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Updated: Jul 10, 2026

Automated Hydrophobic Interaction Chromatography Column Selection for Use in Protein Purification
Published on: September 21, 2011
Determination of the chromatographic hydrophobicity index for ionisable solutes
Elisabet Fuguet1, Clara Ràfols, Elisabeth Bosch
1Departament de Química Analítica, Universitat de Barcelona, Diagonal 647, E-08028 Barcelona, Spain.
Abstract:
A model that relates the chromatographic hydrophobicity index (CHI) to the pH of the mobile phase has been tested in two of the most common high-performance liquid chromatography (HPLC) solvents: methanol and acetonitrile. A set of eight monoprotic acids and nine monoprotic bases of different chemical nature (phenols, benzoic acids, anilines, and pyridine) have been selected for the validation. The variation of CHI values with the pH of the mobile phase shows a good fit to the model for almost all compounds, regardless of their nature, and similar CHI values of the neutral form of the substances are obtained in both organic modifiers. On the contrary, higher differences are observed for the ionic form of the test solutes. The values of the other parameters obtained with the model (s, pK(a)) are discussed according to the nature of the compounds, and the variation of solvent pH and compounds pK(a) along the gradient elution.
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