200804591 (1) 九、發明說明 【發明所屬之技術領域】 本發明係關於具有免疫調節作用之乳酸桿菌屬菌、含 有其之組成物、及有效率地分離具有免疫調節作用之乳酸 菌之方法。 【先前技術】 已知乳酸菌具有整腸作用或免疫賦活作用之有益的生 理活性。另外，對於一般消費者，作爲具有健康效能感之 素材之認知度亦高。如此具有有益的生理活性之乳酸菌大 多爲分離自發酵乳製品或腸管之來自動物素材之乳酸菌。 另外，另一方面，以植物素材爲分離來源之植物性乳 酸菌中，亦發現具有免疫賦活作用之菌株。例如，特開平 10— 167972號公報（公開日：平成10年6月23日）中記 載作爲具有免疫賦活作用之乳酸菌之胚芽乳酸桿菌（ Lactobacillus plantarum) L — 137 株。另外，已知分離自 京都的傳統醃漬物之「酸莖」之短乳桿菌（Lactobacillus brevis ) Labre株，或分離自「Shiba漬」之戊糖乳桿菌（ Lactobacillus pentosus) DA74N株等亦具有免疫賦活作用 (參考：（1)特公平7— 55908公報（公開日：平成6年 7 月 26 日）、（2 ) Pasken Journal 15. 2 卜26，2002,岸惇 子、小久保葵、赤谷薰、扇谷繪里子、藤田晳也、岸田綱 太郎：有效乳酸菌選出用之篩選1，植物性乳酸菌於人類 末梢血液之體外（in vitro )免疫賦活效果，（3 )第6次 -5- 200804591 (2) 腸內細菌學會（ 2002. 〇5· 30-31)要旨，赤谷薰、扇谷顧 里子、小久保葵、藤田晳也、岸田綱太郎）。 已知以植物素材爲分離來源之植物性乳酸菌係與醃漬 物或酒類釀造有極深的關連，自釀造步驟可分離許多菌。 • 認爲除了上述以外，亦存在具有免疫賦活作用等之有效的 - 生理活性之菌株。例如，自酒類之釀造步驟中之葡萄酒釀 造步驟所分離之菌株，可舉例如分離頻率高菌株之胚芽乳 酸桿菌。然而，對於分離自如此的酒類釀造步驟的菌，主 要係關於賦予釀造中的作用或香味等之硏究，對於菌體本 身影響生物體的效能，幾乎未硏究。因此，可期待從分離 自酒類釀造步驟的乳酸菌中，發現具有有效生理活性之菌 株。另外，使用植物性乳酸菌時，會發生獨特的發酵臭， 依據製造食品的種類，必須考慮對於官能上的對策。 【發明內容】 〔發明之揭示〕 本發明係有鑑於上述現狀，以提供具有免疫調節作用 之乳酸菌、含有其之組成物、及分離具有免疫調節作用之 乳酸菌之方法爲目的。 本發明者等爲解決相關課題，關於分離自酒類釀造步 驟的產品之乳酸菌，調查該免疫調節作用，關於選自其中 之屬於胚芽乳酸桿菌及短乳桿菌之乳酸菌，更詳細地調查 免疫調節作用。該結果係發現該乳酸菌具有高免疫調節作 用’並且使用其之發酵物顯示良好的香味，而完成本發明 -6 - 200804591 (3) 具體上，關於分離自酒類釀造步驟的產 比較 Thl 型細胞激素（cytokine )之 Interleukin-1 2 )(以下簡稱爲「I L — 1 2」） 果，發現可高效率地分離具有免疫調節作用 其屬於胚芽乳酸桿菌及短乳桿菌之菌株，對 之腹腔巨噬細胞，進行體外處理時，發現提 生。進一步，對小鼠腹腔內，進行體內之加 亦確認誘導Th 1型細胞激素之IL — 1 2產生。 亦即，本發明係含於酒類製造步驟之微 ，具有免疫調節作用之乳酸桿菌屬菌。尤其 成份爲發酵濁酒，酒類造步驟係葡萄酒釀造: 另外，本發明係具有免疫調節作用之胚 Lactobacillus plantarum )或短乳桿菌（ brevis) 〇[Technical Field] The present invention relates to a Lactobacillus bacterium having an immunomodulatory action, a composition containing the same, and a method for efficiently isolating an immunomodulatory lactic acid bacterium. [Prior Art] Lactic acid bacteria are known to have beneficial physiological activities of intestinal action or immunostimulating action. In addition, for the general consumer, awareness as a material with a sense of health and efficacy is also high. The lactic acid bacteria having such beneficial physiological activities are mostly lactic acid bacteria derived from animal materials separated from fermented dairy products or intestinal tubes. On the other hand, in the plant lactic acid bacteria in which the plant material is isolated, a strain having an immunostimulating action was also found. For example, Lactobacillus plantarum L-137 strain, which is a lactic acid bacteria having an immunostimulating action, is recorded in Japanese Patent Publication No. Hei 10-167972 (Publication Date: June 23, 2004). In addition, it is known that the Lactobacillus brevis Labre strain of the "sour stem" isolated from the traditional pickles of Kyoto, or the Lactobacillus pentosus DA74N strain isolated from "Shiba stain" also has an immunostimulating activity. Role (Reference: (1) Special Fair 7-55908 Bulletin (Publication Day: July 26, 2005), (2) Pasken Journal 15. 2 Bu 26, 2002, Kishiko, Okubo Kui, Akiya Kaoru, Fangu Paint Riko, Fujita Eisan, Kishida Gangtaro: Screening for effective lactic acid bacteria selection, 1 in vitro immunization effect of lactic acid bacteria in human peripheral blood, (3) 6th-5-200804591 (2) Intestine The Keynote of the Society of Bacteria (2002. 〇5· 30-31), Akiya Kaoru, Fan Gu Gu Lizi, Okubo Kui, Fujita Eiji, Kishida Gangtaro). It is known that plant lactic acid bacteria isolated from plant material has a deep connection with pickles or wine brewing, and many bacteria can be isolated from the brewing step. • It is considered that in addition to the above, there are also effective physiologically active strains such as immunostimulating effects. For example, the strain isolated from the wine-making step in the brewing step of the wine may, for example, be a Lactobacillus plantarum having a high frequency of isolation. However, for the bacteria isolated from such a wine-making step, it is mainly concerned with the effect or the fragrance imparted in the brewing, and the effect on the organism itself is hardly affected. Therefore, it is expected that a strain having an effective physiological activity can be found from the lactic acid bacteria separated from the alcohol brewing step. In addition, when a plant lactic acid bacterium is used, a unique fermentation odor occurs, and depending on the type of the food to be produced, it is necessary to take measures against the sensibility. [Disclosure of the Invention] The present invention has been made in an effort to provide a lactic acid bacterium having an immunomodulatory action, a composition containing the same, and a method of isolating a lactic acid bacterium having an immunomodulatory action in view of the above-mentioned state of the art. In order to solve the problem, the inventors of the present invention investigated the immunomodulatory action of the lactic acid bacteria isolated from the product of the alcoholic brewing step, and examined the immunomodulatory effects in more detail with respect to the lactic acid bacteria selected from the group consisting of Lactobacillus plantarum and Lactobacillus brevis. The result is that the lactic acid bacterium has a high immunomodulatory effect and the fermented product exhibits a good aroma, and the present invention is completed. -6 - 200804591 (3) Specifically, the comparative Th1 type cytokine is isolated from the alcohol brewing step. (Cytokine) Interleukin-1 2 ) (hereinafter referred to as "IL - 1 2") fruit, found that it can efficiently isolate the strains which have immunomodulatory effects belonging to Lactobacillus plantarum and Lactobacillus brevis, and peritoneal macrophages When it was treated in vitro, it was found to be lifted. Further, it was confirmed that the intraperitoneal administration of the mouse was induced in the body to induce IL-1 2 production of Th 1 type cytokines. That is, the present invention is a Lactobacillus bacterium which has an immunomodulating effect in the microbial manufacturing step. In particular, the composition is fermented turbid wine, and the wine making step is wine brewing: In addition, the present invention is an immunomodulatory embryo Lactobacillus plantarum or brevis 〇
進一步，本發明係具有免疫調節作用之 爲胚芽乳酸桿菌SAM2446株(FERM ABP-乳桿菌爲短乳桿菌SAM2447株(FERM ABP 另外，本發明係含於酒類製造步驟之微 ，含具有免疫調節作用之乳酸桿菌屬菌之具 用之組成物。尤其，上述組成物係飮食品或1 進一步，本發明係包含自酒類製造步驟 成份分離微生物之步驟，測定所分離之各微 節作用之步驟，及選出免疫調節作用強之乳 品之乳酸菌， 个白質-12 ( 誘導效能之結 之乳酸菌，尤 於調製自小鼠 昇IL— 12產 熱菌液投予， 生物含有成份 ，微生物含有 ，驟。 芽乳酸桿菌（ Lactobacillus 胚芽乳酸桿菌 -1 043 8 )或短 —10439) 〇 生物含有成份 有免疫調節作 I藥品。 之微生物含有 生物之免疫調 酸菌之步驟之 200804591 (4) 分離具有免疫調節作用之乳酸菌的方法。尤其，微生物含 有成份係發酵濁酒，酒類製造步驟係葡萄酒釀造步驟。 〔用以實施發明之最佳形態〕 • <乳酸桿菌屬菌> 乳酸桿菌屬菌係乳酸菌的一種，本發明係含於酒類製 造步驟之微生物含有成份，具有免疫調節作用之乳酸桿菌 屬菌。乳酸桿菌屬菌係以屬於胚芽乳酸桿菌（ Lactobacillus plantarum )、或短乳桿菌（Lactobacillus brevis)之乳酸桿菌屬菌爲宜。 乳酸桿菌屬菌之菌學上性質係記載於 BERGEY’S MANUAL of Systematic Bacteriology (第 1 卷 1 984 年， 第2卷1986年，第3卷1989年，第4卷1989年）。Further, the present invention has the immunomodulatory effect of the Lactobacillus plantarum SAM2446 strain (FERM ABP-Lactobacillus is Lactobacillus brevis strain SAM2447 strain (FERM ABP. In addition, the present invention is contained in the microbial manufacturing step, and has immunomodulatory effects. A composition for use of a bacterium of the genus Lactobacillus. In particular, the above composition is a food or a food. Further, the present invention comprises the steps of separating microorganisms from the components of the alcohol manufacturing step, measuring the action of each of the separated nodules, and selecting A lactic acid bacteria with a strong immunomodulatory effect, a white matter-12 (a lactic acid bacterium that induces potency), especially prepared from a mouse-producing IL-12-producing bacterium, a biologically-containing component, a microbial-containing bacterium, and a lactic acid bacterium. (Lactobacillus lactobacillus-1 043 8 ) or short -10439) 〇 含有 含有 有 200 200 200 200 200 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 804 In particular, the microorganism contains a component which is fermented turbid wine, and the wine making step is a wine brewing step. The best mode for carrying out the invention: <Lactobacillus < A lactic acid bacterium of the genus Lactobacillus, and the present invention is a bacterium of the genus Lactobacillus which contains a microorganism-containing component in the alcohol production step and has an immunomodulating action. The Lactobacillus strain is preferably a Lactobacillus plantarum or a Lactobacillus brevis Lactobacillus. The bacteriological properties of Lactobacillus are described in BERGEY'S MANUAL of Systematic Bacteriology (Section Vol. 1 1984, Vol. 2, 1986, Vol. 3, 1989, Vol. 4, 1989).
作爲本發明相關之乳酸菌代表物，可舉例如胚芽乳酸 桿菌SAM2446株，及短乳桿菌SAM2447株。本菌係寄存 於獨立行政法人產業技術總合硏究所，專利生物寄存中心 ，該委託號碼係 FERM ABP — 1 043 8，及 FERM ABP -10439。胚芽乳酸桿菌 SAM2446 株（FERM ABP — 10438) 之菌學上性質如表1所示，短乳桿菌SAM2447株（FERM * ABP - 1 043 9 )之菌學上性質如表2所示。 -8- (5) 200804591 [表1] 胚芽乳酸桿菌 SAM 2446 株（FERM ABP- 1 043 8) 細胞形態 桿菌 胞子 不形成 格蘭氏染色性 陽性 運動性 Μ j \ \\ 芽胞 Μ ΝΝ 過氧化氫酶反應 陰性 於1 5 °c生育 良好 於45°C生育 糖之營養利用性（陽性：+、陰性：-、弱陽性：w) 未發育 甘油 - 右旋阿拉伯糖 - 左旋阿拉伯糖 + 核糖 + 右旋木糖 - 左旋木糖 - 半乳糖 + 葡萄糖 + 果糖 + 甘露糖 + 鼠李糖 - 甘露糖醇 + 山梨糖醇 + α -甲基-右旋甘露糖甘 - 右旋葡糖苷 纖維二糖 + 乳糖 + 蜜二糖 - 海藻糖 - 棉子糖 - 木糖醇 - (6) 200804591 [表2] 短乳桿菌 SAM 2447 株（FERM ABP-10439) 細胞形態 桿菌 胞子 不形成 格蘭氏染色性 陽性 運動性 並 j\ \\ 芽胞 Μ j \ \\ 過氧化氫酶反應 陰性 於1 5 t生育 良好 於45°C生育 未發育 糖之營養利用性（陽性：+、陰性：-、弱陽性：w) 甘油 右旋阿拉伯糖 W 左旋阿拉伯糖 + 核糖 + 右旋木糖 - 左旋木糖 - 半乳糖 + 葡萄糖 + 果糖 + 甘露糖 - 鼠李糖 - 甘露糖醇 W 山梨糖醇 - α -甲基-右旋甘露糖苷 - 右旋葡糖苷 W 纖維二糖 - 乳糖 - 蜜二糖 - 海藻糖 - 棉子糖 - 木糖醇 - -10- 200804591 (7) 本發明中，所謂「具有免疫調節作用」係指至少具有 於恆定狀態或免疫機能降低時，將其活化之作用（免疫賦 活作用）、或免疫機能過度亢進時，將其抑制成適當的免 疫狀態之作用（免疫抑制作用）、使細胞性免疫及液性免 疫之平衡最適合化作用中之任一種作用。作爲免疫調節作 用，可舉例如細胞激素產生之促進或抑制作用、淋巴球活 化作用、NK (自然殺手細胞）活性增強作用、Thl/Th2平 衡改善作用、免疫降低抑制作用、抗過敏作用等，但並非 侷限於此等者。 本發明之乳酸桿菌屬菌係自酒類製造步驟之微生物含 有成份’分離微生物’測定所分離之各微生物之免疫調節 作用’由選出免疫調節作用強的菌而可得。 本發明中所謂「酒類製造步驟」係指有關酒類製造之 一連串步驟。在此’ 「酒類」亦包含葡萄酒等之水果酒、 啤酒或發泡酒等之麥芽發酵飲料、清酒、威士忌等之蒸餾 酒等。 所謂微生物含有成份係指於酒類製造步驟中之微生物 存在成份，例如加入原料、發酵濁酒等。 <組成物> 本發明相關之乳酸桿菌屬菌係可利用爲含有該乳酸桿 菌屬菌之組成物。尤其，適合利用爲具有免疫調節作用之 藥學上組成物。 -11 . 200804591 (8) 作爲含於上述組成物之乳酸桿菌屬菌係可舉例如乳酸 桿菌屬菌（生菌及死菌）、乳酸桿菌屬菌含有物、乳酸桿 菌屬菌發酵物、乳酸桿菌屬菌處理物等。生菌係可得自該 乳酸桿菌屬菌培養液等之乳酸桿菌屬菌含有物。死菌係可 ‘ 由對生菌進行加熱、紫外線照射、甲醛水處理等而得之。 所得之生菌、死菌係可由進一步的磨碎或破碎等而成處理 物。 亦即，本發明之利用乳酸桿菌屬菌之乳酸桿菌屬菌含 有組成物係只要含有本發明相關之乳酸桿菌屬菌、該乳酸 桿菌屬菌含有物、該乳酸桿菌屬菌發酵物、該乳酸桿菌屬 菌處理物中之至少1種即可。作爲上述之乳酸桿菌屬菌含 有物’可舉例如乳酸桿菌屬菌懸濁液、乳酸桿菌屬菌培養 物（包含菌體、培養上清液、培養基成份）、乳酸桿菌屬 菌培養液（自菌培養物除去固形物者）、乳酸桿菌屬菌發 酵乳（乳酸桿菌屬菌飮料、酸乳、優酪乳等）。作爲使用 於培養屬於乳酸桿菌屬菌之微生物之培養基，只要係可培 育屬於乳酸桿菌屬菌之微生物之培養基即可，並無特別的 限制。另外，作爲乳酸桿菌屬菌處理物，可舉例如磨碎物 、破碎物、液狀物（萃取液等）、濃縮物、糊化物、乾燥 物（噴霧乾燥物、冷凍乾燥物、真空乾燥物、鼓式乾燥物 等）、稀釋物等。另外，因爲本發明相關之胚芽乳酸桿菌 SAM2446株，及短乳桿菌SAM2447株係分離自葡萄酒釀 造步驟者，例如含有將果菜類或穀類，由本發明相關之乳 酸桿菌屬菌發酵之發酵物者，亦適合作爲本發明之利用乳 -12- 200804591 (9) 酸桿菌屬菌發酵物之乳酸菌含有組成物之1種形態。另外 ’如上所述，因爲本發明相關之乳酸桿菌屬菌係原本分離 自食品者，所以安全性高。 本發明之利用乳酸桿菌屬菌發酵物之乳酸桿菌屬菌含 有組成物係適合使用作爲具有免疫調節作用之藥學上組成 物。可舉例如飮食品、醫藥品、飼料等。使用於飲食品時 ’適合實施作爲具有免疫調節作用之健康食品。另外，亦 可與已知之甘味料、酸味料、維生素等之各種成份混合， 製成符合使用者嗜好之製品。可提供例如錠劑、膠囊劑、 口服液劑、口含錠、口香糖、優酪乳等之乳製品、冰淇淋 、飲料、酒精飲料、調味料、加工食品、甜點類、零食類 等之形態。 另外’使用於醫藥品時，使用賦形劑、結合劑、崩壞 劑' 滑澤劑、矯味矯臭劑、助溶劑、懸濁劑、被覆劑等之 醫藥製劑技術領域中通常使用之已知補助劑於主藥而製劑 化。作爲劑型，可舉例如錠劑、膠囊劑、顆粒劑、散劑、 糖漿劑、坐劑、注射劑等，並非特別受侷限者。作爲本醫 藥品之投予途徑，可舉例如經口投予、直腸投予、經腸投 予等，並非特別受侷限者。 使用於飼料時，可混合菌體於例如寵物食品、或家畜 用飼料等之原料。或亦可添加於市售之動物用飼料。其中 ’可適合使用於犬或貓、或雞之鳥類等之食餌。 <具有免疫調節作用之乳酸菌之分離方法> -13- 200804591 do) 測定免疫調節作用係只要該相關業者已知的方法中任 一種皆可。可舉一例，如Thl型細胞激素之介白質12 ( 以下簡稱爲「IL - 1 2」）誘導效能試驗，係藉由對於調製 自小鼠之腹腔巨噬細胞進行體外處理，由測定IL - 1 2產 量而進行。 選出免疫調節作用強之乳酸菌係可依據上述測定免疫 調節作用，選出免疫調節作用高之菌株而進行。作爲選出 基準係與已知免疫調節作用之乳酸菌進行比較、與對照組 比較、或與平均値比較等，適當設定即可。另外，免疫調 節作用之外，亦可加入一般乳酸菌用培養基中是否顯示良 好的增殖於選出基準。 本發明中所謂「酒類製造步驟」係指有關酒類製造之 一連串步驟。在此，「酒類」亦包含葡萄酒等之水果酒、 啤酒或發泡酒等之麥芽發酵飲料、清酒、威士忌等之蒸餾 酒等。 所謂微生物含有成份係指於酒類製造步驟中之微生物 存在成份，例如加入原料、發酵濁酒等。 作爲分離來源，藉由使用上述葡萄酒釀造步驟之發酵 濁酒等’可極有效率地分離具有免疫調節作用之乳酸菌。 由實施例更詳細地說明本發明，但本發明並不侷限於 上述各種實施形態者，可於申請專利範圍進行各種改變， 適當地組合於不同實施形態所分別揭示之技術手段所得之 實施形態亦包含於本發明之技術範圍。 -14- 200804591 (11) 【實施方式】 <實施例> 實施例1 [使用乳酸菌株] 自葡萄酒釀造濁酒，分離出1 04株乳 出Th 1型免疫賦活效能高菌株用之比較/ 1 2 )誘導效能之實驗。 [藉由體外刺激以誘導IL 一 12] 對於 BALB/c小鼠（7週齡公鼠） 4.05 %之硫乙醇酸鹽，4天後，使用PBS ( 回收腹腔內巨噬細胞，使用含l〇%FBS之 爾派克紀念硏究所，Roswell Park Memorial 基，調整成2xl06cells/ml後，接種各0. well )培養皿。添加各菌株之加熱死菌（ 中，測定培養24小時後之培養上清液中之 對照組係未添加死菌者。因爲IL - 1 2之名 p40之次單位鍵結之P70，所以測定IL — 1 ，測定 IL — 12 係使用 OptEIA mouse IL —] BD Pharmingen 社製）。 關於所分離之104株乳酸菌，由體外 生效能之結果，含有許多具有IL - 1 2產 關於此等中幾個之IL - 1 2產生效能，與 用之乳酸菌相比較，認爲仍具有充分高的 酸菌，供應於選 、白質 12 ( IL — ，腹腔內投予 磷酸緩衝溶液） RPMI(羅斯威 Institute)培養 5ml於24孔（ 1 pg/ml )於各孔 :IL— 12 濃度。 5性型係p3 5及 2 ( p70)。另外 I 2測定試劑組（ 評估IL一 12產 生效能之菌株。 已知免疫調節作 活性。其中，關 -15- 200804591 (12) 於具有相對於對照組超過1 0倍之IL - 1 2產生效能之；t 2 株，選出作爲具有局IL- 12產生效能之菌株。 若考量相對於對照組爲1 0倍之極高基準時，認爲此 機率係相S局的値’ 5忍爲葡萄酒發酵濁酒係非常適合作爲 免疫調節乳酸菌之分離來源。 [表 3] _ 菌株記號 IL-12產生量比（*1) 對照組 1 A(*2) 45 B 37 C 36 D 32 E 3 1 F 25 G 25 Η 23 I 20 J 19 K 13 Li*3) 13 * 1 :相對於對照組之各試樣之IL-1 2產生量 *2 ：後述之 SAM 2446 株（FERM ΑΒΡ- 1 043 8) *3 :後述之 SAM 2447 株（FERM ABP-10439) -16- 200804591 (13) 實施例2 [使用乳酸菌株] 將實施例1所選出之1 2株乳酸菌，供予於藉由體內 刺激以比較IL - 1 2誘導效能之實驗。 [藉由體內刺激以誘導IL — 12] 對於BALB/c小鼠（7週齡公鼠），腹腔內投予上述 各菌株之加熱死菌（5 0 0 μ g / 0 · 2 m 1 / m 〇 u s e )之懸濁液（溶劑 :生理食鹽水），6小時後頸椎脫臼後，自心臟抽血。另 外，對於對照組之小鼠，投予等量之生理食鹽水。抽血後 ，藉由離心分離採取血清，使用0 p t E IA m o u s e IL — 1 2測 定試劑組（BD Pharmingen社製）測定血清中之IL — 12濃 度。 結果如圖1所示。血清IL — 1 2濃度係A株及L株特 別強。在此，將此等分別命名爲SAM2446株及SAM2447 株。 實施例3 鑑定SAM2446株、SAM2447株 SAM2446株菌學上性質之檢討結果如表1所示， SAM2447株菌學上性質之檢討結果如表2所示。 藉由此等檢討結果，推定SAM2446株爲胚芽乳酸桿 菌，SAM2447株爲短乳桿菌。已知胚芽乳酸桿菌係具有 及不具有山梨糖醇之營養利用性之兩種，如表1所示’ SAM2446株係具有山梨糖醇之營養利用性之乳酸菌種。另 -17- 200804591 (14) 外，已知短乳桿菌係具有及不具有左旋木糖之營養利用性 之兩種，如表2所示，SAM2447株係不具有左旋木糖之營 養利用性之乳酸菌種。 自SAM2446株及SAM2447株萃取DNA後，對於16S rRNA基因範圍進行解讀。因爲所解讀之 SAM2446株之 1 6S rRNA基因序列（序列號碼1 )係得到與胚芽乳酸桿菌 JCM1 149T ( ATCC14917 )之 1 6 S r RN A 基因序歹[J 9 9 % 相同 性，所以鑑定爲胚芽乳酸桿菌（圖2 )。另外，因爲所解 讀之SAM2447株之16S i:RNA基因序列（序列號碼2 )係 得到與短乳桿菌JCM1059T(ATCC14869)之16S rRNA基 因序列97%相同性，所以鑑定爲短乳桿菌（圖3 )。 實施例4由經口攝取之免疫賦活及免疫調節作用 使C5 7BL/6小鼠（Charles River股份有限公司，7週 齡公鼠，1組4隻），自由攝取SAM2446株、SAM2447 株（死菌體）之飮水（0.2mg/day ) 1週。另外，設有未攝 取SAM2446株、SAM2447株之對照組。1週後摘出脾臟 。自所摘出的脾臟，依據常法調製脾細胞，測定NK活性 及細胞激素產生效能。NK活性係將脾細胞與目標細胞之 Yac-Ι (自然殺手細胞的標的細胞），以 200 : 1、或100 :1之比率，共同培養4小時，以使用流式細胞儀（flow cytometry)之PINK法測定脾細胞之細胞障礙性活性。另 外，使 2.5mg/ml 之 Concanavalin A (刀豆球蛋白 A， Nacalai tesque )作用於脾細胞（5x 1 06個/ml ) 24小時後 -18- 200804591 (15) ，以 ELISA 法（0ptEIA(BD BioScience))測定培養上 清液中所產生之IFN - r濃度。 NK活性之測定結果係如圖4所示。圖4中，NK活性 (% )係表示對於小鼠脾細胞之Yac - 1之細胞傷害性’ E:T比係表示使反應之脾細胞數：Yac— 1數。如圖4所示 ，自經口攝取SAM2446株、SAM2447株之小鼠所調製之 脾細胞之NK活性與對照組相比較（control )，明顯升高 〇 IFN — r濃度之測定結果如圖5所示。如圖5顯示， 自經口攝取SAM2446株、SAM2447株之小鼠所調製之脾 細胞與對照組相比較（control ) ，IFN — τ濃度明顯增加 〇 如上所述，關於經口攝取SAM2446株、SAM2447株 之小鼠，NK活性增加，促進Thl型細胞激素之IFN— τ 產生。因此，顯示SAM2446株、SAM2447株具有免疫賦 活作用及免疫調節作用。 實施例5製造豆漿發酵物 於不受到其他微生物污染下，添加SAM2 446株發酵 菌液於市售豆漿，於30°C靜置24小時而得豆漿發酵物。 發現發酵迅速進行’酸度上升及PH降低。所得之發酵物 與傳統的植物發酵品相比較’未顯示不愉快的發酵臭等， 顯示官能上優異的香味。 -19- 200804591 (16) 實施例6製造乳酸菌粉末添加豆漿 將SAM2447株發酵菌液加熱殺菌後，以離心分離（ 3000rpm，10分鐘）回收菌體，對於所得之菌體進行冷凍 乾燥。添加、攪拌此乳酸菌粉末於市售豆漿，懸濁溶解於 豆漿中所得之乳酸菌粉末添加豆漿係與傳統的乳酸菌含有 ' 發酵食品相比較，未顯示不愉快的發酵臭等，官能上亦顯 示優異的香味。 實施例7製造乳酸菌含有組成物 製造例1 :含有乳酸菌之醫藥品 錠劑： 由如下所示之方法，製造含有乳酸菌之醫藥品（錠劑 )。 將5 0g之SAM2446株菌體粉末，混合25 0g之乳糖及 2g之硬脂酸鎂，由單發式打錠機打錠，製造直徑爲l〇mm ，重量爲3 00mg之錠劑。 顆粒劑： 於100g之SAM2447株菌體粉末，加入〇.5g之硬脂 酸鎂，壓縮、粉碎、整粒、過篩而得20至50網目之顆粒 劑。 製造例2 :含有乳酸菌之各種飲食品 以如下所示之組成，製造加入乳酸菌之各種飮食品。 -20- 200804591 (17)Examples of the lactic acid bacteria representative according to the present invention include Lactobacillus brevisii SAM2446 strain and Lactobacillus brevis strain SAM2447 strain. This strain is deposited in the Institute of Industrial Technology, the Independent Administrative Corporation, and the Patent Bio-Hosting Center. The entrusted numbers are FERM ABP — 1 043 8 and FERM ABP -10439. The bacteriological properties of the Lactobacillus bulgaricus SAM2446 strain (FERM ABP-10438) are shown in Table 1, and the bacteriological properties of the Lactobacillus brevis strain SAM2447 (FERM * ABP - 1 043 9 ) are shown in Table 2. -8- (5) 200804591 [Table 1] Lactobacillus plantarum SAM 2446 strain (FERM ABP-1 043 8) Cellular bacilli do not form Gram-positive motility Μ j \ \\ Sproute Μ Hydrogen peroxide Enzyme reaction negative at 15 °c fertility better than 45 °C tocopherol nutrition utilization (positive: +, negative: -, weak positive: w) undeveloped glycerol - d-arabinose - L-arabinose + ribose + right Xylose - L-xylose - galactose + glucose + fructose + mannose + rhamnose - mannitol + sorbitol + α-methyl-dextromannose - dextran glucoside disaccharide + lactose + Honey disaccharide - Trehalose - Raffinose - Xylitol - (6) 200804591 [Table 2] Lactobacillus brevis strain SAM 2447 strain (FERM ABP-10439) Cellular bacilli do not form Gram stain positive motility And j\ \\ bud Μ j \ \\ catalase reaction negative in 1 5 t fertility good at 45 ° C fertility undeveloped sugar nutrient utilization (positive: +, negative: -, weak positive: w) glycerol Right-handed arabinose W Primary sugar + ribose + dextrose - L-xylose - galactose + glucose + fructose + mannose - rhamnose - mannitol W Sorbitol - α-methyl-dextrose-glycoside - D-glucoside W cellobiose - lactose - melibiose - trehalose - raffinose - xylitol - -10- 200804591 (7) In the present invention, "having an immunomodulatory effect" means having at least a constant state or an immune function When it is reduced, when it is activated (immunization activation) or when the immune function is excessively hyperthyroidized, it is inhibited into an appropriate immune state (immunosuppressive action), and the balance between cellular immunity and liquid immunity is most suitable. Any of the roles. Examples of the immunomodulatory action include promotion or inhibition of cytokine production, lymphocyte activation, NK (natural killer cell) activity enhancement, Th1/Th2 balance improvement, immunosuppression inhibition, anti-allergic action, etc., but Not limited to these people. The Lactobacillus strain of the present invention is obtained by selecting a microorganism having strong immunomodulatory action by measuring the immunomodulatory action of each microorganism isolated from the microorganism containing the component 'separating microorganism' in the alcohol production step. The term "alcohol manufacturing step" as used in the present invention refers to a series of steps relating to the manufacture of alcohol. Here, "alcohol" also includes fruit wine such as wine, malt fermented beverage such as beer or sparkling wine, distilled liquor such as sake and whisky. The microorganism-containing component refers to a microorganism-existing component in the wine-making step, for example, a raw material, a fermented turbid wine, or the like. <Composition> The Lactobacillus strain according to the present invention can be used as a composition containing the bacterium of the genus Lactobacillus. In particular, it is suitably used as a pharmaceutically acceptable composition having an immunomodulatory action. -11. 200804591 (8) Examples of the Lactobacillus strains contained in the above composition include Lactobacillus bacteria (bacteria and dead bacteria), Lactobacillus-containing bacteria, Lactobacillus fermentum, Lactobacillus Phytobacterial treatments, etc. The bacteria-derived strain can be obtained from a Lactobacillus-containing fungus such as the Lactobacillus culture solution. The dead bacteria can be obtained from the heating of the bacteria, ultraviolet irradiation, formaldehyde treatment, and the like. The resulting bacteria and dead bacteria can be processed by further grinding or crushing. In other words, the Lactobacillus-containing composition of the bacterium of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus of the genus At least one of the genus bacteria treatments may be used. Examples of the above-mentioned Lactobacillus-containing substance include, for example, a Lactobacillus suspension, a Lactobacillus culture (including a bacterial cell, a culture supernatant, and a medium component), and a Lactobacillus culture solution (self-bacteria) The culture contains the solid matter), the Lactobacillus fermented milk (Lactobacillus sputum, yogurt, yogurt, etc.). The medium used for cultivating a microorganism belonging to the genus Lactobacillus is not particularly limited as long as it is a medium capable of cultivating a microorganism belonging to the genus Lactobacillus. In addition, examples of the Lactobacillus-treated material include a ground material, a crushed product, a liquid material (extract solution, etc.), a concentrate, a paste, and a dried product (a spray dried product, a freeze-dried product, a vacuum dried product, and Drum dry matter, etc.), dilutions, and the like. In addition, since the Lactobacillus plantarum SAM2446 strain and the Lactobacillus brevis strain SAM2447 strain according to the present invention are isolated from the wine-making step, for example, a fermented product of the Lactobacillus-producing bacteria associated with the present invention, which contains fruit vegetables or cereals, The lactic acid bacterium which is suitable for use as the fermented bacterium of the bacterium of the genus Acid bacterium of the present invention contains one form of the composition. Further, as described above, since the Lactobacillus strain which is related to the present invention is originally isolated from food, it is highly safe. The Lactobacillus bacterium having a fermentation bacterium of the genus Lactobacillus of the present invention contains a composition suitable for use as a pharmaceutically acceptable immunomodulatory agent. For example, foods, medicines, feeds, and the like can be mentioned. When used in food and beverage products, it is suitable for implementation as a health food with immunomodulatory effects. In addition, it can also be mixed with various ingredients such as known sweeteners, sour materials, vitamins, etc., to make products that meet user preferences. Forms such as lozenges, capsules, oral liquids, buccal tablets, chewing gum, yogurt, and the like, ice cream, beverages, alcoholic beverages, seasonings, processed foods, desserts, snacks, and the like can be provided. In addition, when used in pharmaceuticals, known subsidies commonly used in the field of pharmaceutical preparations such as excipients, binders, breakers, slip agents, flavoring agents, solubilizers, suspensions, and coatings are used. The agent is formulated in the main drug. The dosage form may, for example, be a tablet, a capsule, a granule, a powder, a syrup, a sitting, an injection or the like, and is not particularly limited. The administration route of the medical drug may, for example, be oral administration, rectal administration, or enteral administration, and is not particularly limited. When used in feed, the cells may be mixed with raw materials such as pet food or livestock feed. Or it can be added to commercially available animal feed. Among them, it is suitable for use in foods such as dogs or cats, or birds of chickens. <Method for separating lactic acid bacteria having immunomodulating effect> -13- 200804591 do) The immunomodulatory action is measured as long as any one of the methods known to the relevant artisan can be used. For example, a Thl-type cytokine-mediated white matter 12 (hereinafter referred to as "IL-1") induction potency test is performed by measuring IL-1 in vitro by treating peritoneal macrophages modulated from mice. 2 production is carried out. The selection of a lactic acid bacteria strain having a strong immunomodulatory action can be carried out by selecting an immunomodulatory strain according to the above-described immunomodulatory action. The selection criteria may be compared with a lactic acid bacteria having a known immunomodulatory action, compared with a control group, or compared with an average sputum, and the like. Further, in addition to the immunomodulatory action, it is also possible to add a good proliferation to the selection criteria in the medium for general lactic acid bacteria. The term "alcohol manufacturing step" as used in the present invention refers to a series of steps relating to the manufacture of alcohol. Here, "alcohol" also includes fruit wine such as wine, malt fermented beverage such as beer or sparkling wine, distilled liquor such as sake and whisky. The microorganism-containing component refers to a microorganism-existing component in the wine-making step, for example, a raw material, a fermented turbid wine, or the like. As a source of separation, the lactic acid bacteria having immunomodulatory effects can be efficiently separated by using the fermented liquor or the like of the above-described wine-making step. The present invention will be described in more detail by way of examples. However, the present invention is not limited to the various embodiments described above, and various modifications may be made without departing from the scope of the invention. It is included in the technical scope of the present invention. -14-200804591 (11) [Embodiment] <Examples> Example 1 [Using lactic acid strain] Comparison of 1 04 strains of Th 1 type immunostimulating strains isolated from wine brewing turbid wine / 1 2) Experiments to induce efficacy. [In vitro stimulation to induce IL-12] For BALB/c mice (7-week-old male rats) 4.05% thioglycolate, 4 days later, PBS was used (recovering intraperitoneal macrophages, using l〇) %FBS Parker Memorial Research Institute, Roswell Park Memorial Base, adjusted to 2xl06cells/ml, inoculate each 0. well) Petri dish. The heat-killing bacteria of each strain were added (in the control group in the culture supernatant after 24 hours of culture, the dead bacteria were not added. Since the IL-1 2 name p40 subunit bonded P70, the IL was measured. — 1 . Measurement of IL-12 using OptEIA mouse IL —] BD Pharmingen Co., Ltd.). As a result of the in vitro activity of the 104 strains of lactic acid bacteria isolated, there are many IL-1 production products with IL-1 production efficiency, which is considered to be sufficiently high compared with the lactic acid bacteria used. The acid bacteria were supplied to the white matter 12 (IL-, intraperitoneally administered phosphate buffer solution) RPMI (Roswell Institute) to culture 5 ml in 24 wells (1 pg/ml) in each well: IL-12 concentration. 5 sex types p3 5 and 2 (p70). In addition, the I 2 assay reagent group (a strain that evaluates the potency of IL-12 production) is known to be immunomodulatory. Among them, Guan-15-200804591 (12) has an IL-1 production efficiency of more than 10 times relative to the control group. ; t 2 strain, selected as a strain with local IL-12 production efficiency. If the consideration is 10 times the extremely high benchmark relative to the control group, it is considered that this probability is the phase of the S The turbid liquor is very suitable as a source of isolation for immunomodulating lactic acid bacteria. [Table 3] _ strain number IL-12 production ratio (*1) control group 1 A (*2) 45 B 37 C 36 D 32 E 3 1 F 25 G 25 Η 23 I 20 J 19 K 13 Li*3) 13 * 1 : IL-1 2 production amount per sample with respect to the control group *2: SAM 2446 strain (FERM ΑΒΡ-1 043 8) *3 described later *3 : SAM 2447 strain (FERM ABP-10439) -16-200804591 (13) Example 2 [Using lactic acid strain] The 12 strains of lactic acid bacteria selected in Example 1 were supplied for in vivo stimulation to compare IL - 1 2 Experiments to induce efficacy. [Induction of IL-12 by in vivo stimulation] For BALB/c mice (7-week-old male rats), heat-dead bacteria (500 μg / 0 · 2 m 1 / m) of each of the above strains were intraperitoneally administered. 〇use) suspension (solvent: physiological saline), after 6 hours of cervical dislocation, blood was drawn from the heart. Further, for the mice of the control group, an equal amount of physiological saline was administered. After the blood was drawn, serum was taken by centrifugation, and the IL-12 concentration in the serum was measured using a reagent group (BD Pharmingen) of 0 p t E IA m o u s e IL-1. The result is shown in Figure 1. Serum IL-12 concentrations were particularly strong in strains A and L. Here, these are named SAM2446 strain and SAM2447 strain, respectively. Example 3 Identification of the SAM2446 strain and the SAM2447 strain The results of the review of the bacterial properties of the SAM2446 strain are shown in Table 1. The results of the review of the academic properties of the SAM2447 strain are shown in Table 2. Based on the results of the review, it was presumed that the SAM2446 strain was a lactobacillus germ, and the SAM2447 strain was a Lactobacillus brevis. It is known that the Lactobacillus brevis strain has and does not have the nutritional utilization of sorbitol, and the SAM2446 strain has the lactic acid bacteria having the nutritional utilization of sorbitol as shown in Table 1. -17- 200804591 (14) In addition, it is known that the Lactobacillus brevis strain has and does not have the nutritional utilization of L-xylose. As shown in Table 2, the SAM2447 strain does not have the nutritional utilization of L-xylose. Lactic acid bacteria. After extracting DNA from SAM2446 strain and SAM2447 strain, the 16S rRNA gene range was interpreted. Because the 1SS rRNA gene sequence (SEQ ID NO: 1) of the SAM2446 strain was obtained and was obtained from the 1 6 S r RN A gene sequence of the Lactobacillus plantarum JCM1 149T (ATCC14917), it was identified as a germ. Lactobacillus (Figure 2). In addition, since the 16S i:RNA gene sequence (SEQ ID NO: 2) of the SAM2447 strain obtained was 97% identical to the 16S rRNA gene sequence of Lactobacillus brevis strain JCM1059T (ATCC14869), it was identified as Lactobacillus brevis (Fig. 3). . Example 4: Immunological activation and immunomodulation by oral ingestion C5 7BL/6 mice (Charles River Co., Ltd., 7-week-old male rats, 1 group of 4), freely ingested SAM2446 strain, SAM2447 strain (dead bacteria) Body water (0.2mg/day) for 1 week. Further, a control group in which SAM2446 strain and SAM2447 strain were not taken was provided. The spleen was removed after 1 week. From the extracted spleen, spleen cells were conditioned according to the usual method, and NK activity and cytokine production efficiency were measured. NK activity is a combination of spleen cells and target cells of Yac-Ι (target cells of natural killer cells) at a ratio of 200:1, or 100:1 for 4 hours to use flow cytometry. The cytotoxic activity of splenocytes was determined by the PINK method. In addition, 2.5 mg/ml of Concanavalin A (Concanavalin A, Nacalai tesque) was applied to spleen cells (5×1 6 cells/ml) 24 hours later -18-200804591 (15) by ELISA (0ptEIA (BD) BioScience)) The concentration of IFN-r produced in the culture supernatant was determined. The results of the measurement of NK activity are shown in Fig. 4. In Fig. 4, NK activity (%) indicates the cytotoxicity of Yac-1 against mouse spleen cells. The E:T ratio indicates the number of spleen cells to be reacted: Yac-1. As shown in Fig. 4, the NK activity of spleen cells prepared by ingesting SAM2446 strain and SAM2447 strain by oral administration was compared with the control group, and the measurement result of 〇IFN-r concentration was significantly increased as shown in Fig. 5. Show. As shown in Fig. 5, the spleen cells prepared by ingesting the SAM2446 strain and the SAM2447 strain were compared with the control group, and the concentration of IFN-τ was significantly increased. As described above, the oral intake of SAM2446 strain and SAM2447 was observed. In mice of the strain, NK activity is increased, and IFN-τ production of Th1 type cytokines is promoted. Therefore, it was revealed that the SAM2446 strain and the SAM2447 strain have immunostimulatory effects and immunomodulatory effects. Example 5 Production of Fermented Soybean Fermentation A soybean soymilk fermentation product was prepared by adding SAM2 446 strain of fermentation broth to commercially available soybean milk without being contaminated by other microorganisms, and allowing to stand at 30 ° C for 24 hours. The fermentation was found to proceed rapidly with an increase in acidity and a decrease in pH. The obtained fermented product was compared with a conventional plant fermented product. No unpleasant fermentation odor or the like was exhibited, and a functionally excellent flavor was exhibited. -19-200804591 (16) Example 6 Production of lactic acid bacteria powder-added soybean milk After the SAM2447 strain fermentation broth was heat-sterilized, the cells were collected by centrifugation (3000 rpm, 10 minutes), and the obtained cells were freeze-dried. Adding and stirring the lactic acid bacteria powder to the commercially available soybean milk, and the lactic acid bacteria powder obtained by suspending and dissolving in the soybean milk is added to the soybean milk. Compared with the conventional lactic acid bacteria containing the fermented food, the unpleasant fermentation odor is not exhibited, and the functional odor is also excellent. . Example 7 Production of lactic acid bacteria-containing composition Production Example 1 : Pharmaceutical product containing lactic acid bacteria Lozenge: A pharmaceutical product (tablet) containing lactic acid bacteria was produced by the method shown below. 50 g of the SAM2446 strain bacterial powder was mixed with 25 g of lactose and 2 g of magnesium stearate, and ingot was ingot by a single-shot tableting machine to prepare a tablet having a diameter of 10 mm and a weight of 300 mg. Granules: 100 g of the SAM2447 strain of the bacterial powder, added with 5 g of magnesium stearate, compressed, pulverized, sized, and sieved to obtain 20 to 50 mesh granules. Production Example 2: Various foods and drinks containing lactic acid bacteria Various mash foods to which lactic acid bacteria are added are produced by the composition shown below. -20- 200804591 (17)
果凍 (組成） 山梨糖醇粉末 香料 SAM2446株菌體粉末 山梨糠醇薄片 (重量份） 99.7 0.2 0.05 0.05 總量 100.00 (咖啡果凍） (組成） (重量份） 砂糖 15.0 明膠 1.0 咖啡萃取物 5.0 水 78.93 SAM2447株菌體粉末 0.07 總量 100.00 果汁 (組成） (重量份） 冷凍濃縮溫州蜜柑果汁 5.0 果糖葡萄糖液糖 11.0 檸檬酸 0.2 左旋抗壞血酸 0.02 SAM2446株菌體粉末 0.05 -21 - 200804591 (18) 香料 0.2 色素 0.1 7k 83.43 總量 100.00 碳酸飮料： (組成） (重量份） 砂糖 8.0 濃縮檸檬果汁 1.0 左旋抗壞血酸 0.10 檸檬酸 0.09 檸檬酸鈉 0.05 著色料 0.05 香料 0.15 碳酸水 90.55 S AM2447株菌體粉未 0.01 總量 100.00 乳酸菌飮料= (組成） (重量份） 乳固形物爲2 1 %之發酵乳 4.76 果糖葡萄糖液糖 13.31 果膠 0.5 檸檬酸 0.08 -22 (19) 200804591 香料 0.15 水 7 1 · 1 4 SAM2446株菌體粉未_0.06 總量 100.00 乳酸菌飮料： (重量份） 14.82 13.3 1 0.5 0.08 0.15 7 1.14 100.00 (組成） 乳固形物爲21%之SAM2447株發酵乳 果糖葡萄糖液糖 果膠 檸檬酸 香料 ztc_ 總量 咖啡飮料： (組成） (重量份） 砂糖 8.0 脫脂奶粉 5.0 焦糖 0.2 咖啡萃取物 2.0 香料 0.1 聚甘油脂肪酸酯 0.05 食鹽 0.05 -23- 200804591 (20) 水 8 4.56 SAM2446株菌體粉末_0.04 總量 100.00 酒精飲料： (組成） （重量份） 5 0容量％乙醇 3 2.0 砂糖 8.4 果汁 2.4 SAM2447株菌體粉末 0.2 精製水_57.0 總量 100.00 發泡酒： (組成） （重量份） 發泡酒 99.98 S AM2447株菌體粉末_0.02 總量 100.00 製造例3 :含有乳酸菌之飼料 如下所示之組成，製造加入乳酸菌之飼料。 飼料： (組成） （重量份） 市售狗飼料 99.98 -24- 200804591 (21) SAM2446秩菌體粉末_0.02 總量 100.00 〔產業上利用性〕 攝取本發明之乳酸桿菌屬菌等時，藉由賦活化免疫機 能而可抑制免疫機能降低。另外，藉由調節免疫機能平衡 ，亦可抑制對於生物體造成不良影響之免疫機能過度亢進 或進一步衍生對於生物體之各種不愉快症狀等。另外，藉 由本發明，可有效率地得到具有免疫調節作用之乳酸菌。 【圖式簡單說明】 [圖1 ]圖1係表示測定腹腔內投予1 2種乳酸菌於小鼠 後之血清IL 一 1 2濃度之結果圖。 [圖2]圖2係表示所解讀之SAM2446株之16S rRNA 基因序列。劃底線部份係表示與胚芽乳酸桿菌JCM 1 1 49 ( ATCC14917 )序列相異的部份。 [圖3]圖3係表示所解讀之SAM2447株之16S irRNA 基因序列。劃底線部份係表示與短乳桿菌J C Μ 1 0 5 9 T ( ATCC 1 4869 )序列相異的部份。 [圖4]圖4係表示ΝΚ活性之測定結果。 [圖5 ]圖5係表示IFN — r濃度之測定結果。 -25-Jelly (composition) Sorbitol powder flavor SAM2446 strain bacterial powder sorbitol flakes (parts by weight) 99.7 0.2 0.05 0.05 Total 100.00 (coffee jelly) (composition) (parts by weight) Sugar 15.0 Gelatin 1.0 Coffee extract 5.0 Water 78.93 SAM2447 Strains powder 0.07 Total 100.00 Juice (composition) (parts by weight) Frozen concentrated Wenzhou mandarin juice 5.0 Fructose glucose liquid sugar 11.0 Citric acid 0.2 L-ascorbate 0.02 SAM2446 strain bacterial powder 0.05 -21 - 200804591 (18) Perfume 0.2 Pigment 0.1 7k 83.43 Total 100.00 Carbonic acid dip: (composition) (parts by weight) Sugar 8.0 Concentrated lemon juice 1.0 L-ascorbic acid 0.10 Citric acid 0.09 Sodium citrate 0.05 Coloring material 0.05 Perfume 0.15 Carbonic acid water 90.55 S AM2447 strain Cell powder No 0.01 Total 100.00 Lactic acid bacteria = (composition) (parts by weight) Milk solids are 21% fermented milk 4.76 fructose glucose liquid sugar 13.31 Pectin 0.5 Citric acid 0.08 -22 (19) 200804591 Perfume 0.15 Water 7 1 · 1 4 SAM2446 strain Powder not _0.06 Total 100.00 Lactic acid bacteria: (weight 14.82 13.3 1 0.5 0.08 0.15 7 1.14 100.00 (Composition) Milk solids 21% SAM2447 fermented lactulose glucose solution candy gum citric acid spice ztc_ Total coffee beverage: (composition) (parts by weight) Sugar 8.0 Skim milk powder 5.0 Caramel 0.2 Coffee Extract 2.0 Perfume 0.1 Polyglycerol Fatty Acid Ester 0.05 Salt 0.05 -23- 200804591 (20) Water 8 4.56 SAM2446 Strains Powder _0.04 Total 100.00 Alcoholic Beverage: (Composition) (Parts by Weight) 5 0 Capacity % ethanol 3 2.0 Sugar 8.4 Juice 2.4 SAM2447 strain of bacteria powder 0.2 Refined water _57.0 Total amount 100.00 Foamed wine: (composition) (parts by weight) Sparkling wine 99.98 S AM2447 strain of bacterial powder _0.02 Total amount 100.00 Manufacturing example 3 : A lactic acid bacteria-containing feed having the composition shown below, and a feed added to the lactic acid bacteria is produced. Feed: (composition) (parts by weight) Commercial dog feed 99.98 -24- 200804591 (21) SAM2446 rank bacterial powder_0.02 Total amount 100.00 [Industrial use] When ingesting the Lactobacillus sp. of the present invention, etc. It activates immune function and can suppress the reduction of immune function. In addition, by regulating the balance of immune function, it is also possible to suppress excessive hyperactivity of the immune system which adversely affects the living body or further derivatize various unpleasant symptoms to the living body. Further, according to the present invention, lactic acid bacteria having an immunomodulating action can be efficiently obtained. BRIEF DESCRIPTION OF THE DRAWINGS [Fig. 1] Fig. 1 is a graph showing the results of measuring the concentration of serum IL-12 after administration of 12 kinds of lactic acid bacteria in mice in the peritoneal cavity. Fig. 2 is a diagram showing the 16S rRNA gene sequence of the SAM2446 strain thus interpreted. The bottom line portion indicates the portion different from the sequence of the Lactobacillus plantarum JCM 1 1 49 (ATCC14917). Fig. 3 is a view showing the sequence of the 16S irRNA gene of the SAM2447 strain which was interpreted. The bottom line portion indicates the portion different from the sequence of Lactobacillus brevis J C Μ 1 0 5 9 T (ATCC 1 4869). Fig. 4 is a graph showing the results of measurement of hydrazine activity. [Fig. 5] Fig. 5 shows the measurement results of the IFN-r concentration. -25-