Pulsed laser deposition technique has been employed to prepare thin films of WO3 nanoparticles on a glass substrate at 400K under vacuum (1 mbar) by the increasing of the substrate-target distance in the range of about (0.5, 1.5, 2.5 and 3.5 cm). X-Ray diffraction (XRD), Scanning electron microscopy (SEM), Atomic force microscopy (AFM) and UV-Visible investigations have been observed the effect of substrate-target distance on the structural and optical properties of WO3 nanoparticles thin films. Calculations of the transmittance data for all films were given the evidence to increase the optical bandgap values in the range (3.082, 3.046, 3.062and 3.105eV) when increasing the distance between the target and substrate in the range (0.5, 1.5, 2.5, and 3.5 cm), respectively.