N-Butyl-N-(4-hydroxybutyl) nitrosamine (BBN), which selectively induced urinary bladder tumors in several experimental animal species, was found to cause damage of DNA in the bladder epithelium. Wistar strain rats were given 100 mg/kg of BBN intravesically and killed after 2, 6, 12, 24, or 48 hr. DNA damage was examined by measuring the change in sedimentation pattern in an alkaline sucrose gradient. Amount of DNA in each fraction was determined by fluorescence spectrophotometry. At 2 hr, the sedimentation profile shifted from heavier (No 15, control peak) to lighter (Nos. 2 approximately 4) fractions, and the maximum effect appeared at 6 hr as a single peak in the lighter fractions. At 12 hr, damaged DNA was in the process of repair as two peaks were present, one light and one heavy. At 48 hr, the sedimentation profile showed a single peak identical with that of controls indicating complete repair of DNA.