SIZE EXCLUSION CHROMATOGRAPHY OF DNA AND VIRUSES - PROPERTIES OF SPHERICAL AND ASYMMETRIC MOLECULES IN POROUS NETWORKS

This article attempts a general theoretical description of size exclusion chromatography (SEC) derived from novel supportive experiments on some key issues as well as from critically reviewed data from the literature. The elution volume of SEC is stringently derived from thermodynamic principles as the volume accessible to the center of gravity of a solute within a porous matrix. It is thus related to the size of the solute. This size is defined energetically from the interaction energy between the solute and the matrix wall and thus entails a 'hard core contour" of the solute plus interfacial interaction energies and entropic terms. Variation of the ionic strength of the aqueous solvent and comparison of solutes of widely different size and shape demonstrate that electrostatic forces alone are insufficient to explain the experimental results. Instead, retarded van der Waals forces are crucial. Below some 3 nm, hydration forces dominate, and interfacial separation becomes independent of electrostatic forces. Due to the observed huge size dependency of interfacial separation, porous networks, established by these phenomena, are nonetheless permeable to smaller macromolecules, which is crucial for biology. Given the proper description of interfacial separation for a particular mutual orientation between the matrix wall and an arbitrary shaped solute, the average overall configurations, i.e., a proper rotational average of the solute molecule, must be taken. It appears that a hydrodynamic equivalent radius based on intrinsic viscosity is best suited to define the rotational averaging of the hard core contour. The remaining discrepancies, probably related to insufficient definition of some systems and their nonchromatographic reference data, remain to be settled, however. The quest for universal calibration of SEC has thus been generalized into a comprehensive physicochemical description of permeated porous materials.