Crystalline and amorphous calcium carbonate as structural components of the Calappa granulata exoskeleton

The exoskeleton of crustaceans consists of chitin biopolymers where the embedded inorganic biominerals, mainly CaCO₃, affect strongly its mechanical properties. Raman and Near Edge X-ray Absorption Fine Structure (NEXAFS) spectroscopies and Transmission Electron Microscopy (TEM) are applied to investigate the CaCO₃ structure in various parts of the Calappa granulata crab exoskeleton. The shape of the main Raman peak of CaCO₃ reveals the presence of two phases which are identified as calcite and amorphous calcium carbonate (ACC). The relative concentration of the two phases in various parts of the exoskeleton is determined from the area ratio under the corresponding peaks. The results of the Ca L_3,2-edge NEXAFS analysis are in line with the Raman findings, since the energy separation of peaks that appear in the lower frequency region of the main L₂ and L₃ peaks due to crystal field splitting, is directly related to the percentage of the ACC phase in the total CaCO₃ mineral content. The C K-edge spectra are used for the determination of the extent of calcification of the exoskeleton. Furthermore, dark and bright field TEM images reveal the presence of nanocrystallites with an average size of 20 nm. The structure of the nanocrystallites, as derived from the Selected Area Electron Diffraction patterns, is calcite. The results suggest that ACC plays a structural role in the exoskeleton of Calappa granulata.