View on hplc.cloudHEPES is a zwitterionic organic chemical buffering agent. It is a very polar compound which is not retained by traditional reverse phase chromatography. HEPES can be separated by mixed-mode hydrophilic interaction chromatography on the Newcrom AH mixed-mode column, which has both hydrophobic and cationic exchange properties. The mobile phase consists of H2O and Methanesulfonic acid. It can be detected using a conductivity detector.