Bee venom (BV) is a substantial ingredient in secondary metabolic products from honeybees (Apis mellifera L.). Polymers and their varieties are emphasized in modern research for their obvious biological functions in medication. Chitosan nanoparticles (Chs-NPs) are among the anticipations for this approach, which are contingent upon the type of nanoparticles employed for therapeutic purposes. The main objective of this research is to substitute chemicals with environmentally friendly manufacturing and extraction of chitosan from exuviae of black soldier fly, Hermetia illucens L., prepare exuvial chitosan nanoparticles (Chs-NPs), testing the placing of nanoparticles with the honeybee, Apis mellifera venom (BV-placed NPs), and evaluate the antibacterial and anticancer activities of both BV, Chs-NPs, and BV-placed NPs. FTIR spectrum of BV, Chs-NPs and BV-placed NPs were illustrated showed the different functional groups of the tested compounds. Hydrodynamic size, Zeta potential and PDI of synthesized nanoparticles of Chs-NPs 270.6±5.0, +30.1± 3.3 and 0.199±0.02, while for BV-Chs NPs were 380.4±15.0, +25.2±2.5 and 0.514±0.01. BV loaded Chs- NPs has the highest antibacterial impact versus M. smegmatis with inhibition zone of 28.0 ± 0.4 mm. Additionally, BV loaded Chs-NPs has the most promising antitumor impact versus IC50 =274.37±0.1 µg/ml with minimal cytotoxicity versus Vero cells CC50=62.24± 0.2 µg/ml with notable microscopic alterations. Bee venom (BV) when enclosed in the Chs-NPs showed a promising anti- bacterial impact versus Mycobacterium smegmatis as well as antitumor action versus A549 with an acceptable harmful level to be implemented for forthcoming uses.