比菲德氏菌對於高純度果寡糖的醱酵生長與代謝

Growth and Metabolism of Bifidobacteria in the Presence of High-Content Fructooligosaccharides

何金柱(Chin-Chu Ho)

指導教授 ： 許垤棊

大同大學/工程學院/生物工程學系所/碩士(2004年)

摘要

比菲德氏菌存在於人體的大腸和結腸中，可促進人體的健康。消化性糖諸如：葡萄糖、蔗糖、麥芽糖等，均無法到達大腸。寡糖成分具有非消化性，可到達大腸，資化比菲德氏菌。利用不含消化性糖的高純度果寡糖於人體外資化比菲德氏菌，由於沒有葡萄糖和蔗糖的干擾，這樣的研究比前人利用含有大量消化性糖的寡糖的研究更具有意義。
本實驗的高純度果寡糖利用雙酵素系統製造，以蔗糖為原料，經黴菌Aspergillus japonicus的果糖轉移酵素催化，生成果寡糖和葡萄糖；同時，葡萄糖被葡萄糖氧化酵素催化，變成葡萄糖酸，葡萄糖酸和碳酸鈣形成鈣鹽沉澱。反應過程使用95 %氧氣，可得到95 %乾種以上的高純度果寡糖。
在五公升的醱酵槽中，有二公升的培養基，含5 % (w/v)的高純度果寡糖，在厭氧的條件下緩慢攪拌，觀察比菲德氏菌的生長。醱酵過程中，控制pH值為6.0，測定混濁度、生菌數、氧化還原電位(ORP)、醋酸對乳酸的比例，以及用HPLC分析二糖、三糖、四糖等諸寡糖成分的變化。在72小時醱酵中，本研究使用的八株比菲德氏菌是：Bifidobacterium longum (CCRC 14634 and 14602)、B. bifidum (CCRC 14615 and 11844)、B. breve (CCRC 11846)、B. adolescentis (CCRC 14607 and 14609)、B. pseudocatenulatum (CCRC 15476)，其中以B. adolescentis CCRC 14607在16小時內消耗完1-Kestose和B. longum CCRC 14602在20小時內消耗完Nystose為最快。產生最多乳酸的是B. adolescentis CCRC 14609，醋酸則是B. adolescentis CCRC 14607產生最多。但B. bifidum CCRC 11844和B. breve CCRC 11846則在高純度果寡糖裡生長緩慢，因此寡糖的代謝較其他的菌株為差。氧化還原電位在菌體快速生長和消耗寡糖的過程中，急遽下降。生長對數期之後，氧化還原電位緩慢的上升。ORP的變化可反映出比菲德氏菌的生長和寡糖的代謝。

關鍵字

高純度果寡糖 ； 比菲德氏菌 ； 氧化還原電位 ； 葡萄糖氧化酶

並列摘要

Bifidobacteria can promote human health. Bifidobacteria are regularly found in human large intestine and colon, where digestible sugars such as glucose, sucrose and maltose, are not attainable. The investigation of enhancing the in vitro growth of bifidobacteria with such oligosaccharides, which is free of the metabolic interference of glucose and sucrose must be more significant than past research with adding commercial oligosaccharides, in that a large amount of digestible sugars are present.
High-content fructooligosaccharides are produced by a mixed-enzyme system with β-fructofuranosidase from Aspergillus japonicus and a commercial glucose oxidase as biocatalysts. Sucrose is catalyzed by β-fructofuranosidase, forming fructooligosaccharides and glucose, and this glucose is concurrently converted to gluconic acid which is then precipitated to calcium gluconate in solution through the catalysis of glucose oxidase. In such way and under an extreme aerobic condition made by aeration with 95 % (v/v) oxygen, up to 95 %, on a dry weight basis, of high-content fructooligosaccharides can be produced.
Five percent (w/v) of such high-content oligosaccharides are added to a 2-L bifidus culture in a 5-L jar fermenter. The fermentation is carried out under an anaerobic condition with gentle stirring. The optical density and the viable count of the fermentation medium are monitored. Furthermore, the utilization of each sugar component such as disaccharide, trisaccharide or tetrasaccharide is analyzed by HPLC throughout the fermentation process. Other parameters such as ORP, pH and the acetate to lactate ratio are also determined. During 72 h of fermentation, eight bifidobacteria including Bifidobacterium longum CCRC 14634, B. longum CCRC 14602, B. bifidum CCRC 14615, B. bifidum CCRC 11844, B. breve CCRC 11846, B. adolescentis CCRC 14607, B. adolescentis CCRC 14609 and B. pseudocatenulatum CCRC 15476 were investigated. B. adolescentis CCRC 14607 depleted 1-kestose completely at h 16 and B. longum CCRC 14602 consumed nystose completely at h 20, being the best strains among eight bifidobacteria. B. adolescentis CCRC 14609 produced the maximum lactic acid of 40.88 g/L and B. adolescentis CCRC 14607 produced the maximum acetic acid of 27.82 g/L. But B. bifidum CCRC 11844 and B. breve CCRC 11846 did not grow well and consumed fructooligosaccharides poorly. As FOS was being consumed, the ORP of the culture broth declined, until most of the FOS was depleted, and then it would increase very slowly. The decreasing of ORP reflected the consuming of FOS and the growth of bifidobacteria.

並列關鍵字

bifidobacteria ； glucose oxidase ； high-content fructooligosaccharides ； oxidation-reduction potential (ORP) ； β-fructofuranosidase

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