Bioactive alloplastic materials, like beta-tricalcium phosphate (β-TCP) and calcium sulfate (CS), have been extensively researched and are currently used in orthopedic and dental bone regenerative procedures. The purpose of this study was to compare the performance of EthOss versus a bovine xenograft and spontaneous healing. The grafting materials were implanted in standardized 8 mm circular bicortical bone defects in rabbit calvariae. A third similar defect in each animal was left empty for natural healing. Six male rabbits were used. After eight weeks of healing, the animals were euthanized and the bone tissue was analyzed using histology and micro-computed tomography (micro-CT). Defects treated with β-TCP/CS showed the greatest bone regeneration and graft resorption, although differences between groups were not statistically significant. At sites that healed spontaneously, the trabecular number was lower (p < 0.05) and trabecular separation was higher (p < 0.05), compared to sites treated with β-TCP/CS or xenograft. Trabecular thickness was higher at sites treated with the bovine xenograft (p < 0.05) compared to sites filled with β-TCP/CS or sites that healed spontaneously. In conclusion, the novel β -TCP/CS grafting material performed well as a bioactive and biomimetic alloplastic bone substitute when used in cranial defects in this animal model.