Accepted Manuscript Preparation and characterization of nanocomposites of polyvinyl alcohol/cellulose nanowhiskers/chitosan Hong-Zhen Li, Si-Chong Chen, Yu-Zhong Wang PII: DOI: Reference:
S0266-3538(15)00191-8 http://dx.doi.org/10.1016/j.compscitech.2015.05.004 CSTE 6102
To appear in:
Composites Science and Technology
Received Date: Revised Date: Accepted Date:
17 March 2015 1 May 2015 4 May 2015
Please cite this article as: Li, H-Z., Chen, S-C., Wang, Y-Z., Preparation and characterization of nanocomposites of polyvinyl alcohol/cellulose nanowhiskers/chitosan, Composites Science and Technology (2015), doi: http:// dx.doi.org/10.1016/j.compscitech.2015.05.004
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Preparation and characterization of nanocomposites of polyvinyl alcohol/cellulose nanowhiskers/chitosan Hong-Zhen Li, Si-Chong Chen*, Yu-Zhong Wang*
Center for Degradable and Flame-Retardant Polymeric Materials, College of Chemistry, State Key Laboratory of Polymer Materials Engineering, National Engineering Laboratory of Eco-Friendly Polymeric Materials (Sichuan), Sichuan University,
; Fax: +86-28-85410755; Tel: +86-28-85410755.
water-evaporation process. The material properties of these “green” hybrid films were characterized extensively using various techniques. Electrostatic interaction between CNWs and chitosan as well as the hydrogen bonds between the three materials played a very important role in determining the properties of the composites. The antimicrobial property, oxygen barrier property and mechanical property of the nanocomposites were all improved owing to the localized aggregations of CNWs and chitosan driven by the electrostatic interactions, thereby making the nanocomposites potentially useful for many applications including food packaging and antimicrobial packaging. Keywords: A. Nano composites, A. Polymers, B. Mechanical properties 1
1. Introduction In recent times there is growing interest to develop materials with film forming capacity, antimicrobial[1, 2] and oxygen barrier properties which improve food safety and shelf-life. Among the packaging systems, PVA is a promising polymer for food packaging, for its good flexibility, transparency, toughness, biocompatibility, barrier properties, nontoxicity, and biodegradability. To bring antimicrobial and improved oxygen barrier properties to PVA is an effective way to expand its applications in food packaging. Chitosan is a biocompatible natural polymer with strong antimicrobial effects and excellent oxygen-barrier properties due to its high crystallinity and the hydrogen bonds between the molecular chains[6, 7]. So it is a promising way to introduce antimicrobial and improved oxygen barrier properties to PVA by blending with chitosan. Although the blends of PVA/chitosan are the promising materials for food packaging owing to the combination of excellent film forming property of PVA and antimicrobial effects of chitosan, brittleness are still the main obstacles for its application. Percentage elongation at break of PVA/chitosan blends would drastically decrease with an increase in chitosan content, which has been reported by various researchers[9, 10]. Incorporating nanofillers into the PVA/chitosan blended system is one of most facile method for dissolving such problems and expanding the applications. George et al. prepared PVA nanocomposites containing cellulose nanocrystals and silver nanoparticles, and they found that the addition of AgNPs increased the elongation properties without compromising on other mechanical 2
properties and at the same time it significantly reduced the moisture sorption. Uddin et al. prepared fibers of PVA/chitin whiskers (ChWs) by gel spinning with outstanding enhancement in mechanical- and anti-creep properties. Pandele et al. prepared nanocomposites based on chitosan-PVA (CS-PVA) and graphene oxide (GO) by casting method, and their composites were mechanically strong and exhibited improved thermal stability. Huang et al. prepared nanocomposite films of chitosan, multiwalled carbon nanotubes (MWCNTs) and PVA by a solution casting. The results indicated that MWCNTs treated by chitosan dispersed well in the PVA matrix, and the tensile properties and water resistance of nanocomposites were improved greatly compared with neat PVA. However, with the growing environmental and energy problem, finding completely renewable nanofillers having good compatibility with both PVA and chitosan so as to enhance the properties of blend still remains a challenge. In this work, we developed a novel strategy for preparing PVA/chitosan based film with good comprehensive performance by introducing cellulose nanowhiskers (CNWs). The extraction of cellulose whiskers from renewable sources has gained more attention in recent years due to their exceptional mechanical properties (high specific strength and modulus), large specific surface area, high aspect ratio, environmental benefits and low cost[15, 16], and have already been used as nanofillers for many polymeric nanocomposites including PVA[17, 18] and chitosan. Hydrogen bonds and electrostatic interactions between the negatively charged sulfate groups on the whisker surface and the positively charged ammonium groups of chitosan were the driving forces for the 3
good combination of chitosan and CNWs[20-22]. Therefore, incorporating CNWs to PVA/chitosan blends by taking the advantage of interactions between CNWs, chitosan and PVA induced by electrostatic interaction and hydrogen-bond interaction is a feasible and effective method for preparing PVA/chitosan based material with controlled microstructure and enhanced properties. 2. Materials and methods 2.1. Materials Chitosan sample (DD≥90% and molecular weight 5000 Da) was obtained from Golden-Shell Pharmaceutical Co. Ltd, (Zhejiang, China). Ramie was obtained from JB RAMIE Sichuan Co. Ltd, China. PVA (PVA-1799, degree of polymerization 1700, degree of hydrolysis 99%) was obtained from Yunwei Company (Qujing City, Yunnan Province, China) and was rigorously dried at 80 °C in vacuo until a constant weight was obtained, and then stored in a desiccator under vacuum at room temperature over P2O5. 2.2. Preparation of CNWs Sulfuric acid hydrolysis of ramie was performed as described in the literature with minor modiﬁcations. Brieﬂy, the ramie fiber was cut into pieces. Then, 10 g of cellulose was added to 200 mL of 64 wt% sulfuric acid under strong mechanical stirring. Hydrolysis was performed at 55 °C for 30 min. After hydrolysis, the suspensions were then washed with deionized water using repeated centrifuge cyclings. The suspensions that couldn’t be centrifuged were collected, which reached pH ∼5. The ﬁnal concentration of the CNWs dispersions was about 1 wt%. 4
2.3. Preparation of PVA/CNWs/chitosan nanocomposites Chitosan solution (20 wt%) was obtained by dissolving 20 g chitosan powders in 80 g deionized water with continuous stirring at room temperature for 1 h. Acetic acid was not necessary because the low-molecular-weight chitosan was water soluble. PVA solution (5 wt%) was prepared by dissolving 10 g PVA powders in 190 g deionized water with continuous stirring at 90 °C. Then, chitosan and PVA solutions and CNWs dispersions were blended together by a homogenizer to form a homogeneous PVA/CNWs/chitosan suspension. PVA/CNWs/chitosan suspensions with different weight percentages (as indicated in Table 1) were obtained. Then they were poured into the glass plate. After 48 h setting, the PVA/CNWs/chitosan suspensions were de-bubbled and then transferred into a 60 °C oven for about 24 h drying. After that, the blend films were vacuum dried for 24 h at 80 °C in order to remove the residues of
PVA/CNWs/chitosan-x%, where x% is the content of chitosan in PVA/CNWs/chitosan nanocomposite films. As a comparison, PVA/CNWs and PVA/chitosan binary blend films were separately prepared in the same way as described above. For all the PVA/CNWs and PVA/CNWs/chitosan films, the weight content of CNWs is 1%. 2.4. Characterization The morphologies of CNWs and PVA/CNWs/chitosan composite films were respectively observed by scanning electron microscopy (SEM, JSM-5900LV, JEOL, Japan) at an accelerating voltage of 5 kV. CNWs was obtained by evaporating a drop of dilute CNWs aqueous suspensions. Brittle and tensile fracture surfaces were 5
obtained from PVA/CNWs/chitosan films. Before test, all the surfaces were coated with a layer of gold. The substructural morphology of PVA/CNWs/chitosan deposited from a dilute aqueous suspension was studied by transmission electron microscopy (TEM, Tecnai G2F20 S-TWIN electron microscope, FEI, Holland) at an accelerated voltage of 200 kV. 70-80 nm in thickness from films of PVA/chitosan and PVA/CNWs/chitosan composites were sliced using a RMC cryo-ultramicrotome equipped with a diamond knife and mounted on formvar-coated 200-mesh nickel grids. Ultrathin sections of unstained PVA/chitosan and PVA/CNWs/chitosan as well as uranyl acetate-stained PVA/CNWs/chitosan were separately studied by TEM. Surface morphology of PVA/CNWs and PVA/CNWs/chitosan nanocomposites was studied by using Nanoscope Multimode and Explore atomic force microscope (AFM, Veeco Instruments, USA). A definite weight of an aqueous suspension of PVA/CNWs and PVA/CNWs/chitosan was spin coated on a mica sheet. Height and amplitude images of the thin layer film were obtained with a tapping mode. The UV-vis spectra of the aqueous suspensions of PVA/CNWs/chitosan were recorded using a UV-vis spectrophotometer (Varian Cary 50) at a wavelength scan rate of 60 nm/min. And the UV-vis spectra were recorded for every 24 h for 3 days. Fourier transform infrared (FT-IR) analysis for PVA, CNWs, chitosan and PVA/CNWs/chitosan nanocomposites was conducted with a Thermo Nicolet 670 spectrometer from 4000 to 400 cm-1 at a spectral resolution of 4 cm-1 using KBr method. 6
X-ray diffraction (XRD) profiles were carried out on a Philips electronic instrument in steps of 0.03° using Cu Kα radiation at 40 kV and 150 mA between 5° and 60° (2θ). Oxygen permeability measurements were performed by using a gas permeation instrument (VAC-V1, Labthink Instrument Co, China) according to the standard of ASTM standard F2622-08. All samples were cut into circular discs with a diameter of 50 mm and a thickness of 1 mm, and each measurement was continued until a stable oxygen permeability rate was reached. The antimicrobial activity of PVA/CNWs/chitosan composite films was tested by an inhibition zone method. Two different food pathogenic bacteria including Staphylococcus aureus and Escherichia coli were used for testing the antimicrobial activity of the films. For the qualitative measurement of antimicrobial activity, the film samples were punched to make disks (diameter = 10 mm), and the antimicrobial activity was determined using a modified agar diffusion assay (disk test). The plates were examined for possible clear zones after incubation at 30 °C for 2 days. The presence of any clear zone that formed around the film disk on the plate medium was recorded as an indication of inhibition against the microbial species. Tensile tests were carried out according to ASTM D638 with a crosshead speed of 50 mm/min. The sample films (0.4-0.5 mm thickness) were stored at 50% relative humidity (RH) and 25 °C before testing. 3. Results and discussion Fig. 1A shows a scanning electron microscope (SEM) image of the ramie CNWs. 7
Observations of the CNWs obtained from diluted aqueous suspensions show some aggregates. The appearance of laterally aggregated elementary crystallites in SEM images is expected due to the high specific area and strong hydrogen bonds established between the whiskers. These aggregates may exist even in suspension but, when the dispersing medium is removed, as in the case of the SEM sample preparation, bundles of whiskers can be even more numerous than individualized rods. Fig. 1B shows a transmission electron microscopy (TEM) image of CNWs made from a drop of diluted PVA/CNWs/chitosan suspensions. According to the SEM image (Fig. 1A) and TEM image (Fig. 1B), the mean values of the length (L) and diameter (D) of the isolated CNWs were separately determined to be ~200 nm and ~40 nm, respectively, giving an aspect ratio (L/D) of about 5. Aggregations of CNWs in PVA/CNWs/chitosan were discovered in TEM and AFM images in Fig. 1B and Fig. 1C. UV-vis spectroscopy in Fig. 2 shows the light transmittance of PVA/CNWs/chitosan aqueous suspensions at various times after the suspensions were prepared, and it can be seen that the light transmittance determined at 24 h had a distinct reduction compared with the value at 0 h. The electrostatic interaction between positive charges in chitosan and negative charges in CNWs was responsible for the formation of aggregations of chitosan and CNWs. Meanwhile, as shown in Fig. 2, the light transmittance of the suspensions almost keep unchanged after 24 h, suggested that the aggregations were in a stable state in PVA/CNWs/chitosan aqueous suspensions. CNWs and chitosan tend to aggregate and even precipitate out of the solution induced by the electrostatic interaction. 8
Meanwhile, PVA played a role as stabilizer for CNWs and chitosan in the aqueous suspensions because of the hydrogen bonding. Therefore, stable localized aggregations of CNWs and constant light transmittance were obtained when these intermolecular interactions reached equilibrium. FTIR spectra were used to evaluate and identify the interactions between PVA, CNWs and chitosan in the nanocomposites, as shown in Fig. 3. A band of PVA at 3320 cm-1 is attributed to the O-H stretching vibration of hydroxyl group. The main bands of chitosan at 1633 and 1520 cm-1 respectively correspond to the C=O stretching of acetyl groups and N-H bending vibrations (amide and amine groups). The bands at 1070 and 1034 cm-1 are due to the C-O stretching vibrations, whereas the broadband at 3200-3500 cm-1 is assigned to the O-H and N-H stretching. For pure CNWs, the band at 2900 and 1429 cm-1 are characteristic of C-H stretching and bending of -CH2 groups, respectively, whereas the peaks at 1165 and 1059 cm-1 are attributed to the saccharide structure. The broadband around 3200-3450 cm-1 in the nanocomposites moved slightly and became narrower compared to the spectra for pure PVA, CNWs and chitosan, which are due to the interaction between the negatively charged sulfate groups on the CNWs and the positively charged ammonium groups of chitosan[22, 27], as well as the hydrogen bonds between -OH groups of PVA, CNWs and the ammonium groups of chitosan. Moreover, compared to pure chitosan film, the band corresponding to N-H bending of the nanocomposites almost disappeared, suggesting also the formation of hydrogen bonds between chitosan, PVA and CNWs. 9
To further identify the intermolecular interactions in PVA/CNWs/chitosan nanocomposites， X-ray diffraction analysis (XRD) was performed. Fig. 4 presents XRD patterns of PVA, chitosan, CNWs and PVA/CNWs/chitosan nanocomposites. The low-molecular-weight chitosan showed no obvious peaks because of its amorphous structure. All peaks of CNWs disappeared in PVA/CNWs/chitosan nanocomposites, because the content of CNWs was too low. If there were no or weak interaction between chitosan and PVA molecules in the nanocomposites, each component would has its own crystal region in the blend fibers, and XRD patterns would be expressed as simple mixed patterns of chitosan and PVA with the same ratio as those for mechanical blending. In fact, the peak of PVA slightly shifted towards low 2θ from 20.2° to 19.8° with the incorporation of chitosan and CNWs in PVA. Moreover, the intensity of the peaks around 2θ = 19.8° decreased with increasing chitosan content in the blends. These evidences concluded that intermolecular interactions between chitosan and PVA molecule formed in the nanocomposites. Oxygen is the key factor that might cause oxidation, which initiates several food changes such as odor, color, flavor and nutrients deterioration, so obtaining films with proper oxygen barrier can help improving food quality and extending food shelf life. Oxygen permeabilities (OP) of PVA, PVA/CNWs and PVA/CNWs/chitosan films with various load of chitosan were given in Fig. 5. PVA/CNWs film had higher OP than pure PVA film, while PVA/CNWs/chitosan films had much lower OP than pure PVA film. Moreover, increasing chitosan content also slightly decreased OP of PVA/CNWs/chitosan films. Chitosan provides films with good oxygen barrier 10
properties, and polysaccharide content was increased with the increase of chitosan content, which should theoretically lead to a reduction in OP in PVA/CNWs/chitosan films. When comparing with pure PVA film, OP of PVA/CNWs/chitosan films were rather low and they showed excellent oxygen barrier properties. This result indicates the potential of PVA/CNWs/chitosan films to be used as a natural packaging to protect food from oxidation reactions. Antimicrobial property is very important for food packaging, and the antimicrobial property of PVA/CNWs/chitosan nanocomposites was evaluated by an inhibition zone method. All PVA/CNWs/chitosan samples showed a clear antimicrobial property on S. aureus and E. coli. Table 1 shows antimicrobial test results of PVA, PVA/CNWs, PVA/chitosan and PVA/CNWs/chitosan films against S. aureus and E. coli. Fig. 6 shows a picture of antimicrobial test of PVA/chitosan and PVA/CNWs/chitosan films against S. aureus. In comparison with PVA/chitosan samples, synergistic antimicrobial effects occurred for PVA/CNWs/chitosan. One of the reasons for the antimicrobial character of chitosan is its positively charged amino group which interacts with negatively charged microbial cell membranes, leading to the leakage of proteinaceous and other intracellular constituents of the microorganisms. So the concentration of amino groups of chitosan is a major factor that concerns the antimicrobial property of the nanocomposites. For PVA/chitosan blends (Fig. 7A), chitosan dispersed homogeneously in PVA matrix and no obvious phase separation was observed. Therefore, the local concentration of amino groups of this sample was much smaller than that of neat chitosan owing to the dilution effect of PVA matrix. 11
When the negatively charged CNWs were introduced to the polymer blend, obvious difference in contrast between the chitosan phase and PVA matrix could be observed without staining (Fig. 7B), because of the amino groups of the chitosan. The chitosan phase showed darker gray scale than the PVA matrix in the TEM image. In order to observe the dispersity of CNWs, we also used uranyl acetate as staining agent for TEM test of PVA/CNWs/chitosan nanocomposites. The uranyl acetate can characteristically stain the PVA and chitosan. Therefore, some bundles of whiskers (as marked by black arrows), which showed lighter gray scale than PVA and chitosan, could be observed after staining by uranyl acetate (Fig. 7C). This phenomenon also suggested that the electrostatic interaction induced aggregations of CNWs and chitosan, and resulted in a sea-island phase separation morphology of chitosan phase in PVA matrix. The micro-phase separation of chitosan indicated that the PVA/CNWs/chitosan nanocomposites may have relatively high local amino concentration comparing to PVA/chitosan blends. Therefore, the PVA/CNWs/chitosan nanocomposites exhibited improved antimicrobial properties. Fig. 8 shows the tensile strength and elongation at break of PVA, PVA/CNWs, PVA/chitosan-15% and PVA/CNWs/chitosan nanocomposites with various load of chitosan. PVA/chitosan-15% composites had a much lower elongation at break than pure PVA. Since chitosan films are typically very brittle, the addition of chitosan lowered the flexibility of the PVA film. While in contrast, PVA/CNWs nanocomposite films showed an obvious increase in both tensile strength and elongation at break compared with pure PVA films. The reinforced mechanical properties could be 12
ascribed to increased interactions between PVA chains and CNWs via hydrogen bonding, as well as the relative high strength, stiffness and low density of CNWs. PVA/CNWs/chitosan nanocomposites had a significant improvement in flexibility compared with PVA/chitosan films. The elongation at break of PVA/CNWs/chitosan varied little with the addition of chitosan, and at the same time, the tensile strength of PVA/CNWs/chitosan nanocomposites had a slightly reduction. In combination with Fig. 1 and 5, the aggregation structure of CNWs and chitosan driven by the electrostatic interactions could be responsible for the mechanical properties of PVA/CNWs/chitosan films. Fig. 9 shows the SEM images of fracture surfaces of PVA/CNWs/chitosan nanocomposites. A smooth surface and a rough surface were separately obtained by brittle fracture and tensile fracture. The roughness of the tensile fracture surface was owing to the intermolecular interactions in PVA/CNWs/chitosan, like hydrogen-bond interaction and electrostatic interaction, which was in conformity with the properties discussed above. 4. Conclusions PVA/CNWs/chitosan nanocomposites were prepared by an environmentally friendly casting method. A novel strategy was carried out for enhancing with good comprehensive performance of PVA/chitosan based films by incorporating with cellulose nanowhiskers (CNWs) to prepare a ternary nanocomposite system based on intermolecular interactions, including the electrostatic interaction and hydrogen-bond interaction. The antimicrobial property has been confirmed by an inhibition zone 13
method, and a better antimicrobial property was obtained when CNWs was introduced. Chitosan significantly enhanced the oxygen barrier property of the nanocomposite films. The mechanical property of the nanocomposites was improved comparing to those of PVA/chitosan blends owing to the localized aggregations of CNWs and chitosan driven by the electrostatic interactions. With all those excellent properties, PVA/CNWs/chitosan nanocomposites possess a huge potential in the field of food packaging. Acknowledgments This work was financially supported by the National Natural Science Foundation of China (21474066). The Analytical and Testing Center of Sichuan University provided TEM analysis. References  Dutta P, Tripathi S, Mehrotra G, Dutta J. Perspectives for chitosan based antimicrobial films in food applications. Food Chem 2009;114(4):1173-82.  Li M, Li G, Jiang J, Tao Y, Mai K. Preparation, antimicrobial, crystallization and mechanical properties of nano-ZnO-supported zeolite filled polypropylene random copolymer composites. Compos Sci Technol 2013;81:30-6.  Yourdkhani M, Mousavand T, Chapleau N, Hubert P. Thermal, oxygen barrier and mechanical properties of polylactide-organoclay nanocomposites. Compos Sci Technol 2013;82:47-53.  Qiu K, Netravali AN. Fabrication and characterization of biodegradable composites based on microfibrillated cellulose and polyvinyl alcohol. Compos Sci Technol 14
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Legend of figures Table 1 Inhibition of E. coli and S. aureus growth by films of pure PVA, PVA/CNWs, PVA/chitosan and PVA/CNWs/chitosan with various load of chitosan Fig. 1. SEM (A) image of CNWs made from diluted CNWs suspensions, TEM (B) and AFM (C) images of CNWs made from a drop of diluted PVA/CNWs/chitosan suspensions. Fig.
PVA/CNWs/chitosan-20% (right) aqueous suspensions at various times after the suspensions were prepared. Fig. 3. FTIR spectra of PVA (a), CNWs (b), chitosan (c), and PVA/CNWs/chitosan nanocomposites (d). Fig. 4. XRD patterns of PVA, chitosan, CNWs and PVA/CNWs/chitosan nanocomposites with various load of chitosan. Fig. 5. Oxygen permeability of films of PVA, PVA/CNWs and PVA/CNWs/chitosan with various load of chitosan. 18
Fig. 6. Photograph of antimicrobial test results of PVA/chitosan (left) and PVA/CNWs/chitosan
PVA/CNWs/chitosan-25% film; (6) PVA/chitosan-5% film; (7) PVA/chitosan-10% film; (8) PVA/chitosan-15% film; (9) PVA/chitosan-20% film; (10) PVA/chitosan-25% film. Fig. 7. TEM images of ultrathin sections of PVA/chitosan without staining (A), PVA/CNWs/chitosan without staining (B), PVA/CNWs/chitosan stained by uranyl acetate (C). Fig. 8. Tensile strength (right) and elongation at break (left) of PVA, PVA/CNWs, PVA/chitosan-15% and PVA/CNWs/chitosan with various load of chitosan after stored at 50% relative humidity (RH) and 25 °C. Fig. 9. SEM images of fracture surfaces of PVA/CNWs/chitosan with brittle fracture (left) and tensile fracture (right).
Table 1 Inhibition of E. coli and S. aureus growth by films of pure PVA, PVA/CNWs, PVA/chitosan and PVA/CNWs/chitosan with various load of chitosan Samples
Inhibition zone (mm) of Inhibition zone (mm) of E. coli