An efficient, scalable and environmentally friendly separation method for ovoinhibitor from chicken egg white

A separation method for ovoinhibitor from egg white without using toxic chemicals was established. After sequentially removing lysozyme, ovomucin, ovotransferrin and ovomucoid from egg white using Amberlite FPC 3500 ion exchange resin, pH adjustment, acidic ammonium sulfate and ethanol (35% final concentration, v/v) respectively, the resulting ovalbumin-rich precipitant was homogenized with 4 vol of distilled water and then centrifuged. The extraction was done twice and the extracts containing crude ovoinhibitor were pooled, concentrated using ultrafiltration and then subjected to 35% saturation ammonium sulfate (final concentration) precipitation. The precipitated ovoinhibitor was dissolved in acidic water (pH 3.0), desalted and concentrated using ultrafiltration, and then lyophilized. The product had 90.8% purity with a yield of 70.0% and its identity was confirmed by the MALDI-TOF MS/MS. The inhibitory activity of the separated ovoinhibitor showed that it maintained protease inhibitory activities against elastase, trypsin, alpha-chymotrypsin, and subtilisin. This result indicated that the protocol was highly efficient, scalable, and environmentally friendly for the separation of ovoinhibitor from the egg white, and the ovoinhibitor separated has a great potential to be used as a functional protein.