Journal of Biomaterials and Nanobiotechnology, 2011, 2, 55-64
doi:10.4236/jbnb.2011.21008 Published Online January 2011 (http://www.SciRP.org/journal/jbnb)
55
Fabrication of Curcumin Encapsulated
Chitosan-PVA Silver Nanocomposite Films for
Improved Antimicrobial Activity
Kanikireddy Vimala1, Yallapu Murali Mohan1,2, Kokkarachedu Varaprasad1, Nagireddy Narayana
Redd1, Sakey Ravindra1, Neppalli Sudhakar Naidu3, Konduru Mohana Raju1*
1Synthetic Polymer Laboratory, Department of Polymer Science and Technology, Sri Krishnadevaraya University, Anantapur, India;
2Cancer Biology Research Center, Sanford Research/USD, Sioux Falls, USA; 3Polymer Thin Film Laboratory, Department of
Chemical Engineering, Yonsei University, Seoul, South Korea.
Received August 30th, 2010; revised September 22th, 2010; accepted September 28th, 2010.
ABSTRACT
The present study explores the in situ fabrication of chitosan-poly(vinyl alcohol)-silver nanocomposite films in view of
their increasing applications as antimicrobial packaging, wound dressing and antibacterial materials. The reduction of
silver ions into silver nanoparticles (AgNPs ) is achieved in acidic solution of chitosan (C) and poly (vinyl alcohol)
(PVA) using their functional groups (-OH, -COOH, -NH2 groups). The presence of silver nanoparticles in the chito-
san-PVA film is confirmed by UV-Vis spectroscopy, Fourier Transform Infrared (FTIR) spectroscopy and X-ray Dif-
fraction (XRD) analysis. The Scanning Electron Microscopic (SEM) images illustrate the presence of embedded silver
nanoparticles throughou t the films. In addition, the fo rmed silver nanoparticles have an averag e particle size of ~ 16.5
nm as observed by Transmission Electron Microscopy (TEM). The anti-microbial and anti-fungal activity of the chito-
san-PVA silver nanoparticle films have demonstrated significant effects against Escherichia coli (E. coli), Pseudomo-
nas, Staphylococcus, Micrococcus, Candida albicans, and Pseudomonas aeruginosa (P. aeruginosa). To improve fur-
ther their therapeutic efficacy as anti-microbial agents, curcumin encapsulated chitosan-PVA silver nanocomposite
films are developed which showed enormous growth inhibition of E. coli compared to curcumin and chitosan-PVA sil-
ver nanoparticles film alone. Therefore, the present study clearly provides novel antimicrobial films which are poten-
tially useful in preventing/treating infections.
Keywords: Silver Nanoparticles, Chitosan, Poly(Vinyl Alcohol), Curcumin, Wound Dressing, Hydrogel
1. Introduction
Human beings are often infected by micro-organisms
such as bacteria, yeast, mold, virus, etc [1]. Silver and
silver ion based materials are widely known for their
bactericidal and fungicidal activity. Their antimicrobial
effect is due to blockage of respiratory enzyme pathways,
alteration of microbial DNA and the cell wall [2].
Therefore, silver and silver ion containing materials are
used as prostheses, catheters, vascular grafts and as
wound dressings [3,4]. Recent studies have shown that
silver in the form of nanoparticles is very effective as
antimicrobial agent when compared to bulk silver or sil-
ver ions [5,6]. The therapeutic efficacy of silver in
nanoparticles form is several folds greater than conven-
tional silver compounds. The nanoparticles exert their
antimicrobial property by interacting with the sulphur
containing proteins present in bacterial cell membrane as
well as with phosphorous containing DNA [6]. In addi-
tion, silver nanoparticles based antimicrobials have many
advantages due to their thermal stability, health and en-
vironmental safety [7]. As a result, the usage of silver
based commercial products including topical ointments,
bandages, augmentation devices, tissue scaffolds, antim-
icrobial filters and gels have increased for improving
public health care [8,9]. Some commercially available
silver-containing purification systems such as Aqu a pure,
kinetico and QSI-Nano have shown to remove 99.99%
pathogens [10-13].
Many methods have been reported in developing silver
nano-products including chemical reduction method. The
chemical reduction method, using chemicals such as hy-

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 56
drazine hydrate, dimethyl formamide, ethylene glycol,
etc., however causes toxicity or biological hazards. The
current trend in nanotechnology research is to use bio-
compatible or biological friendly polymers which can
provide reductio n as well as stabilization functions in the
preparation of silver nanoparticles [14]. The combination
of silver nanoparticles with water soluble biopolymers
will produce new antimicrobials. Based on this, various
natural polymers such as gum acacia, starch, gelatine,
sodium alginate, carboxy methyl cellulose etc., have
been employed to prepare biocompatible polymeric sil-
ver nanocomposites [ 15-17]. Chitosan, a natural polymer
composed of poly(β-(1-4)-2-amino-2-deoxy-D-glucose),
is one of the structural polysaccharide which is abun-
dantly available in nature after cellulose. Chitosan in-
teracts very easily with bacterium and binds to DNA,
glycosaminoglycans and most of the proteins thereby
enhancing the antimicrobial effect of silver nanoparticles
[18]. Polyvinyl alcohol (PVA), a water soluble synthetic
polymer, having less toxicity, possess excellent wound
dressing bio reactor properties and known to be used as a
stabilizer in nanoparticles synthesis [19,20].
Films and coatings based on biopolymers function as
barriers against moisture, oxygen, aroma flavor as well
as oil and are the materials for future applications [21]. In
addition, they have an interesting application as supports
for antimicrobial, nutritional and antioxidant substances.
Rira Jung et al. [11] prepared hydrated cellulose silver
nanoparticle membranes and investigated their antim-
icrobial activity against S. aureus and E. coli. Durango et
al. [22] developed edible antimicrobial films using yarn
starch and chitosan and shown to exhibit effectively the
growth of S. enteritid is. Tripathi et al. [23] pr epared ch i-
tosan-PVA blend films for food packaging applications
and they have also shown to possess antimicrobial activ-
ity against food pathogenic bacteria.
In view of this, the present investigation involves the
development of a novel, eco-friendly antimicrobial film
containing chitosan, PVA and silver nanaoparticles by in
situ fabrication method, as shown in Figure 1. The de-
veloped chitosan-PVA silver nanoparticles film was
characterized with the help of UV-Vis, FTIR spectro-
photometric, Thermogravimetric Analyzer (TGA), Scan-
ning Electron Microscopy (SEM) and Transmission
Electron Microscopy (TEM) analysis. It’s antimicrobial
activity against gram-positive (Staphylococcus and Mi-
crococcus), gram-negative bacteria (E. coli and Pseudo-
monas), and fungi (Candida alb icans and P. aeruginosa)
has been tested. Curcumin (CUR), a hydrophobic poly-
phenolic compound derived from the rhizome of the herb
curcuma longa, possesses a wide range of biological ac-
tivities including wound healing, anti-bacterial, an-
ti-oxidant, anti-inflammatory and anti-cancer properties.
Therefore this compound is incorporated into chito-
san-PVA nanoparticles film to improve significantly the
therapeutic antib acterial efficacy of the film.
2. Experimental
2.1. Materials
Chitosan (C) (high M.W, > 75% deacetylated) is pur-
chased from Sigma Chemical Company (St. Louis, USA).
Acetic acid (glacial, 99-100%), poly(vinyl alcohol) (PVA)
(MW 125,000 and degree of acetylation, 19.5-22.7%),
silver nitrate (AgNO3) and glutaraldehyde (GA) are pur-
chased from Merck (Mumbai, India). Mineral salt broth
and nutrient agar are obtained from Himedia Chemicals
(Mumbai, India). The Department of Botany (Sri Krish-
nadevaraya University, Anantapur, India) has provided
tandard cultures of the organisms. Curcumin (95% (w/w)
curcuminoids by Spectrophotometry) is a gift sample
from Natural Remedies Private Limited (Bangalore, In-
dia). All the chemicals and reagents are used without
further purification. Double-distilled water is used fo r the
preparation of all solutions throughou t the study.
2.2. Preparation of Chitosan-PVA Blend Films
(CPB)
50 ml of chitosan solution (1% wt./wt. in acetic acid) and
50 ml of PVA solution (1% wt./wt. in water) (1:1) are
mixed in 250 ml beaker and stirred for 1 h at 60oC to
obtain homogeneous solution. To this solution 1 ml of
2% glutaraldehyde solution in water (a cross-linking
agent) is added under stirring at room temperature (25oC).
The solution is transferred immediately into a Teflon
sheet covered glass plate (Dimensions: 100 mm length x
100 mm width x 3 mm height) (Sabean traders, Chennai,
India) and dried at 80oC in an electric oven for 2 h (Ba-
heti Enterprises, Hyderabad, India). The formed
cross-linked chitosan-PVA blend (CPB) films are
washed with double distilled water to neutralization and
dried at room temperature. This film is termed as 1:1
CPB film. Similarly, 50 ml of 2% wt./wt. chitosan solu-
tion + 50 ml of 1% wt./wt. PVA solution (2:1) and 50 ml
of 3% wt./wt. chitosan solution + 50 ml of 1% wt./wt.
PVA solution (3:1) are used to get different CPB films.
2.3. Preparation of Chitosan-PVA Silver
Nanoparticles Films (CPSNP)
AgNO3 (100 mg) is added separately into three beakers
containing 50 ml of different percentages (1%, 2% and
3%) of chitosan solutions at room temperature. The cor-
responding solutions are kept in sunlight for 1 h. The
yellow coloured solution started turning to red, then
brown and brownish indicating the formation of silver
nanoparticles. To this AgNP so lution s, 50 ml o f 1 % PVA

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 57
solution is added and stirrer for 1 h. For all these solu-
tions, 1 ml of 2% glutaraldehyde (cross-linker) is added
under stirring at room temperature. The solutions are
then poured into Teflon covered glass plates and dried as
explained earlier. These films are termed as chito-
san-PVA silver nanoparticles (CPSNPs) films.
2.4. Swelling Studies
The completely dried, pre- weighed CPB and CPSNP
films are equilibrated in 250 ml of phosphate buffer (pH
7.4) at 25oC. The water up take of the films is measured
for every 30 min using analytical balance. The swelling
ratio (Q) of the films is calculated using the following
equation: Q=Ws/Wd, where Ws is the weight of the
swollen film at different time intervals and Wd is the
weight of dry film.
2.5. Characterization
The UV-vis spectroscopic studies are carried out using
Shimadzu 1600 UV-vis sp ectrometer (Kyoto, Jap an) 250
to 600 nm. For this study, silver nanoparticles were ex-
tracted from silver nanocomposite film (10 mg/ml) over
a period of a week, centrifuged at 1000 rpm for 30 min,
and the supernatant was used to measure the absorption
spectra. The distilled water was used as a blank solution .
The UV spectra of the silver nanoparticles solution
showed a characteristic peak between 413-421 nm re-
lated to surface Plasmon resonance absorption peak cor-
responding to silver nanoparticles. The FTIR spectra of
CPB and CPSNP films are recorded on a MB3000 FT-IR
Analyzer (ABB Analytical, Quebec, Canada).To record
the FTIR spectra of films, the samples were completely
dried in an oven(Baheti Enterprises, Hyderabad, India) at
400C for 6h. These samples were read between 600 and
4000 cm-1 using KBr disk method. X-ray Diffraction
(XRD) patterns were carried out for dried and finely
grounded nanocomposite film samples on a Rigaku
D/Max-2550Pc (Tokyo. Japan) using Cu and Ka radia-
tion generated at 40 kV and 50 mA. Thermal studies of
the films were carried out using SDT Q 600 TGA in-
strument (T.A. Instruments-water LLC, Newcastle, DE
19720, USA) at a heating rate of 10oC/min under con-
stant nitrogen flow (100 ml/min). Morphology of the
films are observed by SEM through a JEOL JSM 840A
(Tokyo, Japan).To image the film samples (surface or
cross-sections) were coated with a thin layer of palla-
dium gold alloy after mounting on a double sided carbon
tape. TEM images are recorded using a Technai F 12
transmission electron microscope (Philips Electron Op-
tics, Holland) operating at an acceleration voltage of 15
kV. The samples are prepared for TEM measurements by
extracting the silvernanoparticles from the film while
they are in swollen stage using a soft ball and allowed to
soak for 1 day to come out the silver nanoparticle from
the film network into aqueous phase. From this solution
10-20 µl of the aqueous solution was dropped on a cop-
per grid, removed the excess solution using filter paper
and dried at room temperature. The copper grid was in-
serted into Technai F12 transmission electron micro-
scope.
2.6. Mechanical Properties
Mechanical properties of CPB and CPSNP films are
measured using a INSTRON 3369 Universal Testing
Machine (UTM) (Buckinghamshire, England) running at
a crosshead speed of 5 mm/min. The sample films are cut
into 1cm x10cm size and the gauge length is about 5 cm.
The tensile parameters, maximum stress, young’s
modulus and % elongation at break are measured using
10 kg load cell.
2.7. Curcumin Loading and Encapsulation
Efficiency
Curcumin (CUR) is loaded into CPB or CPSNP films by
swelling method. For loading curcumin, the films (50 mg)
are allowed to swell in 20 ml of CUR solution (5 mg of
CUR in 20 ml, acetone (8 ml) - distilled water (12 ml))
for 24 hrs at 25oC. The loading efficiency of curcumin in
the films is determined spectrophotometrically [24]. The
drug-loaded films are placed in 50 ml of buffer solution
and stirred vigorously for 96 hrs to extract the drug from
the films. The solution is filtered and assayed by UV
spectrophotometer at fixed λ max value of 491.2 nm. The
results of % drug loading and encapsulation efficiency
are calculated using the following equations.
% Drug loading = (Weight of drug in film / Weight of
film) x 100
% Encapsulation efficiency = (% actual loading / %
theoretical loading) x 100
2.8. Release of Curcumin
In order to study the release of curcumin from loaded
films, known weights were placed in a measured volume
(50ml) of 7.4 pH phosphate buffer at room temperatures
and the released amount of curcumin was determined at
different time intervals by recording the absorbance of
release medium by UV-Vis spectrophotometer. The re-
corded absorbance was then related to the amount of re-
leased curcumin using a calibration plot. The absorption
of the solutions of curcumin was measured at λmax 492.2
nm.
2.9. Antimicrobial and Anti-fungal Activity
The antimicrobial and antifungal activity of the devel-
oped CPB and CPSNP films are tested by disc diffusion,
spread plate and viable cell count method against E. coli,

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 58
Pseudomonas, Staphylococcus, Micrococcus, Candida
albicans, an d P. aeru gi n os a, as model bacteria and fungi.
For disc diffusion method, the films are cut into a disc
shape with 5 mm diameter, sterilized by autoclaving for
30 min at 120oC, and placed on different cultured agar
plates. The plates are incubated for 2 days at 37oC in an
incubation chamber maintaining with 5% CO2 flow and
the inhibition zone is then measured. Similarly, for
spread plate method curcumin loaded CPB film and
CPSNP films are placed in Petri plates. Then agar is
poured onto the whole film and allowed to settle on the
top of the film. The inhibition action of films is then
measured. Viable count method measures the bacterial
growth. For this study, 108 colony forming units (CFU)
of E. coli are grown in 10 ml nutrient broth supplement
with film discs (20 mm dia). The bacterial viability is
checked using their O.D values by UV-Vis spectrometer
at 600 nm.
3. Results and Discussion
In the past few decades, a lot of work has been done and
published on the development of synthetic and natural
polymer AgNPs membranes/films as well as their action
against various microorganisms [24,25]. However, these
conventional films have exhibited limited applicability
because of their low rate of fluid absorption and poor
mechanical properties [26]. The films/membranes having
AgNPs that are to be used for wound dressing and water
purification purpo se shou ld po sses good water absor p tion
abilities and mechanical properties. Therefore, due to
greater importance of chitosan and PVA in biomedical
applications, chitosan-PVA silver nanoparticle films are
developed via green process. Chitosan and PVA are well
known polymers with excellent absorption capacities for
a number of metal ions due to the presence of amino
(-NH2) and hydroxyl (-OH) groups in their structures.
The formation of the CPSNP is carried out by two step
process as described in the experimental procedure (Fig-
ure 1).
In the first step, silver nitrate is added into the chitosan
solution and the solution is irrad iated by sunlight thereby
reducing the silver ions into silver nanoparticles. In the
second step, chito san stabilized AgNPs solution is mixed
with PVA solution and then glutaraldehyde cross-linker
is added under stirring. The solution is allowed to form
chitosan-PVA silver nanoparticles (CPSNPs) films. The
AgNPs in these films are uniformly dispersed and en-
trapped throughout the networks via amine (-NH2) and
hydroxyl (-OH) functional groups [27,28]. Similarly
curcumin loaded CPB and CPSNP films are developed
by putting these films in curcumin solution. This ap-
proach is exp ected to improve the flu id absorp tion ability,
mechanical strength as well as antimicrobial activities.
Figure 2 represents the change in physical appearance of
the films during the process of formation of silver nano-
particles and after loading the curcumin within the net-
work. The light yellow colour film turned into dark
brown colour d ue to the prese nce of Ag na no part i cl e.
3.1. Swelling Capacity
The swelling capacity of an antibacterial film/gel and the
nanocomposite plays an important role in the antibacte-
rial activity, wound healing capacity, and for biomedical
application due to their high water/solvent holding ca-
pacity. They can further absorb a slight to moderate
amount of the wound exudates by swelling which helps
in fast healing of the wound. Figure 3 shows the swell-
ing capacity of CPB and CPSNP films with time. The
CPB films showed higher swelling capacity than CPSNP
films. For example, the swelling capacity of 1:1 CPB is
13.92 g/g whereas the corresponding is CPSNP film is
9.8 g/g only. This lowering in the swelling capacity is
attributed due to bind ing of AgNPs with electrons of ‘O’
and ‘N’ atoms of hydroxyl and amine groups present in
chitosan/PVA chains. This produces additional cros-
slink’s within the chain networks [29]. The higher cros-
slink’s within the films restrict the penetration of water
for swelling [30,31].
Figure 1. Schematic diagram of formation of chitosan-PVA silver nanoparticle films and curcumin loaded chitosan
-PVA silver nanoparticle films.

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 59
Figure 2. Representative (a) chitosan-PVA blend (CPB), (b)
chitosan-PVA silver nanoparticle (CPSNP), (c) curcumin
loaded chitosan-PVA blend, and (d) curcumin loaded chi-
tosan-PVA silver nanoparticle films. These films were pre
pared using 1:1 ratio of chitosan and PVA. Similar photo
graphs were obtained for 2:1 and 3:1 ratio of chitosan and
PVA.
The increase of chitosan content in the film (1:1, 2:1,
3:1, chitosan: PVA) increases the swelling capacity sig-
nificantly i.e., 1:1 CPB (13.92 g/g) < 2:1 CPB (16 g/g) <
3:1 CPB (18.1 g/g). Similar trend is maintained in the
case of CPSNP films i.e., 1:1 CPSNP (9.8 g/g) < 2:1
CPSNP (13.1)g/g < 3:1 CPSNP (16 g/g). This is due to
the presence of more hydrophilic groups in the film net-
works which assist in improving the swelling character-
istic of the films [32].
3.2. UV-Vis Spectroscopy
The formation of AgNPs is analyzed by UV-Vis spec-
troscopy. To evaluate the optimum composition of chi-
tosan for the formation of silver nanoparticles, mild con-
ditions are employed to avoid reducing agents. By in-
creasing the concentration of chitosan from 1-3%(w/v) of
films the intensity of surface plasmon resonance absorp-
tion peak corresponding to silver nanoparticles has grad-
ually increased (0.105 to 0.195) with a slight shift in the
wavelength of the peak (413 to 421 nm) (Figure 4).
The surface Plasmon resonance absorption peak is ob-
served at 413-421 nm indicates the formation of silver
nanoparticles of smaller size with narrow size distribu-
tion [33]. The reduction capacity as well as the stabiliza-
tion of the formed nanoparticles increases with increase
of chitosan concentration due to the presence more num-
ber of reducible groups [33]. The solution containing, 3%
wt./v chitosan has shown an intense peak at 421 nm
compared to other compositions. To study the effect of
PVA in the formation of silver nanoparticles a film with
the following composition consisting of chitosan (3%
wt./v.)-PVA (1% wt./v.) solution is prepared. Surpris-
ingly, the intensity of surface plasmon peak absorption
has increased to 0.341 from 0.195. This indicates the
presence of PVA influences the reduction as well as the
stabilization process of silver nanoparticles. From the
foregoing experiments, we choose chitosan(3%)
PVA(1%) AgNP films for the forthcoming experi-
ments. PVA, chitosan and CPB films have not shown any
characteristic peak around 400-450nm in the U.V. spec-
tra (Data not shown).
3.3. FTIR Spectra
The FTIR spectra of CPB and CPSNP films are shown in
Figure 5.
The CPB film (Figure 5a) has shown absorption peaks
at 1664 cm-1 and 1325 cm-1 relating to amide I and III of
C=O stretching, N-H/C-N stretching and CH2 wagging
coupled with OH groups of chitosan respectively. The
peak observed at 1454 cm-1 is due to CH2 bending, and
the peak at 2933 cm-1 is characteristic of –CH2 asymmet-
ric stretching of CS/PVA. The absorption peak observed
at 3414 cm-1 indicates the hydrogen bonding nature of
OH/NH2 stretching. The silver nanoparticles loaded chi-
tosan film (Figure 5b) has shown all the above charac-
teristic peaks with a slight shift of the peak 1325 to
Figure 3. Swelling capacity of chitosan-PVA blend (CPB) and chitosan-PVA silver nanoparticle (CPSNP) films. (A) 1:1 (B)
2:1 and (C) 3:1 ratio of chitosan and PVA.

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 60
Figure 4. Surface plasmon resonanc e peak of silver nano
particles generated using different c o nc e ntr ations of (1-3
wt./v.% ) chitosan solutions and chitosan-PVA solutions.
Figure 5. FTIR spectra of chitosan, PVA, CPB and CPSNP
films.
1352 cm-1 corresponding to amide III band. In addition,
the stretching vibration at 3414 corresponding to
OH/NH2 groups has shifted to 3423 cm-1, indicating that
the silver particles are bounded to the functional groups
present both in chitosan and PVA. The shifting of the
peak is due to formation of co-ordination bond between
the silver atom and the electron rich groups (oxy-
gen/nitrogen) pr esent in ch itosan. This causes an in crease
in bond length and frequency. Blank chitosan and PVA
FTIR spectra are also depicted in Figure 5c and 5d for
comparison purpose. A ll the above observations found in
the IR spectra of films confirm the presence of silver
nanoparticles in the chitosan-PVA film networks.
3.4. XRD Analysis
The X-ray Diffraction (XRD) is used to confirm the na-
ture of crystal structure of the formed silver nano films
(Figure 6A). Broad XRD patterns of CSB and CPSNP
below 30o (arrow mark) indicates the semi-crystalline
nature of the major component i.e., ch itosan. In addition,
CPSNP has exhibited strong reflections around 38o, 44 o,
65 o, and 78 o characteristic of (111) (200) (220) and (311)
planes of face centered cubic (FCC) of the silver nano-
particles [33].
3.5. Thermogravimetric Analysis
Figure 6B illustrates the thermogravimetric analysis of
CPB and CPSNP films. The initial weight loss (below
100oC) observed in the films is due to loss of moisture
present in the fil ms. The weight loss observed in the case
of CPB film is 80% around 490oC where as the weight
loss observed for CPSNP at this temperature is only
73.83%. The weight loss difference between the CPB
and CPSNP represents the presents of silver nanoparti-
cles in CPSNP. The percentage amount of silver nano-
particles present in the CPSNP film can be calculated
from the difference in the weight loss between the CPB
and CPSNP films at 800oC (~6%), which is ~ 5.57%.
3.6. Electron Microscopic Analysis
The SEM analysis of the plain and silver nanoparticles
loaded chitosan-PVA films are shown in Figure 7A. The
plain CPB film (Figure 7A, a-b) has exhibited a dense
and uniform plain microstructure. Whereas CPSNP film
(Figure 7A, c-d) has shown the presence of defined na-
noparticles in the film (arrows). To find out the exact size
and morphology of the formed Ag nanoparticles in the
CPSNP film, TEM analysis is done. The TEM image
Figure 6. (A) X-ray diffraction patterns and (B) thermogra-
vimetric analysis of CPB and CPSNP films.

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 61
Figure 7. (A) Scanning elec tron microscopic images of (a-b)
CPB and (c-d) CPSNP films. Arrows represent silver na-
noparticles. (B) Transmission electron microscopy of silver
nanoparticles obtained from CPSNP film.
of the silver nanoparticles extracted from CPSNP film is
shown in Figure 7(b). The image indicates the particles
are spherical in nature with dispersed morphology. The
TEM results further illustrates that the particles formed
have an average size of ~16nm (Figure 7(b)). As per the
data available the silver nanoparticles generated in the
chitosan-PVA film are essential for antimicrobial appli-
cations.
3.7. Mechanical Properties
Many synthetic and natural polymeric materials are de-
veloped to treat the burn wounds and as antibacterial
materials. However, they have limited applicability due
to poor mechanical properties as well as lower rates of
water absorption. A number of chitosan- silver nano-
composites have been developed for many applications.
To have better mechanical strength silver impregnated
chitosan-PVA nanocomposites are developed in the pre-
sent investigation for higher applicability. The mechani-
cal properties of CPB and CPSNP films are presented in
Table 1. The higher stress at maximum load, modulus,
and elongation at break are noticed for CPSNP film
compared to CPB film. The main objective of this inves-
tigation is to produce high strength to chitosan-PVA
films and achieved by impregnating silver nanoparticles
into Chitosan-PVA films.
Table 1. Mechanical properties of CPB and CPSNP films.
Sample
Code
Stress at
Maximum
Load (MPa)
Modulus
(MPa) Elongation at break (%)
CPB 26.6128 970.2833 4.492
CPSNP 33.274 1195.9382 5.467
3.8. In Vitro Release
If these film materials are used as wound dressings, apart
from absorbing wound exudates during the healing proc-
ess, works as reservoirs to drugs for localized delivery to
prevent bacterial infection. In the present study, curcu-
min (CUR) is selected as a model drug. Curcumin is the
principal curcuminoid of the popular Indian spice tur-
meric, which is a member of the ginger family. It is used
in ayurvedic and in Chinese medicine since long back
[34]. In addition, it may be effective in treating malaria,
prevention of cervical cancer, and may interface with the
replication of HIV virus. Therefore loading efficiency of
curcumin into films is examined (Table 2).
It is observed that the loading efficiency is higher in
the case of CPSNP film compared to CPB films. This is
due to absorption of more number of curcumin molecules
on the silver nanoparticles in addition to entrapment in
the films. Figures 8(a) and 8(b) gives the structure of
curcumin and the drug delivery studies of CPB and
CPSNP respectively.
To confirm whether the loaded curcumin in the films
is in active form for functioning effectively in antibacte-
rial application, the FT-IR analysis is performed. Figure
8(c) supports the presence of curcumin in both plain and
Ag nanocomposites loaded films due to the presence of
additional peak s at 1505 cm-1 and 1265 cm-1 correspond-
ing to curcumin.
3.9. Curcumin Release Kinetics
The drug release kinetics was analyzed by plotting the
cumulative release data verses time and by fitting the
experimental release data into the following power law
equation and determining the exponent,
M1/Meq=ktn
Where Mt is the mass of water uptake at time t, Meq is the
equilibrium water uptake, and k and n are constants,
which are characteristic parameters of the specific (dis
solution medium) system. A value of n=0.5 indicates the
Table 2. Loading efficiency of curcumin into CPB, CPSNP
films, and curcumin release profiles from CPB and CPSNP
films.
Code %E n k (102) R2 Release
I Release II
CPB 69 0.990.58 0.98 12.87 100
CPSNP 81 1.271.47 0.98 6.26 97
(a)
(b)

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 62
Figure 8. (A) Chemical structure of curcumin, (B) Curcu-
min release profile from CCPB and CCP SNP films, and (C)
FTIR spectra of CCPB and CCPSNP films.
Fickian diffusion and n=1 implies case II transport; val-
ues of n between these limits define anomalous or
non-Fickian transports. The values of ‘k’ and ‘n’ have
been calculated by the least-squares method. These re-
sults along with correlation coefficients, r values are
presented in Table 2. The value of CCPB Films expo-
nent ‘n’ is found to 0.99 as calculated from the empirical
equation, respectively, which indicated anomalous nature.
Similarly, the value of CCPSNP film exponent of n is
1.27, which indicated that Zero order transport. The low-
er k values for all the systems indicate a lesser interaction
between the film materials and the curcumin.
3.10. Antimicrobial Activity
The use of Ag-containing chitosan-PVA films as func-
tional wound dressings is assessed by observing their
antimicrobial activity (based on the disc diffusion me-
thod, spread plate method, viable cell count method)
against some common bacteria and fungi found on burn
wounds: E. coli, P. sudomonas, micrococcus, Staphylo-
coccus, Candida albicus, and P. aeroginosa. Figure 9
exhibits the typical antimicrobial test results of films by
the disc method. It is found that the CPSNP films have
exhibited an inhibition zone (blue color arrows) whereas
CPB film does not involve in the inhibition zone process
Figure 9. Anti-microbial and anti-fungal activity of CPB
and CPSNP films against (a) E. coli (b) P. sudomonas (c)
Staphylococcus (d) Micrococcus (e) Candida albicus (f) P.
aeroginosa.
(yellow color arrows).
A second method is used to reveal the biocide action
of curcumin loaded CPB and CPSNP films. The nutrient
agar is spread into the Petri dish over the whole film and,
the E. coli culture is streaked on the solid surface of the
media. A significant difference is observed in the plates
containing CCPB film and CCPSNP film (Figure 10).
The observed results indicates that the CCPB film con-
taining petridish has shown lesser bacterial colonies
where as the CCPSNP film containing petridish exhibited
almost no bacterial colonies. This clearly demonstrates
that curcumin and nanoparticles loaded composite films
have excellent bacterial inhibition growth capabilities.
The killing kinetics of th e E. coli is also tested against
CPB, CPSNP, CCPB and CCPSNP films by performing
in mineral salt broth .The results suggest that a drastic
bacterial growth inhibition is observed for a period of 7
hrs and the results are determined by O.D measurements
(Figure 11). The order of the O.D values are CPB >
CCPB > CPSNP > CCPSNP. The results clearly indi-
cated that the CCPSNP film has exhibited excellent an-
timicrobial activity.
4. Conclusion
The present work demonstrates a simple method in pro-
ducing novel chitosan-PVA silver nanocomposite films.
The developed silver nanocomposite films have exhib-
ited fairly good mechanical strength and superior antim-
icrobial properties. Further, the current work demon-
strates a promising method to combine silver nano-
-composites with a natural compound (curcumin) in de-

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 63
Figure 10. Antibacterial activity of (a) no treatment (control)
(b) CCPB and (c) CCPSNP films.
Figure 11. Antibacterial efficiency of CPB, CPSNP,
CCPBand CCPSNP films against E. coli.
veloping novel antimicrobial agents .These agents may
find potential applications in antimicrobial packaging
materials and wound dressing/wo und burns.
5. Acknowledgements
KMR thanks the Defence Research and Development
Organization (DRDO) and Ministry of Defence, Gov-
ernment of India, New Delhi and KV thank U.G.C-SAP,
New Delhi for the partial financial support.
REFERENCES
[1] D. G. Yu, M. Y. Teng, W. L. Chou, M. C. Yang, “Char-
acterization and Inhibitory Effect of Antibacterial PAN
Based Hollow Fiber Loaded with Silver Nitrate,” Journal
of Membrane Science, Vol. 225, No. 1-2, November 2003,
pp. 115-123. doi:10.1016/j.memsci.2003.08.010
[2] M. Rai, A.Yadav, A.Gade,“Silver Nanoparticles as A
New Generation of Antimicrobial,” Biotechnology Ad-
vances, Vol. 27, No.1, January-February 2009, pp. 76-83.
doi:10.1016/j.biotechadv.2008.09.002
[3] M. Catauro, M. G. Ra ucci, F. de Gaetano and A. Marotta,
“Antibacterial and Bioactive Silver Containing
Na2O·CaO·2SiO2 Glass Prepared by Sol Gel Method,”
Journal of Materials Science: Materials in Medicine,
Vol.15, No.7, July 2004, pp. 831-837.
doi:10.1023/B:JMSM.0000032825.51052.00
[4] J. H. Crabtree, R. J. Burchette, R. A. Siddiqi, I. T. Huen,
L. L. Handott, A. Fishman, “The Efficacy of Silver-Ion
Implanted Catheters in Reducing Peritoneal Dialysis-Re-
lated Infections, ” Peritoneal Dialysis International, Vol.
23, No.4, July- August 2003, pp.368-374.
[5] D.-C. Tien, K.-H. Tseng, C.-Y. Liao, T.-T. Tsung, “Iden-
tification and Quantification of Ionic Silver from Colloi-
dal Silver Prepared by Electric Spark Discharge System
and Its Antimicrobial Potency Study,” Journal of Alloys
and Compounds, Vol. 473, No. 1-2, April 2009, pp.
298-302. doi:10.1016/j.jallcom.2008.05.063
[6] J. S. Kim, E. Kuk, K. N.Yu, J.-H. Kim, S. J. Park, H. J
Lee, S. H. Kim, Y. K. Park, Y. H. Park, C.-Y. Hwang, Y.
K. Kim, Y.-S. Lee, D. H. Jeong, M.-H. Cho, “Antimicro-
bial Effects of Silver Nanoparticles,” Nanomedicine:
Nanotechnology, Biology and Medicine, Vol.3, No.1,
March 2007, pp. 95-101.
doi:10.1016/j.nano.2006.12.001
[7] R. Kumar, H. Munstedt, “Polyamide/Silver Antimicrobi-
als: Effect of Crystallinity on the Silver Ion Release,”
Polymer International, Vol. 54, No. 88, August 2005, pp.
1180-1186. doi:10.1002/pi.1828
[8] J. J. Castellano, S. M. Shafii, F.Ko, G. Donate, T. E.
Wright, R. J. Mannari, W. G. Payne, D. J. Smith, M. C.
Robson, “Comparative Evaluation of Silver Containing
Antimicrobial Dressings and Drugs,” International
Wound Journal, Vol. 4, No.2, June 2007, pp. 114-122.
doi:10.1111/j.1742-481X.2007.00316.x
[9] M. Mishra, H. Kumar, K. Tripathi, “Diabetic Delayed
Wound Healing and the Role of Silver Nanoparticles,”
Digest Journa of Nanomaterials and Biostrctures, Vol. 3,
No. 2, June 2008, pp. 49-54.
[10] Q. Li, S. Mahendra, D. Y. Lyon, L. Brunet, M. V. Liga, D.
Li, P. J. Alvarez, “Antimicrobial Nanomaterials for Water
Disinfection and Microbial Control: Potential Applica-
tions and Implications,’’ Water Research, Vol. 42, No. 18,
November 2008, pp. 4591-4602.
doi:10.1016/j.watres.2008.08.015
[11] R. Jung, Y. Kim, H.-S. Kim, H. J. Jin, “Antimicrobial
Properties of Hydrated Cellulose Membranes with Silver
Nanoparticles,” Journal of Biomaterials Science: Polymer
Edition, Vol. 20, No. 3, 2009, pp. 311-324.
doi:10.1163/156856209X412182
[12] J. Jain, S. Arora, J. M. Rajwade, P. Omray, S. Khandel-
wal, K. M. Paknikar, “Silver Nanoparticles in Therapeu-
tics: Development of an Antimicrobial Gel Formulation
for Topical Use,” Molecular Pharmaceutics, Vol. 6, No.
5, May 2009, pp. 1388-1401. doi:10.1021/mp900056g
[13] A. Petica, S. Gavriliu, M. Lungu, N. Buruntea, C. Pan-
zaru, “Colloidal Silver Solutions with Antimicrobial
Properties,” Materials Science and Engineering: B. Vol.
152, No. 1-3, August 2008, pp. 22-27.
doi:10.1016/j.mseb.2008.06.021
[14] S. K. Bajpai, Y.M. Mohan, M. Bajpai, R. Tankhiwale, V.
Thomas, “Synthesis of Polymerstabilized Silver and Gold
Nanostructures,” Journal of Nanoscience and Nanotech-
nology, Vol. 7, No. 9, September 2007, pp. 2994-3010.
doi:10.1166/jnn.2007.911
[15] H. Kong, J. Jang, “Antibacterial Properties of Novel
Poly(methyl methacrylate) Nano Fiber Containing Silver
Nanoparticles,” Langmuir, Vol. 24, No.5, January 2008,
pp. 2051-2056. doi:10.1021/la703085e

Fabrication of Curcumin Encapsulated Chitosan-PVA Silver Nanocomposite Films for Improved Antimicrobial Activity 64
[16] V. K. Sharma, R. A. Yongard, Y. Lin, “Silver Nanoparti-
cles: Green Synthesis and Their Antimicrobial Activi-
ties,” Advances in Colloid and Interface Science, Vol.
145, No. 1-2, January 2009, pp. 83-96.
doi:10.1016/j.cis.2008.09.002
[17] V. Thomas, M. Namdeo, Y. Murali Mohan, S. K. Bajpai,
M. Bajpai, “Review on Polymer Hydrogel and Microgel
Metal Nanomposites: A Facile Nanotechnological Ap-
proach,” Journal of Macromolecular Science, Part A:
Pure Applied Chemistry, Vol. 45, No. 1, January 2008, pp.
107-119. doi:10.1080/10601320701683470
[18] M. Zeng, Z. Fang, C. X, “Novel Method of Preparing
Microporous Membrane by Selective Dissolution of Chi-
tosan/Poly Ethylene Glycol Blend Membrane,” Journal of
Applied Polymer Science, Vol. 91, No. 5, March 2004, pp.
2840-2847. doi:10.1002/app.13469
[19] H-S. Kim, K-H. Lee, S-G. Kim, “Growth of Monodis-
perse Silver Nanoparticles in Polymer Matrix by Spray
Pyrolysis,” Aerosol Science and Technology, Vol. 40, No.
7, July 2006, pp. 536-544.
doi:10.1080/02786820600714361
[20] S. Liu, J. He, J. Xue, W. Ding, “Efficient Fabrication of
Transparent Antimicrobial Poly (Vinyl Alcohol) Thin
Films,” Journal Nanooparticle Research, Vol. 11, No.3,
October 2007, pp. 553-560.
doi:10.1007/s11051-007-9321-8
[21] S. Lu, W. Gao, H. Y. Gu,“Construction, Application and
Biosafety of Silver Nanocrystalline Chitosan Wound
Dressing,” Burns, Vol. 34, No. 5, August 2008, pp.
623-628. doi:10.1016/j.burns.2007.08.020
[22] A. M. Durango, N. F. F. Soares, S. Benevides, J. Teixeira,
M. Carvalho, C. Wobeto, N. J. Andrade, “Development
and Evaluation of an Edible Antimicrobial Film Based on
Yam Starch and Chitosan,” Packaging Technology Sci-
ence, Vol. 19, No. 1, January-February 2006, pp. 55-59.
doi:10.1002/pts.713
[23] S. Tripathia, G. K. Mehrotra and P. K. Dutta, “Physico-
Chemical and Bioctivity of Crosslinked Chitosan-PVA
Film for Food Packaging Applications,” International
Journal of Biological Macromolecules, Vol. 45, No. 4,
November 2009, pp. 372-376.
doi:10.1016/j.ijbiomac.2009.07.006
[24] O. Suwantong, P. Opanasopit, U. Ruktanonchai, P. Su-
paphol, “Electrospun Cellulose Acetate Fiber Mats Con-
taining Curcumin and Release Characteristic of the Her-
bal Substance,” Polymer, Vol. 48, No. 26, December
2007, pp. 7546-7557. doi:10.1016/j.polymer.2007.11.019
[25] S. K. Bajpai, Y. Murali Mohan, “Recent Advances in
Nanoscience and Technology”, Bentham Science Pub-
lishers, Dubai, 2009.
[26] T. R. Thatiparti, A. Kano, A. Maruyama, A. Takahara,
“Novel Silver-Loaded Semi-Interpenetrating Polymer
Network Gel Films with Antibacterial Activity,” Journal
of Polymer Science Part A: Polymer Chemistry, Vol. 47,
No. 19, October 2009, pp.4950–4962.
doi:10.1002/pola.23546
[27] A. Findon, G. McKay, H. S. Blair “Transport Studies for
the Sorption of Copper Ions by Chitosan,” Journal of En-
vironmental Science and Health. Part A, Environmental
science and engineering, Vol. 28, No. 11, 1993, pp.
173-185. doi:10.1080/10934529309375870
[28] N. G. Kandile, A. S. Nasr, “Environment Friendly Modi-
fied Chitosan Hydrogels as A Matrix for Adsorption of
Metal Ions, Synthesis and Characterization,” Carbohy-
drate Polymers, Vol. 78, No. 4, November 2009, pp
753-759. doi:10.1016/j.carbpol.2009.06.008
[29] K. Vimala, K. Samba Sivudu, Y. Murali Mohan, B.
Sreedhar, K. Mohana Raju, “Controlled Silver Nanoparti-
cles Synthesis in Semi-Hydrogel Networks of
Poly(Acrylamide) and Carbohydrates: A Rational Meth-
odology for Antibacterial Application,” Carbohydrate
Polymers, Vol. 75, No. 4, February 2009, pp. 463-471.
doi:10.1016/j.carbpol.2008.08.009
[30] K.Vimala, Y. Murali Mohan, K. Samba Sivudu, K. Vara-
prasad, S. Ravindra, N. Narayana Reddy, Y. Padma, B.
Sreedhar, K. MohanaRaju, “Fabrication of Porous Chito-
san Films Impregnated with Silver Nanoparticles: A Fac-
ile Approach for Superior Antibacterial Applications,”
Colloids and Surfaces B: Biointerfaces, Vol. 76, No. 1,
March 2010, pp. 248-258.
doi:10.1016/j.colsurfb.2009.10.044
[31] J. Ostrowska-Czubenko, M. Gierszewska-Drużyńska,
“Effect of Ionic Crosslinking on the Water State in Hy-
drogel Chitosan Membranes,” Carbohydrate Polymers,
Vol. 7, No. 3, July 2009, pp.590-598.
doi:10.1016/j.carbpol.2009.01.036
[32] S. Ekici, D. Saraydin, “Interpenetrating Polymeric Net-
work Hydrogels for Potential Gastrointestinal Drug Re-
lease,” Polymer International, Vol. 56, November 2007,
No. 11, pp. 1371-1377.
[33] V. Thomas, Y. Murali Mohan, B. Sreedhar, S. K. Bajpai,
“Fabrication, Characterization of Chitosan/Nanosilver
Film and Its Potential Antibacterial Application,” Journal
of Biomaterials Science, Part A: Polymer Edition, Vol. 20,
No.14, 2009, pp. 2129-2144.
[34] H. Hatcher, R. Planalp, J. Cho, F. M. Torti, S. V. Torti,
“Curcumin: From Ancient Medicine to Current Clinical
Trials,” Cellular and Molecular Life Sciences, Vol. 65,
No. 11, June 2008, pp. 1631-1652.
doi:10.1007/s00018-008-7452-4