Since the last few decades, the green synthesis of metal nanoparticles was one of the most thrust areas due to its widespread application. The study proposed using wasted and unusable Humulus lupulus (Hops) extract to synthesize silver nanoparticles for biomedical application. The environment around us gives us many scopes to use the waste from environmental sources and turn it into something valuable. The spent Hops extract was used to synthesize silver nanoparticles (AgNP@HOPs), and the synthesized product exhibited an excellent therapeutic effect in terms of anti-bacterial and anti-cancer agents. The synthesis was optimized considering different factors like time and the concentration of AgNO 3. The silver nanoparticles were characterized in detail using different characterization techniques XRD, DLS, TEM, BET, XPS, Raman Spectroscopy, SEM, EDAX, AFM, which revealed the uniqueness of the silver nanoparticles. The average hydrodynamic size was found to be 92.42 ± 2.41 with a low polydispersity index. The presence of Ag–C and Ag–O bonds in the AgNP@HOPs indicated that it is composed of organo-silver and silver oxides. The nanoparticles were found to be spherical with an average size of 17.40 nm. The AgNPs were lethal to both E. coli and S. aureus with a MIC-50 of 201.881 μg/mL and 213.189 μg/mL, respectively. The AgNP@HOPs also exhibited an anti-cancer effect with an IC-50 of 147.175. The AgNP@HOPs exhibited less cytotoxicity and genotoxicity against normal cells and exhibited superior haemocompatibility (major criteria for drug selection). There are indeed various reports on the synthesis of silver nanoparticles, but this study proposes a green method for producing non-genotoxic, non-hemolytic organometallic silver nanoparticles using waste material with considerable therapeutic index from the environmental source with potential application in the medical industry. This work could be taken forward for in-vivo studies and for pre clinical studies. [Display omitted] • Use of wasted and spent Hops for synthesis of organometallic silver nanoparticles. • The nanoparticles obtained are spherical in shape with a size range of 10–50 nm. • AgNP@Hops exhibited superior Anticancer Activity on MCF7 breast cancer cell line. • The biosynthesized silver nanoparticles were haemocompatible and non-genotoxic. [ABSTRACT FROM AUTHOR]