- Email: [email protected]
A new bioactive material which transforms in situ into hydroxyapatite
Acta ma&r. Vol. 46, No. 7, pp. 2541-2549, 1998 0 1998 Acta Metallurgica Inc. Published bv Elsevier Science Ltd. All rights reserved Printed in &eat Britain PII: S1359-6454(97)00413-S 1359-6454/98 $19.00 + 0.00
Pergamon
A NEW BIOACTIVE IN SITU
MATERIAL WHICH TRANSFORMS INTO HYDROXYAPATITE
P. N. DE AZA’, F. GUITIAN’ ‘Instituto
and S. DE AZA2
de Ceramica, Universidad de Santiago, Santiago de Compostela, Spain and *Institute Ceramica y Vidrio (CSIC), Arganda del Rey, Madrid, Spain
de
Abstract-A new route of obtaining bioactive ceramic materials, which improve the ingrowth of new bone into implants (osseointegration), is presented. This involves obtaining eutectic structures from selected systems, bearing in mind the different bioactive behaviour of the phases. In this work the eutectic binary system wollastonite-tricalcium phosphate was chosen because wollastonite is bioactive and tricalcium phosphate is resorbable. The eutectic material is formed by spherical colonies composed of alternating radial lamellae of wollastonite and tricalcium phosphate. The eutectic material, in in vitro experiments, reacts by dissolving the wollastonite phase and forming a porous structure of hydroxyapatite by a pseudomorphic transformation of the tricalcium phosphate lamellae, which in turn mimic porous bone. Later, a hydroxyapatite layer is formed by precipitation on the surface of the material. The procedure developed by the authors opens the opportunity to obtain a new family of bioactive materials for which the general name of bioeutectica is proposed. 0 1998 Acta Metallurgica Inc. RbumLUne
nouvelle methode d’obtention de materiaux ceramiques bioactifs qui favorisent l’integration d’implants avec 1’0s est presentee. Le fondement de cette mithode reside dans l’obtention de structures eutectiques au sein de systemes choisis en tenant compte du different comportement bioactif des phases. Dans ce travail, le systeme wollastonite-phosphate tricalcique a Cte choisi du fait que la premiere phase est bioactive et la seconde reabsorbible. Le materiel eutectique est compose de colonies spheriques de lamelles radiales alternes de wollastonite et de phosphate tricalcique respectivement. Dans les experiences in vitro, le materiel eutectique se transforme en diluant la phase wollastonite et en formant une structure poreuse d’hydroxyapatite, pseudomorphique avec le phosphate tricalcique, qui ressemble a 1’0s poreux. I1 se forme ensuite par precipitation une couche d’hydroxyapatite qui recouvre la surface du materiel. Le pro&de developpe par les auteurs ouvre une voie originale pour le dtveloppement d’une nouvelle famille de materiaux ceramiques bioactifs pour laquelle il est propose le nom general de bioeutectiques@t 0 1998 Acta Metallurgica Inc.
Zusammenfassung-Eine neue Methode fur die Erzeugung von keramischen bioaktiven Materialien, die die Knochenintegrierung der Implantate begtinstigen, wird beschrieben. Sie sttitzt sich auf das Erzielen von eutektischen Strukturen in ausgesuchten Systemen beim Beriicksichtigen des verschiedenen Verhaltens der bioaktiven Phasen. In vorliegender Arbeit wurde das System WollastonittKalziumphosphat aus Grund der Bioaktivitat der Wollastonit und der Resorption des Kalziumphosphates ausgewlhlt. Das eutektische Material besteht aus rundfiirmigen Anhaufungen, die von abwechselnden radialen Plattchen beider Phasen gebildet sind. In den in vitro durchgeftihrten Versuchen verlndert sich das eutektische Material, lost sich die Wollastonitphase auf, und bildet sich eine poriise Struktur aus Hydroxylapatit, die pseudomorph mit dem Kalziumphosphat ist und dem poriisen Knochen sich Lhnelt. Nachher wachst eine Schicht beim Niederschlagen von Hydroxylapatit, der die Oberfllche des Materials iiberzieht. Das durch die Verfasser entwickelte Verfahren (i&et einen originellen Weg fur die Entwicklung einer neuen Familie von keramischen bioaktiven Materialien fur die den allgemeinen Name von Bioeutektikas vorgeschlagen wird. 0 1998 Acta Metallurgica Inc.
1. INTRODUCTION Ceramic
biomaterials
becoming
increasingly
outstanding
problems
for
bone
important still remain
replacement
[ 11, however unsolved.
are many Among
them, and probably the most important, is the total osseointegration in the human body of ceramic implants. When bioactive materials are implanted in a living body the interaction between the bone tissue and these materials only takes place on their surfaces, with the remaining bulk of the material unchanged. This commonly results in a harmful shear stress. To improve the ingrowth of new bone into implants (osseointegration) the use of materials
with an appropriate interconnected porous structure has been recommended [2-51. However, porous materials always have poor mechanical properties. To overcome this problem the idea of creating artificial bone, making use of coral, has been developed [c-9]. Coral, the limestone material that is made up of external skeletons of tiny creatures called polyps, has a microstructure similar to porous bone, and is therefore a useful starting point. Coral is normally baked in a diammonium phosphate solution to convert its limestone deposits into hydroxyapatite (HA). At present, however, there is no in viva evidence to suggest that the highly orga-
2542
De AZA et al.:
BIOACTIVE
MATERIAL
WHICH
nized macropore structure of the coral-derived HA is better, with regard to bone ingrowth and bioresorption, than the random pore structures of the more easily prepared synthetic porous implants. In the present work, another new approach is presented. This is based on designing dense bioactive ceramic materials with the capacity to develop an in situ porous HA structure when they are implanted in a living body. To satisfy this purpose the material should be comprised of at least two phases, one bioactive and the other resorbable. Taken into account the bone structure, the most appropriate microstructure to be designed is an irregular lamellar eutectic microstructure [lo], because it becomes similar to porous bone when one of the phases is dissolved. Bearing in mind all these premises, and after searching all the possible binary systems that could satisfy the previous mentioned conditions, the system wollastonite (W))tricalcium phosphate (TCP) was selected and studied (Fig. 1) [ll]. The wollastonite, a simple-chain calcium silicate (CaSiOs), is bioactive [12-141 and the tricalcium phosphate (Ca,(PO&) is resorbable [15, 161. It is apparent that
TRANSORMS
INTO HYDROXYAPATITE
the system is of the eutectic type with an invariant point at 1402” k 3°C and a composition of 60 wt% W and 40 wt% TCP. Additionally, due to the volume fraction of the two phases at the eutectic point (IV = 0.61 and TCP = 0.38) and their high dimensionless entropy of fusion (c( = [Sr/R]>2) (tlw= 3.71 and @rCp= 9.66), the expected microstructure of the eutectic composition should be of irregular lamellar type [lo, 17-191.
2. EXPERIMENTAL
To obtain eutectic morphologies, similar to porous bone, slow cooling and radial heat extraction from the eutectic liquid were selected. To this purpose, pellets of the eutectic composition were heated up to 1500°C/2 h, in platinum foil crucibles, to obtain homogeneous liquids which were then cooled to 1410°C at a rate of 3”C/min. From this temperature the samples were further cooled inside the furnace down to 1390°C 12°C below the eutectic temperature, at rates of 2, 1, O.YC/h. After that, the specimens were furnace cooled.
T (“C) 1700
1600
d TCP + Liq.
/
I
\ \
psW + ctTCP
psW
+fiTCP
\ 1 125f10°C
\
W +pTCP
I
W
10
I
I
I
20
30
40
I
50
I
I
60
70
I
80
I
90
TCP
Wt% Fig.
1. The system
wollastonite-tricalcium phosphate (W = wollastonite; TCP = tricalcium phosphate) [l 11.
psW = pseudowollastonite;
De AZA et al.:
BIOACTIVE
MATERIAL
WHICH
TRANSORMS
2543
INTO HYDROXYAPATITE
Fig. 2. SEM images of eutectic materials, after light etching in citric acid, obtained at cooling rates of: (A) 2”C/h; (B) l”C/h; (C) O.S”C/h. (D) High magnification image of a colony belonging to sample (C).
Plates of the eutectic material, ing rate of O.YC/h, thick were immersed lated
body
ionic
fluid
3 mm in diameter
(SBF),
which
to
that
[20], in polyethylene
body temperature
had
and 2 mm simu-
almost
of
equal
human
bottles
blood
at the human
(36.5”C). The immersion
periods
varied from 1 day to 3 weeks. Changes coupled AES),
in ionic concentration,
plasma
atomic
In addition,
the pH at the interface
material
an ion-sensitive
intervals,
transistor
(SEM)
energy-dispersive
using and
(EDS area
electron
microscope
p-Raman
fitted
allows
a spatial
with
in a transmission
(Jeol Jem 2010 TEM on thinned
samples were also studied
resolution
operated
specimens.
The
by thin film X-ray diffrac-
spectroscopy an
with spec-
at 20 kV. In addition,
(SAD)
a 200 kV) was obtained
The samples exhibited by SEM imaging (Fig. 2) are comprised of quasi-spherical colonies formed by alternating radial lamellae of W and TCP in an irregular eutectic microstructure. EDS microanalysis differentiates between the white phase as W and the darker phase as TCP. The major difference between the samples cooled at the different rates, is the greater size of the colonies for samples that are cooled slowly (Fig. 3). However, the width of the lamellae of W and TCP, -1.3 and -0.9 pm, respectively, seems to remain practically constant. As can be seen in all the microphotographs (Fig. 2) the phase which closes the colonies and constitutes the outer part of them is always the W, which is in agreement with its greater volume fraction in the eutectic composition. Through this phase the colonies join themselves.
electron
6400 fitted
wavelength-dispersive
diffraction
T64000
were removed
by scanning
a Jeol
and WDS)
selected
and
the using
of Si3N4 type
the specimens
the fluid and analyzed
microscopy trometers
between
[21,22].
At periodic
1 ml.
(ICP-
and the SBF was monitored field-effect
(ISFET-Meter)
tion
using inductively spectroscopy
and pH of the SBF were daily determined.
eutectic
from
emission
3. RESULTS
at a cool-
in 100 ml of an acellular
concentration
plasma
obtained
optical
(Jobin
microscope
on the samples
Ivon which close to
20 t
01 0
I OJ
I
0.4.
I
o,*
I
o,*
I 1
Cooling rate Fig. 3. Colony
I I,1
I 1.4
(“C/h)
size vs cooling
rates
I 1,o
I I,0
I 2
2544
De AZA et al.:
Fig. 4. Triple junction
BIOACTIVE
between
colonies.
MATERIAL
Sample
WHICH
cooled at
a rate of 0.5”C/h. Table
1. WDS mean area microanalysis (wt%) in the triple junction of Fig. 4
SiO*
cao
54.1
40.2
.&O, 3.9
PZOS 1.8
Small quantities of impurities drastically affect the way the colonies are joined together. Impurities are segregated to the periphery of the colonies as
TRANSORMS
INTO
HYDROXYAPATITE
they grow and are mainly concentrated at triple junctions. This fact breaks the structure of alternating lamellae of W and TCP to form a continuous matrix of W joining the colonies with dispersed TCP inclusions in it. Figure 4 exhibits clearly this fact and Table 1 lists the mean area composition of the triple junction showing the presence of alumina impurities. The TEM studies confirm the presence of W and TCP phases as shown in Fig. 5. The light phase is W and the darker phase is TCP. The W phase contains a high density of dislocation loops. In the same figure the SAD patterns of the two phases indicate their crystalline nature. The material appears to be dense and free of any cracks at interfaces. However, at a higher resolution, some misfit dislocations were observed in the phase boundaries, indicating a semicoherent interface between the two phases. The eutectic microstructure, belonging to the sample obtained at a cooling rate of O.YC/h, has been etched with diluted HCl acid to eliminate the TCP and is shown in Fig. 6. The morphology of this microstructure is similar to that of porous human bone. Figure 7 shows the microstructure of a polished cross-section of the eutectic material,
De AZA et al.:
BIOACTIVE MATERIAL WHICH TRANSORMS INTO HYDROXYAPATITE
Fig. 6. SEM images of the eutectic material after etching with diluted HCI acid.
2545
obtained a rate of 0.5”C/h, after soaking in SBF for one week as well as elemental X-ray maps of silicon, calcium and phosphorous. Selective dissolution of one phase has occurred to a depth of -50 pm. The dissolved phase is the W, which closed the colonies and joined them in the original microstructure (Fig. 2). This is confirmed by the elemental Xray maps and EDS microanalyses. These show that the remaining phase is a calcium phosphate. pRaman spectroscopy of the reaction zone [Fig. 8(c)] confirms that the calcium phosphate corresponds to a hydroxyapatite (HA) of low crystallinity. It appears that the W dissolves in the SBF and the TCP transforms into HA. The surface of the same sample is shown in Fig. 9. Small spherical deposits of HA are observed and confirmed by p-Raman spectroscopy [Fig. 8(d)]. A polished cross-section of a sample of the eutectic material, (prepared at a cooling rate of O.S’C/h), that had been immersed for three weeks in SBF, is shown in Fig. 10. A 5 pm thick layer of HA has formed on the surface and has been confirmed by p-Raman spectroscopy [Fig. 8(d)]. This thick layer of HA is built up over the HA lamellae previously formed and even penetrates between them. Daily monitoring of pH and of the ionic concentration of calcium, silicon and phosphate are presented in Fig. 11. A substantial increase in the Si +4 concentration, at a rate of - + 0.42 ppm/day, is observed during the first seven days of immersion,
Fig. 7. SEM image of a polished cross-section of the eutectic plate after one week soaking in SBF and elemental X-ray maps of Si, P, and Ca.
2546
De AZA ef al.:
BIOACTIVE MATERIAL WHICH TRANSORMS INTO HYDROXYAPATITE
b
C
i
d
100
300
500
700.
900
1100
100
300
100
700
9
Raman shift (cm-i) Fig. 8. p-Raman spectra of: (a) standard TCP; (b) standard HA; (c) the reaction zone in Fig. 7; (d) the surface of the sample after one week soaking in SBF (Fig. 9).
after which it increases very slowly. This result justifies the dissolution of the W in the SBF. The (P04)3- concentration diminishes slowly, during the same period of time, at a constant rate of - - 0.23 ppm/day. The Cat2 concentration slowly increases at a constant rate of - + 0.13 ppm/day. Lastly, the pH of the SBF remains essentially constant during the entire experiment, changing only from 7.25 to 7.35. In Fig. 11 is also shown the pH variation at the interface between the eutectic material and the SBF. At the beginning of the reaction, the pH quickly increases to 8.5 and then stabilizes.
Fig. 9. SEM image of the surface
4. DISCUSSION
After
immersion
of the eutectic
in the SBF, two different periphery
of the sample.
tion of the material solution formation
of the
stituted
with the SBF causing
the dis-
lamellae
and
the
subsequent
by a pseudomorphic
of the TCP, and the second
by a superficial reaction,
on the by reac-
deposit
sition of HA is less abundant of the
material
The first is formed
wollastonite
of HA
transformation
ceramic
zones are formed
but
becomes
during
the first stage
a thick
layer.
of the sample after one week soaking
is con-
of HA. This depo-
in SBF.
and
dense
De AZA et al.:
Fig.
BIOACTIVE
MATERIAL
WHICH
TRANSORMS
INTO
HYDROXYAPATITE
10. SEM image of a polished cross-section of the eutectic material after three weeks soaking SBF. (A) A 5 pm HA layer; (B) HA lamellae; (C) original eutectic ceramic material.
It is worth emphasizing that no layer of amorphous silica was observed, as is normally detected in reactions of silica-based bioactive materials with SBF. The results obtained can be explained through the following mechanism (Fig. 12): At the beginning of the experiment two nearly simultaneous reactions occur. First, the W phase in contact with the SBF starts to react via an ionic exchange of 2H30+ from the SBF for one Ca*+ from the W network. This exchange transforms the crystalline W into an
2547
in
amorphous silica phase as has been demonstrated in previous works [12-14,211. This reaction starts at the surface of the material, thereby causing the pH at the material/SBF interface to increase to 8.5 and releasing calcium and silicon ions into the SBF solution. The reaction progresses deeply into the material, in the confined channels between the TCP lamellae as W is dissolved. This reaction triggers the next events. Immediately, the TCP lamellae start to react with the Ca*+ and OH- ions present in the confined
Si” @pm) 1
10
8 6
(PO.)’
(Ppm) 30
loo_.
___‘_-
-
1:
-
95 _~
90
2
4
6
8
10 12
time(days)
14
16
25
z
I
18
2
4
6
8
10
12
14
16
16
time (days)
Fig. 11. Changes in Ca’+, (P0,)3- and Si4+ concentration in SBF during soaking of the eutectic material. Changes in pH during exposure to SBF at the interface between the material and the SBF (0) and away from the interface (u).
2548
De AZA et al.:
BIOACTIVE MATERIAL WHICH TRANSORMS INTO HYDROXYAPATITE
SBF 2H,O+
2H,O’
: Ca2
: Ca’+
0- spi-OH
O- i-OH P 0
b
0
P OH-j?i-0
OH- i-0 1 0
b pH=lOS
W
pH=lOS
TCP
W
TCP
W
TCP
W
TCP
3 [Ca,(PO,),]+Ca"+ 2(OH)I)Ca,,(P04),(OH), 6 [HPOi]+lO Ca2't8(0.H)I)Ca,,[P0,],[OH],+6H20 Fig. 12. Schematic representation of HA formation mechanism. channels, at pH = 10.5 [21]. Thus a pseudomorphic transformation of TCP into HA occurs as shown in Fig. 12. The silicon and calcium ions, not involved in the reaction, migrate through the SBF solution away from the interface and are monitored as an increase in concentration (Fig. 11). Calculations, based on the weight percent of both phases in the eutectic and on the quantity of W dissolved in the SBF, determine that -85 wt% of the calcium contained in the dissolved W should be in the SBF. Since a large increase was not detected (Fig. ll), most of Ca2+ must react with the phosphate of the SBF at the material/SBF interface and lead to the precipitation of HA on the surface of the material. This is the second reaction schematically shown in Fig. 12. Initially the precipitation occurs on the newly formed HA lamellae (Fig. 9). Later, as the precipitation increases (Fig. lo), the HA eventually covers the entire surface of the sample. The final effect of the combined reactions is to produce a large increase in Si+4 concentration in the SBF solution, a small increase in Ca2+ and a decrease of (P04)3- as detected in Fig. 11. The first step of the reaction mechanism, which at the beginning is very fast, is slowed down due to the combined effect of the difficulty of diffusion of protons from the SBF to reach the reaction interface as the distance is increased and due to the closure of channels as HA is formed on the surface of the sample (Fig. 11). 5. CONCLUSIONS Eutectic microstructures have been obtained from the system wollastonite-tricalcium phosphate by radial heat extraction and slow cooling inside the furnace. The microstructure is comprised of quasi-
spherical colonies of alternating radial lamellae of both phases. This microstructure may be modified for specific applications of this material. The eutectic ceramic material, when soaked in SBF, reacts first by dissolving the wollastonite phase and immediately forming a porous structure of hydroxyapatite by a pseudomorphic transformation of the tricalcium phosphate lamellae. The microstructure obtained is similar to that of porous bone. Later, a dense hydroxyapatite layer is formed by precipitation on the outer surface of the material. It is expected that this material will behave similarly in in viva experiments facilitating the osseointegration of the implant. The procedure developed by the authors opens the opportunity to obtain a new family of bioactive materials for which the general name of bioeutectics@ is proposed. Acknowledgements-This work was supported by CICYT under Project MAT97-1025. P.N.D.A. thanks the Ministry of Education and Science of Spain for the Fellowship given to her. In addition, the authors wish to thank A. P. Tomsia and E. Saiz and also Z. B. Luklinska and M. R. Anseau for the facilities given to P.N.D.A. during her respective stays at Lawrence Berkeley Laboratory (CA, U.S.A.) and at IRC in Biomedical Materials, The London Hospital, Medical College, University of London. Finally, thanks also are given to L. Artds and R. Cusch for the pRaman spectra and useful discussions.
REFERENCES 1. Hulbert, S. F., Bohros, J. C., Hench, L. L., Wilson, J. and Heimke, G., in High Tech Ceramics, ed. P. Vicenzini. Elsevier Amsterdam, The Netherlands, 1987, pp. 189-213. 2. Klawitter, J. J. and Hulbert, S. F., J. Biomed. Mater. Res. Symp., 1971, 2, 161. 3. Jarcho, M., Chin. Orthop. Rel. Res., 1981, 157, 259.
De AZA et al.:
BIOACTIVE
MATERIAL
WHICH
4. De Groot, K. and Le Geros, R., in Bioceramics Materials Characteristics vs in Vivo Behavior, ed. P. Ducheyne. J. Ann. N.Y. Acad. Sci., New York, 1988, PQ. 268-277. 5. D-e Groot, K., in Bioceramics Material Characteristics vs In Vivo Behavior. ed. P. Duchevne. J. Ann. N.Y. Acad. Sci., New York, 1988, pp. 227-235. 6. Roy, D. M. and Linnehan, S. K., Nature, 1974, 214, 220. 7. Sayler, K., Holmes, R. and Johns, D., J. Dent. Res., 1977, 56B, 173. 8. Tencer, A. F., Shors, E. C., Woodard, P. L. and Holmes, R. E., in Handbook of Bioactive Ceramics, Vol. II. Calcium Phosphate and Hydroxylapatite Ceramics, ed. T. Yamamuro, L. L. Hench and J. Wilson. CRC Press Inc., Boca Raton, FL 33431, U.S.A., 1990, pp. 209-221. 9. Sivakumar, M., Dampath Kumar, T. S., Dhantha, K. L. and Panduranga Rao, K., Biomaterials, 1996, 17, 1709. 10. Kurz, W. and Fisher, D. J., Fundamental of Solidification. Trans. Tech. Publications, Switzerland, 1984. 11. De Aza, P. N., Guitian, F. and De Aza, S., J. Am. Ceram. Sot., 1995, 78, 1653.
TRANSORMS
INTO
HYDROXYAPATITE
2549
12. De Aza, P. N., Guitian, F. and De Aza, S., Scripta metall. mater., 1991, 31, 1001. 13. De Aza, P. N., Guitian, F. and De Aza, S., in Advances in Science and Technology, 12. Materials in Clinical Applications, ed. P. Vicenzini, Techna Sri., Florenze, 1995, pp. 19-27. 14. De Aza, P. N., Luklinska, Z. B., Anseau, M. R., Guitian, F. and De Aza, S., J. Microsc., 1996, 182, 24-31. 15. De Groot, K., in Bioceramics of Calcium Phosphate, ed. K. De Groot. Press Inc., Boca Raton, FL 33431, U.S.A., 1983. 16. Hench, L. L., J. Am. Ceram. Sot., 1991, 74, 1487. 17. Flemings, M. C., Sci. Am., 1974, 231, 88. 18. Elliot, R., ht. Metals Rev., 1977, R219, 161. 19. Ashbrook, R. L., J. Am. Ceram. Sot., 1977, 60, 428. 20. Gamble, J., Chemical Anatomy, Physiology and Pathology of Extracellular Fluid. Harvard University Press, Cambridge, 1967. 21. De Aza, P. N., Guitian, F., Merlos, A., Lora-Tamayo, E. and De Aza, S., J. Mater. Sci. Mater. Med., 1996, 7, 399. 22. Merlos, A., Garcia, I., Cane, C., Esteve, J., Bartroli, J., Jimenez, C., Proc. 5th Conf. on Sensors and Their Applications. Edimburg, U.K., 1991, pp. 127-132.
Pergamon
A NEW BIOACTIVE IN SITU
MATERIAL WHICH TRANSFORMS INTO HYDROXYAPATITE
P. N. DE AZA’, F. GUITIAN’ ‘Instituto
and S. DE AZA2
de Ceramica, Universidad de Santiago, Santiago de Compostela, Spain and *Institute Ceramica y Vidrio (CSIC), Arganda del Rey, Madrid, Spain
de
Abstract-A new route of obtaining bioactive ceramic materials, which improve the ingrowth of new bone into implants (osseointegration), is presented. This involves obtaining eutectic structures from selected systems, bearing in mind the different bioactive behaviour of the phases. In this work the eutectic binary system wollastonite-tricalcium phosphate was chosen because wollastonite is bioactive and tricalcium phosphate is resorbable. The eutectic material is formed by spherical colonies composed of alternating radial lamellae of wollastonite and tricalcium phosphate. The eutectic material, in in vitro experiments, reacts by dissolving the wollastonite phase and forming a porous structure of hydroxyapatite by a pseudomorphic transformation of the tricalcium phosphate lamellae, which in turn mimic porous bone. Later, a hydroxyapatite layer is formed by precipitation on the surface of the material. The procedure developed by the authors opens the opportunity to obtain a new family of bioactive materials for which the general name of bioeutectica is proposed. 0 1998 Acta Metallurgica Inc. RbumLUne
nouvelle methode d’obtention de materiaux ceramiques bioactifs qui favorisent l’integration d’implants avec 1’0s est presentee. Le fondement de cette mithode reside dans l’obtention de structures eutectiques au sein de systemes choisis en tenant compte du different comportement bioactif des phases. Dans ce travail, le systeme wollastonite-phosphate tricalcique a Cte choisi du fait que la premiere phase est bioactive et la seconde reabsorbible. Le materiel eutectique est compose de colonies spheriques de lamelles radiales alternes de wollastonite et de phosphate tricalcique respectivement. Dans les experiences in vitro, le materiel eutectique se transforme en diluant la phase wollastonite et en formant une structure poreuse d’hydroxyapatite, pseudomorphique avec le phosphate tricalcique, qui ressemble a 1’0s poreux. I1 se forme ensuite par precipitation une couche d’hydroxyapatite qui recouvre la surface du materiel. Le pro&de developpe par les auteurs ouvre une voie originale pour le dtveloppement d’une nouvelle famille de materiaux ceramiques bioactifs pour laquelle il est propose le nom general de bioeutectiques@t 0 1998 Acta Metallurgica Inc.
Zusammenfassung-Eine neue Methode fur die Erzeugung von keramischen bioaktiven Materialien, die die Knochenintegrierung der Implantate begtinstigen, wird beschrieben. Sie sttitzt sich auf das Erzielen von eutektischen Strukturen in ausgesuchten Systemen beim Beriicksichtigen des verschiedenen Verhaltens der bioaktiven Phasen. In vorliegender Arbeit wurde das System WollastonittKalziumphosphat aus Grund der Bioaktivitat der Wollastonit und der Resorption des Kalziumphosphates ausgewlhlt. Das eutektische Material besteht aus rundfiirmigen Anhaufungen, die von abwechselnden radialen Plattchen beider Phasen gebildet sind. In den in vitro durchgeftihrten Versuchen verlndert sich das eutektische Material, lost sich die Wollastonitphase auf, und bildet sich eine poriise Struktur aus Hydroxylapatit, die pseudomorph mit dem Kalziumphosphat ist und dem poriisen Knochen sich Lhnelt. Nachher wachst eine Schicht beim Niederschlagen von Hydroxylapatit, der die Oberfllche des Materials iiberzieht. Das durch die Verfasser entwickelte Verfahren (i&et einen originellen Weg fur die Entwicklung einer neuen Familie von keramischen bioaktiven Materialien fur die den allgemeinen Name von Bioeutektikas vorgeschlagen wird. 0 1998 Acta Metallurgica Inc.
1. INTRODUCTION Ceramic
biomaterials
becoming
increasingly
outstanding
problems
for
bone
important still remain
replacement
[ 11, however unsolved.
are many Among
them, and probably the most important, is the total osseointegration in the human body of ceramic implants. When bioactive materials are implanted in a living body the interaction between the bone tissue and these materials only takes place on their surfaces, with the remaining bulk of the material unchanged. This commonly results in a harmful shear stress. To improve the ingrowth of new bone into implants (osseointegration) the use of materials
with an appropriate interconnected porous structure has been recommended [2-51. However, porous materials always have poor mechanical properties. To overcome this problem the idea of creating artificial bone, making use of coral, has been developed [c-9]. Coral, the limestone material that is made up of external skeletons of tiny creatures called polyps, has a microstructure similar to porous bone, and is therefore a useful starting point. Coral is normally baked in a diammonium phosphate solution to convert its limestone deposits into hydroxyapatite (HA). At present, however, there is no in viva evidence to suggest that the highly orga-
2542
De AZA et al.:
BIOACTIVE
MATERIAL
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nized macropore structure of the coral-derived HA is better, with regard to bone ingrowth and bioresorption, than the random pore structures of the more easily prepared synthetic porous implants. In the present work, another new approach is presented. This is based on designing dense bioactive ceramic materials with the capacity to develop an in situ porous HA structure when they are implanted in a living body. To satisfy this purpose the material should be comprised of at least two phases, one bioactive and the other resorbable. Taken into account the bone structure, the most appropriate microstructure to be designed is an irregular lamellar eutectic microstructure [lo], because it becomes similar to porous bone when one of the phases is dissolved. Bearing in mind all these premises, and after searching all the possible binary systems that could satisfy the previous mentioned conditions, the system wollastonite (W))tricalcium phosphate (TCP) was selected and studied (Fig. 1) [ll]. The wollastonite, a simple-chain calcium silicate (CaSiOs), is bioactive [12-141 and the tricalcium phosphate (Ca,(PO&) is resorbable [15, 161. It is apparent that
TRANSORMS
INTO HYDROXYAPATITE
the system is of the eutectic type with an invariant point at 1402” k 3°C and a composition of 60 wt% W and 40 wt% TCP. Additionally, due to the volume fraction of the two phases at the eutectic point (IV = 0.61 and TCP = 0.38) and their high dimensionless entropy of fusion (c( = [Sr/R]>2) (tlw= 3.71 and @rCp= 9.66), the expected microstructure of the eutectic composition should be of irregular lamellar type [lo, 17-191.
2. EXPERIMENTAL
To obtain eutectic morphologies, similar to porous bone, slow cooling and radial heat extraction from the eutectic liquid were selected. To this purpose, pellets of the eutectic composition were heated up to 1500°C/2 h, in platinum foil crucibles, to obtain homogeneous liquids which were then cooled to 1410°C at a rate of 3”C/min. From this temperature the samples were further cooled inside the furnace down to 1390°C 12°C below the eutectic temperature, at rates of 2, 1, O.YC/h. After that, the specimens were furnace cooled.
T (“C) 1700
1600
d TCP + Liq.
/
I
\ \
psW + ctTCP
psW
+fiTCP
\ 1 125f10°C
\
W +pTCP
I
W
10
I
I
I
20
30
40
I
50
I
I
60
70
I
80
I
90
TCP
Wt% Fig.
1. The system
wollastonite-tricalcium phosphate (W = wollastonite; TCP = tricalcium phosphate) [l 11.
psW = pseudowollastonite;
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INTO HYDROXYAPATITE
Fig. 2. SEM images of eutectic materials, after light etching in citric acid, obtained at cooling rates of: (A) 2”C/h; (B) l”C/h; (C) O.S”C/h. (D) High magnification image of a colony belonging to sample (C).
Plates of the eutectic material, ing rate of O.YC/h, thick were immersed lated
body
ionic
fluid
3 mm in diameter
(SBF),
which
to
that
[20], in polyethylene
body temperature
had
and 2 mm simu-
almost
of
equal
human
bottles
blood
at the human
(36.5”C). The immersion
periods
varied from 1 day to 3 weeks. Changes coupled AES),
in ionic concentration,
plasma
atomic
In addition,
the pH at the interface
material
an ion-sensitive
intervals,
transistor
(SEM)
energy-dispersive
using and
(EDS area
electron
microscope
p-Raman
fitted
allows
a spatial
with
in a transmission
(Jeol Jem 2010 TEM on thinned
samples were also studied
resolution
operated
specimens.
The
by thin film X-ray diffrac-
spectroscopy an
with spec-
at 20 kV. In addition,
(SAD)
a 200 kV) was obtained
The samples exhibited by SEM imaging (Fig. 2) are comprised of quasi-spherical colonies formed by alternating radial lamellae of W and TCP in an irregular eutectic microstructure. EDS microanalysis differentiates between the white phase as W and the darker phase as TCP. The major difference between the samples cooled at the different rates, is the greater size of the colonies for samples that are cooled slowly (Fig. 3). However, the width of the lamellae of W and TCP, -1.3 and -0.9 pm, respectively, seems to remain practically constant. As can be seen in all the microphotographs (Fig. 2) the phase which closes the colonies and constitutes the outer part of them is always the W, which is in agreement with its greater volume fraction in the eutectic composition. Through this phase the colonies join themselves.
electron
6400 fitted
wavelength-dispersive
diffraction
T64000
were removed
by scanning
a Jeol
and WDS)
selected
and
the using
of Si3N4 type
the specimens
the fluid and analyzed
microscopy trometers
between
[21,22].
At periodic
1 ml.
(ICP-
and the SBF was monitored field-effect
(ISFET-Meter)
tion
using inductively spectroscopy
and pH of the SBF were daily determined.
eutectic
from
emission
3. RESULTS
at a cool-
in 100 ml of an acellular
concentration
plasma
obtained
optical
(Jobin
microscope
on the samples
Ivon which close to
20 t
01 0
I OJ
I
0.4.
I
o,*
I
o,*
I 1
Cooling rate Fig. 3. Colony
I I,1
I 1.4
(“C/h)
size vs cooling
rates
I 1,o
I I,0
I 2
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De AZA et al.:
Fig. 4. Triple junction
BIOACTIVE
between
colonies.
MATERIAL
Sample
WHICH
cooled at
a rate of 0.5”C/h. Table
1. WDS mean area microanalysis (wt%) in the triple junction of Fig. 4
SiO*
cao
54.1
40.2
.&O, 3.9
PZOS 1.8
Small quantities of impurities drastically affect the way the colonies are joined together. Impurities are segregated to the periphery of the colonies as
TRANSORMS
INTO
HYDROXYAPATITE
they grow and are mainly concentrated at triple junctions. This fact breaks the structure of alternating lamellae of W and TCP to form a continuous matrix of W joining the colonies with dispersed TCP inclusions in it. Figure 4 exhibits clearly this fact and Table 1 lists the mean area composition of the triple junction showing the presence of alumina impurities. The TEM studies confirm the presence of W and TCP phases as shown in Fig. 5. The light phase is W and the darker phase is TCP. The W phase contains a high density of dislocation loops. In the same figure the SAD patterns of the two phases indicate their crystalline nature. The material appears to be dense and free of any cracks at interfaces. However, at a higher resolution, some misfit dislocations were observed in the phase boundaries, indicating a semicoherent interface between the two phases. The eutectic microstructure, belonging to the sample obtained at a cooling rate of O.YC/h, has been etched with diluted HCl acid to eliminate the TCP and is shown in Fig. 6. The morphology of this microstructure is similar to that of porous human bone. Figure 7 shows the microstructure of a polished cross-section of the eutectic material,
De AZA et al.:
BIOACTIVE MATERIAL WHICH TRANSORMS INTO HYDROXYAPATITE
Fig. 6. SEM images of the eutectic material after etching with diluted HCI acid.
2545
obtained a rate of 0.5”C/h, after soaking in SBF for one week as well as elemental X-ray maps of silicon, calcium and phosphorous. Selective dissolution of one phase has occurred to a depth of -50 pm. The dissolved phase is the W, which closed the colonies and joined them in the original microstructure (Fig. 2). This is confirmed by the elemental Xray maps and EDS microanalyses. These show that the remaining phase is a calcium phosphate. pRaman spectroscopy of the reaction zone [Fig. 8(c)] confirms that the calcium phosphate corresponds to a hydroxyapatite (HA) of low crystallinity. It appears that the W dissolves in the SBF and the TCP transforms into HA. The surface of the same sample is shown in Fig. 9. Small spherical deposits of HA are observed and confirmed by p-Raman spectroscopy [Fig. 8(d)]. A polished cross-section of a sample of the eutectic material, (prepared at a cooling rate of O.S’C/h), that had been immersed for three weeks in SBF, is shown in Fig. 10. A 5 pm thick layer of HA has formed on the surface and has been confirmed by p-Raman spectroscopy [Fig. 8(d)]. This thick layer of HA is built up over the HA lamellae previously formed and even penetrates between them. Daily monitoring of pH and of the ionic concentration of calcium, silicon and phosphate are presented in Fig. 11. A substantial increase in the Si +4 concentration, at a rate of - + 0.42 ppm/day, is observed during the first seven days of immersion,
Fig. 7. SEM image of a polished cross-section of the eutectic plate after one week soaking in SBF and elemental X-ray maps of Si, P, and Ca.
2546
De AZA ef al.:
BIOACTIVE MATERIAL WHICH TRANSORMS INTO HYDROXYAPATITE
b
C
i
d
100
300
500
700.
900
1100
100
300
100
700
9
Raman shift (cm-i) Fig. 8. p-Raman spectra of: (a) standard TCP; (b) standard HA; (c) the reaction zone in Fig. 7; (d) the surface of the sample after one week soaking in SBF (Fig. 9).
after which it increases very slowly. This result justifies the dissolution of the W in the SBF. The (P04)3- concentration diminishes slowly, during the same period of time, at a constant rate of - - 0.23 ppm/day. The Cat2 concentration slowly increases at a constant rate of - + 0.13 ppm/day. Lastly, the pH of the SBF remains essentially constant during the entire experiment, changing only from 7.25 to 7.35. In Fig. 11 is also shown the pH variation at the interface between the eutectic material and the SBF. At the beginning of the reaction, the pH quickly increases to 8.5 and then stabilizes.
Fig. 9. SEM image of the surface
4. DISCUSSION
After
immersion
of the eutectic
in the SBF, two different periphery
of the sample.
tion of the material solution formation
of the
stituted
with the SBF causing
the dis-
lamellae
and
the
subsequent
by a pseudomorphic
of the TCP, and the second
by a superficial reaction,
on the by reac-
deposit
sition of HA is less abundant of the
material
The first is formed
wollastonite
of HA
transformation
ceramic
zones are formed
but
becomes
during
the first stage
a thick
layer.
of the sample after one week soaking
is con-
of HA. This depo-
in SBF.
and
dense
De AZA et al.:
Fig.
BIOACTIVE
MATERIAL
WHICH
TRANSORMS
INTO
HYDROXYAPATITE
10. SEM image of a polished cross-section of the eutectic material after three weeks soaking SBF. (A) A 5 pm HA layer; (B) HA lamellae; (C) original eutectic ceramic material.
It is worth emphasizing that no layer of amorphous silica was observed, as is normally detected in reactions of silica-based bioactive materials with SBF. The results obtained can be explained through the following mechanism (Fig. 12): At the beginning of the experiment two nearly simultaneous reactions occur. First, the W phase in contact with the SBF starts to react via an ionic exchange of 2H30+ from the SBF for one Ca*+ from the W network. This exchange transforms the crystalline W into an
2547
in
amorphous silica phase as has been demonstrated in previous works [12-14,211. This reaction starts at the surface of the material, thereby causing the pH at the material/SBF interface to increase to 8.5 and releasing calcium and silicon ions into the SBF solution. The reaction progresses deeply into the material, in the confined channels between the TCP lamellae as W is dissolved. This reaction triggers the next events. Immediately, the TCP lamellae start to react with the Ca*+ and OH- ions present in the confined
Si” @pm) 1
10
8 6
(PO.)’
(Ppm) 30
loo_.
___‘_-
-
1:
-
95 _~
90
2
4
6
8
10 12
time(days)
14
16
25
z
I
18
2
4
6
8
10
12
14
16
16
time (days)
Fig. 11. Changes in Ca’+, (P0,)3- and Si4+ concentration in SBF during soaking of the eutectic material. Changes in pH during exposure to SBF at the interface between the material and the SBF (0) and away from the interface (u).
2548
De AZA et al.:
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SBF 2H,O+
2H,O’
: Ca2
: Ca’+
0- spi-OH
O- i-OH P 0
b
0
P OH-j?i-0
OH- i-0 1 0
b pH=lOS
W
pH=lOS
TCP
W
TCP
W
TCP
W
TCP
3 [Ca,(PO,),]+Ca"+ 2(OH)I)Ca,,(P04),(OH), 6 [HPOi]+lO Ca2't8(0.H)I)Ca,,[P0,],[OH],+6H20 Fig. 12. Schematic representation of HA formation mechanism. channels, at pH = 10.5 [21]. Thus a pseudomorphic transformation of TCP into HA occurs as shown in Fig. 12. The silicon and calcium ions, not involved in the reaction, migrate through the SBF solution away from the interface and are monitored as an increase in concentration (Fig. 11). Calculations, based on the weight percent of both phases in the eutectic and on the quantity of W dissolved in the SBF, determine that -85 wt% of the calcium contained in the dissolved W should be in the SBF. Since a large increase was not detected (Fig. ll), most of Ca2+ must react with the phosphate of the SBF at the material/SBF interface and lead to the precipitation of HA on the surface of the material. This is the second reaction schematically shown in Fig. 12. Initially the precipitation occurs on the newly formed HA lamellae (Fig. 9). Later, as the precipitation increases (Fig. lo), the HA eventually covers the entire surface of the sample. The final effect of the combined reactions is to produce a large increase in Si+4 concentration in the SBF solution, a small increase in Ca2+ and a decrease of (P04)3- as detected in Fig. 11. The first step of the reaction mechanism, which at the beginning is very fast, is slowed down due to the combined effect of the difficulty of diffusion of protons from the SBF to reach the reaction interface as the distance is increased and due to the closure of channels as HA is formed on the surface of the sample (Fig. 11). 5. CONCLUSIONS Eutectic microstructures have been obtained from the system wollastonite-tricalcium phosphate by radial heat extraction and slow cooling inside the furnace. The microstructure is comprised of quasi-
spherical colonies of alternating radial lamellae of both phases. This microstructure may be modified for specific applications of this material. The eutectic ceramic material, when soaked in SBF, reacts first by dissolving the wollastonite phase and immediately forming a porous structure of hydroxyapatite by a pseudomorphic transformation of the tricalcium phosphate lamellae. The microstructure obtained is similar to that of porous bone. Later, a dense hydroxyapatite layer is formed by precipitation on the outer surface of the material. It is expected that this material will behave similarly in in viva experiments facilitating the osseointegration of the implant. The procedure developed by the authors opens the opportunity to obtain a new family of bioactive materials for which the general name of bioeutectics@ is proposed. Acknowledgements-This work was supported by CICYT under Project MAT97-1025. P.N.D.A. thanks the Ministry of Education and Science of Spain for the Fellowship given to her. In addition, the authors wish to thank A. P. Tomsia and E. Saiz and also Z. B. Luklinska and M. R. Anseau for the facilities given to P.N.D.A. during her respective stays at Lawrence Berkeley Laboratory (CA, U.S.A.) and at IRC in Biomedical Materials, The London Hospital, Medical College, University of London. Finally, thanks also are given to L. Artds and R. Cusch for the pRaman spectra and useful discussions.
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4. De Groot, K. and Le Geros, R., in Bioceramics Materials Characteristics vs in Vivo Behavior, ed. P. Ducheyne. J. Ann. N.Y. Acad. Sci., New York, 1988, PQ. 268-277. 5. D-e Groot, K., in Bioceramics Material Characteristics vs In Vivo Behavior. ed. P. Duchevne. J. Ann. N.Y. Acad. Sci., New York, 1988, pp. 227-235. 6. Roy, D. M. and Linnehan, S. K., Nature, 1974, 214, 220. 7. Sayler, K., Holmes, R. and Johns, D., J. Dent. Res., 1977, 56B, 173. 8. Tencer, A. F., Shors, E. C., Woodard, P. L. and Holmes, R. E., in Handbook of Bioactive Ceramics, Vol. II. Calcium Phosphate and Hydroxylapatite Ceramics, ed. T. Yamamuro, L. L. Hench and J. Wilson. CRC Press Inc., Boca Raton, FL 33431, U.S.A., 1990, pp. 209-221. 9. Sivakumar, M., Dampath Kumar, T. S., Dhantha, K. L. and Panduranga Rao, K., Biomaterials, 1996, 17, 1709. 10. Kurz, W. and Fisher, D. J., Fundamental of Solidification. Trans. Tech. Publications, Switzerland, 1984. 11. De Aza, P. N., Guitian, F. and De Aza, S., J. Am. Ceram. Sot., 1995, 78, 1653.
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12. De Aza, P. N., Guitian, F. and De Aza, S., Scripta metall. mater., 1991, 31, 1001. 13. De Aza, P. N., Guitian, F. and De Aza, S., in Advances in Science and Technology, 12. Materials in Clinical Applications, ed. P. Vicenzini, Techna Sri., Florenze, 1995, pp. 19-27. 14. De Aza, P. N., Luklinska, Z. B., Anseau, M. R., Guitian, F. and De Aza, S., J. Microsc., 1996, 182, 24-31. 15. De Groot, K., in Bioceramics of Calcium Phosphate, ed. K. De Groot. Press Inc., Boca Raton, FL 33431, U.S.A., 1983. 16. Hench, L. L., J. Am. Ceram. Sot., 1991, 74, 1487. 17. Flemings, M. C., Sci. Am., 1974, 231, 88. 18. Elliot, R., ht. Metals Rev., 1977, R219, 161. 19. Ashbrook, R. L., J. Am. Ceram. Sot., 1977, 60, 428. 20. Gamble, J., Chemical Anatomy, Physiology and Pathology of Extracellular Fluid. Harvard University Press, Cambridge, 1967. 21. De Aza, P. N., Guitian, F., Merlos, A., Lora-Tamayo, E. and De Aza, S., J. Mater. Sci. Mater. Med., 1996, 7, 399. 22. Merlos, A., Garcia, I., Cane, C., Esteve, J., Bartroli, J., Jimenez, C., Proc. 5th Conf. on Sensors and Their Applications. Edimburg, U.K., 1991, pp. 127-132.