Effect of sodium polyacrylate on the hydrolysis of octacalcium phosphate

doi:10.1016/S0162-0134(00)00015-5

Journal of Inorganic Biochemistry

Volume 78, Issue 3, 29 February 2000, Pages 227-233

https://doi.org/10.1016/S0162-0134(00)00015-5 Get rights and content

Abstract

Octacalcium phosphate (OCP) hydrolysis into hydroxyapatite (HA) has been investigated in aqueous solutions at different concentrations of sodium polyacrylate (NaPA). In the absence of the polyelectrolyte, OCP undergoes a complete transformation into HA in 48 h . The hydrolysis is inhibited by the polymer, which is significantly adsorbed on the crystals, up to about 22 wt.%. A polymer concentration of 10⁻² mM is sufficient to cause a partial inhibition of OCP to HA transformation, which is completely hindered at higher concentrations. The small platelet-like crystals in the TEM images of partially converted OCP can display electron diffraction patterns characteristic either of OCP single crystals or of polycrystalline HA, whereas the much bigger plate-like crystals exhibit diffraction patterns characteristic of OCP single crystals. The polyelectrolyte adsorption on OCP crystals is accompanied by an increase of their mean length and by a significant reduction of the coherence length of the perfect crystalline domains along the c-axis direction. It is suggested that the carboxylate-rich polyelectrolyte is adsorbed on the hydrated layer of the OCP (100) face, thus inhibiting its in situ hydrolysis into HA.

Introduction

The inorganic phase deposited in mineralized tissues, such as bone and dentine, is a poor crystalline carbonated apatite. However, it has been proposed that the mechanism of biological apatite formation involves octacalcium phosphate (Ca₈H₂(PO₄)₆·5H₂O, OCP) as a precursor phase [1], [2]. OCP triclinic crystals consist of alternating hydrated and apatitic layers, the latter closely resembling hydroxyapatite (Ca₁₀(PO₄)₆(OH)₂, HA) [3]. OCP has a greater solubility than the thermodynamically more stable HA, and it undergoes a spontaneous and topotactic transformation into HA [4], [5]. The involvement of OCP in the first stages of crystal growth of bone and tooth tissues has been invoked to explain the lack of HA crystals having hexagonal symmetry in the initial stages of biomineralization [6], as well as the crystal morphology of bone and dentine, which is plate or ribbon like, with the thin plates elongated along the crystallographic c-axis [7], [8]. Furthermore, OCP has been found in dental calculi [9], [10], as well as in other pathological calcifications [11]. However, just recently a high resolution transmission electron microscopy study of calcifying dentine reported the first observation of OCP in biological crystals [12]. The presence of OCP in the central part and of HA at the extremities of the same crystal was interpreted as suggesting that the hydrolysis of OCP in the formation of biological crystals develops along the (100) crystal planes. The matrix macromolecules of the vertebrate mineralized tissues contain acidic proteins and glycoproteins, rich in acidic amino acids with charged carboxylate groups [13]. Other negatively charged groups, such as phosphates and sulphates, are often present. These proteins may affect crystal induction and crystal growth regulation, possibly through interactions with the charged crystal surfaces [14]. The results of in vitro experiments carried out on OCP crystallization suggest a specific interaction of the highly phosphorylated acidic protein, phosphophoryn, with the (010) face, whereas carboxylate-rich proteins seem to interact preferentially with the hydrated layer of the (100) face [14]. We have recently found that sodium polyacrylate (NaPA), a carboxylate-rich polyelectrolyte, inhibits the nucleation and growth of synthetic OCP crystals [15]. The results of the structural and morphological investigation suggest a non-specific interaction with the polyelectrolyte, which is not significantly adsorbed on OCP crystals. On the other hand, preliminary data indicate that NaPA can be adsorbed on OCP from aqueous solutions and prevent its hydrolysis into HA [15]. In order to investigate the structural interaction of the polyelectrolyte with OCP, we have carried out a structural and morphological investigation on OCP submitted to hydrolysis in aqueous solutions at different NaPA concentrations.

Section snippets

Materials and methods

Octacalcium phosphate was synthesized by dropwise addition of 250 mL of 0.04 M Ca(CH₃COO)₂ (Carlo Erba) into 750 mL of a phosphate solution containing 5 mmol of Na₂HPO₄ (Carlo Erba) and 5 mmol of NaH₂PO₄ (Carlo Erba) maintained at 60 °C, starting pH 5 [16]. The precipitate was stored in contact with the mother solution for 10 min, filtered, repeatedly washed with bidistilled water and dried at 37 °C.

Hydrolysis of OCP was carried out in distilled water and in solutions at different NaPA

Results

The most frequent morphology exhibited by the crystal aggregates present in the SEM micrographs of the synthesized OCP is that of long blades radiating from a common origin to form spherulitic growths, as that reported in Fig. 1(a). The TEM micrographs of the starting OCP show long plate-like crystals with straight edges, whose mean dimensions are 2×8 μm (Fig. 1(b)). The thickness of the crystals, which can only be measured in a few cases, is around 0.3–0.4 μm. Before treatment in aqueous

Discussion

The hydrolysis of OCP to HA in aqueous solutions is a well-known transformation, which depends on several parameters, such as temperature and ionic composition of the solution [2], [21], [22].

The results of this paper indicate that a concentration of NaPA of 10⁻¹ mM prevents the hydrolysis of OCP to HA in aqueous solution at 60 °C. OCP is partially converted into HA after 48 h of storage in 10⁻² mM polyelectrolyte solution, whereas complete transformation can be obtained when the concentration

Abbreviations

OCP	octacalcium phosphate
HA	hydroxyapatite
NaPA	sodium polyacrylate
SEM	scanning electron microscopy
TEM	transmission electron microscopy
ED	electron diffraction

Acknowledgements

This research was carried out with the financial support of MURST, CNR (PF MSTA II) and the University of Bologna (Funds for Selected Research Topics).

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Effect of sodium polyacrylate on the hydrolysis of octacalcium phosphate

Abstract

Introduction

Section snippets

Materials and methods

Results

Discussion

Abbreviations

Acknowledgements

Nature

J. Cryst. Growth

J. Phys. Chem.

J. Cryst. Growth

Anat. Rec.

Calcif. Tissue Res.

J. Cryst. Growth

J. Dent. Res.

Scanning Microsc.

Calcif. Tissue Int.

Acta Crystallogr., Sect. D