Macrocyclic mononuclear ruthenium (II) complexes have been synthesized by condensation method [Ru (L1, L2, L3) Cl-2] L-1 = (C-36 H-31 N-9), L-2= (C42H36N8), L-3= (C32H32 N-8)]. These ruthenium complexes have been established by elemental analyses and spectroscopic techniques (Fourier transform infrared spectroscopy (FT-IR), H-1-nuclear magnetic resonance (NMR), C-13-NMR and Electrospray ionization mass spectrometry (ESI-MS)). The coordination mode of the ligand has been confirmed and the octahedral geometry around the ruthenium ion has been revealed. Binding affinity and binding mode of ruthenium (II) complexes with Bovine serum Albumin (BSA) have been characterized by Emission spectra analysis. UV-Visible and fluorescence spectroscopic techniques have also been utilized to examine the interaction between ligand and its complexes L1, L2, & L3 with BSA. Chemical parameters and molecular structure of Ru (II) complexes L1H, L2H, & L3H have been determined by DFT coupled with B3LYP/6-311G** functional in both the gaseous and aqueous phases. (C) 2017 Elsevier B.V. All rights reserved.