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A green synthetic approach to synthesize silver nanoparticles (AgNPs) using the stem extract of Piper chaudocanum for highly sensitive colorimetric detection of Hg 2+ with a low limit of detection of 23 nM and easy colorimetric read-out has been reported. In addition, the biosynthesized AgNPs demonstrated efficient antibacterial activity against Escherichia coli , Pseudomonas aeruginosa and Staphylococcus aureus . The morphology and structure of the as-synthesized AgNPs were examined using SEM, TEM, EDX, XRD and FT-IR analyses. The XRD and TEM results confirm the formation of AgNPs with an average particle size of 8–12 nm. The TLC, CC and HPLC revealed that four main compounds, pentacosanoic acid ( 1 ), piperine ( 2 ), β -sitosterol ( 3 ), and campesterol glucoside ( 4 ), isolated from P. chaudocanum extract act as reducing and stabilizing agents for AgNP formation, and piperine plays a vital role in green synthesis. The chemical structures of these compounds were determined by ESI MS, FTIR, and one- and two-dimensional NMR spectroscopic data analysis. This approach is an efficient, green, cost-effective, eco-friendly and promising technique for synthesizing AgNPs with applications in the colorimetric detection of Hg 2+ and antibacterial activity.