• Oat groats were sieved at six different aperture sizes to produce twelve fractions. • Sieved fractions were analyzed for protein, particle size and tribo-charge behavior. • Particle diameters and protein content of sieved fractions were linearly correlated. • Sieved bran fractions were rich in protein and acquired high charges at laminar flow. • Sieved starchy fractions were enriched in protein by tribo-electrostatic separation. The possibility of fractionation of dehulled oat groat particles was assessed through combined sieving and tribo-electrostatic separation approach. Through this process, oat groats were initially sieved at six different aperture sizes of 53 to 425 μm, resulting in the production of twelve different fractions including six starch-rich fractions as well as six bran-rich fractions among which the bran-rich fractions had significantly higher protein concentration (15.5–19.6%) compared to the starch-rich fractions (9.7–14.0%). A linear relationship (R 2 = 0.9554) was found between protein content and mean particle diameters (D 50 ) of all sieved fractions. The tribo-charging behavior of all sieved fractions was assessed as a function of particle size and protein content. Bran-rich particles despite their large particle sizes were acquired higher charge-to-mass (nC/g) ratios compared to starch-rich fractions at the laminar air flow rate (Re = 2091) in contact with polytetrafluoroethylene (PTFE) tribo-charging tube. To exclusively evaluate the tribo-charging behavior of the fractions as a function of protein content, the specific charge (nC/g) values were recalculated to charge density (nC/m 2 ) values and a three-stage incremental trend was observed between charge density and protein content of the fractions at the laminar air flow rate. Starch-rich fractions exhibited totally different tribo-charging behavior at the turbulent (Re = 4182) air flow rate due to the large surface-to-volume ratio of their fine small particles that resulted in enhanced particle–wall collisions and eventually acquisition of higher specific charge values compared to their equivalent sieved bran-rich fractions. The starch-rich fraction sieved at aperture size of 106 μm with protein content of 10.1% was further fractionated through single- and multi-stage tribo-electrostatic separation processes where its protein content was sharply enhanced by ~80% equivalent to over 55% protein separation efficiency.