Sequential Separation of Value-Added Components from Egg Yolk

Abstract

Egg yolk contains a number of components that can play significant functional roles in foods and human. Although some of the egg yolk components are already isolated and commercialized, the separation methods are largely limited to a single component and the rest are discarded. Therefore, majority of vital natural resources can be wasted. At present, most of the developed methods for the separation of value-added components from egg yolk are for laboratory scale and cannot be used in commercial scales. The objectives of this research are 1) to compare currently available separation methods for value-added elements from egg yolk, 2) to develop an efficient scale-up preparation method for each element for the practical use of egg components, and 3) to develop a sequential separation method for IgY, phosvitin, and phospholipids and neutral lipids from egg yolk. Because the separated components will be used for humans, we try to minimize or eliminate the use of harmful chemicals and organic solvents in the separation methods. The main tools used for the separation of value-added components from egg yolk include dilution with water, pH adjustment, centrifugation, ultrafiltration, and salt precipitation. First, the phosvitin-containing granules were separated from the rest of the egg yolk components using 2 volumes of water (3x dilution) and centrifuged. The precipitant was collected and phosvitin was extracted using 10% NaCl (w/v) in 0.05N NaOH solution and homogenization. The homogenate was diluted with DW and the pH adjusted to 4.0, and then precipitant was removed by centrifugation. The resulting supernatant was desalted and concentrated using ultrafiltration, and the impurities were removed by heat treatment. After centrifugation, the phosvitin-containing supernatant was freeze-dried. The supernatant of the 3x-diluted egg yolk was used to separate IgY and lipids. The supernatant fraction was diluted with 3 volumes of water (4x dilution) and centrifuged. Phospholipids and neutral lipids were extracted from the solid fraction using ethanol and hexane, respectively. Phospholipids were collected after removing ethanol from the ethanol extract, and neutral lipids was recovered after removing hexane using a rotary evaporator. The liquid fraction was concentrated using ultrafiltration and then IgY was precipitated using 20% saturated (NH4)2SO4+15% NaCl (w/v). The precipitant containing IgY was dissolved in water, desalted using ultrafiltration, and then freeze dried. The SDS PAGE and Western Blot analyses indicated that the phosvitin separated had over 90% purity and 91% yield, and IgY had over 90% purity and 80% yield. The method developed protocol can not only be applied for individual component but also for sequential separation of multiple components, can be immediately applicable for scale-up processing, and the separated components can be used for humans as well as drug industry because no toxic compounds or organic solvents were used in the separation processes.