Vitamin D-deficient pigs, rats and chicks were repleted with four daily oral doses of crystalline ergocalciferol (vitamin D2) and cholecalciferol (vitamin D3) containing equal concentrations of each. At 24 h after the last dose, the plasma of each species was analysed for vitamin D and 25-hydroxyvitamin D by standard methods. The mean (+/- S.D.) ratios of plasma cholecalciferol to ergocalciferol concentration were 1.5 +/- 0.1 (pig). 1.7 +/- 0.1 (rat) and 6.3 +/- 1.2 (chick). The mean ratios of plasma 25-hydroxycholecalciferol to 25-hydroxyergocalciferol concentration were 4.0 +/- 0.1 (pig), 0.4 +/- 0.02 (rat) and 10.7 +/- 3.4 (chick). The mean plasma cholecalciferol/ergocalciferol ratios for the 24,25-dihydroxy-, 25,26-dihydroxy- and 1,25-dihydroxy-derivatives in the pig were 2.6 +/- 0.6, 5.8 +/- 1.3 and 5.8 +/- 0.8 respectively. This is the first evidence that mammals other than the New World monkey, like birds, discriminate between ergocalciferol and cholecalciferol. These data, therefore, suggest that species discrimination between the different forms of vitamin D is probably a general phenomenon in mammals. Moreover, this is the first indication of a species (rat) that discriminates against a cholecalciferol metabolite in favour of an ergocalciferol metabolite. Species discrimination against particular forms of vitamin D may be important to the choice of experimental animal models for studying the regulation of vitamin D metabolism and may also be an important consideration in dietary vitamin supplementation.