The aim of this module is to explain the purpose of the mobile phase in HPLC. The basic concepts of the liquid chromatographic process is outlined, as well as discussing the best way to prepare mobile phase components. Potential problems caused by improper preparation and use of mobile phases will be highlighted. We will also describe the way that mobile phase composition effects retention and selectivity in reverse phase HPLC. The various physicochemical properties of the typical solvents used for HPLC are described, and the various methods for degassing HPLC mobile phases introduced.

At the end of this module you will be able to demonstrate an understanding of the purpose of the mobile phase in HPLC and describe suitable methods to prepare and degas your mobile phase. The liquid chromatographic process will be easy for you to describe and you will be able to relate retention and selectivity of analytes to different mobile phase compositions. You should also be able to demonstrate an understanding of how certain physicochemical properties of mobile phase constituents may affect an analysis.

Topics include:

• The mobile phase - introduction
• The liquid chromatographic process
• Analyte retention processes
• The partition coefficient
• Retention and selectivity
• Solvent type and selectivity
• Optimizing selectivity using retention
• Gradient elution
• Mobile phase delivery
• Solvent miscibility
• UV cut off
• Other solvent considerations
• Mobile phase preparation
• Degassing
• Outlet degassing
• Degassing/column damage
• Methods of degassing
• Helium sparging
• Vacuum degassing