ZnO and Mo-doped ZnO thin films were grown using the spray pyrolysis process with variations in Mo doping concentrations. The films were characterized with a comprehensive compositional analysis, including electron dispersive X-ray (EDX), X-rays photoelectron spectroscopy (XPS), and time-of-flight secondary ion mass spectrometry (ToF-SIMS), along with grazing incidence X-ray diffraction(GIXRD), atomic force microscopy (AFM), scanning electron microscopy (SEM) and UV–Vis spectroscopy to investigate their structural, morphological, optical and chemical compositions. Micro-hardness analysis was performed on films as a function of the Mo content. GIXRD analysis detected the presence of characteristic peaks of ZnO along the (100), (002) and (101) planes. The presence of Zn, O, and Mo in the synthesized samples was revealed by EDX, XPS, and ToF-SIMS results, which matched the experimental values of a pure and Mo:ZnO-x composition. XPS analysis revealed the Mo6+ oxidation state of molybdenum present in Mo:ZnO films. From UV–Vis absorption spectroscopy, the bang gap was observed to increase with Mo content, up 5 at.%, and located in the 3.18 – 3.24 eV interval. These results will be helpful in the synthesis of Mo:ZnO films for relevant applications, including sensing, optoelectronics and antimicrobial.