Study of chitosan with different degrees of acetylation as cardboard paper coating

The biodegradability of chitosan is significant for packaging systems. Another relevant property of chitosan is its degree of acetylation (DA), which affects other properties, such as crystallinity and hydrophobicity. The DA can be modulated by chitin deacetylation or even chitosan reacetylation.

Carbohydrate Polymers 210 (2019) 56–63 Contents lists available at ScienceDirect Carbohydrate Polymers journal homepage: www.elsevier.com/locate/carbpol Study of chitosan with different degrees of acetylation as cardboard paper coating T ⁎ Mariane Gatto, Deise Ochi, Cristiana Maria Pedroso Yoshida , Classius Ferreira da Silva UNIFESP - Federal São Paulo University, Institute of Environmental, Chemical and Pharmaceutical Sciences, SP, Brazil A R T I C LE I N FO A B S T R A C T Keywords: Coating Chitosan Chemical reacetylation Acetylation degree The biodegradability of chitosan is significant for packaging systems Another relevant property of chitosan is its degree of acetylation (DA), which affects other properties, such as crystallinity and hydrophobicity The DA can be modulated by chitin deacetylation or even chitosan reacetylation The novelty of this paper is the application of reacetylated chitosan as a coating for cardboard paper surfaces to improve the barrier and mechanical properties of the paper Chitosan with 2% DA was reacetylated to yield chitosan with 48% DA Both samples were applied as cardboard paper coating, and the coated materials were characterized The paper-film system of chitosan with 2% DA had better water barrier and mechanical resistance Heterogeneous deacetylation of chitin reduced the solubility of chitosan because molecular groups were distributed in blocks, increasing the hydrophobicity of the polymer Introduction In the 20th century, petrochemical-based materials were extensively used as packaging materials In general, the barrier properties of the paper packaging material can be controlled by coating with petroleumbased derivatives polymers (e.g., polyolefins and ethylene vinyl alcohol) and waxes (Rastogi & Samyn, 2015) However, there is a demand for product manufacturers to focus on ecofriendly packaging solutions, as sustainability concerns are increasing as well as the replacement of the nonrenewable sources (mainly from petroleum) with renewable sources (Robertson, 2012) Examples of such renewable materials include biopolymers such as starch, cellulose, animal or plant-based proteins, lipids, and chitin/chitosan (Cutter, 2006) Paper is a biodegradable material widely applied as packaging material in many industries such as electronics, food, and pharmaceuticals because of its low cost, biodegradability, good resistance, light weight, and recyclability However, hydrophilicity and porosity limit their packaging applications (Rastogi & Samyn, 2015) Cellulose chains are linked by hydrogen bonds, which directly affect the physicochemical properties (i.e., solubility and crystallinity) and mechanical properties of paper (Kondo, Koschella, Heublein, Klemm, & Heinze, 2008) The hydrogen bonds favor adsorption and transport of water molecules, gases, and oils through the cellulose network, resulting in poor barrier properties Moreover, water adsorption causes fiber swelling, spoiling material shape and reducing mechanical properties, which is not ⁎ desirable in packaging applications (Rastogi & Samyn, 2015) Therefore, despite the versatile advantages of paper, it is still important to improve its mechanical properties; water absorption; and barrier to gas, moisture, and fat (Reis, Yoshida, Reis, & Franco, 2011) The required properties of paper depend on its application; for example, some food packaging applications require high permeability of oxygen or even low water permeability; on the other hand, paper for fried potatoes package has to be oil-impermeable The barrier and mechanical properties of paper can be modified by coating using a nonrenewable polymer; however, this transforms an ecofriendly material in a more polluting material On the other hand, biopolymers coatings offer environmental advantages such as biodegradability, better recyclability, nontoxicity, and biocompatibility, compared to conventional synthetic polymers (Tang, Kumar, Alavi, & Sandeep, 2012) Therefore, coating paper surface with renewable biopolymers is an interesting method to improve the barrier and wettability of paper as packaging materials (Khwaldia, Basta, Aloui, & El-Saied, 2014) For packaging applications, chitosan films are characterized by selective permeability to gases (CO2 and O2), good mechanical properties, and biodegradability However, high permeability to water vapor limits their use, as this is not desirable for packaging applications Promising results have been obtained by the addition of lipids, waxes, and clays to improve hydrophobicity, although this often reduces chemical and mechanical properties (Elsabee & Abdou, 2013; MacIel, Yoshida, & Franco, 2012) Composite coatings or multilayer coatings on paper Corresponding author at: UNIFESP Federal University of São Paulo, Laboratory of Biotechnology and Natural Products, Rua São Nicolau, 210, Diadema, SP, Brazil E-mail address: cristiana.yoshida@unifesp.br (C.M.P Yoshida) https://doi.org/10.1016/j.carbpol.2019.01.053 Received 15 October 2018; Received in revised form 11 January 2019; Accepted 16 January 2019 Available online 17 January 2019 0144-8617/ © 2019 Elsevier Ltd All rights reserved Carbohydrate Polymers 210 (2019) 56–63 M Gatto et al distribution can be obtained by chitosan reacetylation under homogeneous conditions Chitosan reacetylation in homogeneous conditions produces more uniform structures (Chitosan 3, Eq (3)) If the starting chitosan is extensively deacetylated, the obtained copolymer will have randomly distributed GlcN and GlcNAc units (Aiba, 1991) This aim of this work is to evaluate the coating application of deacetylated chitosan with 2% DA and reacetylated chitosan with 48% DA on the cardboard paper surface and test the effects of these different chitosan coatings on the final material properties surface have been prepared to combine the good mechanical resistance of paper material with the barrier properties of coatings materials Khwaldia et al (2014) observed that caseinate and caseinate/chitosan bilayer coatings improved the water vapor barrier and mechanical properties of paper packaging Atkinson et al (2017) applied chitosan as paperboard coating and obtained better water wicking properties and thus less moisture absorption, compared to using paper coated with phenol formaldehyde resin Kuusipalo, Kaunisto, Laine, and Kellomäki (2005) applied chitosan as a wet-end additive in paperboard to improve the mechanical and barrier properties and bending strength Chitosan with low molar mass (0.768 × 105g/mol) coating yielded paper with lower water absorption, more smoothness, and higher dry strength than chitosan with medium molar mass (2.375 × 105 g/mol) (Habibie, Hamzah, Anggaravidya, & Kalembang, 2016) Chitosan is the main chitin derivative and consists of 2-amino-2deoxy-D-glucopyranose (GlcN) units and 2-acetamido-2-deoxy-D-glucopyranose (GlcNAc), the latter in proportions below 50% of the biopolymer chain The GlcNAc proportions in the macromolecule are represented by the degree of acetylation (DA) (Chatelet et al., 2001) Chitin is insoluble in most organic acids, while chitosan is soluble in dilute acidic solutions Chitosan is a strong base because of primary amino group presents in a GlcN unit At low pH, these amino groups (pKa 6.3) become protonated and positively charged Chitosan becomes a water-soluble cationic polyelectrolyte, increasing the pH (above 6), which causes loss of charge, reducing solubility This soluble-insoluble transition occurs at a pKa value of 6.0–6.3 Chitosan solubility is strongly dependent on chitosan DA (presence of GlcN and GlcNAc units) Increasing the acetyl groups promotes changes in chitosan structure; it transforms into a more crystalline chain Therefore, chitosan solubility can be controlled by manipulating the DA (Pillai, Paul, & Sharma, 2009) A sample of chitin with 100% acetylation (i.e., 100% of the residues are GlcNAc) can be deacetylated to obtain “Chitosan 1″ with 60% acetylation degree; that is, 60% of the residues are GlcNAc (Eq 1) In parallel, another partial deacetylation can be performed to obtain “Chitosan 2″ with 3% acetylation degree; that is, 3% of the residues are GlcNAc (Eq 2) However, if “Chitosan 2″ is subjected to a reacetylation reaction, “Chitosan 3″, also with 60% acetylation degree, can be obtained (Eq 3) Although Chitosan samples and have the same amount of GLcNAc residues, the properties of these two samples can be extremely different since the distributions of these residues are different Block-like residues of GlcNAc and GlcN are present in Sample 1; however, such residues are statistically distributed in Sample (nonblock residues), promoting different properties in both samples When the GlcN residues are in blocks (Chitosan 1), they are less accessible to protonation, and therefore, the solubilization of chitosan is more difficult Chitin (DA = 100%) Chitin (DA = 100%) Deacetylation → Deacetylation → Chitosan (DA = 60%) (1) Chitosan (DA = 3%) (2) Chitosan (DA = 3%) + Acetic Anhydride = 60%) Reacetylation → Materials and methods 2.1 Materials Original chitosan with 2% DA, provided by Mahtani Chitosan (Veraval, India) 2.2 Homogeneous reacetylation Reacetylation was carried out as described by Vachoud, Zydowicz, and Domard (1997) Chitosan with 2% DA (C-2) was dissolved (1% w/ w) in aqueous acetic acid (AcA) solution in stoichiometric proportions ([AcA] = [-NH2]) and was agitated overnight Afterward, 1,2-propanediol was added in two portions to the chitosan/AcA solution The first portion was added in its pure form (in an amount corresponding to 90% v/v of the chitosan solution), and the solution was agitated for h A solution was prepared by mixing the second portion of 1,2-propanediol (in an amount corresponding to 10% v/v of initial chitosan solution) and acetic anhydride (AAn) The AAn amount, mAAn(g), was determined by Eq (4), where mc(g) is the amount of chitosan, DA1 and DA2 are initial and desired DAs, respectively (DA2 = 0.48), WH2O is water content determined through thermogravimetric analysis, MAAn is the molar mass of acetic anhydride (102.1 g/mol), and M0 average mass of the repetitive unit of chitosan mAAn = mc (DA2 − DA1 )(1 − w H2 O ) MAAn MO (4) The AAn solution was homogenized for 45 under agitation and was added dropwise to the chitosan solution The mixture was kept under agitation overnight The resulting mixture was precipitated using concentrated NH4OH, washed 10 times with distilled water, frozen, and then freeze-dried The resulting reacetylated chitosan with 48% DA is referred as C-48 in this paper 2.3 Chitosan characterization 2.3.1 Proton nuclear magnetic resonance spectroscopy Proton nuclear magnetic resonance (H NMR) spectroscopy was performed before and after reacetylation to determine the DA Here, mg of chitosan was dissolved in mL of D2O acidified with μL of HCl (37%) Spectra were recorded on an ALS300 300 MHz (Bruker) spectrometer at 300 K The DA (%) was calculated using (5): Chitosan (DA ⎛ ICH ⎞ DA (%) = ⎜ ⎟ × 100 I ⎝ (H − H 6) ⎠ (3) Furthermore, the distribution of N-acetyl groups is related to chitosan solubility (Franca, Freitas, & Lins, 2011) Chitosan is a linear copolymer obtained from chitin deacetylation or partial chitosan reacetylation to obtain chitosan with higher DA These reactions can occur in homogeneous (one phase reaction) or heterogeneous (more than one phase, i.e., solid and liquid) conditions, resulting in copolymers with a different distribution of GlcN and GlcNAc units Chitosan produced from chitin deacetylation under heterogeneous conditions occurs preferentially in amorphous regions of the molecule, forming block copolymers and resulting in a more hydrophobic polymer (Chitosan 1, Eq (1)) Chitosan with a higher DA and different N-acetyl group (5) where ICH3 represents the signal of methyl hydrogen in the acetyl group, and I(H2-H6) is the signal of hydrogen connected to carbon and of the glucopyranosyl ring 2.3.2 Thermogravimetric analysis Samples of chitosan powder (3–10 mg) were placed in open platinum capsules in a thermogravimetric analysis equipment (DTG-60, Shimadzu, Japan) The temperature range was from 30 to 200 °C (10 °C/min) under N2 flow at 100 mL/min The water content was calculated according to the mass loss up to 105 °C 57 Carbohydrate Polymers 210 (2019) 56–63 M Gatto et al 1994a) Ten samples of dimensions 0.125 m × 0.125 m were preconditioned in a desiccator containing silica for 72 h at room temperature (25 ± °C) Samples were individually weighed on a semianalytical scale with a precision of 0.01 g and attached to the Cobb test equipment (Regmed, Brazil) Moreover, 100 mL of water was added to contact with the surface delimited by the apparatus ring (internal diameter of 11.28 ± 0.02 cm) for 120 s Soon after the specimen was removed, it was placed between two sheets of absorbent paper and pressed by a conditioning roller (Regmed, Brazil) to remove excess water Lastly, the samples were weighed Water absorption capacity (Abs, g/m²) was determined using (7), where Mf and Mi(g) are final and initial sample mass, respectively 2.3.3 Size exclusion chromatography Average molar mass (Mw) was determined by a size exclusion chromatograph attached to a refraction index and multi-angle scattering measuring equipment (Wyatt Dawn ES multi-angles, United States) The used column was TSKgel G6000PW + G2500PW (30 cm length and 7.8 mm diameter) The mobile phase was an acid acetic buffer solution (0.15 mol.L−1 ammonium acetate/0.2 mol.L−1) flowing through the column at 0.5 mL/min 2.3.4 Fourier transform infrared spectroscopy Fourier transform infrared (FTIR) spectroscopy was performed at room temperature using a universal attenuated total reflectance accessory (ATR), Nicolet iN10MXDA (Thermo Fisher, United States) Each sample was evaluated by taking 64 scans with spectral range of 650–4000 cm−1 and cm−1 resolution Abs = (Mf − Mi ) A × 100 (7) 2.6.5 Moisture content Based on ASTM D644-99 (ASTM, 2007), the moisture content evaluation experiment was executed in triplicate The samples were cut into squares of cm × cm and dried in a TE-393 forced air circulation oven (Tecnal, Brazil) at 105 ± °C for 24 h Moisture content (g H2O/ 100 g paper) was calculated according to (8), where Mw and Md are wet and dried mass, respectively 2.3.5 Differential scanning calorimetry Powdered chitosan samples (2.8–3 mg) were placed in aluminum capsules hermetically closed in a differential scanning calorimeter DSC60 (Shimadzu, Japan) The capsules were exposed to a temperature of 20 °C–200 °C at 10 °C/min 2.4 Chitosan suspension M − Md ⎞ Moisture Content = ⎛ w × 100 Md ⎠ ⎝ Chitosan suspension was prepared as described by Yoshida (2009) First, 1.0% (w/w) of chitosan was dispersed in the aqueous acid solution Acetic acid was then added in stoichiometric proportions, according to the DA (DA = 2% or DA = 48%) and chitosan weight The system was kept under agitation overnight ⎜ ⎟ (8) 2.6.6 Water vapor transmission rate The procedure was based on ASTM E96-00 (ASTM, 2000) Five discshaped samples were cut from each sample and fixed to the top of permeation cells containing silica gel The cells were conditioned in desiccators containing saturated saline (sodium chloride) at relative humidity of 75 ± 5% and kept in a temperature-controlled chamber (25 ± 0.2 °C) Using an analytical scale, the mass gain of the system (cell and sample) was determined at 24 h intervals over 120 h The water vapor transmission rate (WVTR) (gH2O/m2day) was calculated using (9), where G (g) is the weight gained by the system, A (m²) is the area exposed to vapor transmission, and t (day) is the number of days the sample spent in the chamber 2.5 Chitosan coating application on paper First, mL of chitosan suspension was dispersed on a cardboard surface (Triplex TP 250 g/m2 Suzano Papel e Celulose Ltda., Brazil) using a 60 μm bar (TKB Ericken, Brazil) Coated sheets were dried in a forced air circulation oven (MA035/1000, Marconi Equipamentos, Brazil) at 100 °C for 90 s Each sheet sample was coated three times (MacIel et al., 2012) The studied systems were paper coated with chitosan of 2% DA (CP-2) and 48% DA (CP-48) and paper without coating (P) WVTR = G /(A t ) (9) 2.6 Chitosan-coated cardboard paper characterization 2.6.7 Taber stiffness Stiffness was determined according to T489 om-92 (TAPPI, 1994b) Ten specimens of each system were cut in both directions of cellulosic fibers (machine direction MD and cross-machine direction CD) in dimensions of 38.1 mm × 70.0 mm using a pneumatic guillotine (Regmed, Brazil) Samples were evaluated using an RI-5000 stiffness meter (Regmed, Brazil) at an angle of 15° 2.6.1 Coating evaluation: colored solution penetration The evaluation procedure was adapted from Marcy (1995) and executed in triplicate Erythrosine in isopropanol solution (0.5% w/w) was applied to uncoated (matte side) and coated paper samples (coated side) covering the entire surface Samples were held upright for 60 and dried in a TE-393 forced air circulation oven (Tecnal, Brazil) at 50 °C for 30 2.6.8 Fat barrier The methodology was based on Ham-Pichavant, Sèbe, Pardon, and Coma, (2005) Different test solutions containing castor oil, toluene, and n-heptane were prepared One drop of test solution was applied to the sample paper surface, and the sample was kept for 15 s Excess solution was removed, and the appearance of the opposite surface was observed The highest kit number for the test solutions that did not cause blemishes was the adopted as the kit rating value of fat repellency Kit nº1 and Kit nº12 were respectively the least and the most aggressive solutions for producing a stain on the opposite surface 2.6.2 Grammage Following ASTM D646-96 (ASTM, 1996), 10 samples of each system were cut into squares of 0.016 m² (0.125 m × 0.125 m) and weighed on an analytical scale Grammage G (g/m²) was calculated using (6), where M (g) is the paper weight, and A (m²) is the area G = M /A (6) 2.6.3 Thickness Thickness was measured using a digital micrometer (0.001 mm) (Mitutoyo, Japan) in quintuplicate For each sample, six measurements were carried out at different points 2.6.9 Scanning electron microscopy Samples were cut in squares of cm × cm and kept in a silicacontaining desiccator for 48 h at 25 °C The metallic coating was conducted using a K450 sputter coater (Emitech, France), gold layer thickness estimated at 200 Å Micrographs of the surface (1000× 2.6.4 Cobb test Cobb test was executed as described in T-441 om-90 (TAPPI, 58 Carbohydrate Polymers 210 (2019) 56–63 M Gatto et al band at 1313 cm−1; and a CeCH3 deformation band at 1380 cm−1 The C-2 spectra presented angular deformation bands of NH (amide II) at 1560 cm−1 (Focher, Naggi, & Torri, 1992) A less intensity of C]O (1630 cm−1), amide III (1313 cm−1) and CeCH3 (1380 cm−1) peaks were observed in the C-2 spectra, compared to C-48; this was associated with the lower content of GlcNAc units Focher et al (1992) observed that the bands at 1420–1435 cm−1were related to CH2 Modifications in the CH2OH group were observed in the C-2 and C-48 samples Duarte, Ferreira, Marvão, and Rocha (2002) reported CO stretching at 1159, 1074, and 1025 cm−1 The asymmetric absorbance of CO at 1159 cm−1 could indicate changes promoted by depolymerization during the deacetylation process In the differential scanning calorimetry (DSC) thermograms (Fig 3), an endothermic peak was identified between 20 °C and 100 °C, with an enthalpy of 242.60 J/g at 58.37 °C for C-2 and 251.39 J/g at 58.03 °C for C-48 These peaks are attributed to the evaporation of water bounded to chitosan at hydroxyl groups and free amine in the amorphous region of chitosan (Ghosh, Azam Ali, & Walls, 2010) The C-48 endothermic peak was slightly higher than that of C-2, despite that C48 had a significantly higher WC, as shown in Table Although the water evaporation enthalpies for both samples were practically equal, their WCs were quite distinct It is essential to consider that the analytical conditions were very different for both techniques In DSC analysis, the evaporation of water took place in a few minutes and at temperatures below that used to determine the WC Moreover, in the WC determination tests, the sample was subjected to 24 h drying at 105 °C Thus, water strongly bonded to the chitosan chain could be removed, whereas in the DSC, not all the water molecules were removed, but only those weakly bonded to the chitosan chain Kittur, Harish Prashanth, Udaya Sankar, and Tharanathan (2002) observed that a higher number of amine groups in chitosans with lower DA allowed more bondings with water molecules, and therefore, lower DA was related to higher endothermic peaks They prepared chitosan with different DAs using sodium hydroxide and observed that the water evaporation enthalpy was inversely proportional to the DA when the chitosan samples were obtained directly from chitin; for example, the sample with 52% DA presented enthalpy 20% lower than the sample with 11% DA Our results showed that the difference between the enthalpies of the sample with 2% DA, obtained directly from chitin, was only 3% higher that of the reacetylated sample with 48% DA Therefore, the distribution of the chitosan residues, in blocks or not, affects the water evaporation enthalpy probably due to the accessibility of the water to the residues The exothermic peaks observed between 280 °C and 340 °C were associated with chitosan degradation (Ng, Cheung, & McKay, 2002) Guinesi and Cavalheiro (2006) evaluated different chitosan samples with different DAs and observed a similar exothermic peak related to GlcN group degradation in a temperature range of 296 °C–299 °C In their study, GlcNAc group degradation occurred near 404 °C, but this was not observed in our samples Moreover, they assessed the heterogeneous N-deacetylation of α-chitin to obtain chitosan with different DAs, where the produced blocks of GlcNAc and GlcN along the chitosan chain were probably due to the deacetylation The block-like chitosan could present different thermal events, compared to the chitosan with statistical distribution of acetylated and non-acetylated residues In our study the GlcNAc content in C-2 was shallow; thus, an exothermic peak at 404 °C was not expected Nam, Park, Ihm, and Hudson, (2010) observed two separate peaks in DSC thermograms, which showed that a block-like structure was obtained in the chitosan backbone Our results did not show the separate peaks probably due to the random backbone structure, as seen in C-48 The enthalpy for C-2 was 223.67 J/g at 311.00 °C and for C-48 was 134.15 J/g at 302.53 °C Since high energy and high temperature are required to degrade 2% DA chitosan, C-2 had higher thermal stability than C-48 Hamdi et al (2019) also observed high thermal stability for lower DA However, they did not evaluate GlcN and GlcNAc magnification) were obtained using a scanning electron microscope with energy dispersive X-ray detector (Leo 440i, LEO Electron Microscopy/Oxford) 2.6.10 Differential scanning calorimetry Samples of the paper systems (2.8–3.0 mg) were packed in hermetically sealed aluminum capsules The capsules were exposed to a heating rate of 10 °C/min under a temperature of 20–550 °C and N2 flow of 100 mL/min in a DSC-60 equipment (Shimadzu, Japan) 2.7 Statistical analysis The statistical analysis was ANOVA for one criterion using Assistat7.7 version developed by Universidade Federal de Campina Grande-UFCG Results and discussion 3.1 Chitosan characterization The initial chitosan sample had a DA of 2% After the reacetylation process, a sample with high DA (48%) and higher content of GlcNAc groups was obtained Chitosan with 48% DA was expected to exhibit hydrophobic properties similar to chitin while maintaining partial solubility in acidic conditions to enable posterior paper coating Nonetheless, random distribution of GlcN and GlcNAc related to homogeneous acetylation affected the hydrophilic/hydrophobic property of chitosan to a greater extent than the quantity of these groups The increase in water content in C-48 illustrates this effect Reacetylation also caused an expected increase in molar mass, as seen in Table 1, since acetyl groups were added, converting GlcN units into GlcNAc As stated by Kasaai (2009), chemical modification such as acetylation of chitosan results in a change in H NMR spectra Patterns observed in the H NMR spectra of samples were also identified by Hirai, Odani, and Nakajima (1991) The signal at 2.1 ppm indicates the presence of hydrogen of the methyl group from N-acetyl, and it is more intense in the reacetylated chitosan C-48 sample spectrum (Fig 1b) than in the C2 sample (Fig 1a) This may be associated with the higher quantity of GlcNAc units obtained from the reacetylation reaction The signals at 3.3 and 4.8 ppm represent the hydrogen atoms attached to carbon and carbon of D-glucosamine ring, respectively A higher signal was observed in this interval (Fig 1); this could indicate a higher quantity of GlcNAc units in C-48 chitosan than in C-2 chitosan According to Vårum, Antohonsen, Grasdalen, and Smidsrød (1991), peaks between 3.5 and 4.5 ppm are related to hydrogen atoms attached to carbon and carbon of GlcNAc and GlcN units and hydrogen atom attached to carbon of GlcNAc unit; the signal between 4.5 and 5.0 ppm was related to hydrogen atoms attached to carbon of GlcN unit and carbon of GlcNAc The FTIR spectra of C-2 and C-48 shown in Fig present the typical bands for chitosan samples The C-48 spectra presented specific bands of chitosan: NH (amide II) at 1560 cm−1; CO band at 1660 cm−1, which is related to C]O of intermolecular hydrogen bond with NH groups; C]O band at 1623 cm−1, which is related to hydrogen bonds of NH group and OH attached to carbon at position of the glucopyranoside ring; amide III Table Degree of acetylation (DA), water content (WC), and molar mass (Mw) of chitosan with 2% DA (C-2) and reacetylated chitosan (C-48) Chitosan Samples DA (%) WC (%) Mw (105g/mol) C-2 C-48 48 5.05 16.37 5.238 6.186 59 Carbohydrate Polymers 210 (2019) 56–63 M Gatto et al Fig H-RMN spectra of (a) C-2 and (b) C-48 Table Properties of cardboard paper samples coated with three layers of C-2 and C-48 suspensions and uncoated cardboard paper (P) Properties Cardboard Paper Samples P Average thickness (μm) Grammage (g/m²) Moisture content (gH2O/100 g paper) Water absorption Cobb test (g/m²) WVTR (g H2O/m²day) Taber stiffness MD (mN.m) Taber stiffness CD (mN.m) CP-2 a CP-48 b 3.78 ± 0.01 252.23 ± 1.79a 8.20 ± 0.01b 3.90 ± 0.01 252.40 ± 1.93a 8.80 ± 0.10b 4.08 ± 0.01c 259.93 ± 2.86b 9.86 ± 0.06a 45.21 ± 2.75a 39.74 ± 1.44b 52.16 ± 1.51c 298.17 ± 3.34a 18.10 ± 0.61a 280.69 ± 2.92b 18.91 ± 0.71b 291.44 ± 8.81ab 17.26 ± 0.60c 7.05 ± 0.22a 8.34 ± 0.49b 7.66 ± 0.27c Fig FTIR spectra of C-2 and C-48 samples Note: In the same row, superscript with different letters indicates that the mean values are statistically different Criteria: p < 0.01 for G, E, MD, and CD For moisture content, Cobb test, and WVPR, p < 0.05 Average thickness increased by about 3% and 8% for CP-2 and CP48, respectively, compared with uncoated cardboard paper (P) (Table 2) The slight increase of coated cardboard paper thickness may be related to the penetration of chitosan solution into the cellulose network Khwaldia et al (2014) found that the thickness of caseinate and caseinate/chitosan bilayer coatings on paper ranged between 3.70 and 16.88 μm, applying coatings from to 16 g/m2 Thickness is directly related to the physical and optical characteristics of the paper (tensile strength, transparency, color, whiteness, and electrical resistance) Furthermore, the grammage of CP-48 was statistically higher than those of P and CP-2 This is because C-48 had the highest WC and molar mass (Table 1) However, the difference between the grammages of P and CP-2 was not statistically significant, despite the application of three chitosan layers This result suggests that the water affinity property has an essential effect on the grammage of coated cardpaper The moisture content of CP-2 was not statistically different from that of the uncoated sample The moisture content of CP-48 was about 12% and 20% higher than those of CP-2 and P, respectively Moreover, CP-48 was observed to be more hydrophilic than the CP-2 Despite having a high content of GlcNAc (hydrophobic) groups, the random distribution of molecules in the reacetylated chitosan caused the predominance of hydrophilic property, favoring the hydrogen bonds with water molecules These hydrogen bonds reduce the water vapor barrier property, increasing the interaction between paper and environmental humidity The water absorption capacity (Cobb test) of CP-2 was 12% less than that of P, which is favorable to packaging material application In cellulosic materials, water absorption is related to the water resistance Fig DSC thermograms of C-2 and C-48 distribution On the other hand, according to Wanjun, Cunxin, and Donghua (2005), the random distribution of acetyl group in chitosan with 85% deacetylation degree stabilized the main chain, slowing scissions caused by thermal treatment 3.2 Chitosan-coated paper characterization Application of three layers of C-2 and C-48 chitosan coatings (estimating 2.3 g/m2 of weight coating) resulted in a smooth, uniform, and yellowish cardboard paper surface The systems did not exhibit delamination after rigorous handling, indicating good compatibility between chitosan and cellulose fibers MacIel et al (2012) applied chitosan coating containing anthocyanin and observed similar characteristics 60 Carbohydrate Polymers 210 (2019) 56–63 M Gatto et al and depends on the type of cellulose and the coating material Reis et al (2011) obtained similar results for chitosan-coated Kraft paper Cellulosic fibers are highly hydrophilic because of the hydrogen bonding of cellulose molecules with water molecules, and this reduces the mechanical properties of paper sheets However, the Cobb test showed that the water absorption of CP-48 was 15% and 31% higher than those of P and CP-2, respectively This also confirmed that CP-48 presented higher hydrophilicity than CP-2 As previously mentioned, the random distribution of the GlcN and GlcNAc groups in the chain favored the solubility of chitosan, which may have increased the hydrophilicity of the reacetylated chitosan Habibie et al (2016) observed that the grammage values of the control sample and paper sheet coated with 1% (w/w) of chitosan were similar, but the coated paper sheet had lower water absorption However, the sample treated with 1% of chitosan and 20% of filler presented higher grammage but a slightly lower water absorption Furthermore, the transport properties of coated packaging materials are influenced by the type of coating material, composition, and weight (Aloui, Khwaldia, Slama, & Hamdi, 2011) Chitosan coating significantly reduced WVTR to 6% in CP-2; however, the WVTR difference between CP-48 and P was not statistically significant Considering that paperboard has poor water barrier property, the reduction of WVTR should be due to the chitosan coating The pores between the cellulose fibers network were probably filled with chitosan suspension, which reduced the permeation of the water vapor molecules through the cellulose fibers network Bordenave, Grelier, Pichavant, and Coma (2007) observed a reduction of WVTR of chitosan-coated paper, which was associated with the decrease of the preferential pathway for water across the cellulose network filled with chitosan suspension In the present study, the diffusion rate of water molecules was higher in reacetylated chitosan coating The hydrophilic character of the reacetylated chitosan CP-48 was again reflected by the non-reduction of the WVTR, as its value was similar to that of the uncoated cardboard paper Stiffness tests were performed in the machine direction, MD (that is, the fibers were aligned along the direction of travel in a papermaking machine) and in the cross-machine direction, CD (fibers were transversely aligned) (Brodnjak, 2017) For all samples, Taber stiffness for MD test exceeded the double of that for CD The resistance of fibers in MD was always high due to fibers alignment (Reis et al., 2011) Moreover, CP-2 presented the highest Taber stiffness in MD and CD The resistance and flexibility of chitosan film may have improved the bonds of cellulose fibers in the coated papers, compared to the uncoated paper (MacIel et al., 2012) In addition, CP-48 showed lower Taber stiffness than P Since CP-48 had the highest moisture content, water molecules might have weakened the bonds between the cellulose fibers, reducing the mechanical properties of paper As for coating process evaluation, red-colored aqueous solution penetrated through the sample, which presented colored spots on the opposite surface (Fig 4) In CP-2 and CP-48, the opposite surfaces were clean, indicating that chitosan suspensions formed a uniform and homogeneous coating on the cardboard paper surface Improvement of barrier properties may be related to the solids content in the coated paper, which fills the fibrous structure of the paper Prevention of aqueous solution penetration through the material matrix is a desired property for packaging (Reis et al., 2011) Fig Samples P, CP-2, and CP-48 paper systems during visual analysis 3.2.2 Morphology Application of three layers of chitosan resulted in a more homogeneous paper surface (Fig 5b and c), compared to the uncoated paper (Fig 5a) Chitosan suspension penetrated the cellulose fibers network, filling the paper pores Furthermore, a continuous and uniform film was not observed on the surfaces of CP-2 and CP-48 Although three coating layers were used, chitosan suspension was still absorbed by cellulose fibers, filling empty spaces, and thus, a continuous film was not formed on the surface Anyway, there was some difference between Fig 5(a) – (c), which is probably due to the rugosity Fernandes et al (2009) concluded that the first layers of chitosan solution penetrate the cardboard paper sheets progressively, and at least three chitosan layers are required for film formation due to saturation of the cellulose fibers matrix 3.2.3 DSC analysis In the DSC thermogram shown in Fig 6, the endothermic peaks between 20 °C and 100 °C can be attributed to the evaporation of water attached to paper cellulose chain in P and also that attached to chitosan and cellulosic chains in CP-2 and CP-48 samples The exothermic peak between 330 °C and 420 °C was associated with the degradation of the paper-film system of the samples Chitosan as coating affects the thermal properties of cardboard paper samples The degradation enthalpy of CP-2 was 91.67 J/g at 375.1 °C, while that of CP-48 was 81.18 J/g at 375.0 °C, and that of P was 78.62 J/g at 371.5 °C Acetylated chitosan with 2% DA had better thermal stability, reinforcing the results regarding the higher resistance of CP-2 Habibie et al (2016) attributed the thermal property variation to the greater interaction between the amino groups in chitosan and the hydroxyl groups in cellulose Conclusion 3.2.1 Fat barrier Storage of high-fat content foodstuffs requires packaging materials with a fat barrier In the uncoated cardboard paper (P), the fat barrier was represented by kit test number Papers CP-2 and CP-48 achieved absolute fat barrier, indicating that chitosan is resistant to fat diffusion Cationic groups (NH3+) in chitosan interacted electrostatically with anionic groups of lipid, retaining fat and preventing the appearance of spots on the opposite surface (Ham-Pichavant et al., 2005) Cardboard paper coated with chitosan of 2% DA is a potential packaging material because of the following advantages: reduced water absorption capacity due to absolute fat barrier, decreased WVTR, and higher resistance (higher Taber stiffness in CD and MD), compared to uncoated cardboard paper Moreover, C-48 coating presented a higher hydrophilicity than the original chitosan C-2 coating Block distribution 61 Carbohydrate Polymers 210 (2019) 56–63 M Gatto et al Fig DSC thermogram for P, CP-2, and CP-48 of molecules unit in C-2 decreased solubility because of the unavailability of the NH3+ groups to interact with water The high availability of free NH3+ in C-48, attributed to the random distribution of units, is related to its higher solubility Although C-48 is expected to exhibit hydrophobicity because of a higher amount of acetyl groups compared to C-2, the results of this work indicate the opposite In addition, GlcN and GlcNAc distribution in chitosan influenced hydrophilicity/hydrophobicity more than DA For coating cardboard papers, chitosan of 2% DA obtained from heterogeneous chitin deacetylation can be considered an advantageous alternative to films from synthetic polymers The paper-chitosan packaging system is environmentally friendly due to its high recyclability and biodegradability Acknowledgments This work was financially supported by Brazilian entities FAPESP (grant #2016/25120-7, São Paulo Research Foundation - FAPESP), CAPES (Coordination for the Improvement of Higher Education Personnel) and CNPq (grant #249270/2013-7, National Council for Scientific and Technological Development - CNPq) We want to thank Prof Dr Laurent David and Prof Dr Thierry Delair from the Département Matériaux et Ingénierie des Surfaces - Ingénierie des Matériaux Polymère - Université Claude Bernard Lyon References Aiba, S I (1991) Studies on chitosan: Evidence for the presence of random and block copolymer structures in partially N-acetylated chitosans International Journal of Biological Macromolecules, 13(1), 40–44 https://doi.org/10.1016/0141-8130(91) 90008-I Aloui, H., Khwaldia, K., Slama, M B., & Hamdi, M (2011) Effect of glycerol and coating weight on functional properties of biopolymer-coated 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decrease of the preferential pathway for water across the cellulose network filled with chitosan suspension In the present study, ... spectra of (a) C-2 and (b) C-48 Table Properties of cardboard paper samples coated with three layers of C-2 and C-48 suspensions and uncoated cardboard paper (P) Properties Cardboard Paper Samples