Basic Research—Technology

Environmental Scanning Electron Microscopy Connected

with Energy Dispersive X-ray Analysis and Raman Techniques
to Study ProRoot Mineral Trioxide Aggregate and Calcium
Silicate Cements in Wet Conditions and in Real Time
Maria Giovanna Gandolfi, DBiol, DSc, MSc, PhD,*,† Kirsten Van Landuyt, DMD, PhD,‡
Paola Taddei, DChem, PhD,§ Enrico Modena, DChem,§ Bart Van Meerbeek, DMD, PhD,‡
and Carlo Prati, MD, DDS, PhD*

Abstract
Introduction: ProRoot mineral trioxide aggregate
(MTA) and calcium silicate cements are able to set in
a moist environment. The aim of the study was to
examine the surface structure and composition of
a cement paste under wet conditions and in real time
during setting by environmental scanning electron
microscopy connected with energy dispersive x-ray
analysis (ESEM-EDX) and micro-Raman techniques.
Methods: White ProRoot MTA and experimental white
tetrasilicate cement (wTC) and wTC containing bismuth
oxide (wTC-Bi) were studied. Cement disks were
analyzed 10 minutes after powder-liquid mixing (freshly
prepared samples) and after immersion in Dulbecco
phosphate-buffered saline at 37 C for 24 hours
(24-hour-aged samples). Results: Freshly prepared
wet cements at ESEM-EDX exposed an irregular surface
(displaying calcium, silicon, aluminum, chlorine reflexes,
and bismuth traces in MTA and wTC-Bi) with needle-like
and cubic-hexagonal shaped crystals. Aggregates of
spheroidal Ca-P–rich crystals (spherulites) appeared on
the surface of 24-hour-aged samples. The starting unhy-
drated powders displayed the typical Raman bands of
Portland cement components: alite, belite, and calcium
sulfate (only as anhydrite in MTA and as both anhydrite
and gypsum in wTC and wTC-Bi). MTA powder showed
higher amount of calcium carbonate and lower quanti-
ties of anhydrite and higher crystallinity of the silicate
component, leading to a slower hydration reaction.
Products/markers of hydration reactions were present
on fresh samples; ettringite formed on the surface of
all the cements; calcium hydroxide (portlandite) was de-
tected only on the surface of wTC, but no conclusion can
be drawn on wTC-Bi and MTA because of the interfer-
ence of bismuth oxide. Calcium phosphate and calcite/aragonite bands were detected
on all 24-hour-aged cements; portlandite was no longer detected on wTC. Conclu-
sions: ESEM and micro-Raman are powerful and suitable techniques to investigate
endodontic calcium silicate hydrated cements in real time and in their humid state
without inducing artifacts by sample preparation. The formation of apatite spherulites
on calcium silicate cements might have clinical relevance. (J Endod 2010;36:851–857)
Key Words
Aapatite, bioactive materials, calcium-silicate cements, endodontic cements, environ-
mental scanning electron microscopy (ESEM), micro-Raman spectroscopy, MTA,
Portland cement, root-end filling materials
C alcium silicate cements possess hydraulic properties and spontaneously set in the
presence of water (1). In addition, they release calcium during their hydration and
setting reaction (2–5) and induce the formation of apatite deposits when immersed in
phosphate-containing solutions (4–12). Calcium silicate cements, mainly Portland-
derived cements such as mineral trioxide aggregate (MTA) and others, have been
designed and are used for dental clinical applications such as root-end filling and
root-repair materials, pulp capping, or dentin hypersensitivity (3, 12–17).
The self-setting properties of calcium silicate cements are due to the progressive
hydration reaction of the orthosilicate ions (SiO44–). When calcium silicate particles
react with fluids (water), a nanoporous amorphous calcium silicate hydrate gel
(CSH gel) forms on the cement particles, whereas calcium hydroxide (Ca(OH)2)
(portlandite) nucleates and grows in the available voids and pore spaces (12). CSH
gel polymerizes and hardens over time, forming a solid network, which is associated
with an increased mechanical strength (12). Soluble Ca(OH)2 is released by the cement
surface and increases the alkalinity of the surrounding environment (5, 11).
The setting and hardening of calcium silicate cements are hydration reactions and
require water (18, 19). The preparation procedures for scanning electron microscopy
analysis take time (dehydration procedures and coating procedures) and might create
artifacts that might affect and alter the surface morphology of the specimens.
Environmental scanning electron microscopy connected with energy dispersive x-
ray analysis (ESEM-EDX) has been proposed to study calcium silicate Portland cement

*Laboratory of Biomaterials and Oral Pathology, Endodontic Clinical Section, Department of Odontostomatological Science, Alma Mater Studiorum - University of
Bologna, Bologna, Italy; †Department of Earth Sciences , Alma Mater Studiorum - University of Bologna, Bologna, Italy; ‡Department of Conservative Dentistry, Leuven BIO-
MAT Research Cluster, Catholic University of Leuven, Leuven, Belgium; and §Department of Biochemistry - Alma Mater Studiorum - University of Bologna, Bologna, Italy.
Address requests for reprints to Maria Giovanna Gandolfi, PhD, Department of Odontostomatological Science, University of Bologna, Via san Vitale 59, 40125
Bologna, Italy. E-mail address: mgiovanna.gandolfi@unibo.it.
0099-2399/$0 - see front matter
Copyright ª 2010 American Association of Endodontists.
doi:10.1016/j.joen.2009.12.007

JOE — Volume 36, Number 5, May 2010 Investigation of Endodontic Calcium Silicate Hydrated Cements 851
Basic Research—Technology
building materials (1, 19, 20). Recent investigations have described the
use of ESEM and its application in dentistry (21, 22). The hydration of
cements for biomaterial characterization might be also investigated
in situ and in real time by micro-Raman spectroscopy, with no sample
manipulation and minimal interference from environmental water (7,
23). Micro-Raman configuration allows the detection of the changes
in chemical composition on a microscale, because the laser spot (ie,
the excitation source) size is of the order of few microns.
In this study both ESEM-EDX and micro-Raman spectroscopy have
been innovatively proposed to investigate in real time the surface
morphology of wet endodontic cements (wet calcium silicate cements
for dentistry) in their humid ‘‘natural’’ state and to monitor the chemical
transformations of their dynamic surface immediately after preparation
and 24 hours of soaking in a simulated body fluid solution (Dulbecco
phosphate-buffered saline [DPBS]).
Materials and Methods
Cement Preparation and Characterization
Three calcium silicate cements were studied: (1) white ProRoot
MTA (Dentsply Maillefer, Tulsa, OK; lot no. 08003395), (2) white
tetrasilicate cement containing bismuth oxide (wTC-Bi), and (3) white
tetrasilicate cement without bismuth oxide (wTC). The powder of wTC
was constituted of a white Portland cement (CEM I, Aalborg, Denmark)
subjected to thermal and mechanical treatments and added with calcium
chloride, anhydrite, and montmorillonite. The bismuth-free wTC cement
was used as reference (1) to avoid the interferences of bismuth oxide
(added as radio-opacifier agent) with micro-Raman analysis (in partic-
ular with the detection of the portlandite band) and (2) to study the effect
of bismuth on bioactivity of calcium silicate cements.
The cements were mixed with DPBS (liquid/powder ratio of 0.3)
and layered on a plastic coverslip with 13-mm diameter (Thermanox
Plastic; Nalge Nunc International, New York, NY) to obtain standard
disks. The exposed surface area of the disks was 1.9  0.1 cm2, and
their thickness was approximately 0.9 mm. Mechanical vibrations
were used to obtain a flat and regular surface. ESEM-EDX and micro-
Raman analyses were carried out on the samples 10 minutes after their
preparation (freshly prepared group) and after aging in 5.2 mL soaking
solution (DPBS) at 37 C for 24 hours (24-hour-aged group).
ESEM-EDX
All samples were examined by the environmental scanning elec-
tron microscope ESEM Phillips FEG XL30 (Eindhoven, The
Netherlands) and by the ESEM Zeiss EVO 50 (Carl Zeiss, Oberkochen,
Germany) equipped with EDX by using an accelerating voltage of 20–25
kV. At 25 kV acceleration, the x-ray electron beam penetration of ESEM-
EDX (inside a material with a density of about 3 g/cm3) resulted in 2.98
mm, and consequently the volume excited and involved in the emission
of characteristic x-rays from the constituting elements must be consid-
ered 10 mm3.
Cement disks were placed directly onto the ESEM stub and exam-
ined without any form of preparation (the samples were not coated for
this analysis). All samples were initially analyzed at low-vacuum condi-
tions (9.9 torr, 500 Pa), 100% relative humidity (RH), and 4 C. After
the initial examination each sample was inspected at a higher vacuum
(2.9 torr, 74 Pa), 40% RH, 4 C.
Micro-Raman Spectroscopy
Micro-Raman spectra were obtained by using a Jasco NRS-2000C
instrument connected to a microscope with 20 magnification. In
these conditions the laser spot size (ie, the excitation source) was of
the order of a few microns. All the spectra were recorded in back-scat-
852 Gandolfi et al.
tering conditions with 5 cm 1 spectral resolutions by using the 488 nm
laser line (Innova Coherent 70; Coherent Inc, Santa Clara, CA) with
a power of 50 mW. A 160 K frozen CCD detector from Princeton Instru-
ments Inc (Trenton, NJ) was used.
The cements were analyzed as wet samples, when maintained in
their storage media. Regarding the specimens stored for 24 hours in
DPBS, at least 5 spectra were recorded on their upper surface to mini-
mize variability deriving from the possible sample inhomogeneity.
Raman spectra were also recorded on unhydrated cements.
Results
ESEM-EDX Analyses
Freshly Prepared Samples (Analyzed Ten Minutes after
Mixing). The experimental wTC cement was initially inspected at
9.9 torr (low vacuum), 100% RH, and 4 C. The surface was formed
by a hydrogel matrix (gel-like phase) covered by a thin water layer.
Calcium silicate granules (alite and belite) and needle-like crystals
(ettringite) were only partially visible because they were immersed in
the hydrogel structure (Fig. 1A). EDX showed Ca, Si, and O peaks, sug-
gesting the presence of calcium silicate particles and high amounts of
water (Fig. 1B). Samples were then examined at 4.1 and 3.0–2.9 torr
at low temperature (approximately 4 C) and at 40% RH. The increase
of vacuum caused environmental dehydration after physical squeezing
and reabsorption of gel phase. Water and amorphous hydrogel disap-
peared from the surface, and a granular superficial layer appeared,
composed by irregular rounded calcium silicate particles and elongated
needle-shaped calcium-aluminum-silicate hydrate crystals (ettringite)
that formed a complex network (Fig. 1C,D).
The experimental wTC-Bi cement showed sporadic reflective gran-
ules (bismuth particles) immersed in the needle-like crystals network
(2.9 torr). Water and hydrogel matrix were absent from the surface.
EDX analysis of the area displayed Ca, Si, Bi, and O peaks (figures
not reported).
ProRoot MTA observed at 400 Pa was covered by a water film, and
the hydrogel matrix was visible and masked the mineral phase of the
surface. At 300 Pa pressure a rough cement surface appeared after
the removal of the coating hydrogel. Superficial round-shaped
calcium-silicate mineral particles (alite, belite) with different dimen-
sions and shapes and small needle-like ettringite crystals and porosities
were visible (Fig. 1E). EDX analysis of the area revealed Ca, Si, Cl, Bi, and
O peaks (Fig. 1F).
Samples Aged 24 Hours in DPBS. At 9.9 torr only a flat wet
surface was visible on wTC as a superficial gel-like layer. At higher
vacuum, wTC surface showed different precipitates with different
dimensions and shapes and many nanospherulites (0.2–0.5 mm diam-
eter) packed to form clusters of aggregated spheroidal bodies evenly
distributed throughout the cement surface. At high magnifications
(25,600) they appeared as small apatite spherulites (Fig. 2A). EDX
showed Ca, Si, and P peaks, suggesting the presence of calcium phos-
phate deposits. Punctual microanalyses on spherulite clusters detected
high Ca and P peaks, whereas on prismatic granules only Ca peaks
(Fig. 2B).
At low vacuum (9.9 torr) a water film covered the surface of wTC-
Bi cement, and the mineral particles different in dimensions and shape
were embedded in a hydrogel matrix (Fig. 2C). The surface observed at
3.1 torr showed numerous porosities, deposits, and aggregates of small
round-shaped crystals, and residual hydrogel inside the niches was
present (Fig. 2D). EDX analysis of the area displayed Ca, Cl, O, Si,
and traces of Bi. Phosphorus peak appeared, and the presence of Si
demonstrated the uneven and not homogeneous distribution of spher-
ulitic deposits (Fig. 2E). At 3.0 torr, numerous small cloud-like deposits
JOE — Volume 36, Number 5, May 2010
 Basic Research—Technology

Figure 1. ESEM-EDX analyses of freshly prepared (10-minute mixed) cement surface. wTC cement (A–D). (A) Surface observed at low vacuum (9.9 torr) without
dehydration (1500, original magnification). A transparent water layer covers the cement surface. Calcium silicate particles (alite, belite) with different dimensions
and shapes and needle-like ettringite crystals (hydrated calcium-aluminum-sulfate hydroxide) are embedded in an amorphous hydrogel. (B) EDX analysis of the
area displayed the presence of Ca, Si, O, Cl, and traces of S (sulfur) and Al (aluminum). (C) Cement surface observed at 2.9 torr pressure. Round-shaped calcium
silicate particles were present, and elongated needle-like crystals created a network on the cement surface. Empty porosities not filled by the hydrogel were visible
on the surface. (D) High magnification of ettringite crystal. ProRoot MTA cement (E, F). (E) Surface observed at 300 Pa pressure. Round-shaped calcium silicate
particles (alite, belite) with different dimensions and shapes were present. Porosities were visible. (F) EDX analysis of the area showed the presence of Ca, Bi, Si, and O.

of aggregate apatite spherulites were distributed on the entire surface
(Fig. 2F).
The ProRoot MTA surface observed at low vacuum (500 Pa)
showed an amorphous hydrogel matrix (Fig. 3A). Needle-like crystals
and apatite spherulites were evenly distributed on the entire surface,
were visible at 300 Pa (Fig. 3B), and showed Ca, P, and traces of Si
and Cl (Fig. 3C). At low pressure (74 Pa) and high magnification
(10,000) the cloud-like deposits appeared composed of aggregates
of spherulites (Fig. 3D).
Micro-Raman Spectroscopy
Fig. 4 shows the Raman spectra recorded on the 3 freshly prepared
cements and after 24 hours of storage in DPBS. The bands assigned to
calcium carbonate, anhydrite, gypsum, belite, alite, bismuth oxide, and
apatite (7, 23) have been indicated.
The starting powders displayed the typical bands of Portland
cement components: alite, belite, and calcium sulfate. Calcium sulfate
was present only as anhydrite in Pro-Root MTA and as both anhydrite
and gypsum in wTC and wTC-Bi; moreover, MTA powder showed
a higher amount of calcium carbonate and lower quantities of calcium
sulfate (only as anhydrite). No phosphate band (965 cm 1) was
detected in the unhydrated powder of any of the cements.
The silicate component resulted in more crystalline structure
(higher definition of the 854, 839, and 827 cm 1 bands) in Pro-
Root MTA than in wTC and wTC-Bi, suggesting that the hydration reac-
tion of silicate component (alite and belite with consequent formation of
CSH phase) should be slower in MTA. This characteristic was confirmed
by the results obtained at longer storage times (unpublished results).
The micro-Raman spectra recorded on the surface of the freshly
prepared cements revealed the formation of ettringite on all cements,
as revealed by the appearance of the 990 cm 1 band. The portlandite

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Basic Research—Technology

Figure 2. ESEM-EDX analyses of cement surface stored 24 hours in DPBS at 37 C. wTC cement (A, B). (A) At 4.9 torr numerous small (0.2–0.5 mm) spheroidal
bodies of apatite deposited on the surface were packed and fused together to form clusters of aggregated spherulites evenly distributed throughout the cement
surface. High magnification (25,600) of apatite spherulites on cement surface. (B) EDX on apatite aggregates displayed prominent Ca, Si, and P peaks and traces
of Cl and Na (sodium). wTC-Bi cement (C–F). (C) At low vacuum (9.9 torr) a water film covered the cement surface. (D) Cement surface observed at 3.1 torr
showed the residual hydrogel inside the niches. (E) EDX analysis of the area displayed Ca, Cl, O, Si, and traces of P and Bi. (F) At higher magnification (25,600)
and 3.0 torr pressure the deposits appeared distributed on the entire surface and composed by aggregates of spherulites.

band at 360 cm 1 was detected only on the surface of wTC, but no
conclusion can be drawn on wTC-Bi and MTA as a result of the inter-
ference of bismuth oxide (strong wide bands of bismuth oxide super-
imposed to portlandite band).
The spectra recorded on the surface of the cements stored for 24
hours in DPBS (Fig. 4) showed the formation of a thin and homoge-
neous deposit of apatite (band at about 965 cm 1) and calcite (bands
at about 1085 and 280 cm 1) and/or aragonite (1085 cm 1). At 24
hours, the marker band of portlandite disappeared from the surface
of wTC, suggesting that at this stage it was released into the surrounding
water medium.
Discussion
The investigation of wet cements in real time in early setting stage is
difficult if not impossible with conventional SEM analysis. However,
ESEM can record specimens under moderate pressure in the chamber
(10–2.0 torr), at low temperature (ie, 4 C), and in 100% RH during the
observation period (18, 22). Under these conditions, wet hydrated
cements can be observed in their natural environment (ie, without
drying) in real time (for example, immediately after cement prepara-
tion). At low vacuum, a humid atmosphere prevents desiccation of
the samples, allowing them to remain in their wet state (22).
The study intended to evaluate the morphology and composition of
the cement surface produced by the interaction of ProRoot MTA and
calcium silicate cements with DPBS. Portland cement and MTA contain
the same active components, mainly tricalcium and dicalcium silicates.
When Portland cement is mixed with water, anhydrous oxides
(mainly CaO and SiO2) react with water to form a solution that contains
Ca2+, OH– and silicate ions. As the pH rises above 10 and the Ca2+
concentration exceeds 1 mmol/L, this solution becomes supersaturated
with respect to the precipitation of a CSH nanoporous phase. The

854 Gandolfi et al. JOE — Volume 36, Number 5, May 2010


Figure 3. ESEM-EDX analyses of ProRoot MTA surface after 24 hours of storage in DPBS at 37 C. (A) The amorphous hydrogel matrix was observed at low vacuum
(500 Pa). (B) Elongated structures and needle-like crystals (ettringite) and aggregates of apatite spherulites were noticed at 300 Pa. The deposits were evenly
distributed on the entire surface. (C) EDX analysis of the area displayed Ca, P, and traces of Si and Cl. (D) At high magnification (10,000) aggregates of spherulites
form the cloud-like deposits on the cement surface (74 Pa pressure).
transport process such as the permeation, diffusion, electromigration,
combined transport and binding involve CSH phase and surrounding
fluids (24). Nanoparticles of CSH form and grow around the cement
grains and form a network that fills the spaces between them. Also por-
tlandite Ca(OH)2 nucleates and grows in the available voids and pore
spaces (12). This network is established a few minutes after mixing
and progressively gains strength as hydration proceeds and more
CSH particles are formed. The gain in strength resulting from the
increase in the amount of CSH and the forces between CSH particles
are the key to the cohesion of early cement paste (25). Ettringite crystals
(hexacalcium aluminate trisulfate hydrate), rapidly formed by reaction
of tricalcium aluminate and calcium sulfate, retard the setting reaction
by the delay of calcium aluminate hydrates (CAH). Ettringite formation
generates expansive forces related to swelling. The formation of CSH by
the hydration of calcium silicate particles is related to the hardening
reaction, and the forces between CSH particles are the key to the cohe-
sion of early cement paste. Portlandite Ca(OH)2 is a soluble, mobile,
and reactive compound (5, 11, 24) and is released over time by the
cement and increases the alkalinity of the environment.
In the present study the formation of ettringite and CSH phase were
examined because they are related to the progress (development) of the
hydration reaction. The hydration reaction of studied cements
produced a calcium silicate hydrogel (CSH) and calcium hydroxide.
The former component is not easily detectable by Raman spectroscopy,
because it is characterized by a weak spectrum as a result of the amor-
phous nature of CSH. On the other hand, calcium hydroxide can be
easily identified through its marker band at 360 cm 1, provided that
the cement is devoid of bismuth oxide. Accordingly, portlandite was de-
tected only on the surface of wTC, 10 minutes after preparation, at the
early stage of the hydration process. At longer storage times (ie, 24
JOE — Volume 36, Number 5, May 2010
Basic Research—Technology
hours and more (5, 7)), this phase was no longer detected on the
surface of the cement. In fact, during the reaction the soluble calcium
hydroxide dissolves from the cement surface by a continuous washout
in solution. During the hydration and setting reactions, amorphous CSH
gel is deposited on the original sites of calcium silicate particles. ESEM
at low vacuum showed the presence of a semitransparent hydrogel that
fills all the porosities between ettringite, bismuth oxide, and calcium
carbonate granules. The removal of the hydrated gel by relatively higher
vacuum (ie, 3.1–2.9 torr) discovered an irregular surface with many
porosities and defects. Many investigations reported similar features
by SEM analysis (4, 19, 20, 26–31).
It is probable that during 24 hours of immersion in DPBS, the
superficial Ca-rich layer of cement adsorbs phosphate from the soaking
solution, creating the conditions for the formation of calcium phosphate
precipitates, well-detected by ESEM-EDX analysis and confirmed by
micro-Raman spectroscopy. At ESEM-EDX these precipitates appeared
as small microspherulites and aggregates of spherulites that partially fill
the walls of the superficial porosities. These findings were observed in
all 3 cements.
The mechanisms of fast apatite formation on calcium silicate
cements immersed in simulated body fluids (SBF) involve the leaching
of calcium (with local Ca supersaturation) from silicate hydrates of CSH
matrix and from the dissolution/leaching of portlandite. Apatite might
form by reaction of supplied calcium ions with the environmental phos-
phate ions; a dissolution-precipitation process and nucleation mecha-
nisms on cement surface (with a solid state diffusion-controlled
growth) are involved. Actually, functional groups such as Si-OH on
wet fresh cement surface have been shown to act as nucleation centers
for apatite precipitation (31–34). Moreover, calcium ions provided by
portlandite and cement matrix can react with the environmental
Investigation of Endodontic Calcium Silicate Hydrated Cements 855
Basic Research—Technology
Figure 4. Micro-Raman spectra of the 3 cements: unhydrated cements, freshly prepared (after 10 minutes) and after 24 hours of storage in DPBS. The bands due to belite
(B), alite (A), ettringite (E), anhydrite (An), gypsum (G), calcium carbonate (C), calcium phosphate (Ap), portlandite (P), and bismuth oxide (Bi) have been indicated.
carbonate ions to form a biologically active calcium carbonate superfi-
cial layer (8). Actually, micro-Raman spectroscopy showed the pres-
ence of calcium carbonate on the cement surface, mainly as calcite
and/or aragonite (7).
The bioactivity of calcium silicate cements and the in vitro apatite
layer formation on Portland cement immersed in SBF have been recently
demonstrated (4–7, 10, 11, 33). Silicon-rich materials such as calcium
silicate cements exhibited excellent ability to induce bone-like apatite
formation (12). Tay et al (5) observed the formation of calcium-deficient
apatite precipitates in Portland cement–PBS system. Gallego et al (30)
observed the presence of apatite-like and calcite crystals on set Portland
cement exposed to phosphate solution and CO2 and the apatite-like
phase formation on the carbonated substrates. Ding et al (33) reported
the simultaneous formation of apatite and calcium silicate hydrates
after mixing cement with a phosphate-buffered solution and observed
precipitated apatite spherulites after 1 hour of cement immersion.
The fast formation of apatite nanoparticles after 24 hours of soak-
ing in DPBS might support the biologic behavior of these cements when
in contact with bone tissue.
An essential requirement for a bioactive biomaterial is the forma-
tion of a biologically active bone-like apatite layer or carbonate-contain-
ing calcium-deficient apatite on its surface in a biologic environment
(35, 36). Bioactivity might give a significant clinical advantage and
might be correlated with optimal biocompatibility, osteconductivity,
and osteoinductivity (35, 36). Biomineralization is the process of form-
ing minerals in organisms, in which minerals nucleate and grow from
a supersaturated aqueous solution under biomimetic conditions. Bio-
mineralization widely exists in nature, and several important biomate-
rials such as calcium phosphates and calcium silicates with good
osteoconductivity have been reported (37, 38).
Hydraulic cements with bioactive properties due to the formation
of apatite and calcite might have many important clinical applications in
dentistry, enhancing fast bone regeneration and pulp healing. These
data might explain the excellent clinical behavior reported for ProRoot
MTA and other calcium silicate cements (12–15, 17, 39).
Conclusions
The study demonstrated that ESEM is a powerful tool for the obser-
vation of hydraulic cements like ProRoot MTA and Portland cements at
856 Gandolfi et al.
very early hydration/setting stages and allows the investigation of the wet
surface morphology in real time, without modifying the cement struc-
ture (1, 18, 20). In this way no artifacts related to sample preparation
and dehydration are present. The combination of low temperature and
low vacuum (ie, 9.9 torr) in the specimen chamber permits a high
relative humidity to be maintained, allowing the cement surface to
stay wet. ESEM demonstrated that wet calcium silicate cements possess
a superficial gel-like structure that envelops the surface particles
(calcium silicates and portlandite). Calcium phosphate nanoparticles
appear as deposits after 24 hours of immersion in DPBS. The presence
of bismuth did not alter this dynamic interaction.
Micro-Raman spectroscopy allowed the analysis in real time of the
chemical structure of the wet cements and the appearance of calcite/
aragonite and calcium phosphate deposits in all 24-hour-stored
samples.
This study showed the possibility to monitor in real time the trans-
formations of dynamic/moldable surface of endodontic calcium silicate
hydraulic cements in constant dynamic/active interaction with the envi-
ronment, as well as the unique bioactivity properties of calcium silicate
cements that are the ability to produce calcium phosphate spherulites,
precursors of apatite deposits on their surface. Their special properties
make them excellent materials for clinical use in contact with bone
tissue and blood as root-end filling, root-repair, and root-canal sealers.
Acknowledgments
The authors thank Simit Dental srl (Mantova, Italy) for
providing gift packs of ProRoot MTA. Many thanks to Professor
Rudy De Voz for the qualified and valued technical support and
to Dr Fabiola D’Amato for the secretarial support.
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