It has been suggested that shift work, night work in particular, affects worker's psychophysical health. However, the effects of night work on the health of pregnant women are not physiologically well elucidated. The effects of night work on the biological function of pregnant women were studied in the present study. Three pregnant and six non-pregnant nurses that engaged in fastrotating shift system cooperated for the study. The 24-h urine samples were collected in two time frames, daytime (07:00 to 23:00) and nighttime (23:00 to 07:00) on the day shift, the night shift and the days off. Urinary concentrations of 6-sulfatoxymelatonin and norepinephrine were measured by enzyme-linked immunosorbent assay and estriol by radioimmunoassay. The excretion profiles of urinary 6-sulfatoxymelatonin in the pregnant and the non-pregnant women were altered on the night shift, which might relate the derangement of circadian rhythm of melatonin secretion. The urinary norepinephrine level during the night work was considerably higher in the pregnant women, which indicated the presence of higher physical burden in them due to the night work. The urinary estriol level was not significantly affected by shift, day/night and individual factors, and the effect of night work on urinary estriol level of the pregnant women remained uncertain. The results of the present study suggested that the night work of the pregnant shift workers should be much more relieved.