亲和介质及溶液条件对蛋白质溶液中内毒素去除的影响

Effect of affinity medium and solution conditions on endotoxin removal from protein solutions

摘要:

生物制品中内毒素的去除是一项十分重要的工作。为了更好地去除各种生物制品中的内毒素，采用合成的多粘菌素B琼脂糖亲和介质，通过静态吸附的方法去除蛋白质溶液中的内毒素。重点考察了介质的间臂长度、配基密度以及各种溶液条件 (pH值、盐种类和浓度、蛋白质种类和浓度、内毒素浓度、添加剂等) 对内毒素去除率及蛋白质回收率的影响。分别采用动态浊度法和Lowry法检测内毒素含量和蛋白质浓度。结果表明该介质具有载量高、去除速度快、去除率高、可重复使用的特点。此外，配基密度、pH值、盐浓度和蛋白质特性 (等电点和疏水性) 对内毒素去除效果均有重要影响。在优化的条件下，血红蛋白、人血清白蛋白和溶菌酶的回收率分别达到87.2％、73.4％和97.3％，相应的内毒素去除率分别达到99.8％、97.9％和99.7％。阐明了各种因素对内毒素去除率和蛋白质回收率的影响规律，为生物制品中内毒素的高效去除提供了参考。

Abstract:

Endotoxin removal is essential for the safety of biological products. To remove endotoxin efficiently, we used polymyxin B (PMB) affinity adsorbent to remove endotoxin from protein solutions by static adsorption. We studied the effects of spacer length and ligand density of the affinity adsorbent, pH, salt type and concentration, protein type and concentration, endotoxin concentration, and additive on endotoxin removal and protein recovery. Endotoxin content and protein concentration were determined by test and Lowry assay respectively. The results showed that PMB affinity adsorbent had high capacity, high adsorption speed, high removal efficiency and good reusability. In addition, ligand density, pH, salt concentration and the isoelectric point and hydrophobicity of protein all had remarkable influence on the endotoxin removal. Under the optimal conditions, the recoveries of hemoglobin, human serum albumin and lysozyme were 87.2%, 73.4% and 97.3%, respectively, and the corresponding endotoxin removal rates 99.8%, 97.9% and 99.7%, respectively. This study illustrated the effects of solution conditions on the efficiency of endotoxin removal and protein recovery, and would provide useful reference for the efficient removal of endotoxin from biological products.