Lactoferrin (LF) is a multifunctional protein found in various biological fluids. However, the peripheral action of lactoferrin remains unknown. In this study, peripherally applied bovine lactoferrin showed antinociceptive effect that was reversed by a mu-opioid, receptor antagonist, D-Phe-Cys-Tyr-D-Tip-Orn-Thr-NH2 (CTOP), or by a nitric oxide synthase (NOS) inhibitor, N-G-nitro-L-arginine methyl ester (L-NAME), but not by an inactive enantiomer of L-NAME, N-G-nitro-D-arginine methyl ester (D-NAME), during phase 1 and phase 2 in the rat formalin test. Peripheral coadministration of a mu-opioid receptor agonist, morphine, with subeffective dose of bovine lactoferrin produced a potentiated antinociceptive effect compared to that of morphine alone during both phases in the formalin test. This potentiated antinociception by morphine with bovine lactoferrin was reversed by CTOP or by L-NAME. These results suggest that bovine lactoferrin exerts an antinociceptive activity via potentiation of the peripheral mu-opioidergic system, and that nitric oxide (NO) is involved in this potentiation. (C) 2003 Elsevier B.V All rights reserved.