Assessment of intra-particle diffusion in hydrophilic interaction liquid chromatography and reversed-phase liquid chromatography under conditions of identical packing structure

A recently developed stripping protocol to completely remove the stationary phase of reversed-phase liquid chromatography (RPLC) columns and turn them into hydrophilic interaction liquid chromatography (HILIC) columns with identical packing characteristics is used to study the underlying mechanisms of intra-particle diffusion in RPLC and HILIC. The protocol is applied to a column with a large geometrical volume (250 x 4.6 mm, 5 mu m) to avoid extra-column effects and for compounds with a broad range in retention factors (k" from similar to 0.6 to 8). Three types of behavior for the intra-particle diffusion (D-part/D-m) in RPLC versus HILIC can be distinguished: for nearly unretained compounds (k" < 0.6), intra-particle diffusion in HILIC is larger than in RPLC; for compounds with intermediate retention behavior (k" similar to 0.9-1.2), intra-particle diffusion in HILIC and RPLC are similar; and for well retained compounds (k" > 1.8), intraparticle diffusion in RPLC is larger than in HILIC. To explain these observations, diffusion in the stationary phase (gamma D-s(s)) and in the stagnant mobile phase in the mesopore zone (gamma D-mp(m)) are deduced from experimentally determined values of the intra-particle diffusion, using models derived from the Effective Medium Theory. It is demonstrated that the larger intra-particle diffusion obtained for slightly retained compounds under HILIC conditions is caused by the higher mesopore diffusion in HILIC (gamma(mp) = 0.474 for HILIC versus 0.435 for RPLC), while the larger intra-particle diffusion obtained for strongly retained compounds, under RPLC conditions can be related to the much higher stationary phase diffusion in RPLC (gamma D-s(s)/D-m = 0.200 for RPLC versus 0.113 for HILIC). (C) 2017 Elsevier B.V. All rights reserved.