An efficient, sensitive and reliable method for the simultaneous determination of dabsyl derivatives of numerous proteinogenic and physiological amino acids and biogenic amines in complex matrices by reversed-phase high-performance liquid chromatography (RP-HPLC) is described. A linear relation between peak area and concentration was observed from 1.25 to 1250 pmol and the detection limit was between 0.12 and 0.52 pmol. The average repeatability ranged between 1.3 and 3.1%, and the recovery values between 98 and 104%. An automated derivatization/injection unit clearly improved the performance of the method. Triethylamine was found to be a very effective additive to optimize separation efficiency. The method was successfully applied to the analysis of amino acids from protein hydrolyzates and of amino acid and biogenic amines of biological samples and food. More than 40 compounds could be separated simultaneously.