* Corresponding author. Tel.: #[phone]; fax: #[phone]-

1138.

E-mail address: [e-mail] (L. Cle`ries).

Biomaterials 21 (2000) 1861}1865

Behavior in simulated body #uid of calcium phosphate coatings

obtained by laser ablation

L. Cle`ries*, J.M. FernaHndez-Pradas, J.L. Morenza

Departament de Fn

&sica Aplicada i O"ptica, Universitat de Barcelona, Avda. Diagonal 647, E-08028 Barcelona, Spain

Received 28 July 1999; accepted 20 February 2000

Abstract

Three types of calcium phosphate coatings onto titanium alloy substrates, deposited by the laser ablation technique, were immersed

in a simulated body #uid in order to determine their behavior in conditions similar to the human blood plasma. Neither the
hydroxyapatite coating nor the amorphous calcium phosphate coating do dissolve and the

a-tricalcium phosphate phase of

the coating of

b-tricalcium phosphate with minor a phase slightly dissolves. Precipitation of an apatitic phase is favored onto the

hydroxyapatite coating and onto the coating of

b-tricalcium phosphate with minor a phase. Onto the titanium alloy substrate

reference there is also precipitation but at larger induction times. However, onto the amorphous calcium phosphate coating no
precipitate is formed.

2000 Elsevier Science Ltd. All rights reserved.

Keywords: Calcium phosphate; HA; Pulsed laser deposition; SBF

1. Introduction

Laser ablation is a technique employed for the depo-

sition of calcium phosphate coatings onto metallic
substrates that will be used as implants for bone recon-
struction. With this technique, calcium phosphate
coatings with tailored phases and structures have suc-
cessfully been produced [1,2] and their dissolution
properties in undersaturated conditions have been
assessed [3,4].

However, the real body #uid conditions are saturated

with respect to the hydroxyapatite phase, that is, the
concentration of calcium ions is higher than the one in
equilibrium with this phase. Consequently, it is interest-
ing to also test the calcium phosphate coatings in condi-
tions closer to the in vivo situation, in order to know
their integrity in these conditions and the catalytic}react-
ive properties of their surfaces towards precipitation pro-
cesses.

Therefore, amorphous calcium phosphate coatings

(ACP), hydroxyapatite coatings (HA), and coatings of
b-tricalcium phosphate with minor a phase (ba-TCP)
deposited by laser ablation were immersed in a saturated

solution for di!erent time periods and the evolution of
their constitutive properties was determined. The
saturated solution used was the simulated body #uid
(SBF), a solution whose ion concentrations and pH are
almost equal to those of human blood plasma [5]. This
solution is also the one utilized in the biomimetic (pre-
cipitation) process for the production of apatite layers
onto the sol}gel activated titanium substrates [6].

2. Experimental

The simulated body #uid [5] was prepared by dissolv-

ing reagent grade chemicals strictly in the following or-
der: NaCl, NaHCO, KCl, KHPO ) 3HO, MgCl )

6HO, CaCl ) 2HO, and NaSO into deionized

water. The #uid was bu!ered at pH"7.4 at 373C with
tris-hydroxymethyl-aminomethane

and

hydrochloric

acid. The inorganic composition of SBF emulates that of
human blood plasma, as shown in Table 1. No precipita-
tion was observed during #uid preparation.

Coatings of HA,

ba-TCP, and ACP deposited onto

titanium alloy (Ti}6Al}4V) substrates by laser ablation
of HA with an excimer laser at 248 nm and bare titanium
alloy substrates degreased in ultrasonic baths with
triclorethylene, acetone and ethanol, were used. Details
on the preparation and characterization of the coatings

0142-9612/00/$ - see front matter

2000 Elsevier Science Ltd. All rights reserved.

PII: S 0 1 4 2 - 9 6 1 2 ( 0 0 ) 0 0 0 6 0 - 0

Table 1
Milimolar concentrations (m

M

) of the ions of the SBF solution and comparison to those of human plasma [5]

Na

>

K

>

Ca

>

Mg

>

Cl

\

HCO

\

HPO

\

SO

\

SBF

142.0

5.0

2.5

1.5

147.8

4.2

1.0

0.5

Human plasma

142.0

5.0

2.5

1.5

103.0

27.0

1.0

0.5

Bu!er (TRIS solution): tris-hydroxymethyl-aminomethane (CHOH)CNH 50 m

M

#

HCl 45 m

M

.

Fig. 1. Scanning electron micrographs for (a) the HA coating, (b) the

ba-TCP coating, (c) the ACP coating, after 8 days immersion, and (d) the titanium

alloy substrate after 28 days immersion.

selected for this study, can be found in previous papers
[1}4]. All the coatings had a thickness around 1

lm.

Each sample, with an area of 1 cm

, was soaked in 24 ml

of the SBF solution into separate stopped vials at 373C in
a thermostatic oven, for di!erent periods: 1, 4, 8, and 28
days. At that point, each representative sample was care-
fully washed with distilled water, air dried, weighted and
characterized by X-ray di!raction, Raman spectroscopy
and scanning electron microscopy (SEM).

3. Results

3.1. HA coating

The SEM micrograph after 8 days immersion, depicted

in Fig. 1a, shows that over the columnar coating struc-
ture, a quite dense precipitated layer appears. This layer
is not strongly adhered to the underlying HA coating
since it cleanly detaches from the HA coating in certain

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L. Cle

% ries et al. / Biomaterials 21 (2000) 1861}1865

Fig. 2. X-ray di!raction spectra for (a) the HA coating, (b) the

ba-TCP coating, (c) the ACP coating, before and after 8 days immersion, and

(d) the titanium alloy substrate before and after 28 days immersion.

areas, underexposing the original HA surface which does
not seem to have degraded. The X-ray di!raction spectra
before and after 8 days in SBF are shown in Fig. 2a. The
initial spectrum contains, apart from the substrate peaks,
only HA peaks that do not diminish after 8 days in
solution. It is distinguishable at day 8 that the apparition
of a broad band centered around 323 superimposed to
the HA peaks, with an additional peak at 25.93, which are
attributed to a non-well crystallized apatite [7] with
a preferred (0 0 2) orientation. A representative Raman
spectrum after 8 days (Fig. 3a) shows the 962 cm

\ peak

of the HA coating and a relatively small, almost indis-
tinct, shoulder towards lower wavenumbers that is also
attributable to a non-well crystallized apatite structure
[8]. The gains in mass of the coating-substrate system,
tabulated in Table 2, indicate the growth of the precipi-
tate with time.

3.2.

ba-TCP coating

The SEM micrograph in Fig. 1b shows that there is the

formation of a thick precipitate at day 8. There are some
parts where this precipitate has detached, underexposing
the coating surface, which does not seem to be that

degraded. The X-ray di!raction peaks attributed to the
minor

a-TCP slightly diminish after immersion and those

of

b-TCP do not change (Fig. 2b). Additionally, the

broad band centered around 32 and the 25.93 peak,
attributed to a non-well crystallized apatite structure
with a preferred (0 0 2) orientation, appear. The represen-
tative "nal Raman spectrum (Fig. 3b) shows a broad
band that contains the TCP peaks at 947 and 972 cm

\,

but the existence of an intermediate peak at around
960 cm

\ suggests also the presence of a non-well

crystallized apatite structure. The mass changes for the
4 and 8 day period, which are also tabulated in Table 2,
are similar, indicating the early saturation of the precipi-
tation process.

3.3. ACP coating

In the SEM image (Fig. 1c) no substantial change is

detected, neither dissolution nor precipitation, since the
original morphology with droplets is maintained. The
ACP coating does not show any X-ray di!raction peak
other than the substrate ones after 8 days immersion
(Fig. 2c). The representative "nal Raman spectrum
(Fig. 3c) has only a broad band which is centered towards

L. Cle

% ries et al. / Biomaterials 21 (2000) 1861}1865

1863

Fig. 3. Raman spectra for (a) the HA coating, (b) the

ba-TCP coating,

(c) the ACP coating, after 8 days immersion, and (d) the titanium alloy
substrate after 28 days immersion.

Table 2
Mass variations in mg/cm

and changes observed with the character-

ization techniques

1 day

4 days

8 days

28 days

SEM

XRD

HA

0.41

Ppte

No

1.11

Ppte

NCA

ba-TCP

1.17

Ppte

NCA#

a

1.19

Ppte

NCA#

a

ACP

!

0.03

No

No

!

0.03

No

No

!

0.01

No

No

Titanium

0.23

Ppte

*

alloy

0.85

Ppte

NCA

No: non-detected change; NCA: non-well crystallized apatite struc-

ture;

a : decrease in a-TCP content; Ppte: precipitate observed.

950 cm

\, characteristic of an amorphous structure [6].

Table 2 con"rms that there is no uptake or loss of mass.

3.4. Bare titanium alloy substrate

The SEM image shows the precipitate found after 28

days (Fig. 1d). The X-ray di!raction spectrum (Fig. 2d)
indicates that it has also a non well crystallized apatite
structure with a (0 0 2) preferred orientation. A broad
band centered towards 960 cm

\ in its representative

"nal Raman spectrum (Fig. 3d) also suggests that the

precipitate has a non well crystallized apatite structure.
Table 2 shows the mass uptake for this sample.

4. Discussion

In a previous study [3] the same type of calcium

phosphate coatings used in the present work were tested
in Ca-free undersaturated conditions, and it was found
that the HA coatings were stable, that the

a-TCP phase

in the

ba-TCP coatings completely dissolved leaving

a microporous coating and that the ACP coatings com-
pletely dissolved.

In the saturated conditions of the SBF neither the HA or

the

b-TCP phase of the ba-TCP coatings do dissolve and

on both coatings a precipitate is found. The fact that these
phases do not dissolve is not an unexpected result if we take
into account that this solution is saturated for the di!erent
calcium phosphates, with di!erent degrees of supersatura-
tion represented by di!erent negative Gibbs free energies
(for instance, !7.94 kJ/mol for HA and !3.72 kJ/mol for
b-TCP [9]). The precipitate that is formed has a non-well
crystallized apatite structure with a (0 0 2) preferred orienta-
tion. Indeed, apatite is the most thermodinamically favored
phase to precipitate (has the highest negative Gibbs free
energy value), and the preferred (0 0 2) orientation is
a common "nding among other studies [10,11].

We have found that the precipitation rate is in the

order

ba-TCP 'HA'Ti' ACP. The precipitate is

then formed faster onto the

ba-TCP coating than onto

the HA coating. Radin et al. [12] reported that in a SBF
solution the

a-TCP phase dissolved during an induction

time before precipitation took place and suggested that it
was this initial phase dissolution leading to supersatura-
tion that consequently helped precipitation. Weng et al.
[13] have also suggested that the HA crystalline struc-
ture is not critical in the nucleation process. Following
these interpretations, the dissolution of the

a-TCP phase

of the

ba-TCP coating could favor the precipitation of

the non-well crystallized apatite phase on this coating
while for the HA coating the absence of dissolution
would delay this precipitation.

Onto the bare titanium alloy the precipitation of the

non-well crystallized apatite phase is observed although
the induction time towards precipitation is larger than
for the HA coating, and is similar to those reported in the
literature [14]. Ducheyne et al. [15] have also reported
that precipitation in a SBF solution occurs earlier onto
HA than onto titanium disks. Taking into account that
neither the HA coating not the titanium alloy do release
into the solution the calcium or phosphate ions [3,4] that
could help precipitation, this would suggest that in this
case the crystalline structure is indeed important in dic-
tating the di!erences in the rate of nucleation.

Remarkably, there is no dissolution of the ACP coat-

ing nor precipitation processes onto its surface. Contra-
dictory reports have been found regarding the behavior
in a SBF solution of ACP coatings obtained by RF
magnetron sputtering: Wolke et al. [16] reported that
these ACP coatings during immersion were maintained

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L. Cle

% ries et al. / Biomaterials 21 (2000) 1861}1865

and in some areas a calcium phosphate precipitate was
found. However, Yoshinari et al. [17] observed dissolu-
tion of ACP coated disks within one day without precipi-
tation of a calcium phosphate phase. They attributed this
di!erence in precipitation processes to the variations on
the ratio of solution volume to sample area, the Ca/P
ratio, the grain size of the coating, and the impurities. The
absence of dissolution for our ACP coating could
be explained on the basis that the SBF solution could be
saturated with respect to this particular ACP phase. This
degree of saturation cannot be easily calculated for
amorphous calcium phosphate phases since it largely
depends on their Ca/P ratio which can #uctuate. There is
no precipitation onto the ACP coating even when onto
the titanium alloy substrate a precipitate is found. This is
quite surprising since, in the absence of dissolution, the
ACP coating should provide a better substrate for the
nucleation of apatite compared to the titanium alloy
substrate since it already has calcium and phosphate ions
in its structure. Therefore, other factors intervening in the
mechanisms of the formation of this precipitate, apart
from the existence of a calcium phosphate surface or
calcium and phosphate release, should be considered.

5. Conclusion

The HA coating deposited by excimer laser ablation

con"rms its stability in saturated conditions towards
dissolution. The ACP coating and the

b-TCP phase of

the

ba-TCP coating are also stable under these condi-

tions. The HA and

ba-TCP coatings favor earlier the

precipitation of an apatitic phase with a (0 0 2) preferred
orientation. Onto the titanium alloy substrate reference
there is also precipitation but at larger induction times.
However, onto the amorphous calcium phosphate coat-
ing no precipitate is formed.

Acknowledgements

This work is part of a research program "nanced by

DGESIC of the Spanish Government (Project MAT98-
0334-C02-01) and DGR of the Catalan Government.

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