Plasma concentrations of 1,25-dihydroxyvitamin D [1,25-(OH)2D], 24,25-dihydroxyvitamin D [24,25-(OH)2D], and 25,26-dihydroxyvitamin D [25,26-(OH)2D] were determined in 80 healthy infants of 4 days, 6 weeks, 6 months, and 12 months of age. The 4-day-old babies received breast milk, while the 6-week-old infants were either exclusively breast or formula fed. The older infants were on mixed diets and received daily vitamin D supplements. The levels were analyzed with regard to age and the concentrations of 25-hydroxyvitamin D (25OHD), calcium, phosphate, magnesium, and alkaline phosphatase and were compared with adult levels of vitamin D metabolites. The median 1,25-(OH)2D concentration was highest at 4 days of age and lowest at 6 weeks, but, except for the 6-week-old group, all had higher levels than the adults (6 weeks, P less than 0.1; others, P less than 0.01). 1,25-(OH)2D and 25OHD levels showed significant correlation only at 4 days (r = 0.74; P less than 0.0005), and there were no consistent relationships between 1,25-(OH)2D and the other variables. The median concentration of 24,25-(OH)2D was lower (P less than 0.01), while the 25,26-(OH)2D value was similar to that in the adults. Both were, however, positively related to the 25OHD level [24,25-(OH)2D, r = 0.82; 25,26-(OH)2D, r = 0.65; P less than 0.0005], as in the adults. The ratio of 24,25-(OH)2D to 25OHD was lower beyond 4 days of life than in the adults (medians, 3.4% vs. 5.1%; P less than 0.02). The data suggest that 1,25-(OH)2D synthesis has relative priority over 24,25-(OH)2D production during infancy compared with that in adulthood.