Background <p>Exercise offers many benefits, but its role in ketone body (KB) metabolism is unclear in children. Aerobic exercise enhances KB uptake into muscle and improves mitochondrial function. Reduced ketone body concentration may indicate improved mitochondrial function. We hypothesized that physical activity would reduce circulating ketone levels in children.</p> Methods <p>Healthy young Black girls (<i>n</i> = 201) were randomized to a control condition or a daily, after-school exercise intervention. Participants (<i>n</i> = 53) with missing blood samples at baseline or follow up were excluded, 148 (75 intervention and 73 control) participants analyzed. The intervention entailed 30 min of homework time and 80 min of physical activity, including 35 min of moderate to vigorous physical activity over 10 months. Circulating ketone concentration was measured using nuclear magnetic resonance spectroscopy (NMR) and maximal oxygen consumption (VO₂ max) assessed oxidative capacity.</p> Results <p>Participants had a mean age of 9.5 years and BMI percentile of 69.9 at baseline. After adjusting for age, BMI percentile and breast development, decreased circulating ketone levels (mean = 248–170 μM, 31.3% change, Cohen’s D = 0.402, <i>p</i> = 0.017) were observed in the treatment group.</p> Conclusion <p>Exercise reduces KB levels in young girls, highlighting metabolic adaptation to physical activity programming in children consistent with improved mitochondrial function.</p> Impact <p><UnorderedList Mark="Bullet"> <ItemContent> <p>Existing literature does not consider the effects of long-term exercise on basal ketone concentration.</p> </ItemContent> <ItemContent> <p>Exercise intervention decreased serum ketone body concentration in girls.</p> </ItemContent> <ItemContent> <p>This is evidence for changes in nutrient utilization with exercise in active children.</p> </ItemContent> <ItemContent> <p>Puberty does not impact basal ketone utilization</p> </ItemContent> <ItemContent> <p>This change could serve as indication of changes in mitochondrial function</p> </ItemContent> </UnorderedList></p>