Three-Dimensional Pore Structure of Chromatographic Adsorbents from Electron Tomography

Abstract

The pore structure of chromatographic adsorbents directly influences macromolecular partitioning and transport in chromatography. Quantitative structural characterization of chromatographic media has generally been performed in terms of the mean pore size or, at best, the pore size distribution (PSD), but more detailed information on, e.g., connectivity has been lacking. We have applied electron tomography, a 3D TEM technique that views a sample from multiple perspectives and allows reconstruction of the volumetric structure, to capture the internal details of microporous chromatographic media with nanometer-scale resolution. Visualization of reconstructions of three adsorbents, Toyopearl SP-650 C, SP-550 C, and CM Sepharose FF, provides thorough and direct information on the geometry and the interconnectivity of the pore network. The structures are qualitatively consistent with in situ AFM images, and quantitative data for the porosities and PSDs from the analysis of tomographic data agree reasonably well with inverse size-exclusion chromatography results. For a more straightforward representation of the networking and size features of the disordered pore space, a 3D thinning algorithm was used to derive pore skeletons and consequently quantitative data on distributions of local path lengths, widths, tortuosities, and connectivities. Such enriched structural information can be instrumental in more discriminate structural evaluation and construction of engineered pore models for the study of solute intraparticle transport.

High ionic strength narrows the population of sites participating in protein ion-exchange adsorption: A single-molecule study

“…A variety of sources can lead to the observed heterogeneity of protein adsorption, including intrinsic variance of the adsorbate molecule population due to differences in post-translational modifications, translational inaccuracy, and post-synthesis modifications such as deamidation and oxidation [6,16,17], inter-protein electrostatic repulsion and steric overlap [6,8], and irregularities in the adsorbent surface [18,19]. Because adsorption heterogeneity is observed even with uniform adsorbate populations and at very low loadings [20], it is likely that inherent heterogeneity of the adsorbent surface commonly plays the larger role.…”

Protein adsorption and transport in polymer-functionalized ion-exchangers

“…Furthermore, since uniformity of pore size is difficult to achieve, pore occlusion is also a possibility if the pore size is too small. A suggested optimum [14] is a pore size of order ten times the adsorbate size, although additional characteristics of the different adsorbent morphologies, including the porosity and the pore connectivity, are important considerations not discussed in detail in this simplistic analysis [15]. …”

“…The topography is generally defined by the base matrix [12], leading to pronounced differences between such cases as the fibrous structure of agarose and the more extended sphere-like surface of polymethacrylates [15,16]. However, introduction of the functional ligands can alter the local topography and hence the adsorbate-adsorbent interaction.…”

Modeling of dispersion in a polymeric chromatographic monolith

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