S C

IE N

T IF

IC H

IG H

LI G

H T

S S

T R

U C

T U

R E

O F

M A

T E

R IA

L S

1 2 3 I H I G H L I G H T S 2 0 2 2

Fig. 115: a) Micrograph of a juvenile Pinctada margaritifera specimen. The black rectangle indicates the investigated shell growth zone. b) Scanning electron microscopy image of the shell edge, showing the

isolated, disc-like early mineralisation units at the top and the more developed prism structure that is slowly forming on the periostracum, an organic membrane. c) Stimulated Raman scattering map, showing the

symmetric contrast between ACC (red) and calcite (blue). The presence of more ACC in young disc-like units and the accumulation of ACC in the inter-prismatic space is noteworthy.

Despite the importance of this amorphous phase for the shell formation, comparatively little is known regarding its structural organisation, spatial localisation and the potential presence of multiple, distinct ACC phases. This lack of knowledge stems from the difficulty in characterising the ACC phase properly, mostly due to the concomitant presence of crystalline phases with significantly stronger scattering power than the amorphous phase. This investigation used beamline ID15A and a 60 keV nanobeam for X-ray total scattering, involving a novel approach to separate crystalline and amorphous scattering contributions as a part of the ERC-funded research project 3D-BioMat (No. 724881).

This approach provided the sub-micron, spatially resolved atomic pair distribution function (PDF) of the pearl oyster shell Pinctada margaritifera. By acquiring data over a large portion of the reciprocal space made accessible by sample rotation in the beam, it was possible to exploit the fact that the crystalline scattering contribution is sharply distributed into Bragg peaks,

whereas that of the amorphous phase is distributed isotropically, meaning that the two can be reliably separated by filtering. In this way, it was possible to obtain sub-micron, spatially resolved atomic pair distribution function (PDF) of the amorphous phase in the Pinctada margaritifera shell.

Three ACC states were distinguished in the shell, along the line scan shown in Figure 116a, and assigned to different growth zones by using principal component analysis (Figure 116b). 3D atomic structural models refined against PDF data provided bond parameters showing that the ACC structures along the line scan differ mostly in their Ca-O atomic pair distances but are otherwise remarkably similar (Figure 116c). Together with spectroscopic techniques (Raman spectroscopy, coherent Raman microscopy, SEM-EDX) showing the localised presence of Mg in the oyster shell, including in the crystalline structure, these results demonstrate the key role Mg plays in the transition between different amorphous states.

Fig. 116: a) X-ray fluorescence image indicating the location of the scan presented (red line). b) Principal components analysis of the PDF data, showing the presence of three spatially separated contributions. The red lines indicate the border of the disc-like unit. c) Ca-O pair distance extracted from the structural model, showing the contraction of the Ca-O pair towards the centre of the disc.

The green lines indicate the border of the disc-like unit.