Implementations of temperature gradients in temperature-responsive liquid chromatography.

Abstract Temperature Responsive Liquid Chromatography (TRLC) offers an alternative and environmentally friendly way to perform reversed-phase like separations. Its use of temperature responsive polymers to control retention based on column temperature, instead of the fraction of organic modifier in the mobile phase mobile, eliminates the need for solvent composition gradients and allows, for example, for purely aqueous separations. In principle this temperature induced retention should allow for gradient elutions to be performed using downward temperature gradients to control retention and refocus the analyte peaks. Yet, the unavailability of dedicated commercial temperature controlling systems allowing suitable temperature control in TRLC limits implementations thereof often to isothermal or step gradient applications. In this work we study the potential of 1) a simple yet programmable water bath and of 2) a modified HPLC system allowing column temperature programming through controlled mixing of a warm and cold mobile phase streams. The performance of both systems was evaluated under both isocratic and gradient applications, resulting in a more thorough understanding of the influence of temperature gradients in TRLC. This knowledge is then applied to a sample of phenolic solutes, illustrating that, although both systems have some flaws, both are able to impose temperature gradients in TRLC resulting in significantly reduced retention and enhanced refocusing of the analyte peak.