In situ transmission electron microscopy observations of CaCO3 crystallization onto polysaccharide-coated nanoparticles

Abstract

Polysaccharides and proteoglycans are widely associated with the organic matrix at sites of CaCO₃ biomineralization, and previous studies indicate that these macromolecules may confer greater roles in mineral nucleation than previously recognized. This investigation uses in situ liquid-phase transmission electron microscopy (LP-TEM) to observe CaCO₃ nucleation onto aminated silica (SiO₂–NH₃⁺) nanoparticles treated with a layer of chitosan (near-neutral derivative of chitin) or heparin (a carboxylated and highly sulfated glycosaminoglycan). In the absence of polysaccharides, few CaCO₃ particles formed and exhibited mobility. However, the SiO₂–NH₃⁺ nanoparticles were enveloped in a region of higher mass density relative to the bulk solution, suggesting the development of a local solute-rich environment that surrounds the charged NH₃⁺ groups. The heparin- or chitosan-coated silica particles also exhibited regions of higher mass density around the nanoparticles. In the presence of these polysaccharide coatings, we observed the nucleation of abundant CaCO₃ particles whereby the polyanionic heparin promoted more nucleation than the weakly cationic chitosan. Many crystallites appeared to form at the polysaccharide–TEM cell membrane–solution interface, further indicating interfacial and macromolecule-specific control on crystallization. The combined results demonstrate that chitosan and heparin have an appreciable effect on the timing, size, and location of CaCO₃ nucleation compared to the polysaccharide-free nanoparticles.