Biomimetic scaffold is promising for repairing bone defects that are resulted from diseases or injuries. An ideal scaffold should be similar to native bones. Thus, hydroxyapatite (HAp) composite microparticles in which HAp closely resembles the mineral component in native bones were fabricated. Casein/CaCO3 microspheres, precipitated in the presence of casein, were used as starting material and an in situ ion-exchange method was applied under mild conditions. In order to obtain hierarchical structure in which casein/CaCO3 core was wrapped with HAp layer, various conditions, including starting material (resuspended powder or mother solution), reaction temperature, concentration, and addition rate of Na2HPO4 solution, were intensively studied and optimized. The products were characterized using X-ray diffractometer (XRD), Fourier transform infrared (FTIR) spectroscopy, and scanning electron microscope (SEM). After the in situ transformation, the presence of casein was confirmed and its amount was not significantly changed. The formed microparticles could be in only HAp phase of irregular shape or in HAp/CaCO3 composite phase of well-dispersed microspheres. The cytotoxicity of samples was investigated using direct culturing method and its bioactivity was also evaluated by seeding cells on poly(lactic-co-glycolic acid) (PLGA)/HAp films. Here, adipose-derived stem cells (ADSCs) were used as model cells. The results demonstrated that the hierarchical HAp/CaCO3 composite microparticles were cytocompatible and could be potentially used for bone tissue engineering.