Melatonin, discovered in 1958, is secreted by the pineal gland primarily during the night. Its secretion is controlled by the light/dark cycle of the environment. Melatonin is also produced in and secreted by various extrapineal organs, tissues and cells and its synthesizing enzyme arylalkylamine N-acetyltransferase (AANAT) is expressed in various extrapineal organs, tissues and cells. Recently, it was reported that melatonin is present in saliva, but it is not certain where melatonin was synthesized and whether it was secreted into saliva and what function it may have in saliva. The present study was performed to investigate where melatonin was synthesized and whether it was secreted by salivary glands into saliva. We performed immunohistochemical analysis of the expression of AANAT in rat parotid, submandibular and sublingual glands and the expression of both AANAT and hydroxyindole-O-methyltransferase (HIOMT) in human submandibular glands. We evaluated the expression of AANAT and HIOMT mRNA in rat submandibular glands by quantitative reverse transcription-polymerase chain reaction. As a result, we observed expression of AANAT in epithelial cells of striated ducts in rat salivary glands and expression of AANAT, HIOMT and melatonin in epithelial cells of striated ducts in human submandibular glands. In addition, we also confirmed the expression of the most potent melatonin receptor, melatonin 1a receptor, in rat buccal mucosa. Our findings suggest that melatonin might be produced and secreted by salivary glands directly into saliva and that it might play some physiological role in the oral cavity.