Growing evidence support the role of vitamin D in brain function and behavior. This study investigated the relationship between 25-hydroxyvitamin D-3 [25(OH)D-3] levels, biochemical profile and symptoms of depression and anxiety in healthy individuals. Symptoms of depression were assessed by the Beck Depression Inventory (BDI) and anxiety was evaluated with the State-Trait Anxiety Inventory (STAI). Our sample included 36 individuals, mostly women 27(75%), 36.39 +/- 9.72years old, non-smokers 31(86.1%), body mass index of 26.57 +/- 3.92kg/m(2), 27.95 +/- 7.50% body fat. Participants were divided into those with 25(OH)D-3 levels lower than 40ng/mL (mean 28.16 +/- 7.07) and equal or higher than 40ng/mL (mean 53.19 +/- 6.32). Those with lower 25(OH)D-3 had higher levels of triacylglycerol, triacylglycerol/high density lipoprotein (HDL) ratio, high glucose and homeostatic model assessment of insulin resistance (HOMA-IR) index. No changes were observed in sociodemographic variables, body composition, inflammatory parameters and cortisol. Additionally, in the groups with low and high 25(OH)D-3 levels, STAI state, STAI trait and BDI scores were not statistically different. Levels of 25(OH)D-3 were inversely and independently associated with glucose and HOMA-IR, but not associated with triacylglycerol, depression and anxiety scores. Lower levels of 25(OH)D-3 were associated with dysfunction in glucose metabolism but not with depression and anxiety in healthy individuals.