Experimental and clinical studies suggest that endogenous melatonin plays an important role in pediatric sleep regulation. This finding led to the introduction of exogenous melatonin to treat sleep disturbances. Optimizing the treatment algorithm involves a review of melatonin measurements and interpretations in clinical practice. Diurnal patterns of salivary melatonin and urinary metabolite 6-sulfatoxymelatonin were investigated in 29 children and adolescents (age, 5.5-17.3 years) by measuring concentrations every 3 hours. Relationships between melatonin parameters (peak concentrations and area under the time curve) and anthropometric measures (height, weight, and body mass index), age, and sleep scores (Sleep Disturbance Scale for Children) were investigated. High interindividual variability was evident in melatonin diurnal profiles. Melatonin production (adjusted to body weight) decreased with age and sexual maturation (P < 0.00). Both salivary melatonin and its urinary metabolite measurements can be used to evaluate the melatonin system in children. However, the high interindividual variability of diurnal melatonin concentrations challenges clinical applications in regard to diagnostic purposes and the criteria for initiating exogenous melatonin therapy. Further investigations and the development of criteria for clinical evaluations of the pediatric melatonin system are needed.