This document describes a novel preparation of magnetic hydroxyapatite nanotubes (MHAnt). Magnetite nanoparticles were embedded in a poly(caprolactone) nanofiber template using electrospinning. The nanofiber surface was activated and deposited with an apatite mineral phase through a solution-mediated process. After heat treatment, hollow nanotubes formed with hydroxyapatite as the outer shell and magnetite nanoparticles lining the inner shell. The nanotubes exhibited a ferromagnetic property with a saturation magnetization of 27.20 emu/g. The magnetic hydroxyapatite nanotubes could be useful for biomedical applications such as hyperthermia treatment of bone cancer.

1. A novel preparation of magnetic hydroxyapatite nanotubes
Rajendra K. Singh a,b
, Ahmed M. El-Fiqi a,b
, Kapil D. Patel a,b
, Hae-Won Kim a,b,c,
⁎
a
Institute of Tissue Regeneration Engineering (ITREN), Dankook University, South Korea
b
Department of Nanobiomedical Science, WCU Research Center, Dankook University Graduate School, South Korea
c
Department of Biomaterials Science, School of Dentistry, Dankook University, South Korea
a b s t r a c t
a r t i c l e i n f o
Article history:
Received 13 June 2011
Accepted 29 January 2012
Available online 4 February 2012
Keywords:
Ceramics
Nanomaterials
Magnetic materials
We report here the novel preparation of magnetic hydroxyapatite nanotubes (MHAnt). Poly(caprolactone)
(PCL)–magnetite nanoparticles (MNPs) composite nanofiber was used as a template for the MHAnt. The sur-
face of the composite nanofiber was activated in an alkaline solution and then an apatite mineral phase was
deposited through a series of solution-mediated processes. After heat-treatment at 500 °C, a hollow tube of
HA-MNPs was created in which HA formed an outer shell and most of the MNPs lined the inner shell surface.
The inner shell size was about 650 nm and the shell thickness was about 137 nm. The developed MHAnt
showed a saturation magnetization of 27.20 emu/g, exhibiting a ferromagnetic property. The newly devel-
oped MHAnt may be useful in biomedical applications such as hyperthermia treatment of bone cancer.
© 2012 Elsevier B.V. All rights reserved.
1. Introduction
Nanomaterials containing drugs and acting as drug-delivery sys-
tems offer great promise in medical applications [1,2]. Providing addi-
tional functionality to such systems including magnetic properties
opens novel opportunities to their therapeutic efficacy. Magnetic ma-
terials such as iron oxide nanoparticles have commonly been used in
medical fields such as magnetic resonance imaging (MRI) and drug
delivery vehicles for cancer treatment [3,4]. Additionally, some poly-
mer microspheres or microtubes with ferromagnetic properties have
recently been developed for cell separation and analysis, hyperther-
mia treatment of tumors and selective killing of cancer cells [5,6]. Of
particular interest for these magnetic nanomaterials is the possibility
to use external magnetic fields to guide the drug carriers to precisely
target areas of the body and thus significantly reduce unnecessary
damage to healthy tissue [7]. Moreover, applying the magnetic force
will facilitate hyperthermia therapy of target tissues.
For the treatment of bone repair and related cancer therapy, the
materials and carriers that are compatible to bone tissue is preferred.
Hydroxyapatite (HA) exhibits unique advantages when used in bone
reconstruction because its chemical structure is similar to the inor-
ganic composition of human bone; therefore, it has been the most
commonly used biomaterial for targeting bone repair [8,9]. Develop-
ment of HA into specific nanomaterial forms, including nanospheres,
nanofibers and nanotubes, has also shown promise for the develop-
ment of materials for use as delivery systems of drugs and proteins
and as matrices for bone cell regulation [10,11]. When therapeutic
agents are delivered through HA nanocarriers, the healing potential
of the defective or diseased bone should be greatly enhanced [10,11].
Here, we attempted to develop a nanotubular form of HA that also
has magnetic properties. The combination of magnetic properties with
the biocompatible HA composition is considered to open the door to a
new class of biofunctional nanomaterials targeting bone. The nanotub-
ular form of HA has previously been exploited in our group by using a
polymer nanofiber as a template [12]; therefore, we utilized that meth-
odology here. To impart magnetic properties, magnetite nanoparticles
were embedded within the HA nanotubes. The processing route to pro-
duce the magnetic HA nanotubes (MHAnt) is described and their useful
characteristics, including magnetic properties, were investigated.
2. Experimental
Magnetite nanoparticles were prepared according to the following
procedure: ferrous chloride tetrahydrate (FeCl2·4H2O) in 1 M HCl and
ferric chloride hexahydratate (FeCl3·6H2O) were mixed at room tem-
perature (Fe2+
/Fe3+
=½). The mixture was then dropped into 200 ml
of 1.5 M NaOH solution while stirring vigorously for about 30 min. The
resulting precipitate was then isolated using a magnetic field, after
which the solution was decanted by centrifugation at 8000 rpm.
The separation procedure was conducted twice, after which 200 ml
of 0.02 M HCl solution was added to the precipitate with continuous
agitation. The product was then separated by centrifugation and
dried at 40 °C. All steps were conducted under nitrogen gas. The
magnetite nanoparticles were dispersed in citric acid solution (0.05 M)
under magnetic stirring, and the pH was adjusted to 5.5 using NH3 solu-
tion (28 wt.%). After 4 h, the nanoparticles were precipitated in acetone
and then washed with acetone by magnetic decantation to remove the
redundant citric acid. The samples were then dried at 40 °C.
Materials Letters 75 (2012) 130–133
⁎ Corresponding author at: Institute of Tissue Regeneration Engineering (ITREN),
Dankook University, South Korea. Tel.: +82 41 550 3081; fax: +82 41 550 3085.
E-mail address: kimhw@dku.edu (H.-W. Kim).
0167-577X/$ – see front matter © 2012 Elsevier B.V. All rights reserved.
doi:10.1016/j.matlet.2012.01.129
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2. Nanofiber templates were prepared according to our previous
study, with slight modification [12]. A 10 w/v% suspension of
poly(caprolactone) (PCL, Mw=80,000) in dichromethane/ethanol
solvent which containing magnetite nanoparticles (20 wt.% relative
to PCL) was prepared. The mixture suspension was electrospun into
a random nanofiber mesh at a field strength of 15 kV/10 cm. The
nanofiber surface was activated in 2 M NaOH solution for 4 h, after
which they were washed and alternatively soaked in 150 mM CaCl2
and Na2HPO4 solution. This process was followed by an apatite miner-
alization within 1.5 times simulated body fluid (1.5× SBF) at 37 °C for
seven days. The specimen was then heat-treated at 500 °C for 5 h to
remove out the polymer phase and to form hollow nanotubes.
The crystal phase and chemical bond status of the samples were
characterized by X-ray diffraction (XRD, Rigaku) and Fourier trans-
form infrared (FT-IR, Varian 640-IR) spectroscopy, respectively. The
surface electrical potential of the nanotubes was examined using a
zeta potential measurement (Malvern Zetasizer Nano) at pH=7.0
and 25 °C. The instrument determines the electrophoretic mobility
of the particles automatically and converts it to the zeta (ζ) potential
using Smoluchowski's equation. The morphology of the samples was
characterized by scanning electron microscopy (SEM, Hitachi
S-3000H) and transmission electron microscope (TEM, JEOL-7100).
Based on the TEM images, the size of the magnetic nanoparticles
(MNPs) and the apatite nanotubes was measured, and the existence
of MNPs within the nanotubes was also confirmed. The magnetic
properties of the samples were studied using a superconducting
quantum interference device (SQUID, Quantum Design MPMS-XL7)
in an applied magnetic field of ±20 kOe at room temperature. The
magnetic properties of the particles were evaluated in terms of satu-
ration magnetization and area.
3. Results and discussion
The crystalline phases of the MHAnt were investigated with XRD
as shown in Fig. 1. For references, the MNPs and their composite
nanofiber with PCL before and after the mineralization were also
characterized. As expected, typical magnetite peaks (‘M’) were
Fig. 1. XRD patterns of the samples: (a) MNPs, (b) PCL-MNPs composite nanofiber,
(c) mineralized composite nanofiber and (d) MHAnt after heat-treatment at 500 °C. M:
MNPs; P: PCL; A: apatite. Dashed lines in (d) are HA reference peaks (JCPDS card No. 24–33).
Fig. 2. (a) TEM image of MNPs. (b) SEM image of composite nanofiber and inset TEM image showing the presence of MNPs on the surface region. SEM images of the mineralized
nanofibers (c) before and (d) after heat-treatment. Inset in (d) shows a hollow tubular structure.
131
R.K. Singh et al. / Materials Letters 75 (2012) 130–133

3. shown in the as-prepared iron oxide nanoparticles (Fig. 1(a)) and ad-
ditional PCL peaks (‘P’) were also shown in the composite nanofiber
(Fig. 1(b)). After mineralization, the poorly crystallized apatite
peaks at 2θ≈26° and 32° (‘A’) were well developed (Fig. 1(c)).
After heat-treatment at 500 °C, the apatite peaks became more pro-
nounced (Fig. 1(d)), and the magnetite peaks were also preserved.
There were no other peaks observed except those of apatite and
magnetite.
The morphologies of the produced samples were examined by SEM
and TEM, as shown in Fig. 2. The particle size of the magnetite nano-
particles was found to be 12±1.34 nm, as deduced from the TEM
image (Fig. 2(a)). The nanoparticles were observed to be easily dis-
persed in the dichloromethane/ethanol solvent where the PCL could
also be dissolved to prepare the composite suspension for electrospin-
ning. A nanofiber mesh with sizes of hundreds of nanometers was well
generated without any bead formation from SEM image (Fig. 2(b)). A
high magnification TEM image shown in the inset revealed that the
black dotted MNPs were primarily positioned at the surface region of
the nanofiber. This was likely responsible for the inorganic particles
being segregated at the border region of the fiber during the electro-
spinning process. Previously, we observed a similar phenomenon dur-
ing the electrospinning of a tricalcium phosphate sol and a polymer
mixture, where the inorganic sol was distributed outer surface region
while the polymer was inner [13]. This distribution of MNPs in large
quantity at the surface region is highly beneficial to obtain MHAnt.
The surface of the composite nanofibers was then functionalized
with apatite mineral as shown in the SEM image (Fig. 2(c)). Surface
activation with alkaline solution and further solution-mediated nucle-
ation and crystallization processes led to the successful deposition of
an apatite mineral phase on the nanofiber surface. Further heat-
treatment of the mineralized nanofiber at 500 °C revealed a hollow tu-
bular structure (Fig. 2(d)).
The high resolution image of the MHAnt was revealed by TEM
(Fig. 3). Hollow-structured long tubes were well developed and con-
sisted of a number of highly elongated mineral nanocrystallites with
a morphology similar to bone mineral and that generally observed in
apatite obtained by biomimetic processes. The width of the inner hol-
low part of the tube was measured to be 650 nm (±80 nm) and the
wall thickness was 137 nm (±24 nm). Although magnetite nanopar-
ticles were not clearly discerned from the apatite nanocrystallites,
some particulate forms (representative regions are indicated by ar-
rows) in dark-colored area are considered to indicate their presence.
The nanocrystallite morphology of the apatite embedded with the
MNPs was actually slightly different from the pure apatite without
the nanoparticles, as previously reported; a more dotted structure
was observed MHAnt than in pure HA nanotubes which have a more
plate-like structure. Therefore, we examined the atomic composition
of the areas with EDS. In addition to the Ca and P peaks, a high
level of Fe peaks was found, confirming the existence of iron oxide
MNPs.
The magnetic properties of the MHAnt were investigated using
SQUID magnetometry. The magnetization curve as a function of an
applied magnetic field (Fig. 4) shows a ferromagnetic property
(i.e. the narrow hysteresis loop and low coercivity) and a saturation
magnetization (Ms) of 27.20 emu/g. In fact, the Ms of pure magnetite
nanoparticles was measured to be around 71 emu/g, while that of
bulk magnetite is known to be 92 emu/g [14]. When compared to the
pure magnetite materials, the currently developed magnetic HA had a
much lower Ms value, which is easily understandable when considering
the decrease in the relative mass fraction of the magnetite. Although the
Ms value of the MHAnt did not reach that of pure magnetite, the prop-
erties revealed in this study including the ferromagnetic behavior and
fairly high Ms should indicate that the MHAnt can be used as biocom-
patible magnetic materials. Another important parameter for the mag-
netic characterization is the magnetic loss/cycle or the area of the
hysteresis loop. The integrated hysteresis loop area calculated for a
maximum applied field of 20 kOe was found to be 57.22×103
erg/g.
Since the area under the loop is proportional to the energy loss and
hence the heat generated by a sample under a magnetic field it is sug-
gested the nanotubes find possible uses in hyperthermia treatment.
Taken together, this study provides useful information of the applicabil-
ity of the developed HA hollow magnetic nanotubes in biomedicine, ei-
ther used directly as a delivery system of therapeutic molecules or as a
secondary phase in artificial bone matrices, to treat the bone disease
and trauma via hyperthermia treatment or other therapeutic efficacies
potentiated under magnetic fields.
4. Conclusions
Magnetic hydroxyapatite nanotubes (MHAnt) were produced
using a template made of magnetite nanoparticles/PCL polymer and
Fig. 3. TEM nanostructure image of the MHAnt and the EDS profile in the inset showing
the existence of Fe and thus MNPs. Arrows pointing to dark dotted areas indicate the
possible presence of MNPs.
Fig. 4. Magnetic properties of the magnetic HA nanotubes as determined by SQUID
magnetometry. The magnetization versus magnetic field curve shows that the nano-
tubes exhibit a ferromagnetic property with a saturation magnetization of
27.20 emu/g. The inset shows an enlargement of the curve in the initial range.
132 R.K. Singh et al. / Materials Letters 75 (2012) 130–133

4. its subsequent surface mineralization and thermal treatment. The
MHAnt showed a hollow tubular structure with an inner shell size
of about 650 nm and a shell thickness of about 137 nm. The MHAnt
exhibited a ferromagnetic property with a saturation magnetization
of 27.20 emu/g. The MHAnt developed herein may find usefulness
in many biomedical applications, including the treatment of bone re-
pair and related diseases.
Acknowledgments
This work was supported by the Priority Research Centers Pro-
gram (No. 2009–0093829) and WCU program (R31-10069) through
the National Research Foundation of Korea (NRF) funded by the Min-
istry of Education, Science and Technology. Authors highly thank the
help of IBST, Dankook University.
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