UV absorption‐based inverse modeling of protein chromatography
UV absorption‐based inverse modeling of protein chromatography
复制标题
基于紫外吸收的蛋白质色谱反演模型
DOI：
10.1002/elsc.201400247
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发表时间：
2016
影响因子：
2.7
通讯作者：
J. Hubbuch
中科院分区：
文献类型：
--
作者：
Tobias Hahn;P. Baumann;Thiemo C Huuk;V. Heuveline;J. Hubbuch
紫外吸光度测量在生物过程开发中起着重要作用。在分析SEC或离子交换色谱法中，收率和纯度通常以峰百分比进行评价。此外，工业色谱步骤通常基于UV数据进行控制，并根据吸光度阈值进行合并决策。一旦模型被校准到特定的工艺步骤，基于模型的工艺开发将使精心设计的筛选实验变得多余。到目前为止，吸光度测量不能直接用于建模色谱步骤，因为常用的模型依赖于质量或摩尔浓度。这项研究提出了一个工业相关的色谱设置没有任何知识的饲料组合物的机械建模。重写模型方程，采用边界条件的UV吸光度单位，吸收系数转移到等温线，和标准的参数估计程序可以应用。大肠杆菌中表达的目标蛋白和11个集总杂质峰的阴离子交换色谱案例研究证明了实用性。根据色谱图估计进料中的目标蛋白浓度。使用这种方法，可以从单组分吸光度曲线后验确定离子交换和多峰色谱的初始未知进料浓度。
UV absorbance measurements play an important role in bioprocess development. Yield and purity are often evaluated in terms of peak percentages in analytical SEC or ion‐exchange chromatography. Also, industrial chromatography steps are usually controlled based on UV data with pooling decisions according to absorbance thresholds. Model‐based process development would make elaborate screening experiments redundant, once the model has been calibrated to the specific process step. So far, absorbance measurements could not be used directly for modeling chromatography steps as the commonly applied models rely on mass or molar concentration. This study presents mechanistic modeling of an industrially relevant chromatography setting without any knowledge of the feed composition. The model equations were rewritten to employ boundary conditions in UV absorbance units, the absorption coefficients were shifted into the isotherm, and standard parameter estimation procedures could be applied. An anion‐exchange chromatography case study of a target protein expressed in Escherichia coli and 11 lumped impurity peaks demonstrated practical applicability. The target protein concentration in the feed material was estimated from chromatograms. Using this method, initially unknown feed concentrations can be determined a posteriori for ion‐exchange and multimodal chromatography from single‐component absorbance curves.