Carbon and oxygen isotope values reveal resource partitioning among the large mammal fauna from three contemporaneous Middle Pleistocene hominid-bearing localities within the Sierra de Atapuerca (northern Spain). Carbon isotope values sampled from the tooth enamel of fauna present during Atapuerca Faunal Unit 6 show that a C₃-dominated ecosystem surrounded the area where fossils were preserved during this time. For the herbivores, Fallow deer isotope values are significantly different from Red deer and horses and show that this species did not forage in open environments at this locality. Red deer and horses show similar feeding strategies with less negative carbon values implying use of more open environments for these taxa. For the carnivores, carbon isotope values for Ursus deningeri are significantly different from either lions (Panthera leo) or foxes (Vulpes vulpes) and support the contention that this species is herbivorous. Special metabolic mechanisms involved in hibernation in U. deningeri might also have influenced its isotope values. The carbon isotope values of remaining carnivores were similar and suggest that each was typically a generalist carnivore, eating a wide variety of prey items. While the isotopic results generally correspond to ecology indicated by previous techniques, this study shows that isotope analyses can provide further insights that alternate techniques do not provide. Isotope analyses can help elucidate the ancient ecology of taxa present in the Sierra de Atapuerca during the Middle Pleistocene allowing for an accurate portrayal of the setting in which humans lived.