Reversed Phase Chromatography
Starting in the mid 1970s reversed phase liquid chromatography (RPC) has become the standard technique to analyze small molecular weight compounds in industrial, academic and governmental laboratories. More recently, RPC has become the accepted tool for the separation of peptides, proteins and other biopolymers in the pharmaceutical, chemical and biochemical industries.
The opposite of normal phase chromatography, RPC requires a non-polar stationary phase and a polar mobile phase. Typically the mobile phase consists of a mixture of water (buffer) and acetonitrile, methanol, THF, or 2-propanol.
Silica particles are most commonly used as the support, which then is derivatized with octadecylsilane (ODS). Polymer-based supports have been introduced as an alternative to silica-based reversed phase columns, particularly for analyzing basic compounds in their neutral state at high pH.
RPC columns can be applied to the analysis of a wide variety of compounds, ranging from neutral polar and non-polar solutes to acidic, basic, and amphoteric compounds. RPC is also an efficient technique for the analysis of derivatized amino acids, peptides and proteins, although protein structure is not always maintained due to the high concentration of organic solvent required for their elution.
Tosoh Bioscience offers 14 distinct Reversed Phase column types which are either based on silica or methacrylate particles.