Nanoscale identification of organic pigments in biogenic calcite by atom probe tomography

Abstract

Spatially-resolved mass spectrometry could contribute to the understanding of biological processes. Atom probe tomography (APT) is a technique which could, in principle, fill an analytical gap for nanoscale mass spectrometry. Yet, because of a limited understanding of the process of fragmentation of biomolecules, biological or biochemical interpretation of the obtained data has so far not been successful. Here, we report on analyses of synthetic and biogenic pigmented calcite minerals. The purple coloration of Sphaerechinus granularis sea urchin spines is due to the occlusion of polyhydroxylated naphthoquinone (PHNQ) molecules, mainly spinochrome A, within the calcite phase. Peaks relating to these pigments as well as to other intra-crystalline macromolecules were identified by APT analysis of the purple spine region with comparison to measurements of synthetic calcite single crystals, calcite crystals doped with PHNQ, and the uncoloured (white) region of sea urchin spines. To facilitate the molecular identification of peaks appearing solely in purple-coloured spines, which are possible fragments from PHNQ, we introduce novel APT data filtering techniques based on spatial and temporal correlations in the detection of ions of targeted mass-to-charge state ratios. This allows us to discuss the fragmentation of organic molecules under field conditions and improve the identification of peaks in the mass spectrum, while also determining nanoscale spatial distribution of fragments related to specifically targeted biomolecules in 3D and elucidate the relationship these organic pigment molecules have to the calcite lattice and inorganic elements such as magnesium. The study aims to improve identification of complex molecular structures by APT, including organic molecules, with the potential ability to provide structural information at a near-atomic scale for a range of materials.