KR20140023462A - Extracts from smilacina japonica a. gray, or arabis glabra bernh having potent anti-inflammatory activity - Google Patents

Abstract

The present invention relates to a Smilacina japonica A. Gray extract or an Arabis glabra Bernh extract having anti-inflammatory activity, more specifically to a composition for preventing, curing, and treating inflammatory diseases containing a Smilacina japonica A. Gray extract or a fraction thereof, or an Arabis glabra Bernh extract or a fraction thereof as an effective component. [Reference numerals] (AA) Smilacina japonica A. Gray CHCI3(ug/mL); (BB) Arabis glabra Bernh CHCI3(ug/mL); (CC) Arabis glabra Bernh Hexane(ug/mL)

Description

Herb extracts or extracts of pole herb with anti-inflammatory activity {Extracts from Smilacina japonica A. Gray, or Arabis glabra Bernh Having Potent Anti-Inflammatory Activity}

The present invention relates to a grass swam extract or an extract of Jangnamul with anti-inflammatory activity, and more particularly, extracts or fractions of Smilacina japonica A. Gray, extracts or fractions of Arabis glabra Bernh It relates to a composition for the prevention, treatment and improvement of inflammatory diseases containing as a component.

Inflammation refers to a pathological condition of the abscess formed by the invasion of foreign bacteria. Inflammation reaction is a biological defense reaction process for repairing and regenerating damage caused by invasion that brings about a fundamental change in the cell or tissue of a living body. It is a process of damaging tissues (cells) or infecting external sources (bacteria, fungi, Allergen-inducing substances), various inflammatory mediators and immune cells are involved in local blood vessels and body fluids, and a series of complex physiological responses such as enzyme activation, inflammatory mediator secretion, body fluid infiltration, cell migration, and tissue destruction, And fever pain. In normal cases, the inflammatory reaction removes the external infectious agent and regenerates the damaged tissue to regenerate the organism's function. However, if the antigen is not removed or the internal substance causes the inflammatory reaction to occur excessively or continuously, the major pathological condition of the disease (irritable disease, Chronic inflammation), and become obstacles in the treatment process such as transfusion, drug administration, or organ transplantation.

Inflammatory factors involved in the inflammatory response is nitric oxide (NO). NO is produced by NO synthase (NOS) through L-arginine, an intermediate metabolite, hydroxyarginine, and NO and citrulline. NO is biologically active and acts on the vascular system in vivo and is involved as a mediator in vasodilation, platelet adhesion and aggregation, neurotransmission, and digestive system movement. In addition, NO has been shown to be responsible for various metabolic and physiological functions (neurotransmission, coagulation, blood pressure control, immunity to cancer cells, etc.), and NO is produced in inflammatory cells as well as non-immune cells to protect against microbial infection. It works. In general, NO exists in the form of a gas in the atmosphere, and because it is a free radical structure, it is not only very toxic, but also has an unstable half life of 6 to 10 seconds, and is known to be rapidly converted into the stabilized final products NO ₃ and NO ₂ . Snyder, SH, et al., Scientific American, May, pp 28-35, 1992.

On the other hand, NO synthase (NOS) involved in NO production is largely classified into cNOS (constitutive NOS) and iNOS (inducible NOS) depending on whether calcium (Ca ²⁺ ) or calmodulin (camodulin) dependence. cNOS is calcium or calmodulin dependent and is mainly present in brain, epithelial, neutrophil, and gastrointestinal mucosa cells. iNOS is calcium or calmodulin independent. iNOS is present in macrophages, hepatocytes, cancer cells, IL-1β, Cytokines such as IFN- [gamma], TNF- [alpha], and bacterial endogenous bacterial lipopolysaccharide (LPS). [Dinerman, JL, et al., Circ. Res., 73, pp 217-222, 1993]. This stimulation also activates cyclooxygenase-2 (COX-2) to produce prostaglandins (PGs), which are inflammatory mediators. Therefore, iNOS expression and COX-2 expression are closely related. It also affects COX-2 expression [Robert C., et al., J. Immunol., 165, pp 1582-1587, 2000; Daniela S., et al., Proc. Natl. Acad. Sci. USA, 90, pp 7240-7244, 1993. Therefore, various efforts have been made to secure a substance that inhibits the production of NO by iNOS and develop it as a medicine.

Inflammatory diseases caused by these inflammatory reactions include atherosclerosis, gastritis, colitis, arthritis, nephritis, hepatitis, cancer or degenerative diseases [Gobert A. P. et al., J. Immunol. 168 (12), pp 6002-6006, 2002; Dijkstra G. et al., Scand. J. Gastroent erol. 37 (5), pp 546-554, 2002; Sakac V. and Sakac M. Med. Pregl. 53, pp 463-474, 2000; Albrecht E. W. et al., Am. J. Transplant, 29 (5), 448-453, 2002; Ramachandran A. et al., Free Radical Biol. Med. 33 (11), pp 1465-1474, 2002; Sartor L. et al., Biochemical Pharmacol. 64, pp 229-237, 2002; Sadowska-Krowicka H. et al., Proc. Soc. Exp. biol. Med. 217 (3), pp 351-357, 1998, Lo A-H. et al., Carcinogenesis, 23 (6), pp 983-991, 2002].

On the other hand, Korean Patent No. 442243 discloses a pharmaceutical composition for preventing and treating inflammatory diseases, which contains an extract of a powder (tinnitus, cotton). Phyllostachys nigra var.henonis is a perennial evergreen tree belonging to the family Asteraceae , clearly distinguished from Smilacina japonica A. Gray var. Japonica to be.

To date, no document discloses that extracts or fractions from Smilacina japonica A. Gray or Arabis glabra Bernh have anti-inflammatory activity.

Therefore, the present inventors activated the lipopolysaccharide (LPS), an inflammation-inducing substance, to investigate the anti-inflammatory effect on extracts or fractions isolated from Smilacina japonica A. Gray or Arabis glabra Bernh. The inhibition of NO production and the inhibition of production of prostaglandin E ₂ on one macrophage was measured. As a result, the present invention was completed by confirming that extracts or fractions separated from these plants were excellent in inhibiting the production of inflammatory metabolites.

Therefore, an object of the present invention is to provide a composition comprising an extract or fraction of grass pads or rods as an active ingredient excellent in inhibiting the production of inflammatory metabolites.

In order to solve the above problems, the present invention is characterized by a pharmaceutical composition for the prevention and treatment of inflammatory diseases containing an extract of Smilacina japonica A. Gray or Pole Herb ( Arabis glabra Bernh) as an active ingredient.

In another aspect, the present invention is characterized by a health functional food composition for the prevention and improvement of inflammatory diseases containing an extract of Smilacina japonica A. Gray or Arabis glabra Bernh as an active ingredient.

In addition, the present invention is characterized by a cosmetic composition for the prevention and improvement of inflammatory diseases containing an extract of Smilacina japonica A. Gray or extract of Arabis glabra Bernh as an active ingredient.

In addition, the present invention is characterized in that the extract of Smilacina japonica A. Gray or extract of Arabis glabra Bernh is an ethanol extract.

In addition, the present invention is characterized in that the fraction obtained by sequentially sequencing the extract of Smilacina japonica A. Gray or extract of Arabis glabra Bernh with n-hexane, chloroform, ethyl acetate, n-butanol .

Extracts or fractions of Smilacina japonica A. Gray or Arabis glabra Bernh of the present invention are useful for the prevention, amelioration and treatment of inflammatory diseases.

In the present invention, the term "inflammatory disease" may be defined as any condition specified by a local or systemic biological defense response against external physical and chemical stimuli or infection of an external infectious agent such as bacteria, fungi, and viruses. These reactions activate various inflammatory mediators and enzymes associated with immune cells (eg iNOS, COX-2, etc.), secretion of inflammatory mediators (eg, NO, TNF-α, IL-6, IL-1β, PGE2) It is accompanied by a series of complex physiological reactions, such as fluid infiltration, cell migration, and tissue destruction, and is manifested externally by symptoms such as erythema, pain, swelling, fever, deterioration or loss of certain functions of the body. The inflammatory disease may be acute, chronic, ulcerative, allergic or necrotic, so long as it is included in the definition of inflammatory diseases as above, it may be acute, chronic, ulcerative, allergic, Whether it is necrotic or not. Specifically, the inflammatory diseases include asthma, allergic and non-allergic rhinitis, chronic and acute rhinitis, chronic and acute gastritis or enteritis, ulcerative gastritis, acute and chronic nephritis, acute and chronic hepatitis, chronic obstructive pulmonary disease, pulmonary fibrosis Irritable bowel syndrome, inflammatory pain, migraines, headache, back pain, fibromyalgia, fascia disease, viral infections (eg, type C infections), bacterial infections, fungal infections, burns, surgical or dental wounds, Hyperprostaglandin E excess syndrome, atherosclerosis, gout, arthritis, rheumatoid arthritis, ankylosing spondylitis, Hodgkin's disease, pancreatitis, conjunctivitis, irisitis, scleritis, uveitis, dermatitis, eczema, atopic inflammation, acne, multiple sclerosis, etc. This includes.

Therefore, the extract or fraction of the present invention is useful as an active ingredient of a pharmaceutical composition, nutraceutical composition or cosmetic composition for the purpose of preventing, ameliorating and treating an inflammatory disease.

1 is a graph confirming the NO efficacy for extracts or fractions isolated from Smilacina japonica A. Gray or Arabis glabra Bernh.
Figure 2 is a graph confirming the efficacy of PGE ₂ inhibition on extracts or fractions isolated from Smilacina japonica A. Gray or Arabis glabra Bernh.

The present invention relates to a composition for the prevention, treatment and improvement of inflammatory diseases containing extracts or fractions of grass pads or pole herbs as an active ingredient.

As a plant used in the present invention, Smilacina japonica A. Gray is a perennial herb belonging to the family Liliaceae. The grass pad is similar to the fragrance used in temples, so it is sometimes called 'Jijang-namul'. The grass swam is 20-50 ㎝ in size, the root stem is fleshy, grows laterally, and the main stem comes out obliquely from the end. Leaves are 6-15 cm long, 2-5 cm wide, in two rows along the stem, with long ovals, narrow in tip. Flowers are white and small flowers gather together at the end of the main stem to form a single flower. Fruits are round and red in September. At first glance, the leaves appear to resemble a round, but if you look at the size and stem of the leaf, you can see a distinct difference. Young sprouts are slightly boiled and eaten as herbs, which are sweet and dried. In oriental medicine, grass pads have the effect of strengthening the kidneys, activating blood and smoothing menstruation.

On the other hand, grass pads are sometimes called bands in different regions, but they are clearly different from the plant classifications of the phyllostachys nigra var. Henonis , a perennial evergreen tree belonging to the family Asteraceae .

As another plant used in the present invention, Arabis glabra Bernh is a biennial herb belonging to the cruciferaceae. The growing environment grows in sunny areas with good drainage. The height is about 70 cm, the lower leaves are hairy with the leaves from the root, but the upper leaves are hairless, and the leaves from the stem are long oval. The flower is white, cruciform and runs at the end of the main stem. Fruits are nuts, 4-6 cm long, in the form of strings. Young shoots are slightly blanched and eaten as herbs.

As a plant used in the present invention, grass pads and rods may be used without limitation, such as cultivated, harvested or commercially available. In addition, grass pads and rods may be used for the whole plant, it is preferable to use leaves, stems or roots, but is not limited thereto. The grass swab and pole herb can be used at any time during the growth stage, but preferably using young sprouts.

The composition according to the present invention may include a grass swam extract, a rod extract, or a mixture thereof as an active ingredient.

For the preparation of the extract, as the extraction solvent, water, C _{1 ~} C ₃ alcohol or a mixture thereof may be used, and preferably extracted with methanol or ethanol, but is not limited thereto. The amount of the extraction solvent is preferably 2 to 20 times by weight based on the dry weight of the plant, more preferably 5 to 10 times by weight, but is not limited thereto.

The method of extracting the plant is a method using an extraction device such as hot water extraction, immersion extraction, supercritical extraction, subcritical extraction, high temperature extraction, high pressure extraction, reflux cooling extraction and ultrasonic extraction or adsorption resin including XAD and HP-20. Conventional extraction methods in the art may be used, such as the use of the method. It is preferred that the extract is heated to reflux or at room temperature, but is not limited thereto. The number of extraction is preferably 1 to 5 times, and more preferably three times of extraction, but is not limited thereto. It may be 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 to 7 days, but is not limited thereto. The extract is preferably concentrated under reduced pressure, but is not limited thereto. The vacuum concentration is preferably a vacuum rotary evaporator, but is not limited thereto. The drying is preferably carried out under reduced pressure, vacuum drying, boiling, spray drying, room temperature drying or freeze drying, but not always limited thereto.

In addition, the composition according to the present invention may include a fraction obtained by sequentially fractionating the extract of the above-mentioned grass swabs or Jangnamul as n-hexane, chloroform, ethyl acetate, n-butanol as an active ingredient.

The fractionation process from the extract of grass swabs or rods may be repeated 1 to 5 times, preferably 3 times, to obtain a fraction, and the obtained fraction is preferably concentrated under reduced pressure, but not always limited thereto. The vacuum concentration is preferably a vacuum rotary evaporator, but is not limited thereto. The drying is preferably carried out under reduced pressure, vacuum drying, boiling, spray drying, room temperature drying or freeze drying, but not always limited thereto.

In order to verify the anti-inflammatory effect on the extracts or fractions of grass swabs or rods of the present invention, RAW 264.7 macrophage macrophages through LPS induction was used as an inflammation model. MTT search was performed on the total 10 extracts or fractions extracted from grass pads or pole sprouts to determine the cytotoxicity as IC ₈₀ and set the effective concentration range without toxicity. Set. In addition, the treatment of inflammatory lipopolysaccharide (LPS) induced the production of NO, an inflammatory marker, and the treatment of each extract or fraction confirmed the NO production inhibitory effect, based on the results induced by LPS Inflammatory metabolite prostaglandin E ₂ (PGE ₂ ) was confirmed to inhibit the production.

According to the experimental results, it can be seen that the extract or fraction of the present invention is low in cytotoxicity and excellent in inhibiting NO production, thereby preventing, improving and treating inflammatory diseases. In other words, the extract or fraction of the grass pad and pole herbs of the present invention can be usefully used as an active ingredient of drugs, health functional food, or cosmetics for the prevention, improvement, treatment of inflammatory diseases.

The present invention provides a pharmaceutical composition containing an extract or fraction of grass pads or rods as an active ingredient.

The pharmaceutical composition of the present invention preferably comprises 0.01 to 50% by weight of the extract or fraction based on the total weight of the composition, but is not limited thereto.

Pharmaceutical compositions of the present invention may further comprise suitable carriers, excipients and diluents conventionally used in the manufacture of a medicament.

The pharmaceutical compositions according to the invention can be used in the form of oral dosage forms, external preparations, suppositories, and sterile injectable solutions, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., according to conventional methods, respectively. Can be. Carriers, excipients and diluents which may be included in the pharmaceutical composition of the present invention include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate , Cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil.

In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may be formulated into the compositions of the present invention with at least one excipient such as starch, calcium carbonate, (sucrose), lactose, gelatin and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. As a suppository base, witepsol, macrogol, tween 61, cacao paper, laurin, glycerol gelatin and the like can be used.

The pharmaceutical composition of the present invention may be administered orally or parenterally, any parenteral administration method may be used, and systemic or topical administration is possible, but systemic administration is more preferable, and intravenous administration is most preferred.

Preferred dosages of the pharmaceutical compositions of the present invention vary depending on the condition and weight of the patient, the extent of the disease, the form of the drug, the route of administration and the duration, and may be appropriately selected by those skilled in the art. However, for the desired effect, the composition of the present invention is preferably administered at 0.0001 to 1 g / kg per day, preferably 0.001 to 200 mg / kg per day. The administration may be carried out once a day or divided into several times. The dose is not intended to limit the scope of the invention in any way.

In another aspect, the present invention provides a health functional food composition containing an extract or fraction of the grass pads or rods as an active ingredient.

The composition according to the present invention may be added to the functional food, such as food, beverages, extracts or fractions of grass pads or rods for the purpose of preventing or improving inflammatory diseases. The present invention is not particularly limited to the type of food. Examples of foods to which the extract or fraction may be added include drinks, meat, sausages, breads, biscuits, rice cakes, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum products, dairy products including ice cream, various Soups, beverages, alcoholic beverages and vitamin complexes, dairy products and dairy products, etc., and all the health functional foods in the conventional sense.

The extract or fraction of the wadding or barley sprout of the present invention may be added to a food as it is or used with other food or food ingredients, and may be appropriately used according to a conventional method. The content as an active ingredient can be suitably determined according to the purpose of use (prevention or improvement). In general, the amount of the extract or fraction in the dietary supplement may be added at 0.1 to 90% by weight of the total food weight. However, in the case of long-term intake intended for health and hygiene purposes or for the purpose of controlling health, the amount may be less than the above range, and since there is no problem in terms of safety, the active ingredient may be used in an amount exceeding the above range.

The health functional beverage composition of the present invention contains the extract or fraction as an essential ingredient in the indicated ratio, and there are no particular limitations on other ingredients used for the manufacture of other beverages, and various flavors or natural ingredients such as ordinary drinks Carbohydrates and the like may be included as additional components. Examples of such natural carbohydrates include monosaccharides such as glucose, fructose, and the like; Disaccharides such as maltose, sucrose and the like; And polysaccharides, for example, conventional sugars such as dextrin, cyclodextrin and the like, and sugar alcohols such as xylitol, sorbitol and erythritol. Natural flavors and synthetic flavors and the like can be used as the flavors other than those described above. The proportion of such natural carbohydrates is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the composition of the present invention.

In addition, extracts or fractions of the present invention are various nutrients, vitamins, minerals (electrolytes), flavors such as synthetic and natural flavors, coloring and neutralizing agents (such as cheese, chocolate), pectic acid and salts thereof, alginic acid and Salts, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated drinks, and the like. In addition, the extracts or fractions of the present invention may contain fruit flesh for the production of natural fruit juices and fruit juice beverages and vegetable beverages. These components may be used independently or in combination. The proportion of such additives is not so critical but is generally selected in the range of from 0.1 to about 20 parts by weight per 100 parts by weight of the bee-eater extract or fractions thereof.

In another aspect, the present invention provides a cosmetic composition containing an extract or fraction of grass swab or Jangnamul as an active ingredient.

The cosmetic composition of the present invention preferably comprises an extract or fraction in an amount of 0.01 to 50% by weight based on the total weight of the composition, but is not limited thereto.

The cosmetic composition of the present invention may be in the form of an oil dispersion in an aqueous phase using an aqueous, aqueous-alcoholic or oily solution, oil or water or multi-emulsion, aqueous or oily gel, liquid, pasty or solid anhydrous products, And more preferably in the form of a lipid vesicle in ionic and / or non-ionic form.

The cosmetic composition of the present invention may be somewhat fluid and may have the appearance of a white or colored cream, ointment, milk lotion, serum, essence, paste or mousse. It may optionally be applied to the skin in the form of an aerosol, or in the form of a solid, for example in the form of a stick, in various forms such as creams, lotions, skins, essences, emulsion soaps, foam cleansing, packs, shampoos, rinses, body washes, etc. It can be formulated. It may be used as a skin care product or makeup product. In a known manner the compositions according to the invention are also useful in the cosmetic field as conventional auxiliaries such as hydrophilic or lipophilic gelling agents, hydrophilic or lipophilic active agents, preservatives, antioxidants, solvents, fragrances, fillers, blockers, pigments, odorants and It may contain dyes. The amounts of these various adjuvants are amounts commonly used in the art, such as in the range of 0.01 to 20% by weight relative to the total weight of the cosmetic composition. Depending on its nature, such adjuvants may be incorporated into the fatty phase, aqueous phase, lipid vesicles and / or nanoparticles. In any case, the adjuvants and their proportions will be chosen so as not to adversely affect the desired properties of the cosmetic composition according to the invention.

The present invention as described above will be described in more detail through the following examples, which are only illustrative of the present invention and the scope of the present invention is not limited to these examples.

[Example]

Example 1. Preparation of Extracts and Fractions

The grass swam or pole sprouts were collected in mid-April (young) and mid-August (adult) in Pyeongchang, Gangwon-do. 1 g of ethanol was added to 200 g of the sample of lyophilized grass swab or pole sprouts, and the extract was filtered under reflux for 3 hours. After the filtrate was filtered, 2 L of ethanol was further added to the remaining residue, followed by further repeated heating and reflux extraction for 3 hours, thereby obtaining 6 L of the total extract. 6 L of the extract was concentrated under reduced pressure at 35 ° C. to obtain an extract of grass swab or Jangnamul.

In addition, the grass swab extract or Jangnamul extract was suspended in 0.5 L of distilled water and then fractionated with n-hexane to obtain an n-hexane fraction. The water layer fractionated with n-hexane was fractionated with chloroform to obtain a chloroform fraction. The water layer fractionated with chloroform was fractionated with ethyl acetate to obtain an ethyl acetate fraction. Then, the remaining water layer fractionated with ethyl acetate was fractionated with n-butanol to obtain an n-butanol fraction. Each fraction obtained above was concentrated under reduced pressure at 35 ° C.

Example 2. Cytotoxicity Test

Prior to searching for anti-inflammatory efficacy for each of the extracts and fractions of the present invention, an MTT assay was performed by the following method to set the optimal dose without toxicity.

RAW 264.7 cell lines were incubated at 37 ° C. and 5% CO 2 in DMEM medium (Dulbeco's modified Eagle medium) containing 10% FBS and penicillin (100 μg / mL) and streptomycin (100 U / mL). After dispensing 300 μL of cells at a density of 1 × 10 ⁵ cells / mL in a 96 well plate and incubating for 24 hours, extracts or fractions were added to the medium at various concentrations. After 24 hours of sample treatment, MTT reagent was added and incubated in the shaded area for 4 hours to form MTT Formazan (Formazan). After removing the supernatant, 100 μL of dimethyl sulfoxide (DMSO) was used to dissolve MTT formazan formed. After 10 minutes the absorbance was measured at 540 nm. The results are shown in Table 1 below.

sample IC ₈₀

Grass pad  Ethanol extract > 100  n-butanol fraction > 100  Ethyl acetate fraction > 100  Chloroform fraction > 100  n-hexane fraction > 50

Pole Herb  Ethanol extract > 12.5  n-butanol fraction > 100  Ethyl acetate fraction > 100  Chloroform fraction > 100  Hexane fraction > 50

Cytotoxicity was expressed as IC ₈₀ as the concentration of> 80% survival was determined to be the optimal dose. According to the results of Table 1, the ethanol extract of the coarse sprouts had an IC ₈₀ value at the concentration of 12.5 μg / mL or more and the hexane fraction of the grass swab and the hexane fraction of the coarse sprouts, respectively, at a concentration of 50 μg / mL or more. All samples were not toxic at high concentrations of 100 μg / mL.

Example 3. Determination of anti-inflammatory activity (NO search)

The amount of NO produced in the culture medium of RAW 264.7 cells activated by LPS at a concentration that showed no cytotoxicity in the MTT assay was measured using a Greries reagent. NIL was used as a positive control.

RAW 246.7 macrophage cell lines were placed in Dulbeco's modified Eagle medium containing 10% FBS penicillin (100 μg / mL) and streptomycin (100 μg / mL) and incubated in a 37 ° C., 5% CO ₂ incubator at 37 ° C. It was. After dispensing 300 μL at a density of 1 × 10 ⁵ cells / mL in a 96 well plate and incubating for 24 hours, the extract or fraction sample was treated at a concentration of 50 ppm. After 1 hour of treatment, 1 μg / mL of LPS was treated. After 18 hours, 100 μL of grease reagent was reacted with the amount of NO released in the medium for 10 minutes, and the absorbance was measured at 540 nm. It was. The amount of NO is NO _2- ^?? Measured as

NO release inhibition ability for each extract and fractions are shown in Figure 1 and Table 2, respectively, as an IC ₅₀ value.

sample IC ₅₀ (μg / mL)

Grass pad  Ethanol extract > 100  n-butanol fraction > 100  Ethyl acetate fraction > 100  Chloroform fraction 53.3  n-hexane fraction > 50

Pole Herb  Ethanol extract > 20  n-butanol fraction > 100  Ethyl acetate fraction > 100  Chloroform fraction 83.9  Hexane fraction 21.0 Positive control group (NIL) 10

The chloroform fraction of grass swabs, chloroform fraction of rodents, and hexane fractions of rodents showed the highest concentration-dependent inhibition of NO production with IC ₅₀ of 53.3, 83.9 and 21.0 μg / mL, respectively, and were superior to NIL, the positive control. Efficacy was shown.

Example 4. Determination of anti-inflammatory activity (PGE ₂ search)

Inhibition of the production of PGE ₂ , another inflammatory metabolite induced by LPS, against the chloroform fraction of grass swab, the chloroform fraction of the larvae and the hexane fraction of the larvae, which were determined to have excellent activity in the NO screening of Example 3 It searched by the following method. NS398 was used as a positive control.

That is, the RAW 246.7 macrophage cell line was placed in Dulbeco's modified Eagle medium containing 10% FBS penicillin (100 μg / mL) and streptomycin (100 μg / mL), and the cells were incubated at a temperature of 37 ° C. and a 5% CO ₂ incubator. Was incubated. After dispensing 300 μL at a density of 1 × 10 ⁵ cells / mL in a 96 well plate and incubating for 24 hours, the extract or fraction sample was treated at a concentration of 50 ppm. PGE ₂ was quantified by taking the cell culture solution of each cell group and control group treated with the sample.

PGE ₂ amounts for each extract and fractions are shown in Figure 2 and Table 3, respectively, as an IC ₅₀ value.

sample IC ₅₀ (μg / mL) Chloroform fraction of grass pad 32.5  Chloroform Fraction of Pole Herb 18.0  Hexane Fraction of Pole Sprouts 45.4 Positive control group (NS398)  4

The chloroform fraction of grass swabs, the chloroform fraction of the rods and the hexane fractions of the rods showed concentration-dependent PGE ₂ inhibitory effects.

[Example]

On the other hand, the pharmaceutical composition comprising an extract or fraction according to the present invention can be prepared in various forms according to the purpose. Formulations 1 to 4 below illustrate the preparation of the pharmaceutical preparation containing the extract or fraction according to the present invention as an active ingredient, but the present invention is not limited thereto.

Formulation 1: Tablets (direct pressurization)

After 5.0 mg of the active ingredient was sieved, 14.1 mg of lactose, 0.8 mg of crospovidone USNF and 0.1 mg of magnesium stearate were mixed and pressurized to make tablets.

Formulation 2: Tablet (wet assembly)

After 5.0 mg of the active ingredient was sieved, 16.0 mg of lactose and 4.0 mg of starch were mixed. 0.3 mg of Polysorbate 80 was dissolved in pure water, and an appropriate amount of this solution was added, followed by atomization. After drying, the granules were sieved and mixed with 2.7 mg of colloidal silicon dioxide and 2.0 mg of magnesium stearate. The granules were pressurized to make tablets.

Formulation 3: Powder and Capsule

5.0 mg of the active ingredient was sieved and mixed with 14.8 mg of lactose, 10.0 mg of polyvinylpyrrolidone and 0.2 mg of magnesium stearate. The mixture was filtered through a hard No. 5 gelatin capsules.

Formulation 4: Injection

100 mg as an active ingredient, 180 mg of mannitol, 26 mg of Na ₂ HPO ₄ .12H ₂ O and 2974 mg of distilled water were added to prepare an injection.

In addition, the cosmetic composition comprising an extract or fraction according to the present invention can be prepared in various forms according to the purpose. Formulations 5 to 7 below illustrate cosmetic preparation methods containing an extract or fraction according to the present invention as an active ingredient, but the present invention is not limited thereto.

Formulation 5: Cosmetic Cream

2 mg of polyethylene glycol monostearate, 5 mg of self-emulsifying glycerin monostearate, 6 mg of squalene, 6 mg of tri2-ethylhexaneglyceryl, 1 mg of sphingoglycolipids, 1,3-butylene glycol 7 mg was mixed in an appropriate amount of purified water to prepare a cosmetic cream.

Formulation 6: Cosmetic Lotion

Active ingredient 0.0015 mg, L-ascorbic acid-2-magnesium phosphate 1 mg, water-soluble collagen (1% aqueous solution) 1 mg, sodium citrate 0.1 mg, citric acid 0.05 mg, licorice extract 0.2 mg, 1,3-butylene glycol 3 mg was mixed in an appropriate amount of purified water to prepare a cosmetic lotion.

Formulation 7: Cosmetic Skin

Active ingredient 0.0018 mg, hydroxyethylene cellulose (2% aqueous solution) 12 mg, xanthan gum (2% aqueous solution) 2 mg, 1,3-butylene glycol 6 mg, concentrated glycerin 4 mg, sodium hyaluronate (1% aqueous solution) ) 5 mg was mixed in an appropriate amount of purified water to prepare a cosmetic skin.

In addition, the health functional food composition comprising an extract or fraction according to the present invention can be prepared in various forms according to the purpose. Formulations 8 to 12 below illustrate a method for preparing a health functional food containing the extract or fraction according to the present invention as an active ingredient, but the present invention is not limited thereto.

Formulation 8. Granular Health Food

Active ingredient 1000 mg, Vitamin A Acetate 70 ㎍, Vitamin E 1.0 mg, Vitamin 0.13 mg, Vitamin B ₂ 0.15 mg, Vitamin B ₆ 0.5 mg, Vitamin B ₁₂ 0.2 ㎍, Vitamin C 10 mg, Biotin 10 ㎍, Nicotinamide 1.7 Mg, folate 50 ㎍, calcium pantothenate 0.5 mg, ferrous sulfate 1.75 mg, zinc oxide 0.82 mg, magnesium carbonate 25.3 mg, potassium monophosphate 15 mg, dibasic calcium phosphate 55 mg, potassium citrate 90 mg, calcium carbonate 100 Mg and 24.8 mg of magnesium chloride were mixed, and then granular health food was prepared according to a conventional method.

Formulation 9. Health Drink

1000 mg of active ingredient, 1000 mg of citric acid, 100 g of oligosaccharide, 2 g of plum concentrate, and 1 g of taurine were mixed and purified water was added thereto to adjust the total volume to 900 ml. After stirring and heating at 85 ° C. for about 1 hour, the resulting solution was collected by filtration into a sterilized 2 L container and sealed and sterilized to prepare a healthy beverage.

Although the composition ratio is a composition that is relatively suitable for a preferred beverage in a preferred embodiment, the composition ratio may be arbitrarily modified according to regional and ethnic preferences such as demand hierarchy, demand country, and usage.

Formulation 10. Flour Food

0.5 to 5 g of the active ingredient was added to 100 g of wheat flour, and the mixture was used to prepare bread, cake, cookies, crackers and noodles to prepare health promoting food.

Formulation 11.Dairy Products

5-10 g of active ingredient was added to 100 g of milk, and the milk was used to prepare various dairy products such as butter and ice cream.

Formulation 12. Wire Type

30 g of brown rice, 20 g of barley, 10 g of glutinous rice, and 15 g of yulmu were alphalized by a known method and dried, and the powder was prepared into a powder having a particle size of 60 mesh by a pulverizer. 7 g of black beans, 7 g of black sesame, and 7 g of perilla were steamed and dried in a known manner, and the resulting mixture was used as a powder having a particle size of 60 mesh. The grains and seeds prepared above were mixed with 3 g of the active ingredient of the present invention to prepare a wire.

As described above, the extracts or fractions of the wadding or rod herbs according to the present invention are the production of NO (Nitric oxide) and prostaglandin E ₂ (PGE ₂ ), an inflammatory metabolite induced by lipopolysaccharide (LPS). It can be confirmed that the inhibitory effect in the production.

Therefore, extracts or fractions of grass pads or pole herbs are useful as an active ingredient of drugs, health functional foods, cosmetics for the purpose of preventing, improving or treating inflammatory diseases.

Claims (7)

Pharmaceutical composition for the prevention and treatment of inflammatory diseases containing an extract of Smilacina japonica A. Gray or Arabis glabra Bernh as an active ingredient.
Health functional food composition for the prevention and improvement of inflammatory diseases containing an extract of Smilacina japonica A. Gray or extract of Arabis glabra Bernh as an active ingredient.
Cosmetic composition for the prevention and improvement of inflammatory diseases containing an extract of Smilacina japonica A. Gray or Arabis glabra Bernh as an active ingredient.
The method according to any one of claims 1 to 3,
The extract is a composition characterized in that the extract of Smilacina japonica A. Gray, rods ( Arabis glabra Bernh), or a mixture thereof with an alcohol of C _{1 ~} C ₄ .
The composition of claim 4 wherein the alcohol is methanol or ethanol.
5. The composition according to claim 4, wherein the extract comprises a fraction obtained by sequentially fractionating n-hexane, chloroform, ethyl acetate, n-butanol as an active ingredient.
7. The composition of claim 6, wherein the fraction is an n-hexane fraction or a chloroform fraction.

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