Separation of Ovotransferrin and Ovomucoid from Chicken Egg White
Ovotransferrin and ovomucoid were separated using two methods after extracting ovotransferrin- and ovomucoid-containing fraction from egg white. Diluted egg white (2x) was added with Fe3+ and treated with 43% ethanol (final conc.). After centrifugation, the supernatant was collected and treated with either a high-level ethanol (61%, final conc.) or an acidic salt combination (2.5% ammonium sulfate and 2.5% citric acid) to separate ovotransferrin and ovomucoid. For the high-level ethanol method, ovotransferrin was precipitated using 61% ethanol. After centrifugation, the precipitant was dissolved in 9 vol. distilled water and the residual ethanol in the solution was removed using ultrafiltration. The supernatant, mainly containing ovomucoid, was diluted with 4 vol. water, ethanol removed, concentrated, and then used as ovomucoid fraction. For the acidic salt precipitation method, the ethanol in the supernatant was removed, first. The ethanol-free solution was concentrated and treated with 2.5% ammonium sulfate and 2.5% citric acid combination. After centrifugation, the precipitant was used as ovotransferrin and the supernatant as ovomucoid fraction. The ovomucoid fraction from both of the protocols was further purified by heating at 65 oC for 20 min and the impurities were removed by centrifugation. The yields of ovomucoid and ovotransferrin were > 96% and > 92%, respectively. The purity of ovomucoid was > 89% and that of the ovotransferrin was > 88% purity. ELISA results confirmed that the activity of the separated ovotransferrin was > 95%. Both of the protocols separated ovotransferrin and ovomucoid effectively and the methods were simple, fast and easy to scale-up.