The exploration of milk fat crystallization in milk fat globules by confocal Raman microscopy

Crystallization behavior within oil-in-water emulsion is a key factor for the properties and stability of many food products. Confocal Raman microscopy is a promising method to study such complex structures in situ. This study aimed at evaluating the feasibility of confocal Raman microscopy for visualizing milk fat crystallization and establishing a reliable data acquisition and processing methodology. Milk fat globules from raw milk were fixated in an agarose gel and crystallized at different temperatures. Confocal Raman microscopy was applied to collect two-dimensional area scans and supporting images were obtained by polarized light microscopy. The results revealed differences in lipid characteristics, crystal formation, and spatial distribution as a result of crystallization. Specific C-C stretching vibrations at 1063, 1083, and 1125 cm-1 were found to indicate lipid chain mobility and provide quantitative information on crystallinity. Additionally, the study successfully identified the triple-layered milk fat globule membrane. Different approaches for processing spectroscopic data were compared, emphasizing the importance of proper data handling. This novel spectroscopy imaging approach has significant potential in enhancing our understanding of structural heterogeneities of crystallized structures within complex food matrices.