JP2013158248A - New lactobacillus - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a new lactobacillus capable of producing a substance exhibiting pancreatic islet cell protecting effect and an insulin resistance improving effect, to provide an extracellular released substance derived from the lactobacillus, a pancreatic islet cell protecting agent, and an insulin resistance improving agent containing the extracellular released substance, and to provide a method for producing pickles by using the lactobacillus.SOLUTION: There is provided a new lactobacillus strain, Lactobacillus sp.TK63404 (Accession Number: NITE P-930).

Description

  The present invention relates to a novel lactic acid bacterium, an extracellularly-released substance derived from the lactic acid bacterium, an islet cell protective agent and an insulin resistance improving agent containing the extracellularly-released substance, and a method for producing pickles using the lactic acid bacterium. .

  In recent years, lifestyle-related diseases such as excessive intake of nutrients, lack of exercise, smoking, drinking, and stress have become problems in Japan. Examples of lifestyle-related diseases include obesity, gout, hypertension, diabetes, hyperlipidemia, heart disease, stroke, and cancer. Among them, diabetes causes various complications such as diabetic nephropathy, diabetic retinopathy, diabetic neuropathy, and the number of patients has been steadily increasing, which is a problem.

  Normally, blood sugar level rises after ingestion of food or the like, but insulin is secreted from the islet cells, and the sugar in the blood is taken into cells by the action, and the blood sugar level decreases. However, if insulin is not secreted from the islet cells, the blood glucose level is difficult to decrease. It is insulin-dependent diabetes (type I diabetes) that causes various disorders. The causes include autoimmune diseases and viral infections.

  In recent years, there has been an increase in non-insulin-dependent diabetes (type II diabetes) in which insulin is secreted, but the amount thereof is not sufficient, or the blood sugar level does not decrease due to low sensitivity to insulin. Of these, the latter is closely related to lifestyle disturbances. That is, when nutrients exceeding the basal metabolism are continuously taken up by cells and fat cells swell, substances that improve insulin function (adiponectin) are not released, but free fatty acids and inflammatory properties that make insulin function worse Cytokines such as TNF-α are secreted and insulin resistance becomes apparent.

  In addition, if you continue to eat and drink, the islet cells that are insulin-secreting cells may become inflamed and the amount of insulin released may decrease. In some cases, islet cells die and insulin cannot be released. Therefore, protecting pancreatic islet cells is very important for suppressing blood glucose levels.

  For insulin-dependent diabetes mellitus, no therapeutic means has been established other than injecting insulin, but various drugs have been researched and developed for non-insulin-dependent diabetes mellitus. However, synthetic drugs have side effects, and cannot be used for preventive purposes by taking them constantly, for example, from the stage where symptoms do not become apparent. Therefore, safer drugs are demanded from microorganisms that can be applied to foods.

  For example, Patent Document 1 discloses a therapeutic agent for diabetic complications, which contains a specific lactic acid bacteria culture or cells as active ingredients.

  Patent Document 2 discloses an adipocyte differentiation promoter containing a fermented wheat product produced by a specific lactic acid bacterium as an active ingredient.

  Patent Document 3 describes that a fermented product of grape skin and seeds by lactic acid bacteria has an effect on type II diabetes.

  Patent Document 4 discloses a hypoglycemic agent containing lactic acid bacteria as an active ingredient.

  Patent Document 5 discloses an antidiabetic agent comprising a fermented product of rice bran or brown rice flour by lactic acid bacteria as an active ingredient.

  Patent Document 6 describes that lactic acid bacteria and the like contained in the culture supernatant of lactic acid bacteria exhibit an action to suppress an increase in blood sugar after a meal.

  Patent Documents 7 to 8 disclose islet cell (β cell) protective agents containing serine, glycine, flavonoid compounds, and tannic acid as active ingredients.

JP 2003-252770 A JP 2008-179595 A International Publication No. 2008/093670 Pamphlet Japanese Patent Laid-Open No. 10-7777 Japanese Patent Laid-Open No. 9-40566 JP 2003-116486 A International Publication No. 2007/060924 Pamphlet JP 2008-7452 A

  As described above, drugs related to diabetes and pancreatic islet cell protective agents are being searched for from lactic acid bacteria that can also be used in foods and their cultures. However, since it is not easy to put it into practical use, a new one is always required.

  Then, this invention aims at providing the novel lactic acid bacteria which can produce the substance which shows an effect with respect to protection of an islet cell and insulin resistance. In addition, the present invention also provides an extracellular release substance derived from the lactic acid bacteria, an islet cell protective agent and an insulin resistance improving agent containing the extracellular release substance, and a method for producing pickles using the lactic acid bacteria. Objective.

  The inventors of the present invention have made extensive studies to solve the above problems. As a result, the inventors found that the novel lactic acid bacteria found by the present inventors released a substance having an extremely excellent effect on islet cell protection and insulin resistance into the culture solution, and completed the present invention. did.

  The novel lactic acid bacteria according to the present invention are Lactobacillus sp. It is TK63404 strain (Accession number: NITE P-930).

  The extracellularly released substance according to the present invention is Lactobacillus sp. It is derived from TK63404 strain (Accession number: NITE P-930). The extracellularly released substances are mainly Lactobacillus sp. TK63404 strain (accession number: NITE P-930) is contained in the culture solution after culturing.

  The extracellularly released substance exhibits a protective action on insulin cells secreting insulin and an insulin resistance improving action, and is derived from lactic acid bacteria and is safe. Therefore, the islet cell protective agent and insulin resistance improving agent Can be used as

  In addition, the method for producing pickles according to the present invention includes Lactobacillus sp. Adding raw vegetables to the seasoning liquid containing TK63404 strain (Accession Number: NITE P-930), or Lactobacillus sp. It includes a step of attaching raw material vegetables to a seasoning liquid containing an extracellular substance released from TK63404 strain (Accession Number: NITE P-930).

  The novel lactic acid bacterium and its extracellular release substance according to the present invention are safe ones that can be used in the production of foods, and exhibit excellent islet cell protecting action and insulin resistance improving action. Therefore, by continually ingesting foods using these, it becomes possible not only to suppress or treat diabetes but also to prevent it. As described above, the present invention is industrially excellent as an effective improvement means and prevention means for diabetes, which has been a problem in recent years.

FIG. 1 is a graph showing experimental results of the effect of the extracellularly released substance according to the present invention on blood insulin concentration. FIG. 2 is a graph showing experimental results of the effects of the extracellularly released substance according to the present invention on the blood glucose level. FIG. 3 is a graph showing the experimental results of the effect of the extracellularly released substance according to the present invention on the blood insulin concentration. FIG. 4 is a graph showing the experimental results of the effect of the extracellularly released substance according to the present invention on the blood glucose level. FIG. 5 is a fluorescent photograph showing the activity of pancreatic islet cells in untreated rats, control rats, and rats administered with the extracellular substance according to the present invention.

Lactobacillus sp., A novel lactic acid bacterium according to the present invention. The TK63404 strain is deposited with the depository as follows.
(I) Name and address of depositary institution Name: National Institute of Technology and Evaluation, Patent Microorganism Depositary Center Address: 2-5-8 Kazusa Kamashi, Kisarazu City, Chiba Prefecture, Japan
(Ii) Date of deposit: April 9, 2010 (iii) Deposit number: NITE P-930

  The morphological characteristics and biochemical properties of the novel lactic acid bacteria according to the present invention are as follows.

  As the results of Examples described later, it can be said that the novel lactic acid bacteria according to the present invention are most closely related to Lactobacillus parabrevis from the base sequence information of 16S rDNA. However, according to the results of the DNA-DNA hybridization test, it can be concluded that the genus species of the novel lactic acid bacteria according to the present invention are different from Lactobacillus parabrevis. That is, the species of lactic acid bacteria according to the present invention is a novel one that is not classified into any known genus species.

  The extracellularly released substance according to the present invention is Lactobacillus sp. It is derived from TK63404 strain (Accession number: NITE P-930). The novel lactic acid bacterium according to the present invention produces a substance having an action of protecting pancreatic islet cells and improving insulin resistance, and releases it outside the microbial cell. Such extracellularly released substances can be collected from the culture medium after culturing the lactic acid bacteria according to the present invention. Alternatively, the culture solution can be used as it is.

  The term “derived from the bacterium of the present invention” means that it is produced from the bacterium gene of the present invention that directly or indirectly encodes a substance having an action of protecting pancreatic islet cells or improving insulin resistance. Therefore, for example, not only the secretion released spontaneously from the bacterium of the present invention, but also the secretion released by the bacterium of the present invention modified to promote the expression of the gene, Secretions released from the transformed microorganism into which the above gene has been introduced are also included in the extracellularly released substance according to the present invention. However, from the viewpoint of the confirmed safety, the substance released from the cells according to the present invention is preferably a substance that is produced by the untreated bacteria of the present invention and released outside the cells.

  What is necessary is just to select suitably the component which should be added to the culture solution of the novel lactic acid bacteria based on this invention. For example, carbon sources such as glucose and fructose; general nutritional components such as yeast extract and protein hydrolysates; amino acids and salts thereof such as sodium glutamate; mineral components such as magnesium sulfate; pH adjusters such as lactic acid and sodium acetate What is necessary is just to add. However, the present inventors have found that the novel lactic acid bacteria according to the present invention grow well in a medium containing glutamic acid.

  In addition, although a solid culture medium can be used for culture | cultivation of the novel lactic acid bacteria based on this invention, it is preferable to use a liquid culture medium (culture solution) for isolation | separation of a substance released outside a microbial cell.

  The pH of the culture solution of the novel lactic acid bacteria according to the present invention is preferably adjusted to about 4.0 or more and 6.0 or less, and more preferably 4.5 or more and 5.5 or less.

  The culture conditions for the novel lactic acid bacteria according to the present invention may be adjusted as appropriate. For example, the culture may be performed at 20 ° C. or higher and 40 ° C. or lower for 10 hours or longer and 50 hours or shorter. The culture temperature is more preferably about 25 ° C. or more and 35 ° C. or less. Either stationary culture or shaking culture may be used, but stationary culture is more preferable.

  The culture solution after cultivating the novel lactic acid bacteria according to the present invention is a substance released from the novel lactic acid bacteria to the outside of the microbial cells, and includes substances that have a protective action on islet cells and an action to improve insulin resistance. . Therefore, the culture solution can be used as it is as an islet cell protective agent or an insulin resistance improving agent. Alternatively, it may be used after filtering insoluble matter such as bacterial cells, or may be used after processing such as concentration, freeze drying, spray drying and the like. In addition, the culture solution includes a seasoning solution for producing pickles, in which the novel lactic acid bacteria according to the present invention are cultured. The pickles produced with such a seasoning liquid contain the above-described extracellularly released substance, and thus exhibit an action for protecting pancreatic islet cells and an action for improving insulin resistance.

  Further, the extracellularly released substance according to the present invention may be further purified by a conventional method such as salting out, column chromatography, ion exchange resin, recrystallization and the like.

  The extracellular release component of the novel lactic acid bacterium according to the present invention has an action of protecting islet cells and improving insulin resistance, and therefore can be used as an active ingredient of an islet cell protecting agent and an insulin resistance improving agent. As described above, since the extracellularly released component is released into the culture solution during the culture of the novel lactic acid bacteria, the culture solution itself, or the concentrated solution or dried product of the culture solution is used to improve the islet cell protective agent or insulin resistance. It can be used as an agent. The dried culture solution may be further formulated.

  The form of such a preparation is not particularly limited and may be appropriately selected. For example, solid preparations such as powders, granules, tablets, capsules and coating preparations; liquid preparations such as solutions, suspensions and aerosols can be used. Depending on the formulation, excipients, binders, disintegrants, lubricants, coloring agents, flavoring agents, flavoring agents, coating agents, solvents such as distilled water and physiological saline, emulsifiers, antioxidants, etc. Stabilizers and pH adjusters may be added.

  The novel lactic acid bacteria according to the present invention may be used in the production of pickles.

  Vegetables used as a raw material for such pickles are not particularly limited as long as they are common as materials for shallow pickles. For example, cucumbers, bitter gourd, zucchini, cucumber fruits such as winter melon; solanaceous fruits such as pepper, tomato, eggplant, pepper; Ginger family stem vegetables such as bamboo shoots; Brassicaceae root vegetables such as turnips, zaisai and radishes; Aceraceae root vegetables such as carrots; Ginger family vegetables such as ginger; Examples include cruciferous leafy vegetables such as cabbage, komatsuna, santosai, tarsai, takana, chingensai, Nozawana, Chinese cabbage, spinach, mizuna, and fresh greens; lily family leafy vegetables such as leek;

  As a raw material vegetable, naturally, what was washed after harvesting is preferable. In addition, the raw vegetables may be cut into an appropriate size in advance.

  Next, the raw material vegetables that have undergone the above steps may be submerged before being added to the seasoning liquid. Although the said process is arbitrary, the cell of a raw material vegetable is spin-dry | dehydrated by a soaking process, a structure | tissue will become flexible, and a seasoning liquid will osmose | permeate vegetables easily. An example of the soaking process is to cover the raw vegetables with sodium chloride and leave it still for a whole day and under pressure.

  Next, the raw vegetables are added to the seasoning liquid to make pickles. In that case, the novel lactic acid bacteria based on this invention are used. Specifically, the lactic acid bacteria of the present invention may be added to the seasoning liquid, or the lactic acid bacteria of the present invention may be pre-cultured and applied to the raw material compound together with the culture liquid. Alternatively, the lactic acid bacteria of the present invention may be pre-cultured, separated into solid and liquid, and the liquid part containing the extracellularly released substance according to the present invention may be used as part of the seasoning liquid. The addition of the raw material vegetable to the seasoning liquid means that the raw material vegetable is in sufficient contact with the seasoning liquid. For example, the raw material vegetable may be completely immersed in the seasoning liquid. It may be soaked, or the mixture of raw vegetables and seasoning liquid may be shaken or stirred.

  The seasoning liquid is not particularly limited as long as it is used for the production of shallow pickles. For example, salt and sodium chloride; amino acids such as sodium glutamate, glycine and alanine; nucleic acids such as guanylic acid and inosinic acid; sugar, isomerized liquid sugar, starch syrup, oligosaccharide, stevia, Sweeteners such as saccharin, sorbitol, erythritol, xylitol, maltitol; pH adjusters such as citric acid, lactic acid, acetic acid, sodium acetate; soy sauce, fish sauce, acid-decomposed amino acid solution, protein hydrolyzate, animal and plant extracts, yeast extract And seasonings such as mirin.

  The shallow pickles obtained above may be packaged in small quantities while the vegetables are attached to the seasoning liquid, or may be products by removing the seasoning liquid from the raw vegetables. Since the extracellularly released substance according to the present invention is contained in the seasoning liquid or penetrates into the vegetables, in any aspect, the protective action of islet cells and the action of improving insulin resistance are exhibited.

  EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples. However, the present invention is not limited by the following examples, but may be appropriately modified within a range that can meet the purpose described above and below. Of course, it is possible to implement them, and they are all included in the technical scope of the present invention.

Example 1
(1) Isolation of lactic acid bacteria according to the present invention During the production of pickled products, a unique example was seen in which the pH of the seasoning liquid increased from the standard product as the fermentation progressed. Several strains were isolated from the pickled product of this example and added to a standard product to identify lactic acid bacteria exhibiting the above-mentioned unique properties.

(2) Molecular phylogenetic analysis According to a conventional method, the base sequence of 16S rDNA of lactic acid bacteria identified in (1) above was determined.
Using the obtained base sequence information, a homology search was performed, and 30 types of bacterial groups with high homology rates and presumed to be closely related were selected. At this time, Apollon 2.0 (Techno Suruga Lab) is used as software, and Apollon DB-BA 5.0 (Techno Suruga Lab) and international base sequence database (GenBank / DDBJ / EMBL) are used as databases. It was.

  Next, 16S rDNA information of the reference strain of the fungal group estimated to be closely related by homology search was obtained, and molecular phylogenetic analysis of lactic acid bacteria according to the present invention was performed by the neighborhood binding method. At that time, CLUSTAL W (Thompson, JD, et al., Nucleic Acids Research, 22, pp. 4673-4680 (1994)), which is an alignment software, is used for multiple alignment, and a computer for creating a molecular phylogenetic tree is used. The software MEGA 3.1 (Kumar, R. et al., Briefings in Bioinformatics, 5, pp. 150-163 (2004)) was used.

  As a result of the homology search, the 16S rDNA base sequence of the lactic acid bacterium according to the present invention shows high homology to the same base sequence of the genus Lactobacillus. Parabrevis strain LMG11984 showed the highest homology. In addition, as a result of molecular phylogenetic analysis, the lactic acid bacteria according to the present invention are L. A cluster was formed with parabrevis. Lactic acid bacteria according to the present invention and L. The cluster with parabrevis was supported with a bootstrap value as high as 98%, indicating that both are closely related. However, a distance was recognized between the two.

(3) DNA-DNA Hybridization Test Next, it was concluded that L. Whether or not the lactic acid bacterium according to the present invention is the same species with respect to ATCC 53295, which is a reference strain of parabrevis, was confirmed by a DNA-DNA hybridization test.

  Specifically, the total DNA of both was extracted and purified, Yoshiaki Kawamura, Bacterial phylogeny and identification method, 18th Annual Meeting of the Japanese Society for Bacteriology, 55, pp. 545-584 (2000) and edited by Kenichiro Suzuki et al., “Microbial Classification / Identification Experiment Method” Springer Fairlark Tokyo, pp. 11-28. With reference to 34-47 (2001), a DNA-DNA hybridization test was performed by the microplate method. A fluorescence plate reader (Genios, TECAN) was used for the measurement of fluorescence intensity. The measurement was performed 3 times, and the average value was calculated.

  As a result, the homology value was 14 to 18%, and the average value was 16%. Bacteria are defined as homologous when the DNA-DNA homology value is 70% or more (Wayne LG et al., Int. J. Syst. Bacteriol., 37, pp. 463-464 (1987)). ). Therefore, it was demonstrated that the lactic acid bacteria according to the present invention are a new species.

Example 2 Extracellular Release Substances According to the Present Invention (1) Culture A solution (200 μL) containing the lactic acid bacteria specified in Example 1 (1) above was added to a seed culture solution (10 mL) shown in Table 2, and 30 ° C. For 24 hours. The culture solution (1 mL) was added to the same-type mother culture solution (100 mL) and cultured at 30 ° C. for 18 hours. The culture broth (total amount) was added to the main culture broth (1900 mL) shown in Table 2 and cultured at 30 ° C. for 24 hours.

(2) Extracellular release substance 50% fermented lactic acid aqueous solution (73.1 mL) was added to the culture solution (1800 mL) obtained in (1) above. The mixture was stirred several times to cause foaming. The temperature of the mixed solution was raised to 85 ° C., kept as it was for 10 minutes, and then cooled to room temperature. The mixed solution was filtered with a filter paper (Advantech Toyo Co., Ltd., No. 101) to obtain a clear filtrate. The filtrate was concentrated under reduced pressure at 40 ° C. and 30 to 60 hPa until the Brix sugar content reached 50 ± 2. The obtained solution was used as an extracellularly released substance solution in the following experiments.

Example 3 Protective effect test on islet cells (single administration)
10 healthy rats (SPF) male rats (manufactured by SLC Japan, 8 weeks old) were preliminarily raised for 1 week. The group was divided into two groups: a control group and a group administered with an extracellular substance.

  First, approximately 100 μL of blood was collected from the tail vein of Wistar rats without anesthesia. Subsequently, a mixture of 5 mL of a sample and 5 mL of 40% glucose (manufactured by Wako Pure Chemical Industries, Ltd.) (glucose equivalent: 2 g) was orally administered using a plastic sonde (FUCHIGAMI oral administration sonde). As samples, water was administered to the control group, and 5 mL of the extracellularly released substance solution obtained in Example 2 was administered to the extracellularly released substance-administered group. The time immediately after administration was set to 0 minutes, and blood was collected over time at 30, 60, 90, and 120 minutes thereafter, and the serum insulin concentration and blood glucose level were measured. The insulin concentration was quantified by ELISA using an insulin measurement kit (manufactured by Morinaga Biochemical Laboratories) after centrifugation of the collected blood and collecting the serum. The blood glucose level was measured using a self-test glucose measuring device (Arkray, Glucocard G + Meter GT-1820). The measurement result of serum insulin concentration is shown in FIG. 1, and the measurement result of blood glucose level is shown in FIG.

  As shown in FIG. 1, in the control rats, the insulin concentration significantly increases immediately after glucose administration. That is, in the control rat, a large amount of insulin is secreted in response to administration of glucose. On the other hand, the amount of insulin secreted is almost constant in the rats administered with the extracellularly released substance solution according to the present invention. However, as shown in FIG. 2, the blood glucose level is almost the same between the control group and the extracellular substance-released substance administration group. From the above results, when the extracellularly released substance according to the present invention was administered, insulin secretion was suppressed and pancreatic islet cells were protected despite administration of a large amount of glucose, and the blood glucose level was suppressed. It can be seen that the resistance is relaxed. Therefore, it was demonstrated that the extracellularly released substance according to the present invention exhibits an action for protecting pancreatic islet cells and improving insulin resistance.

Example 4 Protective effect test on islet cells (long-term administration)
(1) Measurement of blood insulin concentration and blood sugar level After pre-breeding 18 Wistar (SPF) male rats (manufactured by SLC Japan, 8 weeks old) which are healthy model animals, the health status was confirmed. Nine animals were divided into two groups, a control group and an extracellularly administered substance administration group, so that the body weight was equal.

  Using a plastic sonde (FUCHIGAMI orally administered sonde), a mixture of 5 mL of sample and 5 mL of 40% glucose (manufactured by Wako Pure Chemical Industries, Ltd.) (glucose equivalent: 2 g) was orally administered every day and reared for 6 weeks. As samples, water was administered to the control group, and 5 mL of the extracellularly released substance solution obtained in Example 2 was administered to the extracellularly released substance-administered group. In the first week, the third week, and the fifth week from the start of breeding, a glucose tolerance test in which glucose alone was administered once was separately performed. Specifically, the time immediately after administration of the same amount of glucose as described above was set to 0 minutes, and thereafter blood was collected over time of 30, 60, 90, and 120 minutes, and the serum insulin concentration was determined in the same manner as in Example 3 above. The blood glucose level was measured. The measurement result of serum insulin concentration is shown in FIG. 3, and the measurement result of blood glucose level is shown in FIG.

  As shown in FIG. 3, compared to the control group, the amount of insulin secreted in the extracellularly administered substance administered group was reduced as the breeding period was increased. However, as shown in FIG. 4, the blood glucose level is almost the same between the control group and the extracellular substance-released substance administration group. From the above results, when the extracellularly released substance according to the present invention was administered, insulin secretion was suppressed and pancreatic islet cells were protected despite administration of a large amount of glucose, and the blood glucose level was suppressed. It can be seen that the resistance is relaxed. Therefore, it was demonstrated that the extracellularly released substance according to the present invention exhibits a protective action on islet cells and an improvement action on insulin resistance even in a long-term administration test.

(2) Confirmation of islet cell protective effect Furthermore, in order to directly evaluate the islet cell protective effect of the extracellularly released substance according to the present invention, an islet cell activity test was performed.

Specifically, after 9 healthy Wistar (SPF) male rats (manufactured by SLC Japan, 8 weeks old), which are healthy model animals, were preliminarily raised for 1 week, the health condition was confirmed and the body weight was equalized. The animals were divided into three groups, that is, an untreated rat, a control group, and an extracellular release substance administration group. The control group and the exogenous substance-administered group were treated in the same manner as in Example 4 (1). The representative rat was sacrificed on the sixth week of breeding, and the pancreas was removed. The pancreas was also removed from the untreated rats after one week of preliminary breeding. After the excised pancreas was sliced to a thickness of 400 μm, the pancreas was immersed in an aeration solution, and a substrate on which a GABA amino group transferase (GABase) was previously loaded was placed to advance the enzyme reaction. GABase catalyzes GABA to succinic acid and produces NADPH in the presence of nicotinamide adenine dinucleotide phosphate (NADP ⁺ ) and α-ketoglutaric acid. Since NADPH emits fluorescence, the activity of pancreatic islet cells can be measured by measuring the fluorescence intensity. The results are shown in FIG.

  In the photograph of FIG. 5, the portion with high fluorescence intensity is represented with short wavelength light, and the portion with low fluorescence intensity is represented with long wavelength light. Therefore, the low activity portion becomes blue and the high activity portion becomes red. As shown in FIG. 5, in the pancreas slices of untreated rats, there are many portions showing red to green, and it can be said that the whole is highly active. On the other hand, in the control rat, most of the fluorescence is blue to green. The cause is considered to be that the activity of the islet cells is decreased due to the excessive secretion of insulin probably due to a high glucose load. However, even in the case of rats administered with an extracellular release substance solution, a highly active portion from green fluorescence to red fluorescence is observed even though a high glucose load is applied. Thus, it has been proved that the extracellular substance released according to the present invention can directly protect the islet cells.

Claims (7)

  Lactobacillus sp. TK63404 strain (Accession number: NITE P-930).   Lactobacillus sp. Extracellular release substance characterized by being derived from TK63404 strain (Accession number: NITE P-930).   Lactobacillus sp. The extracellularly released substance according to claim 2, which is contained in a culture solution after culturing TK63404 strain (Accession No .: NITE P-930).   An islet cell protective agent comprising the extracellular substance released according to claim 2 or 3.   An insulin resistance-improving agent comprising the extracellular substance released according to claim 2 or 3.   Lactobacillus sp. A method for producing pickles characterized by including a step of adding raw material vegetables to a seasoning liquid containing TK63404 strain (accession number: NITE P-930).   Lactobacillus sp. A method for producing pickles characterized by including a step of attaching raw vegetables to a seasoning liquid containing an extracellular substance released from TK63404 strain (accession number: NITE P-930).