We investigated the influence of sex and puberty stage on the circadian urine production and levels of antidiuretic hormone (AVP) in healthy children. Thirty-nine volunteers (9 prepuberty boys, 10 prepuberty girls, 10 midpuberty boys and 10 midpuberty girls) were included. All participants underwent a 24-hours circadian in-patient study under standardized conditions regarding sodium and fluid intake. Blood samples were drawn every four hours for measurements of plasma AVP, serum 17-β-estradiol and testosterone and the urine was fractionally collected for measurements of electrolytes, aquaporin 2 (AQP2) and prostaglandin E2 (PGE2). We found a marked night-time decrease in diuresis (from 1.69±0.08 to 0.86±0.06ml/kg/h,p<0.001) caused by a significant night-time increase in solute-free water reabsorption(TcH2O)(Day-Night ratio 0.64±0.07,p<0.001) concurrent with a significant decrease in osmotic excretion (Day-Night ratio 1.23±0.06,p<0.001). Plasma AVP expressed a circadian rhythm (p<0.01) with a night-time increase and peak levels at midnight (0.49±0.05 pg/ml). The circadian plasma AVP rhythm was not influenced by gender (p=0.56) or puberty stage (p=0.73). There was significantly higher night-time TcH2O in pre-puberty children. This concurred with increased night-time urinary AQP2 excretion in pre-puberty children. Urinary PGE2 exhibited circadian rhythm independent of sex or puberty stage. The levels of serum 17-β-estradiol and testosterone were as expected for sex and pubertal stage and no effect on AVP-AQP2- TcH2O axis was observed. This study finds a circadian rhythm of plasma AVP independent of sex and puberty stage, though night-time solute-free water reabsorption was higher and AQP2 excretion more pronounced in pre-puberty children suggesting higher pre-puberty renal AVP sensitivity.