Microstructure and energy dispersive diffraction reconstruction of 3D patterns of crystallographic texture in a shark centrum

Abstract

Purpose: Tomography using diffracted x-rays produces reconstructions mapping quantities such as crystal lattice parameter(s), crystallite size, and crystallographic texture, information quite different from that obtained with absorption or phase contrast. Diffraction tomography is used to map an entire blue shark centrum with its double cone structure (corpora calcerea) and intermedialia (four wedges).Approach: Energy dispersive diffraction (EDD) and polychromatic synchrotron x-radiation at 6-BM-B, the Advanced Photon Source, were used. Different, properly oriented Bragg planes diffract different x-ray energies; these intensities are measured by one of ten energy-sensitive detectors. A pencil beam defines the irradiated volume, and a collimator before each energysensitive detector selects which portion of the irradiated column is sampled at any one time. Translating the specimen along X; Y, and Z axes produces a 3D map. Results:We report 3D maps of the integrated intensity of several bioapatite reflections from the mineralized cartilage centrum of a blue shark. The c axis reflection's integrated intensities and those of a reflection with no c axis component reveal that the cone wall's bioapatite is oriented with its c axes lateral, i.e., perpendicular to the backbone's axis, and that the wedges' bioapatite is oriented with its c axes axial. Absorption microcomputed tomography (laboratory and synchrotron) and x-ray excited x-ray fluorescence maps provide higher resolution views. Conclusion:The bioapatite in the cone walls and wedges is oriented to resist lateral and axial deflections, respectively. Mineralized tissue samples can be mapped in 3D with EDD tomography and subsequently studied by destructive methods.

Bioapatite in shark centra studied by wide-angle and by small-angle X-ray scattering

“…For blocks Tg2c and Tg2d, the axis of the pair of D -period arcs are roughly normal to the axis of the centrum, with the deviation from 90° being attributable to tilt of the blocks within the tubes. In other words, the bioapatite c -axes of the tiger shark cone walls are perpendicular to the vertebral column axis, in agreement with EDD mapping results for a blue shark centrum [11,12] (another carcharhiniform). The SAXS pattern of L2d (figure 5) also appears to be from cone wall material and has the same c -axis orientation.…”

“…[10] found that the mineral in both tissues is a form of hydroxyapatite (hAp) similar to the bioapatite reinforcing bone, a biocomposite with a different matrix, type I collagen. Since the data of this paper were collected, energy-dispersive (X-ray) diffraction (EDD) of intact shark centra confirmed recently that bioapatite was the mineral phase present and revealed interesting crystallographic texture [11,12]. These studies did not report lattice parameters, and it is unclear whether the mineral is hAp (a relatively pure phase containing low levels of impurities) or carbonated apatite like that found in bone and tooth.…”

“…Even though the ancestors of sharks and of mammals diverged approximately 420 Ma [25], the mineralized tissue of sharks and of bones are both bioapatite nanoparticle-reinforced composite materials based on collagen matrices. Previous studies showed shark centra and tesserae contained a bioapatite similar to that in bone [8–11,18], but quantitative data on the mineral phase (precise and accurate lattice parameters and crystallite size) and its nanoscale organization within the cartilage matrix were lacking. Given the different collagen-based matrices (bone's type I collagen fibrils—triple helices of two α 1 chains and one α 2 chain versus shark's type II collagen fibrils—triple helices of three α 1 chains [26]), one could only speculate whether there were significant differences in the two bioapatites' crystal lattice parameters, nanoparticle sizes and nanoparticle organization, characteristics which can affect tissue mechanical properties.…”

“…[12] and Stock et al . [11] found that the intermedialia bioapatite also contained strong crystallographic texture, with c -axes being oriented along the centrum axis, i.e. at 90° to that in the cone walls.…”

“…[2,4,5,7,8,11,12,22,34], this tissue remains much less investigated than bone or tessellated tissue. Although shark tesserae are the closest mineralized tissue to those of centra tissue, tesserae contain just under 10 vol% soft tissue/porosity [41] whereas centra cones contain approximately 60 vol% and centra intermedialia approximately 75 vol% soft tissue/porosity [11], suggesting that it would be inaccurate to extrapolate centra tissue materials properties from tesserae data. Centrum bioapatite mineral's lattice parameters a and c , crystallite size L c and nanoparticle spacing D are close enough to those of bone (less than 1% and less than 5% differences for lattice parameters and D , respectively, and L c 50% larger than that the authors typically measure for bone at this beam line but within the range reported for bone), that it may be possible, as a zero-order approximation, to extrapolate trends observed in bone tissue to shark centrum tissue.…”