In this work, ficin extract (3.4.22.3, >1.0 u/mg) was immobilized for the first time in poly (vinyl alcohol) (PVA) electrospun nanofibers in order to preserve its tridimensional molecular structure. The analysis of the morphology of the electrospun nanofibers was carried out using scanning electron microscopy (SEM) showing a diameter in the range of 124-194 nm. The interaction of the ficin extract with the nanofibers structure was analyzed by infrared spectroscopy (FTIR) analysis. The immobilization step was achieved through crosslinking involving the exposure to glutaraldehyde (GA) vapor. The enzyme catalytic behavior was followed by ultraviolet spectroscopy (UV) using the Earlanger method using N-alpha-benzoyl-L-arginine-4-nitroanilide hydrochloride (BAPA) as substrate. The maximum catalytic activity was reached with 1 h of crosslinking, 20% of enzyme loading and pH 8. The immobilized ficin extract showed 92% of the enzyme activity of the crude ficin extract. The enzymatic activity of the immobilized ficin extract was conserved after a total of nine reuse cycles and maintained after being stored for 25 days. Finally, both the glass transition (T-g) and heat of fusion (H-f) were affected by number of enzyme molecules inside the polymeric nanofibers matrix according to the study of the thermal properties by differential scanning calorimetry (DSC). 2018 Elsevier B.V. All rights reserved.