Abstract

       Organic/inorganic bulk nanocomposites of poly(ethyl methacrylate-co-hydroxyethyl acrylate) 70/30 wt%/silica, P(EMA-co-HEA)/SiO2, were obtained with varying proportions of silica up to 30 wt%. The synthesis procedure consisted in the copolymerization of the organic monomers EMA and HEA during the simultaneous acid-catalyzed sol-gel polymerization of tetraethoxysilane, TEOS, as silica precursor.
       The structure of SiO2 in the P(EMA-co-HEA) polymeric matrix was inferred from infrared spectroscopy, energy dispersive X-ray spectroscopy, thermogravimetry, pyrolysis, transmission electron microscopy and swelling measurements. TEOS was efficiently hydrolyzed and condensed to silica during the sol-gel process, and presented a homogeneous distribution in the polymeric matrix, in the form of disconnected aggregates of elementary silica nanoparticles in the hybrids with low silica contents (below 10 wt%) or continuous network interpenetrated with the organic network after coalescence of these silica aggregates (above 10 wt%). The small size of the silica aggregates explained the optical transparency of the obtained nanohybrids. The organic polymeric network occurred in the pores left within the elementary silica nanoparticles produced by the liquids acting as a template in the sol-gel polymerization, and those left between the aggregates of silica nanoparticles. Heterocondensation reactions between silica and the organic copolymer, if any, were negligible.
       The physico-chemical properties of the hybrids were studied by density assessments and differential scanning calorimetry, the surfaces were featured by contact angle measurements, and the mechanical properties were analyzed by dynamic-mechanical analyses and compression tests. Intermediate silica contents (10-20 wt%) exhibited the most interesting balanced properties: i) mechanical reinforcement of the glassy organic matrix attained by continuous interpenetrated silica networks, ii) good swelling ability due to the organic network expansion still not hindered by a rigid silica skeleton and to the relatively high number of hydrophilic boundary silanol groups (inorganic concentrations close to coalescence), and iii) enhanced surface reactivity or hydrophilicity due to the still elevated relative content of polar silanol terminal groups available at the surfaces.
       The ‘bioactivity’ or ability of the bulk samples to form hydroxyapatite (HAp) on their surfaces was tested in vitro by soaking them in a simulated body fluid (SBF) for different times up to 35 days, the ion concentrations, temperature and pH of which were adjusted to almost equal those of human blood plasma. After 7 days, the SBF solution was changed to a 2xSBF with periodical renovation, providing more favourable conditions for apatite coating. The composition and morphology of the apatite formed and the structural changes taking place in the nanohybrids when immersed in SBF were analyzed. The formation of the apatite layer was controlled by the mechanism and induction time of the nucleation, which depended in turn on the structure of silica. In the nanohybrids with intermediate silica contents, the dissolution of silica at the surface was facilitated by the relatively large number of silanol groups, rendering an interface layer rich in silanol groups bounding a reaction zone depleted of silica. In this reaction zone calcium and phosphate ions were adsorbed and interacted with the polar silanol groups to form calcium phosphates. The apatite growth continued at the interface hybrid-solution leading to a strongly adhered apatite layer. After 5 days, the growth of the apatite layer and formation of successive ones occurred very rapidly, consuming calcium and phosphate ions from the SBF. The initially amorphous calcium phosphate, containing other ions such as CO32-, Na+, K+ or Mg2+, stabilized leading to low-crystalline needle-shaped polycrystals of calcium-deficient carbonated-hydroxyapatite, with Ca/P ratios close to the physiological HAp ratio.
       A surface modification treatment was applied with the purpose of reducing the induction time for apatite nucleation, combining a NaOH attack to increase the number of surface nucleating sites, and an alternate soaking process in Ca and P solutions to form apatite precursors prior to the immersion in SBF (CaP treatment). The NaOH treatment was not effective by itself in shortening the HAp nucleation induction time. It introduced sodium carboxylates in the copolymer but hydrolyzed the silica network excessively, thus eliminating the silanols nucleating potential. Bioactivity was only due to the carboxylate groups of the organic phase. Maybe a suitable dissolution extent of the silica network so as to improve bioactivity could be attained by controlling the duration of the NaOH treatment. This would be interesting in the nanohybrids with superpercolating silica concentrations, where the density and continuity of the silica network with fewer silanol terminal groups hinder the polymer network swelling, the silica hydrolysis and the diffusion of ions from the SBF. Thus, at these silica concentrations, the bioactivity of the nanohybrids is only due to the preexisting HAp nucleating functional groups available at the surface. The posterior CaP treatment was able to coat the surfaces of the different samples with a calcium phosphate layer within minutes. These amorphous calcium phosphates acted as HAp precursors, skipping the induction period in SBF.
       The simplicity of the synthesis procedure offers the possibility of controlling the architecture of the obtained nanocomposites. Nanohybrid tubular pores scaffolds with silica contents up to 20 wt% could be prepared by polymerization in a fibres template, aiming to mimic the structure and properties of the extracellular mineralized matrix of natural dentin. Although some synthetic scaffolds have been proposed as alternative tooth therapies to induce the regeneration of dentin/pulp tissues, this is, up to our knowledge, the first time a bioactive synthetic scaffold mimicking natural dentin is proposed. Intermediate percentages of silica resulted in scaffolds with mechanical properties compatible with the application while conferred bioactivity to the surfaces. This is expected to facilitate the integration in the host mineralized tissue and stimulate the differentiation of pulpal cells and the invasion of the tubules by new odontoblast prolongations when implanted in vivo, thereby guiding dentin tissue regeneration.

Keywords: Nanocomposite, hybrid, hard tissue repair, ethyl methacrylate, hydroxyethyl acrylate, tetraethyl orthosilicate, silica, sol-gel, bioactive, hydroxyapatite (HAp), simulated body fluid (SBF), scaffold, dentin.