COMPOSITION FOR PREVENTING, IMPROVING, OR TREATING MENOPAUSAL SYMPTOMS, CONTAINING POLYGONUM MULTIFLORUM FERMENTED WITH SHIITAKE HYPHAE

The purpose of the invention to achieve, the present invention refers to dropping possibility five (Polygonum Multiflorum) Fermented ingredient containing female menopause preventives or non-allergic composition number [...] substrate.

In one embodiment of the present invention according to health nutraceutical compositions in, said possibility five KCTC18583P oak mushroom strain is in fermentation comprises dropping (LentinusEdodes) But can be a corresponding motor mycelium of fermentation, the number is not one. Said shiitake mushroom (Lentinus Edodes) To strain (strain: KCTC18583P) was prepared in the lower half 3 July 2017 bio-technology Institute.

In addition, dropping said possibility five fermentation comprises

1) Get hot air-dried and dripping [...] makes who does not pass through a dropping possibility five to 25 mesh sieve to yield a powder; and

2) Inoculating said strain is inoculated with a mushroom mycelium powder obtained after dropping possibility five KCTC18583P in carbon won of glucose, malt as nitrogen won performance by culturing possibility five 35 - 45 days 24 provided 26 °C and pH 6 - 7 conditions obtain dropping fermentation product; including a fermentation method can be a possibility five-dropping and obtained from fermentation,

Preferably, said step 2) in, 25 °C and pH 6 after inoculating strain is inoculated with a mushroom mycelium powder dropping possibility five KCTC18583P in culturing possibility five 40 conditions obtain dropping during fermentation product; including a fermentation method can be a possibility five-dropping but obtained from fermentation, the number is not one.

In addition, fluoxetine use of said diseases, perspiration, skin dryness, qualitative xeransis, qualitative atrophy, lower urethral atrophy, Vaginitis, effective, as a result of urination, class [nyo, concentration disorders, Huntington disorders, anxiety, nerve 5801, pharmaceutical compositions containing decay, for the treatment of, one or more selected from the group consisting of osteoporosis can be a arthralgia, the number is not one.

Said composition powder, granules, ring, positive number, capsule, candy, beverage syrup of selected number-one of the tank can be a number, the number is not one.

Health food composition of the present invention when used for food additives, either adding said second health food composition can be use with other food or food ingredients, can be suitable for use according to the conventional method. (Preventing or ameliorating) is suitable for use in accordance with the aim of use as the active ingredient can be. Generally, health food composition of the present invention hereinafter provided to maintain a number of morning fair food or beverage 15 parts by weight, preferably hereinafter are included in an amount of 10 parts by weight. However in the case of ingestion of the amount said health is hereinafter can have long said range, said range in number because the safety door insofar as the active ingredient can be used in a quantity.

Said special number smooth free type of health food. Said food health food composition can be added examples of meat, sausage, bread, chocolate, candy, snack, confectionery, pizza, if, other noodles, gum type, ice cream acid number a dairy product, various sprocket, beverage, tea drink number, such as alcoholic beverages and vitamin composite number may be, health food comprising both in conventional meaning.

In addition, health food composition of the present invention in food, in particular ethanol bath number can be disclosed. Of the present invention includes the component of a functional food is typically can be added. For example, protein, carbohydrate, fat, nutrient and seasoning comprising the following number. For example, if in addition to the natural carbohydrate or flavor number number number bath to drink ingredient capable of further active ingredient. Natural the carbohydrate (e.g., glucose, such as fructose) said pathogenic agent mono, d company kara id (for example, liver, sucrose or the like), oligosaccharide, polysaccharide (e.g., dextrin, cyclodextrin or the like) or sugar alcohol (e.g., xylitol, sorbitol, such as erythritol) preferably in. The natural flavor (e.g., TAU staining, such as Stevia extract) on said flavor number number number (e.g., saccharin, aspartame or the like) can be using synthetic flavor.

Said health nutraceutical compositions in addition to various nutritional number, vitamin, electrolyte, flavor number, colored number, [pheyk the [thu it buys and salts thereof, alginic acid and salts thereof, organic acid, protective colloidal thickening number, pH control number, stabilizing number, antiseptic number, glycerin, alcohol, further number of calcium hydroxide can be used containing carbonated beverages. The triazines of the present invention the amounts of ingredients added to said large but not significant health nutraceutical compositions 100 parts by weight, 0. 01 To 0. 1 Parts by weight of generally is selected from the range are disclosed.

In addition, the present invention refers to dropping possibility five (PolygonumMultiflorum) A pharmaceutical composition for the prevention or treatment of female menopause diseases containing fermented ingredient number [...] substrate.

Pharmaceutical compositions of the present invention commonly used pharmaceutical composition and production number appropriate carrier, excipient or dilution number can be further includes a number.

The present invention according to composition pharmaceutical dosage forms are combined with suitable set alone or other pharmaceutical active compounds as well as can be used.

The present invention according to pharmaceutical compositions, each an acid number according to conventional method, granules number, positive number, number capsule, suspension, emulsion, syrup, aerosol such as oral number type, external number, number number can be in the form of a sterile injection solution and elder brother anger left on a window. Dropping said fermentation product can be included in the pharmaceutical compositions including possibility five carrier, excipient peak number and dilution number include lactoperoxidase, dextran lactose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methyl hydroxy benzoate, propyl hydroxy benzoate, talc, magnesium stearate and mineral oil or the like various compound or mixture is cited. If filling normally used under anger number number number, a specific number, number coupled, wet number, number disintegrating, dilution number number number or excipients such as surfactants using tank number with each other. Positive number number number is solid for oral administration, ring number, acid number, number granules, such as included in a capsule number, said at least one excipient is possibility five such solid number number number for example dropping or diabetes, starch, calcium purification, sucrose or lactose, gelatin tank pressure is lowered number with each other. In addition simple excipients such as talc and magnesium stearate in addition to lubricating number number are also are used. For liquid oral suspension include number number number, number content, oil number, which number syrup like a simple dilution number corresponding commonly used in water, in addition to the paraffin droplet number various excipients, for example wet number, sweetener number, direction number, number and the like can be preserved. For parental administration number number is a sterile aqueous solution, non-aqueous for number, number suspension, oil number, lyophilized left number number number and multiple myelomas are included. Non-aqueous for number, number suspension include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, such as the [ley which comes the [thu and ethyl ester is used as the injectable can be. As above [theyp brush left number for number, mark with goal, twin 61, carcass five fingers, that it soaked, can be preparing gelatin is used as the alkali.

Pharmaceutical compositions of the present invention the preferred dosage to a patient's state and body weight, degree of disease, forms, according to the stated time routes of different but, can be appropriately selected by one skilled.

Hereinafter, the present invention broadcast receiver using more detailed in the embodiment. In the embodiment of the present invention is more specifically describe the present invention only these for the number one is a person with skill in the art that these range by not notified to the reduced in the art are disclosed.

Material and method

1. Optimum Mushroom mycelium Search

1) Dropping possibility five Inoculation The spawn of fermentation Measuring growth type

Corresponding motor mushroom mycelium possibility five-dropping for establishing optimum condition of fermentation, get hot air-dried and dripping [...] mushroom after same ratio (1:1, w/w) 200 mm diameter 24 mm and height respectively by one selected from a transparent cylindrical in vitro 50g placed next, 15 minutes then heated with the inoculating 121 °C in sterilizing, cordyceps sinensis, stores and skirt 4 mushroom species which code table 1 disclosure to each surface portion dripping possibility secret principles seeding each maltose as oak, box and when the cultured in incubator of the maltose skirt 26 °C, 30 °C him as in the extraction of the spawn situation. Measuring the length of the spawn grown below (mm) from mycelium growth can also comprise a transparent conductive layer, said 4 oak of mushroom mycelium density during the spawn species most transmission option. For improving efficiency of oak mushroom product, total 5 species cultured in industrial Institute (material) changhung group mushroom shiitake mushroom spawn (JMI10075, JMI10076, JMI10077, JMI10078 and KCTC18583P) possibility five pitch using fermentation result, it has been determined that highly order KCTC18583P > JMI10078 > JMI10075 > JMI10076 > JMI10077 spawn density. The use of the present invention based on said result of dripping possibility five KCTC18583P oak maltose decided to spawn final fermentation.

2. Dropping possibility secret principles Searching for fermentation

After grinding and screening body 50 mesh (mesh) and hot air-dried dropping [...], possibility secret principles as to which code table 2 in order to make a mushroom dropping ratio disclosure is mixed. Diameter 200 mm to 24 mm and height adjusting each medium water is 60% transparent cylindrical in vitro by whereupon the 50g each medium, in 30 minutes then heated with the sterilizing 121 °C, shiitake mushroom mycelium seeded onto a surface portion which is bright pitch possibility secret principles 3 disclosure to him as conditions. Mycelia of growth can also measure the length of the leaf is grown to have shown from below.

3. E - screen analysis (E-a screen assay)

E - screen analysis of estrogen agonist to measuring method, estrogen receptor (ER) breast cancer cell lines (MCF-a 7) is inherently expression is the estrogenic material growth promoted by utilizing the experimental method are disclosed. In the present invention 5 species (JMI10075, JMI10076, JMI10077, JMI10078 and KCTC18583P) inoculating mushroom mycelium of fermentable matter whether each possibility five fermented E - screen analysis confirmed that the estrogenic activity of dropping.

Specifically, DMEM containing 10% FBS (Dullbecco's modified Eagle's medium) and adhesive 5% CO MCF-a 7 cells₂ In incubator of 37 °C and him as. When the washed with PBS before cells derived hormones, sample a number for all prior to the estrogenic source (estrogenic source) stand-alone, 2 containing phenol red (phenol red) during 10% CD non-FBS(charcoal-a dextran stripped FBS) without culturing the cells cultured in DMEM 0. Processing 1000 rpm 5 minutes in 25% trypsin - EDTA-centrifuging, number of cells measured 5 × 10³ 96 - Well plate was divided to cell/well. 24 Of the present invention 5 species has been completed (JMI10075, JMI10076, JMI10077, JMI10078 and KCTC18583P) inoculating of mushroom mycelium possibility five 7 and corresponding motor dropping each handling a fermentation product (144 time) after analysis of the MTT proliferation of degree during culturing were measured.

4. Estrogen receptor binding assay (Estrogen receptor binding assay)

Estrogen receptor binding assay (estrogen receptor binding assay) in analysis kits [...] number when the manual proceeds accordingly, binding ability in the presence of sample by polarization value have been determined.

5. Measuring cell survival rate (cell viability)

Mouse macrophage cells (primary) 96 well plate 5 × 10 primary hepatocyte⁵ Cell culturing of mushroom mycelium/ml concentration of corresponding motor dropping a preparation method of the present invention KCTC18583P inoculating each possibility five [...] was dropping. 24 After time, 0. 5 Micro g/ml of MTT (3 - (4, 5 - dimethyl - 2 - thiadiazole and one) - 2, 5 - pen reel [lu the bud which will doze pro mid) 4 so as to form a formazan (formazan) with respect to time. The dimethyl sulfoxide (dimethyl sulfoxide; DMSO) and substances are dissolved in a stand-alone medium number e cup 540 nm absorbance measuring process to an RFID reader in ELISA result controls (control) value was calculated as a ratio.

6. It came - animal model and the [hey e [thok it was recorded (Hematoxylin- Eosin) and May [sunTree chromium(Masson's trichrome) dyeing

The myelin basic protein 7 - 8 week zero detecting BAX female SD does not have to constant temperature (23 ± 1 °C) on humidity (50 ± 5%) during weeks 1 in human use of conditions to produce induced model simulating an adapting piece after surgery when the embodiment under anesthesia both ovarian die number, number of special female menopause model information is normal group (con) treating ovarian, the wetting ability of the present invention KCTC18583P (S) treatment group is the group consisting of ovarian number female menopause model experiment inoculating mushroom mycelium (hereinafter 'sample' multi-) possibility five corresponding motor dropping fermentable matter when the powder into the through oral administration, positive controls are sample for comparative analysis (ascorbic acid) and α - tocopherol (α-a tocopherol) orally administering ascorbic acid instead of a transparent conductive layer, and a group of voice controls are sample executes normal saline feed and his product. Tissue for dyeing, sacrificial animal serum after separation to uterine tissue, ovarian and vaginal separating his order. Tissue observations are uterine, ovarian and vaginal tissue phosphoric acid buffer solution (phosphate buffered paraformaldehyde) 3 hours after formaldehyde content [...] frusta-number to the leucorrhea it fixed to a thickness of 5 μm is formed in paraffin after washing and dehydration in the process of the [hey e [thok it was recorded after cutting it came (Hematoxylin-a Eosin) and conducting dyeing may [sun - chromium (Masson's trichrome) tree.

In the embodiment 1. Dropping possibility five Product Optimum particle size Establishing optimum temperature and

Optimum fermentation dripping possibility five grain size for irradiating, 25 mesh (mesh), through 50 mesh and 100 mesh sieve powder does not pass through the mesh sieve 25 the spawn of residue growth visit from the police. As a result, the particle size according to the disclosure also 1 as dropping a 25 mesh sieve is not passed to the inoculating maltose growth possibility five fastest at the residues 24.4, the first copper layer 25 pass through the mesh in dropping high possibility five appeared. 100 Mesh pass through plant powder possibility five-dropping substantially happened. Culture growth temperature according to the measured results, as disclosure to table 4 to 25 °C spawn density in highest we shall be. The spawn order 27 °C and 23 °C highly density 24.4, 29 °C spawn density in a direction normal to the spawn of the plant rather billion number has been confirmed (2 also). Inoculating the fermentation [...] dropping when using maltose, cultured in the culturing of 25 mesh sieve preferably does not pass through the residual sewage hill 25 °C were confirmed.

¹⁾ +: Poor, + +: Thin, + + +: Sparse, + + + +: Compact, + + + + +: Very compact.

In the embodiment 2. Dropping possibility five Product establishing optimum pH

The inoculating maltose growth in the embodiment 2 to replace the possibility five pH according to dropping in a long time. As a result, the growth of mycelium in pH 6 and 7 to table 3 and 5 when the disclosure to also as good, in the case of pH 4, more drooping spawn growth up. Through these results, pH 6 and 7 has been confirmed that the grid oak maltose growth in acidity one queue.

¹⁾ +: Poor, + +: Thin, + + +: Sparse, + + + +: Compact, + + + + +: Very compact.

In the embodiment 3. Dropping possibility five Product Carbon won And nitrogen won according to confirming the growth of mycelium

Fungi carbon source in carbohydrate, protein, lipid, nucleic acid synthesis and energy as a source such as case as intermediate strain growth are disclosed. The dripping in selecting optimal growth of carbon source in the embodiment 3 to replace the possibility five oak maltose was intended. As a result, 4 to 6 and also to table (glucose, lactose, maltose and sucrose) as disclosure all processing device in growth is preferably 24.4, in particular glucose processing device to replace the possibility five confirmed excellent growth of maltose inoculating dripping in queue.

In addition, PM source tartaric acid ammonium (ammonium tartrate), glutamic acid (glutamic acid), malt extract (malt extract) and potassium nitrate (potassium nitrate) in malt extract processing device to replace the dripping in confirming the crucial possibility five oak mushroom spawn duplication steel pipe were (table 7 and also 5).

In the embodiment 4. Shiitake mushroom mycelium possibility five-dropping of the present invention corresponding motor Fermentable matter Processing according to identifying estrogenic activity

Estrogen receptor (Estrogen receptor) having breast cancer cells MCF-a 7 (breast carcinoma cell) to 5 species (JMI10075, JMI10076, JMI10077, JMI10078, KCTC18583P) inoculating the fermentation product of mushroom mycelium culture during each handling a possibility five corresponding motor dropping 7 after MTT cell proliferation rate analysis measures when the comparative analysis, positive controls to 1 nm of estradiol (Estradiol) was used. E - screen (E-a screen) method measured by estrogenic activity, the disclosure also 6 as 5 species of mushroom mycelium inoculating mushroom mycelium KCTC18583P inoculating corresponding motor dropping during fermentation when possibility five processing, confirming the crucial were MCF-a 7 cell proliferation rate.

5 Species (JMI10075, JMI10076, JMI10077, JMI10078, KCTC18583P) based on said result of dripping water treatment by inoculating each corresponding motor mushroom mycelium fermentation possibility five MCF-a 7 cell proliferation estrogen receptor mediated through for proliferative identify whether, estrogen antagonist ICI 182 in number. 780 (Fulvestrant, 100 nm) were measured changes in cell proliferation rate processing together. As a result, the disclosure also 7 as 5 species of mushroom mycelium possibility five estrogen antagonists in ICI number corresponding motor each dropping inoculating together when it has processed a preparation method, when each corresponding motor mycelium KCTC18583P JMI10078 and dripping possibility five of fermentation, ICI (ici) number reduction is compared to voice processing only regulated billion can be confirmed, in particular of the present invention KCTC18583P fermentation mycelium of corresponding motor-dropping number reduction when it has processed the remarkable effect superior possibility five billion were confirmed. Through, of the present invention treatment or the estrogen receptor activity by dripping possibility five cell proliferation has been confirmed that the MCF-a 7 that were to occur.

In addition, oak mushroom mycelium fermentable matter dropping said 5 species corresponding motor each for anti-estrogen receptor activity was possibility five work performed by comparative analysis. As a result, the disclosure of the present invention KCTC18583P also 8 as of mycelium possibility five corresponding motor dropping when fermentation processing, all concentration (10, 50 and 100 micro g/mL) 4 estrogenic activity in different species each corresponding motor mycelium dropping significantly increased compared fermentation possibility five were confirmed.

In the embodiment 5. Of the present invention KCTC18583PofTo achieve an inoculating mushroom mycelium theFermentation with the non-cytotoxic comparison possibility secret principles dropping pleuropterus multflorus fermentation

The results of said in the embodiment 4 5 species (JMI10075, JMI10076, JMI10077, JMI10078, KCTC18583P) inoculating mushroom mycelium of mushroom mycelium of inoculating corresponding motor dropping possibility five KCTC18583P during fermentation when it has processed, or an estrogenic activity can be confirmed but most, food and pharmaceuticals of the present invention for use in a fermentation and a preparation method for the non-dripping possibility five estrogenic stability function key measures was dropping cytotoxic possibility secret principles of comparison analysis. Breast cancer cells (MCF-a 7) to make said in the embodiment 4 cells use in evaluating stability to the granulocyte-macrophage colony stimulating stem cells and not subject to cytotoxic cells into a suitable conducting experiments. As a result, the disclosure also 9 [...] stem cells and macrophages when it has processed as dropping, depending on concentration significantly decreases while cell survival rate, of the present invention KCTC18583P shiitake mushroom mycelium of fermentation processing comprises the corresponding motor-dropping possibility five indicating cell survival rate (con) similar to making sure that the first call request. Through these results, depending on cell survival rate of the present invention reduced dripping possibility five concentration exhibits cell toxicity but making sure that the capable of dropping possibility five fermentation comprises cells without toxicity.

In the embodiment 6. A flexural rigidity around the number use with treating ovarian A detecting BAX In carrying out animal model using uterine, ovarian and vaginal identifying changes of organization

In the embodiment 6 of the present invention KCTC18583P shiitake mushroom mycelium of fermentation in a corresponding motor-dropping piece of tissue treatment possibility five animal model (uterine, ovarian and vaginal) observed changes. As a result, the disclosure 10 and 11 also as in the case of uterine (Urerus) (Ovary) and ovary, uterus and ovary normal group (Con) voice controls (NC) around the reinforcing treatment when compared with the number, size and thickness has been a brute-regulated overexpression of voice, and ligating uterine tissue necrosis than or discoloration (ligation) portion from a volume reduction and the like ovarian between inhibin receptor. While, and processing of the present invention treating ovarian die number (S) fermentable matter dropping possibility five wine when the magnitude of the normal group (Con) and easily maintained similarly uterine, ovarian necrotic billion were making sure that some number. In the case of vaginal (Vagina) tissue, voice controls (NC) epidermal tissue treating ovarian die number wine in some of the present invention treating ovarian die number (S) fermentable matter and thinned but dropping possibility five positive controls (PC) and wine in ascorbic acid (ascorbic acid) and α - tocopherol (α-a tocopherol) in each handling a group view similar to normal group (Con) making sure that the first call request.