Iron oxides have become increasingly popular for their use as a diagnostic and therapeutic tool in oncology. This study aimed to improve pharmacological valuable of Fe3O4, which may be use to diagnosis colorectal cancers (CRC). Here, we have developed chitosan (CS) coated Fe3O4 through a cost-effective procedure. First, we determined the characterization of OA-C-Fe3O4 by FTIR, UV-Vis spectra, and TEM. Then, we evaluated the photodynamic therapeutic (PDT) activity of OA-C-Fe3O4 in human colorectal carcinoma cell lines (HCT 116). Current results revealed that the light-induced enhanced reactive oxygen species (ROS) activity of the nanoparticles (NPs) and caused cell death via the activity of caspase 9/3. The in vitro magnetic resonance imaging (MRI) experiments in (HCT 116) and human embryonic kidney cells (HEK 293) illustrated that nanohybrid is an effective MRI contrasting agents for the diagnosis of colorectal cancer.