© 2022 Elsevier B.V.Vitamin C&E (VtC&VtE)-loaded bilayer wound dressings were prepared using bacterial cellulose (BC) synthesized by Acetobacter species and pullulan (PUL). VtC-containing PUL hydrogels (100 μg/mL) were immobilized onto BC by crosslinking. BC/PUL-VtC was loaded with VtE (100 μM in ethanol) by immersion for 2 h. No delamination between the layers was observed via SEM. Despite the porous inner PUL side, the outer BC side exhibited nanofibrous morphology serving as barriers to prevent microorganism invasion. Equilibrium water content of BC/PUL was above 85 % due to the hydrogel characteristics of PUL side, suitable to absorb exudate in wound bed. PUL layer lost >90 % of its weight in simulated wound fluid and > 99 % in lysozyme solution within 14 days, mediating co-release of VtC&VtE. Thin BC side possessed adequate strength (⁓22 MPa) and strain (>30 %) to endure against tensile stress generated by bending on wound surface without rupture, whereas thick PUL side was flexible (>70 % strain) to fit into wound bed under compressive stress without causing harm. In vitro studies using L929 fibroblasts elucidated PUL side was anti-adhesive and removable. Synergistic effect of VtC&VtE on antioxidant activity, wound closure, and collagen synthesis was observed. Thus, BC/PUL-VtC/VtE hold promise as cheap and eco-friendly temporary wound dressing.