This study investigates the fabrication and optimization of electrospun shellac fibers loaded with Senna alata (SA) leaf extract. Single-fluid electrospinning was used to create nanofibers from a polymer solution containing active ingredients. A combination of fractional factorial design and Box–Behnken design was employed to optimize the electrospinning conditions that affect fiber morphology and the entrapment efficiency of SA leaf extract. The results showed that shellac content had the most significant impact on fiber diameter and bead formation. The optimal conditions identified were a voltage of 24 kV, a solution feed rate of 0.8 mL/h, and a shellac-extract ratio of 38.5:3.8. These conditions produced nanosized fibers with a diameter of 306 nm, a low bead-to-fiber ratio of 0.29, and an entrapment efficiency of 96%. The biphasic release profile of rhein from the optimized nanofibers was confirmed in vitro, showing an initial burst release of 88% within the first hour followed by a sustained release of over 90% for the next 12 hours. The fibers demonstrated antimicrobial efficacy against various pathogens, suggesting potential applications in wound dressings and protective textiles.This study investigates the fabrication and optimization of electrospun shellac fibers loaded with Senna alata (SA) leaf extract. Single-fluid electrospinning was used to create nanofibers from a polymer solution containing active ingredients. A combination of fractional factorial design and Box–Behnken design was employed to optimize the electrospinning conditions that affect fiber morphology and the entrapment efficiency of SA leaf extract. The results showed that shellac content had the most significant impact on fiber diameter and bead formation. The optimal conditions identified were a voltage of 24 kV, a solution feed rate of 0.8 mL/h, and a shellac-extract ratio of 38.5:3.8. These conditions produced nanosized fibers with a diameter of 306 nm, a low bead-to-fiber ratio of 0.29, and an entrapment efficiency of 96%. The biphasic release profile of rhein from the optimized nanofibers was confirmed in vitro, showing an initial burst release of 88% within the first hour followed by a sustained release of over 90% for the next 12 hours. The fibers demonstrated antimicrobial efficacy against various pathogens, suggesting potential applications in wound dressings and protective textiles.