BackgroundSilver nanoparticles (AgNPs) are renowned for their broad-spectrum antibacterial properties. Various synthesis methods, particularly green synthesis using biogenic agents, have garnered significant attention. However, the detailed impact of green-synthesized AgNPs on the antibacterial mechanism against Staphylococcus aureus remain unclear, limiting the full potential of green synthesis compared to chemical methods. MethodsAgNPs were synthesized via chemical (sodium citrate, NaBH4) and green synthesis (green tea leaves, cassia seed extract) methods. The synthesized AgNPs were evaluated for toxicity and antibacterial activity against Staphylococcus aureus. Significant findingsThis study revealed a strong correlation (R2>0.9) between minimal inhibitory concentration (MIC) and AgNP size for both synthesis methods, with a similar exponential trend. MIC values were 45, 40, 25, and 5 µg mL-1 for citrate-, NaBH4-, green tea extract-, and cassia seed extract-assisted synthesis, respectively. Green-synthesized AgNPs showed higher antibacterial activity than chemical AgNPs at comparable sizes. Chemical AgNPs exhibited low and fluctuating scavenging activity, while green methods were more consistent. Cytotoxicity was noted in chemical AgNPs and at high concentrations of green tea extract-assisted AgNPs. Bacterial membrane disruption and ROS accumulation were also observed, contributing to the enhanced antibacterial activity of green-synthesized AgNPs.