Bio-ethanol is used worldwide in fuel mixtures such as E10 gasoline. In this study, an onboard fuel system employing a pervaporation membrane was investigated to separate E10 into high-octane-number fuel (high-concentration ethanol fuel) and low-octane-number fuel (low-concentration ethanol fuel). The optimal operation conditions and size of the membrane unit for the separation system were determined in consideration of the separation rate and vehicle installation. This system can supply separated ethanol with sufficient speed and quantity to improve engine performance under practical driving conditions. In addition, the study was conducted to confirm that separated fuels have properties sufficient for use in automobiles. This separation rate enabled 5-cycle-mode driving without temporary shortage of permeated fuel.