KR102141462B1 - Antibiotic composition containing extract of Acori Gramineri Rhizoma and extract of Magnoliae Cortex - Google Patents

Abstract

The present invention relates to a composition comprising a Seokchangpo extract and a post-bak extract as an active ingredient, a composition comprising a Seokchangpo extract and a post-bak extract as an active ingredient has a synergistic antibacterial activity against various microorganisms, thus proliferation of various bacteria and fungi or It can be usefully used for antibacterial purposes to suppress growth.

Description

Antimicrobial composition containing extract of Acori Gramineri Rhizoma and extract of Magnoliae Cortex}

The present invention relates to an antimicrobial composition comprising a Seokchangpo extract and a thick pepper extract as an active ingredient, and more particularly, to a composition comprising a Seokchangpo extract and a thick pepper extract having synergistic antibacterial activity.

Direct or indirect damage caused by pathogenic microorganisms causes many problems in the economy, environment and medicine. For example, the overall loss of food during the food distribution process in the food industry, the hazards and environmental pollution from the use of excess chemical pesticides for agricultural products in the agricultural industry, the emergence of antibiotic-resistant strains due to the misuse of antibiotics, etc. Causes damage by pathogenic microorganisms.

An antibacterial agent means a substance that has an antimicrobial effect against microorganisms, specifically, a substance that has an antimicrobial effect by inhibiting a system in which a microorganism synthesizes cell walls or proteins, or is produced from such a substance. Antibacterial agents are widely used to treat diseases caused by bacterial infections, and have been mainly produced by a method of chemically synthesizing or separating from natural products such as mold. Most of the antibiotics currently being used are manufactured through chemical synthesis, and have a high cost and have many limitations such as causing side effects.

Recently, in order to solve various problems caused by the use of chemical antibacterial agents, the main task is to develop an antimicrobial composition using natural extracts, but most natural extracts have microorganisms that have antibacterial effects. It is limited, and even if it has an antibacterial effect, its function is exhibited at a very high concentration, and thus it has to be used in combination with an additional antibacterial agent. Therefore, it is necessary to develop an antibacterial substance derived from nature that reduces side effects, which is a major disadvantage of antibacterial agents, and simultaneously has antibacterial activity against various bacteria.

Accordingly, the present inventors have completed the present invention regarding an antimicrobial composition by confirming that the composition of the present invention has an antibacterial activity synergistically compared to an individual stool extract or a thick pepper extract while having no side effects and simultaneously having antimicrobial effects on various microorganisms.

Republic of Korea Patent Publication No. 10-2005-0068283

An object of the present invention is to provide an antimicrobial composition comprising a Seokchangpo extract and a thick pepper extract as an active ingredient.

One aspect of the present invention provides an antimicrobial composition comprising a Seokchangpo extract and a thick pepper extract as an active ingredient.

In the present invention, the term "success (厚朴, Magnoliae Cortex)" refers to the stem or root bark of the afternoon tree ( Magnolia officinalis Rehd. et Wils.), alpha, beta-eudesmol (α, β) It is known to contain -eudesmol, magnolol, honokiol, and obovatol (Biopharmaceutical Research Society, Modern Pharmacy, Hakchangsa, 567-571, 2003). Hubak is bitter, has a warm nature, and has been used in oriental medicine to treat indigestion, sputum, and asthma. Recently, various effects have been reported for anti-cancer, neurological diseases (anxiety, depression, Alzheimer's disease, stroke, etc.), inflammatory diseases, cardiovascular diseases, gastrointestinal diseases, asthma and liver diseases.

The aftertaste used in the present invention can be purchased from nature or commercially available, but is not limited thereto.

In the present invention, "Seokchangpo (石菖蒲, Acori Gramineri Rhizoma)" is a rhizome of rhododendron ( Acorus gramineus Solander), a perennial herbaceous plant, and digestive problems, depression, epilepsy, and central nervous system diseases. It has been used for a long time to treat various diseases. Oriental medicine, Seokchangpo's properties are warm and very tasteful, and it is said to return to the heart and stomach, and it is said to treat the secretion of wind and sweat. It is known that it has the effect of regulating the circulation of the qi, qi 胃祛 지 and swelling vascular system, or blood circulation, releasing the wind, and removing moisture. Pharmacologically, it contains components such as β-asarone, caryophyllene, α-humulene, and sekishone, has a sedative effect, promotes secretion of digestive fluids, inhibits abnormal fermentation of the stomach, and at the same time relieves spasm of intestinal smooth muscle It is said to be.

Seokchangpo used in the present invention can be purchased in nature or commercially available, but is not limited thereto.

In the present invention, "antibacterial" means the ability to resist microorganisms and the ability to inhibit the growth or proliferation of microorganisms, and refers to all mechanisms that are made to defend against the action of microorganisms such as bacteria, fungi, yeast, and the like.

In the present invention, "bacteria" is a creature belonging to bacteria, the most prosperous species among living organisms, most of which are pathogenic bacteria and have the characteristics of prokaryotes, but do not have a nuclear membrane, mitochondrial, or chloroplast-like structure. The size varies from 0.5㎛ to 0.5mm.

In the present invention, "fungus" refers to a group of microorganisms composed of more than 72,000 species including fungi, yeast, and mushrooms, belongs to eukaryotic organisms with a nuclear membrane, and cell organelles such as mitochondria, vesicles, etc. are developed, and chitin , Glucan and the like. Most of them are characterized by forming a mycelium, which is cellular, to grow, develop, and reproduce sexually and asexually. It forms a spore as a propagation body, but some fungi have unicellular proliferation. As a by-product, it is mainly involved in organic decomposition in nature, but some parasitic or symbiotic with plants and animals.

A composition comprising a stool extract and a thick slice extract according to an embodiment of the present invention has superior synergistic antibacterial activity, and has a wide range of antimicrobial activity against microorganisms including bacteria and fungi compared to individual stool extracts or thick slice extracts. .

In the present invention, "synergistic (synergistic) antibacterial activity" or "synergistic (synergistic) antimicrobial effect", the effect of the composition of the mixture of Seokchangpo extract and thick pepper extract is a combination of antimicrobial effects of a single extract that is reasonably expected It means that it shows better antibacterial activity than the additive effect. Therefore, the antimicrobial composition according to one embodiment of the present invention has a synergistic antibacterial activity by mixing a spear pear extract and a thick pepper extract, and can effectively inhibit the growth or growth of microorganisms even at low concentrations without additional antibacterial agents.

According to one embodiment of the present invention, the Seokchangpo extract and the aftertaste extract may be prepared according to conventional methods known in the art. For example, after crushing the dried changchangpo and thick foil, a solvent commonly used for extraction may be added, respectively, and extracted at an appropriate temperature and pressure to prepare a changchangpo extract and a thick foil extract.

According to one embodiment of the present invention, the stool extract is water, alcohol having 1 to 4 carbon atoms, propylene glycol, butylene glycol, glycerin, acetone, ethyl acetate, butyl acetate, chloroform, diethyl ether, dichloromethane, hexane, and these It may be obtained from a solvent extraction method using an extraction solvent selected from the group consisting of a mixture or a supercritical extraction method.

According to one embodiment of the present invention, the aftertaste extract is water, alcohol having 1 to 4 carbon atoms, propylene glycol, butylene glycol, glycerin, acetone, ethyl acetate, butyl acetate, chloroform, diethyl ether, dichloromethane, hexane and these It may be obtained from a solvent extraction method using an extraction solvent selected from the group consisting of a mixture or a supercritical extraction method.

According to an embodiment of the present invention, the alcohol having 1 to 4 carbon atoms may be ethanol or methanol.

In the present invention, the "extract" is an extract obtained by the extraction treatment of a crude drug, a diluent or concentrate of the extract, a dried product obtained by drying the extract, a crude product or a purified product of the extract, or a mixture thereof, using the extract itself and the extract It contains extracts of all formulations that can be formed.

According to one embodiment of the present invention, the extract of Seokchangpo and the aftertaste can be prepared in powder form by additional processes such as distillation under reduced pressure and freeze drying or spray drying.

According to one embodiment of the present invention, Seokchangpo extract and thick pepper extract include not only extracts by the above-described extraction method, but also extracts that have undergone a conventional purification process. For example, separation using an ultrafiltration membrane having a constant molecular weight cut-off value, separation by various chromatography (made for separation according to size, charge, hydrophobicity or affinity), etc. Fractions obtained through various purification methods may also be included in the extract of the present invention.

In the present invention, the "supercritical extraction method" refers to an extraction system in which the principles of the conventional distillation and extraction techniques are combined, and extracts a substance using the supercritical fluid solubility. A "supercritical fluid" is a substance that does not change its state when heat and pressure are applied to a substance, and when heat and pressure are applied beyond a specific temperature (critical temperature) and pressure (critical pressure), the supercritical fluid is a liquid. It is a state in which the state of and gas cannot be distinguished, and has unique characteristics different from that of a general liquid or gas. The supercritical fluid is a compressible fluid that has a medium property between gas and liquid, so it can diffuse well like a gas and dissolve other substances like a liquid, and can also adjust the physical properties to a desired state by changing pressure and temperature.

According to one embodiment of the present invention, the supercritical fluid may be carbon dioxide, water, methane, ethane, propane, ethylene, propylene, methanol, ethanol, or acetone, but is not limited thereto. The conditions of the supercritical fluid should be chemically safe, no corrosion in the device, low critical temperature and pressure, and good solubility. In this regard, carbon dioxide is mainly used in the process. Supercritical carbon dioxide is relatively inexpensive, is harmless to the human body, and is non-flammable and chemically stable. In addition, it is easy to change the properties with a low critical point (73.8 bar, 31.1°C) and can be used by recirculation.

The supercritical extraction method according to an embodiment of the present invention can extract a temperature-sensitive material without denaturation and decomposition, and enables rapid extraction and phase separation.

According to one embodiment of the present invention, the antimicrobial composition may include extracts of Seokchangpo and Hubak extracts respectively extracted by the above extraction method, or may include extracts obtained by mixing the herbal extracts of Seokchangpo and Hubak by the above extraction method.

According to one embodiment of the present invention, the extract of Seokchangpo and the aftertaste extract have a weight ratio of 100:1 to 1:1000, a weight ratio of 100:1 to 1:500, a weight ratio of 100:1 to 1:100, and a 50:1 to 1 :50 weight ratio, 30:1 to 1:30 weight ratio, 20:1 to 1:20 weight ratio, 20:1 to 1:15 weight ratio, 20:1 to 1:10 weight ratio, 15:1 to 1 : 10 weight ratio, 15:1 to 1:5 weight ratio, 10:1 to 1:10 weight ratio, 2:1 to 1:10 weight ratio, or 1:1 to 1:10 weight ratio. have.

According to an embodiment of the present invention, the composition may contain 0.001% by weight to 99.99% by weight, and preferably 0.1% by weight to 99% by weight, of the changchangpo extract and the aftertaste extract, based on the total weight of the composition, which is the use of an antimicrobial composition Depending on the method and purpose of use, it may be appropriately selected by those skilled in the art to which the present invention pertains.

According to one embodiment of the present invention, the antimicrobial composition is Escherichia sp., Pseudomonas sp., Staphylococcus sp., Candida sp., Malassezia sp., Salmonella sp., Acinetobacter sp., Corynebacterium sp., Mycobacterium sp., And it may be to inhibit the growth of one or more microorganisms selected from the group consisting of Micrococcus sp.

According to one embodiment of the present invention, the microorganisms are Escherichia coli , Pseudomonas aeruginosa , Candida albicans , Candida tropicalis , Malesia Pachidermatis ( Malassezia pachydermatis), Salmonella typhimurium (Salmonella typhimurium), Acinetobacter John Sony (Acinetobacter johnsonii), Corynebacterium Ako Lawrence (Corynebacterium accolens), Corynebacterium Army cola Tomb (Corynebacterium amycolatum), Corynebacterium Corynebacterium xerosis , Corynebacterium jeikeium , Staphylococcus aureus , Staphylococcus haemolyticus , Staphylococcus haemolyticus , Staphylococcus war Staphylococcus warneri , Staphylococcus saprophyticus , Staphylococcus cohnii , Staphylococcus hominis , Staphylococcus hominis , Staphylococcus xylos Staphylococcus xylosus , Staphylococcus simulans , Mycobacterium fortuitum , and Micrococcus luteus .

In the present invention, " Escherichia sp. ( Escherichia sp.) strain" is a Gram-negative strain, which is a type of intestinal bacteria and is a causative strain such as food poisoning. For the purposes of the present invention, Escherichia sp. strains include, without limitation, microorganisms that can be used as targets of the antimicrobial composition provided by the present invention. In the present invention, the strain of Escherichia genus may be, for example, E. coli, but is not limited thereto. Escherichia coli constitutes a normal bacterial layer in the intestine and can be a causative agent of nonpathogenic or biliary tract infection, cystitis, pyelonephritis, and urinary tract infection.

By using the antimicrobial composition according to an embodiment of the present invention it is possible to inhibit the growth or growth of the strain of the genus Escherichia, thereby preventing or improving the disease caused by the strain of the genus Escherichia.

In the present invention, " Pseudomonas sp. ( Pseudomonas sp.) strain" is a gram-negative strain, which may be a pathogen or a rot, but is also used in the production of amino acids by amino acid fermentation. For the purposes of the present invention, the strain of Pseudomonas genus includes, without limitation, microorganisms that can be used as targets of the antimicrobial composition provided by the present invention. The strain of the genus Pseudomonas of the present invention may be, for example, Pseudomonas aeruginosa, but is not limited thereto. Pseudomonas aeruginosa is a Gram-negative aerobic bacillus, which has weak pathogenicity to humans, but is widely distributed in the natural environment, causing mixed infections and secondary infections to exacerbate the disease state. In particular, when bio-resistance, such as surgery and burns, is lowered, infection causes sepsis and diarrhea by invading the digestive mucosa.

By using the antimicrobial composition according to an embodiment of the present invention can suppress the growth or growth of the strain Pseudomonas genus, it is possible to prevent or improve the disease caused by the strain Pseudomonas genus.

In the present invention, " Staphylococcus sp. ( Staphylococcus sp.) strain" belongs to the family Micrococcaceae (Micrococcaceae) and is present in the intestinal tract of humans and animals, as well as in the skin and mucous membranes. S. aureus , S. epidermidis, S. saprophyticus , S. haemolyticus , S. hominis, or S. warneri are isolated from human diseases. For the purposes of the present invention, strains of the genus Staphylococcus include, without limitation, microorganisms that can be used as targets of the antimicrobial composition provided by the present invention. The strains of the genus Staphylococcus of the present invention include, for example, Staphylococcus aureus, Staphylococcus hemolyticus, Staphylococcus warnery, Staphylococcus saprophyticus, Staphylococcus Couscous, staphylococcus hominis, staphylococcus xylos or staphylococcus simulations, but is not limited thereto. Staphylococcus aureus is a gram-positive, anaerobic bacterium, which is present on the skin and nasal surfaces of healthy people or livestock, and produces food poisoning by producing heat-resistant exotoxins, purulent skin, otitis media, cystitis, etc. It is known to cause disease.

By using the antimicrobial composition according to an embodiment of the present invention can inhibit the growth or growth of strains of the genus Staphylococcus, the disease caused by strains of the genus Staphylococcus can be prevented or improved.

In the present invention, " Candida sp. ( Candida sp.) strain" is a genus of incomplete bacteriophage fungi, and is a single cell spherical or oval, and is present in the mouth, skin, etc. of the human body or animal, and is harmless to the human body under normal conditions, but has a long life of antibiotics. When used or when the body's resistance to immunity is weakened, it multiplies abnormally in the body and causes candidiasis. In areas of high infection, the mucous membranes of the mouth and genitals, etc., cause itching and pain that the mucous membranes rub.

For the purposes of the present invention, Candida genus strains include, without limitation, microorganisms that can be used as targets of the antimicrobial composition provided by the present invention. The Candida genus strain of the present invention may be, for example, Candida albicans or Candida tropicalis, but is not limited thereto. Candida albicans is a Gram-positive fungus, a kind of skin fungus that is present in the oral, vaginal, intestinal, and digestive tracts of healthy people, but when immunity decreases, proliferation increases, causing blistering inflammation, and inflammation in the digestive tract and oral cavity However, representatively, it causes inflammation in the female genitals, resulting in candidal vaginitis. It has been reported that about 75% of women suffer from candidal vaginitis more than once in the vagina and vulva, and 45% of the female population is a common disease that occurs more than once a year. Candida tropicalis is the leading causative agent of sepsis and spread candidiasis in people with lymphoma, leukemia and diabetes, and is usually found on the gastrointestinal tract and skin surface.

By using the antimicrobial composition according to one embodiment of the present invention can inhibit the growth or growth of the genus Candida strains, it is possible to prevent or improve the disease caused by the genus Candida strains.

In the present invention, the “ Malassezia sp.” strain is mainly a lipid-affecting bacterium and belongs to the normal microflora of the skin, and naturally exists on the skin surface. Survives mainly by forming colonies. Colonization by Malassezia is known to occur most often from late adolescence to early adulthood, when the sebaceous glands are active from birth. Malassezia grows by eating fat and protein from the skin, and thus has high lipase activity and increases the production and secretion of inflammatory cytokines, thereby causing pityrosporum folliculitis, pityriasis versicolor, seborrheic dermatitis, and It can cause dandruff.

For the purposes of the present invention, strains of the genus Malassezia include, without limitation, microorganisms that can be used as targets of the antimicrobial composition provided by the present invention. The strain of the genus Malassezia of the present invention may be, for example, Malesacea pakidermatis, but is not limited thereto.

By using the antimicrobial composition according to an embodiment of the present invention can inhibit the growth or growth of the strain of Malassezia, it is possible to prevent or improve the disease caused by the strain of Malassezia.

In the present invention, " Salmonella sp. ( Salmonella sp.) strain" is a Gram-negative strain, which is not only cold and frozen but also resistant to drying and is a causative strain such as typhoidosis, acute gastroenteritis, or food poisoning.

For the purposes of the present invention, the strain of Salmonella genus includes, without limitation, microorganisms that can be used as targets of the antimicrobial composition provided by the present invention. The strain of Salmonella genus of the present invention may be, for example, Salmonella typhimurium, but is not limited thereto. Salmonella typhimurium is mainly found in the intestine of the small intestine, and its toxicity is caused by LPS present in the outer membrane of the bacteria, causing acute gastroenteritis and food poisoning.

By using the antimicrobial composition according to an embodiment of the present invention can suppress the growth or growth of the Salmonella genus strain, it is possible to prevent or improve the disease caused by the Salmonella genus strain.

In the present invention, " Corynebacterium sp." is a genus of Actinomycetales, which is a rod-shaped Gram-positive strain that parasitizes the mucous membrane and is well-detected from healthy bodies. It usually combines pathogenicity and toxin production. do.

For the purposes of the present invention, strains of the genus Corynebacterium include, without limitation, microorganisms that can be used as targets of the antimicrobial composition provided by the present invention. The strain of the genus Corynebacterium of the present invention may be, but is not limited to, Corynebacterium acorens, Corynebacterium amicolatum, Corynebacterium xerosis, Corynebacterium jakium, for example.

By using the antimicrobial composition according to an embodiment of the present invention can inhibit the growth or growth of the strain of genus Corynebacterium, the disease caused by the strain of Salmonella can be prevented or improved.

The antimicrobial composition according to one embodiment of the present invention has a wide range of antimicrobial activity against bacteria and fungi, and includes an active ingredient extracted from natural substances, causing toxicity or skin allergies, skin irritation, and environmental hormones compared to chemical synthetic products. There are few side effects, such as potential and causing resistant bacteria.

According to one embodiment of the present invention, the antimicrobial composition disclosed herein may be a composition for skin, scalp, oral, ocular or vaginal. According to one embodiment of the present invention, the antibacterial composition is shampoo, body cleanser, soap, foam cleanser, skin, lotion, cream, ointment, spray, toothpaste, wipes, animal wipes, kitchen cleaner, bathroom cleaner, pet It can be used as a cleaning agent, an antibacterial agent for textiles, a lens cleaner, and a female cleaner.

According to one embodiment of the present invention, the antimicrobial composition may be used as a preservative, preservative, detergent, or cleaner.

In the present invention, preservatives inhibit or sterilize the proliferation of one or more harmful microorganisms or spoilage microorganisms selected from the crowd consisting of bacteria, fungi, and yeast to prevent deterioration or spoilage, chemical changes in food, feed, cosmetics, and medicines, thereby improving nutritional value and freshness. It means the additive used to maintain the. In a narrow sense, it means a substance that can stop or sterilize the growth or growth of decaying microorganisms such as bacteria, mold, and yeast.

As an ideal condition as a preservative, the composition should be non-toxic and should be effective even in trace amounts. Since the antimicrobial composition according to one embodiment of the present invention is a natural ingredient and is not harmful to the human body, it can be safely applied to food, medicine, cosmetics or animal feed.

In one embodiment of the present invention, the preservative may further include additional additional additives appropriately selected according to the purpose and purpose of use, and other active ingredients such as additives commonly used, pigments, surfactants, preservatives, etc. It can be used in admixture with additives. The additive may be prepared by pulverizing, granulating, tableting or liquefying depending on the purpose and use of the additive, and a conventional method for packaging for productization may be used.

In one embodiment of the present invention, the amount of the antimicrobial composition used in the preservative may be appropriately used depending on the purpose of use and the type of application, the application site, the object to be applied, and the like, for example, 0.01 to 50 parts by weight based on the total composition. It can, but is not limited to. Since the antimicrobial composition according to one embodiment of the present invention is a natural component extracted from herbal medicines and is not harmful to the human body, it can be used in various amounts if it is possible to achieve the application purpose due to the characteristics of the product.

Since the antimicrobial composition according to one embodiment of the present invention has excellent antimicrobial activity against various microorganisms, it can be used as a preservative added to foods, quasi-drugs or cosmetic compositions by replacing synthetic preservatives.

The antimicrobial composition according to one embodiment of the present invention may be used as a cleaning agent, and the cleaning agent may be a detergent for clothes, a dishwashing agent, a food cleaning agent, and a household appliance cleaning agent. Since the antimicrobial composition according to one embodiment of the present invention contains natural ingredients extracted from herbal medicines and is not harmful to the human body, it can be used for the purpose of washing or imparting antimicrobial properties to various household products, including kitchenware or food washing. have.

In one embodiment of the present invention, the cleaning agent may further include an appropriately selected additive according to the purpose and purpose of use, and mixed with additives such as other active ingredients, pigments, surfactants, and preservatives, such as commonly used cleaning agents. Can be used.

According to one embodiment of the present invention, the cleaning agent may be manufactured by pulverizing, granulating, tableting or liquefying according to the purpose and use of the cleaning agent, and a conventional method for packaging for productization may be used.

In one embodiment of the present invention, the amount of the antimicrobial composition used in the cleaning agent may be appropriately used according to the purpose of use and application form, application site, object to be applied, etc., for example, 0.01 to 50 parts by weight based on the total composition However, the content of the antimicrobial composition is not limited thereto. Since the antimicrobial composition according to one embodiment of the present invention is a natural component and is not harmful to the human body, it can be used in various amounts if it is possible to achieve the application purpose due to the characteristics of the product.

According to one embodiment of the present invention, the present invention can be applied as an antimicrobial composition as a cleaning agent. The cleaning agent according to one embodiment of the present invention may be used for a hand, mouth or female cleaning agent in a simple manner in order to prevent infection in the body of a pathogenic microorganism, which is often susceptible to infection in everyday life. Since the antimicrobial composition according to one embodiment of the present invention is a natural ingredient and is not harmful to the human body, it can be safely applied even when used as a hand, mouth or female cleanser.

The antimicrobial composition according to one embodiment of the present invention causes inflammation in the female genital organs, which can cause female genitourinary system diseases such as vaginitis, itching and odor, for example Candida albicans and Candida tropicalis It has excellent antibacterial activity against ( Candida tropicalis ) and does not cause skin irritation, so it can be used as a cleanser for women.

The composition comprising the Seokchangpo extract and the aftertaste extract as an active ingredient has a synergistic antibacterial activity against various microorganisms, and thus can be usefully used as an antibacterial application to suppress the growth or growth of bacteria and fungi.

Figure 1 shows the results of evaluating the growth inhibitory effect on C. albicans according to the concentration of Seokchangpo supercritical extract.
Figure 2 shows the results of evaluating the growth inhibitory effect on C. albicans according to the concentration of super thin supercritical extract.
Figure 3 shows the results of evaluating the growth inhibitory effect on C. albicans according to the concentration of methanol extract in Seokchangpo.
Figure 4 shows the results of evaluating the growth inhibitory effect on C. albicans according to the concentration of thick methanol extract.
5 and 6 show a synergistic antibacterial effect on C. albicans when mixing Seokchangpo supercritical extract and thick thin supercritical extract. Specifically, FIG. 5 shows the C. albicans cell density measured when each of Sukchangpo supercritical extract and thick thin supercritical extract is added or mixed, and FIG. 6 shows the growth rate of C. albicans strain when each extract is added to the control group.
FIG. 7 shows a synergistic antibacterial effect on C. albicans when mixing Seokchangpo methanol extract and aftertaste methanol extract, and specifically shows the growth rate of C. albicans strain when each extract is added to the control group.
8 and 9 show a synergistic antibacterial effect on C. tropicalis when mixing Seokchangpo supercritical extract and thick thin supercritical extract. Specifically, FIG. 8 shows the C. tropicalis cell density measured when each of Sukchangpo supercritical extract and thick thin supercritical extract is added or mixed, and FIG. 9 shows the growth rate of the C. tropicalis strain of each extract compared to the control group.

Hereinafter, the present invention will be described in more detail through examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited to these examples.

Example 1. Preparation of herbal extracts

1-1. Preparation of Seokchangpo supercritical extract

Supercritical extraction was performed by requesting EnSNature from Seok Chang-po (Jiyundang Herbal Medicine, Gyeongdong Market, Seoul, Korea). 3220 g of Seokchangpo was added to a 10L supercritical extractor, and extracted for 6 hours using supercritical carbon dioxide at 350 bar and 50°C. After extraction, impurities were removed using a filter paper (whatman™ grade 4 qualitative filter paper; GE Healthcare Life Sciences). The amount of supercritical extract obtained from 3,220 g of Seokchangpo was 50.4 g, and the yield was 1.57%. 0.1007 g of the supercritical extract was dissolved in 1 ml of 50% tween 20 diluted in RPMI 1640 medium to prepare a stock at a concentration of 100 g/L, and then vortexed to uniformly dissolve the extract and used in later experiments.

1-2. Preparation of thick supercritical extract

Supercritical extraction was performed by commissioning Hubak (Jiyundang Herbal Medicine, Gyeongdong Market, Seoul, Korea) to EnSNature. 3200 g of thick foil was introduced into a 10 L supercritical extractor, and extracted for 5 hours using supercritical carbon dioxide under conditions of 350 bar and 50°C. After extraction, impurities were removed using a filter paper (whatman™ grade 4 qualitative filter paper; GE Healthcare Life Sciences). The amount of supercritical extract obtained from 3,200 g of thick foil was 55.0 g, and the yield was 1.72%. 0.1012 g of the supercritical extract was dissolved in 1 ml of 50% tween 20 diluted in RPMI 1640 medium to prepare a stock at a concentration of 100 g/L, and then vortexed to uniformly dissolve the extract and used in later experiments.

1-3. Preparation of Seokchangpo and Hubak extract using methanol solvent

After crushing the changchangpo and thick foil to a size of 5cm or less, 30g or 30g of crushed squash cloth was placed in a flask, and extracted by immersing in 300ml of 99.5% methanol as a solvent in a constant temperature water bath at 50°C for 3 hours. The flask was shaken 4 to 6 times at 30 minute intervals to mix the solvent and the herbal medicine. After extraction, the solids were filtered off using a pressure reducing device using a filter paper (Whatman™ No.1 qualitative filter paper; GE Healthcare Life Sciences). The filtered extract was concentrated using a rotary concentrator and left to stand in a dryer at 60° C. for up to 2 weeks to remove methanol and obtain an extract of solid content. The solid content of 2.46 g of Sukchangpo extract and the solid content of 4.45 g of Shunchang extract were obtained from 30 g of Sukchangpo and 30 g of Sukbak, respectively.

200mg, 100mg, 50mg, and 25mg solids of Seokchangpo methanol extract and 100mg, 50mg, 25mg, 13mg, 6mg, 3mg, and 2mg solids of Mt. After dissolving and dissolving, the extract was dissolved in a solvent as much as possible through a water bath at 50°C for 5 minutes. After vortexing to sufficiently dissolve, after centrifugation at 13,000 rpm for 10 minutes, supernatant excluding insoluble matter was taken and used for further experiments.

Example 2. Measurement of antibacterial activity according to the concentration of Seokchangpo extract or Hubak extract

2-1. Preparation of strain and culture medium

In order to measure the antimicrobial effect of the herbal extract of Candida albicans (Candida albicans, C. albicans or less), KCTC 7965 received pre-sale from the Biological Resources Center (Korean Collection for Type Cultures, Republic of Korea), Candida Tropical faecalis (Candida tropicalis, C. below tropicalis ) KCCM 50075 was purchased from the Korean Culture Center of Microorganisms (Korea).

The composition of the medium used in the experiment is shown in Table 1 below.

SDA medium is used to make C. albicans or C. tropicalis fungi stored at -80°C in an active growth state, and RPMI 1640 medium is C. albicans or C. tropicalis fungus that has been actively grown through SDA medium. Was used when culturing. The finished medium was stored at 4°C.

YPD medium made of solid medium including agar is a medium that makes C. albicans bacteria stored at -80°C active growth state, and YPD liquid medium prepared without agar is C. albicans which has become active growth state. It was used when culturing the bacteria.

2-2. Antibacterial effect of Seokchangpo supercritical extract concentration

CLSI M27-A2 method was used to measure the antibacterial effect on C. albicans according to the concentration of Seokchangpo supercritical extract.

C. albicans KCTC 7965 strain stock was streaked on SDA agar plates and cultured for 2 days at 35°C. A single colony was then taken, suspended in 0.145 mol/L saline and vortexed for 15 seconds. Measure the absorbance of the suspension at 530 nm using an Optizen 2120uv spectrophotometer (Mecasys Co., Korea) to check if the cell density is 0.5 McFarland standard, and if necessary, 0.5 McFarland turbidity. Saline was added as much as possible. Saline was added to a suspension having a McFarland turbidity of 0.5, diluted 100-fold, and further diluted 10-fold using RPMI 1640 medium to dilute to a concentration of 1x10 ³ to 5x10 ³ CFU/mL immediately before being processed on the microplate. One C. albicans fungus suspension was dispensed in 100 µl each on a microplate.

Seokchangpo supercritical extract stock prepared in Example 1-1 (100 g/L) was centrifuged at 13,000 rpm for 10 minutes and supernatant was taken and diluted 50 times with RPMI 1640 medium to have a concentration of 2 g/L. Sukchangpo supercritical extract at a concentration of 2 g/L was serially diluted twice with 1% tween 20 to 1 g/L, 0.5 g/L, 0.25 g/L, 0.126 g/L, 62.5 mg/L, 31.3 mg/L The concentration of Seokchangpo supercritical extract was dispensed in 100 μl each on a microplate containing C. albicans fungus suspension, and 100 μl of RPMI 1640 medium containing 1% tween 20 was not included as a control, without a thick supercritical extract. Finally, the tween20 concentration in the medium was adjusted to 0.5%.

As described above, the microplate containing C. albicans bacteria and Seokchangpo supercritical extract was placed at 35° C. for 48 hours to cultivate and suspend the sinking bacteria, and absorbed at 540 nm with a microplate reader (Dynex Opsys MR™ Microplate Reader). It was measured and expressed as the cell density of C. albicans . This cell density value reflects the proliferation or growth results of C. albicans .

Figure 1 shows the results of evaluating the growth inhibitory effect on C. albicans according to the concentration of Seokchangpo supercritical extract. As shown in FIG. 1, it was confirmed that the growth inhibitory effect of C. albicans increased as the concentration of the extract increased, and showed an inhibitory effect of about 35% compared to the control at the maximum concentration of 1 g/L.

2-3. Antibacterial effect according to the concentration of thick supercritical extract

In the same manner as in Example 2-2, 100 µl of the C. albicans fungus suspension diluted to a concentration of ¹ ×10 ³ to 5×10 ³ CFU/ml immediately before treatment on the microplate was dispensed into the microplate.

The super thin supercritical extract stock prepared in Example 1-2 (100 g/L) was diluted 50-fold with RPMI 1640 medium to a concentration of 2 g/L. 2 g/L super thin supercritical extract at a concentration of 1 g/L, 0.5 g/L, 0.25 g/L, 0.125 g/L, 62.5 mg/L, 31.25 mg/L by serially diluting them twice with 1% tween 20 By dispensing 100 µl each of the super thick supercritical extracts in a microplate containing C. albicans fungus suspension, and 100 µl of RPMI 1640 medium containing 1% tween 20 without the super thin supercritical extract as a control. Finally, the tween20 concentration in the medium was adjusted to 0.5%.

As described above, the microplate containing C. albicans bacteria and super thin supercritical extract was placed at 35°C for 48 hours, cultivated by pipetting the suspended bacteria, and absorbed at 540 nm with a microplate reader (Dynex Opsys MR™ Microplate Reader). It was measured and expressed as the cell density of C. albicans .

Figure 2 shows the results of evaluating the growth inhibitory effect on C. albicans according to the concentration of super thin supercritical extract. As shown in FIG. 2, it can be confirmed that the growth inhibitory effect of C. albicans increases as the concentration of the supercritical superthin extract increases, and the growth inhibition of C. albicans of about 22% compared to the control at the maximum concentration of 1 g/L. It showed an effect.

2-4. Antibacterial effect of Seokchangpo methanol extract or thick methanol extract

C. albicans KCTC 7965 strain stock was streaked on YPD agar plates and cultured at 26°C for 2 days. Thereafter, a single colony was taken and inoculated into 5 ml of YPD medium, followed by pre-incubation for 18 to 24 hours under conditions of 250 rpm and 26°C in a test tube. Using the Optizen 2120uv spectrophotometer (Mecasys Co., Korea), the absorbance was measured at 600 nm to obtain the pre-cultured C. albicans cell density, and using this, the C. albicans strain was inoculated into each test tube so that the OD ₆₀₀ was 0.05. do.

In each test tube, the concentration of methanol extract in Seokchangpo was 1.98g/L, 0.99g/L, 0.5g/L, or 0.25g/L, respectively, and the concentration of the thick methanol extract was 0.99g/L, 0.5g/L, Sukchangpo methanol extract or thick-shell methanol extract prepared in Example 1-3 was added to be 0.25 g/L, 0.13 g/L, 0.06 g/L, 0.03 g/L, or 0.02 g/L. Methanol was added to 5 ml of YPD medium to set YPD medium containing 1% methanol as a control.

The strain was inoculated into the test tube as described above, and the extract was added to incubate at 26° C. for 18 hours. The culture was measured by absorbance at 600 nm using an Optizen 2120uv spectrophotometer (Mecasys Co., Korea), which was expressed as the cell density of C. albicans (left graph in FIGS. 3 and 4), compared with a 1% methanol treatment control group. The growth rate of C. albicans according to the concentration of Seokchangpo methanol extract or thick methanol extract was shown (right graph in FIGS. 3 and 4).

Figure 3 shows the results of evaluating the growth inhibitory effect on C. albicans according to the concentration of methanol extract in Seokchangpo. As shown in FIG. 3, the growth of C. albicans at a concentration of 0.99 g/L was suppressed by 18.35%, and thus a methanol extract of Seokchangpo at a concentration of 0.99 g/L was used in a subsequent experiment.

Figure 4 shows the results of evaluating the growth inhibitory effect on C. albicans according to the concentration of thick methanol extract. As shown in FIG. 4, since the growth of C. albicans was suppressed by 22.44% at the concentration of 0.06 g/L, the thick methanol extract at the concentration of 0.06 g/L was used in future experiments.

Example 3. Mixing of Seokchangpo extract and Hubak extract C. albicans Synergistic antibacterial effect

3-1. Synergistic effect of mixing Seokchangpo supercritical extract and thick thin supercritical extract

Based on the results of Examples 2-2 and 2-3, even if the concentration of the extract is doubled, the effect of growth inhibition is not more than 50% and does not show growth inhibition even at the concentration, Seokchangpo supercritical extract 0.125g /L, superthin supercritical extract 0.25g/L was used to examine whether there is a synergistic antibacterial effect on C. albicans according to the mixing of Seokchangpo supercritical extract and superthin supercritical extract. The concentration of each extract used in the experiment is listed in Table 2.

Each extract was treated at the concentration shown in Table 2, and absorbance was measured at 540 nm in the same manner as in Example 2-2, and this was expressed as the cell density of C. albicans . Sukchangpo supercritical extract 0.125g/L or thick thin supercritical extract 0.25g/L contains 0.25% tween 20, so Sukchangpo supercritical extract 0.125g/L or thick thin supercritical extract 0.25g/L adds 0.25% tween 20 The included RPMI 1640 medium was used as a control (indicated as 0.25% tween 20 in FIG. 5) to calculate C. albicans growth rate as a percentage. In addition, Sukchangpo supercritical extract 0.25g/L, Sukchang supercritical extract 0.5g/L, or a mixture of Sukchangpo extract and Sukbak extract contains 0.5% tween 20, so Sukchangpo supercritical extract 0.25g/L or Sukchang supercritical extract RPMI 1640 medium containing 0.5 g/L or 0.5% tween 20 when the mixture was added was used as a control (indicated as 0.5% tween 20 in FIG. 5) to calculate the growth rate of C. albicans as a percentage. The relative strain growth rate (Relative cell density, %) was calculated by Equation 1 below and the results are shown in Table 2.

When the growth rate (%) of the microorganism when the extract of Seokchangpo was treated was A, and the growth rate (%) when treated with the aftertaste extract was B, the expected growth rate (%) was calculated by Equation 2 below.

When the growth rate (%) when treating the composition containing the extract of thick and spear pear is A+B, if the value of A+B is lower than the expected growth rate calculated by Equation 2, the growth of microorganisms or Since the growth was suppressed than expected, it is judged that there is a synergistic antibacterial effect by mixing the extract.

5 and 6 show a synergistic antimicrobial effect on C. albicans according to mixing Seokchangpo supercritical extract and thick thin supercritical extract. Specifically, FIG. 5 shows the C. albicans cell density measured when each of Sukchangpo supercritical extract and thick thin supercritical extract is added or mixed, and FIG. 6 shows C. albicans growth rate (%) of each extract compared to the control group.

As shown in FIGS. 5 and 6, when Sukchangpo supercritical extract (0.125g/L) or thick supercritical extract (0.25g/L) is added individually, it does not show a C. albicans growth inhibitory effect compared to the control group. However, when treating the composition of a mixture of supercritical superthin extract and Seokchangpo supercritical extract, it showed an effect of inhibiting C. albicans growth of about 30% compared to the control group. The A+B value at this time is 70%, which is 34.4% lower than the expected growth rate of 106.7% due to the mixing of the supercritical extract of Seokchangpo and the supercritical extract of Sukchangpo . It was confirmed that it exhibits an excellent synergistic antibacterial effect.

Moreover, the supercritical superthin extract and Seokchangpo supercritical extract do not show a growth inhibitory effect on C. albicans , but may exhibit antibacterial activity by increasing the concentration of the extract according to the mixing of the supercritical superthin extract and Seokchangpo supercritical extract, As shown in Table 2, Fig. 5 and Fig. 6, compared to the case where the Sukchangpo supercritical extract was treated at a double concentration (0.25 g/L) and the thick thin supercritical extract was treated at a double concentration (0.5 g/L). A composition comprising Seokchangpo supercritical extract (0.125g/L) and thick thin supercritical extract (0.25g/L) shows excellent C. albicans growth inhibitory effect. Therefore, it can be seen from this that the composition of the present invention has an excellent synergistic antimicrobial effect and effectively inhibits growth or proliferation of C. albicans even at low concentrations.

3-2. Synergy of Mixing Seokchangpo Methanol Extract and Thick Methanol Extract

Based on the results of Example 2-4, the antibacterial effect on C. albicans according to the mixture of Seokchangpo methanol extract and Hubakmethanol extract was examined using 0.99g/L of Seokchangpo methanol extract and 0.06g/L of Hubakmethanol extract. The concentration of each extract used in the experiment is listed in Table 3.

As in the method described in Example 2-4, each extract and C. albicans fungus suspension at the concentrations shown in Table 3 were dispensed, and absorbance was measured at 600 nm after cultivation to indicate the cell density of C. albicans . The growth rate (%) of C. albicans was calculated and described in Table 3 as described in Example 3-1.

Figure 7 shows a synergistic antimicrobial effect on C. albicans when mixing Seokchangpo methanol extract and thick methanol extract, and specifically shows the growth rate of C. albicans when each extract is added to the control group.

As shown in FIG. 7, when the methanol extract of Seokchangpo and the methanol extract of Hubakin were treated individually, the growth of C. albicans was inhibited by 42% of the extract of Seokchangpo and 18% of the extract of Hukchang compared to the control group. From the above results, it can be seen that a mixture of Seokchangpo methanol extract and Hubakmethanol extract has a growth inhibitory effect on C. albicans of 2 times and 5 times, respectively, than when treated with Sukchangpo extract and Afternoon extract individually. there was.

At this time, when the expected growth rate is calculated by the method described in Example 3-1, it is 47.56%. On the other hand, when mixed with Seokchangpo methanol extract and thick methanol extract, it showed about 93% of C. albicans growth inhibition effect compared to the control group. At this time, the A+B value was 7%, which was significantly lower than the expected growth rate value (47.56%), and thus it was confirmed that it exhibited excellent synergistic antibacterial effects of Seokchangpo methanol extract and thick methanol extract.

Antibacterial activity may be exhibited by increasing the concentration of the extract according to the mixing of the thick methanol extract and the Seokchangpo methanol extract, but when the concentrations of the Seokchangpo methanol extract and the thick methanol extract are doubled, respectively, 78% and 67% of the control the growth inhibitory effect was observed, since the composition of the solid comprises a seokchangpo methanol extract and magnolia methanol extract than inhibiting the growth of C. albicans, the composition of the present invention and the excellent synergistic antimicrobial effect is present, effective even at low concentration C. It can be seen that albicans has an inhibitory effect on growth or proliferation.

Example 4. According to the mixing of Seokchangpo extract and thick pepper extract C. tropicalis Synergistic antibacterial effect

To examine the antibacterial effect on C. tropicalis , 0.5 g/L of supercritical extract from Seokchangpo and 0.5 g/L from superthin supercritical extract were used to treat C. tropicalis during extract treatment according to the modified CLSI M27-A2 method. Cell density was examined.

C. tropicalis KCCM 50075 strain stock was streaked on SDA agar plates and cultured for 2 days at 35°C. A single colony was then taken, suspended in 0.145 mol/L saline and vortexed for 15 seconds. Measure the absorbance of the suspension at 530 nm using an Optizen 2120uv spectrophotometer (Mecasys Co., Korea) to check if the cell density is 0.5 McFarland standard, and if necessary, 0.5 McFarland turbidity. Saline was added as much as possible. Saline was added to a suspension having a McFarland turbidity of 0.5, diluted 100-fold, and further diluted 10-fold using RPMI 1640 medium to dilute to a concentration of 1x10 ³ to 5x10 ³ CFU/mL immediately before being processed on the microplate. One C. tropicalis fungus suspension was dispensed in 100 µl each on a microplate.

Each plate was dispensed with 100 µl each.

Sukchangpo supercritical extract (2g/L) or 50 µl mixture of thick supercritical extract (2g/L) and 50µl of RPMI 1640 medium, and 50µl of Sukchangpo supercritical extract (2g/L) and supercritical superthin extract (2g/L) ) A composition containing 50 µl (total 100 µl) was dispensed into a microplate containing C. tropicalis fungus suspension, respectively, and the absorbance was measured at 540 nm as in the method described in Example 2-2, and the cells of C. tropicalis . Density was examined, and the growth rate of C. tropicalis was calculated by the method described in Example 3-1 to examine the antibacterial effect of C. tropicalis on the composition of the present invention. Table 4 shows the concentration of the final extract used in the experiment and the growth rate of C. tropicalis .

8 and 9 show a synergistic antibacterial effect on C. tropicalis when mixing Seokchangpo supercritical extract and thick thin supercritical extract. Specifically, FIG. 8 shows the C. tropicalis cell density measured when each of Sukchangpo supercritical extract and thick thin supercritical extract is added or mixed, and FIG. 9 shows the growth rate of the C. tropicalis strain of each extract compared to the control group.

At this time, the relative strain growth rate and expected growth rate were calculated by the method described in Example 3-1. Seokchangpo seconds to extract the critical process of C. tropicalis growth rate is 48.6%, when the supercritical extract of Magnolia treatment of C. tropicalis The growth rate is 63.1%. Therefore, the expected growth rate when these two extracts are mixed is 30.7%. The experimental results of the growth rate when mixed with the seokchangpo magnolia extract of 27.7% because, the growth of C. tropicalis than the expected growth rate inhibition because excellent even for seokchangpo supercritical extract of magnolia and a second C. tropicalis by mixing the extract rising threshold ( synergistic) It was confirmed that it exhibited an antibacterial effect.

Claims (7)

Contains Seokchangpo extract and Hubak extract as active ingredients,
The Seokchangpo extract and the aftertaste extract are obtained from a supercritical extraction method using carbon dioxide or a solvent extraction method using methanol,
Antibacterial composition for inhibiting the growth of one or more strains selected from the group consisting of Candida albicans and Candida tropicalis .
delete delete The method according to claim 1, wherein the extract of Seokchangpo and the aftertaste extract is an antimicrobial composition that is included in a weight ratio of 20:1 to 1:10.
delete delete The method according to claim 1 or claim 4, The antimicrobial composition is an antimicrobial composition that is used as a preservative, preservative, detergent, or cleaner.

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