Multidrug-resistant strains of Acinetobacter baumannii are pathogenic agents of hospital infections, especially in the intensive care unit (ICU). The purpose of this study is to investigate the antibacterial effects and biosynthesis of arsenic nanoparticles (ARn) using pink rose petals on multi-drug resistant strains of A. baumannii. In this study, plant bioactive molecules present in rose petals are responsible for the synthesis of ARn. The characteristics of ARn were determined through FTIR, EDX, DLS, SEM, UV-Vis, and XRD techniques. In addition, the antibacterial activity of biosynthesized ARn was investigated against multi-drug resistant A. baumannii. The results of SEM revealed that the ARn were spherical with an average diameter of 75.71 nm. The results of XRD confirmed the crystal structure of arsenic oxide phase. The results of antibacterial effects exhibited that the lowest MIC of ARn was 800 μg/ml on the standard strain of A. baumannii and 800 μg/ml against clinical strains of multi-drug-resistant A. baumannii. The results of this study provided an new green synthesis method for ARn, which have a high potential on multi-drug-resistant A. baumannii. This study has the potential to pave the way for further research and developments in this field.