The green synthesis of selenium nanoparticles (Se NPs) had been synthesized by pomegranate peel extract (PPE). The antimicrobial, antioxidant, and anticancer activities of the synthesized Se NPs, as well as their hemocompatibility, were investigated. Se NPs were characterized by UV-Vis., SEM, XRD, HR-TEM, DLS, EDX, FTIR, and mapping techniques. HR-TEM image represented the spheroidal forms with moderately monodispersed NPs with a mean diameter 14.5 nm. The SEM image of Se NPs, incorporated with PPE, exhibits uniform NP surfaces, and the appearance was clear. The antimicrobial results confirmed the potential of Se NPs to hinder the growth of some tested pathogenic microbes. Results revealed that Se NPs exhibited promising antibacterial activity against Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis, Staphylococcus aureus, and Streptococcus mutans where inhibition zones were 29, 16, 41, 22, and 54 mm, respectively. Likewise, it exhibited antifungal activity where the values of inhibition zones were 41, 40, 38, and 36 mm against Candida albicans, Cryptococcus neoformans, Aspergillus fumigatus, and A. niger, respectively. The antioxidant activities of Se NPs at concentrations 250-4000 µg/mL were greater than 90% in all cases. Se NP concentrations of 500 µg/mL or less are safe in usage according to hemocompatibility study. Se NPs had an IC of 113.73 µg/mL in a cytotoxicity experiment. Results revealed that Se NPs have promising anticancer activities against MCF7 and Mg63 cancerous cell line, where IC was 69.8 and 47.9 μg/mL, respectively. In conclusion, Se NPs were successfully biosynthesized using PPE for the first time; these Se NPs had promising antimicrobial, antioxidant, and anticancer activities.