KR101532833B1 - Antimicrobacterial composition comprising oil from Chamaecyparis obtusa for growth inhibition of microbacteria-antimicrobial resistance - Google Patents

Abstract

The present invention relates to a composition for antibiotics resistant to an antibiotic resistant strain comprising an essential oil component of an opacifying oil as an active ingredient and is useful as an antimicrobial composition for preventing or treating diseases caused by methicillin resistant Staphylococcus aureus (MRSA), vancomycin resistant enterococci (VRE) or Escherichia coli, A pharmaceutical composition capable of inhibiting the growth of an antibiotic-resistant strain containing an antimicrobial oil essential oil component that can inhibit the growth of an antibiotic-resistant strain by acting on Salmonella typhimurium and Enterococcus faecalis, and an antibiotic-resistant strain The therapeutic agent for infectious diseases may be provided as a solution, a suspension, a spray, a patch, a pad, a cream, an ointment, a gel, a tablet or a pill.

Description

[0001] The present invention relates to a pharmaceutical composition having antimicrobial-resistant growth inhibitory activity against microbacterial-antimicrobial resistance against Chamaecyparis obtusa,

The present invention relates to an antimicrobial composition having an antimicrobial composition for inhibiting the growth of an antibiotic-resistant strain comprising an antimicrobial essential oil component as an active ingredient, which is selected from the group consisting of methicillin-resistant Staphylococcus aureus (MRSA), vancomycin resistant enterococci (VRE) A composition for antimicrobial antibacterial activity, which comprises an antimicrobial composition comprising an antimicrobial oil essential oil component capable of inhibiting the growth of an antibiotic resistant strain by acting on Escherichia coli, pneumococcal bacteria, Salmonella typhimurium, and Enterococcus faecalis, and antibiotic resistant strains .

Antibiotics are medicines for treating infections caused by pathogens, and show excellent efficacy against infection. However, when antibiotics are frequently used, strains resistant to antibiotics survive, or mutants may develop strains resistant to antibiotics, resulting in pathogens that are increasingly resistant to antibiotics. For this reason, more powerful antibiotics , Which eventually leads to bacteria that can resist any powerful antibiotic, called superbacteria.

Super bacteria means bacteria that can resist the frequent use of antibiotics and do not die by strong antibiotics. In 1961, MRSA (methicillin-Resistant Staphylococcus aureus, methicillin-resistant Staphylococcus aureus) in the UK and VRSA (Vancomycin -Resistant Staphylococcus Aureus, vancomycin-resistant Staphylococcus aureus) was first reported.

MRSA [methicillin resistant staphylococcus aureus] is called methicillin resistant Staphylococcus aureus. It is found in a large general hospital which uses antibiotics as a main cause of hospital infection. It has a strong survival ability and fertility such as a body part of the air, a doctor, a nurse, a scalpel, a hospital blanket, It infiltrates into the respiratory system and the affected part of the operation patient, causing symptoms such as high fever, chills and low blood pressure.

Mycobaxterium smegmatis is a non-pathogenic acid-fast bacillus. It is a fungus that resides in the vulvar mucosal surface, jiggle, and is also present in the armpit, earwax, skin of soil, and water. It is a thin and long bacterium. It has branching and polymorphism. It is similar to mycobacteria, so it needs to be distinguished especially in urine.

Micrococcus [micrococcus] is a genus of aerobic, gram-positive, spherical bacteria. Catalase-positive, producing acid oxidatively in sugars. Yellow, orange, etc. carotenoid pigments are produced. It grows in a relatively high salt concentration environment and is resistant to drying. Soil, sewage, food, air and so on. Staphylococcus is present in the sputum, and the oxidation-fermentation (OF) test is used for the differentiation between Micrococcus and Staphylococcus. In other words, Micrococcus oxidatively degrades sugar, but Staphylococcus decomposes fermentatively. The base composition of DNA of Micrococcus is distributed between 66 ~ 75%, but the base composition of Staphylococcus DNA is distributed between 30 ~ 40% . The standard species Micrococcus luteus .

Vancomycin-resistant enterococci infection is a type of germ-positive germ (bacterial) that is gram-positive (observed as a deep purple when stained with Gram stain) and has the form of a bacillus or short chain. It also grows well on medium without blood.

Enterococcus is a strain of E. faecalis, E. faecium, E. avium, E. gallinarum, Enterococcus kassel, 19 species such as E. casseliflavus are known. Enterococcus pyricalis is the most common intestinal bacteria detected in humans, followed by Enterococcus peum and Enterococcus avium. Enterococcus pemphigus is more important in terms of hospital infection than antibiotic resistance in Enterococcus faecalis.

Staphylococcus aureus is a type of Staphylococcus aureus that causes infection. This bacterium was made possible by the invention of penicillin in 1941. Since then, a stronger antibiotic, methicillin, has emerged, but once again a strong Staphylococcus aureus strain has emerged that does not listen to other antibiotics as well as methicillin. In 2006, more than 94,000 people in the United States were infected with MRSA, of whom more than 19,000 people died.

Vancomycin, the most potent antibiotic developed, has been used for the treatment of bacteria since the 1950s, but VRSA with resistance has emerged in 1996. VRSA is a super-strong bacterium that threatens life because it infects all kinds of infectious diseases and does not listen to any antibiotics and eventually causes sepsis. Experts warn that if VRSA is widespread, simple surgery, such as appendicitis and caesarean section, will not be easy.

Since the cancer is gathered in the hospital, surgical patients and various infected patients, the main infection site of the fungus is the hospital. Even a healthy person can move if the patient's skin or patient touches the object. The rate of resistance is expressed by the number of bacteria surviving per 100 bacteria after administration of antibiotics. Antibiotic-resistant strains In order to treat infectious diseases, it is necessary to develop antibacterial treatment agents as natural products rather than antibiotics.

On the other hand, Chamaecyparis obtusa, also known as cypress tree, is an evergreen tree of the ginseng plant, a Japanese plant, and is widely grown as a tree planting species in the southern part of Korea through its development, Deodorant, antibacterial agent and so on.

Phytoncide is a self-defense substance of plants. It is a substance that plants excrete or secretes to resist pathogens, insects and fungi, and contains sterilization and insecticides. The constituent material of phytoncide is an organic compound composed of a phenol compound such as terpene, an alkaloid component, a glycoside, etc., and is known to have antimicrobial action, sedative action, deodorizing action and stress relieving action. Such phytoncide is a natural substance, not a chemical synthetic substance, and selectively absorbs bacteria which are absorbed in human body and harmful to human being.

In addition, phytoncide has been known to perform a number of functions such as antimicrobial action, deodorizing action, sedative action, and stress relieving action. In addition, phytoncide has excellent sterilization, antibacterial and cleansing action, cleansing the skin, moisturizing and enhancing the immune function in the body.

It has been reported that when phytoncide is inhaled, the concentration of cortisol, which is a stress hormone, is drastically reduced, and it is known that even when the carcass of an animal rotates in the forest, the smell does not occur due to phytoncide. It also helps to calm the central nervous system and make you feel comfortable, extend your sleeping time and help you get a good night's sleep.

It has also been shown to improve blood circulation and to be effective in hyperlipidemia and thromboembolism, and it has also been known as a powerful antimicrobial effect that does not cause resistance in the human body. Phytoncide, a natural substance, is known to be most effective at preventing house dust mites by almost 90% without any side effects.

In this way, phytoncide is not only antimicrobial, but also relieves stress, strengthens intestines and cardiopulmonary functions, and sterilizes the interior of the wood. Thus, various studies on the application of phytoncide have been conducted.

The present invention is based on the finding that the monoaxial oil essential oil component extracted from the Japanese white almond tree acts as an antibiotic resistant methicillin-resistant Staphylococcus aureus (MRSA), vancomycin resistant enterococci (VRE) or Escherichia coli, pneumococcal bacteria, Salmonella typhimurium and Enterococcus faecalis The present invention provides a pharmaceutical composition having a function of inhibiting the growth of an antibiotic-resistant strain by using an extract of Clostridium perfringens by confirming a function capable of inhibiting the growth of the strain, and an antibiotic-resistant strain infection disease preventive and therapeutic agent using the same.

Korean Patent Laid-Open Publication No. 10-2009-0029431 relates to an antimicrobial composition comprising an essential oil extract of Chamaecyparis obtusa having antimicrobial activity, and a whitening essential oil has a strong antimicrobial activity against Legionella pneumophila, The present invention relates to an antimicrobial composition for Legionella including a whitening essential oil which can be very useful for prevention or treatment of air circulation and pneumonia. Korean Patent Registration No. 10-0548501 discloses an evaluation of antimicrobial, antifungal effect, deodorizing ability and skin toxicity against essential oil and leachate by using white flour and leaf, Antimicrobial agents, and deodorizers. Korean Patent Laid-Open Publication No. 10-2013-023992 discloses an antimicrobial composition for inhibiting bacterial pathogenic bacteria and a livestock feed using the same as an active ingredient of a white essential oil as an active ingredient. The antimicrobial composition for suppressing livestock pathogenic bacteria, Bacteria causing respiratory diseases of livestock are disclosed with respect to an antimicrobial composition for inhibiting Actinobacillus pleuropneumo-niae or Pasteurella multocida and a livestock feed using the antimicrobial composition. Korean Patent Laid-Open Publication No. 10-2009-0039458 discloses a method for preventing dandruff caused by dandruff caused by dandruff, which is a natural substance having a growth inhibitory effect of Pityrosporum Ovale which is a main cause of dandruff, Which exhibits a specific antifungal action against Pityrosporum Ovale. This composition is added to hair composition to make dandruff using hair composition such as shampoo, rinse, aerosol spray, hair styling gel and soap. And which is capable of inhibiting the side effects of the human body through the use of conventional antibiotics. However, the prior art has been shown to be effective in inhibiting the growth inhibition of antibiotic-resistant strains by acting on methicillin-resistant Staphylococcus aureus (MRSA), vancomycin resistant enterococci (VRE) or antibiotic resistant Escherichia coli, pneumococcal bacteria, Salmonella typhimurium and Enterococcus faecalis Which is different from the configuration of the present invention.

The present invention relates to a composition for antibiotics resistant to an antibiotic resistant strain comprising an essential oil component of an opacifying oil as an active ingredient and is useful as an antimicrobial composition for preventing or treating diseases caused by methicillin resistant Staphylococcus aureus (MRSA), vancomycin resistant enterococci (VRE) or Escherichia coli, An antimicrobial composition capable of inhibiting the growth of an antibiotic resistant strain containing an essential oil component of linseed oil which can inhibit the growth of an antibiotic resistant strain by acting on Salmonella typhimurium and Enterococcus faecalis, and antibiotic resistant strains Thereby providing a preventive and therapeutic agent for infectious diseases.

In order to accomplish the above object, the present invention provides an antimicrobial composition comprising an antimicrobial composition comprising a monoaxial oil essential oil component separated and prepared as a solvent for extracting Leaves of Leafy leaf, as an antibiotic-resistant strain (Methicillin-resistant (Staphylococcus aureus) Staphylococcus aureus MRSA 693E, MRSA 4-5, MRSA5-3, MRSA S1, MRSA S3, MRSA U4, MRSA P8, MRSA B15, MRSA2-32, a vancomycin- resistant Staphylococcus aureus (VRSA), and another vancomycin VRE 3, VRE 4, VRE 5, VRE 6, VRE 82, VRE 89, VRE 98, and imipenem resistant resistant bacteria prouding carbapenemase (IMP) resistant vancomycin resistant enterococci (VRE) , IMP 100, IMP 102, IMP 123, IMP 129, antibiotic resistant E. coli, pneumoniae bacillus, Salmonella typhimurium, and Enterococcus faecalis, To provide verification by cypress five days pharmaceutical compositions and antibiotic resistant strains preventing infectious diseases using the same treatment with the growth inhibitory function of the antibiotic resistant strains containing the essential oil.

The simplex oil essential oil component comprises the steps of: a) drying the senescence leaves in a low temperature drier, then cutting the dried senescence leaves into particles of about 1 to 3 mm, and pulverizing the cut senescence leaves through 1,000 to 2,000 meshes; b) feeding the cut wheatgrass leaves in step a) into a collection net and feeding the distilled water into a distillation tank; c) heating the distillation tank to supply steam to pass through the leaved tree leaves, and transferring the steam containing the essential oil component extracted from the steam to the cooler to separate the leathery oil and the white water. Which can inhibit the growth of an antibiotic-resistant strain, which is selected from the group consisting of an antibiotic-resistant strain, a suspension, a spray, a patch, a pad, a cream, an ointment, a gel, a tablet or a pill .

The antibacterial detergent of the present invention inhibits the growth of Gram-negative bacteria and Gram-positive bacteria resistant to beta-lactam-based or non-beta-lactam antibiotics such as vancomycin, penicillin or methicillin, The present invention is useful as an antibiotic composition capable of inhibiting the growth of a superbacterial antibiotic resistant strain capable of preventing and treating various infectious diseases and an antibiotic resistant strain having antibiotic resistant strains using the same.

Fig. 1 shows a schematic view of a monoaxial oil extract.

The detergent composition having the function of inhibiting the growth of the antibiotic-resistant strain comprising the extract of A. falciparum leaves as an active ingredient according to the present invention is characterized in that it is a component of a simplex oil, and the component of the simplex oil is a) Followed by cutting the dried perilla leaves into particles of about 1 to 3 mm; b) pulverizing the cut whey leaves so as to pass 1,000-2,000 meshes; c) feeding the cut whey leaves of step a) into a collection net and feeding the distilled water into a distillation tank; d) The distillation tank is heated to supply steam to pass through the leaved tree leaves, and the water vapor containing the essential oil component extracted from the steam is transferred to the cooler to separate the leaven oil and the white water.

The antibiotic-resistant strain of the antimicrobial composition having at least one function selected from the group consisting of Escherichia coli, pneumococcus, Salmonella typhimurium, Staphylococcus or Enterococcus, and the antibiotic-resistant strain having a growth inhibitory function are selected from the group consisting of methicillin-resistant Staphylococcus aureus Or vancomycin-resistant enterococci, and imipenem-resistant carbapenemase-producing bacteria. The detergent composition may be contained in an amount of 1 to 5% (V / V).

The prophylactic and therapeutic agent for an infectious disease of an antibiotic-resistant strain having a growth-inhibiting function of an antibiotic-resistant strain including an antimicrobial composition is selected from a solution, a suspension, a spray, a patch, a pad, a cream, an ointment, a gel, It can be provided in any one of the forms. Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings.

1. Manufacture of hydrothermal extract of cottonwood

1 shows a process for producing a phytoncide component of the present invention. Mixing of the leaves and the distilled water is carried out at a rate of 1kg: 10 ~ 20L and heated at 100 ℃ for 2-3 hours. In the present example, 10 kg of the white waxy tree was washed with distilled water, mixed with 200 L of distilled water, and heated at 100 캜 for 3 hours in an electric kettle to conduct hot water extraction.

The extract was filtered with a filter cloth of 400 mesh and then concentrated with a rotary evaporator under reduced pressure. The filtrate was further extracted twice with the same amount of distilled water, and then filtered and concentrated under reduced pressure. The concentrated hot water extract was lyophilized in a freeze dryer to obtain 1 kg (10%

2. Production of Phytoncide Cotton White Water and Essential Oil Extract

The leaves were dried for 28 hours in a low temperature drier at 35 ℃ to give a moisture content of 5%. Then, the dried planthopper leaves were cut into particles of about 1-3 mm.

The cut wheatgrass leaves were pulverized so as to pass 1,000 to 2,000 mesh, and 7 kg of the cut wheatgrass leaves were placed in a collection net, and a collection net containing the white birch leaf was placed in the distillation tank. Then, 20 L of distilled water Respectively.

The distillation tank was heated to 120 DEG C and steam was supplied for 4 hours to pass through the senna leaf. The water vapor containing the essential oil component extracted by the steam was transferred to a cooler at -5 ° C and separated into a white oil and a white water.

When the solvent used in the process of extracting or extracting essential oils with distilled water, lower alcohol, or a mixed solvent thereof, or separating the essential oil extract into purified water and monoaxial oil is distilled water, Deg.] C to 120 [deg.] C, preferably 80 to 110 [deg.] C for 1 to 6 hours, preferably 3 to 6 hours,

When the solvent is distilled water, a lower alcohol, or a mixed solvent thereof, the solvent is extracted by a solvent extraction method, a leaching method, a carbon dioxide gas extraction method or a low-temperature extrusion method to 5 to 10 times, preferably 5 to 15 times, (W / v) distilled water, a lower alcohol such as methanol, ethanol, butanol, or a mixed solvent thereof, and then the mixture is allowed to stand for about 10 minutes until the oil of the collecting tube is completely separated from the solvent layer, And recovered as a white oil.

3. Separation of essential oil components from white oil by GC / MS

The analytical data of the oil-extracted oil components were analyzed using a gas chromatograph / mass spectrometer (GC / MS, HP agilent GC: 7890A MS: 5975C). Table 1 shows the gas chromatograph (GC) and mass spectrometer (MS) system conditions. The analysis identifies the GC conditions that optimize the separation and sensitivity of the analyte, and applies this condition to all samples (background sample, actual sample) identically.

Gas Chromatograph (GC) and Mass Spectrometer (MS) System conditions Control variable Gas chromatograph (GC) conditions [1] column J & W Scientific, DB-5 crosslinked 5% phenylmethyl silicone Lib. Search Wiley 275 Carrier gas helium Split / Splitless Split (1:10) Sample injection amount 1 μL Detector MS condition MS source temperature 230 ( ^o C) MS Quad temperature 150 ( ^o C) Oven temperature program Rising temperature Temperature Stop time ( ^o C / min) ( ^o C) (min) Early Stage 1 50 5 Step 2 4 250 4 Step 3 Total operating time 59 min EL conditions 70 eV Mass range 20500amu Mass search method Full Scan [1] Analytical instrument: HPagilentGC: 7890AMS: 5975C

The sample was analyzed by injecting 1.0 μL of sample into GC and electron impact ionization (EI) was used for ionization of the sample. Qualitative analysis was based on full scan mode and selective ion retention time. In order to analyze the components, the consistency with the main ion (m / z) was analyzed using Wiley 275 library search method.

4. Assessment of growth inhibitory antimicrobial efficacy against common infectious and antibiotic-resistant strains of leavened oil

The antimicrobial efficacy against the common infectious strain and the antibiotic resistant strain (super bacteria) of the elixir oil essential oil component was evaluated. Table 2 shows the results of the minimum growth inhibition concentration (MIC) of antimicrobial agents inhibiting the growth of common infectious strains and antibiotic resistant strains of the unilamellar essential oil component.

The strains used in the experiments of the present invention are available from the BRC or ATCC (www.atcc.org). The methicillin resisitant Staphylococcus aureus (MRSA), vancomycin resistant enterococci (VRE), and IMP (imipenem resistant carbapenemase producing bacteria) among the antibiotic resistant bacteria used in the experiment of the present invention are used by the inventors of the present invention Respectively.

The essential oil components were diluted 6 times with two-step dilution method with tween 20 qs (1: 1), mixed with 15.0 mL of meuller hinton agar medium, and the essential oil concentration in the final medium was 2, 1, 0.5, 0.25, 0.125 , 0.0625% (v / v). In the case of tween 20 used for the dilute white oil, a test medium was prepared by mixing 15 ml of meuller hinton agar medium with 0.3 ml of tween 20, 0.6 ml, to confirm that there was no influence on the growth of the test strain.

Each strain was cultured on Mueller hinton agar (DIFCO) medium and cultured at 1 * 10 ^ 6 CFU / mL at the culture temperature of each strain. Each test strain was inoculated on a black plate and incubated at 37 ° C for 18 hours and then visually observed to determine the minimum inhibitory concentration (MIC) of the antimicrobial agent in which the growth of the test strains was inhibited.

The minimum growth inhibitory concentration (MIC) of antimicrobial agents inhibiting the growth of general infectious strains and antibiotic resistant strains of the oil - NO Gram Organisms MIC (%) One + Enterococcus faecalis ATCC 29212 0.25 2 + Staphylococcus aureus KCTC 1928 0.125 3 + Micrococcus luteus ATCC 9341 0.0625 4 + Mycrobacterium smegmatis ATCC 9341 0.0625 5 - Salmonella typhimrium KCTC 1925 One 6 - Pseudomonas aeruginosa 2 7 MRSA 693E 0.25 8 + MRSA 4-5 0.0625 9 + MRSA 5-3 0.0625 10 + VRE 82 0.25 11 + VRE 89 0.25 12 + VRE 98 0.25 13 + VRSA (MRSA2-32) 0.125 14 + MRSA S1 0.25 15 + MRSA S3 0.25 16 + MRSA U4 0.25 17 + MRSA P8 0.25 18 + MRSA B15 0.25 19 IMP 100 0.5 20 IMP 102 0.5 21 IMP 123 2 22 IMP 129 One 23 + VRE 3 0.125 24 + VRE 4 0.125 25 + VRE 5 0.125 26 + VRE 6 0.25 27 - Enterococcus cloacea 2 28 Staphylococcus aureus subsp. Aureus ATCC 10537 0.5 29 Staphylococcus aureus KCTC1 1928 (R209) 0.0625 30 Staphylococcus aureus ATCC 6538P 0.0625

As shown in the results of the experiment shown in Table 2, the waxy essential oil component is a common infectious strain, Enterococcus Faecalis ATCC 29212, Micrococcus luteus ATCC 9341, Mycrobacterium smegmatis ATCC 9341, Salmonella typhimrium KCTC 1925, Pseudomonas aeruginosa , Enterococcus cloacae , Staphylococcus aureus subsp . aureus ATCC 10537, Staphylococcus aureus KCTC 1928 (R209), and Staphylococcus aureus ATCC 6538P in the range of 0.0625 to 2% (v / v).

In addition, the antimicrobial activity against MRSA, VRE, and IMP among antibiotic resistant bacteria isolated from clinical patients was excellent in the range of 0.125 to 2% (v / v). From the above results, it was confirmed that the antibacterial activity against the growth of general infectious and antibiotic resistant strains (super bacteria) of the oil-repellent oil component was inhibited.

5. Manufacture of Antibiotic-Resistant Staphylococcal Infectious Disease Therapeutics by Using Essential Oil Components

As the minimum growth inhibitory concentration (MIC) of antibacterial agents inhibiting the growth of generic infectious strains and antibiotic resistant strains (superbacteria) against bacterial strains of leukocyte essential oil was confirmed, the preparation of antibiotic resistant strains infectious disease therapeutic agent .

1 to 5 parts by weight of an oil-repellent oil essential oil component is mixed with 1 part by weight of glycerin 94 to 98 and other substances such as vitamins, flavonoids, alkaloids and polysaccharides with excellent free radical scavenging ability to prepare antibiotic resistant strain infection disease therapeutic agents.

The antibiotic resistant strain infection disease therapeutic agent prepared according to the present invention can be prepared as a solution, a suspension, a spray, a patch, a pad, a cream, an ointment, a gel, a tablet or a pill, A method of application can be used, and a preferable administration method can be selected according to the formulation.

The daily dose of the composition of the present invention is determined depending on the subject to be administered, the method of administration, and symptoms, and the administration frequency is preferably at least twice per day, but the administration frequency can also be adjusted depending on the symptom level.

The antimicrobial composition containing the whitening essential oil component of the present invention inhibits the growth of Gram-negative bacteria and Gram-positive bacteria resistant to beta-lactam-based or non-beta-lactam antibiotics such as vancomycin, penicillin or methicillin, Can be used for the prophylaxis and treatment of various infectious diseases caused by infection of bacteria, and can be usefully used as a superbacterial antibiotic.

Claims (7)

A pharmaceutical composition for inhibiting the growth of an antibiotic-resistant strain, characterized in that it contains an essential oil component of the unilamellar oil extracted from a Leaves of Leaves of Clove Leaf, as an active ingredient, and has a growth inhibitory function against an Imipenem-resistant carbapenemase-
The pharmaceutical composition for inhibiting the growth of an antibiotic-resistant strain according to claim 1, characterized in that the essential oil component of the secreted oil is contained in an amount of 0.0625 to 5% (v / v)
A pharmaceutical composition for inhibiting the growth of an antibiotic-resistant strain according to claim 2; Wherein the antibiotic-resistant strain is one selected from the group consisting of a solution, a suspension, a spray, a patch, a pad, a cream, an ointment, a gel, a tablet or a pill. delete delete delete delete

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