Purpose: This study investigated the osteoconductivity of natural calcium carbonate-derived bone substitutes, hen eggshell (ES), and compared with those of commercial bone substitutes. Materials and Methods: Osseous defects created in the rat calvaria were filled with particulated ES(ES-1), ES with calcium-deficient hydroxyapatite surface layer (ES-2), Biocoral(Inoteb, France), and Bio-Oss(Geistlich Pharma, Wolhusen, Switzerland). After 4 and 8 weeks of healing, histomorphometic analysis was performed to evaluate the amount of newly formed mineralized bone area (NB%). Results: Histologic and histomorphometric analysis showed new bone formation and direct bony contact with the grafted materials in all groups. At 4 weeks, Biocoral group showed greater NB% compared to Bio-Oss and ES-1 groups (P<0.05). At 8 weeks, Biocoral and ES-2 groups showed significantly greater NB% compared to Bio-Oss group (P<0.05). Conclusion: These results indicate that natural calcium carbonate-derived bone substitutes with microporous calcium-deficient hydroxyapatite surface layer may be an effective materials treating osseous defects.