Chapter 17 - Mixed-mode HPLC

Abstract

Separation in most common types of HPLC is dominated by few main types of interactions, such as hydrophobic in RP-HPLC, polar in HILIC, ionic in IC, etc. However, even in these cases, the interactions between the analyte, stationary phase, and mobile phase involve a mixture of other interactions. For example, RP-HPLC is also associated with some polar contributions, HILIC is associated with some hydrophobic interactions, etc. A proof of mixed mechanisms is, for example, the nonlinear or even the U-shape dependence of retention factor on the mobile phase composition that can be interpreted as a switch of the retention mechanism involving different interactions between the analytes and stationary phase under the influence of the mobile phase composition. A mixed mechanism is also encountered in case of SEC HPLC, which separates analytes by hydrodynamic volume but cannot eliminate some hydrophobic interactions with the stationary phase. The so-called mixed-mode HPLC is related to special stationary phases having in their compositions at least two different functionalities which can be involved in different main interactions with the analytes.