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In the original version of chapter 2, references 5 and 12 were incorrect. In this version, references 5 and 12 are corrected. The corrected references have now been added in the Chapter which reads as follows:

In order to understand the mineralogical difference between biogenic aragonites and their geological or synthetic ones, the transition temperature from aragonite to calcite by heating and the cell lengths of a number of biogenic aragonites have been measured using conventional and high-temperature XRD, as well as those of abiotic ones. Among 21 specimens, most biogenic aragonites showed a transition temperature 60–100 °C lower than that for abiotic ones. However, the shells of land snails showed almost similar transition temperatures. The temperature range from the beginning to the completion of the transition was also varied among the biogenic aragonites. On the other hand, the axial ratios ( and ) of aragonites in marine molluscan species were considerably larger than those of abiotic ones. However, aragonites in freshwater molluscan species and land snails showed axial ratios similar to abiotic ones. X-ray microanalysis suggested that the origin of such abnormal cell lengths was sodium incorporated in the aragonite crystals, not due to lattice distortion induced by the intracrystalline organic molecules proposed in previous researches.

The radula chiton teeth, , were examined using transmission electron microscopy (TEM). After cutting into segments corresponding roughly to three developmental stages from the onset of tooth development, the middle and the fully matured stages, toluidine blue staining has given the posterior side three different color patterns, colorless, reddish-brown, and black colors, respectively. At the colorless stage, the microvilli attached along the surface of the tooth cusp appeared to be dissembled and convert into the lamellar structure in the tooth interior. At the reddish-brown stage, the electron density between fibrous layers increased. A complex of tiny clusters of grains appeared along the fibrous layers. They seemed to aggregate each other to become larger. At the black stage, multiple layers consisting of irregular-shaped and various size of iron minerals were formed. After treating with an aqua regia solution, organic substances have remained between iron minerals, suggesting the abrasion-resistant role at the posterior side of chiton teeth during feeding. In addition, these minerals were randomly arranged. The lattice intervals of the ion minerals varied at an approximate range from 4.8 to 10.2 Å. Also, we have confirmed clearly the lattice fringe of apatite crystal in the core region.

In this study we investigate pristine and experimentally incinerated bovine bone material with differing annealing times and temperatures from 100 to 1000 °C to analyse the crystallographic change of natural bone mineral during cremation. We used X-ray powder diffraction (XRPD) and Fourier transform infrared (FTIR) spectroscopy as complementary methods. We observe a structural change of bone mineral during cremation. Our study highlights that there are only few or even no hydroxyl ions in pristine bone mineral (bioapatite), which is a carbonate-hydro-apatite rather than a hydroxyapatite. A significant recrystallization reaction from bioapatite to hydroxyapatite takes place at elevated temperatures from 700 °C (after 30 min cremation time). This process is associated with a significant increase of crystallite size, and it involves an increase of hydroxyl in the apatite lattice that goes along with a depletion of water and carbonate contents during cremation. Our first results highlight the importance of both time and temperature on the recrystallization reaction during cremation.

The eggshell membranes (ESM) serve as the first interface with the inorganic phase during eggshell formation. During mineral growth, crystals nucleate on the outer side of the ESM at specialized sites called mammillae, mainly consisting of mammillan, a keratan sulfate proteoglycan together with the activity of carbonic anhydrase (CA).

In order to get insight into the mechanisms of chicken eggshell mineralization, ESM was used as a biotemplate for immobilizing carbonic anhydrase (CA) and study in vitro calcite crystallization. Here, we showed that when the eggshell membrane supplemented with immobilized or dissolved carbonic anhydrase is located at the gas-liquid interface, calcite nucleation and growth are sequestered by the ESM scaffold from solution, thus affecting the morphology and size of the crystals formed.

Venomous animals have specialized venom delivery apparatus such as nematocysts, stings, and fangs in addition to the poisonous organs consisting venom gland or sac, which produce and stock the venom. Snake is one of the major venomous animals, of which fangs are connected to the venom gland to inject the venom into prey. Snake’s venomous fangs showed the unique characteristics including mechanical strength and chemical stability. Especially, (habu) snake fangs showed the resistance against its venom digestive proteases, whereas the bones and teeth of mouse were completely digested in the gastrointestinal tract, although habu fangs were also drawn into the body with the . These observations suggest that structural differences exist between venomous fangs and mammalian bones and teeth.

In this study, to reveal the molecular properties of venomous snake fangs, the matrix proteins of (habu) snake venom fang were analyzed by using proteomics experiments using 2D-PAGE and TOF MS/MS analyses. As a result, several biomineralization-related proteins such as vimentin, tectorin, adaptin, and collagen were identified in the venomous fang matrix proteins. Interestingly, the inhibitory proteins against venomous proteins such as metalloproteinase and PLA2 were also identified in fang’s matrix proteins.

Scales of bony fishes are calcified tissues that contain osteoblasts, osteoclasts, and bone matrix, all of which are similar to those found in mammalian bone. The scales are composed of hydroxylapatite (HAP) and extracellular matrix which is mainly type I collagen fibers. We investigated the scales from goldfish by Fourier transform infrared (FTIR) and Raman spectroscopic analyses to characterize the components in the scales. The attenuated total reflection (ATR)-FTIR spectrum obtained from the surface coat of the normal goldfish scale was quite different from that of the backside coat of the same scale. The former showed a strong band at 1013 cm, which was assignable to HAP, and weak bands at about 1643, 1415, and 870 cm, whereas the latter showed a typical protein profile: strong bands at about 1631, 1550, and 1240 cm, which were assignable to amide-I, amide-II, and amide-III of collagen, respectively. We also investigated the local structure of goldfish scales by using micro-Raman spectroscopy. The Raman spectrum of the normal scale showed the amide-I band at about 1640 cm from the collagen and the PO symmetric stretching band at about 961 cm from the HAP. We discuss the implication of Raman and FTIR profiles for normal and regenerating goldfish scales.

Calcium (Ca) storage in bone has a relationship with eggshell production. Forced molting by feeding restriction for older hens improves eggshell quality but leads to a decline in the bone quality. Dietary minerals improve the eggshell and bone quality; however, the effect on bone quality post-molting has not been clarified. This study evaluated the effects of dietary Ca and minerals on the eggshell and bone quality during both pre- and post-molting periods. The bone quality was evaluated by measurement of bone density and Fourier transform infrared spectroscopy (FT-IR) analysis. The eggshell quality was evaluated by morphological observation of the mammillary cores by scanning electron microscope and FT-IR analysis. The high Ca concentration feed group showed a low bone density post-molting and a high carbonate/phosphate ratio pre-molting. In high mineral concentration feed group, the eggshell strength, the thickness, and the proportion of large mammillary core areas were significantly higher ( < 0.05) than control group post-molting. These results suggest that the eggshell strength increases as the proportion of mammillary core areas increases. Furthermore, the high carbonate/phosphate ratio promoted a decrease in bone density. Therefore, the concentration of dietary mineral is strongly related with the maintenance of eggshells and bone quality post-molting.

Molluscan shells display a wide variety of pigmentation patterns. The diversity in molluscan shell color reflects the variety of different chemical species in the shell surface. Chemical characteristics of molluscan shell pigments have been extensively investigated, and compounds including porphyrins, polyenes, and melanins were identified as shell pigments. Here, we investigated shell pigments in 24 species in the family Neritidae using Raman spectroscopy. An excitation wavelength of 514.5 nm revealed two types of Raman spectra. One was characterized by two peaks ranging in wavenumber from 1100–1200 to 1500–1600 cm, which indicate the presence of polyenes. Another type remained unassigned, implying the presence of other pigments such as porphyrins or melanins. The Raman spectra indicated a different distribution of the two types of pigments in shells. The patterns of the Raman spectra had no obvious relationship with taxonomical classification lower than the genus level and typical habitats. Measurement of the Raman spectrum at an excitation wavelength of 442 nm suggested that the wavelength can distinguish polyenes from other types of pigments.

Three-dimensional (3D) reconstruction is an essential approach in morphological studies in biology and paleontology. Seeking an optimized protocol for nondestructive observations, we attempted 3D visualization of various molluscan shells and animals with X-ray micro-computed tomography (micro-CT). Calcified parts of molluscs were easily visualized except for cases with marked differences in thickness heterogeneity. 3D imaging of shell microstructure was difficult. Visualization of soft tissue requires staining to enhance the image contrast. Especially for soft tissues, synchrotron X-ray microtomography is the most advanced method to generate clear 3D images. 3D data facilitates morphological quantification, enabling calculations of length and volume even for very complex forms. X-ray micro-CT is extremely useful in the morphologic examination of mineralized and soft tissues, although microstructural and histological details should be supplemented by other microscopic techniques.