Simultaneous separation of the four major allergens of hen egg white

Hen egg is a worldwide top-consumed food that has attracted public health concerns because it can induce allergic reactions in sensitized individuals. Food allergy investigations need highly purified egg allergens. However, a limited number of purification methods have been described for the combined separation of more than two egg allergens and only few of them have evaluated the immunological activity of these purified proteins. The aim of this work was to develop a chromatographic method for the separation of the four major egg allergens (ovomucoid, ovalbumin, ovotranferrin, and lysozyme) with a demonstrated immunological activity. After a pre-processing step for ovomucin precipitation and pH adjustment, remaining egg white proteins were loaded onto CM-Sepharose column and major egg allergens were separated using cation-exchange chromatography. Yield of ovomucoid, ovalbumin, ovotranferrin, and lysozyme was 60.0%, 52.1%, 29.6%, and 90.2%, respectively. Purified allergens were compared with their commercial standards, showing a high purity as well as a maintained antigenicity. The protocol described in this work is simple, quick, low-cost, and suitable for the study of the immunological properties of these allergens. For higher ovalbumin demand in the lab, 2.1 g ovalbumin can be produced in a single process with high purity.