Alzheimers disease is characterized by cognitive decline associated with neurodegeneration and accumulation of amyloid beta. Here, experimenting with in vitro models of AD using SHSY5Y cell line after exposing them with Aβ1-40. The natural derivatives like neem oil, plays a potential role in neuroprotection and reduction of tau aggregates in animal models, eventually becoming a alternative treatment beneficiaries in neurodegenerative disease. Formulating the neem oil into a nanoemulsion with Tween 80® and Brij 30™ for stabilization, can be a beneficial drug development for AD. The optimization of this neem oil nanoemulsion was performed with Response Surface Methodology - Box Behnken Design, showed the S mix 10, Sonication power 55%, and Sonication time 15%; as the optimized value to synthesize a stable neem oil nanoemulsion. Secondly, they subjected to thermodynamic stability analysis for estimating the shelf life duration with different temperature conditions. Amyloid beta induced SHSY5Y tend to show severe cell oxidative stress, to estimate that, the neem oil and neem oil nanoemulsion were subjected to DPPH assay. Further, the neem oil and neem oil nanoemulsion analyzed with the Aβ1-40 induced SHSY5Y, resulted in good cytotoxicity, where the nanoemulsion exhibited more reduction in cell viability than neem oil. This trend shows that synthesizing the natural oils into nanocarriers can potentially increase the drug release rate, enhanced stability and good biomedical application.