Green-Synthesized Copper Oxide Nanoparticles as an Adjuvant to Quinine in Experimental Cerebral Malaria: Mechanistic and Safety Evaluation in Mice

Background: Cerebral malaria remains one of the most severe complications of Plasmodium infection and is associated with excessive oxidative stress, neuroinflammation, and high mortality. This study investigated the therapeutic and mechanistic effects of green-synthesized copper oxide nanoparticles (CuO-NPs), administered alone or in combination with quinine (QN), in mice infected with Plasmodium berghei ANKA strain. Methods: Eighty-six BALB/c mice were randomly assigned to infected and noninfected groups. Treatments were administered orally once daily beginning 3 h postinfection for 7 consecutive days. Malaria-infected mice received QN (5 or 10 mg/kg), CuO-NPs (5 or 10 mg/kg), or combinations of CuO-NPs with low-dose QN (5 mg/kg). Survival was monitored for 28 days. Parasitemia suppression, oxidative stress biomarkers, antioxidant gene expression, and inflammatory mediators were evaluated in brain tissue homogenates. For toxicity evaluation, healthy mice received treatments for 14 days. Statistical analyses were performed using one-way analysis of variance followed by Tukey’s post hoc test. Results: Green-synthesized CuO-NPs exhibited spherical morphology with an average diameter of approximately 40 nm. Combined administration of CuO-NPs with QN significantly improved survival and reduced parasitemia suppression, ranging from 93.8% to the absence of microscopically detectable parasitemia relative to infected untreated controls ( p < 0.001). The combinations also significantly reduced malondialdehyde and nitric oxide levels while upregulating Glutathione peroxidase (GPx) and superoxide dismutase expression. In addition, combination therapy modestly downregulated nuclear factor-kappa B p65, toll-like receptor 4, and tumor necrosis factor-alpha Messenger ribonucleic acid (mRNA) expression. Biochemical toxicity analyses revealed no significant alterations in hepatic or renal function markers following 14 days of administration. Conclusion: The combination of green-synthesized CuO-NPs with low-dose QN may improve antimalarial efficacy through modulation of oxidative stress and inflammatory responses in an early-intervention murine model of cerebral malaria without inducing significant short-term biochemical toxicity. However, additional mechanistic, pharmacokinetic, histopathological, and long-term safety studies are required before clinical translation can be considered.