Bioactive Films Based on Arrowroot Starch, Passion Fruit Peel Pectin, and Rosemary Essential Oil: Structural and Functional Characterization for Food Packaging Applications

Abstract

Biodegradable polymeric films with natural active compounds offer a sustainable approach to improving food safety and self-life. This study developed bioactive films using arrowroot starch (AS) and pectin extract (PE) from yellow passion fruit peel via solution casting. Sucrose (SU) and rosemary essential oil (REO) were incorporated to enhance functional properties. Films were characterized by optical, mechanical, barrier, antioxidant, and antimicrobial proprieties. PE acted as a plasticizer and increased antioxidant potential due to its high phenolic content, confirmed by DPPH and FRAP assays. REO exhibited moderate antimicrobial activity against Staphylococcus aureus and Escherichia coli, mainly attributed to α-pinene. Although water vapor permeability of formulation F2 (0.5% REO, 1.4% SU) doubled compared to control, transparency, color, and tensile strength remained stable. F2 also showed a 5.23% increase in DPPH inhibition. These findings highlight the potential of films developed as active, sustainable packaging materials.

Keywords:
Active packaging; antioxidant and antimicrobial properties; arrowroot starch; passion fruit peel pectin; rosemary essential oil; natural polymers

1. Introduction

Most of the currently produced packaging is composed of non-renewable and non-biodegradable polymers, from petroleum sources. The use of these materials has generated serious environmental problems, mainly due to the long time required for their decomposition, which, combined with improper disposal, results in the accumulation of plastic waste on the planet¹.

In response to these questions, in recent decades, researchers have dedicated themselves to the development of biodegradable packaging materials². In this sense, bioplastics derived from plant polysaccharides, such as starch and pectin, are biodegradable under suitable conditions and can replace conventional polymers, especially in single-use applications such as in the food packaging sector³.

Many studies have used starch to produce continuous matrices due to its gelatinization potential, low cost and abundance in nature⁴. Starch is usually found in the form of semicrystalline granules, consisting of two chemical structures: amylose, composed of linear chains with few branches, and amylopectin, with highly branched chains. The proportion of those molecules depends on the growing conditions. The main raw materials for obtaining starch are corn, potatoes and wheat. However, new sources of starch, such as arrowroot, are being explored, mainly due to the scarcity of traditional starch and competition with the food chain⁵,⁶.

Arrowroot (Maranta arundinacea L.) is a perennial herb native to tropical regions and widely recognized as a valuable source of starch. The starch extracted from its rhizomes has been reported to exhibit an amylose content reaching approximately 35.20%, a comparatively high value among tropical starchy crops⁵,⁶. This relatively high amylose content promotes the formation of more compact structures and enables the production of films with enhanced physical and structural properties compared to those obtained from other starch sources. Despite this, films made exclusively by starch are brittle and not very resistant, so the mixture of starch with other biomass polymers, such as pectin, has been a viable and economical alternative for the optimization of their properties⁷-⁹.

Pectin is formed by galacturonic acids, such as homogalacturonan in larger proportions, as well as rhamnogalacturonans (RG-I and RG-II) and other sugars¹⁰. Agro-industrial residues, such as fruit and vegetable peels, can present significant amounts of pectin, which is widely used as a gelling agent, stabilizer and emulsifier in the food industry¹¹. However, despite these applications, a large part of this waste is still disposed of incorrectly, contributing to environmental pollution, greenhouse gas emissions, and the proliferation of pests¹².

A promising source for pectin extraction is the peel of yellow passion fruit (Passiflora Edulis Flavicarpa), especially in Brazil, which is the world's largest producer of this crop, with a total production of more than 600 thousand tons¹³. The peel, which correspond to 50% to 55% of the fruit, are usually discarded by the pulp processing industry, representing a large amount of untapped biomass and a huge potential resource loss since in addition to being rich in pectin (12–18%), it contains valuable nutrients such as polyphenols (5 mg gallic acid equivalent (GAE)/g dry weight (DW)), carotenoids (5 mg β-carotene/100 g DW) and ascorbic acid (40 mg/100 g DW)¹⁴.

Recently, several studies have investigated the combination of starch and pectin, including blends of potato starch and pectin from apple peel⁷, pea starch and different types of citrus pectin and beet pectin⁸, as well as sweet potato starch and pectin films from lemon peel⁹. Based on microstructural analyses, Sani et al.⁷ observed a smooth, uniform and cavity-free surface for the starch/pectin films (3:1 ratio), suggesting good adhesion between the natural polymers. Bai et al.⁸, also using the 3:1 ratio, reported that the addition of any type of pectin resulted in a reduction of up to 35% of water vapor permeability, due to its hydrophobic fractions. In addition, the tensile strength and Young's modulus of starch have increased by up to 3 times. Dash et al.⁹ identified changes in the viscoelastic behavior of starch after the addition of pectin (3:1) and reported a reduction in peak loss factor (tanδ), which is the ratio of loss modulus (G'') to storage modulus (G'). This result suggests that pectin can act as a reinforcing agent, improving the properties of the films.

On the other hand, incorporating natural compounds, such as essential oils, into polymeric matrices to develop an active packaging emitting antioxidant and/or antimicrobial agent has also recently become a trend¹⁵. These packages are designed to interact with food to extend its shelf life and, consequently, represent an important approach to minimizing food waste, which reached alarming levels in 2022, with more than 1 billion tons discarded, according to the Food Waste Index Report published in 2024 by the United Nations Environment Programme (UNEP)¹⁶. In this context, it is important to highlight that essential oils are classified as GRAS (Generally Recognized as Safe) by the U.S. Food and Drug Administration (FDA), which reinforces their safety for use in food contact materials, supporting their application in active food packaging¹⁷.

Among the various essential oils, rosemary essential oil (REO) stands out for being composed mostly of eucalyptol or 1,8-cineole (40.1%), α-pinene (12.9%) and camphor (12.4%)¹⁸, substances that act by damaging the outer membrane of several pathogenic bacteria and, simultaneously, have antioxidant effects, helping in the preservation of packaged foods¹⁹.

In this context, the hypothesis of this work is that arrowroot starch (AS)/pectin extract films obtained from the peel of yellow passion fruit (PE) added with REO can act as active packaging with the ability to release functional compounds and minimize ecological problems in terms of accumulation of agro-industrial waste and plastics. To improve the mechanical properties of the films, we considered using sucrose (SU) as a plasticizer. Therefore, the aim of the study was to study the effect of the incorporation of REO and SU on the antioxidant capacity and structural, optical, mechanical and water vapor barrier properties of films.

2. Materials and Methods

2.1. Materials

Arrowroot starch (Torres brand, São Paulo Brazil), sucrose (refined crystal sugar of Olho d’água brand, Recife Brazil) and rosemary essential oil (Live Aloe brand, Goiânia Brazil), all commercially obtained. To obtain PE, yellow passion fruit (very firm and ripe - degree of ripeness defined by the yellow color of the peel) and lemon were used, purchased from local stores in the city of Recife/PE - Brazil (8° 04′ 03″ S and 34° 55′ 00″ W).

2.2. Obtaining the pectin extract (PE)

To obtain the PE, the passion fruit was washed and sanitized and then cut in half to remove the pulp (endocarp). After pulp removal, the epicarp was removed and the mesocarp (part of interest) was cut into small pieces. Then, 200 g of the chopped mesocarp was mixed with 500 mL of distilled water and heated to 100 ºC until it softened and became translucent. Subsequently, 20 mL of lemon juice was added still under heat for another 10 min. Then, the solution was left to cool until room temperature (25 °C). The solution was filtered, and the pectin extract (PE) was stored in amber glass under refrigeration (5 °C) until use.

2.3. Film preparation

The films were prepared by the solution casting method from a filmogenic solution of AS (4%, w/w, wet basis) and PE, according to the work of Souza, Silva and Druzian²⁰ with adaptations. The AS concentration refers to its proportion relative to the total mixture. The total mass of each filmogenic solution was 200 g, composed of 8 g of AS and 192 g of the remaining components, including PE, REO, and SU.

Film formulations were defined based on a 2² full factorial design, considering the concentrations of SU (0–1.4%, w/w) and REO (0.5–1.5%, w/w) as the independent variables. These percentages were also calculated relative to the total mass of the filmogenic solution (200 g) and were expressed in grams accordingly. In addition, a control film (F7) was produced without the addition of SU and REO. The values in percentages and grams of each formulation are presented in Table 1.

Table 1
Independent variables and their respective levels used in the factorial design 2² with duplicate at center point.

Formulations	Levels		Composition of the film-forming solution
	REO(%)	Sucrose (%)	REO (%) / (g)	Sucrose (%) / (g)	AS (%) / (g)	PE (%) / (g)
F1	-1	-1	0.50/ 1.00	0.00	4/8.00	95.5/191.00
F2	-1	1	0.50/1.00	1.40/2.80	4/8.00	94.1/188.20
F3	1	-1	1.50/3.00	0.00	4/8.00	94.5/189.00
F4	1	1	1.50/3.00	1.40/2.80	4/8.00	93.1/186.20
F5	0	0	1.00/2.00	0.70/1.40	4/8.00	94.3/188.60
F6	0	0	1.00/2.00	0.70/1.40	4/8.00	94.3/188.60
F7	-----------------------------------------------------------				8.00		192.00

• Rosemary Essential Oil (REO); Arrowroot Starch (AS); Pectin Extract (PE).

To prepare the filmogenic solution, the materials were mixed in a beaker and then mechanically stirred for 5 min at 500 rpm. After homogenization, the solution was heated in a microwave, with intermittent stirring, until it reached a temperature between 72 and 78 ºC for complete starch gelation. The solutions were kept at rest to remove the air bubbles formed during the process, and subsequently approximately 45 g were poured into petri dishes (140 x 15 mm). Drying was carried out in an air circulation oven at 40 °C for a period of 7 h. After being removed from the oven, the films were conditioned at 25 °C and 75% relative humidity (RH) until they were characterized.

2.4. Characterization of the REO and PE

2.4.1. Profile of phenolic compounds

To determine the chemical composition of the REO, a Trace 1330 gas chromatograph was used, coupled to an ISQ Single Quadrupole mass spectrophotometer. The separation of volatile compounds was performed by means of a DB5-MS capillary column of TGM fused silica, using helium as the carrier gas. The injector temperature was set to 270 °C, and the thermal programming followed the following profile: 60 °C for 3 min, heating at 10 °C/min up to 240 °C, and 300 °C maintained for 4 min.

2.4.2. Antimicrobial activity

The antimicrobial activity was determined by the disk-diffusion technique according to the methodology described by Silva et al.²¹ against three bacterial species: Staphylococcus Aureus, Escherichia coli and Enterobacter Aerogenes. The culture medium (Nutritive Agar) was prepared in a sterile Petri dish and after its solidification. A 0.1 mL aliquot of the bacterial suspension (visually standardized according to the 0.5 standard of the McFarland scale) was spread on its surface. The filter paper discs, impregnated with the sample, were aseptically transferred to the surface of the inoculated culture medium. The plates were incubated at 35 °C for 48 h, and after this period, the diameter of the inhibition halo formed was measured.

2.4.3. Total phenolic compounds content

The total phenolic compounds content (TPC) was determined by the spectrophotometric method using the Folin-Ciocalteau reagent, according to the methodology of Singleton and Rossi²². A total of 90 μL of sample and 60 μL of Folin-Ciocalteu reagent were mixed in a test tube and stirred for 60 s; then 2700 μL of distilled water and 180 μL of sodium carbonate solution were added and stirred for another 30 s. After 2 h of reaction time, in the absence of light and rest, the reading was performed in a spectrophotometer (Edutec) at 760 nm. The results were calculated using a standard curve of gallic acid (y = 0.092x + 0.122, R² 0.986), at concentrations from 1 to 20 mg/L. The test was performed in triplicate and the results were expressed in mg of GAE (gallic acid equivalent) per g of sample.

2.4.4. Antioxidant activity

The antioxidant activity of REO and PE was determined by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical capture method following the methodology proposed by Brand-Williams, Cuvelier and Berset²³ by the iron reduction capacity (FRAP) according to the methodology of Rufino et al.²⁴.

Antioxidant Activity by the DPPH radical scavenging capacity method: The reaction mixture was performed by the addition of 2700 μL of the ethanolic solution of DPPH, 90 μL of sample and 210 μL of ethanol, in triplicate. The absorbances were read after 30 min of reaction at 517 nm in a UV-visible spectrophotometer of the Edutec brand. The ethanolic solution of DPPH was used as a control and the DPPH radical capture activity was expressed as the percentage of inhibition (% of Inhibition).

Antioxidant Activity by the Iron Reduction Method (FRAP): The FRAP reagent was prepared by mixing 25 mL of 0.3 M acetate buffer with 2.5 mL of TPTZ solution (2,4,6-tri(2-pyridyl-1,3,5-triazine) at 10 mM and 2.5 mL of aqueous solution of ferric chloride at 20 mM. Then, 90 μL of the sample, 270 μL of distilled water and 2700 μL of the FRAP reagent were added in a test tube. The mixture was homogenized and incubated at 37 °C in a water bath for 30 min. The absorbance reading was measured at 595 nm in a UV-visible spectrophotometer (Edutec). The results were obtained from a standard curve of ferrous sulfate (y = 0.0006x - 0.0294, R² 0.9979), at concentrations of 500 to 2000 μM, and expressed in mmol of ferrous sulfate (Fe₂SO₄) per g of sample.

2.5. Characterization of films

The films were evaluated by investigating the antioxidant capacity, optical (color and transparency), physical (Fourier transform infrared spectroscopic analysis, humidity), mechanical (tensile strength and elongation) and barrier (water vapor permeability) properties.

2.5.1. In-vitro antioxidant activity

The in vitro antioxidant activity of the films was carried out according to the methodology of Lima et al.²⁵ with adaptations. Film samples were cut into 9 cm² square parts, which were immersed in 10 mL of distilled water and centrifuged at 4400 rpm for 30 minutes at 5 ºC. From the supernatant, aliquots were removed to determine the antioxidant activity by the DPPH and FRAP methods, following the same procedure used to determine the antioxidant activity of the REO and PE described in item 2.4.4.

2.5.2. Fourier Transform Infrared Spectroscopy (FTIR) and principal component analysis (PCA)

FTIR spectra were acquired in a SHIMADZU IR TRACER 100 spectrometer in the region of 4000-400 cm^-1 according to the methodology of Medina Jaramillo et al.²⁶. The films were placed directly on the reading surface. Forty-five scans were performed at a resolution of 4 cm^-1. The measurements were performed at 5 different points of the film for mathematical treatment applying PCA. Unscrambler 9.7 software was used for data processing. For comparison purposes, the FTIR analysis of the REO was performed with a horizontal attenuated total reflectance (ATR) sensor and the same parameters described above.

2.5.3. Optical properties (color and transparency)

The visual aspect of the films was measured through color variation and transparency. The transmittance of the films at the wavelength of 600 nm was determined using a UV-visible spectrophotometer. The test was performed in triplicate and the transparency obtained through Equation 1²⁷:

$Transparency=\frac{\log \left(\%T\right)}{b}$ (1)

Where, %T, is the percentage of transmittance and b, is the thickness of the film (mm).

The determination of the color of the films was carried out following ASTM E308-17²⁸. The analysis was performed in a GRETAG MACBETH - COLOR-EYE 2180 colorimeter, using the CIELAB Ttan D65 system, a reading angle of 10° and a viewing area of one square inch (0.00064516 m²). The color parameters, L* (luminosity), a* (red and green) and b* (yellow and blue) were obtained at 6 random points of each film and the total difference of color (ΔE) in relation to the standard white plate (L* = 95.83; a* = -0.22; b* = 2.35) was calculated by Equation 2:

$\Delta \text{E}={\left[{\left(\Delta {\text{L}}^{*}\right)}^2+{\left(\Delta {\text{a}}^{*}\right)}^2+{\left(\Delta {\text{b}}^{*}\right)}^2\right]}^{1/2}$ (2)

2.5.4. Moisture and water vapor permeability (WVP)

To determine the moisture content, 2 cm diameter disc film samples were weighed and then dried in an air circulation oven at 105 °C until constant weight. The results were obtained by weight difference by gravimetry.

The water vapor permeability (WVP) of the films was determined gravimetrically following the specifications of the ASTM - E96-9 standard²⁹. The samples, in triplicate, were properly arranged in permeation cells filled with calcium chloride (CaCl₂) as desiccant. Subsequently, these systems were stored in desiccator containing a saturated sodium chloride (NaCl) solution (75% relative humidity). Mass gain was monitored every 12 h for a period of 8 days, and WVP was calculated according to Equation 33.

$WVP=\frac{\left(\frac{\text{Ci}}{A}\right).\text{X}}{\text{Ps}.\left(\text{RH}1-\text{RH}2\right)}$ (3)

Where WVP is water vapor permeability, Ci is slope of the line generated by the weight loss of the system as a function of time, A is the exposed surface area of the film (m²), X is the average film thickness, Ps is the saturation pressure of the water, RH1 is the relative humidity inside the desiccator and RH2 is the relative humidity inside the container.

2.5.5. Mechanical properties

The tensile mechanical properties of the films were evaluated according to the specifications of ASTM D882-12³⁰ using a SHIMADZU static testing instrument – AGS Line - X 10 Kn with initial separation of the grips of 50 mm and rate of 12 mm/min. For the test, the films were cut into rectangular strips with a dimension of 100 x 15 mm and the thickness was measured at 5 random points for each sample (the mean values of the thicknesses ranged from 0.12 mm to 0.19 mm). Tensile strength (TS) (MPa) was determined by the ratio between the maximum measured tractive force (σ) and the initial cross-sectional area of the specimen (Ao). The percentage elongation (ε) at break (%) was determined by the ratio between the distance traveled in the displacement of the grips to the break [final length (Lf) − initial length (Lo)] and initial length of the specimen (Lo).

2.6. Statistical analysis

The results were expressed in mean (n = 3) ± standard deviation or accompanied by their respective 95% confidence intervals (IC95%), calculated from the error mean square (EMS) of the Analysis of Variance (ANOVA). Duncan's statistical test was applied to determine the difference in the significance level of 5% (p < 0.05) using ASSISTAT software version 7.7.

3. Results and Discussion

3.1. Characterization of the rosemary essential oil (REO)

Based on the chromatogram analysis, the main components of REO were identified and are presented in Table 2. It was found that the REO consists mainly of monoterpenes and monoterpenoids. The majority constituents were α-Pinene (26.53%), Camphor (20.32%) and 1.8 Cineole (19.36%), which represent 63.21% of the REO. In agreement, Micić et al.³¹ evaluated REO of different origins, which also showed a predominance of these constituents.

Table 2
Chemical composition of Rosemary Essential Oil (REO).

Compound	Relative Percentage (%)
Monoterpenes
á-Pinene	26.53
Camphene	9.04
D-Limonene	3.64
o-Cymene	2.61
Monoterpenoids
Camphor	20.32
1.8 Cineol	19.36
Endo-Borneol	4.60
á-Terpineol	2.55
Sesquiterpenes
Caryophyllene	2.21
Others	9.14

The chromatographic profile is one of the most important characteristics of EO's, as their biological activities are directly linked to the chemical structure of their components and the proportions in which these compounds are present. These, in turn, can vary according to the environmental and agronomic conditions of the region where the plant was harvested, the stage of development of the plant, the season of the year in which the collection was carried out, in addition to the method and time of extraction³². To understand the correlation between chemical composition and antimicrobial and antioxidant activities, the diameters of the inhibition halos against the bacteria tested, as well as the total phenolic compounds content (TPC) and antioxidant activity by the DPPH and FRAP methods, were obtained and can be observed in Figure 1 and Table 3.

Figure 1
Halos of inhibition observed for Rosemary Essential Oil (REO). Diameter of inhibition halo in mm (mean±SD).

Table 3
Total phenolic content and antioxidant activity of Rosemary Essential Oil (REO) and Pectin Extract (PE).

Sample	TPC (mg of GAE/g of sample)	DPPH (% inhibition)	FRAP (mmol Fe2SO4/g of sample)
REO	33.12±1.53	38.11±0.62	6.24±0.23
PE	230.09± 29.08	70.83± 1.82	53.20± 1.95

• Values expressed in mean (n = 3) ± standard deviation.

According to the National Committee for Clinical Laboratory Standards protocol M02-A10³³, REO was found to exhibit intermediate antimicrobial activity against S. aureus and E. coli, with inhibition halos between 15 and 19 mm. On the other hand, low activity was observed against E. aerogenes strains, with inhibition halos equal to or less than 14 mm. The antimicrobial efficacy of REO is mainly attributed to the terpene compounds present in its composition, such as α-pinene, which causes structural and functional changes in the outer membrane of these pathogens³⁴.

Although the mechanism of antibacterial action of essential oils (EOs) is not yet fully elucidated in the literature, it is known that hydrophobicity facilitates penetration into the bacterial cytoplasmic membrane, promoting its destructuring. Thus, the outer membrane that surrounds the cell wall of gram-negative bacteria restricts the permeability of EOs, making their action less efficient. However, in this study, the REO presented similar inhibition halos for S. aureus (gram-positive) and E. coli (gram-negative). This result suggests that REO is a promising candidate as an antimicrobial agent in active packaging, since these strains, in addition to promoting food spoilage, are among the main causes of foodborne diseases.

In the TPC analysis, REO presented 33.1 mg GAE/g sample, in accordance with the results reported by Baydar et al.³⁵. However, compared to other EOs, such as turmeric and thyme, REO contains a lower content of phenolic compounds³⁶,³⁷. According to the GC/MS quantification, the main chemical constituents of REO (α-pinene, camphor and 1,8-cineole) do not have phenolic structures, which contributes to a moderate antioxidant activity with a percentage of DPPH radical inhibition of 38.1% and its iron reduction power of 6.24 mmol Fe₂SO₄/g, especially when compared to the synthetic antioxidant butylated hydroxytoluene (BHT), which exhibits 51.6% DPPH inhibition and an iron-reducing power of 10.58 mmol Fe₂SO₄/g³⁸.

3.2. Characterization of the pectin extract (PE)

As shown in Table 3, PE showed a high TPC value and good antioxidant activity by the two methods used, presenting a higher action than the synthetic antioxidant BHT. This result can be associated with the fact that the peel of the yellow passion fruit has phenolic compounds such as isoorientin³⁹ in its composition. Isoorientin, also known as homoorientin, is a C-glucosyl flavone that shows greater scavenging capacity of free radical species than well-known antioxidants like ascorbic acid and quercetin⁴⁰. Data from the study by Zeraik et al.³⁹ suggest a direct correlation between antioxidant capacity and isoorientin content in passion fruit peels.

Phenols exhibit antioxidant activity due to their ability to release hydrogen molecules to neutralize free radicals, and due to their ability to chelate transition metals, such as iron, which are catalysts for the formation of free radicals⁴¹. In addition, lemon juice was used as an acidifier to obtain PE. Lemon juice contains ascorbic acid, a powerful antioxidant, as reported by Wanjiku et al.⁴² who evaluated the antioxidant activity of the juice of different citrus fruits and observed the highest results for lemon juice, which also had the highest level of ascorbic acid (71.29 ± 3.52% DPPH radical inhibition). This discovery demonstrates that PE obtained from passion fruit peel is a source of phenolic compounds and high antioxidant activity, with the potential to be used as a natural additive to delay lipid oxidation in foods. Thus, it is an effective strategy to extend the shelf life of foods susceptible to oxidative deterioration, in addition to contributing to the reduction of the use of synthetic additives.

PE was also characterized for its antimicrobial activity by the disk-diffusion method against the bacteriological species E. coli, E. aerogenes and S. aureus. However, no halo of inhibition visible to the naked eye was found for any of the microorganisms tested.

3.3. In-vitro antioxidant activity of films

Gelatinization and drying temperatures of films can cause degradation of active components of REO and PE. Thus, the in vitro antioxidant activity of the produced films was analyzed in order to evaluate whether the antioxidant agents remained active after the film production process. The results of the antioxidant activity of the films by the DPPH and FRAP methods are presented in Figure 2.

Figure 2
Antioxidant activity of films by DPPH radical-scavenging ability and Iron reducing capacity (FRAP). Different letters in the columns indicate statistical difference by Duncan's test (P<0.05). F1 (0.50%REO - 0.00%sucrose); F2 (0.50% REO - 1.40% sucrose); F3 (1.50% REO - 0.00% sucrose); F4 (1.50% REO - 1.40% sucrose); F5 (1.00% REO - 0.70% sucrose); F6 (1.00% REO - 0.70% sucrose); F7 (0.00% REO - 0.00% sucrose). Rosemary Essential Oil (REO).

The first method is based on reducing the alcoholic solution of the DPPH radical. Because of the location of the free electron along the DPPH molecule, the radical is violet in color with an absorption in ethanol or methanol solution at 515-517 nm. In the presence of an antioxidant substance, DPPH receives H⁺, and is reduced to the DPPH-H form, with the loss of violet color to pale yellow or light violet. The higher the rate of DPPH radical scavenging, the stronger the antioxidant activity⁴³. The FRAP method, on the other hand, is based on the antioxidant's ability to reduce Fe³⁺ to Fe²⁺. In the presence of antioxidants and under acidic medium, the ferric complex tripyridyltrazine [Fe(TPTZ)₂]³⁺ receives an electron and is reduced to its ferrous form [Fe²⁺(TPTZ)₂]²⁺ with an intense blue color and maximum absorption at 595 nm⁴⁴.

By the DPPH method, the F2 film (0.50%REO-1.40%Sucrose) showed greater antioxidant action (34.32% DPPH inhibition); by the FRAP method, the film with the highest antioxidant activity was F1 (0.50%REO-0.00%Sucrose) with an iron reduction power of 1.06 mmol Fe₂SO₄/g. This shows that the formulations with lower incorporated concentrations of REO were the ones with the best antioxidant response by the two methods analyzed. In addition, the films added with REO and the control film (F7) showed similar antioxidant activity. These results suggest that the antioxidant action of the films is mainly due to PE and not to REO. As discussed in topics 3.1 and 3.2, REO has lower antioxidant activity than BHT, while that of PE is higher. Possibly, at a lower mass concentration of REO, the phenolic compounds present in the PE structure were more available for the elimination of free radicals. Thus, when in a smaller amount of REO, PE can act more efficiently in controlling oxidation.

The phenolic profiles and antioxidant capacity of pectin solutions and ethanoic extracts from passion fruit peel were evaluated in the study by Huo et al.¹⁴, with variation in the extraction conditions. Based on the DPPH and ABTS+ (2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonate) methods, the authors concluded that the antioxidant activity of PE is comparable to that of standard antioxidants available on the market and superior to that of PP. However, both materials can be used as efficient natural antioxidants in industry, depending on the yield obtained in the extraction step. In the work of Lima et al.²⁵, for example, films of myofibrillar proteins from fish by-products and pectin extracted from yellow passion fruit showed ABTS+ radical scavenging activity, which was associated, in part, with the presence of functional hydroxyl groups in the pectin structure.

Another important observation is that the action of F2 was greater than that of F1, and that of F4 greater than that of F3, showing that the addition of 1.4% of SU increases the antioxidant activity of the films. Possibly, this is because the source of sucrose was commercial refined sugar that also contains phenolic compounds, as reported by Iqbal et al.⁴⁵ who obtained a TPC content in refined sugar of 23.81 μg GAE/g. In the work of Yang et al.⁴⁶ alginate films containing the bioactive and natural antioxidant agent, S. cerevisiae, also showed greater antioxidant action in the presence of sucrose.

3.4. Fourier Transform Infrared Spectroscopy (FTIR) and principal component analysis (PCA)

As can be seen in Figure 3, all the films showed similar spectra with defined bands under the same wavelengths. The broad and strong band in the region of 3300 cm^-1 corresponds to the stretching of the hydroxyl groups (O-H), the band found in 2922 cm^-1 refers to the stretching of the C-H bond and in 1014 cm^-1 there is a spectral band attributed to the stretches of C-O-C ethers present in the polymeric chains of starch. The band at 1639 cm^-1 is resulting from water absorbed due to hydroxyl bending vibrations⁴⁷. At 1732 cm^-1, a band that may be related to the vibrational stretching C=O of esterified carboxyl methyl groups in pectin is seen. This band does not appear in AS films plasticized with glycerol⁴⁷.

Figure 3
Medium infrared spectra.

The similarity between the spectra of films with and without the addition of REO occurs because it has characteristic bands in the same regions as the AS/PE film (F7) (see Figure S1), resulting in overlapping bands in the spectrum. Thus, as the FTIR spectra were not sufficient to indicate whether REO was incorporated into the polymer matrix, Principal Component Analysis (PCA) was performed to show differences between the AS/PE film (F7) and the films with the addition of REO (F1 to F6).

The score graphs (PC1 X PC2) for the films are shown in Figure 4 and were divided into films with and without the addition of sucrose (Figure 4a and 4b, respectively). In both graphs, the films with different concentrations of REO are found in distinct groupings, indicating that there was chemical differentiation of the AS/PE film (F7) for the films with REO and, therefore, giving indications of the presence of REO in the polymeric matrix of the additive films.

Figure 4
Analysis of the main components. PC1 versus PC2 scores plot for films with sucrose (a) and without sucrose, (b) compared to the control film (F7), loadings plot to the clustering in the first component for films with sucrose (c) and without sucrose (e), loadings plot to the clustering in the second component for films with sucrose (d) and without sucrose (f). F1 (0.50% REO - 0.00% sucrose); F2 (0.50% REO - 1.40% sucrose); F3 (1.50% REO - 0.00% sucrose); F4 (1.50% REO - 1.40% sucrose); F5 (1.00% REO - 0.70% sucrose); F6 (1.00% REO - 0.70% sucrose); F7 (0.00% REO - 0.00% sucrose). Rosemary Essential Oil (REO).

Figure 4c to 4f presents the loading graphs for the grouping in the first and second main components (PC1 and PC2). From the loading graphs it is possible to identify the signals responsible for the formation of the different groupings observed in the score graphs. Analyzing the graphs, it is noted the existence of signs in the range of 1700 cm^-1 to 1800 cm^-1, the stretch region of the functional group C=O, which can be attributed to the presence of camphor, present in REO.

3.5. Optical properties (color and transparency) of films

The visual aspect of packaging films is an important factor, as it influences the presentation of the packaged product and, consequently, consumer acceptance. Most of the time, in the packaging area, transparent films are recommended to allow the visualization of the product. The photographs of the films produced (see Figure S2), show that all films have a yellow-ochre color and are visually transparent, and can be applied to products that do not need light protection.

To quantify the intensity of transparency, the films were analyzed in a UV-Vis spectrophotometer in the visible region of the spectrum (wavelength range of 600 nm), comparing the transparency result of the control film with the other formulations. Table 4 shows the transparency results, where a statistical difference was found only with the F4 formulation (1.50%REO-1.40%Sucrose), so it can be said that this statistical difference is not associated with REO or sucrose, since the transparency value of the F3 film (1.50%REO-0.00%Sucrose) and the F2 film (0.50%REO-1, 40% Sucrose) did not show significant difference for the control film (F7).

Table 4
The parameters of color and transparency in films.

Formulation (%REO - % Sucrose)	Color parameters				Transparency (log(%T)/mm)
	L* (luminosity)	a* (green-red)	b* (blue-yellow)	ÄE
F1 (0.50% - 0.00%)	82.01 (80.72–83.30)c	2.65 (2.08–3.22)a	27.61 (25.99–29.23)a.b	29.06 (27.03–31.07)a.b	14.74 (13.50–15.98)a.b
F2 (0.50% - 1.40%)	86.99 (85.63–88.35)a	0.70 (0.10–1.30)c	23.47 (21.76–25.18)c	22.92 (20.77–25.05)c	15.42 (14.12–16.72)a.b
F3 (1.50% - 0.00%)	82.87 (81.47–84.27)b.c	2.18 (1.57–2.80)a.b	29.00 (27.24–30.76)a	29.74 (27.60–31.87)a.b	16.71 (15.37–18.05)a
F4 (1.50% - 1.40%)	84.47 (83.04–85.90)a.b	1.37 (0.74–1.99)b.c	25.36 (23.56–27.16)b.c	25.72 (23.47–27.97)b.c	12.60 (11.24–13.96)c
F5 (1.00% - 0.70%)	82.80 (81.34–84.26)b.c	2.18 (1.54–2.82)a.b	30.25 (28.42–32.08)a	30.90 (28.61–33.19)a	13.68 (12.30–15.06)b.c
F6 (1.00% - 0.70%)	85.17 (83.69–86.65)a.b	1.25 (0.60–1.90)b.c	26.90 (25.05–28.75)a.b	26.80 (24.49–29.13)a.b	12.88 (11.49–14.27)b.c
F7 (0.00% - 0.00%)	86.73 (85.25–88.21)a	0.57 (−0.08–1.22)c	24.43 (22.58–26.28)b.c	23.90 (21.58–26.22)c	14.94 (13.55–16.33)a.b

• Values expressed in mean (n = 3) accompanied by IC95%. The means followed by the same letter on the same column do not differ statistically from each other by the Duncan Test at the level of 5% probability (p < 0.05). Rosemary Essential Oil (REO).

Regarding the color parameters, the L* parameter, which represents the luminosity index, ranged from 82.01 to 86.73, indicating the achievement of clear films. The positive values for parameter b* indicate that yellow color predominates in the material. For parameter a* positive means were obtained, although close to the value 0, which indicates that there is a low tendency to red color. When analyzing the total color difference (ΔE), a statistically significant difference (p<0.05) was observed between the samples, with values ranging from 22.92 in F2 (0.50%OEA-1.40%Sucrose) to 30.90 in F5 (1.00%OEA-0.70%Sucrose) (Table 4). Except for the F2 sample, all the others presented ΔE higher than the control film (F7), thus, in general, there was a reduction in the total color difference with the incorporation of REO. A similar result was found by Yeddes et al.⁴⁸ who found that the incorporation of REO resulted in a significant decrease in the ΔE value in gelatin-chitosan-pectin films.

3.6. Moisture and water vapor permeability (WVP) of films

Polysaccharide films, due to their natural hydrophilic characteristics, absorb moisture from the environment until they reach equilibrium. The amount of water absorbed can vary according to the composition of the film and the water present in the polymeric matrix also acts as a plasticizer⁴⁹. As seen in Table 5, the moisture content of the films ranged from 12.18 to 19.54%. Generally, when EO is incorporated into starch films, hydrogen bonds are formed between the starch and EO functional groups, limiting polysaccharide-water interactions and resulting in a decrease in the moisture content of the films⁵⁰. In this study, we found no evidence that the presence of REO influenced the moisture content of the films, since the moisture content of F4 (1.5% of REO) was statistically equal to that of the control film (F7). Possibly, the amount of REO was insufficient to change the water absorption behavior of the AS/PE system. Basil seed gum and oregano essential oil (OEO) films produced by Hashemi et al.⁵¹ showed no variation in moisture content with the addition of 1% OEO. For concentrations above 2%, a statistically significant increase in moisture content was detected, as also reported for quince seed mucilage films and thyme essential oil⁵².

Table 5
The physical properties of films.

Formulation (% REO - %Sucrose)	Moisture content (%)	WVP (10-8 g/h•m•Pa)	TS (MPa)	å (%)
F1 (0.50% - 0.00%)	14.56 (11.89 – 17.23) a	4.86 (4.01–5.70)a	4.50 (4.34–4.67)d	70.78 (69.45–72.11)e
F2 (0.50% - 1.40%)	12.18 (7.54 – 16.83) a	5.07 (4.18–5.96)a	9.17 (9.00–9.34)a	68.76 (67.36–70.16)e
F3 (1.50% - 0.00%)	12.71 (7.74 – 17.67) a	5.37 (4.45–6.28)a	7.86 (7.68–8.03)b	57.31 (55.87–58.75)f
F4 (1.50% - 1.40%)	17.24 (14.53 – 19.94) b	4.69 (3.76–5.62)a	4.40 (4.22–4.58)d	77.24 (75.77–78.71)d
F5 (1.00% - 0.70%)	18.68 (15.81 – 21.55) b	4.07 (3.13–5.01)a.b	5.76 (5.58–5.94)c	89.75 (88.26–91.24)b
F6 (1.00% - 0.70%)	19.54 (18.99 - 20.08) c	4.32 (3.37–5.27)a.b	5.54 (5.36–5.72)c	82.64 (81.13–84.15)c
F7 (0.00% - 0.00%)	17.11 (12.80 – 21.43) b	2.56 (1.62–3.51) b	9.42 (9.24–9.61)a	104.66 (103.16–106.18)a

• Values expressed in mean (n = 3) accompanied by IC95%. The means followed by the same letter on the same column do not differ statistically from each other by the Duncan Test at the level of 5% probability (p < 0.05). Rosemary Essential Oil (REO); Water Vapor Permeability (WVP); Tensile Strength (TS); percentage elongation (ε) at break.

For some applications, such as dry food, the barrier properties of the films are necessary to preserve the consistency of the packaged food, so it is essential to minimize moisture transfer between the ambient atmosphere and the product. In this work, we produced AS and PE (F7) films with WVP lower than that found for AS films laminated with 20% glycerol (2.56X10^-8 g/h•m•Pa)⁴⁷.

The addition of EOs is generally expected to reduce the WVP of hydrophilic polymeric matrices due to their hydrophobic characteristics. However, as reported by Atarés and Chiralt⁵³, as EOs are complex mixtures of numerous chemical compounds, their hydrophobicity is a variable characteristic and can therefore cause different effects on the WVP of the polymer matrix. In fact, in this study, it was found that the control film (F7) presented the lowest WVP (Table 5), suggesting that the addition of REO is responsible for the decrease in the barrier properties of the films. The increase in WVP with the addition of EOs has also been verified by other authors, Li et al.⁵⁴ observed a slight increase in WVP of chitosan films when incorporating turmeric essential oil, for the authors such an increase could be associated with the formation of a small amount of micropores on the surface of the films that facilitate the diffusion of water to a certain extent. Similarly, Klangmuang and Sothornvit⁵⁵ noted that the addition of Zingiber cassumunar essential oil significantly increased (p<0.05) the WVP of hydroxypropyl methylcellulose-based films. The authors suggested that EO could be acting as a plasticizer by reducing the compaction of the film structure and therefore facilitating water permeation.

3.7. Mechanical properties of films

As shown in Table 5, the TS values ranged from 4.40 MPa for F4 (1.50%REO-1.40%Sucrose) to 9.42 MPa for F7 (control film), being comparable with those presented by low-density polyethylene (6.90-16.00 MPa), a material commonly used in commercial packaging⁵⁶.

In addition, the TS value of the control film, manufactured only with AS and PE, is higher than the 2.15 MPa value found for AS films plasticized with 20% (w/w) glycerol⁴⁷, showing that PE also contributed to improve the mechanical properties of the films, which agrees with what was previously discussed by our group in the article by Araújo et al.⁵⁷ in which PE was used in conjunction with glycerol in the production of corn starch films to improve the stability of the films, allowing the obtention of films without bubbles and cracks, which was not possible when only glycerol was used. Nascimento et al.⁴⁷ reported that it was not possible to prepare pure AS films by casting, and that it was necessary to laminate them. Here we show that it is possible to produce AS films with PE, which confirms that PE can be used as an effective plasticizer for AS films. Although widely recognized as an effective plasticizer for starch matrices, glycerol has a high susceptibility to migration, which can compromise some final properties of the films, as observed by Wu et al.⁵⁸. In this work, both sucrose and PE demonstrate plasticizing potential and present themselves as natural and more environmentally friendly alternatives to conventional plasticizers. This result highlights the essential role of PE as a structural component of the film matrix. Its presence contributed significantly to the integrity and mechanical stability of the material.

The TS results also showed that the control film (F7) showed greater resistance, suggesting that the trend of TS reduction is related to the concentration of REO used. This agrees with previous studies that demonstrate that the inclusion of lipid components in films results in a less resistant matrix⁵⁹. The decrease in mechanical strength with the addition of EO is likely due to the partial replacement of stronger polymer-polymer interactions with weaker polymer-EO interactions, generating discontinuities in the polymer matrix⁶⁰. It is important to emphasize that this behavior has already been widely observed in the literature, which reports that the mechanical properties of starch-based films may vary depending on the type of essential oil used, due to differences in their reactivity and interaction with the polymeric network. As a result, some oils can negatively affect the film matrix, compromising tensile strength. In this context, our findings are consistent with such previous observations⁶¹,⁶².

In the analysis of the percentage of elongation, values ranging from 57.31% in F3 (1.50%REO-0.00% Sucrose) to 104.66% in F7 (control film) were observed. In general, compared to the other formulations with the control film, a reduction of up to 47.35% in elongation is observed. A parameter that can influence the percentage elongation is the moisture content, since water can also act as a plasticizing agent. As shown in Table 5, the formulations F1, F2 and F3 presented lower moisture content and differed significantly from the others (F4, F5, F6 and F7), agreeing with the results of percentage elongation, since, in fact, these formulations with lower moisture content showed lower percentage elongation.

Comparing the F2 and F4 formulations, a 12% increase in the percentage elongation of the formulation with the highest concentration of REO was observed. A similar effect was observed by Qin et. al⁶³ that used essential oils of different species, such as bergamot, clove, lemongrass, and rosemary, to confer mainly antimicrobial action to poly(lactic acid) (PLA) films and found that the percentage of elongation at break was significantly increased (p <0.05) by the addition of essential oil.

Sucrose (SU) has been widely used as a plasticizer additive of polymer matrices, increasing intermolecular distances and reducing existing cohesive forces⁴⁶. From the tabulated data, it can be seen that the plasticizing effect of SU seems to be correlated with the amount of REO inserted. Comparatively, formulations with 1.50% REO (F3 and F4) showed an increase of almost 35% in elongation at break due to the addition of 1.4% SU. However, in the formulations with 0.50% REO (F1 and F2), no statistically significant difference in this property was found with the inclusion of SU. Possibly, the action of SU was effective only in formulations with higher concentrations of REO. Often, additives inserted to add specific characteristics to the packaging can act synergistically or not, depending on the quantities used. According to Veiga-Santos et al.⁶⁴, with the increase in the concentration of SU as a plasticizer, there was an increase in the elongation of cassava starch films. Here, this correlation was not direct, due to the presence of REO above 0.5% and possible physical interactions with the SU. The existence of hydrogen bonds between SU and REO, for example, may have an influence on the differences cited for the property under study.

4. Conclusion

In this study, it was demonstrated that PE obtained from passion fruit peel, in addition to being a source of phenolic compounds with the potential to be used as an antioxidant additive in active packaging, also works as an effective plasticizer for AS films and can replace other synthetic plasticizers. In addition, the PCA results of the FTIR spectra of the films produced suggest the probable incorporation of REO in the polymeric matrix, allowing the films to also present antimicrobial action, since REO showed significant inhibition against the bacteria E. coli and S. aureus.

In general, the addition of REO caused a reduction in the properties of tensile strength and percentage elongation, and an increase in the water vapor permeability of AS/PE films. The plasticizing effect of SU, in turn, was dependent on the amount of REO inserted, and for higher concentrations of REO (above 0.5%), results show possible physical interactions between both, which resulted in an increase in percentage elongation.

In view of the results, it was possible to conclude that the formulation (F2), with 0.5% REO and 1.4% SU, was the one that had the best performance among the formulations studied, as it statistically maintained the transparency, color and tensile strength of the AS/PE film, in addition to having the highest antioxidant activity. Therefore, it is possible to state that there may have been a synergistic effect of REO and PE promoting antioxidant and antimicrobial activity for the packaging material.

Supplementary Material

The following online material is available for this article:

Figure S1

Medium infrared spectra of rosemary essential oil (REO).

Figure S2

Photographs of the films.

5. Acknowledgments

The authors acknowledge the Coordination for the Improvement of Higher Education Personnel (CAPES) for the scholarship to the first author (C.R. Duarte). They also thank the Paraiba State Research Foundation (FAPESQ) for providing the scholarship for the second author (C. V. B. Grisi) [grant 2023/3390.18].

Data Availability

Availability of data and materials data are available by contacting the authors.

6. References

• ¹ Panou A, Karabagias I. Biodegradable packaging materials for foods preservation: sources, advantages, limitations, and future perspectives. Coatings. 2023;13(7):1176. https://doi.org/10.3390/coatings13071176
  » https://doi.org/10.3390/coatings13071176
• ² Walker TR, Fequet L. Current trends of unsustainable plastic production and micro(nano)plastic pollution. Trends Analyt Chem. 2023;160:116984. https://doi.org/10.1016/j.trac.2023.116984
  » https://doi.org/10.1016/j.trac.2023.116984
• ³ Kaplan DL. Biopolymers from renewable resources. Berlin: Springer; 1998. https://doi.org/10.1007/978-3-662-03680-8
  » https://doi.org/10.1007/978-3-662-03680-8
• ⁴ Panou A, Karabagias I. Biodegradable packaging materials for foods preservation: sources, advantages, limitations, and future perspectives. Coatings. 2023;13(7):1176. https://doi.org/10.3390/coatings13071176
  » https://doi.org/10.3390/coatings13071176
• ⁵ Nogueira GF, Fakhouri FM, de Oliveira RA. Extraction and characterization of arrowroot (Maranta arundinaceae L.) starch and its application in edible films. Carbohydr Polym. 2018;186:64-72. https://doi.org/10.1016/j.carbpol.2018.01.024 PMid:29456010.
  » https://doi.org/10.1016/j.carbpol.2018.01.024
• ⁶ Tarique J, Sapuan SM, Khalina A, Sherwani SFK, Yusuf J, Ilyas RA. Recent developments in sustainable arrowroot (Maranta arundinacea Linn) starch biopolymers, fibres, biopolymer composites and their potential industrial applications: a review. J Mater Res Technol. 2021;13:1191-219. https://doi.org/10.1016/j.jmrt.2021.05.047
  » https://doi.org/10.1016/j.jmrt.2021.05.047
• ⁷ Sani IK, Geshlaghi SP, Pirsa S, Asdagh A. Composite film based on potato starch/apple peel pectin/ZrO₂ nanoparticles/microencapsulated Zataria multiflora essential oil; investigation of physicochemical properties and use in quail meat packaging. Food Hydrocoll. 2021;117:106719. https://doi.org/10.1016/j.foodhyd.2021.106719
  » https://doi.org/10.1016/j.foodhyd.2021.106719
• ⁸ Bai W, Vidal NP, Roman L, Portillo-Perez G, Martinez MM. Preparation and characterization of self-standing biofilms from compatible pectin/starch blends: effect of pectin structure. Int J Biol Macromol. 2023;251:126383. https://doi.org/10.1016/j.ijbiomac.2023.126383 PMid:37595713.
  » https://doi.org/10.1016/j.ijbiomac.2023.126383
• ⁹ Dash KK, Ali NA, Das D, Mohanta D. Thorough evaluation of sweet potato starch and lemon-waste pectin based-edible films with nano-titania inclusions for food packaging applications. Int J Biol Macromol. 2019;139:449-58. https://doi.org/10.1016/j.ijbiomac.2019.07.193 PMid:31374275.
  » https://doi.org/10.1016/j.ijbiomac.2019.07.193
• ¹⁰ Yun D, Liu J. Recent advances on the development of food packaging films based on citrus processing wastes: a review. J Agric Food Res. 2022;9:100316. https://doi.org/10.1016/j.jafr.2022.100316
  » https://doi.org/10.1016/j.jafr.2022.100316
• ¹¹ Zhang X, Chen X, Dai J, Cui H, Lin L. Edible films of pectin extracted from dragon fruit peel: effects of boiling water treatment on pectin and film properties. Food Hydrocoll. 2024;147:109324. https://doi.org/10.1016/j.foodhyd.2023.109324
  » https://doi.org/10.1016/j.foodhyd.2023.109324
• ¹² Sharma K, Mahato N, Cho MH, Lee YR. Converting citrus wastes into value-added products: economic and environmently friendly approaches. Nutrition. 2017;34:29-46. https://doi.org/10.1016/j.nut.2016.09.006 PMid:28063510.
  » https://doi.org/10.1016/j.nut.2016.09.006
• ¹³ IBGE: Instituto Brasileiro de Geografia e Estatística. Produção mundial de maracujá. Rio de Janeiro: IBGE; 2022.
• ¹⁴ Huo D, Dai J, Yuan S, Cheng X, Pan Y, Wang L, et al. Eco-friendly simultaneous extraction of pectins and phenolics from passion fruit (Passiflora edulis Sims) peel: process optimization, physicochemical properties, and antioxidant activity. Int J Biol Macromol. 2023;243:125229. https://doi.org/10.1016/j.ijbiomac.2023.125229 PMid:37301339.
  » https://doi.org/10.1016/j.ijbiomac.2023.125229
• ¹⁵ Sharma S, Barkauskaite S, Jaiswal AK, Jaiswal S. Essential oils as additives in active food packaging. Food Chem. 2021;343:128403. https://doi.org/10.1016/j.foodchem.2020.128403 PMid:33268167.
  » https://doi.org/10.1016/j.foodchem.2020.128403
• ¹⁶ UNDESA: United Nations Department of Economic and Social Affairs. The waste crisis: accelerating national to local policy action. evidence-based strategies for sustainable solutions [Internet]. 2024 [cited 2024 Oct 19]. Available from: https://wedocs.unep.org/handle/20.500.11822/45230
  » https://wedocs.unep.org/handle/20.500.11822/45230
• ¹⁷ Kobacy W, Gong X, Elshamandy NKF, Zahran HA, Osman AI, Rashwan AK. ATR-FTIR, DSC, GC–MS of garlic (Allium sativum), parsley (Petroselinum crispum), and thyme (Thymus vulgaris) essential oils and their synergistic effects on antioxidant and antimicrobial properties. Food Biosci. 2025;71:107262. https://doi.org/10.1016/j.fbio.2025.107262
  » https://doi.org/10.1016/j.fbio.2025.107262
• ¹⁸ Christopoulou SD, Androutsopoulou C, Hahalis P, Kotsalou C, Vantarakis A, Lamari FN. Rosemary extract and essential oil as drink ingredients: an evaluation of their chemical composition, genotoxicity, antimicrobial, antiviral, and antioxidant properties. Foods. 2021;10(12):3143. https://doi.org/10.3390/foods10123143 PMid:34945695.
  » https://doi.org/10.3390/foods10123143
• ¹⁹ Yahyaoui M, Gordobil O, Herrera Díaz R, Abderrabba M, Labidi J. Development of novel antimicrobial films based on poly(lactic acid) and essential oils. React Funct Polym. 2016;109:1-8. https://doi.org/10.1016/j.reactfunctpolym.2016.09.001
  » https://doi.org/10.1016/j.reactfunctpolym.2016.09.001
• ²⁰ Souza CO, Silva LT, Druzian JI. Estudo comparativo da caracterização de filmes biodegradáveis de amido de mandioca contendo polpas de manga e de acerola. Quim Nova. 2012;35(2):262-7. https://doi.org/10.1590/S0100-40422012000200006
  » https://doi.org/10.1590/S0100-40422012000200006
• ²¹ Silva AJ, Oliveira LP, Resende JCP, Saraiva IS. Avaliação da atividade antimicrobiana de óleos essenciais obtidos de diferentes fabricantes. Sinapse Múltipla. 2019;8:33-40.
• ²² Singleton VL, Rossi JA Jr. Colorimetry of total phenolics with phosphomolybdic-phosphotungstic acid reagents. Am J Enol Vitic. 1965;16(3):144-58. https://doi.org/10.5344/ajev.1965.16.3.144
  » https://doi.org/10.5344/ajev.1965.16.3.144
• ²³ Brand-Williams W, Cuvelier ME, Berset C. Use of a free radical method to evaluate antioxidant activity. Lebensm Wiss Technol. 1995;28(1):25-30. https://doi.org/10.1016/S0023-6438(95)80008-5
  » https://doi.org/10.1016/S0023-6438(95)80008-5
• ²⁴ Rufino MSM, Alves RE, Brito ES, Morais SM, Sampaio CG, Pérez-Jiménez J, et al. Metodologia científica: determinação da atividade antioxidante total em frutas pelo método de redução do ferro (FRAP). Fortaleza: Embrapa Agroindústria Tropical; 2006.
• ²⁵ Lima DAS, Grisi CVB, Florentino GIB, Santos MMF, Madruga MS, Silva FAP. Preparation and characterization of sustainable active packaging based on myofibrillar proteins and protein hydrolysates from the cutting by-product of Scomberomorus brasiliensis filleting on the band saw machine. Food Chem. 2024;460(Pt 1):140490. https://doi.org/10.1016/j.foodchem.2024.140490 PMid:39033637.
  » https://doi.org/10.1016/j.foodchem.2024.140490
• ²⁶ Medina Jaramillo C, Gutiérrez TJ, Goyanes S, Bernal C, Famá L. Biodegradability and plasticizing effect of yerba mate extract on cassava starch edible films. Carbohydr Polym. 2016;151:150-9. https://doi.org/10.1016/j.carbpol.2016.05.025 PMid:27474554.
  » https://doi.org/10.1016/j.carbpol.2016.05.025
• ²⁷ Sothornvit R, Hong S-I, An DJ, Rhim J-W. Effect of clay content on the physical and antimicrobial properties of whey protein isolate/organo-clay composite films. Lebensm Wiss Technol. 2010;43(2):279-84. https://doi.org/10.1016/j.lwt.2009.08.010
  » https://doi.org/10.1016/j.lwt.2009.08.010
• ²⁸ ASTM: American Society for Testing and Materials. ASTM E308-17: standard practice for computing the colors of objects by using the CIE system. West Conshohocken: ASTM; 2017.
• ²⁹ ASTM: American Society for Testing and Materials. ASTM E96-95: standard test methods for water vapor transmission of materials. West Conshohocken: ASTM; 2014.
• ³⁰ ASTM: American Society for Testing and Materials. ASTM D882-12: standard test method for tensile properties of thin plastic sheeting. West Conshohocken: ASTM; 2012.
• ³¹ Micić D, Đurović S, Riabov P, Tomić A, Šovljanski O, Filip S, et al. Rosemary essential oils as a promising source of bioactive compounds: chemical composition, thermal properties, biological activity, and gastronomical perspectives. Foods. 2021;10(11):2734. https://doi.org/10.3390/foods10112734 PMid:34829014.
  » https://doi.org/10.3390/foods10112734
• ³² Barak TH, Bölükbaş E, Bardakci H. Evaluation of marketed rosemary essential oils (Rosmarinus officinalis L.) in terms of European Pharmacopoeia 10.0 criteria. Turk J Pharm Sci. 2023;20(4):253-60. https://doi.org/10.4274/tjps.galenos.2022.78010 PMid:37606010.
  » https://doi.org/10.4274/tjps.galenos.2022.78010
• ³³ CLSI: Clinical and Laboratory Standards Institute. Performance standards for antimicrobial disk susceptibility tests: approved standard. Wayne: CLSI; 2015.
• ³⁴ Ebrahimi E, Haghjou M, Nematollahi A, Goudarzian F. Effects of rosemary essential oil on growth performance and hematological parameters of young great sturgeon (Huso huso). Aquaculture. 2020;521:734909. https://doi.org/10.1016/j.aquaculture.2019.734909
  » https://doi.org/10.1016/j.aquaculture.2019.734909
• ³⁵ Baydar H, Özkan G, Erbaş S, Altındal D. Yield, chemical composition and antioxidant properties of extracts and essential oils of sage and rosemary depending on seasonal variations. Acta Hortic. 2009;(826):383-90. https://doi.org/10.17660/ActaHortic.2009.826.54
  » https://doi.org/10.17660/ActaHortic.2009.826.54
• ³⁶ Antunes SA, Robazza WS, Schittler L, Gomes GA. Synergistic and antimicrobial properties of commercial turmeric (Curcuma longa) essential oil against pathogenic bacteria. Food Sci Technol. 2012;32(3):525-30. https://doi.org/10.1590/S0101-20612012005000082
  » https://doi.org/10.1590/S0101-20612012005000082
• ³⁷ Aljabeili HS, Barakat H, Abdel-Rahman HA. Chemical composition, antibacterial and antioxidant activities of thyme essential oil (Thymus vulgaris). Food Nutr Sci. 2018;9(5):433-46. https://doi.org/10.4236/fns.2018.95034
  » https://doi.org/10.4236/fns.2018.95034
• ³⁸ Živković J, Ćebović T, Maksimović Z. In vivo and in vitro antioxidant effects of three Veronica species. Open Life Sci. 2012;7(3):559-68. https://doi.org/10.2478/s11535-012-0041-4
  » https://doi.org/10.2478/s11535-012-0041-4
• ³⁹ Zeraik ML, Yariwake JH, Wauters J-N, Tits M, Angenot L. Analysis of passion fruit rinds (Passiflora edulis): isoorientin quantification by HPTLC and evaluation of antioxidant (radical scavenging) capacity. Quim Nova. 2012;35(3):541-5. https://doi.org/10.1590/S0100-40422012000300019
  » https://doi.org/10.1590/S0100-40422012000300019
• ⁴⁰ Ziqubu K, Dludla PV, Joubert E, Muller CJF, Louw J, Tiano L, et al. Isoorientin: a dietary flavone with the potential to ameliorate diverse metabolic complications. Pharmacol Res. 2020;158:104867. https://doi.org/10.1016/j.phrs.2020.104867 PMid:32407953.
  » https://doi.org/10.1016/j.phrs.2020.104867
• ⁴¹ Zeb A. Concept, mechanism, and applications of phenolic antioxidants in foods. J Food Biochem. 2020;44(9):e13394. https://doi.org/10.1111/jfbc.13394 PMid:32691460.
  » https://doi.org/10.1111/jfbc.13394
• ⁴² Wanjiku MM, Nyambura MB, Kirwa ME, Mercy M. Anti-oxidative potential of honey and ascorbic acid in yoghurt fortified with omega-3 fatty acids. J Microbiol Biotechnol Food Sci. 2016;6:702-6. https://doi.org/10.15414/jmbfs.2016.6.1.702-706
  » https://doi.org/10.15414/jmbfs.2016.6.1.702-706
• ⁴³ Wu H, Lei Y, Zhu R, Zhao M, Lu J, Xiao D, et al. Preparation and characterization of bioactive edible packaging films based on pomelo peel flours incorporating tea polyphenol. Food Hydrocoll. 2019;90:41-9. https://doi.org/10.1016/j.foodhyd.2018.12.016
  » https://doi.org/10.1016/j.foodhyd.2018.12.016
• ⁴⁴ Benzie IFF, Strain JJ. The ferric reducing ability of plasma (FRAP) as a measure of "antioxidant power": the FRAP assay. Anal Biochem. 1996;239(1):70-6. https://doi.org/10.1006/abio.1996.0292 PMid:8660627.
  » https://doi.org/10.1006/abio.1996.0292
• ⁴⁵ Iqbal M, Afzal Qamar M, Bokhari TH, Abbas M, Hussain F, Masood N, et al. Total phenolic, chromium contents and antioxidant activity of raw and processed sugars. Inf Process Agric. 2017;4(1):83-9. https://doi.org/10.1016/j.inpa.2016.11.002
  » https://doi.org/10.1016/j.inpa.2016.11.002
• ⁴⁶ Yang Z, Zhai X, Li M, Li Z, Shi J, Huang X, et al. Saccharomyces cerevisiae-incorporated and sucrose-rich sodium alginate film: an effective antioxidant packaging film for longan preservation. Int J Biol Macromol. 2022;223(Pt A):673-83. https://doi.org/10.1016/j.ijbiomac.2022.11.039 PMid:36368365.
  » https://doi.org/10.1016/j.ijbiomac.2022.11.039
• ⁴⁷ Nascimento JAA, Santos AF, Silva IDL, Falcão EHL, Britto D, Vinhas GM. Physico-chemical, mechanical and morphological properties of biodegradable films based on arrowroot starch and poly(vinyl alcohol). J Macromol Sci Part B Phys. 2021;60(12):1045-68. https://doi.org/10.1080/00222348.2021.1949836
  » https://doi.org/10.1080/00222348.2021.1949836
• ⁴⁸ Yeddes W, Djebali K, Aidi Wannes W, Horchani-Naifer K, Hammami M, Younes I, et al. Gelatin-chitosan-pectin films incorporated with rosemary essential oil: optimized formulation using mixture design and response surface methodology. Int J Biol Macromol. 2020;154:92-103. https://doi.org/10.1016/j.ijbiomac.2020.03.092 PMid:32173440.
  » https://doi.org/10.1016/j.ijbiomac.2020.03.092
• ⁴⁹ Cazón P, Puertas G, Vázquez M. Characterization of multilayer bacterial cellulose-chitosan films loaded with grape bagasse antioxidant extract: insights into spectral and water properties, microstructure, and antioxidant activity. Int J Biol Macromol. 2024;268(Pt 2):131774. https://doi.org/10.1016/j.ijbiomac.2024.131774 PMid:38663700.
  » https://doi.org/10.1016/j.ijbiomac.2024.131774
• ⁵⁰ Song X, Cheng L, Tan L. Edible iron yam and maize starch convenient food flavoring packaging films with lemon essential oil as plasticization. Food Sci Technol. 2019;39(4):971-9. https://doi.org/10.1590/fst.13118
  » https://doi.org/10.1590/fst.13118
• ⁵¹ Hashemi SMB, Mousavi Khaneghah A, Ghaderi Ghahfarrokhi M, Eş I. Basil-seed gum containing Origanum vulgare subsp. viride essential oil as edible coating for fresh cut apricots. Postharvest Biol Technol. 2017;125:26-34. https://doi.org/10.1016/j.postharvbio.2016.11.003
  » https://doi.org/10.1016/j.postharvbio.2016.11.003
• ⁵² Jouki M, Mortazavi SA, Yazdi FT, Koocheki A. Characterization of antioxidant–antibacterial quince seed mucilage films containing thyme essential oil. Carbohydr Polym. 2014;99:537-46. https://doi.org/10.1016/j.carbpol.2013.08.077 PMid:24274540.
  » https://doi.org/10.1016/j.carbpol.2013.08.077
• ⁵³ Atarés L, Chiralt A. Essential oils as additives in biodegradable films and coatings for active food packaging. Trends Food Sci Technol. 2016;48:51-62. https://doi.org/10.1016/j.tifs.2015.12.001
  » https://doi.org/10.1016/j.tifs.2015.12.001
• ⁵⁴ Li Z, Lin S, An S, Liu L, Hu Y, Wan L. Preparation, characterization and anti-aflatoxigenic activity of chitosan packaging films incorporated with turmeric essential oil. Int J Biol Macromol. 2019;131:420-34. https://doi.org/10.1016/j.ijbiomac.2019.02.169 PMid:30831165.
  » https://doi.org/10.1016/j.ijbiomac.2019.02.169
• ⁵⁵ Klangmuang P, Sothornvit R. Barrier properties, mechanical properties and antimicrobial activity of hydroxypropyl methylcellulose-based nanocomposite films incorporated with Thai essential oils. Food Hydrocoll. 2016;61:609-16. https://doi.org/10.1016/j.foodhyd.2016.06.018
  » https://doi.org/10.1016/j.foodhyd.2016.06.018
• ⁵⁶ Coutinho FMB, Mello IL, Santa Maria LC. Polietileno: principais tipos, propriedades e aplicações. Int J Biol Macromol. 2003;13(1):1-13. https://doi.org/10.1590/S0104-14282003000100005
  » https://doi.org/10.1590/S0104-14282003000100005
• ⁵⁷ Araújo MNP, Grisi CVB, Duarte CR, Almeida YMB, Vinhas GM. Active packaging of corn starch with pectin extract and essential oil of Turmeric Longa Linn: preparation, characterization and application in sliced bread. Int J Biol Macromol. 2023;226:1352-9. https://doi.org/10.1016/j.ijbiomac.2022.11.248 PMid:36455820.
  » https://doi.org/10.1016/j.ijbiomac.2022.11.248
• ⁵⁸ Wu Y, Gao S, Zhao J, Kong S, Wang H, Wang W, et al. Sugar/sugar alcohol with glycerol as co-plasticizers for high-content starch/PBAT blown films: from fine structure to physicochemical properties. J Sci Food Agric. 2025;105(2):1105-15. https://doi.org/10.1002/jsfa.13901 PMid:39300042.
  » https://doi.org/10.1002/jsfa.13901
• ⁵⁹ Spotti ML, Cecchini JP, Spotti MJ, Carrara CR. Brea Gum (from Cercidium praecox) as a structural support for emulsion-based edible films. Lebensm Wiss Technol. 2016;68:127-34. https://doi.org/10.1016/j.lwt.2015.12.018
  » https://doi.org/10.1016/j.lwt.2015.12.018
• ⁶⁰ Lee JY, Garcia CV, Shin GH, Kim JT. Antibacterial and antioxidant properties of hydroxypropyl methylcellulose-based active composite films incorporating oregano essential oil nanoemulsions. Lebensm Wiss Technol. 2019;106:164-71. https://doi.org/10.1016/j.lwt.2019.02.061
  » https://doi.org/10.1016/j.lwt.2019.02.061
• ⁶¹ Shen Z, Kamdem DP. Development and characterization of biodegradable chitosan films containing two essential oils. Int J Biol Macromol. 2015;74:289-96. https://doi.org/10.1016/j.ijbiomac.2014.11.046 PMid:25542174.
  » https://doi.org/10.1016/j.ijbiomac.2014.11.046
• ⁶² Chavez-Marquez E, Bernedo MSB, de Jara EM, Quequezana-Bedregal MJ, Gutierrez-Oppe EE, Pessôa PA Fo. Development of intelligent and active potato starch films based on purple corn cob extract and molle essential oil. Int J Biol Macromol. 2023;242(Pt 3):125080. https://doi.org/10.1016/j.ijbiomac.2023.125080 PMid:37245765.
  » https://doi.org/10.1016/j.ijbiomac.2023.125080
• ⁶³ Qin Y, Li W, Liu D, Yuan M, Li L. Development of active packaging film made from poly (lactic acid) incorporated essential oil. Prog Org Coat. 2017;103:76-82. https://doi.org/10.1016/j.porgcoat.2016.10.017
  » https://doi.org/10.1016/j.porgcoat.2016.10.017
• ⁶⁴ Veiga-Santos P, Oliveira LM, Cereda MP, Scamparini ARP. Sucrose and inverted sugar as plasticizer. Effect on cassava starch–gelatin film mechanical properties, hydrophilicity and water activity. Food Chem. 2007;103(2):255-62. https://doi.org/10.1016/j.foodchem.2006.07.048
  » https://doi.org/10.1016/j.foodchem.2006.07.048

Edited by

• Associate Editor:
  Elisabete Frollini.

• Editor-in-Chief:
  Luiz Antonio Pessan.

Publication Dates

• Publication in this collection
  27 Feb 2026
• Date of issue
  2026

History

• Received
  23 Apr 2025
• Reviewed
  18 Nov 2025
• Accepted
  16 Jan 2026