固定化细胞生物转化半纤维素水解液生产木糖醇
摘要
本论文的研究工作包括；玉米芯酶水解液的制备，玉米芯半纤维素水解液发酵生产木糖醇，微胶囊固定化方法发酵玉米芯水解液等内容进行了综述和分析。通过比较深入的研究工作，取的研究成果包括：
选用了高温蒸煮使木聚糖溶出，对蒸煮时间，蒸煮温度，固液比等条件做了研究，得出的结论是：将的玉米芯经5％氨水在60℃条件下浸泡12h，滤去浸泡液，水洗至中性，加水至固液比为1：10的比例，于170℃下密封蒸煮2h。将高温蒸煮液，加木聚糖酶水解，对酶解的条件进行了优化，最佳条件是高温蒸煮液加6％的酶量，在pH4.0，在45℃下搅拌速度为200r／min条件下，酶解8h。
研究结果表明，以玉米芯酶解液为底物摇瓶发酵生产木糖醇，最适的条件为：实验条件下20h的种子龄为最佳，初糖的浓度为40g／L、装液量为250mL的三角瓶装100mL的培养基、接种量为5％(体积比)、起始pH6.0，氮源组成：酵母膏和蛋白胨各2.5g／L、温度为30℃。在此条件下，木糖醇的得率可达到66.35％。在此基础上本文还研究了发酵罐中通气量对酵母发酵玉米芯酶水解液生产木糖醇的影响。结果表明：其中0h-20h，2vvm的通气量；20h-80h，1.0vvm的通气量，有利于木糖醇的积累。
本文采用壳聚糖-海藻酸钠微胶囊化酵母细胞，采用的微囊化的条件是，2.5％海藻酸钠溶液以1：5(v／v)的比例混和，通过注射器滴入浓度为1.5％的氯化钙溶液，钙化4小时，制得的海藻酸钙胶珠置于0.5％pH=6的壳聚糖溶液(溶于1.0％乙酸)中成膜15min；洗净后，最后在0.055mol／ml的柠檬酸溶液中液化25min。对影响微胶囊化细胞发酵的因素进行了正交试验，确定了在摇瓶中固定化细胞发酵发酵玉米芯半纤维素水解液的过程中，摇床的转速对木糖醇产量的影响最大，经过优化较好的发酵条件是，摇床转速其中0-20h，200r／min，20-64h，140r／min氮源含量3g／L酵母膏，3g／L的蛋白胨，固定化细胞凝胶珠与玉米芯半纤维素水解液体积比为1：3。
对壳聚糖-海藻酸钠微胶囊固定化，游离细胞，海藻酸钙固定化细胞发酵进行了比较，发现微胶囊固定化具有游离细胞和海藻酸钙固定化细胞发酵两者的特点。微胶囊固定化的发酵木糖醇得率比海藻酸钙固定化细胞发酵高为60.1％，而
浙江工业大学硕士论文
摘要
海藻酸钙固定化细胞发酵的木糖醇得率为49.3%。另外，微胶囊细胞发酵的发酵
周期与固定化细胞相似发酵周期为64h，游离细胞发酵周期为72h。比游离发酵
缩短10h左右。最后在摇瓶条件下，采用分批发酵方式，对微胶囊细胞的发酵玉
米芯酶水解液进行重复8批25d发酵，木糖醇得率平均为61.4ry0。
通过本文的研究，初步探讨了玉米芯酶解液发酵木糖醇的可行行，为进一步
的研究提供了依据。
The research work in this thesis includes: preparation corn cob hemicellulosic enzymatic production and xylitol fermentation from corn cob hemicllucellulosic, enzymatic hydrolysate , microcapsule was applied in xylitol of corn cob hemicellulosic enzymatic hydrolysate
Corn cob is composed of lignin cellulose and hemicellulose.It is difficulte to directly enzymatic hydrolysate.we must be pretreated .we select high temperature cooking solubility of the xylan . the optimum conditions were as follows:corn cob was pretreated by 0.1% acid at 60℃ for 12h. The best proportion of corncob with the water is 1 :10, high temperature cooking temperature is 170℃, reaction time is 2h. the optimum conditions of enzymatic hydrolysate as follows: xylanase 6% , at pH4.0 ,45℃, 200r/min reaction 8h in xylitol fermentation of corn cob hemicellulosic enzymatic production, the optimum conditions were as follows:xylose 40g/L, culture medium 100 ml in 250 ml flask, inoculum size, 5%; inoculum age, 20 h; nitrogen source. 2.5 g/L Bacto-yeast extract and 2.5 g/L Bacto-tryptone,temperature 30℃,In this condition ,the highest xylitol yield 66.35%.In order to evaluate the effect of aeration rate on the xylitol fermentation on corn cob hemicellulosic enaymatic production in fermentator ,batch fermentation
were carriec out two-stage aeration strategies,which provided relative higher aeration rate in the early stage but reduced it in the later stage,and including a one-stage aeration strategy that it provbided a constant aeration rate.The two-stategy oxygen supply strstegy is 2.0wm for the fist 20h,then lowered it to 1 .0vvm.
The preparation process of chitosan-alginate microcapsule is 0.5%chitosan solution (dissolved in 1% HAc, pH 6) and a simple droplet generator were employed to form chitosan-alginate microcapsules The best reaction time of the membrane's construction of microcapsules was found to be 15min, and the best reaction time of the liquefying of microcapsules to be 25min.
Chitosan-alginate microcapsule was applied in xylitol batch fermentation. The
microcapsules were used repeatedly in four batches. It was found that the fermentation with microcapsules had better and fixed conversion (55.17%), comparing with free-cells fermentation (changed from 67.5% to49.87%) and Ca-alginate immobilizing fermentation (changed from 41.23% to 45.67%), while the end time of microcapsule fermentation was enhanced and fixed to 64h.
The chitosan-alginate microcapsuleused for xylitol productin on corn cob hydrolysates. The optimum batch fermtentation conditions was as follows:shaking speed was divided into two stages:200r/min 0-20h,140r/min at 20-64h. nitrogen source. 2.0 g/L Bacto-yeast extract and 2.0g/L Bacto-tryptone,the volume propotion between gel beads of immobilized cells and hydrolysates.
The research work of thesis developed an xylitol fermentation process using corn cob hemicellulosic enzymatic hydrolysate in stead of acid hydrolysate .The technology is of importang value.
选用了高温蒸煮使木聚糖溶出，对蒸煮时间，蒸煮温度，固液比等条件做了研究，得出的结论是：将的玉米芯经5％氨水在60℃条件下浸泡12h，滤去浸泡液，水洗至中性，加水至固液比为1：10的比例，于170℃下密封蒸煮2h。将高温蒸煮液，加木聚糖酶水解，对酶解的条件进行了优化，最佳条件是高温蒸煮液加6％的酶量，在pH4.0，在45℃下搅拌速度为200r／min条件下，酶解8h。
研究结果表明，以玉米芯酶解液为底物摇瓶发酵生产木糖醇，最适的条件为：实验条件下20h的种子龄为最佳，初糖的浓度为40g／L、装液量为250mL的三角瓶装100mL的培养基、接种量为5％(体积比)、起始pH6.0，氮源组成：酵母膏和蛋白胨各2.5g／L、温度为30℃。在此条件下，木糖醇的得率可达到66.35％。在此基础上本文还研究了发酵罐中通气量对酵母发酵玉米芯酶水解液生产木糖醇的影响。结果表明：其中0h-20h，2vvm的通气量；20h-80h，1.0vvm的通气量，有利于木糖醇的积累。
本文采用壳聚糖-海藻酸钠微胶囊化酵母细胞，采用的微囊化的条件是，2.5％海藻酸钠溶液以1：5(v／v)的比例混和，通过注射器滴入浓度为1.5％的氯化钙溶液，钙化4小时，制得的海藻酸钙胶珠置于0.5％pH=6的壳聚糖溶液(溶于1.0％乙酸)中成膜15min；洗净后，最后在0.055mol／ml的柠檬酸溶液中液化25min。对影响微胶囊化细胞发酵的因素进行了正交试验，确定了在摇瓶中固定化细胞发酵发酵玉米芯半纤维素水解液的过程中，摇床的转速对木糖醇产量的影响最大，经过优化较好的发酵条件是，摇床转速其中0-20h，200r／min，20-64h，140r／min氮源含量3g／L酵母膏，3g／L的蛋白胨，固定化细胞凝胶珠与玉米芯半纤维素水解液体积比为1：3。
对壳聚糖-海藻酸钠微胶囊固定化，游离细胞，海藻酸钙固定化细胞发酵进行了比较，发现微胶囊固定化具有游离细胞和海藻酸钙固定化细胞发酵两者的特点。微胶囊固定化的发酵木糖醇得率比海藻酸钙固定化细胞发酵高为60.1％，而
浙江工业大学硕士论文
摘要
海藻酸钙固定化细胞发酵的木糖醇得率为49.3%。另外，微胶囊细胞发酵的发酵
周期与固定化细胞相似发酵周期为64h，游离细胞发酵周期为72h。比游离发酵
缩短10h左右。最后在摇瓶条件下，采用分批发酵方式，对微胶囊细胞的发酵玉
米芯酶水解液进行重复8批25d发酵，木糖醇得率平均为61.4ry0。
通过本文的研究，初步探讨了玉米芯酶解液发酵木糖醇的可行行，为进一步
的研究提供了依据。
The research work in this thesis includes: preparation corn cob hemicellulosic enzymatic production and xylitol fermentation from corn cob hemicllucellulosic, enzymatic hydrolysate , microcapsule was applied in xylitol of corn cob hemicellulosic enzymatic hydrolysate
Corn cob is composed of lignin cellulose and hemicellulose.It is difficulte to directly enzymatic hydrolysate.we must be pretreated .we select high temperature cooking solubility of the xylan . the optimum conditions were as follows:corn cob was pretreated by 0.1% acid at 60℃ for 12h. The best proportion of corncob with the water is 1 :10, high temperature cooking temperature is 170℃, reaction time is 2h. the optimum conditions of enzymatic hydrolysate as follows: xylanase 6% , at pH4.0 ,45℃, 200r/min reaction 8h in xylitol fermentation of corn cob hemicellulosic enzymatic production, the optimum conditions were as follows:xylose 40g/L, culture medium 100 ml in 250 ml flask, inoculum size, 5%; inoculum age, 20 h; nitrogen source. 2.5 g/L Bacto-yeast extract and 2.5 g/L Bacto-tryptone,temperature 30℃,In this condition ,the highest xylitol yield 66.35%.In order to evaluate the effect of aeration rate on the xylitol fermentation on corn cob hemicellulosic enaymatic production in fermentator ,batch fermentation
were carriec out two-stage aeration strategies,which provided relative higher aeration rate in the early stage but reduced it in the later stage,and including a one-stage aeration strategy that it provbided a constant aeration rate.The two-stategy oxygen supply strstegy is 2.0wm for the fist 20h,then lowered it to 1 .0vvm.
The preparation process of chitosan-alginate microcapsule is 0.5%chitosan solution (dissolved in 1% HAc, pH 6) and a simple droplet generator were employed to form chitosan-alginate microcapsules The best reaction time of the membrane's construction of microcapsules was found to be 15min, and the best reaction time of the liquefying of microcapsules to be 25min.
Chitosan-alginate microcapsule was applied in xylitol batch fermentation. The
microcapsules were used repeatedly in four batches. It was found that the fermentation with microcapsules had better and fixed conversion (55.17%), comparing with free-cells fermentation (changed from 67.5% to49.87%) and Ca-alginate immobilizing fermentation (changed from 41.23% to 45.67%), while the end time of microcapsule fermentation was enhanced and fixed to 64h.
The chitosan-alginate microcapsuleused for xylitol productin on corn cob hydrolysates. The optimum batch fermtentation conditions was as follows:shaking speed was divided into two stages:200r/min 0-20h,140r/min at 20-64h. nitrogen source. 2.0 g/L Bacto-yeast extract and 2.0g/L Bacto-tryptone,the volume propotion between gel beads of immobilized cells and hydrolysates.
The research work of thesis developed an xylitol fermentation process using corn cob hemicellulosic enzymatic hydrolysate in stead of acid hydrolysate .The technology is of importang value.
引文
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