Food Hydrocolloids 99 (2020) 105322

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Food Hydrocolloids
journal homepage: www.elsevier.com/locate/foodhyd

Development of biodegradable films based on seaweed polysaccharides and T

Gac pulp (Momordica cochinchinensis), the waste generated from Gac oil
production
Thuy T.B. Trana,b,∗, Paul Roacha, Minh H. Nguyena,c, Penta Pristijonoa, Quan V. Vuonga,∗∗
a
School of Environmental and Life Sciences, The University of Newcastle, Ourimbah, NSW, 2258, Australia
b
Faculty of Food Technology, Nha Trang University, Khanh Hoa, Viet Nam
c
School of Science and Health, Western Sydney University, Penrith, NSW, 2751, Australia

A R T I C LE I N FO A B S T R A C T

Keywords: Gac (Momordica cochinchinensis) is a tropical fruit. It belongs to the Cucurbitaceae family. Gac pulp (or meso-
Gac pulp carp) that accounts for 40–50% of fruit weight is commonly discarded during the processing of Gac fruit.
Sodium alginate However, this by-product is a rich source of nutrients and bioactive compounds, which are potential to produce
Carrageenan edible films. This study aimed to determine the effect of sodium alginate, kappa-carrageenan, Gac pulp and
Glycerol
glycerol on film properties and optimise the formula of this composite film for further applications using a
Edible film
Optimisation
response surface methodology (RSM).
The results showed that sodium alginate, kappa-carrageenan, Gac pulp, and glycerol affected physical and
barrier properties, colour parameters, and mechanical properties of the films. The optimal formulation to gen-
erate a composite film from Gac pulp include sodium alginate 1.03%, kappa-carrageenan 0.65%, Gac pulp 0.4%,
and glycerol 0.85% (w/v), where this film produces high mechanical properties, low water vapour permeability
and acceptable physical properties. This optimised film formulation demonstrates a potential for food applica-
tion.

1. Introduction against moisture transmission (Embuscado & Huber, 2009). Recent

Edible films are an innovative packaging solution in the food in-
dustry. They are commonly produced from major materials including
polysaccharides, proteins and lipids or their composite. Polysaccharides
that are obtained from animal, plant and algal sources can be used to
prepare edible films and coatings (Han, 2014). Polysaccharide based
films often have poor water vapour barrier properties, but they are
efficient oxygen blockers due to their well-order hydrogen bonding
(Bourtoom, 2008), and allow selective permeability for carbon dioxide
and resist lipid migration (Hassan, Chatha, Hussain, Zia, & Akhtar,
2018). Protein based materials, such as corn zein, wheat gluten, soy
protein or casein, collagen, gelatine and egg albumin have been applied
to produce films, which have an excellent barrier for oxygen and carbon
dioxide but poor for moisture barrier (Baldwin, 2007; Dursun & Erkan,
2014; Umaraw & Verma, 2017). Unlike polysaccharides and proteins,
lipids are not biopolymers and have limited film-forming ability
(Elsabee & Abdou, 2013). Lipid based films have excellent barriers
studies have developed edible films based on composite materials to
improve the film properties, while minimising the negative impact of
individual materials (Cazón, Velazquez, Ramírez, & Vázquez, 2017).
Gac (Momordica cochinchinensis), a tropical fruit, belongs to the
Cucurbitaceae family which includes cucumbers, squash, luffa and
bitter melon (Parks, Nguyen, Gale, & Murray, 2013). Peel, pulp (or
mesocarp) and skin are discarded during Gac oil processing as well as
current utilization of Gac fruit. Gac pulp accounts for 40–50% of fruit
weight (Parks et al., 2013). This by-product is a rich source of pectin,
essential oil, and carotenoids (Kubola & Siriamornpun, 2011), thus this
material could be potential for development of edible films. However,
none of previous studies have developed composite films from Gac
pulp. Therefore, this study aimed to determine the effect of sodium
alginate, kappa-carrageenan, Gac pulp and glycerol on film properties
and then optimise the film formulation for further applications using a
response surface methodology (RSM).

∗
Corresponding author. School of Environmental and Life Sciences, University of Newcastle, 10 Chittaway Road, Ourimbah, NSW, 2258, Australia.
∗∗
Corresponding author.
E-mail addresses: thi.b.tran@uon.edu.au (T.T.B. Tran), paul.roach@newcastle.edu.au (P. Roach), minh.nguyen@newcastle.edu.au (M.H. Nguyen),
penta.pristijono@newcastle.edu.au (P. Pristijono), vanquan.vuong@newcastle.edu.au (Q.V. Vuong).

https://doi.org/10.1016/j.foodhyd.2019.105322
Received 12 June 2019; Received in revised form 16 August 2019; Accepted 19 August 2019
Available online 22 August 2019
0268-005X/ © 2019 Elsevier Ltd. All rights reserved.
T.T.B. Tran, et al.
2. Materials and methods
2.1. Materials
Sodium alginate (E410, NaC6H7O6) and kappa-carrageenan (E407,
extracted from Chondrus crispus) were purchased from the Melbourne
Food Ingredient Depot, Brunswick East, Melbourne, Australia. Glycerol
was purchased from Ajax Finechem Pty Ltd. Australia. Mature Gac fruit
was purchased from local market in Nha Trang, Vietnam. The Gac pulp
was then separated from the fruit, sliced and dried using a freeze-dryer
(LyoBeta 35 Telstar Technologies, S.L, Spain) for 24 h. The freeze-dried
Gac pulp was then ground and sieved to obtain particle size of
0.25–0.5 mm and well mixed into one uniform lot.
2.2. Preparation of Gac pulp edible film
The film-forming solution was prepared by a casting process.
Suspension solutions were prepared by dissolving sodium alginate
(0.5–1.5% w/v), kappa-carrageenan (0.5–1% w/v), Gac pulp (0.2–0.8
w/v) in 100 mL deionized water under control heating (65 °C) and
continuous stirring. The range of film-forming materials was selected
based on previous studies (Benavides, Villalobos-Carvajal, & Reyes,
2012; Heydari, Bavandi, & Javadian, 2015) and a series of trial ex-
periments. The film forming solution was cooled to 50 °C and glycerol
(0.5–1.5% w/v) was then added as plasticizers. The suspension solution
was stirred for further 5 min to allow completion of mixing and removal
of air bubbles. All films were prepared by casting 20 g of the suspension
solution in a Petri dish (10 cm in diameter) and then dried in an oven at
30 °C for 24 h. Dried films were peeled off from the petri dishes and
conditioned at 30 °C, relative humidity (RH) 75% for 72 h for further
analysis (Thakur et al., 2017).
The film was formed with seaweed hydrocolloids, including sodium
alginate, kappa-carrageenan; Gac pulp and glycerol (Fig. 1). As can be
visually seen, the films obtained were colourless, smoothly and flexible
with glossy surfaces.
2.3. Characteristics of Gac pulp edible film
2.3.1. Physical properties
2.3.1.1. Thickness. The thickness of Gac pulp edible films was
measured according to Thakur et al. (2017) using a digital micro-
meter (Mitutoyo, Co., Model ID-F125, Japan). Precision of the
instrument was 0.001 mm. Film samples were measured individually
and at least ten different measurements were randomly taken for each
film, including a central point and nine around the perimeter. The film
thickness was a mean value from ten measurements. The film thickness
was also used for further calculation of water vapour permeability of
the film samples.
Fig. 1. Actual image of the Gac edible film.
2
Food Hydrocolloids 99 (2020) 105322
2.3.1.2. Colour. Colour values of edible films were assessed using a
colorimeter (Minolta, Model CR-300, Japan). A white colour plate was
used as a background for measurement of edible films. The lightness
(L), chromaticity parameters ‘a' (redness) and ‘b' (yellowness) were also
recorded (Saberi et al., 2016a,b). Numerical value of ‘L', ‘a' and ‘b'
parameters were employed to calculate the total colour difference (ΔE)
(Eq. (1)), Chroma (Eq. (2)) and Hue angle (Eq. (3)) were calculated
from value of ‘L', ‘a', and ‘b' and used to assess the colour changes due to
different film formulations (Oliveira, Ramos, Brandão, & Silva, 2015)
Δ E= (L∗ − L)2 + (a∗ − a)2 + (b∗ − b)2 (1)
Chroma = a2 + b2 (2)
Hue angle = Degrees (Atan (b/a)) (3)
Where L∗, a∗, b∗ are the standard colour parameter values and L , a , b are
the colour parameter values of the film samples (Saberi et al., 2016a,b).
A white standard colour plate (L∗ = 94.08, a∗ = −0.4, b∗ = 9.83) for
the instrument calibration was used as a control for colour measure-
ment of the films.
2.3.1.3. Opacity. Films opacity was examined according to Thakur
et al. (2016), with minor modification. Rectangular film samples
(10 × 50 mm) was placed individually in a quartz cuvette loaded to a
Cary 50 Bio UV–vis Spectrophotometer (Varian Australia Pty. Ltd.,
Melbourne, VIC Australia). A blank cuvette was measured as a
reference. Measurements were conducted by absorbance at 560 nm.
The value of opacity was inversely proportional to level of
transparency. Opacity if the film samples was calculated using Eq. (4).
O= Abs560 x (4)
Where.
O = Film opacity
Abs560 = Absorbance of the film at 560 nm
x = thickness of the film in mm
2.3.1.4. Moisture content. Moisture content was determined using a
method mentioned in a previous report as described by Alves et al.
(2018). Specifically, film samples were cut into 15 × 40 mm strips, and
then dried at 105 °C for 24 h using a hot-air oven (Labec Laboratory
Equipment, Marrickville, NSW, Australia). Moisture content (MC) was
calculated using the following equation (Eq. (5)):
Mi − Mf
Moisture content (%) = × 100
Mi (5)
Where Mi and Mf were the masses of initial and final dried samples,
respectively. Three replicates were conducted for each sample.
2.3.1.5. Solubility. Solubility of film was measured according to
Farahnaky, Saberi, and Majzoobi (2013). Initial dry weight of the
film specimen (15 × 40 mm) was separately recorded. Then each film
was immersed in a beaker containing 50 mL of distilled water with a
periodical mild agitation for 24 h. Undissolved parts were collected and
dried at 110 °C for 24 h to obtain the final dry weight of the film. The
percentage of the total soluble matter (FS) was determined using the
following equation (Eq. (6)):
(initial dry wt − final dry wt )
FS (%) = × 100
initial dry wt (6)
2.3.2. Barrier properties
2.3.2.1. Water vapour permeability (WVP). Water vapour permeability
was determined using ASTM procedure (ASTM, 2013) according to
T.T.B. Tran, et al.
Thakur et al. (2016), with slight modifications. Permeation cells (cup
containing anhydrous CaCl2 granules with 0% RH) were sealed tightly
by the sample film and placed under controlled RH conditions (NaCl
saturated solution; 75% RH) at 25 °C. Water vapour transport was
determined using the weight gain of the cell at a steady state of transfer.
Changes in weight of individual permeation cells were reported every
2 h. Measurements were made for 24 h and plotted as a function of
time. The slope of individual line was estimated by linear regression
(r2 > 0.99). The water vapour transmission rate was calculated
through the slope of each straight line (g/s) divided by area of the
tested film surface (m2). The WVP was calculated as following equation
(Eq. (7)):
Δm T
WVP (gPa −1s−1m−1) =
A Δt ΔP
Where;
Δm/Δt = weight of moisture gain per time unit (gs−1) and can be
determined by the slope of the straight line
A = area of tested film surface (m2)
T = thickness of the tested film (mm)
ΔP = represents the water vapour pressure difference between in-
side and outside of the film (Pa) (Thakur et al., 2016).
2.3.3. Mechanical properties
Tensile strength (TS), elongation at break (EAB) and elastic modulus
(EM) of the films were determined using a Texture Analyser (LLOYD
Instrument LTD, Fareham, UK) according to Saberi et al. (2016a,b),
with some modifications. Preconditioned film specimens (15 × 40 mm)
were griped between two jaws to measure the maximum load (N) and
extension (mm) curves at break point of the test samples. The mea-
surement was conducted at crosshead speed of 1 mm/s and initial grip
distance 40 mm. For each sample, seven replicates were performed.
2.4. Experimental design and statistical analysis
2.4.1. Response surface methodology (RSM)
Response Surface Methodology (RSM) was used to determine the
optimum formulation of Gac pulp edible film. A Box-Behnken design
with three central points replicates was employed. Twenty-seven edible
film formulations consisting of sodium alginate (X1: 0.5%–1.5%),
kappa-carrageenan (X2: 0.5%–1.0%), Gac pulp (X3: 0.2%–0.8%) and
glycerol (X4: 0.5%–1.5%) were chosen as independent variables. The
optimum levels were selected from preliminary single factor tests. The
design is shown in Table 1. Effect of Gac pulp and seaweed poly-
saccharides with plasticizer (independent variables) on properties of
the film (responses) was employed through the model equations, to
graph 2D contour plots of the responses. The optimum conditions of the
independent variables were estimated using the JMP software.
The experimental data for relation between responses and in-
dependent variables was fitted to the following second order poly-
nomial equation (Eq. (8)) (Saberi et al., 2016a):
k k−1 k k
Y = β0 + ∑ βi Xi + ∑ ∑ βij Xi Xj + ∑ βii Xi2
i=1 i=1 j=2 i=1
Where, Xi are independent variables affecting the responses Y ; and β0 ,
βi , βii , βij are the regression coefficients for intercept, linear, quadratic
and interaction terms, respectively. k represents the number of vari-
ables.
2.4.2. Statistical analysis
Experimental design was performed using JMP software (Version
11, SAS, Cary, NC, USA). The program was used to establish the model
equations, to graph the 2D contour plots of the responses and to predict
the optimum values for the four independent variables. The validity of
Food Hydrocolloids 99 (2020) 105322
the second-order polynomial model was assessed by the lack of fit and
the coefficient of determination (R square). The Student's T-test using
SPSS software (version 20, IBM, Armonk, NY, USA) was applied to
compare the sample means. The differences between the sample means
were chosen at the significance level at p < 0.05.
3. Results and discussion
3.1. Fitting of the models
Based on the Box-Behnken experimental design, experiments were
conducted to produce the films. After analysis of the film properties, the
data were placed in the table accordingly (Table 1). It is important to
(7) check reliability of the models for analysing the interactive effects of
the variables on the responses, and for prediction of the optimal film
formulation. ANOVA analysis was applied to check the adequacy of the
quadratic models and the results are shown in Fig. 2 and Table 2.
The results showed that the values of the coefficient of determina-
tion (R square) were in range of 0.71–0.96, signifying that all the ex-
perimental data can be predictably matched with the actual values
(Fig. 2 and Table 2). Especially, the R square for mechanical properties
(Elastic modulus (EM), Elongation at break (EAB) and Tensile strength
(TS)) closed to 1.0, indicating that the high proportion of fitting be-
tween the predicted values and the actual values. Furthermore, the
predicted residual sum of square (PRESS) for the models showed the
predicted models matched well in the design for all the mechanical
properties, thickness, and solubility and certain colour parameters.
In terms of lack of fit test, the results also demonstrated that the
models fit for all the physical (thickness, moisture content, opacity,
solubility and colour parameters) and mechanical (EM, EAB, and TS)
properties because p-values of lack of fit were non-significant
(p > 0.05) for all tested physical and mechanical properties, con-
firming that predicted models were reliable. In addition, the values of F
for all mechanical properties including EM, EAB, and TS were sig-
nificantly different, further confirming reliability of the models for
prediction and optimisation.
3.2. Development of second order polynomial mathematical models
A second order polynomial mathematical equation (Eq. (8)) was
used to express the experimental data obtained from Box-Behnken de-
sign through applying multiple regression analysis. The relationship
between variables and the responses on the physical and mechanical
properties of edible films with Gac pulp was demonstrated in twelve
following empirical models with coded factors.
Ythickness = 16.84 + 33.53X1 − 36.97X2 + 71.04X3 + 28.44X2 − 1.4X1 X2
− 0.2X1 X3 − 16X1 X 4 + 28X2 X3 + 40.6X2 X 4 − 47X3 X 4 − 0.24X12
− 0.02X22 − 0.21X32 + 0.64X42 (9)
YMC = 0.057 − 0.116X1 − 0.075X2 − 0.007X3 + 0.671X 4 + 0.174X1 X2
− 0.025X1 X3 + 0.323X2 X3 − 0.006X1 X 4 + 0.02X2 X 4 − 0.283X3 X 4
− 0.015X12 − 0.269X22 + 0.019X32 − 0.142X42 (10)
(8)
Yopacity = 4.628 − 4.351X1 − 0.647X2 + 2.876X3 + 0.073X 4 + 1.964X1 X2
− 0.442X1 X3 − 1.258X2 X3 + 0.076X1 X 4 − 0.88X2 X 4 + 1.733X3 X 4
+ 1.099X12 − 0.481X22 − 1.465X32 − 0.41X42 (11)
Ysolubility = 109.049 − 55.428X1 − 159.129X2 − 35.269X3 + 25.929X 4
+ 35.147X1 X2 + 3.519X1 X3 + 24.235X2 X3 − 6.69X1 X 4
+ 10.746X2 X 4 + 11.241X3 X 4 + 11.743X12 + 55.254X22 + 0.726X32
− 5.634X42 (12)
3
T.T.B. Tran, et al. Food Hydrocolloids 99 (2020) 105322

Table 1
Box-Behnken experimental design for formulation of Gac pulp edible film.
Run Independent variables Dependent variables

Responses

X1 (%) X2 (%) X3 (%) X4 (%) T MC O S WVP x 10−10 L E C Hue angle EM EAB TS

1 1.5 0.75 0.8 1 100.9 32.42 2.91 22.19 1.82 95.43 8.06 11.14 94.26 17947.8 21.14 1599.92
2 1.5 0.75 0.5 1.5 103.3 40.81 2.05 30.35 1.92 96.57 3.24 6.47 94.01 7893.46 25.37 1346.16
3 1.5 0.5 0.5 1 91.8 34.38 2.33 25.08 2.79 96 5.37 8.52 95.14 8366.96 21.08 1245.14
4 1.5 0.75 0.5 0.5 85 19.75 2.65 12.02 2.67 95.52 9.94 13.07 95.75 23168.56 17.34 2503.86
5 1.5 1 0.5 1 106.1 28.1 1.75 26.94 3.39 95.38 10.45 13.09 94.96 14745.25 19.33 1590.03
6 1.5 0.75 0.2 1 76.9 34.87 1.47 25.3 1.41 96.46 1.32 4.38 92.92 9097.49 20.6 1233.13
7 1 0.75 0.8 0.5 104 21.85 2.41 13.72 1.23 94.34 14.25 17.24 91.75 26520.29 15.78 1625.06
8 1 1 0.8 1 101.3 29.81 2.48 24.43 2.02 94.44 15.19 18.22 93.58 23890.12 18.58 1688.35
9 1 0.5 0.8 1 91.4 33.3 3.09 29.45 1.99 95.54 8.92 12.05 94.13 11247.45 16.4 943.55
10 1 0.75 0.8 1.5 103.3 38.59 2.32 36.17 1.99 95.2 7.86 12.74 94.66 18105.59 14.88 836.03
11 1 0.75 0.5 1 88.5 35.67 2.43 25.22 1.32 96.22 4.94 8.12 95.07 15100.36 17.13 1225.84
12 1 0.75 0.5 1 87 36.1 2.58 22.98 1.44 95.91 5.75 9.42 94.85 16525.79 18.69 1379.86
13 1 1 0.5 0.5 73.5 17.11 2.61 11.83 0.93 95.86 6.79 9.93 94.31 33736.69 13.46 1667.49
14 1 1 0.5 1.5 105.9 44.46 2.25 38.68 2.14 95.97 4.71 7.84 93.52 12614.54 19.35 1164.66
15 1 0.75 0.5 1 85.6 34.62 2.25 27.46 1.31 96.11 5.44 8.61 94.62 13674.92 15.57 1071.81
16 1 0.5 0.5 1.5 91.9 43.51 2.35 43.26 1.96 95.8 7.1 10.25 94.98 6375.555 18.53 664.16
17 1 0.5 0.5 0.5 79.8 17.14 2.27 21.78 0.83 96.03 7.14 10.01 95.31 18638.49 14.46 1176.59
18 1 0.5 0.2 1 67.6 43.39 1.81 37.03 1.28 96.12 5.16 8.33 92.75 3938.49 19.61 530.08
19 1 0.75 0.2 1.5 91.7 49.79 1.29 27.59 1.77 96.67 1.81 5 93.07 8046.54 20.82 860.07
20 1 0.75 0.2 0.5 64.2 16.04 2.43 11.88 1.1 96.55 2.47 5.66 94.9 25067.35 12.58 1169.66
21 1 1 0.2 1 69.1 30.24 1.58 24.74 1.2 96.54 2.07 5.25 94.44 18922.58 18.77 1521.09
22 0.5 0.75 0.8 1 90.3 37.01 3.95 33.07 1.98 96.01 8.75 10.49 94.04 19771.09 11.94 819.38
23 0.5 1 0.5 1 87.2 27.01 2.36 26.53 1.72 95.73 7.5 10.64 95.17 56612.51 11.35 1259.45
24 0.5 0.75 0.5 0.5 61.8 19.83 3.14 19.9 0.99 96.01 7.4 10.58 95.81 33254.17 10.27 997.49
25 0.5 0.5 0.5 1 72.2 41.98 3.93 42.24 1.46 96.12 7.72 10.9 94.28 10813.18 15.17 603.72
26 0.5 0.75 0.5 1.5 96.1 46.79 2.47 44.92 2.33 96.27 4.53 7.73 93.69 11633.2 12.42 628.51
27 0.5 0.75 0.2 1 61 37.98 2.25 38.29 1.28 96.65 2.02 5.23 93.34 27761.29 8.72 555.07

Independent variables: X1, Sodium alginate (0.5–1.5%); X2, Kappa-carrageenan (0.5–1.0%); X3, Gac pulp powder (0.2–0.8%); X4, glycerol (0.5–1.5%) Responses (Y):
T, thickness (μm); MC, moisture content (%); S, solubility (%); O, Opacity (%); WVP, water vapour permeability (gs−1 m−1 pa−1); L, brightness; ΔE, total colour
difference; C, Chroma; Hue angle; EM, Elastic modulus (N/m); EAB, Elongation at break (mm); TS, tensile strength (N/m).

YWVP = 2.974 − 1.991X1 − 8.071X2 + 1.125X3 + 2.002X 4 + 0.68X1 X2 YE = 26.535 − 5.589X1 − 112.56X2 − 4.814X3 + 6.061X 4 + 10.6X1 X2
− 0.483X1 X3 + 0.367X2 X3 − 2.09X1 X 4 + 0.16X2 X 4 + 0.15X3 X 4 + 0.017X1 X3 + 31.2X2 X3 − 3.83X1 X 4 − 4.08X2 X 4 − 9.55X3 X 4
+ 2.26X12 + 4.94X22 − 0.236X32 + 0.31X42 (13) + 0.768X12 + 26.72X22 + 4.33X22 + 1.24X42 (19)

YEM = 2314.398 − 5634.371X1 + 103844.218X2 − 30439.995X3 YL = 94.327 − 1.118X1 + 7.33X2 + 2.987X3 − 0.955X 4 − 0.46X1 X2
− 31445.049X 4 − 78842.093X1 X2 + 28067.527X1 X3 − 7804.741X2 X3 − 0.65X1 X3 − 5.067X2 X3 + 0.79X1 X 4 + 0.68X2 X 4 + 1.233X3 X 4
+ 6345.866X1 X 4 − 17718.42X2 X 4 + 14343.506X3 X 4 + 15636.997X12 + 0.38X12 − 3.72X22 − 2X32 − 0.3X42 (20)
+ 20221.682X22 + 729.31X32 + 7631.734X42 (14)

3.3. Effects of independent variables on film properties

YEAB = 9.37 + 8.848X1 − 29.899X2 + 13.344X3 + 9.488X 4 + 4.142X1 X2
− 4.457X1 X3 + 10.071X2 X3 + 5.882X1 X 4 + 3.634X2 X 4
− 15.236X3 X 4 − 3.222X12 + 9.849X22 − 1.868X32 − 2.95X42
YTS = −2362.525 + 1856.382X1 + 3031.179X2 + 2994.011X3 + 240.24X 4
− 621.687X1 X2 + 170.792X1 X3 − 820.713X2 X3 − 788.725X1 X 4
+ 19.22X2 X 4 − 799.07X3 X 4 + 34.491X12 − 517.16X22 − 1293.758X32
+ 163.423X42
Ychroma = 28.58 − 3.684X1 − 54.58X2 − 9.525X3 + 4.271X 4 + 9.664X1 X2
+ 2.5X1 X3 + 30.84X2 X3 − 3.75X1 X 4 − 4.66X2 X 4 − 6.4X3 X 4
− 0.44X12 + 23.82X22 + 3.625X32 + 1.7X42
YHue angle = 91.121 + 0.417X1 + 7.814X2 + 9.599X3 − 3.074X 4 − 2.14X1 X2
+ 1.067X1 X3 − 7.467X2 X3 + 0.38X1 X 4 − 0.92X2 X 4 + 7.9X3 X 4
+ 0.196X12 − 0.816X22 − 12.689X32 − 0.608X42
3.3.1. Effect of independent variables on thickness
(15) The results showed that all the four film-forming materials, in-
cluding sodium alginate, kappa-carrageenan, Gac pulp and glycerol had
significant effects on the thickness of the edible film (p < 0.05)
(Table 3). The effects of materials were in the decreasing order gly-
cerol > Gac pulp > sodium alginate > kappa-carrageenan. The re-
sults revealed that thickness of the films increased when increasing
(16) concentration of sodium alginate, kappa-carrageenan, Gac pulp and
glycerol (Fig. 3). These can be explained by the films, which were
casted with the same volume of film-forming solution with various dry
matters, with their thickness depending on quantity of the adding
materials. The results also showed that only interaction between Gac
(17) pulp and glycerol had a negative and significant effect on film thickness
(p < 0.05) (Table 3). Therefore, it can be concluded that all four film-
forming materials directly affected film thickness, and Gac pulp and
glycerol had major influence on the film thickness. The thickness results
were reported to affect the structural properties, including water va-
pour permeability and film opacity (Maran, Sivakumar, Sridhar, and
(18)
Thirugnanasambandham (2013).

4
T.T.B. Tran, et al. Food Hydrocolloids 99 (2020) 105322

Fig. 2. Correlation between predicted and experimental values for thickness (a), moisture content (b), opacity (c), solubility (d), water vapour permeability (e), L (f),
E (g), chroma (h), Hue angle (i), elastic modulus (k), elongation at break (l), and tensile strength (m).

5
T.T.B. Tran, et al. Food Hydrocolloids 99 (2020) 105322

Table 2
Analysis of variance for determination of model fitting.
Parameters T MC O S WVP L ΔE Chroma Hue angle EM EAB TS

2
R 0.936 0.965 0.817 0.946 0.745 0.854 0.906 0.878 0.711 0.88 0.911 0.905
PRESS 1897.166 489.249 10.279 689.255 14.031 7.419 167.553 211.371 39.843 > 210 × 106 187.708 26 × 106
Lack of fit 0.062 0.067 0.143 0.34 0.021 0.174 0.056 0.111 0.071 0.053 0.52 0.398
F ratio of model 12.592 23.592 3.822 14.886 2.505 5.026 8.273 6.157 2.111 6.306 8.767 8.125
p of model F < 0.001 < 0.001 0.013 < 0.001 0.06 0.004 < 0.001 0.002 0.101 0.001 < 0.001 < 0.001

Table 3 moisture content of the films, whereas, increasing glycerol concentra-

Analysis of variance for effects of different factors on the responses. tion resulted in lower moisture content of the films. Positive effect of
P Thickness Moisture Opacity Solubility WVP L
glycerol on film moisture content was in agreement with findings re-
content ported by Fundo, Quintas, and Silva (2011). This could be explained
based on the hydrophilic nature of glycerol, which assisted in the for-
Prob > |t| Prob > |t| Prob > |t| Prob > |t| Prob > |t| Prob > |t| mation of hydrogen bonding with free OH groups (Cerqueira, Souza,
Teixeira, & Vicente, 2012). The results also showed that Gac pulp and
βo < .0001 < .0001 < .0001 < .0001 0.0002 < .0001
β1 0.0002 0.048 0.0032 0.0001 0.0191 0.2367
glycerol had a negative and interactive influence on the film moisture
β2 0.0209 0.0017 0.0647 0.0015 0.4946 0.1668 content (Table 3). This means that moisture content of the films re-
β3 < .0001 0.0571 0.0005 0.6155 0.0814 < .0001 markably increased with higher concentration of glycerol together with
β4 < .0001 < .0001 0.0611 < .0001 0.0164 0.0831 lower amount of Gac pulp.
β12 0.9480 0.1281 0.2309 0.0189 0.7118 0.7346
β13 0.6235 0.7829 0.739 0.7501 0.7524 0.5672 3.3.3. Effect of independent variables on opacity
β23 0.4401 0.0934 0.6364 0.2837 0.9024 0.0407 Transparency of the edible films in this study was determined by
β14 0.1541 0.2894 0.9234 0.3221 0.0387 0.2564 opacity value. Lowest opacity value corresponds to high transparency.
β24 0.0776 0.8541 0.5821 0.4232 0.9245 0.6173
Opacity was relatively crucial property for application of film to coat
β34 0.0200 0.0077 0.2063 0.3183 0.9245 0.2861
food as it affects consumer acceptability (Maran et al., 2013). As can be
β11 0.6826 0.7531 0.1287 0.0582 0.018 0.5205
seen from Table 3, while carrageenan and glycerol did not show any
β22 0.8960 0.1698 0.8613 0.0254 0.1801 0.1312
0.3291 0.8874 0.449 0.9636 0.7991 0.2336
significant impact on opacity of the films, sodium alginate and Gac pulp
β33
β44 0.2878 0.0096 0.55 0.3351 0.8138 0.6108 showed significant effects on film opacity (p < 0.05). Gac pulp had the
greatest impact, followed by sodium alginate and glycerol, and carra-
P ΔE Chrome Hue angle EM EAB TS geenan had the least impact on film opacity. This can be explained by
Prob > |t| Prob > |t| Prob > |t| Prob > |t| Prob > |t| Prob > |t| existence of the oil in Gac pulp that caused a film transparency re-
duction. Atarés and Chiralt (2016) reported that existence of the oil
βo < .0001 < .0001 < .0001 0.0006 < .0001 < .0001
caused a decrease of light transmission. This was possibly due to the
β1 0.9337 0.8599 0.7927 0.0016 < .0001 < .0001
β2 0.3468 0.4366 0.8213 0.0002 0.4747 0.0002
light scattering caused by the film matrix imbedded with oil. As a result,
β3 < .0001 < .0001 0.7116 0.2294 0.6957 0.0372 an increase in opacity of films was obtained together with higher
β4 0.0047 0.0194 0.1656 0.0004 0.0006 0.0002 amount of Gac pulp. The results from Fig. 3 indicated that Gac pulp had
β12 0.1156 0.1954 0.4963 0.0043 0.5586 0.4577 positive impact on film opacity. There was no significant impact be-
β13 0.9975 0.6778 0.6822 0.1598 0.4522 0.8046 tween interaction of the tested variables (p > 0.05).
β23 0.0112 0.0221 0.1676 0.8384 0.3973 0.5546
β14 0.2438 0.308 0.8075 0.5827 0.1132 0.0753 3.3.4. Effect of independent variables on solubility
β24 0.5262 0.5208 0.7681 0.4458 0.6072 0.9815 Water solubility plays an important role in film properties because it
0.0916 0.297 0.0091 0.4586 0.021 0.2595
β34 reveals the film's water affinity, especially films intend to come in
β11 0.7813 0.8877 0.8841 0.1341 0.301 0.9233
contact with high moisture food products (Bourbon et al., 2011). As can
β22 0.0296 0.0746 0.8802 0.6129 0.425 0.7189
be seen from Table 3 and Fig. 3, sodium alginate, kappa-carrageenan,
β33 0.5754 0.6764 0.0047 0.9789 0.8253 0.209
and glycerol significantly affected film solubility (p < 0.05), whereas
β44 0.6554 0.5876 0.6534 0.4481 0.3419 0.6496
Gac pulp did not significantly affect solubility. Sodium alginate, car-
Significantly different at p < 0.05; P: parameter; βo : intercept; β1, β2 , β3 , and β4 : rageenan and Gac pulp had negative effects, while glycerol had positive
linear regression coefficients for sodium alginate, carrageenan, Gac pulp and effects. The Gac pulp was reported to contain oil (Kubola and
glycerol; β12 , β13 , β23 , β14 , β24 , and β34 : regression coefficients for interaction Sirimornpun, 2011) that significantly affects the solubility of edible
between sodium alginate × carrageenan, sodium alginate × Gac pulp, films. These results were in agreement with the studies conducted by
carrageenan × Gac pulp, sodium alginate × glycerol, carrageenan × glycerol, Nur Fatin and Nur Hanani (2017) and Cerqueira et al. (2012) related to
and Gac pulp × glycerol; β11, β22 , β33 and β44 : quadratic regression coefficients the concentration of plant oil. The increase of oil concentration or Gac
for sodium alginate × sodium alginate, carrageenan × carrageenan, Gac pulp amount leads to a significant decrease in water solubility. This was
pulp × Gac pulp, glycerol × glycerol. due to the presence of aliphatic groups in the film when Gac pup
containing oil was added. As a result, the hydrophobic property of Gac
3.3.2. Effect of independent variables on moisture content oil changed the film structure that became less soluble (Nur Fatin and
Results for the independent variables on moisture content are pre- Nur Hanani 2017). Besides, the combination of Gac oil and the seaweed
sented in Table 3 and show that sodium alginate, Gac pulp and glycerol hydrocoloids (sodium alginate and carrageenan) can be seen as an in-
had significant effects on the moisture content of the edible film teraction between the hydroxyl groups of polysaccharide chains and the
(p < 0.05). Gac pulp did not significantly affect moisture content of plant oil components. This interaction declined the availability of the
the film. Effects of independent variables are in decreasing order as polysaccharide-water interactions and caused a decrease in film solu-
follows: glycerol > kappa-carrageenan > sodium alginate > Gac bility (Shojaee-Aliabadi et al., 2014). There was also an interactive
pulp. It can be observed from Fig. 3 that increasing concentration of effect observed between sodium alginate and carrageenan on film so-
kappa-carrageenan, sodium alginate and Gac pulp resulted in higher lubility (p < 0.05). Solubility of the edible films increased with higher

6
T.T.B. Tran, et al. Food Hydrocolloids 99 (2020) 105322

Fig. 3. Interactive effects of factors on film properties.

7
T.T.B. Tran, et al.
glycerol level applied. Particularly, the solubility increased from 20% to
around 40% together with 0.6% and 1.4% respectively in the propor-
tion of glycerol. It was explained by Singh, Chatli, and Sahoo (2015)
that three hydrophilic hydroxyl groups that present in glycerol may
enhance film solubility in water.
3.3.5. Effect of independent variables on water vapour permeability
Water vapour permeability (WVP) is important to understand the
mechanisms of the possible mass transfer through the film surface.
According to Ma, Chang, and Yu (2008), this parameter should be low
to prevent moisture loss during preservation of fresh food products. The
results showed that sodium alginate, glycerol and interaction of sodium
alginate × glycerol had significant effects on the water vapour perme-
ability of the edible films (p < 0.05) (Table 3). Observation showed
that increasing glycerol concentration resulted in increasing WVP va-
lues. According to Maran et al. (2013), addition of glycerol increased
the free volume and chain movements as well as the molecular mobility
of the films, which contributed to the diffusion of water vapour into
films. However, WVP initially reduced with higher sodium alginate
level but later on increased after reaching to a certain minimum at 1.62
(gs−1m−1pa−1) (Fig. 3). Interaction between sodium alginate and
glycerol was significant due to changes in concentration of these in-
gredients that caused fluctuations in film density, which resulted in
films with pores and cracks in their structure, and this facilitated the
water vapour permeability of the films (Wu, Geng, Chang, Yu, & Ma,
2009).
3.3.6. Effect of independent variables on colour
Colour of edible films or coatings play a crucial role in film appli-
cation as it influences food products' appearance and consumer ac-
ceptability. Film colour attributes can be determined by instrumental
colour parameters, including Lightness (L), total colour difference (ΔE),
Chroma, and Hue angle (Ramos et al., 2013). The colour brightness
coordinate ‘L' indicated the whiteness value (Oliveira et al., 2015). This
value ranges from black at 0 to white at 100. The results in Table 3
indicated that only the Gac pulp had significant impact on L value of the
films (< 0.05). In addition, interaction of carrageenan and Gac pulp
also had a significant effect on L value, while sodium alginate, carra-
geenan, and glycerol and their interaction did not show a significant
influence. The results from Fig. 3 showed that increasing concentration
of Gac pulp powder resulted in lower L value. This can be explained by
the yellow in colour of Gac pulp, which was contributed by carotenoids
in the pulp (Chuyen, Nguyen, Roach, Golding, & Parks, 2015). There-
fore, transparency of edible films can be affected by level of Gac pulp
powder.
Total colour difference (ΔE) was established based on colour dif-
ference between the control and the experimental samples. The results
showed that Gac pulp and glycerol significantly affected ΔE of the films
(Table 3), where increasing Gac pulp concentration increased ΔE,
whereas increasing glycerol concentration decreased ΔE of the films
(Fig. 3). In addition, interaction between carrageenan × Gac pulp
showed positive significant effect on ΔE.
Hue angle is the colour description in language (red, yellow, green,
etc.) and Chroma performs the colour intensity (Cardoso et al., 2016).
Both parameters provide details spatial distribution of colour than di-
rect value of chromaticity measurements (Oliveira et al., 2015). The
results showed that all four independent variables, including sodium
alginate, carrageenan, Gac pulp and glycerol had no significant effects
on Hue angle. However, interaction between Gac pulp and glycerol had
a positively significant effect on Hue angle of the films (p < 0.05)
(Table 3). This result could be due to changes in reflection of the light at
the film surface in higher solid concentration, leading to film samples
with high hue angle value (Martins et al., 2012). For Chroma, Gac pulp
and glycerol significantly affected Chroma of the films (p < 0.05)
(Table 3). Gac pulp had significant positive effect, whereas glycerol had
a negative significant impact on Chroma of the films (Fig. 3). These
8
Food Hydrocolloids 99 (2020) 105322
findings were in agreement with previous studies, which also found that
addition of glycerol reduced the Chroma values of the edible films
(Cardoso et al., 2016; Saberi et al., 2016a,b; Veiga-Santos, Suzuki,
Cereda, & Scamparini, 2005).
3.3.7. Effect of independent variables on mechanical properties: elastic
modulus, elongation at break, and tensile strength
Mechanical properties of edible coatings play important roles in
packaging that protects food from external stress and maintains its in-
tegrity during transportation and display (Rao, Kanatt, Chawla, &
Sharma, 2010). Mechanical properties are measured on the basic of
tensile strength (TS), elongation at break (EAB), and elastic modulus
(EM). As can be seen from Table 3, sodium alginate, carrageenan, Gac
pulp and glycerol had a significant influence on TS (p < 0.05), of
which, sodium alginate, Gac pulp and carrageenan had positive effect,
while glycerol had a negative impact on TS of the films. Only interac-
tion between sodium alginate and Gac pulp had a significant impact on
TS of the films.
The results indicated that sodium alginate, carrageenan and glycerol
had a significant influence on EM (p < 0.05), whereas Gac pulp did not
show a significant impact on EM of the films (Table 3). Carrageenan
had positive impact while sodium alginate and glycerol had negative
impact on EM of the films. Only sodium alginate and carrageenan
showed a significant interactive impact on EM of the films. The results
also revealed that only sodium alginate and glycerol had significant
positive influence on EAB of the films.
This study observed that sodium alginate had a significant influence
on mechanical properties including TS, EM and EAB. These findings are
in agreement with previous study and can be explained due to forma-
tion of existing inter-molecular hydrogen bonds that contain NH2 and
OH groups in structure of polymer film-forming materials. This would
improve stability and integrity of the edible films, leads to increases in
TS, EM and EAB (Thakur et al., 2017). This study found that carra-
geenan majorly contributed to improve TS and EM, but not EAB and
these findings are in agreement with data in a previous study (Paula
et al., 2015). This study also revealed that glycerol improved TS and EM
while decreased EAB of the films and the similar findings also supported
by the results reported by Paula et al. (2015).
3.4. Optimisation and validation of the models
Good edible films should have high mechanical properties, low
water vapour permeability and acceptable physical properties as these
enable them to be suitable for coating or using as food packaging ma-
terials. Therefore, this study optimised the film formula from sodium
alginate, carrageenan, Gac pulp and glycerol in order to obtain film
with low water vapour permeability and acceptable physical properties
as well as strong mechanical properties. Based on the prediction pro-
filers of the JMP software, the optimal film formula included sodium
alginate 1.03%, kappa-carrageenan 0.65%, Gac pulp 0.4%, and glycerol
0.85%. Under these conditions, a film with low thickness, low water
vapour permeability, acceptable colour and transparency and good EM,
EAB and TS can be prepared for further applications.
In comparison with previous studies, thickness of the Gac pulp
contained edible films is similar to results reported by Thakur et al.
(2017) using pea starch and chitosan, however produced thinner the
films than that was made of sodium alginate (Norajit, Kim, & Ryu,
2010). This study observed that the films had lower solubility and
permeability values than polysaccharide-based edible coating reported
by Norajit et al. (2010) and Thakur et al. (2017). Thus, the observation
of Gac pulp films describe the better barrier properties. Gac pulp con-
tained edible films had significantly higher in mechanical properties,
tensile strength and elongation at break than the biopolymer films
based on pea starch and chitosan Thakur et al. (2017). Low solubility
and better mechanical and barrier properties are beneficial values of
this study, which meet the important requirements in food applications.
T.T.B. Tran, et al. Food Hydrocolloids 99 (2020) 105322

Table 4 M. A. C., et al. (2011). Physico-chemical characterization of chitosan-based edible

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a
Thickness (mm) 0.078 ± 0.006 0.074 ± 0.002 a Cardoso, G. P., Dutra, M. P., Fontes, P. R., Ramos, A.d. L. S., Gomide, L. A.d. M., & Ramos,
Moisture content (%) 32.66 ± 2.98 a 33.06 ± 1.24 a E. M. (2016). Selection of a chitosan gelatin-based edible coating for color pre-
Opacity (%) 2.34 ± 0.44 a 2.67 ± 0.17 a servation of beef in retail display. Meat Science, 114, 85–94.
Solubility (%) 24.29 ± 3.64 a 26.30 ± 1.67 a Cazón, P., Velazquez, G., Ramírez, J. A., & Vázquez, M. (2017). Polysaccharide-based
WVP (gs−1 m−1 pa−1) 1.21 ± 0.51 a 1.67 ± 0.11 a films and coatings for food packaging: A review. Food Hydrocolloids, 68, 136–148.
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and corn oil on physicochemical properties of polysaccharide films – a comparative
ΔE 4.74 ± 1.76 a 6.27 ± 0.28 a
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Chroma 8.07 ± 1.98 a 11.13 ± 0.19 a
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Hue angle 94.82 ± 0.86 a 95.02 ± 0.33 a (momordica cochinchinensis spreng.): A rich source of bioactive compounds and its
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Elastic modulus 13434.47 ± 6326 14797.94 ± 3040 potential health benefits. International Journal of Food Science and Technology, 50(3),
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physical and mechanical properties, whereas Gac pulp showed sig- Development of model for barrier and optical properties of tapioca starch based
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contributed significant effect on most of the film properties, except for Martins, J. T., Cerqueira, M. A., Bourbon, A. I., Pinheiro, A. C., Souza, B. W. S., & Vicente,
A. A. (2012). Synergistic effects between κ-carrageenan and locust bean gum on
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coating materials and future studies are recommended to apply this film Nur Fatin, N. R., & Nur Hanani, Z. A. (2017). Physicochemical characterization of kappa-
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Conflicts of interest temperature on the quality and bioactive characteristics of dried galega kale (Brassica
oleraceaL. Var. Acephala). Journal of Food Processing and Preservation, 39(6),
The authors declare no conflict of interest. 2485–2496.
Parks, S. E., Nguyen, M. H., Gale, D., & Murray, C. (2013). Assessing the potential for a gac
(Cochinchin gourd) industry in Australia.
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the Ministry of Education and Training, Vietnam and The University of (2013). Effect of whey protein purity and glycerol content upon physical properties of
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