Aromatic glycinonitriles and methylamines as pre-column fluorescence derivatization reagents for catecholamines

The reactivity of some catecholamines with aromatic glycinonitriles (AGN) (four species) and aromatic methylamines (AMA) (five species) was investigated in detail, to find pre-column fluorescence derivatization reagents for catecholamines. Of the nine reagents tested, 2-phenylglycinonitrile (PGN) and benzylamine (BA) were shown to be the best reagents in terms of selectivity and sensitivity. The reagents react selectively with catecholamines under mild conditions in the presence of ammonium molybdate and sodium periodate for PGN and potassium hexacyanoferrate(III) for BA to give fluorescent derivatives. The derivatives of four catecholamines (epinephrine (E), norepinephrine (NE), dopamine (DA) and isoproterenol (IP)) could be separated within 13 min by reversed-phase liquid chromatography with isocratic elution and measured fluorimetrically. The detection limits (signal-to-noise ratio = 3) are in the range 5.2–11.0 fmol for PGN and 1.6–100 fmol for BA in a 50 μl injection volume. The liquid chromatographie (LC) methods with PGN and BA were successfully applied to the determination of some catecholamines in human urine.