米曲霉综合利用与固定化氨基酰化酶拆分乙酰-DL-蛋氨酸-化学工程专业毕业论文.docxVIP

摘要本文系统地研究了应用固定化氨基酰化酶拆分乙酰．DL．蛋氨酸这一课 摘要 本文系统地研究了应用固定化氨基酰化酶拆分乙酰．DL．蛋氨酸这一课 题。首先研究了从实验室液相培养的米曲霉菌体中综合提纯胞内氨基酰化酶 和胞外。一淀粉酶的具体步骤及操作条件。确定了氨基酰化酶提取液的组成， 通过分步盐析、透析和浓缩使氨基酰化酶纯化了2．I倍，单位酶活可达 1000wm]，总收率为77．9％。确定了a～淀粉酶是米曲霉产的胞外酶，使用丙 酮沉淀法从发酵液中提取0～淀粉酶，得到的酶粉(含高岭土)酶活29．12u／g， 收率为89。8％。 使用实验室制各的氨基酰化酶溶液进行了固定化研究，对国产弱碱性大 孔阴离子交换树脂D380、D392及新研制的甲基丙烯酸酯类弱碱型树脂WB、 GMA、E／I-I和DEAE-E／H的固定化效果进行了比较，测得DEAE，E／H载体的 酶活收率和吸附效果最佳，固定化酶活力可达1400—1500u／g，且近似于定向 固定化。考察了固定化酶的最适反应温度、pH值、热稳定性和表观动力学常 数K：和吒。研究了该新型固定化氨基酰化酶拆分乙酰．DL一蛋氨酸反应的动 力学性质，并从理论上探讨了有关机理。将实验结果与Eadie-Hofstee图对照， 判断出反应体系中产物L-蛋氨酸的抑制类型为竞争性抑制，副产物乙酸的抑 制类型为非竞争抑制，并求解了底物、产物和副产物的抑制常数。 设计了固定化酶填充床反应器，研究了料液流速及底物浓度对反应器出 口转化率的影响。对固定床反应器中的连续拆分反应进行了模拟，模型计算 结果与实验值吻合较好。乙酰．L。蛋氨酸在料液流速为6ml／h，底物浓度为 0．1mol／L情况下出口转化率为98．48％。考察了系统的操作稳定性，37C下其 半衰期为27day。 关键词：提纯，固定化，氨基酰化酶，蛋氨酸，光学拆分 ABSTRACTThe ABSTRACT The optical resolution of Nmcetyl-·DL—methionine using the immobilized aminoacylase is systematically studied in the paper．Endocellu[ar aminoacylase of Aspergillus oryzae has been purified b(NH4)2S04 fractionation followed by dialysis and concentration．The components of extraction solvent are confirmed， and the purification fold is 2．1，the enzyme activity is 1000u／ml，the recovery is 77．9％．Aspergillus oryzae is found to secreteⅡ-amylase in liquid culture．By acetone fractionation the a．amylase powder’s activity is 29．12u／g and the recovery is 89．8％． A series of weak base type anioo exchangers have been used to adsorb aminoacylase purified above．The result is that the new prepared DEAE—E／H obtains excellent immobilized enzyme activity and retention percentage．The o皿mlml temperature and pH of the immobilized enzyme are measured and compared with the flee enzyme．The kinetic characters K。and吒of the tmsymmetricat hydrolysis of N-aoetyl-DL—methiordne are explored theoretically and experimemally．By compared the experimental results wi也Eadie—Hofstee figures．it is shown that the hrld of inhibition of L-methionine was competitive and the by-product acetic acid was noncompetitive，and the kinetic