Dark chocolate provides enormous health benefits owing to its enriched antioxidants, such as polyphenols and flavonoids, which can encounter free radicals in the body. These antioxidants can lower blood cholesterol, lowering your risk of heart attacks, cancer, high blood pressure, and stroke. This study was aimed to investigate both the free radical scavenging property and inhibitory property of dark chocolate mediated zinc oxide nanoparticles on activity of cytochrome P450 (CYP) isoform. The DC-ZnO was evaluated against free radicals formation using DPPH and ABTS free radical assays. Besides, the ZnO nanoparticles were also evaluated for CYP34A inhibitory activity. From the results it was demonstrated that there was a dose dependent increase in the inhibition of DPPH and ABTS free radicals and CYP3A4 inhibitory activity by the dark chocolate formulation mediated zinc oxide nanoparticles. The maximum inhibitory activity was observed at maximum concentration of 160µL. It may be thus concluded that the synthesized dark chocolate mediated zinc oxide nanoparticles possess potent antioxidant activity with significant inhibitory effect on Cytochrome P3A4 enzyme.