An electrospinning process was used to fabricate silk fibroin (SF) nanofiber nonwovens for wound dressing applications. The electrospinning of regenerated SF was performed with formic acid as a spinning solvent. For crystallization, as-spun SF nanofiber nonwovens were chemically treated with an aqueous methanol solution of 50%. The morphology, porosity and conformational structures of as-spun and chemically treated SF nanofibers were investigated by scanning electron microscopy (SEM), mercury porosimetry, wide angle X-Ray diffraction (WAXD), attenuated total reflectance infrared spectroscopy (ATR-IR), solid state C-13 CP/MAS nuclear magnetic resonance (NMR) spectroscopy. SEM micrograph showed that the electrospun SF nanofibers had an average diameter of 80 nm and a distribution in diameter ranging from 30 to 120 nm. The porosity of as-spun SF nanofiber nonwovens was 76.1%, indicating it was highly porous. Conformational transitions of the as-spun SF nanofibers from random coil to beta-sheet by aqueous methanol treatment occurred rapidly within 10 min, confirmed by solid-state C-13 NMR, ATR-IR, and X-Ray diffraction.