The metabolism of benzo(a)pyrene in cultured human colon has been investigated. Nontumorous colonie tissue was collected at the time of either surgery or "immediate autopsy" from patients with or without colonic cancer. After 24 hr in culture the expiants were exposed to [3H]benzo(a)pyrene for another 24 hr and the binding to cellular DMA and protein was measured. Two adducts, formed between benzo(a)pyrene and DMA, have been isolated. The major adduct (72 to 100%) was formed between the 10-position of benzo(a)pyrene diol-epoxide I and the 2-amino group of guanine, and the minor adduct was formed between benzo(a)pyrene diol-epoxide II and the 2-amino group of guanine. The major metabolites of benzo(a)pyrene extracted with ethyl acetate/acetone from the tissue culture media were (7,10/8,9)-tetrahydroxy-7,10,8,9-tetrahydrobenzo(a)pyrene, trans-7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene, and a peak containing (7,9,10/8)-tetrahydroxy-7,10,8-9-tetrahydrobenzo(a)-pyrene, (7/8,9)-trihydroxy-7,8-dihydrobenzo(a)pyrene, and frans-9,10, dihydroxy-9,10-dihy drobenzo(a )pyrene. The relative distribution of benzo(a)pyrene metabolites formed by cultured colon varied among individuals. About 10% of the metabolites remained in the water phase after extraction with ethy(acetate/acetone. frans-7,8-Dihydroxy-7,8-dihydrobenzo(a)pyrene and quiÃ±oneswere the major metabolites released when the water-soluble metabolites were treated with /3-glucuronidase and arylsulfatase. The binding levels of benzo(a)pyrene to DNA in cultured colon showed a unimodal distribution (56 cases). A slight difference (p < 0.1) in binding levels between nontumorous tissues from cancer patients (40 cases) and tissues from noncancer patients (16 cases) was observed, the binding level being highest in the latter. The binding level of benzo(a)pyrene to DNA also showed approximately a 5-fold variation among the different anatomical segments of the colon from the same patient; the binding level was generally highest in the ascending colon. Coincubation of the expiants with benzo(a)pyrene and either taurodeoxycholic acid or lithocholic acid increased the binding levels of benzo(a)pyrene to DNA. An increased level of frans-7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene in the culture media was also observed when expiants were coincubated with either taurodeoxycholic acid or lithocholic acid. These results indicate that cultured human colon can metabolize benzo(a)pyrene by pathways similar to those found in human bronchus and in cells of experimental animals.