The mechanism of formation of 5-S-cysteinyldopamine (5-S-CDA), a putative index of oxidative stress in dopaminergic regions of the brain, was investigated by comparing the ability of a number of neurochemically relevant oxidising systems to promote the conjugation of dopamine with cysteine in vitro. Autoxidation of the catecholamine proceeds at relatively slow rate in the physiological pH range, and is little affected by 1 mM Fe(2+)-EDTA complex. In the presence of cysteine, however, the Fe(2+)-induced autoxidation is hastened, affording little amounts of 5-S-CDA. Formation of the adduct is completely suppressed by ascorbic acid. Hydrogen peroxide, in the presence of Fe(2+)-EDTA (Fenton-type oxidation) or peroxidase, promotes a relatively efficient conversion of dopamine to 5-S-CDA and the minor isomer 2-S-CDA. Noteworthy, 15-hydroperoxyeicosatetraenoic acid (arachidonic acid hydroperoxide, HPETE), in the presence of Fe(2+)-EDTA complex, can also mediate 5-S-CDA formation, whilst superoxide radicals are little effective. Overall, these results suggest that ferrous ions, hydrogen peroxide and lipoperoxides may play an important role in 5-S-CDA generation.