Physicochemical properties of the edible films from the blends of high methoxyl apple pectin and chitosan

Accepted Manuscript Physicochemical properties of the edible films from the blends of high methoxyl apple pectin and chitosan

Heba G.R. Younis, Guohua Zhao PII: DOI: Reference:

S0141-8130(19)30136-9 https://doi.org/10.1016/j.ijbiomac.2019.03.096 BIOMAC 11926

To appear in:

International Journal of Biological Macromolecules

Received date: Revised date: Accepted date:

6 January 2019 5 March 2019 14 March 2019

Please cite this article as: H.G.R. Younis and G. Zhao, Physicochemical properties of the edible films from the blends of high methoxyl apple pectin and chitosan, International Journal of Biological Macromolecules, https://doi.org/10.1016/j.ijbiomac.2019.03.096

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ACCEPTED MANUSCRIPT Physicochemical properties of the edible films from the blends of high methoxyl apple pectin and chitosan Heba G. R. Younis a,b, Guohua Zhao a,c* College of Food Science, Southwest University, Chongqing, 400715, China

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Agricultural Engineering Department, Faculty of Agriculture, Cairo University, Giza

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Chongqing Engineering Research Centre of Regional Foods, Chongqing 400715,

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12613, Egypt

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People’s Republic of China

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Corresponding author College of Food Science, Southwest University

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2 Tiansheng Road, Chongqing, 400715, PR China

Fax: +86 23 68 25 19 47

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Tel.: +86 23 68 25 19 02

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E-mail address: [email protected] (G. Zhao)

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ACCEPTED MANUSCRIPT ABSTRACT Chitosan (CH) and pectin (PE) are considered as promising biomaterials in developing eco-friendly films due to their film-forming, biodegradable, and non-toxic characteristics, the films from pure CH or PE have obvious defects such as poor barrier and mechanical

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properties. In this study, the blend films of CH and PE at varying mass ratios were

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characterized. Structurally, numerous small pores evenly distributed in PE film while big

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caves unevenly scattered in CH film. CH film is semicrystalline but PE and blend films are totally amorphous, the two individual films presented comparable values in water

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content and solubility to blend films. The CH film showed lower water vapor

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permeability and surface wettability and these parameters of the blend films decreased with CH level, the blend films exhibited high transparence as PE film did, which is much

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higher than that of CH film. Mechanically, the PE film presented higher values in

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stretchability and tensile strength than CH film. Moreover, in a different blending ratios, synergistic effects were found with several characters of the CH/PE blend film, especially

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in transparence and mechanical properties. These synergistic effects were ascribed to the

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intermolecular electrostatic interactions between CH and PE. Keywords: Biodegradable film, additive effects, synergistic effects, intermolecular

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interactions, transparence

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ACCEPTED MANUSCRIPT 1. Introduction Plastic films made from non-renewable resources often cause environmental problems after disposal. To this end, researches are increasingly focusing on alternative materials with aims to reduce the use of nonbiodegradable and non-renewable films [1,2]. In recent

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years, the innovation of packaging materials has focused more on the films from

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biomaterials, which can be used as substitutes for synthetic polymers. Generally,

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biomaterials are non-toxic, biocompatible, renewable and always present reasonable filmforming ability. Thus, they are important for the food safety, especially the issues resulted

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from the migration of chemicals from plastic packaging materials into foods. The

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common biomaterials applied in formulating bridgeable films include polysaccharides, protein, lipids and the combination of these components. Among these materials,

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polysaccharides are usually cheaper than others, such as pectin (PE), starch, alginate,

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cellulose, chitosan (CH) and agar [3-5].

PE and CH are promising materials that could be used for the development of edible

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coatings and films on an industrial level due to their film-forming, biodegradable and

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non-toxic nature. Specifically, on one hand, CH-based film was proven to possess good antimicrobial activity as well as excellent barrier to gases (CO2 and O2) and good

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mechanical performance [6-9]. However, its brittleness substantially limited its actual application [10,11]. On the other hand, PE-based film was evidenced with excellent oxygen barring capacity, but its major defects lie in the weak moisture barrier and poor mechanical performance [12,13]. In essence, the physicochemical properties of a film highly depended on the intermolecular interaction occurring in the film matrix. The tensile strength of the

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ACCEPTED MANUSCRIPT resultant film is directly proportional to the intermolecular interaction [14,15]. In view of this principle, it is hypothesized that the blending of PE and CH certainly result in a better film than their individual constituents, especially in terms of tensile strength. This is based on the fact that, at an appropriate pH, PE and CH molecules are oppositely

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charged and the electrostatic attraction between them will favor the intermolecular

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interaction within the film matrix and thus enhance the film properties. Therefore, PE and

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CH were often blended to generate various composite materials, such as Chitosan/starch [6,16,17]. Chitosan/Honey/Gly [18], Chitosan/gelatin [8,19], Chitosan/alginates [20],

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Chitosan/collagen [10], Pectin/chitosan [21], and Pectin/alginate [13]. However, the

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specific PE or CH materials applied in these cases and varied from case to case, according to the deacetylation degree of CH and degree of esterification of PE.

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Consequently, the CH becomes more hydrophilic with the increase in its deacetylation

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degree [22], while the hydrophobicity of PE is enhanced when its degree of esterification increased [23]. In this sense, the selection of two polymers is crucial to their

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compatibility in the resultant film and thus finally affects the physicochemical properties

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of the composite materials. Unfortunately, this was not well addressed in the previous literature. As theoretically supposed, CH is liable to be blended with a high methoxyl PE,

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which certainly benefits the performance of the composite materials. Therefore, with an aim to develop a better edible film, the mechanical, barrier, and optical properties as well as structural characters of blend films from a high methoxyl apple PE and CH were investigated in the present study. 2. Materials and methods 2.1. Materials

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ACCEPTED MANUSCRIPT High methoxyl apple pectin (PE, with a degree of esterification of 64%) from apple peel and chitosan (CH, with a deacetylation degree 67.9%) were purchased from Sea Bioengineering company (Tsinan, China). Tween 80, glycerol and citric acid monohydrate are of analytical grade and obtained from Keshi Reagent Chemical Co. Ltd.

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(China).

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2.2. Film preparation

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Edible films were prepared by a casting method according to our previous method [24]. Specifically, fifteen grams of polysaccharide matrix (PE, CH or thereof blend) were

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dissolved in 1.0 L water containing 1% (w/w) citric acid by exhaustively agitating at 80

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°C until a clear solution was obtained. After the resultant solution cooled to 30 °C, glycerol (1.6%, w/w) and (0.5%, w/w) tween 80 were added. The mixture was

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homogenized for 10 min at 600 rpm to obtain the film-forming solution. Subsequently,

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the film-forming solution (200 mL) was cast onto a square porcelain plate (30 cm × 20 cm) and dried at 45 °C for 48 h. Finally, the dried film was peeled off and stored at room

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temperature for further evaluations. For blend films, three specimens were fabricated

respectively.

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with a PE-CH mass ratio of 1:1 (PE1-CH1), 1:2 (PE1-CH2) and 2:1 (PE2-CH1),

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2.3. Structural characterization of films 2.3.1. Fourier transform infrared spectroscopy The Fourier transform infrared (FT-IR) spectra of the films were recorded in an IR Spectrometer (Perkin–Elmer, Model 2000, USA). Prior to measurement, the films were converted to powder using nitrogen liquid, then an aliquot of the film (3.0 mg) was coground with KBr (90 mg) and dried at 105 oC for 2 h to make it a pellet. The

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ACCEPTED MANUSCRIPT measurements were performed at room temperature and the results were recorded in the frequency range of 4000-400 cm–1 with a resolution of 4 cm–1. 2.3.2. X-ray diffraction The X-ray diffraction pattern of each film was recorded by using an Ultima IV X-ray

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diffractometer (Rigaku Corporation, Japan) in the grazing incidence X-ray diffraction

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mode. The film powder was dried as did in FT-IR measurement and crushed to pass a

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100-mesh sieve. The equipment was operated at the CuKα radiation generated at 40 kV and 40 mA. The scanning scope of 2θ was from 3 to 80° at a scanning rate of 0.02°/min.

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The relative crystallinity was quantitatively calculated using Debye-Scherrer equation of

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L=(Kλ)/(βcosθ), where L is crystallinity size (nm), λ is the X-ray wavelength in nanometer (1.54060 nm for CuKα), β is the peak width of the diffraction peak profile at

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half maximum height resulting from small crystallite size (in radians), and K is a constant

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related to crystallite shape, which is normally taken as 0.9 [25]. 2.3.3. Microstructure characterization

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The surface and cross-section microstructures of films were observed by using JEOL

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JSM 5800LV SEM scanning electron microscopy (Phenom Pro, Phenom-World Ltd., USA). After coated with gold, the sample was observed under vacuum at 10 kV at a

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magnification of 2000 ×.

2.4. Determination of the physical properties of films 2.4.1. Film thickness The thickness was measured at nine randomly selected locations of each film using a micrometer caliper (Shan Brand, Guilin, China) with a precision of 0.001 mm. The

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ACCEPTED MANUSCRIPT resultant mean value was used in calculating its density, mechanical and barrier properties. 2.4.2. Water solubility (WS) and water content (WC) of the films The film moisture content was defined as the percentage of the mass loss of a film upon

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complete drying. It was determined according to the method reported by Šuput et al. [26]

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with some modifications. The specimen of a film was cut into square pieces (2 cm×2 cm)

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and accurately weighed (W0). Then, the film pieces were dried at 105 °C for 24 h and the dried samples were weighed again (W1). The water content (WC, g/100g) was calculated

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via the equation of WC= [(W0-W1)/W0] ×100 [9]. Water solubility was defined as the

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percentage of dry mass loss ratio when a film immersed in water. It was determined according to the method reported by Romani et al. [5], a portion of film pieces was

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immersed in 50 mL distilled water contained in a Petri dish. After covered with parafilm,

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the dish was continuously shaken (100 r/min) at 25 °C for 24 h. When the settled time elapsed, the content in the dish was filtered through a Xinxing filter paper (No 101). The

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film residues on the filter paper were carefully collected, dried at 105 °C for 24 h and

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then weighted (W2). The water solubility (WS, %) was calculated by using the equation of WS=[(W1-W2)/W1] ×100.

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2.4.3. Water vapor permeability and contact angle The water vapor permeability (WVP) was measured by using W3/060 Water Vapor Transmission Test System (Labthink Instruments Co., Ltd., China) using the gravimetric cup. A method based on ASTM Standard Method E96-00 [27] was applied. The films were tested up to 16 h and the WVP results were given by the system. The contact angle

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ACCEPTED MANUSCRIPT was recorded by using a contact angle meter (Powereach, Shanghai, Zhongchen digital technology apparatus co., Ltd. China). 2.4.4. Optical properties The opacity (OP) of the films was measured according to the method used by Li et al.

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[16]. The film specimen was trimmed into strips (9 mm × 40 mm), vertically put in a

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quartz cuvette of a spectrophotometer (UV-2450, Shimadzu Corporation, Japan) and the

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absorbance was measured at 600 nm. An empty cuvette was used as a blank control. The opacity (mm-1) of the film was calculated using the equation of OP= -logT600 / Thickness.

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The color parameters of the films were measured by using an Ultra Scan pro1166

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colorimeter (Hunterlab color quest XE, USA). The color parameters of L (luminosity), a (greenness; redness) and b (blueness; yellowness) were recorded. A standard plate CQX

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3477 was used for calibration and as the background for the following measurement.

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After that, Chroma (C*) and hue angle (hue) were calculated according to using the equations of C*= (a2 + b2)1/2 and hue = arctan (b/a) if a ˃ 0, or hue = arctan (b/a) + 180˚ if

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a < 0 [28, 29]. The color changes were expressed as the total color difference (ΔE) and

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calculated accordingly to the equation of ΔE = [(L*- L)2 + (a*- a)2 + (b*- b)2]1/2 [30, 31]. L*, a*, and b* are the values of color parameters of the standard plate. Namely, they are

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97.51, 0.1 and -0.19, respectively. 2.5. Determination of the mechanical properties of the films The mechanical properties of the films, in terms of tensile strength (TS), elongation at break (E) and Elastic modulus (EM), were measured by using an XLW (G)-PC Auto Tensile Tester (Labthink Instruments Co., Ltd., China) based on the ASTM D-882 method [32] with some modifications [33]. In this determination, the tested film

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ACCEPTED MANUSCRIPT specimen was cut into 100 mm × 10 mm strips. The initial grip separation and cross-head speed were set at 50 mm and 50 mm/s, respectively. Prior to the test, all film strips were conditioned at 25 °C and 70 % RH for 24 h. 2.6. Evaluation of the interaction effects between pectin and chitosan

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Regarding the blending effects of pectin and chitosan at different ratios, additive or

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non-additive effect could be concluded specifically. For the present binary system,

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assumedly, the property P of PE and CH was determined as PPE and PCH, respectively. For a specific blend in which PE and CH were mixed in mass percentages of FPE % and

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FCH % (FPE%=1-FCH%), its property P was experimentally determined as dP. In this case,

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the predicted property P of the blend (pP) can be obtained by proportionally combining PPE and PCH as pP=PPE×FPE+PCH×FCH. If pP approached dP, the blending effect was

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defined as additive. In contrast, a non-additive effect is concluded when pP significantly

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differed from dP [34]. The non-additive effect implies that the individual PE and CH in the blends influence each other.

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2.7. Statistical analysis

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Data were expressed as the mean ± standard deviation (SD) of three technical measurements unless specified. Turkey’s test and one-way analysis of variance

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(ANOVA) were used for multiple comparisons by SPSS statistic 20.0 software. The difference was statistically significant at p ˂ 0.05 level. 3. Results and discussion 3.1. Structural characterization of the films 3.1.1. FT-IR analysis

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ACCEPTED MANUSCRIPT Infrared spectroscopy was performed to discover and examine the interactions between the components (mainly PE and CH) in film matrix. This was done by comparing the characteristic region of FT-IR spectra among PE film, CH film and three blend films were shown in Fig. 1, for pure PE film, the absorption at 2929 cm-1 corresponds to C-H

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stretching of CH2 groups [35,36]. With the support from the results reported by Manrique

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and Lajol [37], the band at 1739 cm-1 is attributed to the absorption of the esterified

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carboxylic groups while the one at 1634 cm-1 has resulted from the absorption of the carboxylate anions. The shoulder at 1015 cm-1 is characteristic of C-O-C stretching

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vibrations in polygalacturonic acid [38-40]. The peak at 955 cm-1 is characteristic of

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rhamnogalacturonan I molecules (uronic acids) while the one at 923 cm-1 referred to the absorption of D-glucopyranosyl [35,38]. The shoulders at 890 cm-1 and 852 cm-1 are

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characteristic of α- and β- glycosidic linkage, respectively. The peak at 832 cm-1 is

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assigned to the absorption of a-D-mannopyranose. Regarding the pure CH film, the peak at 2927 cm-1 is assigned to symmetric C-H

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stretching [41, 42]. Due to the absence of any peak around 1730 cm-1 in the FTIR for

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pure chitosan materials, this is a peak at 1736 cm-1 is possibly ascribed to tween 80 added in the film matrix, originating from the C=O stretching of the ester group [43]. The peak

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at 1626 cm-1 indicated the vibrations of carbonyl group of the carboxylate ion COO stretching of secondary amide group (amide I) of the acetylated units of chitosan, while the one at 1530 cm-1 revealed to asymmetric deformation of Vibrations of N-H (N-acetylated residues of the amide II band [44, 45], but the peaks located at 1353 and 1395 cm-1 belong to N–H in plane deformation coupled with C=N stretching (amide III

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ACCEPTED MANUSCRIPT band) [46-48]. Pyranose ring was found at 955 cm-1 [51,19], While the C-N fingerprint band appears at 890 cm-1 [49]. When the FT-IR spectra of the blend films were compared to the individual CH and PE films, clear differences in intensity or shift were observed with the peaks of PE at 1634

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cm-1, 832 cm-1, and 764 cm-1 as well as the peak of CH at 1530 cm-1. In blend films, the

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intensities of peaks at 832 cm-1 and 764 cm-1 are proportional to the level of PE in the

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film matrix, while the intensity of the peak at 1541 cm-1 obviously was determined by the CH level. The changes in the intensity of these substrate specific peaks well reflected the

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blending ratio applied. In blend films, the PE specific peak at 1634 cm-1 was shifted to

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1626-1629 cm-1 and overlapped with the CH specific peak at 1626 cm-1. Meanwhile, the CH specific peak at 1530 cm-1 was shifted to 1541 cm-1 and it even disappeared in the

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blend film of PE2-CH1. These shifts were possibly the consequences of the polymer

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interaction in film matrix. In view of the above assignments of these peaks, the shifts and decreases in intensity should be an outcome of the interactions between the two polymers

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via their charged groups (-NH3+ and -COO-). These interactions not only partially

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consumed the charged groups but also certainly changed their original vibrations. 3.1.2. X-ray diffraction

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XRD analysis was used to explore the crystalline structure and evaluate the interactions of major film components (Fig. 2). Obviously, the PE-based film did not show any diffraction peak, while two peaks although not so intensive were observed with CH-based film at 24.4° (2θ) and 26.12° (2θ), respectively. In this sense, the PE-based film is totally in an amorphous form while the CH-based film is mostly amorphous and weakly crystallized. The crystalline nature of chitosan has been reported with several

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ACCEPTED MANUSCRIPT crystalline polymorphic forms, which shared an extended two-fold helical structure but differed in packing density and water content [50]. Regarding the blend films, they are totally amorphous irrespective of the blending ratio. This disclosed that strong interactions occurred between PE and CH in the matrix of the blend films, which

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destroyed the close packing for the CH molecules for the formation of regular crystallites

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[51]. This is in line with the results reported by Chetouani et al. [49] that the pectin

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addition to chitosan resulted in an amorphous material. Similar results were reported for the incorporation of starch or honey into CH-based films [17,18]. In considering the pH

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values of the forming solution (PE, 2.41; CH 2.10; PE1-CH1, 2.14; PE2-CH2, 2.20 and

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PE1-CH2, 2.12) and the pKa values of CH (6.5) and PE (3.5) [50], the chitosan chains were fully charged while the pectin chains were much less charged. In this scenario, it

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was speculated that hydrogen bonding, hydrophobic interactions as well as ionic

3.1.3. Film morphology

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complexation involve in the interactions of PE and CH in film matrix [52].

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The scanning electron microscopy (SEM) is often used to display the homogeneity and

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structural imperfects (pores, cracks, and fissures) existing in the films [21]. The SEM images for the surface and cross sections of films were illustrated in Fig. 3. In contrast,

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PE-based film presents a smoother surface than CH-based film. Obviously, many lumps present on the surface of the CH-based film are possibly resulting from the poor dispersibility of CH molecules in the film forming solutions leading to the formation of large agglomerates. Regarding the cross-section, the PE-based film showed a typical porous structure, in which holes with a similar sizes are evenly and individually distributed in the film matrix with some occasional combination or connection. This

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ACCEPTED MANUSCRIPT would be ascribed to the highly homogenous dispersibility of pectin molecules in the film forming solution and the strong gas-holding capacity possibly derived from its interfacial activity [53]. As for CH-based film, a heterogeneous cross-section was observed, in which big cavities irregularly distributed in a solid matrix.

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Regarding the blend films, the surface smoothness of PE1-CH1 and PE1-CH2 are

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comparable to that of PE-based film, while, clearly, the surface of PE2-CH1 was

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decorated with enormous embossments of serval micrometer-scale. This may be a consequence of the intensity difference in the molecular interactions involved in these

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three systems. Due to the presence of highly charged CH molecules and less charged PE

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molecules, the highest intermolecular interactions would happen with the system having more PE molecules, namely, PE2-CH1. The surficial embossments observed with PE2-

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CH1 were assumed as the coagulates of oppositely charged PE and CH molecules by

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electrostatic interaction. For the cross-sections of blend films, they were much more homogenous than the individual PE- or CH-based film. In contrast, they are less porous

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than PE-based film and were not founded with any big cavities as did in CH-based film.

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In terms of the overall evaluation of cross-section structure among three blend films, the PE1-CH2 gained the highest score due to its highest homogeneity and the absence of

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obvious cracks or fissures. In contrast to PE2-CH1, PE1-CH1 displayed approximately solid film matrix but present obvious longitudinal and transverse fissures. However, PE2CH1 showed a much looser film matrix, which contained a lot of small cavities. The microstructure and molecular interactions occurring in the film matrix can also be reflected by the viscosity of the film forming solutions. Due to the poor dispersibility of CH, the viscosity of the forming solution of CH film was too low to be accurately

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ACCEPTED MANUSCRIPT determined. The corresponding viscosity values of PE, PE1-CH1, PE2-CH1 and PE1CH2 were determined as 32.41 mPa. s, 22.97 mPa. s, 77.39 mPa. s, 103 mPa. s, and 80.49 mPa. s, respectively. this clearly attested that the viscosity of the film forming solution sharply increased at any blending ratio. In terms of viscosity, the strongest

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molecular interactions occurred in PE2-CH1, then followed by PE1-CH2 and PE1-CH1.

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3.2. Physical Properties of films

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3.2.1. The thickness and density of the films

The physical properties of blend films were summarized in Table 1. Clearly, the CH-

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based film (54.37 μm) presented a smaller thickness than PE-based film (87.00 μm),

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which was in agreement with the facts observed by SEM that PE-based film was much more porous than CH-based film. Similarly, Zuo et al. [54] witnessed a positive relation

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between the porosity and the thickness for zein-starch blend films. Furthermore, the

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results of Hallik et al. [55] showed that, for polyppyrrole-dodecysulfate films, a higher average diameter of pores always accompanied with a higher film thickness. Irrespective

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of blending ratio, the blend films of CH and PE showed intermediate thickness and were

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located in between PE and CH-based films. No significant difference was evidenced among blend films with varying CH:PE ratios. In terms of density, CH-based film (2.82 g

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cm-3) presented a bigger value than PE-based film (1.63 g cm-3) but no significant difference was concluded. Statistically, no significant difference was found among the density values of PE, CH and their blend films, the differences in film thickness and density are possibly co-results of the water content and structure compactness of the films [16]. For the thickness and density of blend films, the determined and predicted values

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ACCEPTED MANUSCRIPT were comparable and no significant difference was concluded, indicating an additive nature of the blending for these parameters. 3.2.2. The water solubility and water content of the films Water content indicates the water holding capacity of film matrix upon drying and

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water solubility applauds the disintegration of formed structure or the water resistance of film sheet when contacting with aqueous media. Zavareze et al. [56] reported that the

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applications of biodegradable films sometimes require low solubility or insolubility to

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reinforce moisture barrier properties and shelf-life stability. Although they are not Statistically different, the value in water solubility of PE film is somewhat higher than

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that of CH film. This cannot be understood in considering the stronger hydrophilic nature

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or water binding capacity of PE than CH. After a careful review, it was speculated that the lower water content of PE could be caused by its porous structure, which gives a

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bigger moisture evaporating surface and tunnels to escape. Statistically, no significant

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difference was observed among the determined values in water content or water solubility of PE, CH, PE1-CH1, PE2-CH1 and PE1-CH2 films (Fig. 4). The determined values in

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water content and water solubility were similar to the predicted ones indicating that

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additive blending effects occurred. This disclosed that the blend films presented comparable dehydration rate with the control films under the same drying regime. These are results of the joint actions of the hydrophobicity/hydrophilicity of PE and CH, their interactions and film structures. As an example, Aljawish et al. [57] suggested the high moisture content of CH film was due to the strong hydrogen bond interactions between water molecules and the functional groups (e.g. hydroxyl and amino groups) of chitosan. Contextually, the water content of the blend films was not substantially affected by the

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ACCEPTED MANUSCRIPT molecular interaction of PE and CH. Moreover, a previous study evidenced that the interactions of a positively charged polyelectrolyte and a negatively charged polyelectrolyte usually form a soluble complex [8]. 3.2.3. Water vapor permeability and contact angle for the films

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Water vapor permeability (WVP) is a vital parameter for food package indicating the

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moisture exchange between food and surrounding atmosphere [8,20]. WVP values of the

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PE, CH and blend films were presented in Fig. 4. The WVP value of PE film (5.53×10-12 g. cm-1. s-1. Pa-1) was much higher than that of CH film (3.30×10-12 g. cm-1. s-1. Pa-1). This

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was firstly ascribed to the higher hydrophilicity of PE than CH, which made the PE more

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liable to adsorb moisture from the environment and favored the transportation of water molecules across the film. Secondly, in contrast to CH film, the more porous structure of

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PE film also is a major contributor to its higher WVP. For the present three blend films,

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their WVP values were not significantly different and comparable to that of PE film, while the PE1-CH1 and PE1-CH2 films presented lower WVP values than PE film. The

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significant reduction in WVP makes the films more suitable for food packaging as it

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reduces moisture loss or uptake of food products over long-term storage resulting in a longer shelf life [58]. For all blend films, the determined WVP is similar to the predicted

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one indicating an additive blending effects. The hydrophobicity of the film surface was evaluated by analyzing the water contact angle [59,60], the results are presented (Table 1). In contrast, CH film displayed a bigger contact angle than PE film indicating a more hydrophobic surface of CH film than its PE counterpart. In other words, PE film is more liable to be wet than CH film in contacting hydrous media. In terms of contact angle, all blend films stand in between PE and CH films. Interestingly, the determined and

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ACCEPTED MANUSCRIPT predicted contact angles were observed almost same for PE2-CH1 and PE1-CH2 films, but they are significantly different in the case of PE1-CH1. This reflected that a nonadditive blending effect could be expected on contact angle to enhance the surface hydrophobicity of a blend film by applying a propriate blending ratio.

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3.3. Optical properties

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The optical properties are considered as important factors of packaging film in which

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the visual characteristic of the product is obtained by the consumers. The optical properties of the present films were presented in (Table 2). Clearly, in contrast to CH

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film, the PE film had a lower luminosity (L) indicating its darker nature. All the blend

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films showed the comparable L values to PE film. By judging their absolute values in a, PE film is of a weak green color (a =2.82 > 0) but the CH film is of a weak red color (a =

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-1.57 < 0). All the blend films are of a very weak red color with their determined a values

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varying from 0.63 to 1.37. Overall, all films, the individual or the blend, showed a yellowness nature indicated by their positive b values. This is in agreement with the fact

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that their Hue angles are located in the range of 75-95. As reported previously, the

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yellowish nature of the polysaccharide-based materials is related to the presence of repeat units of D-glucopyranose [61]. Interestingly, at a blending PE/CH ratio of 1:1, significant

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differences were observed in b, ΔE, Chroma, and Hue angle between their determined and predicted values. The opacity (OP) is an inverse measure of transparency, the change in transparency of films might be related to the thickness, i.e. a high opacity is always in line with a low transparency [62]. The opacity values of the presented were shown in (Table 2). Clearly, the CH film is more opaque than the PE film, this is due to the nature of the chitosan [61].

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ACCEPTED MANUSCRIPT Also, this was possibly due to the poor solubility, lower thickness, and higher water content of CH in film forming solution, in which CH was dispersed as micro particles and less dissolved than PE. Our results are in agreement with the results obtained from Ren et al. [6] where the values of the opacity for the films changed due to the water solubility

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and luminosity. The presence of these particulates eventually caused the reflection and

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dispersion of the incident light at the two-phase interface, thus gave rise to a high opacity

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in CH film. Importantly, all the blend films displayed lower values in opacity than CH film, which are similar to that of PE film. This would be interpreted by the electrostatic

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interactions of CH and PE, by which a soluble complex was formed and therefore the

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dispensability of CH was improved [8]. As a consequence, in contrast to CH film, less particulates presented in the forming solutions of the blend films and a better light

M

transmittance was achieved [63]. More importantly, at any blending ratio, the determined

ED

opacity was much lower than its predicted counterpart, indicating that a synergistic effect occurred. This claimed that the blending with PE is an efficient way to improve the

PT

transparency of CH film.

CE

3.3. Mechanical properties

Mechanical properties were considered important parameters in evaluating the strength

AC

and flexibility of thin films. They also attested the durability and stability of such films during handling, shipping and storage [20,64]. Tensile strength (TS) is defined as the ability of a film to resist fracture against tensile stress, reflecting its capacity in maintaining the structural integrity. The elongation at break (EAB) is weighed as the maximum stretchability of a film under tensile stress prior to breakage. The Young’s modulus, also called tensile modulus or elastic modulus, predicts the liability of a film to

18

ACCEPTED MANUSCRIPT undergo extension upon tension or shortening upon compression. For a given film, its mechanical properties ultimately depended on the physicochemical properties of the applied polymers, the types and additions of adjuvants, the intermolecular interactions, water content, and the microstructure formed [65, 66].

T

As shown in (Table 3), in contrast to PE film, CH film presented lower values in TS

IP

and Young’s modulus but a high value in EAB. In comparing with the data reported by

CR

Ojagh et al. [66], the present CH film displayed a much lower value in TS (10.97 MPa → 1.22 MPa) but a comparable value in EAB (24.73% → 22.67%). However, the CH film

US

reported by Pereda et al. [19] had both much higher values in TS (17.34 MPa) and EAB

AN

(44.2%) than the present one. The reasons for these discrepancies are really hard to be discerned. Although the water content of the film by Pereda et al. [19] was not reported,

M

the discrepancies of the CH films by Ojagh et al. [67] and ourselves were assumed to be

ED

mainly caused by the much higher water content of the present film (35.92 g/100g) than that of the reported one (20.82 g/100g).

PT

Regarding the TS of blend films, the PE2-CH1 was regarded as the best one, which

CE

presented a much higher value (6.49 MPa) than the other blend films (3.55 and 5.06 MPa) and individual films (1.22 and 3.63 MPa). In terms of EAB, all blend films

AC

displayed comparable results, which are similar to the case of CH film but much lower than the case of PE film. As for Young’s modulus, the PE1-CH1 (24.07 MPa) and PE2CH1 (24.23 MPa) films were much excellent than PE1-CH2 (13.64 MPa), PE (8.79 MPa) and CH (5.29 MPa) films. Combining the three mechanical parameters together, obviously, the PE2-CH1 film is the best one. It must be noted that, for blend films, their determined mechanical parameters are much higher (TS and Young’s modulus) or lower

19

ACCEPTED MANUSCRIPT (EAB) than the corresponding predicted ones in most cases. This applauded that synergistic effects were widely observed in improving mechanical properties of the edible films by lending CH and PE. These synergistic effects were certainly ascribed to the intermolecular interactions occurring between CH and PE, which highly strengthened

T

the network of polymers in film matrix, thus facilitated the TS and Young’s modulus but

IP

decreased the EAB. As a consequence, the optimal blending ratio of CH and PE was

CR

selected as 1:1. This possibly resulted from the difference in the degrees in CH protonation (-NH3+) and PE deprotonation (-COO-) under film formation. As

US

aforementioned, the pH values of blend film forming solutions were located in the range

AN

of 2.10 to 2.20. In this case, the CH molecules were more intensively charged than the PE

molecules than CH counterparts.

ED

4. Conclusions

M

molecules. Thus, from a charge equivalent view, the system should provide more PE

Chitosan (CH) and pectin (PE) are excellent representatives of natural polymers in

PT

fabricating biodegradable or edible packaging materials. Although both of CH and PE

CE

can be applied solely, the present study revealed that the joint application or blending of CH and PE could generate an edible film with superior performance to their own

AC

individual constituents. These achievements are the ultimate consequences of the electrostatic interactions occurring between CH and PE molecules. More importantly, the blending of CH and PE could bring about synergistic effects in improving several properties of the resultant films, especially in transparency and mechanical properties. Fundamentally, the intermolecular interactions of CH and PE depend on the structure of involved polymers and environmental conditions applied. Contextually, in view structural

20

ACCEPTED MANUSCRIPT diversity of both CH and PE, the dependence of the film performance on the polymer structure and environmental factors should be elaborately addressed in further investigations on this topic. Acknowledgment

T

This research was funded by National Key Research and Development Plan

IP

(2016YFD0400204-2) and National Natural Science Foundation of China (31771932).

CR

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122

(1)

(2010)

161-166,

ACCEPTED MANUSCRIPT Figure captions Fig.1. FT-IR spectra of the films prepared from apple pectin (PE), chitosan (CH) and thereof blends at mass ratios of 1:1 (PE1-CH1), 2:1 (PE2-CH1) and 1:2 (PE1-CH2).

T

Fig. 2. X-ray diffractions of the films prepared from apple pectin (PE), chitosan (CH) and

IP

thereof blends at mass ratios of 1:1 (PE1-CH1), 2:1 (PE2-CH1) and 1:2 (PE1-CH2).

CR

Fig. 3. SEM micrographs of the surface (left column) and cross-section (right column) of the films prepared from apple pectin (PE), chitosan (CH) and thereof blends at mass

US

ratios of 1:1 (PE1-CH1), 2:1 (PE2-CH1) and 1:2 (PE1-CH2) at a magnification of ×

AN

2000.

Fig. 4. Water solubility (WS), Water content (WC), and Water vapor permeability

M

(WVP) for the films prepared from apple pectin (PE), chitosan (CH) and thereof blends at

ED

mass ratios of 1:1 (PE1-CH1), 2:1 (PE2-CH1) and 1:2 (PE1-CH2), the black and white

AC

CE

PT

color column refers to the determined and predicted values respectively.

32

ACCEPTED MANUSCRIPT Table1. Physical Properties of films prepared with different ratios of pectin (PE) and chitosan (CH).

Thickness (µm)

Density (g·cm-3)

Contact angle (°)

PE

87.00 ± 6.53c

1.63 ± 0.53a

31.69 ± 0.72a

CH

54.37 ± 2.95a

2.82 ± 0.45a

46.05 ± 1.41d

PE1-CH1-D

68.44 ± 3.53ab

1.63 ± 0.61a

34.44 ± 1.26ab

PE1-CH1-P

70.68 ± 3.69

2.23 ± 0.43

PE2-CH1-D

81.22 ± 4.27bc

2.21 ± 0.09a

PE2-CH1-P

76.23 ± 4.56

2.02 ± 0.45

PE1-CH2-D

70.85 ± 9.19b

2.14 ± 1.02a

PE1-CH2-P

65.14 ± 3.05

AN

IP

US

CR

38.87 ± 0.63*

2.43 ± 0.42

36.28 ± 0.46b 36.43 ± 0.49 41.87 ± 1.00c 41.31 ± 0.86

Data bearing different superscript lowercase letters within the same column are

M

a-d

T

Film code e

ED

significantly different (p ˂ 0.05). e The films were made by a casting method from a film forming solution, which was prepared by dissolving 15 g polysaccharide matrix (sum of

PT

pectin and chitosan) in 1 L water containing 16 g glycerol, 5 g tween 80 and 10 g citric

CE

acid. PE and CH referred to the films prepared with pure pectin and chitosan, respectively. PEx-CHy referred to the film from a blend of pectin and chitosan at a ratio

AC

of x/y. For the film codes with a suffix of D, the values in its row are experimentally determined, while for the film codes with a suffix of P, the values in its row are calculated from the corresponding data of PE and CH according to their blend ratios, called predicted values.

*

The asterisk means the predicted value was significantly

different from the determined one (p<0.05).

33

ACCEPTED MANUSCRIPT Table 2. Optical properties of the films prepared with different ratios of pectin (PE) and chitosan (CH). Film

a

∆E

b

PE

16.64 ± 89.46 ± 2.01

a

96.19 ± 0.87

b

2.82 ± 1.05

c

14.11 ± 3.16

ab

10.26 ± 1.02

ab

13.99 ± 2.82

ab

CH

10.08 ± -1.57 ± 0.26

a

CH1-D

15.52 ± 91.29 ±1.79

a

0.76 ± 0.84

b

0.63 ± 0.65

12.18 ± 1.25

PE290.69 ±0.73

a

1.37 ± 0.43

bc

14.15 ± 2.57

PE291.68 ±1.23

1.37 ± 0.79

PE190.92 ±1.39

0.74 ± 0.39

b

17.29 ± 1.96

PE1-

93.97 ±0.62

PT

CH2-P

-0.12 ± 0.52

14.02 ± 2.86

ab

12.39 ±

13.07 ±

2.59

1.97

18.71 ±

17.31 ±

b

ED

CH2-D

a

1.03

a

14.47 ±

12.84 ± 1.88

M

CH1-P

10.38 ±

14.21 ±

ab

AN

CH1-D

3.34

ab

13.36 ± 92.82 ±0.87

3.30

ab

US

PE1CH1-P

0.18

a

14.39 ±

CR

PE1-

1.95

1.30

15.93 ± 2.66

ab

2.33

b

Opacity

(hue)

(mm-1)

78.93 ±

1.76 ±

1.61

a

98.68 ±

2.59

ab

3.35

b

0.95

c

87.26 ± 2.74

b

0.42a 10.82 ± 0.80b 2.54 ± 0.25a

88.81 ±

6.29 ±

1.24

0.34*

84.53 ± 0.79

b

1.64 ± 0.31a

85.45 ±

4.75 ±

1.36

0.26*

87.61 ± 1.04

b

2.82 ± 0.12a 7.83 0.

12.24 ±

11.53 ± 0.70

*

1.30

*

11.70 ± 0.72

*

92.16 ± 1.14

*

48 *

Data bearing different superscript lowercase letters within the same column are

CE

a-C

3.84

ab

C

angle

T

L

*

IP

code

Hue

d

significantly different (p ˂ 0.05). d The films were made by a casting method from a film

AC

forming solution, which was prepared by dissolving 15 g polysaccharide matrix (sum of pectin and chitosan) in 1 L water containing 16 g glycerol, 5 g tween 80 and 10 g citric acid. PE and CH referred to the films prepared with pure pectin and chitosan, respectively. PEx-CHy referred to the film from a blend of pectin and chitosan at a ratio of x/y. For the film codes with a suffix of D, the values in its row are experimentally determined, while for the film codes with a suffix of P, the values in its row are calculated from the corresponding data of PE and CH according to their blend ratios, called predicted values.

*

The asterisk means the predicted value was significantly

34

ACCEPTED MANUSCRIPT different from the determined one (p<0.05). L (luminosity), a (greenness; redness) and b

AC

CE

PT

ED

M

AN

US

CR

IP

T

(blueness; yellowness), ∆E (Color change), C* (Chromaticity).

35

ACCEPTED MANUSCRIPT Table 3. Mechanical properties of the films prepared with different ratios of pectin (PE) and chitosan (CH). Film code d

Elongation at break

Youngs modulus

(MPa)

(%)

(MPa)

PE

3.63 ± 0.26b

43.77 ± 5.51b

8.79 ± 1.44ab

CH

1.22 ± 0.17a

22.67 ± 1.53a

5.29 ± 0.53a

PE1-CH1-D

5.06 ± 0.74bc

22.09 ± 3.05a

24.07 ± 2.49c

PE1-CH1-P

2.25 ± 0.47*

33.22 ± 3.51*

PE2-CH1-D

6.49 ± 1.15c

24.73 ± 2.15a

PE2-CH1-P

2.83 ± 0.19*

36.80 ± 4.19*

7.64 ± 0.80*

PE1-CH2-D

3.55 ± 0.41b

30.04 ± 5.56a

13.64 ± 2.63b

PE1-CH2-P

2.01 ± 0.16

29.63 ± 2.83

IP

CR

US

6.21 ± 1.82* 24.23 ± 4.04c

6.45 ± 0.19*

Data bearing different superscripts lowercase letters within the same column are

significantly different (p ˂ 0.05).

d

AN

a-c

T

Tensile strength

The films were made by a casting method from a

M

film forming solution, which was prepared by dissolving 15 g polysaccharide matrix (sum of pectin and chitosan) in 1 L water containing 16 g glycerol, 5 g tween 80 and 10 g

ED

citric acid. PE and CH referred to the films prepared with pure pectin and chitosan, respectively. PEx-CHy referred to the film from a blend of pectin and chitosan at a ratio

PT

of x/y. For the film codes with a suffix of D, the values in its row are experimentally determined, while for the film codes with a suffix of P, the values in its row are

CE

calculated from the corresponding data of PE and CH according to their blend ratios, called predicted values.

*

The asterisk means the predicted value was significantly

AC

different from the determined one (p<0.05).

36

Figure 1

Figure 2

Figure 3

Figure 4