title: Further investigations of the effect of pressure on retention in ultra-high-pressure liquid chromatography.
abstract: In this study, we investigated further the large increases in retention with pressure that we observed previously in RP-LC especially for ionised solutes. These findings were initially confirmed on a conventional silica C(18) column, which gave extremely similar results to the hybrid C(18) phase originally used. Large increases in retention factor of approximately 50% for a pressure increase of 500 bar were also shown for high MW polar but neutral solutes. However, experiments with the same bases in ionised and non-ionised forms suggest that somewhat greater pressure-induced retention increases are found for ionised solutes. Retention increases with pressure were found to be considerably smaller for a C(1) column compared with a C(18) column; decreases in retention with increasing pressure were noted for ionised bases when using a bare silica column in the hydrophilic interaction chromatography (HILIC) mode. These observations are consistent with the partial loss of the solvation layer in RP-LC as the solute is forced into the hydrophobic environment of the stationary phase, and consequent reduction in the solute molar volume, while the water layer on the surface of a HILIC packing increases the hydration of a basic analyte. Finally, retention changes with pressure in RP-LC can also be observed at a mobile phase pH close to the solute pK(a), due to changes in pK(a) with pressure. However, this effect has no influence on the results of most of our studies.
authors.affiliation: Centre for Research in Biomedicine, University of the West of England, Frenchay, Bristol BS16 1QY, UK.
authors.forename: Morgane M
authors.initials: MM
authors.lastname: Fallas
authors.forename: Uwe D
authors.initials: UD
authors.lastname: Neue
authors.forename: Mark R
authors.initials: MR
authors.lastname: Hadley
authors.forename: David V
authors.initials: DV
authors.lastname: McCalley
mesh_terms: D000814:Aniline Compounds / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D016014:Linear Models; D011312:Pressure
chemical_list: D000814:Aniline Compounds