KR20160006877A - Method of reducing stink in epicarp of ginkgo biloba using enterococcus faecium - Google Patents

Abstract

The present invention relates to Enterococcus faecium which can reduce offensive odor in an exocarp of Ginkgo biloba, and to a method for reducing offensive odor in an exocarp of Ginkgo biloba using the Enterococcus faecium. According to the method of the present invention, butyric acid and hexanoic acid contents are reduced, wherein the butyric acid and hexanoic acid are main offensive odor ingredients in the exocarp of Ginkgo biloba, and thus offensive odor in the exocarp of Ginkgo biloba can be reduced. In addition, a fermented product using the exocarp of Ginkgo biloba can be produced, wherein the offensive odor is reduced in the fermented product.

Description

[0001] METHOD OF REDUCING STINK IN EPICARP OF GINKGO BILOBA USING ENTEROCOCCUS FAECIUM [0002]

The present invention relates to a method for reducing the aversion of bank steroids using Enterococcus faecium .

Banks ( Ginkgo biloba ) have been used as medicines for various diseases in herbal medicine, folk medicine and roots, and have long been regarded as food for longevity because of the long life of ginkgo. The bank is a deciduous tree belonging to the Ginkgo biloba. It is only 1 and 1 genus in the world. It was once distributed all over the world, but now it is growing in East Asia. Ginkgo has a flavonoid-based insecticide and germicide component, so it does not eat sickness or pests, and has a good growth characteristic even in polluted soil environment. Bank extracts consist of 24% of ginkgo flavone glycosides and 6% of terpenes (ginkgolides and bilobalides) and have been used in China for bronchial asthma and bronchitis for thousands of years. It is also known to have an excellent effect in the treatment of peripheral blood flow disorder and brain dysfunction in Western societies.

According to the data of Korea Agricultural Yearbook (2001), the total production of Korean banks in 2000 was about 1,760 M / T, which is continuously increasing. Domestic banks use about 20% of total production and 80% of them are exported to Japan at very low cost with no processing, so it is urgent to increase the value added of banks.

The bank is composed of a yellow fruit of ginkgo, a multifocal suture, a white hard muscle, an inner coat of a brown coat inside the muscle, and a turquoise bank egg (endosperm and embryo) in the endosperm. Bank eggs are nutritious foods containing 55% moisture, 38% carbohydrates, 1.7% fat, 5% protein, carotene, vitamin C, calcium, potassium, phosphorus, iron and other lecithin, asparaginic acid and ergosterol.

Butyric acid (CH ₃ CH ₂ CH ₂ COOH), which is a saturated fatty acid of carbon number 4, exists in the bank steroids as a major anomaly. In addition, Alkyl Phenolic acid and Alkyl Phenols, which are allergenic substances in leaves and scalp, Generally containing less than 10 ppm, see Ginkgo Biloba, Edited by Teris A. van Beek, Harwood Academic Publishers, in Introduction p3, 2005].

In addition, the volume ratio of the fruit (bank eggs) and the sphincter is about 1: 4. The ratio of the sphincter to be discarded is large in the process of separating the flesh of the sphincter to obtain the bank eggs. In addition, since the bank fruit contains a large amount of overpotential, a large amount of overproduced water is produced during the banking and the yield is low.

Up to now, excessive fluid and surgical blood contained in the sphincter have been discarded despite being contained in large quantities of efficacious substances, which is not only a waste of resources but also pollutes the water and the environment of the surplus blood and the blood contained in the discarded ginkgo fruit, There is a problem that the specific odor of the scar can spread and pollute the atmospheric environment.

On the other hand, fermentation using useful microorganisms such as lactic acid bacteria and yeast is known to have effects such as elimination of odor, purification of water and prevention of oxidation. In this connection, Korean Patent Laid-Open Publication No. 10-2008-0100535 has proposed a method for producing a malodor removing agent using organic materials and microorganisms. However, there is no new technology specialized in bank steroids, which can effectively remove odor-causing substances from bank steroids and utilize them as food materials.

Accordingly, it is an object of the present invention to provide a strain capable of reducing the aversion of the bank's outer membrane and a method of reducing the aversion of the bank's outer membrane using the same.

The present inventors have conducted intensive studies to achieve the above object. As a result, the present inventors have found that when fermented by inoculating Enterococcus faecium into a bank suture, the content of butyric acid and hexanoic acid, , And as a result, it is possible to reduce the abstinence of the bank sphincter, thereby completing the present invention.

According to the method of the present invention, it is possible to reduce the content of butyric acid and hexanoic acid, which are major aversive ingredients of the bank's sphincter, as a result, to reduce the aversion of the bank's sphincter, Can be manufactured. As a result, it is expected to increase the value added of banks by greatly increasing industrial utilization of bank sutures.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a graph showing the results of MTT analysis of fermented bank sacks according to an embodiment of the present invention. FIG.
FIG. 2 is a graph showing the NO production results of fermented bank sacks according to an embodiment of the present invention.

Hereinafter, the present invention will be described in detail. However, it should be understood that the present invention may be embodied in many other specific forms without departing from the spirit or essential characteristics thereof.

In one aspect, the present invention relates to a method for reducing anxiety in a bank suture including inoculating the bank suture with an Enterococcus faecium inoculation.

The present invention relates to a method for reducing the aversion of bank steroids, comprising the step of inoculating Enterococcus faecium into the bank suture to reduce the content of butyric acid and / or hexanoic acid in the bank suture, . The butyric acid and hexanoic acid are major components of the aphrodisiac occurring in the bank steroids, and especially in the case of butyric acid, the minimum detectable concentration is 0.000019 ppm, which is perceivable by humans, and because of the unpleasant rotten smell, It is essential to reduce the content of butyric acid in the bank's sphincter to reduce irritation. When fermentation is carried out by inoculating Enterococcus faecium according to the present invention into the bank's skins, the content of such butyric acid and / or hexanoic acid can be reduced.

In addition, the present invention provides a method for reducing the aversion of bank steroids by inoculating Enterococcus faecium into the bank sphincter to increase the content of volatile organic acids in the bank steroid. This method uses the principle of the masking method. The masking method means a method of making the aesthetic sensation less noticeable by making it impossible to perceive the anomaly component sensitively by using a strong directional substance other than the aversion ingredient. When fermenting Enterococcus faecium according to the present invention, the content of volatile organic acids such as acetic acid, propionic acid, butyric acid and formic acid having a boiling point of 150 ° C or less in the bank sterilization is increased, Or the odor generated from hexanoic acid can be reduced or alleviated. In particular, when the enterococcus faecium according to the present invention is fermented by inoculating the bank's scrotal, the content of acetic acid and / or formic acid in the volatile organic acid may be increased.

As yet another method for reducing the aversion of the bank's suture, the present invention provides a method of admixing one or more additives to the bank's suture prior to inoculating the Enterococcus faecium. It is preferable to use at least one additive selected from the group consisting of sodium bicarbonate, potassium bicarbonate and calcium bicarbonate. When such an additive is mixed with a bank suture, the effect of reducing the content of butyric acid in the bank suture can be further obtained. Specifically, 0.03 to 0.5% by weight calcium bicarbonate, 0.05 to 1.0% by weight potassium bicarbonate, or 0.05 to 1.0% by weight sodium bicarbonate, based on the total weight of the bank sterile fermentation product, But are not limited thereto. In order to maximize the effect of reducing the butyric acid content, it is preferred to add 0.03% to 0.5% by weight calcium bicarbonate to the bank suture, based on the total weight of the bank sperm fermentation product, prior to inoculating the Enterococcus faecium, % To 0.5% by weight calcium bicarbonate.

Particularly, the present invention relates to a method for reducing an aversion of bank steroids using Enterococcus faecium KFCC11584P as the Enterococcus faecium.

In another aspect, the present invention relates to a method for preparing a bank sphincter fermentation product comprising fermenting the bank sphincter by inoculating the enterococcus faecium, and a fermentation product of the bank sphincter prepared according to such method. Particularly, the present invention relates to a method for producing a fermentation product of bank Surgery using Enterococcus faecium KFCC11584P as the above Enterococcus faecium, and a fermentation product of a bank sphincter prepared according to such method.

In another aspect, the present invention relates to a pharmaceutical composition for preventing or treating a cancer or an immune-related disease, which comprises a fermented product of a bank sphincter prepared according to the above-described method for producing a fermented product of a bank Surgery as an effective ingredient.

The pharmaceutical composition according to the present invention may be formulated into oral compositions such as powders, granules, tablets, capsules, suspensions, emulsions, syrups and aerosols, external preparations, suppository syringes, pre-filled syringes ) Solution form, or a lyophilized form. However, the present invention is not limited thereto.

In the case of formulation, it may be prepared using diluents or excipients such as fillers, extenders, binders, humectants, disintegrants, surfactants and the like which are usually used. Solid formulations for oral administration include tablets, pills, powders, granules, capsules and the like, which may contain at least one or more excipients such as starch, calcium carbonate, sucrose, lactose, gelatin, . In addition to simple excipients, lubricants such as magnesium stearate and talc may also be used.

Liquid preparations for oral administration include suspensions, solutions, emulsions, and syrups. Various excipients such as wetting agents, sweetening agents, fragrances, preservatives, etc. may be included in addition to water and liquid paraffin, which are simple diluents commonly used. have. Formulations for parenteral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, freeze-dried preparations, suppositories, and the like.

The preferred dosage of the pharmaceutical composition of the present invention can be appropriately selected depending on the condition of the patient, the body weight, the degree of symptoms, the type of disease, the drug form, the administration route, It is preferable for the pharmaceutical composition of the present invention that the bank steril fermented product, which is an effective ingredient, is administered 0.2 to 200 mg / kg per day for optimal efficacy. The administration may be carried out once a day or divided into several doses, but is not limited thereto.

The pharmaceutical composition of the present invention can be used for prevention or treatment of cancer or immune-related diseases. The cancer may be, but is not limited to, breast cancer, melanoma, gastric cancer, liver cancer or lung cancer. In addition, the above-mentioned immune related diseases may be selected from the group consisting of systemic lupus erythematosus, rheumatoid arthritis, chronic arthritis, spondyloarthropathies, systemic sclerosis (scleroderma), idiopathic inflammatory muscle disorders (dermatomyositis, polymyositis), Sjogren's syndrome, , Autoimmune thrombocytopenia (autoimmune thrombocytopenia, paroxysmal nocturnal hemoglobinuria), autoimmune thrombocytopenia (idiopathic thrombocytopenic purpura, immune mediated thrombocytopenia), thyroiditis (Graves' disease, Hashimoto's thyroiditis, Dehydration of central and peripheral nervous system such as atrophic thyroiditis, diabetes mellitus, immune-mediated kidney disease (glomerulonephritis, interstitial nephritis), multiple sclerosis, idiopathic dehydrating polyneuritis or Gilrange-Barre syndrome and chronic inflammatory dehydration multinodular neuritis Diseases, infectious hepatitis (hepatitis A, B, C, D, hepatitis and other non-hepatitis) (Inflammatory bowel disease) such as inflammatory bowel disease (ulcerative colitis: Crohn's disease), gluten-sensitive intestinal disease, and whey, such as hepatic biliary diseases such as immune chronic active hepatitis, primary biliary cirrhosis, granulomatous hepatitis and sclerosing cholangitis. An allergic disease such as psoriasis, asthma, allergic rhinitis, atopic dermatitis, food hypersensitivity and dermatitis, eosinophilia, dyspepsia, Pneumonia, idiopathic pulmonary fibrosis, and hypersensitivity pneumonitis.

In another aspect, the present invention relates to a health functional food for enhancing immunity, which contains, as an active ingredient, a fermented product of a bank sphincter prepared according to the above-described method for producing a fermented product of a bank sphincter.

The health functional food of the present invention can be manufactured in various forms such as, but not limited to, various drinks, gum, tea, confectionery, vitamin complex, health supplement and the like.

The preferred intake amount of the health functional food of the present invention varies depending on the condition of the recipient, the body weight, the degree of symptoms, the type of food, and the period of consumption, and can be appropriately selected. It is preferable for the health functional food of the present invention to consume 0.2 to 200 mg / kg of the bank steril fermented product as an effective ingredient for one day for optimum effect.

In another aspect, the present invention relates to a novel lactic acid bacterium strain Enterococcus faecium KFCC11584P.

Hereinafter, the present invention will be described in more detail with reference to examples. However, the following examples are intended only to illustrate the present invention and are not intended to limit the scope of the present invention.

[ Example ]

1. Preparation of sample

The bank used in this example was harvested from the Doosung Bank Farming Association in late October and early November 2011 and used yellow ripe fruit.

2. Selection of strain

Among strains of food, lactic acid bacteria and yeast were selected as strains capable of reducing the aversion of bank steroids. In this example, experiments were conducted on 60 strains of lactic acid bacteria and 40 strains of yeast. 60 kinds of lactic acid bacteria used and 40 kinds of yeast used in the experiment are shown in the following Tables 1 and 2.

60 species of lactic acid bacteria No. Strain Strain number Separation source One Bifidobacterium longum KFRI 744 - 2 Bifidobacterium breve KFRI 743 Intestine of infant 3 Enterococcus faecalis KCTC 3195 - 4 Enterococcus faecalis there . liquefaciens KFRI 685 - 5 Enterococcus faecium KFRI 826 Kimchi 6 Enterococcus faecium KFRI 823 Kimchi 7 Enterococcus faecium KFRI 822 - 8 Enterococcus feacalis there . liquefaciens KFRI 685 - 9 Enterococcus feacium KFRI 1182 Fermaenting apple juice 10 Eubacterium lymosum KFRI 753 - 11 Lactobacillus amylophilus KFRI 238 - 12 Lactobacillus brevis KCTC 3102 Fermenting Sevillano variety olives 13 Lactobacillus brevis KCTC 3498 human feces 14 Lactobacillus brevis KFRI 146 - 15 Lactobacillus brevis KFRI 812 Kimchi 16 Lactobacillus brevis KFRI 805 Kimchi 17 Lactobacillus casei KFRI 228 - 18 Lactobacillus corniformis subsp. corniformis KCTC 3505 Fermentating olives 19 Lactobacillus curvatus KFRI 654 Milk 20 Lactobacillus brevis KFRI 145 Fermented beets 21 Lactobacillus hilgardii KFRI 229 - 22 Lactobacillus honohiechii KFRI 234 - 23 Lactobacillus lactis KCTC 2181 - 24 Lactobacillus pentosus KFRI 1183 - 25 Lactobacillus pentosus KFRI 481 26 Lactobacillus plantarum KCTC3099 - 27 Lactobacillus plantarum KFRI 464 Pickled cabbage 28 Lactobacillus plantarum KFRI 144 - 29 Lactobacillus plantarum KFRI 814 Kimchi 30 Lactobacillus plantarum KFRI 813 Kimchi 31 Lactobacillus plantarum KFRI 660 Pickled cabbage 32 Lactobacillus plantarum KFRI 471 - 33 Lactobacillus plantarum KFRI 470 - 34 Lactobacillus plantarum KFRI 464 Pickled cabbage 35 Lactobacillus sakei KFRI 815 Kimchi 36 Lactococcu lactic sp . lactis KFRI 669 - 37 Lactococcu lactic sp . lactis KFRI 670 - 38 Lactococcu lactic sp . lactis 　 KFRI 671 - 39 Lactococcus plantarum KFRI 1186 Frozen peas 40 Lactococcus plantarum KCTC 1048 - 41 Leuconostoc carnosum KCTC 3252 Soil 42 Leuconostoc lacits KFRI 232 - 43 Leuconostoc mesenteroides sp . mesenteroides KCTC 3505 Fermentating olives 44 Leuconostoc carnosum KCTC 3524 - 45 Leuconostoc carnosum KCTC 3525 Vacuum-packaged meats 46 Leuconostoc lactis KCTC 3528 Milk 47 Leuconostoc mesenteroides KCTC 3530 - 48 Leuconostoc mesenteroides KCTC 3505 Fermentating olives 49 Leuconostoc mesenteroides sp . mesenteroides KFRI 145 Fermented beets 50 Leuconostoc creoris KFRI 241 - 51 Leuconostoc menseteroides KFRI 818 - 52 Leuconostoc menseteroides sp . menseteroides KFRI 819 Kimchi 53 Leuconostoc menseteroides sp . menseteroides KFRI 820 Kimchi 54 Leuconostoc menseteroides sp . menseteroides KFRI 821 Kimchi 55 Pediococcus cerevisiae KCTC 3101 - 56 Pediococcus pentosaceus KCTC 3507 Dried American beer yeast 57 Pediococcus pentosaceus KFRI 832 - 58 Pediococcus pentosaceus KFRI 833 Kimchi 59 Pediococcus pentosaceus KFRI 834 Kimchi 60 Weissella confusa KFRI 1184 -

40 kinds of yeast No. Strain Strain number Separation source One Saccharomycopsis fibuligera KFRI 00106 Ragi 2 Candida edax KFRI 00113 Fermaented swine manure 3 Candidaintermedia KFRI 00115 Feces 4 Kluyveromyces marxianus there . KFRI 00137 Whey fermentation 5 Rhodotorula glutinis KFRI 00172 - 6 Candida tropicalis KFRI 00178 - 7 Schizosaccharomyces pumice KFRI 00259 Rum 8 Saccharomyces cerevisiae KFRI 00265 - 9 Saccharomyces heterogenicus KFRI 00270 - 10 Rhodosporidium toruloides KFRI 00316 - 11 Saccharomycopsis fibuligera KFRI 00441 Ragi 12 Crytococcus albidus KFRI 00457 soil 13 Torulaspora fermentation KFRI 00550 Sherry 14 Saccharomycodes ludwigii KFRI 00551 - 15 Rhodosporidium toruloides KFRI 00552 - 16 Filobasidium capsuligenum KFRI 00555 - 17 Candida rugosa KFRI 00557 - 18 Torulaspora hansenii KFRI 00560 - 19 Hansenula sp. KFRI 00588 - 20 Candida sp. KFRI 00589 - 21 Citeromyces sp. KFRI 00590 - 22 Cantharellus cibarius KFRI 00597 - 23 Pleurotus sajor - caju KFRI 00600 - 24 Hericium erinaceus KFRI 00606 - 25 Saccharomyces cerevisiae KFRI 00631 Wine 26 Schizosaccharomyces pumice KFRI 00635 - 27 Rhodotorula gracillis KFRI 00636 - 28 Sporobolomyces holsaticus KFRI 00637 - 29 Sporobolomyces holsaticus KFRI 00638 - 30 Torulopsis candida KFRI 00639 - 31 Saccharomyces cerevisiae KFRI 00795 - 32 Candida lipolytica KFRI 00909 - 33 Candida pseudotropicalis KFRI 00910 Yogurt 34 Saccharomyces cerevisiae KFRI 00920 - 35 Saccharomyces lactis KFRI 00922 Gassy cheese 36 Saccharomyces rosei KFRI 00926 - 37 Saccharomyces cerevisiae KFRI 00937 - 38 Schizosaccharomyces pumice KFRI 00938 Rum 39 Saccharomyces cerevisiae KFRI 01013 Wine 40 Saccharomyces cerevisiae KFRI 01019 -

2-1. Disgust  analysis

The finely pulverized bank suture was added to TSB to prepare a 10% bank sterilized liquid medium. After 100% of 1% strain was inoculated into the liquid medium, the yeast was incubated at 25 占 폚 and the lactic acid bacteria at 48 占Lt; / RTI >

The thus-obtained fermented product was centrifuged at 4,000 rpm at 4 ° C for 10 minutes, and the recovered supernatant was diluted with methanol for HPLC at a ratio of 1: 9 (w: v), followed by filtration through a 0.2 μm PVDF syringe filter (Whatman) And then used for analysis. The organic acid analysis was performed using high performance liquid chromatography (HPLC), and the analysis conditions are shown in Table 3 below.

HPLC  Instrument Condition Instrument Waters, E2695 Column Aminex HPX-87H (300 x 7.8 mm) Elution 0.008 NH ₂ SO ₄ Flow rate 0.6 mL / min Detector UV 210 nm 오빈 온도 50 ℃ Injection volume 20 μl

Changes in the content of butyric acid, hexanoic acid, acetic acid and formic acid by the strains were confirmed and are shown in Table 4 below.

No. Strain Strain number content butyric acid
(ppm) hexanoic acid
(ppm) acetic acid
(ppm) formic acid
(ppm) One control (10% bank sack badge) - 54,199 17,045 1,211 1,381 2 Bifidobacterium longum KFRI 744 39,432 9,651 3,221 1,311 3 Bifidobacterium breve KFRI 743 50,218 15426 1,242 1,320 4 Enterococcus faecalis KCTC 3195 54,162 15,452 1,324 1,351 5 Enterococcus faecalis there . liquefaciens KFRI 685 51,357 17,065 1,240 1,384 6 Enterococcus faecium KFRI 826 52,394 16,532 1,242 1,320 7 Enterococcus faecium KFRI 823 41,469 16,523 1,652 1,302 8 Enterococcus faecium KFRI 822 48,253 12,354 1,452 1,375 9 Enterococcus feacalis there . liquefaciens KFRI 685 51,558 17,263 1,320 1,304 10 Enterococcus feacium KFRI  1182 31,362 7,292 2,248 1,329 11 Eubacterium lymosum KFRI 753 41,734 16,267 1,246 1,276 12 Lactobacillus amylophilus KFRI 238 48,437 16,826 1,652 1,324 13 Lactobacillus brevis KCTC 3102 52,203 19,562 1,685 1,210 14 Lactobacillus brevis KCTC 3498 37,472 15,362 1,642 1,342 15 Lactobacillus brevis KFRI 146 49,290 16,428 1,520 1,324 16 Lactobacillus brevis KFRI 812 55,859 12,375 1,652 1,375 17 Lactobacillus brevis KFRI 805 54,943 16,845 1,423 1,365 18 Lactobacillus casei KFRI 228 47,125 14,562 1,652 1,354 19 Lactobacillus corniformis subsp. corniformis KCTC 3505 40,768 16,542 1,667 1,324 20 Lactobacillus curvatus KFRI 654 54,732 14,521 1,542 1,395 21 Lactobacillus brevis KFRI 145 36,881 6,554 2,432 3,241 22 Lactobacillus hilgardii KFRI 229 43,228 10,324 1,272 1,327 23 Lactobacillus honohiechii KFRI 234 46,672 16,247 1,264 1,248 24 Lactobacillus lactis KCTC 2181 51,202 12,522 1,320 1,249 25 Lactobacillus pentosus KFRI 1183 38,864 16,592 1,624 1,375 26 Lactobacillus pentosus KFRI 481 42,157 11,246 1,726 1,392 27 Lactobacillus plantarum KCTC 3099 54,218 16,275 1,246 1,346 28 Lactobacillus plantarum KFRI 464 37,181 15,894 1,325 1,270 29 Lactobacillus plantarum KFRI 144 44,678 15,782 1,342 3,200 30 Lactobacillus plantarum KFRI 814 51,120 15,410 1,279 1,276 31 Lactobacillus plantarum KFRI 813 46,984 16,242 1,360 1,138 32 Lactobacillus plantarum KFRI 660 54,123 10,325 1,249 1,356 33 Lactobacillus plantarum KFRI 471 54,621 12,024 1,247 1,301 34 Lactobacillus plantarum KFRI 470 41,785 12,530 1,368 1,395 35 Lactobacillus plantarum KFRI 464 49,523 12,062 1,369 3,642 36 Lactobacillus sakei KFRI 815 32,245 7,501 2,032 2,039 37 Lactococcu lactic sp . lactis KFRI 669 56,782 12,071 1,652 1,207 38 Lactococcu lactic sp . lactis KFRI 670 43,582 12,620 1,624 1,295 39 Lactococcu lactic sp . lactis 　 KFRI 671 49,503 12,842 1,579 1,348 40 Lactococcus plantarum KFRI 1186 48,265 13,265 1,326 1,369 41 Lactococcus plantarum KCTC 1048 48,325 13,225 1,346 1,302 42 Leuconostoc carnosum KCTC 3252 39,237 16,220 1,262 1,327 43 Leuconostoc lacits KFRI 232 39,546 12,207 1,276 2,290 44 Leuconostoc mesenteroides sp . mesenteroides KCTC 3505 38,324 14,208 1,620 1,246 45 Leuconostoc carnosum KCTC 3524 42,651 10,325 1,462 1,203 46 Leuconostoc carnosum KCTC 3525 43,628 10,426 1,552 1,276 47 Leuconostoc lactis KCTC 3528 52,012 11,652 1,324 1,320 48 Leuconostoc mesenteroides KCTC 3530 46,823 10,328 1,320 1,267 49 Leuconostoc mesenteroides KCTC 3505 44,125 13,207 1,326 1,203 50 Leuconostoc mesenteroides sp . mesenteroides KFRI 145 41,032 12,039 1,342 1,380 51 Leuconostoc creoris KFRI 241 40,326 11,032 1,624 1,271 52 Leuconostoc menseteroides KFRI 818 38,230 16,121 1,607 1,235 53 Leuconostoc menseteroides sp . menseteroides KFRI 819 37,210 15,421 1,362 1,276 54 Leuconostoc menseteroides sp . menseteroides KFRI 820 41,265 11,451 1,380 1,320 55 Leuconostoc menseteroides sp . menseteroides KFRI 821 42,562 16,520 1,427 1,346 56 Pediococcus cerevisiae KCTC 3101 50,232 12,842 1,271 1,390 57 Pediococcus pentosaceus KCTC 3507 49,213 10,301 1,427 1,357 58 Pediococcus pentosaceus KFRI 832 44,032 9,984 1,268 1,304 59 Pediococcus pentosaceus KFRI 833 34,921 12,201 1,491 1,290 60 Pediococcus pentosaceus KFRI 834 46,366 11,065 1,274 1,302 61 Weissella confusa KFRI 1184 32,210 9,881 3,104 1,371 62 Saccharomycopsis fibuligera KFRI 00106 36,219 11,012 1,274 1,620 63 Candida edax KFRI 00113 42,689 10,482 1,650 1,652 64 Candidaintermedia KFRI 00115 46,865 10,113 2,111 1,384 65 Kluyveromyces marxianus there . KFRI 00137 53,265 13,024 1,241 1,271 66 Rhodotorula glutinis KFRI 00172 51,035 10,325 1,234 1,249 67 Candida tropicalis KFRI 00178 43,253 11,039 1,032 1,230 68 Schizosaccharomyces pumice KFRI 00259 47,856 12,745 1,652 1,901 69 Saccharomyces cerevisiae KFRI 00265 47,965 8,452 1,745 1,320 70 Saccharomyces heterogenicus KFRI 00270 50,368 10,128 1,248 1,384 71 Rhodosporidium toruloides KFRI 00316 46,823 14,712 1,352 1,368 72 Saccharomycopsis fibuligera KFRI 00441 39,236 15,520 1,492 1,342 73 Crytococcus albidus KFRI 00457 39,520 14,403 1,614 1,420 74 Torulaspora fermentation KFRI 00550 38,652 16,210 1,268 1,562 75 Saccharomycodes ludwigii KFRI 00551 39,626 14,149 1,241 1,421 76 Rhodosporidium toruloides KFRI 00552 34,685 15,170 1,354 1,074 77 Filobasidium capsuligenum KFRI 00555 41,652 11,032 1,657 1,203 78 Candida rugosa KFRI 00557 45,469 12,071 1,492 1,273 79 Torulaspora hansenii KFRI 00560 51,253 11,682 1,542 1,032 80 Hansenula sp. KFRI 00588 42,656 12,302 1,642 1,024 81 Candida sp. KFRI 00589 52,315 12,212 1,426 1,065 82 Citeromyces sp. KFRI 00590 42,352 11,087 1,234 1,420 83 Cantharellus cibarius KFRI 00597 49,562 10,071 1,625 1,243 84 Pleurotus sajor - caju KFRI 00600 48,522 11,802 2,034 1,652 85 Hericium erinaceus KFRI 00606 47,326 13,260 1,121 1,275 86 Saccharomyces cerevisiae KFRI 00631 46,523 8,940 1,652 1,359 87 Schizosaccharomyces pumice KFRI 00635 48,216 13,301 1,349 1,243 88 Rhodotorula gracillis KFRI 00636 47,326 12,032 1,362 1,652 89 Sporobolomyces holsaticus KFRI 00637 43,956 11,072 1,592 1,460 90 Sporobolomyces holsaticus KFRI 00638 38,962 13,017 1,397 1,246 91 Torulopsis candida KFRI 00639 41,356 11,420 1,032 1,320 92 Saccharomyces cerevisiae KFRI 00795 43,579 9,052 3,626 1,324 93 Candida lipolytica KFRI 00909 43,692 11,073 1,624 1,320 94 Candida pseudotropicalis KFRI 00910 44,326 12,469 1,846 1,652 95 Saccharomyces cerevisiae KFRI 00920 33,261 12,920 2,487 1,485 96 Saccharomyces lactis KFRI 00922 42,953 13,402 2,610 1,320 97 Saccharomyces rosei KFRI 00926 43,625 12,321 1,231 1,623 98 Saccharomyces cerevisiae KFRI 00937 43,262 12,709 3,210 1,462 99 Schizosaccharomyces pumice KFRI 00938 49,562 10,478 1,624 1,954 100 Saccharomyces cerevisiae KFRI 01013 48,623 9,548 2,340 1,452 101 Saccharomyces cerevisiae KFRI 01019 31,742 8,221 4,120 2,641

In the case of Enterococcus faecium (KFRI 1182), the content of butyric acid and hexanoic acid, which are disgusting components of bank steroids, was remarkably reduced and the content of acetic acid, a volatile organic acid, was significantly increased compared with the control group. .

2-2. Live birth  Review

For the selection of strains resistant to urushiol, 100 strains were cultured in urushiol supplemented medium. For this purpose, a solid medium was prepared by adding lacquer extract and bank spermatocyte containing urushiol to each concentration. The resistance of the strain to urushiol was determined by the growth of the strain and the size of the colonies (size of colony diameter, mm) Respectively. The results are shown in Table 5 below.

No. Strain Strain number Solid medium composition control
(TSA) lacquer
0.1% lacquer
One% Bank Surgery
One% Bank Surgery 10% Bank Surgery 50% One Bifidobacterium  longum KFRI 744 15 15 10 12 8 3 2 Bifidobacterium breve KFRI 743 30 0 0 0 0 0 3 Enterococcus faecalis KCTC 3195 8 0 0 0 0 0 4 Enterococcus faecalis there . liquefaciens KFRI 685 30 0 0 0 0 0 5 Enterococcus faecium KFRI 826 18 20 3 0 0 0 6 Enterococcus faecium KFRI 823 5 50 One 0 0 0 7 Enterococcus faecium KFRI 822 6 0 0 0 0 0 8 Enterococcus feacalis there . liquefaciens KFRI 685 18 8 0 0 0 0 9 Enterococcus feacium KFRI  1182 22 16 9 19 8 0 10 Eubacterium lymosum KFRI 753 11 0 0 0 0 0 11 Lactobacillus amylophilus KFRI 238 25 11 0 4 2 2 12 Lactobacillus brevis KCTC 3102 33 22 0 0 0 0 13 Lactobacillus brevis KCTC 3498 19 6 0 0 0 0 14 Lactobacillus brevis KFRI 146 8 6 4 One One One 15 Lactobacillus brevis KFRI 812 8 0 0 One One One 16 Lactobacillus brevis KFRI 805 7 3 0 One 2 One 17 Lactobacillus casei KFRI 228 8 6 0 One 2 One 18 Lactobacillus corniformis subsp. corniformis KCTC 3505 15 2 2 2 One One 19 Lactobacillus curvatus KFRI 654 10 0 0 0 0 0 20 Lactobacillus brevis KFRI 145 19 11 11 3 2 One 21 Lactobacillus hilgardii KFRI 229 26 26 One 0 0 0 22 Lactobacillus honohiechii KFRI 234 14 8 2 0 0 0 23 Lactobacillus lactis KCTC 2181 39 30 23 20 20 One 24 Lactobacillus pentosus KFRI 1183 14 One 0 0 0 0 25 Lactobacillus pentosus KFRI 481 10 0 0 0 0 0 26 Lactobacillus plantarum KCTC 3099 22 10 0 5 0 0 27 Lactobacillus plantarum KFRI 464 11 One 0 0 0 0 28 Lactobacillus plantarum KFRI 144 22 0 0 0 0 0 29 Lactobacillus plantarum KFRI 814 20 3 0 0 0 0 30 Lactobacillus plantarum KFRI 813 4 2 0 0 0 0 31 Lactobacillus plantarum KFRI 660 4 4 2 0 0 0 32 Lactobacillus plantarum KFRI 471 4 2 0 0 0 0 33 Lactobacillus plantarum KFRI 470 4 2 0 0 0 0 34 Lactobacillus plantarum KFRI 464 4 0 0 0 0 0 35 Lactobacillus sakei KFRI 815 15 15 12 15 8 One 36 Lactococcu lactic sp . lactis KFRI 669 4 2 0 0 0 0 37 Lactococcu lactic sp . lactis KFRI 670 5 0 0 0 0 0 38 Lactococcu lactic sp . lactis KFRI 671 23 11 0 0 0 0 39 Lactococcus plantarum KFRI 1186 23 One 0 0 0 0 40 Lactococcus plantarum KCTC 1048 4 3 0 0 0 0 41 Leuconostoc carnosum KCTC 3252 28 0 0 0 0 0 42 Leuconostoc lacits KFRI 232 5 2 0 0 0 0 43 Leuconostoc mesenteroides sp . mesenteroides KCTC 3505 5 One 0 0 0 0 44 Leuconostoc carnosum KCTC 3524 27 11 0 0 0 0 45 Leuconostoc carnosum KCTC 3525 7 3 0 0 0 0 46 Leuconostoc lactis KCTC 3528 13 3 0 3 One One 47 Leuconostoc mesenteroides KCTC 3530 8 3 One One One One 48 Leuconostoc mesenteroides KCTC 3505 7 3 0 0 0 0 49 Leuconostoc mesenteroides sp . mesenteroides KFRI 145 24 3 0 0 0 0 50 Leuconostoc creoris KFRI 241 4 3 0 One One One 51 Leuconostoc menseteroides KFRI 818 23 2 0 0 0 0 52 Leuconostoc menseteroides sp . menseteroides KFRI 819 23 8 0 0 0 0 53 Leuconostoc menseteroides sp . menseteroides KFRI 820 19 6 One 19 19 One 54 Leuconostoc menseteroides sp . menseteroides KFRI 821 4 5 0 3 0 0 55 Pediococcus cerevisiae KCTC 3101 7 6 One One One One 56 Pediococcus pentosaceus KCTC 3507 11 5 0 2 One One 57 Pediococcus pentosaceus KFRI 832 21 5 One 2 One One 58 Pediococcus pentosaceus KFRI 833 6 5 0 2 One One 59 Pediococcus pentosaceus KFRI 834 6 0 0 0 0 0 60 Weissella confusa KFRI 1184 12 10 0 8 6 3 61 Saccharomycopsis fibuligera KFRI 00106 30 0 0 0 0 0 62 Candida edax KFRI 00113 5 0 0 0 0 0 63 Candidaintermedia KFRI 00115 8 0 0 0 0 0 64 Kluyveromyces marxianus there . KFRI 00137 16 0 0 5 0 0 65 Rhodotorula glutinis KFRI 00172 5 0 0 0 0 0 66 Candida tropicalis KFRI 00178 27 0 0 0 0 0 67 Schizosaccharomyces pumice KFRI 00259 8 0 0 0 0 0 68 Saccharomyces cerevisiae KFRI 00265 9 0 0 0 0 0 69 Saccharomyces heterogenicus KFRI 00270 14 0 0 0 0 0 70 Rhodosporidium toruloides KFRI 00316 7 0 0 0 0 0 71 Saccharomycopsis fibuligera KFRI 00441 5 0 0 0 0 0 72 Crytococcus albidus KFRI 00457 6 0 0 0 0 0 73 Torulaspora fermentation KFRI 00550 5 0 0 0 0 0 74 Saccharomycodes ludwigii KFRI 00551 4 0 0 0 0 0 75 Rhodosporidium toruloides KFRI 00552 7 0 0 0 0 0 76 Filobasidium capsuligenum KFRI 00555 8 0 0 0 0 0 77 Candida rugosa KFRI 00557 3 0 0 0 0 0 78 Torulaspora hansenii KFRI 00560 16 0 0 0 0 0 79 Hansenula sp. KFRI 00588 9 0 0 0 0 0 80 Candida sp. KFRI 00589 7 0 0 0 0 0 81 Citeromyces sp. KFRI 00590 11 0 0 0 0 0 82 Cantharellus cibarius KFRI 00597 10 0 0 0 0 0 83 Pleurotus sajor - caju KFRI 00600 20 0 0 0 0 0 84 Hericium erinaceus KFRI 00606 18 0 0 0 0 0 85 Saccharomyces cerevisiae KFRI 00631 3 0 0 0 0 0 86 Schizosaccharomyces pumice KFRI 00635 3 0 0 0 0 0 87 Rhodotorula gracillis KFRI 00636 9 0 0 0 0 0 88 Sporobolomyces holsaticus KFRI 00637 3.5 0 0 2 0 0 89 Sporobolomyces holsaticus KFRI 00638 11 0 0 0 0 0 90 Torulopsis candida KFRI 00639 8 0 0 0 0 0 91 Saccharomyces cerevisiae KFRI 00795 9 0 0 0 0 0 92 Candida lipolytica KFRI 00909 21 0 0 0 0 0 93 Candida pseudotropicalis KFRI 00910 9 0 0 0 0 0 94 Saccharomyces cerevisiae KFRI 00920 2 0 0 0 0 0 95 Saccharomyces lactis KFRI 00922 11 0 0 7 2 0 96 Saccharomyces rosei KFRI 00926 13 0 0 6 3 0 97 Saccharomyces cerevisiae KFRI 00937 5 0 0 0 0 0 98 Schizosaccharomyces pumice KFRI 00938 8.5 0 0 0 0 0 99 Saccharomyces cerevisiae KFRI 01013 10 0 0 0 0 0 100 Saccharomyces cerevisiae KFRI 01019 9 0 0 5 2 0

In the case of Enterococcus faecium (KFRI 1182), survival was observed in the medium containing 0.1% of lacquer, 1% of addition medium, 1% of bank sachet, and 10% of addition. For more detailed breeding studies, Enterococcus faecium (KFRI 1182) was again inoculated into the liquid medium to measure viable counts after 24 hours of culture and 48 hours of culture. The results are shown in Tables 6 and 7 below.

After 24 hours of culture, viable cells Strain Strain name control lacquer
0.1% lacquer
One% Bank
Surgical
0.5% Bank
Surgical
5% Bank
Surgical
10% Log
(cfu / ml) Log
(cfu / ml) Log
(cfu / ml) Log
(cfu / ml) Log
(cfu / ml) Log
(cfu / ml) Enterococcus feacium KFRI 1182 9.14 9.14 8.87 9.04 8.97 8.74

After 48 hours of culture, viable cells Strain Strain name control Lacquer 0.1% lacquer
One% Bank
Surgical
0.5% Bank
Surgical
5% Bank
Surgical
10% Log
(cfu / ml) Log
(cfu / ml) Log
(cfu / ml) Log
(cfu / ml) Log
(cfu / ml) Log
(cfu / ml) Enterococcus feacium KFRI 1182 9.83 9.83 9.58 9.79 9.71 9.44

As shown in Tables 6 and 7, enterococcus faecium (KFRI 1182) showed similar growth to that of the control group in all experimental groups after 24 hours and 48 hours of culture, And a cluster size of about log 9.4 cfu / mL or more.

Through such anoxicological analysis and breeding experiment, Enterococcus faecium (KFRI 1182) resistant to uriciol, which has the effect of reducing butyric acid and hexanoic acid, was selected and named GB2. In addition, Enterococcus faecium (GB2) was deposited with the Korean Microorganism Conservation Center and granted accession number KFCC11584P.

3. Bank Surgical Fermentation product

3-1. Bank Surgical Fermented  Manufacturing and checking the content of butyric acid

The bank sperm was finely ground and four solid mediums were prepared as shown in Table 8 below. Four types of bank sperm fermentations were prepared by inoculating 1% of activated Enterococcus pheumium (GB2) in each of the thus prepared bank spermatogonial solid medium and culturing at 30 DEG C for 48 hours.

No. Medium composition Water content (%) One Bank Surgery: Water = 1: 1 (w / v) 80% 2 Bank Surgery: Water = 1: 2 (w / v) 90% 3 Bank Surgery: Rice: Water = 1: 1: 2 (w / w / v) 80% 4 Bank Surgery: Rice: Water = 1: 1: 4 (w / w / v) 90%

The thus-prepared fermented bank sphincter was centrifuged at 4,000 rpm at 4 ° C for 10 minutes, and the recovered supernatant was diluted with methanol for HPLC at a ratio of 1: 9 (w: v), followed by filtration through a 0.2 μm PVDF syringe filter ) And then used for analysis. The organic acid analysis was carried out by using high performance liquid chromatography (HPLC) to analyze the butyric acid content, and the same assay conditions as those shown in Table 3 were used. The results are shown in Table 9 below.

No. 기판 tR Quantity One control 1 28.92 36745.41 2 1-GB2 28.87 21933.14 3 control 2 28.81 15264.24 4 2-GB2 28.76 9720.88 5 control 3 28.68 8417.54 6 3-GB2 28.64 4679.43 7 control 4 28.60 4478.57 8 4-GB2 28.56 1501.05

As shown in Table 9, it can be confirmed that the content of butyric acid in the bank sterilized fermented broth cultured by inoculation with Enterococcus faecium (GB2) was decreased in all experimental groups compared to the control.

3-2. Measurement of cell survival rate

Cell viability was assayed by measuring MTT (3- (4,5-dimethylthiazol-2yl) -2,5-diphenyl tetrazolium bromide) (Sigma-Aldrich). Raw 264.7 cells were seeded at a density of 2 x 105 cells / mL in a 96-well plate, cultured for 24 hours, treated with 500 μg / mL of bank sterile fermented broth at a concentration of 50 μg / mL and then cultured for 20 hours. well media was removed and MTT reagent at a concentration of 5 mg / mL was mixed with serum-free DMEM and 200 μL / well was added to each well and cultured for 4 hours. After incubation, the supernatant was removed and DMSO was added to dissolve the cells. The formazan produced by MTT reduction was measured for absorbance at 540 nm using a microplate reader.

The results of evaluating cell viability through MTT assay are shown in Fig. All of the bank steril fermentations cultured on the substrate with Enterococcus faecium (1-GB2, 2-GB2, 3-GB2, 4-GB2) showed cell survival rates of over 80%.

3-3. NO  Confirmation of production amount

Nitric oxide production in the culture was evaluated by measuring nitrite, which is an oxide of nitrogen oxide. Raw 264.7 cells were seeded at a density of 2 x 105 cells / mL in a 96-well plate, cultured for 24 hours, and then selected in a Raw 264.7 cell to a concentration of 500 μg / mL. And then cultured for 20 hours. After incubation, 50 μL of the supernatant was transferred to a new 96-well plate, mixed with a griess sample (Sigma-Aldrich) at 1: 1, reacted on a shaker for 10 minutes, and then transferred to a microplate reader (infinite M200 Pro, The absorbance at 540 nm was measured using a spectrophotometer (manufactured by Tecan Co.). The standard curve was measured using sodium nitrite.

The results of measurement of the NO production amount of the sample relative to the LPS treatment group are shown in FIG. In particular, fermented bank sperm cultures cultured in the presence of Enterococcus faecium (1-GB2 and 2-GB2) on the substrate showed higher NO production ability than control 1 and 2.

4. Banks with additives Surgical Fermentation product

4-1. A bank of additives Surgical Fermented  Produce

Bank oegwapi 10% of calcium bicarbonate with the addition of culture medium _{(CaHCO 3) 0.03%, 0.25} %, 0.5%, potassium bicarbonate _{(KHCO 3) 0.05%, 0.5} %, 1%, sodium bicarbonate _{(NaHCO 3) 0.05%, 0.5} % , And 1%, respectively. The thus-prepared bank spermatogonial cells were inoculated with activated Enterococcus faecium (GB2) and cultured at 30 ° C. for 48 hours to prepare a bank sperm fermented product.

4-2. Content analysis of butyric acid

The bank sterilized fermented product mixed with additives was centrifuged at 4,000 rpm at 4 ° C for 10 minutes, and the recovered supernatant was diluted with methanol for HPLC at a ratio of 1: 9 (w: v), followed by filtration through a 0.2 μm PVDF syringe filter Ltd.) and then used for analysis. The organic acid analysis was carried out by using high performance liquid chromatography (HPLC) to analyze the butyric acid content, and the same assay conditions as those shown in Table 3 were used. The results are shown in Table 10 below.

No. Inoculum Furtherance additive additive
content(%) Identification of bacteria (cfu / ml) butyric acid content
(ppm) One Enterococcus feacium
(GB2) Bank
Surgical
10%
adding No additives - 9.67     31,362 2 CaHCO ₃ 0.03 9.98     12,450 3 CaHCO ₃ 0.25 10.14       8,054 4 CaHCO ₃ 0.5 10.38       7,309 5 KHCO ₃ 0.05 8.26       8,521 6 KHCO ₃ 0.5 8.38       7,016 7 KHCO ₃ One 8.53       7,361 8 NaHCO ₃ 0.05 10.54     12,790 9 NaHCO ₃ 0.5 10.57     12,689 10 NaHCO ₃ One 10.47     13,101

As shown in Table 10, it can be seen that the butyric acid content in the bank steril fermented product in all cases in which the additive was added was decreased compared with the case without addition of the additive. In particular, when 0.25% and 0.5% CaHCO ₃ were added , The butyric acid content of the fermented product decreased to the greatest extent.

Korea Microorganism Conservation Center KFCC11584P 20140609

Claims (18)

A method of reducing the aversion of a bank's suture, including fermenting the bank suture with an Enterococcus faecium inoculation. The method according to claim 1,
Wherein the enterococcus pacemaker is Enterococcus faecium KFCC11584P. The method according to claim 1,
Characterized in that the content of butyric acid, hexanoic acid, or butyric acid and hexanoic acid in the bank sphincter is reduced. The method according to claim 1,
Wherein the amount of volatile organic acid in the bank suture is increased. 5. The method of claim 4,
Wherein the volatile organic acid is acetic acid, formic acid, acetic acid and formic acid. The method according to claim 1,
Characterized in that the bank steroid is mixed with at least one additive selected from the group consisting of sodium bicarbonate, potassium bicarbonate and calcium bicarbonate prior to inoculating the Enterococcus faecium. The method according to claim 6,
Characterized in that 0.25% to 0.5% by weight calcium bicarbonate is added based on the total weight of the fermented product of the bank sphincter. A method of producing a bank sphincter fermentation comprising fermenting a bank sphincter by inoculating Enterococcus faecium . 9. The method of claim 8,
Characterized in that the Enterococcus faecium is Enterococcus faecium KFCC11584P. 9. The method of claim 8,
Characterized in that the content of butyric acid, hexanoic acid, or butyric acid and hexanoic acid in the bank sphincter is reduced. 9. The method of claim 8,
Wherein the content of volatile organic acid in the bank suture is increased. 12. The method of claim 11,
Wherein said volatile organic acid is acetic acid, formic acid, or acetic acid and formic acid. 9. The method of claim 8,
Characterized in that before the Enterococcus faecium is inoculated, the bank sphincter is mixed with at least one additive selected from the group consisting of sodium bicarbonate, potassium bicarbonate and calcium bicarbonate. 14. The method of claim 13,
Characterized in that 0.25% to 0.5% by weight calcium bicarbonate is added, based on the total weight of the bank sphincter fermented product. 14. A fermentation product of a bank sphincter prepared according to the method of any one of claims 8 to 14. A pharmaceutical composition for preventing or treating cancer or an immune-related disease, which comprises the fermented product of the bank sphincter of claim 15 as an active ingredient. A health functional food for enhancing immunity comprising the fermented product of the bank sphincter of claim 15 as an active ingredient. Enterococcus faecium ) KFCC11584P.

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