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UV-irradiated gelatin-chitosan bio-based composite film, physiochemical features and release properties for packaging applications
Journal Pre-proofs UV-irradiated gelatin-chitosan bio-based composite film, physiochemical features and release properties for packaging applications Mahsa Rezaee, Gholamreza Askari, Zahra EmamDjomeh PII: DOI: Reference:
S0141-8130(19)35068-8 https://doi.org/10.1016/j.ijbiomac.2019.10.066 BIOMAC 13560
To appear in:
International Journal of Biological Macromolecules
Received Date: Revised Date: Accepted Date:
3 July 2019 24 September 2019 7 October 2019
Please cite this article as: M. Rezaee, G. Askari, Z. EmamDjomeh, UV-irradiated gelatin-chitosan bio-based composite film, physiochemical features and release properties for packaging applications, International Journal of Biological Macromolecules (2019), doi: https://doi.org/10.1016/j.ijbiomac.2019.10.066
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© 2019 Published by Elsevier B.V.
UV-irradiated gelatin-chitosan bio-based composite film, physiochemical features and release properties for packaging applications
Mahsa Rezaee1, Gholamreza Askari1*, Zahra EmamDjomeh1, 1
Transport Phenomena Laboratory (TPL) Department of Food Science and Technology, University College of Agriculture and Natural Resources, University of Tehran, Karaj, Iran *- corresponding author: Gholamreza Askari, Email: [email protected]
Abstract In this study, Chitosan-Gelatin biodegradable films loaded with Gallic acid (GA) were developed. Ethanol, tween, and β-Cyclodextrin (β-CD) were used to enhance the distribution of GA in the film forming solutions. The effect of the exposure time of UV irradiation on the color, mechanical properties (TS, %E), water vapor permeability (WVP) microstructure and release rate of GA to fatty food simulants investigated. UV irradiation reduced darkness of the films markedly and increased the release rate of the GA. Among the non-irradiated samples the highest and lowest release rate were obtained for the films containing β-CD and ethanol respectively. On the other hand incorporation of β-CD could reduce the increasing effect of UV light on release rate. The UV irradiation for 10 minutes reduced mechanical strength and barrier properties of the film against water vapor. Microscopic analysis, showed the consistent and uniform microstructure. However, long-term radiation caused some cracks in the films network. The UV irradiation weakened the _OH bending and enhanced the amide I amide II.
Key words: UV irradiation; biodegradable; chitosan; gelatin; Gallic acid; β-Cyclodextrin 1. Introduction Due to the ecological concerns of plastics as the main packaging materials, Bio-based polymers have been considered recently because of their biodegradable nature and consequently environmental friendly characteristics[1]. Nowadays, active packaging has become more popular. There are two kinds of active packaging systems: active-releasing which target compounds added into the headspace or the packaged food, and activescavenging systems, which undesired compounds removed permanently from the food or its environment[2]. Antioxidants are the most widely used components in the first type of active packaging systems. Undesirable properties of synthetic antioxidants such as toxicity has led to a growing tendency to use natural, nontoxic antioxidants [3]. Gallic Acid (GA) is a common natural antioxidant that generally found in many sources such as grape, tea, staghorn sumac, and strawberry. Based on some studies, GA shows anticancer and anti-tumor properties in addition to its antioxidant and bacteriostatic characteristics[4,5].
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Similar to other natural antioxidants, GA is also sensitive to environmental stresses such as pH, light, and elevated temperatures[6]. On the other hand researchers are looking for ways to control the release of biologically active compounds to the packaged foods, since the release rate and its pattern has a major effect on the efficiency of drugs and biocompounds. The desired result can be achieved when the biological compound is released in controlled manner [7]. In such cases, cyclodextrins (CD’s) are one of the best vehicles to protect GA and preserve its structural integrity through forming inclusion complexes(IC’s)[8]. Cone-shaped structure of CD’s with hydrophobic cavity and hydrophilic surface make them suitable to form IC’s with guest molecules such as bioactive compounds[9]. In addition, it has shown that CD’s are promising agents to ease controlled release of hydrophobic molecules[10]. Meanwhile, Chitosan and Gelatin as biodegradable, nontoxic and compatible biopolymers are good candidates for making packaging materials [11]. They can form excellent poly electric complex with each other because of their potentially opposite charges at specific conditions[12]. There are electrostatic interactions and hydrogen bonds, mainly between the amino groups of Chitosan and the carboxyl groups of Gelatin, and between polymer chains involving their carbonyl, hydroxyl and amino groups [13]. Cross-linking is a known method for modification of biopolymers. It has been reported that cross- linking amplified the characteristics of Chitosan-based biomaterials[14]. Cross- linking could be done through chemical agents, enzymatic treatment or physical methods[15–17]. Due to many benefits such as simplicity, ecofriendly, low cost, and additive free process, radiation induced crosslinking gained more attention in recent years[18,19]. In comparison with the other ionizing radiation, UV radiation is suitable for modifying physical and mechanical properties of protein-based films due to moderate and nondestructive effect of UV light. Some studies reported improvement in cross-linking of collagen and Gelatin, in addition to modifying physical and mechanical properties of protein-based films by UV radiation [20]. Up to know, few works have considered the effects of UV irradiation on some characteristics of edible films such as mechanical and physicochemical properties [20–22]. To the best of our knowledge, no previous study has been reported the effect of UV treatment on release behavior of antioxidant agent from carbohydrate-protein composite films, particularly Chitosan-Gelatin films. Although many studies have been done about the mixture of antioxidant and β-CD, its characteristics and effects on physical and release properties in irradiated edible films has not been investigated [7,23–26]. The objective of current research was to determine the effect of some additives including ethanol, tween and β-CD and different time of UV irradiation on chemical, physical and morphological properties of active films based on blends of Gelatin and Chitosan incorporating GA. Release profile of GA into ethanol solution as the fatty food simulant was also investigated.
2. Materials and methods 2.1. Raw materials Two main film forming materials were medium molecular weight Chitosan (Sigma-Aldrich, Germany) and Gelatin (Merck, Darmstadt, Germany). An emulsifier agent, Plasticizer, and antioxidant used were Tween 20 (Merck, Germany), extra pure (≥99 %) glycerol (Merck, Germany), and Gallic acid (monohydrate, extra pure, molecular weight of 188.14 g mol-1, (Sigma-Aldrich, USA). Glacial acetic acid obtained from Merck, 2
Germany. Ethanol 96% was obtained from Alcohol Pars Co. (Iran). β-CD obtained from Sigma-Aldrich (USA). 2.2. Preparation of Chitosan- Gelatin solution Preparation of film forming solution carried out according to our previous work [3]. At First, in order to achieve 1 % (w/v) Chitosan film forming solution, certain amount of Chitosan powder dissolved in 1 % (v/v) solution of aqueous acetic acid at 40 °C with magnetic stirring. Afterwards, glycerol (10% w/w dry matter) added to the solution under constant rate of stirring. The pH of the solution determined about 4 ± 0.2. Preparation of Gelatin film forming solution (3 % w/v) was performed by adding Gelatin powder to distilled water under stirring condition at 50 °C for 30 min until complete dissolution. Finally, glycerol (10 % w/w dry matter) was incorporated to Gelatin solution. The measured pH was about 5 ± 0.2. In order to obtain clear film-forming solution, each solution was separately filtered through filter paper (No. 4). The final composite film forming solution achieved by mixing equal weights of the Gelatin and Chitosan solutions and stirring for 30 min. To obtain polyelectrolyte complex between Chitosan and Gelatin the pH value of the solution adjusted at 5.6 using sodium hydroxide solution. 2.3. UV irradiation process Time-controlled irradiation of composite solution was carried out using UV lamp (TUV 6W G6T5, Philips, Poland). After pouring composite solution into a graduated cylinder, UV lamp placed in contact with solution into the quartz cylinder. The solution was circulated by blowing air bubbles in order to obtain uniform irradiation. The whole set were in a chamber which its temperature was controlled by a fan at 25ͦ C (ambient temperature). Irradiation time was varied for 0, 5, and 10 minutes. 2.4. Antioxidant addition GA was added to the resulting film forming solution at a concentration of 1 % (g/g total dry weight of film). Antioxidant addition carried out in four different ways: adding GA with ethanol as the solvent of antioxidant, with Tween 20 as an emulsifier agent at concentrations of 100 % (w/w) relative to the weight of the Gallic acid, and adding singular powder of GA and its physical mixture with β-CD to the solution. Homogenization of all the aqueous dispersions carried out at 13,000 rpm for 3 min using the Ultra Turrax (T25, IKA, Germany) homogenizer. 2.5. Film formation and conditioning A volume of 85 mL of the final film forming solutions were poured into Petri dishes with 15 cm diameter and dried at 30 °C for 5 days. The resulting films were peeled off from the casting plates. Prior to any characterization, films conditioned at room temperature over a saturated solution of MgCl2 (32 % RH) in desiccators for 72 h. Name of the film samples and their abbreviation are listed in Table 1. Suggested location for table 1 2.6. Films thickness. The thickness of the films was measured using a micrometer (Mitutoyo, ID C112PM, Kawasaki-shi, Japan). Mean values of five measurements were used for thickness determination and further calculations. 2.7. Color measurements.
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Optical properties of the samples were measured using colorimeter (Minolta CR-300 Series, Minolta, Japan). Color parameters of the films including L for luminosity and b for tonalities from yellow to blue were measured. The reflection spectra of the samples placed on the surface of a standard white plate resulted in the coordinates CIE Lab L, a, and b. The yellowness indexes (YI) of samples was determined using the following equation [27]. YI = 142.86 bL-1 The measurement for all samples for each formulation was carried out in triplicate. 2.8. Mechanical properties Mechanical properties of the fabricated films, including Tensile strength (TS) and elongation at break (%EB) were measured using the texture analyzer (Testometric M350-10CT, Rochdale, Lancs., England) according to the ASTM method D882 [28] with slight modifications described by Weijie, et al [29]. Samples were cut into rectangular shapes with width and length of 10 mm and 80 mm, respectively. Testing of the samples were performed through placing them in the film-extension grips with an initial distance of 50 mm at the speed of 50 mm/min until breakage. Following equations used to calculate TS and EB: TS (MPa) = (Maximum force (N) / Film thickness (m) × Film width (m)) × 10-6 EB (%) = (Final length of the film sample at the point of break )m)/Initial length of the film sample (m)) × 100 2.9. Water vapor permeability (WVP) The water vapor permeability of the films measured according to the standard ASTM E96–95 [30]. The containers which made of glass with internal diameter of 12.3 mm filled with anhydrous calcium chloride to achieve 0% RH. The film samples were placed on the mouth of the containers and sealed with molten paraffin and parafilm. Then, the containers were introduced into a desiccator containing saturated solution of sodium chloride with 75% relative humidity at room temperature and weighted at intervals of 12 h during 13 days to record their mass gain. Linear regression was used to determine the slope of mass changes versus time. WVP was calculated by the following equation: WVTR = Curve slope (g.s-1.mm-2)/Film area (mm2) WVP = Thickness (mm) × WVTR/Pressure difference (KPa) Where WVTR is the transmission rate of water vapor through the film and calculated from the slope of the weight changes versus time using linear regression. 2.10. Fourier transform infrared spectroscopy (FTIR) FTIR spectra of all films was carried out by a FT-IR spectrometer (PerkinElmer instruments, USA). The pellets of samples prepared with KBr powder and the IR spectra were recorded in the wavenumber range of 4000–450 cm−1. 2.11. Microstructure
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Scanning electron microscope (LEO 1455 VP, England) was used to examine microstructure of the films. In order to obtain brittle structure of films to prepare cross-sections, they immersed in liquid nitrogen in advance. 2.12. Antioxidant Release The release properties of GA, was studied according to the method described in our previous work [12]. Briefly, the film sample (2 cm × 2 cm) immersed into 50 mL of ethanol 96%, as fatty food simulant, in a 100 mL Erlenmeyer flask, and shake at 70 rpm and 30 °C. At regular time intervals 4 mL of solutions was collected. Measuring the amount of released GA in the collected samples was performed by a UV–visible spectrophotometer (Spectrum Instruments, Germany) at a wavelength of 259 nm and each collected sample was returned to the flask after each measurement. The simulant containing GA free film sample was used as blank. The standard curve was achieved by measuring the absorbance of different specified concentrations of GA in food simulants at 259 nm. Lastly, the release kinetics obtained based on released amount of GA as a function of time (days). For each certain film the test was carried out in quadruplicate with different flasks. 2.13. Statistical analysis The data analysis was performed using the IBM SPSS Statistics 21.0 software. In order to determine the significant differences at the 95% confidence level, Duncan'smultiple range tests used. 3. Results and discussion 3. 1. Film thickness The average value of films thickness illustrated in Table 2. As illustrated, C-G-GA films, particularly irradiated samples, had the lowest thickness among all formulations. It could be attributed to the possible hydrogen bonding between the NH3 groups of the Chitosan backbone and the -OH groups of Gallic acid. The reducing effect of hydrogen bonding on the volume of Chitosan films through decrease molecular mobility was reported in the literature[31]. In other samples Irradiation did not show significant effect on thickness, which could be the result of simultaneous formation and destruction of some bonds. In samples containing β-CD, UV radiation increased the thickness of the films. It may be attributed to destruction of composed hydrogen bonds between β-CD and polymers’ networks (Chitosan and Gelatin)[9,32]. In the same time, possible formation of further bonds within or between two polymers may lead to insignificant changes in thickness[21,33]. Suggested location for table 2 3. 2. Mechanical properties Tensile strength and elongation at break are the main mechanical properties of composite films. It can be observed that increasing the time of irradiation to 10 minutes leads to significant decrease in the TS value for all samples Table 2. Degradation of polymer at higher irradiation time may be the possible cause for such behavior. The photodegradation of Chitosan by UV irradiation has been reported by Sionkowska et al [22]. It can be observed that short UV irradiation time (5 min), reduced the TS of the samples except films containing β-CD which showed significant increase. This could be explained by formation of a more stable network by intermolecular bonds between β-CD and polymers. The Possibility of chemical banding (hydrogen bonds) between Gelatin and/or Chitosan and β-CD have reported by Dang et al [32] and Jingou 5
et al [34].In samples containing ethanol, the negative effect of 5 minutes irradiation on TS was more than those for 10 minutes treatment. It can be stated that plasticizing effect of incorporated OH groups of ethanol as well as simultaneous formation of loose hydrogen bonds at lower time is followed by destruction of these bonds or groups. Then formation of stronger bonds between polymer chains is occurred. Elongation at break (EB%) for films followed opposite trend compared with TS values. The opposite relation between TS and EB% values in UV treated films has reported previously [35]. 3. 3. Water vapor permeability To avoid moisture transfer between environment and packaged/ coated foods, WVP of the packaging/ coating materials must be kept as low as possible. The measured values of WVP of the fabricated films are shown in Table 2. UV irradiation for 10 minutes caused an increase in WVP values for almost all fabricated films (except films containing β-CD). These results are in accordance with the measured values for mechanical strength where irradiation for long times led to the weakening of the samples and it could be explained by breaking of some bonds during long-time of irradiation. In contrast, irradiation of samples containing β-CD led to reduction in WVP significantly. Meanwhile, the short time of irradiation, as compared to the longer periods, had a greater effect on the reduction of the WVP, which is in agreement with the obtained TS values of the films. Such behavior could be defined by formation of the new bonds between polymer and β-CD and/ or polymer network during short time irradiation and their breakage with increasing of irradiation time[[36], [37], [38], [39]]. In other words, further irradiation results in dominance of destruction of links over their formation. Bhat and Karim (2014), have stated that radiation of aqueous protein solutions generate free radicals like hydroxyl radicals (.OH), these compounds can affect some amino acids like aromatics which are involved in the formation of intermolecular covalent cross-linking bonds[20]. 3. 4. Color measurement Color is an essential factor that determines the quality of both coating and films. As shown in Table 2, UV irradiation significantly decreased the yellowness index (YI) and darkness (increase in L) of all samples. As the yellow color of Chitosan is result of the presence of β-1-4 linked 2-amino-2-deoxy-D-glucopyranose repeated units [40], decrease in yellowish and YI might be related to the destruction of these bonds, especially during the long time of treatment. The irradiated films tended to have a light appearance. Regarding samples containing β-CD and tween, the impact of short time irradiation on reducing darkness was more evident. Decrease in darkness during Irradiation is in contrast with Benbetaeeb et al, where they reported a reduction in L*value in electron beam irradiated Chitosan-Gelatin films that considered as the result of oxidative reactions during irradiation[36]. 3. 5. Release properties of GA The release trends of GA from the irradiated Chitosan-Gelatin composite films to ethanol as a fatty food simulant are shown in Figure 1. As can be seen, the slowest release of GA in non-irradiated films was detected for samples containing ethanol, which could be explained by the role of ethanol as a sufficient solvent of GA consequently more appropriate distribution of GA in such sample. On the opposite, samples with β-CD showed the fastest release. It can be explained by solubility of β-CD in polar solvent (ethanol) due to formation of hydrogen bonds between β-CD and ethanol. In addition, it is proved that these hydrogen bonds can cause structural deformation resulting in release of guest molecules (GA) from host β-CDs[41]. As illustrated for irradiated samples, UV treatment clearly led to increase in release rate (slope 6
of the curve), initial and final release percentage of GA to alcoholic medium. However, the amount of released GA from 5 minutes irradiated films containing β-CD was fewer compared with other irradiated samples. Such behavior could be the result of two effects of UV treatment in these films: help to bond between GA and β-CD molecules as well as contribution of β-CD to create linkage with film network, which is in agreement with mechanical properties where 5 minutes irradiated C-G-GA-β samples showed the highest TS values among all irradiated films. The percentage of accumulative released GA for C-G-GA and C-G-GA-β samples became higher with an increase of irradiation time from 5 to 10 minutes as a result of polymer degradation, while released GA from C-G-GA-T and C-G-GA-E samples which irradiated for 5 minutes were more than those with 10 minutes irradiation. It could be associated with formation of free radicals during short time of irradiation and their further cooperation in bonding formation through more treatment. Such results show that both destruction and formation of cross-links could occur in different parts of the samples during both times of UV treatment in polymers.
Suggested location for figure 1.
3.6. FTIR The FTIR spectra of fabricated samples are illustrated in Figure 2. Incorporation of β-CD to the films with GA caused the enhancement in the intensity of the band within the range of 1020–1030 cm−1 (stretching vibrations and bending vibrations of the phenolic group C–OH at 1400–1000 cm−1). Also, bands between 2700 cm−1and 3000 cm−1 (assigned to the aromatic ring) [24] and absorbance bands in the regions of 30003500 cm−1 (–OH stretching of hydroxyl groups and NH2 groups) became slightly wider for samples containing GA and β-CD. It also shows the enhanced hydrogen bonding action between Chitosan and βCD [12,34,42]. Concerning C-G-GA samples, irradiation weakened the absorption peak at approximately 1423 cm−1 (–OH bending) [34]. Applying UV treatment for 5 minutes weakened the peak and increasing the time of irradiation from 5 to 10 minutes resulted in removing that. In addition, increasing in irradiation time enhanced the sharpness (intensity) of main characteristic bonds at 1630 and 1530 cm−1 associated with C═O stretching (amide I) and N─H bending (amide II) of polymers, respectively [43]. The widest band related to the non-irradiated films especially between 3000-3500 cm−1which shows increase of –OH stretching of hydroxyl groups and NH2 groups [24,34,42]. Among C-G-GA-β samples, UV treatment for 5 minutes leads to formation of intensive peaks specially in 1400 and 1530, while the peak at 1423 cm−1 (– OH bending) disappeared.
Suggested location for figure 2.
3. 7. Scanning electron microscopy SEM micrographs of cross section of irradiated and non-irradiated samples of C-G-GA and C-G-GA-β films with different UV irradiation time are presented in Figure 3. While incorporated GA in C-G-GA films distributed in vast area, its dispersion is almost limited to the areas with β –CD in C-G-GA-β films. Minimum difference in percentage of released GA between non- irradiated and irradiated films with β-CD in 7
comparison with C-G-GA films could be explained by GA absorption into β –CD. Physical mixture of GA and β-CD observed as particles with irregular appearance which could be the result of different amounts of GA absorbed by β-CD particles [12,43]. Non-irradiated and short-time irradiated samples showed uniform and compact network with smooth surface, but fabricated films were exposed to the long-time of irradiation indicated some cracks in their structure. The reduction of tensile strength could be due to such uneven and discontinuities structure. Moreover, long time of UV treatment might induce network degradation. Rupture of covalent bands and degradation of polymers under ionizing radiation have been reported in the literature [36].
Suggested location for figure 3.
4. Conclusions Chitosan-Gelatin composite films including antioxidant, ethanol, tween and β-CD were fabricated. Incorporation of ethanol and tween decreased the film thickness significantly, however such effect did not observed for films containing β-CD. In the case of WVP, no significant effect was observed when ethanol and tween were added, however β-CD significantly increased the WVP. Similar results were observed in the case of mechanical strength, adding those additives with the exception of β-CD did not change the TS of the films. In all cases incorporation of those additives led to form of more yellow color in final products. In comparison with other additives, ethanol significantly deceased the release rate of GA to fatty food simulant. The highest release rate was observed for the films containing β-CD. Exposing an aqueous solution of Chitosan-Gelatin to different time of UV light (5, 10 min) performed in advance. From these observations, it is evident that radiation influenced the release properties of antioxidant. Although films containing β-CD had the highest release rate among all non-irradiated samples, UV treatments for 5 minutes could reduce the rate of antioxidant release. Long-time treatment (10min) reduced TS value and led to an uneven structure. The UV treatment increased the WVP in all samples with the exception of those containing β-CD, which showed the highest resistance against the diffusion of water vapor when irradiated for 5 minutes. Short-time (5min) irradiation also increased the TS of the films containing β-CD. As in our previous study it was noted that addition of β-CD had negative effect on mechanical properties of the gelatin -chitosan films[3], use of UV treatment was able to alleviate the problem to some extent. Besides, C-G-GA-β irradiated films showed better efficiency in WVP, darkness and release properties , and maximum accumulative amount of released GA was just about 20% after a couple of weeks. In order to these benefits, Incorporating of β-CD might be efficient candidate for fatty food preservation. Finally, application of UV irradiation could be appropriate tool to manipulate or modified various properties of composite films such as release rate of incorporated antioxidant, mechanical, and physicochemical characteristics, in order to our priorities.
Acknowledgments The University of Tehran is acknowledged for supporting this research. 8
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Figure captions Figure 1. Release of Gallic acid from fabricated films in 96% ethanol at 30 °C as a function of time. Subscripts show the time of irradiation: (a) C-G-GA, (b) C-G-GA-E, (C) C-G-GA- β, (d) C-G-GA-T. Figure 2. The FTIR spectra of fabricated C-G-GA and C-G-GA-β films. Subscripts show the time of irradiation. Figure 3. SEM micrographs of the cross-section of fabricated films with and without β-CD. Magnification of the figures is shown by scale bar. Subscripts show the time of irradiation.
13
Table captions Table 1. Abbreviated form of the name of different compositions of composite films Table 2. Thickness, WVP, optical and mechanical properties of irradiated and non-irradiated fabricated films
14
Figure 1.
15
Figure 2.
16
Figure 3.
17
Table 1. Film Name Chitosan- Gelatin- Gallic acid Chitosan- Gelatin- Gallic acid- Ethanol Chitosan- Gelatin- Gallic acid- Tween Chitosan- Gelatin- Gallic acid- β-cyclodextrin
Abbreviation C-G-GA C-G-GA-E C-G-GA-T C-G-GA-β
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Table 2.
Film samples
C-G-GA
C-G-GA-E
C-G-GA-T
C-G-GA-β
Irradiatio n time (min)
Thickness( µm)
TS(Mpa)
0
110±8.00bc
35.93±2.21a
5
92±9.83a
10
EB%
L
WVP g/(m.Pa.S) ˣ1010
YI
70.28±3.47a
2.01±0.00a
15.29±2.42a
80.44±3.88b
27.59±1.42b
85.97±13.83a
3.26±0.00a
5.29±0.50b
94.25±0.76a
83±3.11a
15.84±3.27c
69.62±5.10a
2.20±0.00a
4..40±0.43b
95.71±0.95a
0
118±7.95bc
35.18±14.30a
66.54±4.30b
2.40±0.00a
19.22±0.68a
76.22±0.97c
5
113±8.61bc
12.24±4.43b
75.21±7.56b
1.48±0.06a
6.86±0.56b
92.14±0.89b
10
100±9.15ab
20.3±1.82ab
90.54±6.77a
3.31±1.40a
4.82±0.26c
94.71±0.48a
0
118±4.47bc
34.39±1.70a
66.34±1.60c
1.15±0.00a
20.53±0.38a
74.89±0.78c
5
121±8.99c
30.38±0.90a
91.1±6.20a
1.36±0.30a
8±1.20c
89.71±1.79a
10
119±6.42bc
15.62±2.70b
75.64±0.99b
1.36±0.30a
9.88±1.06b
86.71±0.75b
0
110±9.14bc
23.03±4.27b
69.59±2.66a
5.66±0.00b
16.89±2.17a
78.55±2.29c
5
114±7.40bc
33.77±1.38a
70.20±2.50a
1.3±0. 20a
4.42±0.42c
95.42±0.53a
10
116 ±8.62bc
21.19±1.09b
75.29±11.77a
2.11±0.00a
8.83±0.25b
89.85±0.69b
Values are given as mean ± standard deviation. Means with the same superscripted letter in the same column not significantly different at p < 0.05.
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Highlights 1- Chitosan-Gelatin composite films including antioxidant with and without β-cyclodextrin content were fabricated. 2- UV irradiation influenced the release properties of antioxidant. 3- UV irradiation could compensate the negative effect of β-cyclodextrin on the films integrity
S0141-8130(19)35068-8 https://doi.org/10.1016/j.ijbiomac.2019.10.066 BIOMAC 13560
To appear in:
International Journal of Biological Macromolecules
Received Date: Revised Date: Accepted Date:
3 July 2019 24 September 2019 7 October 2019
Please cite this article as: M. Rezaee, G. Askari, Z. EmamDjomeh, UV-irradiated gelatin-chitosan bio-based composite film, physiochemical features and release properties for packaging applications, International Journal of Biological Macromolecules (2019), doi: https://doi.org/10.1016/j.ijbiomac.2019.10.066
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© 2019 Published by Elsevier B.V.
UV-irradiated gelatin-chitosan bio-based composite film, physiochemical features and release properties for packaging applications
Mahsa Rezaee1, Gholamreza Askari1*, Zahra EmamDjomeh1, 1
Transport Phenomena Laboratory (TPL) Department of Food Science and Technology, University College of Agriculture and Natural Resources, University of Tehran, Karaj, Iran *- corresponding author: Gholamreza Askari, Email: [email protected]
Abstract In this study, Chitosan-Gelatin biodegradable films loaded with Gallic acid (GA) were developed. Ethanol, tween, and β-Cyclodextrin (β-CD) were used to enhance the distribution of GA in the film forming solutions. The effect of the exposure time of UV irradiation on the color, mechanical properties (TS, %E), water vapor permeability (WVP) microstructure and release rate of GA to fatty food simulants investigated. UV irradiation reduced darkness of the films markedly and increased the release rate of the GA. Among the non-irradiated samples the highest and lowest release rate were obtained for the films containing β-CD and ethanol respectively. On the other hand incorporation of β-CD could reduce the increasing effect of UV light on release rate. The UV irradiation for 10 minutes reduced mechanical strength and barrier properties of the film against water vapor. Microscopic analysis, showed the consistent and uniform microstructure. However, long-term radiation caused some cracks in the films network. The UV irradiation weakened the _OH bending and enhanced the amide I amide II.
Key words: UV irradiation; biodegradable; chitosan; gelatin; Gallic acid; β-Cyclodextrin 1. Introduction Due to the ecological concerns of plastics as the main packaging materials, Bio-based polymers have been considered recently because of their biodegradable nature and consequently environmental friendly characteristics[1]. Nowadays, active packaging has become more popular. There are two kinds of active packaging systems: active-releasing which target compounds added into the headspace or the packaged food, and activescavenging systems, which undesired compounds removed permanently from the food or its environment[2]. Antioxidants are the most widely used components in the first type of active packaging systems. Undesirable properties of synthetic antioxidants such as toxicity has led to a growing tendency to use natural, nontoxic antioxidants [3]. Gallic Acid (GA) is a common natural antioxidant that generally found in many sources such as grape, tea, staghorn sumac, and strawberry. Based on some studies, GA shows anticancer and anti-tumor properties in addition to its antioxidant and bacteriostatic characteristics[4,5].
1
Similar to other natural antioxidants, GA is also sensitive to environmental stresses such as pH, light, and elevated temperatures[6]. On the other hand researchers are looking for ways to control the release of biologically active compounds to the packaged foods, since the release rate and its pattern has a major effect on the efficiency of drugs and biocompounds. The desired result can be achieved when the biological compound is released in controlled manner [7]. In such cases, cyclodextrins (CD’s) are one of the best vehicles to protect GA and preserve its structural integrity through forming inclusion complexes(IC’s)[8]. Cone-shaped structure of CD’s with hydrophobic cavity and hydrophilic surface make them suitable to form IC’s with guest molecules such as bioactive compounds[9]. In addition, it has shown that CD’s are promising agents to ease controlled release of hydrophobic molecules[10]. Meanwhile, Chitosan and Gelatin as biodegradable, nontoxic and compatible biopolymers are good candidates for making packaging materials [11]. They can form excellent poly electric complex with each other because of their potentially opposite charges at specific conditions[12]. There are electrostatic interactions and hydrogen bonds, mainly between the amino groups of Chitosan and the carboxyl groups of Gelatin, and between polymer chains involving their carbonyl, hydroxyl and amino groups [13]. Cross-linking is a known method for modification of biopolymers. It has been reported that cross- linking amplified the characteristics of Chitosan-based biomaterials[14]. Cross- linking could be done through chemical agents, enzymatic treatment or physical methods[15–17]. Due to many benefits such as simplicity, ecofriendly, low cost, and additive free process, radiation induced crosslinking gained more attention in recent years[18,19]. In comparison with the other ionizing radiation, UV radiation is suitable for modifying physical and mechanical properties of protein-based films due to moderate and nondestructive effect of UV light. Some studies reported improvement in cross-linking of collagen and Gelatin, in addition to modifying physical and mechanical properties of protein-based films by UV radiation [20]. Up to know, few works have considered the effects of UV irradiation on some characteristics of edible films such as mechanical and physicochemical properties [20–22]. To the best of our knowledge, no previous study has been reported the effect of UV treatment on release behavior of antioxidant agent from carbohydrate-protein composite films, particularly Chitosan-Gelatin films. Although many studies have been done about the mixture of antioxidant and β-CD, its characteristics and effects on physical and release properties in irradiated edible films has not been investigated [7,23–26]. The objective of current research was to determine the effect of some additives including ethanol, tween and β-CD and different time of UV irradiation on chemical, physical and morphological properties of active films based on blends of Gelatin and Chitosan incorporating GA. Release profile of GA into ethanol solution as the fatty food simulant was also investigated.
2. Materials and methods 2.1. Raw materials Two main film forming materials were medium molecular weight Chitosan (Sigma-Aldrich, Germany) and Gelatin (Merck, Darmstadt, Germany). An emulsifier agent, Plasticizer, and antioxidant used were Tween 20 (Merck, Germany), extra pure (≥99 %) glycerol (Merck, Germany), and Gallic acid (monohydrate, extra pure, molecular weight of 188.14 g mol-1, (Sigma-Aldrich, USA). Glacial acetic acid obtained from Merck, 2
Germany. Ethanol 96% was obtained from Alcohol Pars Co. (Iran). β-CD obtained from Sigma-Aldrich (USA). 2.2. Preparation of Chitosan- Gelatin solution Preparation of film forming solution carried out according to our previous work [3]. At First, in order to achieve 1 % (w/v) Chitosan film forming solution, certain amount of Chitosan powder dissolved in 1 % (v/v) solution of aqueous acetic acid at 40 °C with magnetic stirring. Afterwards, glycerol (10% w/w dry matter) added to the solution under constant rate of stirring. The pH of the solution determined about 4 ± 0.2. Preparation of Gelatin film forming solution (3 % w/v) was performed by adding Gelatin powder to distilled water under stirring condition at 50 °C for 30 min until complete dissolution. Finally, glycerol (10 % w/w dry matter) was incorporated to Gelatin solution. The measured pH was about 5 ± 0.2. In order to obtain clear film-forming solution, each solution was separately filtered through filter paper (No. 4). The final composite film forming solution achieved by mixing equal weights of the Gelatin and Chitosan solutions and stirring for 30 min. To obtain polyelectrolyte complex between Chitosan and Gelatin the pH value of the solution adjusted at 5.6 using sodium hydroxide solution. 2.3. UV irradiation process Time-controlled irradiation of composite solution was carried out using UV lamp (TUV 6W G6T5, Philips, Poland). After pouring composite solution into a graduated cylinder, UV lamp placed in contact with solution into the quartz cylinder. The solution was circulated by blowing air bubbles in order to obtain uniform irradiation. The whole set were in a chamber which its temperature was controlled by a fan at 25ͦ C (ambient temperature). Irradiation time was varied for 0, 5, and 10 minutes. 2.4. Antioxidant addition GA was added to the resulting film forming solution at a concentration of 1 % (g/g total dry weight of film). Antioxidant addition carried out in four different ways: adding GA with ethanol as the solvent of antioxidant, with Tween 20 as an emulsifier agent at concentrations of 100 % (w/w) relative to the weight of the Gallic acid, and adding singular powder of GA and its physical mixture with β-CD to the solution. Homogenization of all the aqueous dispersions carried out at 13,000 rpm for 3 min using the Ultra Turrax (T25, IKA, Germany) homogenizer. 2.5. Film formation and conditioning A volume of 85 mL of the final film forming solutions were poured into Petri dishes with 15 cm diameter and dried at 30 °C for 5 days. The resulting films were peeled off from the casting plates. Prior to any characterization, films conditioned at room temperature over a saturated solution of MgCl2 (32 % RH) in desiccators for 72 h. Name of the film samples and their abbreviation are listed in Table 1. Suggested location for table 1 2.6. Films thickness. The thickness of the films was measured using a micrometer (Mitutoyo, ID C112PM, Kawasaki-shi, Japan). Mean values of five measurements were used for thickness determination and further calculations. 2.7. Color measurements.
3
Optical properties of the samples were measured using colorimeter (Minolta CR-300 Series, Minolta, Japan). Color parameters of the films including L for luminosity and b for tonalities from yellow to blue were measured. The reflection spectra of the samples placed on the surface of a standard white plate resulted in the coordinates CIE Lab L, a, and b. The yellowness indexes (YI) of samples was determined using the following equation [27]. YI = 142.86 bL-1 The measurement for all samples for each formulation was carried out in triplicate. 2.8. Mechanical properties Mechanical properties of the fabricated films, including Tensile strength (TS) and elongation at break (%EB) were measured using the texture analyzer (Testometric M350-10CT, Rochdale, Lancs., England) according to the ASTM method D882 [28] with slight modifications described by Weijie, et al [29]. Samples were cut into rectangular shapes with width and length of 10 mm and 80 mm, respectively. Testing of the samples were performed through placing them in the film-extension grips with an initial distance of 50 mm at the speed of 50 mm/min until breakage. Following equations used to calculate TS and EB: TS (MPa) = (Maximum force (N) / Film thickness (m) × Film width (m)) × 10-6 EB (%) = (Final length of the film sample at the point of break )m)/Initial length of the film sample (m)) × 100 2.9. Water vapor permeability (WVP) The water vapor permeability of the films measured according to the standard ASTM E96–95 [30]. The containers which made of glass with internal diameter of 12.3 mm filled with anhydrous calcium chloride to achieve 0% RH. The film samples were placed on the mouth of the containers and sealed with molten paraffin and parafilm. Then, the containers were introduced into a desiccator containing saturated solution of sodium chloride with 75% relative humidity at room temperature and weighted at intervals of 12 h during 13 days to record their mass gain. Linear regression was used to determine the slope of mass changes versus time. WVP was calculated by the following equation: WVTR = Curve slope (g.s-1.mm-2)/Film area (mm2) WVP = Thickness (mm) × WVTR/Pressure difference (KPa) Where WVTR is the transmission rate of water vapor through the film and calculated from the slope of the weight changes versus time using linear regression. 2.10. Fourier transform infrared spectroscopy (FTIR) FTIR spectra of all films was carried out by a FT-IR spectrometer (PerkinElmer instruments, USA). The pellets of samples prepared with KBr powder and the IR spectra were recorded in the wavenumber range of 4000–450 cm−1. 2.11. Microstructure
4
Scanning electron microscope (LEO 1455 VP, England) was used to examine microstructure of the films. In order to obtain brittle structure of films to prepare cross-sections, they immersed in liquid nitrogen in advance. 2.12. Antioxidant Release The release properties of GA, was studied according to the method described in our previous work [12]. Briefly, the film sample (2 cm × 2 cm) immersed into 50 mL of ethanol 96%, as fatty food simulant, in a 100 mL Erlenmeyer flask, and shake at 70 rpm and 30 °C. At regular time intervals 4 mL of solutions was collected. Measuring the amount of released GA in the collected samples was performed by a UV–visible spectrophotometer (Spectrum Instruments, Germany) at a wavelength of 259 nm and each collected sample was returned to the flask after each measurement. The simulant containing GA free film sample was used as blank. The standard curve was achieved by measuring the absorbance of different specified concentrations of GA in food simulants at 259 nm. Lastly, the release kinetics obtained based on released amount of GA as a function of time (days). For each certain film the test was carried out in quadruplicate with different flasks. 2.13. Statistical analysis The data analysis was performed using the IBM SPSS Statistics 21.0 software. In order to determine the significant differences at the 95% confidence level, Duncan'smultiple range tests used. 3. Results and discussion 3. 1. Film thickness The average value of films thickness illustrated in Table 2. As illustrated, C-G-GA films, particularly irradiated samples, had the lowest thickness among all formulations. It could be attributed to the possible hydrogen bonding between the NH3 groups of the Chitosan backbone and the -OH groups of Gallic acid. The reducing effect of hydrogen bonding on the volume of Chitosan films through decrease molecular mobility was reported in the literature[31]. In other samples Irradiation did not show significant effect on thickness, which could be the result of simultaneous formation and destruction of some bonds. In samples containing β-CD, UV radiation increased the thickness of the films. It may be attributed to destruction of composed hydrogen bonds between β-CD and polymers’ networks (Chitosan and Gelatin)[9,32]. In the same time, possible formation of further bonds within or between two polymers may lead to insignificant changes in thickness[21,33]. Suggested location for table 2 3. 2. Mechanical properties Tensile strength and elongation at break are the main mechanical properties of composite films. It can be observed that increasing the time of irradiation to 10 minutes leads to significant decrease in the TS value for all samples Table 2. Degradation of polymer at higher irradiation time may be the possible cause for such behavior. The photodegradation of Chitosan by UV irradiation has been reported by Sionkowska et al [22]. It can be observed that short UV irradiation time (5 min), reduced the TS of the samples except films containing β-CD which showed significant increase. This could be explained by formation of a more stable network by intermolecular bonds between β-CD and polymers. The Possibility of chemical banding (hydrogen bonds) between Gelatin and/or Chitosan and β-CD have reported by Dang et al [32] and Jingou 5
et al [34].In samples containing ethanol, the negative effect of 5 minutes irradiation on TS was more than those for 10 minutes treatment. It can be stated that plasticizing effect of incorporated OH groups of ethanol as well as simultaneous formation of loose hydrogen bonds at lower time is followed by destruction of these bonds or groups. Then formation of stronger bonds between polymer chains is occurred. Elongation at break (EB%) for films followed opposite trend compared with TS values. The opposite relation between TS and EB% values in UV treated films has reported previously [35]. 3. 3. Water vapor permeability To avoid moisture transfer between environment and packaged/ coated foods, WVP of the packaging/ coating materials must be kept as low as possible. The measured values of WVP of the fabricated films are shown in Table 2. UV irradiation for 10 minutes caused an increase in WVP values for almost all fabricated films (except films containing β-CD). These results are in accordance with the measured values for mechanical strength where irradiation for long times led to the weakening of the samples and it could be explained by breaking of some bonds during long-time of irradiation. In contrast, irradiation of samples containing β-CD led to reduction in WVP significantly. Meanwhile, the short time of irradiation, as compared to the longer periods, had a greater effect on the reduction of the WVP, which is in agreement with the obtained TS values of the films. Such behavior could be defined by formation of the new bonds between polymer and β-CD and/ or polymer network during short time irradiation and their breakage with increasing of irradiation time[[36], [37], [38], [39]]. In other words, further irradiation results in dominance of destruction of links over their formation. Bhat and Karim (2014), have stated that radiation of aqueous protein solutions generate free radicals like hydroxyl radicals (.OH), these compounds can affect some amino acids like aromatics which are involved in the formation of intermolecular covalent cross-linking bonds[20]. 3. 4. Color measurement Color is an essential factor that determines the quality of both coating and films. As shown in Table 2, UV irradiation significantly decreased the yellowness index (YI) and darkness (increase in L) of all samples. As the yellow color of Chitosan is result of the presence of β-1-4 linked 2-amino-2-deoxy-D-glucopyranose repeated units [40], decrease in yellowish and YI might be related to the destruction of these bonds, especially during the long time of treatment. The irradiated films tended to have a light appearance. Regarding samples containing β-CD and tween, the impact of short time irradiation on reducing darkness was more evident. Decrease in darkness during Irradiation is in contrast with Benbetaeeb et al, where they reported a reduction in L*value in electron beam irradiated Chitosan-Gelatin films that considered as the result of oxidative reactions during irradiation[36]. 3. 5. Release properties of GA The release trends of GA from the irradiated Chitosan-Gelatin composite films to ethanol as a fatty food simulant are shown in Figure 1. As can be seen, the slowest release of GA in non-irradiated films was detected for samples containing ethanol, which could be explained by the role of ethanol as a sufficient solvent of GA consequently more appropriate distribution of GA in such sample. On the opposite, samples with β-CD showed the fastest release. It can be explained by solubility of β-CD in polar solvent (ethanol) due to formation of hydrogen bonds between β-CD and ethanol. In addition, it is proved that these hydrogen bonds can cause structural deformation resulting in release of guest molecules (GA) from host β-CDs[41]. As illustrated for irradiated samples, UV treatment clearly led to increase in release rate (slope 6
of the curve), initial and final release percentage of GA to alcoholic medium. However, the amount of released GA from 5 minutes irradiated films containing β-CD was fewer compared with other irradiated samples. Such behavior could be the result of two effects of UV treatment in these films: help to bond between GA and β-CD molecules as well as contribution of β-CD to create linkage with film network, which is in agreement with mechanical properties where 5 minutes irradiated C-G-GA-β samples showed the highest TS values among all irradiated films. The percentage of accumulative released GA for C-G-GA and C-G-GA-β samples became higher with an increase of irradiation time from 5 to 10 minutes as a result of polymer degradation, while released GA from C-G-GA-T and C-G-GA-E samples which irradiated for 5 minutes were more than those with 10 minutes irradiation. It could be associated with formation of free radicals during short time of irradiation and their further cooperation in bonding formation through more treatment. Such results show that both destruction and formation of cross-links could occur in different parts of the samples during both times of UV treatment in polymers.
Suggested location for figure 1.
3.6. FTIR The FTIR spectra of fabricated samples are illustrated in Figure 2. Incorporation of β-CD to the films with GA caused the enhancement in the intensity of the band within the range of 1020–1030 cm−1 (stretching vibrations and bending vibrations of the phenolic group C–OH at 1400–1000 cm−1). Also, bands between 2700 cm−1and 3000 cm−1 (assigned to the aromatic ring) [24] and absorbance bands in the regions of 30003500 cm−1 (–OH stretching of hydroxyl groups and NH2 groups) became slightly wider for samples containing GA and β-CD. It also shows the enhanced hydrogen bonding action between Chitosan and βCD [12,34,42]. Concerning C-G-GA samples, irradiation weakened the absorption peak at approximately 1423 cm−1 (–OH bending) [34]. Applying UV treatment for 5 minutes weakened the peak and increasing the time of irradiation from 5 to 10 minutes resulted in removing that. In addition, increasing in irradiation time enhanced the sharpness (intensity) of main characteristic bonds at 1630 and 1530 cm−1 associated with C═O stretching (amide I) and N─H bending (amide II) of polymers, respectively [43]. The widest band related to the non-irradiated films especially between 3000-3500 cm−1which shows increase of –OH stretching of hydroxyl groups and NH2 groups [24,34,42]. Among C-G-GA-β samples, UV treatment for 5 minutes leads to formation of intensive peaks specially in 1400 and 1530, while the peak at 1423 cm−1 (– OH bending) disappeared.
Suggested location for figure 2.
3. 7. Scanning electron microscopy SEM micrographs of cross section of irradiated and non-irradiated samples of C-G-GA and C-G-GA-β films with different UV irradiation time are presented in Figure 3. While incorporated GA in C-G-GA films distributed in vast area, its dispersion is almost limited to the areas with β –CD in C-G-GA-β films. Minimum difference in percentage of released GA between non- irradiated and irradiated films with β-CD in 7
comparison with C-G-GA films could be explained by GA absorption into β –CD. Physical mixture of GA and β-CD observed as particles with irregular appearance which could be the result of different amounts of GA absorbed by β-CD particles [12,43]. Non-irradiated and short-time irradiated samples showed uniform and compact network with smooth surface, but fabricated films were exposed to the long-time of irradiation indicated some cracks in their structure. The reduction of tensile strength could be due to such uneven and discontinuities structure. Moreover, long time of UV treatment might induce network degradation. Rupture of covalent bands and degradation of polymers under ionizing radiation have been reported in the literature [36].
Suggested location for figure 3.
4. Conclusions Chitosan-Gelatin composite films including antioxidant, ethanol, tween and β-CD were fabricated. Incorporation of ethanol and tween decreased the film thickness significantly, however such effect did not observed for films containing β-CD. In the case of WVP, no significant effect was observed when ethanol and tween were added, however β-CD significantly increased the WVP. Similar results were observed in the case of mechanical strength, adding those additives with the exception of β-CD did not change the TS of the films. In all cases incorporation of those additives led to form of more yellow color in final products. In comparison with other additives, ethanol significantly deceased the release rate of GA to fatty food simulant. The highest release rate was observed for the films containing β-CD. Exposing an aqueous solution of Chitosan-Gelatin to different time of UV light (5, 10 min) performed in advance. From these observations, it is evident that radiation influenced the release properties of antioxidant. Although films containing β-CD had the highest release rate among all non-irradiated samples, UV treatments for 5 minutes could reduce the rate of antioxidant release. Long-time treatment (10min) reduced TS value and led to an uneven structure. The UV treatment increased the WVP in all samples with the exception of those containing β-CD, which showed the highest resistance against the diffusion of water vapor when irradiated for 5 minutes. Short-time (5min) irradiation also increased the TS of the films containing β-CD. As in our previous study it was noted that addition of β-CD had negative effect on mechanical properties of the gelatin -chitosan films[3], use of UV treatment was able to alleviate the problem to some extent. Besides, C-G-GA-β irradiated films showed better efficiency in WVP, darkness and release properties , and maximum accumulative amount of released GA was just about 20% after a couple of weeks. In order to these benefits, Incorporating of β-CD might be efficient candidate for fatty food preservation. Finally, application of UV irradiation could be appropriate tool to manipulate or modified various properties of composite films such as release rate of incorporated antioxidant, mechanical, and physicochemical characteristics, in order to our priorities.
Acknowledgments The University of Tehran is acknowledged for supporting this research. 8
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Figure captions Figure 1. Release of Gallic acid from fabricated films in 96% ethanol at 30 °C as a function of time. Subscripts show the time of irradiation: (a) C-G-GA, (b) C-G-GA-E, (C) C-G-GA- β, (d) C-G-GA-T. Figure 2. The FTIR spectra of fabricated C-G-GA and C-G-GA-β films. Subscripts show the time of irradiation. Figure 3. SEM micrographs of the cross-section of fabricated films with and without β-CD. Magnification of the figures is shown by scale bar. Subscripts show the time of irradiation.
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Table captions Table 1. Abbreviated form of the name of different compositions of composite films Table 2. Thickness, WVP, optical and mechanical properties of irradiated and non-irradiated fabricated films
14
Figure 1.
15
Figure 2.
16
Figure 3.
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Table 1. Film Name Chitosan- Gelatin- Gallic acid Chitosan- Gelatin- Gallic acid- Ethanol Chitosan- Gelatin- Gallic acid- Tween Chitosan- Gelatin- Gallic acid- β-cyclodextrin
Abbreviation C-G-GA C-G-GA-E C-G-GA-T C-G-GA-β
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Table 2.
Film samples
C-G-GA
C-G-GA-E
C-G-GA-T
C-G-GA-β
Irradiatio n time (min)
Thickness( µm)
TS(Mpa)
0
110±8.00bc
35.93±2.21a
5
92±9.83a
10
EB%
L
WVP g/(m.Pa.S) ˣ1010
YI
70.28±3.47a
2.01±0.00a
15.29±2.42a
80.44±3.88b
27.59±1.42b
85.97±13.83a
3.26±0.00a
5.29±0.50b
94.25±0.76a
83±3.11a
15.84±3.27c
69.62±5.10a
2.20±0.00a
4..40±0.43b
95.71±0.95a
0
118±7.95bc
35.18±14.30a
66.54±4.30b
2.40±0.00a
19.22±0.68a
76.22±0.97c
5
113±8.61bc
12.24±4.43b
75.21±7.56b
1.48±0.06a
6.86±0.56b
92.14±0.89b
10
100±9.15ab
20.3±1.82ab
90.54±6.77a
3.31±1.40a
4.82±0.26c
94.71±0.48a
0
118±4.47bc
34.39±1.70a
66.34±1.60c
1.15±0.00a
20.53±0.38a
74.89±0.78c
5
121±8.99c
30.38±0.90a
91.1±6.20a
1.36±0.30a
8±1.20c
89.71±1.79a
10
119±6.42bc
15.62±2.70b
75.64±0.99b
1.36±0.30a
9.88±1.06b
86.71±0.75b
0
110±9.14bc
23.03±4.27b
69.59±2.66a
5.66±0.00b
16.89±2.17a
78.55±2.29c
5
114±7.40bc
33.77±1.38a
70.20±2.50a
1.3±0. 20a
4.42±0.42c
95.42±0.53a
10
116 ±8.62bc
21.19±1.09b
75.29±11.77a
2.11±0.00a
8.83±0.25b
89.85±0.69b
Values are given as mean ± standard deviation. Means with the same superscripted letter in the same column not significantly different at p < 0.05.
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Highlights 1- Chitosan-Gelatin composite films including antioxidant with and without β-cyclodextrin content were fabricated. 2- UV irradiation influenced the release properties of antioxidant. 3- UV irradiation could compensate the negative effect of β-cyclodextrin on the films integrity