Selenium nanoparticles (SeNPs) attract more and more interest due to good bioavailability and low toxicity, accompanied by various bioactivities consisting of antimicrobial, antioxidant, and anticancer activity. SeNPs could be generated by physical, chemical, or biological methods and their potential depends on the particle diameter, homogeneity, coating agents, etc Up to now, there has been no previous work reporting on the activities of SeNPs produced by electron beam yet. In our work, SeNPs created by electron beam (SeNP/EB) or gamma irradiation (SeNP/G), stabilized by gum arabic were evaluated for the anticancer capacity by MTT assay, the antioxidant activity by DPPH radical scavenging assay, and the antibacterial ability by agar well diffusion assay. The results showed that SeNP/EB and SeNP/G displayed growth inhibition on HeLa cervical cancer cells with IC50 values of 2.83 and 1.54 μg ml−1, while the values on MCF7 breast cancer cells were 27.70 and 38.80 μg ml−1 respectively. The SeNPs affected HeLa cancer cells more selectively than normal fibroblasts as evidenced by the high selectivity index of 7.98 and 26.25. Notably, the results demonstrated that SeNP/G is much safer than SeNP/EB when applying for cancer treatment in the future. Regarding the DPPH assay, SeNPs of both synthetic methods exhibited potential IC50 values (13.5 and 12 μg ml−1) compared with that of ascorbic acid (8.4 μg ml−1). In comparison to previous studies, our results sugessted that gamma and electron beam irradiation methods, accompanied by coating with gum arabic could be novel approaches in SeNP synthesis to enhance the antioxidant activity of the SeNPs. Besides, SeNPs also caused an inhibition towards Listeria monocytogenes and Escherichia coli, which was verified by the inhibition-zone diameter of approximately 8–12 mm, through inducing oxidative stress in bacterial cells. In conclusion, along with the advantages of physical methods such as time-saving, eco-friendly processes, SeNPs in our work could be a promising candidate for the research and development of healthcare products.