JP2007070249A - Immunological function regulator, antiallergic agent, composition for immunomodulation, antiallergic composition and food containing the same - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide an immunological function regulator having sufficient immunomodulation activities and antiallergic activities compared to a lactic bacterium derived from an animal; and to provide an antiallergic agent, a composition for immunomodulation, an antiallergic composition and a food containing them. <P>SOLUTION: The immunological function regulator contains live bacteria, dead bacteria or a biomass-treated product of at least one or more of lactic acid bacteria derived from a plant and belonging to the genus Lactobacillus, lactic acid bacteria derived from a plant and belonging to the genus Leuconostoc, and lactic acid bacteria derived from a plant and belonging to the genus Pediococcus as an active ingredient. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to an immune function regulator, an antiallergic agent, an immunoregulatory composition and an antiallergic composition containing a plant-derived lactic acid bacterium as an active ingredient, and a food containing these.

  In recent years, an increasing number of people are suffering from allergic rhinitis, atopic dermatitis, hay fever, food allergies, etc. with changes in eating habits and lifestyle habits. In particular, hay fever has occurred in 20% of Japanese adults and has become a serious social phenomenon.

  The following mechanism is considered as the mechanism of allergy development. When the causative agent allergen (antigen) enters from the oral cavity, nasal cavity or intestinal tract, antigen-presenting cells such as dendritic cells and macrophages are phagocytosed and present the antigen to T cells and B cells. B cells mature into IgE-producing cells with the help of helper T cells (particularly type II helper T cells (Th2)), and produce IgE antibodies. IgE antibody binds to the surface of mast cells, and when an antigen further binds, the stimulus is transmitted into the mast cells, and inflammatory chemical substances such as histamine and leukotriene are released, causing allergic symptoms.

  Helper T cells (Th) include type I helper T cells (Th1) and type II helper T cells (Th2). Th2 produces interleukin (IL) -4 and IL-5 and promotes IgE production. Th1 cells are activated by IL-12 produced by antigen-presenting cells, produce IFN-γ, and suppress IgE production. The allergen invasion causes a human to become Th2 type and promotes IgE production, but activation of Th1 suppresses IgE production.

  By the way, regarding the immunoregulatory function of lactic acid bacteria, various studies have recently been conducted on lactic acid bacteria in the human intestine and so-called animal lactic acid bacteria derived from fermented milk. For example, the antiallergic action of Lactobacillus paracasei derived from dairy products (Patent Document 1), the antiallergic action of Lactococcus lactis (Patent Document 2), the antiallergic action of Enterococcus faecalis and Lactobacillus reuteri (Patent Document 3), Lactobacillus acidophilus immune function-modulating effect (Patent Document 4), Lactobacillus rhamnosus immune enhancing action (Patent Document 5) and the like are known.

JP 2005-139160 A JP 2004-18469 A JP 2000-95697 A Japanese Patent Laid-Open No. 2004-277381 JP-T-2001-513990

  However, animal-derived lactic acid bacteria have a problem that their biological activity is not sufficient. On the other hand, little research has been conducted on the isolation and physiological functions of plant-derived lactic acid bacteria.

  Therefore, the present invention includes an immune function regulator, an antiallergic agent, an immunoregulatory composition and an antiallergic composition, which have sufficient immunoregulatory action and antiallergic action compared to animal-derived lactic acid bacteria, and these. The purpose is to provide food.

  In order to achieve the above-mentioned object, the present inventors have conducted extensive research, and as a result, the lactic acid bacteria belonging to the genus Lactobacillus derived from plants, the lactic acid bacteria belonging to the genus Leuconostoc derived from plants, and the genus Pediococcus derived from plants. It has been found that at least one live or dead lactic acid bacterium or a treated product thereof has sufficient immune function regulating action or anti-allergic action compared to animal-derived animal lactic acid bacteria. That is, the present invention is a lactic acid bacterium belonging to the genus Lactobacillus derived from a plant, a lactic acid bacterium belonging to the genus Leuconostoc derived from a plant, and a lactic acid bacterium belonging to the genus Pediococcus derived from a plant, It is an immune function regulator and an antiallergic agent, or a composition for regulating an immune regulator function and an antiallergic composition, each comprising a bacterium or a treated product thereof as an active ingredient.

  As described above, according to the present invention, an immune function regulator, an antiallergic agent, an immunoregulatory composition and an antiallergic composition, which have sufficient immunomodulatory action and antiallergic action compared to animal-derived lactic acid bacteria, and these Can be provided.

  Plant-derived plant lactic acid bacteria can grow in an oligotrophic state as compared to animal lactic acid bacteria, and environmental factors such as growth temperature and pH can proliferate under more severe conditions. In addition, animal lactic acid bacteria ferment only milk and livestock meat, whereas plant lactic acid bacteria can ferment a wide range of foods such as vegetables, fruits, grains, beans, and seaweeds.

  It is preferable that the plant lactic acid bacteria contained in the immune function regulator, antiallergic agent, immunoregulatory composition and antiallergic composition according to the present invention are separated from pickles. Sunki-zuke means pickles that are produced and eaten in the Kiso region of Nagano Prefecture. In its manufacture, salt is not used at all and seasoned only by lactic acid fermentation to improve shelf life. Have The plant lactic acid bacteria can be separated from the pickled pickles by a known method. For example, a method of separating lactic acid bacteria liberated in pickles with a medium for separating lactic acid bacteria, or a method of separating pickled soups with a culture medium for separating lactic acid bacteria after enrichment culture, and a culture of pickled pickles that are accumulated and separated with a medium for separating lactic acid bacteria And a method of separating lactic acid bacteria adhering to pickles after stirring, homogenization, ultrasonic treatment, etc., and separating them with a medium for separating lactic acid bacteria, and a method of separating them from pickled pickled seeds.

  In the immune function regulator and the composition for immunoregulation according to the present invention, the immunoregulatory action refers to, for example, the action of enhancing Th1-type cytokine production, and the immune function regulator according to the present invention is compared with animal lactic acid bacteria. Thus, the ability to produce Th1-type cytokines is several times higher. In the antiallergic agent and antiallergic composition according to the present invention, the antiallergic action refers to, for example, the action of suppressing IgE production, and the antiallergic drug according to the present invention inhibits IgE production compared to animal lactic acid bacteria. The effect is 10 times higher.

  In the immune function regulator, the antiallergic agent, the immunoregulatory composition, and the antiallergic composition according to the present invention, the plant lactic acid bacterium is a single or a culture solution obtained by culturing several types of plant lactic acid bacteria, and a collection thereof. Fungi, plant lactic acid bacteria powder, granules, tablets, etc. dried by freeze drying, spray drying, etc., which have the ability to proliferate when inoculated into a medium for lactic acid bacteria growth, and dead bacteria , One or a culture solution of several types of plant lactic acid bacteria, heated, sterilized by physical treatment such as chemical treatment with antibacterial agents, electron beam, ultraviolet rays, radiation, etc. It refers to a pulverized product obtained by destroying several types of plant lactic acid bacteria by a method such as enzyme treatment, homogenization, or ultrasonic treatment, or a fraction of cell wall fractions having immune function regulating activity or antiallergic activity.

  In the immune function regulator, the antiallergic agent, the immunoregulatory composition and the antiallergic composition according to the present invention, the lactic acid bacterium belonging to the genus Lactobacillus derived from the plant is a lactic acid bacterium and a Lactobacillus plan belonging to Lactobacillus delbruecki The lactic acid bacteria belonging to Talam are preferably at least one or more, and the lactic acid bacteria belonging to Lactobacillus delbruecki and the lactic acid bacteria belonging to Lactobacillus plantarum may be used in combination.

  The forms of the immune function regulator, antiallergic agent, immunoregulatory composition and antiallergic composition according to the present invention include liquid, powder, granule and tablet isolates from plants such as pickled vegetables. In addition to those processed into a shape or the like, the plant itself may be processed without being separated from plants such as pickled vegetables. In particular, many immune function regulating functions and antiallergic functions can be obtained depending on the fermentation conditions of the pickled vegetables. As what processed the plant itself, there exist what was dried and cut | disconnected, and what was pulverized, for example.

  Some foods according to the present invention are obtained by fermenting food using a strain or a fermented product containing the strain as an inoculum. Examples of these fermented foods include yogurt, pickles and fermented beverages. In addition to these, the food according to the present invention can be used in many applications such as tablet-like or capsule-like health foods, confectionery such as candy and gum, beverages, bread or noodles.

  Next, examples of the immune function regulator, antiallergic agent, immunoregulatory composition and antiallergic composition according to the present invention will be described. A portion of sunki-zuke is accumulated and cultured in an accumulation medium for lactic acid bacteria, and optionally diluted with sterilized physiological saline, and cultured on an agar medium for isolation of lactic acid bacteria (GYP chalk agar medium) supplemented with drugs to isolate lactic acid bacteria. It was. The colonies detected on the agar medium for lactic acid bacteria isolation are individually purely cultured, and after morphological observation with Gram staining and microscope by a conventional method, the bacteria identification test kit (Api 50CHL (Nihon Biomelieu Co., Ltd.)) In addition, we sequenced the PCR product of SSU rDNA encoding 16S rRNA, and identified lactic acid bacteria from the base sequence.37 strains of lactic acid bacteria isolated and identified by the above method were listed in Table 1. The separated lactic acid bacteria were purely cultured in MRS medium (OXIOID), and the lactic acid bacteria powder according to Examples 1 to 37 was prepared by collecting, washing, sterilizing, and freeze-drying.

Experimental example 1
Next, 11-25 week-old female BALB / c mice were dissected and spleen cells were isolated and prepared. Lactic acid bacteria powders according to Examples 1 to 37 were added to the prepared mouse spleen cells, respectively, and cultured in RPMI 1640 medium. On the third day of culture, each culture supernatant was sampled, and the production amounts of IL-12 and IFN-γ, which are Th1-type cytokines, were measured by ELISA. Furthermore, each culture supernatant was sampled on the 7th day of culture, and the antibody production amounts of IgA, IgG and IgM were measured by ELISA. As a comparative example, a lactic acid bacterium-free addition was designated as Comparative Example 1, LPS (Gram-negative bacterial cell wall lipopolysaccharide) was added instead of lactic acid bacterium as Comparative Example 2, and Con A (Nata Bean Lectin, Concanavalin A) was added. This was designated as Comparative Example 3.

  The production amount of IL-12 is shown in Table 2, the production amount of IFN-γ is shown in Table 3, the production amount of IgA is shown in Table 4, the production amount of IgG is shown in Table 5, and the production amount of IgM is shown in Table 4. It is shown in FIG.

  As shown in Table 2, lactic acid bacteria according to Examples 1 and 2, Examples 4 to 15, Examples 18 and 19, Examples 21 to 23, Examples 25 to 29, and Examples 31 to 37 are IL-12. Promoted the production of As shown in Table 3, lactic acid bacteria according to Examples 2 to 10, Example 12, Examples 15 to 19, Examples 21 to 24, and Examples 26 to 37 promoted the production of IFN-γ. As shown in Table 4, the lactic acid bacteria according to Examples 2 and 3, Examples 5 to 25, Example 28, and Examples 31 to 37 promoted the production of IgA. As shown in Table 5, the lactic acid bacteria according to Examples 2 to 37 promoted the production of IgG. As shown in Table 6, the lactic acid bacteria according to Examples 1 to 37 promoted the production of IgM.

  The lactic acid bacteria according to Examples 2, 5 to 7, 10 and 12, which had particularly high cytokine-inducing ability and antibody-inducing ability in Experimental Example 1, were purely cultured in MRS medium (OXIOID), and collected, washed and sterilized, and then lyophilized. Thus, lactic acid bacteria powders according to Example 2, Examples 5 to 7, Example 10 and Example 12 were produced.

  Ovalbumin (OVA) and adjuvant were intraperitoneally injected into female BALB / c mice of 11 to 25 weeks of age 21 days before dissection and 7 days before dissection, and spleen cells were isolated and adjusted.

  OVA 20 μg / well and lactic acid bacteria powder 20 μg / well according to Example 2, Examples 5 to 7, Example 10, and Example 12 were added to the prepared mouse spleen cells, respectively, and cultured in RPMI 1640 medium. On the third day of culture, each culture supernatant was sampled, and the production amounts of IL-12 and IFN-γ, which are Th1-type cytokines, were measured by ELISA. Furthermore, each culture supernatant was sampled on the 14th day of culture, and the antibody production amount of IgE was measured by ELISA. As Comparative Examples, Comparative Example 2, Comparative Examples 5 to 7, Comparative Example 10 and Comparative Example 12 were added without adding lactic acid bacteria and only OVA.

  The production amount of IL-12 is shown in Table 7, the production amount of IFN-γ is shown in Table 8, and the production amount of IgE is shown in Table 9.

  As shown in Table 7, the lactic acid bacteria according to Example 2, Examples 5 to 7, Example 10, and Example 12 promoted IL-12 production. As shown in Table 8, the lactic acid bacteria according to Example 2, Examples 5 to 7, Example 10, and Example 12 promoted production of IFN-γ. As shown in Table 9, the lactic acid bacteria according to Example 2, Examples 5 to 7, Example 10, and Example 12 suppressed the production of IgE.

Experimental example 3
Next, Lactobacillus ramnosus GG which has high Th1 type cytokine production ability and has IgE production inhibitory effect among lactic acid bacteria according to Example 2, Examples 5 to 7, Example 10 and Example 12, and animal lactic acid bacteria. Comparison of IL-12, IFN-γ and IgE production ability with the strain (LGG) was performed.

  As in Experimental Example 2, each lactic acid bacterium was purely cultured in MRS medium (OXIOID), collected, washed, sterilized, and freeze-dried to give Example 2, Examples 5 to 7, Example 10, and Example 12. The lactic acid bacteria powder which concerns on the comparative example of Example 20 and a LGG strain was produced.

  Ovalbumin (OVA) and an adjuvant were injected intraperitoneally into female BALB / c mice of 11 to 25 weeks of age 21 days and 7 days before dissection, and spleen cells were isolated and adjusted.

  To the prepared mouse spleen cells, OVA 20 μg / well and lactic acid bacteria powders according to Example 2, Examples 5 to 7, Example 10, and Example 12 were respectively 0 mg / ml, 0.001 mg / ml, and 0.01 mg / ml. And 0.1 mg / ml concentration, and cultured in RPMI 1640 medium. On the third day of culture, each culture supernatant was sampled, and the production amounts of IL-12 and IFN-γ, which are Th1-type cytokines, were measured by ELISA. Furthermore, each culture supernatant was sampled on the 14th day of culture, and the antibody production amount of IgE was measured by ELISA.

  The production amount of IL-12 is shown in Table 10, the production amount of IFN-γ is shown in Table 11, and the production amount of IgE is shown in Table 12.

  As shown in Table 10, the lactic acid bacteria according to Example 2, Examples 5 to 7, Example 10, and Example 12 promote the production of IL-12, and the production ability of IL-12 from the LGG bacteria according to the comparative example Turned out to be expensive. As shown in Table 11, the lactic acid bacteria according to Example 2, Examples 5 to 7, and Example 12 promote the production of IFN-γ and have higher IFN-γ production ability than the LGG bacteria according to the comparative example. found. As shown in Table 12, the lactic acid bacteria according to Examples 2, 5 to 7, Example 10, and Example 12 suppress the production of IgE and have a higher IgE production suppression effect than the LGG bacteria according to the comparative example. There was found.

  Next, soymilk yogurt was prepared as a food containing the immune function regulator and antiallergic agent according to the present invention. First, 100 parts of soy milk as raw materials, 20 parts of defatted soybean powder and 50 parts of water are uniformly mixed and dissolved, heat sterilized at 100 ° C. for 30 seconds, cooled to 45 ° C., and then Lactobacillus according to Example 2. 20 parts of a mixed starter of Delbrucky and Leuconostoc citrus according to Example 25 was added, filled into a container, and then fermented at 37 ° C. The fermentation was terminated after about 16 hours and cooled to 10 ° C. The pH at the end of the fermentation was 4.2, and a curd (yogurt) was formed by gelation of soybean protein.

Next, a capsule-shaped health food was prepared as a food containing the immune function regulator and antiallergic agent according to the present invention. First, 50 parts of dextrin is mixed with 100 parts of lyophilized bacterial powder of Lactobacillus plantarum according to Example 7, homogenized, filled into hard capsules mainly composed of gelatin and glycerin, and encapsulated health food Got. The number of lactic acid bacteria per capsule was 1 × 10 ¹⁰ .

Claims (9)

        A lactic acid bacterium belonging to the genus Lactobacillus derived from a plant, a lactic acid bacterium belonging to the genus Leuconostoc derived from a plant, and a lactic acid bacterium belonging to the genus Pediococcus derived from a plant. An immune function regulator comprising a product as an active ingredient.        2. The immune function regulator according to claim 1, wherein the lactic acid bacterium enhances a Th1-type immune response in a living body.         The immune function regulator according to claim 1 or 2, wherein the plant-derived lactic acid bacterium belonging to the genus Lactobacillus is at least one of lactic acid bacteria belonging to Lactobacillus delbruecki and lactic acid bacteria belonging to Lactobacillus plantarum. .         A lactic acid bacterium belonging to the genus Lactobacillus derived from a plant, a lactic acid bacterium belonging to the genus Leuconostoc derived from a plant, and a lactic acid bacterium belonging to the genus Pediococcus derived from a plant. A composition for regulating immune function comprising a product as an active ingredient.         A lactic acid bacterium belonging to the genus Lactobacillus derived from a plant, a lactic acid bacterium belonging to the genus Leuconostoc derived from a plant, and a lactic acid bacterium belonging to the genus Pediococcus derived from a plant. Anti-allergic agent containing a product as an active ingredient.        6. The antiallergic agent according to claim 5, wherein the lactic acid bacterium has an IgE antibody production inhibitory action.         The antiallergic agent according to claim 5 or 6, wherein the lactic acid bacterium belonging to the genus Lactobacillus derived from plants is at least one of lactic acid bacteria belonging to Lactobacillus delbruecki and lactic acid bacteria belonging to Lactobacillus plantarum.         A lactic acid bacterium belonging to the genus Lactobacillus derived from a plant, a lactic acid bacterium belonging to the genus Leuconostoc derived from a plant, and a lactic acid bacterium belonging to the genus Pediococcus derived from a plant. Anti-allergic composition comprising a product as an active ingredient.         The immune function regulator according to any one of claims 1 to 3, the immune function regulation composition according to claim 4, the antiallergic agent according to any one of claims 5 to 7, or the antiallergy composition according to claim 8. A food characterized by containing.