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Egg yolk is well known as a natural oil-in-water emulsion. Because of its multifunctional properties egg yolk is extensively used in the food, medical, pharmaceutical, and cosmetics. The protein content of egg yolk represents about 15.7–16.6% of all hen egg yolk compounds. Livetins fraction with α, β, and γ-livetins is relatively heterogeneous and accounts for about 9.3% of hen egg yolk proteins. The literature is lacking information on the physicochemical and functional properties of the IgY-rich livetins fraction obtained from hen egg yolk. If physicochemical and functional properties of livetins were better known that would pave way for more efficient or more specific use of this fraction (livetins) in food, nutraceutical, medical and pharmaceutical industries. Hence, the objectives of this study were to determine the physicochemical and functional properties of the livetins fraction obtained from egg yolk in a wide range of pH conditions.
Freshly laid white shell eggs were obtained from the PRC of the University of Alberta. Egg yolks were manually separated from the white and carefully rolled on Whatman paper (grade No. 4) to remove all albumen from the vitellin membrane. Egg yolk was diluted ten times (w/w) with distilled water and IgY was collected, concentrated and freeze dried until further testing. Total nitrogen content of the livetins was measured by Dumas Combustion method. Protein content was calculated by multiplying the total nitrogen value by the conversion factor of 6.25. Fat content was determined by extracting with petroleum ether for 8 h using a soxhlet apparatus and the fat content was expressed as g/100 g sample. The average yield was calculated by measuring the amount of freeze-dried livetins recovered as a percentage of 100 g fresh egg yolk used for the experiment. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) under non-reducing conditions was carried out using 4–20% Mini-PROTEAN TGX precast protein gels using Mini-PROTEAN tetra cell unit to check the status of livetins (including IgY). Foaming properties (foam capacity and foam stability), emulsifying activity index (EAI) and emulsion stability index (ESI) were evaluated.
The average yield of the freeze-dried livetin fraction was 2.41±0.10 g/100 g fresh egg yolk. Livetins represented about 4.6% yolk dried matter. The protein and fat contents of the livetin fraction were 86.3±1.10% and 1.92±0.63%, respectively. The protein electrophoretic profile of livetins showed a distinct protein pattern in the range of 33–203 kDa. The isolated fraction containing α, β and γ-livetins clearly shows the presence of 6 major protein bands having MW of 33, 36, 47, 55, 73, and 203 kDa. At pH 6 and 7, livetins exhibited relatively lower solubilities when compared to the solubilities at other pH values (from 2 to 5 and 8–12). High solubility profile of livetins at various pH values could be useful in the preparation of various food/pharmaceutical formulations. In the current investigation, the single endothermic peak (83.3 °C) can be the convolution of various proteins such as α-livetin (albumin), β-livetin (α-2-glycoprotein) and γ-livetin (IgY) that are present in the livetins fraction. In the current study, livetins showed intramolecular (native) β-sheet structures and no detectable intermolecular β-sheets, which indicates denaturation. Livetins showed foam capacity in the range of 21–58% at various pH values. Livetins exhibited foam stability. Not much difference was observed in the foam stability of livetins after 5 and 10 min; however, the foam stability decreased after 30 and 60 min as compared to that at 5 min. Livetins showed higher foam stability (20–56%) after 5 min, whereas the foam was stable to an extent of 5–36% and 3–33% after 30 and 60 min, respectively. The current study results indicate that the livetins are effective emulsifiers at the o/w interface in the formation and stabilization of emulsions. Livetins showed higher emulsifying stability at alkaline pH 12.
For the first time, physicochemical (solubility, surface hydrophobicity, zeta potential, denaturation enthalpy, and protein structural properties) and functional properties (foaming and emulsifying properties) of livetins fraction isolated from hen egg yolk were determined. Livetins exhibited solubility, foaming properties and emulsification activities at various pH values (2–12). Zeta potential results showed the isoelectric point at pH 5.7. Thermal analysis of livetins indicated the denaturation temperature of 83.3 °C. The results suggest that livetins with desirable physicochemical and functional properties could be used as functional ingredients in various food product applications.