J. Biomedical Science and Engineering, 2010, 3, 1117-1124
doi:10.4236/jbise.2010.312145 Published Online December 2010 (http://www.SciRP.org/journal/jbise/ JBiSE ).
Published Online December 2010 in SciRes. http://www.scirp.org/journal/JBiSE
Fabrication and characterization of cross-linked gelatin
electro-spun nano-fibers
Thi-Hiep Nguyen, Byong-Taek Lee
Department of Biomedical Engineering and Materials, College of Medicine, Soonchunhyang University, Cheonan, Korea
Received 5 Octorber 2010; revised 15 Octorber 2010; accepted 18 Octorber 2010.
ABSTRACT
In this study, we developed a fast, simple and novel
process to fabricate cross-linked electro-spun gelatin
with limited amounts of glutaraldehyde (GA) using
trifluoroacetic acid (TFA) as the solvent. Using SEM,
the uncross-linked gelatin fibers were determined to
have diameters between 50-300 nm, while the
cross-linked gelatin electro-spun fibers had diameters
between 100-500 nm. FT-IR revealed that the un-
cross-linked and cross-linked electro-spun gelatin
was fabricated successfully by electro-spinning using
TFA as a solvent, which has not been reported until
now. Stress-strain curves showed that the addition of
small amounts of GA increased the strength of the
gelatin by two fold and allowed for the creation of a
water insoluble gelatin electro-spun membrane.
Keywords: Gelatin, Cross-Linking, Glutaraldehyde
1. INTRODUCTION
Collagen is an important protein for cell proliferation
and is found in the extracellular matrix (ECM). M.D.
Johnson repor ted that collag en is one of the main f ibrous
elements in connective tissue [1]. Gelatin is a natural
biopolymer that is prepared by partial hydrolysis of col-
lagens. Most of these structural proteins are found in the
skin, tendons, cartilage, bones and connective tissues of
animals. Gelatin (type A, obtained from porcine skin,
~300 Bloom) prepared from acid-treated collagens has a
biocompatibility that is similar to collagen but is much
cheaper to produce. Therefore, gelatin is extensively
used in medical products, such as wound dressings [2],
drug delivery systems [3], nerves [4], etc. Gelatin has
been widely applied because it has three excellent fea-
tures; first, it has a high biocompatibility, biodegradabil-
ity and bioactivity [5]. Second, it can improve water
absorption of non-absorptive polymers such as poly-
caprolactone [6], poly (lactic-co-glycolic acid) (PLGA)
[4], etc. Finally, gelatin can react with -OH groups and
positively charged polymers such as polyvinyl alcohol
(PVA) [7], chitosan [8] to create 3-D porous sponges
where gelatin is involved in physical and/or chemical
reactions.
Electro-spinning has recently been shown to be a use-
ful method to generate fibrous scaffolds for tissue engi-
neering applications and has been used in a broad range
of different research fields [9]. Electro-spinning is an
inexpensive, effective, and simple method to produce
non-woven nano-fibrous mats, which have intrinsically
high surface to volume ratios, increased flexibility in
surface functionalities, improved mechanical perform-
ances, and smaller pores than fibers produced using tra-
ditional methods [10]. The advantages of the elec-
tro-spinning technique include the production of very
thin fibers that are on the order of a few nanometers or
micrometers with large surface areas, ease of function-
alization for various applications, superior mechanical
properties and ease of processing [11]. Decreasing the
fiber diameter within these mats causes many beneficial
effects, including increased specific surface area to vo-
lume ratios. Electro-spun scaffolds allow cells to grow
while providing sufficient mechanical support. Therefo re,
various biodegradable as well as synthetic polymers [12],
peptide copolymers [13], and natural proteins [14] have
been electro-spun into micro/nano-fibers for a multitude
of biomedical applications such as scaffolds for use in
tissue engineering [15], wound dressing [16], drug de-
livery [17], and vascular grafts [18]. The necessary com-
ponents of an electro-spinning apparatus include a high
power voltage supply, a capillary tube with a needle or
pipette, and a collector that typically consists of a con-
ducting material. [19] The solvent is the most important
factor in the electro-spinning operation. In previous stu-
dies, electro-spun gelatin was fabricated using various
solvents such as 2,2,2-triflourothanol (TFE) [20], wa-

T.-H. Nguyen et al. / J. Biomedical Science and Engineering 3 (2010) 1117-1124 1118
ter/water in acetic acid or mixed with ethyl acetate [21].
However, Trifluoroacetic acid (TFA) has not yet been
used as a solvent for the fabrication of electro-spun gela-
tin even though collagen [22] and chitosan [10] have
been fabricated using TFA as a co-solvent.
When the inherently beneficial effects of gelatin ma-
terial are combined with the enhanced properties of na-
no-fibrous mats, applications arise in a wide range of
tissue engineering fields. In this work, we developed a
novel one-step process to fabricate cross-linked elec-
tro-spun gelatin nano-fibers that was faster and more
economical than the previously reported two-step va-
por-cross-linking method [20]. In this study, the elec-
tro-spun gelatin was fabricated using TFA as a solvent
and cross-linked electro-spun gelatin was produced by
adding small amounts of glutaraldehyde (GA) to the
gelatin solution. Small amounts of GA were shown to be
nontoxic and increased the mechanical strength by
two-fold, relative to the uncross-linked gelatin elec-
tro-spun mats [3,5,23-25]. SEM morphology, DSC and
FT-IR demonstrated that uncross-linked and cross-linked
electro-spun gelatin was successfully fabricated by elec-
tro-spinning using TFA as a solvent. In the in vitro bio-
degradable studies, the morphology of the gelatin mat
was found to change due to swelling and biodegradabil-
ity. Thus, the cross-linked electro-spun gelatin holds
promise for use in tissue engineering and biomedical
applications.
2. MATERIALS AND METHODS
2.1. Materials
Gelatin (from porcine skin, type A) was purchased from
Sigma-Aldrich USA. Trifluoroacetic acid (TFA, CF3
COOH, 99.0%) was purchased from Duksan Pure
Chemical Co., Korea. Glutaraldehyde (GA) was ob-
tained from DeaJung Co., Korea. Fetal bovine serum
(FBS), P.S. (penicillin/streptomycin (antibiotics)), Dul-
becco’s phosphate buffered saline (D-PBS) without cal-
cium or magnesium, MTT solution and trypsin-EDTA
were purchased from GIBCO (Carlsbad, CA). The
L-929 cell line was obtained from the ATCC Cell Line
(CCL-1TM, NCTC clone 929 [L cell, L-929, derivative
of Strain L], Korea). DMSO (Dimethylsulfoxide 99, 0%)
was purchased from, Samchun Pure Chemical Co., LTD
(Korea).
2.2. Preparation of Polymer Solutions
A 15 wt % gelatin solution was prepared by adding a
TFA solution to 1.5 g of gelatin until the solution
weighed 10g. The gelatin was completely dissolved by
stirring for 6 hours. 1.5 ml of 0.05 % wt/v GA in PBS
was then added to 10 ml of the 15 wt % gelatin.
2.3. Electro-spinning Setting
The gelatin solutions were placed in a plastic syringe
(lure-lock type, 12 ml) that was fitted to a needle with a
25 gauge (inner diameter 0.25 mm) tip diameter. The
flow rate of the CS solutions (0.5 ml/h) was controlled
using a syringe pump (lure-lock type, Korea). The elec-
tro-spinning voltage (10kV) was supplied directly by a
high DC voltage pow er supply (NNC–30 kilovolts–2mA
portable type, Korea). A grounded steel plate located 15
centimeters away from the tip of the syringe needle was
used to collect the nano-fiber mats. The structure of the
gelatin fibrous mats fabricated with TFA as a solvent
was observed by SEM (SM-65F, JEOL, and Japan).
2.4. Characterizations
2.4.1. Fourier Transform-Infrared Spectra (FT-IR)
The cross-linked gelatin was characterized by attenuated
reflectance Fourier transform spectroscopy (Spectrum
GX, PerkinElmer, USA). The infrared spectra of the
samples were measured over a wavelength range of
4000-500 cm-1. All spectra were acquired in the spectral
range through the accumulation of 64 scans with a reso-
lution of 4 cm-1.
2.4.2. Tensil e Str e ngth
Tensile strength of all samples was tested using the R&B
Model Unitech Series (Universal Testing Machine). The
dimensions of the samples (width 2 mm, length 27 mm)
were measured using a digital micrometer and the
thickness (100 µm) was measured by SEM prior to the
measurement of tensile strength . Sample preparation and
measurement of the tensile strength have been described
in our previous reports [26].
2.4.3. Differential Scanning C alorimetr y (DSC)
DSC measurements (METTLER TOLEDO KOREA –
DSC822e) were conducted using a sample weight of 3-5
mg under a nitrogen atmosphere and at a scanning speed
of 10℃/min. The samples were heated from 0 to 250℃
at a rate of 10℃/min.
2.5. Characterization of Gelatin in Vitro Study
To characterize the in vitro properties of the ge latin mat,
9 samples for each group were cut into square shapes (1
cm x 1cm) and then immersed in simulated body fluid
(SBF) at 37℃ for 30 min, 120 min, 720 min and 1440
min.
2.5.1. Swelling Behavior of Gelatin Membranes
The swelling behavior of the gelatin mat was measured
by swelling the electro-spun mat in an SPF solution in a
humid environment (6% CO2, 37℃ in an incubator). The
weight of the dry gelatin mat (approximately 0 .05 g) and
the wet gelatin mat removed from the SPF solution at

T.-H. Nguyen et al. / J. Biomedical Science and Engineering 3 (2010) 1117-1124 111 9
different times was then determined. Their wet weights
were established after first blotting with a filter paper
followed by blowing with a stream of air to remove sur-
face water. The swelling ratio was calculated using the
following equation:
Esr (%) = ((Ws – Wd) / Wd) × 100 (1)
(Where Esr is the water absorption (% wt) of the
membranes, and Wd and Ws are the weights of the sam-
ples in the dry and swollen states, respectively).
2.5.2. Biodegradability of Gelatin in Vitr o
The biodegradability of the gelatin was assessed by
changes in the SEM morphology after different immer-
sion time periods.
3. RESULTS
3.1. SEM Morphology of Uncross-Linked and
Cross-Linked Electro-Spun Gelatin
The nano-morphology of the uncross-linked and cross-
linked electro-spun gelatin fibers is shown in Figure 1.
The electro-spun gelatin mats were fabricated using an
electro-spinning machine (NNC-30kV-2mA portable
type, Nano NC. Korea.). The parameters used for the
fabrication of each polymer fibrous mats were as follows:
voltage 10 kV, distance 15 cm, flow rate 0.5 ml/hour and
nozzle 25 G. The morphology of the gelatin nano-fibers
electro-spun from gelatin in TFA without GA is shown in
Figure 1a. In this case, the fiber diameter was 50-300 nm.
The random arrangement of the fibers mimicked the
behavior of extra cellular matrix (ECM) proteins, which
provides mechanical support and regulates cellular ac-
tivities [27]. However, electro-spun gelatin is water so-
luble [25], which is not suitable for long-term applica-
tions. To overcome this drawback, GA was added to the
gelatin solution as described above. The na-
no-morphology of the gelatin-GA electro-spun mat is
shown in Figure 1(b) and Figure 1(c). Figure 1(b)
shows the SEM morphology of the electro-spun gelatin
fabricated with 15 wt % of gelatin and 1.5 ml of GA
(0.05 wt %) in PBS without storage in a vacuum oven.
Figure 1(c) shows the SEM morphology of sample B,
which was maintained in a vacuum oven at 100℃ for 24
hours. While GA is considered a creep cross-linking
agent [28], and was added to the gelatin before elec-
tro-spinning, GA could not cross-link all gelatin because
of its highly acidic properties. However, when the elec-
tro-spun gelatin was stored overnight, the acid evapo-
rated and GA was able to more extensively cross-link the
gelatin. The electro-spun gelatin maintained at room
temperature was referred to as cross-linked A, while the
electro-spun gelatin maintained in a vacuum oven at
100℃ was referred to as cross-linked B. GA remained
(a)
(b)
(c)
Figure 1. Uncross-linked electro-spun (a) cross-linked A gela-
tin electro-spun after keeping 1 day at room temperature (b)
and cross-linked B gelatin after keeping 1 day in vacuum oven
at 100 °C (c).

T.-H. Nguyen et al. / J. Biomedical Science and Engineering 3 (2010) 1117-1124 1120
on the surface of the gelatin fibers after electro-spinning
and the fibers were cross-linked, as previously reported
for chitosan [10]. The diameter of the electro-spun gela-
tin-GA was between 100-500 nm, which was larger than
the un-cross-linked fibers.
3.2. Characterization of Un-Cross-Linked and
Cross-Linked Electro-Spun Gelatin
Nanofibers through FTIR Analysis
FTIR measurements were conducted on un-cross-linked
gelatin and cross-linked gelatin to determine whether
cross-linking of the electro-spun gelatin affected the
primary gelatin structure (Figure 2). The FTIR spectrum
of the un-cross-linked electro-spun gelatin revealed a
structure that was similar to raw gelatin, which we de-
scribed in a previous report [25]. This demonstrated that
TFA did not alter the structure of raw gelatin. The
un-cross-linked electro-spun gelatin had an amide I peak
(C = O stretch) at1636-1640 cm-1, amide II peak (N-H
bend and C-H stretch) at 1542 -1544 cm-1, amide III peak
(C-N stretch plus N-H in phase bending) at 1240 cm-1
and amide A peak (N-H stretching vibration) at 3300
cm-1, which are the distinguishing features of gelatin.
The spectrum of the cross-linked gelatin is shown in
greater detail in Figure 3. In addition to the previously
mentioned peaks, a strong peak at 1450 cm-1 was ob-
served in the cross-linked gelatin due to aldimine ab-
sorption [3]. In comparison to previous reports [3,5,29],
we found that the amide II peak changed from smooth to
several small peaks. Glutaraldehydes have an aldehyde
group (-CHO) that reacts with the amino group of the
lysine residues of proteins [5]. The un-cross-linked and
cross-linked membranes were discriminated by a slow
change in color from white to yellow. The color change
occurred because the aldimine linkage (CH = N) reac-
tions took place during the cross-linking process.
This is the first report where uncross-linked elec-
tro-spun gelatin and cross-linked electro-spun gelatin,
through the addition of glutaraldehyde, were fabricating
using TFA; however, TFA had been previously u sed as a
solvent in the fabrication of electro-spun collagen [22].
The characteristic absorption of the aldimine groups
occurred at 1450 cm-1. Furthermore, additional peaks
were observed at 1470-1570 cm-1, which increased as
the cross-linking reaction proceeded (Figure 2). To more
conclusively determine if cross-linked gelatin was suc-
cessfully fabricated, we expanded the FT-IR spectra
from 1100-1750 cm-1 (Figure 3). In this region of the
FT-IR spectra, the amount of CH=N groups increased.
Moreover, some peaks appeared in this region was simi-
lar to previous repot [5]. Thus, both un-cross-linked and
cross-linked electro-spun mats gelatin were successfully
fabricated via the electro-spinning method using TFA as
Figure 2. FT-IR of uncross-linked gelatin fibers (a) and cross-
linked-B gelatin fibers (b).
Figure 3. FT-IR of uncross-linked gelatin (a) and cross-linked-
B gelatin electrospun fibers (b).
a solvent and the differences in the spectra may have
resulted from the broad background underneath each
spectrum. This method allows one to control the addition
of very small amounts of glutaraldehyde during the
cross-linking process, which is an essential condition to
limit the insolubility and increase th e biocompatibility o f
gelatin for applications in the biomedical field [23].
3.3. Mechanics of Uncross-Linked and
Cross-Linked Gelatin Nanofiber
Electrospun by Tensile Strength of
Thin Membrane
Stress-strain curves were recorded using a R&B Model
Unitech Series (Universal Testing Machine) and the cells
were loaded following ASTM Standard D882-01 proto-
col [30] for thin membranes. For the tensile strength
measurements, the samples were prepared with strip-
shape dimensions of 0.1 mm thick, 25 mm long and 2

T.-H. Nguyen et al. / J. Biomedical Science and Engineering 3 (2010) 1117-1124 1121
mm wide. The samples were then attached to the paper
frame as described in reference [26]. The ratio of the
stretching process was set at 0.1 mm/min.
The stress of the cross-linked electro-spun gelatin in-
creased approximately 2 times that of the uncross-linked
electro-spun gelatin (Figure 4). The uncross-linked
electro-spun gelatin was fabricated using TFA as a sol-
vent, which is more economical than 2,2,2-trifluoroe-
thanol, and the tensile strength was similar to previous
reports with a stress around 1 MPa and a strain around
20% [20]. Moreover, the strain of the cross-linked elec-
tro-spun gelatin was 140% higher than the uncross-
linked electro-spun gelatin 5 times, which was also
higher than the samples subjected to the immersion me-
thods at the same concentration as described in ref [23].
However, the stress of the cross-linked electro-spun ge-
latin (2 MPa) was not significantly higher as described
in ref [20]. The biocompatible properties of the cross-
linked gelatin depended on the glutaraldehyde concen-
tration, which was why only small amounts of GA were
used in this study. This method is advantageous in that
one can easily and reliably control the glutaraldehyde
concentration, which is important for biomedical and
biocompatibility ap plications.
Controlling the amount of GA not only decreases tox-
icity but also increases the biocompatibility of cross-
linked electro-spun gelatin membranes and the me-
chanical strength, which is essential to applications in-
volving implantation [23,24,31]. Figure 5 shows the
changes in the morphology of the gelatin and the tensile
fracture surfaces of uncross-linked and cross-linked
electro-spun mats. Uncross-linked ge latin had brittle and
abrupt break properties (Figure 5(a)). In contrast, the
cross-linked gelatin had elongated and flexible proper-
Figure 4. Tensile strength of uncross-linked gelatin (a) and
cross-linked-B gelatin electro-spun fibers (b).
(a)
(b)
Figure 5. SEM morphology of uncross-linked gelatin (a) and
cross-linked gelatin mat at the point of fracture after tensile
strength (b).
ties (Figure 5(b)). Remark arrow (P) show the direction
of load cells applied.
3.4. Differential Scanning Calorimetry (DSC)
DSC investigations are widely used to examine changes
in the thermal properties of initial components and new
components created after a reaction.
Figure 6 shows the DSC thermograms of gelatin
powder (a), uncross-linked electro-spun gelatin mats (b)
and cross linked electro spun gelatin mats (c). Two peaks
were observed in the DSC scans of the gelatin powder,
while only one peak was observed at approximately
95.8℃ for the cross-linked gelatin sample, which corre-
sponded to the endothermic peak of cross-linked gelatin
nanofibers. In contrast, the endothermic peak of un-
cross-linked gelatin was observed at approximately 75℃.
Therefore, because of the cross-linking reaction, the Tg
value of the uncross-linked gelatin mat changed from

T.-H. Nguyen et al. / J. Biomedical Science and Engineering 3 (2010) 1117-1124 1122
Figure 6. DSC thermograms of gelatin powder, uncross-linked
electro-spun gelatin mats and cross-linked electro-spun gelatin
mats.
75℃ to 95.8℃.
3.5. Characterization of Gelatin Electro-Spun in
in Vitr o Study
3.5.1. Swelling behavior of gelatin mats
To measure the swelling ratio of uncross-linked and
cross-linked electro-spun mats, the swelling ratio was
determined using the procedures outlined in the Materi-
als and Methods. As shown in Figure 7, the swelling
ratio of the cross-linked electrospun gelatin mat (a) was
twice that of the uncross-linked electro-spun gelatin mat
(b). These results demonstrated that the swelling of the
electro-spun gelatin was restricted due to the cross-
linking reaction. This property is highly important for
implantation applications. The ratio of both cross-linked
and uncross-linked electro-spun gelatin mats demon-
strated that the electro-spun mat was stable between
200-min and 1000- min.
3.5.2. Biod egradability
Figure 8 shows the SEM morphology of the cross-
linked electro-spun mats that had been immersed in the
SBF solution for varies periods of time. The SEM mor-
phologies of the cross-linked electro-spun gelatin that
had been immersed in SBF for 30 min, 120 min, 720
min and 1440 min are shown in Figure 8(a), Figure
8(b), Figure 8(c) and Figure 8(d), respectively. These
images show that most of the cross-linked gelatin elec-
tro-spun fibers remained intact; however, some regions
of the electro-spun gelatin fibers had melted. Neverthe-
less, even after 1440 min, most of the electro- spun fibers
still remained intact. In the biodegradability test, no
changes in biodegradability were observed when GA
was added to the gelatin solution before electro-spinning.
Figure 7. Swelling behavior of uncross-linked electro-spun
gelatin (a) and cross-linked electro-spun gelatin (b) after im-
mersing in the SPF solution for various periods of time.
(a) (b)
(c) (d)
Figure 8. SEM morphology of cross-linked electro-spun gela-
tin after immersing in the SPF solution for 30 min (a), 120 min
(b), 720 min (c) and 1440 min (d).
4. DISCUSSION
Gelatin derived from collagen is a highly effective fi-
brous matrix for cell attachment. In addition , gelatin has
excellent biocompatibility, biodegradability and bio-
absorption, and is widely used in biomedical, biomate-
rial, and tissue engineering applications. Therefore, sci-
entists have focused on using this material as scaffolds
in tissue engineering and biomedical regeneration appli-
cations. Electro-spun gelatin is similar to the collagen
ECM in the human body, which is the main matrix for
cell attachment and ground substance of polysaccharides,

T.-H. Nguyen et al. / J. Biomedical Science and Engineering 3 (2010) 1117-1124 1123
proteins and water [1]. However, gelatin is a water- so-
luble material; hence uncross-linked electro-spun gelatin
is also water-soluble. To maintain the gelatin ECM, ge-
latin electro-spun f ibrous mats should be cr oss-linked. In
this work, glutaraldehyde was used as a cross-linking
agent. However, the amount of glutaraldehyde used for
cross-linking should be limited because glutaraldehyde
is a toxic chemical. Previous studies cross-lin ked gelatin
through the evaporation method. The evaporation me-
thod has two drawbacks: the first is that the cross-link ing
agent used to cross-link the gelatin molecule from the
surface to the inside requires large amounts of GA. Sec-
ond, it is difficult to control the amount of GA, which is
toxic at concentrations higher than 5%. To overcome
these complications, we developed a novel method for
fabricating cross-linking gelatin fibrous mats by adding
low amounts of GA in a controlled manner to the gelatin
solution, which dissolves in strong acid. This acid effect
limits the cross-link ing agent from cross-linking all gela-
tins during the electro-spinning process but this effect
can be eliminated by storing at room temperature or in a
vacuum oven after completion of the electro-spinning
process. Finally, in this study, we obtained uncross-
linked electro-spun gelatin and cross-linked electro-spun
gelatin fibers that had nanometer scale diameters. How-
ever, the diameter of the cross-linked electro-spun gela-
tin fibers increased from 50-300 nm to 100-500 nm
compared with the uncross-linked electro-spun gelatin
mats (Figure 1) Successful cross-linking of the gelatin
membrane was confirmed by FT-IR, as shown in Figure
2 and Figure 3. Stress-strain curves showed that the
cross-linked electro-spun gelatin membrane was stronger
and more flexible. In addition, it was not brittle as we
previously reported for the electro-spun cross-linked
gelatin formed by the evaporated method (Figure 4). To
confirm the results obtained from the stress-strain curves,
surface fractures of the electro-spun gelatin were ob-
served by SEM (Figure 5). In the DSC thermograms of
uncross-linked and cross-linked electro-spun gelatin, an
increase in the Tg of the cross-linked electro-spun mat
was observed, demonstrating that the cross-linking agent
successfully cross-linked the gelatin fiber (Figure 6).
Prior to using biomaterials in implantation applications,
the swelling ratio and biodegradability must be investi-
gated. Therefore, the swelling ratios and biodegradabil-
ity of electro-spun gelatin mats were investigated before
the material was tested in vitro (Figure 7 and Figure 8,
respectively).
5. CONCLUSION
In this study, we developed a novel process to fabricate
cross-linked electro-spun gelatin by adding GA to the
gelatin solution before electro-spinning. In this work,
TFA was used for the first time as the solvent to fabri-
cate the electro-spun gelatin. This was also the first time
that GA was added to the gelatin solution before elec-
tro-spinning to cross-link the gelatin. The FT-IR data
confirmed that TFA did not alter the amide groups of the
gelatin and that the mat was successfully cross-linked.
The advantages of this method are: 1) the amount of GA
added to the solution was easily controlled during the
cross-linking process, which limits the cytotoxicity of
GA activity. 2) GA can disperse onto the gelatin fibers
and be cross-linked inside the fibers, which help main-
tain the ECM structure. 3) The small amount of added
GA increased the mechanical strength of the material by
two fold, and allowed for the formation of a flexible
water-insoluble cross-linked gelatin electro-spun mem-
brane.
6. ACKNOWLEDGEMENTS
This work was supported by the Mid-career Research Program through
a NRF grant funded by MEST (NO 2009–0092808).
REFERENCES
[1] Johnson, M.D. (2000) Human Biology: Concenpts and
Current Issues. San Francisco, California.
[2] Choi, Y.S., Hong, S.R., Lee, Y.M., et al. (1999). Study on
gelatin-containing artificial skin: I. Preparation and char-
acteristics of novel gelatin-alginate sponge. Biomaterials,
20, 409-417.
[3] Akin, H. and Hasircl, N. (1995) Preparation and Charac-
terization of Crosslinked Gelatin Microspheres. Journal
of Applied Polymer Science, 58, 95-100.
[4] Li, X.-K., Cai, S.-X., Liu, B., et al. (2007) Characteristics
of PLGA-gelatin complex as potential artificial nerve
scaffold. Colloids and Surfaces B: Biointerfaces, 57,
198-203.
[5] W., Y.Z., W., Y.L., C., G.X., et al. (2000) Preparation and
characterization of gelatin gel with a gradient structure.
Polymer Internationa, 49, 1600-1603.
[6] Chong, E.J., Phan, T.T., Lim, I.J., et al. (2007) Evalua-
tion of electrospun PCL/gelatin nanofibrous scaffold for
wound healing and layered dermal reconstitution. Acta
Biomaterialia, 3, 321-330.
[7] Yurong, L., Luke, M.G., Kennedy, J.E., et al. (2009)
Thermal behavior and mechanical properties of physi-
cally crosslinked PVA/Gelatin hydrogels. Journal of the
Mechanical Behavior of Biomedical Materials, doi:
10.1016/j.jmbbm.2009.07.001.
[8] Nagahama, H., Maeda, H., Kashiki, T., et al. (2009)
Preparation and characterization of novel chitosan/gelatin
membranes using chitosan hydrogel. Carbohydrate Po-
lymers, 76, 255-260.
[9] Li, M., Guo, Y., Wei, Y., et al. (2006) Electrospinning
polyaniline-contained gelatin nanofibers for tissue engi-
neering applications. Biomaterials, 27, 2705-2715.
[10] Schiffman, J.D., and Schauer, C.L. (2007) One-Step
Electrospinning of Cross-Linked Chitosan Fibers. Bio-

T.-H. Nguyen et al. / J. Biomedical Science and Engineering 3 (2010) 1117-1124 1124
JBiSE
macromolecules, 8, 2665-2667.
[11] Agarwal, S., Wendorff, J.H. and Greiner, A. (2008) Use
of electrospinning technique for biomedical applications.
Polymer, 49, 5603-5621.
[12] You, Y.L., Lee, S.J., Min, B.M., et al. (2006) Effect of
solution properties on nanofibrous structure of elec-
trospun poly (lactic-co-glycolic acid). Journal of Applied
Polymmer Science, 99, 1214-1221.
[13] Mark, M., Naphtali, O.C., Soumen, M., et al. (2005).
Saccharide–Peptide Hybrid Copolymers as Biomaterials.
Angewandte Chemie International Edition, 44, 6529-
6533.
[14] Buttafoco, L., Kolkman, N.G., Engbers-Buijtenhuijs, P.,
et al. (2006) Electrospinning of collagen and elastin for
tissue engineering applications, Biomaterials, 27, 724-734.
[15] Pham, Q.P., Sharma, U. and Mikos, A.G. (2006) Electro-
spinning of polymeric nanofibers for tissue engineering
applications: A review. Tissue Engenineering, 12,
1197-1211.
[16] Duan, B., Yuan, X., Zhu, Y., et al. (2006) A nanofibrous
composite membrane of PLGA–chitosan/PVA prepared
by electrospinning. European Polymer Journal, 42,
2013-2022.
[17] Kenawy, E.R., Bowlin, G.L., Mansfield, K., et al. (2002)
Release of tetracycline hydrochloride from electrospun
poly(ethylene-co-vinylacetate), poly(lactic acid), and a
blend Journal of Controlled Release, 81, 57-64.
[18] Shudong, W., Youzhu, Z., Hongwei, W. et al. (2009)
Fabrication and Properties of the Electrospun Polylac-
tide/Silk Fibroin-Gelatin Composite Tubular Scaffold.
Biomacromolecules, 10, 2240-2244.
[19] Huang, Z.-M., Zhang, Y.Z., Kotaki, M., et al. (2003)
Composite Science Technology, 63, 2223-2253.
[20] Zhang, Y.Z., Venugopal, J., Huang, Z.-M., et al. (2006)
Crosslinking of the electrospun gelatin nanofibers. Po-
lymer, 47, 2911-2917.
[21] Song, J.-H., Kim, H.-E. and Kim, H.-W. (2008) Produc-
tion of electrospun gelatin nanofiber by water-based
co-solvent approach. Journal of Materials Science: Ma-
terials in Medicine, 19, 95-102.
[22] Chen, Z.G., Wang, P.W., Wei, B., et al. (2010) Elec-
trospun collagen–chitosan nanofiber: A biomimetic ex-
tracellular matrix for endothelial cell and smooth muscle
cell. Act a Biomaterialia, 6, 372-382.
[23] Bigi, A. and Cojazzi, G. (2001) Mechanical and thermal
properties of gelatin films at different degrees of glu-
taraldehyde crosslinking. Biomaterials, 22, 763-768.
[24] Gilberto, G., Junior, E.M., Marcantoö nio, R. A. C., et al.
(1999) Biocompatibility studies of anionic collagen
membranes with different degree of glutaraldehyde
cross-linking. Biomaterials, 20, 27-34.
[25] Sisson, K., C.Z., Farach-Carson, M.C., Chase, D.B. and
Rabolt J.F. (2009) Evaluation of cross-linking methods
for electrospun gelatin on cell growth and viability. Bio-
macromolecules, 10, doi: 10.1021/ bm900036s.
[26] Nguyen, T.H. and Lee, B.-T. (2010) Electro-spinning of
PLGA/PCL blends for tissue engineering and their bio-
compatibility. Journal of Materials Science: Materials in
Medicine, 21, 1969-1978.
[27] Min, B.-M., You,Y., Kim , J.-M., at el. (2004) Formation
of nanostructured poly(lactic-co-glycolic acid)/chitin
matrix and its cellular response to normal human kerati-
nocytes and fibroblasts. Carbohydrate Polymers, 57,
285-292.
[28] Martucci, J.F., Ruseckaite, R.A. and V´azquez, A. (2006)
Creep of glutaraldehyde-crosslinked gelatin films. Mate-
rials Science and Engineering A, 435-436, 681-686.
[29] Talebian, A., Kordestani, S.S., Dadashian F. et al. (2007)
World Applied Sciences Journal, 2, 575-581.
[30] ASTM Standard (1991) Standard test method for tensile
properties of thin plastic sheeting. In: American Society
for Testing and Materials, West Conshohocken, Ed.
194-202.
[31] Russell, A.D., Dancer, B.N. and Power, E.G.M. (1991)
Effects of Chemical Agents on Bacterial Sporulation,
Germination and Outgrowth. In: Denyer, S.P. and Hugo,
W. B. E d s. , Mechanisms of Action of Chemical Biocides:
Their Study and Exploitation, Blackwell Scientific Pub-
lications, Oxford, 23-28.