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Evaluation of water diffusion, water vapor permeability coefficients,

physicochemical and antimicrobial properties of thin films of nopal mucilage,
orange essential oil, and orange...

Article in Journal of Food Engineering · December 2023

DOI: 10.1016/j.jfoodeng.2023.111865

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 Journal of Food Engineering 366 (2024) 111865

Contents lists available at ScienceDirect

Journal of Food Engineering

journal homepage: www.elsevier.com/locate/jfoodeng

Evaluation of water diffusion, water vapor permeability coefficients,

physicochemical and antimicrobial properties of thin films of nopal
mucilage, orange essential oil, and orange pectin
Roberta M. Mora-Palma a, Porfirio E. Martinez-Munoz b, *, Margarita Contreras-Padilla c, **,
A. Feregrino-Perez d, Mario E. Rodriguez-Garcia e
a
Maestría en Diseño e Innovación, Facultad de Ingeniería, Universidad Autónoma de Querétaro Centro, Cerro de las Campanas s/n, Las Campanas, CP 76010, Santiago
de Querétaro, Qro., Mexico
b
Posgrado en Ciencia e Ingeniería de Materiales, Centro de Física Aplicada y Tecnología Avanzada, Universidad Nacional Autónoma de México, Campus Juriquilla,
Querétaro, Qro., 76230, Mexico
c
Centro Académico de Innovación y Desarrollo de Productos, Facultad de Ingeniería, Universidad Autónoma de Querétaro, Cerro de las Campanas s/n, Col. Las
Campanas, C.P. 76010, Santiago de Querétaro, Querétaro, Mexico
d
Facultad de Ingeniería, Universidad Autónoma de Querétaro Centro, Cerro de las Campanas s/n, CP 76010, Las Campanas, Santiago de Querétaro, Qro., Mexico
e
Departamento de Nanotecnología, Centro de Física Aplicada y Tecnología Avanzada, Universidad Nacional Autónoma de México, Campus Juriquilla, Querétaro, Qro.,
76230, Mexico

A R T I C L E I N F O A B S T R A C T

Keywords: There is a growing interest in the development of biodegradable, safe materials to extend the shelf life of food
Edible films and replace non-biodegradable plastics. Several materials have been investigated that can be used as edible
Water vapor permeability coatings to extend the shelf life of fruit and vegetable products. In the present work, the design and charac­
Photoacoustic analysis
terization of different films of nopal mucilage, orange pectin, and orange essential oil were investigated for their
Nopal mucilage
Antimicrobial activity
potential use as edible coatings. It was found that the pectin concentration had a significant effect (p < 0.05) on
Orange essential oil the thickness, color, and transparency of the edible films. According to the results, the formulation 0.5%
mucilage-0.25 % pectin-0.25 % essential orange oil was chosen. Infrared spectroscopy showed no changes in the
chemical compounds of the film formulation. The XRD pattern showed that this thin film is amorphous and
contains calcium oxalate of nopal mucilage and oleic acid. In the water vapor permeability tests, the films
containing orange essential oil in their formulation showed a higher effective water diffusion and water
permeability coefficients as compared to the other formulations. In addition, the concentration of orange
essential oil has an antimicrobial effect (p < 0.05) on some bacteria of interest for food safety. This work
demonstrates a new methodology and measurement technique to determine the effective water diffusion and
water permeability coefficients for edible thin films based on the photoacoustic effect.

1. Introduction and Siddiqui, 2018).

Postharvest losses are a major challenge for the food industry. Ac­
cording to the FAO, about one-third of all edible food produced for
human consumption is lost (Gustavsson et al., 2011). Fruits and vege­
tables have intrinsic and extrinsic properties that are associated with
postharvest losses. Intrinsic factors include physiology, size, color, and
ripeness. Mechanical losses, diseases caused by microbes, and lack of
postharvest management, on the other hand, are extrinsic factors (Brasil
As they are not properly managed or disposed of waste generated
during the various stages of agro-industrial production is a major
problem worldwide. Orange processing is a good example of this, as it is
a major sector worldwide that generates a lot of discarded orange peel
(Siles et al., 2016; Mellinas et al., 2020). Another example of losses and
contamination is nopal cladodes (Opuntia ficus indica), which contain
both soluble and insoluble fibers (Rodríguez-Garcia, et al., 2007). Their
overproduction leads to postharvest losses as 60% of them are lost and

* Corresponding author.
** Corresponding author.
E-mail addresses: porfirio_martinez@fata.unam.mx (P.E. Martinez-Munoz), margarita.contrerasp@uaq.mx (M. Contreras-Padilla).

https://doi.org/10.1016/j.jfoodeng.2023.111865
Received 7 July 2023; Received in revised form 22 November 2023; Accepted 29 November 2023
Available online 2 December 2023
0260-8774/© 2023 Elsevier Ltd. All rights reserved.
R.M. Mora-Palma et al. Journal of Food Engineering 366 (2024) 111865

wasted (Sáenz Hernández et al., 2013). 2. Materials and methods

The terms films and coating are sometimes used interchangeably, but
they have different forms of application. Films are discrete, self- 2.1. Mucilage preparation
contained structures that are manufactured and used on the surface of
food. While coatings are applied directly to the food surface by dipping, The mucilage was prepared according to the method proposed by
spraying, or sieving (Otoni et al., 2017). Contreras-Padilla et al. (2016). The nopal cladodes (Opuntia Ficus indica)
The development and application of thin films and coating, whose were harvested when they were 100 days old. The crown and thorns
essential function is to reduce water and gas (e.g., O2, CO2) loss, is an were removed, and the cladodes were cut into cubes and dried at 45 ◦ C
important strategy to extend the shelf life of fruits and vegetables after for 12 h. The dried cladodes cubes were ground in a Pulvex mini-100
harvest. In addition, they prevent the loss of firmness and control the grinder with an outlet restriction of 1 mm. Distilled water was used to
ripening and respiration rate of the food, maintain its quality, improve prepare a 4 L suspension containing 8% of the solid particles of the nopal
the appearance of the product through better color, and even prevent cladodes. This was then centrifuged at 10,000 RPM for 10 min. At 20 ◦ C,
microbial growth (Gutiérrez et al., 2008; Kumar et al., 2016). in a Velocity 14 R bendchtop centrifuge, Metrix brand. Two phases were
Various materials have been used as edible coatings on fruits and obtained (soluble and insoluble fibers). The soluble fraction was
vegetables to extend their shelf life. Al-Tayyar et al. (2020) developed an precipitated with 96 % ethanol in a ratio 1:2 (1 L soluble fraction: 2 L
antimicrobial thin film based on chitosan, poly (vinyl alcohol), and sil­ ethanol). The result of this precipitation was dried at 40 ◦ C for 4 h.
icon dioxide nanoparticles doped with zinc oxide nanoparticles that Finally, the dried mucilage was ground in a Pulvex mini-100 (Pulvex)
exhibited activity against Gram-positive and Gram-negative bacteria. mill with an outlet restriction of 0.5 mm.
Cherry tomatoes were coating with pullulan and ethanolic propolis
extract reduced the number of microorganisms and delayed ripening, 2.2. Oil and pectin extraction
extending its shelf life improving its chemical properties during storage
without affecting its sensory properties (Pobiega et al. (2020). Edible Pectin was extracted from the peels of oranges according to the
films can also provide surface sterility and prevent the loss of other method of Cerón-Salazar and Cardona-Alzate (2011). The hydrolysis of
important ingredients (Embuscado and Huber, 2009). orange peels was carried out with acidified water for 60 min at a con­
The developed antimicrobial films by incorporating essential oils of stant temperature of 80 ◦ C, with a wet peel-to-water ratio of 1:2. The
oregano and thyme into low-density polyethylene (LDPE) as food mixture was cooled for 2 h and filtered to separate the liquid extract
packaging was reported by Valderrama Solano and de Rojas Gante from the peels. The concentrated mixture was precipitated with twice
(2012). They conducted a water vapor transmission rate (WVTR) test the volume of 96% ethanol. The resulting pectin was decanted, pressed,
according to standard method F-1249-06 (ASTM F1249-13, 2013). The and dried for 15 h at 35 ◦ C in a stainless-steel dryer (Super Q, Intertek).
essential oils added to these films had a plasticizing effect and in the case Finally, the pectin was ground in a Pulvex mini-100 mill (Pulvex) with
of oregano, a significant effect on the WVTR. At 1% oil, the WVTR was an outlet constriction of 0.5 mm.
10.2 ± 0.4 and at 4% it increased to 13.2 ± 1.1 (g/m2/day). However,
they did not indicate the effective water coefficient. 2.3. Experimental design
The antibacterial activity and physical properties of chitosan films
added with lysozyme and treated with helium plasma were studied by The experiment in this paper was carried out in two parts (see
Ulbin-Figlewicz et al. (2014). The WVP was determined according to the Table 1): In Part I, we determined the water diffusion and water vapor
ASTM E− 96 (Raheem, 2019) method at 4 ◦ C and 60 % RH. They found permeability coefficients and performed a physicochemical analysis. In
that the WVP of the films was not significantly affected by helium part II, after selecting the thin film with the best properties, the anti­
plasma exposure. The WVP values varied from 1.3 to 1.56 ± 0.22 × microbial analysis was performed by varying the concentration of or­
10− 10 (g/s m Pa). ange essential oil. The experimental design was factorial 2 × 2 x 2; with
Photoacoustic techniques have already been used for various food 3 factors: mucilage with 2 levels, pectin with 2 levels, and orange
applications. Gutiérrez-Cortez et al. (2007) studied water and calcium essential oil with 2 levels, see Table 1, Part I.
diffusion in corn kernels during thermo-alkaline treatment in situ. In
addition, Barrientos-Sotelo et al. (2015) used a simple photoacoustic cell 2.4. Thin film preparation
to determine the permeability coefficient of Capsicum annuum. However,
the use of a simple configuration does not avoid environmental and To prepare the edible film, mucilage, and pectin were rehydrated in
electronic noise. The instrumental noise in the system was reduced by distilled water at 70 ◦ C and kept under constant stirring. The resulting
modulated and differential signals, as is the case with differential pho­ suspension was then cooled to 30 ◦ C. After the addition of orange
toacoustic cells (DPC). Martinez-Munoz et al. (2022) developed a
method based on DCP to evaluate the permeability of thin films. This Table 1
method consists of evaluating the instrumental noise through an elec­ Experimental formulation design for films, Part I and Part II.
tronic calibration of the DPC. This work concludes with a methodology
Treatment Part I Component and concentration (%)
to evaluate permeability by monitoring changes in relative humidity
Mucilage Pectin Orange essential oil
when performing water vapor diffusion.
The aim of this work is to evaluate the physicochemical properties 1 0.25 0.25 0
and microbiological activity of thin films of mucilage, pectin, and or­ 2 0.25 0.5 0
3 0.5 0.25 0
ange essential oil that could be considered as candidates for use as 4 0.5 0.5 0
antimicrobial thin films. In addition, a differential photoacoustic cell 5 0.25 0.25 0.25
was developed to determine the effectiveness of the water diffusion and 6 0.25 0.5 0.25
water vapor permeability coefficients of the films studied. The thin films 7 0.5 0.25 0.25
8 0.5 0.5 0.25
were formulated by varying the concentration of mucilage, pectin, and
orange essential oil. Treatment Part II Component and concentration (%)
Mucilage Pectin Orange essential oil

TA1 0.5 0.25 0

TA2 0.5 0.25 0.25
TA3 0.5 0.25 0.5

2
R.M. Mora-Palma et al.
essential oil and propylene-glycol (0.2%) as plasticizer, all components
(according to the respective treatment in Table 1, Part I) were mixed in a
blender with 300 ml of water in a glass beaker. The resulting suspension
was filtered and allowed to rest for 1 h to remove the bubbles formed.
For film formation, the different treatments were poured into plastic
plates with a diameter of 13 cm (40 ml were added to each plate).
Finally, the suspensions were dried at room temperature for 48 h, which
lead to the formation of the films (Šešlija et al., 2018).
2.5. Edible film characterization
2.5.1. Thickness
Thickness was measured according to the methodology reported by
Imran et al. (2010) at 6 random positions for each film, 4 at the edge and
2 at the center using a Mitutoyo IP 65 (Japan) digital micrometer. The
thickness was expressed in mm and the measurement was carried out in
triplicate.
2.5.2. Morphology
Morphologic analysis of thin film samples was carried out in a Jeol
JSM 6060 LV Scanning Electron Microscope (Japan). The analysis was
performed using high vacuum, 2-kV electron acceleration voltage, and
secondary electron mode. Images were carried out on thin films that
were placed on a sample holder. The mounted samples were sputter-
coated with gold.
2.5.3. X-ray characterization
XRD patterns of a thin film were performed on a Rigaku-Ultima 4
diffractometer. Operating conditions were 35 kV and 15 mA, with a
CuKα radiation wavelength of λ = 0.1540 nm, and the measurements
were made from 5 to 80◦ on a 2θ scale. The pattern was performed with a
step size of 0.01◦ to obtain better intensity and resolution of each one of
the possible diffraction peaks or crystal structures.
2.5.4. Color
The color was evaluated according to the method of Con­
treras-Padilla et al. (2016). The color parameters hue (h◦ ), chroma (C*),
and luminosity (L*) of the different films were determined with the help
of a High-Quality Colorimeter brand colorimeter, model NH310. The
samples were analyzed in triplicate.
2.5.5. Transparency
Transparency was evaluated according to the methodology reported
by González-Sandoval et al. (2019). It was measured using a Genesys
10S spectrophotometer (Thermo Scientific) employing rectangular
samples of 0.5 × 0.4 cm and placed perpendicular to the light path in
cells. In each sample, absorbance was measured at a wavelength of 550
nm. An empty cell was used as a blank. The samples were analyzed in
triplicate.
The transparency of the films was calculated using the following
equation:
T=
A550
(1)
X film-forming suspension was determined by the disk diffusion method
where:
A550 is the absorbance and X is the film thickness (mm).
According to Eq. (1), a high T value indicates a high degree of
transparency.
2.6. Vibrational analysis spectroscopy
FTIR was performed to analyze the vibrational states of mucilage,
pectin, and essential oil, in the different formulations of thin films. A
PerkinElmer Spectrum Two equipment with an attenuated total reflec­
tance accessory with a diamond crystal was used. The spectral range
3
Journal of Food Engineering 366 (2024) 111865
used was 600–4000 cm− 1, for each sample, 36 scans were average with a
spectral resolution of 2 cm− 1 (Contreras-Padilla et al., 2016).
2.7. Water permeability coefficient
A differential photoacoustic system (equipment assembled in the
laboratory) was used to study the effective water diffusion coefficient
and the water permeability coefficient of the thin films, as reported by
Martinez-Munoz et al. (2022). The system consists of a sealed chamber
in which a gas is confined between the sample and the microphone
detector. In this chamber, a modulated laser is used to generate a
pressure change by heating the sample. The heat propagation in the
material is transferred to the gas and produces the fluctuations
mentioned above, which are sound waves. Two chambers are shown in
Fig. 1; the first serves as a reference, in this cell a gold film (500 μm) is
used (Sigma Aldrich 99.99 %, USA). The other chamber is used to
determine the permeability coefficient. The samples were stabilized in a
desiccator at 39% RH for 24 h prior to photoacoustic analysis (PA). A
saline solution was added to the reservoir to achieve a relative humidity
(RH) of 70%.
A time scan was performed for 40 min to determine the permeability
coefficient. The modulated frequency of the laser was 33 Hz. The curve
obtained was fitted using Eq. (2), which determines the photoacoustic
signal (PAS); Martinez-Munoz et al. (2022) described this equation.
[ (
t
)]
S = S0 + ΔS 1 − exp − (2)
τd
l2s
τd = (3)
2D
where S is the PAS amplitude, S0 is the initial signal amplitude, ΔS is the
progression of the PA signal until it reaches the saturation value in the
new environment and τd is the water vapor diffusion time. In Eq. (3), D is
the effective water vapor diffusion coefficient. Finally, to obtain the
water vapor permeability coefficient Eq. (4) was used to determine the
value in mPerm, which is defined as the flux of 1 g of water vapor per
day per m2 per mmHg (Martinez-Munoz et al., 2022).
D
Π= (4)
(RAT0 ls )
where A is the relative molecular mass of water, R is the universal gas
constant, and T0 is the temperature of the ambient temperature.
2.8. Antimicrobial activity of films
In the second characterization phase (Part II), the focus is on anti­
microbial activity. For this reason, the formulation that showed the best
properties in the first characterization phase was used (0.5 mucilage-
0.25 pectin). Table 1, Part II shows the three treatments; whose varia­
tion was the concentration of orange essential oil used.
The film was prepared according to the procedure described in sec­
tion 2.3 (Preparation of the thin film). The antimicrobial activity of the
using the Kirby Bauer technique, with some modifications (Imran et al.,
2010; Torres-Alvarez et al., 2017). The microorganisms used were
bacterial strains of Escherichia coli ATCC 25922 and Salmonella typhi­
murium ATCC 14028.
To active of the bacterial strains, 3 mL of trypticasein soy broth so­
lution was prepared and the broth was sterilized in test tubes. 100 μL of
inoculum was taken from each strain, added to the test tube containing
trypticase-soy broth, vortexed, and incubated at 37 ◦ C for 48 h. To
prepare the antibiogram, 100 μL of each bacterial strain was inoculated
with 7 mL of Müller-Hinton agar, which had previously been sterilized.
This medium mixed with the microorganism was poured into a Petri dish
and allowed to dry for 10 min. A sterile filter paper disk with a diameter
R.M. Mora-Palma et al. Journal of Food Engineering 366 (2024) 111865

Fig. 1. Differential photoacoustic equipment for determining the effective water diffusion and water permeability coefficients, a) schematic diagram of the dif­
ferential photoacoustic cell, b) reservoir lid with gas inlet and outlet and c) differential photoacoustic cells.

of 5 mm was then placed on the Petri dishes and 25 μL of the film-

Table 2
forming suspension was added to the disk. The antibacterial activity
Thickness values, water vapor permeability (WVP) and effective water diffusion
was determined after 48 h of incubation at 37 ◦ C. The diameter of the
coefficients (Deff) for the samples of the different treatments. Different letters
inhibition zones was measured in mm. All samples were analyzed in
indicate statistical difference (p < 0.05).
triplicate.
Treatment Thickness WVP Deff (m2/s)
(μm) (mol⋅kg⋅cm− 2⋅s− 1⋅Pa− 1)
2.9. Statistical analysis
9
T1 56.17 ± 14.80a 2.90 ± 0.04 × 10− 2.45 ± 0.33 ×
10− 8
The data was analyzed using trifactorial and variance (ANOVA) T2 69.67 ± 11.91 1.98 ± 0.36 × 10 − 10
1.67 ± 0.73 ×
analysis at a probability of p< 0.05. Differences between the means ab 10− 9
were determined using the Tukey test. All the measurements were car­ T3 68.75 ± 8.01a 1.78 ± 0.15 × 10− 10
1.50 ± 0.28 ×
ried out in triplicates. The Statgraphics XV software was used. 10− 9
10
T4 92.12 ± 24.0b 1.39 ± 0.05 × 10− 1.18 ± 0.37 ×
10− 9
3. Results and discussion T5 69.53 ± 8.23 ab 3.76 ± 0.28 × 10 − 11
3.18 ± 0.40 ×
10− 10
3.1. Sample thickness T6 66.16 ± 5.71a 2.95 ± 0.45 × 10− 11
2.49 ± 0.16 ×
10− 10
11
T7 62.53 ± 9.53a 2.71 ± 0.08 × 10− 2.29 ± 0.52
In this study, the thickness of the films is given in Table 2. The values
×
10− 10
given are satisfactory as they do not exceed the recommended value of T8 76.62 ± 14.33b 4.33 ± 0.14 × 10 − 12
3.66 ± 0.06 ×
0.3 mm (Embuscado and Huber, 2009). 10− 11
According to the three-factorial analysis (pectin, mucilage, and
essential oil concentration), the pectin concentration had a significant
3.2. SEM morphology
effect (p < 0.05). As the concentration increases, the thickness of the
film also increases. Some studies have shown that the thickness of the
Fig. 2a and b shows the morphology of the thin films of mucilage,
film depends mainly on the type of biopolymers used and their con­
pectin, and essential oil. It is obvious that the films are layered, and their
centration (Imran et al., 2010). Galus and Lenart (2013) studied edible
surfaces are uniform. The morphology of the thin films is due to the fact
films with different concentrations of sodium alginate and pectin. There
that it is a mixture of the components, and these thin films have no pores
was a statistically significant difference between the treatments in terms
or microporosity. However, it is necessary to determine if these struc­
of thickness (p < 0.001). As the concentration in the films increased, the
tures are crystalline or amorphous.
thickness also increased. This was associated with the colloidal prop­
erties of this compound, including thickening, suspension, and interac­
tion between the components. 3.3. XRD thin film characterization

XRD analysis of the thin film provides valuable information about

4
R.M. Mora-Palma et al. Journal of Food Engineering 366 (2024) 111865

Fig. 2. a) and b) Micrographs of the morphology of thin films consisting of mucilage, pectin and essential oil.

the internal order or structural composition. As mentioned above, the

WVP value is called the effective coefficient for isotropic materials,
whereas this parameter must be defined as the effective coefficient for
amorphous materials.
Fig. 3 shows a characteristic XRD pattern of sample treatment 5 (T5),
0.25 mucilage, 0.25 pectin, and 0.25 oral essential oil. According to this
pattern, this thin film is amorphous. The presence of oleic acid coming
from the orange peel (black continuous vertical lines) was identified
with ICDD Card 11–0802, using PDF-4 software (Gates-Rector and
Blanton, 2019).
Calcium oxalate was identified with ICDD Card 20–0231 and shown
with dashed blue lines. The crystalline phase of the oxalate is originated
from the mucilage component as reported by Contreras-Padilla et al.,
2011. This result indicates that the thin films shown in Fig. 2 consist of
continuous but amorphous layers, which means that the water diffusion
and water permeability coefficients must be referred to as effective
values. All tested samples show the same amorphous properties.

3.4. Color

3.4.1. Chroma (C*)

The Analysis of variance, which evaluates the effects of the factors
mucilage, pectin, and concentration of orange essential oil on the
chroma value, shows that the pectin concentration is the only factor that
has a statistically significant effect (p< 0.05). As the pectin concentra­
tion increases, the chroma value also increases, i.e., the color of the film
becomes more intense (Fig. 4a). However, it is important to note that the

Fig. 4. Values of the color parameters of the treated samples. a) Chroma values,
b) Hue values and c) Lightness values, and d) transparency values.

Fig. 3. shows the XRD pattern of sample T5, which consists of 0.25 mucilage,
0.25 pectin, and 0.25 oral essential oil.

5
R.M. Mora-Palma et al.
Chroma values of all tested samples were low (4.38–7.09). This means
that the color of all samples is very pale. This is due to the amorphous
character of the films, as you can see in Fig. 3.
3.4.2. Hue (h◦ )
According to the statistical analysis, there was no difference between
the treatments for the hue value (Fig. 4b). The range of h◦ values was
between 77 and 85, indicating that the hue tone of the films tended to be
slightly yellow.
3.4.3. Lightness (L*)
In the analysis of variance for this variable, there is no significant
difference between the treatments (Fig. 4c), i.e., the concentration of the
components has no influence on the luminosity of the films. The L *
values ranged from 89 to 92, indicating that the films have a high lu­
minosity. The analysis showed that the overall color of the film had no
effect on the appearance of the coated food in the samples. Galus &
Lenart (2013), reported that there were no statistically significant dif­
ferences in the color parameters of sodium alginate and pectin films.
The color of food is a very important sensory attribute and should
therefore be monitored when using technologies such as films and edible
coatings. For this reason, films and coatings should ideally be as color­
less as possible (Galus and Lenart, 2013).
3.5. Transparency
According to the analysis of variance, the pectin concentration factor
is the only one that has a statistically significant influence on the
transparency value (p < 0.05), see Fig. 4d. The higher the pectin con­
centration, the lower the transparency of the film. This is related to the
thickness, because when the film is thinner, the light can partially pass
through it, resulting in higher transparency. However, as the thickness
increases, the material becomes opaque due to lower light transparency
(Fakhouri et al., 2012).
In this case, the films with the highest concentration of pectin
showed less transparency. It would be best to have a coating with higher
transparency so that it does not affect the color or optical appearance of
the coated product (Imran et al., 2010).
3.6. Vibrational analysis
First, it is necessary to determine the molecular nature of orange
essential oil, pectin, and mucilage. The main components of orange oil
are fatty acids, including palmitic acid (26.5–28%), oleic acid (27–30%),
and linoleic acid (34–36.5%) (Reazai et al., 2014). As can be seen in
Fig. 5. IR spectra of samples T1-T8 (a) in the range 4000-2400 cm-1 and (b) in the range 2600-600 cm-1.
6
Journal of Food Engineering 366 (2024) 111865
Fig. 3, oleic acid was found in this figure.
Pectin is mainly composed of galacturonic acid (C6H10O7). Mucilage
is a complex polysaccharide (Cx(H2O)x-1). The band at 3300 cm− 1 is due
to the O–H stretching (ν) of the structure and ambient humidity (Smith,
2011; Silverstein et al., 2005).
In Fig. 4, a shift at 3274 cm− 1 is observed for samples T2 (1:2 ratio of
mucilage to pectin) and T3 (2:1 ratio of mucilage to pectin), which is due
to a different stoichiometric ratio of 1:1 mucilage to pectin. This shift
indicates a change in the chemical environment and type of binding. In
samples T5-T8, which contained orange oil, the O–H band remained at
3300 cm− 1. The changes in intensity observed in all samples are due to
variations in the concentration of the components. For samples T1-T4,
the intensity increases with increasing concentration. Fig. 5 and the IR
analysis show that mucilage contains more O–H molecules than pectin.
In samples T5-T8, the addition of orange oil changes the O–H concen­
tration by altering the chemical environment, so that samples T6 and T7
do not show the same trend as samples T2 and T3, which contain equal
concentrations of mucilage and pectin. An analogous interpretation is
possible for the O–H bending (δ) at 1600 cm− 1. The bands at 2923 and
2888 cm− 1 correspond to asymmetric (νA) and the symmetric (νS)
H–C–H stretching, respectively, which correspond to the mucilage and
pectin bonds.
In samples T5-T8 two new bands appear in the vicinity of H–C–H νA
and νS. These bands are located at 2963 and 2855 cm− 1, which are
associated with H–C–H νA and νS, respectively, due to the inclusion of
the orange oil in the matrix of the films. The band at 1371 cm− 1, present
in all samples, corresponds to H–C–H rocking (ρ) (Smith, 2011; Silver­
stein et al., 2005). The band at 1142 cm− 1 is associated with C–O ν in all
samples. The band at 1070 cm− 1 corresponds to C–C ν and the band at
1016 cm− 1 is due to C–O–C νS) (Smith, 2011; Silverstein et al., 2005).
Samples T5-T8 have two bands that are not present in samples T1-T4,
and that occurred when orange oil is present in the films. In this sense,
the bands at 2963 and 2855 cm− 1 are associated with palmitic acid
present in the orange oil (Galus and Lenart, 2013). The presence of oleic
acid and linoleic acid cannot be detected in the spectra because the
mucilage and pectin molecules which form a new complex system can
screen their vibrational states. The other bands can be seen in all
samples.
As it was showed in Fig. 3, the XRD-ray pattern reveals the presence
of calcium oxalate. In Fig. 5 b the bands 1632, 1316 and 779 cm− 1
corresponds with ν8 (νA (C–O)) and ν5 (νA (C–O)), respectively. These
bands were founded by Shippey (1980) and associated with calcium
oxalate monohydrate.
R.M. Mora-Palma et al.
3.7. Effective water diffusion and water vapor permeability coefficients of
thin films
The effective water vapor coefficient is a kinematic property that
provides valuable information about the diffusion of a particle or
molecule through a thin film. For the design and functionality of a thin
film for food applications, the effective water vapor diffusion and
permeability coefficients are crucial factors. Water protection should be
considered when developing edible thin films for long-term food
preservation.
One of the problems in fitting the effective water vapor and perme­
ability coefficients is the selection of the range in which Eq. (2) can be
applied. This problem is solved by analyzing the behavior of ln[1 − (S −
S0 ) /ΔS] = τtD . This means that Eq. (2) can be adjusted in the range in
which a linear relationship between these parameters arises. In this way,
the experimental data used for the fit guarantee the determination of the
permeability coefficients. The permeability process with PA can be
evaluated with different thin films.
Fig. 6a shows the behavior of ln[1 − (S − S0 ) /ΔS] = τtD , as can be seen
there is an almost linear relationship. The fluctuation of the PA signal
can be attributed to the fact that the thin film consists of different layers.
Fig. 6b shows a characteristic PA amplitude signal as a function of time,
which is used to determine the effective water vapor and permeability
coefficients. The black squares represent the experimental data, and the
solid red line corresponds to the best fit of Eq. (2). The PA character­
ization for determining water vapor diffusion and water permeability
analyzes the amplitude signal as a function of the time it takes for water
to cross the sample. The characteristic exponential regimen is achieved.
For long times, the signal tends to stabilize when the sample is saturated
Fig. 6. Shows the characteristic amplitude of the PA signal as a function of time for sample T4. a) the behavior of ln[1 − (S − S0 ) /ΔS] = t/τD , b) a characteristic PA
amplitude signal, c) the temperature and d) the relative humidity.
7
Journal of Food Engineering 366 (2024) 111865
with water. Fig. 6c and d shows the values of temperature and relative
humidity as a function of time. These values were determined using a
BME280 detector, as previously mentioned. The temperature was
monitored to observe a possible increase in temperature due to radiation
exposure. As shown in Fig. 6c, the temperature was kept constant at
28.9 ◦ C. The permeability coefficient measurement was performed at
this temperature. Since the temperature remained constant, it had no
effect on the sample other than producing the photoacoustic effect. The
same procedure was used for all samples tested. In Fig. 6d, region I
shows the increase to stabilize the RH after the addition of the salt so­
lution. Region II corresponds to the diffusion water process where the
RH is constant, and the parameters can be calculated. This new method
allows the determination of temperature and RH to obtain the effective
water diffusion and water permeability coefficients for the thin films.
Table 2 shows the effective diffusion coefficient (Deff) and the water
vapor permeability coefficient (WVP) for the tested samples. The sam­
ples without oil show the highest Deff and WVP coefficients (samples T1
to T4). The thin film matrix was modified after oil inclusion (samples T5
to T8). The oil content prevents water diffusion, so that these coefficients
decrease by almost one order of magnitude. Vázquez-Luna et al. (2019)
determined the water vapor permeability (WVP) values of starch and
g
glycerol thin films in the range of 1.624 × 10− 10 − 3.033×10− 10 m⋅Pa⋅s
which is higher than the values reported in this work due to the use of
essential oil in the formulation of the films.
3.8. Antimicrobial activity
The addition of the pectin component showed statistically significant
effects on several physicochemical properties evaluated in this study.
R.M. Mora-Palma et al. Journal of Food Engineering 366 (2024) 111865

These observations were taken into account to select the edible coating
formulation with a pectin concentration of 0.25 % and a mucilage
concentration of 0.5% for the evaluation of antimicrobial activity. Fig. 7
shows the interval chart of the inhibition halo of the coating-forming
suspension with orange essential oil for the two bacterial strains of in­
terest. The treatments containing orange essential oil are the ones that
exhibit inhibitory aureole. In contrast, TA1 did not contain orange
essential oil and did not exhibit an inhibitory halo.
The analysis of variance of the inhibitory halo showed that there was
no statistically significant difference between TA2 and TA3 (Fig. 7),
which contained 0.25 and 0.5% orange essential oil in the film-forming
suspension. However, there was a significant difference between the
suspension with orange essential oil and the suspension without orange
essential oil (TA1).
Torres-Alvarez et al. (2017) determined the inhibitory effect of or­
ange essential oil (undiluted) on Salmonella typhimurium. They reported
a diameter of 16 mm, twice as much as in this experiment at a con­
centration of 0.5% (7.5 mm). However, it should be mentioned that, in
this study, the orange essential oil was present as a component of the
edible film/coating, so the low inhibition halo has to do with the diluted
concentration in the film-forming suspension/coating. In another study,
the inhibitory effect of various citrus essential oils against Escherichia coli
and Salmonella typhimurium was determined using the same technique as
in this work (Bozkurt et al., 2017). They found that S. typhimurium was
the most resistant bacterium, as a small zone of inhibition developed in
all tests with different citrus oils and different bacteria. This result is
consistent with the results found at the concentrations used in the film.
The halo that developed in the S. typhimurium test was smaller than the
halo of E. coli.
Pobiega et al. (2020) used a similar test for antimicrobial activity in
the film they developed and found similar results for the diameter of the
inhibition halo of S. typhimurium and E. coli.

4. Conclusions

The thickness, color, and transparency parameters evaluated for the

different edible coating formulations studied showed the best properties
(low thickness and high transparency) for formulations with a lower
concentration of pectin. The XRD pattern showed that these thin films
are amorphous and have inclusions of oleic acid and calcium oxalate.
The concentration of orange essential oil significantly affects the
inhibition of the microorganisms studied at both concentrations used.
Infrared spectroscopy shows no changes in the chemical composition of
the thin film formulation.
A methodology and a measurement procedure were developed to
determine the effective water diffusion coefficient and the water
permeability coefficient of the thin film samples. The effective water
diffusion coefficient and water permeability coefficient decreased with
the addition of orange oil. Therefore, samples T5-T8 have the lowest
effective water diffusion coefficient, a property that an edible thin film
needs to improve durability of the product. For this reason, sample T7
with 0.5% mucilage, 0.25 % pectin, and 0.25% orange essential oil was
selected as the formulation, which has a Deff = 2.29 ± 0.52 × 10− 10
cm2/s. The photoacoustic technique is a suitable and rapid method for
determining the water diffusion and water permeability coefficients in
edible films. By-products (orange peels and nopal cladodes) that have
potential in the post-harvest industry can be used to develop value-
added products, such as edible films and coatings. This is an excellent
alternative to offer quality products to consumers.
Statements and declarations
Funding Declaration: The authors did not receive external support
from any organization for the submitted work.
Fig. 7. Inhibition halo for two bacterial strains as a function of essential oil
content. a) Escherichia coli and b) Salmonella typhimurium. Different letters
indicate a statistical difference (p < 0.05).
CRediT authorship contribution statement
Roberta M. Mora-Palma: Conceptualization, Investigation, Meth­
odology, Writing – original draft. Porfirio E. Martinez-Munoz: Formal
analysis, Investigation, Methodology, Project administration, Visuali­
zation, Writing – original draft, Writing – review & editing. Margarita
Contreras-Padilla: Conceptualization, Formal analysis, Investigation,
Methodology, Project administration, Supervision, Validation, Writing –
original draft, Writing – review & editing. A. Feregrino-Perez: Formal
analysis, Methodology, Validation. Mario E. Rodriguez-Garcia: Formal
analysis, Supervision, Validation, Visualization, Writing – original draft,
Writing – review & editing.
Declaration of competing interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Data availability
Data will be made available on request.
Acknowledgments

Roberta M. Mora Palma and Porfirio Martinez would like to thank

8
R.M. Mora-Palma et al.
CONHACYT Mexico for the financial support of their and Ph. D. studies,
respectively. In addition, authors would like to thank to FONDEC-UAQ
for the financial support of this project. Finally, this work was supported
by Laboratorio Nacional de Caracterización de Materiales at CFATA-
UNAM, Querétaro, México.
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