Chemopreventive effects of bovine lactoferrin (bLF), which is found at high concentrations in colostrum, on rat bladder carcinogenesis were investigated using a rat bladder medium-term bioassay. In experiment 1, a total of 80 F344 male rats, 6 weeks old, were divided into 5 groups. Groups 1 and 2 were treated with 0.05% N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) in the drinking water for 8 weeks and after a 1-week interval, received dietary supplementation with 2% and 0.2% bLF, respectively. Group 3 received 0.05% BBN for 8 weeks and then no treatment. Group 4 was administered 2% bLF alone from week 9, without prior carcinogen exposure. Group 5 was maintained without any treatment throughout the experiment. All rats were killed at the end of week 36. Group 1 demonstrated a significantly decreased multiplicity of the bladder tumors (carcinomas and papillomas) as compared with group 3. Maximum cut surface areas of bladder tumors were also significantly decreased in groups 1 and 2 compared with group 3. No bladder tumors were observed in groups 4 or 5. In experiment 2, a total of 60 rats were divided into two groups (30 rats each); both were treated with 0.05% BBN for 4 weeks and after a 1-week interval, one received 2% bLF (group 1) and the other, basal diet (group 2) for 4 weeks. Group 1 demonstrated a tendency for decrease of the 5-bromo-2'-deoxyuridine (BrdU) labeling index. bLF was detected in the urine of rats fed bLF by ELISA as well as western blot analysis. The findings indicate that 2% bLF can inhibit BBN-induced rat bladder carcinogenesis, and that this may be due to bLF in the urine.