The potential impact of TiO₂ and Fe incorporated TiO₂ nanoparticles at the organelle level has been reported. The toxicity of the samples on mitochondria isolated from chicken liver tissue has been examined through mitochondrial swelling, membrane fluidity, ROS generation capacity, and activity of complex II. The toxic effect of TiO₂ was prevented by incorporating Fe into the TiO₂ matrix at different concentrations. The activity of the succinate dehydrogenase enzyme complex was affected and permeabilization of the mitochondrial inner membrane to H⁺ and K⁺ and its alteration in membrane fluidity at 100 μg mL⁻¹ of nano-TiO₂ dosage were investigated, which showed significant changes in the anisotropy of DPH-labeled mitochondria. Fe incorporation into the TiO₂ matrix makes it more biocompatible by changing its structure and morphology.