Catecholestrogens, 2-methoxyestrogens and "classical" estrogens (estrone, estradiol, estriol) were measured simultaneously in serum and urine samples of 220 pregnant women from the 8th week of pregnancy until to delivery. From these data we established the central 0.80 centile intervals as time specified reference intervals for each substance analyzed. Serum and urinary estradiol rise steadily during the progress of pregnancy, whereas estrone, catecholestrogens and 2-methoxyestrogens reach a plateau during the last trimester. These observations support the hypothesis, that the amount of the latter compounds may be regulated by separate mechanisms. The values of concentration and excretion of 2- and 4-substituted estrogens varied widely throughout pregnancy. Even very high or very low concentrations of these substances had no recognizable relation to the outcome of pregnancy. This supports the assumption that catecholestrogens and their methylethers are metabolites without any regulatory function in pregnancy.