The present study focused on the phytochemical chromatographic isolation of two chemical constituents, sterol and flavonoid (stigmasterol glucoside and luteolin) from Plectranthus asirensis. A novel, eco-friendly, and cost benefit, ultrasensitive sequential injection analysis chemiluminescence (SIA-CL) approach based on the enhanced catalytic activity of silver nanoparticles (AgNPs) was suggested to evaluate the isolated compounds. Silver nanoparticles were synthesized using the biomass of an ethanolic extract of aerial parts of P. asirensis as a reducing agent. Spectroscopic and microscopic characterization of the prepared AgNPs were performed, including UV-Vis spectrometry, XRD and FT-IR as well as TEM and SEM. UVspectroscopic method detected the formed AgNPs at an absorbance wavelength of 420 nm. Furthermore, antioxidant activity of the isolated compounds was also determined using 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity protocol.