3D nanoporous structures of a set of bicontinuous microemulsion-fabricated polymeric membrane films were examined using synchrotron X-ray nanotomography imaging. The interconnected pores across the membrane were displayed as visualized traces by a track analysis, where the ratio of the total length of the trace divided by the membrane thickness represented the tortuosity of the porous structure and indicated a correlation with reduced oxygen transmissibility through the membranes. 3D volume renderings of the membrane provided pore morphology with interconnecting channels and quantitative porosity. The results indicate the potential use of this nanoscale imaging technique in assessing the structure-related transport properties of nanostructured functional materials.