McCue Justin T, Engel Philip, Ng Austen, Macniven Rich, Thömmes Jörg
Biogen Idec Corporation, Bioprocess Development, 14 Cambridge Center, Cambridge, MA 02142, USA.
Bioprocess Biosyst Eng. 2008 Apr;31(3):261-75. doi: 10.1007/s00449-008-0200-1. Epub 2008 Jan 19.
Hydrophobic interaction chromatography (HIC) is commonly used to separate protein monomer and aggregate species in the purification of protein therapeutics. Despite being used frequently, the HIC separation mechanism is quite complex and not well understood. In this paper, we examined the separation of a monomer and aggregate protein mixture using Phenyl Sepharose FF. The mechanisms of protein adsorption, desorption, and diffusion of the two species were evaluated using several experimental approaches to determine which processes controlled the separation. A chromatography model, which used homogeneous diffusion (to describe mass transfer) and a competitive Langmuir binary isotherm (to describe protein adsorption and desorption), was formulated and used to predict the separation of the monomer and aggregate species. The experimental studies showed a fraction of the aggregate species bound irreversibly to the adsorbent, which was a major factor governing the separation of the species. The model predictions showed inclusion of irreversible binding in the adsorption mechanism greatly improved the model predictions over a range of operating conditions. The model successfully predicted the separation performance of the adsorbent with the examined feed.
疏水作用色谱法（HIC）常用于蛋白质治疗药物纯化过程中分离蛋白质单体和聚集体。尽管使用频繁，但HIC的分离机制相当复杂，尚未得到充分理解。在本文中，我们使用苯基琼脂糖凝胶FF研究了单体和聚集体蛋白质混合物的分离。采用多种实验方法评估了两种物质的蛋白质吸附、解吸和扩散机制，以确定哪些过程控制了分离。构建了一个色谱模型，该模型使用均相扩散（描述传质）和竞争性朗缪尔二元等温线（描述蛋白质吸附和解吸）来预测单体和聚集体的分离。实验研究表明，一部分聚集体物质不可逆地结合到吸附剂上，这是控制物质分离的主要因素。模型预测表明，在吸附机制中纳入不可逆结合在一系列操作条件下大大改善了模型预测。该模型成功预测了所研究进料条件下吸附剂的分离性能。
