KR101761186B1 - New lactic acid bacteria isolated from Kimchi and antibacterial composition using the same - Google Patents

Abstract

The present invention relates to a novel lactic acid bacterium isolated from kimchi and an antimicrobial composition using the same. More specifically, the present invention relates to Lactobacillus plantarum DS2 (KCTC12992BP), Leuconostomercenteroid DS3 A composition for antimicrobial use containing as an active ingredient one or two or more strains selected from the group consisting of Leuconostoc mesenteroides DS3 and KCTC13016BP and Lactobacillus sakei DS4 and KCTC13017BP or a culture thereof, Kimchi, pickled foods, feed and feed additives, and cosmetics.
Lactobacillus plantarum DS2, lucconostomercenteroid DS3 and lactobacillus saca DS4, which are novel lactic acid bacteria of the present invention, are lactic acid bacteria derived from kimchi, and are excellent antibacterial and antimicrobial agents for microorganisms and pathogenic microorganisms which are safe for human body, Activity. Therefore, when these microorganisms are cultured in the presence of microbial cells or cultures thereof, they can be used safely without worrying about serious side effects of artificial synthetic compounds which have been mainly used in the past, and can be used effectively for eliminating harmful microorganisms in daily life When they are used for foods, feeds, cosmetics and the like, products having greatly improved preservation ability can be produced only by a very simple method of adding the antimicrobial composition of the present invention. The novel lactic acid bacteria and the antimicrobial composition of the present invention are expected to be useful in various industrial fields.

Description

[0001] The present invention relates to a novel lactic acid bacterium isolated from kimchi and a composition for antibacterial use using the same.

The present invention relates to a novel lactic acid bacterium isolated from kimchi and an antimicrobial composition using the same. More specifically, the present invention relates to Lactobacillus plantarum DS2 (KCTC12992BP), Leuconostomercenteroid DS3 A composition for antimicrobial use containing as an active ingredient one or two or more strains selected from the group consisting of Leuconostoc mesenteroides DS3 and KCTC13016BP and Lactobacillus sakei DS4 and KCTC13017BP or a culture thereof, Additives, feeds, feed additives and cosmetics.

Lactic acid bacteria is one of the most beneficial microorganisms available to humans and has a long history and has a close and direct relationship with human life. Lactic acid bacteria are widely distributed not only in foods but also in natural environments such as oral cavity, digestive tract and soil of mammals, and are useful for human life to perform important roles related to food preservation and sensory characteristics.

The name lactic acid bacteria is conceptual and is not a taxonomic academic name for bacteria. Lactic acid bacteria are bacteria that produce about 50% or more of lactic acid as a metabolite for sugar that is usually consumed. Unlike E. coli, it does not produce a decay product. Therefore, bacteria that produce a large amount of lactic acid from saccharides and that have a beneficial effect such as preventing spoilage without generating substances harmful to human body in human and animal intestines are conceptually referred to as lactic acid bacteria. The usual concept of lactic acid bacteria is Gram positive or bacillus, does not form catalase negative (anaerobic fermentation), spores, and is generally mobility (Axelsson, 1998, Lactic acid bacteria: Classification and physiology, Marcel Dekker Inc., New York, pp. 1-63).

Lactic acid bacteria produce various antibacterials and organic acids such as lactic acid and acetic acid which have antibacterial activity (Barefoot, et al., 1993. Antibiosis revised; bacteriocin produced by dairy starter cultures. J. Dairy Sci. 76: 2366-2379). Lactic acid bacteria produce hydrogen peroxide in the presence of oxygen and do not have catalase activity, so hydrogen peroxide is secreted into the medium to accumulate to microbial inhibitory concentrations (Juven, et al., 1991). Antagonistic effects of lactobacilli and pediococci to control intestinal colonization by human enteropathogens in live poultry, J. Appl. Bacteriol., 70: 95-103). In addition, diacetyl, one of the non-proteinic antimicrobial substances produced by lactic acid bacteria, is one of the representative flavors of lactic acid fermented products and has a strong inhibitory effect on pathogenic microorganisms by interacting with pH (Klaenhammer, 1988. Bacteriocin of lactic acid bacteria. Biochimie. 70: 337-349).

In general, bacteriocin is a protein or protein complex material with a relatively small molecular weight that inhibits other microorganisms by the collapse of the proton motive force (the driving force of ATP synthesis) of the bacterial cell membrane. Most of them are stable to heat and stable at low pH, but most are inactivated by proteolytic enzyme treatment (Abee, 1995. Pore-forming bacteriocins of Gram-positive bacteriocin and self-protection mechanisms of producer organisms FEMS Micobiol. (1): 1-10). A representative bacteriocin-derived bacteriocin, nisin, is produced in Lactococcus lactis and has long been accepted for commercial use due to its broad antibacterial activity (Zendo, 2013, Pore-forming bacteriocins of Gram-positive bacteriocin and self Lacticin, Lactococcin, Pediocin and the like have been known as bacteriocins derived from lactic acid bacteria (see, for example, (Klaenhammer, TR, 1993, Genetics of bacteriocins produced by Lactic acid Bacteria FEMS, Microbiol. Rev. 12: 39-86).

Meanwhile, kimchi is a food that can be manufactured by universal methods of origin, without any special knowledge. It is more representative of Korea's traditional fermented foods developed from the Three Kingdoms period. , Garlic, and other seasonings. It is one of the representative fermented foods in Korea, which is fermented by the microorganisms present around the living environment and aged. Recently, many studies on the usefulness of kimchi have been published. For example, physiological activities such as immunological activity and anticancer effect have been recognized, and they have been recognized as essential food for health maintenance besides nutrition supply. Especially, it is reported that the content of lactic acid bacteria is higher than that of fermented milk products because fresh vegetable is immersed without heat treatment, so that all the nutrients of vegetables are maintained and can provide a good habitat environment for lactic acid bacteria.

Lactic acid bacteria involved in kimchi fermentation include lactic acid bacteria such as Lactobacillus sp., Leuconostoc sp., Pediococcus sp., Streptococcus sp., And more than 200 kinds of yeast such as Lactobacillus sp. (Lee, et al., 1998, Research Report: Control of kimchi fermentation by regulating microbial succession. Korea Food Research Institute, E 1421-0898).

Studies on the above lactic acid bacteria have mainly been conducted on dairy products, and systematic and scientific researches on lactic acid bacteria that are involved in the fermentation process of kimchi over a long period of time have been conducted recently.

The present inventors have found new microorganisms having safety and useful properties, particularly antimicrobial activity against harmful microorganisms in the human body, and applying them to foods, feeds, cosmetics and the like to provide products having excellent effects. As a result, it was possible to find three new lactic acid bacteria having excellent antimicrobial activity against microorganisms and pathogenic microorganisms involved in food corruption. The inventors of the present invention have confirmed that preserving ability can be greatly improved when a food is manufactured using lactic acid bacteria, and the present invention has been completed.

Axelsson, 1998, Lactic acid bacteria: Classification and physiology. Marcel Dekker Inc., New York. PP 1-63. Barefoot, et al. 1993. Antibiosis revisited; bacteriocin produced by dairy starter cultures. J. Dairy Sci. 76: 2366-2379. Juven et al., 1991. Antagonistic effects of lactobacilli and pediococci to control intestinal colonization by human enteropathogens in live poultry. J. Appl. Bacteriol., 70: 95-103. Klaenhammer, 1988. Bacteriocin of lactic acid bacteria. Biochimie. 70: 337-349. Abee, 1995. Pore-forming bacteriocins of Gram-positive bacteriocin and self-protection mechanisms of producer organisms. FEMS Micobiol. Lett. 129 (1): 1-10. Zendo, 2013, Pore-forming bacteriocins of Gram-positive bacteriocin and self-protection mechanisms of producer organisms. Biosci. Biotech. Bioch. 77 (5): 893-899. Klaenhammer, T. R., 1993, Genetics of bacteriocins produced by Lactic acid Bacteria FEMS, Microbiol. Rev. 12: 39-86. Lee, et al., 1998, Research Report: Control of kimchi fermentation by regulating microbial succession. Korea Food Research Institute, E 1421-0898.

Therefore, a main object of the present invention is to provide a novel lactic acid bacteria having excellent antibacterial activity.

It is another object of the present invention to provide a composition for antimicrobial use that utilizes the antimicrobial activity of the novel lactic acid bacterium.

It is still another object of the present invention to provide foods, food additives, feeds, feed additives and cosmetics having improved preservability using the antimicrobial composition.

According to one aspect of the present invention, the present invention provides a method for producing Lactobacillus plantarum DS2 (KCTC12992BP), Leuconostoc mesenteroides DS3 (KCTC13016BP) and Lactobacillus sakei DS4, KCTC13017BP ), Or a culture thereof, as an active ingredient.

According to another aspect of the present invention, there is provided a food product prepared by adding the antimicrobial composition.

In the food of the present invention, it is preferable that the food is a meat-processed food or a kimchi.

According to still another aspect of the present invention, there is provided a food additive prepared by adding the antimicrobial composition.

According to still another aspect of the present invention, there is provided a feed prepared by adding the antimicrobial composition.

According to another aspect of the present invention, there is provided a feed additive prepared by adding the antimicrobial composition.

According to still another aspect of the present invention, there is provided a cosmetic product prepared by adding the above antimicrobial composition.

According to another aspect of the present invention, there is provided a lactobacillus plantarum DS2 ( Lactobacillus plantarum DS2, KCTC12992BP) which shows excellent antibacterial activity as a novel Kimchi lactic acid bacteria isolated from kimchi.

According to another aspect of the present invention, the present invention provides Leuconostoc mesenteroides DS3 (KCTC13016BP), which exhibits excellent antibacterial activity as a novel Kimchi lactic acid bacteria isolated from kimchi.

According to another aspect of the present invention, the present invention provides Lactobacillus sakei DS4 (KCTC13017BP), which exhibits excellent antibacterial activity as a novel Kimchi lactic acid bacteria isolated from kimchi.

Lactobacillus plantarum DS2, lucconostomercenteroid DS3 and lactobacillus saca DS4, which are novel lactic acid bacteria of the present invention, are lactic acid bacteria derived from kimchi, and are excellent antibacterial and antimicrobial agents for microorganisms and pathogenic microorganisms which are safe for human body, Activity. Therefore, when these microorganisms are cultured in the presence of microbial cells or cultures thereof, they can be used safely without worrying about serious side effects of artificial synthetic compounds which have been mainly used in the past, and can be used effectively for eliminating harmful microorganisms in daily life When they are used for foods, food additives, feeds, feed additives, cosmetics, etc., it is possible to produce a product having greatly improved preservation power by simply adding the antimicrobial composition of the present invention. The novel lactic acid bacteria and the antimicrobial composition of the present invention are expected to be useful in various industrial fields.

1 is a phylogenetic relationship analysis based on the 16S rDNA sequence of Lactobacillus plantarum DS2 ( Lactobacillus plantarum DS2, KCTC12992BP) which is a novel lactic acid bacterium of the present invention.
FIG. 2 is a phylogenetic relationship analysis based on the 16S rDNA sequence of Leuconostoc mesenteroides DS3 and KCTC13016BP, which is a novel lactic acid bacterium of the present invention.
FIG. 3 is a phylogenetic relationship analysis based on the 16S rDNA sequence of Lactobacillus sakei DS4 ( Lactobacillus sakei DS4, KCTC13017BP), which is a novel lactic acid bacterium of the present invention.

Lactobacillus plantarum DS2 of the present invention, Leuconostomercenteroid DS3 and Lactobacillus case DS4 are novel lactic acid bacteria isolated from kimchi.

Lactobacillus plantarum DS2 is a Gram-positive bacterium that forms a white milk-colored round colony with negative hemolysis results. And has a 16S rDNA sequence as in SEQ ID NO: 1. 1, and can be classified as Lactobacillus plantarum. Exhibit strong antimicrobial activity against Bacillus cereus ATCC 21768, Listeria monocytogenes ATCC 15313, Lactococcus lactis and Leuconostoc mesenteroides strains, and Staphylococcus aureus ( Escherichia coli ATCC 25922) and exhibits a slightly lower level of antimicrobial activity in Staphylococcus aureus ATCC 6538 and Weissella viridescens strains. This strain has been deposited with KCTC12992BP under the accession number KCTC12992BP of the Korea Biotechnology Research Institute.

Leuconostomercenteroid DS3 is a Gram-positive staphylococcus and forms a transparent milk-colored round colony, which is negative as a hemolysis test. And has the same 16S rDNA sequence as SEQ ID NO: 2. 2, and can be classified as licorostomercenteroid. Listeria monocytogenes and Lactococcus lactis, and exhibits a slightly lower antimicrobial activity than Bacillus cereus, Escherichia coli, Leuconostomercenteroid, and Bicerella viridis strains, and exhibits antimicrobial activity against Staphylococcus aureus Antibiotic activity can also be shown in the mouse. This strain is deposited with KCTC13016BP under the accession number KCTC13016BP of the Korea Biotechnology Research Institute.

Lactobacillus saccharum DS4 is a Gram-positive staphylococcus and forms a transparent milk-colored round colony, which is negative for hemolysis. And has the same 16S rDNA sequence as SEQ ID NO: 3. 3, and can be classified as Lactobacillus sake. Bacillus cereus, Listeria monocytogenes, Lactococcus lactis, Escherichia coli, Leuconostomercenteroid, and Bacella viridis strain, and may exhibit antibacterial activity against Staphylococcus aureus. This strain has been deposited with the Korea Research Institute of Bioscience and Biotechnology Center under accession number KCTC13017BP.

The antimicrobial composition of the present invention may comprise one or two or more strains selected from the three strains or a culture thereof. For better antimicrobial activity, it is preferable to include two or more strains selected from the above three strains or a culture thereof, more preferably all three strains or cultures thereof.

The culture can be obtained through culturing the strain. For example, the strain may be inoculated in a medium such as MRS and cultured at 25 to 35 ° C for 20 to 40 hours. In addition, the culture may be a culture containing cells or cells in which the cells have been removed, and may be in a liquid state or a solid state. The culture in which the microbial cells have been removed can be obtained by inoculating and culturing the above-mentioned strain in an MRS liquid medium, then centrifuging the culture broth to recover the supernatant and filtering, and the solid culture can be obtained by concentrating and spraying Followed by drying.

The antimicrobial composition of the present invention may be the above-mentioned strain or the culture thereof itself, and may further contain commonly used additives such as excipients and diluents required for formulation, and may be added to the strain of the present invention or a culture thereof It may further contain an antibacterial substance.

The content of the strain of the present invention or the culture thereof contained in the antimicrobial composition may be appropriately selected in consideration of the intended use and the method of use, and may be, for example, 1 to 100% by weight, 10 to 100% by weight, more preferably 50 to 100% by weight.

The present invention provides a food, a food additive, a feed, a feed additive, and a cosmetic prepared by adding the antibacterial composition as described above.

The above food products include meat products such as meat products, fish meat products, tofu, mushrooms, porridge, noodles such as noodles and noodles, seasonings such as soy sauce, miso, hot pepper paste, mixed soup, sauces, confectionery, milk products such as fermented milk and cheese, Fruit drinks, vegetables, soybean milk, fermented beverages, etc., and the antibacterial composition may be added during the process of manufacturing the foods. The amount of the antimicrobial composition to be added may be appropriately selected in consideration of the kind, form, purpose, manufacturing method and the like of the food. For example, in the case of a meat product, 0.0001 to 10% May be 0.001 to 5% by weight, more preferably 0.01 to 1% by weight, and in case of kimchi, 0.0001 to 10% by weight, preferably 0.001 to 5% by weight, more preferably 0.01 To 1% by weight.

The feed can be prepared in the form of a fermented feed, a compounded feed, a pellet form, a silage reed, or the like. The fermented feed may be prepared by fermenting an organic material by adding the antimicrobial composition, various other microorganisms or enzymes, and the compound feed may be prepared by mixing various kinds of general feed and the antimicrobial composition. The pellet-shaped feed can be prepared by formulating the fermented feed or the compound feed into a pelletizer, and the silage can be prepared by adding the antimicrobial composition to the fermented feed and fermenting. At this time, it is preferable to use the antimicrobial composition in which the strain of the present invention is contained in the form of living cells. In this case, the amount of the antimicrobial composition to be added may be appropriately selected in consideration of the kind, form, purpose, manufacturing method, etc. of the feed, and the same amount as that in the food may be used.

The cosmetics may be manufactured in a form selected from the group consisting of lotion, nutritional lotion, nutritional cream, massage cream, pack, and nutritional essence, and may be prepared by adding the antimicrobial composition during the process of manufacturing the cosmetics . The amount of the antimicrobial composition to be added may be appropriately selected in consideration of the type, form, purpose, manufacturing method, and the like of the cosmetic, and the same amount as that in the food may be used.

The food additive and the feed additive of the present invention may be the above-described antimicrobial composition itself, and may further include commonly used additives such as excipients and diluents required for formulation. The amount of the antimicrobial composition added may be, for example, 1 to 100% by weight, preferably 10 to 100% by weight, more preferably 50 to 100% by weight, based on the weight of the food additive or the feed additive.

Hereinafter, the present invention will be described in more detail with reference to Examples. These embodiments are only for illustrating the present invention, and thus the scope of the present invention is not construed as being limited by these embodiments.

Experimental Example 1. Kimchi Lactic Acid Bacteria Isolation

After collecting 30 kinds of kimchi immersed in the home, 10 g of each kimchi was sampled, mixed with sterilized saline (90 ml), and aseptically filtered to collect the kimchi filtrate. The kimchi filtrate was diluted stepwise with sterile saline, plated on a BCP selective medium (Bromcresol purple lactose agar) with a steak planting method, and cultured at 30 ° C for 24 hours. Among the cultivated strains, 78 strains were selected which were changed into yellow due to acid production.

Experimental Example 2. Measurement of antimicrobial activity against a harmful microorganism

The antibacterial activity was evaluated by culturing 78 kinds of Kimchi lactic acid bacteria selected and separated in Experimental Example 1 above. Indicative strains for the evaluation of antimicrobial activity were Lactococcus lactis , Leuconostoc mesenteroides , and Weissellae , which were directly isolated from the slime from the meat products as shown in Table 1, a viridescens) and general pathogens of Bacillus cereus (Bacillus cereus ATCC21768), Staphylococcus aureus (Staphylococcus aureus ATCC6538), L. monocytogenes Genesis (Listeria monocytogenes ATCC15313), and E. coli (Escherichia coli ATCC25922) was used.

The antimicrobial activity was confirmed by a paper disc assay. 78 kinds of lactic acid bacteria selected in Experimental Example 1 were inoculated into MRS liquid medium (de Man, Rogosa and Sharpe broth medium) and cultured at 30 ° C for 24 hours with shaking. The cultured medium was centrifuged to remove the supernatant, and the residual strain was completely removed with a 0.2 mu m membrane filter.

The inactivated culture filtrate was concentrated 10 times with a rotary vacuum concentrator to prepare a culture concentrate, which was injected into a sterilized paper disc and dried.

A soft cloth medium containing indicator bacteria was overlaid and a paper disc in which the above concentrate was injected was placed on the medium. The indicator bacteria were cultured for 16 hours under appropriate culture conditions to measure the diameter of transparent rings formed around the paper disc, that is, the inhibitory rings.

Lactic acid bacteria that produced the largest inhibitory ring were selected for each indicator bacteria among 78 kinds of lactic acid bacteria. Finally, three kinds of lactic acid bacteria were selected and designated as Kimchi lactic acid bacteria DS2, DS3, and DS4, respectively.

The characteristics of the indicator bacteria used in Experimental Example 2 are shown in Table 1.

The antimicrobial activities of the lactic acid bacteria DS2, DS3 and DS4 isolated from kimchi are shown in Table 2.

The concentrated culture filtrate of Kimchi-isolated lactic acid bacterium DS2 showed strong antimicrobial activity against Bacillus cereus, Listeria monocytogenesis, Lactococcus lactis, and Leukonostomycesenteroid strains, and Staphylococcus aureus and Baicella viridis strain Showed moderate antimicrobial activity and showed weak antimicrobial activity against E. coli.

Kimchi isolate Lactobacillus DS3 concentrated culture filtrate showed strong antimicrobial activity against Listeria monocytogenes and Lactococcus lactis and showed moderate antimicrobial activity against Bacillus cereus, Escherichia coli, Leuconostomercenteroid, and Bacela viridisense And exhibited weak antimicrobial activity against Staphylococcus aureus.

Kimchi isolate Lactobacillus DS4 concentrated culture filtrate showed moderate antimicrobial activity against Bacillus cereus, Listeria monocytogenes, Lactococcus lactis, Escherichia coli, Leuconostomercenteroid, and Bacella viridisensis, and Staphylococcus aureus And exhibited weak antimicrobial activity.

Indicative strains for antimicrobial activity experiments Classification Indication strain Culture medium Characteristic Corruption
microbe Lactococcus lactis MRS Broth Slime production, gas production, discoloration caused by meat products Leuconostoc mesenteroides MRS Broth Weissella viridescens MRS Broth

Pathogenicity
microbe Bacillus cereus ATCC21768 Nutrient Broth Diarrhea and vomiting caused by food poisoning Staphylococcus aureus ATCC6538 Trypticase Soy Broth Toxin-induced food poisoning Escherichia coli ATCC 25922 Trypticase Soy Broth Food poisoning and hemorrhagic enteritis Listeria monocytogenes ATCC15313 Brain Heart Infusion Broth Hemolytic food poisoning

Results of Antimicrobial Activity of Kimchi Lactic Acid Bacteria DS2, DS3 and DS4 Indication strain Kimchi lactic acid bacteria DS2 Kimchi lactic acid bacteria DS3 Kimchi lactic acid bacteria DS4 Bacillus cereus ATCC 21768 22 mm 16 mm 16 mm Escherichia coli ATCC 25922 10 mm 13 mm 12 mm Staphylococcus aureus ATCC6538 13 mm 10 mm 11 mm Listeria monocytogenes ATCC15313 23 mm 20 mm 17 mm Lactococcus lactis 23 mm 18 mm 16 mm Leuconostoc mesenteroides 20 mm 14 mm 14 mm Weissella viridescens 17 mm 14 mm 13 mm Paper disc size: 8 mm

Experimental Example 3. Identification of lactic acid bacteria

The morphological, physiological and biochemical characteristics of Kimchi lactic acid bacteria DS2, DS3 and DS4 selected in Experimental Example 2 are shown in Table 3.

The 16S rDNA sequence analysis of the isolated lactic acid bacteria showed 99% similarity with Lactobacillus plantarum , Leuconostoc mesenteroides and Lactobacillus sakei , and its phylogenetic classification The relationships are shown in Figs.

Each 16S rDNA sequence was deposited in GenBank, and Lactobacillus plantarum DS2 was assigned KX470811, Leuconostomercenteroid DS3 was named KX470810, and Lactobacillus case DS4 was assigned the number KX470812.

Morphological, physiological and biochemical characteristics of Kimchi lactic acid bacteria DS2, DS3 and DS4 characteristic Lactobacillus plantarum DS2 Leuconostoc mesenteroides DS3 Lactobacillus sakei DS4 Source Kimchi Kimchi Kimchi Morphology (Shape) Bacillus Sweet bacillus Bacillus Gram-stain + + + Catalase test
(catalase test) - - - Hemolysis - - - Colony color and transparency White milk color Transparent milk color White milk color

Example 1 Lactobacillus planta DS2 ( Lactobacillus plantarum  DS2) Culture Powder Production

The Lactobacillus planta DS2 (KCTC12992BP) strain was cultured in an MRS liquid medium at 30 DEG C for 30 hours, centrifuged, and the supernatant was taken and the remaining strain was completely removed by membrane filtration (0.2 mu m membrane filter). 950 g of the Lactobacillus plantarum DS2 culture filtrate and 50 g of maltodextrin were mixed and concentrated and pulverized using a spray dryer to obtain about 50 g of a pale brown dried powder.

Example 2. Reukonovosthamceteroid DS3 ( Leuconostoc mesenteroides  DS3) Production of culture powder

Cultured powder was prepared in the same manner as in Example 1 above using a strain of Ryukono Stokmeensenoid DS3.

Example 3. Lactobacillus casei DS4 ( Lactobacillus sakei  DS4) Production of culture powder

Lactobacillus saca DS4 strain was used to prepare a culture powder in the same manner as in Example 1 above.

Experimental Example 4. Lactobacillus plantarum DS2, Leuconostomercenteroid DS3, Lactobacillus casei DS4 The preservation ability of sausage added with the culture powder

The sausages (Examples 4 to 10) and the sausages (Comparative Example 1) to which the culture powders obtained in Examples 1 to 3 were added were prepared and the degree of increase in the number of slime-producing lactic acid bacteria was observed during the storage period. Table 4 shows the sausage raw material ingredients added with the culture culture powder and the sausage raw material ingredients of Comparative Example 1. Raw meat, fat, ice, and sample were crushed, filled in a natural cage, and heat treated at 72 ℃ or higher. After cooling, it was vacuum packed and treated with hot water (85 ° C, 3 to 5 minutes). The sausages of Comparative Examples 1 and 4 to 10 were stored in a refrigerator (15 ° C) and the number of lactic acid bacteria producing slime was compared.

Sausage raw materials used for preservation experiment Raw material name Comparative Example 1 Example 4 5 6 7 8 9 10 Pork 60 60 60 60 60 60 60 60 Fat 20 20 20 20 20 20 20 20 ice 20 19.7 19.7 19.7 19.7 19.7 19.7 19.7 Example 1 - 0.3 - - 0.15 0.15 - 0.1 Example 2 - - 0.3 - 0.15 - 0.15 0.1 Example 3 - - - 0.3 - 0.15 0.15 0.1

Unit: wt%

Changes in the Number of Lactic Acid Bacteria Generated by Sausage Slime Number of lactic acid bacteria producing slime (log cfu / ml) Primary Comparative Example 1 Example 4 5 6 7 8 9 10 One 4.81 3.25 4.47 4.86 2.88 2.60 2.85 2.55 3 5.62 4.33 5.82 5.78 3.98 3.71 3.82 3.30 8 7.23 5.62 6.43 6.52 5.25 5.30 5.43 4.82 13 7.41 6.32 6.84 6.72 6.24 6.04 5.88 5.22

As shown in Table 5, the increase in the number of slime-producing lactic acid bacteria in the sausage of Example 10 in which Lactobacillus plantarum DS2, Leuconostomercenteroid DS3, and Lactobacillus saccharose DS4 culture powder were mixed 0.1% was the lowest . This indicates that the addition of the three culture culture powders to the product has the highest antimicrobial activity. Also, the slime-forming lactic acid bacteria counts of Examples 7, 8 and 9 were lower than the slime-producing lactic acid bacteria counts of Examples 4, 5 and 6, which were used at least two times Indicating that the antimicrobial activity effect is great.

EXPERIMENTAL EXAMPLE 5. Evaluation of Preservation Capacity of Kimchi Added with Lactobacillus plantarum DS2 Cultured Powder

Experiments were conducted to prepare and compare the kimchi (Examples 11 to 17) and the non-additive kimchi (Comparative Example 2) to which the cultured strains obtained in Examples 1 to 3 were added. Kimchi spices sold in the market were purchased, and the kimchi added with the samples obtained in Examples 1 to 3 and the kimchi not containing the kimchi were stored in the refrigerator (15 ° C) for storage, and the change in pH and the number of bacteria were compared. Table 6 shows the kimchi seasonings added with the samples obtained in Examples 1 to 3 and the kimchi seasoning ingredients of Comparative Example 2.

The raw ingredients of kimchi used in preservation experiment Raw material name Comparative Example 2 Example 11 12 13 14 15 16 17 radish 31.6 31.6 31.6 31.6 31.6 31.6 31.6 31.6 Chili powder 15.8 15.8 15.8 15.8 15.8 15.8 15.8 15.8 Water again 7.4 7.4 7.4 7.4 7.4 7.4 7.4 7.4 Anchovy sauce 7.5 7.5 7.5 7.5 7.5 7.5 7.5 7.5 Shrimp 6.3 6.3 6.3 6.3 6.3 6.3 6.3 6.3 garlic 5.4 5.4 5.4 5.4 5.4 5.4 5.4 5.4 Glutinous rice porridge 4.7 4.4 4.4 4.4 4.4 4.4 4.4 4.4 onion 3.2 3.2 3.2 3.2 3.2 3.2 3.2 3.2 Green onion 3.2 3.2 3.2 3.2 3.2 3.2 3.2 3.2 Anchovy sauce 3.3 3.3 3.3 3.3 3.3 3.3 3.3 3.3 Dried red pepper 2.8 2.8 2.8 2.8 2.8 2.8 2.8 2.8 ship 2.2 2.2 2.2 2.2 2.2 2.2 2.2 2.2 The 1.6 1.6 1.6 1.6 1.6 1.6 1.6 1.6 ginger 1.6 1.6 1.6 1.6 1.6 1.6 1.6 1.6 Sugar 1.6 1.6 1.6 1.6 1.6 1.6 1.6 1.6 lampshade 0.9 0.9 0.9 0.9 0.9 0.9 0.9 0.9 Hwangseok 0.9 0.9 0.9 0.9 0.9 0.9 0.9 0.9 Example 1 - 0.3 - - 0.15 0.15 - 0.1 Example 2 - - 0.3 - 0.15 - 0.15 0.1 Example 3 - - - 0.3 - 0.15 0.15 0.1

Unit: wt%

Changes in pH of kimchi during storage period pH Primary Comparative Example 2 Example 11 12 13 14 15 16 17 One 5.16 5.18 5.16 5.18 5.22 5.20 5.22 5.22 3 5.13 5.16 5.12 5.16 5.18 5.18 5.24 5.22 7 4.88 4.96 4.94 4.94 5.02 5.04 4.96 5.06 10 4.56 4.80 4.76 4.82 4.83 4.85 4.82 4.88

General bacterial changes of kimchi according to storage period General bacterium (log cfu / ml) Primary Comparative Example 2 Example 11 12 13 14 15 16 17 One 2.65 2.85 2.72 2.70 2.64 2.60 2.65 2.66 3 4.30 3.90 3.90 3.70 3.48 3.30 3.52 3.38 7 6.41 4.90 5.34 5.30 3.70 3.60 3.90 3.62 10 6.78 5.50 5.76 5.70 5.11 4.95 5.23 4.74

As shown in Tables 7 and 8, the kimchi of Example 17 in which Lactobacillus plantarum DS2, Leuconostomercenteroid DS3, and Lactobacillus saccharose DS4 culture powders were mixed in an amount of 0.1%, showed a decrease in pH and an increase in general bacteria Was the lowest. This indicates that the addition of the three culture culture powders to the product has the highest antimicrobial activity. Examples 14, 15 and 16 using two or more of Lactobacillus plantarum DS2, Leuconostomercenteroid DS3 and Lactobacillus saccharose DS4 culture powders were used in Examples 11, 12 and 13 The amount of decrease in pH and the increase in general bacteria were lower. This indicates that the use of two or more culture powders at the same time is effective in enhancing the antimicrobial activity and enhancing the preservability of kimchi.

Korea Biotechnology Research Institute KCTC12992BP 20160322 Korea Biotechnology Research Institute KCTC13016BP 20160426 Korea Biotechnology Research Institute KCTC13017BP 20160426

<110> DYNE SOZE CO., LTD <120> New lactic acid bacteria isolated from Kimchi and antibacterial          composition using the same <130> PA-D16375 <160> 3 <170> KoPatentin 3.0 <210> 1 <211> 1623 <212> DNA <213> Unknown <220> <223> Lactobacillus plantarum DS2, KCTC12992BP <400> 1 gagggatttt ttttagtttt tttcccctgc tcaggacgaa cgctggcggc gtgcttaata 60 catgcaagtc gaacgaactc tggtattgat tggtgcttgc atcatgattt acatttgagt 120 gagtggcgaa ctggtgagta acacgtggga aacctgccca gaagcggggg ataacacctg 180 gaaacagatg ctaataccgc ataacaactt ggaccgcatg gtccgagttt gaaagatggc 240 ttcggctatc acttttggat ggtcccgcgg cgtattagct agatggtggg gtaacggctc 300 accatggcaa tgatacgtag ccgacctgag agggtaatcg gccacattgg gactgagaca 360 cggcccaaac tcctacggga ggcagcagta gggaatcttc cacaatggac gaaagtctga 420 tggagcaacg ccgcgtgagt gaagaagggt ttcggctcgt aaaactctgt tgttaaagaa 480 gaacatatct gagagtaact gttcaggtat tgacggtatt taaccagaaa gccacggcta 540 actacgtgcc agcagccgcg gtaatacgta ggtggcaagc gttgtccgga tttattgggc 600 gtaaagcgag cgcaggcggt tttttaagtc tgatgtgaaa gccttcggct caaccgaaga 660 agtgcatcgg aaactgggaa acttgagtgc agaagaggac agtggaactc catgtgtagc 720 gt; ctgacgctga ggctcgaaag tatgggtagc aaacaggatt agataccctg gtagtccata 840 ccgtaaacga tgaatgctaa gtgttggagg gtttccgccc ttcagtgctg cagctaacgc 900 attaagcatt ccgcctgggg agtacggccg caaggctgaa actcaaagga attgacgggg 960 gcccgcacaa gcggtggagc atgtggttta attcgaagct acgcgaagaa ccttaccagg 1020 tcttgacata ctatgcaaat ctaagagatt agacgttccc ttcggggaca tggatacagg 1080 tggtgcatgg ttgtcgtcag ctcgtgtcgt gagatgttgg gttaagtccc gcaacgagcg 1140 caacccttat tatcagttgc cagcattaag ttgggcactc tggtgagact gccggtgaca 1200 aaccggagga aggtggggat gacgtcaaat catcatgccc cttatgacct gggctacaca 1260 cgtgctacaa tggatggtac aacgagttgc gaactcgcga gagtaagcta atctcttaaa 1320 gccattctca gttcggattg taggctgcaa ctcgcctaca tgaagtcgga atcgctagta 1380 atcgcggatc agcatgccgc ggtgaatacg ttcccgggcc ttgtacacac cgcccgtcac 1440 accatgagag tttgtaacac ccaaagtcgg tggggtaacc ttttaggaac cagccgccta 1500 aggtgggaca gatgattagg gtgaagtcgt aacaggggaa accggaaaaa tcttagtttt 1560 tcaaaagttt ccgggcgtca ggcgtcgaat ttgggacctg gatcccccga gttttggccg 1620 tgg 1623 <210> 2 <211> 1539 <212> DNA <213> Unknown <220> <223> Leuconostoc mesenteroides DS3, KCTC13016BP <400> 2 tattttttta tctttttata gattttttta cctcgctcag gatgaacgct ggcggcgtgc 60 ctaatacatg caagtcgaac gcacagcgaa aggtgcttgc acctttcaag tgagtggcga 120 acgggtgagt aacacgtgga caacctgcct caaggctggg gataacattt ggaaacagat 180 gctaataccg aataaaactt agtgtcgcat gacacaaagt taaaaggcgc ttcggcgtca 240 cctagagatg gatccgcggt gcattagtta gttggtgggg taaaggccta ccaagacaat 300 gatgcatagc cgagttgaga gactgatcgg ccacattggg actgagacac ggcccaaact 360 cctacgggag gctgcagtag ggaatcttcc acaatgggcg aaagcctgat ggagcaacgc 420 cgcgtgtgtg atgaaggctt tcgggtcgta aagcactgtt gtatgggaag aacagctaga 480 ataggaaatg attttagttt gacggtacca taccagaaag ggacggctaa atacgtgcca 540 gcagccgcgg taatacgtat gtcccgagcg ttatccggat ttattgggcg taaagcgagc 600 gcagacggtt tattaagtct gatgtgaaag cccggagctc aactccggaa tggcattgga 660 aactggttaa cttgagtgca gtagaggtaa gtggaactcc atgtgtagcg gtggaatgcg 720 tagatatatg gaagaacacc agtggcgaag gcggcttact ggactgcaac tgacgttgag 780 gctcgaaagt gtgggtagca aacaggatta gataccctgg tagtccacac cgtaaacgat 840 gaacactagg tgttaggagg tttccgcctc ttagtgccga agctaacgca ttaagtgttc 900 cgcctgggga gtacgaccgc aaggttgaaa ctcaaaggaa ttgacgggga cccgcacaag 960 cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac cttaccaggt cttgacatcc 1020 tttgaagctt ttagagatag aagtgttctc ttcggagaca aagtgacagg tggtgcatgg 1080 tcgtcgtcag ctcgtgtcgt gagatgttgg gttaagtccc gcaacgagcg caacccttat 1140 tgttagttgc cagcattcag atgggcactc tagcgagact gccggtgaca aaccggagga 1200 aggcggggac gacgtcagat catcatgccc cttatgacct gggctacaca cgtgctacaa 1260 tggcgtatac aacgagttgc caacccgcga gggtgagcta atctcttaaa gtacgtctca 1320 gttcggattg tagtctgcaa ctcgactaca tgaagtcgga atcgctagta atcgcggatc 1380 agcacgccgc ggtgaatacg ttcccgggtc ttgtacacac cgcccgtcac accatgggag 1440 tttgtaatgc ccaaagccgg tggcctaacc ttttaggaag gagccgtcta aggcaggaca 1500 gatgactggg gtgaagtcgt aacaggtgaa accgtaaaa 1539 <210> 3 <211> 1733 <212> DNA <213> Unknown <220> <223> Lactobacillus sakei DS4, KCTC13017BP <400> 3 tattgagtag atttttagag tttttgaacc agctcaggac gaacgctggc ggcgtgccta 60 gattgataaa catttgagtg agtggcggac gggtgagtaa cacgtgggta acctgcccta aagtggggga 180 taacatttgg aaacagatgc taataccgca taaaacctaa caccgcatgg tgtagggttg 240 aaagatggtt tcggctatca ctttaggatg gacccgcggt gcattagtta gttggtgagg 300 taaaggctca ccaagaccgt gatgcatagc cgacctgaga gggtaatcgg ccacactggg 360 actgagacac ggcccagact cctacgggag gcagcagtag ggaatcttcc acaatggacg 420 aaagtctgat ggagcaacgc cgcgtgagtg aagaaggttt tcggatcgta aaactctgtt 480 gttggagaag aatgtatctg atagtaactg atcaggtagt gacggtatcc aaccagaaag 540 ccacggctaa ctacgtgcca gcagccgcgg taatacgtag gtggcaagcg ttgtccggat 600 ttattgggcg taaagcgagc gcaggcggtt tcttaagtct gatgtgaaag ccttcggctc 660 aaccgaagaa gtgcatggga aactgggaaa cttgagtgca gaagaggaca gtggaactcc 720 atgtgtagcg gtgaaatgcg tagatatatg gaagaacacc agtggcgaag gcggctgtct 780 ggtctgtaac tgacgctgag gctcgaaagc atgggtagca aacaggatta gataccctgg 840 tagtccatgc cgtaaacgat gagtgctagg tgttggaggg tttccgccct tcagtgccgc 900 agctaacgca ttaagcactc cgcctgggaa gtacgaccgc aaggttgaaa ctcaaaggaa 960 ttgacggggg cccgcacaag cggtggagca tgtggtttaa ttcgaagcaa cgcgaagaac 1020 cttaccaggt cttgacatcc tttgaccact ctagagatag agctttccct tcggggacaa 1080 agtgacaggt ggtgcatggt tgtcgtcagc tcgtgtcgtg agatgttggg ttaagtcccg 1140 caacgagcgc aacccttatt actagttgcc agcatttagt tgggcactct agtgagactg 1200 ccggtgacaa accggaggaa ggtggggacg acgtcaaatc atcatgcccc ttatgacctg 1260 ggctacacac gtgctacaat ggatggtaca acgagttgcg agaccgcgag gtttagctaa 1320 tctcttaaaa ccattctcag ttcggattgt aggctgcaac tcgcctacat gaagccggaa 1380 tcgctagtaa tcgcggatca gcatgccgcg gtgaatacgt tcccgggcct tgtacacacc 1440 gcccgtcaca ccatgagagt ttgtaacacc caaagccggt gaggtaaccc ttcggggagc 1500 cagccgtcta aggtgggaca gatgattagg gtgaagtcgt aacagggaaa acccgtaaaa 1560 aaaaaaaaaa cacaggaccc ccaaacccgt cgctttggct cctttactca actccccgcc 1620 ctcctcccca gggtgtttct accctaaact ctacatacct gtcgagttcc gcacttggtc 1680 tacacattgt ttgtcatcta gacctccgtt tcttccccta tctaccaatt cac 1733

Claims (10)

The present invention relates to a method for producing a strain of Lactobacillus sakei DS4 or KCTC13017BP which comprises as an active ingredient a strain of Bacillus cereus , Escherichia coli , Staphylococcus aureus , Listeria monocytogenes Wherein the composition has antimicrobial activity against Listeria monocytogenes , Lactococcus lactis , Leuconostoc mesenteroides or Weissella viridescens . A food prepared by adding the composition of claim 1. 3. The method of claim 2,
Wherein the food is a meat-processed food or a kimchi. A food additive prepared by adding the composition of claim 1. 3. The method of claim 2,
Wherein the strain is produced by further adding one or two strains selected from Lactobacillus plantarum DS2 and KCTC12992BP and Leuconostoc mesenteroides DS3 and KCTC13016BP or a culture thereof. 5. The method of claim 4,
Wherein the food additive is prepared by further adding one or two strains selected from Lactobacillus plantarum DS2 and KCTC12992BP and Leuconostoc mesenteroides DS3 and KCTC13016BP or a culture thereof. . delete delete delete Lactobacillus sakei DS4, KCTC13017BP).

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