A simple procedure for the precolumn derivatization of toxic and non-toxic non-protein amino acids occurring in the legume crop Lathyrus sativus and other Lathyrus species with phenyl isothiocyanate and an HPLC method for the separation of the derivatives in the nano- and picomole range are reported. The results are compared with those of conventional ion-exchange chromatography for the separation of non-protein amino acids with postcolumn ninhydrin reaction. The relative standard deviations for the two methods are compared.