Abstract: Emulsion formation during enzyme-assisted aqueous extraction of soybean oil is an important bottleneck restricting the popularization of enzyme-assisted aqueous extraction. The particle size distribution and spatial distribution of lipids and proteins in emulsions obtained after different times (1, 2, 3 h) of enzymatic hydrolysis of soybeans were characterized by dynamic light scattering and laser scanning confocal microscopy. The mechanism for the attenuation of fluorescence intensity in emulsions was analyzed by Raman spectroscopy based on hydrophobic interaction of chromophores, disulfide bonds and oil-protein interactions. The results showed that enzymatic hydrolysis resulted in the cleavage of proteins in emulsions into small peptides, which fell off from the surface of the oil droplets, leading to the aggregation of the oil droplets. The particle size increased with extended hydrolysis time, and the number of oil droplets decreased correspondingly, becoming irregular in shape. Laser scanning confocal microscopy showed that the fluorescence intensity was attenuated with increasing hydrolysis time. The Raman spectral intensity of chromophores from the emulsion resulting from enzymatic hydrolysis of whole soybean flour was dramatically reduced as compared with that from soybean protein isolate (SPI) at the same hydrolysis time and it decreased with the increase in hydrolysis time. These findings led us to conclude that hydrophobic oil-protein interaction in emulsions shielded the intermolecular hydrophobic groups and masked the chromophores, thus resulting in a reduction of fluorescence intensity.

Key words: emulsion, spatial distribution, fluorescence intensity, Raman spectrum, hydrophobic interaction