JP5300772B2 - 新規乳酸菌、並びに新規乳酸菌を含有する医薬、飲食品、及び飼料 - Google Patents
新規乳酸菌、並びに新規乳酸菌を含有する医薬、飲食品、及び飼料 Download PDFInfo
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- JP5300772B2 JP5300772B2 JP2010073674A JP2010073674A JP5300772B2 JP 5300772 B2 JP5300772 B2 JP 5300772B2 JP 2010073674 A JP2010073674 A JP 2010073674A JP 2010073674 A JP2010073674 A JP 2010073674A JP 5300772 B2 JP5300772 B2 JP 5300772B2
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Description
ヒト糞便1gにリン酸緩衝生理食塩水(PBS)9mlを加えて混合し、さらにPBSで段階的に希釈した。希釈液をラクトバチルス選択培地(0.8%Lab−lemco powder(Oxoid製)及び1.5%酢酸ナトリウムを含むLBS寒天培地(BBL製))に塗抹し、嫌気条件下、37℃で2〜3日間の培養を行った。形成されたコロニーを釣菌し、MRS寒天培地(Difco製)に画線接種し同様に培養した。さらに同じ操作を行い、単一のコロニーを形成する乳酸菌を分離した。得られた分離株について、菌学的性質及び16SrRNA遺伝子の塩基配列を決定した。
本試験では、ラクトバチルス・ガセリMCC1183株の低pH環境における生残性を確認することを目的とした。
ラクトバチルス・ガセリMCC1183株、ラクトバチルス・ガセリOLL 2716株(FERM BP−6999)、及びラクトバチルス・ラムノサスGG株(ATCC 53103)を含む110株の乳酸菌を試験試料とした。ラクトバチルス・ガセリOLL
2716株(FERM BP−6999)は高い胃酸耐性を有すると報告されている菌株である。また、塩化ナトリウム(国産化学製)の0.2%水溶液、及びペプシン(Sigma製)の0.35%水溶液を調製したのち、pH1.4〜2.0の範囲となるように混合し、これを人工胃液とした。
各乳酸菌を、MRS培地(Difco製)で培養(37℃、16時間)し、乳酸菌培養液を得た。得られた乳酸菌培養液を人工胃液に0.01%で添加し、37℃で2時間インキュベーションした。インキュベーション前後での生菌数をMRS寒天培地(Difco製)を用いて測定し、生残性を算出した。
ラクトバチルス・ガセリMCC1183株を試験群、ラクトバチルス・ガセリOLL 2716株(FERM BP−6999)を比較群として、人工胃液耐性試験の結果を図1に示す。
本試験では、ラクトバチルス・ガセリMCC1183株のヘリコバクター・ピロリ増殖抑制効果を確認することを目的とした。
乳酸菌として、ラクトバチルス・ガセリMCC1183株、ラクトバチルス・ガセリOLL 2716株(FERM BP−6999)、及びラクトバチルス・ラムノサスGG株(ATCC 53103)を用いた。ラクトバチルス・ガセリOLL 2716株(FERM BP−6999)、及びラクトバチルス・ラムノサスGG株(ATCC 53103)は、いずれもヘリコバクター・ピロリ増殖抑制効果が知られている株である。また、ヘリコバクター・ピロリとして、ヘリコバクター・ピロリ(ATCC 43504T)を用いた。
MRS培地(Difco製)で培養(37℃、16時間)した乳酸菌の培養液を、5%非働化ウシ胎仔血清(FCS)を含有するBHI培地(Difco製)で希釈し、乳酸菌濃度が1×109cfu/mlになるように650nmにおける吸光度に基づいて調製し、乳酸菌懸濁液とした。ヘリコバクター・ピロリを5%FCS含有BHI培地に接種し、10%CO2存在下で37℃、24時間培養し、ヘリコバクター・ピロリ濃度が1×107cfu/mlになるように調製し、ヘリコバクター・ピロリ懸濁液とした。4mlの5%FCS含有BHI培地に、0.5mlのヘリコバクター・ピロリ懸濁液と0.5mlの乳酸菌懸濁液とを添加し、10%CO2存在下で37℃、12時間培養した。
結果を図2に示す。培養前のヘリコバクター・ピロリ菌数は0.946×106cfu/mlであった。対照群では、培養後に5.95×106cfu/mlとなり、培養前と比較して6倍以上の濃度に増殖した。また、ヘリコバクター・ピロリ増殖抑制効果を有することが知られている既知の乳酸菌と混合した比較群1及び2では、培養後のヘリコバクター・ピロリ菌数はそれぞれ1.87×106cfu/ml、2.05×106cfu/mlであり、培養前と比較していずれも2倍以上の濃度に増殖した。
本試験は、ラクトバチルス・ガセリMCC1183株のヘリコバクター・ピロリの抗原特異的Th1反応に対する抑制効果を確認することを目的とした。具体的には、マウスパイエル板由来の抗原提示細胞とヘリコバクター・ピロリ抗原で感作したマウスのT細胞にヘリコバクター・ピロリ加熱菌体を添加することで誘導されるIFN−γの産生量を指標とした。
乳酸菌として、ラクトバチルス・ガセリMCC1183株及びラクトバチルス・ガセリOLL 2716株(FERM BP−6999)を用いた。ヘリコバクター・ピロリとして、ヘリコバクター・ピロリ(ATCC 43504T)を用いた。
各乳酸菌をMRS培地で16時間培養し、菌体をPBSで2回洗浄し、さらに蒸留水で2回洗浄した後に蒸留水で懸濁し、凍結乾燥を行った。凍結乾燥した菌体をPBSで懸濁して10mg/mlに調整し、100℃、30分間の加熱処理を行い、乳酸菌加熱菌体とした。ヘリコバクター・ピロリを5%FCS含有BHI培地で24時間培養し、菌体をPBSで2回洗浄し、PBSで10mg/mlに調整した。さらに、100℃、30分間の加熱処理を行い、ヘリコバクター・ピロリ加熱菌体懸濁液とした。
ヘリコバクター・ピロリ抗原特異的Th1反応に対する乳酸菌による抑制試験の結果を図3に示す。無刺激群ではIFN−γの産生が認められないのに対し、無刺激群にヘリコバクター・ピロリ加熱菌体を添加した対照群では、ヘリコバクター・ピロリ抗原特異的反応によるIFN−γの産生が認められた。乳酸菌加熱菌体として、ラクトバチルス・ガセリOLL 2716株(FERM BP−6999)加熱菌体を添加した比較群のIFN−γ産生量は、対照群に対して有意な差は認められなかった。
Claims (12)
- ラクトバチルス・ガセリ(Lactobacillus gasseri)MCC1183株(受託番号:FERM P−21908)。
- ラクトバチルス・ガセリMCC1183株を含有する医薬。
- ラクトバチルス・ガセリMCC1183株を含有する飲食品。
- ラクトバチルス・ガセリMCC1183株を含有する飼料。
- ラクトバチルス・ガセリMCC1183株を含有する抗菌剤。
- ヘリコバクター・ピロリ(Helicobacter pylori)の抗菌に用いられる、請求項5に記載の抗菌剤。
- ラクトバチルス・ガセリMCC1183株を含有する抗炎症剤。
- ヘリコバクター・ピロリに起因する胃炎の予防又は治療に用いられる、請求項7に記載の抗炎症剤。
- ラクトバチルス・ガセリMCC1183株を含有する抗潰瘍剤。
- ヘリコバクター・ピロリに起因する胃潰瘍又は十二指腸潰瘍の予防又は治療に用いられる、請求項9に記載の抗潰瘍剤。
- ラクトバチルス・ガセリMCC1183株を含有する免疫抑制剤。
- ヘリコバクター・ピロリによって誘発されるTh1細胞の免疫反応を抑制するために用いられる、請求項11に記載の免疫抑制剤。
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