KR102220543B1 - Composition comprising Lactobacillus plantarum BK-021 for preventing or treating diabetes mellitus or vascular disease - Google Patents

Abstract

The present invention relates to a composition for preventing or treating diabetes or vascular disease containing a strain of Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P), wherein the strain is α-glucosidase inhibitory effect Or it has the effect of reducing blood cholesterol, along with diabetes, hypercholesterolemia, hyperlipidemia, hypertension, thrombosis, arteriosclerosis, hypertension, angina, myocardial infarction, ischemic heart disease, heart failure, and milder diseases caused by the deposition of cholesterol in blood vessels. It can be usefully used as a pharmaceutical composition or health functional food capable of complex prevention, improvement or treatment of complications, cerebral infarction, cerebral hemorrhage, stroke, and the like that occur after angioplasty.

Description

Composition comprising Lactobacillus plantarum BK-021 for preventing or treating diabetes mellitus or vascular disease {Composition comprising Lactobacillus plantarum BK-021 for preventing or treating diabetes mellitus or vascular disease}

The present invention relates to a composition for preventing or treating diabetes or vascular disease containing a strain of Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P). In more detail, the present invention relates to a composition for preventing or treating diabetes or vascular disease containing Lactobacillus plantarum BK-021 strain having α-glucosidase inhibitory effect or blood cholesterol reduction effect.

The incidence of cardiovascular diseases, including cancer, and metabolic diseases is also on the rise despite the great advances in modern life sciences in relation to changes in dietary habits, aging population and various stresses according to recent economic development. In particular, among the above metabolic diseases, diabetes is a chronic wasting disease characterized by abnormal glucose metabolic dysfunction due to absolute deficiency of insulin or resistance, and a disease that causes various complications such as systemic lethargy and resistance decrease, vascular disorders, cerebral infarction, myocardial infarction, etc. to be. In addition, according to the World Health Organization (WHO), 350 million people suffer from diabetes, and the number is projected to double by 2030 (Danaei, G. et al., 2011).

Diabetes is largely divided into two types depending on the cause of the onset and the condition of the disease. A specific human lymphocyte antigen (HLA) and viral infection destroy the beta cells of Langerhans islands, resulting in a non-progressive insulin secretion. There is type 1 diabetes, and insulin-independent type 2 diabetes caused by increased resistance to insulin in peripheral tissues such as muscle, liver, and adipose tissue due to lack of exercise, obesity, overeating, and stress. In particular, type 2 diabetes accounts for more than 90% of all diabetic patients due to aging and lifestyle changes in the modern society, and the number is steadily increasing. The main cause of the onset of type 2 diabetes is insulin resistance, which refers to a condition in which the response to a certain amount of insulin concentration is reduced than normal, and generally occurs due to genetic and environmental factors (Hossain, P. et al. , 2007).

α-glucosidase is a digestive enzyme present in the villi of the small intestine. It hydrolyzes disaccharides or small saccharides into monosaccharides, which are digestive and absorbed by carbohydrates. Therefore, α-glucosidase inhibitors inhibit the hydrolysis of carbohydrates in the upper part of the plant including the duodenum (lower the digestive absorption rate), thereby inhibiting the rise of blood sugar levels, and thus can be used for the treatment of adult diseases such as diabetes and fructose. . In addition, many studies have been conducted on the inhibition of α-glucosidase, which is essential for carbohydrate metabolism, for the purpose of controlling blood sugar lowering (Tewari, N. et al., 2003).

The α-glucosidase inhibitor for this action is a substance that inhibits the degradation of oligosaccharides by α-glucosidase and is distributed in some plants and microorganisms. Microbial α-glucosidase inhibitors include pseudooligosaccharide from Actinoplanes strain, 1-deoxynojirimycin produced by bacteria of the genus Streptomyces and Bacillus. Is known, and commercially available α-glucosidase inhibitors such as acarbose, voglibose, and 1-deoxynojirimycin are used, but they not only show gastrointestinal disorders such as abdominal distension, vomiting, and diarrhea, but also undegraded disaccharides are decomposed by bacteria in the colon. It can cause side effects such as gas, diarrhea, and constipation. In addition, when α-glucosidase is excessively inhibited, there is a problem in that the absorption rate of sugar is significantly lowered, resulting in symptoms of hypoglycemia.

On the other hand, in addition to diabetes, various economic and social changes take place along with the progress of economic development and industrialization, and the incidence of different types of adult diseases is remarkably increasing as the dietary habits and lifestyles gradually change. In particular, infectious diseases, digestive and respiratory diseases are gradually decreasing, and hypercholesterolemia, hyperlipidemia, hypertension, thrombosis, arteriosclerosis, hypertension, which are vascular diseases caused by aging population, overeating, drinking, smoking, and lack of exercise. , Angina pectoris, myocardial infarction, ischemic heart disease, heart failure, complications that occur after transvascular angioplasty, cerebral infarction, cerebral hemorrhage, and chronic diseases such as stroke are increasing. These vascular diseases are chronic diseases caused by oxidation of LDL cholesterol among cholesterol in the serum, and the oxidation products accumulate in the subcutaneous tissues in the arteries, and various inflammatory reactions increase.It is the second cause of death in the elderly in Korea after cancer. . In particular, cardiovascular diseases such as hyperlipidemia, hypertension, hypertensive retinopathy, arrhythmia, myocardial infarction, deep vein and angina, and cardiac infarction account for most of the causes of death in the elderly.

According to the guidelines for diagnosing hyperlipidemia according to blood lipid and lipoprotein concentrations, in the case of normal people, total cholesterol is 110-240mg/㎗, LDL cholesterol is 65-160mg/㎗, and triglycerides are 200mg/㎗ or less. If left untreated, serious complications such as high blood pressure, coronary atherosclerosis (angina pectoris, myocardial infarction), and cerebral arteriosclerosis (cerebral infarction) can result (Fire Committee on Treatment Guidelines for Hyperlipidemia). The vascular diseases include thrombosis, arteriosclerosis, high blood pressure, angina pectoris, myocardial infarction, ischemic heart disease, heart failure, complications occurring after transvascular arterioplasty, cerebral infarction, cerebral hemorrhage, and stroke. Patients with vascular disease are continuously given drugs to treat it, but it is not easy to be cured in modern medicine. Also, if you take the drug for a long time, the side effects are great.

On the other hand, anti-obesity studies using probiotics as a new alternative to the treatment of such adult diseases are in progress. The necessity of developing a therapeutic agent using a complex method such as resolving the imbalance of intestinal flora, increasing intestinal beneficial bacteria, and restoring the intestinal tract using probiotics having excellent effects in such treatment is gradually increasing. Lactobacillus is widely distributed in human and animal oral, intestinal, vaginal, feces, and fermented foods such as kimchi, and is closely related to human and animal health, and promotes various health such as intestinal action, suppression of harmful bacteria, immune regulation, and anticancer. It shows the effect.

Accordingly, the present inventors were studying various physiological activities of lactic acid bacteria, and the present inventors' strain, Lactobacillus plantarum BK-021, confirmed that it has excellent intestinal adhesion, α-glucosidase inhibitory effect, and blood cholesterol reduction effect. The present invention has been completed.

Republic of Korea Patent Registration No. 10-1825836 (Name of the invention: novel Lactobacillus plantarum Lb41 strain and pharmaceutical composition for the prevention and treatment of diabetes or insulin resistance syndrome containing the same, Applicant: Korea Food Research Institute, registration date: 2018 January 30) Republic of Korea Patent Registration No. 10-1825837 (Name of invention: novel Lactobacillus plantarum Ln4 strain and pharmaceutical composition for the prevention and treatment of diabetes or insulin resistance syndrome containing the same, Applicant: Konkuk University Industry-Academic Cooperation Foundation/Korea Food Research Institute , Registration date: January 30, 2018) Republic of Korea Patent Publication No. 10-2018-0092178 (Title of invention: New Lactobacillus plantarum FMB #31 strain with excellent anti-cholesterol activity and its use, Applicant: Pukyong National University Industry-Academic Cooperation Foundation, Publication Date: August 17, 2018 )

Danaei, G. et al., National, regional, and global trends in fasting plasma glucose and diabetes prevalence since 1980: systematic analysis of health examination surveys and epidemiological studies with 370 country-years and 2ㅇ7 million participants, Lancet, 378( 9785), 31-40, 2011. Hossain, P. et al., Obesity and diabetes in the developing world-a growing challenge, Engl. J. Med., 356(3), 213-215, 2007. Tewari, N. et al., Synthesis and bioevaluation of glycosyl ureas as alpha-glucosidase inhibitors and their effect on mycobacterium, Bioorg. Med. Chem., 11(13), 2911-2922, 2003.

An object of the present invention is to provide a composition for the prevention or treatment of diabetes or vascular disease containing the Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P) strain. In more detail, an object of the present invention is to provide a composition for preventing or treating diabetes or vascular disease containing Lactobacillus plantarum BK-021 strain having α-glucosidase inhibitory effect or blood cholesterol reduction effect .

The present invention relates to a composition for the prevention or treatment of diabetes or vascular disease containing a Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P) strain or a culture thereof.

The strain or a culture thereof is characterized by having an α-glucosidase inhibitory effect.

In addition, the strain or a culture thereof is characterized in that it has the effect of reducing blood cholesterol.

The vascular disease is hypercholesterolemia, hyperlipidemia, hypertension, thrombosis, arteriosclerosis, hypertension, angina pectoris, myocardial infarction, ischemic heart disease, heart failure, complications that occur after transvascular arterioplasty, which are diseases caused by the deposition of cholesterol in blood vessels, It is characterized in that it is selected from diseases consisting of cerebral infarction, cerebral hemorrhage and stroke.

In addition, the present invention provides a health functional food for preventing or improving diabetes or vascular disease containing the Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P) strain or a culture thereof.

Hereinafter, the present invention will be described in more detail.

The Lactobacillus plantarum BK-021 strain of the present invention is characterized in that it has intestinal adhesion.

The culture of the Lactobacillus plantarum BK-021 strain obtained in the present invention can be prepared in a liquid or solid form, and a process of drying or powdering the culture in a liquid state may be added. Likewise, a solid culture may be dried or powdered according to the moisture content.

In addition, the present invention provides a pharmaceutical composition for the prevention or treatment of diabetes or vascular disease containing Lactobacillus plantarum BK-021 strain or a culture thereof. The Lactobacillus plantarum BK-021 strain or a culture thereof may be added to the pharmaceutical composition of the present invention in an amount of 0.001 to 100% by weight. The pharmaceutical compositions may be formulated and used in the form of oral dosage forms such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., external preparations, suppositories, and sterile injectable solutions, respectively, according to conventional methods. Carriers, excipients and diluents that may be included in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, gum acacia, alginate, gelatin, calcium phosphate, calcium silicate, cellulose , Methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate, and mineral oils. In the case of formulation, it is prepared using diluents or excipients such as fillers, extenders, binders, wetting agents, disintegrants, and surfactants that are usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and these solid preparations contain at least one excipient, such as starch, calcium carbonate, and water in the strain or culture thereof of the present invention. It is prepared by mixing cross or lactose, gelatin, and the like. In addition to simple excipients, lubricants such as magnesium stearate and talc are also used. Liquid preparations for oral use include suspensions, liquid solutions, emulsions, syrups, etc. In addition to water and liquid paraffin, which are commonly used simple diluents, various excipients such as wetting agents, sweetening agents, fragrances, and preservatives may be included. . Preparations for parenteral administration include sterile aqueous solutions, non-aqueous solutions, suspensions, emulsions, lyophilized preparations, and suppositories. As the non-aqueous solvent and suspending agent, propylene glycol, polyethylene glycol, vegetable oil such as olive oil, and injectable ester such as ethyl oleate may be used. As a base for suppositories, witepsol, macrogol, tween 61, cacao butter, laurin, glycerogelatin, and the like may be used.

The dosage of the pharmaceutical composition of the present invention will vary depending on the age, sex, weight of the subject to be treated, the specific disease or pathology to be treated, the severity of the disease or pathology, the route of administration, and the judgment of the prescriber. Dosage determination based on these factors is within the level of one of ordinary skill in the art, and dosages generally range from 0.01 mg/kg/day to approximately 2000 mg/kg/day. A more preferred dosage is from 1 mg/kg/day to 500 mg/kg/day. Administration may be administered once a day, or may be divided several times. The above dosage does not in any way limit the scope of the present invention.

The pharmaceutical composition of the present invention can be administered to mammals such as mice, livestock, and humans by various routes. All modes of administration can be expected and can be administered, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or cerebrovascular injection. The composition containing the strain or a culture thereof of the present invention is edible and has little toxicity and side effects, so it is a drug that can be safely used even when taken for a long time for prophylactic purposes.

In addition, the present invention provides a health functional food for the prevention or treatment of diabetes or vascular disease, comprising the Lactobacillus plantarum BK-021 strain or a culture thereof and food additives acceptable for food. The Lactobacillus plantarum BK-021 strain or a culture thereof may be added to the health functional food of the present invention in an amount of 0.001 to 100% by weight. The health functional food of the present invention includes the form of tablets, capsules, pills or liquids, and as foods to which the strain or culture thereof of the present invention can be added, for example, various drinks, meat, sausages, Breads, candies, snacks, noodles, ice cream, dairy products, soups, ionic beverages, beverages, alcoholic beverages, gum, tea and vitamin complexes.

Accordingly, the present invention can also provide a fermented food for preventing or improving diabetes or vascular disease fermented using a strain of Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P), the fermented food You can choose from kimchi, salted fish, yogurt, or paste. The paste may be soybean paste, soy sauce, red pepper paste, or cheonggukjang.

The culture of the strain of the present invention may be one of Lactobacillus plantarum BK-021 cultivated in a liquid or solid medium for culturing MRS lactic acid bacteria, more preferably, it may be cultured in the following medium, which is a vegetable medium. The vegetable medium may contain Hydrolysed Soy Protein, Yeast Extract, D-Glucose Monohydrate, Tween 80 (Polyoxyethyelene Sorbitan Monooleate), Sodium Acetate, Magnesium Sulfate, Manganese Sulfate, Dipotassium Phosphate. Preferably Hydrolysed Soy Protein 5~20g, Yeast Extract 5~20g, D-Glucose Monohydrate 10~30g, Tween 80 0.5~3g, Sodium Acetate 2~10g, Magnesium Sulfate 0.01~1g, Manganese Sulfate 0.001~0.1g, Dipotassium Phosphate 1~10g/ ℓ may be included, and most preferably in distilled water, Hydrolysed Soy Protein 15g, Yeast Extract 10g, D-Glucose Monohydrate 20g, Tween 80 1g, Sodium Acetate 5g, Magnesium Sulfate 0.1g, Manganese Sulfate 0.05g, Dipotassium Phosphate may be a medium prepared in the range of 2g / L. The pH of the medium may be adjusted in the same manner as the MRS medium.

The present invention relates to a composition for preventing or treating diabetes or vascular disease containing a strain of Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P), wherein the strain is α-glucosidase inhibitory effect Or it has the effect of reducing blood cholesterol, along with diabetes, hypercholesterolemia, hyperlipidemia, hypertension, thrombosis, arteriosclerosis, hypertension, angina, myocardial infarction, ischemic heart disease, heart failure, and milder diseases caused by the deposition of cholesterol in blood vessels. It can be usefully used as a pharmaceutical composition or health functional food capable of complex prevention, improvement or treatment of complications, cerebral infarction, cerebral hemorrhage, stroke, and the like that occur after angioplasty.

Although prior art related to the Lactobacillus plantarum strain, which is effective in treating diabetes or reducing blood cholesterol, has been disclosed, it is a strain that is completely different from the strain of the present invention in genetic or physiological efficacy. And the present invention can be said to have a difference in the technical configuration and effects resulting from it, respectively.

1 is a phylogenetic tree diagram showing the genetic location of the strain of the present invention.
2 is a graph showing the results of confirming the α-glucosidase inhibitory effect after culturing the strain of the present invention for 24 hours in MRS and MRS replacement plant medium.
3 is a graph showing the results of confirming the α-glucosidase inhibitory effect after culturing the strain of the present invention for each time in MRS replacement plant medium.
Figure 4 is a graph of the results confirming the effect of inhibiting total blood cholesterol of the strain of the present invention.
5 is a graph showing the results of confirming the effect of reducing blood low-density lipoprotein cholesterol (LDL-C) of the strain of the present invention.

Hereinafter, a preferred embodiment of the present invention will be described in detail. However, the present invention is not limited to the embodiments described herein and may be embodied in other forms. Rather, it is provided to sufficiently convey the spirit of the present invention to those skilled in the art so that the contents introduced herein are thorough and complete.

Example 1. Lactobacillus plantarum Sympathy of BK-021

A single colony of bacteria obtained by spreading the stock solution of the onion fermentation extract on MRS medium was collected with loops and cultured in MRS broth. DNA extraction and 16S rRNA sequencing were identified by requesting the Korea Microbiological Conservation Center. The strain was identified by analyzing the 16S rRNA sequence using the Basic Local Alignment Search Tool (BLAST) search engine of the National Center for Biotechnology Information (NCBI, www.ncbi.nlm.nih.gov) and the EzTaxon server. The isolated strain was found to have a nucleic acid sequence of SEQ ID NO: 1 as a result of 16S rRNA sequencing analysis for identification and classification of microorganisms.

SEQ ID NO: 1: 16S rRNA of Lactobacillus plantarum BK-021

In addition, as a result of the analysis, the lactic acid bacteria strain is Lactobacillus plantarum subsp. As shown in the phylogenetic tree of Figure 1 shown by the neighbor-joining method. plantarum ACGZ01000098 showed the highest molecular phylogenetic relationship.

The strain thus identified was named Lactobacillus plantarum BK-021 and deposited with the Korea Microbial Conservation Center on November 14, 2017 (accession number KFCC11749P).

In addition, the Lactobacillus plantarum BK-021 strain is anaerobic, the minimum temperature that can be grown is 25 ℃, the maximum temperature that can be grown 45 ℃, the pH that can be grown is 4 to 7, the appropriate culture time is 24 to 48 hours, the appropriate growth temperature is It was confirmed to be 30 ~ 37 ℃. A suitable medium for culturing the strain is Lactobacilli MRS media, and it was confirmed that cultivation is possible using various animal and plant materials as raw materials.

Among them, as a vegetable medium, Hydrolysed Soy Protein 5~20g, Yeast Extract 5~20g, D-Glucose Monohydrate 10~30g, Tween 80 0.5~3g, Sodium Acetate 2~10g, Magnesium Sulfate 0.01~1g, Manganese Sulfate 0.001~0.1g, Dipotassium Phosphate 1~10g/ℓ It grows best in a medium prepared.

Next, in the enzymatic properties of the strain using the ZYM kit, it was confirmed that it has the enzymatic activities of Naphthol-AS-BI-phosphohydrolase, which activates phagocytosis, and α-Glucosidase, which is effective in lactose deficiency, and is shown in Table 1. And, it is shown in Table 2 by examining the sugar utilization ability using the API 50CH kit. In Tables 1 and 2, when each enzyme and carbon source are used, it is represented by +, when not used, it is represented by -, and when a weak reaction is shown, it is represented by w.

Enzyme Lactobacillus plantarum BK-021 Alkaline phosphatase + Esterase (C4) w Esterase Lipase (C8-Lipases) w Lipase (C14) - Leucine arylamidase + Valine arylamidase + Cystine arylamidase (Proteases) + Trypsin - α-Chymotrypsin - Acid phosphatase (Phosphatases) + Naphthol-AS-BI-phosphohydrolase + α-Galactosidase (melibiase) - β-Galactosidase (lactases) + β-Glucuronidase (hyaluronidase) - α-Glucosidase (maltases) + β-Glucosidase (cellulases) + N-acetyl-β-glucosaminidase (chitinases) + α-Mannosidase - α-Fucosidase -

Characteristic (per) Lactobacillus plantarum BK-021 Glycerol - D-arabinose - D-Ribose + D-Xylose + D-Galactose + D-Glucose + D-Fructose + D-Mannose + D-Mannitol + D-Sorbitol + Methyl-α-D-glucopyranozide + N-acetylglucosamine + Amygdalin + Arbutin + Salicin + D-Celobiose + D-Maltose + D-Lactose (of cow origin) + D-Saccharose + D-Trehalose + Inulin + D-Melesitose -

Example 2. Lactobacillus plantarum BK-021's intestinal adhesion ability evaluation

In order for various intestinal physiological effects to be exerted in the body, it is important that lactic acid bacteria have long-term survival in the intestine and have intestinal adhesion (Appl. Environ. Microbiol., 1992, 58, 2034-2039).

In order to confirm the intestinal adhesion ability to Lactobacillus plantarum BK-021, the method of Haghshenas et al. (2015) was modified and carried out using Caco-2 cells. _{Culture (5% CO 2} ) in RPMI-1640 media and spread with 2 × 10 ⁵ cells/well, and then Lactobacillus plantarum BK-021 and control Lactobacillus plantarum VB6 (OD = 1; 10 ⁸ CFU/ml) were added to Caco-2. Cells were co-cultured at a ratio of 1: 1 (2 × 10 ⁵ ; v/v) (37℃, 3 hrs., absented CO ₂ condition), and the intestinal adhesion ability of the lactic acid bacteria was confirmed by microscopic observation and viable cell count. . The control group Lactobacillus plantarum VB6 did not show intestinal attachment ability, but in Lactobacillus plantarum BK-021, ^{it was confirmed that the intestinal attachment ability was 1.3 × 10 3} CFU/ml, and is shown in Table 3.

Identified species Treated (CFU/well) Adhesion (CFU/ml) Lactobacillus plantarum VB6 KCCM11921P 1.7 × 10 ⁵ 0 Lactobacillus plantarum BK-021 1.13 × 10 ⁵ 1.3 × 10 ³

Example 3. α-glucosidase activity inhibition effect measurement experiment

The initial concentration of the plant lactic acid bacteria Lactobacillus plantarum BK-021 is 1 × 10 ⁸ CFU/ml ~ 1 × 10 ¹⁰ CFU/ml, respectively. MRS and its own MRS replacement vegetable medium (Food grade: Hydrolysed Soy Protein 15g, Yeast Extract 10g, D-Glucose Monohydrate 20g, Tween 80 1g, Sodium Acetate 5g, Magnesium Sulfate 0.1g, Manganese Sulfate 0.05g, Dipotassium Phosphate 2g/ℓ) at 30℃ for 12 to 48 hours and centrifuge the bacterial solution at 4,000 rpm for 30 minutes After separation, a culture supernatant sample was prepared.

To inhibit the activity of the sample against α-glucosidase, p-nitrophenyl a-D-glucopyranoside (pNPG) was dissolved in 0.1M potassium phosphate buffer (pH 6.9) and used as a substrate. The extract was mixed well after adding 0.1U/ml of α-glucosidase (Sigma Chemical Co, USA) in an enzyme solution dissolved in 0.1M potassium phosphate buffer (pH 6.9) and 2mM p-nitrophenyl aD-glucopyranoside in a substrate solution, and then mixed well at 37℃. After reacting for 20 minutes, absorbance was measured at 405 nm. As a positive control, acarbose concentrations of 1, 5, and 10 mM were used in the experiment in the same manner.

Inhibition (%) = [1-(Sample absorbance slope/Blank absorbance slope)] × 100

In order to confirm the α-glucosidase inhibitory effect, the strain was cultured in MRS medium and its own plant-based media (PBM) plant medium for MRS replacement for 24 hours, and then the α-glucosidase inhibitory effect was confirmed.

The results are shown in Figure 2, as a result, the culture supernatant of the BK-021 strain in both media showed a high inhibitory effect of 82 to 90% similar to the positive control, acarbose 10mM (92% inhibition).

Example 4. L. plantarum Optimization of culture conditions of BK-021 strain

The α-glucosidase inhibitory effect of the culture supernatant of L. plantarum BK-021 strain for each culture time using plant-based media (PBM) plant medium was measured, and the results are shown in FIG. 3. At this time, Acarbose was cultured for 48 hours at each concentration and presented as a control group.

As a result, when cultured for 48 hours, the best 90% inhibitory effect was confirmed, which is higher than Acarbose 10mM (63% inhibition), which is a positive control, and the effect is maintained when cultured for 24 to 96 hours, more preferably 36 to 48 hours. Was found to be.

Example 5. L. plantarum Measurement of total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) in blood by BK-021 strain

Example 5-1. L. plantarum Preparation of freeze-dried sample of BK-021 strain

The initial concentration of L. plantarum BK-021 strain is 1 × 10 ⁸ CFU/ml ~ 1 × 10 ¹⁰ CFU/ml, respectively, and MRS replacement plant medium (Food grade: Hydrolysed Soy Protein 15g, Yeast Extract 10g, D-Glucose Monohydrate 20g, Tween 80 1g, Sodium Acetate 5g, Magnesium Sulfate 0.1g, Manganese Sulfate 0.05g, Dipotassium Phosphate 2g) incubated for 12 hours at 30℃ and centrifuged for 20 minutes at 4,000rpm, and then the supernatant was removed. The remaining bacteria were washed with physiological saline (0.85% NaCl). After removal of physiological saline, the washed strain was mixed with 0.5 ml of a cryoprotectant solution (10% inulin aqueous solution) and frozen for 1 hour at RT (room temperature, 25°C), 2 hours at -20°C, and 2 hours at -80°C. Then, it was freeze-dried at -80°C for 48 hours using a freeze dryer. The lyophilized test strain was stored at -80°C, then dissolved in 10 ml of DW before administration to experimental animals and administered orally in 1 ml each.

Example 5-2. Preparation of experimental animals and experimental groups

The experimental animals were 5 weeks old SD-rat (Orient Bio, Gwangju City) purchased and used after 2 weeks of acclimatization. Administration was carried out for 8 weeks with twice a day administration for 5 days a week, and L. plantarum BK-021 1 × 10 ¹⁰ CFU/ml strain was ingested as an experimental group. Diet, drinking water, and body weight are measured once a week, and the experimental group is a normal group (normal diet n=10) who consumes a normal diet (Labdiet PicoLab Rodent Diet, #5053), and a high carb diet (Research diets AIN-76A Rodent Diet #D10001). ) Intake control group (n=10), high carbohydrate diet intake and L. plantarum BK-021 administration BK-021 group (n=10), high carbohydrate diet and Acarbose 15 mg/kg positive control group ( It consisted of a total of 4 groups with n=10).

Example 5-3. Blood total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) measurement

A biochemical analyzer (TBA-120FR, Toshiba LTD, Tokyo, etc.) to confirm the effect of reducing cholesterol by measuring the concentration of lipid-related total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) in the serum collected at autopsy. Japan).

The results of measuring the total cholesterol concentration are shown in Figure 4, compared to the normal group (about 76.90±4.59 mg/㎗), the control group (about 103.60±5.11 mg/㎗) increased total cholesterol in the blood, and the BK-021 experimental group It was confirmed that total cholesterol in blood (about 93.40±7.33 mg/㎗) decreased compared to the control group (about 103.60±5.11 mg/㎗).

In addition, the results of measuring the low-density lipoprotein cholesterol (LDL-C) concentration are shown in Figure 5, compared to the normal group (about 14.80 ± 1.41 mg / ㎗) in the blood of the control group low-density lipoprotein cholesterol (about 18.70 ± 1.03 mg / ㎗). ), and it was confirmed that the blood low-density lipoprotein cholesterol (about 16.90±1.55 mg/㎗) in the BK-021 experimental group decreased compared to the control group (about 18.70±1.03 mg/㎗).

<110> Vegut LAB Inc. Agency for Korea National Food cluster (AnFC) <120> Composition comprising Lactobacillus plantarum BK-021 for preventing or treating diabetes mellitus or vascular disease <130> 0504 <160> 1 <170> KoPatentIn 3.0 <210> 1 <211> 1506 <212> DNA <213> Unknown <220> <223> Lactobacillus plantarum <400> 1 tagagtttga tcctggctca ggacgaacgc tggcggcgtg cctaatacat gcaagtcgaa 60 cgaactctgg tattgattgg tgcttgcatc atgatttaca tttgagtgag tggcgaactg 120 gtgagtaaca cgtgggaaac ctgcccagaa gcgggggata acacctggaa acagatgcta 180 ataccgcata acaacttgga ccgcatggtc cgagtttgaa agatggcttc ggctatcact 240 tttggatggt cccgcggcgt attagctaga tggtggggta acggctcacc atggcaatga 300 tacgtagccg acctgagagg gtaatcggcc acattgggac tgagacacgg cccaaactcc 360 tacgggaggc agcagtaggg aatcttccac aatggacgaa agtctgatgg agcaacgccg 420 cgtgagtgaa gaagggtttc ggctcgtaaa actctgttgt taaagaagaa catatctgag 480 agtaactgtt caggtattga cggtatttaa ccagaaagcc acggctaact acgtgccagc 540 agccgcggta atacgtaggt ggcaagcgtt gtccggattt attgggcgta aagcgagcgc 600 aggcggtttt ttaagtctga tgtgaaagcc ttcggctcaa ccgaagaagt gcatcggaaa 660 ctgggaaact tgagtgcaga agaggacagt ggaactccat gtgtagcggt gaaatgcgta 720 gatatatgga agaacaccag tggcgaaggc ggctgtctgg tctgtaactg acgctgaggc 780 tcgaaagtat gggtagcaaa caggattaga taccctggta gtccataccg taaacgatga 840 atgctaagtg ttggagggtt tccgcccttc agtgctgcag ctaacgcatt aagcattccg 900 cctggggagt acggccgcaa ggctgaaact caaaggaatt gacgggggcc cgcacaagcg 960 gtggagcatg tggtttaatt cgaagctacg cgaagaacct taccaggtct tgacatacta 1020 tgcaaatcta agagattaga cgttcccttc ggggacatgg atacaggtgg tgcatggttg 1080 tcgtcagctc gtgtcgtgag atgttgggtt aagtcccgca acgagcgcaa cccttattat 1140 cagttgccag cattaagttg ggcactctgg tgagactgcc ggtgacaaac cggaggaagg 1200 tggggatgac gtcaaatcat catgcccctt atgacctggg ctacacacgt gctacaatgg 1260 atggtacaac gagttgcgaa ctcgcgagag taagctaatc tcttaaagcc attctcagtt 1320 cggattgtag gctgcaactc gcctacatga agtcggaatc gctagtaatc gcggatcagc 1380 atgccgcggt gaatacgttc ccgggccttg tacacaccgc ccgtcacacc gtgagagttt 1440 gtaacatcca atgtcggtgg ggtcacgcta agtcgtacca acgtcgcagt agagatgacg 1500 gacgaa 1506

Claims (5)

α-glucosidase inhibitory effect, blood total cholesterol reduction effect, and low-density lipoprotein cholesterol reduction effect, and Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P) comprising the gene sequence of SEQ ID NO: 1 Composition for the simultaneous prevention or treatment of diabetes and hyperlipidemia, characterized in that it contains a strain or a culture thereof. delete delete delete α-glucosidase inhibitory effect, blood total cholesterol reduction effect, and low-density lipoprotein cholesterol reduction effect, and Lactobacillus plantarum BK-021 (Lactobacillus plantarum BK-021, accession number KFCC11749P) comprising the gene sequence of SEQ ID NO: 1 Health functional food for preventing or improving diabetes and hyperlipidemia, characterized in that it contains a strain or a culture thereof.

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