Covalent labelling of β-casein and its effect on the microstructure and physico-chemical properties of emulsions stabilized by β-casein and whey protein

The objective of this work was to investigate the effect of covalent labelling on the physico-chemical properties of β-casein (β-CN) in solution and in emulsions stabilized by β-CN and whey protein isolate (WPI). β-CN was covalently labelled by 5-(and 6)-carboxytetramethylrhodamine, succinimidyl ester (NHS-Rhodamine). The effect of conjugating β-CN with NHS-Rhodamine on the spectroscopic properties of labelled β-CN (β-CN_labelled) was examined. No significant difference in interfacial tension (p > 0.05) was found between mixture of WPI and β-CN_labelled (0.5% w/w WPI/β-CN_labelled) and of WPI and β-CN (0.5% w/w WPI/β-CN) in 10 mM phosphate buffer (pH 7.0) at 20 °C. Oil-in-water emulsions stabilized with either WPI/β-CN or WPI/β-CN_labelled (0.5% w/w) were also investigated using laser-light scattering, analytical centrifugation, rheometry and CLSM. It was shown that labelling had no significant effect on the physico-chemical properties of emulsions (p > 0.05) in terms of droplet size, creaming stability, viscosity or zeta-potential. Confocal micrographs of emulsions made with WPI/β-CN_labelled showed that both β-CN and whey proteins could be observed simultaneously, and were co-localized at the surface of fat globules. Furthermore, it was found through image analysis that β-CN produced a thicker interfacial layer than WPI.