KR102074558B1 - ANTIBACTERAL COMPOSITION AGAINST PATHOGENIC BACTERIA COMPRISING EXTRACT OF Forsythia suspensa - Google Patents

Abstract

The present invention relates to an antimicrobial composition against Escherichia coli, Salmonella and Staphylococcus aureus comprising a duct extract as an active ingredient.

Description

ANTIBACTERAL COMPOSITION AGAINST PATHOGENIC BACTERIA COMPRISING EXTRACT OF Forsythia suspensa}

The present invention relates to an antimicrobial composition for pathogenic microorganisms, including duct extract, specifically, comprising a duct extract, pathogenic microorganisms causing disease in poultry (e.g., E. coli, Salmonella and Staphylococcus aureus) It relates to a composition for antimicrobial.

Secondary metabolite (natural metabolite) derived from a natural product has a variety of physiological activity functions are actively researched. In the past, it has been used in the form of herbal medicines and drugs using the same for the purpose of treatment, and has been used in foods in the form of spices to increase the flavor of the food and not deteriorate from harmful microorganisms. In addition, by extracting essential oil (essential oil) by a variety of methods, such as distillation, compression, extraction and cold soaking is used for the purpose of inhibiting the growth of microorganisms and expression of harmful factors. To date, more than 200,000 secondary metabolites have been reported in plants. Examples of industrially used secondary metabolites include ginkgo extracted from ginkgo biloba leaves, taxol and yew bark extracts. Aspirin is made by refining salicylic acid.

The domestic poultry industry is taking large-scale, concentrated farming as the industrialization progresses. As a result, various diseases are rapidly transmitted and productivity is lowered. Thus, animal medicines including various antibiotics are used for disease prevention and productivity improvement. Representative animal antibiotics used in the poultry industry include tetracycline series, and are used as additives for formulated feed to promote chicken growth and feed efficiency, including therapeutic prescriptions.

Pathogenic microorganisms that cause disease in poultry, including chickens, include Escherichia coli , Salmonella enteritidis , Salmonella gallinarum , Staphylococcus aureus , Mycoplasma gallisepticum , and these are known to cause sepsis and necrosis. Avian pathogenic Escherichia coli (APEC) infects the poultry's intestine through air and water in the kennel, causing colibacillosis. Chicken E. coli causes respiratory disease and diarrhea, and other pathogenic microbial microbes that cause respiratory diseases in the kennel, and additional respiratory stimuli such as viruses and ammonia gas from feces, can cause a variety of acute sepsis, corditis and enteritis. Accompanied by complications. In addition, infection of chickens in the incubator does not hatch, causing large losses. The genus Salmonella is transmitted through contaminated water or feed in a kennel, and infects poultry and poultry eggs, causing vomiting and diarrhea.

According to the Ministry of Food and Drug Safety, the total sales of animal antibiotics in 2015 was 910 tons, of which 157 tons were used for chickens. In addition, antibiotic susceptibility test showed higher resistance of tetracycline, streptomycin, and ampicillin in E. coli , and higher resistance rate in chickens and chicken carcasses compared to other livestock. Therefore, many studies have been reported on the development of antimicrobial agents using various natural materials to prevent the emergence of multi-drug resistant bacteria and its diseases due to the abuse of antibiotics as described above.

The present invention is to provide an antimicrobial composition for E. coli, Salmonella and Staphylococcus aureus comprising a duct extract as an active ingredient.

However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problem, another task that is not mentioned will be clearly understood by those skilled in the art from the following description.

The present invention provides an antimicrobial composition against Escherichia coli, Salmonella and Staphylococcus aureus comprising a duct extract as an active ingredient.

The duct extract has a total phenolic compound content of 100 to 150 mg gallic acid equivalent (GAE) / g solid; Total flavonoid content 30-80 mg quercetin equivalent (QE) / g solid; And a total saponin content of 100-150 mg quilayapi saponin equivalents (QSE) / g solids.

The concentration of the duct extract may be 0.1 mg / mL to 10 mg / mL.

The extract may further comprise extract.

The Yeongyo extract: The sundae extract: The weight ratio of the individual extract may be 1 to 3: 0 to 3: 0 to 3.

The extract is a total phenolic compound content of 30 to 80 mg gallic acid equivalent (GAE) / g solid; Total flavonoid content 30-80 mg quercetin equivalent (QE) / g solid; And a total saponin content of 50 to 100 mg quillajapi saponin equivalents (QSE) / g solids.

The barberry extract has a total phenolic compound content of 10 to 30 mg gallic acid equivalent (GAE) / g solid; Total flavonoid content 1-20 mg quercetin equivalent (QE) / g solid; And a total saponin content of 50 to 100 mg quillajapi saponin equivalents (QSE) / g solids.

The antimicrobial composition may be selected from the group consisting of pharmaceutical compositions, food additives, feed additives, antiseptic compositions and quasi-drug compositions.

The antimicrobial composition according to the present invention is characterized in that it comprises a duct extract as an active ingredient, pathogenic microorganisms (especially, pathogenic microorganisms causing diseases in poultry, E. coli, Salmonella and Staphylococcus aureus) It can have excellent antimicrobial activity. In addition, by additionally including a capillary extract or barberry extract in a specific weight ratio to the duct extract, against pathogenic microorganisms (especially, pathogenic microorganisms causing diseases in poultry, E. coli, Salmonella and Staphylococcus aureus) It can further increase the antimicrobial activity.

Therefore, the antimicrobial composition according to the present invention can be used in various ways as pharmaceutical compositions, food additives, feed additives, antiseptic compositions and quasi-drug compositions, in particular, as a poultry barn disinfectant and feed additives susceptible to pathogenic microorganisms. Thus, not only the generation of harmful residues in the poultry products can be suppressed, but also the growth and killing of pathogenic microorganisms causing diseases in poultry can be induced within a range of harmless organisms.

The inventors of the present invention, by evaluating the antimicrobial effect on pathogenic microorganisms by using a duct bridge, bokhwa or individual used as a herbal medicinal plant among various natural materials, it can replace the existing animal antibiotics within the harmless range It confirmed and completed this invention.

The present invention provides an antimicrobial composition against Escherichia coli, Salmonella and Staphylococcus aureus comprising a duct extract as an active ingredient.

The antimicrobial composition has an antimicrobial activity against E. coli, Salmonella and Staphylococcus aureus, E. coli, Salmonella and Staphylococcus aureus are considered as a kind of pathogenic microorganisms that cause disease in poultry. Can be. Specifically, E. coli B / 06/63 , Salmonella is S. gallinarum , Staphylococcus aureus is S. aureus SE (A / C / E), TC.

Herein referred to as I "yeongyo (Forsythia suspensa)" is to refer the castle Forsythia or dried fruit of Forsythia per the Oleaceae, and has a hypotensive, antipyretic efficacy and bleeding. In addition, the bridge is characterized by containing a large amount of useful components such as saponin (saponin) and oleanolic acid (oleanolic acid).

The duct extract is dried dry duct (g basis) in water, anhydrous or hydrous lower alcohol of 1-4 carbon atoms, lower ketones of 1-4 carbon atoms, lower esters of 1-4 carbon atoms, chloroform and mixtures thereof (preferably, Water or ethanol) (based on ml) may be prepared by extracting by adding 2 to 20 times of dry bridge, and is preferably prepared by adding 3 to 15 times, but is not limited thereto. At this time, the extraction temperature and pressure may be 30 ℃ to 100 ℃ and 1 atm to 1.5 atm, preferably 70 to 100 ℃ and 1.1 atm to 1.3 atm, but is not limited thereto. The extraction time may be 1 hour to 20 hours, preferably 5 hours to 10 hours, but is not limited thereto. The extraction method may be any one of cold needle, ultrasonic extraction or reflux cooling extraction method, but is not limited thereto. The extraction number is preferably 1 to 5 times, more preferably 2 to 3 times, but is not limited thereto. Optionally, the duct extract can be diluted, concentrated, diluted or concentrated and then dried. Thereafter, the duct bridge extract can be used by freezing, spraying and drying.

The pontoon extract prepared by the method is characterized in that the total polyphenol compound content, the total flavonoid content and the total saponin content is high, specifically, the duct bridge extract is 100 to 150 mg gallic acid equivalent (GAE) / g solid; Total flavonoid content 30-80 mg quercetin equivalent (QE) / g solid; And a total saponin content of 100 to 150 mg quillaillac saponin equivalent (QSE) / g solids, but is not limited thereto.

As described above, the duct extract has a high total phenolic compound content, total flavonoid content and total saponin content, as well as due to useful components such as oleanoic acid, E. coli, Salmonella and Staphylococcus aureus is considered to have an excellent antimicrobial effect Can be.

In addition, the duct extract is condensed tannin content of 0.1 to 1 mg catechin equivalent (CE) / g solids (solid); And reducing sugar content of 1 to 10 mg glucose equivalent (GE) / g solid.

In addition, the concentration of the duct extract may be 0.1 mg / mL to 10 mg / mL, preferably 0.1 mg / mL to 5 mg / mL, more preferably 0.1 mg / mL to 2.5 mg / mL, It is not limited to this. In this case, even if the minimum inhibitory concentration of 2.5 mg / mL, the antibacterial effect against E. coli, Salmonella and Staphylococcus aureus is excellent.

On the other hand, in the antimicrobial composition, it may further comprise a capillary extract or individual extract.

That is, the present invention provides an antimicrobial composition against Escherichia coli, Salmonella and Staphylococcus aureus, which comprises a duct extract, and a capillary extract or barberry extract as active ingredients.

The useful components contained in the duct extract may have antimicrobial activity against Escherichia coli, Salmonella and Staphylococcus aureus, but the useful components contained in the duct extract may include When the useful ingredients are mixed with each other, they exhibit a synergistic effect, further enhancing the antimicrobial activity against Escherichia coli, Salmonella and Staphylococcus aureus.

The term “ inula britannica ” in the present specification is a wildflower found in East Asia such as Korea, China, and Japan, and has been used as a medicinal herb since dried flowers of asteraceae. Condensation is known to be effective in digestive disorders, asthma and bronchitis. Flavonoids such as quercetin, kaempferol and luteolin, and sesquiterpene lactone, It is characterized by containing a large amount of the same useful component.

The sun-dried extract is dried and sun-dried (based on grams) in water, anhydrous or hydrous lower alcohol of 1-4 carbon atoms, lower ketones of 1-4 carbon atoms, lower esters of 1-4 carbon atoms, chloroform and mixtures thereof (preferably May be prepared by extracting by adding 2 times to 20 times the dry sunning of water or ethanol) (based on ml), and preferably by adding 3 times to 15 times to extract, but is not limited thereto. At this time, the extraction temperature and pressure may be 30 ℃ to 100 ℃ and 1 atm to 1.5 atm, preferably 70 to 100 ℃ and 1.1 atm to 1.3 atm, but is not limited thereto. The extraction time may be 1 hour to 20 hours, preferably 5 hours to 10 hours, but is not limited thereto. The extraction method may be any one of cold needle, ultrasonic extraction or reflux cooling extraction method, but is not limited thereto. The extraction number is preferably 1 to 5 times, more preferably 2 to 3 times, but is not limited thereto. Optionally, the mulberry extract can be diluted, concentrated, diluted or concentrated and then dried. Thereafter, the sundae extract can be used by drying, such as freezing, spraying.

Said capillary extract prepared by the method also has a high total polyphenolic compound content, total flavonoid content and total saponin content. Specifically, the capillary extract has a total phenolic compound content of 30 to 80 mg gallic acid equivalent ( GAE) / g solids; Total flavonoid content 30-80 mg quercetin equivalent (QE) / g solid; And a total saponin content of 50 to 100 mg quillajapy saponin equivalent (QSE) / g solids, but is not limited thereto.

In addition, the extract is condensed tannin content of 0.1 to 1 mg catechin equivalent (CE) / g solid (solid); And reducing sugar content of 1 to 10 mg glucose equivalent (GE) / g solid.

The term “brassica juncea ” in this specification refers to mature seeds of mustard belonging to the cruciferaceae. It is native to Eurasia and has been used in many countries for its spice and medicinal ingredients since ancient times. Individuals are characterized by containing a large amount of useful ingredients such as glucosinolate, such as sinigrin and sinalbin, which have a characteristic spicy taste.

The barberry extract may be dried dry (in g) water, anhydrous or hydrous lower alcohols having 1 to 4 carbon atoms, lower ketones having 1 to 4 carbon atoms, lower esters having 1 to 4 carbon atoms, chloroform and mixtures thereof (preferably, Water or ethanol) (based on ml) may be prepared by extracting 2 to 20 times the dry individual, preferably by adding 3 times to 15 times, but is not limited thereto. At this time, the extraction temperature and pressure may be 30 ℃ to 100 ℃ and 1 atm to 1.5 atm, preferably 70 to 100 ℃ and 1.1 atm to 1.3 atm, but is not limited thereto. The extraction time may be 1 hour to 20 hours, preferably 5 hours to 10 hours, but is not limited thereto. The extraction method may be any one of cold needle, ultrasonic extraction or reflux cooling extraction method, but is not limited thereto. The extraction number is preferably 1 to 5 times, more preferably 2 to 3 times, but is not limited thereto. Optionally, the individual extract may be diluted, concentrated, diluted or concentrated and then dried. Thereafter, the extract can be used by freezing, spraying and drying.

The basin extract prepared by the above method is characterized in that the total saponin content is high, specifically, the basin extract contains a total saponin content of 50 to 100 mg quillailla saponin equivalent (QSE) / g solids (solid) Preferably, but not limited to.

In addition, the extract is a total phenolic compound content of 10 to 30 mg gallic acid equivalent (GAE) / g solid; Total flavonoid content 1-20 mg quercetin equivalent (QE) / g solid; Condensed tannin content 0.1-1 mg catechin equivalent (CE) / g solid; And reducing sugar content of 1 to 10 mg glucose equivalent (GE) / g solid.

The duct extract: the extract extract: the weight ratio of the extract of the individual is preferably 1-3: 0-3: 0-3, the duct extract: the extract extract: the weight ratio of the extract of 1-2: It is more preferable that it is 1-2: 0-2, and the weight ratio of the duct extract: the sunk extract: the individual extract is most preferably 1: 1: 1, but is not limited thereto. In this case, the duct bridge extract is an essential natural material material, and does not include it, and if it contains only the extract and the barberry extract, there is a problem that the antimicrobial activity against Salmonella does not appear at all. At this time, when the weight ratio of the duct extract and the ectopic extract is too high, there is a problem that the antimicrobial activity against E. coli is lowered, and if the weight ratio of the individual extract is too high, there is a problem that the antimicrobial activity against Salmonella all decreases.

In addition, the total concentration of the Yeongyo extract, the sundae extract and the barberry extract may be 0.1 mg / mL to 10 mg / mL, preferably 0.1 mg / mL to 5 mg / mL, and from 0.1 mg / mL to More preferably, it is 2.5 mg / mL, but is not limited thereto.

The antimicrobial composition is preferably selected from the group consisting of pharmaceutical compositions, food additives, feed additives, antiseptic compositions and quasi-drug compositions, but is not limited thereto.

Specifically, when the antimicrobial composition is used as an antimicrobial pharmaceutical composition, the antimicrobial pharmaceutical composition may be administered parenterally during clinical administration and may be used in the form of a general pharmaceutical formulation. Parenteral administration may refer to administration via a route other than oral such as rectal, intravenous, peritoneal, muscular, arterial, transdermal, nasal, inhaled, ocular and subcutaneous. When using the antimicrobial pharmaceutical composition as a medicine, it may further contain one or more active ingredients exhibiting the same or similar functions.

That is, the antimicrobial pharmaceutical composition may be administered in various parenteral formulations, and when formulated, it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrating agents, surfactants, etc. that are commonly used. do. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, lyophilized preparations, suppositories. As the non-aqueous solvent and the suspension solvent, 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, Whitepsol, macrogol, Tween 61, cacao butter, uririnji, glycerogelatin and the like can be used.

In addition, the antimicrobial pharmaceutical composition may be used in admixture with various carriers (Carrier) accepted as a drug, such as physiological saline or organic solvent, carbohydrates such as glucose, sucrose or dextran, to increase stability or absorption Antioxidants such as ascorbic acid or glutathione, chelating agents, small molecule proteins or other stabilizers can be used as medicaments.

The effective dose of the antimicrobial pharmaceutical composition is 0.01 to 100 mg / kg, preferably 0.1 to 10 mg / kg, and may be administered once to three times a day.

In the antimicrobial pharmaceutical composition, the total effective amount may be administered to the patient in a single dose by a bolus form or by infusion for a relatively short period of time. Administration may be by this long-term administered fractional treatment protocol. Since the concentration is determined in consideration of various factors such as the age and health status of the patient as well as the route and frequency of treatment of the drug, in view of this, those skilled in the art Appropriate effective dosages for particular applications as pharmaceutical compositions may be determined.

In addition, the antimicrobial adjuvant as the pharmaceutical composition for antimicrobial may be administered at the time of antibiotic administration, with the antibiotic, before the antibiotic treatment, after the antibiotic treatment, and may be used as an adjuvant for enhancing the antimicrobial activity of the antibiotic. The antimicrobial adjuvant preferably has a synergistic effect on antibiotics and antimicrobial activity.

When the antimicrobial composition is used as a food additive, the duct bridge extract may be added as it is, or used together with other food ingredients, and may be appropriately used according to a conventional method. The mixed amount of the active ingredient can be appropriately determined according to the purpose of use thereof. In general, the duct bridge extract is added in an amount of 15 parts by weight or less, preferably 10 parts by weight or less with respect to the raw material. However, in the case of long-term intake, the amount may be below the above range, and since there is no problem in terms of stability, the active ingredient may be used in an amount above the above range.

Examples of the foods include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gums, dairy products including ice cream, various soups, beverages, teas, drinks, alcoholic beverages and vitamin complexes. This includes all foods in the usual sense.

When the antimicrobial composition is used as a feed additive, it replaces the existing antibiotics for animals and suppresses the growth of harmful pathogenic microorganisms to improve the health of the animal body, improve the weight gain and meat quality of the animal body, and improve the milk yield and immunity. There is an increasing effect. The feed composition may be prepared in the form of fermented feed, blended feed, pellet form and silage.

The fermented feed may be prepared by fermenting an organic material by adding various microorganisms or enzymes other than the duct extract, and the compound feed may be prepared by mixing various kinds of general feed and the duct extract. Pellet-type feed may be prepared by applying heat and pressure to the blended feed, etc. in a pellet machine, silage can be prepared by fermenting the green feed to the microorganisms. The wet fermented feed collects and transports organic materials such as food waste, mixes excipients for sterilization and moisture control at a certain ratio, and then ferments at a temperature suitable for fermentation for at least 24 hours so that the water content is about 70%. It can be prepared by adjusting. Fermented dry irrigated feed may be prepared by adjusting the wet fermented feed to contain about 30% to 40% of water by further drying.

When the antimicrobial composition is used as an antiseptic composition, the antiseptic composition includes a food preservative, a cosmetic preservative or a pharmaceutical preservative. The food preservatives, cosmetic preservatives and pharmaceutical preservatives are additives used to prevent the deterioration, decay, discoloration and chemical changes of the pharmaceutical products include fungicides and antioxidants, and inhibits the growth of microorganisms such as bacteria, fungi and yeasts. And functional antibiotics such as inhibiting or sterilizing the growth of decay microorganisms in medicine. Ideal conditions for these preservative compositions should be nontoxic and effective in trace amounts.

When the antimicrobial composition is used as a quasi-drug composition, the duct extract may be added as it is, or used together with other quasi-drugs or quasi-drug components, and may be appropriately used according to a conventional method. The mixing amount of the active ingredient may be appropriately determined depending on the intended use.

Specifically, the quasi-drug composition may be a disinfectant cleaner, shower foam, gagreen, wet tissue, detergent soap, hand wash, humidifier filler, mask, ointment, patch, or filter filler.

In addition, (a) preparing a duct extract; (b) preparing each extract or barberry extract; And (c) it can provide an antimicrobial composition against E. coli, Salmonella and Staphylococcus aureus comprising the step of mixing the extract prepared in step (a), the extract prepared in step (b).

As described above, the antimicrobial composition according to the present invention is characterized in that it comprises a duct extract as an active ingredient, pathogenic microorganisms (especially, pathogenic microorganisms causing diseases in poultry, E. coli, Salmonella and yellow) Staphylococcus aureus) can have excellent antimicrobial activity. In addition, by additionally including a capillary extract or barberry extract in a specific weight ratio to the duct extract, against pathogenic microorganisms (especially, pathogenic microorganisms causing diseases in poultry, E. coli, Salmonella and Staphylococcus aureus) It can further increase the antimicrobial activity.

Therefore, the antimicrobial composition according to the present invention can be used in various ways as pharmaceutical compositions, food additives, feed additives, antiseptic compositions and quasi-drug compositions, in particular, as a poultry barn disinfectant and feed additives susceptible to pathogenic microorganisms. Thus, not only the generation of harmful residues in the poultry products can be suppressed, but also the growth and killing of pathogenic microorganisms causing diseases in poultry can be induced within a range of harmless organisms.

Hereinafter, preferred examples are provided to aid in understanding the present invention. However, the following examples are merely provided to more easily understand the present invention, and the contents of the present invention are not limited by the following examples.

< Example >

Example  One

100 g of the pulverized dry bridge was mixed with 1000 mL of water, and then extracted under a pressure of 80 and about 1.2 atm using a water bath (OCOO Co. Ltd., Boryeong, Korea) for 6 hours. Then, it was lyophilized and stored at -20 to prepare a hot water extract.

Example  2-7 and Comparative example  1 to 3

Yeongyo hot water extract prepared in Example 1; Capillary hot-water extract prepared as follows; Or mixed with hot water extract prepared as follows to the weight ratio as shown in Table 1.

Meanwhile, 100 g of the pulverized dry liquor was mixed with 1000 mL of water, and extracted using a water bath (OCOO Co. Ltd., Boryeong, Korea) for 6 hours at 80 ° C. and a pressure of about 1.2 atm. Then, it was freeze-dried and stored at -20 to prepare a captive hot water extract.

In addition, 100 g of individual pieces were mixed with 1000 mL of water, and then extracted using a water bath (OCOO Co., Ltd., Boryeong, Korea) for 6 hours under pressure of 80 and about 1.2 atm. Then, it was lyophilized and stored at -20 to prepare a hot water extract.

Component and weight ratio Example 1 Yeonkyo Hot Water Extract Sole Example 2 Yeongyo Hot Water Extract: Condensed Hot Water Extract = 1: 1 Example 3 Yeongyo Hydrothermal Extract: Hot Water Extract = 1: 1 Example 4 Yeongyo Hot Water Extract: Convoluted Hot Water Extract: Barberry Hot Water Extract = 1: 1: 1 Example 5 Yeongyo Hydrothermal Extract: Convoluted Hydrothermal Extract: Algae Hydrothermal Extract = 2: 1: 1 Example 6 Yeongyo Hydrothermal Extract: Convoluted Hydrothermal Extract: Barberry Hydrothermal Extract = 1: 2: 1 Example 7 Yeongyo Hydrothermal Extract: Convoluted Hydrothermal Extract: Algae Hydrothermal Extract = 1: 1: 2 Comparative Example 1 Barberry Hot Water Extract Sole Comparative Example 2 Condensed Hot Water Extract Sole Comparative Example 3 Hot Water Extract: Condensed Hot Water Extract = 1: 1

Experimental Example  One

Phenolic compound, flavonoid, saponin, condensation in Yeongyo hot-water extract prepared in Example 1, hot-water hydrolyzed extract prepared in Comparative Example 1, and the abalone hot-water extract prepared in Comparative Example 2 To confirm the presence of condensed tannin and reducing sugar, colorimetric method was used, and the phytochemicals and their contents of each extract are shown in Table 2 below.

Specifically, phenolic compounds were measured using the Folin-Ciocalteu assay. 90 μL of each extract was mixed with 1.8 mL of 2% sodium carbonated solution and 90 μL of Folin-Ciocalteu's reagent for 30 minutes, and then absorbance was confirmed at 752 nm. A standard curve was prepared using gallic acid and its value was calculated and expressed as gallic acid equivalent (GAE) / g solid. Flavonoids were also measured using an aluminum chloride assay. 400 μL of distilled water, 30 μL of 15% sodium nitrate solution, 30 μL of 10% aluminum chloride solution and 400 μL of 4% sodium hydroxide were mixed with 100 μL of each extract, and then 40 μL of distilled water was added to the total volume of 1 mL. After reacting at, the absorbance was measured at 510 nm. Quercetin was used to prepare a standard curve to calculate the value and was expressed as quercetin equivalent (QE) / g solids. In addition, saponin was confirmed using a vanillin assay. 100 μL of each extract was mixed with 100 μL of an 8% vanillin solution dissolved in methanol and 1 mL of 72% sulfuric acid dissolved in methanol. Thereafter, the reaction was carried out at 60 ° C. for 10 minutes, and then the absorbance was measured at 540 nm. A standard curve was prepared using Quillaja saponin to calculate the value and expressed as quillaja saponin equivalent (QSE) / g solid. In addition, condensed tannins were measured using vanillin-HCl assay. 20 μL of each extract was reacted with 600 μL of a 4% vanillin solution dissolved in methanol, 300 μL of concentrated hydrochloric acid in a dark room at 25 ° C. for 20 minutes, and the absorbance was measured at 500 nm. The standard curve was prepared using (+)-catechin and the value was calculated and expressed as catechin equivalent (CE) / g solid. In addition, reducing sugar was confirmed using DNS (3,5-dinitrosalicylic acid) solution. DNS solution was dissolved 2.5 g of 3,5-dinitrosalicylic acid in 25 mL of 80 ℃, 75 g of potassium sodium tartrate and 2 N sodium hydroxide solution, the final volume of 250 mL It was prepared by adding distilled water as possible. After 100 μL of each extract and 100 μL of the DNS solution were reacted in boiling water for 10 minutes, after cooling for 15 minutes on ice, 1 mL of distilled water was added, and the absorbance was measured at 540 nm. The standard curve was prepared using glucose, and the value was calculated and expressed in mg glucose equivalent (GE) / g solid.

Plant chemicals Example 1 Comparative Example 1 Comparative Example 2 Total Phenolic Compound Content
(mg GAE / g solid) 113.3 ± 4.3 17.9 ± 0.1 50.8 ± 2.5 Total flavonoid content
(mg QE / g solid) 40.7 ± 0.2 4.1 ± 0.0 51.6 ± 1.0 Total saponin content
(mg QSE / g solid) 137.1 ± 2.2 62.7 ± 0.3 82.9 ± 2.6 Condensed Tannin Content
(mg CE / g solid) 0.7 ± 0.0 0.4 ± 0.0 0.8 ± 0.0 Reducing Sugar Content
(mg GE / g solid) 7.6 ± 0.2 3.5 ± 0.0 5.0 ± 0.2

As shown in Table 2, both Example 1 and Comparative Examples 1 and 2 were found to have the highest saponin content among the plant chemical components in each extract. In the case of Example 1, it was confirmed that the total phenolic compound content, the total flavonoid content and the total saponin content are significantly higher than in Comparative Examples 1 and 2.

Experimental Example  2

Three kinds of poultry pathogenic microorganisms of Yeon-kyo hot-water extract prepared in Example 1, hot-watered hot-water extract prepared in Comparative Example 1, and capillary hot-water extract prepared in Comparative Example 2 ( E. coli B / 06/63 , S. gallinarum , S. aureus In order to evaluate the antimicrobial effect on SE (A / C / E), TC), a disc diffusion assay was performed, and the results are shown in Table 3 below.

Specifically, each extract was dissolved at a concentration of 100 mg / mL, and then filtered using a 0.45 μm membrane filter, while amoxicillin, erythromycin, gentamicin, and gentamicin, A control group was prepared by dissolving tetracycline and kanamycin in distilled water at a concentration of 0.1 mg / mL. Then, three poultry pathogenic microorganisms cultured in Mueller Hinton Broth (MHB) for 24 hours were plated using a cotton swab sterilized in Mueller Hinton Agar (MHA). Then, a sterile 8 mm disc was placed on the smeared medium, and 50 μL of each extract was dispensed and incubated for 24 hours. After incubation, the antimicrobial effect was evaluated by measuring the size of the inhibition zone generated around the disc.

E. coli B / 06/63 S. gallinarum S. aureus SE
(A / C / E), TC Example 1 10.50 ± 0.41 11.00 ± 0.41 17.17 ± 0.47 Comparative Example 1 - - - Comparative Example 2 - - - Amoxicillin - 11.67 ± 0.47 25.83 ± 0.24 Erythromycin - - 9.50 ± 0.41 Gentamicin 12.50 ± 0.41 13.67 ± 0.24 13.17 ± 0.24 Tetracycline 15.33 ± 0.47 14.67 ± 1.25 21.67 ± 0.47 Kanamycin 10.67 ± 0.47 11.83 ± 0.24 12.17 ± 0.24

As shown in Table 3, in Example 1, unlike Comparative Examples 1 and 2, three poultry pathogenic microorganisms ( E. coli B / 06/63, S. gallinarum , S. aureus It is confirmed to have an antimicrobial effect on SE (A / C / E), TC). This is confirmed to be at or above the level of antibiotics currently available on the market. As discussed above, in Example 1, since the total phenolic compound content, the total flavonoid content and the total saponin content are significantly higher than those of Comparative Examples 1 and 2, three kinds of poultry pathogenic microorganisms ( E. coli B / 06/63 , S. gallinarum , S. aureus SE (A / C / E), TC) have excellent antimicrobial effect. In addition, it can be seen as an effect according to useful components such as oleanolic acid contained in the hot-spring hot water extract prepared in Example 1.

Experimental Example  3

Examples 1 to the extract, and Comparative Example 3 kinds of poultry pathogenic microorganisms of the extract prepared from 1 to 3 prepared in 7 (E. coli B / 06/ 63, S. gallinarum, S. aureus SE (A / C / E In order to evaluate the minimum inhibitory concentrations for), TC), ρ-iodonitrotetrazolium violet (INT) was performed, and the results are shown in Table 4 below.

Specifically, three poultry pathogenic microorganisms ( E. coli B / 06/63 , S. gallinarum , S. aureus SE (A / C / E), TC) were incubated for 24 hours, and then 100 μL of the culture solution and each extract 100 μL was mixed and incubated for 24 hours to determine the minimum inhibitory concentration. Each concentration was tested by diluting twice from 5 mg / mL, and INT reagents were used to stain the viable cells in purple.

Minimum inhibitory concentration (mg / mL) E. coli B / 06/63 S. gallinarum S. aureus SE
(A / C / E), TC Example 1 2.5 2.5 2.5 Example 2 2.5 2.5 1.25 Example 3 5 5 2.5 Example 4 1.25 2.5 1.25 Example 5 2.5 2.5 1.25 Example 6 2.5 2.5 1.25 Example 7 2.5 5 1.25 Comparative Example 1 - - - Comparative Example 2 - - - Comparative Example 3 2.5 - 5

As shown in Table 4, Example for 1-7, three kinds of poultry pathogenic microorganisms (E. coli B / 06/63 , S. gallinarum, S. aureus The minimum inhibitory concentrations for SE (A / C / E), TC) are all determined to be less than 5 mg / mL. In particular, compared to Yeongyo hydrothermal extracts alone, S. aureus when mixed with hot water extracts It was found that the minimum inhibitory concentrations for SE (A / C / E) and TC were lowered, and E. coli was mixed with Yeongyo hot-water extract in a 1: 1: 1 weight ratio of extract and hot water extract. Minimum inhibitory concentration for B / 06/63 is 1.25 mg / mL, minimum inhibitory concentration for S. gallinarum is 2.5 mg / mL, and S. aureus The minimum inhibitory concentration for SE (A / C / E) and TC is 1.25 mg / mL, which is the best antibacterial effect.

The foregoing description of the present invention is intended for illustration, and it will be understood by those skilled in the art that the present invention may be easily modified in other specific forms without changing the technical spirit or essential features of the present invention. will be. Therefore, it should be understood that the embodiments described above are exemplary in all respects and not restrictive.

Claims (8)

Yeongyo hot water extract is included as an active ingredient,
The duct bridge hot water extract is extracted by adding duct bridge to water for 5 hours to 10 hours at 70 ℃ to 100 ℃ and 1.1 to 1.3 atm,
The Yeongyo hot water extract is 100 to 150 mg gallic acid equivalent (GAE) / g solid; Total flavonoid content 30-80 mg quercetin equivalent (QE) / g solid; And a total saponin content of 100 to 150 mg quillajapi saponin equivalents (QSE) / g solid,
Antimicrobial composition against Escherichia coli , Salmonella gallinarum and Staphylococcus aureus .
delete The method of claim 1,
Concentration of the hot-water hot water extract in the antimicrobial composition is 0.1 mg / mL to 10 mg / mL, antimicrobial composition.
The method of claim 1,
An antimicrobial composition further comprising a condensed hydrothermal extract or individual extract.
The method of claim 4, wherein
The Yeongyo hot water extract: The condensed hot water extract: The weight ratio of the hot water extract of the barberry is 1-3: 0-3: 0-3, antimicrobial composition.
The method of claim 4, wherein
Said linearized hot water extract is extracted by adding caped to water at 70 ° C. to 100 ° C. and 1.1 atm to 1.3 atm for 5 hours to 10 hours,
The above-mentioned hot water extract has a total phenolic compound content of 30 to 80 mg gallic acid equivalent (GAE) / g solid; Total flavonoid content 30-80 mg quercetin equivalent (QE) / g solid; And a total saponin content of 50 to 100 mg quillajapi saponin equivalent (QSE) / g solid.
The method of claim 4, wherein
The hot water extract of the barberry is added to the water to extract the water for 5 to 10 hours at 70 ℃ to 100 ℃ and 1.1 to 1.3 atm,
The hot-water extract of the Chinese offspring has a total phenolic compound content of 10 to 30 mg gallic acid equivalent (GAE) / g solid; Total flavonoid content 1-20 mg quercetin equivalent (QE) / g solid; And a total saponin content of 50 to 100 mg quillajapi saponin equivalent (QSE) / g solid.
The method of claim 1,
The antimicrobial composition is selected from the group consisting of pharmaceutical compositions, food additives, feed additives, antiseptic compositions and quasi-drug compositions, antimicrobial compositions.