Latilactobacillus sakei WDGT01 strain and use thereof

Abstract

The present invention relates to a Latilactobacillus sakei WDGT01 strain, and more specifically, the present invention relates to an antioxidant composition, an anti-inflammatory composition, and a food composition, each comprising a Latilactobacillus sakei WDGT01 strain having antioxidant and anti-inflammatory activities, or a culture medium, dead cells or mebabolites thereof, and thus being capable of improving intestinal environment.

Description

Latilactobacillus sakei WDGT01 strain and uses thereof

[0001] This invention applies to cases where research and development costs have been supported by an institution (such as a company) other than the government.

[0002] [Assignment No.] D2403010

[0003] [Project Title] Securing Novel Lactic Acid Bacteria Derived from Green Tripe (Ruminant Stomach) and Developing Functional Synbiotic Compositions for Pets Utilizing Them

[0004] [Specialized Agency] Gyeonggi Economic & Science Promotion Agency

[0005] [Research Project Name] Gyeonggi Technology Development Project (First Step R&D)

[0006] [Principal Research and Development Organization] Weedd Co., Ltd.

[0007] [Joint R&D Institution] Gachon University Industry-Academic Cooperation Foundation

[0008] [R&D Period] 2024.08.01 ~ 2025.07.31

[0009] The present invention relates to a novel microbial strain derived from green tripe and its uses.

[0010]

[0011] Green tripe refers to the unprocessed fourth stomach of grass-eating ruminants, such as sheep, deer, and cattle. It contains not only essential fatty acids and digestive enzymes but also beneficial intestinal bacteria, such as Lactobacillus acidophilus, as the grass consumed by the ruminants is fermented by intestinal microorganisms. Due to these characteristics, green tripe can be used as a good source of probiotics.

[0012] The intestinal flora can change depending on the state of the human body, such as external food intake, stress, and hormone secretion. The enzymes and metabolites produced by these microorganisms are involved in metabolism within the human body and can affect the absorption of nutrients or drugs, as well as the production of toxic substances. According to the World Health Organization (WHO), probiotics refer to "living microorganisms that are beneficial to health when consumed in appropriate amounts." By consuming an appropriate amount of these probiotics, one can improve the intestinal flora by suppressing the proliferation of harmful bacteria and promoting the growth of beneficial bacteria.

[0013] Prebiotics are indigestible components that aid the growth of beneficial bacteria in the gut. They serve as a nutrient source, or food, for probiotics, and by supplying them, they can help beneficial bacteria colonize the environment, thereby contributing to the improvement of the intestinal system. These are substances that humans cannot break down but beneficial bacteria can; examples include fructooligosaccharides, galactooligosaccharides, and xylooligosaccharides.

[0014] Meanwhile, postbiotics refer to inactive microorganisms and / or their components that provide health benefits to a host. According to this definition, postbiotics may include inactive microorganisms, fragments or structures thereof, or metabolites or end products of microorganisms. Compared to live bacteria, postbiotics have a lower risk of side effects and higher safety regarding quality maintenance, making them easy to commercialize into products. They are attracting attention as fourth-generation lactic acid bacteria products, following probiotics, prebiotics, and synbiotics which contain both.

[0015] Meanwhile, while researching green tripe, the inventors confirmed that a novel strain derived from green tripe and the metabolites of the strain have anti-inflammatory effects, thereby completing the present invention.

[0016]

[0017] The present invention aims to provide a novel Latilactobacillus strain derived from green tripe.

[0018]

[0019] 1. Latilactobacillus sakei WDGT01 strain of accession number KCTC 16418BP.

[0020] 2. The Latilactobacillus sakei WDGT01 strain having the 16S rRNA of SEQ ID NO. 1 in the above 1.

[0021] 3. An antioxidant composition comprising the strain of 1, a culture medium thereof, dead cells, or metabolites.

[0022] 4. An antioxidant composition according to 3, wherein the strain, culture medium thereof, dead cells, or metabolites are included at a concentration of 0.3 to 20 mg / mL.

[0023] 5. An anti-inflammatory composition comprising the strain of 1, a culture medium thereof, dead cells, or metabolites.

[0024] 6. An anti-inflammatory composition according to 5, wherein the strain, culture medium thereof, dead cells, or metabolites are included at a concentration of 1 to 100 μg / mL.

[0025] 7. A food composition comprising the strain of 1, a culture medium thereof, dead cells, or metabolites.

[0026]

[0027] The present invention can provide a novel Latylactobacillus sakei strain derived from green tripe.

[0028] The present invention can provide the Latilactobacillus sakei WDGT01 strain of accession number KCTC 16418BP.

[0029] The present invention may provide an antioxidant composition comprising the Latilactobacillus sakei WDGT01 strain of accession number KCTC 16418BP, a culture medium thereof, a dead cell, or a metabolite.

[0030] The present invention may provide an anti-inflammatory composition comprising the Latilactobacillus sakei WDGT01 strain of accession number KCTC 16418BP, a culture medium thereof, a dead cell, or a metabolite.

[0031]

[0032] Figure 1 shows the results of performing CCK-8 to confirm the cytotoxicity of the strain of the present invention (dead cells: Heat-killedLatilactobacillus(HK-L); metabolites: Medtabolites(Met), dead cells + metabolites: Met+HK-L).

[0033] Figure 2 shows the results of a FRAP assay performed to confirm the antioxidant activity of the strain of the present invention (dead cells: Heat-killedLatilactobacillus(HK-L); metabolites: Medtabolites(Met), dead cells + metabolites: Met+HK-L).

[0034] Figure 3 shows the results of an ABTS assay performed to confirm the antioxidant activity of the strain of the present invention (dead cells: Heat-killedLatilactobacillus (HK-L); metabolites: Medtabolites (Met), dead cells + metabolites: Met+HK-L).

[0035] Figure 4 shows the results of a DPPH assay performed to confirm the antioxidant activity of the strain of the present invention (dead cells: Heat-killedLatilactobacillus(HK-L); metabolites: Medtabolites(Met), dead cells + metabolites: Met+HK-L).

[0036] Figure 5 shows the results of an SOD assay performed to confirm the antioxidant activity of the strain of the present invention (dead cells: Heat-killedLatilactobacillus(HK-L); metabolites: Medtabolites(Met), dead cells + metabolites: Met+HK-L).

[0037] Figure 6 shows the results of an NO assay performed to confirm the anti-inflammatory activity of the mixed composition of the strain and metabolite of the present invention. Statistical significance compared to the LPS-induced control group is indicated by * (p<0.05).

[0038]

[0039] The present invention provides a Latilactobacillus sakei WDGT01 strain.

[0040] The present invention provides a Latilactobacillus sakei WDGT01 strain having the 16S rRNA of SEQ ID NO. 1.

[0041] The present invention provides an antioxidant or anti-inflammatory composition and a food composition capable of improving the intestinal environment by including the Lactobacillus sakei WDGT01 strain having antioxidant and anti-inflammatory activity, a culture medium thereof, a dead cell or a metabolite.

[0042] The Latilactobacillus sakei WDGT01 strain was deposited at the Korean Collection for Type Cultures and registered under accession number KCTC 16418BP.

[0043] The Latilactobacillus sakei WDGT01 strain has antioxidant and anti-inflammatory activity.

[0044] In the present invention, the Latilactobacillus sakei WDGT01 strain may be isolated from green tripe, but is not limited thereto.

[0045] A "strain" refers to a group of bacteria or fungi that have been isolated and cultured in a pure manner and have the same genetic composition. Within a single species, there may exist several genetic variants with different genetic characteristics.

[0046] "Culture medium" means a liquid containing either the cells of microorganisms or substances secreted by microorganisms as they grow, obtained by inoculating microorganisms into a medium suitable for microbial growth and inducing their growth in a culture medium for a certain period, and prepared by filtering out impurities.

[0047] In the present invention, the culture medium may be obtained by culturing the Latilactobacillus sakei WDGT01 strain for 12 to 30 hours, 12 to 24 hours, 18 to 30 hours, or 18 to 24 hours, for example, 22 to 26 hours, but is not limited thereto.

[0048] In the present invention, the culture medium may be the culture supernatant obtained after culturing the Latilactobacillus sakei WDGT01 strain and removing the strain, the concentrate thereof, the fraction thereof, or the freeze-dried product thereof.

[0049] The culture medium and other culture conditions used for culturing the strain of the present invention may be any medium commonly used for culturing microorganisms of the genus Laetilactobacillus without any particular limitations. Specifically, the strain of the present invention may be cultured under aerobic or anaerobic conditions while controlling the temperature, pH, etc., in a conventional medium containing a suitable carbon source, nitrogen source, phosphorus, inorganic compound, amino acid, and / or vitamin.

[0050] "Dead cells" can be prepared by heat-treating live cells or treating them with formalin or other disinfectants, and the dead cells may be used even if they are substantially dead. In one embodiment, the dead cells may be prepared by freeze-drying live cells and then heat-treating them.

[0051] "Metabolites" is a collective term for various low or high molecular weight organic or inorganic substances produced and / or secreted by lactic acid bacteria during their growth. Metabolites can be obtained, for example, by filtering the culture medium of the strain.

[0052] The present invention provides a composition comprising the Latilactobacillus sakei WDGT01 strain, a culture medium thereof, a dead cell, or a metabolite.

[0053] The composition of the present invention may contain the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites at a concentration of 5 μg / mL to 20 mg / mL.

[0054] The composition of the present invention may inhibit the growth of harmful bacteria in the intestines and promote the growth of beneficial bacteria in the intestines.

[0055] The composition of the present invention may be intended to improve the intestinal flora.

[0056] The present invention provides an antioxidant composition comprising the Latilactobacillus sakei WDGT01 strain and its metabolites.

[0057] The antioxidant composition of the present invention may contain a strain, a culture medium thereof, a dead cell, or a metabolite at a concentration of 0.3 to 20 mg / mL, 0.5 to 20 mg / mL, 1 to 20 mg / mL, 1 to 15 mg / mL, 1 to 10 mg / mL, 2 to 10 mg / mL, 2.5 to 10 mg / mL, or 2.5 to 5 mg / mL.

[0058] The present invention provides an anti-inflammatory composition comprising the Latilactobacillus sakei WDGT01 strain and a metabolite thereof.

[0059] The anti-inflammatory composition of the present invention may contain its culture medium, dead cells, or metabolites at a concentration of 1 to 100 μg / mL, 5 to 70 μg / mL, 10 to 50 μg / mL, or 25 to 50 μg / mL.

[0060] The composition of the present invention may be a pharmaceutical composition comprising an active ingredient alone, or additionally comprising one or more pharmaceutically acceptable carriers, excipients, or diluents.

[0061] The active ingredient of the composition of the present invention may be the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites.

[0062] The carrier, excipient, or diluent that may be included in the pharmaceutical composition of the present invention may be lactose, dextrose, sucrose, dextrin, maltodextrin, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, microcrystalline cellulose, polyvinylpyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, or mineral oil, but is not limited thereto.

[0063]

[0064] The present invention provides a food composition comprising the Latilactobacillus sakei WDGT01 strain, a culture medium thereof, a dead cell, or a metabolite.

[0065] The present invention provides a food composition for antioxidant or anti-inflammatory purposes comprising the Latilactobacillus sakei WDGT01 strain and metabolites thereof.

[0066] The food composition of the present invention may improve the intestinal flora by inhibiting the growth of harmful bacteria in the intestines and promoting the growth of beneficial bacteria in the intestines.

[0067]

[0068] In the food composition of the present invention, the same applies unless it contradicts the foregoing matters.

[0069] The Lactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites may be added to food for anti-inflammatory purposes. When used as a food additive, the Lactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites may be added as is or used together with other food ingredients, or may be used appropriately according to conventional methods.

[0070] The food composition of the present invention may include conventional food additives, and unless otherwise specified, suitability as a food additive is determined by the specifications and standards for the relevant items in accordance with the general provisions and general test methods of the Food Additives Codex approved by the Ministry of Food and Drug Safety. Items listed in the Food Additives Codex include, for example, chemically synthesized compounds such as ketones, glycine, calcium citrate, nicotinic acid, and cinnamon acid; natural additives such as persimmon dye, licorice extract, crystalline cellulose, sorghum dye, and guar gum; and mixed preparations such as L-sodium glutamate preparations, alkaline agents added to noodles, preservative preparations, and tar dye preparations, but are not limited thereto. In addition, the food composition of the present invention may include a suitable carrier that is conventionally used in the manufacture of food compositions.

[0071] The formulation of the food composition of the present invention is prepared according to conventional methods and can be encapsulated after drying with a carrier, or formulated into other forms such as tablets, granules, powders, beverages, porridge, etc., and can be prepared in any food form other than those described above. For example, the food may be beverages, meat, chocolate, food products, confectionery, pizza, ramen, other noodles, chewing gum, candy, ice cream, alcoholic beverages, vitamin complexes, and health supplements.

[0072] The present invention provides a feed composition comprising the Latilactobacillus sakei WDGT01 strain, a culture medium thereof, a dead cell, or a metabolite.

[0073] The present invention provides a feed composition for antioxidant or anti-inflammatory purposes comprising the Latilactobacillus sakei WDGT01 strain and metabolites thereof.

[0074] The feed composition of the present invention may improve the intestinal flora by inhibiting the growth of harmful bacteria in the intestines and promoting the growth of beneficial bacteria in the intestines.

[0075]

[0076] In the feed composition of the present invention, the same applies to the food composition, provided that it does not contradict the previously mentioned matters.

[0077]

[0078] Ultimately, the present invention can provide antioxidant or anti-inflammatory uses for the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites.

[0079] Accordingly, the present invention also provides an antioxidant or anti-inflammatory method comprising the step of administering the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites to an animal (an animal including humans or an animal excluding humans).

[0080] The above animal may be a mammal.

[0081] The above animal may be an animal that requires the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites.

[0082] In addition, the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites administered above may be an effective amount of the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites.

[0083] The above Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites may be appropriately administered to individuals according to conventional methods and administration routes used in the industry as needed for the purpose or necessity, including oral administration or parenteral administration.

[0084]

[0085] In addition, the present invention provides a use for the preparation of an antioxidant or anti-inflammatory agent of the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites.

[0086]

[0087] In the methods and uses of the present invention, the composition is applied in the same way, provided that it does not contradict the previously mentioned details.

[0088]

[0089] The dosage of the Latilactobacillus sakei WDGT01 strain, its culture medium, dead cells, or metabolites included in the composition of the present invention, or used in the method or use, is, for example, 1 μg / kg / day to 300 μg / kg / day when administered orally, and preferably 1 μg / kg / day to 100 μg / kg / day.

[0090] However, this is not limited to such examples, and appropriate dosages and frequency of administration may be selected according to methods known in the art, and the amount and frequency of administration of the Latilactobacillus sakei WDGT01 strain of the present invention, its culture medium, dead cells, or metabolites actually administered may be appropriately determined by various factors such as the type of symptoms, route of administration, gender, health status, diet, age and weight of the individual, and the severity of the disease.

[0091]

[0092] Hereinafter, in order to specifically explain the present invention, it will be described in detail with reference to examples.

[0093]

[0094] Examples

[0095] Example 1. Identification of Latilactobacillus sakei WDGT01 strain

[0096] To identify the Latilactobacillus sakei WDGT01 strain, the nucleotide sequence (1444 bp) of the 16S rRNA gene of the Latilactobacillus sakei WDGT01 strain was determined, and a homology search was performed with the GenBank database to examine its phylogenetic position.

[0097] The specific nucleotide sequence of the 16S rRNA gene of the Latilactobacillus sakei WDGT01 strain is shown in Sequence List 1.

[0098] Genetic similarity was over 99% with Latilactobacillus sakei, Latilactobacillus graminis, and Latilactobacillus curvatus. Based on phylogenetic and similarity analysis results, the WDGT01 strain is a novel microbial strain belonging to Latilactobacillus sakei.

[0099]

[0100] Example 2. Sample Preparation

[0101] To confirm the physiological activity of the Lactilactobacillus sakei WDGT01 strain, dead cells, metabolites, and mixed samples of dead cells and metabolites were prepared.

[0102] First, to prepare the dead cell samples, 0.5 g of freeze-dried Latilactobacillus sakei WDGT01 strain was dissolved in 50 mL of distilled water to prepare a 10 mg / mL solution. After sterilization in an autoclave at 90°C for 30 minutes, the dead cell solution was serially diluted so that the concentrations of each sample were 10 mg / mL, 5 mg / mL, 2.5 mg / mL, and 1.25 mg / mL.

[0103] To prepare metabolite samples, 0.5 g of freeze-dried metabolite of Latilactobacillus sakei WDGT01 strain was dissolved in 50 mL of distilled water to prepare a 10 mg / mL solution. After sterilization in an autoclave at 90°C for 30 minutes, the dead cell solution was serially diluted to obtain concentrations of 10 mg / mL, 5 mg / mL, 2.5 mg / mL, and 1.25 mg / mL for each sample.

[0104] To prepare a mixed sample of dead cells and metabolites, 0.25 g of the freeze-dried dead cells and metabolites were dissolved in 50 mL of distilled water to prepare a 10 mg / mL solution. After sterilization in an autoclave at 90°C for 30 minutes, the dead cell solution was serially diluted so that the concentrations of each sample were 10 mg / mL, 5 mg / mL, 2.5 mg / mL, and 1.25 mg / mL.

[0105] The dead cells, metabolites, and samples of dead cells and metabolites prepared in this manner can be further diluted to the desired concentration for use in each experiment.

[0106]

[0107] Example 3. Evaluation of cytotoxicity of the strain

[0108] CCK-8 was performed to evaluate the cytotoxicity of dead cells, metabolites, and dead cells + metabolites samples of the Latylactobacillus sakei WDGT01 strain.

[0109] Each sample was prepared at concentrations of 1000, 500, and 100 μg / mL for the evaluation of concentration-dependent cytotoxicity. Raw 264.7 cell lines were used as the target cells and cultured in medium containing DMEM, FBS, and Penicillin-Streptomycin. 100 μL of 5 x 10⁶ 3 After adding 10 Raw 264.7 cells, the cells were cultured in a CO2 incubator for 24 hours. Subsequently, 10 μL of cell counting kit-8 was added to each well, and the cells were cultured in a CO2 incubator for 4 hours. After the reaction was complete, the absorbance was measured at 450 nm.

[0110] As a result, all samples showed cell viability similar to that of the control group, and in particular, the dead cell + metabolite sample showed the highest cell viability at all concentrations. From these results, it was confirmed that the dead cells, metabolites, and mixtures of dead cells and metabolites of the strain of the present invention all have almost no cytotoxicity (Fig. 1).

[0111]

[0112] Example 4. Antioxidant Experiment

[0113] 3-1. FRAP assay

[0114] To evaluate the antioxidant activity of the dead cells, metabolites, and dead cells + metabolites of the Latilactobacillus sakei WDGT01 strain prepared in Example 2, a FRAP assay was performed to confirm antioxidant activity by reducing the Fe(III)-TPTZ complex to Fe(II)-TPTZ.

[0115] First, 30 μL of the suspension of each sample and 70 μL of distilled water were added to 900 μL of a solution prepared by mixing 0.625 g of TPTZ (ferric tripyridyltrizaine) and 0.162 g of ferric chloride in 0.3 M acetate buffer, and the mixed solution was reacted at 37°C for 4 minutes. Ascorbic acid at a concentration of 0.00625 mg / ml was used as a control. After the reaction was completed, the absorbance at 595 nm was measured, and a higher absorbance value indicates higher antioxidant activity.

[0116] Specific measurement results for samples of each concentration are shown in Table 1 below.

[0117] TestSampleAntioxidant activity (Relative %)1.25 mg / mL2.5 mg / mL5 mg / mLLFRAPHKL74.11±0.54114.45±0.47176.14±0.34MET82.41±0.39132.14±0.11197.45±0.18HKL+MET51.12±0.97106.12±0.09167.12±0.08

[0118] Experimental results confirmed that dead cell and metabolite samples had antioxidant activity at a concentration of 2.5 mg / mL, and in particular, at a concentration of 5 mg / mL, all samples of dead cells, metabolites, and dead cells + metabolites showed significant antioxidant activity compared to the control group (Fig. 2).

[0119]

[0120] 3-2. ABTS assay

[0121] To evaluate the antioxidant activity of the dead cells, metabolites, and dead cells + metabolites of the Latylactobacillus sakei WDGT01 strain prepared in Example 2, an ABTS assay was performed.

[0122] 50 μL of suspension for each sample was prepared, and 2,2-azino-bis(3-rthylbenzthiazoline-6-sulfonicacid)diammonium salt (0.038 g / 10 mL) prepared for 16 hours and 7.35 mM potassium persulfate (0.019 g / 10 mL) were added to the suspension in a weight ratio of 1:0.5, followed by a reaction at room temperature for 6 minutes. Ascorbic acid at a concentration of 0.00625 mg / ml was used as a control. After the reaction was complete, absorbance was measured at 734 nm, and a lower absorbance value indicates higher antioxidant activity.

[0123] Specific measurement results for samples of each concentration are shown in Table 2 below.

[0124] TestSampleAntioxidant activity (Relative %)1.25 mg / mL2.5 mg / mL5 mg / mLABTSHKL49.97±1.0475.12±0.88169.12±0.19MET62.47±0.94132.15±1.11227.12±0.07HKL+MET50.11±0.78102.17±0.67204.12±1.12

[0125] Experimental results confirmed that the metabolite sample had antioxidant activity at a concentration of 2.5 mg / mL, and in particular, at a concentration of 5 mg / mL, all samples—dead cells, metabolites, and dead cells + metabolites—showed significant antioxidant activity compared to the control group (Fig. 3).

[0126]

[0127] 3-3. DPPH assay

[0128] To evaluate the antioxidant activity of the dead cells, metabolites, and dead cells + metabolites of the Latylactobacillus sakei WDGT01 strain prepared in Example 2, a DPPH assay was performed.

[0129] Antioxidant activity in a sample can be measured by utilizing the characteristic that the color of the purple DPPH (2,2-diphenyl-1-picrylhydrazyl) radical, which contains reactive oxygen species, changes to yellow when the reactive oxygen species are removed. 600 μL of 0.5 mM DPPH 2,2-diphenyl-1-picrylhydrazyl (in 100% EtOH) was mixed with 200 μL of each sample suspension and reacted in a dark room for 20 minutes. Centrifugation was performed at 13,500 rpm for 5 minutes to precipitate substances that could affect the absorbance of the samples. Ascorbic acid at a concentration of 0.00625 mg / ml was used as a control. After the reaction was complete, absorbance was measured at 490 nm, and a lower absorbance value indicates higher antioxidant activity.

[0130] Specific measurement results for samples of each concentration are shown in Table 3 below.

[0131] TestSampleAntioxidant activity (Relative %)1.25 mg / mL2.5 mg / mL5 mg / mLLDPPHHKL67.45±2.45118.12±1.98203.45±1.00MET87.47±0.08165.45±0.08222.75±0.98HKL+MET72.43±1.85137.45±1.02209.89±2.01

[0132] Experimental results confirmed that the metabolite sample had antioxidant activity at a concentration of 2.5 mg / mL, and in particular, at a concentration of 5 mg / mL, all samples—dead cells, metabolites, and dead cells + metabolites—showed significant antioxidant activity compared to the control group (Fig. 4).

[0133]

[0134] 3-4. SOD assay

[0135] To evaluate the antioxidant activity of the dead cells, metabolites, and dead cells + metabolites of the Latilactobacillus sakei WDGT01 strain prepared in Example 2, an SOD assay was performed. Antioxidant activity was tested using water-soluble tetrazolium salt and enzyme working solution (xanthine oxidase) according to the protocol of the EZ SOD assay kit.

[0136] Specific measurement results for samples of each concentration are shown in Table 4 below.

[0137] TestSampleAntioxidant activity (Relative %)0.3125 mg / mL0.625 mg / mL1.25 mg / mL2.5 mg / mLSODHKL21.97±1.3436.45±1.0137.12±1.1542.45±1.24MET28.87±1.5434.12± 1.1835.12±0.9549.72±1.01HKL+MET15.78±1.6821.45±1.3425.17±0.5737.55±0.95

[0138] Experimental results confirmed that dead cells, metabolites, and dead cells + metabolites samples exhibited SOD activity (Fig. 5).

[0139]

[0140] Example 5. Anti-inflammatory experiment

[0141] To evaluate the anti-inflammatory activity of the mixture of dead cells and metabolites of the Latylactobacillus sakei WDGT01 strain prepared in Example 2, an NO assay was performed.

[0142] A 10 mg / mL dead cell + metabolite stock sample was diluted to prepare concentrations of 10 μg / mL, 25 μg / mL, and 50 μg / mL, respectively. Raw 264.7 cells were placed in 2 x 10 well plates. 5 1 mL / well was dispensed and incubated for 24 hours in an incubator at 37°C and 5% CO2. After incubation was complete, the medium was replaced with serum-free medium containing dead cell + metabolite samples at each concentration, and incubated again for 3 hours in an incubator at 37°C and 5% CO2. Subsequently, LPS was administered to the induction control group and the test substance treatment group (dead cell + metabolite sample treatment group), and inflammation was artificially induced by incubating for 24 hours in an incubator at 37°C and 5% CO2. Once incubation was complete, cell culture supernatants from all groups were collected, and nitrate production was confirmed using the Nitric Oxide Assay Kit Colorimetric (Abcam) (Griess reaction method).

[0143] The same experiment was performed three times and averaged, and statistical analysis was conducted using the SPSS statistical program (Ver. 19.0). Levene's test was performed to evaluate the homogeneity of variances, followed by a one-way ANOVA test to confirm the significance between each test group. If no significance was confirmed between test groups as a result of the one-way ANOVA test, no further statistical processing was performed. If significance was confirmed between test groups, post-hoc tests were conducted depending on the homogeneity of variances (Scheffe multiple test for homogeneous variances, and Dunnett's T3 for heterogeneous variances). The specific results are shown in Table 5 below, and statistical significance (p<0.05) compared to the control group is indicated by *.

[0144] Group Applied Concentration (Final con.) OD5 40 - Blank (Mean ± SD) % of Provocation Control (Mean ± SD) Nitrate concentration (μM) Negative Control - 0.029 ± 0.00 5 6.43 ± 0.34 6 3.35 4 ± 0.18 1 * Provocation Control (LPS) 1 μg / mL 0.44 5 ± 0.01 3 10 0.00 ± 1.66 3 5 2.19 4 ± 0.86 8 Test Substance Application Group 10 μg / mL 0.36 5 ± 0.01 6 8 2.11 ± 1.99 6 4 2.85 6 ± 0.01 6 * 25 μg / mL 0.33 8 ± 0.00 7 7 5.91 ± 0.80 2 39.62 2 ± 0.00 7 5 0 μg / mL0.278±0.00862.57±0.71232.658±0.372

[0145] As a result, compared to the provocation control (LPS) (0.445±0.013), the OD at a concentration of 10 μg / mL 540 The values ​​were confirmed to be (0.365±0.016) as (82.11±1.996)%, (0.338±0.007) as (75.91±0.802)% at a concentration of 25 μg / mL, and (0.278±0.008) as (62.57±0.712)% at a concentration of 50 μg / mL. OD of the negative control group (Control). 540The value was (0.029±0.005), which was confirmed to be (6.43±0.346)% of the inducing control group. Based on this, the nitrate concentration was calculated to be 3.354±0.181 μM for the negative control group and 52.194±0.868 μM for the positive control group, while at 10 μg / mL, 25 μg / mL, and 50 μg / mL, the values ​​were 42.856±0.016 μM, 39.622±0.007 μM, and 32.658±0.372 μM, respectively. As can be seen from these results, it was confirmed that the strain and metabolite of the present invention possess anti-inflammatory activity at all concentrations (Fig. 6).

[0146]

Claims

1. Latilactobacillus sakei WDGT01 strain of accession number KCTC 16418BP.

2. The Latilactobacillus sakei WDGT01 strain of Claim 1 having the 16S rRNA of Sequence No.

1.

3. An antioxidant composition comprising the strain of Claim 1, a culture medium thereof, a dead cell, or a metabolite.

4. An antioxidant composition according to claim 3, wherein the strain, its culture medium, dead cells, or metabolites are included at a concentration of 0.3 to 20 mg / mL.

5. An anti-inflammatory composition comprising the strain of Claim 1, a culture medium thereof, a dead cell, or a metabolite.

6. An anti-inflammatory composition according to claim 5, wherein the strain, culture medium thereof, dead cells, or metabolites are included at a concentration of 1 to 100 μg / mL.

7. A food composition comprising the strain of Claim 1, a culture medium thereof, a dead cell, or a metabolite.

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