UV absorption‐based inverse modeling of protein chromatography

Abstract

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.

Model‐Based Investigation on the Mass Transfer and Adsorption Mechanisms of Mono‐Pegylated Lysozyme in Ion‐Exchange Chromatography

“…The chromatograms resulting from LGE and breakthrough experiments were used to estimate the parameters with the inverse method . The adaptive simulated annealing (ASA) yielding the first guess was followed by the Levenberg–Marquardt (LM) algorithm for the fine adjustment of the parameter estimates.…”

Systematic Interpolation Method Predicts Antibody Monomer-Dimer Separation by Gradient Elution Chromatography at High Protein Loads

“…Other approaches to the determination of the SMA parameters are possible including directly fitting their values to chromatograms at high‐protein loads. This is generally known as the “inverse method” and has been successfully applied to describing protein ion exchange chromatography using the SMA model . While powerful, we did not consider the inverse method here since our goal is to predict high load chromatography directly from isotherm data without fitting high‐load experimental chromatograms.…”

Calibration‐free inverse modeling of ion‐exchange chromatography in industrial antibody purification

“…The SMA isotherm [8], modified in [18], is a commonly used semimechanistic isotherm in ion-exchange chromatography. It is capable of reproducing the influence of counter ions on the retention behavior of protein species using the proteins' characteristic charges ν i .…”