Top-down graphene production via exfoliation from graphite produces a mass of graphene with structural variation in terms of the number of layers, sheet size, edge type, and defect density. All of these characteristics affect its electronic structure. To develop useful applications of graphene, structural separation of graphene is necessary. In this study, we investigate the adsorption behavior of different types of graphene fragments using a multicolumn gel chromatography system with a view to developing an efficient method for separating high-quality graphene. The graphene was dispersed in an aqueous sodium dodecyl sulfate (SDS) surfactant solution and flown through allyl-dextran-based gel columns connected in series. In the chromatographic operation, we observed that the small-sized or oxidized graphene fragments tended to bind to the gel and the relatively large-sized graphene with a low oxygen content eluted from the gel column. In this system, the adsorbed SDS molecules on the graphitic surface prevented graphitic materials from binding to the gel and the oxygen functional groups on the graphene oxide or at the abundant edge of small-sized graphene hindered SDS adsorption. We hypothesize that the reduced SDS adsorption density results in the preferential adsorption of small-sized or oxidized graphene fragments on the gel. This type of chromatographic separation is a cost-effective and scalable method for sorting nanomaterials. The structural separation of graphene based on the adsorption priority found in this study will improve the quality of graphene nanosheets on an industrial scale.