KR101962294B1 - Novel Lactobacillus plantarum K105 with Antioxidant and GABA-producing ability, composition comprising a fermented product of Dendropanax Morbifera extract using the same and manufacturing method therof - Google Patents

Abstract

The present invention relates to a lactobacillus plantarum strain having antioxidative and GABA production promoting activity, a fermented product of the same, and a method for preparing the same. More particularly, the present invention relates to a novel lactose having excellent activity for promoting antioxidation and GABA production Bacillus plutha room strain and a fermented product obtained by inoculating the fermented product with an extract of Fusarium sp.

Description

Lactobacillus plantarum K105 strain having antioxidant activity and GABA production promoting activity, fermented product of Fusarium oxyspermum L. using the same, and a method for producing the fermented product of Dustropanax morphifera extract using the fermented product of Novel Lactobacillus plantarum K105 with antioxidant and GABA-producing ability same and manufacturing method therof}

The present invention relates to a lactobacillus plantarum K105 strain having antioxidative and GABA production promoting activity, a fermented product of the same, and a method for producing the same, and more particularly, to a novel fermented product of a novel Lactobacillus plantarum strain and a fermented product obtained by inoculating the fermented Lactobacillus planta and an extract thereof with an antioxidant and an increased GABA content.

Gamma-amino butyric acid (GABA) is a nonprotein constituent amino acid that acts not only as a neurotransmitter in the brain but also as a brain stimulator, a mental stabilizer, a blood pressure lowering agent, a diuretic agent, , Alcohol metabolism promoting action, and deodorizing action. Especially, it is registered as a medicine and is used for treatment of headache, tinnitus and hypersensitivity due to stroke or cerebral artery sequelae.

GABA is known to be widely distributed in various vegetables, fruits, rice, beans, etc. However, its content is low and it has a disadvantage that it can not show the physiological function.

Dendropanax morbifera is a plant belonging to Araliaceae. It is a medicinal plant with excellent pharmacological efficacy. It has a scientific name Dendro-panax morbifera Nakai , meaning a panacea. In the past, it was used as a natural coating material because it is composed of 66.7% of nonvolatile matter, 10.8% of directional component, 8.1% of moisture, and 14.4% of solid content. However, recent studies have shown that diabetic, hypertensive, , Liver function improvement, antioxidation, osteoporosis, periodontal disease, immunity enhancement, nervous stability, antimicrobial, anticancer and so on.

In order to exert a pharmacological effect, such a method of extracting a useful component and taking it for a long time has been studied. However, the extract is not high in its pharmacological activity per unit weight, is slow in absorption into the human body, It is difficult to change the formulation.

As a part of this research, it has been known that fermented extracts of leafy leaves of Aspergillus oryzae were inoculated and cultured in the leaves of the leaves of Tochigi leaves, In order to increase the content of the present invention.

Therefore, the present inventors have made efforts to find a method of improving antioxidant and GABA content by using the extract of Hwangchujang. As a result, it has been found that a novel Lactobacillus plantarum strain is isolated, By confirming that the GABA content is improved, it is intended to provide a GABA high-content fermentation material and a production method thereof.

Patent Document 1. Korean Patent No. 10-1423433

The present invention has been conceived to solve the above-mentioned problems, and an object of the present invention is to provide a Lactobacillus plantarum strain K105 (accession number KCCM12014P) having antioxidative and GABA production promoting activity.

In addition, another object of the present invention is to provide a fermented yellowtail tree having increased antioxidative and GABA contents obtained by fermenting Lactobacillus plantarum K105 strain (accession number KCCM 12014P) to a perennial tree.

It is still another object of the present invention to provide a fermented product of Yellow-throated tree which is fermented by inoculation with Lactobacillus plantarum K105 strain (Deposit No. KCCM12014P) on a perilla tree.

In order to achieve the above object, the present invention provides Lactobacillus plantarum K105 strain (Accession No. KCCM12014P) having antioxidative and GABA production promoting activity.

The strain is characterized in that it is for fermentation of yellow mulberry tree.

The strain is characterized in that it is cultured in the presence of an external carbon source in addition to Hwangchu.

The strain is characterized in that it has acid resistance and biliary cholesterol.

The strain may be selected from the group consisting of leucine arylamidase,? -Galactosidase, valine arylamidase,? -Glucosidase and N-acetyl-β (N-acetyl-β-glucosaminidase), but is not active for the β-glucuronidase enzyme .

Wherein the strain is inhibited by one or more antibiotics selected from the group consisting of ampicillin, rifampicin, novobiocin and chloramphenicol, and is selected from the group consisting of kanamycin, streptomycin, lincomycin, polymycin B and vancomycin And is resistant to the above antibiotics.

The strain is characterized in that it has antibacterial activity against food poisoning bacteria.

Wherein the food poisoning bacterium is any one or more selected from the group consisting of Escherichia coli , Salmonella Typhimurium , Staphylococcus aureus , and Listeria monocytogenes .

In order to achieve the above-mentioned other objects, the present invention provides an antioxidant obtained by inoculation of Lactobacillus plantarum K105 strain (accession number KCCM12014P) and a fermented product of Ganoderma lucidum with increased GABA content do.

The whitish wood is characterized by being a whitened wood pulverized product or an extract of Hokutogi tree.

 The extract is an extract of water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.

The extract is a hot-water extract.

Wherein the GABA content of the perennial fermented product is 1600 to 2000 占 퐂 / ml or more based on the total weight of the perilla seed oil fermented product.

According to another aspect of the present invention, there is provided a method for producing a fermented Yellowlily of Korea, wherein the fermentation is carried out by inoculating L. benthamiana Lactobacillus plantarum K105 strain (Deposit No. KCCM12014P) .

The whitish wood is characterized by being a whitened wood pulverized product or an extract of Hokutogi tree.

 The extract is an extract of water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof.

The extract is a hot-water extract.

The Hwigolin extract has a total solid content of 0.5 to 5 wt%, a pH of 2 to 7, and a sugar concentration of 1 to 5 Brix.

The extract is a mixture of leaves and stems of Echinochloa crus-galli at 8 to 10: 1 weight ratio.

Wherein the fermentation is performed at 25 to 50 DEG C for 10 to 96 hours.

The fermentation is characterized in that it is cultured in the presence of an external carbon source in addition to Hwasungi.

The novel Lactobacillus plantarum strain K105 according to the present invention can increase the antioxidant and GABA contents in the extract of Wuchu chrysanthemum using the extract of Huangchu tree as a fermentation substrate. This fermented fermented yellowtail has the advantage of enhancing various physiological activities because it contains high concentration of GABA.

In addition, the fermented product according to the present invention can be used as safe medicines, food additives or cosmetic raw materials since it induces apoptosis of normal cells or does not show cytotoxicity, and thus has excellent stability to human body.

The novel Lactobacillus plantarum K105 strain of the present invention is isolated from kimchi. The fermented product of Wuchulia japonica prepared by using the Lactobacillus plantarum strain K105 is a composition containing a high concentration of GABA The problem of toxicity and side effects can be reduced.

Figure 1 is a phylogenic tree of Lactobacillus plantarum strain K105.
2 is a result of measuring the number of viable cells (a) and pH (b) for Lactobacillus plantarum strain K105 for 24 hours at 34 ° C, 37 ° C and 40 ° C using an MRS liquid medium .
Fig. 3 shows the result of measuring the bile resistance of Lactobacillus plantarum strain K105. A total of three experiments were performed at this time, and these data are expressed as mean ± standard deviation; * p < 0.05
FIG. 4 is a graph showing the acid resistance of Lactobacillus plantarum K105, which was cultured in various concentrations of HCl solutions (pH 2.0, 3.0, 4.0, and 6.4) for 3 hours and then viable viable cells were measured. A total of three experiments were performed at this time, and these data are expressed as mean ± standard deviation; * p < 0.05, ** p < 0.01
5 is a graph showing the adhesion ability of Lactobacillus plantarum strain K105 to HT29 cells. At this time, a total of three experiments were carried out and the data were expressed as mean ± standard deviation; * p < 0.05

Therefore, the inventors of the present invention have made extensive efforts to develop a new strain for the fermentation of Fusarium oxysporum, which greatly improves the functionality and functionality of the Fusarium oxysporum extract. As a result, it has been found that the strain for fermentation of Fusarium oxysporum according to the present invention, And a method for producing the same, and have completed the present invention.

One aspect of the invention relates to an antioxidant and Lactobacillus Planta room (Lactobacillu plantarum s) K105 strain with the GABA generation promoting activity (Accession No. KCCM12014P).

Lactobacillus strains belonging to the genus Lactobacillus , commonly referred to as bacilli, are part of lactic acid strains. Lactic acid bacteria are bacteria that produce lactic acid by decomposing saccharides such as glucose, and are often referred to as lactic acid bacteria.

The Lactobacillus Planta room (Lactobacillu plantarum s) K105 strain is separated from Kimchi, hwangchil For wood fermentation strain has a 16S rDNA as shown in SEQ ID NO: 1, which is a base sequence of direction "in the 3 '5.

The Lactobacillus Planta room (Lactobacillu plantarum s) K105 strain accession number is KCCM12014P.

The Lactobacillus Planta room (Lactobacillu plantarum s) K105 strain is Lactobacillus Planta room (Lactobacillu Plantarum s) and may exhibit at least 99% homology.

The Lactobacillus Planta room (Lactobacillu plantarum s) K105 strain has the advantage of being able to prepare a fermented product comprising fermented by inoculation with a hwangchil wood, a high concentration of GABA. In other words, the K105 strain is a strain having an activity of enhancing GABA content through yellowtail milling, that is, an activity of promoting antioxidation and GABA production, and a strain containing high concentration of GABA as compared with other lactic acid bacteria or Lactobacillus plantarum It is possible not only to produce the fermented yellowtail millet, but also to improve the sensibility and functionality of the fermented yellowtail millet.

The strain may be a lactic acid bacterium isolated from kimchi, and the kimchi may be one prepared by mixing with salt-seasoned vegetables and seasoning, specifically, a Kimchi is not particularly limited thereto. More preferably, It is preferable to use those which have been aged for 10 days or more.

The Lactobacillus Planta room (Lactobacillu s plantarum) K105 strain may be cultured with no external carbon source addition hwangchil tree, that is, even if the hwangchil wood is the only nutrient source in a well-grown, the high concentration of GABA since it is excellent antioxidant and GABA generation activity It is preferable to contain Horticultura as a sole nutrient source to produce the fermented Horticulture.

In addition, it is advantageous in that it does not require additional external carbon sources and therefore does not require additional processing and costs for the fermentation substrate.

The strain may be fermented at 25 to 50 캜 for 10 to 96 hours, preferably at 34 to 45 캜 for 20 to 30 hours. And the pH of the finally produced fermentation product is 4.5 or less. The optimum fermentation conditions are not particularly limited to the above-mentioned conditions, but when the fermentation temperature is lower than 34 ° C, the growth rate is significantly lowered, and there is a problem that growth is not achieved at 15 ° C. However, since it is excellent in acid resistance, it can be grown under various pH conditions, so that the fermentation conditions are simple and the cost can be reduced.

Through the K105 strain, the Fusarium oxysporum fermentation product prepared from the Fusarium oxysporum fungi as a fermentation substrate has a pH of 4.5 or lower, preferably 4-4.5, and an antioxidative activity of 92% or higher by radical scavenging activity. , The GABA content was significantly higher than 1400 ㎍ / ㎖ in relation to the total weight of the perilla seedlings.

Therefore, if the pH, antioxidant ability and GABA content of the fermented Yellowlard seedlings are in the range of the upper limit value to the lower limit value, it is possible to obtain a fermented ferulic acid fermented product having significantly improved functionality compared to the extract of Yellowlake tree from other strains belonging to the genus Lactobacillus . Compared with Lactobacillus plantarum isolated from other plant or material, it has excellent growth activity, extreme antioxidant activity and GABA production activity even under the extreme conditions which is the only nutrient source. Sensitivity was 1.5 to 2 times better.

The Lactobacillus plantarum K105 strain is highly resistant to acid because it has bile resistance and stably maintains GABA production promoting activity even at pH 2, and its long-term fixing ability (adhesion ability) 15% to 20% higher than other strains of the strain by more than 5%.

In addition, the Lactobacillus plantarum K105 strain has excellent enzyme activity against leucine arylamidase, beta -galactosidase, and then the valine arylamidase Glucosidase and N-acetyl-β-glucosaminidase, as well as the activity of the enzyme (Valine arylamidase), β-glucosidase and N-acetyl-β-glucosaminidase. However, it does not have the activity of β-glucuronidase. Therefore, it was confirmed that the strain K105 according to the present invention has excellent stability against living organisms.

The Lactobacillus plantarum strain K105 is resistant to growth by ampicillin, rifampicin, novobiocin and chloramphenicol, and is resistant to antibiotics such as kanamycin, streptomycin, lincomycin, polymycin B and vancomycin have.

The Lactobacillus Planta room (Lactobacillus plantarum) K105 strain There have high antibacterial activity against sikjungdokgyun, in particular Escherichia coli (Escherichia coli), Salmonella tie blood head Titanium (Salmonella Typhimurium), Rhodococcus fluorescein mouse stearyl fatigue ( Staphylococcus aureus) and Listeria monocytogenes (30%, 50% or more).

Another aspect of the present invention relates to a Fagus crenata fermented product having increased GABA content obtained by inoculation with Lactobacillus plantarum strain K105 (Accession No. KCCM12014P) on a perennial tree.

The above yellowtail millet fermented product is obtained by inoculating Lactobacillus plantarum K105 strain of the present invention into a woody wood.

The perennial plant may be an extract of Echinochloa crataegus or an extract of Echinochloa crus-galli.

The Hwigaechuki extract includes not only the crude extract obtained by treating the extraction raw material with the extraction solvent but also the crude extract extract. For example, the extract may be prepared in powder form by an additional process such as vacuum distillation and lyophilization or spray drying. The extract also includes fractions obtained by further fractionating the crude extract.

The extract is an extract of water, an alcohol having 1 to 4 carbon atoms or a mixed solvent thereof, and the extraction method is not particularly limited.

When water is used as the solvent, hot water extraction is preferred. For example, at a temperature of 80 to 105 DEG C, preferably 90 to 100 DEG C for 0.5 to 24 hours, preferably 1 to 6 hours. Even if the hot water is not used as a solvent, the components of Hwangryu can be extracted from the diluted solution obtained by diluting the Hwangchu tree powder with cold water or room temperature water. When extracting the woody ingredients using cold water or room temperature water, it may be extracted separately from the fermentation, or may be extracted separately from the fermentation by inoculation with Lactobacillus plantarum K105. You may.

An extract of an aqueous solution of an alcohol having 1 to 4 carbon atoms can be used. For example, extraction is carried out with an alcohol aqueous solution such as ethanol, methanol or isopropanol, preferably 20 to 80% by weight aqueous alcohol solution, more preferably 50 to 70% by weight aqueous alcohol solution. When alcohol extracts are used, before inoculation of Lactobacillus plantarum strain K105, concentrate or dry the alcohol concentrate or alcohol extract, which has lowered the alcohol content by evaporating alcohol from the alcohol extract, and dissolve in water. In addition, in the present invention, the extract of Hwangchu-myeon is broadly composed of a diluted solution of Hwangchu-myeon powder in water.

The hwangchil wood lactic acid nutrient source, such as Lactobacillus Planta room (Lactobacillus plantarum) Lactobacillus Planta room (Lactobacillus plantarum) in order to promote the growth of the K105 strain protein source, a carbohydrate source, vitamins or minerals prior to inoculation the K105 strain Further mixing is possible. The nutrient source of the strain may be a commercially available medium or may be supplemented with only necessary nutrients.

However, the Lactobacillus plantarum strain K105 according to the present invention is active even in the absence of an external carbon source, that is, the only nutrient source, and has excellent antioxidative and GABA production activity. Therefore, In order to produce the GABA-containing yellowtail fermented product, it is preferable that the germinated yellowtail be the only nutrient source.

In addition, by making the Hwangchu tree the sole nutritional source, it is possible to obtain an effect of reducing the additional process and cost for the fermentation substrate, since no additional external carbon sources are added.

In addition, the mixture of the above Eustachia japonica extract or the mixture of the extract and the nutrient may be heat-sterilized before the inoculation of the strain Lactobacillus plantarum K105.

The solid content of the extract is not particularly limited, but is 0.5 to 1% by weight when there is no other concentration step after extraction. The extract may be used directly, or may be concentrated, or may be concentrated or dried and then diluted . When the extract is concentrated and used, the solid content may be 1 to 10% by weight.

It is preferable that the extract is used in an amount of 0.5 to 5% by weight, a pH of 2 to 7, and a sugar concentration of 1 to 5 Brix to increase the growth activity and GABA production promoting activity of the strain. Most preferred.

The fermentation conditions are the same as those of general lactic acid bacteria fermentation, so that the fermentation conditions are not particularly limited, but they may be performed at 25 to 50 ° C for 10 to 96 hours, preferably 34 to 45 ° C for 20 to 30 hours.

The Fusarium oxysulfuric acid fermented product may be a fermented product containing cells or a fermented product from which cells have been removed. The fermented product from which the cells have been removed may be sterilized to include dead cells, or may be a filtrate or a centrifuged supernatant from which cells have been removed by filtration or centrifugation. Since the components contained in the extract of the present invention are biosynthesized or produced by the Lactobacillus plantarum strain K105 to GABA which is an easily available ingredient in vivo, the various physiological activity effects Respectively. This is remarkably higher than the effect obtained by adding the simple lactic acid bacteria themselves. The fermented product can be used as a liquid fermented product itself, or may be dried and pulverized.

The GABA content of the perilla seedlings was 1600 g / ml or more, which was 1.5-1.6 times greater than the GABA content of the perilla seedlings. have.

Yet another aspect of the present invention relates to a method for producing a fermented Yellow Mill tree characterized by fermentation by inoculating L. bolus Lactobacillus plantarum K105 strain (Deposit No. KCCM12014P).

The Lactobacillus plantarum K105 strain of the present invention comprises a crushed cell wall fraction of the strain, a live cell, a dead cell or a dried cell, wherein the culture liquid is a culture medium itself obtained by culturing the strain in a liquid medium, Filtration or centrifugation to remove the strain (centrifuged supernatant), and the like.

The culture solution may be dried (e.g., lyophilized) and then powdered.

The perennial plant may be an extract of Echinochloa crataegus or an extract of Echinochloa crus-galli.

The Hwigaechuki extract includes not only the crude extract obtained by treating the extraction raw material with the extraction solvent but also the crude extract extract. For example, the extract may be prepared in powder form by an additional process such as vacuum distillation and lyophilization or spray drying. The extract also includes fractions obtained by further fractionating the crude extract.

The extract is an extract of water, an alcohol having 1 to 4 carbon atoms or a mixed solvent thereof, and the extraction method is not particularly limited.

When water is used as the solvent, hot water extraction is preferred. For example, at a temperature of 80 to 105 DEG C, preferably 90 to 100 DEG C for 0.5 to 24 hours, preferably 1 to 6 hours. Even if the hot water is not used as a solvent, the components of Hwangryu can be extracted from the diluted solution obtained by diluting the Hwangchu tree powder with cold water or room temperature water. When extracting the woody ingredients using cold water or room temperature water, it may be extracted separately from the fermentation, or may be extracted separately from the fermentation by inoculation with Lactobacillus plantarum K105. You may.

An extract of an aqueous solution of an alcohol having 1 to 4 carbon atoms can be used. For example, extraction is carried out with an alcohol aqueous solution such as ethanol, methanol or isopropanol, preferably 20 to 80% by weight aqueous alcohol solution, more preferably 50 to 70% by weight aqueous alcohol solution. When alcohol extracts are used, before inoculation of Lactobacillus plantarum strain K105, concentrate or dry the alcohol concentrate or alcohol extract, which has lowered the alcohol content by evaporating alcohol from the alcohol extract, and dissolve in water. In addition, in the present invention, the extract of Hwangchu-myeon is broadly composed of a diluted solution of Hwangchu-myeon powder in water.

However, when the degree of mixture of impurities and the degree of expression of flavor are considered in the extract, the most preferred method is to use the extracted water as a fermentation substrate.

The hwangchil tree extract Lactobacillus Planta room (Lactobacillus plantarum) prior to inoculation the K105 strain Lactobacillus Planta room (Lactobacillus plantarum) in order to promote the growth of the K105 strain protein source, a carbohydrate source, such as a vitamin or mineral lactobacillus nutrients Can be further mixed. The nutrient source of the strain may be a commercially available medium or may be supplemented with only necessary nutrients.

However, the Lactobacillus plantarum strain K105 according to the present invention is active even in the absence of an external carbon source, that is, the only nutrient source, and has excellent antioxidative and GABA production activity. Therefore, In order to produce the GABA-containing yellowtail fermented product, it is preferable that the germinated yellowtail be the only nutrient source.

In addition, by making the Hwangchu tree the sole nutritional source, it is possible to obtain an effect of reducing the additional process and cost for the fermentation substrate, since no additional external carbon sources are added.

In addition, the mixture of the above Eustachia japonica extract or the mixture of the extract and the nutrient may be heat-sterilized before the inoculation of the strain Lactobacillus plantarum K105.

The solid content of the extract is not particularly limited, but is 0.5 to 1% by weight when there is no other concentration step after extraction. The extract may be used directly, or may be concentrated, or may be concentrated or dried and then diluted . When the extract is concentrated and used, the solid content may be 1 to 10% by weight.

The components of the extract are selected from the group consisting of leaves, roots, roots, and fruits. However, the components of the extract are not required in the fermentation process of the extract. In order to prevent mixing, it is most preferable to use a mixture of the leaf and stem of the Hwigae-gil tree as a raw material.

Preferably, in the present invention, the extract is prepared from the mixture of leaves and stems of 8% to 10: 1 by weight, and when the leaves are out of the above range or other parts such as roots or fruit are included, Since the composition of the extract varies, the problem of lowering the GABA content in the fermented product may arise.

It is preferable that the extract is used in an amount of 0.5 to 5% by weight, a pH of 2 to 7, and a sugar concentration of 1 to 5 Brix to increase the growth activity and GABA production promoting activity of the strain. Most preferred.

In addition, the method may further include the step of inoculating the strain into a nutrient medium (peptone, yeast extract, glucose / distilled water) before inoculating the extract, and pre-culturing the strain at 20 to 30 DEG C for 18 to 28 hours, And then inoculating the pre-culture solution to the above-mentioned Hokutogi extract.

Preferably, the Lactobacillus plantarum K105 strain is added in an amount of 1 × 10 ⁴ to 1 × 10 ⁸ CFU per g of the extract, and more preferably the Lactobacillus planta Room K105 The strain may be mixed with 1 x 10 ⁷ to 1 x 10 ⁸ CFU.

The fermentation conditions are the same as those of general lactic acid bacteria fermentation, so that the fermentation conditions are not particularly limited, but they may be performed at 25 to 50 ° C for 10 to 96 hours, preferably 34 to 45 ° C for 20 to 30 hours.

The Fusarium oxysulfuric acid fermented product may be a fermented product containing cells or a fermented product from which cells have been removed. The fermented product from which the cells have been removed may be sterilized to include dead cells, or may be a filtrate or a centrifuged supernatant from which cells have been removed by filtration or centrifugation. Since the components contained in the extract of the present invention are biosynthesized or produced by the Lactobacillus plantarum strain K105 to GABA which is an easily available ingredient in vivo, the various physiological activity effects Respectively. This is remarkably higher than the effect obtained by adding the simple lactic acid bacteria themselves. The fermented product can be used as a liquid fermented product itself, or may be dried and pulverized.

The GABA content of the perilla seedlings was 1600 g / ml or more, which was 1.5-1.6 times greater than the GABA content of the perilla seedlings. have.

Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings. The present invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein.

Experimental Method

<Preparation of Yellowtail Tree Concentrate>

10 g of water was added to 1 g of the total solid weight of the mixture, and the mixture was heated at 100 ° C for 3 hours to obtain a hot-water extract. The total solid content contained in the hot water extract was 0.74% by weight and the pH was 5.1.

The concentrate was concentrated by using a vacuum decompressor (rotavator R-124, Buchi, Switzerland) at 100 rpm and 50 ° C to prepare an extract solution suitable for fermentation. The concentrate was 4.0 Brix °, pH 5.0 , And the total solid content contained in the concentrate was 4.2 wt%.

Experimental Example 1. Strain Collection and Separation from Raw Materials (Primary)

 1) Collecting and isolating strains in traditional fermented products

(Lactobacillus MRS agar, BD Difco, Sparks, MD, USA), which is a rich medium, is used as a nutrient-rich liquid medium for collecting and isolating strains from traditional fermentation products such as kimchi, And the ratio of the medium described in the manual was maintained.

Bromocresol purple and sodium azide were further added to the MRS solid medium, and the conventional fermented product was rubbed on each of the MRS solid medium, followed by incubation at 37 ° C. for 48 hours. Then, each of the yellow colonies Colonies were selected and purely isolated.

For pure isolation, the colonies were streaked on MRS solid medium, and the colonies were inoculated on a tryptic soy agar slant and incubated at 37 ° C for 18 hours. Then, Respectively. As a result, 170 kinds of conventional fermentation products were firstly selected.

2) Strain collection and separation in crude oil

Except for traditional fermented products, crude oil was used at each of the farms supported by the crude oil inspection centers (Gyeonggi and Gangwon provinces), Jeonbuk Livestock Research Institute, Gyeongbuk Livestock Hygiene Laboratory, and Cheju Provincial Livestock Promotion Agency in central Seoul, northern and western districts. 1.1) and the process of collecting and isolating strains were performed in the same manner. As a result, 100 kinds of crude oil isolates were firstly selected.

3) Collecting and isolating strains in feces

Experimental example 1.1) and strain collection and isolation were carried out in the same manner, except that feces instead of conventional fermentation products were used after 3 hours of bowel movement. As a result, 120 kinds of fecal isolates were firstly selected.

Experimental Example 2 Selection of Strains Growing in the Yellow Forest Tree Concentrate (Secondary)

In order to isolate strains with phagocytosis, it is necessary to have acid - producing ability. Therefore, each strain was cultivated in a concentrate of Ganoderma lucidum and then a strain with a pH value of 4.5 or less was secondly selected.

The primary cultures (10 ⁹ CFU / ml) inoculated with MRS liquid medium and incubated at 37 ° C for 18 hours were inoculated 1% to the medium concentration and incubated at 37 ° C for 24 hours. A total of 13 kimchi isolates, 11 crude isolates, and 5 fecal isolates were screened by preparing a fermented tree and measuring the pH of the isolate. The strains selected in Table 1 are indicated as raw materials.

Raw material Strain name Kimchi isolate
(Traditional fermentation products) K4 K99 K105 K112 K266 KL69 KL71 KL72 KC3 KC13 KC19 KC20 KC23 Crude oil isolate Y48 Y51 Y106 Y122 Y140 Y142 Y158 Y161 R2 R24 R32 Fecal isolate BB7 BB21 BB32 JU4 JU9

Experimental Example 3. Selection of a strain having GABA production promoting activity

In the previous experiment, the strain was firstly isolated from the raw material, and the strain was inoculated with 1% of the extract in a concentration of yellowtail. After culturing for 24 hours, the pH was measured to secondarily select strains having a pH of 4.0 or less, The results were compared with the results of the DHHP radical scavenging ability and the GABA productivity assay described in 1) and 2) below, and the most active K105 strain was finally selected (Table 2).

At this time, the control group was a sample obtained by measuring the concentration of Hwangchil tree concentrate before the strain was inoculated.

1) Analysis method of DPPH radical scavenging ability

The sample was reacted with 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) to measure the radical scavenging activity of the sample. The DPPH radical scavenging activity was measured by using a spectrophotometer at 515 nm.

2) Analysis method of gamma-aminobutyric acid (GABA) content

Quantitative analysis of GABA was carried out by adjusting Zhang and Bown 's method to 96 well plate. 0.1 g of the sample and 400 μl of methanol were added to an eppendorf tube, mixed thoroughly, and thoroughly dried for about 30 minutes using a water bath preheated to 75 ° C. Then, 1 ml of 70 mM LaCl _{3 was} added, centrifuged at 13,000 × g for 5 minutes, 700 μl of the supernatant was taken, and the precipitate was discarded. 700 μl of the supernatant is taken up in an Eppendorf tube, and then 160 μl of 0.1 M KOH is added thereto. The mixture is stirred for 3-5 minutes, and further centrifuged at 13,000 × g for 5 minutes to obtain a supernatant. This was diluted and used for GABA content measurement. GABA content was determined by using an enzyme method using GABase and the amount of NADPH produced was measured at 340 nm using an ELISA reader.

Raw material Strain DPPH (%) Relative value of DPPH versus control GABA generation
(ug / mL) Relative amount of GABA relative to control Control group 86.20 ± 0.20 100 966.11 + - 35.85 100 Kimchi isolate K4 88.90 ± 0.24 103.1 1244.23 + - 35.17 128.8 K99 91.20 ± 0.18 105.8 1244.23 ± 27.79 128.8 K105 93.02 + - 0.23 107.9 1434.52 + - 31.39 148.5 K112 89.48 ± 0.17 103.8 1127.12 + - 33.80 116.7 K266 91.36 ± 0.14 106.0 1156.4 ± 36.90 119.7 KL69 87.32 + - 0.16 101.3 995.38 ± 25.28 103.0 KL71 87.70 ± 0.60 101.7 1010.02 + - 25.28 104.5 KL72 89.94 + 0.33 104.3 1214.95 ± 23.79 125.8 KC3 89.98 + 0.48 103.2 1097.85 ± 16.90 113.6 KC13 86.82 ± 0.45 100.7 936.83 + - 25.28 97.0 KC19 90.21 + - 0.41 104.5 995.38 +/- 13.79 103.1 KC20 86.90 ± 0.58 100.8 980.74 + - 66.90 101.5 KC23 88.40 +/- 0.39 102.6 1010.02 + - 25.28 104.5 Crude oil isolate Y48 88.47 ± 0.21 102.6 984.34 ± 22.28 101.9 Y51 87.40 ± 0.12 101.4 975.12 + - 17.45 100.9 Y106 88.85 ± 0.06 103.1 965.14 + 32.27 99.9 Y122 86.40 ± 0.21 100.2 954.84 + - 42.47 98.8 Y140 91.14 + 0.14 106.0 975.28 + 15.24 100.9 Y142 87.21 + - 0.45 101.2 992.14 + - 21.14 102.7 Y158 89.42 + 0.15 103.7 981.32 ± 30.14 101.6 Y161 86.78 ± 0.17 100.7 974.26 ± 14.28 100.8 R2 88.20 ± 0.11 102.3 962.74 + - 42.15 99.7 R24 87.08 + - 0.31 101.0 1012.02 + - 22.18 104.8 R32 88.12 + - 0.11 102.2 960.45 + - 39.96 99.4 Fecal isolate BB7 86.40 ± 0.21 100.2 978.40 ± 32.85 101.3 BB21 90.72 + 0.18 105.2 964.34 ± 15.73 99.8 BB32 88.30 ± 0.24 102.4 987.12 + - 24.80 102.2 JU4 87.21 + - 0.32 101.2 977.47 ± 21.52 101.2 JU9 90.02 + - 0.15 104.4 991.21 + 33.47 102.6

As shown in Table 2, it was confirmed that strain K105 was the most excellent in both DPPH scavenging activity and GABA production activity.

Even though the same kimchi was isolated as a raw material, the bacteria such as K99 and K266 showed low DPPH scavenging ability and showed a difference of 200 to 500 ㎍ per 1 ㎖ of GABA production activity even if they were close to 90%.

This difference is only 1.5 to 1.6 times the difference measured in terms of ㎖, but it is a very significant difference in the process unit.

Experimental Example 4. Evaluation of functional (physiological) activity of strain K105

The final selected K105 strain is inoculated 1% in the Hwanyeongkuk concentrate and cultured for 24 hours at 37 ℃ to obtain a Fusarium oxysporum fungus. The fermented products were evaluated for functionality (bioactivity - antioxidant / cognitive function) before fermentation.

Among the isolates isolated from K135, the functionalities of K99 and K266 strains, which have excellent GABA production ability, were also evaluated to compare their functionality with K105 (Table 3).

1) Analysis method of active oxygen (OH) scavenging activity

In order to measure the scavenging activity of the active oxygen (OH) in the yellowtail concentrate, 0.2 ml of the sample to be measured, 10 mM of FeSO ₄ .7H ₂ O, 10 mM of EDTA, 2-deoxyribose (10 mM) 1.8 ml of a phosphate buffer solution (pH 7.4) and ₂ ml of H ₂ O ₂ (10 mM) were added thereto, and the cells were incubated at 37 ° C in an incubator. After completion of the incubation, 1 ml of TCA solution (2.8%) was added to the culture solution to stop the reaction. 1 ml of 1.0% TBA solution was added and the mixture was heated at 100 ° C for 10 minutes. After rapid cooling, Were measured.

2) Superoxide dismutase (SOD) activity assay method

3 ml of Tris-HCl buffer (50 mM Tris + 10 mM EDTA, pH 8.5) and 0.2 ml of 7.2 mM pyrogallol were added to 0.2 ml of the sample to be measured, and the mixture was incubated at 25 ° C for 10 minutes. Ml was added to stop the reaction, and the amount of oxidized pyrogallic acid present in the solution was measured at 420 nm. At this time, the SOD activity was expressed as% of absorbance reduction by comparing the addition of the sample with the addition of the sample.

pH SOD-like activity (%) H OH (%) Control group 4.5 16.86 + - 7.42 82.60 +/- 1.65 K99 3.81 17.27 + - 4.15 83.54 ± 1.28 K105 3.91 22.62 ± 5.39 88.80 ± 1.09 K266 3.85 18.64 + - 4.62 82.75 + - 0.55

As shown in Table 3, the finally selected K105 strain was found to be superior in function to other strains.

Experimental Example 5. Identification of K105 Strain

1) Physiological and biochemical characterization

First, to determine the genus and species of the finally selected strain K105, the following conditions were determined: Gran staining, microscopic observation, spore generation, aerobic and anaerobic growth, catalase generation, growth at 15 ° C and 45 ° C, and ammonia production from arginine. The results of the sugar fermentation test were also shown in Table 4. The results of the fermentation experiments per API are shown in Table 5.

strains K105 Gram stain + Cell morphology rod Spore formation - Motility - Aerobic growth + Anaerobic growth + Catalase - Growth at 15 ℃ - Growth at 45 ° C + Gas from glucose - Ammonia from arginine -

As shown in Table 4, the K105 strain of the present invention was confirmed to be a Gram-positive group, and it was confirmed that it was a rod-shaped bacterium. Also, aerobic and anaerobic growth were possible, and it was not possible to grow at 15 ℃, and it was confirmed that it could grow at 45 ℃.

Strains K105 K105 K105 Control - Inositol - D-Melezitose + Glycerol - D-Mannitol + D-Raffinose + Erythritol - D-Sorbitol + Amidon - D-Arabinose - Methyl-aD-Mannopyranoside - Glycogen - L-Arabinose + Methyl-aD-Glucopyranoside - Xylitol - D-Ribose + N-AcetylGlucosamine + Gentiobiose ± D-Xylose + Amygdalin + D-Turanose + L-Xylose - Arbutin + D-Lyxose - D-Adonitol - Esculin ferric citrate + D-Tagatose - Methyl-β-D-Xylopyranoside - Salicin ± D-Fucose - D-Galactose + D-Celiobiose + L-Fucose - D-Glucose + D-Maltose + D-arabitol - D-Fructose + D-Lactose + L-Arabitol - D-Mannose + D-Melibiose + potassium Gluconate ± L-Sorbose - D-Saccharose + potassium 2-KetoGluconate - L-Rhamnose ± D-Trehalose + potassium 5-KetoGluconate - Dulcitol - Inulin -

2) 16S rDNA sequencing

For the identification of the finally selected K105 strain, 16S rDNA sequencing was performed. For 16S rDNA sequencing, the selected K105 strain was cultured in MRS broth and DNA was extracted using a genomic DNA extraction kit (iNtRON Biotechnology, Korea). The extracted DNA was confirmed using 1% agarose gel. The 16S rRNA gene was amplified by PCR using a universal primer.

The PCR conditions were 30 cycles of denaturation at 95 ° C for 1 min, annealing at 45 ° C for 1 min, and extension at 72 ° C for 1 min 30 sec.

The obtained PCR products were purified using a purification kit (iNtRON Biotechnology, Korea) for sequencing. The nucleotide sequence was determined using ABI PRISM 3799 DNA analyzer (Perkin Elmer, U.S.A), and the K105 strain was identified by comparing it with the GenBank database using NCBI's BLAST.

As a result of analysis of the 16S rDNA sequence, the K105 strain finally isolated from kimchi showed 99% homology with Lactobacillus plantarum . The physiological characteristics of the K105 strain were the antioxidant ability, It was confirmed that the active function was excellent. From the above results, strain K105 was named Lactobacillus plantarum strain K105.

In addition, in the present invention, it was confirmed that the 16S rDNA of Lactobacillus plantarum strain K105 was encoded by the nucleotide sequence of SEQ ID NO: 1, and it was deposited with the Korean Microorganism Conservation Center (KCCM) KCCM 12014P.

A phylogenic tree of the Lactobacillus plantarum strain K105 is shown in FIG.

Name of depository: Korea Microorganism Conservation Center (overseas)

Accession number: KCCM12014P

Checked on: 20170414

Experimental Example 6: Growth test of K105 strain

Growth of Lactobacillus plantarum K105 strain was measured by measuring viable cell count and pH at various temperature conditions. The viable cell counts were obtained by inoculating 1 ml of lactic acid bacteria into 150 ml of MRS liquid medium, 1 ml of each pipette, and culturing at 34 ° C, 37 ° C, and 40 ° C for 3 hours at intervals of 24 hours. Each sample was diluted with 0.1% Plate count agar was poured on a plate, hardened, incubated at 35 ° C for 48 hours and counted by pH and temperature. The measurement results are shown in Figs.

2 is a result of measuring the number of viable cells (a) and pH (b) for Lactobacillus plantarum strain K105 for 24 hours at 34 ° C, 37 ° C and 40 ° C using an MRS liquid medium .

As shown in FIGS. 2A and 2B, the optimum temperature for growth of Lactobacillus plantarum strain K105 was 34 to 45 ° C, and the optimal pH condition of the fermented product reached pH 4.3 to 4.4, which was 13 hours .

Experimental Example 7. Bile tolerance test of strain K105

The bile tolerance test was performed according to the method of Gilliland and Walker (Gilliland & Walker, J. Dairy Sci., 73, 905, 1990), and the measurement results are shown in FIG.

Fig. 3 shows the result of measuring the bile resistance of Lactobacillus plantarum strain K105. A total of three experiments were performed at this time, and these data are expressed as mean ± standard deviation; * p < 0.05

As can be seen in FIG. 3, after the 7-hour incubation, there was a slight difference between without oxgall and with oxgall, but it was basically insensitive to bile .

Experimental Example 7: Acid resistance test of strain K105

The pH tolerance test was carried out according to the method of Clark et al. (Clark et al., Cultured Dairy Products J., 28 (4), 11, 1993).

FIG. 4 is a graph showing the acid resistance of Lactobacillus plantarum K105, which was cultured in various concentrations of HCl solutions (pH 2.0, 3.0, 4.0, and 6.4) for 3 hours and then viable viable cells were measured. A total of three experiments were performed at this time, and these data are expressed as mean ± standard deviation; * p < 0.05, ** p < 0.01

As shown in FIG. 4, when the resistance of Lactobacillus plantarum K105 was compared after 3 hours in a hydrochloric acid solution, the strain of the present invention showed almost no effect on growth even at pH 2, which is stronger than pH 6.4 It was confirmed that it has very strong acid resistance.

Experimental Example 8. Adhesion test of K105 strain to colon epithelial cells (HT cell)

The following experiment was conducted to determine the ability of the finally selected Lactobacillus plantarum strain K105 to coat the colon epithelial cells (HT29-MTX cells).

HT29-MTX cells were cultured in Dulbecco's modified Eagle's medium (DMEM) medium supplemented with 10% (v / v) heat-inactivated fetal bovine serum and antibiotic mixture. At this time, the antibiotics were finally made to be 50 mg / ml penicillin, 50 mg / ml streptomycin, 50 mg / ml gentamicin and 1.25 mg / ml amphotericin B, respectively.

HT29-MTX cells were seeded in a 30 mm culture dish to 1 x 10 ⁵ cells / ml and cultured at 37 ° C for 14 days in a 5% CO ₂ incubator (about 1 × 10 ⁷ cells / ml). All experiments were repeated three times. Lactobacillus plantarum K105 culture broth was collected by centrifugation, washed twice with phosphate buffer, and suspended in DMEM without antibiotics to a bacterial: intestinal cell concentration of 10: 1. To remove antibiotics, the HT29-MTX monolayer was washed twice with Dulbecco's PBS and Lactobacillus plantarum strain K105 was added. After incubation at 37 ° C in a CO ₂ incubator for 1 hour, the supernatant was discarded and each well was gently washed three times with Dulbecco's PBS buffer to remove unattached bacteria. The cells 4% fixing solution (35% formaldehyde 100 ㎖, Na 2 HPO 4 16 g, NaH 2 PO 4 · H 2 O 4 g with distilled water to 1 L) fixed, Lactobacillus Planta room (Lactobacillus plantarum) K105 as The strain was stained with Gram and observed with an optical microscope. In order to count the number of Lactobacillus plantarum K105 cells attached to the cells, cells and Lactobacillus plantarum strain K105 were first removed from the culture dish with 0.1% TritonX-100 and serially diluted 100 [mu] l each were inoculated into the MRS solid medium by the spreading method. CFU was then counted for 2 days in anaerobic conditions. All of the experiments were repeated three times and the data were analyzed using SPSS 11.0 software for Windows (SPSS Inc., Chicago, IL, USA) (Gonzalez de los Reyes-Gavilan, Clara et al., 2011. Adhesion of even-adapted Bifidobacterium Kainulainena V., J. Reunanena, K. Hiippalaa, S. Guglielmettib, and S. Gullielmettib, J. Clin. Endocrinol. S. Vesterlundc, A. Palvaa and R. Satokaria, 2013. BopA does not have a major role in the adhesion of Bifidobacterium bifidum to intestinal epithelial cells, extracellular matrix proteins, and mucus. Appl. Environ Microbiol 79: 6989-6997 ).

5 is a graph showing the adhesion ability of Lactobacillus plantarum strain K105 to HT29 cells. At this time, a total of three experiments were carried out and the data were expressed as mean ± standard deviation; * p < 0.05

As shown in FIG. 5, the Lactobacillus plantarum K105 strain of the present invention was found to be well adhered to the intestinal cell line HT29-MTX cells.

Example 9. Enzyme activity test

For the enzyme activity test, Lactobacillus plantarum K105 strain, which was cultured in MRS liquid medium at 37 ° C for 18 hours, was diluted with physiological saline to prepare a sample of 10 ⁵ to 10 ⁶ cfu / ml, and API ZYM (API bioMerieux, France) for 5 hours at 37 ° C, followed by enzymatic reaction. The enzyme activity was expressed as 0-5 in comparison with the standard color trademark, and the results are summarized in Table 6 below. Table 6 below shows the results of comparing enzyme activities of Lactobacillus plantarum strain K105.

Enzyme K105 Alkaline phosphatase 0 Esterase (C4) 0 Esterase Lipase (C8) 0 Lipase (C14) 0 Leucine arylamidase 5 Valine arylamidase 4 Cystinearylamidase 2 Trypsin 0 alpha-chymotrypsin 0 Acid phosphatase 2 Naphtol-AS-BI-phosphohydrolase 2 alpha -galactosidase 0 β-galactosidase 5 β-glucuronidase 0 α-glucosidase 2 β-glucosidase 4 N-acetyl-β-glucosaminidase 4 alpha-mannosidase 0 alpha-fucosidase 0

*: Standard color is specified in steps from 0 to 5, 0 is negative and 5 is maximum intensity. 1 shows 5 nano moles, 2 has 10 nano moles, 3 has 20 nano moles, 4 has 30 nano moles, and 5 has an enzyme activity of 40 nano moles or more.

As can be seen in Table 6, Lactobacillus plantarum K105 showed high enzyme activity in leucine arylamidase and β-galactosidase.

Followed by the enzyme activity in valine arylamidase, β-glucosidase and N-acetyl-β-glucosaminidase. In the case of β-glucuronidase, a carcinogenic enzyme that converts benzopyrene into a carcinogenic substance, it has been shown that it has no enzyme activity and thus has excellent safety.

Experimental Example 10. Antibiotic resistance test of strain K105

The antimicrobial susceptibility test of Lactobacillus plantarum strain K105 was performed by the diffusion method of the National Committee for Clinical Laboratory Standards (NCCLS). Antibiotic discs (BBL Sensi-disc, Becton Dickinson Co., USA) used were amikacin (30 μg, AN), gentamicin (10 μg, GM), kanamycin (10 μg, AM), ampicillin (10 μg, S), neomycin (30 μg, N), streptomycin , 10 μg, AM), bacitracin (20 μg, AM), rifampicin (10 μg, AM), Novobiocin (10 μg, AM), lincomycin ), Polymyxin B (2 ㎍, L), chloramphenicol (2 ㎍, L) and vancomycin (5 ㎍, L) The final selected strain K105 was inoculated into Mueller Hinton broth (MErck, Germany) and incubated at 37 ° C for 5 hours. The strain was diluted with 0.5 MacFarland (BioMerieux) microflora and plated on Muller-Hinton agar (Difco) After drying, 15 antibiotic discs were placed and incubated at 37 ° C for 1 day. The size of inhibitory ring for each antibiotic was measured and tolerance was determined by CLSI guideline.

R has antibiotic resistance (inhibitory ring size: 0 ㎜), IS has intermediate susceptibility (inhibitory ring size: 1-5 ㎜); S indicates susceptibility (inhibition ring size:> 5 mm).

Anti-microbial agents Antibiotic resistance K105 Aminoglycosides Amikacin IS (1 mm) Gentamycin IS (3 mm) Kanamycin R (0 mM) Neomycin IS (2 mm) Streptomycin R (0 mM) β-lactams Penicillin-G IS (5 mm) Oxacillin IS (5 mm) Ampicillin S (10 mM) Gram-positive spectrum Bacitracin IS (4 mm) Rifampicin S (6 mm) Novobiocin S (7 mM) Lincomycin R (0 mM) Gram-negative spectrum Polymyxin B R (0 mM) Broad spectrum Chloramphenicol S (10 mM) Vancomycin R (0 mM)

As shown in Table 7, the strain K105 according to the present invention was found to be inhibited against growth of ampicillin, rifampicin, novobiocin and chloramphenicol as a result of investigation of susceptibility to antibiotics, and kanamycin, streptomycin, lincomycin, It was confirmed that the antibiotics of vancomycin family are resistant.

Experimental Example 11. Antibacterial Test of K105 Strain

The antimicrobial activity test was carried out according to Gilliland & Speck, J. Food Prot., 40 (12), 820, 1977. The measurement results are shown in Table 8 below. Table 8 below shows the results of measuring the antimicrobial activity of Lactobacillus plantarum strain K105 in the MRS medium. The initial count of Lactobacillus plantarum strain K105 was 4.93 ± 0.25 × 10 ⁶ CFU / ml (*), and a was measured after incubation at 37 ° C. for 6 hours It means.

pathogens Growth pathogens ^a K105 + pathogens ^a Inhibition (%) CFU / mL CFU / mL Escherichia coli 3.23 ± 0.25 × 10 ⁶ 2.00 ± 0.53 × 10 ⁶ 38.14% Salmonella typhimurium 6.46 ± 0.35 × 10 ⁶ 2.93 ± 0.40 × 10 ⁶ 54.64% Listeria monocytogenes 1.57 ± 0.20 × 10 ⁵ 1.02 ± 0.26 × 10 ⁵ 35.38% Staphyloccous aureus 3.46 ± 0.87 × 10 ⁶ 2.27 ± 0.12 × 10 ⁶ 34.62%

As can be seen in Table 8, the Lactobacillus plantarum K105 of the present invention showed a suppressive effect against Escherichia coli of 38%, Salmonella Typhimurium , , The inhibition was 54.64%. Staphylococcus aureus ( Staphylococcus aureus ) was 34.62%, which is similar to that of Staphylococcus aureus .

EXPERIMENTAL EXAMPLE 12. Confirmation of fermentation characteristics of fermented products according to kinds of fermentation medium.

10 parts by weight of water was added to 1 part by weight of the total solid content of these solid components, and the mixture was subjected to hot water extraction at 100 DEG C for 3 hours to prepare a hot water extract. The total solid content contained in the hot water extract was 0.74% by weight and the pH was 5.1.

The concentrate was concentrated using a vacuum decompressor (rotavator R-124, Buchi, Switzerland) at 100 rpm and 50 ° C to obtain an extract solution suitable for fermentation. Five kinds of Bulgogi concentrate having a weight ratio of 7: 1, a weight ratio of 11: 1, and a weight ratio of 15: 1, respectively, were used: .

In order to confirm the fermentation characteristics of the fermented product when the extract was used as a fermentation substrate, Lactobacillus plantarum strain K105 was inoculated to the five yellowtail concentrates, The percentage of DPPH (%) and GABA in the fermented product at 24 hours is shown in Table 9. In this case, in Example 3, K105 strain was fermented by inoculation with an extract of Fusarium oxysporum sp. (Weight ratio of leaves and stems of 9: 1).

number Mixed ratio of weed leaf and stem weight DPPH (%) GABA generation
(ug / mL) Comparative Example 1 Uses only Hwangchilok 85.22 ± 0.20 956.11 + - 31.85 Comparative Example 2 Use only yellow trunk trunks 84.42 ± 0.16 983.42 + - 38.25 Comparative Example 3 7: 1 91.71 + 0.24 914.34 ± 12.73 Comparative Example 4 11: 1 91.38 + - 0.44 972.15 + 26.50 Comparative Example 5 15: 1 85.23 + - 0.12 998.48 ± 21.42 Example 1 9: 1 93.02 + - 0.23 1434.52 + - 31.39

As shown in Table 9, when the concentrated fermented product of the present invention was used in a weight ratio of 8: 10: 1, the highest GABA production activity and DPPH (% ).

Since the components of the extract vary depending on each site, the difference in the raw materials causes the growth and activation of the strain, and thus the functionality and GABA content of the final fermentation product are different from each other .

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the invention is not limited to the disclosed exemplary embodiments, but, on the contrary, is intended to cover various modifications and equivalent arrangements included within the spirit and scope of the appended claims. It is natural.

<110> KOREA FOOD RESEARCH INSTITUTE <120> Novel Lactobacillus plantarum K105 with Antioxidant and          GABA-producing ability, composition comprising a fermented          product of Dendropanax Morbifera extract using the same and          manufacturing method therof <130> HP7344 <160> 1 <170> KoPatentin 3.0 <210> 1 <211> 929 <212> DNA <213> Unknown <220> <223> 16S rDNA of Lactobacillus plantarum K105 <400> 1 ccgtcggggt ctccaggcgg atgcttaatg cgttagctgc agcactgaag ggcggaaacc 60 ctccaacact tagcattcat cgtttacggt atggactacc agggtatcta atcctgtttg 120 ctacccatac tttcgagcct cagcgtcagt tacagaccag acagccgcct tcgccactgg 180 tgttcttcca tatatctacg catttcaccg ctacacatgg agttccactg tcctcttctg 240 cactcaagtt tcccagtttc cgatgcactt cttcggttga gccgaaggct ttcacatcag 300 acttaaaaaa ccgcctgcgc tcgctttacg cccaataaat ccggacaacg cttgccacct 360 acgtattacc gcggctgctg gcacgtagtt agccgtggct ttctggttaa ataccgtcaa 420 tacctgaaca gttactctca gatatgttct tctttaacaa cagagtttta cgagccgaaa 480 cccttcttca ctcacgcggc gttgctccat cagactttcg tccattgtgg aagattccct 540 actgctgcct cccgtaggag tttgggccgt gtctcagtcc caatgtggcc gattaccctc 600 tcaggtcggc tacgtatcat tgccatggtg agccgttacc ccaccatcta gctaatacgc 660 cgcgggacca tccaaaagtg atagccgaag ccatctttca aactcggacc atgcggtcca 720 agttgttatg cggtattagc atctgtttcc aggtgttatc ccccgcttct gggcaggttt 780 cccacgtgtt actcaccagt tcgccactca ctcaaatgta aatcatgatg caagcaccaa 840 tcaataccag agttcgttcg acttgcatgt attaggcacg ccgccagcgt tcgtcctgag 900 ccatgaatca aactctaacg gtgggcagg 929

Claims (21)

The present invention relates to a method for producing an antimicrobial agent against at least one food poisonous bacterium selected from the group consisting of Escherichia coli , Salmonella Typhimurium , Staphylococcus aureus , and Listeria monocytogenes . Active,
The present invention relates to a fermented product of Ganoderma lucidum with an increased GABA content, which is characterized in that it has an antioxidative activity and an activity of increasing GABA, Lactobacillus plantarum K105 strain for manufacture (Accession No. KCCM12014P). delete delete The method according to claim 1,
The strain of Lactobacillus plantarum K105 is characterized in that it has acid-fastness and bile resistance. The method according to claim 1,
The strain may be selected from the group consisting of leucine arylamidase,? -Galactosidase, valine arylamidase,? -Glucosidase and N-acetyl-β (N-acetyl-β-glucosaminidase), but is not active for the β-glucuronidase enzyme Lactobacillus plantarum K105 strain characterized. The method according to claim 1,
Wherein the strain is inhibited by any one or more antibiotics selected from the group consisting of ampicillin, rifampicin, novobiocin and chloramphenicol,
Lactobacillus plantarum K105 strain characterized in that it is resistant to any one or more antibiotics selected from the group consisting of kanamycin, streptomycin, lincomycin, polymyxin B and vancomycin. delete delete Lactobacillus plantarum strain K105 (accession number KCCM12014P) was inoculated to the Horticultural extract and fermented at 25 to 50 DEG C for 10 to 96 hours,
The extract was prepared by mixing the leaves and the stem of Pyrrhotronate brassicae at 8-10: 1 weight ratio,
The Lactobacillus plantarum K105 strain does not contain an external carbon source in addition to the extract of Hwangchilchu tree,
The Lactobacillus plantarum K105 strain is selected from the group consisting of Escherichia coli , Salmonella Typhimurium , Staphylococcus aureus and Listeria monocytogenes Wherein the growth of one or more selected food poisoning bacteria is inhibited. delete 10. The method of claim 9,
Wherein the extract is an extract of water, an alcohol having 1 to 4 carbon atoms, or a mixed solvent thereof. 10. The method of claim 9,
Wherein the extract is a hydrothermal extract. 10. The method of claim 9,
Wherein the GABA content of the perennial fermented product is in the range of 1600 to 2000 g / ml or more based on the total weight of the perennial fermented product. Comprising the step of inoculating L. pergolas Lactobacillus plantarum strain K105 (accession number KCCM12014P) and fermenting at 25 to 50 DEG C for 10 to 96 hours,
The extract was prepared by mixing the leaves and the stem of Pyrrhotronate brassicae at 8-10: 1 weight ratio,
The Lactobacillus plantarum K105 strain does not contain an external carbon source in addition to the extract of Hwangchilchu tree,
The Lactobacillus plantarum K105 strain is selected from the group consisting of Escherichia coli , Salmonella Typhimurium , Staphylococcus aureus and Listeria monocytogenes And inhibiting the growth of at least one selected food poisoning bacterium. delete 15. The method of claim 14,
Wherein the extract is an extract of water, an alcohol having 1 to 4 carbon atoms or a mixed solvent thereof. 15. The method of claim 14,
Wherein the extract is a hot-water extract. 15. The method of claim 14,
Wherein the extract has a total solid content of 0.5 to 5% by weight, a pH of 2 to 7, and a sugar concentration of 1 to 5 Brrix. delete 15. The method of claim 14,
Wherein the fermentation is carried out at 25 to 50 DEG C for 10 to 96 hours. 15. The method of claim 14,
Wherein the fermentation is carried out in the absence of an external carbon source in addition to the woody spruce.

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