Phosphorus and calcium balance was measured prospectively in stable premature infants (less than or equal to 1600 gm) fed human milk or a standard commercial formula. Throughout the study, the P and Ca intakes of the infants fed human milk were two to three times less than those of infants fed formula. Infants fed human milk showed low serum P and normal serum Ca concentrations, complete renal reabsorption of P, and elevated renal Ca excretion. The net effect in infants fed human milk was a 50% reduction in the P and Ca retention, compared with the formula-fed group. Despite the unfavorable P and Ca balance in the group fed human milk, the only evidence of rickets was elevated alkaline phosphatase activity. Nevertheless, based on the biochemical changes in these infants, low serum P values, and excess urinary calcium losses, we conclude that the stable small premature infant fed human milk exclusively is deficient in phosphorus and only slightly more sufficient in calcium.