Karlsson David, Jakobsson Niklas, Axelsson Anders, Nilsson Bernt
Department of Chemical Engineering, Lund University, SE-221 00 Lund, Sweden.
J Chromatogr A. 2004 Nov 5;1055(1-2):29-39. doi: 10.1016/j.chroma.2004.08.151.
A method using a model-based approach to design and optimize an ion-exchange step in a protein purification process is proposed for the separation of IgG from a mixture containing IgG, BSA and myoglobin. The method consists of three steps. In the first step, the model is calibrated against carefully designed experiments. The chromatographic model describes the convective and dispersive flow in the column, the diffusion in the adsorbent particles, and the protein adsorption using Langmuir kinetics with mobile phase modulators (MPM). In the second step, the model is validated against a validation experiment and analyzed. In the third and final step, the operating conditions are optimized. In the optimization step, the loading volume and the elution gradient are optimized with regard to the most important costs: the fixed costs and the feed cost. The optimization is achieved by maximizing the objective functions productivity (i.e. the production rate for a given amount of stationary phase) and product yield (i.e. the fraction of IgG recovered in the product stream). All optimization is conducted under the constraint of 99% purity of the IgG. The model calibration and the analysis show that this purification step is determined mainly by the kinetics, although as large a protein as IgG is used in the study. The two different optima resulting from this study are a productivity of 2.7 g IgG/(s m3) stationary phase and a yield of 90%. This model-based approach also gives information of the robustness of the chosen operating conditions. It is shown that the bead diameter could only be increased from 15 microm to 35 microm with maximum productivity and a 99% purity constraint due to increased diffusion hindrance in larger beads.
提出了一种基于模型的方法来设计和优化蛋白质纯化过程中的离子交换步骤，用于从含有免疫球蛋白G（IgG）、牛血清白蛋白（BSA）和肌红蛋白的混合物中分离IgG。该方法包括三个步骤。第一步，根据精心设计的实验对模型进行校准。色谱模型描述了柱内的对流和扩散流、吸附剂颗粒内的扩散以及使用带有流动相调节剂（MPM）的朗缪尔动力学的蛋白质吸附。第二步，根据验证实验对模型进行验证并分析。在第三步也是最后一步，对操作条件进行优化。在优化步骤中，针对最重要的成本：固定成本和进料成本，对进样体积和洗脱梯度进行优化。通过最大化目标函数生产率（即给定固定相量的生产速率）和产品收率（即产品流中回收的IgG比例）来实现优化。所有优化均在IgG纯度为99%的约束条件下进行。模型校准和分析表明，尽管本研究中使用的是像IgG这样大的蛋白质，但该纯化步骤主要由动力学决定。本研究得出的两个不同的最优结果是生产率为2.7 g IgG/(s·m³)固定相，收率为90%。这种基于模型的方法还给出了所选操作条件的稳健性信息。结果表明，由于较大珠子中扩散阻碍增加，在最大生产率和99%纯度约束条件下，珠子直径只能从15微米增加到35微米。
