Feedlot steers (n = 36) from three biological types (Bos indicus, Bos taurus-Continental, and Bos taurus-English) were used to determine the Ca, P, and vitamin D3 status of feedlot cattle. The USDA yield and quality grade traits were measured at slaughter, and the concentrations of vitamin D3 (VITD) and the metabolites 25-hydroxyvitamin D3 (25-OH D) and 1,25-dihydroxyvitamin D3 (1,25-(OH)2 D) were determined in LM, liver, kidney, and plasma. Plasma and muscle Ca and P concentrations also were determined. Biological type of cattle affected a number of carcass traits. Carcasses from Bos taurus-English cattle had more marbling, resulting in higher quality grades (P < 0.05). Carcasses from Bos taurus-Continental cattle had lower calculated yield grades (P < 0.05) than did carcasses from cattle in the other biological types. In general, differences in carcass traits resulting from biological type were consistent with other reports. Plasma and LM Ca and P concentrations were not affected (P = 0.06) by biological type of cattle, indicating that Ca and P homeostasis is a conserved trait across the different types of cattle. Plasma VITD and 25-OH D concentrations were not affected (P = 0.41) by biological type, whereas plasma 1,25-(OH)2 D concentration was lower (P < 0.05) in Bos taurus-English cattle than in Bos taurus-Continental and Bos indicus cattle. Liver VITD and 25-OH D were not affected by biological type (P = 0.76), but liver 1,25-(OH)2 D concentration was greater (P < 0.05) in Bos indicus cattle than in Bos taurus-Continental cattle. Kidney vitamin D metabolite concentrations were not affected by biological type of cattle (P = 0.21). Muscle VITD concentration was greater (P < 0.05) in Bos taurus-English cattle than in the other two biological types, and muscle 25-OH D concentrations were greater (P < 0.05) in Bos taurus-English cattle than in Bos indicus cattle. Muscle 1,25-(OH)2 D concentration was less (P < 0.05) in the Bos taurus-Continental cattle than in the other two biological types. Cooking eliminated vitamin D metabolite differences among the biological types. Our results suggest that Bos indicus cattle had greater 1,25-(OH)2 D (the biologically active form) in tissues, and greater 1,25-(OH)2 D plasma concentrations than Bos taurus cattle. Thus, the need for VITD supplementation and optimal levels of Ca and P in feedlot diets might differ between Bos indicus and Bos taurus cattle.