The remarkable physicochemical properties of nanomaterials have attracted researchers due to the numerous applications in the field of chemistry, biology, and physics. Despite the various applications, superparamagnetic iron oxide nanoparticles (SPIONs) are harmful to living organisms and to the environment as they are released without any safety testing. In this study, SPIONs were synthesized and further characterized. The aim of the study was to examine the toxicity of synthesized SPIONs against animal models: Zebrafish—Danio rerio, Earthworm—Eudrilus eugeniae, and Drosophila—Drosophila melanogaster through histology using Hematoxylin–Eosin and Prussian Blue staining. The accumulation of SPIONs was further quantified by using Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Through histology images, it was observed that the SPIONs had caused damages and a lower concentration of 0.001–0.002 μg/ml of metal accumulation was detected in the ICP-MS analysis.