JP4603273B2 - 固定化微生物担体または固定化酵素担体の製造方法 - Google Patents
固定化微生物担体または固定化酵素担体の製造方法 Download PDFInfo
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- Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
Description
固定化酵素担体の製造(1):
アルギン酸ナトリウムのマヌロン酸とグルロン酸の比(M/G比)がそれぞれ0.5(ダックアルギン45G:紀文フードケミファ製)、1.2(X−C073T:新田ゼラチン製)、2(X−C074T:新田ゼラチン製)であるものについて、蒸留水を用い、粘度が約8,000cpとなるようにそれぞれの水溶液100gを調製し、殺菌のため90℃で1分間加熱した。次に、酵母スポロボロマイセス シンギュラリス(Sporobolomyces
singularis)を、30gのラクトース、9gのイーストエキストラクト、0.3gのリン酸水素二カリウム、0.15gの硫酸マグネシウムを含む3000mlの培地において25℃で72時間培養した後、遠心分離により集菌した。次いで、これを生理食塩水で洗浄し、得られた湿菌体のうち20gを、先に調製したそれぞれの80gのアルギン酸ナトリウム水溶液と良く混合し、三種のアルギン酸ナトリウム・酵母混合液を調製した。
固定化酵素担体によるオリゴ糖の製造(1):
酵母としてスポロボロマイセス シンギュラリス(Sporobolomyces singularis)を用い、M/G比が0.5(ダックアルギン45G:紀文フードケミファ製)および2(X−C074T:新田ゼラチン製)であるアルギン酸ナトリウムを用いる以外は、実施例1に記述した方法に従い、20gの包括固定化酵母を作成した。得られた包括固定化酵母を、実施例1に記述した方法と同様に高濃度のオリゴ糖溶液により処理し、収縮させた。
ゲル形性におけるリン酸塩の影響:
M/G比が1.2(X−C073T:新田ゼラチン製)および2(X−C074T:新田ゼラチン製)であるアルギン酸ナトリウムを用いる以外は、実施例1と同様に作成した包括固定化酵母を、50%の高濃度オリゴ糖溶液(Brix50)で処理した。この処理された包括固定化酵母を、それぞれ、終濃度5、10、20mMのリン酸ナトリウム緩衝液(pH6.0)およびリン酸カリウム緩衝液(pH6.0)を含む50%のオリゴ糖溶液(Brix50)中、55℃で7日間連続して振盪した。製造直後(0日目)と7日目のせん断応力を測定した結果を表2に示す。
固定化酵素担体によるオリゴ糖の製造(2):
酵母としてスポロボロマイセス シンギュラリス(Sporobolomyces singularis)を用い、M/G比が2(X−C074T:新田ゼラチン製)のアルギン酸ナトリウムを用いる以外は、実施例1に記述した方法に従い、20gの包括固定化酵母を作成した。得られた包括固定化酵母を実施例1に記述した方法と同様にオリゴ糖溶液により収縮させた。この包括固定化酵母を用いて実施例2と同様にオリゴ糖生成反応を行った。更に、この反応を19回繰り返し行い、オリゴ糖の生成率とせん断応力を測定した。その結果を表3に示す。
せん断応力における糖溶液濃度(Brix)の影響:
M/G比が2.0(X−C074T:新田ゼラチン製)であるアルギン酸ナトリウムを用いる以外は、包括固定化酵母を実施例1と同様に作成し、あらかじめ10%、20%、30%、40%および50%の濃度(Brix)となるように調整したオリゴ糖溶液中に浸漬した。次いで、オリゴ糖溶液の濃度を一定に維持するため、同一の溶液で2、3回置換して、上記濃度に対応する5種の包括固定化酵母を作成した。置換前および置換後の包括固定化酵母の直径とせん断応力をそれぞれ測定した。なお、結果を表4に示す。
固定化酵素担体の製造(2):
κ−カラギナン(ニッタカラギーナンk−21:新田ゼラチン製)の4%溶液を調整し、スポロボロマイセス シンギュラリス(Sporobolomyces singularis)酵母菌体と重量比6:4で混合し、液温を60℃以上に維持しながら、2%塩化カリウム溶液(Brix2)に滴下し、ビーズを形成させ、一晩エージングした。次いでこれをオリゴメイト55(ヤクルトマテリアル製)と40mMの酢酸−酢酸カルシウム緩衝液(pH=6.0)とを用いて50%に調整したオリゴ糖溶液(Brix50)に浸漬した。更に、オリゴ糖溶液の濃度を一定に維持するため、同一の溶液で2、3回置換して、包括固定化酵母を作成した。置換後のビーズの直径およびせん断応力を上記実施例と同様に測定した。
以 上
Claims (6)
- アルギン酸ナトリウムおよびκ−カラギーナンから選ばれる多糖類と微生物または酵素とを混合後ゲル化せしめ、次いで当該ゲルを当該多糖類のゲル化に使用した金属塩水溶液、水または緩衝液よりも可溶性固形分の濃度が相対的に高い溶液に浸漬させて収縮させることを特徴とする固定化微生物担体または固定化酵素担体の製造方法。
- 多糖類が、アルギン酸ナトリウムである請求項第1項記載の固定化微生物担体または固定化酵素担体の製造方法。
- アルギン酸ナトリウム中のマヌロン酸とグルロン酸の比(M/G比)が1以上である請求項第2項記載の固定化微生物担体または固定化酵素担体の製造方法。
- 多糖類のゲル化に使用した金属塩水溶液、水または緩衝液よりも可溶性固形分の濃度が相対的に高い溶液が、糖溶液である請求項第1項記載の固定化微生物担体または固定化酵素担体の製造方法。
- 糖溶液に使用される糖類が、ガラクトース、フラクトース、マンノース、キシロース、フコース糖、シュクロース、マルトース、セロビオース、キシロビオース糖、ガラクトオリゴ糖、フラクトオリゴ糖、マルトオリゴ糖、乳化オリゴ糖、キシロオリゴ糖またはラフィノース糖である請求項第4項記載の固定化微生物担体または固定化酵素担体の製造方法。
- 多糖類のゲル化に使用した金属塩水溶液、水または緩衝液よりも可溶性固形分の濃度が相対的に高い溶液が、10〜50質量%の基質を含有するものである請求項第1項記載の固定化微生物担体または固定化酵素担体の製造方法。
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