The incorporation of additional functional groups such as bismuth nanoparticles (Bi NPs) into magnetite nanoparticles (FeO NPs) is critical for their properties modification, stabilization, and multi-functionalization in biomedical applications. In this work, ultrasound has rapidly modified iron oxide (FeO) NPs via incorporating their surface through coating with Bi NPs, creating unique FeO@Bi composite NPs. The FeO@Bi nanocomposites were synthesized and statistically optimized using an ultrasonic probe and response surface methodology (RSM). A face-centered central composite design (FCCD) investigated the effect of preparation settings on the stability, size, and size distribution of the nanocomposite. Based on the numerical desirability function, the optimized preparation parameters that influenced the responses were determined to be 40 ml, 5 ml, and 12 min for Bi concentration, sodium borohydride (SBH) concentration, and sonication time, respectively. It was found that the sonication time was the most influential factor in determining the responses. The predicted values for the zeta potential, hydrodynamic size, and polydispersity index (PDI) at the highest desirability solution (100%) were -45 mV, 122 nm, and 0.257, while their experimental values at the optimal preparation conditions were -47.1 mV, 125 nm, and 0.281, respectively. Dynamic light scattering (DLS) result shows that the ultrasound efficiently stabilized and functionalized FeONPs following modification to FeO@Bi NPs, improved the zeta potential value from -33.5 to -47.1 mV, but increased the hydrodynamic size from 98 to 125 nm. Energy dispersive spectroscopy (EDX) validated the elemental compositions and Fourier transform infrared spectroscopy (FTIR) confirmed the presence of Sumac (Rhus coriaria) compounds in the composition of the nanocomposites. The stability and biocompatibility of FeO@Bi NPs were improved by using the extract solution of the Sumacedible plant. Other physicochemical results revealed that FeONPs and FeO@Bi NPs were crystalline, semi-spherical, and monodisperse with average particle sizes of 11.7 nm and 19.5 nm, while their saturation magnetization (Ms) values were found to be 132.33 emu/g and 92.192 emu/g, respectively. In vitro cytotoxicity of FeO@Bi NPs on the HEK-293 cells was dose- and time-dependent. Based on our findings, the sonochemical approach efficiently produced (and RSM accurately optimized) an extremely stable, homogeneous, and biocompatible FeO@Bi NPs with multifunctional potential for various biomedical applications.