Herein, we have reported the binding interaction between Human Serum Albumin and the titanium dioxide nanoparticles in vitro. Characterization of the nanoparticles physicochemical and morphological properties was done with X-ray Diffraction, Fourier Transform Infra Red, Raman spectroscopic and microscopic methods. XRD and FT-Raman studies have confirmed the anatase phase of the nanoparticles. Surface active functional groups have been identified from FTIR studies. SEMEDAX has figured out the spherical shape of the particles with their metallic constituents. Hyperchromic effect observed against the absorbance and the quenching behavior of the nanoparticles on the intrinsic fluorescence of the HSA has confirmed the complex formation to exist via static mechanism. Circular Dichroism studies have dictated the loss in the secondary structures. Docking studies showed the binding mode of TNPs-HSA complex system. Copyright © 2017 American Scientific Publishers All rights reserved.