Colloidal pathways of amorphous calcium carbonate formation lead to distinct water environments and conductivity

Abstract

CaCO₃ is the most abundant biomineral and a major constituent of incrustations arising from water hardness. Polycarboxylates play key roles in controlling mineralization. Herein, we present an analytical and spectroscopic study of polycarboxylate-stabilized amorphous CaCO₃ (ACC) and its formation via a dense liquid precursor phase (DLP). Polycarboxylates facilitate pronounced, kinetic bicarbonate entrapment in the DLP. Since bicarbonate is destabilized in the solid state, DLP dehydration towards solid ACC necessitates the formation of locally calcium deficient sites, thereby inhibiting nucleation. Magic-angle spinning (MAS) nuclear magnetic resonance (NMR) spectroscopy of poly-aspartate-stabilized ACC reveals the presence of two distinct environments. The first contains immobile calcium and carbonate ions and structural water molecules, undergoing restricted, anisotropic motion. In the second environment, water molecules undergo slow, but isotropic motion. Indeed, conductive atomic force microscopy (C-AFM) reveals that ACC conducts electrical current, strongly suggesting that the mobile environment pervades the bulk of ACC, with dissolved hydroxide ions constituting the charge carriers. We propose that the distinct environments arise from colloidally stabilized interfaces of DLP nanodroplets, consistent with the pre-nucleation cluster (PNC) pathway.

Fig. 1. Potentiometric titration experiments in the presence of polymer additives.

a pH dependency of the scale factor (crystallization time relative to reference) for experiments containing 10 mg/L PAsp (red) and PGlu (blue). b Amount of NaOH added in titration experiments at pH 9.0 containing 10 mg/L polymer. The dotted lines visualize the slope of added NaOH in the prenucleation regime close to nucleation. In experiments with polymer, the extent of NaOH addition was significantly lower (see inset for easier comparison of slopes). c Development of molar amount of bound CO₃²⁻ per added Ca²⁺ in the prenucleation regime for experiments containing 0.1 g/L (PAsp and PGlu) or 0.01 g/L (PAA) polymer at pH 9.8. Values are calculated from the Ca-ISE (green) by assuming a 1:1 Ca²⁺:CO₃²⁻ binding ratio and from the NaOH addition (black) by calculating the amount of bound CO₃^2- from changes in buffer equilibria. d From the values shown in (c), the proportion of bicarbonate binding relative to the total amount of bound carbonate species (CO₃²⁻ + HCO₃⁻) can be calculated (black). Error bars represent ±1 − σ-standard deviation. The scale factors for each experiment are also shown (orange bars). The calculations are described in detail in Section 2 in the Supplementary Information.

Fig. 2. Characterization of isolated polymer stabilized ACC.

The sample was isolated from a titration experiment using 0.1 g/L PAsp at pH 9.8 by quenching the solution in ethanol (see Methods section). a ¹³C direct excitation (DE) and ¹H–¹³C cross-polarization (CP) spectra of 10% ¹³C-carbonate ACC stabilized by PAsp (PAsp_disACC) at a spinning frequency of 10 kHz. The spectra are scaled at the C_α-peak of PAsp. b TGA (red) and DSC (blue) analysis. The exothermic decomposition of the bicarbonate species is highlighted in grey. c ATR-FTIR spectra of polymer-stabilized ACC sample, showing significant amounts of polymer incorporation. Pure ACC and PAsp calcium salt (PAsp_Ca) are shown as references (detailed FTIR spectra are shown in Supplementary Fig. 6). d Normalized QMID for TGA-MS measurement on the PAsp_ACC sample using ¹³C enriched carbonates in the titrations. Due to the natural abundance of carbonate distribution in the polymer, released gases from polymer (¹²CO₂; m/z = 44, black) and from mineral (¹³CO₂; m/z = 45, red) can be distinguished, showing significant amounts of mineral decomposition below 300 °C (highlighted in grey). e TGA-IR analysis of the ¹³C carbonate enriched PAsp_ACC sample confirms the strong ¹³CO₂ release from (bi)carbonate species at around 300 °C.

Fig. 3. ¹H MAS NMR spectra of PAsp-stabilized ACC.

a Directly and ¹³C-carbonate detected ¹H MAS NMR spectra of PAsp-stabilized, 10% ¹³C-carbonate ACC (PAsp_disACC), with accompanying simulations, at a spinning frequency of 10 kHz and room temperature. The ¹Hs from PAsp do not significantly alter the shape of the directly detected spectra, see Supplementary Fig. 12. The MAS NMR probe, however, gives rise to a broad ¹H NMR background signal, which has been removed from the directly detected ¹H spectrum (black) following a procedure outlined in the Supplementary information (Supplementary Fig. 13). b Central region of the directly detected ¹H NMR spectrum of 100% ¹³C-carbonate ACC stabilized by PAsp (PAsp_disACC), at a spinning frequency of 10 kHz and room temperature.

Fig. 4. Conductivity measurements of ACC samples using C-AFM.

a AFM amplitude map recorded in non-contact mode (NCM) during the investigation of a polymer stabilized ACC sample (PAsp_ACC). Individual ACC nanoparticles are visible. b Results of C-AFM spectroscopy measurements of polymer-stabilized ACC of the area as shown in (a). The slope of the I/V diagram close to the origin is shown versus the Z-height of the respective measurement point. The grey part shows the standard deviation of 100 measurements on the gold wafer and is a measure for the highest conductivity that can be measured using this approach. c NCM amplitude map for a different area of the same polymer-stabilized ACC sample showing a large structure of several µm in diameter and d corresponding C-AFM results, showing good conductivity across a measurement distance of several 100 nm (more details are shown in Supplementary Fig. 24). e NCM amplitude map for a polymer-free ACC sample and f corresponding C-AFM results, showing good conductivity up to a Z-height of 100 nm (more details are shown in Supplementary Fig. 23).

Fig. 5. Cartoon depicting the proposed formation mechanism of distinct water environments present in ACC (not to scale).

Solute prenucleation clusters (PNC) undergo phase separation and form nanodroplets of a dense liquid phase (DLP) of the mineral. The polycarboxylate binds to calcium ions on their surface and colloidally stabilizes them. Eventually, the nanodroplets aggregate to reduce their interfacial free energy, and the species present on the surface of the droplets, as well as counter-ions present in the mother solution (mostly bicarbonate, and also hydroxide ions at pH > 9, shown in blue), are entrapped in the growing liquid-like mineral precursor via interface internalization (green areas). Due to the high polymer concentration and the different chemical environments on the surface of the DLP droplets, the distinct chemical environments will remain in the growing DLP and be transferred into solid ACC upon dehydration and solidification. As shown by MAS NMR, one of these environments is rigid and allows only restricted, anisotropic motion of the water molecules. We attribute this environment to the bulk of the original DLP nanodroplets. The other environment remains from the imperfect coalescence of the DLP nanodroplets and hence consists of water molecules undergoing isotropic motion and kinetically entrapped hydroxide ions, which form a network across the mineral precursor.