Analysis of dipeptide mixtures by the combination of ion-pair reversed-phase high-performance liquid chromatographic and gas chromatographic-mass spectrometric techniques☆

Abstract Reversed-phase high-performance liquid chromatography using aliphatic carboxylic acids as surfactants was used to separate basic, acidic and neutral dipeptides. The enhancement of the capacity factors of basic dipeptides versus the chain length of carboxylic acid followed sigmoidal curves. The capacity factor of dipeptides fell precipitously with addition of salts until it decreased to a value which was unaffected by the addition of more salts. For the identification of dipeptides produced by the hydrolysis of proteins by proteases (dipeptidylaminopeptidases), gas chromatographic-mass spectrometric techniques were used for acidic and neutral dipeptides. Basic dipeptides could be positively identified either by normal chromatographic procedures or by collecting the suspected peaks for further characterization by alternate techniques.