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Current Nanoscience, 2013, 9, 308-317
Optimization of Nano-Titania and Titania–Chitosan Nanocomposite to Enhance Biocompatibility
K. Kavithaa, M. Prabhua, V. Rajendrana*, P. Manivasankana, P. Prabua and T. Jayakumarb
a
b
Centre for Nano Science and Technology, K.S. Rangasamy College of Technology, Tiruchengode 637215, Tamil Nadu, India,
Metallurgy and Materials Group, Indira Gandhi Centre for Atomic Research, Kalpakkam 603102, Tamil Nadu, India
Abstract: Nano-structured titania (TiO2) has received considerable attention in the field of biomedical research because of its numerous
advantages and few health concerns. To produce good biocompatible biomaterial, we synthesized different structural nano-TiO2 and
TiO2–chitosan (CH) nanocomposite by precipitation, hydrothermal, and in situ sol–gel methods. The prepared samples were characterized comprehensively. Nano-TiO2 prepared by precipitation and hydrothermal methods (sintered at 673 K) showed an anatase phase with
spherical morphology, whereas precipitation at 393 K confirmed amorphous phase. In situ synthesized composites were amorphous and
had irregular morphology. The samples were further analyzed for in vitro bioactivity, antibacterial activity, and cytotoxicity. The characterization after bioactivity study confirmed formation of stoichiometric ratio of hydroxyapatite on TiO2–CH nanocomposite compared
with other TiO2 samples. No significant cytotoxic effect of all the samples on gastric adenocarcinoma (AGS) human cell line was observed. This study shows that method of synthesis has no significant effect on the bioactivity and antibacterial activity, whereas it influences cytotoxicity. However, TiO2–CH nanocomposite has an important function in conferring better biocompatibility. Thus, TiO2-based
polymer composites prepared with appropriate synthesis methods are recommended for implant and tissue engineering applications.
Keywords: Titania, titania–chitosan, cytotoxicity, biocompatibility, antibacterial activity, in vitro bioactivity.
INTRODUCTION
Because of increasing life expectancy, use of biomaterials for
fixing bone fractures is continuously increasing in orthopedics [1,
2]. Although biomaterials have wide applications in the biomedical
field, a complication arising from implant loosening limits their use
[3]. This complication is mostly due to postoperative infections [4]
and incompatibility with tissues. Appropriate bone fillers or coating
materials not only seal bone defects but also kill residual bacteria.
Hence, orthopedists require a suitable material having specific antibacterial property, good bioactivity, and osseointegration [1, 3].
Staphylococcus aureus is one of the most common pathogens
responsible for postoperative implant-related infections [4, 5].
Clinical pathogens such as S. epidermis, Pseudomonas aeruginosa,
Bacillus sp., Escherichia coli, and Klebsiella pneumoniae are also
associated with bone infections [3, 6]. These bacteria are often resistant to antibiotics and hence there is a need to develop a material
in order to prevent infections [5]. Generally, the antibacterial activity of nano-TiO2 increases when its concentration is increased.
However, a very high nano-TiO2 concentration is not recommended
because it is toxic [7]. Widely used implants (Ti and Ti alloys) also
require modification or coating with materials such as nano-TiO2
and TiO2-based composites to enhance their specific bactericidal
effect and osseointegration [8]. There are many reports on the antibacterial activity of TiO2 [5, 7], but studies on TiO2 with biodegradable polymer-based nanocomposite are scanty.
TiO2 has many applications in various fields, which are based
on surface area, crystallite size, porosity, mechanical properties,
morphology, and stability [9]. Of the various methods available to
synthesize TiO2 [9-12], precipitation and hydrothermal methods are
reliable for cell attachment because of their simple and feasible
process [11, 12]. Although nano-TiO2 has many advantages in implant and tissue engineering applications [13] (such as easy formation of hydroxyapatite (HAp) via Ti–OH site, corrosion resistance,
*Address correspondence to this author at the Centre for Nano Science and
Technology, K.S. Rangasamy College of Technology, Tiruchengode 637215,
Tamil Nadu, India; Tel: +91-4288-274880, 91-4288-274741-4;
Fax: +91-4288-274880; E-mail: [email protected]
1573-4137/13 $58.00+.00
high stability [14], and ability to accelerate bone growth), it has
some associated risk because of its rate of transport and nonbiodegradability [15, 16]. Hence, composites with their beneficial effects
such as less toxicity, biodegradability, and improved osseointegration have attracted much attention in biomedical research [17, 18].
Chitosan, a natural polymer, has considerable resemblance with
extracellular matrix elements and has promising biological properties such as antibacterial property, ability to accelerate wound healing, and antitumor activity [19, 20]. An earlier study reveals that
improved mechanical property is achieved by a strong interaction
between the polymer and nanoparticles [21]. The in situ sol–gel
method is a well-known method to produce composite that can be
used to improve bonding strength, thereby reducing nonselective
voids and controlling size distributions [19]. Thus, because of the
above-mentioned properties, nanoparticles induce osseointegration
and osteoconduction [18].
The objective of this investigation is to synthesize, characterize,
and analyze nano-TiO2 structures and TiO2–CH nanocomposite for
biomedical applications. Nano-TiO2 is synthesized by wet chemical
and hydrothermal methods whereas TiO2–CH nanocomposite is
synthesized using the in situ sol–gel method. To explore the biocompatibility of the prepared materials, we investigated antibacterial effect, bioactivity, and cytotoxicity using 1.5 simulated body
fluid (1.5 SBF), implant-related clinical pathogens, and the gastric
adenocarcinoma (AGS) cell line, respectively. The TiO2 prepared
by wet chemical method is compared with that prepared by hydrothermal method to analyze the biocompatibility. TiO2–CH nanocomposite is compared with pure nano-TiO2 structures to explain
the effect of methodology and preference of composites in enhancing biocompatibility.
MATERIALS AND METHODS
Synthesis of Nano-TiO2 by Precipitation Method
Nano-TiO2 particles were synthesized by precipitation method.
Titanium isopropoxide (Ti(OC3H7)4; Sigma-Aldrich; catalog no.
205273, 97%) was hydrolyzed in isopropyl alcohol (C3H7OH;
Merck, 95%) at 310 K with continuous stirring for 1 h and then
double-distilled water was added drop-wise into the solution with a
© 2013 Bentham Science Publishers
Biocompatibility of Different Titania Nanostructures and Composites
Current Nanoscience, 2013, Vol. 9, No. 3
309
weight ratio of 01:20:80 [12]. A precipitate of hydrous oxide was
formed during vigorous stirring for 4 h at 310 K. The precipitate
was washed and dried in a hot-air oven at 393 K for 1 h and then
ground to reduce the agglomeration. The prepared sample (hereafter
termed as TiA) was sintered at 673 K for 1 h at 5 K/min in a muffle
furnace and the powder (hereafter termed as TiC) was collected.
Nutrient broth (100 ml) was inoculated with 2 ml of the overnight
culture and incubated at 310 K for 3 h. After incubation, 0.1 ml
culture suspension was swabbed uniformly on a nutrient agar plate.
Sterile disks were coated with 50 mg samples predissolved in sterile
distilled water. These disks were placed on the nutrient agar plate
and then incubated at 310 K for 24 h.
Synthesis of Nano-TiO2 by Hydrothermal Method
Wet chemically prepared solution containing a mixture of titanium isopropoxide, isopropyl alcohol, and double-distilled water at
a weight ratio of 01:20:80 was treated using an indigenously designed high-pressure, high-temperature autoclave at 393 K at 60
rpm for 3–4 h for better nucleation of particles. It was then processed and sintered as mentioned in sample TiC (hereafter termed as
TiC1).
In Vitro Bioactivity Study
The bioactivity of the samples was studied using 1.5 SBF prepared according to the formulation and method developed by
Gerhardt et al. [24]. The solution was prepared with analytical
grade chemicals (Sigma-Aldrich and HiMedia). Samples (150 mg
each) were pressed into pellets and placed in 50 ml of 1.5 SBF in
polyurethane bottles and incubated for 3 weeks at 309.6 ± 1 K. The
changes in the pH value and conductivity in the 1.5 SBF were recorded regularly at an interval of 24 h using a pH and conductivity
meter (5 Star; Thermo Scientific Orion, USA). After 21 days, the
pellets were rinsed carefully with distilled water and ethanol. The
rinsed samples were vacuum dried and characterized again using
XRD, FTIR, and XRF studies to confirm the formation of the HAp
layer.
Synthesis of TiO2–Chitosan Nanocomposite by the In Situ Sol–
Gel Method
TiO2–CH nanocomposite was prepared by in situ sol–gel
method to achieve better bonding [19]. Titanium isopropoxide,
acetylacetone (C5H8O2; Merck, 98%), and isopropyl alcohol were
used at a molar ratio of 1.0:0.7:4.0. Addition of acetylacetone controls hydrolysis of titanium isopropoxide. One gram chitosan from
crab shells (catalog no. 41796-3) was dissolved in 2% acetic acid
and the solution was stirred for 5–6 h to achieve uniform dispersion. This solution was added to the above solutions while being
vigorously stirred to obtain gel-like constituents. The constituents
were dried for 3 h at 393 K to evaporate the solvent and the dried
compound was ground to obtain homogeneous powder (hereafter
termed as TiAC).
Characterization
The X-ray diffraction (XRD) patterns of all the prepared samples were evaluated using an X-ray diffractometer (X’Pert PRO;
PANalytical, the Netherlands) using CuK as a radiation source (
= 0.15406 Å). The average crystallite size was determined from the
corresponding X-ray spectral peaks using the following Scherrer
formula:
D=
k
cos
(i)
where D is the crystal size of the particles, k the Scherrer constant, the wavelength of CuK radiation (1.5406 Å), the full width at
half maximum, and the diffraction angle of the sample [12,22].
The specific surface area (SSA) of the crystalline samples, which
were uniformly spherical, was calculated using the formula S = 6 103/L, where S is the SSA (m2/g), L the average crystallite size,
and the density of TiO2 (3.84 g/cm3) [23]. The characteristic
peaks of all the prepared samples were obtained using a Fourier
transform infrared spectrometer (FTIR) (Spectrum 100; PerkinElmer, USA) using potassium bromide as a reference. A stable
colloidal solution was used for particle size distribution (PSD) of
the prepared samples using a particle size analyzer (Nanophox,
Germany) based on the dynamic light scattering technique at a scattering angle of 90°. Qualitative and quantitative elemental analyses
of the samples were carried out using X-ray fluorescence spectrometry (XRF) (EDX-720; Shimadzu, Japan) and energydispersive X-ray spectroscopy (EDX) (JSM 6360; JEOL, Japan)
studies. The surface morphology and primary particle size were
determined by scanning electron microscopy (SEM) (JSM 6390LV;
JEOL, Japan) and transmission electron microscopy (TEM)
(CM200; Phillips, USA).
Antibacterial Assay
Mother cultures of gram-negative bacteria (E. coli and K.
pneumoniae) and gram-positive bacteria (S. aureus and B. subtilis)
were inoculated and incubated overnight in nutrient broth at 310 K.
Cell Culture and Cytotoxicity Study
The cytotoxic effects of the prepared TiO2 and composite
nanoparticles were studied using the human AGS cell line (ATCC
1739). After some time the cells were inoculated in Dulbecco’s
modified Eagle’s medium and nutrient mixture of F12 Ham (1:1)
supplemented with 10% fetal bovine serum, sodium pyruvate, sodium bicarbonate, non-essential amino acids, glutamine, and penicillin (100 g/ml)/streptomycin (100 g/ml).
The toxicity of the samples was estimated using the MTT (3(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay
[17]. After attaining confluent growth, cells were seeded in 96-well
plates at a concentration of 1 103 per well and incubated for 24 h.
Then, the medium was replaced with a medium containing nanoparticles with different concentrations (0.2, 1, 20, and 100 g/ml). The
morphology of the cells was observed every 24 h using an inverted
tissue culture binocular microscope. The MTT solution (80 g) was
added to the well. After incubation for 4 h at 310 K, the solution
was replaced with dimethyl sulfoxide to dissolve the formazan crystals. The optical density (OD) was read at 570/630 nm to predict the
cytotoxicity. The percentage of cell viability with the triplicates was
calculated using the following formula:
OD of the nanoparticle-treated cells
100
OD of the untreated cells
(ii)
Data comparison between nanoparticle-treated and untreated
cell viability was carried out using one-way analysis of variance
followed by Tukey’s and Duncan’s tests with SPSS (version 16)
software. A p-value < 0.05 was considered statistically significant.
RESULTS AND DISCUSSION
Structural Analysis
The XRD patterns of the samples are shown in Fig. (1a). The
high-intensity peak appearing at 25.328° (2) is in accordance with
the characteristic diffraction peak of TiO2 anatase (JCPDS no. 211276). The different synthesis methods and their sample codes are
given in Table 1. The crystallite size is determined by the Scherrer
equation and the SSA calculated from the crystallite size is summarized in Table 2. The results indicate a decrease in crystallite size
and primary particle size with an increase in surface area. Owing to
the processing method, TiC1 shows reduced particle size and higher
surface area [11,25,26]. The addition of chitosan to TiO2 (TiAC)
does not show any shifts or changes in the TiO2 peak. In the XRD
Kavithaet al.
310 Current Nanoscience, 2013, Vol. 9, No. 3
Fig. (1). X-ray diffraction pattern of the different structural nano-TiO2 and TiO2–CH nanocomposite (a) before and (b) after in vitro bioactivity study.
Table 1.
Synthesis Method, Processing Temperature, Sample Name and Nature of the Prepared Samples
Sample name
Material
Synthesis methods
Processing temperature
Existing phase
(K)
TiA
TiO2
Precipitation
393
Amorphous
TiC
TiO2
Precipitation
673
Crystalline
TiC1
TiO2
Hydrothermal
673
Crystalline
TiAC
TiO2–CH
In situ sol gel
393
Amorphous
Table 2.
Crystallite Size, Specific Surface Area, Particle Size and Particle Size Distribution of nano–TiO2 and TiO2–CH
Nanocomposite
Crystallite size
Specific surface area
Particle size distribution (d50)
Primary particle size
(nm)
2
(m /g)
(nm)
(nm)
TiA
–
–
35.58
10.58
TiC
8.0
195.31
19.98
12.65
TiC1
4.6
339.67
32.63
7.50
TiAC
–
–
46.19
7.25
Sample name
pattern, the peak intensity and peak sharpness show that the sintered
samples do not have any impurities.
nucleation because of the release of Ca2+, Na+, or K+ ions [29].
TiAC shows a peak at 1392 cm1 due to the stretching vibrations of
the CH3 bands [19,29].
Functional Group Analysis
Fig. (2) shows FTIR spectra of all the samples. The characteristic peaks of TiO2 and the HAp layer are listed in Table 3. Fig. (2a)
shows that the broad peak obtained at 400–900 cm1 is the characteristic peak of Ti–O–Ti stretching mode [27]. The vibration bands
obtained at 1103, 1109, and 980 and 1117 cm1 are assigned to the
stretching and bending modes of CH3, Ti–O–C, and Ti–OH surface
groups, respectively [27,28]. The Ti–OH groups accelerate apatite
Particle Size and Particle Distribution
Fig. (3) confirms that the PSD of the synthesized samples is in
the range of 20–50 nm. Table 4 shows that TiC1 has a smaller primary particle size than TiC, which can be attributed to the synthesis
of nanoparticles under high pressure and temperature (Fig. 4).
However, the particle size analysis of TiC1 reveals a wider PSD
range than TiC. Similarly, the PSD is wider for the smaller-sized
Biocompatibility of Different Titania Nanostructures and Composites
Current Nanoscience, 2013, Vol. 9, No. 3
311
Fig. (2). Fourier transform infrared spectra of nano-TiO2 and TiO2–CH samples (a) before and (b) after in vitro bioactivity study.
Fig. (3). Particle size distribution of prepared nano-TiO2 and TiO2–CH nanocomposite.
particles. This can be attributed to the agglomeration of crystalline
particles [30]. The diffraction patterns of the prepared samples are
incorporated in TEM images, confirming the amorphous and crystalline nature of the samples as seen in XRD. Among them, the
composite shows a very small particle size, which can be attributed
to the in situ preparation. However, due to the macromolecular
nature of the chitosan, the PSD is higher. This study shows that
methods by which nanoparticles are processed play a dominant role
in determining the PSD.
Surface Morphology and Sample Purity
The SEM images show the particle morphologies. From Fig.
(5a–c), the spherical morphology for TiA, TiC, and TiC1 samples
with uniform size distribution can be confirmed. However, Fig.
(5d) shows an irregular morphology, which may be due to the interaction of chitosan with TiO2 (TiAC). The EDX results confirm
the purity of the chemically synthesized samples. The amorphous
nature of TiO2 (TiA and TiAC) consists of a negligible amount of
Kavithaet al.
312 Current Nanoscience, 2013, Vol. 9, No. 3
Table 3.
Infrared Frequencies and their Assignments of the Structural nano–TiO2 and TiO2–CH Composite
Vibration frequency (cm1)
Before SBF Study
Peak assignments
After SBF Study
TiA
TiC
TiC1
TiAC
TiA
TiC
TiC1
TiAC
607–791
786
657
629
785
745
579
610
Ti–OH surface group [28,27]
1223
1229
1117
980
1229
1231
H–O–H bending [27]
1626
1627
1620
1628
Anti symmetric stretching of C–O [32]
–
–
–
–
1055
1052
1062
1070
CH3 bands [19,29]
–
–
–
1392
–
1369
1384
–
CO3 band [31,33]
–
–
–
–
–
–
–
1415
Asymmetric vibrations of COO bands [29-33]
–
–
–
–
–
–
–
1539
Stretching vibrations of C=O bands [17,19]
–
–
–
–
1683
1639
1683
–
Ti–O–Ti stretching [27]
2–
Table 4.
Percentage of Calcium and Phosphate on nano–TiO2 and TiO2 –CH Nanocomposite
Before in vitro bioactivity
Sample
After in vitro bioactivity
name
Ca (%)
P (%)
Ca (%)
P (%)
Ca/P ratio
TiA
–
–
5.987
3.455
1.732
TiC
–
–
5.823
4.076
1.45
TiC1
–
–
5.567
2.949
1.887
TiAC
–
–
6.534
4.262
1.533
carbon because of improper solvent evaporation. However, after
sintering, the samples had no impurities (TiC1 and TiC). The EDX
results are correlated well with XRD studies.
In Vitro Bioactivity Study
After in vitro bioactivity study (1.5 SBF), HAp and TiO2 peaks
were observed in the XRD pattern (Fig. 1b). The XRD peaks observed at 25.354° (201), 25.879° (002), 31.775° (211), 46.675°
(222), 49.5° (213), and 76.1° (432) represent characteristic peaks of
HAp hexagonal phase (JCPDS no. 090432) [31]. However, owing
to domination of the TiO2 peak, few HAp peaks were overlapping.
From the XRD patterns, we could determine that TiAC had highintensity HAp crystalline peaks than TiA (amorphous) and TiC and
TiC1 (crystalline), which might be due to the variations in the formation of the HAp layer. It concludes that the synthesis method
does not show any significant difference in bioactivity. However,
the arrangement of atoms (crystalline and amorphous) in the sample
had notable difference because of the lattice match.
The characteristic peaks of TiO2 and the HAp layer after the
bioactivity study are shown in Fig. (2b). The stretching modes of
C–O, C=O, COO, and CH3 were observed [17,19,29-33], which
might be due to the deposition of ions on the surface as a result of
the presence of the Ti–OH group [14]. The band obtained at 1415
cm1 was assigned to CO32 because of the HAp layer formation
[31,33]. Earlier studies [25,34] revealed that the absorption bands at
1043, 963, 601–03, 566–68, 599, and 469 cm1 are the characteristic features of phosphate. According to our study results, the phosphate bands are dominated by the broad absorptions (400–900
cm1) of titanium bands. The FTIR results indicate that the compos-
ite sample (TiAC) has appropriate peaks for the HAp layer formation among the prepared samples.
During the bioactivity study, a gradual fluctuation in the pH
values was observed (Fig. 6a). This fluctuation was due to regular
exchanges of ions between the samples and 1.5 SBF. The conductivity measurements were correlated well with the pH measurements (Fig. 6b). It was evident that an increase in ion release in the
1.5 SBF leads to an increase in conductivity. Owing to the solubility of the samples, the pH value of the 1.5 SBF decreased immediately after soaking the sample [35]. On the 14th day, the ion release
was very low, which might be due to the deposition of supplementary ions on the surface of the samples to initiate the HAp layer
formation. After sufficient deposition of ions (14th day), the conductivity was increased gradually for 21 days. This was because of
the saturation in deposition of ions for the HAp layer formation on
the samples.
After the bioactivity study, the dry weight percentage of the
pellets was measured and compared with that measured on the first
day. From Table 5, it can be seen that the weight of the samples
(TiA, TiC, and TiC1) has undergone reasonable reduction. However, instead of weight loss, TiAC showed 1.29% increase in
weight, which can be attributed to the swelling property as a result
of the inclusion of polymer in the sample [36]. This property of
chitosan causes the TiAC sample to be completely swollen within a
few days and degraded gradually when compared with pure TiO2
samples [36].
Elemental Analysis
After the 1.5 SBF studies, each sample was found to contain its
own deposition of calcium and phosphate (Ca/P) according to their
Biocompatibility of Different Titania Nanostructures and Composites
Fig. (4). TEM images and corresponding diffraction pattern of nano-TiO2 and TiO2–CH nanocomposite.
Current Nanoscience, 2013, Vol. 9, No. 3
313
Kavithaet al.
314 Current Nanoscience, 2013, Vol. 9, No. 3
Fig. (5). SEM–EDX images of different structural nano-TiO2 and TiO2–CH nanocomposite.
bioactivity (Table 4). Depending on the Ca/P ratio, different HAp
phases [37] such as calcium-deficient HAp (Ca/P = 1.2–1.56) [22],
Oxy-HAp (Ca/P = 1.5–1.67), and carbonate-substituted HAp (Ca/P
=1.93 –1.67) are formed [22,38]. Generally, Oxy-HAp is the accepted Ca/P ratio for better HAp layer formation [39]. After the 1.5
SBF study, the percentage of Ca/P deposition is higher in TiC than
that in TiC1, whereas the appropriate Ca/P ratio is not attained in
the sample TiC (1.45). During the bioactivity study, TiC1 promotes
carbonate-substituted HAp formation (1.887) whereas TiA shows
calcium-deficient HAp formation (1.732), as evident from Table 4.
When compared with TiO2 samples, the composite has higher levels of deposition as well as appropriate Ca/P ratios (1.5330), which
help accelerate bone growth.
Antimicrobial Activity
The antibacterial effect of the samples is shown in Fig. (7). It
can be inferred that nano-TiO2 and its composite show specific
antimicrobial effect on S. aureus than on E. coli, K. pneumoniae,
and B. subtilis, which can be attributed to the characteristics of the
sample and differences in the bacterial cell wall [20]. Fig. (7a)
shows that TiC1 has a better antibacterial effect than TiC. It may be
due to increase in interaction with crystalline particles because of
the smaller primary particle size and better crystallization
[11,25,26]. The presence of chitosan in TiAC led to better antibacterial effect than TiA. When compared with crystalline TiO2, amorphous TiO2 shows good antimicrobial effect. The synthesis method
and the size of the primary particle also play an important role in
determining the antibacterial property. Owing to the excellent properties of chitosan and specific relationship with surface characteris-
tics of the bacterial cell wall, TiAC exhibits higher antibacterial
effect [20,38] among all pure TiO2 samples.
Cytotoxicity Study
The cell viability and cytotoxic effect of the samples are evaluated in the human AGS cell line. After sample incubation for 24 h,
little or no effect is observed using an inverted phase-contrast microscope, whereas after incubation for 48 h, a little difference in
cell morphology is observed and marked (Fig. 8). The toxic effect
of nanoparticles in the cell lines is estimated using the MTT assay
[17] (Fig. 9). No significant toxic effect is observed in any of the
samples, which is evident by statistical analysis. However, at a
concentration of 1 g/ml, all the prepared TiO2 samples have a
minor inhibitory effect on the viability of the AGS cells, except the
composite (TiAC). The hydrothermally synthesized sample (TiC)
has more viability when compared with other samples at a concentration of 20 g/ml. This can be attributed to the adherent property
of TiC in an optimal concentration and good viability because of
good crystalline formation [25] and higher surface area. In contrast,
TiAC shows no cell death and uniform increase in cell viability as
the concentration of the sample increases. The addition of nanoTiO2 in the chitosan controls and alters the swelling property, depending on the physiochemical condition [18, 40]. This swelling
property regulates the transport and may prevent the accumulation
of the particles in the body than the pure TiO2 nanostructures,
thereby controlling the interaction with the AGS cells.
The swelling property of the composite increases the surface
area, which, in turn, helps to improve the cell attachment and
Biocompatibility of Different Titania Nanostructures and Composites
Table 5.
Current Nanoscience, 2013, Vol. 9, No. 3
315
Weight Percentage of Structural nano–TiO2 and TiO2 –CH Nanocomposite Pellets
Weight percentage of samples
Sample name
Before in vitro bioactivity
After in vitro bioactivity
Weight modulations
(mg)
(mg)
(%)
TiA
143.38
139.77
–2.52
TiC
147.18
145.65
–1.04
TiC1
143.00
141.30
–1.19
TiAC
145.82
147.70
+1.29
Fig. (7). Antibacterial effects of different structural nano-TiO2 and TiO 2–CH
nanocomposite (C–(control sample), TiA, TiC, TiC1, and TiAC).
patibility. Hence, the cytotoxicity of nano-TiO2 depends not only on
the addition of chitosan and its crystalline nature [2] but also on the
formation of crystals and surface property, which are influenced by
synthesis methods.
Fig. (6). Ionic mesurmements of nano-TiO2 and TiO2–CH (1 mg/ml) nanocomposite in the 1.5 SBF: (a) pH values at the period of 21 days and (b)
conductivity vs soaking period.
growth in three-dimensional ways by absorbing more nutrients [18].
Owing to this behavior, the solubility of the sample was affected
during the in vitro study. The observed results conclude that TiO2–
CH nanocomposite and the different synthesis methods of TiO2
nanoparticles are vital in determining the cytotoxicity and biocom-
CONCLUSIONS
The bioactivity and biocompatibility of nano-TiO2 and TiO2–
CH nanocomposite were analyzed in the 1.5 SBF, bone-infecting
microorganisms, and the AGS cell line. The anatase phase of nanoTiO2 was successfully synthesized using wet chemical and hydrothermal methods and analyzed for biocompatibility. The results
show that different methods of synthesis are favorable for cell viability whereas bioactivity and antibacterial property experienced
only a slight effect. This is due to the increased interaction with the
cells because of the surface property, smaller primary particle size,
and better crystalline formation through synthesis. In addition, wellbonded TiO2–CH nanocomposite was obtained by the in situ sol–
gel method and biocompatibility was analyzed along with nanoTiO2. Collectively, these results conclude that the synthesis procedure and addition of constituents (chitosan) are crucial in biological
applications. However, nanocomposite materials facilitate a favor-
Kavithaet al.
316 Current Nanoscience, 2013, Vol. 9, No. 3
CONFLICT OF INTEREST
The authors confirm that this article content has no conflicts of
interest.
ACKNOWLEDGEMENTS
The authors acknowledge the financial support from the UGCDAE Consortium for Scientific Research, Kalpakkam Node, to
carry out this research project (CSR/Acctts/2010–11/1136 dt.
06.01.2011) and thank G. Amarendra, Head, Metal Physics Section,
Indira Gandhi Centre for Atomic Research, Kalpakkam, for his
valuable suggestions. The authors also thank G. Kumaresan and P.
Jayaprakash of the Department of Genetics, School of Biological
Sciences, Madurai Kamaraj University (MKU) for their help in the
cell line study with the support of the Department of Atomic Energy (DAE)–MKU research project on “Cancer Drug Discovery
Assay Development.”
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Fig. (8). Morphology of nano-TiO2 and TiO2–CH nanocomposite-treated
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[7]
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[11]
Fig. (9). Cytotoxicity study of nano-TiO2 and TiO2–CH nanocomposite by
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able environment for the HAp layer formation because of the swelling behavior of chitosan and the Ti–OH group of TiO2. In addition,
no toxic effect was observed in different concentrations of the composite samples. The TiO2–CH nanocomposite has specific antibacterial property toward bone-infecting microorganisms, lack of
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Received: July 1, 2012
Revised: October 4, 2012
Accepted: January 20, 2013
Current Nanoscience, 2013, Vol. 9, No. 3
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