- 學位論文
利用日本麴菌進行液態發酵和固態發酵生產果糖轉移酶
Fructosyltransferase production by Aspergillus japonicus TU-26 in submerged and solid-state fermentation
摘要
利用日本麴菌Aspergillus japonicus TU-26發酵生產果糖轉移酶，以液態和固態發酵生產酵素，最後在進行兩者間的比較。 液態發酵利用5升發酵槽進行發酵，發酵條件如後。發酵液操作體積2~3 L，溫度：30 ℃，通氣量：1 vvm，攪拌速度：500 rpm，pH值控制在6.0。液態發酵培養基包含1%的酵母萃取物，0.5%的KH2PO4，0.05%的MgSO4•7H2O以及不同濃度的蔗糖溶液。 探討液態發酵的部份，利用20%的蔗糖進行批次發酵，於24小時可獲得最大酵素產量359 U/mL；利用25%蔗糖進行批次發酵，於40小時可獲得最大酵素產量534 U/mL。發酵過程蔗糖轉變成果二糖、寡糖 (包括：果三糖、果四糖、蔗果三糖、蔗果四糖) ，最後分解成葡萄糖和果糖。蔗糖和寡糖均會促進果糖轉移酶的生成，這些糖消失後酵素即不再增加。在饋料發酵中，分別於發酵開始的24小時內連續饋入20%蔗糖或25%蔗糖，可分別在36小時或52小時獲得463 U/mL和615 U/mL的酵素產量。探討固態發酵的部份，經缺一試驗得知麥麩、稻殼、酵母萃取物、蔗糖以及礦物質 (包括：磷酸二氫鉀和硫酸鎂) 都是重要的培養基成分。實驗結果顯示固態發酵培養5天可獲得最高酵素產量，添加較多的麥麩亦可提升酵素產量。 控制每瓶固態發酵培養基成本相當於100 mL的液態發酵培養基，比較液態以及固態發酵的差異。用連續饋料液態發酵和固態發酵相比，液態發酵之酵素產量為46310 U/100 mL，而固態發酵為42930 U /瓶，液態發酵的酵素產量，稍優於固態發酵。
並列摘要
The production of fructosyltransferase (FTase) by Aspergillus japonicus TU-26 in submerged and solid-state fermentation was investigated. Submerged fermentation was carried out in a 5-L fermentor at 30 ℃, 1vvm, 500 rpm and pH 6, with a working volume of 2-3 L. The culture medium consisted of 1% yeast extract, 0.5% KH2PO4, 0.05% MgSO4•7H2O and sucrose of varied concentration. A maximum FTase activity of 359 U/mL was obtained at hour 24 when batch fermentation with 200 g/L of sucrose was performed. A maximum enzyme activity of 534 U/mL was achieved at hour 40 when 250 g/L of sucrose was used. As the fermentation was progressed, sucrose was converted to difructoside, fructo-oligosaccharides (FOS, including: trifructoside, tetrafructoside, 1-kestose, nystose and fructosyl nystose) , glucose and fructose. Eventually, all of the di- and oligo-saccharides were hydrolyzed into glucose and fructose. During the fermentation, FTase was continuously produced until both sucrose and FOS disappeared. In fed-batch fermentation, continuous sucrose-feeding to reach a final concentration of 200 g/L or 250 g/L in the first 24 hours was carried out. Maximum FTase activities of 463 U/mL and 615 U/mL at hour 36 and 52, respectively, were achieved. In solid-state fermentation, an omission test revealed that wheat bran, rice hull, yeast extract, sucrose, KH2PO4 and MgSO4•7H2O played important roles for the production of FTase by A. japonicus. During the solid-state fermentation, FTase production peaked at day 5. More wheat bran added resulted in more FTase produced. Solid-state fermentation and submerge fermentation were compared in FTase productivity based on that the cost of the solid culture medium in a 500-mL flask was equivalent to 100 mL of submerged culture medium. A flask of solid-state culture produced 42930 U of FTase, whereas 100 mL of submerged culture in the fed-batch fermentation with continuous sucrose-feeding gave 46310 U of FTase.