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Modulating the properties of polylactic acid for packaging applications using biobased plasticizers and naturally obtained fillers
Journal Pre-proofs Modulating the properties of polylactic acid for packaging applications using biobased plasticizers and naturally obtained fillers Anshu Anjali Singh, Swati Sharma, Mayuri Srivastava, Abhijit Majumdar PII: DOI: Reference:
S0141-8130(19)37183-1 https://doi.org/10.1016/j.ijbiomac.2019.10.246 BIOMAC 13747
To appear in:
International Journal of Biological Macromolecules
Received Date: Revised Date: Accepted Date:
5 September 2019 23 October 2019 26 October 2019
Please cite this article as: A. Anjali Singh, S. Sharma, M. Srivastava, A. Majumdar, Modulating the properties of polylactic acid for packaging applications using biobased plasticizers and naturally obtained fillers, International Journal of Biological Macromolecules (2019), doi: https://doi.org/10.1016/j.ijbiomac.2019.10.246
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© 2019 Published by Elsevier B.V.
Modulating the properties of polylactic acid for packaging applications using biobased plasticizers and naturally obtained fillers Anshu Anjali Singh*, Swati Sharma, Mayuri Srivastava, Abhijit Majumdar* Department of Textile Technology, Indian Institute of Technology Delhi, New Delhi, India, 110 016. *Corresponding author Email: [email protected] *Corresponding author Email: [email protected]
Abstract The properties of PLA films intended for packaging applications have been modulated by using bio-based platicizers and naturally obtained fillers. Triethyl citrate (TEC) and glycerol triacetate (GTA) have been used as platicizers and halloycite nanotubes (HNT) and chitosan have been used as fillers. The addition of 10 wt.% TEC, 10 wt.% GTA and 3 wt.% HNT improves the ductility of PLA films, however, reduces the tensile modulus and tensile strength. Addition of chitosan (1 wt.%), on the other hand, acts as a good reinforcing filler and improves the tensile strength and tensile modulus. PLA-HNT-chitosan film show comparable tensile strength and tensile modulus and ~12 times higher elongation at break compared to pure PLA. Besides, PLA-HNT-chitosan film demonstrates very good barrier properties against moisture and ultraviolet (UV) rays. Additionally, its antibacterial efficacy against E. coli and S. aureus are found to be around 80% and 70%, respectively. The study demonstrates the complementary effects of HNT and chitosan to modulate the properties of PLA film and indicates that the PLAHNT-chitosan film can emerge as a very potent material for packaging applications. Keywords: Chitosan; Halloysite nanotubes; Polylactic acid; Antibacterial; Water vaour transmission.
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1. Introduction Global proliferation of commodities made of plastic materials results in rapid depletion of nonrenewable fossil fuels together with the global-warming caused by the production of carbon dioxide from the combustion. Most of the food packaging materials are manufactured from non-biodegradable and petrochemical based plastics which persist for a long time in the environment and their end-life cannot be anticipated easily [1]. In view of this, there is an impending need to develop materials from renewable resources and therefore, efforts need to be emphasised on the production of environment friendly sustainable packaging materials having the potential to replace petrochemical based plastics. Extensive research works have been performed in this context globally and scientists and researchers have proposed several biopolymers, which are bio-degradable, sustainable and acquire similar properties to that of conventional plastics. Among different biopolymers available, polylactic acid (PLA) is considered as one of the most promising for the production of sustainable and green packaging materials. PLA is a biodegradable, non-toxic, aliphatic thermoplastic polyester, obtained from renewable resources like corn, sugar beet and potato. It is considered as safe by the US Food and Drug Administration (FDA) for food packaging applications [2]. The strength and stiffness of PLA is similar to that of polyamide and polycarbonate. Moreover, PLA has good transparency, biocompatibility and biodegradability. However, the disadvantages of PLA like embrittlement, low thermal stability, poor gas and UV ray barrier properties, antimicrobial properties limit its application. Therefore, these properties need to be improved or tuned adequately in order to develop a suitable packaging material [3, 4]. Several approaches have been adopted to enhance the properties of PLA, for example, use of plasticizers, incorporation of micro or nano fillers etc. However, to keep the material as green as possible, it is important to use all the components which can be obtained naturally or can be degraded easily. 2
One of the most important considerations while developing a food packaging material is to choose non-toxic additives and fillers. Bio-based plasticizers like polyethylene glycol, citrate esters, triacetin, oligomeric lactic acid have been used to improve the ductility of PLA but at the cost of stiffness, tensile strength and glass transition temperature, as these properties generally deteriorate concurrently with the addition of plasticizers [5-7]. In addition, composites of PLA reinforced with naturally obatined additives/fillers are of great interest and the improvement in properties depends on the dispersion of the filler and good interfacial adhesion between the matrix and fillers. However, for the composites having matrix and fillers of different polarities like PLA and cellulose or chitosan, a third component needs to be used to improve the compatibilty between matrix and fillers and hence their properties. For example, Zakaria et al. [8] used epoxidized natural rubber as a compatibilizer for PLA-chitosan composite. Similarly, Tham et al. [3] studied the effect of 5-20 wt.% of epoxidized natural rubber on different properties of PLA-halloysite nanotubes. HNT is an aluminosilicate (Al2Si2O5(OH)4.2H2O) with hollow tubular structure, and is obtained naturally [9]. It has high stiffness and exhibit unique surface property due to the multilayered structure with few hydroxyl (-OH) groups [10]. HNT efficiently improves the strength, stiffness and thermal properties of the polymer matrix and can be considered as a potential nano-reinforcement [11, 12]. Although it is a promising material, however, limited work has been reported on its use in polymer composite. Like HNT, chitosan is also a natural biopolymer and is renewable, non-toxic and biodegrdable. It can be produced after the deacetylation of chitin, which is the second most abundant naturally occurred polysaccharide, after cellulose [13]. It is a linear copolymer composed of N-acetyl-D-glucosamine and D-glucosamine units linked by 1, 4-β-D-glucopyranosamine and is a polycationic polymer with one amino group (NH2) and two hydroxyl (-OH) groups in the repeating unit [14]. Chitosan has excellent antimicrobial behaviour due to the presence of positively charged amino groups, and hence is
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considered good for food packaging applications [15, 16]. Rapa et al. [17] reported good antifungal and antimicrobial properties of biocomposites containing tributyl-o-acetyl citrate plasticized PLA and 1-5 wt.% of chitosan. Chitosan together with nanoclay was also used as a coating material to improve the poor gas barrier properties of PLA [18]. The main objective of this work was to modulate the properties of PLA films with biobased plasticizers and naturally obtained fillers, in order to develop the sustainable packaging material. Triethyl citrate (TEC) and glycerol triacetate (GTA) were used as plasticizers as both are biodegradable and non-toxic. Further to modulate the properties of plasticized PLA, halloysite nanotubes (HNT) and chitosan were used as fillers. PLA films with triethyl citrate, glycerol triacetate, halloysite nanotubes and chitosan, in different combinations, were prepared by solvent casting methods and consequent effect on mechanical, thermal, barrier and antibacterial properties was investigated.
2. Experimental 2.1 Materials Polylactic acid (PLA) pellets, Ingeo 4043D grade was purchased from NatureWorks Co. Ltd. Coimbatore, India. Two bio-based plasticizers, namely triethyl citrate (TEC) and glycerol triacetate (GTA) were purchased from Sigma-Aldrich, India. Table 1 shows the structure and properties and of TEC and GTA. Halloysite nanotubes (HNT) and chitosan powder were purchased from Natural Nano Inc. USA and SRL Pvt. Ltd. India, respectively. The transmission electron micrograph (TEM) was used to study the morphology of HNT (Fig. 1a) while the morphology of chitosan was studied using scanning electron micrograph (SEM) (Fig. 1b). HNT had length in the range of 200 nm to 1300 nm and diameter in the range of 50 nm to 200 nm. The size of chitosan powder varied from 2 µm to 20 µm. Methylene chloride or
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dichloromethane (DCM) was purchased from Fisher Scientific, India. All the chemicals were used as received. Table 1 - Properties of plasticizers Name Triethyl citrate (TEC)
Molecular weight (g/mol) 276.285
Chemica l formula C12H20O7
218.205
C9H14O6
Glycerol triacetate (GTA)
Chemical structure
Figure 1. (a) transmission electron micrograph of HNT and (b) scanning electron micrograph of chitosan
2.2 Preparation of PLA films All the samples were prepared by solvent casting method. At first, PLA, HNT and chitosan were dried at 40 °C for 24 hours in order to remove the moisture. Requisite amount of PLA pellets-plasticizers-HNT-chitosan were added in the dichloromethane (DCM) and the suspension (10 wt.% of solid content) was than continuously stirred at room temperature for 24 hours. Thereafter, the mixed suspension was poured into a glass petri dish and DCM was allowed to evaporate overnight at room temperature in a fume hood followed by drying in an 5
oven at 40 °C for 24 hours. The dried film was then peeled off carefully for further use. In this study, nine different films were prepared and the details about their formulation are reported in Table 2. The amount of plasticizer (TEC or GTA), HNT and chitosan used was 10 wt.%, 3 wt.% and 1wt.%, respectively. The thickness of all the films was approximately 0.3 mm.
Table 2 - Sample coding and formulation of the films Sample coding PLA PLA-TEC PLA-GTA PLA-HNT PLA-TEC-HNT PLA-GTA-HNT PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
PLA (wt.%) 100 90 90 97 87 87 96 86 86
TEC (wt.%) 10 10 10 10 -
GTA (wt.%) -
HNT (wt.%) -
Chitosan (wt.%) -
10 10
3 3 3 3 3 3
1 1 1
3. Testing and Characterizations 3.1 Morphology The surface morphology of HNT and chitosan was characterized by transmission electron microscopy (TEM) and scanning electron microscopy (SEM) using JEM-1400 (JEOL) and LEO 982 (Zeiss), respectively. The samples for TEM was prepared by applying a drop of diluted suspension of HNT on carbon coated grid followed by drying in an oven at 50 °C for overnight. Chitosan powder was coated with gold before the SEM analysis.
3.2 Mechanical properties Tensile test of rectangular shaped samples having a width of 10 mm was performed using universal tensile testing machine (Instron 3365) equipped with a 500 N load cell. Five specimens for each sample were tested with a gauge length of 20 mm and cross-head speed of
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5 mm/min. The tensile modulus, ultimate tensile strength and elongation at break were evaluated.
3.3 Thermal characterization Thermal behaviour of all the samples was examined using differential scanning calorimetry (DSC Q 2000, TA) under nitrogen atmosphere. Approximately 2.5 mg of sample was sealed in an aluminium crucible and heated gradually from 0 °C to 200 °C at a heating rate of 10 °C/min for first heating scan. The samples were then held at 200 °C for 5 minutes to remove the thermal history. Further, they were cooled to 0 °C at a cooling rate of 10 °C/min for the cooling scan before a second heating scan from 0 °C to 200 °C at a heating rate of 10 °C/min. Melting temperature (Tm), cold crystallization temperature (Tcc) and glass transition temperature (Tg) and were determined from the 2nd heating scans.
3.4 X-ray diffraction Crystallinity percentage of the samples was analysed by the Ultima IV X-ray diffractometer (Rigaku) in reflection mode. X-ray diffractograms (XRD) of the film samples were recorded at room temperature with Cu α radiation. The scan was performed at a speed of 3°(2θ)/min and the data were collected at 0.02°/interval over the 2 θ range from 10 to 40°. Percentage crystallinity (߯ %) of the sample was calculated using Eq. 1. [19]. ߯ ሺΨሻ ൌ
௨ௗ௧ ்௧
ൈ ͳͲͲ
(1)
3.5 UV-Vis spectroscopy Light transmittance of the films was measured using a Perkin Elmer UV/VIS Spectrometer Lambda 1050. Rectangular shaped film specimen was placed directly in a test cell and the scan
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was carried out from 300 to 800 nm wavelength with a scan speed of 240 nm/min using air as the reference.
3.6 Water vapour permeability Water vapour transmission rate (WVTR) of the films was measured using an automatic water vapour permeability testing machine (Labthink W3/060). Each sample having an area of 33 cm2, was tested at 38 °C and relative humidity of 80 %. Three specimens for each sample were tested and the average value was taken for analysis.
3.7 Anti-bacterial activity The antibacterial properties of pure PLA film and films with chitosan were studied against Gram-negative Escherichia coli (E. coli) and Gram-positive Staphylococcus aureus (S. aureus). Fresh bacteria of E. coli and S. aureus were first activated by incubating them at 37 °C for 24 hours in a fresh nutrient agar medium. Freshly grown colonies of both the species of bacteria were then placed in the nutrient broth and incubated again at 37 °C for 24 hours. After 24 hours, the bacterial suspension was diluted to a concentration of approximately 105 CFU/ml. The properly sterilized samples (20 mm × 10 mm) were then exposed to the bacterial suspension at 37 °C for 24 hours. Finally, the incubated bacterial suspension with samples were cultured on the petri dishes with agar medium. These petri dishes were incubated at 37 °C for 24 hours and the colony forming units (CFU) were counted. The antibacterial efficiency (AE %) of the samples was calculated using Eq. 2, where B and A are the total number of bacterial colonies in pure PLA (reference) and PLA films with chitosan, respectively [20]. ݕ݂݂݈ܿ݊݁݅ܿ݅݁ܽ݅ݎ݁ݐܾܿܽ݅ݐ݊ܣሺܧܣΨሻ ൌ
8
ି
ൈ ͳͲͲ
(2)
4. Results and Discussion 4.1 Mechanical Properties The effects of addition of plasticizers (TEC and GTA) and fillers (HNT and Chitosan) on the tensile properties are shown in the Fig. 2(a-c) and Table 3. It can be observed from the Fig. 2 and Table 3, that the presence of plasticizers and fillers significantly influence the mechanical properties of PLA. As expected, pure PLA showed brittle failure with an elongation at break of around 6 %, whereas this value is found to be much higher for PLA-TEC (390 %) and PLAGTA (418 %). However, tensile modulus (0.96 GPa and 0.75 GPa, respectively) and ultimate tensile strength (30 MPa and 27 MPa, respectively) decreases, indicating that TEC and GTA efficiently plasticizes the PLA, making it more flexible and ductile [21, 22]. However, subtle difference in the tensile properties can be noticed between PLA-TEC and PLA-GTA with latter showing lower strength and modulus but higher elongation at break. In addition, tensile strength and tensile modulus of PLA-HNT reduce to 40 MPa and 1.6 GPa, respectively, then those of pure PLA i.e., 47 MPa and 2.4 GPa, respectively, while the elongation at break increases to 91 %. Improvement in elongation at break or ductility of PLA after the addition of nano-fillers has been widely reported. Li et al. [23] reported that the stress-whitening induced by the extensive crazing is responsible for the improvement in strain at break of PLA nanocomposite after the addition of 0.5 to 5 wt.% of organically modified rectorite. Jiang et al. [24] reported that the PLA with organically modified montmorillonite and nano-sized precipitated calcium carbonate showed significant improvement in the strain at break compared to pure PLA. Liu et al. [11] studied the effect of HNT content on PLA and reported that the elongation at break of PLA-HNT nanocomposites with less than 20 phr (parts per hundred of PLA) HNT has higher value than that of pure PLA, as the small quantity of HNT could acts as a plasticizer.
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Furthermore, PLA-TEC-HNT and PLA-GTA-HNT show slight improvement in the elongation at break compared to those of PLA-TEC and PLA-GTA, respectively, while insignificant change in ultimate tensile strength and tensile modulus are observed. This might be due to the synergistic effect of plasticization of TEC or GTA and small quantity of HNT in PLA [11]. Rashmi et al. [25] also reported the improved ductility of the PLA/Polyamide11 (PLA/PA11) blend after the addition of 2 to 6 wt.% of HNT compared to PLA/PA11 blend. All the specimens of PLA with HNT show prominent stress-whitening and this phenomenon causes ductile failure during tensile test compared to the brittle failure in case of pure PLA [22]. It is known that the addition of filler usually improves the tensile strength and tensile modulus of the polymer composites [26]. However, in this study the addition of HNT in pure PLA and plasticized PLA (i.e. PLA-TEC and PLA-GTA) increases the elongation at break, which might be due to the immense crazing caused by the stress-whitening as well as plasticization together with slippery effect of HNT in PLA [11, 22-24, 27]. Stress whitening led to the enhanced opacity in the tensile fractured specimens and is attributed to the variations in the refractive index caused either by the formation of micro voids or change in the structure [22, 27].
Table 3 - Tensile properties of PLA and its composites Sample PLA PLA-TEC PLA-GTA PLA-HNT PLA-TEC-HNT PLA-GTA-HNT PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
Tensile modulus (GPa) 2.40 (± 0.10) 0.96 (± 0.05) 0.75 (± 0.05) 1.60 (± 0.03) 0.62 (± 0.03) 0.74 (± 0.03) 2.10 (± 0.30) 1.10 (± 0.20) 0.97 (± 0.20) 10
Ultimate tensile strength (MPa) 47 (± 2.6) 30 (± 0.7) 27 (± 0.5) 40 (± 0.3) 27 (± 0.3) 26 (± 1.0) 48 (± 2.5) 34 (± 3.5) 35 (± 3.2)
Elongation at break (%) 6 (± 0.4) 390 (± 34) 418 (± 17) 91 (± 7) 468 (± 46) 490 (± 78) 58 (± 25) 108 (± 22) 132 (± 27)
Figure 2 - Tensile stress-strain curve of the PLA and its composites
Further, after the addition of 1 wt.% chitosan as filler, tensile properties of PLA-HNTchitosan, PLA-TEC-HNT-chitosan and PLA-GTA-HNT-chitosan were also explored. It is observed (Fig. 2 and Table 3) that the addition of chitosan causes significant improvement in tensile modulus and tensile strength compared to the corresponding samples without chitosan. This indicates that the chitosan acts as a good reinforcing material and establishes good interfacial interactions with the components of the composite [26]. However, elongation at break decreases which might be attributed to the stiffening action of the chitosan that causes substantial local stress concentration and consequently failure at lowered strain [28]. Although, the elongation of PLA-TEC-HNT-chitosan (108 %) and PLA-GTA-HNT-chitosan (132 %) are found to be higher than that of PLA-HNT-chitosan (58 %) due to the predominating plasticizing effects of TEC and GTA, respectively. Of all the composites, PLA-HNT-chitosan exhibited comparable tensile strength and modulus as of pure PLA with higher elongation at
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break, indicating that PLA, HNT and chitosan has good interfacial interaction altogether. Two different kind of hydrogen bonds are formed in the PLA-HNT-chitosan composite; one between the carbonyl groups of PLA and hydroxyl group of HNT and chitosan while other between the carbonyl groups of PLA and amino group of chitosan. The proposed hydrogen bond formation between the PLA, HNT and chitosan are shown in the Fig. 3. Similar findings were also reported by Zakaria et al. [8] and Vasile et al. [29]. Zakaria et al. [8] reported that the composite of PLA, chitosan and epoxidised natural rubber (ENR) had higher tensile modulus and tensile strength than PLA/ENR and also the tensile modulus of the former was higher than that of pure PLA. Vasile et al. [29] reported that PLA, polyethylene glycol (PEG) and chitosan composite had higher modulus than that of the PLA/PEG. Overall, the present study demonstrates the complementary effects of HNT and chitosan on the tensile properties of PLA, by efficiently improving the ductility of the composite without compromising the tensile modulus and tensile strength.
Figure 3 –Proposed hydrogen bond interaction between PLA, HNT and chitosan 12
4.2 Thermal Properties DSC analysis of all the samples was carried out to study the effect of plasticizers and fillers on the thermal behaviour of PLA. The DSC thermograms of all the samples, obtained during the second heating scan are shown in Fig. 4 and the values extracted from the thermograms are reported in Table 4. The second heating scan of DSC thermograms indicate the presence of glass transition (Tg) endotherm, cold crystallization (Tcc) exotherm and melting (Tm) endotherm, which are typical of any semi-crystalline polymer. The Tg, Tcc and Tm of pure PLA film are 61.5 °C, 125 °C, and 149.7 °C, respectively. Values of Tg, Tcc and Tm decreases for PLA-TEC and PLA-GTA due to the enhanced flexibility of polymer chains in presence of TEC and GTA, respectively [21, 30]. The shift in the Tcc to lower temperature is attributed to the increased crystallinity, which is further confirmed by the XRD analysis [31]. In addition, the Tg and Tm recorded for PLA-TEC (44.5 °C and 143.1 °C, respectively) are found to be slightly higher than that of PLA-GTA (42.2 °C and 141.2 °C, respectively). This change in the thermal behaviour can be attributed to the lower molecular weight of GTA (218.205 g/mol) than that of TEC (276.85 g/mol) [32]. Similar observation has been reported by the Ljungberg et al. [33] for PLA plasticized with tributyl citrate and Martin et al. [34] for PLA plasticized with different molecular weight of polyethylene glycol. Miniscule change in the Tg and Tm are observed for PLA-HNT composite with respect to pure PLA, however, the intensity of Tcc decreased compared to pure PLA. Addition of Chitosan does not alter the Tg and Tcc when compared to the respective samples without chitosan but Tm seems to decrease in PLA-HNT-chitosan only (from 149.3 °C to 142.2 °C). The decreased Tm could be attributed to the thinner lamellar thickness of PLA [35, 36].
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Table 4 - Thermal properties and crystallinity percentage of PLA and its composites Sample
PLA PLA-TEC PLA-GTA PLA-HNT PLA-TEC-HNT PLA-GTA-HNT PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
Tg (°C) 61.5 44.5 42.2 61.4 42.5 41.4 60.7 41.2 -
DSC Tcc (°C) 125 112 111 105 110 105 112
Tm (°C) 149.7 143.1 141.2 149.3 142.1 142.7 142.2 142.2 143.7
XRD Crystallinity (%) Amorphous 43.6 41.7 34.7 39.4 38.2 24.4 28.8 22.7
Figure 4 – DSC thermograms of PLA and its composites 4.3 X-Ray diffraction XRD was performed to investigate the effect of plasticizers and fillers on the crystallinity of PLA. X-ray diffractograms of all the samples are presented in Fig. 5 and the calculated
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crystallinity percentage are reported in Table 4. Only one broad XRD spectrum, without any sharp peak, is found to be present in pure PLA (Fig. 5a) at around 2θ = 16.7°, which is characteristic of the (200/110) crystallographic plane, signifying pure PLA to be amorphous [11, 30]. This indicates that the PLA does not get enough time to crystallize during the sample preparation by solvent casting method as the solvent evaporates quickly. The XRD profile of PLA-TEC, PLA-GTA and PLA-HNT exhibit profound diffraction peaks as the intensity is found to be sharper and stronger than that of pure PLA implying higher crystallinity. Sharp diffraction peaks at 2θ = 12.09°, 14.7°, 16.7°, and 22.3° corresponding to (004/103), (010), (200/110) and (105) crystallographic planes, respectively, are present in all the samples except for pure PLA [30]. The diffraction peaks at 2θ = 14.7° and 22.3° reflect the presence of α- form of PLA, while the peak at 2θ = 12.09° could be assigned to the D-form of PLA [30, 37]. Further, it can be noticed that the peak at 2θ = 19.90°, corresponding to (020/110) crystallographic plane, is present only in PLA-HNT and PLA-HNT-chitosan. The shift in the peak was observed at 2θ = 19.0° corresponding to (203) crystallographic plane, in the presence of TEC and GTA, indicating that the addition of plasticizer altered the stable α crystal of PLA [37].
Figure 5 –X ray diffractogram of PLA and its composites It can be seen from the Table 4 that the addition of TEC, GTA and HNT increase the crystallinity percentage of PLA. The degree of crystallinity of PLA-TEC (43.6 %) and PLA-
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GTA (41.7 %) is found to be much higher than that of pure PLA indicating that the plasticizers enhance the segmental molecular chain mobility of PLA and thus facilitates the crystallisation in plasticized PLA [21, 30, 34]. Besides, HNT acts as an effective nucleating agent for PLA as the addition of 3 wt.% of HNT increased the crystallinity of PLA to 34.7 % [11]. Slight decrease in percentage crystallinity is observed for PLA-TEC-HNT (39.4 %) and PLA-GTA-HNT (38.2 %) when compared to PLA-TEC and PLA-GTA, respectively. However, the addition of chitosan decreases the crystallinity of PLA films when compared to the respective samples without chitosan, although the values are still higher than that of pure PLA. Correlo et al. [35] and Bonilla et al. [38] also reported the similar trends. Correlo et al. [35] opined that the decreased crystallinity after the addition of chitosan can be attributed to the hydrogen bond formation between the carbonyl groups of polyester and -OH and -NH2 groups of chitosan in the amorphous phase. Senda et al. [36] reported that difference in the molecular mobility between matrix and chitosan could be another possible reason for lower degree of crystallinity, besides hydrogen bonding interaction.
4.4 Optical transparency Transmission of ultraviolet rays (UV) and visible light through the polymeric film is one of the important parameters in packaging application, in order to store and protect the products which are light sensitive [4]. The transmittance percent at different wavelengths of all the films are shown in the Fig. 6(a) and their visual appearances are shown in the Fig 6. (b-j). It is observed from the Fig. 6 that the pure PLA has the highest transmission in the visible region (400-800 nm) as well as in the UV region (< 400 nm), indicating its good transparency in the visible region but poor UV barrier property. The transparency can also be ratified from the visual appearance of PLA film shown in Fig. 6(b). The percent light transmittance (T %) of the PLA films with TEC, GTA and HNT are found to be lower than that of pure PLA. The recorded
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transmissions for the PLA, PLA-TEC, PLA-GTA and PLA-HNT, in the middle of the visible spectrum (i.e., 600 nm), are 92, 88, 87 and 84 %, respectively. Further, the incorporation of HNT in PLA-TEC and PLA-GTA shows slightly lesser transmission at 600 nm as the transmission recorded are 83 % and 80 % for PLA-TEC-HNT and PLA-GTA-HNT, respectively. It can be seen from the Fig 6 (b-j), that the visual appearance of PLA films with TEC, GTA and HNT are less transparent than that of pure PLA. The decreased transmittance of PLA films in the presence of plasticizers and HNT than that of pure PLA is due to the increased crystallinity and homogeneous dispersion of HNT in the composite [22, 39]. In addition, the result demonstrates (Fig. 6 and Table 5) that the percent transmittance of all the films decrease in the UV region and the films containing HNT shows better UV barrier property than the films of PLA, PLA-TEC and PLA-GTA. A reduction in approximately 32 %, 33 % and 41 % transmittance are observed in PLA-HNT, PLA-TEC-HNT and PLA-GTA-HNT than that of pure PLA at wavelength of 320 nm. This may be attributed to the higher reflection and absorption of UV rays by the HNT. However, interestingly, presence of chitosan shows miniscule change in the percent light transmittance in visible region as well as in UV region. This might be due to the bigger particle size of chitosan than HNT [22, 39]. The results imply that the nanocomposite PLA films possess improved barrier property particularly in the UV region, while maintaining the good transparency in the visible light spectrum. Table 5 –Transmittance % and WVTR of PLA and its composites Sample
PLA PLA-TEC PLA-GTA PLA-HNT PLA-TEC-HNT PLA-GTA-HNT PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
Transmittance % (λ = 600 nm) 92 88 87 84 83 80 84 83 81 17
Transmittance % (λ = 320 nm) 81 74 72 55 54 48 57 48 49
WVTR (g/m2.day) 39 (±2.5) 63 (±2.5) 57 (±1.5) 47 (± 4.5) 55 (±4.0) 50 (±5.6) 40 (± 4.5) 60 (± 5.2) 58 (±6.5)
Figure 6 - (a) Transmittance percent vs wavelength of PLA and its composites and visual appearance of film of (b) PLA, (c) PLA-TEC, (d) PLA-GTA, (e) PLA-HNT (f) PLA-TECHNT, (g) PLA-GTA-HNT, (h) PLA-HNT-chitosan, (i) PLA-TEC-HNT-chitosan and (j) PLA-GTA-HNT-chitosan
4.5 Water vapour transmission Water vapour transmission rate (WVTR) is another important barrier property of food packaging material as it determines the spoilage of moisture sensitive food. WVTR is defined as the amount of water vapour transmitted, under specific temperature and humidity, through unit area of film per unit time. Water vapour barrier property of a polymeric film or composite is mostly influenced by the hydrophobicity or hydrophilicity of matrix and reinforcement, amount and dispersion of the later and the presence of voids or cracks [40]. Table 5 shows the WVTR of all the samples. The WVTR is 39 g/m2/day for pure PLA and the addition of plasticizers and fillers substantially influence the transmission rate of water vapour. The WVTR increases by approximately 62 %, 46 % and 20 % for PLA-TEC, PLA-GTA and PLAHNT, respectively, compared to pure PLA. This may be due to the hydrophilic behaviour of the TEC and GTA as both the plasticizers are soluble in water and free hydroxyl (-OH) groups present on the surface of HNT. Although, the WVTR of PLA-HNT is found to be superior than the plasticized PLA and this could be due to the use of less amount of HNT (3 wt.%) than
18
plasticizers (10 wt.%). The findings are in good agreement with the results reported by the other researchers. Fan et al. [41] and Gao et al. [42] reported that the water vapour permeability of PLA films increased after the addition of acetyl-tributyl citrate (ATBC) and tributyl citrate (TBC) respectively. Dadashi et al. [40] reported that the PLA films with unmodified natural nanoclay showed slightly higher WVTR while the same decreased for the films with organically modified nanoclays. They attributed it to the hydrophilic nature of the former and hydrophobicity of the later. The addition of chitosan improves the WVTR of the films compared to the respective samples without chitosan. The lowest WVTR recorded in this study was 40 g/m2/day for PLA-HNT-chitosan, which is comparable to the hydrophobic pure PLA. The improvement in the water vapour barrier property of the PLA films after the addition of chitosan might be associated with its hydrophobicity due to the presence of acetyl groups of incompletely deacetylated chitosan [43]. The hydrogen bond interaction between PLA, HNT and chitosan reduces the availability of free hydroxyl group for interaction with water molecules and thus decreasing the water vapour transmission rate. The result reveals that the WVTR of the PLA film was influenced by the hydrophilicity/hydrophobicity of the material used, and the use of plasticizers like TEC or GTA is not desirable as they increase the WVTR. However, HNT and chitosan showed complementary effect in improving the water vapour barrier properties.
4.6 Antibacterial activity Two different species of bacteria i.e., E. coli (Gram-negative) and S. aureus (Gram-positive) were used to study the antibacterial property of PLA and its composites. Both the bacteria are considered as a pathogenic infectious and their symptoms range from mild to serious [44]. The images representing the bacterial colony growth are shown in Fig. 7 while the total bacterial colony count and antibacterial efficiency are presented in Table 6. The total number of visible
19
bacterial colonies are more for pure PLA than that of PLA with chitosan and thus the antibacterial efficiency of the later is significantly higher. This is due to presence of positively charged amino (-NH2) group in chitosan that binds easily with the negatively charged peptidoglycan, liposaccharides and proteins present on the cell wall of the bacteria, thus obstructing their biosynthesis and prevent the transportation of nutrient through the cell wall and eventually supress their growth [44]. In addition, it can also be observed that the PLA and PLA-chitosan composites are more resistant to E. coli than S. aureus, as the growth of bacterial colonies are more in case of the latter. Similar findings were also reported by other researchers [4, 45]. Hu et al. [46] also found that the antibacterial activity of chitosan is higher against the E. coli than that against S. aureus and this is due to the difference in the cell wall structure of these bacteria (Fig. 8). The cell wall of S. aureus consists of single thick layer of peptidoglycan while the cell wall of E. coli consists of thin peptidoglycan layer surrounded by an outer membrane.
Outer
membrane
present
in
the
Gram-negative
bacteria
contains
lipopolysaccharides and proteins which increases the negative charge on the surface and consequently confer resistance to hydrophobic compounds [47].
Figure 7 - Antibacterial activity of (a) PLA, (b) PLA-HNT-chitosan, (c) PLA-TEC-HNTchitosan and (d) PLA-GTA-HNT-chitosan, against E. coli and S. aureus
20
Table 6 – Antibacterial efficiency against E. coli and S. aureus Sample
Total bacterial colony
PLA PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
E. coli 32 6 7 7
S. aureus 369 108 102 102
Antibacterial efficiency % E. coli S. aureus 81 71 78 72 78 72
Figure 8. Cell wall structure of Gram-positive and Gram-negative bacteria
5. Conclusions The influence of addition of two bio-based plasticizers, namely triethyl citrate (TEC) and glycerol triacetate (GTA) and two fillers, namely halloysite nanotubes (HNT) and chitosan powder, in different combinations, on mechanical, thermal, barrier and anti-microbial properties of polylactic acid (PLA) films was studied. Tensile results revealed that the TEC and GTA efficiently plasticizes the PLA as the elongation at break increased significantly (~80 times) though tensile modulus and tensile strength reduced significantly. Addition of HNT also improved the elongation at break (~18 times) along with decrease in tensile modulus and tensile strength, though the extents of reduction were much lower as compared to those obtained in case of TEC and GTA. However, marginal improvement in ductility was noticed in PLA-TECHNT and PLA-GTA-HNT compared to those of PLA-TEC and PLA-GTA, respectively. Addition of chitosan in PLA-HNT, PLA-TEC-HNT and PLA-GTA-HNT improved the tensile modulus and tensile strength while reducing the elongation at break. PLA-HNT-chitosan showed comparable tensile strength and tensile modulus and ~12 times elongation at break as 21
compared to those of pure PLA, indicating good interfacial interactions between these components. Thermal study demonstrated that the Tg, Tcc and Tm decreased for PLA-TEC and PLAGTA, as compared to those of pure PLA, as a result of plasticization effect. XRD studies showed that, degree of crystallinity of the PLA films increased significantly with the addition of TEC, GTA and HNT, however, the incorporation of chitosan lowered the degree of crystallinity. UV-vis spectroscopy confirmed that the PLA has good transparency in the visible region but has poor UV barrier property. The overall reduction in the percentage transmittance was observed for PLA-TEC, PLA-GTA, PLA-HNT, PLA-TEC-HNT and PLA-GTA-HNT, when compared to pure PLA, indicating the improved UV barrier property of the PLA composite films. Negligible change in the transparency was observed in the presence of chitosan. Water vapour permeability (WVP) of PLA with HNT, TEC and GTA increased, however, addition of chitosan lowered the WVP. The lowest water vapour transmission rate was obtained in case of PLA-HNT-chitosan film, which was comparable to the hydrophobic pure PLA. Nanocomposite films with chitosan showed good antibacterial properties against E. coli and S. aureus and the former being more resistant due to the difference in the cell wall structure. Overall, the results revealed that the properties of PLA could be easily modulated by using TEC, GTA, HNT and Chitosan in combinations. The results also demonstrated the complementary effect of HNT and chitosan as PLA-HNT-chitosan showed improved ductility without compromising the tensile modulus and tensile strength of PLA. Also, the good UV barrier, anti-microbial properties and comparable water vapour permeability properties of PLA-HNT-chitosan composite intensify their potential application in packaging sector.
22
Acknowledgement The authors are thankful to Defence Research and Development Organization (DRDO), India for
providing
financial
assistance
for
this
research
work
through
Grant
No. DFTM/03/3203/M/01/JATC. The authors are also grateful to the Aayush Sharma and Tushar Bansal for their help with the sample preparation.
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Graphical abstract
29
S0141-8130(19)37183-1 https://doi.org/10.1016/j.ijbiomac.2019.10.246 BIOMAC 13747
To appear in:
International Journal of Biological Macromolecules
Received Date: Revised Date: Accepted Date:
5 September 2019 23 October 2019 26 October 2019
Please cite this article as: A. Anjali Singh, S. Sharma, M. Srivastava, A. Majumdar, Modulating the properties of polylactic acid for packaging applications using biobased plasticizers and naturally obtained fillers, International Journal of Biological Macromolecules (2019), doi: https://doi.org/10.1016/j.ijbiomac.2019.10.246
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© 2019 Published by Elsevier B.V.
Modulating the properties of polylactic acid for packaging applications using biobased plasticizers and naturally obtained fillers Anshu Anjali Singh*, Swati Sharma, Mayuri Srivastava, Abhijit Majumdar* Department of Textile Technology, Indian Institute of Technology Delhi, New Delhi, India, 110 016. *Corresponding author Email: [email protected] *Corresponding author Email: [email protected]
Abstract The properties of PLA films intended for packaging applications have been modulated by using bio-based platicizers and naturally obtained fillers. Triethyl citrate (TEC) and glycerol triacetate (GTA) have been used as platicizers and halloycite nanotubes (HNT) and chitosan have been used as fillers. The addition of 10 wt.% TEC, 10 wt.% GTA and 3 wt.% HNT improves the ductility of PLA films, however, reduces the tensile modulus and tensile strength. Addition of chitosan (1 wt.%), on the other hand, acts as a good reinforcing filler and improves the tensile strength and tensile modulus. PLA-HNT-chitosan film show comparable tensile strength and tensile modulus and ~12 times higher elongation at break compared to pure PLA. Besides, PLA-HNT-chitosan film demonstrates very good barrier properties against moisture and ultraviolet (UV) rays. Additionally, its antibacterial efficacy against E. coli and S. aureus are found to be around 80% and 70%, respectively. The study demonstrates the complementary effects of HNT and chitosan to modulate the properties of PLA film and indicates that the PLAHNT-chitosan film can emerge as a very potent material for packaging applications. Keywords: Chitosan; Halloysite nanotubes; Polylactic acid; Antibacterial; Water vaour transmission.
1
1. Introduction Global proliferation of commodities made of plastic materials results in rapid depletion of nonrenewable fossil fuels together with the global-warming caused by the production of carbon dioxide from the combustion. Most of the food packaging materials are manufactured from non-biodegradable and petrochemical based plastics which persist for a long time in the environment and their end-life cannot be anticipated easily [1]. In view of this, there is an impending need to develop materials from renewable resources and therefore, efforts need to be emphasised on the production of environment friendly sustainable packaging materials having the potential to replace petrochemical based plastics. Extensive research works have been performed in this context globally and scientists and researchers have proposed several biopolymers, which are bio-degradable, sustainable and acquire similar properties to that of conventional plastics. Among different biopolymers available, polylactic acid (PLA) is considered as one of the most promising for the production of sustainable and green packaging materials. PLA is a biodegradable, non-toxic, aliphatic thermoplastic polyester, obtained from renewable resources like corn, sugar beet and potato. It is considered as safe by the US Food and Drug Administration (FDA) for food packaging applications [2]. The strength and stiffness of PLA is similar to that of polyamide and polycarbonate. Moreover, PLA has good transparency, biocompatibility and biodegradability. However, the disadvantages of PLA like embrittlement, low thermal stability, poor gas and UV ray barrier properties, antimicrobial properties limit its application. Therefore, these properties need to be improved or tuned adequately in order to develop a suitable packaging material [3, 4]. Several approaches have been adopted to enhance the properties of PLA, for example, use of plasticizers, incorporation of micro or nano fillers etc. However, to keep the material as green as possible, it is important to use all the components which can be obtained naturally or can be degraded easily. 2
One of the most important considerations while developing a food packaging material is to choose non-toxic additives and fillers. Bio-based plasticizers like polyethylene glycol, citrate esters, triacetin, oligomeric lactic acid have been used to improve the ductility of PLA but at the cost of stiffness, tensile strength and glass transition temperature, as these properties generally deteriorate concurrently with the addition of plasticizers [5-7]. In addition, composites of PLA reinforced with naturally obatined additives/fillers are of great interest and the improvement in properties depends on the dispersion of the filler and good interfacial adhesion between the matrix and fillers. However, for the composites having matrix and fillers of different polarities like PLA and cellulose or chitosan, a third component needs to be used to improve the compatibilty between matrix and fillers and hence their properties. For example, Zakaria et al. [8] used epoxidized natural rubber as a compatibilizer for PLA-chitosan composite. Similarly, Tham et al. [3] studied the effect of 5-20 wt.% of epoxidized natural rubber on different properties of PLA-halloysite nanotubes. HNT is an aluminosilicate (Al2Si2O5(OH)4.2H2O) with hollow tubular structure, and is obtained naturally [9]. It has high stiffness and exhibit unique surface property due to the multilayered structure with few hydroxyl (-OH) groups [10]. HNT efficiently improves the strength, stiffness and thermal properties of the polymer matrix and can be considered as a potential nano-reinforcement [11, 12]. Although it is a promising material, however, limited work has been reported on its use in polymer composite. Like HNT, chitosan is also a natural biopolymer and is renewable, non-toxic and biodegrdable. It can be produced after the deacetylation of chitin, which is the second most abundant naturally occurred polysaccharide, after cellulose [13]. It is a linear copolymer composed of N-acetyl-D-glucosamine and D-glucosamine units linked by 1, 4-β-D-glucopyranosamine and is a polycationic polymer with one amino group (NH2) and two hydroxyl (-OH) groups in the repeating unit [14]. Chitosan has excellent antimicrobial behaviour due to the presence of positively charged amino groups, and hence is
3
considered good for food packaging applications [15, 16]. Rapa et al. [17] reported good antifungal and antimicrobial properties of biocomposites containing tributyl-o-acetyl citrate plasticized PLA and 1-5 wt.% of chitosan. Chitosan together with nanoclay was also used as a coating material to improve the poor gas barrier properties of PLA [18]. The main objective of this work was to modulate the properties of PLA films with biobased plasticizers and naturally obtained fillers, in order to develop the sustainable packaging material. Triethyl citrate (TEC) and glycerol triacetate (GTA) were used as plasticizers as both are biodegradable and non-toxic. Further to modulate the properties of plasticized PLA, halloysite nanotubes (HNT) and chitosan were used as fillers. PLA films with triethyl citrate, glycerol triacetate, halloysite nanotubes and chitosan, in different combinations, were prepared by solvent casting methods and consequent effect on mechanical, thermal, barrier and antibacterial properties was investigated.
2. Experimental 2.1 Materials Polylactic acid (PLA) pellets, Ingeo 4043D grade was purchased from NatureWorks Co. Ltd. Coimbatore, India. Two bio-based plasticizers, namely triethyl citrate (TEC) and glycerol triacetate (GTA) were purchased from Sigma-Aldrich, India. Table 1 shows the structure and properties and of TEC and GTA. Halloysite nanotubes (HNT) and chitosan powder were purchased from Natural Nano Inc. USA and SRL Pvt. Ltd. India, respectively. The transmission electron micrograph (TEM) was used to study the morphology of HNT (Fig. 1a) while the morphology of chitosan was studied using scanning electron micrograph (SEM) (Fig. 1b). HNT had length in the range of 200 nm to 1300 nm and diameter in the range of 50 nm to 200 nm. The size of chitosan powder varied from 2 µm to 20 µm. Methylene chloride or
4
dichloromethane (DCM) was purchased from Fisher Scientific, India. All the chemicals were used as received. Table 1 - Properties of plasticizers Name Triethyl citrate (TEC)
Molecular weight (g/mol) 276.285
Chemica l formula C12H20O7
218.205
C9H14O6
Glycerol triacetate (GTA)
Chemical structure
Figure 1. (a) transmission electron micrograph of HNT and (b) scanning electron micrograph of chitosan
2.2 Preparation of PLA films All the samples were prepared by solvent casting method. At first, PLA, HNT and chitosan were dried at 40 °C for 24 hours in order to remove the moisture. Requisite amount of PLA pellets-plasticizers-HNT-chitosan were added in the dichloromethane (DCM) and the suspension (10 wt.% of solid content) was than continuously stirred at room temperature for 24 hours. Thereafter, the mixed suspension was poured into a glass petri dish and DCM was allowed to evaporate overnight at room temperature in a fume hood followed by drying in an 5
oven at 40 °C for 24 hours. The dried film was then peeled off carefully for further use. In this study, nine different films were prepared and the details about their formulation are reported in Table 2. The amount of plasticizer (TEC or GTA), HNT and chitosan used was 10 wt.%, 3 wt.% and 1wt.%, respectively. The thickness of all the films was approximately 0.3 mm.
Table 2 - Sample coding and formulation of the films Sample coding PLA PLA-TEC PLA-GTA PLA-HNT PLA-TEC-HNT PLA-GTA-HNT PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
PLA (wt.%) 100 90 90 97 87 87 96 86 86
TEC (wt.%) 10 10 10 10 -
GTA (wt.%) -
HNT (wt.%) -
Chitosan (wt.%) -
10 10
3 3 3 3 3 3
1 1 1
3. Testing and Characterizations 3.1 Morphology The surface morphology of HNT and chitosan was characterized by transmission electron microscopy (TEM) and scanning electron microscopy (SEM) using JEM-1400 (JEOL) and LEO 982 (Zeiss), respectively. The samples for TEM was prepared by applying a drop of diluted suspension of HNT on carbon coated grid followed by drying in an oven at 50 °C for overnight. Chitosan powder was coated with gold before the SEM analysis.
3.2 Mechanical properties Tensile test of rectangular shaped samples having a width of 10 mm was performed using universal tensile testing machine (Instron 3365) equipped with a 500 N load cell. Five specimens for each sample were tested with a gauge length of 20 mm and cross-head speed of
6
5 mm/min. The tensile modulus, ultimate tensile strength and elongation at break were evaluated.
3.3 Thermal characterization Thermal behaviour of all the samples was examined using differential scanning calorimetry (DSC Q 2000, TA) under nitrogen atmosphere. Approximately 2.5 mg of sample was sealed in an aluminium crucible and heated gradually from 0 °C to 200 °C at a heating rate of 10 °C/min for first heating scan. The samples were then held at 200 °C for 5 minutes to remove the thermal history. Further, they were cooled to 0 °C at a cooling rate of 10 °C/min for the cooling scan before a second heating scan from 0 °C to 200 °C at a heating rate of 10 °C/min. Melting temperature (Tm), cold crystallization temperature (Tcc) and glass transition temperature (Tg) and were determined from the 2nd heating scans.
3.4 X-ray diffraction Crystallinity percentage of the samples was analysed by the Ultima IV X-ray diffractometer (Rigaku) in reflection mode. X-ray diffractograms (XRD) of the film samples were recorded at room temperature with Cu α radiation. The scan was performed at a speed of 3°(2θ)/min and the data were collected at 0.02°/interval over the 2 θ range from 10 to 40°. Percentage crystallinity (߯ %) of the sample was calculated using Eq. 1. [19]. ߯ ሺΨሻ ൌ
௨ௗ௧ ்௧
ൈ ͳͲͲ
(1)
3.5 UV-Vis spectroscopy Light transmittance of the films was measured using a Perkin Elmer UV/VIS Spectrometer Lambda 1050. Rectangular shaped film specimen was placed directly in a test cell and the scan
7
was carried out from 300 to 800 nm wavelength with a scan speed of 240 nm/min using air as the reference.
3.6 Water vapour permeability Water vapour transmission rate (WVTR) of the films was measured using an automatic water vapour permeability testing machine (Labthink W3/060). Each sample having an area of 33 cm2, was tested at 38 °C and relative humidity of 80 %. Three specimens for each sample were tested and the average value was taken for analysis.
3.7 Anti-bacterial activity The antibacterial properties of pure PLA film and films with chitosan were studied against Gram-negative Escherichia coli (E. coli) and Gram-positive Staphylococcus aureus (S. aureus). Fresh bacteria of E. coli and S. aureus were first activated by incubating them at 37 °C for 24 hours in a fresh nutrient agar medium. Freshly grown colonies of both the species of bacteria were then placed in the nutrient broth and incubated again at 37 °C for 24 hours. After 24 hours, the bacterial suspension was diluted to a concentration of approximately 105 CFU/ml. The properly sterilized samples (20 mm × 10 mm) were then exposed to the bacterial suspension at 37 °C for 24 hours. Finally, the incubated bacterial suspension with samples were cultured on the petri dishes with agar medium. These petri dishes were incubated at 37 °C for 24 hours and the colony forming units (CFU) were counted. The antibacterial efficiency (AE %) of the samples was calculated using Eq. 2, where B and A are the total number of bacterial colonies in pure PLA (reference) and PLA films with chitosan, respectively [20]. ݕ݂݂݈ܿ݊݁݅ܿ݅݁ܽ݅ݎ݁ݐܾܿܽ݅ݐ݊ܣሺܧܣΨሻ ൌ
8
ି
ൈ ͳͲͲ
(2)
4. Results and Discussion 4.1 Mechanical Properties The effects of addition of plasticizers (TEC and GTA) and fillers (HNT and Chitosan) on the tensile properties are shown in the Fig. 2(a-c) and Table 3. It can be observed from the Fig. 2 and Table 3, that the presence of plasticizers and fillers significantly influence the mechanical properties of PLA. As expected, pure PLA showed brittle failure with an elongation at break of around 6 %, whereas this value is found to be much higher for PLA-TEC (390 %) and PLAGTA (418 %). However, tensile modulus (0.96 GPa and 0.75 GPa, respectively) and ultimate tensile strength (30 MPa and 27 MPa, respectively) decreases, indicating that TEC and GTA efficiently plasticizes the PLA, making it more flexible and ductile [21, 22]. However, subtle difference in the tensile properties can be noticed between PLA-TEC and PLA-GTA with latter showing lower strength and modulus but higher elongation at break. In addition, tensile strength and tensile modulus of PLA-HNT reduce to 40 MPa and 1.6 GPa, respectively, then those of pure PLA i.e., 47 MPa and 2.4 GPa, respectively, while the elongation at break increases to 91 %. Improvement in elongation at break or ductility of PLA after the addition of nano-fillers has been widely reported. Li et al. [23] reported that the stress-whitening induced by the extensive crazing is responsible for the improvement in strain at break of PLA nanocomposite after the addition of 0.5 to 5 wt.% of organically modified rectorite. Jiang et al. [24] reported that the PLA with organically modified montmorillonite and nano-sized precipitated calcium carbonate showed significant improvement in the strain at break compared to pure PLA. Liu et al. [11] studied the effect of HNT content on PLA and reported that the elongation at break of PLA-HNT nanocomposites with less than 20 phr (parts per hundred of PLA) HNT has higher value than that of pure PLA, as the small quantity of HNT could acts as a plasticizer.
9
Furthermore, PLA-TEC-HNT and PLA-GTA-HNT show slight improvement in the elongation at break compared to those of PLA-TEC and PLA-GTA, respectively, while insignificant change in ultimate tensile strength and tensile modulus are observed. This might be due to the synergistic effect of plasticization of TEC or GTA and small quantity of HNT in PLA [11]. Rashmi et al. [25] also reported the improved ductility of the PLA/Polyamide11 (PLA/PA11) blend after the addition of 2 to 6 wt.% of HNT compared to PLA/PA11 blend. All the specimens of PLA with HNT show prominent stress-whitening and this phenomenon causes ductile failure during tensile test compared to the brittle failure in case of pure PLA [22]. It is known that the addition of filler usually improves the tensile strength and tensile modulus of the polymer composites [26]. However, in this study the addition of HNT in pure PLA and plasticized PLA (i.e. PLA-TEC and PLA-GTA) increases the elongation at break, which might be due to the immense crazing caused by the stress-whitening as well as plasticization together with slippery effect of HNT in PLA [11, 22-24, 27]. Stress whitening led to the enhanced opacity in the tensile fractured specimens and is attributed to the variations in the refractive index caused either by the formation of micro voids or change in the structure [22, 27].
Table 3 - Tensile properties of PLA and its composites Sample PLA PLA-TEC PLA-GTA PLA-HNT PLA-TEC-HNT PLA-GTA-HNT PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
Tensile modulus (GPa) 2.40 (± 0.10) 0.96 (± 0.05) 0.75 (± 0.05) 1.60 (± 0.03) 0.62 (± 0.03) 0.74 (± 0.03) 2.10 (± 0.30) 1.10 (± 0.20) 0.97 (± 0.20) 10
Ultimate tensile strength (MPa) 47 (± 2.6) 30 (± 0.7) 27 (± 0.5) 40 (± 0.3) 27 (± 0.3) 26 (± 1.0) 48 (± 2.5) 34 (± 3.5) 35 (± 3.2)
Elongation at break (%) 6 (± 0.4) 390 (± 34) 418 (± 17) 91 (± 7) 468 (± 46) 490 (± 78) 58 (± 25) 108 (± 22) 132 (± 27)
Figure 2 - Tensile stress-strain curve of the PLA and its composites
Further, after the addition of 1 wt.% chitosan as filler, tensile properties of PLA-HNTchitosan, PLA-TEC-HNT-chitosan and PLA-GTA-HNT-chitosan were also explored. It is observed (Fig. 2 and Table 3) that the addition of chitosan causes significant improvement in tensile modulus and tensile strength compared to the corresponding samples without chitosan. This indicates that the chitosan acts as a good reinforcing material and establishes good interfacial interactions with the components of the composite [26]. However, elongation at break decreases which might be attributed to the stiffening action of the chitosan that causes substantial local stress concentration and consequently failure at lowered strain [28]. Although, the elongation of PLA-TEC-HNT-chitosan (108 %) and PLA-GTA-HNT-chitosan (132 %) are found to be higher than that of PLA-HNT-chitosan (58 %) due to the predominating plasticizing effects of TEC and GTA, respectively. Of all the composites, PLA-HNT-chitosan exhibited comparable tensile strength and modulus as of pure PLA with higher elongation at
11
break, indicating that PLA, HNT and chitosan has good interfacial interaction altogether. Two different kind of hydrogen bonds are formed in the PLA-HNT-chitosan composite; one between the carbonyl groups of PLA and hydroxyl group of HNT and chitosan while other between the carbonyl groups of PLA and amino group of chitosan. The proposed hydrogen bond formation between the PLA, HNT and chitosan are shown in the Fig. 3. Similar findings were also reported by Zakaria et al. [8] and Vasile et al. [29]. Zakaria et al. [8] reported that the composite of PLA, chitosan and epoxidised natural rubber (ENR) had higher tensile modulus and tensile strength than PLA/ENR and also the tensile modulus of the former was higher than that of pure PLA. Vasile et al. [29] reported that PLA, polyethylene glycol (PEG) and chitosan composite had higher modulus than that of the PLA/PEG. Overall, the present study demonstrates the complementary effects of HNT and chitosan on the tensile properties of PLA, by efficiently improving the ductility of the composite without compromising the tensile modulus and tensile strength.
Figure 3 –Proposed hydrogen bond interaction between PLA, HNT and chitosan 12
4.2 Thermal Properties DSC analysis of all the samples was carried out to study the effect of plasticizers and fillers on the thermal behaviour of PLA. The DSC thermograms of all the samples, obtained during the second heating scan are shown in Fig. 4 and the values extracted from the thermograms are reported in Table 4. The second heating scan of DSC thermograms indicate the presence of glass transition (Tg) endotherm, cold crystallization (Tcc) exotherm and melting (Tm) endotherm, which are typical of any semi-crystalline polymer. The Tg, Tcc and Tm of pure PLA film are 61.5 °C, 125 °C, and 149.7 °C, respectively. Values of Tg, Tcc and Tm decreases for PLA-TEC and PLA-GTA due to the enhanced flexibility of polymer chains in presence of TEC and GTA, respectively [21, 30]. The shift in the Tcc to lower temperature is attributed to the increased crystallinity, which is further confirmed by the XRD analysis [31]. In addition, the Tg and Tm recorded for PLA-TEC (44.5 °C and 143.1 °C, respectively) are found to be slightly higher than that of PLA-GTA (42.2 °C and 141.2 °C, respectively). This change in the thermal behaviour can be attributed to the lower molecular weight of GTA (218.205 g/mol) than that of TEC (276.85 g/mol) [32]. Similar observation has been reported by the Ljungberg et al. [33] for PLA plasticized with tributyl citrate and Martin et al. [34] for PLA plasticized with different molecular weight of polyethylene glycol. Miniscule change in the Tg and Tm are observed for PLA-HNT composite with respect to pure PLA, however, the intensity of Tcc decreased compared to pure PLA. Addition of Chitosan does not alter the Tg and Tcc when compared to the respective samples without chitosan but Tm seems to decrease in PLA-HNT-chitosan only (from 149.3 °C to 142.2 °C). The decreased Tm could be attributed to the thinner lamellar thickness of PLA [35, 36].
13
Table 4 - Thermal properties and crystallinity percentage of PLA and its composites Sample
PLA PLA-TEC PLA-GTA PLA-HNT PLA-TEC-HNT PLA-GTA-HNT PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
Tg (°C) 61.5 44.5 42.2 61.4 42.5 41.4 60.7 41.2 -
DSC Tcc (°C) 125 112 111 105 110 105 112
Tm (°C) 149.7 143.1 141.2 149.3 142.1 142.7 142.2 142.2 143.7
XRD Crystallinity (%) Amorphous 43.6 41.7 34.7 39.4 38.2 24.4 28.8 22.7
Figure 4 – DSC thermograms of PLA and its composites 4.3 X-Ray diffraction XRD was performed to investigate the effect of plasticizers and fillers on the crystallinity of PLA. X-ray diffractograms of all the samples are presented in Fig. 5 and the calculated
14
crystallinity percentage are reported in Table 4. Only one broad XRD spectrum, without any sharp peak, is found to be present in pure PLA (Fig. 5a) at around 2θ = 16.7°, which is characteristic of the (200/110) crystallographic plane, signifying pure PLA to be amorphous [11, 30]. This indicates that the PLA does not get enough time to crystallize during the sample preparation by solvent casting method as the solvent evaporates quickly. The XRD profile of PLA-TEC, PLA-GTA and PLA-HNT exhibit profound diffraction peaks as the intensity is found to be sharper and stronger than that of pure PLA implying higher crystallinity. Sharp diffraction peaks at 2θ = 12.09°, 14.7°, 16.7°, and 22.3° corresponding to (004/103), (010), (200/110) and (105) crystallographic planes, respectively, are present in all the samples except for pure PLA [30]. The diffraction peaks at 2θ = 14.7° and 22.3° reflect the presence of α- form of PLA, while the peak at 2θ = 12.09° could be assigned to the D-form of PLA [30, 37]. Further, it can be noticed that the peak at 2θ = 19.90°, corresponding to (020/110) crystallographic plane, is present only in PLA-HNT and PLA-HNT-chitosan. The shift in the peak was observed at 2θ = 19.0° corresponding to (203) crystallographic plane, in the presence of TEC and GTA, indicating that the addition of plasticizer altered the stable α crystal of PLA [37].
Figure 5 –X ray diffractogram of PLA and its composites It can be seen from the Table 4 that the addition of TEC, GTA and HNT increase the crystallinity percentage of PLA. The degree of crystallinity of PLA-TEC (43.6 %) and PLA-
15
GTA (41.7 %) is found to be much higher than that of pure PLA indicating that the plasticizers enhance the segmental molecular chain mobility of PLA and thus facilitates the crystallisation in plasticized PLA [21, 30, 34]. Besides, HNT acts as an effective nucleating agent for PLA as the addition of 3 wt.% of HNT increased the crystallinity of PLA to 34.7 % [11]. Slight decrease in percentage crystallinity is observed for PLA-TEC-HNT (39.4 %) and PLA-GTA-HNT (38.2 %) when compared to PLA-TEC and PLA-GTA, respectively. However, the addition of chitosan decreases the crystallinity of PLA films when compared to the respective samples without chitosan, although the values are still higher than that of pure PLA. Correlo et al. [35] and Bonilla et al. [38] also reported the similar trends. Correlo et al. [35] opined that the decreased crystallinity after the addition of chitosan can be attributed to the hydrogen bond formation between the carbonyl groups of polyester and -OH and -NH2 groups of chitosan in the amorphous phase. Senda et al. [36] reported that difference in the molecular mobility between matrix and chitosan could be another possible reason for lower degree of crystallinity, besides hydrogen bonding interaction.
4.4 Optical transparency Transmission of ultraviolet rays (UV) and visible light through the polymeric film is one of the important parameters in packaging application, in order to store and protect the products which are light sensitive [4]. The transmittance percent at different wavelengths of all the films are shown in the Fig. 6(a) and their visual appearances are shown in the Fig 6. (b-j). It is observed from the Fig. 6 that the pure PLA has the highest transmission in the visible region (400-800 nm) as well as in the UV region (< 400 nm), indicating its good transparency in the visible region but poor UV barrier property. The transparency can also be ratified from the visual appearance of PLA film shown in Fig. 6(b). The percent light transmittance (T %) of the PLA films with TEC, GTA and HNT are found to be lower than that of pure PLA. The recorded
16
transmissions for the PLA, PLA-TEC, PLA-GTA and PLA-HNT, in the middle of the visible spectrum (i.e., 600 nm), are 92, 88, 87 and 84 %, respectively. Further, the incorporation of HNT in PLA-TEC and PLA-GTA shows slightly lesser transmission at 600 nm as the transmission recorded are 83 % and 80 % for PLA-TEC-HNT and PLA-GTA-HNT, respectively. It can be seen from the Fig 6 (b-j), that the visual appearance of PLA films with TEC, GTA and HNT are less transparent than that of pure PLA. The decreased transmittance of PLA films in the presence of plasticizers and HNT than that of pure PLA is due to the increased crystallinity and homogeneous dispersion of HNT in the composite [22, 39]. In addition, the result demonstrates (Fig. 6 and Table 5) that the percent transmittance of all the films decrease in the UV region and the films containing HNT shows better UV barrier property than the films of PLA, PLA-TEC and PLA-GTA. A reduction in approximately 32 %, 33 % and 41 % transmittance are observed in PLA-HNT, PLA-TEC-HNT and PLA-GTA-HNT than that of pure PLA at wavelength of 320 nm. This may be attributed to the higher reflection and absorption of UV rays by the HNT. However, interestingly, presence of chitosan shows miniscule change in the percent light transmittance in visible region as well as in UV region. This might be due to the bigger particle size of chitosan than HNT [22, 39]. The results imply that the nanocomposite PLA films possess improved barrier property particularly in the UV region, while maintaining the good transparency in the visible light spectrum. Table 5 –Transmittance % and WVTR of PLA and its composites Sample
PLA PLA-TEC PLA-GTA PLA-HNT PLA-TEC-HNT PLA-GTA-HNT PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
Transmittance % (λ = 600 nm) 92 88 87 84 83 80 84 83 81 17
Transmittance % (λ = 320 nm) 81 74 72 55 54 48 57 48 49
WVTR (g/m2.day) 39 (±2.5) 63 (±2.5) 57 (±1.5) 47 (± 4.5) 55 (±4.0) 50 (±5.6) 40 (± 4.5) 60 (± 5.2) 58 (±6.5)
Figure 6 - (a) Transmittance percent vs wavelength of PLA and its composites and visual appearance of film of (b) PLA, (c) PLA-TEC, (d) PLA-GTA, (e) PLA-HNT (f) PLA-TECHNT, (g) PLA-GTA-HNT, (h) PLA-HNT-chitosan, (i) PLA-TEC-HNT-chitosan and (j) PLA-GTA-HNT-chitosan
4.5 Water vapour transmission Water vapour transmission rate (WVTR) is another important barrier property of food packaging material as it determines the spoilage of moisture sensitive food. WVTR is defined as the amount of water vapour transmitted, under specific temperature and humidity, through unit area of film per unit time. Water vapour barrier property of a polymeric film or composite is mostly influenced by the hydrophobicity or hydrophilicity of matrix and reinforcement, amount and dispersion of the later and the presence of voids or cracks [40]. Table 5 shows the WVTR of all the samples. The WVTR is 39 g/m2/day for pure PLA and the addition of plasticizers and fillers substantially influence the transmission rate of water vapour. The WVTR increases by approximately 62 %, 46 % and 20 % for PLA-TEC, PLA-GTA and PLAHNT, respectively, compared to pure PLA. This may be due to the hydrophilic behaviour of the TEC and GTA as both the plasticizers are soluble in water and free hydroxyl (-OH) groups present on the surface of HNT. Although, the WVTR of PLA-HNT is found to be superior than the plasticized PLA and this could be due to the use of less amount of HNT (3 wt.%) than
18
plasticizers (10 wt.%). The findings are in good agreement with the results reported by the other researchers. Fan et al. [41] and Gao et al. [42] reported that the water vapour permeability of PLA films increased after the addition of acetyl-tributyl citrate (ATBC) and tributyl citrate (TBC) respectively. Dadashi et al. [40] reported that the PLA films with unmodified natural nanoclay showed slightly higher WVTR while the same decreased for the films with organically modified nanoclays. They attributed it to the hydrophilic nature of the former and hydrophobicity of the later. The addition of chitosan improves the WVTR of the films compared to the respective samples without chitosan. The lowest WVTR recorded in this study was 40 g/m2/day for PLA-HNT-chitosan, which is comparable to the hydrophobic pure PLA. The improvement in the water vapour barrier property of the PLA films after the addition of chitosan might be associated with its hydrophobicity due to the presence of acetyl groups of incompletely deacetylated chitosan [43]. The hydrogen bond interaction between PLA, HNT and chitosan reduces the availability of free hydroxyl group for interaction with water molecules and thus decreasing the water vapour transmission rate. The result reveals that the WVTR of the PLA film was influenced by the hydrophilicity/hydrophobicity of the material used, and the use of plasticizers like TEC or GTA is not desirable as they increase the WVTR. However, HNT and chitosan showed complementary effect in improving the water vapour barrier properties.
4.6 Antibacterial activity Two different species of bacteria i.e., E. coli (Gram-negative) and S. aureus (Gram-positive) were used to study the antibacterial property of PLA and its composites. Both the bacteria are considered as a pathogenic infectious and their symptoms range from mild to serious [44]. The images representing the bacterial colony growth are shown in Fig. 7 while the total bacterial colony count and antibacterial efficiency are presented in Table 6. The total number of visible
19
bacterial colonies are more for pure PLA than that of PLA with chitosan and thus the antibacterial efficiency of the later is significantly higher. This is due to presence of positively charged amino (-NH2) group in chitosan that binds easily with the negatively charged peptidoglycan, liposaccharides and proteins present on the cell wall of the bacteria, thus obstructing their biosynthesis and prevent the transportation of nutrient through the cell wall and eventually supress their growth [44]. In addition, it can also be observed that the PLA and PLA-chitosan composites are more resistant to E. coli than S. aureus, as the growth of bacterial colonies are more in case of the latter. Similar findings were also reported by other researchers [4, 45]. Hu et al. [46] also found that the antibacterial activity of chitosan is higher against the E. coli than that against S. aureus and this is due to the difference in the cell wall structure of these bacteria (Fig. 8). The cell wall of S. aureus consists of single thick layer of peptidoglycan while the cell wall of E. coli consists of thin peptidoglycan layer surrounded by an outer membrane.
Outer
membrane
present
in
the
Gram-negative
bacteria
contains
lipopolysaccharides and proteins which increases the negative charge on the surface and consequently confer resistance to hydrophobic compounds [47].
Figure 7 - Antibacterial activity of (a) PLA, (b) PLA-HNT-chitosan, (c) PLA-TEC-HNTchitosan and (d) PLA-GTA-HNT-chitosan, against E. coli and S. aureus
20
Table 6 – Antibacterial efficiency against E. coli and S. aureus Sample
Total bacterial colony
PLA PLA-HNT-chitosan PLA-TEC-HNT-chitosan PLA-GTA-HNT-chitosan
E. coli 32 6 7 7
S. aureus 369 108 102 102
Antibacterial efficiency % E. coli S. aureus 81 71 78 72 78 72
Figure 8. Cell wall structure of Gram-positive and Gram-negative bacteria
5. Conclusions The influence of addition of two bio-based plasticizers, namely triethyl citrate (TEC) and glycerol triacetate (GTA) and two fillers, namely halloysite nanotubes (HNT) and chitosan powder, in different combinations, on mechanical, thermal, barrier and anti-microbial properties of polylactic acid (PLA) films was studied. Tensile results revealed that the TEC and GTA efficiently plasticizes the PLA as the elongation at break increased significantly (~80 times) though tensile modulus and tensile strength reduced significantly. Addition of HNT also improved the elongation at break (~18 times) along with decrease in tensile modulus and tensile strength, though the extents of reduction were much lower as compared to those obtained in case of TEC and GTA. However, marginal improvement in ductility was noticed in PLA-TECHNT and PLA-GTA-HNT compared to those of PLA-TEC and PLA-GTA, respectively. Addition of chitosan in PLA-HNT, PLA-TEC-HNT and PLA-GTA-HNT improved the tensile modulus and tensile strength while reducing the elongation at break. PLA-HNT-chitosan showed comparable tensile strength and tensile modulus and ~12 times elongation at break as 21
compared to those of pure PLA, indicating good interfacial interactions between these components. Thermal study demonstrated that the Tg, Tcc and Tm decreased for PLA-TEC and PLAGTA, as compared to those of pure PLA, as a result of plasticization effect. XRD studies showed that, degree of crystallinity of the PLA films increased significantly with the addition of TEC, GTA and HNT, however, the incorporation of chitosan lowered the degree of crystallinity. UV-vis spectroscopy confirmed that the PLA has good transparency in the visible region but has poor UV barrier property. The overall reduction in the percentage transmittance was observed for PLA-TEC, PLA-GTA, PLA-HNT, PLA-TEC-HNT and PLA-GTA-HNT, when compared to pure PLA, indicating the improved UV barrier property of the PLA composite films. Negligible change in the transparency was observed in the presence of chitosan. Water vapour permeability (WVP) of PLA with HNT, TEC and GTA increased, however, addition of chitosan lowered the WVP. The lowest water vapour transmission rate was obtained in case of PLA-HNT-chitosan film, which was comparable to the hydrophobic pure PLA. Nanocomposite films with chitosan showed good antibacterial properties against E. coli and S. aureus and the former being more resistant due to the difference in the cell wall structure. Overall, the results revealed that the properties of PLA could be easily modulated by using TEC, GTA, HNT and Chitosan in combinations. The results also demonstrated the complementary effect of HNT and chitosan as PLA-HNT-chitosan showed improved ductility without compromising the tensile modulus and tensile strength of PLA. Also, the good UV barrier, anti-microbial properties and comparable water vapour permeability properties of PLA-HNT-chitosan composite intensify their potential application in packaging sector.
22
Acknowledgement The authors are thankful to Defence Research and Development Organization (DRDO), India for
providing
financial
assistance
for
this
research
work
through
Grant
No. DFTM/03/3203/M/01/JATC. The authors are also grateful to the Aayush Sharma and Tushar Bansal for their help with the sample preparation.
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Graphical abstract
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