OBJECTIVE In this work, we analysed the mechanism of action of caffeine on peroxidase secretion in the female rat submandibular gland. The signaling molecules cAMP and nitric oxide were monitored as potential mediators. DESIGN The salivary gland peroxidase secretion of female albino Wistar rats was assessed by a spectroscopic method. RESULTS Caffeine was found to exert an increase on peroxidase secretion in a concentration-response manner: the peroxidase secretion stimulation index (SI) (secreted peroxidase from treated/secreted peroxidase from basal) for caffeine 10 microg/ml: 2.2+/-0.18 (P<0.05); caffeine 100 microg/ml alone: 3+/-0.18 (P<0.01); +LNMMA (LN monomethyl arginine): 1+/-0.1 (P<0.05); caffeine 1000 microg/ml alone: 5+/-0.35 (P<0.01); +LNMMA: 2+/-0.2 (P<0.05). These results were associated with an increase in cAMP and total nitrites production. Total nitrites, SI caffeine 100 microg/ml alone: 2.8+/-0.2 (P<0.01); +LNMMA: 1+/-0.08 (P<0.05); caffeine 1000 microg/ml alone: 4.8+/-0.3 (P<0.01); +LNMMA: 2.3+/-0.18 (P<0.05). CONCLUSION It could thus be concluded that cAMP and NO are involved in the mechanism of action displayed by caffeine. This is the first report on the mechanism of action of caffeine on peroxidase secretion.