Green synthesis of copper oxide nanoparticles from Calotropis gigantea of antimicrobial, antidiabetic, and toxicity evaluation in zebrafish.

Calotropis gigantea (giant milkweed or crown flower), a traditional medicinal plant used to treat various ailments, contains toxic compounds that require careful handling. In this study, copper oxide nanoparticles (CuO NPs) were synthesized using an eco-friendly green method with C. gigantea leaf ethanol extract ofsynthesized and characterized copper oxide nanoparticles (CG-CuO NPs) using UV-Vis spectrophotometry, FT-IR, XRD, SEM, and EDX. The study evaluated the in vitro bioactivity of the CG-CuO NPs leaf extract focusing on their antidiabetic, anti-inflammatory, cytotoxic, and antimicrobial properties. The Phytochemical screening of secondary metabolites such as flavonoids, tannins, steroids, saponins, polyphenols which act as efficient bio-reductants and capping agents, leading to smaller, more stable CuO nanoparticles with superior bioactivity. Characterization and confirmation of CG-CuO NPs synthesis by UV-Visible spectroscopy confirmed nanoparticle formation, showing a characteristic absorbance peak at 437 nm. Stabilization analysis of FTIR analysis identified hydroxyl, amine, carboxyl, and aromatic functional groups involved in nanoparticle stabilization, with prominent absorption bands at 1072 cm-1 and 530 cm-1. Morphology and size of SEM revealed predominantly spherical nanoparticles with smooth surfaces and uniform dispersion. The TEM images showed that the particle sizes ranged from 35 to 85 nm, following a narrow quasi-Gaussian size distribution. The crystallinity and Composition of XRD analysis confirmed the crystalline monoclinic phase of the CuO NPs (matching JCPDS No. 48-1548). Elemental analysis indicated that copper oxide was the major component (75.37 wt%), with trace amounts of carbon, sodium, and oxygen. Toxicity analysis of zebrafish embryotoxicity assays showed concentration- and time-dependent developmental toxicity, including deformities and mortality. Decreasing LC₅₀ and EC₅₀ values between 24- and 120-h post-fertilization indicated increasing toxicity over time. Biological Activities of the biogenic CuO NPs exhibited strong biological effects, including potent α-glucosidase inhibition (IC₅₀ = 22 μg/mL). The significant anti-inflammatory activity (IC₅₀ = 109 μg/mL) exceeded that of the chemically synthesized counterparts. The CG-CuO NPs exhibited selective anticancer potential, with an IC₅₀ of 31.5 μg/mL against MCF-7 breast cancer cells compared to 124.8 μg/mL in non-cancerous HaCaT keratinocytes, demonstrating preferential cytotoxicity toward cancer cells. C. gigantea-synthesized CuO nanoparticles demonstrate promising antidiabetic, anti-inflammatory, and selective anticancer properties of green synthesis and biocompatibility supporting potential biomedical applications.