Currently, commercial milk may contain abundant pregnancy-related hormones, the regular consumption of which puts children at a risk of precocious puberty and sex-hormone-associated tumors in adulthood. In this intervention trial, 51 healthy prepubescent children were randomly assigned to the intervention or control arms at a ratio of 3 : 1 to receive 250 or 600 mL m(-2) (body surface area) of milk intervention or matching equienergetic sugar water as the control. On testing cow's milk, progesterone was detected, while estrone, estradiol (E-2), and testosterone (T-2) were not. Cow's milk ingestion did not significantly influence the serum FSH, E-2, PRL, LH, and T-2 levels (P > 0.05) of pre-pubertal children 3 h after the intervention, while it increased their serum progesterone levels (P < 0.05) when compared with that in the control arm. Regarding the urinary hormone levels, cow's milk ingestion increased the urinary pregnanediol level within 4 h (P < 0.05), but not significantly when compared with that of the control (P > 0.05). The level of pregnanediol and E-2 in the morning urine for three consecutive days showed no significant difference between the two arms (P > 0.05). Drinking commercial milk with progesterone influenced the progesterone levels of pre-pubertal children in hours but not days and did not affect other sex hormone levels of pre-pubertal children.