We recently reported fleroxacin significantly affected cell proliferation in a dose-dependent manner in transitional cell carcinoma cell lines. In this study, we investigated the effect of fleroxacin on mouse bladder carcinogenesis with N-butyl-N-(4-hydroxybutyl) nitrosamine (BHBN). Five-week-old C57BL/6 female mice were divided into three groups. The forced oral administrations of fleroxacin (10 or 50 mg/kg/day) were done for 1 week, then the 0.05% BHBN was given for 8 or 12 weeks. Fleroxacin treatments were continued until sacrifice. The mice were sacrificed at 4 weeks or 8 weeks latent period after BHBN administration, and the histological changes in the bladder were examined. Our study suggested that fleroxacin tended to suppress the development of bladder carcinoma or the malignant changes induced by a shorter period of BHBN administration (8 weeks) in mice. However, in the group of the BHBN administration for 12 weeks, there were no significant effects on the prevention of bladder carcinoma in both of the treatment groups with fleroxacin (10 or 50 mg/kg/day). This study did not make it clear that oral administration of fleroxacin suppressed the development of bladder carcinoma induced by BHBN in mice. However, it is very preliminary data and further experiments need to be done with a much higher dose of the fleroxacin or with other kinds of fluoroquinolone antibiotics.