Fluorescein functionalized random amino acid copolymers in the biomimetic synthesis of CaCO3

In nature we find a large variety of biocomposites with complex structures to fulfill structural functions such as skeletal support and protection of soft tissues. For calcium carbonate based biominerals, acidic proteins have been reported to be instrumental in controlling the mineral formation process, including the control over nucleation, growth and polymorph selection. To unravel the crucial physicochemical characteristics of these biopolymers, we have synthesized random amino acid copolymers with varying Glu/Asp/Ala ratios using N-carboxy anhydride ring opening polymerization (NCA ROP) and a post-polymerization modification with fluorescein. As these polymers lack a specific order in their amino acid sequence, as well as a defined secondary structure under mineralization conditions, they can only influence the mineralization reactions through their amino acid composition which decides the Glu/Asp ratio and the hydrophilic/hydrophobic balance. Where P(Asp-co-Ala) produced dog-bone like crystals elongated along the crystallographic c-axis, P(Glu-co-Ala) led to the formation of rounded calcite. Also for P(Glu-co-Asp-co-Ala) the calcite crystals were elongated along the c-axis but they were overgrown with rounded calcite rhombs, crystallographically aligned with the elongated core. Hence, the ter-copolymer combines the effects of the two di-copolymers, leading to single crystals with unusual complex morphologies. The presence of a fluorescent group allowed us to study the location of the polymers in the mineral phase using fluorescence microscopy. This demonstrated that while controlling the nucleation and growth of calcite, all polymers were also incorporated within the crystals