Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/51899
DC FieldValueLanguage
dc.contributor.advisor陳錦樹zh_TW
dc.contributor.author朱文銘zh_TW
dc.date1999zh_TW
dc.date.accessioned2014-06-06T08:55:08Z-
dc.date.available2014-06-06T08:55:08Z-
dc.identifier.urihttp://hdl.handle.net/11455/51899-
dc.description.abstract本研究由台灣中部地區的土壤中篩選出一株真菌NCH-3,能分泌高活性之聚木糖。 將菌株NCH-3培養於CZ及MEA培養基上,於顯微鏡下觀察其特徵並與標準菌株比對後,暫命名為Aspergillus niger NCH-3。 接著以Mandels-Reese培養基為基礎進行A. niger NCH-3生產聚木糖之較適培養條件之探討。結果發現碳源以0.75% 燕麥聚木糖,氮源0.38% 硝酸鉀最佳。培養溫度35℃,起始pH 8.0,孢子接種量105 spores/ml及振盪速率130rpm為最佳培養條件。在上述較適培養條件下A. niger NCH-3 於第五天時有最大酵素活性(26.17U/ml)。另外,培養液中β-xylosidase的最高活性為17.01U/ml。A. niger NCH-3聚木糖經超過濾濃縮、硫酸銨沈澱(飽和度40%-70%)、離子交換層析及膠體過濾層析等步驟純化後,分別以SDS-PAGE及IEF等電焦集電泳分析聚木糖之分子量及pI值,發現此酵素分子量約為35k Da ,而pI值為4.0。另外酵素性質如下:最適反應pH值為pH 5, 在pH 5-7 之間活性仍高,最適反應溫度50℃,溫度超過60℃活性明顯下降。基質特異性高,只對由木糖構成之多醣如樺木、櫸木及燕麥等來源之聚木糖有活性。推測硫氫基(-SH )應位於酵素之活性中心區域或其附近。而酵素之硫醇基應涉及催化反應。錳離子(Mn2+)對酵素活性有提高的作用。zh_TW
dc.description.abstractA high xylanase producing mold, strain NCH-3 was isolated from Taiwan soil. According to morphologic characterization of strain NCH-3 grown on CZ and MEA media, it was identified to be the same as a type culture of Aspergillus niger and was named temporarily as Aspergillus niger NCH-3. The optimal medium composition and cultivation conditions for A. niger NCH-3 was determined in shaking flasks (capacity 500 ml). The results showed that the Mandels-Reese medium with oat spelt xylan at 0.75% as carbon source, 0.38% KNO3 as nitrogen sources, initial pH at 8.0, inoculum size at 105 spores/ml, incubation temperature at 35 ℃, and shaking rate at 130 rpm gave the best result for the enzyme production. The maximum xylanase activity of 26.17 U/ml in the culture broth was obtained after 5 days under above conditions. The broth filtrate from A. niger NCH-3 was purified by ammonium sulfate precipitation (40-70%)、ion exchange chromatography and gel filtration chromatography. The purified enzyme appeared as single protein band on SDS-PAGE with molecular weight of 35 kDa. The isoelectric point of purified xylanase was 4.0. The optimum pH of activity was pH 5.0;howere, the enzyme remained quite active in pH ranging from pH 5.0 to 7.0. Both thermal stability and optimal temperature for purified xylanase were at 50℃. Enzyme showed high specificity on hydrolysis of xylans which came from beech wood, birch wood and oat spelt . The activity of xylanase was inhibited by Hg2+ ion, while Mn2+ and -mercaptoethanol at concentrations of 5mM stimulated the enzyme activity. It was suggested that the sulfhydryl (-SH) group in the protein molecule was located at or near the active site, and probably involved in the hydrolysis reaction catalyzed by the enzyme.zh_TW
dc.description.tableofcontents錄 謝 誌.........................................................................................................................I 中文摘要........................................................................................................................II 英文摘要.......................................................................................................................IV 目 錄........................................................................................................................V 圖 目.......................................................................................................................IX 表 目....................................................................................................................XVI 壹、前 言.................................................................................................................1 貳、文獻整理.................................................................................................................3 (壹)、研究背景…………...........................................................................................3 (貳)、半纖維素的種類與成分...................................................................................4 (參)、關於聚木糖...................................................................................................8 一、作用於聚木糖主鏈之酵素..................................................…………..........8 二、作用於聚木糖支鏈之酵素............................................................................8 (肆)、聚木糖之多樣性.....................................................................................….10 (伍)、Aspergillus spp.聚木糖…............................................................................12 (陸)、Aspergillus niger之介紹............................................................................…..13 (柒)、聚木糖在工業上之應用情況................................................................…...13 參、材料與方法...........................................................................................................22 [壹]、材料.................................................................................................................22 一、實驗菌株......................................................................................................22 二、液態培養基..................................................................................................22 三、化學藥劑......................................................................................................23 [貳]、實驗設備.........................................................................................................24 [參]、實驗方法.........................................................................................................26 一、實驗大綱.......................................................................................................26 二、實驗方法.......................................................................................................28 (一)、孢子懸浮液之製備...........................................................................….28 (二)、聚木糖之生產與粗酵素液之製備.................................................…28 (三)、聚木糖之純化.............................................................................…...28 (四)、添加非聚木糖物質至含聚木糖之培養基菌株誘導特性之影響.....….29 (五)、含聚木糖之培養基對菌株培養特性之影響......................................…31 (六)、非聚木糖物質為碳源對菌株培養特性之探討..................................…33 (七)、不同種類半纖維素之製備及在聚木糖生產上之應用..................…37 (八)、分析方法................................................................................................39 肆、結果與討論...........................................................................................................42 [壹]、添加非聚木糖物質至含聚木糖之培養基對菌株誘導特性之影響.........…42 一、Avicel、cellulose、lactose或sorbose添加時機(一次添加) 對酵素活性之影響......................................................................................42 二、不同添加方式對酵素活性之影響............................................................….48 1、連續二次添加.................................................................................................48 2、連續三次添加.................................................................................................51 三、Avicel、cellulose、lactose或sorbose添加頻率(三次添加) 對酵素活性之影響..........................................................................................59 四、添加混合醣類對酵素活性之影響.......................................................……...64 [貳]、含聚木糖之培養基對菌株培養特性之影響............................................…..67 (一)添加聚木糖及非聚木糖物質..........................................................................67 1.不同乳化劑對酵素活性之影響......................................................................67 2.不同時機追加碳源、氮源及孢子數對酵素活性之影響..................……….75 (二)不同培養方式生產聚木糖.........................................................................76 1.兩階段式培養法生產聚木糖(不同基質)...........................................…..76 2. 兩階段式培養對酵素活性之影響(相同基質)........................................…85 [參]、非聚木糖物質為唯一碳源對菌株培養特性探討............………………….88 一、添加非聚木糖物質對酵素活性之影響.................................................….88 二、添加葡萄糖於不同種類基質之培養基中對酵素活性之影響...........……91 三、聚木糖添加時機對酵素活性之影響.....................................................…99 四、添加混合醣類(二種醣類混合) 對酵素活性之影響............................…104 五、乳糖及纖維素之濃度比例對酵素活性之影響...................................…107 六、添加混合醣類(三種醣類混合)對酵素活性之影響........................…….112 七、不同濃度比例之sorbose、avicel及cellulose對酵素活性之影響..….115 [肆]、不同種類半纖維素之製備及在聚木糖生產上之應用.................................119 一、半纖維素之抽(萃)取與分離....................................................................122 二、半纖維素中聚戊糖含量之分析...............................................................122 三、 半纖維素碳源種類對酵素活性之影響...................................................122 四、 酵素水解產物分析……………………………………………………….129 伍、結論......................................................................................................................131 陸、參考文獻..............................................................................................................134 圖 目 圖一、植物細胞壁之結構.............................................................................................4 圖二、半纖維素之結構.................................................................................................5 圖三、聚木糖之結構.....................................................................................................8 圖四、聚木糖及降解聚木糖協同酵素之假設模式................................................….13 圖五、Cryptococcus albidus之聚木糖降解系統..........................................................17 圖六、合成聚木糖之假設模式................................................................................18 圖七、Avicel、cellulose、lactose或sorbose等之添加物對T. longibrachiatum 185 在含燕麥聚木糖之培養基中聚木糖生產之影響.......................................…44 圖八、Avicel、cellulose、lactose或sorbose等之添加物對T longibrachiatum 185 在含燕麥聚木糖之培養基中纖維素生產之影響...........................................45 圖九、Avicel、cellulose、lactose或sorbose等之添加物對T. longibrachiatum 185 在含燕麥聚木糖之培養基中pH值之影響........................................................46 圖十、Avicel、cellulose、lactose或sorbose等之添加物對T. longibrachiatum 185 在含燕麥聚木糖之培養基中孢子數之影響.....................................................47 圖十一、連續二天添加1% avicel、cellulose、lactose或sorbose至含燕麥聚木糖 碳源之培養基對T. longibrachiatum 185聚木糖之影響............................49 圖十二、連續二天添加1% avicel、cellulose、lactose或sorbose至含燕麥聚木糖之 培碳源養基對T. longibrachiatum 185纖維素之影響................................50 圖十三、連續二天添加1% avicel、cellulose、lactose或sorbose至含燕麥聚木糖之 培碳源養基對T. longibrachiatum 185培養液pH值之影響...........................52 圖十四、連續二天添加1% avicel、cellulose、lactose或sorbose至含燕麥聚木糖碳 源之培養基對T. longibrachiatum 185孢子數之影響....................................53 圖十五、連續三天添加1% avicel、cellulose、lactose或sorbose至含燕麥聚木糖碳 源之培養基對T. longibrachiatum 185聚木糖生產之影響........................54 圖十六、連續三天添加1% avicel、cellulose、lactose或sorbose至含燕麥聚木糖碳 源之培養基對T. longibrachiatum 185纖維素生產之影響........................56 圖十七、連續三天添加1% avicel、cellulose、lactose或sorbose至含燕麥聚木糖碳 源之培養基對T. longibrachiatum 185培養液pH值之影響...........................57 圖十八、連續三天添加1% avicel、cellulose、lactose或sorbose至含燕麥聚木糖碳 源之培養基對T. longibrachiatum 185孢子數之影響....................................58 圖十九、 Avicel、cellulose、lactose或sorbose添加至含燕麥聚木糖碳源之培養基之 頻率對T. longibrachiatum 185聚木糖之影響..........................................60 圖二十、 Avicel、cellulose、lactose或sorbose添加至含燕麥聚木糖碳源之培養基 之頻率對T. longibrachiatum 185纖維素之影響......................................61 圖二十一、Avicel、cellulose、lactose或sorbose添加至含燕麥聚木糖碳源之培養 基之頻率對T. longibrachiatum 185培養液p值之影響........................….62 圖二十二、 Avicel、cellulose、lactose或sorbose添加至含燕麥聚木糖碳源之培養 基之頻率對T. longibrachiatum 185孢子數之影響..................................63 圖二十三、添加混合醣纇(二種醣纇混合)至含燕麥聚木糖碳源之培養基對T. longibrachiatum 185聚木糖之影響...........................................................65 圖二十四、添加混合醣纇(二種醣纇混合)至含燕麥聚木糖碳源之培養基對T. longibrachiatum 185纖維素之影響...........................................................66 圖二十五、添加混合醣纇(二種醣纇混合)至含燕麥聚木糖碳源之培養基對T. longibrachiatum 185培養液pH值之影響......................................................68 圖二十六、添加混合醣纇(二種醣纇混合)至含燕麥聚木糖碳源之培養基對T. longibrachiatum 185孢子數之影響...............................................................69 圖二十七、添加不同脂質至含燕麥聚木糖之培養基對T. longibrachiatum 185聚木 糖之影響.................................................................................................71 圖二十八、添加不同脂質至含燕麥聚木糖之培養基對T. longibrachiatum 185纖維 素之影響................................................................................................72 圖二十九、添加不同脂質至含燕麥聚木糖之培養基對T. longibrachiatum 185培 養液pH值之影響........................................................................................73 圖三十、添加不同脂質至含燕麥聚木糖之培養基對T. longibrachiatum 185孢 子數之影響.....................................................................................................74 圖三十一、不同時機追加碳源、氮源或孢子數至含燕麥聚木糖培養基 中對T. longibrachiatum 185聚木糖之影響.........................................…77 圖三十二、不同時機追加碳源、氮源或孢子數至含燕麥聚木糖培養 基中對T. longibrachiatum 185纖維素之影響.....................................…78 圖三十三、不同時機追加碳源、氮源或孢子數至含燕麥聚木糖培養基 中對T. longibrachiatum 185培養液pH值之影響....................................….79 圖三十四、不同時機追加碳源、氮源或孢子數至含燕麥聚木糖培養 基中對T. longibrachiatum 185孢子數之影響.........................................….80 圖三十五、兩階段培養對T. longibrachiatum 185聚木糖之影響.........................….82 圖三十六、兩階段培養對T. longibrachiatum 185纖維素之影響.........................….83 圖三十七、兩階段培養對T. longibrachiatum 185培養液pH值之影響....................…..84 圖三十八、兩階段培養對T. longibrachiatum 185孢子數之影響.............................…..86 圖三十九、菌絲接種量對T. longibrachiatum 185在含燕麥聚木糖之培養基中 生長之影響..............................................................................................….87 圖四十、基質濃度對T. longibrachiatum 185聚木糖之影響.................................….89 圖四十一、基質濃度對T. longibrachiatum 185纖維素之影響.............................….90 圖四十二、基質濃度對T. longibrachiatum 185培養液pH值之影響.........................….93 圖四十三、基質濃度對T. longibrachiatum 185培養液中孢子數之影響..................….94 圖四十四、添加葡萄糖培養基對T. longibrachiatum 185聚木糖之影響..............….95 圖四十五、添加葡萄糖培養基對T. longibrachiatum 185纖維素之影響................96 圖四十六、添加葡萄糖培養基對T. longibrachiatum 185培養液pH值之影響.......….97 圖四十七、添加葡萄糖培養基對T. longibrachiatum 185孢子數之影響....................98 圖四十八、燕麥聚木糖(0.5%)添加至含1% avicel、cellulose、lactose及sorbose之 培養基對T. longibrachiatum 185聚木糖之影響.................................100 圖四十九、燕麥聚木糖(0.5%)添加至含1% avicel、cellulose、lactose及sorbose之 培養基對T. longibrachiatum 185纖維素之影響.................................101 圖五十、燕麥聚木糖(0.5%)添加至含1% avicel、cellulose、lactose及sorbose之培 養基對T. longibrachiatum 185培養液pH值之影響....................................102 圖五十一、燕麥聚木糖(0.5%)添加至含1% avicel、cellulose、lactose及sorbose之 培養基對T. longibrachiatum 185培養液孢子數之影響.....................…103 圖五十二、以混合性醣類(二種醣類混合)為基質對T. longibrachiatum 185聚木糖 之影響..............................................................................................105 圖五十三、以混合性醣類(二種醣類混合)為基質對T. longibrachiatum 185纖維素 之影響...........................................................................................….106 圖五十四、以混合性醣類(二種醣類混合)為基質對T. longibrachiatum 185培養液 pH值之影響............................................................................................108 圖五十五、以混合性醣類(二種醣類混合)為基質對T. longibrachiatum 185孢子 數之影響.................................................................................................109 圖五十六、以不同濃度之乳糖及纖維素組合為基質對T. longibrachiatum 185生 產聚木糖之影響.................................................................................110 圖五十七、以不同濃度之乳糖及纖維素組合為基質對T. longibrachiatum 185生 產纖維素之影響.................................................................................111 圖五十八、以不同濃度之乳糖及纖維素組合為基質對T. longibrachiatum 培養 液pH值之影響........................................................................................113 圖五十九、以混合性醣類(三種醣類混合)為基質對T. longibrachiatum 185聚木 糖之影響..............................................................................................114 圖六十、以混合性醣類(二種醣類混合)為基質對T. longibrachiatum 185纖維 素之影響..................................................................................................115 圖六十一、以混合性醣類(二種醣類混合)為基質對T. longibrachiatum 185培 養液pH值之影響.............................................................................…….117 圖六十二、以不同濃度之lactose、avicel及sorbose組合為基質 對T. longibrachiatum 185聚木糖之影響........................................….118 圖六十三、以不同濃度之lactose、avicel及sorbose組合為基質 對T. longibrachiatum 185纖維素之影響............................................120 圖六十四、以不同濃度之lactose、avicel及sorbose組合為基質 對T. longibrachiatum 185培養液pH值之影響..................................…..121 圖六十五、不同來源之農產廢棄物中聚戊糖含量之比較..............................…….123 圖六十六、半纖維素碳源之種類對T. longibrachiatum 185生產聚木糖之影響..124 圖六十七、半纖維素碳源之種類對T. longibrachiatum 185生產纖維素之影響..126 圖六十八、半纖維素碳源之種類對T. longibrachiatum 185培養液pH值之影響….127 圖六十九、半纖維素碳源之種類對T. longibrachiatum 185孢子數之影響.............128 圖七十、T. longibrachiatum 185經膠體過濾純化之聚木糖作用於紙漿 及茭白筍殼後產物之經時變化……………………………………130 表 目 表一、半纖維素分解酵素之種類....................................................................10 表二、液態Trichoderma spp.之生產聚木糖之條件.....................................15.....zh_TW
dc.language.isoen_USzh_TW
dc.publisher食品科學系zh_TW
dc.subjectAspergillus niger groupen_US
dc.subjectAspergillus nigerzh_TW
dc.subjectxylanaseen_US
dc.subjectoat spelt xylanen_US
dc.subjectpotassium nitrateen_US
dc.subjectsulfhydryl (-SH)en_US
dc.subject聚木糖zh_TW
dc.subject燕麥聚木糖zh_TW
dc.subject硝酸鉀zh_TW
dc.subject硫氫基zh_TW
dc.titleAspergillus niger NCH-3聚木糖之生產條件及酵素性質研究zh_TW
dc.titleStudies on the production and properties of xylanase from Aspergillus niger NCH-3en_US
dc.typeThesis and Dissertationzh_TW
Appears in Collections:食品暨應用生物科技學系
文件中的檔案:

取得全文請前往華藝線上圖書館



Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.