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ORIGINAL PAPER
In situ production of nanocomposites of poly(vinyl alcohol)and cellulose nanofibrils from Gluconacetobacter bacteria:effect of chemical crosslinking
Cristina Castro • Arja Vesterinen • Robin Zuluaga •
Gloria Caro • Ilari Filpponen • Orlando J. Rojas •
Galder Kortaberria • Piedad Ganan
Received: 25 October 2013 / Accepted: 20 January 2014
� Springer Science+Business Media Dordrecht 2014
Abstract Nanocomposites of poly(vinyl alcohol)
(PVA) reinforced with bacterial cellulose (BC) were
bioproduced by Gluconacetobacter genus bacteria. BC
was grown from a culture medium modified with
water-soluble PVA to allow in situ assembly and
production of a novel nanocomposite that displayed
synergistic property contributions from the individual
components. Chemical crosslinking with glyoxal was
performed to avoid the loss of PVA matrix during
purification steps and to improve the functional
properties of composite films. Reinforcement with
BC at 0.6, 6 and 14 wt% content yielded nanocom-
posites with excellent mechanical, thermal and dimen-
sional properties as well as moisture stability. Young’s
modulus and strength at break increased markedly with
the reinforcing BC: relative to the control sample (in
absence of BC), increases of 15, 165 and 680 % were
determined for nanocomposites with 0.6, 6 and 14 %
BC loading, respectively. The corresponding increase
in tensile strengths at yield were 1, 12 and 40 %,
respectively. The results indicate an exceptional rein-
forcing effect by the three-dimensional network struc-
ture formed by the BC upon biosynthesis embedded in
the PVA matrix and also suggest a large percolation
within the matrix. Bonding (mainly hydrogen bonding)
and chemical crosslinking between the reinforcing
phase and matrix were the main contributions to the
properties of the nanocomposite.
Keywords Nano composites � Poly(vinyl
alcohol) � Bacterial cellulose �Gluconacetobacter medellinensis � Thermal
stability � Mechanical properties
Introduction
Various composite materials reinforced with cellulose
have emerged in light of recent advances in the areas
of nanotechnology and bioengineering (Samir et al.
2004; Habibi et al. 2010; Iwamoto et al. 2007;
C. Castro � R. Zuluaga (&) � G. Caro � P. Ganan
School of Engineering, Universidad Pontificia
Bolivariana, Circular 1 # 70-01, Medellın, Colombia
e-mail: [email protected]
A. Vesterinen
Department of Biotechnology and Chemical Technology,
School of Chemical Technology, Aalto University,
P.O. Box 16100, 00076 Espoo, Finland
I. Filpponen � O. J. Rojas
Department of Forest Products Technology, School of
Chemical Technology, Aalto University, P.O. Box 16100,
00076 Espoo, Finland
O. J. Rojas
Departments of Forest Biomaterials and Chemical and
Biomolecular Engineering, North Carolina University,
Campus Box 8005, Raleigh, NC 27695, USA
G. Kortaberria
‘‘Materials?Technologies’’ Group, Chemical and
Environmental Engineering Department, Universidad del
Paıs Vasco, 20018 San Sebastian, Spain
123
Cellulose
DOI 10.1007/s10570-014-0170-1
Iwamoto et al. 2005). The most abundant source of
cellulose is vascular plants wherein it forms an
intermixed and tight system with hemicelluloses and
lignin. Chemical, enzymatic or mechanical treatments
(or their combinations) are required to deconstruct the
basic components of the cell walls (Janardhnan and
Sain 2006; Zuluaga et al. 2009). Many of these
processes deteriorate the fiber and produce microfi-
brils with a wide size dispersion and high tendency to
agglomerate, which may limit the performance of
composites obtained after their incorporation and
homogenization into a matrix by compression, injec-
tion, extrusion or casting (Bondeson et al. 2006;
Berglund 2005).
One possible solution to this problem is the use of
cellulose of bacterial origin (bacterial cellulose, BC).
BC is excreted extracellularly by bacteria of the genus
Gluconacetobacter (formerly Acetobacter) by enzy-
matic mechanisms that lead to the polymerization of
glucose into chains and to their assembly into bundles
of microfibrils and ribbons (Iguchi et al. 2000; Klemm
et al. 2001). In general, BC has the same chemical
composition as cellulose from vascular plants, but BC
is produced free of other polymers (such as hetero-
polysaccharides or lignin); thus, BC is chemically pure
and forms fibrils of uniform lateral dimensions (Brown
et al. 1976; Nakagaito et al. 2005). In addition, BC has
excellent mechanical strength, high water-holding
capacity and crystallinity (Bielecki et al. 2005).
Bacterial cellulose is produced as a film infinitely
interconnected as a percolating cluster. Such a
network has been the subject of investigations on the
effect of water-soluble substances that are added to the
culture medium, mainly in relation to the rate of
cellulose synthesis and fibril dimensions (Uhlin et al.
1995; Tokoh et al. 2002). Brown and Laborie (2007)
adopted a biomimetic approach for the production of
nanocomposites of BC and poly(vinyl alcohol) (PVA)
as well as polyethylene oxide (PEO). It was found that
the reinforcing BC phase was evenly distributed over
the entire matrix. However, loss or solubilization of
the polymer matrix (PVA or PEO) occurred upon post
processing (washing and purification). This observa-
tion was ascribed to the fact that the PVA or PEO
matrix and the reinforcing BC interacted via hydrogen
bonding, which was insufficient to prevent polymer
solubilization (Laborie 2009). Consequently, the
purity and functional properties of the nanocomposites
were affected by the presence of substances from the
culture medium, such as sugars, organic acids and
proteins. Gea et al. (2010) prepared in situ PVA/BC
composites through the addition of PVA into the
culture media and compared them with materials
obtained after impregnation of BC gels with PVA
solutions. Compared to composites prepared in situ,
those prepared by impregnation have a higher PVA
content (3.7 compared to 1.4 PVA wt%) for the same
initial component composition. Obviously, the differ-
ence was due to the effect of purification. Interest-
ingly, the in situ process resulted in composites with
better mechanical and optical properties due to the
more effective component intermixing and homoge-
nization. Other composites with BC as the primary
component have been obtained after immersion of BC
in solutions of host compounds such as acrylic acid,
gelatin, silanol, phosphate and fibrin (Choi et al. 2004;
Yasuda et al. 2005; Lin et al. 2009; Barud et al. 2007;
Brown et al. 2011; Wan et al. 2006). In situ manufac-
ture of composites of BC and PVA and other
polymers, such as acemannan (from aloe vera),
chitosan and starch, during BC synthesis was recently
reported (Saibuatong and Phisalaphong 2010; Phisal-
aphong and Jatupaiboon 2008; Grande et al. 2008).
However, a distinctive feature in these systems was
again the fact that the host polymer was partially or
fully removed during the purification process; there-
fore, as was the case in impregnation processes, BC
was the main constituent of the final material. Overall,
it is not surprising that the properties that have been
assessed in BC-based composites are mainly attributed
to the BC component and affected only slightly by the
residual polymer.
In the present work, we prepared PVA/BC nano-
composites in situ followed by chemical crosslinking.
The culture medium of a new strain of Gluconacetob-
acter bacteria (Castro et al. 2012, 2013) was modified
with PVA that was added together with a crosslinking
agent to act as the matrix (and main component) of the
composite. Therefore, cellulose ribbons were synthe-
sized by the bacteria in contact with the PVA resulting
in highly crosslinked nanocomposites that resisted
matrix material (PVA) losses during washing and
purification processes. The main morphological and
physical properties of the obtained composites were
determined and discussed as a function of BC content.
To the best of our knowledge, no reports are
available describing the BC synthesis in the presence
of a polymer matrix with simultaneous crosslinking to
Cellulose
123
yield highly reinforced matrices that would otherwise
be solubilized (for example, during purification steps).
Materials and methods
Pre-conditioning of the microbial strain
The recently discovered Gluconacetobacter strain
used in this study was isolated from a pellicle of
homemade vinegar (Castro et al. 2012, 2013). The
purified bacterium was incubated in a static Hestrin-
Schramm culture medium (HS) containing 2 w/v %
glucose, 0.5 w/v % peptone, 0.5 w/v % yeast extract
and 0.27 w/v % Na2HPO4, adjusted to pH 3.5 by
phosphoric acid and autoclaved at 121 �C. The
toxicity of the crosslinking agent (glyoxal) to bacteria
was minimized by a methodology that is currently in
the patent process.
In situ production of PVA/BC nanocomposites
Poly(vinyl alcohol) (Sigma-Aldrich, Saint Louis, MO)
with a reported molecular mass of 146-186 kDa and
98–99 % hydrolysis degree was added to the HS
medium under stirring at 90 �C to obtain aqueous
solutions of 0, 3, 4.5 and 6 wt% PVA concentrations.
The respective solution was allowed to reach room
temperature, and respective volumes of glyoxal aque-
ous solution (40 wt%) were added under stirring until
a final glyoxal content of 10 wt% with respect to PVA
was reached. The modified HS media was inoculated
with 10 v/v % of the preconditioned inoculum and
statically incubated for 8 days at 28 �C. The collected
pellicles were dried at 40 �C for 48 h and then cured
(crosslinked) at 120 �C for 5 min. The crosslinked
pellicles were then washed with distilled water,
immersed for 14 h in 5 wt% aqueous KOH solution
and finally rinsed until reaching pH 7 to remove any
bacteria and residual components from the culture
media.
Six sample batches were prepared for each PVA
concentration; three of the samples were crosslinked,
and the remaining ones were used as reference to
quantify (gravimetrically) the amount of cellulose in
the nanocomposites. Control samples of crosslinked
PVA (in the absence of BC) were also prepared
following the same procedure used for the manufac-
ture of the PVA/BC nanocomposites. A schematic
diagram summarizing the manufacture procedures and
obtained samples is provided in Fig. 1. The different
specimens are referred to as ‘‘PVA X’’ or ‘‘PVA/BC X’’
reference (matrix) or composite samples, respectively;
here ‘‘X’’ refers to the actual reinforcing BC content
on a dry basis of the final solid material, which was
measured gravimetrically, as noted in Table 1 and
elsewhere in the discussion.
Morphology on PVA/BC nanocomposites
Scanning electron microscopy (SEM, Jeol JSM 5910
LV operated at 10 kV) was used to image the fracture
surfaces of dry nanocomposites deformed in tension.
Before SEM analyses, all specimens were precondi-
tioned at 75 % RH (relative humidity) using an
atmosphere saturated with NaCl solution for 8 days
and then coated with gold/palladium using an ion
sputter coater for 5 min.
Chemical characterization
Attenuated total reflection Fourier transform infrared
spectroscopy (ATR-FT-IR) was used to identify the
main chemical features of the PVA/BC nanocompos-
ites. Before the measurement, the nanocomposites were
dried for 2 h at 40 �C to precondition the test samples.
ATR-FT-IR spectra were recorded on a Nicolet 6700
spectrophotometer in the 4,000–400 cm-1 range ATR
with a diamond crystal. The spectra were recorded with
a resolution of 4 cm-1 and an accumulation of 64 scans.
Mechanical properties
Tensile strength was measured on specimens cut into
dimensions according to the ASTM D-1708 standard
and preconditioned at 24 �C and 75 % RH. The
strength at break and elastic modulus were determined
with an Instron Instrument according to ASTM D-882
using a load cell of 200 N at 5 mm/min and 22 mm
grip distance.
Dynamic mechanical analysis (DMA) with humid-
ity control was carried out with a Q 800 DMA (TA
Instruments) equipped with a humidity chamber.
Oscillatory measurements (1 Hz frequency) were
performed at 23 �C using 5.3 mm 9 12-mm strips
cut from the crosslinked nanocomposites and refer-
ence films. Samples were loaded to the chamber and
conditioned in 0 %RH for 240 min. Thereafter, two
Cellulose
123
humidity cycles of varying % RH (between 0 and
90 %) were applied using 480 min as equilibrium
time. The data were collected every 2 s until reaching
equilibrium.
Water absorbency
Water absorption was evaluated after immersion of the
samples in pure water at room temperature for 48 h.
All samples were cut into the same dimensions (circles
with 20 mm in diameter), dried (40 �C for 48 h) and
weighed before immersion. The weight gain after
immersion and removal of excess water with an
absorbent paper were determined. The water uptake
per gram of dry sample (swelling fraction) wc was
calculated using the equation:
wc ¼ ðws � wdÞ=wd ð1Þ
where ws and wd are the weights of the samples after
swelling and drying, respectively.
Thermal properties
Thermogravimetrical analysis (TGA, Mettler Toledo)
was performed to study the thermal degradation
behavior of the composite samples. The TGA appa-
ratus was flushed with nitrogen atmosphere, and
10 mg of sample was used. Each specimen was heated
from room temperature to 800 �C at a rate of 10 �C/min.
Differential scanning calorimetry (DSC, Metter Toledo)
was used to acquire thermograms under N2 flow.
Samples (5 mg) were placed in hermetically closed
Fig. 1 Scheme for the
production of
nanocomposites PVA/BC
during cellulose synthesis
by Gluconacetobacter
bacteria
Table 1 Nanocomposite composition and reference names
Nanocomposite Reference PVA in
culture
medium
(wt%)
BC
content
(wt%)
Glyoxal
in culture
medium
(wt%)
PVA/BC 0.6 PVA 0.6 6 0.6 0.60
PVA/BC 6 PVA 6 4.5 6.0 0.60
PVA/BC 14 PVA 14 3 14.0 0.30
Cellulose
123
DSC crucibles and heated from -40 to 250 �C at
10 �C/min to erase the thermal history, after which
they were cooled to -40 �C at 10 �C/min. The
second heat scan was conducted from -40 to 250 �C
at 10 �C/min, and the glass transition temperature (Tg)
was taken as the inflection point of the specific heat
increment at the glass-rubber transition, while the
melting temperature (Tm) was taken as the peak
temperature of the melting endotherm.
Two crystallinity parameters were determined from
Eqs. 2 and 3; the first one was calculated from the
sample weight (Xc), and the other one (Xp) took into
account the amount of matrix material in the compos-
ite. In these equations DHm� = 161.6 Jg-1 is the heat
of fusion for 100 % crystalline PVA (Roohani et al.
2008), and w is the weight fraction of polymeric
matrix material in the composite.
Xc ¼ DHm=DH0m ð2Þ
Xp ¼ Xc=w ð3Þ
Dielectric spectroscopy
Dielectric spectroscopy measurements were carried
out with a Novocontrol Alpha high-resolution dielec-
tric analyzer performing temperature sweeps from
-50 to 150 �C at a constant frequency of 1 kHz with a
heating rate of 3 �C/min. The instrument was inter-
faced to a computer and equipped with a Novocontrol
Novocool cryogenic system for temperature control.
Results and discussion
Poly(vinyl alcohol)/bacterial cellulose nanocompos-
ites were produced during the biosynthesis of cellulose
in static conditions by bacteria of the genus Glu-
conacetobacter. PVA added to the culture medium
was crosslinked with the reinforcing BC fibrils by
glyoxilation to improve bonding and prevent losses of
the PVA matrix during material washing and purifi-
cation. Table 1 includes the composition of the
nanocomposites obtained from different PVA and
BC ratios (glyoxal was always added at 10 % based on
PVA mass). The precursor PVA concentration in the
culture media was initially 3, 4.5 or 6 wt% resulting in
nanocomposites with BC weight percent based on
total dry mass of 14, 6 and 0.6 wt%, respectively.
Therefore, the composites were manufactured with
PVA as the main component and BC as the minority,
reinforcing phase. As noted, a decrease in BC
production was observed in the presence of larger
amounts of PVA in the culture medium, mainly owing
to the increased viscosity of the medium, which
hindered the transfer of microorganisms to the surface
where they consume oxygen for metabolism. Addi-
tional factors that may prevent BC production include
the possibility of PVA acting as a barrier to oxygen
diffusion through the medium and the relatively large
concentration of glyoxal used, which can limit the cell
growth. More importantly, there is an indication that
nanocomposites with different components can be
manufactured from a wide range of reinforcing/matrix
compositions, for example, through the addition of
water-soluble polymers to the culture medium at
different concentrations.
It is well known that light scattering in composite
materials increases with the amount of reinforcing
material caused by differences in the refractive indexes.
Furthermore, composites reinforced with BC have been
shown to display optical properties that depend heavily
on the refractive indexes of the components (Yano
et al. 2005; Iwamoto et al. 2005). In the present case,
the transparency of films produced with the PVA
matrix after crosslinking was not affected by the
presence of BC (Fig. 2a, b). The composite retains the
transparency of the PVA matrix even at cellulose
concentrations of 14 wt%, largely because of the
intimate contact and strong interfacial adhesion
between the ultrafine cellulose nanofibrils and PVA
matrix. Compared to films of pure BC (Fig. 2c), the
crosslinked composites exhibit yellowing. This is due
to the presence of glyoxal. Thus, during the crosslink-
ing reaction, the samples take on a yellowish appear-
ance. Figure 3 includes ATR-FT-IR spectra of PVA,
BC and PVA/BC nanocomposites after the respective
washing and purification steps. Nanocomposite spectra
(Fig. 3b–d) include the characteristic bands of PVA
observed in Fig. 3a. The characteristic cellulose signals
of C–O–C pyranose ring skeletal vibration at 1,060 and
1,030 cm-1 (indicated by the dotted line) is observed in
the spectrum of BC (Fig. 3e), but only observed in the
composite highly loaded with BC. This is mainly
because of the amount of reinforcing BC relative to
PVA in the nanocomposite; when the cellulose content
decreases the intensity of the band also decreases. In
addition, PVA presents characteristic bands at the same
wavelength, which creates overlap bands (see spectra
Cellulose
123
for PVA/BC 6 and PVA/BC 0.6). Unfortunately, the
esterification band from carbonyl groups at 1,730 cm-1
generated by the reaction of PVA and glyoxal was not
evident, mainly because this reaction yields a five-
member ring whose bands overlap with those of PVA
(Choi et al. 1999). Interestingly, the relative intensity of
the band at 1,140 cm-1 corresponding to symmetric
C–C stretching increases with BC content from the
PVA/BC 0.6 to the PVA/BC 6 composite but decreases
in PVA/BC 14. This band is strongly related with the
PVA crystallinity (Choi et al. 1999; Ngui and Malla-
pragada 1998; Peppas and Hansen 1982; Kenney and
Willcockson 1966). Therefore, it is possible that the
presence of BC in the matrix improves the PVA
crystallinity up to a given concentration, after which it
decreases. This decrease in crystallinity with cellulose
addition has been observed in the case of PVA
reinforced with CNCs (Peresin et al. 2010).
Results from tensile tests of PVA and PVA/BC
composite films are included in Fig. 4. Under the same
testing conditions, the reference matrices display a
highly elastic behavior. The nanocomposite remains
ductile after incorporation of BC; for example,
samples PVA/BC 0.6 have a yield point at an
elongation close to that of the reference film. The
nanocomposite eventually becomes very stiff when
the BC content increases to 14 % (PVA/BC 14). It is
observed that both Young’s modulus (YM) and
strength at break increase markedly with the reinforc-
ing BC: relative to the control sample (in absence of
BC), YM increases of 15, 165 and 680 % are
determined for nanocomposites with 0.6, 6 and 14 %
BC concentration, respectively. The corresponding
increase in tensile strengths at yield are 1, 12 and
40 %. The results indicate an exceptional reinforcing
effect by the three-dimensional network structure
formed by the BC upon biosynthesis embedded in the
PVA matrix and also suggest a large percolation
within the matrix (Samir et al. 2005). In addition, it is
likely that after crosslinking covalent bonds were
formed between the reinforcing BC and the PVA
matrix.
The observed increases in mechanical strength
have not been reported for PVA reinforced with
different cellulosic elements (Zimmermann et al.
2004; Cheng et al. 2009; Lu et al. 2008; Roohani
et al. 2008; Lee et al. 2009; Zhang et al. 2011; Qiu and
Fig. 2 Visual appearance of PVA 14 without BC (a), PVA/BC 14 nanocomposite (b) and bacterial cellulose (c) crosslinked films
Fig. 3 ATR-FT-IR spectra of crosslinked nanocomposites with
different amounts of bacterial cellulose produced from cellulose
synthesis: a PVA, b PVA/BC 0.6, c PVA/BC 6, d PVA/BC 14
and e bacterial cellulose
Cellulose
123
Netravali 2012). This behavior is closely tied to the
manufacturing method of the nanocomposite, wherein
microorganism bioengineering allows developing
materials in which there are intimate contacts between
the reinforcing phase and matrix with a maximum
percolation.
Scanning electron microscopy images of the frac-
tured surfaces of crosslinked PVA reference films
(prepared from 3 wt% aqueous solution), PVA 14 (see
also Table 1), the corresponding BC-reinforced nano-
composite (PVA/BC 14) and films of BC are shown in
Fig. 5. Figure 5a shows a uniform fracture of PVA 14
film with flow lines in the matrix indicating that the
rupture was caused by a crack propagation on the
smooth brittle surface. The respective nanocomposite
fracture shows no evidence of agglomerates indicating
that the BC ribbons were homogeneously distributed
throughout the matrix (Fig. 5b). Moreover, good
compatibility between the reinforcing PVA and the
BC matrix (good fiber-matrix bonding) is suggested by
the absence of pull-out BC ribbons. Likewise, the
typical delamination behavior of the BC films
observed in Fig. 5c is absent in the composite
material, suggesting that there is improved contact
and adhesion between the layers comprising the BC
network and not only between the reinforcing and matrix
components. The enhancement over nanocomposite
delamination promotes better stress transfer within the
material (Quero et al. 2010, 2011). All in all, the
results indicate strong interactions between the two
components, which has a direct influence on the final
mechanical characteristics of the composite after
crosslinking.
In previous work, it was verified that crosslinking
reactions of BC-glyoxal-BC occur even at low con-
centrations of glyoxal in the culture medium (docu-
ment in the patent process). Furthermore, the
insolubility of PVA films after crosslinking with
glyoxal confirms effective PVA-glyoxal-PVA reac-
tions. Therefore, it is also likely that PVA-glyoxal-BC
crosslinking takes place in the nanocomposite.
Thermograms of PVA films and their nanocom-
posites with BC are shown in Fig. 6. TGA profiles of
PVA exhibit two degradation peaks, while the nano-
composites display only a single peak. This suggests
that a high chemical compatibility and entanglement
exist between PVA and BC in the nanocomposite. The
first weight loss at 40–200 �C is due to water
evaporation absorbed during the preconditioning at
75 % RH. The PVA matrices have a higher percentage
of water loss compared with their respective nano-
composites; therefore, they have a greater tendency to
absorb water, and this trend is decreased with increasing
BC content. More important to this discussion is the fact
Fig. 4 Stress-strain
behavior of tensile test result
on crosslinked
nanocomposites and their
matrices
Cellulose
123
that compared to the PVA reference film, the thermal
stability of the nanocomposite is higher and increases
with the cellulose content. The maximum temperature
of thermal degradation for PVA/BC 6 and PVA/BC
14 nanocomposites increases from around 5–10 and
7–16 �C, respectively. This is explained by the effect of
BC in the polymer matrix, the good dispersion of BC, and
strong chemical and mechanical interactions with PVA.
Fig. 5 SEM images of the tensile fracture surface of crosslinked materials: a PVA 14, b PVA/BC 14 nanocomposite and c bacterial
cellulose. The SEM images are shown with two magnifications, as indicated
Fig. 6 Thermal
degradation profiles of
crosslinked materials. The
black lines correspond to the
nanocomposite and the gray
lines to their matrices
Cellulose
123
The thermal behavior of the samples was also
studied by DSC (Table 2). The glass transition tem-
peratures of PVA/BC 6 and 14 composites are shifted
to higher values compared to the respective matrices.
This effect suggests that good miscibility exists
between the PVA matrix and the reinforcing BC,
and the presence of the BC network restricts the
movement of the polymer chains. Moreover, the
melting point (Tm) and degree of crystallinity (Xc) of
the composites are slightly increased when compared
to the corresponding matrices (Table 2). These results
are in agreement with the FTIR data: the cellulose
surface acts as a nucleating agent for PVA crystalli-
zation, and the molecular mobility of PVA chains
decreases in the interfacial zone. Similar results were
reported for PCL and PVA with cellulose nanocrystals
(Roohani et al. 2008; Zoppe et al. 2009).
Dielectric spectroscopy was performed in order to
investigate further the effect of temperature on the
molecular interaction of PVA and BC. PVA/BC 14
samples as well as the respective matrix were
analyzed, and the results are shown in Fig. 7 for the
evolution of dielectric modulus M with temperature at
1 kHz. Due to the fact that some dielectric relaxations
may be obscured by the conductivity contribution, it is
more convenient to use the electric modulus formal-
ism, which shifts the loss peaks to the region of
frequencies where most of the measuring equipment
operates and at the same time diminishes the values of
the abscissa because of the definition of the electric
modulus (M* = 1/e*, where e* is complex permittiv-
ity) (Kortaberria et al. 2011). Two peaks are observed
for PVA/BC 14 and PVA 14: the main relaxation a
associated with the glass transition and b relaxation
assigned to local motions around main-chain bonds
and relaxation in crystalline domains (De la Rosa et al.
2001). The shifting of a relaxation to higher temper-
atures when BC is added to the composite indicates an
increase in the glass transition temperature of the
matrix. In fact, the Tg increases by 20 �C with 14 wt%
BC reinforcement. This was also observed in the DSC
experiments discussed previously (Tg increase 8 �C
with 14 wt% BC reinforcement), which highlights the
PVA-BC interactions and related reduction of polymer
chain mobility. The temperature at which b relaxation
appears does not seem to be markedly affected by
the reinforcement. This indicates that local motions
are not affected by the presence of reinforcement
Table 2 Thermal transitions and crystallinity (*) of matrix
and in situ nanocomposite reinforced with different amounts of
bacterial cellulose
Sample Tg
(�C)
Tm
(�C)
DHm
(Jg-1)
Xc
(%)
Xp
(%)
PVA 6 63.78 212.01 40.27 0.25 0.25
PVA/BC 6 87.05 215.76 43.78 0.27 0.29
PVA 14 68.33 209.47 39.62 0.25 0.25
PVA/BC
14
76.30 210.81 36.70 0.23 0.26
(*) Xc = DHm/DHm� and Xp = Xc/w; where
DHm� = 161.6 Jg-1 is the heat of fusion for 100 %
crystalline PVA (Roohani et al. 2008), and w is the weight
fraction of polymeric matrix material in the compositeFig. 7 Dielectric spectroscopy results of crosslinked materials.
The black lines correspond to nanocomposite PVA/BC 14 and
the gray lines to its matrix PVA 14
Fig. 8 Bulk water uptake capacity of matrices and nanocom-
posites after 48 h of immersion
Cellulose
123
presenting the same mobility. The reinforcement
affects only the main chain motions, reducing their
mobility.
Water absorption measurements were carried out in
order to study the swelling capacity of the composites,
and the results at equilibrium are shown in Fig. 8.
Compared to PVA reference samples, it is observed
that the water absorption (measured as the amount of
water retained by the material) of the nancomposites
decreases by 24, 26 and 36 wt% in the case of PVA/
BC 0.6, PVA/BC 6 and PVA/BC 14, respectively. This
is explained by the decrease in the free volume and
chain flexibility upon addition of BC, which restricts
swelling of the matrix. PVA and BC interactions
through hydrogen-bonding and crosslinking are
expected to decrease the availability of hydroxyl
groups to bind with water molecules, as suggested
elsewhere (Peresin et al. 2010).
In order to investigate the effect of absorbed water
on the mechanical behavior of the nanocomposites,
composite films were exposed to humidity cycles
between 0 and 90 % RH in a DMA unit (see Fig. 9).
Changes observed in storage modulus were accompa-
nied by shifts in material strain as the relative humidity
of the surrounding environment changed. As expected,
the modulus decreases with increasing humidity
(Fig. 9). However, compared with the BC-free films,
the changes were smaller for the nanocomposites.
Thus, as the amount of reinforcing BC increases,
better mechanical stability is achieved. Likewise,
changes in the dimensions of the materials were
observed with the gain of moisture: Fig. 9b indicates
an increase in the strain of the system as the % RH
increases due to the plasticizing effect of water.
Moreover, a higher dimensional stability was
observed for the composites with increased content
of reinforcing BC. This is in agreement with the
strength and swelling measurements of composites
(Figs. 4, 8). The highest increase in dimensional
stability was observed with PVA/BC 6 and PVA/BC
14. These findings suggest that cellulose restrains the
flow of PVA polymer chains. However, the high
mobility remains with PVA/BC 0.6, which can also be
observed in tensile tests (Fig. 4), where the mobility of
PVA/BC 0.6 resists a breakage resulting in high strain.
The storage modulus and strain of the PVA
matrices and the respective nanocomposites fully
and reversibly recovered after humidity cycles. This
elastic behavior is not affected by dehydration and
hydration of the specimens. Consequently, the net
effect of cellulose network coupled with strong
reinforcing-matrix interactions reduces moisture
uptake and restricts the movement of PVA chains,
providing mechanical and dimensional stability to the
composite.
Conclusions
Poly(vinyl alcohol) nanocomposites were produced
in situ during cellulose synthesis by bacteria of the
Gluconacetobacter genus. The loss of the hydrosoluble
Fig. 9 Variation of the a storage modulus and b strain of matrices and nanocomposites after two humidity cycles from 0 to 90 % RH at
room temperature
Cellulose
123
PVA polymer matrix during post-processing was
prevented by chemical crosslinking and the benefits
in mechanical and thermal properties assessed. Com-
posites with different BC reinforcing levels were
produced. Both Young’s modulus and tensile strength
markedly increased with the BC loading: improve-
ments of 680 and 40 % were obtained when compared
with the respective BC-free systems. SEM of fractured
films, DSC, TGA and dielectric spectroscopy results
suggest that effective dispersion occurred between the
PVA matrix and reinforcement BC phase. The
molecular mobility of PVA chains decreased in the
interfacial zone because of the presence of the network
of cellulose fibrils. This effect led to an increase in the
crystallinity of the matrix in the nanocomposite. The
TGA analysis also indicated that the nanocomposites
had a higher thermal stability compared to the
respective matrix. This further supports the evidence
of a high entanglement between the two components
produced as a three-dimensional network of nanoscale
BC after biosynthesis by the microorganisms. Overall,
the BC network is postulated to afford an improved
percolation within the matrix accompanied by an
improved bonding between the reinforcing BC and
PVA matrix (hydrogen bonds and crosslinking). This
can explain the exceptional mechanical performance
of the nanocomposites. Finally, the BC reinforcement
was shown to reduce water uptake and to improve
moisture stability of the composites under cyclic
humidity conditions.
Acknowledgments The authors would like to acknowledge
Colombia’s COLCIENCIAS and SENA for financial support as
well as Prof. Janne Laine of the Department of Forest Products
Technology of Aalto University (Finland).
References
Barud HS, Ribeiro CA, Crespi MS, Martines MAU, Dexpert-
Ghys J, Marques RFC, Messaddeq Y, Ribeiro SJL
(2007) Thermal characterization of bacterial cellulose-
phosphate composite membranes. J Therm Anal Calorim
87:815–818
Berglund L (2005) Cellulose based nanocomposites. In: Moh-
anty A, Misra M, Drzal L (eds) Natural fibers, biopolymers,
and biocomposites. CRC Press, Boca Raton, pp 807–832
Bielecki S, Krystynowicz A, Turkiewicz M, Kalinowska H
(2005) Bacterial cellulose. In: Steinbuchel A, Doi Y (eds)
Biotechnology of polymer: from synthesis to patents.
Wiley, Weinheim, pp 381–434
Bondeson D, Mathew A, Oksman K (2006) Optimization of the
isolation of nanocrystals from microcrystalline cellulose
by acid hydrolysis. Cellulose 13:171–180
Brown E, Laborie M-P (2007) Bioengineering bacterial cellu-
lose/poly(ethylene oxide) nano composites. Biomacro-
molecules 8:3074–3081
Brown JR, Willison J, Richardson C (1976) Cellulose biosyn-
thesis in Acetobacter xylinum: visualization of the site of
synthesis and direct measurement of the in vivo process.
Proc Natl Acad Sci USA 73:4565–4569
Brown EE, Zhang J, Laborie M-P (2011) Never-dried bacterial
cellulose/fibrin composites: preparation, morphology and
mechanical properties. Cellulose 18:631–641
Castro C, Zuluaga R, Alvarez AL, Putaux J-L, Caro G, Rojas O,
Mondragon I, Ganan P (2012) Bacterial cellulose produced
by a new acid-resistant strain of Gluconacetobacter genus.
Carbohydr Polym 89:1033–1037
Castro C, Cleenwerck I, Trcek J, Zuluaga R, De Vos P, Caro G,
Aguirre R, Putaux J-L, Ganan P (2013) Gluconacetobacter
medellinensis sp. nov., cellulose- and non-cellulose-pro-
ducing acetic acid bacteria isolated from vinegar. Int J Syst
Evol Microbiol 63:1119–1125
Cheng Q, Wang S, Rials T (2009) Poly(vinyl alcohol) nano-
composites reinforced with cellulose fibrils isolated by
high intensity ultrasonication. Compos Part A Appl S
40:218–224
Choi H-M, Kim JH, Shin S (1999) Characterization of cotton
fabrics treated with glyoxal and glutaraldehyde. J Appl
Polym Sci 73:2691–2699
Choi Y, Ahn Y, Kang M, Jun H, Kim IS, Moon S (2004)
Preparation and characterization of acrylic acid-treated
bacterial cellulose cation-exchange membrane. J Chem
Technol Biotechnol 79:79–84
De la Rosa A, Heux L, Cavaille JY (2001) Secondary relax-
ations in poly(allyl alcohol), PAA, and poly(vinyl alcohol),
PVA. II. Dielectric relaxations compared with dielectric
behaviour of amorphous dried and hydrated cellulose and
dextran. Polymer 42:5371–5379
Gea S, Bilotti E, Reynolds C, Soykeabkeaw N, Peijs T (2010)
Bacterial cellulose-poly(vinyl alcohol) nanocomposites
prepared by an in situ process. Mater Lett 64:901–904
Grande C, Torres F, Gomez C, Troncoso O, Canet-Ferrer J,
Martinez-Pastor J (2008) Morphological characterisation
of bacterial cellulose-starch nano composites. Polym
Polym Compos 16:181–185
Habibi Y, Lucia L, Rojas O (2010) Cellulose nanocrystals:
chemistry, self-assembly, and applications. Chem Rev
110:3479–3500
Iguchi M, Yamanaka S, Budhiono A (2000) Bacterial cellulose-
a masterpiece of nature’s arts. J Mater Sci 35:261–270
Iwamoto S, Nakagaito A, Yano H, Nogi M (2005) Optically
transparent composites reinforced with plant fiber-based
nanofibers. Appl Phys A Mater Sci Process 81:1109–1112
Iwamoto S, Nakagaito A, Yano H (2007) Nano-fibrillation of
pulp fibers for the processing of transparent nano com-
posites. Appl Phys A Mater Sci Process 89:461–466
Janardhnan S, Sain M (2006) Isolation of cellulose microfibrils:
an enzymatic approach. BioResources 1:176–188
Kenney JF, Willcockson GW (1966) Structure-property rela-
tionships of poly(vinyl alcohol). III. Relationships between
Cellulose
123
stereo-regularity, crystallinity, and water resistance in
poly(vinyl alcohol). J Polym Sci Pol Chem 4:679–698
Klemm D, Schumann D, Udhardt U, Marsch S (2001) Bacterial
synthesized cellulose—artificial blood vessels for micro-
surgery. Prog Polym Sci 26:1561–1603
Kortaberria G, Arruti P, Mondragon I, Vescovo L, Sangermano
M (2011) Dynamics of in situ synthetized silver–epoxy
nanocomposites as studied by dielectric relaxation spec-
troscopy. J Appl Polym Sci 120:2361–2367
Laborie M-P (2009) Bacterial cellulose and its polymeric
nanocomposites. In: Lucia L, Rojas OR (eds) The nano-
science and technology of renewable biomaterials. Wiley,
Chichester, pp 231–271
Lee S-Y, Mohan DM, Kang I-A, Doh G-H, Lee S, Han SO
(2009) Nanocellulose reinforced PVA composite films:
effects of acid treatment and filler loading. Fiber Polym
10:77–82
Lin SB, Hsu CP, Chen LC, Chen HH (2009) Adding enzymat-
ically modified gelatin to enhance the rehydration abilities
and mechanical properties of bacterial cellulose. Food
Hydrocoll 23:2195–2203
Lu J, Wang T, Drzal LT (2008) Preparation and properties of
microfibrillated cellulose polyvinyl alcohol composite
materials. Compos Part A Appl 39:738–746
Nakagaito A, Iwamoto S, Yano H (2005) Bacterial cellulose: the
ultimate nano-scalar cellulose morphology for the pro-
duction of high-strength composites. Appl Phys A Mater
Sci Process 80:93–97
Ngui MO, Mallapragada SK (1998) Understanding isothermal
semicrystalline polymer drying: mathematical models and
experimental characterization. J Polym Sci Part B Polym
Phys 36:2771–2780
Peppas NA, Hansen PJ (1982) Crystallization kinetics of
poly(vinyl alcohol). J Appl Polym Sci 27:4787–4797
Peresin MS, Habibi Y, Vesterinen AH, Rojas OJ, Pawlak JJ,
Seppala JV (2010) Effect of moisture on electrospun
nanofiber composites of poly(vinyl alcohol) and cellulose
nanocrystals. Biomacromolecules 11:2471–2477
Phisalaphong M, Jatupaiboon N (2008) Biosynthesis and char-
acterization of bacteria cellulose–chitosan film. Carbohydr
Polym 74:482–488
Qiu K, Netravali A (2012) Bacterial cellulose-based membrane-
like biodegradable composites using cross-linked and
noncross-linked polyvinyl alcohol. J Mater Sci 47:6066–
6075
Quero F, Nogi M, Lee K-Y, Yano H, Abdulsalami K, Holmes
SM, Sakakini BH, Eichhorn SJ (2010) Optimization of the
mechanical performance of bacterial cellulose/poly(l-lac-
tic) acid composites. ACS Appl Mater Inter 2(1):321–330
Quero F, Nogi M, Lee K-Y, Vanden Poel G, Bismarck A,
Mantalaris A, Yano H, Eichhorn SJ (2011) Cross-linked
bacterial cellulose networks using glyoxalization. ACS
Appl Mater Interfaces 3:490–499
Roohani M, Habibi Y, Belgasem NM, Ebrahim G, Karimi AN,
Dufresne A (2008) Cellulose whiskers reinforced polyvi-
nyl alcohol copolymers nano composites. Eur Polym J
44:2489–2498
Saibuatong O, Phisalaphong M (2010) Novo Aloe vera-bacterial
cellulose composite film from biosynthesis. Carbohydr
Polym 79:455–460
Samir S, Alloin F, Paillet M, Dufresne A (2004) Tangling effect
in fibrillated cellulose reinforced nano composites. Mac-
romolecules 37:4313–4316
Samir MASA, Alloin F, Dufresne A (2005) Review of recent
research into cellulosic whiskers, their properties and their
application in nanocomposite field. Biomacromolecules
6:612–626
Tokoh C, Takabe K, Sugiyama J, Fujita M (2002) Cellulose
synthesized by Acetobacter xylinum in the presence of
plant cell wall polysaccharides. Cellulose 9:65–74
Uhlin IK, Atalla RH, Thompson NS (1995) Influence of hemi-
celluloses on the aggregation patterns of bacterial cellu-
lose. Cellulose 2:129–144
Wan YZ, Hong L, Jia SR, Huang Y, Zhu Y, Wang YL, Jiang HJ
(2006) Synthesis and characterization of hydroxyapatite-
bacterial cellulose nanocomposites. Compos Sci Technol
66:1825–1832
Yano H, Sugiyama J, Nakagaito AN, Nogi M, Matsuura T,
Hikita M, Handa K (2005) Optically transparent compos-
ites reinforced with networks of bacterial nanofibers. Adv
Mater 17:153–155
Yasuda K, Gong JP, Katsuyama Y, Nakayama A, Takanabe Y,
Kondo E, Ueno M, Osada Y (2005) Biomechanical prop-
erties of high-toughness double network hydrogels. Bio-
materials 26:4468–4475
Zhang W, Yang X, Li C, Liang M, Lu C, Deng Y (2011)
Mechanochemical activation of cellulose and its thermo-
plastic polyvinyl alcohol ecocomposites with enhanced
physicochemical properties. Carbohydr Polym 83:257–263
Zimmermann T, Pohler E, Geiger T (2004) Cellulose fibrils for
polymer reinforcement. Adv Eng Mater 6:754–761
Zoppe JO, Peresin MS, Habibi Y, Venditti RA, Rojas OJ (2009)
Reinforcing poly(e-caprolactone) nanofibers with cellulose
nanocrystals. ACS Appl. Mater. Inter. 1:1996–2004
Zuluaga R, Putaux J-L, Cruz J, Velez J, Mondragon I, Ganan P
(2009) Cellulose microfibrils from banana rachis: effect of
alkaline treatments on structural and morphological fea-
tures. Carbohydr Polym 76:51–59
Cellulose
123