Eggshell membrane removal for subsequent extraction of intermineral and intramineral proteins

Biominerals are composite materials with organic and inorganic components. The organic components influence nucleation, growth, and the physical and material properties of the biomineral. Calcium minerals account for about 50% of all biominerals, and polymorphic calcium carbonate is dominant with calcite and aragonite being the most common natural forms. The eggshell of the domestic fowl Gallus gallus comprises calcite, in association with organic components, precipitated on membranes. Environmental scanning electron microscopy (E-SEM) analysis displays this intimate mineral-organic intergrowth between the inner membrane and the calcite mammillary caps. Before analysis of organic components within the eggshell, the inner membrane must be removed. Membrane removal is achieved by either plasma ashing (etching), sodium hypochlorite, or acid (HCl) treatment. These three methods are compared with the aim of subsequent extraction of intra- and intermineral proteins. Plasma ashing is suitable for subsequent SEM of the inner surface of the eggshell but entirely unsuitable for subsequent protein extraction. Removal of the inner membrane by sodium hypochlorite or acid (HCl) treatment are both suitable methods. Sodium hypochlorite treatment is the preferred method since the protein yield is highest here, and, unlike acid treatment, it does not remove part of the calcite fraction.