Measurements of dietary selenium absorption and retention were obtained after administration of a single dose of the extrinsic stable isotope tag 74Se in 20 appropriate for gestational age premature infants with birth weights between 720 and 1,630 g and gestational ages between 26 and 33 weeks. Infants were assigned randomly to receive a standard premature formula (1.34 microgram of Se/dl) or a selenium-supplemented version of that formula (2.03 micrograms of Se/dl). Each study consisted of one feeding that had been extrinsically labeled with 74Se (1.03 microgram/kg) and a timed stool and urine collection. The percent 74Se absorption was 91.2 +/- 5.4% (mean +/- SD) from the standard formula and 86.2 +/- 3.0% from the selenium-supplemented formula (p less than 0.05), but the percent of the absorbed 74Se retained was not different, i.e., 96.6 +/- 2.1% and 95.0 +/- 2.8%, respectively. The percent net absorption and net retention were also not different between the standard and selenium-supplemented formulas; net absorption was 72.7 +/- 18.1% vs. 67.8 +/- 18.8% and net retention was 57.2 +/- 17.6% vs. 53.3 +/- 20.2%, respectively. The percent 74Se absorption and true selenium absorption were significantly correlated with the percent net selenium absorption and net selenium absorption, respectively. We conclude that an extrinsically administered dose of 74Se can be used to study selenium nutrition in growing premature infants.