CaCO3 (Sethmann)

Calcium carbonate biomineral formation and bio-inspired mineralisation in hydrogel matrices

Ingo Sethmann, Hans-Joachim Kleebe

Biomineralised hard parts, such as skeletons, teeth, and shells, are produced by organisms under cellular control, i.e., under physiological conditions. In many cases they show elaborate morphologies and ultrastructures that are adapted to specific functions. CaCO3-minerals, predominantly calcite and aragonite, are among the most common biomineral phases. These mineral phases are usually intimately associated with a so-called organic matrix to form organic-inorganic composites. These organic matrices must already be present while mineral precipitation takes place. Hence, they are expected to influence crystal growth and may play a role in the controlled formation of complex, often hierarchically organised structures. However, the factors that control this morphogenesis and structure formation in biomineralisation are still not well-known. The aim of this investigation is to learn more about the role of the organic matrices.

As an example of biominerals, we investigated the sclerites that form a flexible supporting skeleton of octocoral (Sinularia sp.) colonies. The sclerites are mostly spindle-shaped millimetre-sized objects with tubercles on their surfaces (Fig. 1a). Structurally, the sclerites are composed of densely packed Mg-calcite fibres, which are embedded in a network-like organic matrix and largely oriented parallel to the morphological axis of the sclerite. The sclerite surfaces, however, were found to consist of nano-granular layers with a transition towards the internal bulk structure by aligned aggregation and fusion of nanogranules to form crystal fibres (Fig. 1b). This finding strongly suggests that the sclerites grow by precipitation of nanoparticles onto the sclerite surfaces in the presence of an organic matrix and subsequent transformation of the particles into calcite crystal fibres by self-assembly. Although the driving force of this elegant self-assembly of hierarchically structured material remains unclear, we assume that the organic matrix is involved in the modification of the crystal growth pattern.

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Fig. 1: Octocoral sclerites, (a) complete sclerite, (b) transition from nanoparticles to crystal fibres by aligned fusion; scanning electron micrographs (SEM).

As an experimental approach to a better understanding of the role of organic network matrices in biomineral formation, we carried out bio-inspired CaCO3 precipitation in pH-buffered polyacrylamide (pAAm) hydrogel by counter-diffusion of calcium and carbonate ions in U-shaped glass tubes. The resulting calcite crystals resembled hierarchically structured aggregates of aligned rhombohedral microcrystals (Fig. 2a). Cleaved and 'premature' crystals revealed that the aggregate structure developed only after the initial crystal had been grown to a certain size. Correspondingly, the crystal surfaces of aggregate blocks showed nanogranular compositions (Fig. 2b), while those of 'premature' non-aggregate crystals were almost flat. In cleaved aggregate blocks, the internal nanogranular appearance is more pronounced directly below its growth surface than in its core. Thermo-gravimetric and differential thermal analyses (TGA/DTA) revealed that small amounts of the gel matrix became incorporated into the aggregate structure, probably wedged between the aggregate blocks. These experimental observations suggest that with progressive growth of the crystals, the gel matrix was pushed outwards and became compressed at the crystal surface, where it then formed an obstacle for crystal growth and gave rise to crystal growth with an aggregate pattern on a micro- and nanometre scale as well as to incorporation of parts of the organic network. During crystal growth, nanogranular surface aggregates seem to fuse and form the compact crystalline material of the aggregate blocks.

biomineral

Fig. 2: Calcite crystal grown in pAAm hydrogel, (a) complete crystal (SEM), (b) nanogranular crystal surface (atomic force micrograph, AFM).

Although the experimental precipitation system is rather simple compared to biological systems as in octocorals, the resulting mineralised structures seem to resemble each other surprisingly well in their crystal growth mechanism, their hierarchical structure, and their composite nature. Therefore, we assume that, in addition to being a structural component of the final material, one role of organic network matrices in biomineralisation is to modify the mineral precipitation and crystal growth pattern to facilitate the formation of hierarchical structures. Additionally, controlled precipitation and aggregation of nanoparticles enable the construction of mineralised hard parts with complex and rounded morphologies.