Abstract
A combinational approach to antimicrobial design emerges from a nanoemulsified nanoplatform from the union of two Egyptian essential oils-orange peel and clove-whose distinct chemistries were revealed by GC-MS. Orange EO is dominated by D-limonene (63.25%), with supporting alkyl-benzenes (22.92%) and β-myrcene (1.41%), while clove EO centers on Eugenol (36.20%), Caryophyllene (19.30%), Eugenol acetate (18.71%), along with alkyl-benzenes (11.86%) and humulene (2.41%). The chemical consonance between Eugenol and Limonene hints at a cooperative assembly: their lipophilic profiles enable the formation of nanoemulsified nanoparticles from their emulsion blend, potentially amplifying bioavailability and multi-target action. Biologically, Eugenol/Limonene demonstrates tangible antimicrobial activity. In a direct DHFR inhibition assay, the nanoformulation inhibits DHFR with an IC(50) of 8075 ± 0.96 μg/mL, while the benchmark drug methotrexate (MTX) shows far stronger inhibition ( IC(50) = 0.81 ± 0.07 μg/mL). Across pathogenic strains, the nanoemulsion exhibits broad antibacterial reach, producing inhibition zones of 24.0-29.0 mm and delivering bactericidal effects with MICs of 8.0-32.0 μg/mL and MBCs of 64.0-512.0 μg/mL. Explorations into in vivo-like tissue responses reveal nuanced outcomes. The nano-treated cohort shows dramatic reductions in bacterial load, yet also manifests airway changes consistent with bronchiolar irritation and alveolar remodelling, signalling both therapeutic potential and safety considerations. Histological comparisons indicate that nanoparticle-treated groups preserve some alveolar integrity, with Type II pneumocytes and lamellar bodies present. Overall, the study underscores the promise of combining EO components to enhance antimicrobial performance via nanostructured carriers, while highlighting the delicate balance between efficacy and pulmonary safety.