Mechanistic Insights Into ROS‐Mediated Apoptotic Toxicity of Achillea millefolium L. Fabricated Manganese Oxide Nanoparticles in Human Neuroblastoma

ABSTRACT

Green nanotechnology offers a sustainable strategy for synthesising functional nanomaterials with enhanced biomedical potential. In this study, manganese oxide nanoparticles (MnO NPs) were biosynthesised using Achillea millefolium L . leaf methanolic extract and evaluated for their antioxidant and anticancer activities against human neuroblastoma IMR‐32 cells. Phytochemical screening and GC–MS analysis of the extract revealed the presence of diverse bioactive compounds, with major constituents including stigmast‐5‐en‐3‐ol (23.64%), 2‐hexadecen‐1‐ol, 3,7,11,15‐tetramethyl‐ (6.81%), 2,6,10‐trimethyl,14‐ethylene‐14‐pentadecne (5.95%) and 9‐octadecenoic acid (5.70%), and terpenoid derivatives (~6.81%). Structural characterisation confirmed the successful formation of crystalline MnO NPs with an average crystallite size of 49.18 nm and an optical band gap of 2.056 eV, while FTIR and electron microscopy analyses indicated phytochemical capping and predominantly spherical morphology. The biosynthesised MnO NPs exhibited significantly higher antioxidant activity than the crude extract ( p  < 0.05). The nanoparticles showed 60.37 ± 0.57% DPPH radical scavenging activity at 500 μg mL ⁻¹ (IC₅₀ = 480 ± 1.00 μg mL ⁻¹ ) and 56.1 ± 1.00% ABTS scavenging activity (IC₅₀ = 440 ± 1.00 μg mL ⁻¹ ). Enhanced ferric reducing power (45.3 ± 0.54%) and metal‐chelating activity (44.0 ± 0.4%) further confirmed their superior antioxidant potential. Cytotoxicity evaluation using the MTT assay revealed pronounced antiproliferative activity against IMR‐32 neuroblastoma cells with a GI₅₀ value of 54.02 μg mL ⁻¹ . Mechanistic investigations indicated that MnO NP treatment significantly elevated intracellular reactive oxygen species (43.2% increase) and induced apoptosis, with 84% apoptotic cell population detected by Annexin V‐FITC/PI staining. Taken together, these findings suggest that A. millefolium ‐mediated MnO NPs exhibit notable antioxidant properties along with ROS‐mediated anticancer activity. This indicates their potential as a promising and environmentally friendly nanotherapeutic approach for the treatment of neuroblastoma, although further investigation is needed to confirm their efficacy and safety.