The nanostructure of biomaterial samples of Hydroxyapatite (HAp) was treated by microwave radiation treatment with different additives (1, 10, 20, 30 and 40 wt. in percent) of Lithium Iodide (LiI) using the known sol–gel process. The sample crystal structure was achieved through the analysis of X-ray diffraction and some associated parameters like that strain, grain size, and dislocation. The FTIR and Raman spectroscopy investigations identified effective functional groups and their modes. SEM conforms to the sample morphology's fine low-dimensional structure (10–28 nm). The various preparations were studied for their anti-proliferative/anticytotoxicity/stimulatory ability against normal and activated splenic cells of the mouse in vitro and for the lytic effects on red blood cells (RBCs). Results showed that various LiI additives have inhibitory effects on normal splenic cells of the mouse. The pure LiI and Li 10 had relaxing effects on splenic cells activated by the mouse while the other formulations (LiI 20, LiI 30 and LiI 40) displayed inhibitory/cytotoxic results besides that all preparation samples were safe effects to RBCs.