The association between light exposure at night and sex hormone levels in utero has scarcely reported. We assessed the associations between sleep duration or being awake in the late evening hours, which can be as indicator of light exposure at night, and the maternal and umbilical blood hormone levels during pregnancy and at delivery among Japanese women. The data for 236 women and their newborns who visited a maternal clinic in Gifu, Japan, between May 2000 and October 2001 were analyzed. Maternal blood samples were obtained at approximately the 10th weeks, 29th weeks of gestation, and at delivery. Umbilical cord artery blood was immediately drawn after birth. Information for sleep during pregnancy was obtained by a self-administered questionnaire. The levels of estradiol and testosterone were measured using radioimmunoassay. Maternal serum testosterone level in the 10th week was higher among those who were awake at or after 1:00 a.m. than among those who were asleep at that time (P = 0.032). Maternal estradiol level in the 29th week was inversely associated with sleep duration on weekends (P = 0.043). Umbilical testosterone level at delivery inversely correlated with sleep duration on weekdays (P = 0.030). These associations were somewhat stronger among mothers with female offspring than those with male offspring. These results suggested that exposure to light at night might increase sex hormone levels during pregnancy.