Recent advances in molecular biology and separation science have resulted in large-scale production of diagnostics and biotherapeutics. In downstream purification of the produced biomacromolecules, desirable bioseparation performance can be achieved using chromatography processes based on suitable supported matrix and affinity ligands. We have first carried out molecular dynamics (MD) simulations of biomolecular transport and adsorption onto a charged solid surface in an electrolytic solution to study bioseparation by ion-exchange chromatography. Simulation results confirmed the important role that electrophoretic migration plays in addition to thermal diffusion and demonstrated the greater importance of partial charge distribution over that of net charge in biomolecular adsorption and transport. Our study also uncovered the existence and effect of a critical lower bound of pore radius in a porous medium for effective bioseparation that has received little attention to date. Although these results can be expected to be still valid, the better selectivity of bioseparation by affinity chromatography results mostly from affinity ligands, whose functioning, however, is still not completely understood. In this respect, we have considered in our MD simulations an N-benzoylglycine-derived ligand which has recently been shown to be of great promise. We investigate the resultant potential energy fields of this ligand and its artificial isomers in order to obtain a better understanding of affinity chromatography and re-evaluate the conventional wisdom of lock-and-key mechanism. The implications of our simulation results in the selection of affinity ligands and in the scientific design of chromatography processes for effective bioseparation will also be discussed.