The effect of temperature and mobile phase composition on separation mechanism of flavonoid compounds on hydrosilated silica-based columns

We investigated the effects of mobile phase composition on the retention of flavones on four different hydrosilated C silica-based columns in buffered aqueous acetonitrile. Cogent UDC cholesterol™ and Cogent bidentate C18™ columns show significant dual reversed-phase/normal-phase retention behavior, while Cogent Diamond hydride™ and Cogent Silica hydride™ columns show negligible retention in the reversed-phase mode. The effect of the aqueous acetate buffer concentration on retention factors of flavones over the full mobile phase composition range, including both aqueous normal-phase (ANP) and reversed-phase mechanisms, can be described by a four-parameter equation for dual-retention mechanism. At increasing temperature, the retention factors and peak widths decrease both in the aqueous normal phase and in the reversed phase mobile phase range. In agreement with van’t Hoff model, linear ln k versus 1/T plots were observed, showing a single retention mechanism in the highly organic normal-phase and in highly aqueous reversed-phase mobile phase ranges. From among the stationary phases tested, Cogent UDC cholesterol™ column has high temperature stability (up to 100 °C) and provides most selective and efficient separations of flavones both in the ANP and in the RP modes with almost reversed elution order.

► Flavonoid compounds were separated on four hydrosilated columns.
► Flavonoids show dual ANP–RP mode retention mechanism.
► Flavonoid glycosides and aglycones show different retention behavior.
► Increasing temperature decreases the retention of flavonoid compounds.
► Entropic and enthalpic contributions to the retention were determined.