Titanium and Strontium-doped
Phosphate Glasses as Vehicles
for Strontium Ion Delivery
to Cells
NILAY LAKHKAR,* ENSANYA A. ABOU NEEL, VEHID SALIH AND
JONATHAN CAMPBELL KNOWLES
University College London, Eastman Dental Institute, London, UK
ABSTRACT: This study investigated the use of a Ti-containing quaternary
phosphate glass system P2O5–Na2O–CaO–TiO2 as a vehicle for strontium ion
delivery to cells. Four glass compositions were manufactured: 0.5P2O5–
0.15Na2O–0.05TiO2–(0.3 x)CaO–xSrO (x ¼ 0, 0.01, 0.03, and 0.05). Structural
characterization revealed that sodium calcium phosphate is the dominant phase
in all the glasses. Degradation studies demonstrated highly linear glass
degradation, with Sr-containing glasses degrading at higher rates than the
Sr-free glass. Biocompatibility studies using MG63 cells showed that the
Sr-containing glasses possess excellent cell attachment and growth, particularly
over short periods (4 days).
KEY WORDS: phosphate glasses, titanium oxide, strontium oxide, biocompatibility, dental applications.
INTRODUCTION
T
he role of strontium in biological processes is a topic that has
received increasing attention in recent years, particularly from the
standpoint of its complementary relationship with calcium in vivo.
*Author to whom correspondence should be addressed. E-mail: [email protected]
Figure 1 appears in color online: http://jba.sagepub.com
JOURNAL OF BIOMATERIALS APPLICATIONS Vol. 25 — May 2011
0885-3282/11/08 0877–17 $10.00/0
DOI: 10.1177/0885328210362125
ß The Author(s), 2010. Reprints and permissions:
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N. LAKHKAR ET AL.
The ionic forms of strontium and calcium are similar in that both are
divalently charged, and their ionic radii are very close to each other
(Ca2þ: 1.00 A.U.; Sr2þ: 1.18 A.U.). From a physiological perspective,
calcium and strontium follow the same absorption and excretion
pathways (through the gastrointestinal tract and the urinary systems,
respectively) and both are primarily concentrated in bone tissue [1].
Several studies have demonstrated that strontium facilitates osteoblast
function, thereby inducing bone formation and inhibiting bone resorption [2–5]. Further research on the biological properties of strontium has
been stimulated by two major developments, namely, the emergence of
strontium ranelate as a potent drug for the treatment of osteoporosis [4]
and the application of Sr89 isotope compounds in the palliation of pain
from bone metastases of prostate cancer [6].
In this context, the development of materials as suitable carriers for
in vivo strontium ion delivery assumes particular importance. To this
day, the materials that have been investigated for this purpose include
glass ionomer cements [7], brushite-forming -TCP cements [8], calcium
silicate CaSiO3 ceramics [9], and bioactive glasses such as SiO2–CaO–
SrO glasses [10] and SrO–CaO–ZnO–SiO2 glasses [11]. Experimental
evaluations of these materials have yielded encouraging results in terms
of strontium release and biocompatibility. Ternary P2O5–Na2O–CaO
phosphate glasses, which have been extensively researched in our
laboratory on account of their highly controllable degradation properties
and compositional affinity to the mineral phase of bone [12–14], can be
considered very promising candidates to serve as vehicles for in vivo
strontium ion release. Ternary phosphate glasses have hitherto been
doped with many metal oxides such as Fe2O3 [15,16], ZnO [17], Ag2O
[18], and TiO2 [19–21]. It has been demonstrated that ternary
phosphate glasses doped with metal oxides can degrade at rates ranging
over several orders of magnitude, from 2–3 h to over 1 year, and that the
released metal ions exert antimicrobial and therapeutic effects. In
particular, TiO2 addition significantly reduces the degradation rate of
otherwise highly soluble ternary glasses, and phosphate glasses containing TiO2 elicit a more favorable response from bone cells as compared to
ternary glasses; consequently, they are of great interest for developing
synthetic bone substitutes to combat bone tissue loss caused by injury,
disease, or congenital defects.
In a previous study, Abou Neel et al. [22] investigated the structure
and properties of quaternary P2O5–Na2O–CaO–SrO glasses; however,
these glasses tended to degrade at excessively high rates and therefore
did not possess favorable biocompatibility. To reduce the degradation
rates to more acceptable levels, Lakhkar et al. [23] doped ternary glasses
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Ti and Sr-doped Phosphate Glasses as Vechicles for Sr Ion Delivery 879
with both SrO and TiO2 in a preliminary study, and they investigated
the physicochemical properties and biocompatibility of the five-component P2O5–Na2O–CaO–TiO2–SrO glasses thus formed. Their glasses
degraded much more slowly than the quaternary P2O5–Na2O–CaO–SrO
glasses and exhibited considerably enhanced cell viability. The end
objective of their study was to produce glasses that could simultaneously
exert effective therapeutic effects for orthopaedic, dental, and maxillofacial applications. At the time, their results indicated that the
properties of the glasses could be further optimized by changing the
metal oxide content (particularly that of TiO2, which was 3 mol%). To
that end, in the present study, we increased the TiO2 content from 3 to
5 mol% and investigated the effects of adding varying amounts of SrO on
the structural properties, degradation behavior, and biocompatibility of
the resulting P2O5–Na2O–CaO–TiO2–SrO glasses.
MATERIALS AND METHODS
Preparation of Glass Samples
The glass samples were prepared in the form of rods with diameters of
15 mm by a previously described melt quenching method [21]. The
following precursors (all with purity of 98% or greater) were used:
phosphorus pentoxide (P2O5) (Fisher Scientific, Loughborough, UK),
calcium carbonate (CaCO3), sodium dihydrogen orthophosphate
(NaH2PO4), titanium dioxide (TiO2), and strontium carbonate (SrCO3;
BDH, Poole, UK) (see Table 1 for glass codes and compositions). The
required amounts of all the precursors were weighed, mixed, and heated
in two different types of crucibles depending on the glass composition.
For the Sr-free glass, a Pt/10% Rh type 71040 crucible (Johnson
Matthey, Royston, UK) was used, while each Sr-containing glass was
heated in a separate vitreous silica crucible (Saint-Gobain Quartz, Tyne
& Wear, UK). The precursor mixture was initially preheated at 7008C
Table 1. Glass compositions and codes used.
Glass composition (mol%)
Glass code
P50C30N17Ti3Sr0
P50C29N17Ti3Sr1
P50C27N17Ti3Sr3
P50C25N17Ti3Sr5
(Sr0)
(Sr1)
(Sr3)
(Sr5)
P2O5
CaO
Na2O
TiO2
SrO
50
50
50
50
30
29
27
25
15
15
15
15
5
5
5
5
0
1
3
5
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N. LAKHKAR ET AL.
for 10 min and then melted at 13008C for 3 h. After melting, annealing
was carried out in a preheated graphite mould at 4258C for 2 h to remove
residual stresses, followed by cooling overnight at ambient temperature.
A Testbourne diamond saw (Testbourne, Basingstoke, UK) was used to
cut the glass rods into discs with thicknesses of approximately 2 mm;
methanol was used as the coolant and lubricant.
Structural Characterization
The structural characterization of the glasses was carried out by
differential thermal analysis (DTA), X-ray diffraction (XRD), and hightemperature XRD (HTXRD). The methods and experimental parameters
used for DTA and XRD have been described elsewhere [21]. The DTA
experiments were carried out using a Setaram Differential Thermal
Analyzer (Setaram, France), and the following parameters were
measured: glass transition temperature (Tg), crystallization temperature
(Tc), and melting temperature (Tm). The XRD analyses were performed
using a Brüker D8 Advance Diffractometer (Brüker, Coventry, UK) and a
LynxEye detector (Brüker, Coventry, UK) in flat-plate geometry with
Ni-filtered Cu K radiation. For the HTXRD analyses, the general
method described by Pickup et al. [24] was followed. Data were collected
at 2 values in the range 208–408 for a step size and counting time of 0.028
and 0.1 s per point, respectively. The temperature range used was from
ambient to 11008C at a heating rate of 218C min1, and helium was used
as the purge gas to reduce incident and scattered X-ray absorption.
Degradation and Ion Release
Degradation behavior was studied by measuring weight loss and ion
release over a 15-day period using a simple aqueous model in accordance
with a previously described method [21]. The following time points were
used: 25, 50, 95, 166, 264, and 351 h. Glass disks immersed in plastic
containers holding 25 mL of high-purity deionized water (resistivity
¼ 18.2 M cm1) were incubated at 378C. At each time point, the solution
from each container was removed and stored for further analyses.
The discs were dried, weighed, immersed in fresh deionized water, and
reintroduced in the incubator. Polyphosphate anion and Naþ and Ca2þ
cation release were measured by ion chromatography; a Dionex ICS-2500
ion chromatography system (Dionex, Surrey, UK) was used for the
phosphate anions, whereas a Dionex ICS-1000 ion chromatography
system was used for the cations [21]. Further, inductively coupled
plasma mass spectroscopy (ICP-MS; Spectromass 2000 ICP mass
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Ti and Sr-doped Phosphate Glasses as Vechicles for Sr Ion Delivery 881
spectrometer, Spectro, Germany) was employed to measure Ti(IV) and
Sr2þ ion release [21]. Anion chromatography and ICP-MS did not require
the analyte to be purified further; however, for cation chromatography, it
was necessary to filter the solution using a Dionex OnGuard IIA cartridge
to eliminate anions that bind to the cation column.
Cell Culture Studies
Cell culture studies were conducted in accordance with Chen et al.’s
methods with some modifications [25]. Briefly, MG63 osteoblast-type
cells were seeded onto polished and sterilized glass discs at a seeding
density of 5000 cells per disc, followed by incubation at 378C in a 5% CO2
atmosphere for 7 days; standard glass cover slips were used as the control.
Cell proliferation experiments were conducted at time points of 1, 4, and 7
days using the AlamarBlueTM assay (AbD Serotec, UK). Cells were
prepared for scanning electron microscopy (SEM) by dehydration in a
graded series of alcohols and hexamethyldisilazane prior to gold/platinum
coating. SEM images were obtained at 1 and 7 days using a JEOL model
JSM 5410LV SEM (JEOL, USA) at an accelerating voltage of 25 kV [25].
RESULTS
Structural Characterization
Figure 1(a) and (b) show combination plots of Sr0 and Sr1 glasses,
respectively, where for each composition, the DTA trace, normal XRD
profile, and contour HTXRD plot have been included in the same graph.
The combination plots of the Sr3 and Sr5 glasses (not included in this
study) are similar to the Sr1 glass plot, and the results obtained for the
Sr3 and Sr5 glasses are briefly explained later in the text.
The DTA traces of the investigated glasses revealed that the Tg values
increased with the Sr content: the Sr0–5 glasses showed Tg values of
4368C, 4378C, 4408C, and 4438C, respectively (note that the difference in
Tg values is not significant). The Sr-free glass showed two crystallization
peaks at temperatures of 6268C and 6838C. In contrast, the Srcontaining glasses showed only one crystallization peak, which was
notably broader than the corresponding two peaks in the Sr-free glass,
at Tc values in the range 6608C–6688C (Tc ¼ 6608C for the Sr1 glass).
Both Sr-free and Sr-containing glasses showed melting peaks at Tm
values of 7638C and 7188C–7258C, respectively (Tm ¼ 7258C for the Sr1
glass). All the investigated glasses showed a small crystallization peak
immediately after the melting peak.
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N. LAKHKAR ET AL.
(a)
1100
HTXRD plot
1000
900
Tc3
Temperature (°C)
800
Tm
Tc2
700
600
Tc1
DTA trace
500
Tg
400
XRD plot
300
200
110
20
21
22
23
24
25
26
27
28
29
31
30
32
33
34
35
36
37
38
39
35
36
37
38
39
2q Scale
(b)
HTXRD plot
1000
900
Temperature (°C)
800
Tc2
Tm
700
Tc1
600
500
Tg
DTA trace
400
300
XRD plot
200
100
60
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
2q Scale
Figure 1. Combination plots of (a) Sr0 and (b) Sr1 glasses. For both compositions,
DTA trace, normal XRD profile, and contour HTXRD plot are included in the same graph.
Normal XRD studies were performed using powders of glass samples
crystallized at temperatures close to the respective Tc values obtained in
the DTA results, while uncrystallized glass samples were utilized for the
HTXRD studies. (Two HTXRD peaks at approximately 298 and 398 and
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Ti and Sr-doped Phosphate Glasses as Vechicles for Sr Ion Delivery 883
shifting toward lower 2 values can be observed; these can be attributed
to the underlying platinum electrode, and they tend to fluctuate
according to the thickness of the glass specimen on top of the electrode.)
The results of the DTA, XRD, and HTXRD studies were closely related to
each other for all the glass samples. To explain using the Sr0 plot, the first
crystallization peak at 6268C corresponded to a series of crystallization
events in the HTXRD plot at 2 values of 23.48, 25.68, 26.58, 28.08, 29.98,
and 30.78. Normal XRD analysis of Sr0 powder crystallized at 6208C
revealed that sodium calcium phosphate (NaCa(PO3)3; ICDD no. 231699) is formed at approximately the same 2 values. Thus, it is
reasonable to consider that the crystallization peak at 6268C corresponds
to the formation of NaCa(PO3)3 as the dominant phase in the glass.
Using a similar approach, we found that the second crystallization
peak at 6838C corresponded to the formation of sodium titanium
phosphate (Na4TiO4; ICDD no. 25-1297) and a third, smaller crystallization peak at 8108C to titanium phosphate (TiPO4; ICDD no. 381468), although the correspondence between the HTXRD and DTA plots
was not clear for the third peak. Similar analyses of the Sr-containing
glasses revealed that the same phases are formed but at higher
temperatures. Glasses with higher Sr contents recorded fewer events
in the HTXRD contour plots.
Degradation and Ion Release
For the reasons mentioned in a previous study on phosphate glasses
[21], the percentage weight loss per unit surface area was calculated as
follows and analyzed as a function of time:
½ðM0 Mt Þ=M0 A 100,
ð1Þ
where M0 is the original weight of the glass disc (mg), Mt is the weight of
the disc at time t, and A is the surface area of the glass disc (mm2).
In all the investigated glasses, the degradation process increased with
time; more interestingly, the Sr-containing glasses showed greater
degradation than the Sr-free glass over the same time period (Figure 2).
The slope of the trend line plotted for each composition provides the
degradation rate (Table 2). The addition of 1 mol% Sr to a Sr-free glass
caused the degradation rate to increase by close to an order of
magnitude, from 0.6 105% mm2 h1 to 4.6 105% mm2 h1.
Further addition of Sr decreased the degradation rate to 2.7 105%
mm2 h1 for the Sr3 glass and 2.5 105% mm2 h1 for the Sr5 glass.
This trend could be algebraically represented as Sr05Sr55Sr35Sr1.
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884
Cumulative weight loss per SA (%mm–2)
N. LAKHKAR ET AL.
0.020
0.018
y = 4.6127E–05x
R2 = 9.9704E–01
0.016
0.014
0.012
y = 2.7212E–05x
R2 = 9.9647E–01
0.010
y = 2.4893E–05x
R2 = 9.9498E–01
0.008
0.006
0.004
y = 6E–06x
R2 = 09811
0.002
0.000
0
50
100
150
Sr1
250
200
Time (h)
Sr3
Sr5
300
350
400
Sr0
Figure 2. Degradation presented as percentage cumulative weight loss per unit surface
area (%mm2) as a function of time for Ti-containing glasses with different Sr contents.
Table 2. Cumulative degradation rate (%mm2 h1) and anion, cation
(Naþ and Ca2þ), Ti4þ, and Sr2þ release rates (ppm h1) determined
from the slopes of the linear fit against time for Ti-containing phosphate
glasses with different SrO contents.
Glass code
2
Degradation rate (%mm
Anion (ppm h1)
Cation (ppm h1)
Ti4þ (ppm h1)
Sr2þ (ppm h1)
1
h
5
10 )
PO3
4
P3 O3
9
P2 O4
7
P3 O5
10
2þ
Ca
Naþ
Sr0
Sr1
Sr3
Sr5
0.6
0.0192
0.0413
0.0039
0.0161
0.0439
0.0271
0.0067
N.A.
4.6
0.0943
0.1714
0.0910
0.1646
0.2383
0.1314
0.0291
0.0127
2.7
0.0892
0.0974
0.0974
0.1562
0.1566
0.0962
0.0306
0.0355
2.5
0.0834
0.0800
0.0902
0.1449
0.1274
0.0877
0.0245
0.0443
All the glasses were found to degrade in a highly linear fashion, with the
goodness-of-fit (R2) values exceeding 0.98.
A number of similarities were observed between the cumulative
release profiles of Naþ and Ca2þ cations (Figure 3(a) and (b)), metallic
Ti(IV) and Sr2þ ions (Figure 4(a) and (b)), and polyphosphate anions
3
4
5
(PO3
4 , P3 O9 , P2 O7 , and P3 O10 ). (The results for the polyphosphates
were obtained in a manner similar to those of the cations and metallic
ions; the polyphosphate release rates have been tabulated in Table 2).
For all the investigated ions, the release rates, as given by the slope of
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Ti and Sr-doped Phosphate Glasses as Vechicles for Sr Ion Delivery 885
(a) 50
y = 0.131x
R2 = 0.912
Cumulative Na+ release (ppm)
45
40
35
y = 0.096x
R2 = 0.925
30
y = 0.087x
R2 = 0.988
25
20
15
y = 0.027x
R2 = 0.957
10
5
0
0
50
100
150
200
250
300
350
400
Time (h)
Sr1
Sr3
Sr5
Sr0
Cumulative Ca2+ release (ppm)
(b) 90
y = 0.238x
R2 = 0.927
80
70
60
y = 0.156x
R2 = 0.930
50
y = 0.127x
R2 = 0.987
40
30
20
y = 0.043x
R2 = 0.971
10
0
0
50
100
150
200
250
300
350
400
Time (h)
Sr1
Sr3
Sr5
Sr0
Figure 3. Cumulative release (ppm) of (a) Naþ and (b) Ca2þ ions as a function of time for
Ti-containing glasses with different Sr contents.
the linear fit of ion release over time, increased with time. In general, the
release rates of the Sr-containing glasses were very close to each other
and were substantially higher than the release rate of the Sr-free glass.
Similar to the weight loss study, the ion release studies revealed
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886
N. LAKHKAR ET AL.
Cumulative Ti4+ release (ppm)
(a) 12
y = 0.030x
R2 = 0.915
10
y = 0.029x
R2 = 0.980
8
y = 0.024x
R2 = 0.950
6
4
y = 0.006x
R2 = 0.933
2
0
0
50
100
150
Sr1
200
Time (h)
Sr3
250
Sr5
300
350
400
Sr0
Cumulative Sr2+ release (ppm)
(b) 18
y = 0.044x
R2 = 0.949
16
14
y = 0.035x
R2 = 0.897
12
10
8
6
y = 0.012x
R2 = 0.959
4
2
0
0
50
100
150
200
250
300
350
400
Time (h)
Sr1
Sr3
Sr5
Figure 4. Cumulative release (ppm) of (a) Ti(IV) and (b) Sr2þ ions as a function of time
for the investigated glasses.
considerably high R2 values, in excess of 0.9. The Sr05Sr55Sr35Sr1
trend was evident for all the ionic species other than the metallic ions
2þ
(Ti(IV) and Sr2þ) and the P2 O4
ions, the ion release
7 anion. For the Sr
rate increased with the Sr content of the glass as given in Table 2.
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Ti and Sr-doped Phosphate Glasses as Vechicles for Sr Ion Delivery 887
% Change over control
250
200
150
100
50
0
Sr0
Sr3
Sr1
Sr5
Sample
Day 1
Day 4
Day 7
Figure 5. Histogram showing percentage change in cell growth relative to the control.
Cell Culture Studies
The results of the cell proliferation assay can be represented in the
form of a histogram (Figure 5) that shows the percentage change in cell
growth relative to the control, where cell growth on the control is fixed
as 100%, by using the following formula for each time point:
%Ci ¼ 100 þ ðCsi Cci Þ=Cci 100 , i ¼ 1, 4, 7,
ð2Þ
where %Ci is the percentage change in cell growth relative to the
control, Csi is the cell growth on the sample, Cci is the cell growth on the
control, and i is the time point under consideration.
Clearly, as early as day 1, all the investigated glass specimens showed
cell growth at levels very close to that of the control. On day 4, the Srfree sample showed somewhat larger cell growth than the control, but
the Sr-containing glass samples showed a remarkably superior capacity
to nurture cells on the sample surface. The cell growth values on day 4
showed the trend Sr05Sr55Sr35Sr1. By day 7, all the samples again
showed cell growths comparable to that of the control.
Figure 6 shows SEM images demonstrating the attachment of MG63
cells on days 1 and 7, respectively, when seeded on (a, f) Sr0 glass, (b, g)
Sr1 glass, (c, h) Sr3 glass, (d, i) Sr5 glass, and (e, j) control glass cover
slips. The SEM images provided clear evidence of a marked increase in
cell growth and cell attachment over the 7-day time period. Visual
examination of the cell morphology on each specimen revealed that the
cells were already spread out on the surface on day 1, and by day 7, the
surfaces of all the specimens were covered with a confluent layer of cells.
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888
N. LAKHKAR ET AL.
Figure 6. SEM images showing attachment of MG63 cells on days 1 and 7, respectively,
when seeded on (a, f) Sr0 glass, (b, g) Sr1 glass, (c, h) Sr3 glass, (d, i) Sr5 glass, and (e, j)
control glass cover slips. White shapes in some of the images correspond to precipitates
from sodium cacodylate used for fixing the specimens.
DISCUSSION
In a preliminary study on five-component glasses having compositions
of 0.5P2O5–0.17Na2O–0.03TiO2–(0.3 x)CaO–xSrO (x ¼ 0, 0.01, 0.03,
and 0.05), Lakhkar et al. [23] had obtained encouraging results in terms
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Ti and Sr-doped Phosphate Glasses as Vechicles for Sr Ion Delivery 889
of controlled degradation and biocompatibility. Their Sr-containing
glasses degraded at a rate that was optimal for use in dental applications
and showed favorable capabilities to nurture cells on the glass surface.
To that end, the present study endeavored to further optimize the
properties of these glasses by increasing the TiO2 content, at the expense
of the Na2O content, from 3 to 5 mol%.
The increase in the Ti content of the glasses offers some interesting
results. Most notably, the 5 mol% Ti glasses exhibit, on average, about
2–4 times lower degradation and ion release rates than the 3 mol% Ti
glasses. In addition, a comparison of the cell proliferation assay results
reveals that the 5 mol% Ti glasses have a greater number of attached
cells than the 3 mol% Ti glasses over the 7-day time period. Both
observations serve as proof of a well-known fact: glasses that degrade
more slowly tend to exhibit greater number of attached cells.
A major development in this study is the identification of the
dominant phases present in the glass at elevated temperatures (up to
slightly above the melting point of the glass). As mentioned by Pickup
et al. [24], the DTA/XRD/HTXRD combined plots facilitate an accurate
assessment of the thermal behavior of the glass. The correlation
between three distinct sets of experimental results is remarkable and,
to our knowledge, has not been demonstrated for phosphate-based
glasses thus far. The occurrence of the same dominant phases in all the
investigated glasses can be attributed to two facts: (A) the maximum
SrO content is only 5 mol% and (B) Sr2þ ions are added to the glass at
the expense of equivalently charged Ca2þ ions. Thus, the glass phases
will not be particularly affected at such low levels of oxide addition
involving the replacement of equivalently charged ions.
Several salient observations from all the experimental studies point to a
significant alteration of the network structure of the quaternary P2O5–
Na2O–CaO–TiO2 glass by the addition of 1 mol% SrO. These include (A)
the transformation of the two sharp crystallization peaks in the Sr0 glass
to a single, broader peak in the Sr1 glass with a considerable increase in
the Tc value; (B) the significant decrease in Tm from 7638C for the Sr0
glass to 7258C for the Sr1 glass; (C) the formation of the same
crystallization phases in the Sr1 glass as those in the Sr0 glass, but at
higher temperatures; (D) the considerably lower solubility and ion release
rates of the Sr0 glass as compared to those of the Sr1 glass; and (E) the
substantial increase in the cell growth value from Sr0 to Sr1 on day 4.
As explained by Abou Neel et al. [21], four-component P2O5–Na2O–
CaO–TiO2 systems are characterized by the presence of TiO5 or TiO6
titanate polyhedra between adjacent phosphate glass chains in the glass
network. These titanate polyhedra strengthen the glass network because
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N. LAKHKAR ET AL.
of two reasons. First, the Ti(IV) ion has a smaller ionic radius than either
the Naþ or Ca2þ ions (Ti(IV): 0.61 A.U.; Naþ: 1.02 A.U.; Ca2þ: 1.00 A.U.).
Second, the combined charge of the Naþ and Ca2þ ions is less than
the charge of the Ti(IV) ion by þ1. However, when SrO (Sr2þ: 1.18 A.U.)
is added to this system, the larger ionic radius of Sr2þ as compared to
the other ions plays a role in perturbating the atomic arrangement of
the glass network, thereby weakening the glass structure and increasing
its solubility.
The above five observations serve as a reference for another indication:
increases in the SrO content of the glass from 1 to 3 and 5 mol% did not
alter the glass properties to the same extent. The Sr05Sr55Sr35Sr1
trend is conspicuous in many of our experimental results. Further study
is required in order to elucidate the exact mechanism governing this
trend. However, the results clearly suggest that 1 mol% SrO serves as a
threshold concentration for maximum alteration of the structure and
properties of the quaternary P2O5–Na2O–CaO–TiO2 glass.
The results of the cell proliferation assay (and also the SEM images)
offer conclusive evidence of the utility of these glasses in short-term
biomedical applications (of the order of 4 days). Furthermore, the
high R2 values obtained in the degradation and ion release studies
confirm that our glasses can definitely be considered as potential
candidate materials for use in controlled and sustained antimicrobial ion
delivery systems. Although the overall results indicate that 1 mol% SrO
would be the ideal Sr content for the five-component system, the choice
of SrO content is dependent on the objectives of the end application. If
the objective is to, as much as possible, accelerate glass degradation
and provide enhanced biocompatibility over a short time span, then
1 mol% SrO seems to be the optimum amount of dopant for the
investigated quaternary system. On the other hand, if the objective is to
exert a therapeutic, or specifically antimicrobial, effect through the
release of Sr2þ ions, then further SrO addition is advisable since, as
observed earlier, the Sr2þ ion release increases with the SrO content of
the glass.
CONCLUSIONS
In conclusion, the effect of SrO addition on the structure, degradation,
and biocompatibility of quaternary P2O5–Na2O–CaO–TiO2 phosphate
glasses was studied. Structural characterization revealed the transition
of the main phase in the glasses with an increase in temperature, from
sodium calcium phosphate (NaCa(PO3)3) at 6268C onwards to sodium
titanium phosphate (Na4TiO4) at 6838C onwards and titanium
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Ti and Sr-doped Phosphate Glasses as Vechicles for Sr Ion Delivery 891
phosphate phase (TiPO4) at 8108C onwards. Degradation studies
confirmed that Sr-containing glasses degrade at much greater rates
than glasses doped with TiO2 alone, with 1 mol% SrO being the optimum
dopant content for increasing the degradation rate; furthermore, the
degradation process was highly linear, thereby raising interesting
possibilities for using these glasses in applications involving controlled
and sustained delivery of antimicrobial ions. Cell culture studies
conducted using MG63 cells demonstrated that over a 7-day period, all
the Sr-containing glasses possess improved biocompatibility in comparison with standard glass cover slips used as controls. The
Sr05Sr55Sr35Sr1 trend is conspicuous in many of the results and
is deserving of further study. Taken together, the results demonstrate
that four-component P2O5–Na2O–CaO–TiO2 phosphate glasses doped
with SrO can serve as effective delivery vehicles for releasing Sr ions. In
view of the antimicrobial properties of SrO, the investigated glasses offer
considerable potential for use in various orthopaedic, dental, and
maxillofacial applications.
REFERENCES
1. Wren, A., Boyd, D. and Towler, M.R. The Processing, Mechanical Properties
and Bioactivity of Strontium Based Glass Polyalkenoate Cements, J. Mater.
Sci. Mater. Med., 2008: 19: 1737–1743.
2. Guo, D., Xu, K., Zhao, X. and Han, Y. Development of a Strontium Containing
Hydroxyapatite Bone Cement, Biomaterials, 2005: 26: 4073–4083.
3. Wong, C.T., Chen, Q.Z., Lu, W.W., et al. Ultrastructural Study of
Mineralization of a Strontium-containing Hydroxyapatite (Sr-HA) Cement
In Vivo, J. Biomed. Mater. Res., 2004: 70A: 428–435.
4. Marie, P.J., Ammann, P., Boivin, G. and Rey, C. Mechanisms of Action and
Therapeutic Potential of Strontium in Bone, Calcif. Tissue Int., 2001: 69:
121–129.
5. Verberckmoes, S.C., De Broe, M.E. and D’Haese, P.C. Dose-Dependent
Effects of Strontium on Osteoblast Function and Mineralization, Kidney Int.,
2003: 64: 534–543.
6. Kraeber-Bodéré, F., Campion, L., Rousseau, C., Bourdin, S., Chatal, J.-F. and
Resche, I. Treatment of Bone Metastases of Prostate Cancer with Strontium89 Chloride: Efficacy in Relation to the Degree of Bone Involvement, Eur. J.
Nucl. Med., 2000: 27: 1487–1493.
7. Guida, A., Towler, M.R., Wall, J.G., Hill, R.G. and Eramo, S. Preliminary
Work on the Antibacterial Effect of Strontium in Glass Ionomer Cements,
J. Mater. Sci. Lett., 2003: 22: 1401–1403.
8. Pina, S., Torres, P.M., Goetz-Neunhoeffer, F., Neubauer, J. and Ferreira,
J.M.F. Newly Developed Sr-substituted -TCP Bone Cements,
Acta Biomater., 2009: 6: 928–935.
Downloaded from jba.sagepub.com at PENNSYLVANIA STATE UNIV on September 16, 2016
892
N. LAKHKAR ET AL.
9. Wu, C., Ramaswamy, Y., Kwik, D. and Zreiqat, H. The Effect of Strontium
Incorporation Into CaSiO3 Ceramics on their Physical and Biological
Properties, Biomaterials, 2007: 28: 3171–3181.
10. Lao, J., Jallot, E. and Nedelec, J.M. Strontium-Delivering Glasses With
Enhanced Bioactivity: A New Biomaterial for Antiosteoporotic Applications?
Chem. Mater., 2008: 20: 4969–4973.
11. Towler, M., Boyd, D., Freeman, C., Brook, I.M. and Farthing, P. Comparison
of In Vitro and In Vivo Bioactivity of SrO-CaO-ZnO-SiO2 Glass Grafts,
J. Biomater. Appl., 2009: 23: 561–572.
12. Franks, K., Abrahams, I. and Knowles, J.C. Development of Soluble Glasses
for Biomedical Use Part I: In Vitro Solubility Measurement, J. Mater. Sci.
Mater. Med., 2000: 11: 609–614.
13. Salih, V., Franks, K., James, M., Hastings, G.W., Knowles, J.C. and Olsen, I.
Development of Soluble Glasses for Biomedical Use Part II: The Biological
Response of Human Osteoblast Cell Lines to Phosphate-based Soluble
Glasses, J. Mater. Sci. Mater. Med., 2000: 11: 615–620.
14. Knowles, J.C. Phosphate Based Glasses for Biomedical Applications,
J. Mater. Chem., 2003: 13: 2395–2401.
15. Yu, X.Y., Day, D.E., Long, G.J. and Brow, R.K. Properties and Structure of
Sodium-Iron Phosphate Glasses, J. Non-Cryst. Solids, 1997: 215: 21–31.
16. Ray, C.S., Fang, X., Karabulut, M., Marasinghe, G.K. and Day, D.E. Effect of
Melting Temperature and Time on Iron Valence and Crystallization of Iron
Phosphate Glasses, J. Non-Cryst. Solids, 1999: 249: 1–16.
17. Abou Neel, E.A., O’Dell, L.A., Smith, M.E. and Knowles, J.C. Processing,
Characterisation, and Biocompatibility of Zinc Modified Metaphosphate
Based Glasses for Biomedical Applications, J. Mater. Sci. Mater. Med., 2008:
19: 1669–1679.
18. Valappil, S.P., Knowles, J.C. and Wilson, M. Effect of Silver-doped
Phosphate-based Glasses on Bacterial Biofilm Growth, Appl. Environ.
Microbiol., 2008: 74: 5228–5230.
19. Navarro, M., Ginebra, M.-P. and Planell, J.A. Cellular Response to Calcium
Phosphate Glasses with Controlled Solubility, J. Biomed. Mater. Res. A,
2003: 67A: 1009–1015.
20. Rajendran, V., Gayathri Devi, A.V., Azooz, M. and El-Batal, F.H.
Physicochemical Studies of Phosphate Based P2O5–Na2O–CaO–
TiO2 Glasses for Biomedical Applications, J. Non-Cryst. Solids, 2006:
353: 77–84.
21. Abou Neel, E.A. and Knowles, J.C. Physical and Biocompatibility
Studies of Novel Titanium Dioxide Doped Phosphate-based Glasses for
Bone Tissue Engineering Applications, J. Mater. Sci. Mater. Med., 2008: 19:
377–386.
22. Abou Neel, E.A., Chrzanowski, W., Pickup, D.M., et al. Structure and
Properties of Strontium-Doped Phosphate-Based Glasses, J. R. Soc.
Interface, 2008: 6: 435–446.
23. Lakhkar, N.J., Abou Neel, E.A., Salih, V. and Knowles, J.C. Strontium Oxide
Doped Quaternary Glasses: Effect on Structure, Degradation and
Cytocompatibility, J. Mater. Sci. Mater. Med., 2009: 20: 1339–1246.
Downloaded from jba.sagepub.com at PENNSYLVANIA STATE UNIV on September 16, 2016
Ti and Sr-doped Phosphate Glasses as Vechicles for Sr Ion Delivery 893
24. Pickup, D.M., Guerry, P., Moss, R.M., Knowles, J.C., Smith, M.E. and
Newport, R.J. New Sol–Gel Synthesis of a (CaO)0.3(Na2O)0.2(P2O5)0.5
Bioresorbable Glass and Its Structural Characterisation, J. Mater. Chem.,
2007: 17: 4777–4784.
25. Chen, Q.Z., Efthymiou, A., Salih, V. and Boccaccini, A.R. BioglassÕ Derived Glass-Ceramic Scaffolds: Study of Cell Proliferation and
Scaffold Degradation In Vitro, J. Biomed. Mater. Res. A, 2008: 84A:
1049–1060.
Downloaded from jba.sagepub.com at PENNSYLVANIA STATE UNIV on September 16, 2016