extract of milk mushroom fungus or fractions thereof has the activity of inhibiting the production of inflammatory cytokines without cytotoxicity, and thus it can be seen that there is an excellent anti-inflammatory action, an active ingredient of anti-aging, immune enhancing or anti-inflammatory cosmetic composition It can be usefully used as.

The present invention also provides a topical skin preparation for the prevention and improvement of an inflammatory disease containing a milk mushroom fungus extract or a fraction thereof as an active ingredient.

The inflammation may include chronic inflammation that has progressed to a disease state, an inflammatory response by a generally harmless substance such as pollen, or an autoimmune response such as asthma or rheumatoid arthritis.

The inflammatory diseases include edema, dermatitis, allergies, atopic, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer, gastritis, Crohn's disease, ulcerative colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatic fever, lupus fever , Fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, periarthritis, tendinitis, hay salt, peritonitis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome, multiple sclerosis, and various acute and chronic inflammatory diseases It is preferably one selected from the group consisting of, but is not limited thereto.

The external preparation for the skin is preferably, but not limited to, 0.1 to 50 parts by weight of the extract of milk mushroom fungi or fractions thereof based on the total weight.

The external preparations for the skin may further comprise fatty substances, organic solvents, solubilizers, thickening and gelling agents, emollients, antioxidants, suspending agents, stabilizers, foaming agents, fragrances, surfactants, water, ionic or nonionics. With emulsifiers, fillers, metal ion sequestrants and chelating agents, preservatives, vitamins, blockers, wetting agents, essential oils, dyes, pigments, hydrophilic or lipophilic actives, lipid vesicles or any other ingredients commonly used in external preparations for the skin. It may contain adjuvants commonly used in the same dermatological field. In addition, the ingredients may be introduced in amounts generally used in the field of dermatology.

Milk mushroom fungus extract of the present invention effectively inhibits IFN-γ production of inflammatory cytokines and is a safe substance that does not have cytotoxicity to cells, and as an active ingredient of an external preparation for skin that can prevent or improve inflammatory diseases. It can be usefully used.

In addition, the present invention provides a pharmaceutical composition for the prevention and treatment of inflammatory diseases containing milk mushroom fungus extract or fractions thereof as an active ingredient.

The inflammatory diseases include dermatitis, allergy, atopic dermatitis, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer, gastritis, Crohn's disease, colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatoiditis, lupus, fibromyalgia ), Psoriatic arthritis, osteoarthritis, rheumatoid arthritis, shoulder periarthritis, tendinitis, hay salt, periarthritis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome, multiple sclerosis, and acute and chronic inflammatory diseases It is preferably one, but is not limited thereto.

The pharmaceutical composition containing the milk mushroom fungus extract or a fraction thereof as an active ingredient may be used in the form of a general pharmaceutical preparation.

The pharmaceutical composition may further include a pharmaceutically acceptable additive, wherein the pharmaceutically acceptable additive may include starch, gelatinized starch, microcrystalline cellulose, lactose, povidone, colloidal silicon dioxide, calcium hydrogen phosphate, and lactose. , Mannitol, malt, gum arabic, pregelatinized starch, corn starch, powdered cellulose, hydroxypropyl cellulose, opiodry, sodium starch glycolate, lead carnauba, synthetic aluminum silicate, stearic acid, magnesium stearate, aluminum stearate, calcium stearate , Sucrose, dextrose, sorbitol, talc and the like can be used. The pharmaceutically acceptable additive according to the present invention is preferably included 0.1 to 90 parts by weight based on the composition.

The pharmaceutical composition may be administered in various dosage forms, oral and parenteral, in actual clinical administration, and when formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, surfactants, etc., which are commonly used, may be used. It is prepared by. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and the solid preparations may contain at least one excipient such as starch, calcium carbonate, sucrose, or the like. (Sucrose) or lactose (Lactose), gelatin, etc. are mixed and prepared. In addition to simple excipients, lubricants such as magnesium styrate talc are also used. Oral liquid preparations include suspensions, solvents, emulsions, and syrups, and may include various excipients, such as wetting agents, sweeteners, fragrances, and preservatives, in addition to commonly used simple diluents such as water and liquid paraffin. . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. As the non-aqueous solvent and the suspension solvent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like can be used. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

The pharmaceutical composition may be administered orally or parenterally according to a desired method, and when administered parenterally, external skin or intraperitoneal injection, rectal injection, subcutaneous injection, intravenous injection, intramuscular injection or intrathoracic injection It is preferable to select. Dosage varies depending on the patient's weight, age, sex, health condition, diet, time of administration, method of administration, rate of excretion and severity of disease.

The dosage of the pharmaceutical composition varies according to the weight, age, sex, health condition, diet, time of administration, method of administration, excretion rate and severity of the disease of the patient, the daily dosage is the extract of the present invention or It is 0.0001 to 100 mg / kg, preferably 0.001 to 10 mg / kg, based on the amount of its fraction, and may be administered 1 to 6 times a day.

The pharmaceutical composition of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy and biological response modifiers for the prevention and treatment of inflammatory diseases.

The fungus extract of the present invention or a fraction thereof is a safe substance that effectively inhibits the production of inflammatory cytokines IFN-γ and does not have cytotoxicity to cells, and thus is useful as an active ingredient of a pharmaceutical composition that can prevent or treat inflammatory diseases. Can be used.

In addition, the present invention provides a health food for the prevention and improvement of inflammatory diseases containing milk mushroom fungus extract or fractions thereof as an active ingredient.

The inflammatory diseases include dermatitis, allergy, atopic dermatitis, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer, gastritis, Crohn's disease, colitis, hemorrhoids, gout, ankylosing spondylitis, rheumatoiditis, lupus, fibromyalgia ), Psoriatic arthritis, osteoarthritis, rheumatoid arthritis, shoulder periarthritis, tendinitis, hay salt, periarthritis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome, multiple sclerosis, and acute and chronic inflammatory diseases It is preferably one, but is not limited thereto.

Lactobacillus extract of the present invention or a fraction thereof may be added as it is, or used with other foods or food ingredients, and may be appropriately used according to conventional methods.

The health food of the present invention includes ingredients that are commonly added in food production, and include, for example, proteins, carbohydrates, fats, nutrients and seasonings.

There is no particular limitation on the kind of food. Examples of foods to which the milk mushroom extract or fractions thereof may be added include meat, sausage, bread, chocolate, candy, snacks, confectionary, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, Drinks, teas, drinks, alcoholic beverages and vitamin complexes and the like, and includes all of the health food in the usual sense.

The health beverage composition of the present invention may contain various flavors or natural carbohydrates as an additional ingredient such as ordinary beverages. Such natural carbohydrates are monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, and polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol and erythritol. Examples of sweeteners include natural sweeteners such as tau martin and stevia extract, synthetic sweeteners such as saccharin and aspartame, and the like. 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, the extract of the mushroom fungus of the present invention or fractions thereof are various nutrients, vitamins, electrolytes, flavors, coloring agents, pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloid thickeners, pH regulators, stabilizers, Preservatives, glycerin, alcohols, carbonation agents used in carbonated drinks, and the like. In addition, the milk mushroom fungus extract of the present invention or a fraction thereof may contain a flesh for preparing natural fruit juice, fruit juice beverage and vegetable beverage. These components can be used independently or in combination. The proportion of such additives is not critical 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.

Milk mushroom fungus extract of the present invention is a safe substance that effectively inhibits the production of inflammatory cytokines IFN-γ and does not have cytotoxicity to cells, and thus is useful as an active ingredient of health foods that can prevent or improve inflammatory diseases. Can be used.

Hereinafter, the present invention will be described in detail by examples and production examples.

However, the following Examples and Preparation Examples are merely illustrative of the present invention, and the content of the present invention is not limited to the following Examples and Preparation Examples.

Milk mushroom  Preparation of Extract

Lactarius obtained from the National Agrobiodiversity Center (Suwon, Korea) 6 kg of fruiting bodies of hatsudake were extracted with 30 liters of methanol three times for three days (at each extraction) at room temperature. 240 g of a residue was prepared by concentrating under reduced pressure the methanol solution collected and collected three times.

Milk mushroom  From extract Fraction  Produce

In order to prepare a fraction from the milk mushroom fungus extract, 240 g of the methanol extract prepared in Example 1 was suspended in water, and then 1 L of normal-hexane (n-hexane) was added thereto, followed by partitioning. The ethyl acetate layer was separated by adding 1 l of ethyl acetate (ethylacetate, EtOAc) to the remaining water layer. In this way, 1 l of normal-butanol (n-butanol, n-BuOH) was added to normal-butanol. The layers were separated to give 17 g, 15 g and 60 g of n-hexane, ethylacetate (EtOAc) and n-butanol (n-BuOH) fractions, respectively. .

Milk mushroom  Extract or its Fraction  Interferon-gamma Interferon -γ, IFN -γ) inhibitory effect

<3-1> Milk mushroom  Extract or its Fraction  process

In order to examine the anti-inflammatory action through the IFN-γ inhibitory activity of the extract of the mushroom extract of the present invention or IFN-γ, IFN- treated with NK92 cells by treating the extract of the extract of the mushroom fungus or fractions prepared in Examples 1 and 2 The effect on the regulation of γ was confirmed. Specifically, NK92 cells (human NK lymphoma) (American type culture collection) 5.0 × 10 ⁵ cells / ml, which are IFN-γ-dependent natural killer cell lines, were placed in a 48 well culture plate with 20% fetal bovine serum ( fetal calf serum) (HyClone, Logan, UT, USA), 2 mM L-glutamate, 100 μg- ¹ penicillin, 100 μg- ¹ streptomycin (Life Technologies) and 100 U In α-MEM medium containing ml ⁻¹ IL-2 (Chiron, Emeryville, Calif., USA) was incubated in 37 ° C. and 5% CO ₂ humidity environment. The NK92 cells cultured in this way were treated with mushroom extracts or fractions thereof at various concentrations (4, 20, 100 or 400 μM). At this time, the negative control group cultured NK92 cells by treating only the culture medium, the positive control group containing 100 ng of phorbol-12-myristate-13-acetate (PMA) The media was used to cause inflammation.

<3-2> IFN Quantification of -γ

Quantification of human IFN-γ was performed according to the manufacturer's protocol using commercially available mAb pairs (Endogen, Woburn, MA, USA). After treatment with NK92 cells for 18 hours at 37 ℃ milk mushroom extract or fractions thereof, the supernatant except the cells were obtained, IFN-γ using an Enzyme Linked Immunosorbent Assay (ELISA) kit Detected. The results are shown as the mean value of three replicate wells ± s.e.m.

As a result, as shown in Table 1 and Figure 1, the production of IFN-γ in NK92 cells treated with methanol extract of milk mushroom or organic solvent fractions thereof (normal-hexane, ethyl acetate or normal-butanol fraction), respectively It was found to be suppressed and showed significantly lower IFN-γ production compared to the inflammation-induced positive control group (1.4 ng / ml), which was found to be concentration-dependent (Table 1 and FIG. 1). Thus, it was confirmed that the milk mushroom fungus extract or its fraction inhibits the production of IFN-γ in NK cells.

Milk mushroom fungus sample (μM) IFN-γ production amount (ng / ml) Methanol extract


4 1.35 20 1.25 100 1.22 400 0.95 Normal-butanol Fraction

4 1.30 20 1.22 100 1.20 400 0.65 Ethyl acetate fraction


4 1.32 20 1.25 100 1.20 400 0.85 Normal-butanol Fraction
4 1.30 20 1.24 100 1.22 400 0.90 Positive control group 1.4 Negative Control 0.1

Milk mushroom  Extract or its Fraction  Cytotoxicity Check

In order to determine the cytotoxicity of the milk mushroom extract or fractions thereof, the milk mushroom fungus extract or fractions thereof were treated with NK92 cells to analyze cell proliferation. Specifically, cell proliferation was analyzed using WST-1 (catalog no. 11644807001; Roche Applied Science, Indianapolis, IN, USA) according to the manufacturer's instructions. 2.5 × 10 ³ NK92 cells were cultured in 96 well plates, and the absorbance was measured after 12 hours treatment with 4, 20, 100, or 400 μM concentrations of the extract of milk mushroom fungi or fractions thereof.

As a result, as shown in Figure 2, all concentrations of milk mushroom extract, including 400 μM or fractions thereof was confirmed to be a safe substance with little cytotoxicity to NK92 cells (Fig. 2).

< Manufacturing example  1> Manufacture of cosmetics

Milk mushroom fungal extract of the present invention or a fraction thereof may be prepared cosmetics for the prevention and improvement of inflammatory diseases containing as an active ingredient. The present inventors prepared the cosmetic of solubilized formulations such as cosmetics and soft cosmetics of the emulsion formulations such as nutrient cosmetics, creams, essences and the like as the cosmetics.

<1-1> Cosmetic Preparation of Emulsifying Formulations

To the cosmetic composition of the emulsifier type was prepared in the composition shown in Table 2. The manufacturing method is as follows.

1) The mixture which mixed the raw materials of 1-9 was heated at 65-70 degreeC.

2) 10 was added to the mixture of step 1).

3) The mixture of the raw materials of 11-13 was heated to 65-70 degreeC, and was fully dissolved.

4) Through step 3), the mixture of 2) was slowly added to emulsify for 2-3 minutes at 6,000 rpm.

5) The raw material of 14 was dissolved in a small amount of water, and then added to the mixture of step 4) and emulsified for 2 minutes.

6) After weighing 15 to 17 raw materials, the mixture of step 5) was further emulsified at 40 ° C. for 30 seconds.

7) After the emulsification of the mixture of step 6) through a degassing process was cooled to 25 ~ 35 ℃ to prepare an emulsion cosmetic.

Composition of Emulsifying Formulations 1, 2, 3 Furtherance Emulsifier 1 Emulsifier 2 Emulsifier 3 One Stearic acid 0.3 0.3 0.3 2 Steali alcohol 0.2 0.2 0.2 3 Glyceryl Monostearate 1.2 1.2 1.2 4 Beeswax 0.4 0.4 0.4 5 Polyoxyethylene sorbitan
Monolauric acid ester 2.2 2.2 2.2 6 Methyl paraoxybenzoate 0.1 0.1 0.1 7 Paraoxybenzoic Acid Profiles 0.05 0.05 0.05 8 Cetylethylhexanoate 5 5 5 9 Triglyceride 2 2 2 10 Cyclomethicone 3 3 3 11 Purified water To 100 To 100 To 100 12 Concentrated glycerin 5 5 5 13 Triethanolamine 0.15 0.15 0.15 14 Polyacrylic acid polymer 0.12 0.12 0.12 15 Pigment 0.0001 0.0001 0.0001 16 incense 0.10 0.10 0.10 17 Ethanol Extract of Example 1 0.0001 One 10

<1-2> Solubilization  Cosmetic preparation of the formulation

A cosmetic of a solubilized formulation was prepared with the composition described in Table 3 below. The manufacturing method is as follows.

1) Raw materials 2 to 6 were placed in raw material 1 (purified water) and dissolved using a still mixer.

2) The raw materials of 8 to 11 were put into the raw material of 7 (alcohol) and completely dissolved.

3) The mixture of step 2) was solubilized with slow addition to the mixture of step 1).

Composition of Solubilized Formulations 1, 2, 3 Furtherance Solubilized Formulation 1 Solubilized Formulation 2 Solubilized Formulation 3 One Purified water To 100 To 100 To 100 2 Concentrated glycerin 3 3 3 3 1,3-butylene glycol 2 2 2 4 EDTA-2Na 0.01 0.01 0.01 5 Pigment 0.0001 0.0002 0.0002 6 Normal-hexane fraction of Example 2 0.1 5 5 7 Alcohol (95%) 8 8 8 8 Methyl paraoxybenzoate 0.1 0.1 0.1 9 Polyoxyethylene
Hydrogenated Ester 0.3 0.3 0.3 10 incense 0.15 0.15 0.15 11 Cyclomethicone - - 0.2

< Manufacturing example  2> Preparation of Pharmaceutical Formulations

<2-1> Powder  Produce

2 g of ethyl acetate fraction of Example 2 of the invention

1 g lactose

The above components were mixed and packed in airtight bags to prepare powders.

<2-2> Preparation of Tablet

100 mg of the normal-butanol fraction of Example 2 of the present invention

Corn starch 100 mg

100 mg of milk

2 mg magnesium stearate

After mixing the above components, tablets were prepared by tableting according to a conventional method for producing tablets.

<2-3> Preparation of Capsule

100 mg of the normal-hexane fraction of Example 2 of the present invention

Corn starch 100 mg

100 mg of milk

2 mg magnesium stearate

After mixing the above components, the capsule was prepared by filling in gelatin capsules according to the conventional method for producing a capsule.

<2-4> Preparation of the ring

1 g of normal-butanol fraction of Example 2 of the invention

Lactose 1.5 g

1 g of glycerin

Xylitol 0.5 g

After mixing the above components, it was prepared to be 4 g per ring according to a conventional method.

<2-5> Preparation of Granules

150 mg of ethyl acetate fraction of Example 2 of the present invention

Soybean Extract 50mg

Glucose 200 mg

600 mg of starch

After mixing the above components, 100 mg of 30% ethanol was added and dried at 60 ° C. to form granules, which were then filled into fabrics.

< Manufacturing example  3> Manufacture of food

The food containing the extract of the mushroom fungus of the present invention or a fraction thereof was prepared as follows.

<3-1> Production of flour food

0.5 to 5.0 parts by weight of the extract of milk mushroom fungus of the present invention was added to flour, and bread, cake, cookies, crackers and noodles were prepared using this mixture.

<3-2> soup  And juicy ( gravies Manufacturing

0.1-5.0 parts by weight of the normal-hexane fraction of the milk mushroom fungus extract of the present invention was added to soups and broths to prepare meat products for health promotion, soups and broths of noodles.

<3-3> Ground Beef  Produce

Health-enhanced ground beef was prepared by adding 10 parts by weight of the normal-butanol fraction of the milk mushroom fungus extract of the present invention to the ground beef.

<3-4> Dairy products ( dairy products Manufacturing

5-10 parts by weight of the ethyl acetate fraction of the extract of the mushroom fungus extract of the present invention was added to milk, and various dairy products such as butter and ice cream were prepared using the milk.

<3-5> Solar  Produce

Brown rice, barley, glutinous rice, and yulmu were dried by a known method and dried, and the mixture was granulated to a powder having a particle size of 60 mesh.

Black soybeans, black sesame seeds, and perilla seeds were steamed and dried by a conventional method, and then they were prepared into powder having a particle size of 60 mesh by a pulverizer.

Ethyl acetate fractions of the extract of milk mushroom fungus of the present invention was concentrated under reduced pressure in a vacuum concentrator, dried by spraying and drying with a hot air dryer, and ground to a particle size of 60 mesh by a grinder to obtain a dry powder.

Cereals, seeds and milk mushrooms prepared as described above were prepared by combining the ethyl acetate fractions in the following ratios.

Cereals (30 parts by weight brown rice, 15 parts by weight brittle, 20 parts by weight of barley),

Seeds (7 parts by weight of perilla, 8 parts by weight of black beans, 7 parts by weight of black sesame seeds)

Milk mushroom fraction ethyl acetate fraction (3 parts by weight),

(0.5 part by weight),

(0.5 parts by weight)

< Manufacturing example  4> Manufacture of beverage

<4-1> Health drink  Produce

Instant sterilization by homogeneously mixing 5 g of milk mushroom fungus extract of the present invention with subsidiary materials such as liquid fructose (0.5%), oligosaccharide (2%), sugar (2%), salt (0.5%) and water (75%) After it was prepared by packaging in a small packaging container such as glass bottles, plastic bottles.

<4-2> Production of vegetable juice

Vegetable juice was prepared by adding 5 g of an ethyl acetate fraction of the extract of the mushroom fungus of the present invention to 1,000 ml of tomato or carrot juice.

<4-3> Preparation of fruit juice

A fruit juice was prepared by adding 1 g of a normal-hexane fraction of the milk mushroom fungus extract of the present invention to 1,000 ml of apple or grape juice.

Claims (13)

Lactarius hatsudake ) Anti- aging, immuno-enhancing, or inflammatory disease prevention and a cosmetic composition containing 0.01 to 50 parts by weight of the extract or fractions thereof as an active ingredient.
The cosmetic composition according to claim 1, wherein the extract is extracted with water, C ₁ -C ₂ alcohol or a mixture thereof.
The cosmetic composition according to claim 2, wherein the C ₁ to C ₂ alcohol is methanol or ethanol.
The method of claim 1, wherein the fraction is normal-hexane, ethyl acetate, normal-butanol fractions obtained by sequentially sifting the extract of milk mushroom fungus with normal-hexane, ethyl acetate, normal-butanol, and water fractions or water fractions Cosmetic composition, characterized in that.
The cosmetic composition according to claim 1, wherein the inflammatory disease is any one selected from the group consisting of edema, dermatitis, allergy, atopy, acne, pruritus, multiple sclerosis and acute inflammatory disease.
The cosmetic composition of claim 1, wherein the cosmetic composition improves or prevents wrinkles and aging resulting from an inflammatory disease.
According to claim 1, wherein the cosmetic composition is prepared in one or more formulations selected from the group consisting of emulsion, cream, essence, essence, body lotion, body gel, body essence, body cleanser, cleansing foam, pack and cleansing cream Cosmetic composition, characterized in that.
Skin external preparation for the prevention and improvement of inflammatory diseases containing Lactarius hatsudake extract or fractions thereof as an active ingredient.
The method of claim 8, wherein the inflammatory disease is edema, dermatitis, allergy, atopy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer, gastritis, Crohn's disease, ulcerative colitis, hemorrhoids, gout, ankylosing Spondylitis, rheumatic fever, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, periarthritis, tendinitis, hay salt, peritonitis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome, multiple sclerosis, and a variety of The external preparation for skin, which is any one selected from the group consisting of acute and chronic inflammatory diseases.
Lactarius hatsudake ) A pharmaceutical composition for the prevention and treatment of inflammatory diseases containing an extract or a fraction thereof as an active ingredient.
The method of claim 10, wherein the inflammatory disease is edema, dermatitis, allergy, atopic, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer, gastritis, Crohn's disease, ulcerative colitis, hemorrhoids, gout, ankylosing Spondylitis, rheumatic fever, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, periarthritis, tendinitis, hay salt, peritonitis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome, multiple sclerosis, and a variety of The external preparation for skin, which is any one selected from the group consisting of acute and chronic inflammatory diseases.
Lactarius hatsudake ) Health food for the prevention and improvement of inflammatory diseases containing extract or fractions thereof as an active ingredient.
The method of claim 12, wherein the inflammatory disease is edema, dermatitis, allergy, atopy, asthma, conjunctivitis, periodontitis, rhinitis, otitis media, sore throat, tonsillitis, pneumonia, gastric ulcer, gastritis, Crohn's disease, ulcerative colitis, hemorrhoids, gout, ankylosing Spondylitis, rheumatic fever, lupus, fibromyalgia, psoriatic arthritis, osteoarthritis, rheumatoid arthritis, periarthritis, tendinitis, hay salt, peritonitis, myositis, hepatitis, cystitis, nephritis, sjogren's syndrome, multiple sclerosis, and a variety of Health food, characterized in that any one selected from the group consisting of acute and chronic inflammatory diseases.

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