Green synthesized nanoparticles (GSNPs) display fascinating properties compared to physical and chemical synthesized ones. GSNPs are currently used in numerous applications such as food packaging, surface coating agents, environmental remediation, antimicrobial, and medicine. In the present study, the aqueous leaf extract of Perilla frutescens L. having suitable capping, reducing, and stabilizing compounds was used for green synthesis of silver nanoparticles (Pf-AgNPs). The bioreductant capacity of aqueous leaf extract of P. frutescens for Pf-AgNPs was determined by different confirmatory techniques including UV-Visible spectroscopy, XRD, FESEM, EDX, zeta potential, DLS, SERS, and FTIR analysis. The results exhibited that Pf-AgNPs had optimal size (< 61nm), shape (spherical), and stability (-18.1 mV). The antioxidant activity of Pf-AgNPs with both DPPH and FRAP assays was significantly higher compared to P. frutescens extract. Furthermore, Pf-AgNPs had high antimicrobial activity against Escherichia coli and Staphylococcus aureus (MIC= 0.78 mg/mL), and Candida albicans (MIC= 8 mg/mL) while the plant extract showed low antimicrobial activity against both bacterial strains and the fungus tested. Pf-AgNPs and P. frutescens extract also exhibited moderate toxicity on MCF-7 cancer cells with IC50 values of 346.2 and 467.4 µg/mL, respectively. The results provide insights into using the biosynthesized Pf-AgNPs as an eco-friendly material for a wide range of biomedical applications.