potentially lead to the formation of non-apatitic calcium phosphate
phases or salts other than calcium phosphates.46 The
Ca:F ratio of that method was calculated to be about 5:1, which is the
appropriate stoichiometry for FAp.34 The incubation of
different substrates inside the fluoride-rich supersaturated calcium
phosphate solutions, are thought to drive the occurrence of
heterogeneous nucleation on the surfaces, which is known to take place
faster than the homogenous nucleation, as the substrates decrease the
energy required to form a nucleus.47,48 The
morphology, coverage, and crystal sizes of apatite coatings precipitated
on the substrates, is known to be dependent on the substrate’s
chemistry, topography, charge, grain size, lattice geometry, and
architecture.47,49 Concave surfaces usually have high
spatial charge, enhance the assembling of ions in three dimensions, thus
they are considered as good nucleation sites. While convex surfaces
dissipate the charge, which then decrease the
nucleation.47 Moreover, planar surfaces were also
known to act as two dimensions nucleation sites with equal distribution
of charge depending on the substrates’ isoelectric point itself rather
than its topographical properties.47
After mineralization, we have observed multiple organized structures
made of apatite nanocrystals. For example, dandelion shaped nanocrystals
(Chart 1c) were seen on all the substrates at 37◦C and 70◦C (Figs. 4a,
b, d, e, g, h, j, and k). As well, bowknot-like structures made of
assembled nanocrystals (Chart 1d), were observed on NE-Ti at 37◦C on the
E-Ti disc at 70◦C (Figure 4a, e). Furthermore, smooth ball-like
structures either with spikes (Chart 1a, Figs 4a, b, d, e, g, j, h, and
k) or without spikes (Chart 1b, Figs. 4a, b, d, e, g, j, h, and k) were
observed on all substrates, except on the NE-Ti, E-Ti, P-Sa at 90◦C. On
the other hand, disordered nanocrystals (Chart 1e) were detected on all
substrates at 90◦C except the U-Sa (Figure 4c, f, i, and l).
The difference in the morphology between the ball-like structures with
and without the spiky features, is potentially due to the outward growth
of nanocrystals from inside the ball-like structures. Kinetically, it is
proposed that the ball-like structures with the spiky features, can
transform at later stages into bowknots. This then leads to the
morphological transformation into the dandelion structures. Therefore,
those morphologies which are found mainly at 37ᵒC could be tested as
potential biocompatible material at the bone-implant interface.
Activation energy for nucleation is known to be dependent on the
nucleus-substrate interactions. Crystal growth is favoured at low
energies, as it decreases the overall free energy of the critical
nucleus formation, thus promoting the sequential phase
transformations.50 In our scenario, spherical
structures (Chart1a, b, f) were mostly seen at lower temperature as
these structures are energetically favoured with less energy needed for
their formation. With increasing the temperature to 70ᵒC, self-assembled
organized nanocrystals start to grow out of the ball-like structures
(Chart 1c,d), with further formation of ordered apatite features.
However, at high temperatures (90ᵒC), the nanocrystals seem to lack the
hierarchical organization and appeared to be arranged randomly.
Effect of Substrate and Temperature on the Coverage of the
Apatite Coatings
Fujishiro et al.51 reported deposited HAp on various
substrates; they found that the coverage, morphology, and adhesive
strength of the coatings. For example, they found that the coverage
changes considerably between different substrates, as titanium was
entirely coated compared to sparsely coated alumina substrate.
Furthermore, it is well known that at pH of 6, titanium substrate acts
as a negatively charged surface,52 while sapphire
substrate is either neutral or exhibits a slightly positive surface at
values of the pH below the isoelectric point, sapphire has a positive
charge.53 Zhu et al.54 observed that
the nucleation of HAp is favoured on negatively charged substrates
compared to the positively charged ones. Using image analyses, we
quantified the extent of the coating coverage on the different
substrates at 37ᵒC, 70ᵒC and 90ᵒC.
This quantification was conducted, to assess the effect of the different
surface properties including the surface chemistry and topography on the
coverage. The highest apatite coverage was found on P-Sa
37ᵒC and U-Sa at 70ᵒC with a
percentage of 45.5 ± 0.03% and 35±0.02% respectively. All other
substrates at different temperatures showed moderate coverage between
12-29%. The enhanced apatite coverage, which is seen on the E-Ti
surfaces, gives an indication that the apatite nucleation and growth is
favoured due to increased grooves and pits on the surface, which in turn
increase the nucleation sites.55 and overall coverage.
Holbrough et al.56, who confirmed that the surface
topography can control efficiently the nucleation sites and in turn the
coverage, stated that some cavities on the substrate surface could have
the same dimensions of a critical nucleus. Moreover, our results agree
with Dennig and Stevenson, and Campbell et al.57,58who stated that the nucleation events prefer rougher surfaces such as
scratches.