Study on Submerged Culture of Mushroom Mycelium by Bubble Column Fermentors

本研究由鼓泡塔生物反应器深层培养蘑菇菌丝体的研究和反胶束分离蛋白质的研究两部分组成。I.鼓泡塔生物反应器深层培养蘑菇菌丝体：我们设计了一种新型的气升塔，该塔具有锥形提升管，有望适用于蘑菇菌丝体的深层培养。对气升塔内的流体力学(气含率和液体循环速度)和氧气传输率进行了实验和理论研究。研究了在标准鼓泡塔中悬浮培养蘑菇菌丝体时常形成的大球形颗粒的悬浮条件。其次，利用裂殖酵母生产L-苹果酸和规模化培养松茸两种蘑菇菌丝体进行深层培养。确定了各菌种液体深层培养的最适培养基组分、温度和通风量。反胶束提取蛋白质：由于蘑菇菌丝体能产生许多有用的蛋白质，因此对反胶束提取蛋白质进行了研究。找到了反胶束提取溶菌酶的最小AOT浓度和最佳AOT浓度。结果表明，含有细胞色素C和溶菌酶的二元蛋白质混合物的最小AOT浓度小于单一蛋白质体系中每种蛋白质的最低AOT浓度之和。在料液中加入低浓度的胍盐作为前处理，降低了AOT的最低浓度，增加了正向萃取率，使蛋白质能够从胶束有机相中成功回收，而海藻糖的加入则降低了正向萃取率。使用非离子Span 60反胶束，在高于CMC的浓度下成功地提取了细胞色素c。

This study is composed of two parts ; the study on submerged culture of mushroom mycelium by bubble column bioreactors, and the study on separation of protein by reverse micelles.I.Submerged culture of mushroom mycelium by bubble column bioreactors : We devised a novel type of airlift column with a tapered riser which is anticipated to be suitable to the submerged culture of mushroom mycelium. The hydrodynamics (gas holdup and liquid circulation velocity) and the oxygen transfer rate in the airlift column were investigated empirically and theoretically. Conditions to suspend large spherical pellets, which are often formed for the submerged culture of mushroom mycelium, in the standard bubble column were investigated. Next, submerged culture of mushroom mycelium was carried out using two species of mushroom : one is productipn of L-malic acid by Schizophyllum commune, and the other is large-scale culture of Tricholoma matsutake. The optimum conditions of the submerged culture on medium composition, temperature, and air flow rate were found for each mushroom.2. Extraction of protein by reverse micelles : Since mushroom mycelium produces many useful proteins, extraction of protein by reverse micelles was investigated. We found the minimal and optimal AOT concentration for the extraction of lysozyme using reverse micelles. The minimal AOT concentration of a binary protein mixture containing cytocbrome c and lysozyme was found to be smaller than the sum of the minimal AOT concentration of each protein obtained in single protein systems. The addition of low concentration of guanidium salts into the feed solution as a pretreatment decreased the minimal AOT concentration, increased the forward extraction, and enabled the successful recoverry of the protein from the micellar organic phase, while the trehalose addition reduced the forward extraction. Using non-ionic Span 60 reverse micelles, cytochrome c was successfully extracted at concentration higher than the CMC.