In this study, we aimed to determine the relationship between chronotype, sleep problems and serum levels of GABA (gamma amminobutyric acid), glutamate and homocysteine in children and adolescents diagnosed with attention deficit and hyperactivity disorder (ADHD) as well as factors affecting this relationship. Sleep problems of 46 children and adolescents aged 7-18 years diagnosed with ADHD and 30 healthy volunteers aged 7-18 years were evaluated with Children's Sleep Habits Questionnaire (CSHQ) while chronotypes were evaluated with Children's Chronotype Questionnaire (CCQ). Serum glutamate, GABA and homocysteine levels were measured using immunosorbent test (ELISA) kits. Sleep problems were significantly more common in the ADHD group compared to the control group (p < .001). Serum GABA, glutamate and homocysteine levels were found to be predictor biomarkers for ADHD, independent of total sleep problem score. When the homocysteine levels were above the cut-off point of 9.445 mu mol/L, the sensitivity in early diagnosis of ADHD was 84.8% and the specificity was 70.0%. Although ADHD is a disorder in which sleep problems are common, increased serum GABA, glutamate and homocysteine are important in diagnosing ADHD independent of ADHD-related sleep problems. Homocysteine levels may be an important predictor for the presence of ADHD.