We have investigated the effect of sulfur concentration ([S]/[Mo] = 0–10 at.%) on nanostructural, optical and electrical properties of MoO3 thin films grown on glass substrate by spray pyrolysis technique. X-ray diffraction analysis showed that the films were crystallized with mixed structures of MoO3 orthorhombic and MoS2 hexagonal structure. According to FESEM images, the studied films have a sponge-type structure on the order of nanometers. Optical measurements revealed that two distinct inflexions indicative two transitions correspond to MoO3/MoS2 phases in agreement with the XRD analysis. The Hall effect and thermoelectric measurements have shown p-type conductivity, and the free hole density increases with increasing sulfur to molybdenum ratio, in agreement with reflectance spectra of the layers. The obtained value of Seebeck coefficient for MoO3 film with [S]/[Mo] ratio of 8 at.% was as high as 169 μV/K.