The rates of metabolism of the carcinogenic 2-hydroxybenzo[a]pyrene (2-OH-B[a]P) and the non-carcinogenic 3- and 9-hydroxybenzo[a]pyrenes in cultured cell systems have been studied and compared. While 70-80% of the non-carcinogens are converted to water-soluble derivatives by hamster embryo fibroblasts in 24 h, carcinogenic 2-OH-B[a]P is metabolized at a slower rate (45% in 24 h), comparable to that for the parent hydrocarbon, benzo[a]pyrene (B[a]P). Analysis of extracellular organic solvent-soluble metabolites of 2-OH-B[a]P in cultured hamster embryo fibroblasts, using h.p.l.c., indicates the presence of a single major metabolite, which has been identified by mass spectroscopy as a dihydroxy derivative of B[a]P. At least one additional major organic solvent-soluble metabolite is formed in cultures of either mouse epidermal epithelial cells or human foreskin fibroblasts, indicating a different balance of metabolic pathways in these cell systems. The greater persistence of carcinogenic 2-OH-B[a]P in cells and its higher concentration in the cell cytoplasm compared with the non-carcinogenic phenols may be related to its relatively high biological activity. Differences in metabolism of 2-OH-B[a]P in several cultured cell systems indicate the importance of an appropriate choice of activating system in understanding the relationship between metabolism and carcinogenesis.