Analytical Determination of the Lipid Fraction of Nigella sativa Fatty Oil by GC and NMR Analysis and Evaluation of Its Cytotoxic and Antioxidant Activity

<i>Nigella sativa</i>, or black cumin, is used as a spice in cooking and as a food supplement like seeds or oil for its biological properties, including antioxidant capacity, anti-inflammatory action, and support for the immune system. In the present study, the chemical composition and biological activities of the <i>Nigella sativa</i> seeds' fatty oil (<i>NS</i>) were investigated. The analytical composition was carried out by several techniques, such as GC-MS spectrometry and ¹H- and ¹³C-NMR spectroscopies using appropriate internal standards. The GC-MS analysis highlighted the presence of palmitic and linoleic acid as major compounds. The antioxidant potential was evaluated through the DPPH radical-scavenging assay, and, furthermore, the <i>NS</i> effect on intracellular reactive oxygen species (ROS) levels was assessed in HaCaT cells (non-tumorigenic human keratinocytes) under oxidative stress induced by hydrogen peroxide. The cytotoxic and genotoxic profiles were evaluated on Caco-2 cells (human colorectal adenocarcinoma cells) using the CCK-8 viability assay and the Comet assay, respectively. Overall, the results demonstrated that <i>NS</i> possessed antioxidant activity, as evidenced by concentration-dependent DPPH radical scavenging and reduced intracellular ROS levels in HaCaT cells under oxidative stress. In Caco-2 colorectal cancer cells, <i>NS</i> induced significant cytotoxicity and DNA damage at higher concentrations, suggesting potential genotoxic effects. These findings support the dual role of <i>NS</i> as a natural antioxidant and a promising candidate for nutraceutical and dermatological applications, including those targeting oxidative stress-related conditions and cancer.