The rapid growth of industry and economic development for the last few decades has significantly contributed to increasing water pollution, primarily due to factors such as oil spills and leaks, leading to substantial environmental and ecological damage. This study explored the synthesis of ultrasound-assisted preparation of chitosan-based microspheres under varying ultrasonic power (90, 180, 270 W) for marine diesel oil–seawater separation. Scanning Electron Microscope (SEM), Fourier Transform Infrared Spectroscopy (FTIR), X-ray Diffraction (XRD), and X-ray Fluorescence (XRF) were used to characterize the microspheres. Batch experiments were conducted to investigate the effect of oil–seawater concentration and contact time on the adsorption performance of the microspheres. Models were used to describe in detail the adsorption kinetics (Pseudo-first order and Pseudo-second order) and isotherms (Langmuir and Freundlich). Maximum removal efficiencies were observed at 99.49% for lower oil–seawater concentration (10 g/L) and 90.54% at longer contact time (4 h). The adsorption process also followed a pseudo-first order kinetic model (R2 = 0.991) and fitted the Langmuir isotherm (R2 = 0.9957), indicating a monolayer adsorption. The results obtained in this study will offer valuable insight into the adsorption mechanism and support future applications in the design of treatment systems for real-world oily wastewater.