Oral 25-hydroxycholecalciferol (25OHD3) has been used to prevent early neonatal hypocalcemia. Thirty premature neonates were studied prospectively from birth: 22 premature infants constituted a control group, and eight were treated orally with 25OHD3 (2 microgram/kg/day) for five days beginning within the first 12 hours of life. Eleven of the 22 premature infants in the control group became hypocalcemic, with serum ionized calcium (Ca++) level less than 3.0 mg/dl and total calcium (CaT) level less than 7.0 mg/dl on day 2 of life. Only one of the eight premature infants treated with 25OHD3 became hypocalcemic. In treated neonates, serum 25-hydroxyvitamin D, as well as serum Ca++ and CaT levels, increased during the first five days of life and were considerably greater than levels in control neonates. No untoward effects of this treatment were observed.