ARTICLE IN PRESS

Biomaterials 26 (2005) 509–517

Improvement in biocompatibility of ZrO2–Al2O3 nano-composite

by addition of HA
Young-Min Kong, Chang-Jun Bae, Su-Hee Lee, Hae-Won Kim, Hyoun-Ee Kim*
School of Materials Science and Engineering, Seoul National University, Seoul 151-744, South Korea
Received 22 November 2003; accepted 24 February 2004

Abstract

The biocompatibility of zirconia–alumina (ZA) nano-composites in load-bearing applications such as dental/orthopedic implants
was significantly enhanced by the addition of bioactive HA. The ZA matrix was composed of nano-composite powder obtained
from the Pechini process and had higher flexural strength than conventionally mixed zirconia–alumina composite. Because the ZA
nano-composite powder effectively decreased the contact area between HA and zirconia for their reaction during the sintering
process, the HA-added ZA nano-composites contained biphasic calcium phosphates (BCP) of HA/TCP and had higher flexural
strength than conventionally mixed ZA–HA composite. From the in vitro test with osteoblastic cell-lines, the proliferation and the
differentiation (as expressed by the alkaline phosphatase activity) of the cellular response on the HA-added ZA nano-composites
gradually increased as the amount of HA added increased. From the mechanical and biological evaluations of the HA-added ZA
nano-composites, 30HA (30 vol% HA+70 vol% ZA) was found to be the optimal composition for load-bearing biological
applications.
r 2004 Elsevier Ltd. All rights reserved.

Keywords: Hydroxyapatite (HA); Zirconia–Alumina (ZA); Nano-composite; In vitro; Alkaline phosphatase

1. Introduction of this approach is the susceptibility of the HA coating

Hydroxyapatite {HA; Ca10(PO4)6(OH)2}, a synthetic
analog of the calcified tissues of vertebrates, has often
been considered for use in load-bearing applications,
because HA bonds to bone directly and promotes the
new bone formation necessary for implant osseointegra-
tion [1–4]. However, its poor mechanical properties have
limited its use to non-load-bearing parts. Therefore, in
order for it to be used in load-bearing parts, the
mechanical properties (mainly the strength and fracture
toughness) of HA need to be improved [5–9].
There are two possible approaches to achieve this
goal; one approach is to place a coating layer on the
metallic implants, while the other involves the use of
composites. Metallic implants coated with HA, which
are widely used as load-bearing implants, are a good
example of the former approach. One of the limitations
*Corresponding author. Tel.: +82-2-880-7161; fax: +82-2-884-
1413.
E-mail address: kimhe@snu.ac.kr (H.-E. Kim).
to debond from the metallic substrate [10].
The latter approach can be subdivided into two
distinct strategies depending on whether an HA-based
composite or an HA-added composite is used. The
reinforcement of HA, which is accomplished by the
addition of other materials in the form of powders,
platelets, or fibers, belongs to the former strategy. In
studies involving the use of these materials, the
mechanical properties of the HA-based composites were
found to be enhanced by a factor of 3 (from 100 MPa,
1 MPam1/2 to 300 MPa, 3 MPam1/2) [8]; at the same
time, in vivo tests with screw-shaped dental implants
also showed that HA-based composites were superior to
those made with commercially pure Ti (two times higher
removal torque values) [11]. However, further improve-
ments are needed before such materials can be employed
for load-bearing applications such as dental/orthopedic
implants. The second strategy involves the improvement
of the biocompatibility of various strong materials
through the incorporation of bioactive materials. The
zirconia–alumina system (ZA, 80 wt% ZrO2–20 wt%
Al2O3) is well-known for its high flexural strength [12],

0142-9612/$ - see front matter r 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.biomaterials.2004.02.061
 ARTICLE IN PRESS
510 Y.-M. Kong et al. / Biomaterials 26 (2005) 509–517
and is classified as a bioinert ceramic [1]. Therefore, ZA
is considered to be one of the most suitable materials to
be used as a matrix for the HA-added strong composites.
This study focused on improving the biocompatibility
of strong ZA nano-composites through the addition of
bioactive HA. The strength and biocompatibility of the
composites were measured as a function of the HA
content and correlated with the compositional and
structural variations. This study verifies the potential
of HA-added ZA nano-composites to be used in load-
bearing applications.
2. Materials and methods
2.1. ZA nano-composite powder by Pechini process
The Pechini process is based on the formation of a
network which is composed of a polymeric precursor
and a cationic network modifier [13]. In the Pechini
process, the metallic ions are distributed homogeneously
on an atomic scale throughout the polymeric network.
Such a structure eliminates the need for long-range
diffusion during the subsequent formation of metal
oxides. Therefore, in conventional Pechini processes, the
precursor forms a homogeneous single phase oxide of
precise stoichiometry at a relatively low temperature. In
this experiment, we used Zr and Al as the metallic ions
in the Pechini process to make a nano-composite
powder. Since zirconium and aluminum are unlikely to
form a compound, the intimacy between the Zr and Al
ions captured in a polymeric network enables them to
form a nano-sized composite powder.
Metal chlorides (ZrCl2O
8H2O, AlCl3
6H2O,
YCl3
6H2O) were used as the cationic sources; citric
acid monohydrate (C6H8O7
H2O, CAM) and ethylene
glycol (C2H6O2, EG) were used as the polymeric matrix.
A stoichiometric mixture of Zr and Y source solution
(3 mol% Y2O3 doping in ZrO2) and Al source solution
were used as the starting materials. The compositional
ratio of ZrO2 to Al2O3 was 4:1 by weight, because this
composition showed the highest flexural strength [12].
After mixing the metallic source solutions with CAM-
EG solution, the resultant mixture was heated to 130 C
to promote esterification between CAM and EG. Finally,
the gel was dried, crushed and calcined at 800 C. The
obtained powders were ball-milled in ethanol for 48 h
with ZrO2 balls to reduce the secondary particle size.
The total polymer content in the powder precursor
was as high as 90 wt%. When 90% of polymer was
added to the solution, the particles began to coalesce so
as to form a nano-composite powder composed of ZA.
All of the particles were composed of 100–200 nm
clusters, which contained zirconia nano-crystallites
whose size was about 10 nm, as shown in Fig. 1(A)
and (B). Since a greater number of nucleation sites were
formed in the gel as the polymer content increased, the
zirconia gradually grew into nano-sized particles and
eventually became a nano-composite powder through
their merging with alumina.
2.2. Preparation and characterization of HA-added
composites
The starting powders were commercial HA (Alfa
Aesar Co., Ward Hill, MA, USA) and the above-
mentioned ZA nano-composite powder. In addition,
commercially available ZrO2 (3Y-TZP, Tosoh Co.,
Tokyo, Japan) and Al2O3 (AKP50, Sumitomo Co.,
Osaka, Japan) were used for the purpose of comparison.
The ZA nano-composite powders (composition of
80 wt% ZrO2–20 wt% Al2O3) were mixed with HA
(specific compositions of up to 40 vol%) by ball milling
for 24 h. After the mixed slurry was dried, crushed and
sieved, the resulting powders were hot-pressed in a
graphite mold at 1400 C with an applied pressure of
30 MPa for 1 h. The phase of the hot-pressed bodies was
analyzed with an X-ray diffractometer (M18XHF, MAC
Science, Yokohama, Japan), and the density of the
specimens was measured by the Archimedes method. The
fractured and polished surfaces were observed with a
SEM (JSM-6330F, JEOL, Tokyo, Japan). The flexural
strength was measured by means of the four-point flexure
test with a cross-head speed of 0.5 mm/min, and inner-
and outer-spans of 20 and 40 mm, respectively (number
of samples=7) [8]. The tensile surface of the specimens
was polished using diamond slurries (1 mm finish).
2.3. Biocompatibility
The specimens used for the cell tests (proliferation and
differentiation) were prepared from the hot-pressed
specimens, so as to have a square shape with dimensions
of 10 mm  10 mm  1 mm. After polishing both sides
with diamond slurries down to 1 mm, the specimens were
washed and sterilized at 121 C for 20 min.
(A) 50 nm (B) 50 nm
Fig. 1. TEM images of ZrO2–Al2O3 nano-composite powder; (A)
bright field image and (B) dark field image.
 ARTICLE IN PRESS
Y.-M. Kong et al. / Biomaterials 26 (2005) 509–517
In order to investigate the proliferation behavior on the
ZA–HA composites, MG63 cell lines were cultured in a
dish containing Dulbecco’s modified Eagle’s medium
(DMEM, Invitrogen Co., Carlsbad, CA, USA) supple-
mented with 10% fetal bovine serum (FBS, Invitrogen
Co., Carlsbad, CA, USA). The fabricated specimens,
including pure HA as a positive control, were placed into a
24-well plate, which was followed by plating 3  104 cells/
cm2 onto the discs. The cells were cultured for 7 days in a
humidified incubator under an atmosphere of 5% CO2 at
37 C. After detaching the cells from the specimens with a
0.05% trypsin-EDTA solution and staining with 0.4%
trypan blue, the living cells were counted using a
hemocytometer (Paul Marienfeld GmbH & Co., KG,
Lauda-K.onigshofen, Germany). Each set of tests was
performed in triplicate, and the data were normalized so
as to take the surface area into consideration. For the
SEM observations, the 5 day-cultured cells (1  104 cells/
cm2) were fixed with 2.5% glutaraldehyde, and then
dehydrated with graded ethanol (70%, 90%, and 100%).
In order to measure the differentiation level expressed
by the alkaline phosphatase (ALP) activity of the HOS
cell-line on the specimens, the cells were plated at a
density of 1  104 cells/cm2 in a 24-well plate, and then
cultured for 10 days. At harvest, the culture media were
decanted, and the cell layers were washed once with
HBSS, followed by trypsinization. After centrifugation
at 1200 rpm for 7 min, the cell pellets were washed with
PBS and resuspended by shaking them in 100 ml of 0.1%
Triton X-100. The pellets were disrupted by 5 cycles of
sequential freezing/thawing. After centrifugation at
13,000 rpm for 20 min at 4 C, the cell lysates were
quantified using the BioRad DC protein assay (Bio-Rad
Laboratories Inc., Hercules, CA, USA) and assayed
colorimetrically for ALP activity using p-nitrophenyl
phosphate as the substrate (Sigma, St. Louis. MO,
USA). Each reaction was initiated with p-nitrophenyl
phosphate and lasted for 10 min at 37 C, before being
stopped by quenching on ice. The p-nitrophenol
produced was measured at 410 nm using a spectro-
photometer (UV-1700, Shimadzu Co., Kyoto, Japan).
2.4. Statistical analysis
A one-way analysis of variance (ANOVA) was used
to compare the means of the different groups, and
statistical significance was accepted at the 0.05 con-
fidence level.
3. Results
3.1. Evaluation of HA-added ZA nanocomposites
Figs. 2(A)–(D) show the microstructures of the ZA
nano-composites containing different amounts of HA.
511
Fig. 2(A) shows the ZA nano-composite without HA, in
which dark alumina grains were homogeneously dis-
persed in a bright zirconia matrix. The homogeneous
microstructure and very small grain size (about 250 nm)
are well illustrated in this micrograph. When 10 vol%
HA was added, the microstructure was changed, as
shown in Fig. 2(B); the small dark grains of alumina
became coarser, and large dark grains (B1 mm) of HA
appeared in the microstructure. When 20 and 30 vol%
HA were added (Figs. 2(C) and (D)), the resulting
alumina grains had an acicular shape, while the HA
grains were plate-like in shape. The pores (about the
same size as the zirconia grains) were thought to be
formed during the polishing and thermal etching
processes.
Fig. 3 shows the densities of the ZA nano-composites
containing various amounts of HA. Compared to the
theoretical density, which was calculated by the rule of
mixture, the density of the specimens with a low HA
content almost matched with the theoretical value.
However, as the amount of HA was increased up to
40 vol%, the discrepancy between the measured and
theoretical densities increased.
The composition of the specimens was analyzed
by means of their XRD patterns. When no HA was
added, only peaks corresponding to ZrO2 and Al2O3
were detected, as shown in Fig. 4(A). With the
addition of 20% and 30% HA (Figs. 4(B) and (C),
respectively), as well as the peaks corresponding to HA,
peaks were also detected for TCP, which originated
from the reaction between HA and ZrO2. These XRD
patterns illustrate that sintering the composite at 1400oC
led to the appearance of BCP of HA/TCP in the
specimen.
The flexural strengths of the HA-added ZA nano-
composites are shown in Fig. 5. The strength of the pure
ZA nano-composite was 1460 MPa when it was hot-
pressed at 1400 C. When 5 vol% of HA was added to
the ZA nano-composite powder and it was hot-pressed
under the same conditions, the flexural strength
decreased to 1270 MPa. As the HA content was
increased up to 40 vol%, the flexural strength decreased
steadily (ANOVA, po0.01). The hot-pressed specimens
obtained from the conventionally mixed ZA powder
showed a similar trend in terms of their strength (open
circle in Fig. 5) with respect to the addition of
HA, however, their strengths were B than those of
B powders.
Fig. 6 shows the fracture surfaces of the specimens.
When the pure ZA nano-composite was hot-pressed
at 1400 C, the fracture proceeded mostly in an
intergranular pattern and the fracture surface was
very rough, as shown in Fig. 6(A). When 20 vol%
of HA was added to ZA nano-composite (Fig. 6(B)),
the fracture also occurred in an intergranular
mode. However, the fracture morphology was slightly
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512 Y.-M. Kong et al. / Biomaterials 26 (2005) 509–517

(A) (B)

HA

(C) (D)

HA
HA

Fig. 2. SEM images of ZA (80 wt%ZrO2–20 wt% Al2O3) composites; (A) without HA, (B) containing 10 vol% HA, (C) containing 20 vol% HA, and
(D) containing 30 vol% HA. All specimens were hot-pressed at 1400 C for 1 h, followed by thermal etching at 1300 C. The bright and dark grains in
the image belong to zirconia and alumina, respectively.

5.6
Z

(A) Z
A A
Z A
5.2
Intensity [Arb. Unit]
Density [g/cm ]
3

measured density theoretical density

4.8
(B)

4.4

4.0 (C)

0 20 40
25 30 35 40 45
HA content [vol%] 2θ [o]
Fig. 3. Density of ZA–HA composites hot-pressed at 1400 C as a Fig. 4. XRD data of the ZA composites; (A) without HA, (B)
function of the HA content. containing 10 vol% HA, and (C) containing 30 vol% HA. All
specimens were hot-pressed at 1400 C: () HA, (~) TCP, (A)
alumina, and (Z) zirconia.

different, due to the large size of the HA grains.
In the case of pure HA (hot-pressed at 1200 C),
the fracture morphology was much different from
that of the other specimens, with the fractures
mostly occurring a transgranular pattern, as shown in
Fig. 6(C).
3.2. Biological evaluations
SEM images of the MG63 cells which were cultured
for 5 days on various specimens are shown in Fig. 7; (A)
pure ZA nano-composite, (B) 10 vol% HA+90 vol%
ZA (10HA), (C) 30 vol% HA+70 vol% ZA (30HA),
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Y.-M. Kong et al. / Biomaterials 26 (2005) 509–517 513

and (D) pure HA. The cells on all specimens evolved
stellately and, in each case, the cells came into contact
with each other through the elongation of the cytosols.
There seemed to be no significant morphological
differences between the various ZA nano-composites
containing different amounts of HA.
The number of cells on each specimen after prolifera-
tion for 7 days was plotted, and the resulting graph is
shown in Fig. 8. The cells proliferated actively on all of
the specimens including on pure ZA. However, when
2000
(ANOVA, p < 0.01)
1600
Flexural Strength [MPa]
10% HA was added, the cells proliferated more actively
on the composite as compared to that on the pure ZA
(ANOVA, po0.05). As the HA content increased, the
proliferation rate increased steadily (ANOVA,
po0.001). Interestingly, when 40% HA was added to
the ZA, the proliferation rate on the corresponding
specimen was comparable with that on the pure HA
specimen.
Figure 9 shows the differentiation behavior expressed
in terms of the ALP activity of the HOS cell lines on the
specimens after 10 days culture. The ALP activity
exhibited a similar trend to that of the proliferation.
However, in contrast to the proliferation behavior, the
specimen containing 10% HA had a similar expression
level compared to the pure ZA (ANOVA, p=0.062).

The ALP expression level became significantly different

1200
from that of the pure ZA when 20% HA was added to
nano-composite powder the ZA (ANOVA, po0.05). The expression level
increased steadily as the amount of HA added was
800 increased (ANOVA, po0.005). As in the case of the
proliferation rate, the ALP expression level on the
mixed powder specimen containing 40% HA was comparable with that
400
on the pure HA (ANOVA, p>0.05).
0

0 20
HA content [vol%]
Fig. 5. Flexural strength of the composites containing various
amounts of HA. The error bars represent 71 standard deviations.
The mean values are significantly different (ANOVA, po0.01).
40 4. Discussion
As a bioinert implant material, ZA nano-composite is
considered to have sufficient flexural strength, however
its biocompatibility has to be improved by the addition

(A) (B)

(C)

Fig. 6. Fractured surface images of the specimens; (A) pure ZA, (B) ZA containing 20 vol% HA, and (C) pure HA.
 ARTICLE IN PRESS
514 Y.-M. Kong et al. / Biomaterials 26 (2005) 509–517

(A) 100 µm (B) 100 µm

(C) 100 µm (D) 100 µm

Fig. 7. Morphologies of proliferated MG63 cells on (A) pure ZA, (B) 10HA (10 vol% HA+90 vol% ZA) composite, (C) 30HA (30 vol%
HA+70 vol% ZA) composite, and (D) pure HA after 5 days culture.

10
ALP Activity [µmol p-nitrophenol/h/mg protein]

( p > 0.05) (ANOVA, p < 0.005)

1.0 (p = 0.91)
(ANOVA, p < 0.001)
Number of Cells [x 10 /cm ]
2

8 (p < 0.05)
5

0.8 (p = 0.062)
( p < 0.05)

6
0.6

4 0.4

2 0.2

0.0
0 ZA 10HA 20HA 30HA 40HA HA
ZA 10HA 20HA 30HA 40HA HA
Specimen
Specimen
Fig. 9. Differentiation level as expressed by the ALP activity on the
Fig. 8. Proliferation results on the specimens after 7 days culture. The specimens after 10 days culture. The ALP expression level of the ZA–
number of cells was significantly increased by the addition of 10% HA HA composite was significantly different from that of pure ZA when
to the ZA (ANOVA, po0.05). The number of cells increased steadily 20% HA was added (ANOVA, po0.05). The expression level
with further addition of HA (ANOVA, po0.001). When 40% HA was increased steadily with increasing HA content (ANOVA, po0.001).
added to the ZA, the proliferation rate on that specimen became The expression level of the composite became comparable with that of
comparable to that on the pure HA. pure HA when 40% HA was added. (ANOVA, p>0.05).

of calcium phosphates such as HA or TCP, in order for ceramics. The reaction products, such as tricalcium
satisfactory osseointegration and faster bone regenera- phosphate [TCP; Ca3(PO4)2], dissolve in the body fluid
tion to be achieved [14,15]. much faster than HA. As a bioresorbable material, TCP
When HA-based composites are made using other might be ideal for some biomedical applications [1].
ceramics as reinforcing agents, chemical reactions have However, as a constituent for the composite material for
been found to occur between the HA and the other load-bearing applications, the use of TCP is not
 ARTICLE IN PRESS
Y.-M. Kong et al. / Biomaterials 26 (2005) 509–517 515

preferable because of the consequent difficulty in meet- 100 nm

ing the requirements in terms of mechanical perfor-
mance of the implants.
Recently, biphasic calcium phosphate (BCP) of HA/
TCP was reported to have the potential for being a
superior biomaterial; HA and TCP act as a seed for new
bone and a supplier of the Ca and P ions required to
accelerate new bone formation, respectively. In fact,
BCP of HA/TCP was reported to induce faster bone
regeneration than HA [14], to show higher strength than
HA [15], to induce new bone more effectively than TCP
[16], and to show higher bone/bone marrow formation
[17]. Therefore, the decomposition reaction of HA into
TCP, which has been thought until now to be
deleterious for the HA-composite body, could in fact
be beneficial, if the extent of the reaction is controllable
(then, there would be BCP of HA/TCP present in the
composite).
If the reaction which takes place in the HA-added ZA
composite mainly occurs at the contact area between
HA and zirconia, then the main factor affecting the
reaction is the surface area of zirconia. In this study, the
use of ZA nano-composite powder was thought to
decrease the contact area between HA and zirconia,
which would hinder the reaction and consequently leave
BCP of HA/TCP. As schematically shown in Fig. 10(A),
100 nm ZA nano-composite powder contains approxi-
mately 10 nm-sized zirconia crystallites (strongly clus-
tered together). Only those zirconia crystallites situated
in the surface regions of the 100 nm-sized clusters
(composition of ZA is 80 wt% zirconia+20 wt% alumi-
na) are in contact with the HA particles and are
therefore available for subsequent reaction. On the other
hand, conventionally mixed zirconia–alumina has about
300 nm-sized zirconia agglomerates dispersed in the
zirconia–alumina powder mixture (Fig. 10(B)) and,
therefore, all of the zirconia particles can freely come
into contact with the HA particles after the mixing
process. In other words, conventionally mixed ZA
powder has more surface area in contact with HA than
ZA nano-composite powder.
After the hybridization of the HA and ZA powders,
the results of the mechanical property evaluation
and phase analysis of the specimens (conducted by
means of XRD) support the above hypothesis. In the
conventionally mixed ZA–HA composite, severe reac-
tions between HA and zirconia decreased the strength of
the specimen considerably (as shown in Fig. 5, from
990 MPa to 240 MPa, in the case of 40 vol% HA
addition) compared with the HA-added ZA nano-
composite. Therefore, in order to reduce the extent of
the reaction, the use of ZA nano-composite powder
would be preferable.
As shown in the XRD results of the HA-added ZA
nano-composite (Fig. 4), the intensities of the HA and
TCP peaks increased steadily with increasing HA
ZrO2
ZrO2
Al2O3 ZrO2 Al2O3
ZrO2 ZrO2
10 nm 300 nm
(A) (B)
Fig. 10. Schematic diagram of (A) ZA nano-composite powder and
(B) conventionally mixed ZA powders. The bright and gray circles
represent ZrO2 and Al2O3 particles, respectively.
100
I TCP / ( I HA + I TCP ) [%] 75
50
25
0
10 20 30 40
HA content [vol%]
Fig. 11. Relative amount of TCP in ZA–HA composites hot-pressed
at 1400 C as a function of the HA content.
content. The relative amounts of TCP to HA in the
HA–ZA nano-composites were calculated from the
XRD data and are plotted in Fig. 11. The relative am-
ounts of TCP to HA in all compositions were in the
range of 40–60%; this result further supports the
hypothesis that the reaction between the ZA nano-
composite powder and the HA powder only occurs to a
limited extent.
For the SEM observation of the cell morphology,
cells were cultured for 5 days with low-cell density.
No significant morphological difference was observed
among the ZA–HA composites as the HA content in the
ZA nano-composite increased (as shown in Fig. 7).
However, the cells proliferated more actively on the
specimens with a higher HA content, as shown in Fig. 8.
Similar enhanced proliferation behaviors were observed
when HA was added to various polymeric biomaterials
[18–20]. This improved proliferation behavior is thought
to be due to the osteogenic effect of the HA component
dispersed within the materials. It is noted that the
 ARTICLE IN PRESS
516 Y.-M. Kong et al. / Biomaterials 26 (2005) 509–517

in the formation of BCP of HA/TCP within the

1600 1.0
composites. The resultant body has high mechanical

[µmol p-nitrophenol/h/mg protein]

Alkaline Phosphatase Activity
0.9
strength, while maintaining the excellent biocompat-
1200 ALP activity ibility. As the amount of HA was increased, the
Strength [MPa]

0.8 proliferation and differentiation behaviors of the osteo-

800
blast-like cells on the composites increased.
0.7

400 0.6
Strength
Acknowledgements
0.5
0
This work was supported by a grant from the Korea
0 20 40 60 80 100 Health 21 R&D Project, the Ministry of Health &
HA content [vol%] Welfare, Republic of Korea (02-PJ3-PG6-EV11-0002).
Fig. 12. Effect of HA addition on the flexural strength and bioactivity
(ALP activity) of the ZA composites. The error bars represent 71
standard deviations (for flexural strength) and 71 standard errors (for
ALP activity). References

[1] Hench LL. Bioceramics: from concept to clinic. J Am Ceram Soc

number of cells proliferated on the ZA with 40% HA 1991;74(7):1487–510.
was not significantly different from that on the pure HA. [2] Garcia R, Doremus RH. Electron microscopy of the bone-
The differentiation behavior as expressed by the ALP implant interface from a human dental implant. J Mater Sci
activity after 10 days culture on the specimens (Fig. 9) Mater Med 1992;3:154–6.
followed a trend similar to that of the proliferation [3] Webster TJ, Ergun C, Doremus RH, Bizios R. Hydroxylapatite
with substituted magnesium, zinc, cadmium, and yttrium. II.
behavior. As in the case of the proliferation behavior, Mechanism of osteoblast adhesion. J Biomed Mater Res 2002;59:
the ALP activity increased steadily with increasing HA 312–7.
content in the ZA–HA composite. When 40% HA was [4] Lee IS, Whang C, Kim H, Park J, Song JH, Kim S. Various Ca/P
added, the ALP activity of the cells proliferated on the ratios of thin calcium phosphate films. Mat Sci Eng C 2002;22:
composite specimen was about the same as that on the 15–20.
[5] Ducheyne P, Marcolongo M, Schepers E. Bioceramic composites.
pure HA (ANOVA, p>0.05). These ALP expression In: Hench LL, Wilson J, editors. An Introduction to bioceramics.
levels, along with the proliferation behaviors, provide a Singapore: World Scientific Publishing Co; 1993. p. 281–97.
good illustration of how the biocompatibility of bioinert [6] Choi JW, Kong YM, Kim HE, Lee IS. Reinforcement of
ceramics could be improved through the addition of HA. hydroxyapatite bioceramic by addition of Ni3Al and Al2O3.
Lastly, in order to estimate the optimal bio-mechan- J Am Ceram Soc 1998;81(7):1743–8.
[7] Tamari N, Mouri M, Kondo I. Mechanical properties and
ical property of the HA-added ZA nano-composites, the existing phases of composite ceramics obtained by sintering of a
flexural strength and ALP activity of the composites mixture of hydroxyapatite and zirconia. Yogyo-Kyokai-Shi 1987;
were simultaneously plotted, as shown in Fig. 12. With 95(8):806–9.
increasing HA content in the composite, the strength [8] Kong YM, Kim S, Kim HE, Lee IS. Reinforcement of
hydroxyapatite bioceramic by addition of ZrO2 coated with
decreased, while the ALP activity steadily increased.
Al2O3. J Am Ceram Soc 1999;82(11):2963–8.
There is a wide range of compositions that can be [9] Kim S, Kong YM, Kim HE, Lee IS. Effect of calcinations of
utilized for actual load-bearing biomedical applications starting powder on mechanical properties of hydroxyapatite-
[21]. If the emphasis is placed on the mechanical alumina bioceramic composite. J Mater Sci Mater Med 2002;
properties, composites containing less than 30% HA 13(3):307–10.
would be preferable. On the other hand, if the [10] MacDonald DE, Betts F, Stranick M, Doty S, Boskey AL.
Physicochemical study of plasma-sprayed hydroxyapatite-coated
bioactivity is more important, more than 30% HA implants in humans. J Biomed Mater Res 2001;54:480–90.
should be added to the composite. In this case, the [11] Kong YM, Kim DH, Kim HE, Heo SJ, Koak JY. Hydroxyapatite
strength of the nano-composite specimen is much higher based composite for dental implants: an in vivo removal torque
than that of pure HA, while the bioactivity is about the experiment. J Biomed Mater Res 2002;63:714–21.
same. [12] Tsukuma K, Ueda K, Shimada M. Strength and fracture
toughness of isostatically hot-pressed composites of Al2O3 and
Y2O3—partially stabilized ZrO2 tetragonal ZrO2. J Am Ceram
Soc 1985;68(1):C4.
5. Conclusion [13] Brinker CJ, Scherer GW. Sol-gel science: the physics and
chemistry of sol–gel process. Boston: Academic Press; 1990.
p. 291.
Strong zirconia–alumina (ZA) nano-composites were
[14] Harada Y. Experimental studies of healing process on compound
fabricated with the addition of bioactive HA for the blocks of hydroxyapatite particles and tricalcium phosphate
purpose of improving their biocompatibility. Hot powder implantation in rabbit mandible. J Tokyo Dent College
pressing the nano-composite powder at 1400oC resulted Soc 1989;89:263–97.
 ARTICLE IN PRESS
Y.-M. Kong et al. / Biomaterials 26 (2005) 509–517
[15] Monma H. Tricalcium phosphate ceramics complexed with
hydroxyapatite. J Ceram Soc Jpn 1987;96:60–4.
[16] Alam MI, Ashhina I, Ohmamiuda K, Enomoto S. Comparative
study of biphasic calcium phosphate ceramics impregnated with
rhBMP-2 as bone substitutes. J Biomed Mat Res 2001;54:129–38.
[17] Kurashina K, Kurita H, Wu Q, Ohtsuka A, Kobayashi H.
Ectopic osteogenesis with biphasic ceramics of hydroxyapatite
and tricalcium phosphate in rabbits. Biomaterials 2002;23:407–12.
[18] Moursi AM, Winnard AV, Winnard PL, Lannutti JJ, Seghi RR.
Enhanced osteoblast response to a PMMA-HA composite.
Biomaterials 2002;23:133–44.
517
[19] Huang J, Di Silvio L, Wang M, Tanner KE, Bonfield W. In vitro
mechanical and biological assessment of hydroxyapatite rein-
forced polyethylene composite. J Mater Sci Mater Med 1997;8:
775–9.
[20] Dalby MH, Di Silvio L, Harper EJ, Bonfield W. In vitro
evaluation of a new PMMA cement reinforced with hydroxyapa-
tite. J Mater Sci Mater Med 1999;10:793–6.
[21] Li J, Hastings GW. Oxide bioceramics: inert ceramic materials in
medicine and dentistry. In: Black J, Hastings GW, editors.
Handbook of biomaterials properties. London: Chapman & Hall;
1998. p. 340–54.