JP2007126399A - Composition for increasing glutathione - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a composition for oral ingestion capable of increasing in vivo glutathione concentration of human or animals utilizing a food, a feed, a cosmetic or a medicine by adding the composition to the food, the feed, the cosmetic or the medicine. <P>SOLUTION: The present invention provides the composition for oral ingestion containing a lactic acid bacterium-containing material as an active ingredient. Glutathione concentration in the tissue/in the cell can be maintained or increased by ingesting the composition, and oxidative stress deeply involving lifestyle-related diseases, their precursor state, etc., can be protected or alleviated thereby. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to a composition for oral consumption that can contain a lactic acid bacterium-containing substance as an active ingredient and can increase glutathione concentration in vivo. More specifically, the composition for oral consumption has the effect of increasing the in vivo glutathione concentration of humans or animals by adding lactic acid bacteria, which are the active ingredients of the present invention, to foods, feeds, cosmetics, and pharmaceuticals. Related to things.

  In recent years, with rapid changes in social life and aging, there are concerns about an increase in diseases including lifestyle-related diseases such as diabetes and its complications, hypertension, arteriosclerotic diseases, Alzheimer's disease and the like. It has been clarified that oxidative stress is involved in processes such as the establishment, onset, and progression of these many diseases.

  In addition, the following are known as diseases considered to be caused by in vivo oxidation. In other words, brain / neurological diseases include cerebral edema, traumatic epilepsy, cerebral ischemia, Parkinson's disease, spinal cord injury, etc., and eye diseases include cataract, retinal degeneration, retinopathy of prematurity, etc., respiratory system Diseases include bronchial asthma, pulmonary emphysema, pulmonary fibrosis, adult respiratory distress syndrome (ARDS), respiratory distress syndrome (IRDS), etc., cardiovascular diseases such as myocardial infarction, and digestive diseases Ulcerative colitis, stress gastric ulcer, pancreatitis, gastrointestinal mucosal disorder, etc., skin diseases include ultraviolet rays, atopic dermatitis, burns, frostbite, bed sores, etc., kidney diseases include nephritis, renal failure Other diseases include cancer, collagen disease, rheumatism, autoimmune disease, Behcet's disease and the like. Aging, skin wrinkles and dementia due to aging are also considered to be due to in vivo oxidation.

  Substances that prevent or reduce such in vivo oxidation include natural antioxidants such as ascorbic acid (vitamin C) and α-tocopherol (vitamin E), and phenols such as BHA (butylhydroxyanisole) and BHT (butylhydroxytoluene). Synthetic antioxidants are known. Antioxidant effects have also been found in lactic acid bacteria extracts, including the antioxidant action of Streptococcus lactis lister cell digests (Patent Document 1), the antioxidant action of Lactobacillus gasseri cells and their cultures. (Patent Document 2).

  However, a major problem in the treatment of diseases with these antioxidants is that there are many doses that can show efficacy, and with such large doses, side effects occur with a higher probability. These antioxidants are expected to be able to suppress the occurrence and exacerbation of diseases by reducing oxidative stress. Development of a highly effective in vivo oxidative stress reducing agent (antioxidant) has been desired.

  On the other hand, it is known to measure the degree of in vivo glutathione concentration reduction as an index for evaluating the oxidative stress state. Glutathione, which is a peptide composed of glutamic acid, cysteine, and glycine, is a multifunctional biological component that is contained in cells at a high concentration of 1 to 10 mM, and is involved in the redox system in the living body and reactive oxygen species in the living body. It has an antioxidant action to trap Furthermore, glutathione plays an important role in eliminating hydrogen peroxide and lipid peroxide as a substrate of glutathione peroxidase, and is also involved in detoxification metabolism as a substrate of glutathione-S-transferase. In addition, in aging accompanying aging, a decrease in tissue / intracellular glutathione concentration is observed, so increasing the glutathione concentration in vivo can prevent or treat diseases caused by many oxidative stresses, It is considered significant for the prevention of aging.

  As a method for increasing the glutathione concentration in vivo, it is easily considered that glutathione is orally ingested, but it is rapidly degraded by glutathione-degrading enzyme (transpeptidase) in the digestive tract or liver, or a high dose of glutathione is ingested. However, it is known that glutathione taken orally does not directly contribute to the increase of glutathione in the living tissue because no increase in blood glutathione concentration is observed (Non-patent Document 1).

  Thus, in response to the fact that it is pointed out that administration of glutathione itself is not effective, the following drug development attempts can be cited for the purpose of solving this problem. That is, an acyl derivative of γ-L-pyroglutamyl-L-cysteine, which is a compound that promotes biosynthesis of endogenous glutathione, is an intracellular oxidation state caused by heavy alcohol consumption, xenobiotics, radiation damage, liver disease, pharmaceuticals And other compound intoxications, as well as reductions and deficiencies in glutathione, such as acute and chronic neurological diseases, immune system dysfunction, and other oxidative tissue disorders, especially pathological conditions involving disorders caused by excessive free radicals There is a disclosure that it is effective in the treatment of various diseases caused by (patent document 3).

  Patent Document 4 discloses that γ-L-glutamyl-L-cysteine methyl ester is a compound that increases the tissue or intracellular glutathione concentration as a compound for treating and preventing various diseases including cataract, liver disease and kidney disease. It is disclosed to be effective.

  Further, tissue glutathione is disclosed as an S-lower fatty acid glutathione derivative as an anti-inflammatory, antiallergic, liver disorder inhibitor (Patent Documents 5 and 6), and a sugar-containing glutathione derivative as a liver protectant (Patent Document 7). Has been shown to be effective in the treatment and prevention of a wide range of diseases.

  However, the main purpose of such treatment is to deal with urgent or severe cases. From the viewpoint of daily health management aimed at improving the quality of life (QOL), safe food that can maintain health by measuring the increase in glutathione in tissues that have become low levels using familiar foods as a raw material There is an urgent need to develop ingestion compositions.

  Compounds derived from natural products that increase intracellular glutathione concentration include silymarin (Non-patent document 2), cyanohydroxybutene (Non-patent document 3), apocynin (Non-patent document 4), rubrofusarin glycoside (patent) Document 8) is already known.

  On the other hand, an antioxidant effect has also been found in lactic acid bacteria extracts. That is, it is disclosed as an antioxidant action of Streptococcus lactis lister cell digest (Patent Document 9), an anti-oxidant action of Lactobacillus gasseri cell culture and its culture (Patent Document 10). Regarding the extract of lactic acid bacteria, the effect of increasing glutathione concentration has not been known so far.

Various problems also exist with respect to in vivo glutathione increasing agents derived from natural products. Conventionally, plant extracts such as those known as Kampo, herbs, or folk medicines have medicinal effects other than glutathione increase, and side effects due to the medicinal effects may occur. Therefore, a safe glutathione increasing agent that has been conventionally used as a food is desired. Moreover, since the conventional natural glutathione increasing agent was cultivated and used for the purpose of extraction, there also existed a problem that cost was high.
JP-A-3-4768 JP 2003-253262 A WO94 / 12527 pamphlet JP-A-1-26516 WO91 / 12262 pamphlet WO91 / 16337 pamphlet Japanese Patent Laid-Open No. 9-25292 Japanese Patent Laid-Open No. 2004-2231 JP-A-3-4768 JP 2003-253262 A Eur. J. Clin. Pharmacol. 43, 667, 1992 Planta Med. 55, 420, 1989 Toxicol. Appl. Pharmacol. 123, 257, 1993 FEBS Lett. 443, 235, 1999

  The object of the present invention is to increase the intracellular glutathione concentration by using highly safe and familiar food materials, and is effective in preventing and treating various organ functions and various diseases caused by glutathione deficiency. An object of the present invention is to provide an orally ingestible composition for increasing tissue or intracellular glutathione, which is objective, safe and can be produced at low cost.

  The present inventors have extensively searched for an active ingredient that increases the intracellular glutathione concentration suitable for the above purpose using materials having dietary experience, and found that the lactic acid bacteria-containing material exhibits an effective effect. The present invention has been completed.

  The present invention can maintain or increase glutathione concentration in tissues / cells by ingesting lactic acid bacteria-containing substances, thereby protecting or reducing oxidative stress that is deeply related to lifestyle-related diseases and their precursor states. Can be relaxed. That is, it is effective for preventing and treating various diseases caused by this oxidative stress, and also effective for preventing aging.

BEST MODE FOR CARRYING OUT THE INVENTION

  In the composition for oral intake for increasing glutathione of the present invention, a lactic acid bacterium-containing material is used as an active ingredient. The lactic acid bacterium-containing material includes lactic acid bacterium cells, lactic acid bacterium suspension, lactic acid bacterium culture (including microbial cells, culture supernatant, and medium components), lactic acid bacterium culture solution (obtained by removing solid content from the bacterium culture), lactic acid bacterium Examples thereof include fermented products (lactic acid bacteria beverages, sour milk, yogurt, fermented fruit vegetables, fermented cereals, etc.). The lactic acid bacteria may be cultured under known conditions using an arbitrary medium, and need not be based on a special culture method. Moreover, as this microbial cell, any of dead cell body, a microbial cell destruction material, a microbial cell extract, etc. other than a living microbial cell can be used.

  Lactic acid bacteria have been widely used in food production since ancient times, and when producing lactic acid bacteria beverages, dairy products such as fermented milk, cheese and fermented butter, agricultural food products such as pickles and kimchi, and livestock meat products such as fermented sausages and fermented salami. It is used for. It is also used as a bread starter. Furthermore, recently, physiological effects such as the intestinal regulating effect of lactic acid bacteria have been clarified one after another, and development has been made to use lactic acid bacteria cells themselves or lactic acid bacteria cultures as materials for health foods and pharmaceuticals. Thus, the utilization of lactic acid bacteria is diverse, and in particular, the Lactobacillus pentosus S-PT84 strain used in the examples of the present invention is one of the traditional pickles in Kyoto, “Shiba. It is a type of lactic acid bacterium isolated from “Kizuke” and has been shown to have immunomodulatory effects such as immunostimulatory action and antiallergic action.

  Examples of lactic acid bacteria that can be used in the present invention include Lactobacillus, Lactococcus, Streptococcus, Bifidobacterium, Leuconostoc and the like. I can do it.

  The active ingredient of the present invention is used as a composition for oral intake having an action of increasing glutathione, and it is particularly preferable to use it as a pharmaceutically formulated composition. For example, food / beverage products, pharmaceuticals, etc. can be mentioned. When used in foods and drinks, it is preferable to implement as a health food having a glutathione increasing action, a food for specified health use, or a dietary supplement. In formulating, known additives commonly used in pharmaceutics can be blended. These additives include sweeteners, acidulants, stabilizers, preservatives, colorants, fragrances, vitamins, etc., and these additives are appropriately blended, mixed, and tablet, granule, granule, etc. by a conventional method. It can be provided in the form of powder, capsule, liquid, jelly, cream, beverage, seasoning, processed food and the like.

  Moreover, when using as a pharmaceutical, a dosage form can be selected according to a therapeutic purpose, an administration route, etc. For example, in addition to oral preparations such as tablets, pills, capsules, granules, powders, solutions, suspensions, emulsions, syrups and the like, suppositories, injections and the like can be mentioned, but the invention is particularly limited to these. Is not to be done. In addition, for formulation as a pharmaceutical, a diluent, excipient, etc. that are usually used such as a filler, a bulking agent, a binder, a moisturizer, a disintegrant, a surfactant, and a lubricant are added as necessary. Can be used. In addition, additives such as coloring agents, preservatives, fragrances, flavors, sweeteners, and other active ingredients may be contained in the pharmaceutical preparation. For example, oral administration is simple as an administration route of the pharmaceutical product, but rectal administration, enteral administration, and the like can be mentioned, and the administration route is not particularly limited.

  In the composition of the present invention, in addition to the active ingredient lactic acid bacteria-containing material, nutrients, vitamins, antioxidants, oligosaccharides, dietary fiber, amino acids, fatty acids, enzymes, energy supply substances, etc. are blended, Combined effects can also be achieved.

  The composition for oral ingestion of the present invention may be ingested before or after a meal, during a meal, or between meals, and the frequency of ingestion is not particularly limited, but is usually 1 to 3 times a day, preferably, Take three times a day. The amount of intake is not particularly limited, but it is usually sufficient that an active ingredient of 0.02 mg to 50,000 mg, preferably 0.2 to 200 mg per day can be ingested.

EXAMPLES The present invention will be specifically described below with reference to examples, but the present invention is not limited to these examples.
[Example 1] Effect of increasing intracellular reduced glutathione by in vitro stimulation Three types of lactic acid bacteria (Lactobacillus pentosus, Lactobacillus casei or Lactobacillus paracasei, of which Lactobacillus pentosus, FERM ABP-10028 (Lactobacillus pentosus SAM2336, S-PT84) deposited at the Deposit Center was cultured in MRS medium, and the cells were collected. After washing the cells, they were suspended in distilled water and sterilized by heating (95 ° C., 5 minutes), and the cells after lyophilization were regarded as dead cells.

The effect of increasing intracellular reduced glutathione was measured using mouse bone marrow-derived dendritic cells (BMDC). Specifically, bone marrow cells were collected from BALB / C mice (10-week-old male) and cultured in a 5% CO ₂ incubator for 5 days using RPMI medium containing 10% FBS. Differentiated. From the beginning of the culture, 20 ng / ml granulocyte macrophage colony stimulating factor (GM-CSF) and 20 ng / ml interleukin 3 (IL-3) were added, and the medium was exchanged as necessary. After inoculating 1 × 10 ⁶ cells (1 mL) of immature dendritic cells into a 12-well plate, lactic acid bacteria dead cells (0.1 μg / ml, 1 μg / ml, 10 μg / ml) were added to each well, Cells were harvested 48 hours later using a rubber policeman. 80 μl of 10 mmol / l HCl was added to 1 × 10 ⁶ cells washed with PBS, and the cell membrane was broken by repeating freezing and thawing twice. Furthermore, 20 μl of 5% sulfosalicylic acid (SSA) was added, centrifuged at 8,000 × g for 10 minutes, and the supernatant was collected to obtain a glutathione concentration measurement sample. The reduced glutathione concentration was measured by an enzyme recycling method using a Total Glutathione Quantification Kit (manufactured by Dojindo).

The measurement result of the reduced glutathione concentration is shown in FIG. Thereby, the effect of increasing the intracellular reduced glutathione concentration of lactic acid bacteria was recognized.
[Example 2] Intracellular / intracellular reduced glutathione increasing effect by oral ingestion Lactobacillus pentosus S-PT84 strain (dead cells) equivalent to 0.2 mg / day, equivalent to 2 mg / day, or equivalent to 20 mg / day As such, it was suspended in drinking water and allowed to freely ingest 5 each of C57BL / 6 mice (7 weeks old, male). In addition, a control group (5 animals) that did not take the S-PT84 strain was provided. Three days after the start of ingestion, 4.05% thioglycolate was intraperitoneally administered. After four days, blood was collected from the carotid artery, and peritoneal macrophages were collected and the liver was removed. Serum (100 μl) was collected from the blood by centrifugation, 50 μl of 5% SSA was added, centrifuged at 8,000 × g for 10 minutes, and the supernatant was collected to obtain a reduced glutathione concentration measurement sample. The liver was homogenized in 10 times the amount of 5% SSA, centrifuged at 8,000 × g for 10 minutes, and the supernatant was collected to obtain a reduced glutathione concentration measurement sample. The peritoneal macrophages were washed with PBS, 80 μl of 10 mmol / l HCl was added to 1 × 10 ⁶ cells, and the cell membrane was broken by repeating freezing and lysis twice. Further, 20 μl of 5% SSA was added and centrifuged at 8,000 × g for 10 minutes, and the supernatant was collected to obtain a reduced glutathione concentration measurement sample. Serum, liver, and peritoneal macrophage intracellular reduced glutathione concentrations were measured by an enzyme recycling method using Total Glutathione Quantification Kit (manufactured by Dojindo).

The measurement results of the reduced glutathione concentration are shown in FIG. 2, FIG. 3, and FIG. As is clear from these figures, the tissue / intracellular glutathione concentration increased in the S-PT84 intake group compared to the control group, and the effect was most prominent in the 2 mg / day equivalent intake group. .
[Example 3] Inhibitory effect on reduction of tissue / intracellular reduced glutathione accompanying restraint stress Nine C57BL / 6 mice (7-week-old male) were divided into 4 groups so that the average body weight of each group was almost the same. It was. The group composition was a control group (3 mice), a stress load group (3 mice), an S-PT84 strain intake + stress load group (3 mice). Lactobacillus pentosas S-PT84 strain S-PT84 strain intake group was allowed to ingest S-PT84 strain (dead cells) freely with drinking water (equivalent to 2 mg / day) for 7 days. On day 4 after ingestion, 4.05% thioglycolate was intraperitoneally administered to induce peritoneal macrophages. On the seventh day after ingestion, the stress load group and the S-PT84 strain intake + stress load group were immersed in water once, then placed in a 50 mL polyethylene tube with an air hole at the tip and restrained for 24 hours. The control group was fasted and fasted for 24 hours. Thereafter, blood was collected from the carotid artery of the animal, and peritoneal macrophages were collected. Serum and peritoneal macrophage intracellular reduced glutathione concentrations were measured by the same method as in Example 2.

  As shown in FIGS. 5 and 6, when restraint stress was applied, the reduced glutathione concentration in both serum and peritoneal macrophage cells decreased. On the other hand, it was revealed that the reduced glutathione concentration similar to that in the control group was maintained in the S-PT84 strain intake group + stress load group, which was significantly improved as compared with the stress load group.

  The present invention provides a composition for oral ingestion containing a lactic acid bacterium-containing substance as an active ingredient. By ingesting this composition, the concentration of glutathione in a tissue / cell can be maintained or increased. Can protect or alleviate oxidative stress that is deeply related to lifestyle-related diseases and their precursors. That is, the composition of the present invention is effective for the prevention and treatment of various diseases caused by this oxidative stress as food and drink, food additives, health foods, or pharmaceuticals, and is also effective for the prevention of aging. A highly safe composition is provided.

It is a graph which shows the mouse | mouth dendritic cell glutathione density | concentration increase by in vitro stimulation. It is a graph which shows the glutathione content in serum after ingesting S-PT84. It is a graph which shows the liver glutathione content after ingesting S-PT84. It is a graph which shows the glutathione content in a macrophage cell after ingesting S-PT84. It is a graph which shows that S-PT84 oral intake acts inhibitory with respect to the fall of the glutathione level in serum by restraint stress. It is a graph which shows that S-PT84 ingestion acts suppressively with respect to the fall of the macrophage intracellular glutathione level by restraint stress.

Claims (8)

A composition for oral consumption for maintaining or increasing the in vivo glutathione concentration containing a lactic acid bacterium-containing substance as an active ingredient. The composition according to claim 1, wherein the lactic acid bacterium-containing material is a lactic acid bacterium cell, a lactic acid bacterium suspension, a lactic acid bacterium culture, a lactic acid bacterium culture solution, or a lactic acid bacterium fermentation product. The composition for oral consumption according to claim 1 or 2, wherein the daily intake of lactic acid bacteria per adult is 0.02 mg to 50,000 mg. The composition for oral consumption according to claim 3, wherein the daily intake for adults is 0.2 mg to 200 mg. The composition for oral consumption according to any one of claims 1 to 4, which is used for prevention or treatment of a disease caused by glutathione deficiency. The composition for oral consumption according to any one of claims 1 to 4, which is used for prevention or treatment of a disease caused by oxidative stress. The composition for oral intake according to any one of claims 1 to 4, which is used for preventing aging. The composition for oral consumption according to any one of claims 1 to 7, which is a food, drink, food additive, health food or pharmaceutical product.

Images