Description of the retention behavior and chromatographic measurement of the change in pKa with temperature of a diastereomeric pair of isoleucine derivatives

The retention of a diastereomeric pair of peptide analogs was investigated as a function of temperature using reversed-phase HPLC. The retention of these compounds showed anomalous behavior with retention increasing with increasing temperature. The change in the compounds' pK(a) as temperature was varied was measured chromatographically and decreases with increasing temperature. The change in degree of ionization as a function of temperature was used to explain this anomalous retention behavior.