Purpose: The purpose of this study was to investigate the colloidal structures formed on digestion of medium chain triglyceride (MCT) with a specific objective of identifying and characterizing a previously reported vesicular phase, which has been linked to supersaturation and anomalous digestion kinetics, and to evaluate the influence of lipid mass and enzyme inhibition on self assembled structure. Methods: MCT was digested in vitro and nanostructure was monitored in real time using synchrotron small angle X-ray scattering (sSAXS), and morphology was studied using cryogenic transmission electron microscopy (cryo-TEM). Results: Formation of the putative vesicular phase formed on digestion of MCT was confirmed and its structural attributes were determined. Vesicle formation was dependent on lipid mass and bile salt concentration. The use of enzyme inhibitor for offline analysis of lipolysis samples did influence structural aspects of the digestion medium when compared to real time evaluation. Conclusions: The formation of a vesicular phase was directly linked to the kinetics of lipid digestion. Vesicle formation is linked to lipid mass, or more specifically the ratio of lipid to bile salts present in the digestion mixture. Inhibition of lipase to halt digestion during sampling for offline analysis must be done with caution as structural aspects were shown to differ for the MCT digests with and without inhibitor present. ? 2013 Springer Science+Business Media New York.