Microscopic characterisation and composition of proteins from lupin seed (Lupinus angustifolius L.) as affected by the isolation procedure

Two important techniques for protein isolation from vegetable sources are well-established, the alkaline extraction with subsequent isoelectric precipitation and the salt-induced extraction followed by dilutive precipitation. Both techniques provide protein isolates with different properties. In the case of lupin the isoelectric protein isolate is commercially applied as an egg-substitute in diverse food products, whereas a specific dilutive lupin protein isolate exhibits fat-like properties and shows the high potential of lupin proteins for fat replacement. However, the reason for this behaviour has not been investigated up to now. Therefore, the influence of common precipitation parameters on structure formation of proteins from Lupinus angustifolius L. Vitabor was investigated using bright field light microscopy, fluorescence microscopy and cryo-scanning electron microscopy. The structure formation upon application of nine different procedures revealed that precipitation at pH. 4.5 always led to an unfolding of the protein indicating an irreversible denaturation. Applying the dilutive precipitation, formation of flexible but well-ordered globular aggregates was observed. This effect persisted after combining dilutive and pH mediated precipitation procedures. Only the isoelectric lupin protein isolate showed autofluorescence properties at three wavelengths. A number of indications evidenced that the physical protein structure was responsible for differences in protein behaviour rather than variations in the chemical composition of the protein isolates.