Fundamentals of reversed phase chromatography: Thermodynamic and exothermodynamic treatment

Reversed phase chromatography (RPC) is the most popular branch of HPLC for the analysis and purification of a wide variety of substances. Despite significant advances in both our knowledge and understanding of the fundamental principles governing the retention behavior in RPC, there is considerable debate in the literature regarding the mechanism of retention. This review addresses the theoretical foundation of the chromatographic technique, with an emphasis on thermodynamic and exothermo-dynamic treatment of retention equilibrium, as well as their implication on the mechanistic aspects of RPC retention. A unified and rigorous treatment based on the solvophobic theory is reviewed in terms of its ability to shed light on the physicochemical underpinnings of the retention in RPC, and to quantitatively predict the retention behavior of nonpolar compounds, acids and bases, and peptides and proteins. Also highlighted are areas of future challenges in the theory and practice of RPC, potentially leading to a better quantitative understanding and use of the popular technique.