For decades the prevailing perception was that satisfactory re-equilibration of reversed-phase columns following gradient elution took a long time. In the early 2000s we showed that this perception was not well founded, and demonstrated that adequate re‑equilibration could be achieved in seconds. Recently, we have shown the same for HILIC columns. All of this work so far has been with small molecules. In this article, we present an overview of this work, and summarize the practical utility of it all.
Many users have questions about how long LC columns should be re-equilibrated following solvent gradient elution separations. Here we show satisfactory results with short re-equilibration periods, for both reversed-phase and HILIC separations.
When preparing buffers for hydrophilic interaction chromatography (HILIC) separations, is the buffer concentration considered relative to the aqueous portion of the eluent only, or to the aqueous-organic mixture?