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Τhe effect of silica nanoparticles on the thermomechanical properties and degradation behavior of
polylactic acid
P Georgiopoulos, E Kontou, A Meristoudi, S Pispas and M Chatzinikolaidou
J Biomater Appl 2014 29: 662 originally published online 4 August 2014
DOI: 10.1177/0885328214545351

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Biomaterials Processing
Journal of Biomaterials Applications
2014, Vol. 29(5) 662–674
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thermomechanical properties and DOI: 10.1177/0885328214545351
jba.sagepub.com
degradation behavior of polylactic acid

P Georgiopoulos1, E Kontou1, A Meristoudi2, S Pispas2 and M Chatzinikolaidou3,4

Abstract
In this work a series of polylactic acid/SiO2 nanocomposites have been prepared by a melt mixing procedure. The
dispersion quality was examined by scanning electron microscopy. To study the degradation behavior of the polylactic
acid/nanocomposites prepared, the samples were immersed in a buffer solution at a temperature of 37 C with a pH of
7.4 for a time period of up to 23 weeks. These conditions simulate those in the human body, appropriate in medical
applications. In order to assess their suitability in biomedical applications, we investigated the biocompatibility of these
materials in terms of cell viability, growth, and morphology. A good initial cell adhesion has been detected, supporting
their potential use in bone tissue engineering applications. The hydrolytic degradation of polylactic acid, under the
prescribed conditions, was studied by the molecular weight reduction in terms of size exclusion chromatography,
whereas the progress of thermal stability of polylactic acid and polylactic acid/nanocomposites during aging was
tested by thermogravimetric analysis. The evolution of the materials’ thermomechanical properties during aging was
studied by differential scanning calorimetry, dynamic mechanical analysis, and tensile testing. The crystallization behavior
in polylactic acid and the way it is affected by the presence of nanofillers during degradation procedure has been studied
and values of 44% crystallinity increment have been found. At the specific aging conditions studied, silica nanoparticles
accelerate the degradability of polylactic acid, having a higher impact on Young’s modulus, under the specified aging
conditions, for 7 weeks and hereafter this acceleration is retarded, due to the crystallinity increment, as a result of the
molecular weight reduction.

Keywords
Biodegradation, cytotoxicity testing, nanocomposites, mechanical properties, crystallinity

Introduction polymers also create undesirable effects which must

Polylactic acid (PLA) is nowadays one of the most
important polymers, due to its combined properties,
such as biocompatibility, biodegradability, and ade-
quate mechanical performance. PLA consists of an
alternative to petrochemical-based polymers, because
it can be produced from natural sources such as corn
and wheat and is therefore related to a wide range of
medical, textile, and packaging applications. The
(bio)degradability of PLA though has both negative
and positive aspects. The positive aspect is the (bio)deg-
radation to form nonhazardous products when PLA
polymers or articles are discarded or composted after
completing their useful life.1 The negative aspect is the
degradation of PLA polymers during processing. Thus,
the same properties that make PLA polymers desirable
as replacements for nondegradable petrochemical
be overcome. For several applications the degradation
rate of PLA is still low as compared to the waste accu-
mulation rate. On the other hand, the fact that PLA
1
School of Applied Mathematical and Physical Sciences, Department of
Mechanics, National Technical University of Athens, Athens, Greece
2
Theoretical and Physical Chemistry Institute, National Hellenic Research
Foundation, Athens, Greece
3
Department of Materials Science and Technology, University of Crete,
Heraklio, Greece
4
Institute of Electronic Structure & Laser, Foundation for Research &
Technology Hellas, Heraklio, Greece
Corresponding author:
E Kontou, School of Applied Mathematical and Physical Sciences,
Department of Mechanics, National Technical University of Athens, 5
Heroes of Polytechnion, 15773 Athens, Greece.
Email: ekontou@central.ntua.gr

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Georgiopoulos et al.
degrades slowly—over a period of several weeks up to
about 1 year—is advantageous for some other applica-
tions such as biomedical, since it leads to a relatively
good shelf life.
PLA degrades during thermal processing or under
hydrolytic conditions, giving a reduction of molecular
weight that affects the final properties of the material,
such as the mechanical strength.2,3 The degradation
behavior depends strongly on the molecular weight
and the crystallinity of the PLA.4 Previous studies
have shown that degradation of PLA is enhanced by
an increase in temperature and relative humidity (RH).5
The temperature, in presence of oxygen, triggers ther-
mal oxidation, while the moisture promotes hydrolytic
degradation. PLA degradation driven by hydrolysis
needs higher temperature in order to take place
(T > 50 C).
Chemical hydrolysis of the hydrolytically unstable
backbone is the primary mechanism for degradation
of the PLA polymer. Degradation occurs first by
water penetrating the bulk of the polymer and hydro-
lyzing the ester bonds, preferentially those in the
amorphous phase, and converting long chains into
shorter water-soluble fragments.6
On the other hand, the thermomechanical and bar-
rier properties of PLA can be enhanced by the incorp-
oration either of biocompatible fibers7 or of a fairly
low amount of nanosized fillers (1–5 wt%).8–10 Many
studies have been reported on the preparation and
characterization of a wide range of PLA nanocompo-
sites.10–12 A large variety of inorganic nanoparticles,
including silica,13 layered silicates,14 hydroxyapatite,15
nanotubes,16 have been investigated for this purpose.
Among the various types of nanoparticles, layered sili-
cates, which are naturally abundant, economic, and
benign to the environment, have been most extensively
used for the preparation of PLA nanocomposites.17
Besides the thermomechanical properties, the hydro-
lytic degradation behavior of PLA is also greatly
affected by the presence of nanofillers. The manner in
which the thermomechanical properties of PLA and
PLA/nanocomposites alter over time is still an issue
of great importance. Depending on the applications,
acceleration of degradation (medical devices) is
required, while in some other cases (packaging) sup-
pression of degradation is needed. The effect of various
nanofiller types on the PLA degradation behavior has
been extensively studied18–22 and it has been reported
that the hydrolytic degradation of PLA takes place first
at the interface between PLA matrix and nanofillers
rather than in the bulk polymer.18 A complex effect
on the degradation behavior of PLA has been revealed
in Kontou et al.,19 where the effect of two different
nanofiller types (silica and montmorillonite) and their
mixtures has been studied on the degradability of PLA.
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663
The present work is complementary to our previous
works1,19 on the degradation effects on crystalline and
thermomechanical properties of PLA/nanocomposites.
In the present work, the degradation ability of PLA
reinforced with nanosilica particles at three different
contents, 2, 3, and 5% wt, under specific environmental
conditions, namely immersion in a buffer solution at
temperature of 37 C with a pH of 7.4 has been studied
experimentally. These conditions simulate those in the
human body, appropriate in medical applications. The
majority of prior art studies on the stability of PLA have
been carried out at similar physiological conditions,4,6,23
but there are still some issues that need to be addressed.
A matter of importance has been proved to be the crys-
tallization behavior in PLA and the way it is affected by
the presence of nanofillers during degradation proced-
ure. As it is mentioned in Chen et al.18 the effect of silica
on the hydrolytic degradation behavior of PLA is less
researched. Apart from this, the role of nanofillers on the
evolution of thermomechanical properties of PLA/nano-
composites at various stages of degradation has been less
examined, and this work is a contribution toward this
direction. The stability of PLA and PLA/nanocompo-
sites was monitored by a number of techniques, includ-
ing size exclusion chromatography (SEC), scanning
electron microscopy (SEM), thermogravimetric analysis
(TGA), differential scanning calorimetry (DSC),
dynamic mechanical analysis (DMA), and tensile meas-
urements. In order to assess their suitability in biomed-
ical applications, we investigated the biocompatibility of
the materials tested in terms of cell viability, growth, and
morphology. We therefore cultured MC3T3-E1 preos-
teoblastic cells on films made either from PLA or PLA
with 2% wt silica nanocomposite, visualized the morph-
ology of the adhered cells, and quantified the cell prolif-
eration on them.
Materials
PLA was employed for the preparation of the nano-
composites. The selected grade of PLA (supplied by
Nature works) has a D content of 4.25%, a residual
monomer content of 0.3%, and a density of 1.24 g/
cm3. The material in pellets form was dried at 45 C
for a minimum of 8 h prior to use in a desiccating dryer.
For the preparation of the PLA nanocomposites,
Silica Aerosil R972 (supplied by Degussa Chemicals)
has been employed, which is hydrophobic fumed
silica after treated with dimethyldichlorosilane and
based on hydrophilic fumed silica, with a specific sur-
face area of 130 m2/g. The average primary size is
16 nm. A series of PLA nanocomposites with a filler
content of 2, 3, and 5 wt.% of silica (Si) have been
prepared and designated as PLA/Si/2, PLA/Si/3, and
PLA/Si/5.
664
Experimental
For the preparation of the materials, melt mixing of
nanofillers with matrix was performed with a
Brabender mixer at a temperature of 180 C, while the
rotation speed of the screws was 40 r/min. The tempera-
ture for melt mixing was kept as low as possible in
order to minimize the effect of degradation.
Hereafter, the material was compression molded at
130 C, using a thermo-press and a special mold of
2 mm thickness. The material was then cooled slowly
down at ambient temperature. For each material type,
two samples were prepared in the same manner, which
provided reproducible results.
To study the effect of nanofillers on the degradability
of PLA, PLA/nanocomposites were exposed to specific
aging conditions, namely immersed in a buffer solution
at a temperature of 37 C with a pH of 7.4, for a time
period of up to 23 weeks. The effect of the above-men-
tioned aging conditions has been studied at time inter-
vals of this period.
For the in vitro biocompatibility assessment, we
employed thin films on round glass substrates of
15 mm in diameter, by drop casting the materials’
PLA or PLA with 2% per weight nanosilica. The
glass substrate surface was covered with the material
and dried in an oven at 60 C. Samples were disinfected
in 70% ethanol and air-dried in a laminar flow bench
prior to cell seeding on them.
The evolution of molecular weight of the PLA at vari-
ous aging time intervals was recorded by SEC. All aged
and nonaged samples were dissolved in tetrahydrofuran
(THF) without any insoluble residues being visible to the
naked eye, suggesting that no crosslinking took place.
The molecular weights and molecular weight distribu-
tions of the PLA samples were determined by SEC
using a Waters system, composed of a Waters 1515 iso-
cratic pump, a set of three m-Styragel mixed bed col-
umns, with a porosity range of 102–106 Å, a Waters
2414 refractive index detector (at 40 C) and operated/
controlled through Breeze software. THF, containing
3% v/v triethylamine, was the mobile phase used at a
flow rate of 1.0 ml/min at 30 C. The setup was calibrated
with linear polystyrene standards having weight average
molecular weights in the range 1250–900,000 g/mol. The
PLA samples were dissolved directly to the carrier solv-
ent overnight at concentrations ca. 0.1% w/v and char-
acterized the next day.
TGA was performed by using a TGA Q500 V20.2
Build 27 instrument by TA Instruments (New Castle,
DE) in an inert atmosphere of nitrogen. In a typical
experiment, 10 mg of the material was placed in the
sample pan, and the temperature was equilibrated at
25 C. The temperature was then increased to 600 C
at a rate of 20 C/min.
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Journal of Biomaterials Applications 29(5)
SEM images were obtained by direct observation of
the topography of the sample’s surface with a NovaTM
NanoSEM 230 (FEI, Hillsboro, Oregon, USA) scan-
ning microscope operated at an acceleration voltage
of 5 or 15 kV. All the studied samples were coated
with gold to avoid charging under the electron beam.
She studied surface of the samples was the fractured
cross-section obtained from the tensile test.
Cell culture
Minimum essential Eagle’s medium (a-MEM), penicillin/
streptomycin, fetal bovine serum (FBS), and trypsin/ethy-
lenediaminetraacetic acid (EDTA) were purchased from
Sigma-Aldrich. PrestoBlueTM reagent and carboxyfluores-
cein diacetate succinimidyl ester (CFSE) cell viability assay
were purchased from Invitrogen Life Technologies. Cell
culture plates were obtained from Corning.
Early passages of MC3T3-E1 preosteoblastic cells
(DSZM, Germany) were cultured in alpha-MEM sup-
plemented with 10% FBS and 1% penicillin/strepto-
mycin and maintained in an incubator (Thermo
Scientific) at 37 C in a humidified atmosphere with
5% CO2. Cultures were medium changed every 2–3
days and subcultured using trypsin/EDTA. Prior to
seeding on the different substrates, confluent cells
were harvested using trypsin/EDTA and counted in a
Neubauer chamber.
In vitro cytotoxicity testing/assessment
Optical and fluorescence microscopy
A suspension of 2  104 cells in a-MEM cell culture
medium was seeded on the PLA-based material films
and placed in the cell culture incubator at 37 C. The
cells on the samples were examined daily and visualized
by means of a Zeiss Axiovert 200 microscope. Images
were taken by a ProgResÕ CFscan Jenoptik camera
(Jena, Germany) using the ProgResÕ CapturePro 2.0
software and an objective lens for the 20-fold magnifi-
cation. The visualization of the living cells by means of
the CFSE fluorescent dye was performed in the same
microscope using a fluorescence lamp and a filter for
emission at 520 nm. Images were taken with the above-
mentioned software with an exposure time set at
152 ms.
Cell viability and proliferation on the materials
Cell viability and proliferation of MC3T3-E1 preosteo-
blasts were assessed using the Presto BlueTM assay,
which is a resazurin-based non-toxic metabolic indica-
tor for viable cells.24 PLA-coated samples were disin-
fected with ethanol and transferred to an empty plate
Georgiopoulos et al.
before assaying. A total of 2  104 cells in complete
alpha-MEM culture medium were seeded on the sam-
ples and placed in the cell culture incubator at 37 C. On
days 1, 3, and 7 postseeding cell viability and prolifer-
ation assay was performed with the PrestoBlueTM
reagent according to the manufacturer’s instructions.
The absorbance was measured in a spectrophotometer
(Molecular Devices SpectraMax M2) and cell number
quantification was performed by means of a calibration
curve. Error bars represent the average of tripli-
cates  standard deviation in two independent experi-
ments for each time point.
Examination of cell metabolic activity on the mater-
ials was performed by staining with CFSE, which is
a nonfluorescent, cell-permeable dye that diffuses into
cells and the acetate groups are cleaved by intracellular
esterases to yield highly fluorescent carboxyfluorescein
succinimidyl ester. After cleavage of the acetate
groups, it has a peak excitation of 494 nm and peak
emission of 521 nm. Cells (2  104) were cultured on
PLA-containing samples for 7 days, then washed with
phosphate buffered saline (PBS), stained for 15 min with
20 mM CFSE dye in PBS and viewed by epifluorescence
microscopy.
DSCs were carried out using a Setaram DSC 141
instrument, calibrated with an Indium standard. Each
PLA sample (aged and nonaged) was heated at a con-
stant heating rate of 10 C/min from 20 C up to 170 C,
and the thermogram was recorded. The degree of crys-
tallinity was calculated by considering a melting
enthalpy of 93.1 J/g for 100% crystalline PLA.25
Three sheet specimens of 2 mm thickness were tested
for every aging procedure. DSC samples were taken
from both the center and the tip of every PLA sheet,
and it was found that the DSC results were repeatable,
with an average scatter lower than 5%. In our study,
especially for DSC tests, the aging time period was 36
weeks.
The DSC samples were washed with distilled water
two times and dried in the ambient for 2 h, before
testing.
DMA experiments were performed using the TA
Instruments DMA Q800 instrument. The mode of
deformation applied was the single cantilever beam,
and the mean dimensions of sample plaques were
12.6 mm  2 mm  17.5 mm. In order to obtain a
direct measure of the storage modulus evolution
versus aging time, the test temperature was kept con-
stant equal to 20 C, so no further thermal effects could
take place. The experiments were performed at a con-
stant frequency of 1 Hz.
Tensile measurements were performed with an
Instron 1121 type tester, at room temperature. The
dumbbell type specimens were of a gauge length of
30 mm, and the applied crosshead speed was 0.2 mm/
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665
min. This value corresponds to an effective strain rate
of 1.1  10–4 s–1. The deformation could be measured
very accurately with an experimental procedure,
which is based on a noncontact method with a laser
extensometer, described in detail in a previous work.26
Tensile stress–strain curves were then obtained up to
the breaking point. Five specimens were tested for
each aging procedure, and the scatter between experi-
mental data was lower than 8%.
Results and discussion
SEC results
The degradation of PLA matrix has been tested by
examining the molecular weight distribution after
exposure to the prescribed aging conditions, for differ-
ent time periods. The weight average molecular weight
M w versus aging time is given in Table 1. A substantial
molecular weight decrement is obtained after 163 days
of aging.
As can be seen in Table 1, after aging at 37 C for 65
days, PLA underwent little change in its molecular
weight. Significantly lower molecular weights were
measured for samples after 163 days (23 weeks) of
aging. This is exemplified in Figure 1, which shows
the elution curves of PLA samples for different time
intervals. As can be seen in Figure 1, the elution peak
shifted to lower molecular weight with increasing aging
time. Furthermore, the elution peaks of the aged sam-
ples display a higher contribution of low molecular
weight fractions, which is attributed mainly to chain
scission due to hydrolysis.
The degradation rate was quantified in terms of the
average hydrolytic degradation rate constant (kt). The
(kt) values were evaluated assuming an exponential
decrease of Mn using the following equation4
lnMn ðt2 Þ¼ lnMn ðt1 Þkt t ð1Þ
where Mn(t2) and Mn(t1) are the Mn values at the
hydrolytic degradation times of t2 and t1, respectively.
In Figure 2, the molecular weight changes are plotted
logarithmically with varying the aging time. In the same
Table 1. PLA molecular weight variation with aging time.
Aging time (weeks) Molecular weight (g/mol)
0 194,873
9.0 169,439.5
23 89,609
48 69,600
666 Journal of Biomaterials Applications 29(5)

4
results.2 Regarding the aging temperature effect, the
degradation process was very slow at temperatures
3
equal or below 40 C, but the speed was greatly
0 weeks enhanced at 50 C. The hydrolytic degradation rate at
the buffer solution 37 C appears to be almost the same
2 with the conditions of 40 C and 80% RH.
MV

23 weeks
9 weeks
1 SEM results
The morphological changes incurred in the PLA/nano-
0
48 weeks composites after exposure to the prescribed aging con-
ditions were analyzed by SEM micrographs and shown
16 18 20 22
Minutes
24 26 28 30 representatively in Figure 3(a) to (d) for PLA/Si/3 and
PLA/Si/5 nanocomposites. From these photos, the
Figure 1. Elution curves for PLA at different aging times good quality of nanofiller’s dispersion is apparent,
obtained by size exclusion chromatography. given that the nanoparticles are visible and quite homo-
geneously dispersed. Nanoparticles of a few nano-
meters size coexist with larger aggregates, with the
12.4 aggregates being larger with increasing the nanofiller
12.2 content. For PLA/Si/3 after 23 weeks of aging, an
12.0 increasing surface roughness is observed, as well as
11.8
the existence of holes (dark regions) especially around
11.6
nanoparticles. Analogous trend is obtained for PLA/Si/
5 where, in some cases, holes connecting neighboring
Ln(Mw)

11.4
nanoparticles are also created.
11.2

11.0 80% RH/20°C

80% RH/40°C
80% RH/50°C
10.8
buf.soI/37°C
In vitro cytotoxicity testing/assessment
10.6

10.4
0 20 40 60 80 100
Time (Days)
Figure 2. PLA molecular weight variation versus time at
various aging conditions.
plot, analogous results for PLA, degraded under differ-
ent conditions1 are depicted for comparison. The
experimental procedure applied for aging PLA at
80% RH and various temperatures is presented in
detail in Niaounakis et al.1 Following Figure 2, an aver-
age value kt for the hydrolytic degradation rate has
been calculated. The estimated kt values for the differ-
ent time intervals were 3.73  10–4, 2.96  10–3, and
20  10–3 days–1 at 20, 40, and 50 C, and 80% RH,
respectively. The sample immersed in the buffer
solution at 37 C had an average degradation rate
equal to 4.9  10–3 days–1 for a similar time period of
aging with the other samples. When the time period is
extended to 340 days, the value of kt is reduced to
3.185  10–3 days–1. The results showed that the degrad-
ation rate was higher at the beginning (first 60 days) for
every temperature and decreased significantly after 30
days. This finding was in agreement with previous
Initial cell adhesion on PLA and PLA þ SiO2 coatings
120 140 160 180 The optical microscopy images indicate a good initial
cell adhesion 2 h postseeding on both materials, the
PLA (Figure 4(b)) and PLA/Si/2 (Figure 4(c)), which
is comparable to the tissue culture treated polystyrene
control polystyrene (PS) (Figure 4(a)). Cells extend elon-
gated protrusions, displaying a spindle-shape morph-
ology, and attach in a similar number and manner on
both PLA-containing coatings as on the preferable PS
control. These data depict a characteristic morphology
of preosteoblastic cells seeded on a biocompatible mater-
ial surface without showing any adverse effects.
Cell proliferation on PLA and PLA þ SiO2 coatings
The graph in Figure 5 shows the number of living cells
on the tissue culture treated polystyrene control sur-
face, PLA and PLA/Si/2 after 1, 3, and 7 days in cul-
ture. Beginning with an initial number of 2  104 cells
seeded on each sample type, we observe a significantly
increasing cell proliferation after 3 and 7 days in culture
on both PLA and PLA/Si/2 materials, which is similar
to the PS control surface. Among the different material
surfaces at each experimental time point, we do not
observe any statistically significant proliferation
increase.

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Georgiopoulos et al. 667

Figure 3. SEM micrographs of PLA/Si/3 after 23 weeks of aging. SEM micrographs of PLA/Si/5.

Figure 4. Initial preosteoblastic cell adhesion on PS (control) (a), PLA (b), and PLA/Si/2 (c) after 2 h in culture shown by optical
microscopy indicating a comparable number of cells with a similar spindle-shaped morphology attached on all surfaces, 20-fold
magnification. Scale bar represents 50 mm.

cells appear to have increasingly proliferated and

Cell metabolic activity
Figure 6 shows the epifluorescence of living preosteo-
blastic cells stained with the CFSE dye on either tissue
culture treated polystyrene control (a), or PLA (b), or
PLA/Si/2 (c) after 7 days in culture. We observe a com-
parable number of living cells appearing in green, with
a similar flattened morphology on both PLA and PLA/
Si/2 materials, as on the polystyrene control. The
depicted high cell density and the polygonal cell morph-
ology are evident for well-attached living cells on the
material coatings. After 7 days in culture, and com-
pared to the initial adhesion stage shown in Figure 4,
formed a confluent live cell layer on all three surfaces.
Additionally, these data corroborate with the quantita-
tive cell proliferation analysis shown in Figure 5.
TGA results
The TGA results are summarized in Table 2, where the
10 wt% loss temperature with varying aging time is pre-
sented for all samples. From this table it is extracted
that the thermal stability of PLA and PLA/Si/5 nano-
composite remains almost unaffected with aging time,
while for PLA/Si/2 and PLA/Si/3 it is reduced and
shifted toward lower temperatures. This effect denotes

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668 Journal of Biomaterials Applications 29(5)

that the good quality dispersion of silica at those sam- restriction of silica nanoparticles on the segmental
ples, as postulated by SEM, increases the interfacial and long-range chain mobility of the matrix takes
region, which is more sensitive to the aging conditions. place, which would lead to a Tg increment of the nano-
The formation of large agglomerates in PLA/Si/5 does composites. On the other hand, no indication of poor
not have the same effect. adhesion between matrix and nanofillers exists, since in
that case a Tg reduction should be obtained.27 A simi-
lar effect, where the addition of 2% vol. of fumed silica
DSC results on PLA does not significantly affect the Tg of the
The DSC results of the PLA nanocomposites and their material has been found.28
evolution with aging time are summarized in Table 3. In the following paragraphs, the evolution of ther-
PLA nanocomposites exhibit three transitions, the mal properties with aging time of PLA/nanocomposites
glass transition temperature Tg, the cold crystallization is analytically presented. Following Table 3, the Tg of
temperature Tcc (for PLA matrix), and the melting PLA matrix is slightly decreasing during the first 12
regime Tm. weeks of aging, and hereafter starts increasing. The
The Tgs of the PLA/nanocomposites were not Tg of PLA/nanocomposites does not substantially
affected by the presence of silica, meaning that no change with aging time, exhibiting a slight increment
after 36 weeks of aging. Generally, the Tg decrement is
attributed to the molecular weight reduction of the
samples. In previous works it has been found that the
80000 Tg is either not significantly affected by aging20 or it is
PS decreased with exposure time.29 In our case, the Tg is
70000
PLA generally increased after a long time period of aging for
60000 PLA+SiO2 PLA and PLA/nanocomposites, with this increment
being more essential in the PLA matrix. The Tg incre-
50000
ment with aging has also been detected for similar
Cell number

40000
materials in Georgiopoulos et al.30
From Figures 7 to 10 where DSC curves are illu-
30000 strated for PLA and PLA/nanocomposites correspond-
ingly, it is shown that the Tg region is expressed by an
20000

10000

0 Table 2. TGA 10 wt% loss temperature in  C at various aging

Day 1 Day 3 Day7 times.
Figure 5. Cell proliferation on tissue culture treated polystyr- Aging time
ene (control), PLA and PLA/Si/2 after 1, 3, and 7 days in culture (weeks) PLA PLA/Si/2 PLA/Si/3 PLA/Si/5
by means of the PrestoBlueTM assay. Cells proliferate on both
PLA-containing materials similarly to the PS control surface. 0 335.8 342 332.7 333
Error bars represent the average of triplicates  STDV for each 14 333 332.5 331.5 331
time point. Optical density values are normalized to the cell 24 330.5 327.5 325 330.8
number according to a calibration curve.

Figure 6. Epifluorescence of living cells stained with the CFSE dye on tissue culture treated polystyrene control (a), PLA
(b), and PLA/Si/2 (c) after 7 days in culture. A comparable number of living cells with a similar flattened morphology is observed on
both PLA-containing materials as on the PS control. Images are taken in a 20-fold magnification, exposure time is 152 ms. Scale bar
represents 50 mm.

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Georgiopoulos et al. 669

Table 3. DSC properties of PLA/nanocomposites with varying aging time.

Material/aging Degree of Percentage change

time (weeks) Tg ( C)
Har (J/g)
Hcc (J/g)
Ham (J/g) crystallinity XC (%) crystallinity (%) Tm ( C) Tm1/Tm2

PLA
0 58.42 3.49 5.31 22 18.0 0 146.1/154.4
3.0 56.8 5.99 8.16 22.49 15.4 14.4 146.25/154.52
12.0 57.1 7.86 6.9 23.77 18.1 0.55 146/155.2
18.0 58.4 8.32 9.05 24.66 16.8 6.6 146/155.8
23.0 60.24 7.9 9.36 22.45 14.0 22.2 146.8/157.8
36.0 63.7 8.35 9.33 27.72 19.8 10.0 146/156.0
PLA/Si/2
0 58.81 1.06 22.55 24.24 0 151/155.3
3.0 59.29 3.88 24.18 26.0 7.26 149.7/155.29
12.0 58.1 4.22 25.52 27.44 13.2 149.54/155.9
18.0 58.15 3.7 27.33 29.38 21.2 149.56/156.4
23.0 60.15 5.0 24.71 26.56 9.57 150/157.74
36.0 59.0 6.25 31.55 33.92 40.0 149.47/156.81
PLA/Si/3
0 58.22 1.15 21.89 23.53 0 150/155.3
3.0 61.0 3.13 24.64 26.49 12.6 150.38/154.8
12.0 58.2 4.11 25.62 27.54 17.0 149.7/155.9
18.0 58.82 4.51 26.39 28.37 20.6 150.1/156.5
23.0 59.6 4.75 27.34 29.39 25.0 149.5/156.3
36.0 62.4 6.0 30.1 32.36 37.52 150/156.54
PLA/Si/5
0 58.15 0.92 20.88 22.45 0 150.3/155.8
3.0 61.5 3.76 23.47 25.23 12.38 149.8/154.7
12.0 57.9 4.0 23.72 25.5 13.58 150.38/156.12
18.0 60.31 3.78 22.79 24.5 9.13 150.1/157.1
23.0 59.8 5.37 27.63 29.7 32.3 150.8/156.19
36.0 62.34 6.67 30.2 32.36 44.14 149.5/156.67
a
Enthalpy values are normalized with respect to pure polymer.

endothermic peak, corresponding to an enthalpy relax-

PLA
ation
Hr. The existence of such an endothermic peak
is related to the melting of a mesophase, which is a
36 weeks
region with some kind of molecular ordering.31,32
As presented in Table 3, for the unaged materials, a
23 weeks
substantial
Hr decrement with the addition of nano-
Heat Flow (mw)

18 weeks particles, which implies a confinement of a fraction of

exo PLA chains around nanofillers surface, exists, which
0.00 cannot contribute to a relaxation around Tg.33 In con-
12weeks
trast,
Hr of all samples is increased with aging time,
3 weeks
and this increment is enlarged at PLA/nanocomposites
0 weeks compared to that of PLA matrix. This effect denotes
that after the sample has been hydrolyzed for a certain
120
period of time, the formation of this locally ordered
40 60 80 100 140 160 180
°Temperature (°C)
structure is enhanced.
As shown in Figure 7, PLA matrix exhibits the exo-
Figure 7. DSC curves of PLA at various aging times. thermic effect, attributed to the cold crystallization of

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670 Journal of Biomaterials Applications 29(5)

PLA/Si2 PLA during the DSC heating process. It has been found
36 weeks that the cold crystallization temperature Tcc is decreas-
ing with aging time, while the corresponding enthalpy
23 weeks

Hcc is increasing. When cold crystallization occurs,
Heat Flow (mw)

18 weeks imperfect crystals are formed, which melt during the

0.0 exo DSC heating scan. In addition, the
Hcc increment,
12 weeks related to the enlarged cold crystallization area of
PLA matrix after 36 weeks of aging (Figure 7), indi-
3 weeks cates the presence of more polymeric chains involved in
the cold crystallization process, thus strengthening the
0 weeks interpretation of morphological changes previously
drawn from the
Hr assessment. Regarding crystallin-
40 60 80 100 120 140 160 180
ity, the PLA/nanocomposites, in the unaged stage,
Temperature (°C) exhibit a percentage change crystallinity, namely 34.7,
30.7, and 24.7% for PLA/Si/2, PLA/Si/3, and PLA/Si/5
Figure 8. DSC curves of PLA/Si/2 at various aging times. correspondingly. These values are calculated with
respect to the crystallinity content of PLA matrix,
and this effect shows that silica nanoparticles act as
nucleating agents.
PLA/Si3 As far as crystalline behavior of PLA and PLA/
nanocomposites under aging conditions is concerned,
36 weeks it must be noted that for a semicrystalline polymer,
23 weeks
the initial crystalline structure strongly affects the
hydrolytic degradation of PLA.18,34
Generally, hydrolytic degradation occurs first in the
Heat Flow (mw)

18 weeks
exo
12 weeks amorphous region. The degree of crystallinity Xc(%) of
0.0 PLA with increasing aging time exhibits a nonmono-
3 weeks
tonic behavior, but generally has a tendency to increase
up to a value of 10% (Table 3). PLA/Si nanocompo-
sites, on the other hand, appear as monotonic incre-
0 weeks
ment of the degree of crystallinity with aging time, up
60 80
to values of the order of 40–44% with respect to the
40 100 120 140 160 180
Temperature (°C) initial value before aging. This effect consists of a
strong indication that silica is an effective nucleating
Figure 9. DSC curves of PLA/Si/3 at various aging times. agent, promoting the crystallization of PLA matrix,
during the hydrolytic degradation.18,34 It has been
reported that the increased heat of fusion (and there-
fore degree of crystallinity increment) is attributed to
plasticization of PLA by water molecules as well as by
PLA/Si5 lactic oligomers that would give sufficient mobility to
the polymer chains to organize and further crystallize.22
36 weeks
This is the result of two competitive effects: The
silica content leads to the increase of the hydrophilicity
23 weeks
of the sample, which results in the damage of more
Heat Flow (mw)

exo crystalline structure during the hydrolytic degradation

process. On the other hand, due to degradation, we
0.0 18 weeks
have easier crystal formation, and the nucleation of
3 weeks
new crystalline regions, where smaller chains are
involved.
0 weeks Analyzing further the evolution of crystalline struc-
ture during aging, the samples obtained at various
40 60 80 100 120 140 160 180
aging time periods were examined by DSC. From
Temperature (°C) Figures 7 to 10, illustrating PLA and PLA/nanocom-
posites, as far as the melting region is concerned, two
Figure 10. DSC curves of PLA/Si/5 at various aging times. endothermic peaks appear in all material types.

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Georgiopoulos et al. 671

The melting point generally is directly related to the Table 4. Tensile properties of PLA/nanocomposites with
quality of the crystalline region, and consequently the varying aging time.
thickness of the lamellae; the bigger the lamellae thick-
Material/aging Young’s Yield Strain
ness, the higher the melting peak temperature. time (weeks) modulus (GPa) stress (MPa) at break
Therefore, double melting peaks are generally asso-
ciated with the presence of two main crystal popula- PLA
tions, with different size and quality.35–37 Following 0 3.5 54.9 0.055
Table 3, the low temperature peak Tm1 for PLA/nano- 3.0 3.0 52.65 0.074
composites is higher than that of PLA matrix, whereas 7.0 3.03 54.4 0.033
the second melting peak Tm2 is almost the same for all 14.0 3.3 54.3 0.04
materials examined. The first peak Tm1 has been related 23.0 3.11 49.6 0.025
to the fusion of rather imperfect crystals, formed by PLA/Si/2
primary crystallization, whereas the second melting
0 4.87 56.2 0.055
peak Tm2 can be related to the melting of more perfect
3.0 3.6 53.32 0.062
crystals, which were formed during a reorganization
procedure upon heating. 7.0 3.6 54.3 0.048
With increasing aging time, it can be observed from 14.0 3.64 51.0 0.02
Figures 7 to 10 and Table 3 that Tm1 is almost constant 23.0 3.75 0.012
with varying aging time (with an exception for PLA/Si/ PLA/Si/3
2, where Tm1 slightly decreases), while Tm2 exhibits a 0 4.83 59.0 0.044
slight increment with aging time, for all material types. 3.0 3.67 53.2 0.061
It has been reported38 that when Tm decreases with 7.0 3.78 54.35 0.075
aging, crystalline regions are partially degraded, spe- 14.0 4.0 49.5 0.016
cially the interlamellar amorphous phases into the crys- 23.0 3.7 37.2 0.012
talline regions, even to a less extent than in the PLA/Si/5
amorphous regions.
0 4.85 58.7 0.07
In our case, following Figures 7 to 10, the Tm2 peak
3.0 3.58 55.9 0.032
becomes more intense with aging time for PLA/nano-
composites. It probably happens, due to a mechanism 7.0 3.62 53.62 0.06
of melting and recrystallization of less perfect crystal- 14.0 3.65 47.2 0.02
lites into thicker ones, which hereafter melt at a higher 23.0 3.87 39.3 0.012
temperature. Aging procedure seems to facilitate this
mechanism. This fact combined with the enhancement
of the total heat of fusion with aging, denotes that silica
can promote the crystallization of PLA matrix during 60

the hydrolytic degradation process for the specific aging 14 weeks 0 weeks
50
conditions applied. 7 weeks
3 weeks
23weeks
40
Stress (MPa)

Tensile testing results

30
The experimental tensile properties of PLA/Si nano-
PLA
composites are extensively analyzed in Kontou et al.19 20
and Georgiopoulos et al.30 The corresponding evolu-
tion of the tensile properties of pristine PLA and its 10
nanocomposites with aging at the specific conditions
examined, at various time periods are presented in 0
0.08
0.00 0.01 0.02 0.03 0.04 0.05 0.06 0.07
Table 4 and in Figures 11 and 12 representatively for Strain
PLA and PLA/Si/3. From Table 4, PLA/Si nanocom-
posites exhibit a 39% Young’s modulus increment com- Figure 11. Tensile stress–strain curves for PLA at various aging
pared to the Young’s modulus of PLA matrix. From conditions.
Figure 11 of pristine PLA, it can be observed that in all
stages of aging, a yield stress and a subsequent strain PLA/nanocomposites exhibit a 26% reduction.
softening is exhibited. Following Table 4, it can be Hereafter, all studied samples exhibited almost the
noticed that after 3 weeks of aging, PLA matrix under- same trend in tensile properties during aging, i.e. their
goes a 14% reduction in the Young’s modulus, while Young’s modulus appears to be not further greatly

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672
60
0 weeks
3 weeks
50
14 weeks 7 weeks
40
Strees (MPa)
23 weeks
30
PLA/Si/3
20
10
0
0.00 0.01 0.02 0.03 0.04 0.05 0.06
Strain
Figure 12. Tensile stress–strain curves for PLA/Si/3 at various
aging conditions.
affected, with increasing aging time. Finally, after 23
weeks of aging, the Young’s modulus reduction for
PLA is about 11%, while it is about 23% for PLA/
nanocomposites. Therefore, it can be extracted that
silica enhances degradability of PLA, while this
enhancement slows down with increasing aging time,
probably due to the crystallinity increment for PLA/
nanocomposites. Similar trend is obtained for the
yield stress variation. Initially, PLA/nanocomposites
have a slightly yield stress increment (of the order of
7%) compared to that of PLA matrix. With varying
aging time, the yield stress of all samples is gradually
decreased, with the PLA/nanocomposites demonstrat-
ing higher decrement (up to 37% for PLA/Si/3). On the
other hand, after 18 weeks of aging, yield stress of
the nanocomposites turns to be lower than that of the
matrix. It can be extracted that in this case, the entire
load is carried by the polymeric matrix and the load-
bearing cross-section is lowering with increasing
nanofiller content.28 This effect is an indication that
the reinforcing effect, observed initially, is no longer
active, revealing that degradation is more intense in
the matrix–nanofiller interface. This effect is in agree-
ment with SEM results, where the degradation appears
to be more intense around nanoparticles. In our previ-
ous study,19 with the same materials system, and differ-
ent aging conditions, namely 40 C and 80% RH, yield
stress of PLA/Si nanocomposites was generally higher
than that of the PLA matrix. Comparing the two dif-
ferent aging conditions, it is revealed that at 40 C and
80% RH, the mechanical properties substantially
decrease in the first 3 weeks and hereafter they are
recovering, while under immersion in a buffer solution
at 37 C, the same properties after the exhibited decre-
ment in 3 weeks are slightly recovered and remain
almost stable for a long time period of aging. It must
also be noted that PLA exhibits a lower degradability in
all cases, compared to the PLA/nanocomposites. In the
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Journal of Biomaterials Applications 29(5)
Stain amplitude =0.2% PLA
4.0
frequency=1 Hz PLA/Si/2
Temperature= 37°C PLA/Si/3
3.8
PLA/Si/5
Storage Modulus (GPa)
3.6
3.4
3.2
3.0
2.8
0.08 2.6
0.07
0 5 10 15 20 25 30 35
weeks
Figure 13. Storage modulus at various aging times for PLA and
PLA/nanocomposites.
present study, the degradation under immersion in a
buffer solution at 37 C is enhanced, compared to
40 C and 80% RH, which is also related to the
higher hydrolytic degradation rate, calculated in the
previous paragraph.
It can be concluded that silica nanoparticles acceler-
ate the degradability of PLA, having a higher impact on
Young’s modulus, under the specified aging conditions,
for 7 weeks and hereafter this acceleration is retarded,
due to the crystallinity increment, as a result of the
molecular weight reduction.
DMA
In Figure 13 the storage modulus at a frequency of 1 Hz
and 37 C is depicted with varying aging time. From this
plot it is revealed that the storage modulus is always
higher for PLA/nanocomposites compared to that of
PLA matrix, while the reinforcing effect is higher for
PLA/Si/2, probably due to the finer dispersion of nano-
particles. The evolution of storage modulus with aging
time is monotonically decreasing, appearing to be dif-
ferent than that of the Young’s modulus. This is due to
the different type of deformation applied. The slope of
the storage modulus decrement is lower for PLA
matrix, while PLA/Si/5 exhibits the higher slope of deg-
radation compared to PLA/Si/2 and PLA/Si/3.
Conclusions
A series of biobased polymer matrix nanocomposites
based on PLA with varying nanosilica content have
been prepared and studied experimentally. A 39%
Young’s modulus enhancement has been found for
PLA/nanocomposites with respect to the pure PLA
matrix. In addition, 33% crystallinity increment in
average has been detected for PLA/nanocomposites,
indicating that nanosilica fillers act as nucleating
Georgiopoulos et al.
agents. Hereafter, this enhancement does not change
with varying nanofiller loading.
The cell culture investigation demonstrates that both
materials PLA and the PLA/nanocomposites facilitate
a good initial cell adhesion, increase preosteoblastic cell
proliferation after 3 and 7 days in culture, and therefore
support their potential use in bone tissue engineering
applications.
A study of the change of the thermomechanical
properties of PLA and PLA/nanocomposites with
aging time after exposure at a buffer solution at a tem-
perature of 37 C with a pH of 7.4, simulating the
human body, has been performed. Comparative study
with various aging conditions, previously executed has
also been made. Significantly lower molecular weights
for PLA matrix were measured for samples after 23
weeks of aging. The aged samples display a higher con-
tribution of low molecular weight fractions, which is
attributed mainly to chain scission due to hydrolysis.
The crystallization behavior in PLA and the way it is
affected by the presence of nanofillers during degrad-
ation procedure has been studied and values of 44%
crystallinity increment have been found. Double melt-
ing endotherms at 146 and 156 C, associated with dif-
ferent morphologies have been detected for all material
types studied. The high temperature melting peak was
more intense with increasing aging time, and this effect
has been attributed to a recrystallization procedure fol-
lowed by a subsequent melting.
At the particular aging conditions studied, silica
nanoparticles accelerate the degradability of PLA,
having a higher impact on Young’s modulus, under
the specified aging conditions, for 7 weeks and hereafter
this acceleration is retarded, due to the crystallinity
increment, as a result of the molecular weight reduction.
Declaration of conflicting interests
None declared.
Funding
This research has been co-financed by the European Union
(European Social Fund, ESF) and Greek national funds
through the Operational Program ‘‘Education and Lifelong
Learning’’, Research Funding Program Aristeia (E.K.), and
Aristeia II ‘Osteobiomimesis’ (MC).
References
1. Niaounakis M, Kontou E and Xanthis M. Effects of aging
on the thermomechanical properties of poly(lactic acid).
J Appl Polym Sci 1966; 119: 472–481.
2. Signori F, Coltelli M-B and Bronco S. Thermal
degradation of poly(lactic acid) (PLA) and poly(butylene
adipate-co-terephthalate) (PBAT) and their blends upon
melt processing. Polym Degrad Stab 2009; 94: 74–82.
Downloaded from jba.sagepub.com at UCSF LIBRARY & CKM on November 28, 2014
673
3. Pillin I, Montrelay N, Bourmaud A, et al. Effect of
thermo-mechanical cycles on the physic-chemical proper-
ties of poly(lactic acid). Polym Degrad Stab 2008; 93:
321–328.
4. Saha SK and Tsuji H. Effects of molecular weight and
small amounts of D-lactide units on hydrolytic degrad-
ation of poly(L-lactic acid)s. Polym Degrad Stab 2006; 91:
1665–1673.
5. Ho KLG, Pometto AL and Hinz PN. Effects of tempera-
ture and relative humidity on polylactic acid plastic deg-
radation. Polym Environ 1999; 7: 83–92.
6. Middleton JC and Tipton AJ. Synthetic biodegradable
polymers as orthopedic devices. Biomaterials 2000; 21:
2335–2346.
7. Felfel RM, Ahmed I, Parsons AJ, et al. Investigation of
crystallinity, molecular weight change and mechanical
properties of PLA/PBG bioresorbable composites as
bone fracture fixation plates. J Biomater Appl 2012; 26:
765–789.
8. Fukushima K, Abbate C, Tabuani D, et al.
Biodegradation of poly(lactic acid) and its nanocompo-
sites. Polym Degrad Stab 2009; 94: 1646–1655.
9. Ray SS and Bousmina M. Biodegradable polymers and
their layered silicate nanocomposites: In greening the 21st
century materials world. Prog Mater Sci 2005; 50:
962–1079.
10. MacNeill I and Leiper H. Degradation studies of some
polyesters and polycarbonates-2. Polylactide:
Degradation under isothermal conditions, thermal deg-
radation mechanism and photolysis of the polymer.
Polym Degrad Stab 1985; 11: 309–326.
11. Feijoo J, Cabedo L, Gimenez E, et al. Development of
amorphous PLA-montmorillonite nanocomposites.
J Mater Sci 2005; 40: 1785–1788.
12. Jungraithmayr W, Laube I, Hild N, et al. Bioactive nano-
composite for chest-wall replacement: Cellular response
in amurine model. J Biomater Appl 2014; 29: 36–45.
13. Wen X, Lin Y, Han C, et al. Thermomechanical and
optical properties of biodegradable poly(L-lactide)/silica
nanocomposites by melt compounding. J Appl Polym Sci
2009; 114: 3379–3388.
14. Ray SS, Maiti P, Okamoto M, et al. New polylactide/
layered silicate nanocomposites. 1. Preparation,
characterization, and properties. Macromolecules 2002;
35: 3104–3110.
15. Delabarde C, Plummer CJG, Bourban PE, et al.
Accelerated ageing and degradation in poly-L-lactide/
hydroxyapatite nanocomposites. Polym Degrad Stab
2011; 96: 595–607.
16. Zhao Y, Qiu Z and Yang W. Effect of functionalization
of multiwalled nanotubes on the crystallization and
hydrolytic degradation of biodegradable poly(L-lactide).
J Phys Chem B 2008; 112: 16461–16468.
17. Ray SS, Yamada K, Okamoto M, et al. New polylactide-
layered silicate nanocomposites. 2. Concurrent improve-
ments of material properties, biodegradability and melt
rheology. Polymer 2003; 44: 857–866.
18. Chen H, Wang Y, Chen J, et al. Hydrolytic degradation
behavior of poly(L-lactide)/SiO2 composite. Polym
Degrad Stab 2013; 98: 2672–2679.
674
19. Kontou E, Georgiopoulos P and Niaounakis M. The role
of nanofillers on the degradation behavior of polylactic
acid. Polym Compos 2012; 33: 282–294.
20. Chen H, Chen J, Chen J, et al. Effect of organic mont-
morillonite on cold crystallization and hydrolytic degrad-
ation of poly(L-lactide). Polym Degrad Stab 2012; 97:
2273–2283.
21. Paul MA, Delcourt C, Alexandre M, et al. Polylactide/
montmorillonite nanocomposites: Study of the hydrolytic
degradation. Polym Degrad Stab 2005; 87: 535–542.
22. Fukushima K, Tabuani D, Dottori M, et al. Effect of
temperature and nanoparticle type on hydrolytic degrad-
ation of poly(lactic acid) nanocomposites. Polym Degrad
Stab 2011; 96: 2120–2129.
23. Zhang X, Espiritu M, Bilyk A, et al. Morphological
behavior of poly(lactic acid) during hydrolytic degrad-
ation. Polym Degrad Stab 2008; 93: 1964–1970.
24. Terzaki K, Kissamitaki M, Skarmoutsou A, et al.
Pre-osteoblastic cell response on three-dimensional,
organic-inorganic hybrid material scaffolds for bone
tissue engineering. J Biomed Mater Res Part A 2013;
101A: 2283–2294.
25. Fischer EW, Sterzel HJ and Wegner G. Investigation of
the structure of solution grown crystals of lactide copoly-
mers by means of chemical reactions. Colloid Polym Sci
1973; 251: 980–990.
26. Kontou E and Farasoglou P. Determination of the true
stress-strain behaviour of polypropylene. J Mater Sci
1998; 33: 147–153.
27. Dacko P, Kowalczuk M, Janeczek H, et al. Physical
properties of the biodegradable polymer compositions
containing natural polyesters and their synthetic ana-
logues. Macromol Symp 2006; 239: 209–216.
28. Dorigato A, Sebastiani M, Pegoretti A, et al. Effect of
silica nanoparticles on the mechanical performance of
poly(lactic acid). J Polym Environ 2012; 20: 713–725.
29. Vasanthan N and Ly O. Effect of microstructure on
hydrolytic degradation studies of poly(L-lactic acid) by
Downloaded from jba.sagepub.com at UCSF LIBRARY & CKM on November 28, 2014
Journal of Biomaterials Applications 29(5)
FTIR spectroscopy and differential scanning calorimetry.
Polym Degrad Stab 2009; 94: 1364–1372.
30. Georgiopoulos P, Kontou E and Niaounakis M.
Thermomechanical properties and rheological behavior
of biodegradable composites. Polym Compos 2014; 35:
1140–1149.
31. Chen HM, Feng CX, Zhang WB, et al. Hydrolytic deg-
radation behavior of poly(L-lactide)/carbon nanotubes
nanocomposites. Polym Degrad Stab 2013; 98: 198–208.
32. Zhang TP, Hu J, Duan YX, et al. Physical aging enhanced
mesomorphic structure in melt-quenched poly(L-lactic
acid). J Phys Chem B 2011; 115: 13835–13841.
33. Pluta M, Murariu M, Alexandre M, et al. Polylactide
compositions. The influence of ageing on the structure,
thermal and viscoelastic properties of PLA/calcium
sulfate composites. Polym Degrad Stab 2008; 93:
925–931.
34. Tsuji H, Ikarashi K and Fukuda N. Poly(L-lactide): XII.
Formation, growth, and morphology of crystalline resi-
dues as extended-chain crystallites through hydrolysis of
poly(L-lactide) films in phosphate-buffered solution.
Polym Degrad Stab 2004; 84: 515–523.
35. Huang JW, Hung YC, Wen YL, et al. Polylactide/nano-
and microscale silica composite films. II. Melting behav-
ior and cold crystallization. J Appl Polym Sci 2009; 112:
3149–3156.
36. Shieh YT and Liu GL. Effects of carbon nanotubes on
crystallization and melting behavior of poly(L-lactide) via
DSC and TMDSC studies. J Polym Sci Part B Polym
Phys 2007; 45: 1870–1881.
37. Chen HM, Zhang WB, Du XC, et al. Crystallization kin-
etics and melting behaviors of poly(L-lactide)-graphene
oxides composites. Thermochim Acta 2013; 566: 57–70.
38. Ray SS, Yamada K, Okamoto M, et al. Polylactide-
layered silicate nanocomposite: A novel biodegradable
material. Nano Lett 2002; 2: 1093–1096.