Shape-Selective Separation of Model Analytes in Normal-Phase Liquid Chromatography: A Combined Simulation-Experimental Study

In this study, we demonstrate that liquid chromatographic separation can be effectively achieved based on shape differences, even for analytes of very similar chemical characters. Using a combined experimental-theoretical approach, we investigated the retention behavior of spherical buckminsterfullerene C60 and disk-shaped coronene at a hydroxylated silica stationary phase, with a mobile phase composed of toluene and n-hexane at varying compositions. High-performance liquid chromatography (HPLC) measurements revealed that increasing the n-hexane content enhances the separability of the two analytes, primarily due to coronene's stronger retention. Molecular simulations, coupled with a two-state model, attributed this effect to the structured layering of toluene at the stationary phase, which differentially influences analyte-wall interactions. Our analysis of Henry coefficients further identified the second solvent layer as the primary region governing adsorption, providing a thermodynamically consistent description of the separation process. These findings highlight the role of shape anisotropy in chromatographic retention and suggest new avenues for designing shape-selective separation strategies for molecules and nanoparticles in liquid chromatography.