This paper reports the effects of hydrogen bonding on the starch granule dissolution, spinnability of the starch solution, and the properties of the electrospun starch fibers. Dissolution behaviors of the starch granules in dimethyl sulfoxide (DMSO) and water mixed solvents are characterized by confocal laser microscope, differential scanning calorimetry (DSC), and rheologic measurement. The starch granules are dissolved in the 100% DMSO or 6% H2O/94%DMSO solvent, resulting in good electrospinnability. On the other hand, the gelatinization behavior is observed for the starch granules in the 22%H2O/78%DMSO solvent, resulting in poor fiber formation during electrospinning. Infrared spectroscopy, DSC, x-ray diffraction, and mechanical property characterizations of the electrospun starch nanofibers suggest that the water content in the solvent plays an important role in the properties of the nanofibers. The dissolution behavior, electrospinnability, and the properties of the electrospun nanofibers are explained by the effects of the DMSO and H2O molecules on the ubiquitous hydrogen bonds in starch.