We have previously demonstrated that oral administration of bovine lactoferrin (bLF) markedly increases CD4+ and CD8+ T cells and NK (asialoGM1+) cells in the blood of tumor-bearing mice and enhances anti-metastatic activity. In this paper, we document that oral administration of bLF and bLF-hydrolysate (bLFH) is associated with strong increases in CD4+ and CD8+ T, as well as asialoGM1+ cells in lymphoid tissues and lamina propria of the small intestine in mice, especially in tumor-bearing animals in which Co26Lu cells were implanted subcutaneously. Moreover, IgM+ and IgA+ B cells in lamina propria of the small intestine were also significantly increased by bLF and bLFH. Bovine apo-transferrin (bTF) did not exhibit such activity. In the colon, only CD8+ cells were significantly increased by treatment with bLF, while asialoGM1+ cells were significantly decreased. bLF and bLFH induced cytokines to activate T, B and asialoGM1+ cells. Administration of bLF and bLFH, but not bTF, increased production of interleukin-18 (IL-18), interferon-gamma (IFN-γ ) and caspase-1 in the mucosa of the small intestine. Particularly high levels of IL-18 were found in the epithelial cells of the small intestine. Moreover, administration of bLF and bLFH, but not bTF, induced IFN-γ presenting cells in the small intestine. Caspase-1, which processes proIL-18 to mature IL-18, was also induced in the epithelial cells of the small intestine following treatment with bLF and bLFH, but not with bTF. These results suggest that enhanced production of IL-18 and IFN-γ and caspase-1 induction by treatment with bLF may be important for elevation of intestinal mucosal immunity.