Reconstituted oil bodies characterization at the air/water and at the air/oil/water interfaces

Oil bodies (OBs) are micelle-like structures with an outer phospholipid monolayer embedding some specific proteins (oleosins) and surrounding a hydrophobic core of triacylglycerols (TAGs). Oleosins are alkaline hairpin-like proteins that are anchored into the OBs structure with their hydrophilic domains covering the surface. We performed surface pressure (Π) and Brewster Angle Microscopy investigations of reconstituted OBs (ROBs) and of trypsin digested ROBs. The obtained Π vs time isotherms clearly show the formation of a surface layer. Upon ROBs suspension injection into the subphase, a clear-cut Π enhancement is recorded, followed by a long plateau region for ROBs suspensions more concentrated than 12.5μg/ml. The BAM analysis highlighted the presence of a dark background, ascribable to a 2D layer due to free components rearrangement and brilliant circular 3D domains, due to unaltered ROBs or small aggregates of ROBs. Increasing ROBs concentration, large domains appeared. We hypothesize that the presence of an excess of free TAGs in the 2D layer is crucial for the generation of such domains. We verified the generation of such typical structures, studying the behavior of a ROBs suspension (concentration of 12.5μg/ml) with two different approaches: after injection under a concentrated TAGs floating layers and after digestion with trypsin. These two procedures resulted in similar effects since proteinase digestion is like to induce the same morphology of a TAGs excess.