The eco-friendly bio-mediated synthesis of metal oxide nanostructures is found to be gaining research attention towards various applications. In this manuscript, Aerva lanata (A. lanata)-mediated bio-treated production of copper oxide NPs (CuO NPs) were achieved. Also, BBD–RSM method is being used for the optimization of synthesized CuO NPs. Further, the bio-treated CuO NPs were well studied by various characterizations, such as UV–Vis, FTIR, SEM with EDAX, BET and TEM analysis. The answers have been found to infer that the synthesized bio-mediated CuO NPs with high crystallinity were in the spherical form ranging from 10 to 50 nm. Further, the synthesized bio-mediated CuO NPs is found to act as an excellent larvicidal agent against A. ageypti which results in significant activity against dengue vector at higher concentration of 200 ppm of 95{\%} of mortality rate. Overall, the proposed synthesis route of CuO NPs can be utilized by pharmaceutical industry to develop effective drugs against microbial infections.