苔蘚桿菌聚木糖酶之協同性及生產木寡醣.doc
苔蘚桿菌聚木糖酶之協同性及生產木寡醣最適條件之探討Synergistic effect of endoxylanase from Bacillus licheniformis and optimal production of xylooligosaccharides研究生：孫文仁 We-Len Sun指導教授：廖遠東 Ean-Tun Liaw【摘要】本研究以自行純化之Bacillus licheniformis內切聚木糖酶，分別進行纖維酵素協同性、降解產物分析及木寡醣益生特性之探討。首先將聚木糖酶 (xylanase)、纖維素酶 (cellulase) 及木糖苷酶 (xylosidase) 作複合酵素組合，並以樺木聚木醣 (birchwood xylan)、麥木聚木醣 (oat-spelts xylan)、玉米穗軸、未預處理之蔗渣、不同濃度硫酸處理之玉米穗軸及蔗渣粗半纖維素為受質，測定複合酵素間之協同性。另外以麥木聚木醣及樺木聚木醣為基質，於不同水解時間下進行木寡醣之製備，並以高效液相層析儀進行產物分析，且選擇Bifidobacterium bifidum、Lactobacillus acidophilus、Clostridium perfringens為對象，評估聚木糖酶水解產物對腸道益生菌及有害菌生長之影響。實驗結果顯示，1.5% 硫酸處理之玉米穗軸及未預處理之蔗渣，以cellulase/xylanase降解，其加乘效果最好，DSE值分別可達1.67及2.17，而樺木聚木醣、麥木聚木醣、蔗渣粗半纖維素則以xylanase/xylosidase協同性最佳，DSE值分別可得1.59、1.94及1.89。在降解產物分佈方面，麥木聚木醣以聚木糖酶反應48小時，其二醣至六醣所佔比例分別為18、25、7、18及13%，而樺木聚木醣之寡醣分佈則以二、三醣為主，兩者約可佔總寡醣含量8成以上。另外益菌作用結果指出，木寡醣於低濃度下，其促進有益菌增殖之效果並不顯著，然而提高二、三、四醣濃度總和至30,000 ppm及延長木寡醣與菌體培養時間達72小時，則麥木聚木醣降解產物可抑制C. perfringens及促進B. bifidum之增殖。關鍵字：苔蘚桿菌、聚木糖酶、協同性、益生作用【Abstract】The objective of the present study is to conduct the hydrolysate profiles of xylan, prebiotic effect of xylooligosaccharides and synergistic effect by a purified endoxylanase derived from Bacillus licheniformis. First all, a synergistic effect on birchwood xylan, oat-spelts xylan, corn cob, untreated sugar cane bagasse, H2SO4 treated corn cob and hemicellulose prepared from sugar cane bagasse by single xylanase, cellulase and xylosidase or combined enzymes among them were systematically established. Xylooligosaccharides prepared from birchwood and oat-spelts xylan was colleted for prebiotic study using Bifidobacterium bifidum, Lactobacillus acidophilus, Clostridium perfringens as subjects. The results indicated that cellulase/xylanase exhibited the optimal synergistic effect on 1.5% H2SO4 treated corn cob and untreated sugar cane bagasse, where were DSE of 1.67 and 2.17, respectively. Furthermore, xylanase/xylosidase displayed the most efficient hydrolysis in Birchwood xylan, oat-spelts xylan and hemicellulose (from sugar cane bagasse), where were DSE of 1.59, 1.94 and 1.89, respectively. Hydrolysate profiles of oat-spelts xylan showed that the relative ratio of xylobiose to xylohexaose is 18, 25, 7, 18 and 13% respectively, but the major hydrolysates of birchwood xylan are xylobiose and xylotriose which are composed of 80% of total oligomers. Prebiotic study illustrated that little acceleration on microbial growth was observed if low concentration of xylooligomers was employed. However, a moderate effect was found in C. perfringens inhibition and B. bifidum stimulation when increase the total xylobiose, xylotriose and xylotetraose concentration to 30,000 ppm and extend the incubation period up to 72 hours.Key words: Bacillus licheniformis, xylanase, synergism, prebiotic effect
