Direct loading versus chitosan nanoparticles encapsulation of 1,8-cineole and eucalyptus oil in PVA-based nanofibers: Box-Behnken optimization, sustained delivery, antibacterial, and wound healing activities.

Chronic wound healing remains challenging because many dressings fail to maintain long-lasting antimicrobial activity while promoting tissue regeneration. To address this limitation, two natural agents with reported antibacterial and pro-healing potential; 1,8-cineole (CIN) and eucalyptus oil (EU), were incorporated into electrospun poly(vinyl alcohol) (PVA) nanofibers (NFs) using two encapsulation levels: direct loading into PVA fibers or dual encapsulation via chitosan nanoparticles (CS NPs) embedded within the PVA matrix. A Box-Behnken design optimized the representative dual system, identifying electrospinning conditions of 0.5 mL·h-1, 22 kV, and 5 mg·mL-1 NP loading; these settings were then applied to all formulations. FTIR spectroscopy, SEM/TEM, contact angle, TGA, and tensile testing confirmed incorporation and strengthened polymer-NP interactions, supporting improved stability and controlled delivery. At pH 7.4, release was sustained up to 72 h and followed Weibull-controlled multi-diffusion kinetics, with improved retention in dual systems. Antibacterial activity was demonstrated against E. coli, P. aeruginosa, S. aureus, and MRSA, with strain-selective responses. High cytocompatibility was maintained about 97 % of L929 viability, and scratch assays showed accelerated fibroblast migration at 48 h, with dual systems achieving near-complete closure. In a rat wound model, wound contraction approached near-complete closure by day 10 and reached ∼100 % by day 15, accompanied by improved epithelialization and collagen deposition. Moreover, Molecular docking supported the antibacterial and pro-regenerative outcomes. Among formulations, dual-encapsulated CIN NFs (PVA-CS-CIN NFs) showed the most promising performance. Overall, dual encapsulation in PVA NFs provides a proof-of-concept strategy to stabilize volatile natural agents and improve wound healing.