Protease and xylanase activities and thermophilic populations as potential process monitoring tools during thermophilic aerobic digestion

Thermophilic aerobic digestion (TAD) of potato process waste slurry was carried out in a batch process in a continuously stirred tank reactor at 55 °C for a total of 156 h. The pH of the slurry was either unregulated, or regulated at 6.0, 7.0, 8.0, 9.0 and 9.5, and aeration rate was 0.5 vvm. The effect of aeration rate on the digestion process was studied at 0.1, 0.25, 0.5 and 1.0 vvm at pH 7.0 and 55 °C. The development of thermophiles (55 and 65 °C populations) and hydrolytic enzyme activities (protease and xylanase) in the process were monitored. Thermophiles developed rapidly to reach peak populations in 24h or earlier, and remained stable at all fixed pH reactions. The thermophile population was only minimally affected by the aeration rate, and they were present mostly as spores after 96h of digestion. Xylanase enzyme appeared rapidly, reached a peak in approximately 60h, and declined rapidly thereafter. Highest activities were produced in neutral reactions and higher aeration rates. Aeration rates affected protease activity profoundly. The profile of both enzymes closely reflected the development of microbial activity and the overall progress of TAD, in a medium where their substrates were not predominant and not the preferred carbon sources. The relatively simple process of measuring the activities of these enzymes is potentially a more direct measure of the progress of TAD than enumeration of the microbial populations and thus, has greater potential in process monitoring. Copyright © 2003 Society of Chemical Industry     

实验设计法在优化表达Fc融合蛋白的CHO细胞发酵培养工艺中的应用

目的通过实验设计(Design of Experiments,DOE)分析优化重组CHO细胞在生物反应器中的培养工艺。方法以2 L生物反应器控制参数温度(T)、pH及溶氧(dissolved oxygen,DO)作为变量,以蛋白表达量、蛋白纯度及蛋白唾液酸含量为响应变量,应用Design Expert 10软件进行3因素2水平DOE拟合分析,筛选最适表达Fc融合蛋白的CHO细胞发酵培养工艺参数,并采用5 L生物反应器进行验证。结果蛋白表达量仅与T有关,且呈成正相关,pH和DO对其无显著影响,T与pH对其有交互作用;蛋白纯度受T和pH影响显著,T呈负相关,pH呈正相关,DO对其无显著影响,T与pH对其有交互作用;唾液酸含量受T和pH影响显著,T呈正相关,pH呈负相关,DO对其无显著影响。最适工艺参数确定T为(34±0.5)℃,pH为(7.05±0.02),DO为(45±5)%。采用最适工艺参数的反应器与优化前工艺比较,活细胞密度(viable cell density,VCD)增加;蛋白表达量、蛋白纯度、唾液酸含量分别提高了29.7%、8%和87.9%。结论通过DOE方法快速优化了生物反应器培养表达Fc融合蛋白的CHO细胞的工艺参数,在不降低蛋白产量及纯度的前提下,极大提高了融合蛋白唾液酸含量。