Hydrophobicity of ionisable compounds studied by countercurrent chromatography.

Abstract

Countercurrent chromatography (CCC) is a liquid chromatography technique in which the stationary phase is also a liquid. The main chemical process involved in solute separation is partitioning between the two immiscible liquid phases: the mobile phase and the support-free liquid stationary phase. The octanol–water partition coefficients (Po/w) is the accepted parameter measuring the hydrophobicity of molecules. It is considered to estimate active principle partitioning over a biomembrane. It was related to the substance biological activity. CCC is able to work with an octanol stationary phase and an aqueous mobile phase. In this configuration, CCC is a useful and easy alternative to measure directly the Po/w of the molecules compared to other methods including the classical and tedious shake-flask method. Three ketones are used as model compounds to illustrate the CCC protocol of Po/w measurement. The focus of this work is put on ionisable molecules whose apparent Po/w is completely changed by ionization. β-Blockers, diuretics and sulfonamides are compound classes that were studied. Some of the experimentally determined Po/w coefficients of the molecular forms disagreed with calculated and experimental values available in the literature. The Po/w coefficients of the ionic forms and the acidity constants were also calculated using a theoretical model. Relationships between biological properties and hydrophobicity are also discussed.