Relation of protein structure and adsorbent pore architecture to adsorption and transport in chromatography

Chromatography is commonly used in the recovery and purification of therapeutic biologics. An improved mechanistic understanding of protein chromatography would be helpful for the development of effective adsorbents and efficient processes.; Protein sets with varying degrees of structural differences were studied to relate protein structure to retention in ion-exchange chromatography. The modeling of protein-adsorbent electrostatic interactions at the continuum level yields semi-quantitative descriptions of the retention of highly homologous protein variants. It also helps to rationalize differences in the observed adsorbent-dependent retention of two fibroblast growth factors based on different binding mechanisms. Comparative analysis for proteins with different arginine-lysine compositions suggests that hydration effects must be incorporated for full quantitative modeling of retention in ion-exchange. The trajectories of water molecules near proteins were obtained using molecular dynamics simulations, which can provide the basis for probing its influence on retention by considering the restructuring of water during protein binding.; The pore structure of chromatographic adsorbents has a direct influence on performance parameters. Inverse size-exclusion chromatography (ISEC) was used to determine the pore size distributions (PSD) of a number of commercial adsorbents, which could then be used to provide approximate predictions of the binding capacities. Analysis of transport and breakthrough behavior indicates that both the PSD and connectivity are necessary for an accurate description of the intraparticle transport.; For a more thorough investigation of structural properties of porous adsorbents, transmission electron microscopy (TEM) techniques were exploited. 2D TEM studies yield valuable information on the extent of structural heterogeneity, and electron tomography captures the 3D internal structures at nanometer resolution. The PSDs and porosities from the analysis of tomographic data agree reasonably well with ISEC results. Furthermore, a number of structural descriptors at the pore level, such as the path length, tortuosity, and connectivity, were made available by the medial axis analysis. Such enriched information is valuable for a better comparative evaluation of pore structure and the construction of simulated pore networks for the modeling of molecular transport in microporous adsorbents.