KR100757219B1 - Antibacterial composition containing the extract of moutan cortex radicis or the compounds isolated therefrom - Google Patents

Abstract

An antibacterial composition containing the extract of Moutan Radicis Cortex or the compounds isolated therefrom is provided to inhibit growth of zoonosis-occurring bacteria with reduced toxicity and tolerance. A composition for inhibiting growth of zoonosis-occurring bacteria contains the extract of Moutan Radicis Cortex or the compounds isolated therefrom including mudanpioside-H or galloyl-oxypaeoniflorin as an effective ingredient, wherein the zoonosis-occurring bacteria include Shigella dysenteriae, Shigella flexneri, Salmonella typhi, Salmonella paratyphi, Salmonella gallinarum, Salmonella typhimurium, Salmonella enteritidis, Vibrio cholerae, Escherichia coli and methicillin-resistant S. aureus(MRSA).

Description

Antibacterial composition containing the extract of Moutan Cortex Radicis or the compounds isolated therefrom

The present invention relates to an antimicrobial composition comprising an extract of the bark or a compound isolated therefrom having an antimicrobial activity.

It is urgent to develop new antimicrobial drugs to cope with 15% market share of medicines and 1.5 trillion won antibiotic market. In addition, in Korea, economic and cultural international exchanges are active, foreign pathogenic bacteria are more likely to be introduced, and livestock imports are increasing, which will accelerate domestic spread of new bacteria. Currently, most of the bacteria present in domestic livestock are resistant to conventional antibiotics, and the treatment is becoming increasingly difficult. These therapeutic problems will be severely transmitted to the human body through livestock products infected with these bacteria. According to recent reports, more than 50% of human infectious diseases appear to be common infectious diseases, so safe livestock supply is a prerequisite for public health. Antibiotic resistance expression of bacteria is not present in some localized antibiotics, but is expressed in almost all drugs currently in use, and the physiological and genetic characterization of the bacteria is expected to have the ability to resist any drug. Among the various bacteria resistant to antibiotics, methicillin-resistant S. aureus ( MRSA), vancomycin-resistant enterococci (VRE), vancomycin-mediated S. aureus. aureus (vancomycin intermediate S. aureus; VISA) , and Escherichia coli (E. coli) with the antibiotic resistance of at least 6 kinds of O157: H7 including a. In Korea alone, MRSA is a nosocomial infection agent in hospitals, and it is almost 80% in large general hospitals. VRE and VISA have also been rampant since the 1990s and have spread throughout the country. Escherichia coli O157: H7 is also resistant to multiple antibiotics due to multiple agent resistance and there is no complete therapeutic antibiotic yet. E. coli has already been a frequent social problem in the United States, Europe, and nearby Japan, and has occurred sporadically in humans and livestock in Korea. And since the import of cattle has been liberalized since 2001, it is certain that E. coli will prevail in Korea when livestock products or live cattle are imported from the E. coli Sangje region. These resistant bacteria do not have special antibiotic selection system in Korea, and the factor of rapid increase in resistance is much larger than in other countries. Increasing antibiotic resistance is a vicious cycle of increasing use of new expensive antibiotics, increasing treatment duration, and mortality, which in turn increases economic and social problems. Therefore, in order to solve this problem, it is urgently necessary to understand the mechanism of resistance that is occurring and to develop a new antibiotic with a structure that does not cause resistance.

In order to cope with the rapidly increasing bacterial drug resistance and effectively treat various bacteria, new drugs different from the existing antibiotics (molecular structure, efficacy of drugs, etc.) are urgently needed. In particular, due to the emergence of super bacteria that are resistant to antibiotics, the development of new antibiotics is urgently required. In addition, the condition of the new drug should be a safe drug with no side effects on human and animal cells as well as bactericidal effect. No matter how good the antibiotic is, the resistance develops over time, and natural medicines are expected to solve this problem. The reason for this is that natural products are new components that have not been known so far, and since individual actions are mixtures of various single components, it is considered that the pharmacological action of the complexes will reduce side effects such as resistance.

Many antibiotics currently used for livestock are at severe levels of resistance and some are close to 100%. In livestock, antibiotics are mainly used as feed additives or for cleaning purposes regularly between 7 and 10 days for broilers. According to the report, the antibiotic resistance rate of food poisoning bacteria such as Salmonella and enterococci isolated from cattle, pigs, and chickens from all slaughterhouses in the country was tested. As such, currently used for livestock, most antibiotics are resistant to bacteria, so it is urgent to develop new antibiotics that can be effectively used for livestock.

Recently, the research trends of medicines are getting serious side effects, toxicity, and resistance, so researches on the development of medicines from natural products are actively being carried out worldwide. On the other hand, in the East, including our country, we have a background that has promoted the steady development of traditional medicine, one of natural medicines, through long periods of time. Antibiotics have been discovered since 1941 when penicillin was clinically used, and more than 5,500 species have been discovered, of which only 100 are fermented and mostly chemically synthesized. In order to be effectively used for the treatment of diseases, the therapeutic dose should not be toxic to the host and the antibacterial effect to the infectious agent should be very large, but it has poisoning or unusual side effects. It is a phenomenon. Therefore, there is a need for research and development of new antibiotics having low toxicity and effective against resistant bacteria, and researches for reducing the appearance and toxicity of resistant bacteria by methods such as co-administration.

The most common incidence of bacterial diseases in Korea is Salmonellosis, which is especially mediated in food for humans. More than 2,300 serotypes have been reported, and their symptoms are similar. Salmonella (Salmonella) can cause a variety of clinical symptoms and there is a variety of toxins such as endotoxins, cell toxin, enterotoxin cells exist. In particular, the pathogen can be transmitted through the intestinal epithelial cells invade host cells, and almost all mammals are susceptible to a representative strains of the most common strains of pathogens that cause food poisoning in people causing zoonotic epidemic Salmonella typhimurium (Salmonella typhimurium ) . It is spread to humans mainly through contaminated meat and usually causes symptoms such as acute enteritis, high fever and sepsis.

Antibiotics have been continuously used everywhere for the control of bacteria, and antibiotic-resistant bacteria have recently emerged due to the misuse and abuse of such antibiotics and antimicrobial substances, which has become a big problem in society. As the awareness of this problem spreads, social atmospheres that prevent the use of antibiotics or antimicrobial substances are being created. In addition, research on substances that can replace antibiotics is being actively conducted, and plants and probiotics that have been used as folk medicine for a long time ( Probiotics have been actively used to obtain antimicrobial and immune enhancing effects.

Accordingly, the present inventors completed the present invention by confirming that the extract of the bark of the present invention and the compound separated therefrom have excellent antibacterial effect while identifying a substance having an antimicrobial effect from natural drug resources.

It is an object of the present invention to provide an antimicrobial composition comprising bark extract and compounds isolated therefrom having low toxicity and resistance to living organisms and having antimicrobial activity against common infectious disease strains.

In order to achieve the above object, the present invention provides an antimicrobial composition against a common infectious disease strain comprising Moutan Cortex Radicis extract as an active ingredient having low toxicity and resistance to living organisms and exhibiting antimicrobial activity.

The present invention also provides mudanpioside-H or galloyl-oxypaeoniflorin and pharmaceutically acceptable salts thereof, which are isolated from the bark skin, which exhibits low toxicity and antimicrobial activity and exhibits antimicrobial activity. It provides an antimicrobial composition comprising.

The present invention provides an antimicrobial adjuvant for the prevention and treatment of common infectious diseases, including the extract of the neck skin as an active ingredient.

In another aspect, the present invention provides an antimicrobial adjuvant for the prevention and treatment of common infectious diseases including mudanpioside-H or galloyl-oxypaeoniflorin isolated from the bark skin as an active ingredient. .

The extract of the bark and the compound isolated therefrom is characterized in that it is extracted and separated from the peony root peel (목 丹 牧, Moutan Cortex Radicis).

In addition, the extract is a crude extract, a polar solvent soluble extract or a non-polar solvent soluble extract, the crude extract is extracted with a solvent selected from water, a lower alcohol having 1 to 4 carbon atoms or a mixed solvent thereof, preferably methanol The polar solvent soluble extract is a solvent selected from water, methanol, butanol or a mixed solvent thereof, preferably an extract soluble in methanol, and the nonpolar solvent soluble extract is a solvent selected from hexane, chloroform, dichloromethane and ethyl acetate, preferably Means extracts soluble in chloroform or ethyl acetate.

Animals of the acquired common infectious diseases include mammals such as pigs, cattle and goats; Fish such as carp and goldfish; And poultry such as pheasants, chickens, ducks, turkeys and the like.

In addition, the acquired common infectious disease strain is Mycobacterium tuberculosis , Mycobacterium bovis , Bacillus anthracis ), Brucella genus abortus, B. melitensis, B. suis, B. canis), Leptospira disease isolates (Leptospira canicola, L. pomona, L. icterohaemorrhagiae, L. ballum), Visionary strain (Pseudomonas mallei), the analogy that the strain (Pseudomonas Pseudomallei ), Erysipelothrix rhusiopathiae (E. insidiosa), Shigella disenteriae, S. flexneri, S. boydii, S. sonnei, Hersinia pestis, Francicella tularensis, Ersinia pseudotuberculosis, Ersinia enterrocerisis enterocolitis, Listeria monocytogenes, Pasteurella multocida, Streptobacillus moniliformis, Spirilium minor (Spirillum minus) Borrelia recurrentis, Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus equimyris, Streptococcus facal S. faecalis, Clostridium tetani, Clostridium perfringens, Clostridium novi, Cl. Novyi, Clostridium septicum, Clostri Cl. Histoliticum, Clostridium sporogenes, Fusobacterium necrophorum, Salmonella typhi, Salmonella paratyphi, Salmonella Salmonella gallinarium, Salmonella typhimurium, Salmonella enteritidis, Champylobacter fetus (subsp.fetus, venerealis), Champilobacter jejuni (C. jejuni, C. coli, Actinomyces bovis, Actinomyces isralii, Nocardia asterioides, Nocardia N. madurae, N. farcinica, Rabies virus, Hantan virus, Seoul virus, Pumara virus, herpes Viruses from the genus Herpesvirus simiae (herpesvirus B), Filoviridae (Marburg virus), Paramyxovirus, Newcastle virus, and Orthopoxvirus. , Pseudocowpoxvirus in Parafaxvirus, Contagious pustular dermatitis virus in Parafaxvirus, Foot-and-mouth disease virus and mouth disease virus, Vesicular stomatitis virus, Lymphocytic choriomeningitis virus of the genus Arenavirus, Lassa fever genus of the Arenavirus virus, Influenza virus of Orthomyxoviridae, Japanese B encephalitis virus of the genus Flavivirus, Aspergillus fumigata ), Cryptococcus neoformans, Candida albicans, Sportoricum scheckii, Coccidioides immitis, Blastomas dermatitis Blastomyces dermatitidis, Histoplasma capsulatum, Haplosporangium parvum, Haflospo Clear Kress sense (H. crescens, Trichphyton schoenleini, Richettia typhi (R. mooseri), Ricketta prowazekii, Rickettsia tsutsugamusi (R. orientali), R. akari, Coxiella burnetii, Chlamydia psittaci, Entamoeba histolytica, Toxoplasna gondii, Barantidium colli (Balantidium coli), Fasciola hepatica, Fasciola gigantica, Clonorchis sinensis, Paragonimos westermanii, Dibotherio cephalas latum (Dibothriocephalus latum), Taenia saginatus, Taenia solium, Echinococcus glanulosus, Ascaris lumbricoides, Ascaris Suum (Ascaris suum), Vibrio cholera (Vibrio cholerae), yikolrayi (Escherichia coli) and S. aureus (methicillin-resistant S. aureus; MRSA ), preferably Vibrio cholerae O1 (Vibrio cholerae O1), Vibrio cholerae O139, Shigella decenteria dysenteriae ), Shigella flexeneri , E. coli O157, Salmonella typhi, Salmonella paratypy A ( Salmonella) paratyphi A), methicillin resistance S. aureus 104 (MRSA 104), methicillin resistance S. aureus 6 (MRSA 6), E. coli O157 ( E. coli O157), methicillin resistance S. aureus ( MRSA), Salmonella Galina Leeum (Salmonella gallinarium), Salmonella typhimurium (Salmonella includes typhimurium), Salmonella Entebbe utility disk (Salmonella enteritidis), E. coli K99 (E. coli K99) and the E. coli K88 (E. coli K88).

Hereinafter, a method of obtaining the extract and the compound of the present invention will be described in detail.

Bark bark extract of the present invention, the dried peony root bark neck bark chopped to about 1 to 20 times, preferably about 2 to 6 times the dry weight of water, C ₁ to C ₄ lower alcohol or a mixed solvent thereof , Preferably with water or methanol, cold extraction, hot water extraction, ultrasonic extraction, for about 1 hour to 10 days, preferably about 2 hours to 5 hours at an extraction temperature of 20 to 150 ℃, preferably 70 to 120 ℃ An extraction method such as reflux cooling extraction, preferably 1 to 10 times, preferably 2 to 7 times repeated extractions obtained using a reflux cooling extraction method, followed by concentration under reduced pressure to obtain a crude extract; A second step of suspending the crude extract in distilled water and extracting each with chloroform and ethyl acetate; A third step of distilling the ethyl acetate solvent extract obtained in the second step into chloroform: methanol (8: 1 → 1: 1 (v / v)) by elution solvent and dividing into 5 fractions; A fourth step of separating the third of the fractions of the third step by Sephadex column chromatography to obtain stepless pioside-ace; Separation of the fourth fraction of the third fraction into seven primary lower fractions by Sephadex column chromatography, the fifth fraction is separated by chromatography using a Bakerbond SPE C-18 column galloyloxy paeoniflorin Through the manufacturing process of the fifth step to obtain the bark bark extract of the present invention and purely separated compounds therefrom can be obtained.

In addition, conventional fractionation processes can also be carried out (Harborne JB, Phytochemical methods: A guide). to modern techniques of plant analysis , 3 rd Ed. , pp. 6-7, 1998).

In another aspect, the present invention provides an antimicrobial composition against the acquired common infectious disease strain comprising the stepless Pioside-AH or galloyloxy paeooniflorin isolated from the bark skin obtained by the above production method.

The extract of the bark of the present invention and the compound isolated therefrom are characterized by low toxicity and resistance and antibacterial activity against the common infectious disease strain.

The compounds of the present invention can be prepared with pharmaceutically acceptable salts and solvates according to methods conventional in the art.

As the pharmaceutically acceptable salt, acid addition salts formed by free acid are useful. Acid addition salts are prepared by conventional methods, for example by dissolving a compound in an excess of aqueous acid solution and precipitating the salt using a water miscible organic solvent such as methanol, ethanol, acetone or acetonitrile. Equimolar amounts of the compound and acid or alcohol (eg, glycol monomethylether) in water can be heated and the mixture can then be evaporated to dryness or the precipitated salts can be suction filtered.

In this case, organic acids and inorganic acids may be used as the free acid, hydrochloric acid, phosphoric acid, sulfuric acid, nitric acid, tartaric acid, etc. may be used as the inorganic acid, and methanesulfonic acid, p -toluenesulfonic acid, acetic acid, trifluoroacetic acid, Citric acid, maleic acid, succinic acid, oxalic acid, benzoic acid, tartaric acid, fumaric acid, manderic acid, propionic acid, citric acid, lactic acid, glycolic acid, gluconic acid ( gluconic acid), galacturonic acid, glutamic acid, glutaric acid, glucuronic acid, glucuronic acid, aspartic acid, ascorbic acid, carbonic acid, vanic acid, hydroiodic acid, and the like.

Bases can also be used to make pharmaceutically acceptable metal salts. An alkali metal or alkaline earth metal salt is obtained by, for example, dissolving a compound in an excess alkali metal hydroxide or alkaline earth metal hydroxide solution, filtering the insoluble compound salt, and then evaporating and drying the filtrate. At this time, as the metal salt, it is particularly suitable to prepare sodium, potassium or calcium salt, and the corresponding silver salt is obtained by reacting an alkali metal or alkaline earth metal salt with a suitable silver salt (for example, silver nitrate).

Pharmaceutically acceptable salts of the compounds of the invention include salts of acidic or basic groups which may be present in the compounds of the invention, unless otherwise indicated. For example, pharmaceutically acceptable salts include sodium, calcium and potassium salts of the hydroxy group, and other pharmaceutically acceptable salts of the amino group include hydrobromide, sulfate, hydrogen sulphate, phosphate, hydrogen phosphate, Hydrogen phosphate, acetate, succinate, citrate, tartrate, lactate, mandelate, methanesulfonate (mesylate) and p -toluenesulfonate (tosylate) salts. It can be prepared through the process.

The antimicrobial composition of the present invention comprises 0.1 to 50% by weight of the extract or compound based on the total weight of the composition.

Antimicrobial compositions comprising the extract or compound of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions.

Pharmaceutical dosage forms of the extracts or compounds of the present invention may be used in the form of their pharmaceutically acceptable salts, and may be used alone or in combination with other pharmaceutically active compounds, as well as in a suitable collection.

Compositions comprising extracts or compounds according to the invention, in the form of powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols and the like, external preparations, suppositories and sterile injectable solutions, respectively, according to conventional methods Can be formulated and used. Carriers, excipients and diluents that may be included in the composition comprising the compound include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate , Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. When formulated, diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, and surfactants are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and such solid preparations may contain at least one excipient such as starch, calcium carbonate, sucrose, or the like. ) Or lactose, gelatin and the like are mixed. In addition to simple excipients, lubricants such as magnesium stearate and 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 sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories. As the non-aqueous solvent and suspending agent, 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.

Preferred dosages of the extracts or compounds 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 preferred effect, the extract or compound of the present invention is preferably administered at 0.0001 to 100 mg / kg, preferably 0.001 to 10 mg / kg per day. Administration may be administered once a day or may be divided several times. The dosage does not limit the scope of the invention in any aspect.

The extracts or compounds of the present invention can be administered to mammals such as mice, mice, livestock, humans, etc. by various routes. All modes of administration can be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection.

Below, The invention is illustrated in detail by the following examples and experimental examples.

However, the following Examples and Experimental Examples are only illustrative of the present invention, the contents of the present invention is not limited by the following Examples, Reference Examples and Experimental Examples.

Example  1. Preparation of Bark Extract

1-1. Neck skin Crude extract  Produce

After drying the dried peony root bark obtained from the University of Korea Medicine in Iksan-si using a grinder to the particle size of less than 30 mesh (mesh) to obtain a wood peel powder, the dried dried wood peel powder (2 kg) 100% methanol solution corresponding to three times the mass was added, reflux cooled extraction at 100 ° C. for 3 hours, filtration and concentration under reduced pressure yielded 217 g of the bark extract.

1-2. Neck skin polar solvent and Nonpolar Solvent  Preparation of Soluble Extracts

Suspended crude bark extract (217 g) obtained in Example 1-1 was suspended in distilled water (1 L) and placed in a separatory funnel, and chloroform (800 mL) and ethyl acetate (800 mL) were sequentially added thereto. After shaking, each of the soluble parts was concentrated using a vacuum concentrator to obtain chloroform soluble extract (56 g), ethyl acetate soluble extract (16 g), and water soluble extract (140 g).

Example  2. Isolation of Compounds from Bark Skin

2-1. Mudanpioside - H of  detach

Chlorophyll ethyl acetate soluble extract (16 g) obtained in Example 1-2 was subjected to chromatography using silica gel column using chloroform: methanol (8: 1 → 1: 1 (v / v)) as the elution solvent. Then, those showing the same aspect in thin layer chromatography were combined, concentrated, and divided into five fractions (A to E). The third fraction, C (1.1 g), was separated by Sephadex LH-20 column chromatography (eluent; Chloroform: Methanol (6: 1 (v / v)) was added, and the solvent fractions eluted between 280 mL and 330 mL were combined and concentrated to give 60 mg of mudanpioside-H (compound 1) having the following physical properties. Was obtained (Ding et al., Chem . Pharm . Bull . , 47 , pp. 652-655, 1999).

Colorless powder; (-)-ESI-MS m / z 615 [M ⁻ H] ⁻

¹ H-NMR (Pyridin- d ₅ , 500 MHz): 2.28 (1H, d, J = 12.8 Hz, H-3a), 2.44 (1H, d, J = 12.8 Hz, H-3b), 3.06 (1H, d, J = 6.4 Hz, H-5), 2.23 (1H, d, J = 8.2 Hz, H-6a), 2.85 (1H, dd, J = 8.2, 6.4 Hz, H-6b), 5.01 (1H, d, J = 12 Hz, H-8a), 5.16 (1H, d, J = 12 Hz, H-8b), 5.92 (1H, s, H-9), 1.67 (3H, s, H-10), 5.14 (1H, d, J = 7.8 Hz, H-1 '), 4.05 (1H, t, J = 7.8 Hz, H-2'), 4.24 (1H, t, J = 7.8 Hz, H-3 ') , 4.09 (1H, t, J = 7.8 Hz, H-4 '), 4.10 (1H, dd, J = 7.8, 5.1 Hz, H-5'), 4.96 (1H, dd, J = 11.2, 5.1 Hz, H-6′a), 5.19 (1H, d, J = 11.2 Hz, H-6′b), 8.13 (2H, d, J = 8.7 Hz, H-2 ′, 6 Hz), 7.06 (2H, d , J = 8.7 Hz, H-3 ˝, 5 ˝), 8.27 (2H, d, J = 8.7 Hz, H-2, 6), 7.18 (2H, d, J = 8.7 Hz, H-3, 5) .

¹³ C-NMR (Pyridin- d ₅ , 125 MHz): 88.8 (C-1), 85.9 (C-2), 44.6 (C-3), 105.9 (C-4), 43.7 (C-5), 22.7 (C-6), 71.7 (C-7), 60.7 (C-8), 101.6 (C-9), 19.7 (C-10), 100.2 (C-1 ′), 74.8 (C-2 ′), 78.1 (C-3 '), 71.8 (C-4'), 75.1 (C-5 '), 64.6 (C-6'), 121.2 (C-1 '), 132.3 (C-2', 6 ') , 116.0 (C-3˝, 5˝), 163.5 (C-4˝), 166.4 (C-7˝), 121.5 (C-1), 132.3 (C-2, 6), 116.1 (C-3, 5), 163.6 (C-4), 166.6 (C-7).

2-2. Of galloyloxypaeoniflorin  detach

Fourth fraction D (2.3 g) of Example 2-1 was subjected to Sephadex LH-20 column chromatography (elution solvent; chloroform: methanol (4: 1 (v / v)) to give 7 The second fraction was subdivided into (D1 ~ D7) Bakerbond SPE C-18 using the fifth fraction, D5 (70 mg), as acetonitrile: water (1: 4 (v / v)) as eluent. Chromatography using a column gave 22 mg of galloyl-oxypaeoniflorin (Compound 2) having the following physical properties (Yoshikawa et al., Chem. Pharm . Bull . , 40 , pp.2248- 2250, 1992).

Colorless powder; (-)-ESI-MS m / z 647 [M ⁻ H] ⁻

¹ H-NMR (CD ₃ OD, 500 MHz): 1.69 (1H, d, J = 12.4 Hz, H-3a), 1.90 (1H, d, J = 12.4 Hz, H-3b), 2.51 (1H, d , J = 6.4 Hz, H-5), 1.73 (1H, d, J = 10.5 Hz, H-6a), 2.43 (1H, dd, J = 10.5, 6.4 Hz, H-6b), 4.64 (2H, br s, H-8), 5.36 (1H, s, H-9), 1.24 (3H, s, H-10), 4.53 (1H, d, J = 7.8 Hz, H-1 ′), 3.24 (1H, t, J = 7.8 Hz, H-2 '), 3.54 (1H, t, J = 7.8 Hz, H-3'), 3.28 (1H, t, J = 7.8 Hz, H-4 '), 3.29 (1H , dd, J = 7.8, 6.8 Hz, H-5 '), 4.44 (1H, dd, J = 12.0, 6.8 Hz, H-6'a), 4.50 (1H, d, J = 12.0 Hz, H-6 B), 7.88 (2H, d, J = 8.7 Hz, H-2 Hz, 6 Hz), 6.82 (2H, d, J = 8.7 Hz, H-3 Hz, 5 Hz), 7.07 (2H, s, H-2, 6).

¹³ C-NMR (CD ₃ OD, 125 MHz): 88.0 (C-1), 85.9 (C-2), 43.1 (C-3), 105.0 (C-4), 42.5 (C-5), 21.6 ( C-6), 70.7 (C-7), 59.8 (C-8), 100.9 (C-9), 18.2 (C-10), 98.7 (C-1 ′), 73.6 (C-2 ′), 76.5 (C-3 '), 70.7 (C-4'), 73.9 (C-5 '), 63.4 (C-6'), 120.7 (C-1 '), 131.7 (C-2', 6 '), 114.9 (C-3 ˝, 5 ˝), 162.4 (C-4 ˝), 166.8 (C-7 ˝), 120.0 (C-1), 108.8 (C-2, 6), 145.3 (C-3, 5 ), 138.6 (C-4), 166.8 (C-7).

Experimental Example  1. Antibacterial test

1-1. Disclosure strains and medium and strain culture

Strains used for the determination of antimicrobial activity are isolated outdoor strains and ATCC standard strains in humans or animals, and the strain list is shown in Table 1. The strain was inoculated in TSB (Tryptic Soy broth; Difco, USA) medium and incubated for 24 hours in a 37 ° C thermostat and used for the experiment.Muller-Hinton (Difco, USA) agar medium was used for visual observation. It was.

List of strains for antimicrobial activity test Name origin Vibrio cholerae O1 JOL375 Vibrio cholerae O139 JOL376 Shigrlla dysenteriae JOL377 Shigella flexeneri JOL378 E. coli O157 ATCC 43890 Salmonella typhi JOL380 Salmonella paratyphi A JOL381 MRSA 104 Person isolate MRSA 6 Animal (chicken) isolate E. coli O157 Animal isolate MRSA ATCC 700698 Shigella dysenteriae ATCC 49557 Salmonella gallinarium Water-based standard strain E. coli O157 ATCC 35150 Salmonella typhimurium Water-based Standard Strains (Sal-13) Salmonella enteritidis Water-based standard strain (Sal-36) E. coli K99 Water-based Standard Strains (E-125) E. coli K88 Water-based Standard Strain (E-126)

1-2.  Check disk ( Bauer - Kirby test )

Bacterial culture was inoculated on the Muller-Hinton agar medium, and the antimicrobial test sample solution was injected into the disk to produce an antibiotic concentration gradient on the medium. The inhibitory ring appears in size depending on the solubility of the antimicrobial substance and the degree of sensitivity of the bacteria, and the sensitivity of each antimicrobial substance is sensitive (S), intermeditely sensitive (I), and resistant ( resistant, R).

A. Determination of Antimicrobial Activity of Crude Bark Extracts

After treating the crude extract with mokdanpi 1 ㎎ / ㎖ concentration, as to results of testing the antibacterial activity by the disk diffusion method shown in Table 2 Vibrio cholerae It was sensitive to seven bacteria including O1 (JOL 375).

Strain Diameter of clean area when processing crude extracts (mm) Vibrio cholerae O1 JOL375 12 Vibrio cholerae O139 JOL376 16 Shigrlla dysenteriae JOL377 11 Shigella flexeneri JOL378 16 E. coli O157 ATCC 43890 11 Salmonella typhi JOL380 - Salmonella paratyphi A JOL381 - MRSA 104 Person Isolation 13 MRSA 6 animal (chicken) isolate 11 E. coli O157 animal isolate -

B. Determination of Antimicrobial Activity of Soluble Extracts of Polydermal and Nonpolar Solvents

After treating the chloroform, ethyl acetate and water soluble extracts of the bark bark obtained from Example 1-2 at a concentration of 1 mg / ml, respectively, the antimicrobial activity was examined by a disk diffusion method. As shown in Table 3, the ethyl acetate soluble extract was All strains showed strong susceptibility.

 Strain Diameter of clean areas when treating polar and non-polar solvent soluble extracts Bark chloroform soluble extract Bark Ethyl Acetate Soluble Extract Bark Water Soluble Extract Vibrio cholerae O1 JOL375 7 15 7 Vibrio cholerae O139 JOL376 11 16 - Shigrlla dysenteriae JOL377 8 14 - Shigella flexeneri JOL378 13 15 - E. coli O157 ATCC 43890 8 10 - Salmonella typhi JOL380 - 11 - Salmonella paratyphi A JOL381 - 13 - MRSA 104 Person Separation 7 12 - MRSA 6 animal (chicken) isolate 7 11 - E. coli O157 animal isolate - 10 -

1-3. Minimum inhibitory concentration ( Minimum inhibitory concentration , MIC ) Measure

Muller-Hinton agar medium was used to determine the minimum inhibitory concentration for 18 test strains of Compound 1 and Compound 2 obtained in Example 2. The bacteria enriched in TSB were re-sterilized using the same medium, and the absorbance was measured at 590 nm to calculate CFU (dog / ml), and then diluted to 1 × 10 ⁵ cells / ml using 0.85% saline.

The measured concentrate was dissolved in Muller-Hinton broth at 55-60 ° C., incubated with diluted bacteria, and incubated in a 37 ° C. thermostat for 24 hours. The cultured bacteria were diluted 100-fold with sterile distilled water, and then 100 μl diluted solution was added to Muller-Hinton agar medium and incubated at 37 ° C for 18 hours, followed by visual observation. Determined by the inhibitory concentration is shown in Table 4. As a result, the compounds obtained in Example 2 (Compound 1, Compound 2) showed excellent antimicrobial activity with a minimum inhibitory concentration of 100 ~ 300 ㎍ / ㎖, respectively, for all 18 species causing the common infectious disease.

Name origin Antimicrobial Activity (Minimum Growth Concentration: μg / ml) Compound 1 Compound 2 P / S ^{a )} K ^{b )} Vibrio cholerae O1 JOL375 200 300 NA ^{c )} 10 Vibrio cholerae O139 JOL376 100 100 NA 10 Shigrlla dysenteriae JOL377 100 100 NA 10 Shigella flexeneri JOL378 100 100 NA 10 E. coli O157 ATCC 43890 100 200 NA 10 Salmonella typhi JOL380 200 200 NA 10 Salmonella paratyphi A JOL381 100 100 NA 10 MRSA 104 Person isolate 300 300 NA 10 MRSA 6 Animal (chicken) isolate 300 300 NA 10 E. coli O157 Animal isolate 100 100 NA 10 MRSA ATCC 700698 300 300 One NA Shigella dysenteriae ATCC 49557 300 300 NA 10 Salmonella gallinarium Water-based standard strain 100 100 NA 10 E. coli O157 ATCC 35150 200 100 NA 10 Salmonella typhimurium Water-based Standard Strains (Sal-13) 200 100 NA 10 Salmonella enteritidis Water-based standard strain (Sal-36) 200 200 NA 10 E. coli K99 Water-based Standard Strains (E-125) 200 200 NA 10 E. coli K88 Water-based Standard Strain (E-126) 200 200 NA 10

^a) : positive control drug (penicillin / streptomycin equivalent mixture), ^b) positive control drug (kanamycin), ^c) : no antimicrobial activity

Although formulation examples of the composition comprising the extract or compound of the present invention will be described, the present invention is not intended to be limited thereto but merely to be described in detail.

Formulation example  One. Powder  Produce

Short skin extract 20 mg

Lactose 100 mg

Talc 10 mg

The above ingredients are mixed and filled in an airtight cloth to prepare a powder.

Formulation example  2. Preparation of Tablets

Stepless Pioside-A Hitch 10 mg

Corn starch 100 mg

Lactose 100 mg

2 mg magnesium stearate

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

Formulation example  3. Manufacture of capsule

Galoyloxypaeoniflorin 10 mg

3 mg of crystalline cellulose

Lactose 14.8 mg

Magnesium Stearate 0.2 mg

According to a conventional capsule preparation method, the above ingredients are mixed and filled into gelatin capsules to prepare capsules.

Formulation example  4. Preparation of Injectables

Stepless Pioside-A Hitch 10 mg

Mannitol 180 mg

Sterile distilled water for injection 2974 mg

Na ₂ HPO ₄ 12H ₂ O 26 mg

According to the conventional method for preparing an injection, the amount of the above ingredient is prepared per ampoule (2 ml).

Formulation example  5. Liquid  Produce

Galloyloxypaeoniflorin 20 mg

10 g of isomerized sugar

5 g of mannitol

Purified water

According to the conventional method of preparing a liquid solution, each component is added and dissolved in purified water, lemon flavor is added to the mixture, and then the above ingredients are mixed, purified water is added to adjust the total amount to 100 ml, and then filled in a brown bottle. The solution is prepared by sterilization.

As mentioned above, the bark extract of the present invention and the compound isolated from the bark of the present invention can be used as an antimicrobial composition by showing an excellent antimicrobial effect against the common common infectious disease strains.

Claims (10)

delete delete delete delete delete Shigella disenteriae, a common infectious disease strain containing mudanpioside-H or galloyl-oxypaeoniflorin isolated from Muton Cortex Radicis as an active ingredient ), S. flexneri, Salmonella typhi, Salmonella paratyphi, Salmonella gallinarium, Salmonella typhimurium, Salmonella enteritidis Salmonella enteritidis), Vibrio cholera (Vibrio cholerae), yikolrayi (Esche r ichia coli) and S. aureus (methicillin-resistant S. aureus; antimicrobial composition of the selected strain from MRSA). delete delete delete Shigella disenteriae, Shigella flexe, containing mudanpioside-H or galloyl-oxypaeoniflorin as an active ingredient, isolated from Mutan Cortex Radicis S. flexneri, Salmonella typhi, Salmonella paratyphi, S. paratyphi, Salmonella gallinarium, Salmonella typhimurium, Salmonella enteritidis, Salmonella enteritidis cholera (Vibrio cholerae), yikolrayi (Esche r ichia coli) and S. aureus; antibiotic supplements for the prevention and treatment of infectious diseases caused by zoonotic strains selected from (methicillin-resistant S. aureus MRSA) .