polymers

Article
Synthesis of Intelligent pH Indicative Films from
Chitosan/Poly(vinyl alcohol)/Anthocyanin Extracted
from Red Cabbage
Thuy-Vi Vo 1 , Tan-Hiep Dang 2 and Bing-Hung Chen 1, *
1 Department of Chemical Engineering, National Cheng Kung University, Tainan 70101, Taiwan
2 Department of Chemical Technology, Ho Chi Minh City University of Food Industry,
Ho Chi Minh City 70000, Vietnam
* Correspondence: bkchen@mail.ncku.edu.tw; Tel.: +886-6-275-7575 (ext. 62695); Fax: +886-6-234-4496




Received: 14 May 2019; Accepted: 24 June 2019; Published: 26 June 2019


Abstract: In this study, pH indicative films were successfully synthesized from hydrogels made by
blending 1% poly(vinyl alcohol) (PVA) and 1% chitosan (CS) with anthocyanin (ATH) and sodium
tripolyphosphate (STPP). Particularly, ATH extracted from red cabbage was used as the pH indicator,
while STPP was utilized as the cross-linking agent to provide better mechanical properties of the
cast films. FT-IR spectra confirmed the existence of the ATH in the cast films. Moreover, the tensile
strength, the elongation-at-break, and the swelling indices of the cast films were measured. In general,
these properties of pH indicative films were profoundly influenced by the compositions of PVA/CS
and the STPP dosage applied in the hydrogels. For example, the tensile strength could change from
43.27 MPa on a film cast from pure PVA hydrogel to 29.89 MPa, if 35% of the PVA hydrogel was
substituted with CS. The cast films were applied as a food wrap that could be used to monitor visually
the quality of the enwrapped food via the color change of the film upon the variation in pH values of
the enwrapped food. In practice, a sequential change in color was successfully observed on the pH
indicative films partially enwrapping the pork belly, indicating the spoilage of the meat.

Keywords: poly(vinyl alcohol); chitosan; anthocyanin; pH indicative film; food wrap

1. Introduction
The United Nations adopted 17 Sustainable Development Goals (SDGs) as part of the 2030 Agenda
for Sustainable Development on September 25 2015. As an example of one of these goals, SDG 2 is Zero
Hunger, which aims to end hunger, to achieve food security, and to promote sustainable agriculture.
Indeed, food spoilage and security have always been a predominant concern for human beings since
the early days, as the adverse effects of food poisoning could lead to disastrous consequences for human
survival. Consequently, relentless efforts have been invested in studying and in the development of
proper packaging and preservation of food all around the world [1–4]. For example, the invention of
synthetic polymers has resulted in the mass production of plastic bags and thin-film wraps that are
widely applied as food packaging materials. In general, these plastic films have relatively low gas
permeability. As such, they often serve as passive barriers to moisture, oxygen and microorganisms to
protect the enwrapped food from spoilage. In addition, several SDGs have been proposed to encourage
more sustainable consumption and production patterns as well as more sustainable development
of our world. Therefore, renewable packaging substitutes, e.g., plastics from renewable biomass
sources, are under extensive development to replace the synthetic components in food packaging [1–4].
Even more, additional components such as antioxidants and colorants are processed into the plastic
films [2]. One example is shown in the inclusion of anthocyanins, commonly found as antioxidants

Polymers 2019, 11, 1088; doi:10.3390/polym11071088 www.mdpi.com/journal/polymers

Polymers 2019, 11, 1088 2 of 12

and pigments in plants [5], to actively indicate the variation in pH values near the contact between
food and package [6].
One aim of this work is to develop a food packaging film made as much as possible from renewable
and biodegradable components. Chitosan (CS) is consequently chosen as the substituent to partially
replace poly(vinyl alcohol) (PVA) in the packaging film. In addition, anthocyanin (ATH), commonly
used as food colorants [5] and extracted from red cabbage in this work, is applied to the cast film for
actively monitoring the change in pH values of the enwrapped food.
Chitosan (CS) is a natural polymer derived from chitin and found abundantly in the shells
of crabs and shrimps [7,8]. It possesses special characteristics, such as a polycationic nature and
chelating properties, as well as a film-forming capability, due to the affluent amino and hydroxyl
functional groups in chitosan. Chitosan is found to exhibit many interesting biological activities, such
as antimicrobial activity, biodegradability, etc. [7–11]. Adversely, chitosan is not always preferable
in polymer processing for film production owing to its inferior mechanical properties, such as lower
tensile strength, less elongation-at-break, and Young’s modulus. Hence, chitosan is commonly blended
with other polymers with more flexible chains, e.g., poly(vinyl alcohol) (PVA), for further processing of
polymer films [8–10,12–16].
Poly(vinyl alcohol) (PVA) is a water-soluble biodegradable synthetic polymer with large tensile
strength and flexibility, which also allows very low permeability of gases, including O2 and CO2 .
PVA can possess a moderate water solubility at an elevated temperature [9,10,16]. With excellent film
forming properties, PVA has been widely used in applications as thin films, e.g., for food packaging
and packaging for laundry detergent tablets. The composites blended with PVA and chitosan are
known to have improved stability, biocompatibility, and mechanical strength, relative to those of pure
PVA and pure CS polymers [8–14,16].
Anthocyanins are natural colorants, water-soluble pigments abundant in plants, such as red
cabbage, blueberry, eggplants and flowers. They could be used as a color indicator depending on pH
values. In particular, the anthocyanins obtained from red cabbage can vary in color from red to purple
and even blue at different pH values, which facilitates the recognition of food quality in terms of pH
changes [5,6,8,10,14,15,17–21]. Therefore, the indicative film synthesized from PVA and CS with ATH
as the color indicator could express good spectroscopic and physicochemical characteristics and could
be applied in intelligent food packaging [14].
In processing thin polymer films, the addition of cross-linking substances, such as sodium
tripolyphosphate (STPP), glyoxal, and glutaraldehyde, is necessary to improve the mechanical strength
of cast films [22]. These cross-linkers could connect one polymer chain to different macromolecules to
eventually form polymer mats. These polymer mats, compared to the less cross-linked ones, exhibit
better membrane durability and reduced water absorption [10,14]. Specifically, STPP is capable of
forming an ether or ester by cross–linking hydroxyl groups on the same molecules or multiple polymer
molecules [14,23].
Solvent casting and hot melt extrusion are currently two of the most popularly applied processing
methods for the production of polymer thin-films. The hot melt extrusion process often requires the
input of high energy, such as mechanical energy in the form of very high shear stress and/or thermal
energy, to ensure the processed polymer in the melt state. As such, this method is not suitable for
processing a polymer with heat and/or shear sensitive molecules. In contrast, solvent casting is the
predominant method for manufacturing films containing temperature-sensitive ingredients, such as
anthocyanin, because the temperature required to evaporate the solvent is often lower than the process
temperature of the hot melt extrusion [24].
As aforementioned, PVA/CS composites were chosen as model polymers for the study of pH
indicative thin-film wraps in this work. PVA/CS hydrogels have recently been extensively studied for
their potential applications, such as in biomedical applications and biomaterials [13,16]. Owing to
the polysaccharide nature of chitosan, the percent elongation-at-break decreases significantly with
increasing CS content in hydrogels [13]. In this report, the influence of the cross-linking agent, e.g.,
Polymers 2019, 11, 1088 3 of 12

STPP, and the mixing ratio of PVA/CS on the mechanical properties of the resulted thin films, especially
in the changes of tensile strength, the elongation-at-break and water absorption (swelling index), were
investigated. The pH indicative films were examined by FT-IR to determine the chemical interaction
among polymer components and anthocyanin as well.

2. Materials and Methods

2.1. Materials
Chitosan (CS, 75%–85% deacetylated), poly(vinyl alcohol) (PVA, Mw = 89,000–98,000), and sodium
tripolyphosphate (STPP) were purchased from Sigma-Aldrich. Hydrochloric acid (37%) and glacial
acetic acid were supplied by Merck. All chemicals were of analytical grade and used as received.
Deionized water with resistivity greater than 18.2 MΩ-cm was used in sample preparation in this study.
The anthocyanin (ATH) used in this work was extracted from red cabbage (Brassica oleracea), a
produce harvested from the Lâm Đồng Province of Vietnam, chiefly following the solvent extraction
method reported by Fuleki and Francis [25]. Accordingly, the extraction procedure for ATH from
red cabbage has been modified in this work and is given as follows. Typically, 150 g of minced red
cabbage is macerated and soaked in ca. 303 mL of solvents, composed of absolute ethanol (150 mL)
and concentrated HCl (3 mL) in deionized water (150 mL), in an ultrasonic bath for 60 min [10,18,26].

2.2. Preparation of pH Indicative Films

The pH indicative films were synthesized with the solvent (solution) casting method using the
hydrogels prepared from the mixed solutions of chitosan (CS)/poly(vinyl alcohol) (PVA) incorporated
with anthocyanin (ATH) by a procedure revised from that of Stefani et al. [14,19]. STPP was used as
the crosslinking agent to the polymer films. All ingredients for making up hydrogels as precursors
to the pH indicative films were previously prepared as pure solutions. Notably, the concentrations
of polymers and STPP in the parental solutions were 1 wt% for both PVA and CS, and 0.1 wt% for
STPP, respectively. Furthermore, the concentration of anthocyanin in the extract was maintained at
86.67 mg/L.
The parental solution of 1 wt% PVA was obtained by dissolving 1 g of PVA in 99 g of deionized
water with continuous magnetic stirring at 70 ◦ C. The PVA solution was then allowed to cool down to
the ambient temperature, at which it became transparent and quite viscous. Similarly, the 1 wt% CS
solution was prepared by dissolving 1 g of chitosan flake in 99 g of 1 wt% acetic acid solution under
continuous agitation for 24 h at room temperature.
To study the effect of chitosan and the crosslinking agent STPP on the mechanical properties of
the finished pH indicative films, various volume ratios of 1 wt% PVA solution and 1 wt% chitosan
solutions were mixed and agitated constantly for 30 min to give the PVA/CS solutions. Subsequently, a
proper volume of ATH extract was added into the aforementioned mixed solutions of PVA/CS and
stirred for another 30 min at room temperature. The volume of the added ATH solution was fixed at a
quarter of the total volume of the final PVA/CS/ATH solutions. For example, 100 mL of PVA/CS/ATH
solution was composed of 75 mL PVA/CS solution and 25 mL ATH solution. Afterwards, a suitable
volume of 0.1% STPP solution was added to the solutions of PVA/CS/ATH in order to promote the
cross-linkage in the resultant hydrogels.
Explicitly, the formulation used for the study on the effect of chitosan dosage on the mechanical
properties of pH indicative films was given in proportion as follows: (1) 75 mL PVA/CS solution, in
which various volume ratios of PVA solution to CS solution were used; (2) 25 mL ATH extract solution;
and (3) 6 mL of 0.1% STPP solutions. Similarly, the formulation for the study on the effect of the
crosslinking agent STPP on the mechanical properties of pH indicative films was given in proportion
as follows: (1) 75 mL PVA/CS solution, i.e., the volume ratio of PVA solution to CS solution kept at
75/25; (2) 25 mL ATH extract solution; and (3) the predetermined amount of 0.1% STPP solutions.
Polymers 2019, 11, 1088 4 of 12

Polymers 2019, 11, 1088 4 of 12

The pH value of the hydrogel, consisting of PVA/CS/ATH/STPP, was adjusted to 6 with 1.0 M
dishes (120
NaOH. mmthe
Finally, diameter), followed
pH indicative filmbywastheattained
removal byofcasting
solventthefrom the cast
obtained film by
hydrogel (50placing
mL) inthese
Petri
Petri
dishesdishes
(120 mm in an oven at 45
diameter), °C for 72
followed by h.
theAll the experiments
removal reported
of solvent from in this
the cast filmwork were performed
by placing these Petri
at least in
dishes inan
triplicate.
oven at 45 ◦ C for 72 h. All the experiments reported in this work were performed at least
in triplicate.
2.3. Characterization of pH Indicative Films
2.3. Characterization of pH Indicative Films
A universal tensile testing instrument, Model RTC-1210A, from A&D Company (Tokyo, Japan)
A universal
was employed totensile
measure testing instrument,
the tensile Model
strength (TS)RTC-1210A, from A&D
and the percent Company (Tokyo,
elongation-at-break Japan)
of the pH
was employed
indicative to measure
film using the tensile
the method strength
described in (TS)
ASTM and the percent
D882 [27]. Theelongation-at-break
specimens for the of the pH
universal
indicative
tensile film were
testing using prepared
the method indescribed
rectangular in ASTM
strips D882
with a[27]. The specimens
dimension of 100 for
mmthe universal
(length) tensile
× 10 mm
testing were prepared in rectangular strips with a dimension
(width) × 50 μm (thickness). The speed of testing was chosen at 50 mm/min. of 100 mm (length) × 10 mm (width) ×
50 µm (thickness).
The The speed
swelling index of testing was
was determined bychosen at 50 mm/min.
the gravimetric method. The prepared films were cut
into coupons of 30 × 30 mm . These strips of pH indicative method.
The swelling index was 2 determined by the gravimetric films were The prepared
weighed films wereand
individually cut
into coupons of 30 × 30 mm 2 . These strips of pH indicative films were weighed individually and
subsequently immersed in 100 mL of deionized water for various periods of time at room
subsequently At
temperature. immersed
each time in 100
intervalmL of deionized
(0.5, water
1, 2, 5, 10, for various
15 and 20 min),periods
sampleofspecimens
time at room
weretemperature.
taken out,
At each time interval (0.5, 1, 2, 5, 10, 15 and 20 min), sample specimens
dried and weighed. Consequently, the swelling index (SI) was calculated using Equation (1): were taken out, dried and
weighed. Consequently, the swelling index (SI) was calculated using Equation (1):
m m
SI (%)  m11 − m00 100 (1)
SI (%) = m0 × 100 (1)
m0
where SI is the welling index, namely the percentage of water adsorption by the film, as well as m0
where SI is the welling index, namely the percentage of water adsorption by the film, as well as m0 and
and m1 are the weights (g) of the film initially and at the given time t, respectively.
m1 are the weights (g) of the film initially and at the given time t, respectively.
The functional groups of as–prepared indicative films were characterized by using the FT-IR
The functional groups of as–prepared indicative films were characterized by using the FT-IR
Spectrometer (Bruker Tensor 27, Germany) in the Institute of Chemical Technology (Ho Chi Minh
Spectrometer (Bruker Tensor 27, Germany) in the Institute of Chemical Technology (Ho Chi Minh City,
City, Vietnam). The FT-IR spectra of the samples diluted in KBr pellets were recorded in the 4000–
Vietnam). The FT-IR spectra of the samples diluted in KBr pellets were recorded in the 4000–400 cm−1
400 cm−1 range with a resolution−1of 4 cm−1. In addition, the morphology of the films was observed
range with a resolution of 4 cm . In addition, the morphology of the films was observed using a
using a Field Emission Scanning Electron Microscope (FE–SEM, Hitachi S–4800, Tokyo, Japan) at the
Field Emission Scanning Electron Microscope (FE–SEM, Hitachi S–4800, Tokyo, Japan) at the Research
Research Laboratories of Saigon High-Tech Park (SHTPLABS, Ho Chi Minh City, Vietnam)
Laboratories of Saigon High-Tech Park (SHTPLABS, Ho Chi Minh City, Vietnam)

2.4.
2.4. Coloration
Coloration of
of pH
pH Indicative
Indicative Films
Films
The colorationof
The coloration ofthe
thepHpH indicative
indicative films
films waswas demonstrated
demonstrated in the inbuffer
the buffer solutions
solutions with
with pH pH
values
values from 1 to 13. The films were cut into small coupons of 22 × 2 cm 2 and submerged in buffer
from 1 to 13. The films were cut into small coupons of 2 × 2 cm and submerged in buffer solutions.
solutions. Usually,
Usually, the the color
color change change
could could beand
be observed observed andwithin
recorded recorded30 s.within 30 s.

3.
3. Results
Results

3.1. Synthesis
Synthesis and
and Chemical
Chemical Characteristics
Characteristics of
of pH
pH Indicative
Indicative Films
Films

Anthocyanin
Anthocyanin (ATH)
(ATH) was
was successfully
successfully extracted
extracted from
from red
red cabbage.
cabbage. Figure
Figure 11 showed
showed clearly
clearly the
the
color change in buffer solutions with an ATH concentration at 86.67 mg/L from pH 1 to pH 13. One
color change in buffer solutions with an ATH concentration at 86.67 mg/L from pH 1 to pH 13. One
advantage
advantage associated
associated with
with the
the use
use of
of anthocyanin
anthocyanin as as aa pH
pH indicator
indicator isis reflected
reflected from
from their
their obvious
obvious
change
change in color upon
in color upon contact
contactwith
withdifferent
differentpH pHvalues.
values.For
For example,
example, their
their colors
colors areare
redred or pink
or pink in
in the
the acidic solutions, and blue or green in alkaline solutions. Consequently, the ATH was used as a
acidic solutions, and blue or green in alkaline solutions. Consequently, the ATH was used as a pH
pH indicator
indicator andand formulated
formulated to cast
to cast the pH
the pH indicative
indicative filmsfilms composed
composed of PVA,
of PVA, CS and
CS and STPP.
STPP.

Figure 1. Color change of anthocyanin extract from pH 1 (left) to pH 13 (right).

Figure 1. Color change of anthocyanin extract from pH 1 (left) to pH 13 (right).

Polymers 2019, 11, 1088 5 of 12

Prior 2019,
Polymers to the addition
11, 1088 of the cross-linker STPP, the solution consisting of PVA/CS/ATH in5 aof 12
volume ratio of 26.25/48.75/25, viz. (75 × 0.35)/(75 × 0.65)/25, at pH 6.1 exhibited a pink-magenta 5color
Polymers 2019, 11, 1088 of 12

(Figure 2a),
Priorsimilar
to thetoaddition
that of theof ATH extract at pHSTPP,
the cross-linker 6 (Figure 1). The hydrogel
the solution consistingwasofgradually
PVA/CS/ATH formedin a
in the aforementioned
volume
Prior toratio
the of PVA/CS/ATH
26.25/48.75/25,
addition solution after
viz. (75 × 0.35)/(75
of the cross-linker the slow
STPP, the×solution addition
0.65)/25,consistingof STPP
at pH 6.1 exhibitedin a dose at the
a pink-magenta
of PVA/CS/ATH volume
in a volume color
ratio
ratio of
of STPP
(Figure 2a),solution
26.25/48.75/25, toviz.
similar to PVA/CS/ATH
that of the
(75 × ATH solution
extract
0.35)/(75 atpH
at 6%.6 The
× 0.65)/25, pH6.1
(Figure
at pH indicative
1). The film
hydrogel
exhibited a was
wascast after water
gradually
pink-magenta formed
color
hadin been
(Figurethe2a), removed
similar to from
aforementioned ofthe
the hydrogel
thatPVA/CS/ATH with
solution
ATH extract an 6 oven-drying
after
at pH the slow
(Figure process
1).addition
The at was
of STPP
hydrogel 45inC for at72the
a dose
gradually h.volume
formedAs
aforementioned,
ratio
in the the
of STPP solution
aforementioned film was cast from
to PVA/CS/ATH
PVA/CS/ATH 50
solution mL of
solution hydrogels
after theat slow
6%. The in a Petri dish
pH indicative
addition of
of STPP infilm 12 mL in
wasatcast
a dose diameter.
the after
volumewater
Explicitly,
had
ratio of STPP thissolution
been dry film
removed to was
fromcomposed
PVA/CS/ATH approximately
the hydrogel
solution with
at 6%.anThe of 123.8 mg PVA,
oven-drying
pH indicative 230
process
film mg
was CS,after
at cast
45 1.02for
C mg72ATH
water h. As
had
and
been 2.83 mg STPP.
aforementioned,
removed the hydrogel
from the film was with
cast from 50 mL of hydrogels
an oven-drying process at in45 a◦ CPetri
for 72dish of 12
h. As mL in diameter.
aforementioned,
theExplicitly, thisfrom
film was cast dry 50
film
mLwas composedinapproximately
of hydrogels of 123.8
a Petri dish of 12 mL inmg PVA, 230
diameter. mg CS,this
Explicitly, 1.02 mg
dry ATH
film
and 2.83 mg STPP.
was composed approximately of 123.8 mg PVA, 230 (b)mg CS, 1.02 mg ATH and 2.83 mg STPP.
(a)

(a) (b)

= 120 mm

Figure 2. Poly(vinyl alcohol)= (PVA)/

120 mmchitosan (CS)/ anthocyanin (ATH) hydrogel and the cast film: (a)
before and (b) after the evaporation of water.
Figure 2. Poly(vinyl alcohol) (PVA)/chitosan (CS)/anthocyanin (ATH) hydrogel and the cast film: (a)
Figure
before and2.(b)
Poly(vinyl alcohol) (PVA)/
after the evaporation chitosan (CS)/ anthocyanin (ATH) hydrogel and the cast film: (a)
of water.
Interestingly,
before and (b)theafter
color
the of the resultant
evaporation film changed from pink-magenta as a wet hydrogel
of water.
(Figure 2a), to sea-green
Interestingly, the color as a of
dried
thecast film (Figure
resultant 2b). It is from
film changed surmised that color changes
pink-magenta as a wetduring hydrogelthe
oven-drying process
Interestingly, are
the mainly
color the
of theresult of
resultant the pH
film change
changed in the
from
(Figure 2a), to sea-green as a dried cast film (Figure 2b). It is surmised that color changes during thehydrogel
pink-magenta owing asto athe
wet gradual
hydrogel
evaporation
(Figure 2a),
oven-drying of water
to
process from
sea-green
are the
as aPVA/CS/ATH/STPP
mainly dried cast film
the result of the(Figure
pH hydrogel, insurmised
2b). It is
change in which
the both
thatsodium
hydrogel color hydroxide
owingchanges andthe
during
to the gradual
acetic acid existed
oven-drying
evaporation of watersimultaneously
process from arethemainly inthetheresult
PVA/CS/ATH/STPP gel. of thehydrogel,
pH change in the both
in which hydrogelsodium owing to the gradual
hydroxide and
The coloration
evaporation
acetic acid existed of
of water the cast pH indicative
from the PVA/CS/ATH/STPP
simultaneously in the gel. films submerged in buffer solutions
hydrogel, in which both sodium hydroxide and with pH values
from 1 tocoloration
acetic
The 12 is
acid shown
existed incast
Figure
of simultaneously
the 3. These
in the films
pH indicative films were
gel. submergedsensitive to thesolutions
in buffer pH values withofpH thevalues
solutions.
from
1 to 12 is shown in Figure 3. These films were sensitive to the pH values of the solutions. Commonly,than
Commonly, The the color
coloration change
of the on these
cast pH pH indicative
indicative films
films occurred
submerged very
in quickly,
buffer in
solutions usually
with less
pH values
the
30s.
color The
from coloration
1 toon
change 12these of
is shownthese films
pH indicative could
in Figure be
3. These
films attributable
occurred filmsvery to
were the transformation
sensitive
quickly, to the
in usually in
pH
less the
than chemical
values of the
30s. The structure
solutions.
coloration
of ATH
of these molecules
Commonly, the with
films could color respect
changeto
be attributable onthe pHpH
these
to the values of the films
indicative
transformation solution [5,14,26].
occurred
in the chemical veryAtquickly,
pH 1,ofthis
structure in film
usually
ATH shows a
less than
molecules
reddish
30s. color,
The and
colorationmarkedly
of thesechanges
films its
could color
be from reddish
attributable to to
the purple as the
transformation
with respect to the pH values of the solution [5,14,26]. At pH 1, this film shows a reddish color, and pH is
in increased
the chemical to 6. It is
structure
blueofatATH
markedly pHchanges
7molecules
and 8,its but changes
with
color from to
respect sea
to the
reddish greenpH
to at pH as
values
purple 9.of
With
thethe
pH anisincrease
solution of to
pH
[5,14,26].
increased 6.toIt 12,
At ispH this film
1, this
blue at pH becomes
film shows
7 and 8, a
yellow-green
reddish fast.
color, The
and vivid
markedly color changes
changes itsof the
color prepared
from films
reddish to against
but changes to sea green at pH 9. With an increase of pH to 12, this film becomes yellow-green fast.It is
purple the
as pH
the values
pH is have
increased enabled
to 6.
their
Theblueapplications
vividat color
pH 7changes
andas 8,
pH but
ofindicators
changes
the in
preparedto food
sea packaging,
green
films at pHthe
against and this
9. With
pH anwill
values be demonstrated
increase
have of pH to
enabled 12,inapplications
their the film
this latterbecomes
part
as
of this report.
pHyellow-green
indicators in foodfast. The vivid color
packaging, and changes
this will of bethe prepared films
demonstrated in theagainst
latter the
partpH of values have enabled
this report.
their applications as pH indicators in food packaging, and this will be demonstrated in the latter part
of this report.

pH 1 pH 2 pH 3 pH 4 pH 5 pH 6

pH 1 pH 2 pH 3 pH 4 pH 5 pH 6

pH 7 pH 8 pH 9 pH 10 pH 11 pH 12

Figure
Figure 3.
3. Coloration
Coloration of
of the
the cast
cast pH
pH indicative
indicative films
films submerged
submerged in
in solutions
solutions at
at different pH values.
different pH values.
pH 7 pH 8 pH 9 pH 10 pH 11 pH 12
The FT-IR spectra of PVA, CS, ATH and the PVA/CS/ATH film are shown in Figure 4. The film
Figure 3. Coloration of the cast pH indicative films submerged in solutions at different pH values.
examined was synthesized from the ATH-containing hydrogel with a volume ratio of PVA/CS=35/65.
On the FT-IR spectrum of PVA, peaks corresponding to 3337.6 cm−1 and 1427.5 cm−1 are attributed
 Polymers 2019, 11, 1088 6 of 12

The FT-IR spectra of PVA, CS, ATH and the PVA/CS/ATH film are shown in Figure 4. The
Polymers 2019, 11, 1088
film
6 of 12
examined was synthesized from the ATH-containing hydrogel with a volume ratio of PVA/CS=35/65.
On the FT-IR spectrum of PVA, peaks corresponding to 3337.6 cm–1 and 1427.5 cm–1 are attributed to
tothe
thestretching
stretchingand andthe bending
the bending vibrations
vibrations of of
–OH–OHgroup,
group,while the the
while bands at 1331.5
bands cm−1cm
at 1331.5 and 2941.3
−1 and

cm are the result of –CH2 stretching

2941.3–1 cm −1 are the result of –CH 2 vibrations. The peak appearing at 1656.6 cm arises from the
stretching vibrations. The peak appearing at –1
1656.6 cm −1 arises
crystallized
from water molecules
the crystallized in the in
water molecules polymer. The bands
the polymer. The bands fromfrom
919.2 to 1093.9
919.2 cmcm
to 1093.9 feature
–1 −1 feature the
oscillations
the oscillations ofofthe theC–O–H
C–O–Hgroup group[12,28].
[12,28]. For
For pure chitosan (CS),
pure chitosan (CS), the
theabsorption
absorptionatat3447.48
3447.48 cmcm −1 –1

correspondstotothe
corresponds thestretching
stretching–OH –OHgroup,
group,and andthe
theband
bandatat2925.58
2925.58cm cm−1 is one
–1 is one characteristic
characteristic ofof the
the –
CH22 group
–CH group bonded
bonded to to–OH.
–OH.The Thepeak
peakatat1636.65
1636.65cm
cm isisattributed
−1–1 attributedtotoC–O C–Ostretching
stretchingofofthe
theacetyl
acetyl
group(amide
group (amide I).I). Furthermore,
Furthermore, thethe peak
peak atat 1421.34
1421.34 cmcm is the bending vibration of the –C–OH group,
−1–1is the bending vibration of the –C–OH group,

whilethat
while thatofof 1384.18
1384.18 cm cm is the result of the knife in the plane of the –OH group. The regions from
−1–1is the result of the knife in the plane of the –OH group. The regions from

1153.51
1153.51 toto 1038.69
1038.69 cm–1are
cm −1
arethe
thecharacteristics
characteristicsofofthethesymmetric
symmetricoscillation
oscillationofofthetheC–O–C
C–O–Cgroup
group[14,28].
[14,28].
AsAsforfor the FT-IR spectrum of anthocyanin, the uptake at 3299.85 cm relates to the –OH group, and
the FT-IR spectrum of anthocyanin, the uptake at 3299.85 cm −1–1relates to the –OH group, and

thepeak
the peakatat 1639.02
1639.02 cmcm represents the C=C aromatic ring, bands from 1044.58 to 1084.89 cm arise
−1–1represents the C=C aromatic ring, bands from 1044.58 to 1084.89 cm−1 –1
arise
fromthe
from theoscillations
oscillationsofofthe the C–O–C
C–O–C group
group [14].
[14].

(a) PVA (b) CS

(c) ATH (d) PVA/CS/ATH Film

Figure4.4.FT-IR
Figure FT-IRspectra
spectraofof(a)
(a)PVA,
PVA,(b)
(b)chitosan,
chitosan,(c)(c)anthocyanin,
anthocyanin,(d)
(d)PVA/CS/ATH
PVA/CS/ATH film.
film.

TheFT-IR
The FT-IRspectra
spectraofofthe
thePVA/CS/ATH
PVA/CS/ATH pHpH indicative
indicative film,
film, shown
shown inin Figure4d,
Figure 4d,obviously
obviouslycontaincontain
somecharacteristics
some characteristics ofof PVA,
PVA, chitosan
chitosan andand anthocyanin.
anthocyanin. TheThe broadened
broadened peak
peak at at 3262.77
3262.77 cmcm isisa aresult
−1 –1
result
ofofthe
thestretching
stretching ofof
thethe
–OH–OH group.
group. The
The band
band atat 1555.92
1555.92 cmcm isisattributable
−1 −1 attributabletotothe thebending
bendingofofthe the-NH
-NH
bondofofamine,
bond amine,which
whichcan canbebeexplained
explained bybythe
thefact
factthat
thatthe
theamine
aminegroup
group –NH
–NH 2 in
2 in chitosanhas
chitosan hasreacted
reacted
totoform
formthetheamine
amine-NH -NH(II).
(II). The
The bands
bands from
from 1149.08
1149.08 toto 1406.66
1406.66 cmcm stand for the deformation of the
−1 –1stand for the deformation of the

–CH
–CH 2 group
2 group inin
–CH–CH 2OH.
2 OH. InInaddition,
addition,the
the adsorption
adsorption bands
bands from
from 926.46
926.46 to to 1064.36
1064.36 cmcm are typical of
−1 –1are typical of

theoscillation
the oscillationofofthetheC–O–C
C–O–Cgroup.group.
Thesurface
The surfacemorphology
morphologyofofthe thepolymer
polymerfilm filmwas
wasprobed
probedbybySEM SEM(Figure
(Figure5).5).The
Thesurface
surfaceofofthe the
PVA/CS/ATH
PVA/CS/ATH film
film waswas quite
quite smooth.
smooth. NoNo crystal
crystal grains
grains could
could bebedetected
detectedatata amagnification
magnificationofof60,000 60,000
times,suggesting
times, suggestingthatthatthe
themembrane
membranecomponents
components were
were uniformly
uniformly blended.
blended.
Polymers 2019, 11, 1088 7 of 12
Polymers2019,
Polymers 2019,11,
11,1088
1088 77 of
of 12
12

Figure5.
Figure
Figure 5.SEM
5. SEMmicrograph
SEM micrographof
micrograph ofPVA/CS/ATH
of PVA/CS/ATHfilm
PVA/CS/ATH filmsynthesized
film synthesizedfrom
synthesized fromthe
from theATH-containing
the ATH-containing
ATH-containing hydrogel
hydrogel
hydrogel with
with
with a
aa volume
volume
volume ratio
ratio
ratio of
ofof PVA/CS
PVA/CS
PVA/CS == 35/65.
35/65.
= 35/65.

3.2. Effect
3.2.
3.2. Effect of
Effect of STPP
of STPP Crosslinking
STPP Crosslinking Agent
Crosslinking Agent on
Agent on Physical
on Physical Properties
Physical Propertiesof
Properties ofpH
of pHIndicative
pH IndicativeFilm
Indicative Film
Film
PVA/CS hydrogels
PVA/CS
PVA/CS hydrogels have
hydrogels have recently
haverecently
recentlybeenbeen
been extensively
extensively
extensively studied
studied
studied forfor
for their
their potential
their applications,
potential
potential applications,
applications, e.g.,
e.g.,
in biomedical
e.g., in biomedical applications and
applications biomaterials
and biomaterials [13,16]. Chitosan
[13,16]. is
Chitosan
in biomedical applications and biomaterials [13,16]. Chitosan is a kind of natural polymer, deriveda kind is of
a natural
kind of polymer,
natural derived
polymer,
from sustainable
derived
from sustainable
from sustainableresources
resources [8]. Owing
resources
[8]. Owing
[8]. Owing to the
to the polysaccharide
to the polysaccharide
polysaccharide nature
nature
nature ofofchitosan,
of chitosan,
chitosan,the the percent
the percent
percent
elongation-at-break
elongation-at-break decreases
decreases significantly
significantly with with
an an increasing
increasing
elongation-at-break decreases significantly with an increasing CS content in hydrogels [13]. CS contentCS content
in hydrogelsin hydrogels
[13]. [13].
Therefore,
Therefore,
addition of addition of
biodegradable biodegradable
and flexible and
PVA flexible
to CS PVA
becomes to CSan becomes
alternative
Therefore, addition of biodegradable and flexible PVA to CS becomes an alternative for the cast films. an alternative
for the cast for the
films. cast films.
Stefani et
Stefani et
al. [14]
Stefani et al.
al. [14]
studied [14] studied
the studied the
PVA/CS/ATHthe PVA/CS/ATH
PVA/CS/ATH film
film as a film asas aa time-temperature
time-temperature time-temperature indicator.
indicator. In indicator. In
their work, In their
their work,
the work, the
composition the
composition
of CS and PVA of inCS and
the PVA
cast filmin the
was cast
not film
varied was but,not varied
instead, but, instead,
maintained
composition of CS and PVA in the cast film was not varied but, instead, maintained constant at 70/30 maintained
constant at constant
70/30 along at 70/30
with a
along
constant withdosea constant
of STPP dose
[14]. of
It STPP
is known[14]. It
that is known
the that
crosslinker the crosslinker
could influence
along with a constant dose of STPP [14]. It is known that the crosslinker could influence the structural could theinfluence
structural the structural
strength of
strength
strength
the hydrogelof the
of the
and hydrogel
hydrogel and the
and
the resultant the resultant
resultant
films. films.
Hence,films. Hence,
Hence,
the effect the effect
the
of STPP effect
dosage of STPP
of STPP dosage
on thedosage on the
on
mechanical theproperties
mechanical
mechanical of
properties
the resultant of the resultant
PVA/CS/ATH PVA/CS/ATH
films was studiedfilms inwas
thisstudied
work in this
properties of the resultant PVA/CS/ATH films was studied in this work (Figure 6a). Similarly, Figure
(Figure work
6a). (Figure
Similarly, 6a). Similarly,
Figure 6b showsFigure
the
6b shows
6b shows
effect of STPP the effect
the effect of
dosage ofonSTPP
STPP dosage on
dosage
the time-dependent on the
the time-dependent
time-dependent
swelling of the cast swelling
swelling
films.of of the cast
the
Notably, castthe
films.
films.
resultsNotably,
Notably,
shownthe the
in
results6shown
results
Figure shown in Figure
in
were obtained Figurefrom
66 were
were obtained with
obtained
experiments fromPVA/CS
from experiments
experimentsat 25/75. with
with PVA/CS
InPVA/CS
general,at at 25/75.
the25/75.
tensile Instrength
In general,and
general, the
the
tensile
the strength
percent and the
elongation-at-breakpercent elongation-at-break
increased with an increased
tensile strength and the percent elongation-at-break increased with an increasing amount of STPP
increasing amountwith an
of increasing
STPP added amount
to the of STPP
hydrogels.
added to
added
Similarly,to the
the hydrogels.
the hydrogels.
swelling Similarly,
Similarly,
ratio of the cast the swelling
the swelling
films ratio
wasratio of the
of the cast
significantly cast films was
films
suppressed waswith significantly
significantly
more STPP suppressed
suppressed
initially
with more
with
presentmore STPP
STPP
in the initially present
initially
hydrogel. present in in the
the hydrogel.
hydrogel.

(a) Tensile
(a) Tensilestrength andand
strength elongation-at-break
elongation-at- (b)Swelling
(b) Swellingindex
index(SI)
(SI)
break40 20 100
100
Elongation (%)
40 Tensile Strength (MPa) 20
Elongation (%)
(MPa)

16 8080
Swelling Index (%)
Swelling Index (%)
(%)(%)

30 Tensile Strength (MPa)

(MPa)

16
30
Elongation

12
strength

6060
Elongation

20 12
strength

20 8 4040
Tensile

8
10
Tensile

4 2020 2%STPP
2% STPP 4%
4% STPP
STPP
10 6%STPP
6% STPP 8%
8% STPP
STPP
4 10%STPP
10% STPP
0 0 00
2 4 6 8 10 00 55 10
10 15
15 20
20
0 0
Volume
2 4Ratio of6 STPP(aq)
8 to 10 Time(mins)
Time (mins)
Volume Ratio of STPP (%)
PVA/CS/ATH Solution to
(aq)
PVA/CS/ATH
Figure 6. Solution
Effect of sodium (%)
tripolyphosphate (STPP) dosage on the tensile strength and the percent
elongation-at-break of the cast films (a) and the swelling index (SI) of the cast films (b).
Figure 6. Effect of sodium tripolyphosphate (STPP) dosage on the tensile strength and the percent
elongation-at-break
Figure of the cast
6. Effect of sodium films (a) and the
tripolyphosphate swelling
(STPP) index
dosage on(SI)
theof the cast
tensile films (b).
strength and the percent
elongation-at-break of the cast films (a) and the swelling index (SI) of the cast films (b).
Polymers 2019, 11, 1088 8 of 12

The addition of the cross-linking agent STPP to the hydrogels is required in casting PVA/CS/ATH
films. In practice, films cast without STPP added to the hydrogels were easily smashed into species
when they were peeled out from petri dishes after solvent vaporization. This could be probably
attributable to very weak cross-linking bonds formed in the cast films. Thus, cast films without STPP
added could not be examined with the tensile testing instrument.
With the amount of STPP added increasing from 2% to 10%, the tensile strength is enhanced
from 4.9 MPa to 29.4 MPa, i.e., a sextuple increment in tensile strength, and the elongation-at-break
is increased from 10.2% to 17.2%. STPP with a chemical formula of Na5 P3 O10 is a penta-anion,
and can possibly create electrostatic bonds with some polycations, such as chitosan [23]. As such,
STPP could enhance the molecular entanglement among PVA and chitosan entrapped with ATH
molecules, thus leading to a more-difficult-to-break film that possesses increasing tensile strength.
On the other hand, with more STPP they tend to form more tightly integrated blocks with larger
structures of polymer complexes resulting in a more plastic-like nature of the synthesized films and
an increased elongation-at-break of the cast films. In brief, the addition of the crosslinking agent will
increase the entanglement of chitosan and PVA, namely leading to larger tensile strength and more
elongation-at-break [14].
The swelling index (SI) could be used as an indicator for the water absorptivity of the pH indicative
film. In general, a greater extent of cross-linkage in pH indicative films would improve the mechanical
properties and the water resistance of the films. Therefore, it is expected that the cast films would
absorb less water with more STPP initially introduced into the CS/PVS/ATH hydrogels. For example,
the resultant film with 2% STPP was shown to have the largest water absorptivity as its SI increased to
94% in 2 min. Afterwards, the film was broken down into small pieces. Therefore, it is impossible to
measure the weight of the water-adsorbed films submersed in water for more than 3 min. Increasing
the dosage of STPP to 10%, the water absorptivity of the film could decrease significantly, while its
SI was increased from 30% to 43.5% after being immersed in water from 0.5 min to 20 min. After a
20-min duration of submersion in water, this pH indicative film remained almost in shape. In brief,
with addition of 4%–10% STPP, the resultant pH indicative films remained intact after a prolonged
duration of submersion, e.g., 20 min. Notably, the synthetic films became less translucent with the
initial STPP dosage more than 6%, possibly owing to the increasing coherent linkage of molecules.
Consequently, STPP dosages of 2% and 6% were chosen for further study on the effect of the volume
ratios of PVA/CS in the hydrogel on the mechanical properties of synthesized pH indicative films.
In general, as the dosage of the crosslinking agent STPP used in PVA/CS/ATH films increased, the
SI values of these films decreased. Alternatively speaking, as the cross-linking grids inside these cast
films get stronger with an increasing STPP dosage, the process of water molecules imbibed into cast
films could become more difficult. Fewer ATH molecules would be leached out from the cast films.
Likewise, when the duration of the submersion of cast films in water increased, the absorption amount
of water and the ATH quantity leached out from cast films increased accordingly.

3.3. Effect of the PVA/CS Ratio on Mechanical Properties of the pH Indicative Film
As aforementioned, the aim of this work is to explore how to use more sustainable components, e.g,
chitosan (CS), in packaging films. Accordingly, the effect of the CS dosage in the PVA/CS mixed solutions
on the resultant pH indicative films was studied. The mechanical properties of the pH indicative films
with 2% and 6% STPP, including the ultimate tensile strength (TS), the elongation-at-break and the
swelling index, are summarized in Figure 7.
Polymers 2019, 11, 1088 9 of 12

Likewise, the tensile strength of pure PVA films is larger than that of pure CS [9,10]. In general, the
differences
Polymers 2019,between
11, 1088 CS films and PVA films in the elongation-at-break and the tensile strength9 of 12
become more obvious with more STPP initially introduced to the PVA/CS hydrogels.

50 50
222% 73.8% Elongation, 6% STPP (%)
Elongation, 2% STPP (%) 45
40 Tensile Strength, 6% STPP (MPa) 40
Tensile Strength, 2% STPP (MPa)

Tensile strength (MPa)

35

Elongation (%)
Tensile strength (TS)
30 30
25
20 20
15
10 10
5
0 0
100/0 50/50 35/65 25/75 0/100
Volume Ratio of PVA/CS

Figure
Figure7.7.Effect
Effectof
of mixed volumeratios
mixed volume ratiosofofPVA/CS
PVA/CSonon
thethe tensile
tensile strength
strength andand percent
percent elongation-at-
elongation-at-break
break
of theofsynthesized
the synthesized pH indicative
pH indicative films.films.

Interestingly,
It would seemthe trends
that filmsincast elongation
from PVA and
onlytensile strength
possessed of pHelongation-at-break,
a larger indicative films arenamely not so clear
222%
with the CSwith
and 73.8% doses in PVA/CS
initial hydrogels
STPP dosages at 6% (Figure
and 2%, 7). respectively,
Generally, the in elongation-at-break
contrast to 13.9% anddecreases 4.62% forwithfilms
an increasing CS amount in the PVA/CS films. In contrast, the tensile strength of PVA/CS films was
cast from chitosan only. Chitosan is a linear polysaccharide and, specifically, a random copolymer of
lower than thoseand
d-glucosamine of pure PVA films. With 2%while
N-acetyl-d-glucosamine, STPP, PVA theistensile
often an strength
atacticof PVA/CSFrom
polymer. filmsa cast from
molecular
mixed hydrogels with PVA/CS mixed ratios at 50/50, 35/65 and 25/75 were found at 5.18 MPa, 5.54
point of view, the rigid 6-membered ring structures of d-glucosamine and N-acetyl-d-glucosamine in
MPa and 4.94 MPa, respectively. That is, the film cast from PVA/CS at 35/65 possesses the relatively
the main chain make chitosan more difficult to rotate and less flexible than linear PVA [16,29]. Hence,
highest tensile strength.
the elongation of pure CS This matbecomes more to
is expected obvious
be lessin thanthe that
tensile strength
of the PVA mat.of PVA/CSLikewise,filmsthewith 6%
tensile
STPP. At a PVA/CS mixed ratio at 35/65, the tensile strength was measured as 29.9 MPa, compared
strength of pure PVA films is larger than that of pure CS [9,10]. In general, the differences between CS
to 21.2 MPa and 17.1 MPa for those films with a PVA/CS ratio at 50/50 and 25/75.
films and PVA films in the elongation-at-break and the tensile strength become more obvious with
more AsSTPP
shown in Figure
initially 6b, filmstocast
introduced the with
PVA/CS 2% hydrogels.
STPP swelled quickly. Hence, only films cast with 6%
STPP proceeded to the swelling test. The absorption
Interestingly, the trends in elongation and tensile amount
strength of ofwater by pH indicative
pH indicative films arefilms varied
not so clear
and
withwas the dependent
CS doses inon the mixed
PVA/CS volume
hydrogels ratios7).
(Figure of Generally,
PVA/CS inthe theelongation-at-break
films. In general, the adsorption
decreases with
kinetics of water by these films grows exponentially with more submersion time. Moreover, was
an increasing CS amount in the PVA/CS films. In contrast, the tensile strength of PVA/CS films an
increasing
lower thanCS content
those of purein indicative
PVA films. filmWithresults in a smaller
2% STPP, the tensileswelling indexofatPVA/CS
strength the same submersion
films cast from
time
mixed (Figure 8). For
hydrogels with example,
PVA/CSmore mixed water
ratios was imbibed
at 50/50, 35/65 intoand the25/75
PVAwere film found
than others,
at 5.18 as reflected
MPa, 5.54 MPain
the relatively larger SI values of PVA films ranging from 61.5% to 123.7%. This can be attributed to
and 4.94 MPa, respectively. That is, the film cast from PVA/CS at 35/65 possesses the relatively highest
the hydrophilic
tensile strength.nature of PVA more
This becomes polymer.obviousThe in lowest SI values
the tensile wereoffound
strength PVA/CS around
films 34.8%
with 6% to STPP.
47.7%At ona
the films containing
PVA/CS mixed ratiochitosan
at 35/65,only. The SI values
the tensile strength of was
PVA/CS indicative
measured films
as 29.9 are generally
MPa, comparedlarger to 21.2when
MPa
more PVA is present in the mixed hydrogels of PVA/CS. For example, the SI values of PVA/CS film
and 17.1 MPa for those films with a PVA/CS ratio at 50/50 and 25/75.
with aAs mixed
shown PVA/CS
in Figure ratio
6b,offilms25/75 forwith
cast hydrogel
2% STPP increase
swelled from 43% at
quickly. 0.5 min
Hence, only to films
62.3%castat 20
withmin,
6%
while the SI values of films cast from the hydrogel (PVA/CS = 35/65) increase from 50.1% at 0.5 min
STPP proceeded to the swelling test. The absorption amount of water by pH indicative films varied and
to
was69.2% at 20 min.
dependent on the That is, the
mixed waterratios
volume absorptivity
of PVA/CS of PVA/CS
in the films. filmsInincreases
general, the with increasing
adsorption PVA
kinetics
content
of waterinby hydrogels.
these filmsGenerally, the as-synthesized
grows exponentially with more PVA/CSsubmersionfilms could absorb water
time. Moreover, quickly and
an increasing CS
possessed good tensile strength. Taking the mechanical properties into account, the film cast from
content in indicative film results in a smaller swelling index at the same submersion time (Figure 8).
the mixed hydrogel of PVA/CS = 35/65 is deemed as suitable for a pH indicative food-packaging
For example, more water was imbibed into the PVA film than others, as reflected in the relatively larger
application
SI values ofsince PVAthefilms food quality
ranging could
from be visually
61.5% to 123.7%. observed
This can by betheattributed
color change to theof the PVA/CS films.
hydrophilic nature
of PVA polymer. The lowest SI values were found around 34.8% to 47.7% on the films containing
chitosan only. The SI values of PVA/CS indicative films are generally larger when more PVA is present
in the mixed hydrogels of PVA/CS. For example, the SI values of PVA/CS film with a mixed PVA/CS
ratio of 25/75 for hydrogel increase from 43% at 0.5 min to 62.3% at 20 min, while the SI values of films
cast from the hydrogel (PVA/CS = 35/65) increase from 50.1% at 0.5 min to 69.2% at 20 min. That is, the
water absorptivity of PVA/CS films increases with increasing PVA content in hydrogels. Generally, the
as-synthesized PVA/CS films could absorb water quickly and possessed good tensile strength. Taking
the mechanical properties into account, the film cast from the mixed hydrogel of PVA/CS = 35/65 is
Polymers 2019, 11, 1088 10 of 12

deemed as suitable for a pH indicative food-packaging application since the food quality could be
Polymers
Polymers 2019,
visually2019, 11, 1088
11,
observed 1088 by
the color change of the PVA/CS films. 10 of
10 of 12
12

140
140

120
120

(%)
Index(%)
100
100
SwellingIndex
80
80
Swelling

60
60

40
40
PVA/CS(100/0)
PVA/CS(100/0) PVA/CS (50/50)
PVA/CS (50/50)
20
20 PVA/CS (35/65)
PVA/CS (35/65) PVA/CS (25/75)
PVA/CS (25/75)
PVA/CS (0/100)
PVA/CS (0/100)

00
00 55 10
10 15
15 20
20
Time (min)
Time (min)

Figure
Figure 8.
Figure 8. Effect
8. Effect of
Effectof volume
ofvolume ratios
volumeratios of
ratiosof PVA/CS
ofPVA/CS on the
PVA/CS on
on the swelling
the swelling index
swelling index of
index of the
of the synthesized
the synthesized pH
synthesized pH indicative
pH indicative
indicative
films
films dosed
filmsdosed with
dosedwith 6%
with 6% STPP.
6% STPP.
STPP.

3.4. Application
3.4. Application of
of PVA/CS/ATH
PVA/CS/ATH pH pHIndicative
IndicativeFilm
Filmfor forFood
FoodPackaging
Packaging
3.4. Application of PVA/CS/ATH pH Indicative Film for Food Packaging
The
The cast PVA/CS/ATH
The cast
cast PVA/CS/ATHpH
PVA/CS/ATH pH
pH indicative
indicative
indicative films
films
films
were were
were applied
applied to partially
partially
to partially
applied to
wrapwrap pork slices,
pork belly
wrap pork belly slices,
belly slices,
which
which were
were
which were covered
covered covered with
with with Petri
Petri Petri dishes
dishes dishes and
and and exposed
exposed exposed to
to ambient to ambient
ambient air
air for 12 hair for
for 12
and 12 hh and
24h. and 24h.
The 24h. The
covered The covered
covered
portions by
portions
films are by
portions films are
highlighted
by films areand
highlighted
enclosed and
highlighted enclosed
in the in the
red rectangles
and enclosed in the red
red rectangles
on Figure on Figure
9, whereas
rectangles on Figure 9, whereas
the rest
9, whereas the rest
were exposed
the rest
to
were exposed
air.
were exposed to
The to air.
partially air. The
The partially
wrappedpartially wrapped
meat wrapped meat
was meat was
intentionally was intentionally
intentionally spoiled
spoiled in the spoiled in
ambient in the
the ambient
ambient conditions
conditions to conditions
observe the
to observe
successive the successive
color change ofcolor change
the pH of the
indicative pH
filmsindicative
during the films during
spoilage of the
the spoilage
meat. of the meat.
to observe the successive color change of the pH indicative films during the spoilage of the meat.

(a) Exposed
(a) Exposed to
to ambient
ambient air
air for
for 12
12 h.
h. (b) Exposed
(b) Exposed to
to ambient
ambient air
air for
for 24
24 h.
h.
Figure
Figure
Figure9:9: Color
9.Color change
Colorchange of
changeof PVA/CS/ATH
ofPVA/CS/ATH films
PVA/CS/ATH films in
films in contact
incontact with
contactwith raw
withraw pork
rawpork belly
porkbelly slices
bellyslices exposed
slicesexposed to
exposedto ambient
toambient
ambient
air for 12 h and 24
air for 12 h and 24 h.
air for 12 h and 24 h.
h.

The cast
Thecast
The film
castfilm was
film was initially
wasinitially translucently
initially translucently sea-green,
translucently sea-green, as
sea-green, as shown
as shown
shown in in Figure
in Figure 2. After
Figure 2.
2. After being
Afterbeing exposed
beingexposed
exposedtoto
to
ambient air
ambientair
ambient for
airfor 12
for 12 h,
12 h, the
h, the film
the film wrapping the
film wrapping
wrapping the meat becomes
the meat
meat becomes pink,
becomes pink, indicating
pink, indicating
indicating an an acidic
an acidic condition
acidic condition near
condition near
near
pH 5–6 on the surface of pork slices. The mucilaginous surfaces of the meat slices
pH 5–6 on the surface of pork slices. The mucilaginous surfaces of the meat slices appeared as with appeared as with
dense layers
dense layers ofof bacteria
bacteria and
and fungi.
fungi. The
The funky
funky and
and sour
sour flesh
flesh odor
odor could
could bebe smelled,
smelled, asas the
the meat
meat was
was
contaminated with microorganisms that could decompose proteins and fat into
contaminated with microorganisms that could decompose proteins and fat into polypeptides, amino polypeptides, amino
acids, fatty
acids, fatty acids
acids and
and other
other compounds
compounds with with characteristic
characteristic odors,
odors, such
such as
as hydrogen
hydrogen sulfide,
sulfide, indole,
indole,
Polymers 2019, 11, 1088 11 of 12

pH 5–6 on the surface of pork slices. The mucilaginous surfaces of the meat slices appeared as with
dense layers of bacteria and fungi. The funky and sour flesh odor could be smelled, as the meat was
contaminated with microorganisms that could decompose proteins and fat into polypeptides, amino
acids, fatty acids and other compounds with characteristic odors, such as hydrogen sulfide, indole,
skatole, butyric acid, etc. With further exposure in the ambient conditions for another 8 h, the pork
meat turned dark-brown and softer, while the pH indicative film became yellowish with pale green,
showing it in the slight alkaline range. In brief, the pH indicative film applied to wrap the meat
successfully showed the sequential change in color during the meat spoilage.

4. Conclusions
The pH indicative films of PVA/CS/ATH have been successfully cast from hydrogels by blending
1% PVA solution and 1% CS solution in various volume ratios, doped with ATH as an indicator, and
mixed with STPP as a cross-linker. The tensile strength, the elongation-at-break, and the swelling
indices of the cast films have been measured. These films possess the satisfactory mechanical properties
and exhibit a rapid color change in pH buffers. For example, the film synthesized from the mixed
hydrogels made of 1 wt% PVA solution of 35 mL, 1 wt% CS solution of 65 mL and 0.1 wt% STPP of 6 mL,
was found to have the tensile strength at 29.89 MPa and a percent elongation-at-break near 21.13%.
FT-IR spectra of the cast films confirmed the existence of the ATH in the cast film. The PVA/CS/ATH
pH indicative films were applied to partially wrap slices of pork belly, which were exposed to ambient
air for 12 h and 24 h. These films have successfully shown a series of color changes with regard to
exposure time, corresponding to the observation on food spoilage.

Author Contributions: Experimental work and writing—original draft preparation, T.-V.V.; Experimental set-up,
methodology, and critical review and editing of the manuscript, T.-H.D.; Conceptualization, critical review and
editing of the manuscript, supervision, funding acquisition, B.-H.C.
Funding: This work was also financially supported by the Ministry of Science and Technology of Taiwan (MoST
107-2923-E-006-004-MY3).
Acknowledgments: The authors would like to acknowledge the Vietnam–Germany Technology Center at the Ho
Chi Minh City University of Food Industry for the instrument support as well as Minh Tu, Hoang Nam and Anh
Thu for their assistance in sample preparations.
Conflicts of Interest: The authors declare no conflict of interest.

References
1. Ali, A.; Ahmed, S. Recent Advances in Edible Polymer based Hydrogels as a Sustainable alternative to
Conventional Polymers. J. Agri. Food Chem. 2018, 66, 6940–6967. [CrossRef] [PubMed]
2. Domínguez, R.; Barba, F.J.; Gómez, B.; Putnik, P.; Kovačević, D.B.; Pateiro, M.; Santos, E.M.; Lorenzo, J.M.
Active packaging films with natural antioxidants to be used in meat industry: A review. Food Res. Inter.
2018, 113, 93–101.
3. Szente, L.; Fenyvesi, É. Cyclodextrin-Enabled Polymer Composites for Packaging. Molecules 2018, 23, 1556.
[CrossRef] [PubMed]
4. Youssef, A.M.; El-Sayed, S.M. Bionanocomposites materials for food packaging applications: Concepts and
future outlook. Carbohydr. Polym. 2018, 193, 19–27. [CrossRef] [PubMed]
5. Williams, M.; Hrazdina, G. Anthocyanins as food colorants: Effect of pH on the formation of anthocyanin-rutin
complexes. J. Food Sci. 1979, 44, 66–68. [CrossRef]
6. Prietto, L.; Mirapalhete, T.C.; Pinto, V.Z.; Hoffmann, J.F.; Vanier, N.L.; Lim, L.T.; Dias, A.R.G.; da Rosa
Zavareze, E. pH-sensitive films containing anthocyanins extracted from black bean seed coat and red cabbage.
LWT-Food Sci. Technol. 2017, 80, 492–500. [CrossRef]
7. Halász, K.; Csóka, L. Black chokeberry (Aronia melanocarpa) pomace extract immobilized in chitosan for
colorimetric pH indicator film application. Food Packag. Shelf Life 2018, 16, 185–193. [CrossRef]
8. Yoshida, C.M.; Maciel, V.B.V.; Mendonça, M.E.D.; Franco, T.T. Chitosan biobased and intelligent films:
Monitoring pH variations. LWT-Food Sci. Technol. 2014, 55, 83–89. [CrossRef]
Polymers 2019, 11, 1088 12 of 12

9. Casey, L.S.; Wilson, L.D. Investigation of chitosan-PVA composite films and their adsorption properties.
J. Geosci. Environ. Prot. 2015, 3, 78. [CrossRef]
10. De Souza Costa-Júnior, E.; Pereira, M.M.; Mansur, H.S. Properties and biocompatibility of chitosan films
modified by blending with PVA and chemically crosslinked. J. Mater. Sci. Mater. Med. 2009, 20, 553–561.
[CrossRef]
11. Rogovina, S.Z.; Vikhoreva, G.A. Polysaccharide-based polymer blends: Methods of their production.
Glycoconj. J. 2006, 23, 611. [CrossRef] [PubMed]
12. Bonilla, J.; Fortunati, E.; Atarés, L.; Chiralt, A.; Kenny, J.M. Physical, structural and antimicrobial properties
of poly vinyl alcohol–chitosan biodegradable films. Food Hydrocoll. 2014, 35, 463–470. [CrossRef]
13. Figueroa-Pizano, M.; Vélaz, I.; Peñas, F.; Zavala-Rivera, P.; Rosas-Durazo, A.; Maldonado-Arce, A.;
Martínez-Barbosa, M. Effect of freeze-thawing conditions for preparation of chitosan-poly (vinyl alcohol)
hydrogels and drug release studies. Carbohydr. Polym. 2018, 195, 476–485. [CrossRef] [PubMed]
14. Pereira, V.A., Jr.; de Arruda, I.N.Q.; Stefani, R. Active chitosan/PVA films with anthocyanins from Brassica
oleraceae (Red Cabbage) as time–temperature indicators for application in intelligent food packaging. Food
Hydrocoll. 2015, 43, 180–188. [CrossRef]
15. Pourjavaher, S.; Almasi, H.; Meshkini, S.; Pirsa, S.; Parandi, E. Development of a colorimetric pH indicator
based on bacterial cellulose nanofibers and red cabbage (Brassica oleraceae) extract. Carbohydr. Polym. 2017,
156, 193–201. [CrossRef] [PubMed]
16. Teodorescu, M.; Bercea, M.; Morariu, S. Biomaterials of Poly(vinyl alcohol) and Natural Polymers. Polym.
Rev. 2018, 58, 247–287. [CrossRef]
17. Bento, L.; Silva-Pereira, M.; Chaves, K.; Stefani, R. Development and evaluation of a smart packaging for the
monitoring of ricotta cheese spoilage. MOJ Food Process. Technol. 2015, 1, 3–5.
18. Choi, I.; Lee, J.Y.; Lacroix, M.; Han, J. Intelligent pH indicator film composed of agar/potato starch and
anthocyanin extracts from purple sweet potato. Food Chem. 2017, 218, 122–128. [CrossRef]
19. Silva-Pereira, M.C.; Teixeira, J.A.; Pereira-Júnior, V.A.; Stefani, R. Chitosan/corn starch blend films with
extract from Brassica oleraceae (red cabbage) as a visual indicator of fish deterioration. LWT-Food Sci. Technol.
2015, 61, 258–262. [CrossRef]
20. Wei, Y.C.; Cheng, C.H.; Ho, Y.C.; Tsai, M.L.; Mi, F.L. Active gellan gum/purple sweet potato composite films
capable of monitoring pH variations. Food Hydrocoll. 2017, 69, 491–502. [CrossRef]
21. Zhang, X.; Lu, S.; Chen, X. A visual pH sensing film using natural dyes from Bauhinia blakeana Dunn. Sens.
Actuators B Chem. 2014, 198, 268–273. [CrossRef]
22. Liang, J.; Chen, R. Impact of cross-linking mode on the physical properties of zein/PVA composite films.
Food Packag. Shelf Life 2018, 18, 101–106. [CrossRef]
23. Bhumkar, D.R.; Pokharkar, V.B. Studies on effect of pH on cross-linking of chitosan with sodium
tripolyphosphate: A technical note. AAPS PharmSciTech 2006, 7, E138–E143. [CrossRef] [PubMed]
24. Karki, S.; Kim, H.; Na, S.J.; Shin, D.; Jo, K.; Lee, J. Thin films as an emerging platform for drug delivery. Asian
J. Pharm. Sci. 2016, 11, 559–574. [CrossRef]
25. Fuleki, T.; Francis, F. Quantitative methods for anthocyanins. 1. Extraction and determination of total
anthocyanin in cranberries. J. Food Sci. 1968, 33, 72–77. [CrossRef]
26. Chigurupati, N.; Saiki, L.; Gayser, C., Jr.; Dash, A.K. Evaluation of red cabbage dye as a potential natural
color for pharmaceutical use. Int. J. Pharm. 2002, 241, 293–299. [CrossRef]
27. ASTM International. ASTM D882-18. Standard Test Method for Tensile Properties of Thin Plastic Sheeting; ASTM
International: West Conshohocken, PA, USA, 2018.
28. Choo, K.; Ching, Y.C.; Chuah, C.H.; Julai, S.; Liou, N.-S. Preparation and characterization of polyvinyl
alcohol-chitosan composite films reinforced with cellulose nanofiber. Materials 2016, 9, 644. [CrossRef]
29. Oelschlaeger, C.; Cota Pinto Coelho, M.; Willenbacher, N. Chain flexibility and dynamics of polysaccharide
hyaluronan in entangled solutions: A high frequency rheology and diffusing wave spectroscopy study.
Biomacromolecules 2013, 14, 3689–3696. [CrossRef]

© 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).