The cellular concentration of reduced glutathione (GSH) modulates the sensitivity of human melanoma cells to alkylating drugs in vitro. To investigate whether the membrane-associated enzyme gamma-glutamyl transpeptidase (gamma-GTP) involved in GSH breakdown was expressed in melanoma cells, the enzymatic activity of gamma-GTP as well as the secretion of GSH were measured in human melanoma cells from four different cell lines (Me8, JUSO, GLL19, Swift). All the cells showed low gamma-GTP activities (0-1 mU/mg protein) and released GSH in culture supernatants at significant rates. After incubation for 24 h in growth medium containing 0.1 mmol/L cystine, the levels of GSH in supernatants ranged from 56 to 111 nmol GSH/mg protein. The GSH metabolism of melanoma cells was also evaluated by measuring the levels of the melanogenesis intermediate 5-S-cysteinyldopa under different experimental conditions. The results of these experiments suggest that melanoma cells have a low ability to metabolize the tripeptide GSH, which appears to be responsible for GSH secretion and accumulation in culture supernatants.