Three groups of 10-days-old chicks were fed on one of three diets having phosphorus contents of 0.08 mol/kg, 0.14 mol/kg or 0.21 mol/kg. Ten days later duodenal calcium absorption by the ligated loop technique in vivo, and plasma calcium and phosphorus concentrations, were measured. In addition the metabolism in vitro of 25-hydroxycholecalciferol [25-(OH)D3] by kidney homogenates was studied. 2. In the low phosphorus group (0.08 mol/kg) calcium absorption and the activity of 25-(OH)D3-1-hydroxylase were significantly higher than those of the high phosphorus group (0.21 mol/kg). However, in the medium phosphorus group (0.14 mol/kg), calcium absorption was significantly higher although the activity of 25-(OH)D3-1-hydroxylase was not significantly higher when compared with the high phosphorus group (0.21 mol/kg). 3. It is concluded that in phosphorus deprivation, unlike in calcium deprivation, a diet very low in phosphorus is required to stimulate the renal 25-(OH)D3-1-hydroxylase activity.