Lactiplantibacillus plantarum strain and composition for preventing or treating metabolic diseases comprising same

Abstract

Disclosed is a novel Lactiplantibacillus plantarum strain having a DPP-4 activity inhibitory effect and a beneficial metabolite secretory effect while stimulating GLP-1 secretion. Also, provided is a pharmaceutical composition for treating or inhibiting metabolic diseases, comprising the strain or a culture thereof.

Description

Lactibacillus plantarum strain and composition for preventing or treating metabolic diseases containing the same

[0001] The present invention relates to a strain of Lactibacillus plantarum. The present invention also relates to a composition for preventing or treating metabolic diseases comprising such a strain.

[0002] Probiotics are microorganisms with antimicrobial and enzymatic activity that help maintain intestinal microbial balance. Furthermore, probiotics are defined as live microorganisms, either single or complex strains, administered to humans or animals in the form of dried cells or fermented products to improve intestinal flora. Probiotics must be able to inhabit the human intestine, be non-pathogenic and non-toxic, and survive their passage through the intestines. Furthermore, they must maintain viability and activity before consumption in food, be sensitive to antibiotics used for infection prevention, and not harbor antibiotic-resistant plasmids. Furthermore, they must be resistant to acids, enzymes, and bile in the intestinal environment. Recently, probiotics have been reported to improve various health functions, including intestinal health, and are emerging as a key therapeutic agent that can replace existing compound-based treatments.

[0003] Meanwhile, among the various metabolites secreted by symbiotic microorganisms, including lactic acid bacteria, outside the microbial cells during their life cycle, some are known to have beneficial metabolic effects on host cells. For example, 3-phenyllactic acid is known to have the effect of preventing obesity or type 2 diabetes in the host by regulating intestinal lipid metabolism. α-Ketoglutaric acid, which is also an intermediate in the Krebs cycle, has been reported to have lipid metabolism-improving effects, such as limiting weight gain, suppressing inflammation, and promoting browning of adipocytes, through its intestinal microflora-regulating function of reducing the proportion of bacteria belonging to the Firmicutes phylum and increasing the proportion of bacteria belonging to the Bacteroidetes phylum.

[0004] In relation to probiotic lactic acid bacteria related to the effects of preventing and treating obesity, Korean Patent No. 10-1494279 discloses Lactobacillus plantarum KY1032 strain (Accession No.: KCCM-10430) having an effect of inhibiting adipocyte differentiation, Korean Patent No. 10-0996577 discloses Lactobacillus curbetus HY7601 (Accession No.: KCTC 11456BP) having an effect of inhibiting obesity, and Korean Patent No. 10-1394348 discloses Lactobacillus plantarum DSR920 strain (Accession No.: KCCM 11210P) having an effect of inhibiting adipocyte differentiation, but the effects have not reached a level sufficient for commercial success.

[0005] The technical task of the present invention is to develop a new lactic acid bacteria strain with excellent effects in preventing and treating metabolic diseases.

[0006] To achieve the above purpose, one aspect of the present invention provides Lactiplantibacillus plantarum strain GBCC_F0227 (Accession No.: KCTC 15450BP).

[0007] Another aspect of the present invention provides a pharmaceutical composition for preventing or treating a metabolic disease, comprising Lactibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or a culture thereof as an active ingredient.

[0008] Another aspect of the present invention provides a food composition comprising Lactiplantibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or a culture thereof.

[0009] In another aspect of the present invention, a health functional food comprising Lactiplantibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or a culture thereof is provided.

[0010] Another aspect of the present invention provides a feed composition comprising Lactiplantibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or a culture thereof.

[0011] In another aspect of the present invention, a pharmaceutical composition for treating or preventing a metabolic disease that increases GLP-1 secretion or suppresses appetite is provided, comprising as an active ingredient at least one selected from a strain of the species Lactibacillus plantarum, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, and a cell-free culture supernatant of the strain.

[0012] The Lactibacillus plantarum GBCC_F0227 strain of the present invention (Accession No.: KCTC 15450BP) has excellent GLP-1 secretion stimulation efficacy, such as being able to secrete a large amount of metabolites that are effective in stimulating GLP-1 secretion in the host into the cell, and further, can effectively inhibit DPP-4 enzyme activity and promote the expression of GLP-1 receptors, thereby doubling the beneficial effects of GLP-1 action. The GBCC_F0227 strain of the present invention is effective in improving fat metabolism. In particular, it has the effect of reducing white fat accumulation that may accompany obesity in visceral adipose tissues such as the liver and restoring adiponectin secretion, and can also suppress the expression of genes that contribute to visceral neutral fat accumulation, such as CD36 and PPARγ. Therefore, the GBCC_F0227 strain and a composition containing it can contribute to the prevention and treatment of fatty liver and non-alcoholic fatty liver disease. In addition, the GBCC_F0227 strain of the present invention also exhibits an excellent blood sugar improvement effect, and in particular, since it can reduce fasting blood sugar levels, it can effectively improve insulin resistance by improving glucose tolerance and increasing insulin sensitivity. Furthermore, the GBCC_F0227 strain of the present invention has the effect of preventing the increase of blood triglycerides and blood cholesterol accompanying diet-induced obesity. In addition, the GBCC_F0227 strain of the present invention can secrete a large amount of metabolites that suppress weight gain, and in addition to the effect of enhancing GLP-1 secretion, it can also promote the expression of GLP-1 receptor, POMC, and CART genes that contribute to suppressing appetite under obese conditions, and therefore has an excellent anti-obesity effect of suppressing weight gain and improving body fat composition. Therefore, the GBCC_F0227 strain of the present invention and a composition containing the same can be usefully used to prevent or treat metabolic diseases such as obesity and metabolic syndrome.

[0013] Figure 1a is a graph evaluating the degree to which the culture supernatant of Lactobacillus plantarum strains, among lactic acid bacteria strains isolated from fermented foods, stimulates GLP-1 secretion of the NCI-H716 cell line. In the graph, NS indicates an experiment in which no sample was added to the NCI-H716 cell line, and MRS indicates an experiment in which an untreated culture without culturing the lactic acid bacteria strain was added.

[0014] Figure 1b is a graph evaluating the degree to which the culture supernatant of the Lactobacillus paracasei strain, among lactic acid bacteria strains isolated from fermented foods, stimulates GLP-1 secretion of the NCI-H716 cell line. In the graph, NS indicates an experiment in which no sample was added to the NCI-H716 cell line, and MRS indicates an experiment in which an untreated culture without culturing the lactic acid bacteria strain was added.

[0015] Figure 1c is a graph evaluating the degree to which the culture supernatant of the Lactobacillus coryniformis strain, among lactic acid bacteria strains isolated from fermented foods, stimulates GLP-1 secretion in the NCI-H716 cell line. In the graph, NS indicates an experiment in which no sample was added to the NCI-H716 cell line, and MRS indicates an experiment in which an untreated culture without culturing the lactic acid bacteria strain was added.

[0016] Figure 1d is a graph evaluating the degree to which the culture supernatant of the Lactiplantibacillus sakei strain, among lactic acid bacteria strains isolated from fermented foods, stimulates GLP-1 secretion of the NCI-H716 cell line. In the graph, NS represents an experiment in which no sample was added to the NCI-H716 cell line, and MRS represents an experiment in which an untreated culture without culturing the lactic acid bacteria strain was added.

[0017] Figure 1e is a graph evaluating the degree to which the culture supernatant of Lactiplantibacillus fermentum strains, among lactic acid bacteria strains isolated from fermented foods, stimulates GLP-1 secretion of the NCI-H716 cell line. In the graph, NS indicates an experiment in which no sample was added to the NCI-H716 cell line, and MRS indicates an experiment in which an untreated culture without culturing the lactic acid bacteria strain was added.

[0018] Figure 1f is a graph evaluating the degree to which the culture supernatant of other isolated strains, which are strains of the genus Lactobacillus other than the aforementioned species among the isolated lactic acid bacteria strains, stimulates GLP-1 secretion of the NCI-H716 cell line. In the graph, NS indicates an experiment in which no sample was added to the NCI-H716 cell line, and MRS indicates an experiment in which an untreated culture without culturing the lactic acid bacteria strain was added.

[0019] Figure 1h is a genetic tree constructed by comparing the GBCC_F0227 genome sequence with the genomes of other Lactiplantibacillus plantarum strains and the Lactiplantibacillus argentoratensis genome sequence.

[0020] Figure 1g is a graph evaluating the degree to which the culture supernatant of strains of the genus Bifidobacterium, among lactic acid bacteria strains isolated from fermented foods, stimulates GLP-1 secretion in the NCI-H716 cell line. In the graph, NS represents an experiment in which no sample was added to the NCI-H716 cell line, and MRS represents an experiment in which an untreated culture without culturing the lactic acid bacteria strain was added.

[0021] Figure 2a is a graph comparing the GLP-1 secretion stimulation efficacy of the Lactobacillus plantarum GBCC_F0227 strain with that of other known, distributed strains of the same species and other isolated strains of the same species obtained from fermented foods to determine whether the GLP-1 secretion stimulation efficacy of this strain is specific to this strain. In the graph, NS indicates an experiment in which no sample was added to the NCI-H716 cell line, and MRS indicates an experiment in which an untreated culture without culturing a lactic acid bacteria strain was added.

[0022] Figure 2b is a graph comparing the GLP-1 secretion stimulation efficacy of Lactobacillus plantarum isolated strains, including GBCC_F0227, with that of the known plantarum strain GB104.

[0023] Figure 3 is a graph showing the relationship between administration of the GBCC_F0227 strain and blood GLP-1 concentration in mice.

[0024] Figure 4a is a graph showing the DPP-4 enzyme activity inhibitory effect of selected Lactobacillus plantarum strains.

[0025] Figure 4b is a graph showing the results of an experiment comparing the DPP-4 enzyme activity inhibitory effect of selected Lactibacillus plantarum strains with that of known Lactibacillus plantarum strains.

[0026] Figure 4c is a graph showing the results of an experiment comparing the DPP-4 enzyme activity inhibition effect of the GBCC_F0227 strain and other strains of the genus Lactiplantibacillus.

[0027] Figure 5 is a graph showing the correlation between GLP-1 secretion stimulation activity and DPP-4 activity inhibition efficacy for each selected isolated strain of the present invention.

[0028] Figure 6a is a graph showing the extent to which the GBCC_F0227 strain inhibits weight gain in high-fat diet mice.

[0029] Figure 6b is a graph showing how body fat and lean body mass composition change in high-fat diet-induced obese mice when administered the GBCC_F0227 strain.

[0030] Figure 7a is an oral glucose tolerance test in mice administered the GBCC_F0227 strain, Figure 7b shows fasting blood glucose measurements, and Figure 7c is a graph showing the area under the graph between blood glucose and time.

[0031] Figures 8a and 8b are tables showing the results of quantitative analysis of secreted metabolites after MRS medium culture for the L. platanrum strain of the present invention (indicated as F0227) and other lactic acid bacteria strains (L. platanrum strains (GBCC_F0053 (indicated as F0053), GBCC_F0058 (indicated as F0058), KCTC3108), Limosilactobacillus fermentum strains (GB102, KCTC3112), and Lactobacillus paragasseri strains (H0213, KCTC3163)).

[0032] Figure 9a is a graph of the results of an in vitro experiment evaluating the efficacy of seven types of lactic acid bacteria metabolites to stimulate GLP-1 secretion from NCI-H716 cells in a concentration-dependent manner.

[0033] Figure 9b is a graph showing the quantitative analysis of GLP-1 secreted by NCI-H716 cells by seven types of lactic acid bacteria metabolites at a concentration of 25 mM.

[0034] Figure 10a shows photomicrographs of fragments of the left lobe of the liver and epididymal fat from obese negative control mice (G1) fed a normal diet and a cryoprotectant daily for 8 weeks, obese positive control mice fed a high-fat diet and a cryoprotectant (G2), or experimental mice fed a high-fat diet and a cryoprotectant concurrently with Lactoplantibacillus plantarum GBCC_F0227 (G3). The scale bar indicates 1000 μm in the top row of images, 500 μm in the bottom row of images, and 100 μm in the images below that.

[0035] Figure 10b shows the area fraction (%) of fat droplets in fat cells in the left lobe of the liver or the average size (μm) of fat droplets in the epididymal fat for the mice in groups G1, G2, and G3 of Figure 10a. 2 ) is a graph showing the

[0036] Figure 10c is a photomicrograph of tissue sections from the left lobe of the liver of mice in groups G1, G2, and G3 of Figure 10a, stained with Oil Red O. The scale bar in the photograph indicates 100 μm.

[0037] Figure 10d is a graph showing the oil red-O staining intensity measured in the experiment of Figure 10c as a percentage of the control group (G1) staining results. In the graph, * indicates significance for the p value calculated by performing a one-way ANOVA Tukey test on the corresponding measurement value (p * <0.05,p ** <0.01,p *** <0.001). Figure 10e is a graph showing that the GBCC_F0227 strain suppresses the increase in blood triglycerides and cholesterol caused by a high-fat diet in a mouse model.

[0038] Figure 11 is a graph measuring the triglyceride concentration in the culture supernatant before and after culture for 31 probiotic strains including Lactobacillus plantarum GBCC_F0227.

[0039] Figure 12 shows the absorbance OD at 600 nm for five strains of L. plantarum species, including Lactobacillus plantarum GBCC_F0227. 600 This is a graph measuring the triglyceride concentration in the supernatant obtained by culturing until reaching 3 or 6 for each strain (Fig. 12a) and a graph organizing this based on the absorbance value (Fig. 12b). In the graph of Fig. 12, the *** mark indicates that the p-value calculated by performing a one-way ANOVA test on the corresponding triglyceride measurement value is less than 0.001.

[0040] Figure 13 shows the absorbance OD at 600 nm of Lactobacillus plantarum GBCC_F0227 and Lactobacillus plantarum WCFS1 strains. 600 This is a graph measuring the relative expression rate of genes belonging to the α / β hydrolase gene group when cultured to 3 (Fig. 13a) or 6 (Fig. 13b). In the graph of Fig. 13, the ** mark indicates that the p-value calculated by performing an unpaired Student's t-test on the relative expression rate measurement value of the corresponding gene is less than 0.01, and the *** mark indicates that the calculated p-value is less than 0.001.

[0041] Figure 14 shows the results of an experiment measuring the expression rate of genes related to fat metabolism and obesity in three groups of obese negative control mice (G1) administered a normal diet and a cryoprotectant daily for 8 weeks, or obese positive control mice (G2) administered a high-fat diet and a cryoprotectant daily, or experimental group mice (G3) administered a high-fat diet and GBCC_F0227 simultaneously, or in the case of Figures 14e to 14g, four groups of control mice (G4) administered a high-fat diet and the same amount of KCTC 3108 strain as G3. In the graph of Figure 14, the * mark indicates that the p-value calculated by performing a one-way ANOVA test with the comparative measurement value for the corresponding measurement value is less than 0.05, the ** mark indicates that the p-value is less than 0.01, and the *** mark indicates that the p-value is less than 0.001, respectively.

[0042] Figure 14a shows the change in the expression rate of the adiponectin gene in the mouse epididymis, Figure 14b shows the change in the expression rate of the leptin gene in the mouse epididymis, Figure 14c shows the change in the expression rate of the CD36 gene in the left lobe of the liver, Figure 14d shows the change in the expression rate of the PPARγ gene in the left lobe of the liver, Figure 14e shows the change in the expression rate of the GLP-1 receptor gene in the hypothalamus, Figure 14f shows the change in the expression rate of the POMC gene in the hypothalamus, and Figure 14g shows the change in the expression rate of the CART gene in the hypothalamus.

[0043] Figure 15 is a graph showing the change in hindlimb skeletal muscle mass in three groups of mice: obese negative control mice (G1) that were administered a normal diet and a cryoprotectant daily for 8 weeks, obese positive control mice (G2) that were administered a high-fat diet and a cryoprotectant, or experimental group mice (G3) that were administered a high-fat diet and Lactobacillus plantarum GBCC_F0227 concurrently. In the graph of Figure 15, the * mark indicates that the p-value calculated by performing a one-way ANOVA test on the corresponding measurement value and the comparison target measurement value is less than 0.05, the ** mark indicates that the p-value is less than 0.01, the *** mark indicates that the p-value is less than 0.001, and ns indicates that the difference is not statistically significant. Figure 15a is a graph showing the muscle mass of the tibialis anterior (TA), Figure 15b is an extensor digitorum longus (EDL), Figure 15c is an graph showing the muscle mass of the soleus (Sol), and Figure 15d is an graph showing the muscle mass of the gastrocnemius (GA) of the hind leg, respectively.

[0044] Figure 16 is a graph showing the results of a grip strength measurement test on mice fed the 8-week diet of Figure 15. In the graph of Figure 16, the ** mark indicates that the p-value calculated by performing a one-way ANOVA test on the corresponding measurement value and the comparison target measurement value is less than 0.01, and the *** mark indicates that the p-value is less than 0.001.

[0045] Hereinafter, embodiments of the present invention will be described in detail. Prior to this, it should be noted that the terms and words used in this specification and claims should not be construed as limited to their conventional or dictionary meanings. Based on the principle that the inventor can appropriately define the concept of a term to best explain his or her invention, they should be interpreted in a way that aligns with the technical spirit of the present invention.

[0046] Therefore, the embodiments described in this specification are merely examples presented for the purpose of helping to understand the present invention and do not represent all of the technical ideas of the present invention. Therefore, it should be understood that there may be various equivalents and modified examples that can replace them at the time of filing this application.

[0047] Unless otherwise defined, all technical terms used herein have the same meaning as commonly understood by those skilled in the art. While preferred methods and samples are described herein, similar or equivalent methods are also included within the scope of the present invention. Numerical values ​​described herein are considered to include the meaning of "about" even if not explicitly stated. The numerical range indicated using the term "to" herein includes the ranges that include the values ​​described before and after the term "to" as the lower and upper limits, respectively.

[0048] As used herein, “metabolic syndrome” refers to a condition characterized by metabolic risk factors including abdominal obesity, insulin resistance or glucose intolerance, atherosclerotic dyslipidemia, prothrombotic state, proinflammatory state, and hypertension. The Adult Treatment Panel defines metabolic syndrome as a condition in which a patient exhibits at least three of the following symptoms:

[0049] Waist measurement of 40 inches or more for men and 35 inches or more for women;

[0050] Serum triglyceride level greater than 150 mg / dL;

[0051] HDL cholesterol levels less than 40 mg / dL for men and 50 mg / dL for women;

[0052] Blood pressure of 135 / 80 mmHg or higher and blood sugar of 110 mg / dL or higher.

[0053] In the United States, it's estimated that more than 25% of the population suffers from metabolic syndrome. One cause of metabolic syndrome is thought to be insulin resistance, a condition in which insulin's ability to absorb sugar from the blood is impaired. This causes elevated blood sugar levels after meals in the pancreas, which secretes insulin. If left untreated, metabolic syndrome significantly increases the risk of type 2 diabetes, cardiovascular disease, and other conditions associated with plaque buildup in arterial walls.

[0054] As used herein, "diabetes mellitus" refers to a chronic disease characterized by a relative or absolute deficiency of insulin, resulting in glucose intolerance. Diabetes mellitus includes all types of diabetes, including, but not limited to, type 1 diabetes, type 2 diabetes, and hereditary diabetes. Type 1 diabetes is insulin-dependent diabetes, primarily caused by the destruction of β-cells. As used herein, the term "type 2 diabetes" refers to non-insulin-dependent diabetes, caused by insulin resistance. It develops when the body fails to detect an increase in insulin in muscle and adipose tissue, or when it detects it, insulin does not work effectively.

[0055] As used herein, "insulin resistance" refers to a condition in which an individual's metabolic system fails to respond to insulin, which lowers blood sugar levels, preventing cells from effectively burning glucose. High insulin resistance results in excessive insulin production compared to optimal insulin responsiveness, leading to conditions such as hyperinsulinemia, hypertension, or dyslipidemia, as well as heart disease and diabetes.

[0056] In this specification, "insulin resistance syndrome" refers to a general term for diseases caused by the aforementioned insulin resistance. Insulin resistance syndrome is characterized by cellular resistance to insulin action, hyperinsulinemia, increased very low density lipoprotein (VLDL) and triglycerides, decreased high density lipoprotein (HDL), and hypertension, and is recognized as a risk factor for cardiovascular disease and type 2 diabetes.

[0057] In the present invention, the term "culture" means a product obtained by culturing lactic acid bacteria in a known medium, and the product may include lactic acid bacteria. The medium may be selected from known liquid media or solid media, and may be, for example, MRS liquid media, GAM liquid media, MRS agar media, GAM agar media, and BL agar media, but is not limited thereto. The term "cell-free culture supernatant" in the present invention refers to a liquid mixture obtained by culturing a lactic acid bacteria strain for a certain period of time and removing bacterial cells from the culture, and includes a liquid mixture containing metabolites, excess nutrients, etc. produced during the culturing process of the strain. In addition, the term "cell-free culture supernatant" in the present invention encompasses all of the concentrate of the cell-free culture supernatant obtained immediately after culturing the lactic acid bacteria strain, or the extract or fraction thereof. As the cell-free culture supernatant, a liquid obtained by leaving the culture solution still for a certain period of time and taking only the liquid in the upper layer excluding the portion that has settled to the lower layer, a liquid obtained by removing the bacterial cells through filtration, a liquid obtained by centrifuging the culture solution to remove the sediment at the bottom and taking only the liquid in the upper layer, etc. can be used. In the present invention, the term "lysate" means lactic acid bacteria that have been destroyed by enzyme treatment, homogenization, or ultrasonic treatment. In addition, the term "extract" in the present invention means a product obtained by extracting lactic acid bacteria with a known extraction solvent. In addition, the term "live cells" in the present invention means the novel lactic acid bacteria of the present invention themselves, and "dead cells" mean lactic acid bacteria that have been sterilized by heating, pressurization, or drug treatment.

[0058] In the present invention, the term “prevention” means any act of suppressing symptoms or delaying progression of a disease, such as a metabolic disease, in a host administered the pharmaceutical composition of the present invention.

[0059] In the present invention, the term “treatment” means any act of improving or beneficially altering the symptoms of a disease, such as a metabolic disease, in a host administered the pharmaceutical composition of the present invention.

[0060] In the present invention, when referring to Lactibacillus plantarum, the term “secretion” refers to a phenomenon in which a Lactibacillus plantarum strain secretes a substance such as a metabolite or a signaling substance out of its cell, for example, into a medium or a host body.

[0061] In the present invention, "3-phenyllactic acid" in the context of Lactibacillus plantarum secreting substances into the extracellular space is a term encompassing not only the undissociated neutral molecular form (i.e., 2-hydroxy-3-phenylpropanoic acid in its IUPAC name) but also the dissociated anion or salt form (i.e., 2-hydroxy-3-phenylpropanoic acid anion or a salt of the anion), unless specifically indicated otherwise.

[0062] In the present invention, "α-ketoglutaric acid" in the context of Lactibacillus plantarum secreting substances into the extracellular space is a term encompassing not only the undissociated neutral molecular form (i.e., 2-oxopentanedioic acid in its IUPAC name) but also the dissociated anion or salt form (i.e., 2-oxopentanedioic acid anion or a salt of the anion), unless specifically indicated otherwise.

[0063] In the present invention, the term "host" refers to a person who has been administered a pharmaceutically effective amount of the Lactobacillus plantarum strain disclosed in the present invention, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, or an extract of the culture.

[0064] In the present invention, "2-hydroxy-3-methylbutyric acid" in the context of Lactobacillus plantarum secreting substances into the extracellular space is a term encompassing not only the undissociated neutral molecular form (i.e., the neutral molecule of 2-hydroxy-3-methylbutanoic acid in the IUPAC name) but also the dissociated anion or salt form (i.e., 2-hydroxy-3-methylbutyrate anion or a salt of the anion), unless specifically indicated otherwise.

[0065] In one aspect of the present invention, a Lactiplantibacillus plantarum GBCC_F0227 strain (Accession Number: KCTC 15450BP) is provided. Lactiplantibacillus bacteria are gram-positive rod-shaped bacteria that are widely distributed in nature and are oxygen-tolerant anaerobic microorganisms, and are a type of lactic acid bacteria. Lactiplantibacillus was previously widely known by the name Lactobacillus. Microorganisms belonging to the genus Lactiplantibacillus include Lactiplantibacillus plantarum and Lactiplantibacillus sakei. The present inventors selected a novel Lactiplantibacillus plantarum strain exhibiting both excellent GLP-1 secretion stimulation activity and DPP-4 enzyme inhibition activity, and named it "Lactiplantibacillus plantarumGBCC_F0227." The strain was deposited with the Biological Resource Center of the Korea Research Institute of Bioscience and Biotechnology on May 22, 2023, under the deposit number KCTC 15450BP. In addition, the strain corresponds to a probiotic strain, is harmless to the human body, and can be used without side effects. In the present invention, the "Lactiplantibacillus plantarumGBCC_F0227" strain is L. plantarum GBCC_F0227 or Lactibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or it may be simply abbreviated as GBCC_F0227 without the name of the bacterial species, and in the drawings and graphs attached to the specification, it may be expressed more simply as F0227 or f0227.

[0066] The GBCC_F0227 strain of the present invention, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has an effect of stimulating the secretion of glucagon-like peptide-1 (GLP-1), which has the function of increasing energy consumption and suppressing appetite in a host.

[0067] The GBCC_F0227 strain of the present invention, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has an effect of promoting the expression of a glucagon-like peptide-1 (GLP-1) receptor gene in a host. More specifically, the GBCC_F0227 strain of the present invention has an effect of promoting the expression of a GLP-1 receptor gene in a host under diet-induced obesity conditions.

[0068] Dipetidyl peptidase-4 (DPP-4) enzyme is known to degrade GLP-1 secreted from enteroendocrine cells (L cells) within 5 minutes, and therefore inhibition of this enzyme is beneficial for the maintenance and action of GLP-1 in vivo. The GBCC_F0227 strain of the present invention has the ability to promote GLP-1 secretion and GLP-1 receptor expression while inhibiting DPP-4 activity, thereby preventing the destruction of secreted GLP-1 and maintaining the activity of GLP-1 for a long time, and thus a strong GLP-1 action effect on the host can be expected.

[0069] The GBCC_F0227 strain of the present invention, the lysate of the strain, the extract of the strain, the dead cells of the strain, the culture of the strain, the extract of the culture or the cell-free culture supernatant of the strain, has an effect of stimulating appetite-suppressing neurons of a host. More specifically, the GBCC_F0227 strain of the present invention, the lysate of the strain, the extract of the strain, the dead cells of the strain, the culture of the strain, the extract of the culture or the cell-free culture supernatant of the strain has an effect of enhancing the expression of genes capable of suppressing appetite, particularly the proopiomelanocortin (POMC) gene and the CART (cocaine- and amphetamine-regulated transcript) gene, under diet-induced obesity conditions.

[0070] The GBCC_F0227 strain of the present invention, the lysate of the strain, the extract of the strain, the killed cells of the strain, the culture of the strain, the extract of the culture or the cell-free culture supernatant of the strain, has an effect of promoting the expression of the adiponectin gene, which has a protective effect against hepatic fat accumulation under diet-induced obesity conditions. The GBCC_F0227 strain of the present invention, the lysate of the strain, the extract of the strain, the killed cells of the strain, the culture of the strain, the extract of the culture or the cell-free culture supernatant of the strain, further has an effect of suppressing the expression of CD36 and PPARγ, which are genes contributing to visceral neutral fat accumulation under diet-induced obesity conditions.

[0071] The GBCC_F0227 strain of the present invention, the lysate of the strain, the extract of the strain, the dead cells of the strain, the culture of the strain, the extract of the culture, or the cell-free culture supernatant of the strain has an effect of reducing blood triglycerides of a host. More specifically, the GBCC_F0227 strain of the present invention, the lysate of the strain, the extract of the strain, the dead cells of the strain, the culture of the strain, the extract of the culture, or the cell-free culture supernatant of the strain has an effect of reducing an increase in blood triglycerides of a host that occurs under diet-induced obesity conditions.

[0072] The GBCC_F0227 strain of the present invention, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has an effect of reducing visceral fat accumulation in a host. According to one embodiment of the present invention, the GBCC_F0227 strain of the present invention can suppress fat accumulation in white visceral fat caused by a high-fat diet. More specifically, the GBCC_F0227 strain of the present invention can suppress an increase in the fraction and size of fat fraction in white visceral fat tissue caused by a high-fat diet. Most specifically, the GBCC_F0227 strain can suppress fat accumulation in the liver, particularly white fat accumulation, and thus can contribute to the prevention and treatment of fatty liver and non-alcoholic fatty liver disease.

[0073] According to one embodiment of the present invention, the strain can reduce blood sugar levels. Preferably, it can reduce fasting blood sugar levels and improve glucose tolerance. Specifically, in one embodiment of the present invention, when the strain was orally administered to a mouse model fed a high-fat diet, the fasting blood sugar levels and post-glucose administration blood sugar levels of the mouse model were reduced. On the other hand, when a general lactic acid bacteria was orally administered to a mouse model fed a high-fat diet, the fasting blood sugar levels and post-glucose administration blood sugar levels of the mouse model did not decrease. Through this, it was confirmed that the strain reduces fasting blood sugar levels and improves glucose tolerance (Figures 7a to 7c). This effect may lead to improved insulin resistance.

[0074] The GBCC_F0227 strain of the present invention has the effect of alleviating the skeletal muscle loss accompanying obesity in a host. In particular, the GBCC_F0227 strain of the present invention has the effect of alleviating the skeletal muscle mass loss and / or skeletal muscle strength loss accompanying obesity in a host.

[0075] The GBCC_F0227 strain of the present invention has the advantageous effect of secreting a large amount of α-ketoglutaric acid and 3-phenyllactic acid, which are metabolites that can help prevent type 2 diabetes by regulating fat metabolism and intestinal flora and have the function of suppressing weight gain, into the extracellular space, for example, in a larger amount than the Lactibacillus plantarum type strain under the same culture conditions. In one embodiment, the Lactibacillus plantarum GBCC_F0227 strain of the present invention secretes 1.5 to 2.5 times more α-ketoglutaric acid and 3-phenyllactic acid than the Lactibacillus plantarum type strain KCTC 3108 when cultured in MRS medium. In one specific embodiment, when the Lactibacillus plantarum GBCC_F0227 strain of the present invention is cultured in MRS medium for 12 to 16 hours, the secretion amounts of α-ketoglutaric acid and 3-phenyllactic acid are each in the range of 1.5 to 2.5 times compared to the corresponding secretion amounts of the Lactibacillus plantarum KCTC 3108 strain inoculated and cultured under the same conditions. In a more specific embodiment, when the Lactobacillus plantarum GBCC_F0227 strain of the present invention is inoculated into an MRS medium in an amount such that the 600 nm absorbance is 0.01 to 0.02 and cultured for 16 hours, the secretion amounts of α-ketoglutaric acid and 3-phenyllactic acid are both in the range of 1.5 to 2.5 times that of the corresponding secretion amounts of the Lactobacillus plantarum KCTC 3108 strain inoculated and cultured under the same conditions. In the most specific embodiment, when the Lactibacillus plantarum GBCC_F0227 strain of the present invention is inoculated into MRS medium in an amount such that the 600 nm absorbance is 0.01 to 0.02 and cultured for 16 hours, the secretion amounts of α-ketoglutaric acid and 3-phenyllactic acid are both in the range of 1.95 to 2.05 times compared to the corresponding secretion amounts of the Lactibacillus plantarum KCTC 3108 strain inoculated and cultured under the same conditions.In a more specific embodiment, in addition to the above-described secretion characteristics, when the Lactobacillus plantarum GBCC_F0227 strain of the present invention is inoculated into an MRS medium in an amount such that the 600 nm absorbance is 0.01 to 0.02 and cultured for 16 hours, the secretion amount of 2-hydroxy-3-methylbutyric acid is in the range of 2.0 to 3.0 times that of the corresponding secretion amount of the Lactobacillus plantarum KCTC 3108 strain inoculated and cultured under the same conditions.

[0076] The GBCC_F0227 strain of the present invention, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has at least one of the effects of suppressing weight gain in a host, reducing body fat in a host, and suppressing appetite in a host.

[0077] In another embodiment, the GBCC_F0227 strain of the present invention is a Lactobacillus plantarum strain comprising a mutation of a naturally occurring Lactobacillus plantarum.

[0078] In another aspect of the present invention, a pharmaceutical composition for preventing or treating a metabolic disease is provided, which comprises at least one selected from the group consisting of Lactobacillus plantarum GBCC_F0227 strain, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, and a cell-free culture supernatant of the strain as an active ingredient.

[0079] In the pharmaceutical composition for preventing or treating a metabolic disease of the present invention, the metabolic disease is any one of obesity, obesity-induced anorexia, metabolic syndrome, hypertension, arteriosclerosis, hyperlipidemia, fatty liver, nonalcoholic fatty liver disease, hyperinsulinemia, diabetes, impaired glucose tolerance, hypercholesterolemia, hypertriglyceridemia, cardiovascular disease, and insulin resistance syndrome. More specifically, the metabolic disease is any one of obesity, obesity-induced anorexia, metabolic syndrome, diabetes, impaired glucose tolerance, nonalcoholic fatty liver disease, hypercholesterolemia, hypertriglyceridemia, and insulin resistance syndrome. In the most specific embodiment, the metabolic disease is obesity, metabolic syndrome impaired glucose tolerance, or diabetes.

[0080] The above Lactiplantibacillus plantarum GBCC_F0227 strain (Accession Number: KCTC 15450BP) is the same as described above. At this time, the strain may be a live cell or a dead cell, and is preferably a live cell. In addition, the culture of the strain may or may not contain the strain, and is preferably a culture containing the strain.

[0081] In one embodiment, the GBCC_F0227 strain of the present invention comprises a recA gene sequence set forth in SEQ ID NO: 2.

[0082] The pharmaceutical composition of the present invention may further include, in addition to the active ingredient, one or more additives selected from the group consisting of preservatives, dyes, emulsifiers, sweeteners, stabilizers, flavor enhancers, flavoring agents, and acidulants.

[0083] The pharmaceutical composition of the present invention may further comprise at least one pharmaceutically acceptable excipient and / or a lyophilizer.

[0084] “Pharmaceutically acceptable” means physiologically acceptable and does not typically cause severe gastrointestinal upset, dizziness, allergic reactions or similar reactions when administered to humans.

[0085] The pharmaceutical composition according to the present invention may further comprise at least one pharmaceutically acceptable excipient in addition to the novel lactic acid bacteria. Excipients that may be included in the composition of the present invention include, but are not limited to, lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methylcellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oil. The composition of the present invention can be formulated into a formulation for oral administration or a formulation for parenteral administration by a conventional method, and when formulated, it can be prepared using commonly used fillers, bulking agents, binders, wetting agents, disintegrants, surfactants, cryoprotectants, etc.

[0086] For example, a cryoprotectant selected from the group consisting of sucrose, maltose, maltodextrin, trehalose, mannitol, sorbitol, inulin, glycerol, DMSO, ethylene glycol, propylene glycol, 2-methyl-2,4-pentanediol, polyethylene glycol, polyvinylpyrrolidone, polyvinyl alcohol, polyglycerol, skim milk, milk protein, whey protein, betaine, adonitol, lactose or any combination thereof may be used.

[0087] Preferably, the cryoprotectant may include at least one selected from the group consisting of sucrose, skim milk, and sorbitol. More preferably, it may include sucrose, skim milk, and sorbitol, and specifically, it may include 2 to 20 wt% of sucrose, 2 to 20 wt% of sorbitol, and 5 to 30 wt% of skim milk based on the total weight of the composition. By adding the cryoprotectant, the freeze-dried lactic acid bacteria can exhibit significantly increased survival rate, storage stability, acid resistance, and bile resistance. In addition, the addition of an antioxidant such as riboflavin, riboflavin phosphate or a physiologically acceptable salt thereof, glutathione, ascorbate, and cysteine ​​to the freeze-dried composition according to the present invention can further increase the survival rate of the strain during storage.

[0088] When the composition of the present invention is formulated as a solid preparation for oral administration, it includes tablets, pills, powders, granules, capsules, etc., and such solid preparations may include at least one excipient, such as starch, calcium carbonate, sucrose, lactose, or gelatin, in addition to the active ingredient. In addition, in addition to simple excipients, lubricants such as magnesium stearate and talc may be included, but are not limited thereto.

[0089] The dosage of the pharmaceutical composition according to the present invention must be a pharmaceutically effective amount. "Pharmaceutically effective amount" means an amount sufficient to prevent or treat a neurological disease at a reasonable benefit / risk ratio applicable to medical treatment. The effective dosage level can be variously selected by those skilled in the art according to factors such as the formulation method, the patient's condition and body weight, the patient's sex, age, the extent of the disease, the drug form, the route and duration of administration, the rate of excretion, and the sensitivity of the reaction. As will be recognized by those skilled in the art, the effective amount may vary depending on the route of administration, the use of excipients, and the possibility of use with other drugs. However, for a desirable effect, in the case of oral administration, the composition of the present invention can generally be administered to adults at a daily dose of 0.001 to 1000 mg / kg of body weight per day, preferably 0.01 to 100 mg / kg.

[0090] When administering the dosage form as described above, the Lactobacillus plantarum GBCC_F0227 strain of the present invention is administered at 1×10 per day. 3 Colony forming units (CFU) / g to 1×10 16 It can be administered in CFU / g. For example, the strain of the present invention can be administered in 10 3 10 inland 16 CFU / g, 10 3 10 inland 15 CFU / g, 10 3 10 inland 14 CFU / g, 10 3 10 inland 13 CFU / g, 10 3 10 inland 12 CFU / g, 10 4 10 inland 16 CFU / g, 10 4 10 inland 15 CFU / g, 10 4 10 inland 14 CFU / g, 10 4 10 inland 13 CFU / g, 10 4 10 inland12 CFU / g, 10 5 10 inland 16 CFU / g, 10 5 10 inland 15 CFU / g, 10 5 10 inland 14 CFU / g, 10 5 10 inland 13 CFU / g, 10 5 10 inland 12 CFU / g, 10 6 10 inland 13 CFU / g, 10 6 10 inland 12 CFU / g, 10 7 10 inland 13 CFU / g, 10 7 10 inland 12 CFU / g, 10 8 10 inland 13 CFU / g or 10 8 10 inland 12 It can be administered at a dosage of 100 CFU / g. The daily dosage can be administered all at once or divided into several doses. The above dosage does not limit the scope of the present invention in any way.

[0091] When the composition of the present invention is formulated as a liquid preparation for oral administration, it includes suspensions, solutions, emulsions, syrups, etc., and in addition to commonly used simple diluents such as water and liquid paraffin, it may include various excipients such as wetting agents, sweeteners, fragrances, preservatives, etc., but is not limited thereto. When the composition of the present invention is formulated as a preparation for parenteral administration, it may include sterilized aqueous solutions, non-aqueous solvents, suspensions, emulsions, suppositories, etc. Non-aqueous solvents and suspensions may include, but are not limited to, propylene glycol, polyethylene glycol, vegetable oils such as olive oil, and injectable esters such as ethyl oleate. Base materials for suppositories may include witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin, etc. The content of the novel lactic acid bacteria, etc., which are the active ingredients of the pharmaceutical composition of the present invention, can be adjusted within a wide range depending on the specific form, purpose of use, and aspect of the composition. The content of the active ingredient in the pharmaceutical composition according to the present invention is not particularly limited, and may be, for example, 0.01 to 99 wt%, specifically 0.1 to 75 wt%, and more specifically 0.5 to 50 wt%, based on the total weight of the composition. The cryoprotectant used in the present invention is used to preserve the probiotic formulation during lyophilization and improve the shelf life. The cryoprotectant used in the present invention may include a common saccharide. The saccharide may be a monosaccharide, a disaccharide, an oligosaccharide, a polysaccharide, or a mixture of at least two or more saccharides.

[0092] The pharmaceutical composition according to the present invention can be administered to humans, mammals, and livestock, specifically monkeys, mice, rats, rabbits, sheep, cows, dogs, horses, pigs, etc., via various routes. Specifically, the pharmaceutical composition of the present invention can be administered orally or parenterally (e.g., by topical application or intravenous, subcutaneous, or intraperitoneal injection), but oral administration is preferred. Solid preparations for oral administration may include powders, granules, tablets, capsules, soft capsules, pills, etc.

[0093] The pharmaceutical composition of the present invention may be provided, in particular, as an enteric-coated preparation in the form of an oral unit dosage form. The term "enteric coating" as used herein includes all types of pharmaceutically acceptable known coatings that are not decomposed by gastric acid to maintain the coating, but are sufficiently decomposed in the small intestine to release the active ingredient therein. The "enteric coating" of the present invention refers to a coating that remains intact for at least 2 hours when brought into contact with an artificial gastric fluid, such as an HCl solution having a pH of 1, at 36°C to 38°C, and preferably, is subsequently decomposed within 30 minutes in an artificial intestinal fluid, such as a KH2PO4 buffer solution having a pH of 6.8.

[0094] In one embodiment of the pharmaceutical composition of the present invention, the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain contains at least one selected from the group consisting of α-ketoglutaric acid, 3-phenyllactic acid, cystathionine, 1-monopalmitin, glycerol monostearate, and 2-hydroxy-3-methylbutyric acid.

[0095] In the pharmaceutical composition of the present invention, the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has an effect of inhibiting DPP-4 enzyme activity.

[0096] In the pharmaceutical composition of the present invention, the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has an effect of stimulating GLP-1 secretion in the host. The pharmaceutical composition of the present invention has an effect of inhibiting DPP-4 enzyme activity.

[0097] In the pharmaceutical composition of the present invention, the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has an effect of promoting the expression of the GLP-1 receptor in the host. The pharmaceutical composition of the present invention has an effect of inhibiting DPP-4 enzyme activity.

[0098] In the pharmaceutical composition of the present invention, the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has an effect of alleviating skeletal muscle loss in a host accompanying obesity.

[0099] In the pharmaceutical composition of the present invention, the GBCC_F0227 strain, the lysate of the strain, the extract of the strain, the killed cells of the strain, the culture of the strain, the extract of the culture, or the cell-free culture supernatant of the strain has an effect of reducing visceral fat accumulation in the host. In particular, the GBCC_F0227 strain, the lysate of the strain, the extract of the strain, the killed cells of the strain, the culture of the strain, the extract of the culture, or the cell-free culture supernatant of the strain has an effect of reducing hepatic fat accumulation in the host.

[0100] In the pharmaceutical composition of the present invention, the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain has an effect of reducing blood neutral fat of the host.

[0101] In another aspect of the present invention, a pharmaceutical composition for treating or preventing a metabolic disease that increases GLP-1 secretion or suppresses appetite is provided, comprising as an active ingredient at least one selected from a strain of the species Lactibacillus plantarum, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, and a cell-free culture supernatant of the strain.

[0102] In one embodiment of the pharmaceutical composition for treating or preventing the metabolic disease, the strain of the Lactibacillus plantarum species, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain exhibits at least one of the effects of suppressing weight gain in the host, reducing body fat in the host, and suppressing appetite in the host.

[0103] In one specific embodiment of the pharmaceutical composition for treating or preventing the above metabolic disease, the strain of the Lactibacillus plantarum species may be a strain having an average nucleotide identity (ANI) of 69% or more to less than 100% with respect to the genome of Lactibacillus plantarum GBCC_F0227 (Accession No.: KCTC 15450BP). A strain of Lactibacillus plantarum species having an ANI value of 69% or more and less than 100% of the genome of Lactibacillus plantarum GBCC_F0227 is advantageous in enhancing the GLP-1 action of the host, as it can induce host GLP-1 secretion to a level similar to GBCC-F0227. In a more specific embodiment of the pharmaceutical composition for treating or preventing the metabolic disease, the strain of Lactibacillus plantarum species is a strain having a genome in the ANI range of 69% or more and less than 100% of the genome of Lactibacillus plantarum GBCC_F0227, and the strain of Lactibacillus plantarum species, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain comprises α-ketoglutaric acid, Contains at least one selected from the group consisting of 3-phenyllactic acid, cystathionine, 1-monopalmitin, glycerol monostearate and 2-hydroxy-3-methylbutyric acid.

[0104] In another aspect of the present invention, a food composition comprising the Lactibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or a culture thereof is provided. The Lactibacillus plantarum GBCC_F0227 is the same as described above. The food composition includes all forms such as functional food, nutritional supplement, health food, and food additives, and the food composition of the above type can be prepared in various forms according to conventional methods known in the art. In addition, the "food composition" in the present invention is used to mean not only the food itself as an example of the food composition described above, but also the "food additive" or "food additive composition" added to such food. When the strain is used as a food additive, the strain can be added as is or used together with other foods or food ingredients, and can be used appropriately according to conventional methods. The amount of the active ingredient can be appropriately determined depending on the intended use (prevention, health, or therapeutic treatment). Generally, when manufacturing a food or beverage, the strain can be added in an amount of 0.0001 wt% to 1 wt%, specifically 0.001 wt% to 0.1 wt%, of the raw material composition containing the strain. However, in the case of long-term intake for the purpose of health and hygiene or health regulation, the amount can be used even below the above range. The food composition of the present invention can be used for the prevention or suppression of metabolic diseases.

[0105] In another aspect of the present invention, a health functional food is provided, which comprises as an active ingredient at least one selected from the group consisting of Lactobacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP), a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, and a cell-free culture supernatant of the strain. The health functional food may be a health functional food for preventing or improving obesity.

[0106] In this specification, "health functional food" refers to a food manufactured or processed using a specific ingredient as a raw material or a specific ingredient contained in a food raw material by methods such as extraction, concentration, purification, or mixing for the purpose of health supplementation, and refers to a food designed and processed so that the above-mentioned ingredient can sufficiently exert biological regulation functions such as biological defense, biological rhythm regulation, disease prevention, and recovery on the body, and refers to a food that can perform functions related to disease prevention or health recovery.

[0107] When the Lactibacillus plantarum GBCC_F0227 strain according to the present invention is used as a health functional food, it can be added as is or used together with other foods or food ingredients, and these can be selected and used appropriately as needed. In addition, the mixed amount of the added GBCC_F0227 strain can be appropriately determined depending on the intended use. The formulation of the health functional food comprising the Lactibacillus plantarum GBCC_F0227 strain according to the present invention is not particularly limited, and can be in the form of powder, granules, pills, tablets, capsules, as well as any form of general food or beverage. There are no specific restrictions on the types of these foods, and examples of foods to which the above substances can be added include meat, sausage, bread, chocolate, candy, snacks, confectionery, pizza, ramen, other noodles, gum, dairy products including ice cream, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes, and encompass foods in the general sense.

[0108] As described above, the GBCC_F0227 strain of the food composition or the GBCC_F0227 strain used in the culture thereof is a strain in which, when cultured in MRS medium for 12 to 16 hours, the secretion amount of α-ketoglutaric acid and 3-phenyllactic acid is 1.5 to 2.5 times higher than the corresponding secretion amount of the Lactobacillus plantarum KCTC 3108 strain inoculated and cultured under the same conditions. In a more specific embodiment, the GBCC_F0227 strain of the food composition of the present invention or the GBCC_F0227 strain used in the culture thereof is a strain in which, when inoculated into an MRS medium in an amount such that the 600 nm absorbance is 0.01 to 0.02 and cultured for 16 hours, the secretion amounts of α-ketoglutaric acid and 3-phenyllactic acid are both 1.5 to 2.5 times higher than the corresponding secretion amounts of the Lactobacillus plantarum KCTC 3108 strain inoculated and cultured under the same conditions. In the most specific embodiment, the GBCC_F0227 strain of the food composition of the present invention or the GBCC_F0227 strain used in the culture thereof is inoculated into MRS medium in an amount such that the 600 nm absorbance is 0.01 to 0.02 and cultured for 16 hours, and the secretion amounts of α-ketoglutaric acid and 3-phenyllactic acid are both in the range of 1.95 to 2.05 times compared to the corresponding secretion amounts of Lactobacillus plantarum KCTC 3108 strain inoculated and cultured under the same conditions. In a more specific embodiment, the GBCC_F0227 strain of the food composition of the present invention or the GBCC_F0227 strain used in the culture thereof, in addition to the above-described secretion characteristics, when inoculated into MRS medium in an amount such that the 600 nm absorbance is 0.01 to 0.02 and cultured for 16 hours, the secretion amount of 2-hydroxy-3-methylbutyric acid is in the range of 2.0 to 3.0 times that of the corresponding secretion amount of Lactobacillus plantarum KCTC 3108 strain inoculated and cultured under the same conditions.

[0109] Another aspect of the present invention provides a feed composition comprising the Lactibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or a culture thereof. The Lactibacillus plantarum strain (Accession No.: KCTC 15450BP) is the same as described above. In the present invention, the feed composition means not only the feed itself but also a feed additive (feed additive composition). The feed composition for preventing or improving the metabolic disease can be prepared by adding the GBCC_F0227 strain in an appropriate effective concentration range according to various feed preparation methods known in the art. The feed composition of the present invention may be used for preventing or suppressing metabolic diseases in livestock.

[0110] Another aspect of the present invention provides a method for preventing and treating metabolic diseases, comprising administering to a subject a Lactibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or a culture thereof. The subject may be a subject having a metabolic disease. In addition, the subject may be a mammal, preferably a human. In this case, the Lactibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) is the same as described above. In addition, the route of administration, dosage, and frequency of administration of the strain or the culture thereof may be administered to a subject in various ways and amounts depending on the condition of the patient and the presence or absence of side effects, and an ordinary skilled person may select the optimal administration method, dosage, and frequency of administration within an appropriate range. In addition, the types of metabolic diseases are as described above.

[0111] In another aspect of the present invention, there is provided a use of Lactiplantibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) or a culture thereof for treating metabolic diseases. In this case, the Lactiplantibacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP) is the same as described above. In addition, the type of metabolic disease is the same as described above.

[0112] Hereinafter, the present invention will be described in more detail through the following examples and experimental examples. However, these examples and experimental examples are intended only to aid understanding of the present invention and are not intended to limit the scope of the present invention in any way. Various changes and modifications may be made to the present examples, and such changes and modifications also fall within the scope of the appended claims.

[0113] Manufacturing example: Isolation and identification of lactic acid bacteria from a source

[0114] Candidate lactic acid bacteria strains were selected from human fecal samples and fermented foods. The fermented foods used as bacterial sources were sauerkraut (German pickled cabbage), cabbage kimchi, green onion kimchi, young radish kimchi, ulgal kimchi, and white kimchi. Samples were collected from sauerkraut (9, 12, and 13 days), cabbage kimchi (3 days), young radish kimchi (1, 2, 10, and 23 days), white kimchi (5 days), and ulgal kimchi (3 days) after pickling for a specified period of time. Samples collected from human feces and fermented foods were ground in phosphate buffered saline (PBS), serially diluted by one-tenth, and plated on MRS (deMan, Rogosa, Sharpe) plates, a lactic acid bacteria culture medium. These media were cultured in an anaerobic chamber at 37°C for more than two days, and bacterial colonies were observed. A single colony was subcultured onto a new MRS plate medium for pure isolation. The pure strain was identified by PCR and sequencing of the 16S rRNA gene. For long-term storage of the strain, 30% (v / v) glycerol was added to the culture broth that reached the end of the exponential growth phase and stored in a -70°C ultra-low temperature freezer. Cultivation after pure isolation and identification was also performed on MRS plate medium and liquid medium.

[0115] Identification of the strain was based on the sequence similarity of the 16S rRNA gene and recA gene. DNA was extracted from the bacterial culture using the FastDNA® SPIN Kit for soil (MP Bio, 6560200). Polymerase chain reaction (PCR) was performed on the extracted DNA to obtain genetic information for the identification of the bacteria. For 16S rDNA PCR, forward primer 27F (5'-AGAGT TTGAT CMTGG CTCAG-3') and reverse primer 1492R (5'-TACGG YTACC TTGTT ACGAC TT-3') were used, and for recA PCR, forward primer L. p_recA_F (5'-CTGAT GCACG GAAAG CAGC-3') and reverse primer L. p_recA_R (5'-CGCTT AAATG GYGGY GCMAC C-3') were used.

[0116] As a result of analyzing the strain species using the bacterial genetic information obtained in this way, 324 strains of Lactiplantibacillus and Bifidobacterium were identified. Specifically, 205 strains of Lactiplantibacillus plantarum, 37 strains of Lactiplantibacillus paracesei, 45 other strains of Lactiplantibacillus, and 45 strains of Bifidobacterium were isolated.

[0117] The GLP-1 secretion effect and DPP-4 activity inhibition effect were measured for these isolated strains to select superior strains.

[0118] [Example 1]

[0119] Glucagon-like peptide-1 (GLP-1) secretion stimulation effect

[0120] To select strains that stimulate intestinal GLP-1 secretion among the aforementioned isolated lactic acid bacteria strains, the GLP-1 secretion stimulation properties of the culture supernatants of the isolated strains were evaluated. 114 of the isolated strains were anaerobically cultured in MRS broth medium at 37°C for 24 h, then centrifuged to precipitate the strains, and only the supernatant was collected. The culture supernatant was filtered through a 0.22 μm syringe filter to remove cells, and used in the experiment.

[0121] NCI-H716 cell line (Korea Cell Line Bank (KCLB) No. 10251), a human enteroendocrine cell line known to secrete GLP-1, was seeded at 4 × 10 cells per well of a 96-well plate. 4 After overnight inoculation with a dog, the cells were washed three times with DPBS (Dulbecco's phosphate-buffered saline) containing 0.2% bovine serum albumin (BSA) to remove medium components and cultured for 2 hours to adjust the level of GLP-1 secretion between cells. Afterwards, the culture supernatants of various isolated strains were diluted to a concentration of 20% v / v in DPBS containing 0.2% BSA and treated with the cells for 2 hours. The GLP-1 concentration of each strain was measured for the supernatant collected after the reaction using a GLP-1 ELISA kit (Human GLP-1 EIA, Raybiotech).

[0122] The results of the GLP-1 secretion stimulation performance of the 114 strains of lactic acid bacteria isolated in this way are shown in Figs. 1a to 1g. As can be clearly seen in Figs. 1a to 1g, among the isolated lactic acid bacteria strains, the GLP-1 concentration in the culture supernatant of the L. plantarum strain (Fig. 1a) was significantly higher than that of L. paracasei (Fig. 1b) and Bifidobacterium strains (Fig. 1g). In addition, it was confirmed that the GBCC_F0227 strain isolated from sauerkraut among the L. plantarum strains had the highest GLP-1 secretion stimulation effect.

[0123] [Example 2]

[0124] Comparison of strain-specific GLP-1 secretion stimulation effects

[0125] To determine whether the intestinal GLP-1 secretion stimulation effect of the L. plantarum GBCC_F0227 strain isolated from sauerkraut is strain-specific, the GLP-1 secretion stimulation effect of the culture supernatants of the KCTC3108 strain, a type strain of the same L. plantarum provided by the Korea Center for Biological Resources (KCTC), L. plantarum GBCC_F0053 isolated from radish kimchi, and L. plantarum GBCC_F0058 strain isolated from radish kimchi was compared with that of GBCC_F0227 using the same method as in Example 1.

[0126] As shown in Fig. 2a, the standard strain KCTC3108 of the same Plantarum species as the L. plantarum GBCC_F0227 strain, as well as the Plantarum isolates GBCC_F0053 and GBCC_F0058 isolated from other fungal sources other than sauerkraut, exhibited GLP-1 secretion-stimulating efficacy significantly lower than that of GBCC_F0227. This confirms that the excellent GLP-1 secretion-stimulating efficacy of the GBCC_F0227 strain is specific to this strain and is not common to all Plantarum species.

[0127] In order to compare the GLP-1 secretion stimulation efficacy of the isolated strains of Plantarum species, including the F0227 strain, with the known Plantarum strains, a comparative experiment was conducted with the known Lactibacillus plantarum strain GB104 (Korea Research Institute of Bioscience and Biotechnology Deposit No. KCTC15450BP) disclosed in Patent Publication No. 2021-0086540 in the same manner as in Example 1. The experimental results are shown in Fig. 2b. Among the isolated strains tested, nine strains, including GBCC_F0227, exhibited better GLP-1 secretion stimulation efficacy than the known Plantarum strain GB104.

[0128] [Example 3]

[0129] Comparison of the individual-level GLP-1 secretion stimulation effects of F0227

[0130] We evaluated whether the F0227 strain had the GLP-1 secretion stimulation effect at the individual level. C57BL / 6 mice (Orient Bio) were divided into positive control group (normal diet without strain administration), negative control group (high-fat diet with 60% fat composition of calories without strain administration), and negative control group (high-fat diet with 5×10 strains daily) 9 The mice were divided into three groups and administered GBCC_F0227 strain with colony forming units (CFU) for 8 weeks, and then autopsied to examine the GLP-1 concentration in the blood. In this case, the mice in the high-fat diet group were injected with 5×10 5 hours before autopsy. 9Colony forming units (CFU) of L. plantarum GBCC_F0227 were administered orally. Blood was obtained through cardiac puncture, and the obtained blood was immediately transferred to a tube containing EDTA containing 500 KIU of aprotinin and 1 μg / mL of diprotin A to prevent biodegradation of GLP-1, and stored on ice. The obtained blood was centrifuged to separate plasma, and the GLP-1 level in the plasma was measured using a GLP-1 ELISA kit (GLP-1 EIA, Raybiotech) to confirm the effect of the strain on increasing GLP-1 concentration. As a result, it was confirmed that the experimental group (G3_F0227 in Figure 3) administered GBCC_F0227 together with a high-fat diet increased the blood GLP-1 concentration to a significantly higher level than the positive control group (G1_NCD in Figure 3) and the negative control group (G2_HFD in Figure 3).

[0131] [Example 4]

[0132] Inhibitory effect of isolated strains on dipeptidyl peptidase-4 (DPP-4) activity

[0133] DPP-4 enzyme is known to degrade GLP-1 secreted from enteroendocrine cells (L cells) within 5 minutes, so inhibition of this enzyme is considered beneficial for the maintenance and action of GLP-1. To confirm whether the culture supernatant of the strains isolated above had the ability to inhibit DPP-4 activity, an enzyme inhibition experiment was conducted as follows. Gly-Pro-p-nitroanilide (0.8 mM), a substrate of DPP-4 enzyme, was diluted with phosphate buffer and cultured in the culture supernatant (5% v / v) of the isolated lactic acid bacteria strain or in MRS medium (5%) as a culture medium control at 37°C for 10 minutes, and then DPP-4 enzyme (0.001 enzyme unit) was added thereto and the enzyme and substrate were reacted at 37°C for 60 minutes. After the reaction was terminated by adding sodium acetate buffer (1 M, pH 4.0), the decrease in absorbance due to substrate degradation was measured at a wavelength of 405 nm using a microplate reader. The degree of DPP-4 enzyme inhibition of the isolated lactic acid bacteria culture supernatant was evaluated and compared, and the results are shown in Fig. 4a. In the experiment shown in Fig. 4a, the Lactobacillus plantarum GBCC_F0227 strain also had the highest DPP-4 enzyme inhibition ability among the isolated strains.

[0134] Figure 4b is a graph comparing the DPP-4 activity inhibition efficacy of selected Lactibacillus plantarum strains with that of the known plantarum strain GB104. The comparative experiment shown in Figure 4b was performed in the same manner as in Figure 4a. In a comparative analysis where the DPP-4 activity inhibition efficacy of GB104 was set at 100%, GBCC_F0227 showed the highest inhibition efficacy among the isolated strains, with an inhibition efficacy of 117.92%.

[0135] Comparison of inhibitory efficacy with other lactic acid bacteria species

[0136] The DPP-4 inhibitory efficacy of F0227 was compared with that of non-plantarum strains of lactic acid bacteria and known plantarum strains. The tested strains were L. plantarum GBCC_F0227, Limosilactobacillus fermentum (GB102) disclosed in Patent Publication No. 2021-0086537, the known Lacticaseibacillus rhamnosus GG (LGG), Latilactobacillus curvatus (GBCC_F0007), Ligilactobacillus acidipiscis (GBCC_F0023), Latilactobacillus sakei (GBCC_F0025), Lactiplantibacillus mudanjiangensis (GBCC_F0172), Lacticaseibacillus porcinae (GBCC_F0339), Lentilactobacillus kefiri (GBCC_F0351), Loigolactobacillus coryniformis (GBCC_F0257), Schleiferilactobacillus isolated from food by the present applicant harbinensis (GBCC_F0372) was compared. For the DPP-4 activity inhibition test, all strains were anaerobically cultured for 16 hours under the same conditions, and the culture supernatant obtained was diluted to 5% in phosphate buffered saline (PBS) and the DPP-4 activity inhibition efficacy was tested as described above. As a result of the experiment shown in Fig. 4b, GBCC_F0227 was the most effective with an absolute activity inhibition rate of approximately 60%, LGG showed approximately 40%, and all other strains showed inhibition rates of around 30%, confirming that L. plantarum GBCC_F0227 had the best DPP4 activity inhibition efficacy.

[0137] Summary of GLP-1 secretion stimulation and DPP-4 inhibition effects of plantarum isolates

[0138] Figure 5 is a graph showing the correlation between the GLP-1 secretion stimulation efficacy and DPP-4 activity inhibition efficacy of GBCC_F0227 and other selected Plantarum isolates. Figure 5 confirms that GBCC_F0227 is superior in the comprehensive efficacy evaluation of GLP-1 and DPP-4 among the selected lactic acid bacteria isolates.

[0139] [Example 5]

[0140] GBCC_F0227 Gene and Genome Analysis

[0141] The full-length 16S rDNA and recA gene sequences of GBCC_F0227 are represented by the nucleotide sequences 1 and 2, respectively. GBCC_F0227 showed a sequence similarity of over 98.5% with 11 standard strains of the genus Lactiplantibacillus, including Lactiplantibacillus plantarum, based on the 16S rRNA sequence, confirming that GBCC_F0227 corresponds to one of these 11 strains. The results of the 16S rDNA similarity calculation are summarized in Table 1.

[0142]

[0143] For more accurate species identification, similarity analysis was performed based on the sequence of the recA gene, which is widely used as a taxonomic biomarker for Lactobacillus bacteria. The results confirmed that GBCC_F0227 is the closest strain to Lactiplantibacillus plantarum. The identification of GBCC_F0227 strain as a Lactiplantibacillus plantarum strain was also supported by a comparison of the average nucleotide identity (ANI) values ​​of the genome (the cutoff threshold was 95, and the ANI was 98.78 among Lactiplantibacillus plantarum species).

[0144]

[0145] Genome-based phylogenetic analysis to confirm the intraspecific position of GBCC_F0227

[0146] To determine the evolutionary relationships of GBCC_F0227 within the Lactiplantibacillus plantarum species to which it belongs, a genome-based phylogenetic analysis was performed. The GBCC_F0227 genome sequence was compared with the complete genome sequences of 82 strains of Lactiplantibacillus plantarum species published in GenBank and one strain of Lactiplantibacillus argentoratensis, a closely related species. Lactiplantibacillus paraplantarum DSM 10667 was used as an outgroup for the genome-based phylogenetic tree. T The fully sequenced genomes of the strains were used. Ortholog clustering of protein-coding genes present in the strain genomes was performed using the orthoMCL program. 1,714 genes were selected, which were present in all 85 strain genomes, including the outgroup, and each strain had only one gene in its genome. The amino acid sequences of the selected genes were aligned using the MUSCLE alignment algorithm, and a phylogenetic tree was constructed using the RAxML program based on the aligned amino acid sequences. The constructed phylogenetic tree is shown in Figure 1h. This phylogenetic tree shows that strain GBCC_F0227 is a member of the Lactobacillus plantarum reference strain DSM 20174. T Or, it indicates that the strain is of a different lineage from the existing patented strains, ATG-K2, ATG-K6, and ATG-K8.

[0147] [Example 6]

[0148] Anti-obesity effect of F0227

[0149] The anti-obesity efficacy of L. plantarum GBCC_F0227 strain was confirmed using a high-fat diet mouse model. Specifically, C57BL / 6 mice were fed a 60% high-fat diet (HFD) and simultaneously orally administered GBCC_F0227 and the L. plantarum standard strain KCTC3108 daily, and these mice were set as the experimental group. In addition, mice fed only the 60% high-fat diet were set as the negative control group, and mice fed a normal chow diet (NCD) were set as the positive control group. At this time, GBCC_F0227 and KCTC3108 lactic acid bacteria were administered to each mouse at a concentration of 5 × 10 9 CFU was administered orally daily for 8 weeks, and the anti-obesity effect was confirmed by comparing the body weight gain of the negative control group and the experimental group. As shown in the experimental results in Fig. 6a, when comparing the body weight gain at week 8, it was confirmed that the mouse group administered the L. plantarum GBCC_F0227 strain (HFD+F0227 in Fig. 6a) had a very significantly lower body weight gain than the KCTC3108 administration group (HFD+KCTC3108 in Fig. 6a) and the negative control group fed only a high-fat diet (HFD in Fig. 6a).

[0150] Additionally, in the 8th week of the experiment, the body fat mass and lean mass of the mice were measured using a body composition analyzer (LF50 Body composition mice analyzer), and the ratio of body fat to lean mass (Fat / Lean) was compared. As shown in Fig. 6b, it was confirmed that the mice administered with GBCC_F0227 lactic acid bacteria (G3 GBCC_F0227 in Fig. 6b) showed a very significant decrease in body fat accumulation compared to the negative control mice (G2 HFD in Fig. 6b).

[0151] [Example 7]

[0152] F0227's effect on improving blood sugar control

[0153] The effect of L. plantarum GBCC_F0227 strain on improving glucose tolerance was confirmed through an oral glucose tolerance test (OGTT) using a high-fat diet-induced obese mouse model. C57BL / 6 mice (Orient Bio) were fed a 60% high-fat diet for 8 weeks. One week before the end of the animal model experiment, the experimental mice were fasted for 12 hours, their body weights were measured, and blood was collected from the tail vein to measure blood glucose levels at fasting conditions immediately before administration and at 30, 60, 90, and 120 minutes after oral administration of 2 g / kg glucose. In addition, a blood glucose change trend graph was drawn with time (minutes) on the horizontal axis based on the measured values, and the area under the OGTT curve (AUC) was calculated by multiplying the blood glucose value and time on the graph.

[0154] The results of the oral glucose tolerance test are shown in Fig. 7a. The group of mice administered L. plantarum GBCC_F0227 together with a high-fat diet (HFD+F0227 in Fig. 7a) showed a significantly lower peak blood glucose level 30 minutes after oral administration of 2 g / kg glucose compared to the negative control group of the high-fat diet (HFD in Fig. 7a). The fasting blood glucose measurement values ​​shown in Fig. 7b also confirmed that the GBCC_F0227-administered group (G3 GBCC_F0227 in Fig. 7b) had a significantly lower fasting blood glucose level than the negative control group of the high-fat diet (HFD in Fig. 7b). The AUC behavior shown in Fig. 7c also showed that the GBCC_F0227-administered group mice showed a statistically significant decrease in the AUC value compared to the negative control group. This suggests that L. PlantarumGBCC_F0227 was confirmed to have a positive effect in reducing fasting blood sugar levels and improving glucose tolerance.

[0155] Although the present invention has been described with reference to limited embodiments as described above, the technical idea and scope of the present invention are not limited to these aforementioned embodiments, and the invention of the patent claims described below, as well as various modifications or variations of the invention described in the patent claims that are obvious to a person having ordinary skill in the art to which the present invention pertains, are also included in the scope of the present invention within the equivalent scope of the invention described in the patent claims.

[0156] [Example 8]

[0157] Metabolite analysis of F0227

[0158] In this example, the metabolites secreted extracellularly by the L. plantarum GBCC_F0227 strain were compared with those of other lactic acid bacteria strains. As other lactic acid bacteria strains for comparison of metabolites, the metabolites of L. platanrum strains (F0053 isolated from green onion kimchi, KCTC 3108, a commercially available probiotic lactic acid bacteria), Limosilactobacillus fermentum strains (MG901 and KCTC 3112, commercially available probiotic lactic acid bacteria), Lactobacillus gasseri strain (KCTC 3163, a commercially available probiotic lactic acid bacteria), and Lactobacillus paragasseri strain (strain M0213 isolated herein) were quantified by gas chromatography-mass spectrometry (GC-MS).

[0159] Specifically, a single colony of each strain obtained by culturing the lactic acid bacteria strain on MRS agar medium was inoculated into MRS medium and seed cultured for 24 hours. Then, the seed culture was inoculated into 50 mL of new MRS medium in an amount that gave an absorbance (600 nm) value of 0.01 or 0.02 and cultured at 37°C for 16 hours. Then, the culture solution was centrifuged at 8,000 rpm for 5 minutes to obtain a culture supernatant, which was then filtered through a 0.22 μm syringe filter and stored in the refrigerator until test analysis.

[0160] 200 μL of each strain culture supernatant obtained in this way was extracted with cold methanol for 10 minutes in a mixer mill (MM400, Retch, Germany) at a frequency of 30 Hz, followed by sonication for 10 minutes. Then, the layers were separated using a centrifuge (centrifuge 5427R, Eppendorf, Germany) at 12,000 rpm and 4°C for 10 minutes, and the supernatant was filtered through a 0.2 μm polytetrafluoroethylene (PTFE) filter (Korea Chrome Disk) and evaporated using a high-speed vacuum concentrator (Korea Lavex). Then, each dried sample was redissolved in 80% methanol to obtain a methanol redissolution with a final concentration of 20,000 ppm. 100 μL of each methanol redissolution was dried using a high-speed vacuum concentrator and derivatized using the following method. First, 50 μL of methoxyamine hydrochloride (20 mg / mL in pyridine) was added to the dried samples and incubated in a thermomixer at 30°C for 90 min. Then, 50 μL of N-methyl-N-(trimethylsilyl)trifluoroacetamide (MSTFA) was added and incubated at 37°C for 30 min to convert them into silylated derivatives. 1 μL of the silylated derivatives from each sample was injected into a CHroZen GC system (Youngin Chromass, Anyang, Korea) equipped with an L-PAL3 autosampler system and a YL9600 mass scan spectrometer. The metabolites were separated on a DB-5MS column (30 m length × 0.25 mm inner diameter × 0.25 μm particle size; Agilent, Santa Clara, CA, USA) using helium as the carrier gas at a flow rate of 1.0 mL / min. The GC oven temperature was maintained at 60°C for 3 minutes, then increased by 10°C per minute to 320°C, and this final temperature was maintained for an additional 3 minutes. The mass spectrometry data acquisition rate was set to 2750 amu / s over a scan range of 50–600 amu, and then set to electron ionization (EI) mode at -70 eV. The ion source and transfer line temperatures were set to 230°C and 250°C, respectively.

[0161] Figure 8 is a table that organizes a heatmap of the amount of metabolites for each strain obtained by GC-MS, color-coded with relative numerical ratios. The values ​​shown in Figure 8 represent the ratio of the secretion amount to the average value (chromatogram peak area) of all strain samples tested for each metabolite.

[0162] The L. plantarum GBCC_F0227 strain exhibited mycological characteristics in that it secreted large amounts of organic acids such as α-ketoglutaric acid and 3-phenyllactic acid as metabolites, the amino acid cystathionine, and lipids and fatty acids such as 1-monopalmitin, glycerol monostearate, and the short-chain fatty acid 2-hydroxy-3-methylbutyric acid. Among these, the secretion of α-ketoglutaric acid and 3-phenyllactic acid, metabolites that may play a beneficial role in preventing weight gain and fat metabolism in the host, was prominent, and the secretion amount of these metabolites was 1.95 to 2.05 times that of KCTC 3108, the standard strain of L. plantarum. In addition, the secretion of 2-hydroxy-3-methylbutyric acid was 2.0 to 3.0 times that of KCTC 3108.

[0163] [Example 9]

[0164] GLP-1 secretion stimulation effect of metabolites of F0227

[0165] In this example, we confirmed in vitro whether the characteristic metabolites secreted by L. plantarum GBCC_F0227 strain into the extracellular space have the ability to stimulate GLP-1 secretion themselves. In order to compare the stimulating effects of GBCC_F0227 metabolites with those of other lactic acid bacteria strains, including KCTC 3108, a standard strain of L. plantarum, we examined the effects of seven metabolites: α-ketoglutaric acid, 3-phenyllactic acid, and 2-hydroxy-3-methylbutyric acid, which are secreted in large quantities by L. plantarum GBCC_F0227 strain, and short chain fatty acids (SCFAs) known to stimulate GLP-1 secretion: acetic acid, propionic acid, butyric acid, and lactic acid.

[0166] Similar to Example 1, NCI-H716 cell line (KCLB No. 10251) was seeded at 4 × 10 cells per well of a 96-well plate. 4 After overnight incubation after inoculation with a dog, the cells were washed three times with DPBS containing 0.2% bovine serum albumin to remove the medium components and incubated for 2 hours to adjust the level of GLP-1 secretion between cells. Afterwards, the seven substances mentioned above were diluted to concentrations of 5 mM, 10 mM, 15 mM, 20 mM, or 25 mM in DPBS containing 0.2% BSA and treated with NCI-H716 cells for 2 hours. The GLP-1 concentration in the supernatant collected after the reaction was measured using a GLP-1 ELISA kit (Human GLP-1 EIA, Raybiotech).

[0167] Figures 9a and 9b show the results of the GLP-1 secretion stimulation performance experiment. As shown in Figure 9a, all of the seven metabolites had the effect of stimulating the GLP-1 secretion of NCI-H716 in a concentration-dependent manner. In the case of α-ketoglutaric acid and 3-phenyllactic acid, which are abundantly secreted metabolites of GBCC_F0227, their secretion efficacy was overwhelmingly superior to that of short-chain fatty acids, and even lactic acid, which had the highest secretion efficacy among short-chain fatty acids, was found to have a significantly inferior secretion efficacy to 2-hydroxy-3-methylbutyric acid, a metabolite of GBCC_F0227. For example, in the stimulation results at 25 mM metabolite concentrations shown in Fig. 9b, it was confirmed that α-ketoglutaric acid had a GLP-1 secretion stimulation effect that was about 46 times superior to butyric acid, 3-phenyllactic acid had a GLP-1 secretion stimulation effect that was about 36 times superior to butyric acid, and 2-hydroxy-3-methylbutyric acid had a GLP-1 secretion stimulation effect that was about 12 times superior to butyric acid. In addition, it was confirmed that 3-phenyllactic acid had a GLP-1 secretion stimulation effect that was about 5 times superior to lactic acid, a common metabolite of lactic acid bacteria, and 2-hydroxy-3-methylbutyric acid had a GLP-1 secretion stimulation effect that was about 12 times superior to butyric acid. These results indicate that the excellent human GLP-1 secretion stimulation effect of the F0227 strain of the present invention was supported by the metabolites of GBCC_F0227.

[0168] [Example 10]

[0169] F0227's effect on reducing visceral fat and blood lipids

[0170] In this example, it was confirmed that the L. plantarumGBCC_F0227 strain has the effect of reducing white visceral fat fraction and blood triglyceride and cholesterol levels in a mouse model induced with high-fat diet obesity. Specifically, 5-week-old C57BL / 6 mice (Orient Bio, Korea) were acclimatized for 1 week, and then divided into three groups and orally administered the following feeds daily for 8 weeks: 5×10 per mouse daily with a high-fat diet (D12492, Research Diets, USA) with a fat ratio of 60 kcal% 9 The experimental group (G3) administered CFU of L. plantarumGBCC_F0227 strain in parallel, the obese positive control group (G2) administered cryoprotectant at the same mass as the strain administered in G3 along with the same high-fat diet, and the obese negative control group (G1) administered cryoprotectant along with a general feed diet (TEKLAD 2018S, Inotiv, USA).

[0171] After 8 weeks, the left lateral lobe of the liver and epididymal fat of the experimental mice were excised through necropsy and fixed in 10% neutral buffered formalin for 3 days. Then, they were dehydrated in ethanol, infiltrated with xylene, and paraffin-embedded using a tissue processor. Paraffin sections of 5 μm thickness were mounted on slides and visualized by hematoxylin-eosin staining. Histological quantification of the fat fraction in the liver and epididymal fat was performed using the ImageJ program. For the left lateral lobe of the liver, the area fraction occupied by lipid droplets in the tissue (Area fraction %) was measured, and for the epididymal fat, the size of lipid droplets accumulated in adipocytes (μm) was measured. 2 ) were quantified and the average of the values ​​obtained from three different parts per individual was used to plot the graph.

[0172] Blood analysis of mice was performed by obtaining blood through cardiac puncture at autopsy, separating serum through centrifugation, and measuring total cholesterol and triglyceride (neutral fat) levels in the blood using an automatic biochemical analyzer (clinical chemistry analyzer, DRI-CHEM NX700, Fujifilm, Japan).

[0173] As shown in Fig. 10a, the left lobe liver tissue fragments of high-fat diet mice (G3) administered L. plantarum GBCC_F0227 showed little difference in morphology and staining patterns in white fat compared to those of the obese negative control group (G1), but the difference in observed white fat was more pronounced compared to the obese positive control group (G2) induced by high-fat diet without strain administration. The strain-administered group (G3) also showed improved fat accumulation in epididymal adipose tissue compared to the obese positive control group (G2).

[0174] The area fraction occupied by fat droplets within adipocytes in liver tissue and the average size (area) of fat droplets in epididymal adipose tissue also supported these microscopic observations. As shown in Figure 10b, GBCC_F0227 administration significantly suppressed the increase in fat fraction induced by a high-fat diet, as measured by area fraction and adipocyte size.

[0175] As another method of observing the amount of fat in visceral adipose tissue, oil red O, a reagent specific for neutral fat and fatty acids, was used to stain and visualize fat in the left lobe tissue sections collected from the three groups above. Fat appears red in the oil red O staining, and tissue staining photographs of G1, G2, and G3 are shown in Fig. 10c. As shown in the staining results in Fig. 10c, the liver of the normal diet group (G1) was stained close to purple, but the liver of the high-fat diet group (G2) was stained bright red, indicating fat accumulation in the liver tissue due to the high-fat diet. In the group (G3) that was administered a high-fat diet and the strain in combination, the liver tissue was stained a lighter red than the high-fat diet group. This tendency regarding fat accumulation in liver tissue can be better understood by comparing the staining results by quantifying them. In the graph of Fig. 10d, which compares the staining intensity of Fig. 10c relative to the staining intensity value of the control group (G1), the staining intensity of the strain administration group (G3) was statistically significantly reduced compared to the high-fat diet group (G2).

[0176] These results indicate that fat accumulation in the left lobe of the liver caused by a high-fat diet can be alleviated by concurrent administration of a high-fat diet and the GBCC_F0227 strain.

[0177] In addition, when comparing the total cholesterol and triglyceride levels in the blood, the GBCC_F0227 strain of the present invention had the effect of reducing the extent of the increase in total cholesterol due to a high-fat diet (G3). In addition, when the GBCC_F0227 strain was administered in combination with a high-fat diet, the effect of statistically significantly reducing the increase in triglycerides in the blood that occurred when only a high-fat diet was administered (G2) was shown, and the blood triglyceride content of the parallel administration group (G3) was lowered to below the level of the normal diet group (Fig. 10e).

[0178] Therefore, when the GBCC_F0227 strain of the present invention was administered to a high-fat diet-induced obese mouse model, it was confirmed that it had not only an anti-obesity effect of reducing body weight but also an effect of lowering blood triglyceride and cholesterol levels, thereby confirming that the GBCC_F0227 strain has a positive effect on indicators of various metabolic diseases, including obesity.

[0179] [Example 11]

[0180] Strain-specific neutral fat catabolism of GBCC_F0227

[0181] In this example, we selected an effective strain that can inhibit the absorption of neutral fat, i.e., triglyceride-type fat, and examined its fat absorption characteristics. Using the fat absorption inhibition ability as a selection criterion, the triglyceride consumption rate before and after culture was measured to determine how effectively various types of probiotic bacteria metabolize triglycerides present in the culture medium. The tested strains included GBCC_F0227 of the present invention, WCFS1, which is a reference strain of L. plantarum, and KCTC3108, which is a standard strain, KCTC3112, which is a standard strain of L. fermentum, and KCTC3163, which is a standard strain of L. gasseri, and a total of 31 strains of 10 species. The consumption rate test was performed by comparing the triglyceride amount measured in the culture medium before cultivation with the triglyceride measurement in the sample filtered with a 0.22 μm syringe filter after centrifuging the culture supernatant obtained by anaerobically culturing the test strains in the MRS culture medium at 37°C for 16 hours using an automatic biochemical analyzer (Clinical chemistry analyzer, DRI-CHEM NX700). Figure 11 shows the triglyceride measurement values ​​after cultivation for each test strain. The neutral fat in the culture supernatant of most of the test strains decreased after cultivation (the measured neutral fat in the MRS culture medium before cultivation was about 100 mg / dL, see the gray bar on the far left of Figure 11), but in particular, the GBCC_F0227 and WCFS1 strains of the present invention were superior to the other strains in neutral fat consumption, confirming that they catabolized and consumed all the neutral fat in the medium.

[0182] Since the growth rate and other factors are different for each strain, there may be differences in the degree of material metabolism, which may affect the measured value of neutral fat consumption rate. Therefore, to control the variables, optical density (OD) at 600 nm was used as a surrogate for cell density. 600) was used to measure neutral fat. In this experiment, OD was measured using four other strains of the same L. plantarum species as GBCC_F0227 (GBCC_F0053, GBCC_F0001, GBCC_F0404, and WCFS1). 600 Triglyceride concentration was measured for the supernatant cultured with one of the test strains under the same conditions until reaching 3 or 6. As a result, GBCC_F0227 showed the best neutral fat catabolism, as in the experiment of Fig. 11, especially OD 600 In the 6 culture, it was observed that the neutral fat consumption rate was significantly faster than that of the other Plantarum strains, and almost all neutral fat was consumed (Figs. 12a and 12b).

[0183] [Example 12]

[0184] Neutral fat catabolic gene activation of GBCC_F0227

[0185] In this example, it was verified at the genetic level that GBCC_F0227 of the present invention is excellent in neutral fat catabolism. The expression of genes of the α / β hydrolase gene family including lipase was compared and analyzed at the same culture level for WCFS1, a Lactobacillus plantarum strain similar to GBCC_F0227. Specifically, the two strains GBCC_F0227 and WCFS1 were cultured in MRS medium at OD 600After anaerobically culturing at 37°C to a level of 3 or 6, RNA was extracted from the pellet precipitated by centrifugation, cDNA was synthesized using random hexamer primers, and α / β hydrolysis genes shared by the two strains were selected through genome analysis, and their expression levels were quantified using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). RNA polymerase subunit beta gene (rpoβ) was selected as a housekeeping gene as a criterion for comparing gene expression levels, and Moraxella lipase 2-like (symbol abH04), α / β hydrolase (symbol abH08_1), α / β fold hydrolase (symbol abH08_2), and Proline iminopeptidase (symbols abH11_1 and abH11_2) were selected as α / β hydrolase genes shared by the two strains, and their expression was quantified by constructing primers. The relative expression rate of the α / β hydrolase gene group gene (abH) quantified in this way is shown in Figure 13a (OD 600 3) and Fig. 13b (OD 600 6) is shown in GBCC_F0227. OD is compared to WCFS1. 600 At 3 cell concentrations, gene expression of abH08_1, abH08_2, and abH11_2 was significantly higher (Fig. 13a), and OD 600 Under the 6 conditions, the expression of abH04, abH08_1, abH08_2, and abH11_1 was significantly higher (Fig. 13b). In addition, the abH08 gene showed a significantly higher expression level than WCFS1 in both culture conditions, and it was confirmed that abH04 and abH11 showed different increases and decreases in expression levels depending on the culture level conditions. In summary, GBCC_F0227 showed a higher OD than WCFS1. 600 In both conditions 3 and 6, the expression of most α / β hydrolase gene groups was significantly higher, confirming that neutral fat could be catabolized and consumed more quickly.

[0186] [Example 13]

[0187] F0227's regulatory effect on lipid metabolism and obesity-related gene activity

[0188] In this example, it was confirmed that the GBCC_F0227 strain has the effect of regulating the gene activity of adiponectin, leptin, PPARγ, CD36, GLP-1 receptor, POMC, and CART, which are proteins related to fat metabolism and obesity, toward anti-obesity in a mouse model in which obesity was induced by a high-fat diet.

[0189] Adiponectin is a hormone involved in regulating energy metabolism, which has the effect of lowering blood triglycerides, increasing high-density lipoprotein (HDL), and inhibiting fat accumulation in the liver. Leptin is an adipokine secreted by adipocytes that regulates food intake and energy expenditure. Blood leptin concentrations increase in obesity, and in patients with non-alcoholic fatty liver disease (NAFLD), higher blood leptin concentrations are known to worsen the severity of NAFLD. Cluster of differentiation 36 (CD36) is a cell surface receptor that binds to oxidized low-density lipoprotein (LDL) or long-chain fatty acids, and is known to promote triglyceride accumulation by taking up free fatty acids into cells. PPARγ (peroxisome proliferator-activated receptor gamma) is a nuclear receptor with transcription factor function, mainly present in adipose tissue, and is known to induce the transcription of fatty acid absorption-related proteins, including CD36. Pro-opiomelanocortin (POMC) is a hormone precursor synthesized in the pituitary gland. When cleaved, it produces hormones involved in energy metabolism homeostasis and appetite regulation, including ACTH and MSH. CART (cocaine- and amphetamine-regulated transcript) is a neuropeptide secreted in response to dopamine release, which regulates energy homeostasis and interacts with appetite-regulating neural circuits in the hypothalamus. The GLP-1 receptor is a G protein-coupled receptor that suppresses appetite when bound to its ligand, GLP-1.

[0190] The mice were divided into three groups (an experimental group (G3) administered with the L. plantarum GBCC_F0227 strain in parallel, an obese positive control group (G2) administered with the same high-fat diet and a cryoprotectant at the same amount as the strain administered in G3, and an obese negative control group (G1) administered with a cryoprotectant along with a normal feed diet (TEKLAD 2018S, Inotiv, USA)) in the same manner as Example 10. For some genes, an additional control group (G4) was added to which the strain in G3 was administered with a high-fat diet and the same amount of the L. plantarum standard strain KCTC 3108, instead of the strain in G3, and the mice were fed for 8 weeks. After the feeding period, the mice were necropsied, and epididymal white adipose tissue (eWAT), the left medial lobe of the liver, and the hypothalamus tissue of the brain were obtained, and total RNA was extracted from each. Only the hypothalamus tissue was collected from the G4 control group. From this RNA, cDNA was synthesized using oligodeoxythymidine (Oligo dT), and the expression of adiponectin and leptin genes (epididymis), PPARγ and CD36 genes (left medial lobe of the liver), and POMC and CART genes (hypothalamus) was quantified by RT-qPCR. The housekeeping gene glyceraldehyde 3-phosphate dehydrogenase (GADPH) gene was selected as an internal standard for expression rate quantification, and RT-qPCR was performed using primers designed by referencing the base sequences of genes including adiponectin from the gene database of the National Center for Biotechnology Information (NCBI) in the United States. The results of expression rate measurement are summarized in Figures 14a to 14g.

[0191] In the epididymis, which is a visceral white adipose tissue, the expression level of the adiponectin gene was significantly reduced to half the level of the normal weight diet group (G1) in the obesity-induced group (G2) induced with a high-fat diet, but in the experimental group (G3) administered with the GBCC_F0227 strain of the present invention in conjunction with the high-fat diet, the expression of adiponectin significantly increased to a level close to that of the normal diet group (Fig. 14a). The expression rate of leptin, which had been significantly increased in the obesity-induced group (G2), also statistically significantly decreased when the GBCC_F0227 strain was administered (Fig. 14b). The expression rates of CD36 (Fig. 14c) and PPARγ (Fig. 14d), which worsen hepatic steatosis and hepatic fat accumulation, in the left lobe of the liver were also increased in the obesity-induced group (G2) compared to the normal diet group (G1), but administration of GBCC_F0227 statistically significantly reduced the expression rates, dropping them to a level lower than that of the normal diet group (G1) and beyond restoration.

[0192] Administration of GBCC_F0227 statistically significantly increased the hypothalamic expression rates of GLP-1 receptors (Fig. 14e), POMC (Fig. 14f), and CART (Fig. 14g), which have appetite-suppressing effects, compared to the high-fat diet-induced obesity group (G2) or the normal diet group (G1). In addition, the expression rate-enhancing effect of GBCC_F0227 was statistically significantly superior to that of the standard strain of the same species, KCTC 3108, in the case of the GLP-1 receptor, or was statistically equivalent to that of POMC or CD36 with a higher average value.

[0193] Therefore, when the GBCC_F0227 strain of the present invention was administered to a high-fat diet-induced obese mouse model, it was confirmed that it not only had an anti-obesity effect of reducing body weight, but also had an effect of regulating the expression of obesity genes related to fat accumulation, energy metabolism, or appetite in an anti-obesity direction.

[0194] [Example 14]

[0195] F0227's effect on preventing muscle mass loss

[0196] In this example, we examined whether GBCC_F0227 has the efficacy to reverse muscle mass loss in an animal model of diet-induced obesity.

[0197] Mice were divided into three groups (G1, G2, G3) in the same manner as in Example 10 and fed for 8 weeks. After the feeding period, the skeletal muscles in the hind limbs of the mice, namely the tibialis anterior (TA), extensor digitorum longus (EDL), soleus (Sol), and gastrocnemius (GA), were removed and weighed. The measured skeletal muscle weights were normalized as a ratio (mg / g) to the mouse body weight to compare the weight differences among the experimental groups. In addition, the percentages for each skeletal muscle were calculated based on the muscle mass of the high-fat diet group (G2).

[0198] The experimental results showed that mice (G2) induced to have obesity with a high-fat diet showed a statistically significant decrease in muscle mass in all four types of skeletal muscles compared to the normal diet group (G1). Administration of GBCC_F0227 lactic acid bacteria had a statistically significant effect in restoring muscle mass compared to G2. Specifically, administration of GBCC_F0227 had the effect of increasing skeletal muscle mass by 1.24 times in the tibialis anterior (Fig. 15a), 1.05 times in the extensor digitorum longus (Fig. 15b), 1.18 times in the soleus (Fig. 15c), and 1.24 times in the gastrocnemius (Fig. 15d) compared to G2.

[0199] [Example 15]

[0200] Anti-muscle loss effect of F0227

[0201] In this example, we examined whether GBCC_F0227 has the efficacy to reverse muscle loss observed in an animal model of diet-induced obesity.

[0202] In the same manner as in Example 14, grip strength tests were performed on the four paws of mice before dissection of mice fed for 8 weeks. The grip strength test was performed in a blinded manner so that the tester did not know which experimental group the mouse belonged to, and the grip strength (in g weight) was measured three times for each individual at intervals to prevent the mice from becoming exhausted. The average value was normalized and expressed as the value (g / g) divided by the mouse body weight (in g).

[0203] As a result of the experiment, mice (G2) induced to be obese by a high-fat diet showed a significant decrease in grip strength compared to the normal diet group (G1) (p< 0.001), but when GBCC_F0227 lactic acid bacteria was administered, grip strength was statistically significantly restored compared to G2. Specifically, GBCC_F0227 administration had the effect of increasing the grip strength of all four paws of the mice by 1.13 times compared to G2, and this was a statistically significant recovery (p< 0.01).

[0204]

Claims

1. A pharmaceutical composition for treating or preventing a metabolic disease that increases GLP-1 secretion or suppresses appetite, comprising at least one selected from among a strain of the species Lactiplantibacillus plantarum, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, and a cell-free culture supernatant of the strain as an active ingredient.

2. A pharmaceutical composition for treating or preventing a metabolic disease, characterized in that the strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain in claim 1 exhibits at least one effect of suppressing weight gain in the host, reducing body fat in the host, and suppressing appetite in the host.

3. Lactiplantibacillus plantarum GBCC_F0227 strain (Accession number: KCTC 15450BP).

4. In the third paragraph, the GBCC_F0227 strain is a Lactiplantibacillus plantarum strain characterized in that the recA gene sequence includes the sequence described in SEQ ID NO:

2.

5. In the third paragraph, the GBCC_F0227 strain is a Lactobacillus plantarum strain characterized in that it contains a naturally occurring mutation of Lactobacillus plantarum.

6. A pharmaceutical composition for preventing or treating a metabolic disease, comprising as an active ingredient at least one selected from among Lactobacillus plantarum GBCC_F0227 strain (Accession No.: KCTC 15450BP), a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, and a cell-free culture supernatant of the strain.

7. A pharmaceutical composition for preventing or treating a metabolic disease, characterized in that the metabolic disease in claim 6 is any one selected from the group consisting of obesity, obesity-induced appetite suppression, metabolic syndrome, hypertension, arteriosclerosis, hyperglycemia, hyperlipidemia, hypertriglyceridemia, fatty liver, non-alcoholic fatty liver disease, hyperinsulinemia, diabetes, impaired glucose tolerance, hypercholesterolemia, cardiovascular disease, and insulin resistance syndrome.

8. A pharmaceutical composition for preventing or treating a metabolic disease, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, and an extract of the culture inhibit DPP-4 enzyme activity in the 6th paragraph.

9. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain stimulates GLP-1 secretion in the host in accordance with paragraph 6.

10. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain promotes expression of a GLP-1 receptor in a host.

11. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain stimulates appetite-suppressing neurons of the host in accordance with paragraph 6.

12. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain increases the expression of proopiomelanocortin or CART (cocaine- and amphetamine-regulated transcript) of the host in the 12th paragraph.

13. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain exhibits at least one effect of suppressing weight gain in the host, reducing body fat in the host, and suppressing appetite in the host, in accordance with paragraph 6.

14. A pharmaceutical composition for preventing or treating a metabolic disease, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain inhibits DPP-4 enzyme activity in the 9th or 10th paragraph.

15. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain reduces a decrease in skeletal muscle mass and / or a decrease in skeletal muscle strength in a host accompanied by obesity, according to claim 6.

16. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain reduces visceral fat accumulation in the host.

17. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain reduces liver fat accumulation in the host.

18. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, a dead cell of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain reduces blood neutral fat of the host in the 6th paragraph.

19. A pharmaceutical composition for preventing or treating metabolic diseases, characterized in that in paragraph 6, the GBCC_F0227 strain, a lysate of the strain, an extract of the strain, dead cells of the strain, a culture of the strain, an extract of the culture, or a cell-free culture supernatant of the strain contains at least one selected from the group consisting of α-ketoglutaric acid, 3-phenyllactic acid, cystathionine, 1-monopalmitin, glycerol monostearate, and 2-hydroxy-3-methylbutyric acid.

20. A food composition comprising Lactiplantibacillus plantarum GBCC_F0227 strain (accession number: KCTC 15450BP) or a culture thereof.

21. Health functional food containing Lactibacillus plantarum GBCC_F0227 strain (Accession number: KCTC 15450BP) or a culture thereof.

22. Feed composition comprising a Lactiplantibacillus plantarum strain (Accession No.: KCTC 15450BP) or a culture thereof.

Images