Hybrid films of gold nanoparticles and graphene oxides (GOs) were prepared by directly growing gold nanoparticles on supported thin layers of GO films on a glass slide. The gold/GO nanohybrid films were thoroughly investigated using various analytical methods, including Raman spectroscopy and atomic force microscopy. The hybrid film was then applied to laser desorption/ionization (LDI) of small molecules, which enabled mass spectrometric analysis of analytes. After a series of detailed mechanistic studies and systematic investigations, we found that the gold/GO hybrid films serve as a successful LDI platform for small-molecule analysis because of the high desorption efficiency of analytes from the hybrid films without inducing significant fragmentation of analytes. We suggest that the underlying GO films may effectively dissipate excess thermal energy generated by laser irradiation of Au to prevent undesirable analyte fragmentation.