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Introduction Fibroblast Growth Factor 21 (FGF21) is a novel metabolic factor with effect on glucose and lipid metabolism, and shown to be elevated in diseases related to metabolic syndrome. Due to the increasing frequency of metabolic syndrome in the pediatric population, and as FGF21 studies in children are limited, we investigated baseline serum levels of FGF21 in healthy children during an oral glucose tolerance test. Methods A total of 179 children and adolescents from the COPENHAGEN Puberty Study were included. An OGTT with glucose and insulin measurements, a dual energy X-ray absorptiometry (DXA) scan and a clinical examination including pubertal staging were done on all subjects. Serum levels of FGF21, adiponectin, and leptin were determined by immunoassays at baseline. Results The girls had significantly higher levels of FGF21 compared with boys (155 pg/mL vs. 105 pg/mL, P = 0.04). 38 children (21%) had levels below detection limit of assay. Baseline levels of FGF21 showed positive correlation with triglycerides, but no significant correlations were found between FGF21-concentration and body mass index (BMI), DXA-derived fat percentage, LDL- HDL- and non-HDL cholesterol, leptin or adiponectin levels, respectively. Neither was any correlation found between baseline FGF21-levels and the dynamic changes in glucose and insulin levels during the OGTT. Conclusion FGF21 is independent of adiposity in children, and the significant metabolic effect seems to be limited to pathological conditions associated with insulin resistance. The higher levels of triglycerides in the girls may explain the significantly higher levels of FGF21 in girls compared with boys. [ABSTRACT FROM AUTHOR]