In this study, lactobionic acid (LBA) was incorporated into poly(vinyl alcohol) (PVA) and poly(ε-caprolactone) (PCL) by electrospinning. The antimicrobial effects of the nanofibers were tested using the agar diffusion method. Only the PVA formulations showed antimicrobial activity against Staphylococcus aureus. The PVA and PCL nanofibers containing LBA showed antioxidant activity ranging from 690.33 to 798.67µM TEAC when tested by the ABTS method. The characterization of nanofibers was performed by scanning electron microscopy, Fourier transform infrared spectroscopy, thermogravimetric analysis, differential scanning calorimetry, and mechanical analysis. The nanofibers showed a uniform morphology and their average diameters ranged from 295.5 to 2778.2nm. The LBA addition induced a decrease in the enthalpy of fusion (ΔHm) of PVA and PCL nanofibers, while the Young's modulus was reduced from 20 to 10MPa in PCL and PCL-LBA nanofibers, respectively. No relevant differences were observed between the FTIR spectra of the control nanofibers and the nanofibers containing LBA. All nanofibers presented hemolysis rate below 2%, thus can be considered as non-hemolytic materials. Further toxicological assessment was performed with the selected formulation PVA10 + LBA. The evaluations by mutagenicity assay, cell survival measurement, cell viability analysis and agar diffusion cytotoxicity test indicated that there are no significant toxic effects. Electrospun nanofibers PVA-LBA and PCL-LBA were successfully produced, showing good thermal and mechanical properties and non-toxic effects. Furthermore, the nanofibers showed antimicrobial activity and antioxidant activity. The findings of this study indicate that PVA and PCL electrospun nanofibers incorporating LBA are promising for use in packaging applications.