Schmidt Beata, Wandrey Christine, Freitag Ruth
Laboratory of Chemical Biotechnology, Institute of Chemical and Biological Process Sciences, Faculty of Basic Sciences, Swiss Federal Institute of Technology Lausanne, Ecublens, Lausanne CH-1015, Switzerland.
J Chromatogr A. 2003 Nov 14;1018(2):155-67. doi: 10.1016/s0021-9673(03)01326-8.
The overall topic of the investigation was the separation of basic proteins by cation exchange displacement chromatography. For this purpose two principal column morphologies were compared for the separation of ribonuclease A and alpha-chymotrypsinogen, two proteins found in the bovine pancreas. These were a column packed with porous particles (Macro-Prep S, 10 microm, 1000 A) and a monolithic column (UNO S1). Both columns are strong cation exchangers, carrying -SO3(-)-groups linked to a hydrophilic polymer support. Poly(diallyl-dimethylammonium chloride) (PDADMAC), a linear cationic polyelectrolyte composed of 100-200 quaternary pyrrolidinium rings, was used as displacer. The steric mass action (SMA) model and, in particular, the operating regime and dynamic affinity plots were used to aid method development. To date the SMA model has been applied primarily to simulate non-linear displacement chromatography of proteins using low molar mass displacers. Here, the model is applied to polyelectrolytes with a molar mass below 20000 g mol(-1), which corresponds to a degree of polymerization below 125 and an average contour length of less than 60 nm. The columns were characterized in terms of the adsorption isotherms (affinity, capacity) of the investigated proteins and the displacer.
该研究的总体主题是通过阳离子交换置换色谱法分离碱性蛋白质。为此，比较了两种主要的柱型用于分离核糖核酸酶A和α-胰凝乳蛋白酶原，这两种蛋白质存在于牛胰腺中。这两种柱分别是填充有多孔颗粒（Macro-Prep S，10微米，1000埃）的柱和整体柱（UNO S1）。两种柱都是强阳离子交换剂，带有连接到亲水性聚合物载体上的-SO3(-)-基团。聚（二烯丙基二甲基氯化铵）（PDADMAC），一种由100 - 200个季铵吡咯烷环组成的线性阳离子聚电解质，用作置换剂。空间质量作用（SMA）模型，特别是操作模式和动态亲和图被用于辅助方法开发。迄今为止，SMA模型主要应用于模拟使用低摩尔质量置换剂的蛋白质非线性置换色谱。在此，该模型应用于摩尔质量低于20000 g·mol⁻¹的聚电解质，这对应于聚合度低于125且平均轮廓长度小于60 nm。根据所研究蛋白质和置换剂的吸附等温线（亲和力、容量）对柱进行了表征。
