Improving the extensibility of paper: Sequential spray addition of gelatine and agar

Transcription

1 Improving the extensibility of paper: Sequential spray addition of gelatine and agar Alexey Vishtal, Alexey Khakalo, Orlando J. Rojas and Elias Retulainen KEYWORDS: Agar, Gelatine, Extensibility, Formability, Spraying, Papermaking SUMMARY: High extensibility of paper is of key importance for production of novel 3D-packaging materials. The application of agar onto a wet web has been shown to significantly improve the extensibility of dry paper as a result of shrinkage during drying while addition of gelatine strengthens inter-fibre bonding. In this work, these two bio-based materials were applied sequentially to yield paper with higher extensibility compared to that obtained by single component application. We studied the interactions between agar, gelatine and cellulose by using quartz crystal microgravimetry and atomic force microscopy. Agar adsorption was significantly improved after priming the cellulose surface with gelatine. This synergistic effect on extensibility only occurred if the protein was added first. It is hypothesized that the gelatine strengthens the interfibre bonds while the polysaccharide forms a film on the web surface, and reinforces it. The extensibility of webs treated with gelatine (4%) and agar (4%) was ca. 15% after unrestrained drying. Such remarkable level of extensibility allows production of tray-like shapes via conventional thermoforming machine to depths of up to 2 cm. Overall, a protocol based on the sequential application of two abundant biopolymers is proposed to enhance formability of paper. ADDRESSES OF THE AUTHORS: Alexey Vishtal (alexey.vishtal@vtt.fi), VTT Technical Research Centre of Finland/Tampere University of Technology, Laboratory of Paper Converting and Packaging Technology, Koivurannantie 1, P.O. Box 1603, 40101, Jyväskylä, Finland. Elias Retulainen (elias.retulainen@vtt.fi), VTT Technical Research Centre of Finland, Alexey Khakalo (alexey.khakalo@aalto.fi), Aalto University School of Chemical Technology, Department of Forest Products Technology, P.O. Box 16300, Aalto, Finland, Orlando J. Rojas (orlando.rojas@aalto.fi), Aalto University School of Chemical Technology, Department of Forest Products Technology, P.O. Box 16300, Aalto, Finland Corresponding author: Alexey Vishtal The traditional market for printing grades of paper is declining; however, the use of paper in packaging applications is increasing steadily. Highly-extensible paper and paperboard for 3D-forming have recently gained increased attention both from academia and packaging industry (Hauptmann, Majschak 2011, Svensson et al. 2013). So-called formable paper can be used for production of 3D-shapes, for instance in the form-fill-seal machines. The primary requirements for such paper include high extensibility, sufficient strength and minimal elastic recovery (Vishtal et al. 2014). The extensibility of paper can be improved by different treatments and additives and such effect comes via modification of the fibre structure or its bonding (Vishtal, Retulainen 2014b). Recently, spray application of agar (Vishtal, Retulainen 2014a) and gelatine (Khakalo et al. 2014) on wet webs was found to improve extensibility and concomitant formability of paper. The speculated mechanisms of action of these two polymers with respect to extensibility improvement can be combined for a more positive effect if compared to that of the single component application. In fact, it has been shown that the mechanical properties of the gelatine-based films can be improved by addition of relatively small amount of carbohydrate polymers, such as gellan gum and k- carrageenan (Pranoto et al. 2007). Agar resembles carrageenan with respect to the structure and gel-forming properties and thus might also improve gelatine performance in paper. Both agar and gelatine are gel-forming substances; however, besides the obvious differences in chemical structures, they differ in the gel-forming mechanism, gel strength and gelling temperature (Banerjee, Bhattacharya, 2012). At present, these polymers have very limited use in papermaking, for instance in finishing of photographic paper. However, gelatine was a commodity papermaking additive in the past. It was used as an animal glue for sizing of writing grades starting from ca. 16 th century till the middle of 19 th century; the gelatine content was as high as 10-15% of dry weight of paper (Barrett et al. 2014). The fact that gelatine easily penetrates into the paper (Khakalo et al. 2014) while agar forms a film on the surface (Vishtal, Retulainen, 2014a) highlights the difference in their action and opens the possibility for an additive or synergistic effect. Moreover, cross-linking of gelatine and agar enhances tensile strength of films and reduces their extensibility due to the depression of molecular mobility. However, if the crosslinker is applied when the gelatine or agar are added to paper, adhesion between the film and cellulose is improved, which results in increased overall extensibility (Khakalo et al. 2014). The aim of this study is thus to verify whether gelatine and agar can be used together and whether they have an additive action for improved paper extensibility and formability. This was carried out by evaluation of the mechanical properties of paper and by investigating adsorption phenomena via quartz crystal microbalance with dissipation (QCM-D). In addition, the influence of crosslinker addition to polymer solutions on the properties of the treated paper was investigated. Finally, formability of the paper samples was evaluated using a 2D-formability tester. 452

2 Materials and Methods Pulp The fibre raw material used in this study was once-dried first-thinning bleached softwood kraft pulp produced by the Pietarsaari mill of UPM-Kymmene. Trimethylsilyl cellulose (TMSC) was synthetized as described elsewhere (Kontturi et al., 2003) and used in experiments with QCM-D. Agar Agar used in this study was typical food grade agar (also referred as E406) obtained from the Gourmetologia Oy, Finland. Gelatine Gelatine from porcine skin (Type A, ~300 g Bloom gel strength, # ) was obtained from Sigma-Aldrich (US). Crosslinker Ammonium zirconium (IV) carbonate (AmZrCarb) was used as a water solution with 1-2% of tartaric acid stabilizer (CAS Reg. No ) and purchased from Sigma-Aldrich. Mechanical treatment of fibres Pulp fibres were mechanically processed using a combination of high-consistency wing defibrator treatment and a subsequent low-consistency refining in a Valley beater. The detailed procedure and effects of the treatment on the fibre properties can be found elsewhere (Zeng et al. 2013, Vishtal, Retulainen 2014a). High consistency treatment creates micro-compressions and dislocations in fibres, while low-consistency refining straightens the fibres and improves bonding (Zeng et al. 2013). After refining, the fibre suspension was washed into Na-form in order to remove adsorbed metal ions and water-soluble substances, according to Laine et al. (2000). Handsheet preparation Handsheets were prepared according to SCAN-C26 standard with a target grammage of 60 g/m 2. Unrestrained handsheets were dried between two synthetic wires with a ca. 1-3 mm gap to avoid extensive cockling of paper. Due to drying shrinkage and addition of agar and gelatine the final grammage of the handsheets was higher, in the range of g/m 2. High-grammage (250 g/m 2 ), A4-sized sheets were prepared using sheet former Juupeli developed by VTT. These handsheets were used for the preparation of the 3D shapes in Coveris Rigid Packaging in Hämeenlinna, Finland. Polymer solution preparation and spraying Polymer solutions were prepared by adding either agar or gelatine powder into deionized hot water (ca C for agar and C for gelatine) under vigorous mechanical stirring. The stirring and heating of the resulting mixture was extended until polymer dissolution was complete. The concentration of polymer solutions was 2%. Crosslinker (AmZrCarb) was added into the agar and gelatine solutions about 30 s before spraying. This short mixing time before spraying was required in order to avoid extensive evaporation of ammonia from AmZrCarb and to prevent agar from self-crosslinking. Crosslinker was added at 3% relative to the mass of polymer in solution. Polymer solutions were sprayed onto the wet webs using commercial household electrospray gun followed by wet pressing. The amount of added polymers was controlled gravimetrically. The dryness of paper after wet pressing was the same for non-treated handsheets and handsheets containing gelatine added by spraying. Samples treated with agar had 1-2% lower dryness due to high water retention of the agar gel layer. In case of combined addition, gelatine was sprayed first, followed by agar application after 2 min. Gelatine and agar were added only onto one side of the web (top side) since addition to both sides did not bring any apparent benefit if compared with single-side addition. Drying shrinkage measurement The drying shrinkage was evaluated by measuring the perimeter between four dots on the paper before and after drying: 100% [1] where P w and P d are the perimeter of the square in the wet and dry handsheet, respectively. Scanning electronic microscopy (SEM) A FEI Quanta 200 scanning electron microscope (SEM) was used for surface and cross section imaging of the paper samples. The SEM was operated in back scattered electron mode at an emission current of approx. 100μA, and using an accelerating voltage of 12.5 kv or 15 kv, at distance 10 mm and a spot size of 5 to 6 was used. The samples were spun-coated using BALZERS SCD 050. The cross sections were prepared by a Hitachi IM4000 cross section cutter prior to the SEM investigation. Quartz Crystal Microbalance with Dissipation (QCM-D) Gelatine and agar adsorption onto cellulose and the properties of adsorbed layers were investigated by using a QCM-D instrument (model E4, QSense AB, Sweden). The QCM instrument simultaneously measures changes in the resonance frequency (Δf) and energy dissipation (ΔD) of an oscillating piezoelectric crystal upon increase/decrease of the mass on the crystal surface. Dissipation refers to the frictional losses that lead to damping of the oscillation frequency depending on the viscoelastic properties of the material. For a rigid adsorbed layer that is uniformly distributed on the surface and small compared to the mass of the crystal, the Sauerbrey relation can be applied to calculate the mass change upon adsorption ( m): [2] where, C is the Sauerbrey constant (17.7 ng/cm 2 for 5 MHz crystal), Δf is change in frequency and n is the overtone number (n=1, 3, 5, etc.). Note that this calculation is limited to rigidly adsorbed, thin layers. If the adsorbed layer is soft (i.e., viscoelastic) or highly hydrated, the Sauerbrey relation is not valid due to the high energy dissipation during the crystal oscillation. In such cases the Voigt viscoelastic model can be used to 453

3 more accurately calculate the extent of adsorption. The damping (or dissipation; D) is defined as: [3] where, E diss and E stor are the energy dissipated and total energy stored in the adsorbed film during one oscillation cycle, respectively. Preparation of model cellulose surfaces Cellulose-gelatine-agar interactions were investigated by using gold-coated sensors in QCM-D experiments. The sensors were first cleaned with UV/ozone treatment for 15 min followed by spin coating of 0.1% polystyrene in toluene (4000 rpm, 60 s). The prepared polystyrenecoated sensors were dried in an oven at 60 C for 10 min to ensure a uniform hydrophobic layer formation suitable for trimethylsilyl cellulose (TMSC) deposition. TMSC was deposited on the polystyrene-coated sensors by using the Langmuir Schaeffer (LS) horizontal lifting deposition technique as described by Tammelin et al., (2006). The TMSC film (30 layers) was then converted to cellulose via desilylation with hydrochloric acid vapour as described elsewhere (Schaub et al., 1993). The crystallinity degree, thickness, and roughness of the LScellulose films prepared in the same manner have previously been observed to be 54 %, 17.8 nm, and 0.5 nm, respectively (Aulin et al., 2009). Before QCM-D experiments the cellulose films were allowed to swell overnight in Milli-Q water (Millipore, USA). Polymer solution preparation and QCM-D monitoring Prior to QCM-D adsorption experiments, gelatine and agar were dissolved in Milli-Q water and dialyzed using 1 kda mesh membrane tubes (SpectraPor, Spectrumlabs) at 35 C and 85 C, respectively, followed by freezedrying. Purified gelatine and agar were dissolved for 30 min in Milli-Q water at 35 and 85 C, respectively to obtain solutions of 0.5 mg/ml concentration. The ph of prepared gelatine and agar solutions were 7.8 and 13.8 while the conductivities were 5.6 and 10.8 µs/cm, respectively. Finally, polymer solutions were filtered using 0.45 µm filters and degassed. QCM-D experiments were performed at a constant flow rate of 100 µl/min and the temperature was maintained at 25 C until adsorption plateau was reached. Thereafter, rinsing with Milli-Q water was applied to ascertain the irreversible binding of the polymers to pre-adsorbed layers on the QCM-D sensors. Finally, washed sensors were stored in desiccator until further investigation. Each set of experiments was performed at least by duplicate. The Voigt viscoelastic model (Q-Tools software, version 2.1, Q-Sense, Västra Frölunda, Sweden) was applied for calculating the adsorbed mass and film thickness of the gelatine- and agar-coated cellulose layers. In this model, the adsorbed layer is treated as a viscoelastic layer between the quartz crystal and a semiinfinite Newtonian liquid layer and can be represented as a system consisting of a spring and a dashpot filled with Table 1 - Testing conditions investigated in this study and respective nomenclature used in the text (X= crosslinker, U= Unrestrained, R= Restrained) Gelatine, Agar, % X, % to Designation Drying % to fibre to fibre polymer HCLC No No No U/R A4 No 4 No U/R G4 4 No No U/R G2A2 2 2 No U/R G4A4 4 4 No U/R G4X 4 No 3 U/R A4X No 4 3 U/R G4A4X U/R G4A4X (250 g/m 2 ) U viscous fluid connected in parallel (Höök et al., 1998; Voinova et al., 1999). For data evaluation or fitting frequency and dissipation data from several overtones (n = 3, 5, 7, 9, 11 and 13) of each adsorbed layer were used. The densities of the adsorbed gelatine and agar layers were assumed to be 1250 and 1300 g/m 3, respectively. Frequency shifts (Δf) at the third overtone (15 MHz) are presented since this overtone usually has the best signal-to-noise ratio. Atomic Force Microscopy (AFM) The surface morphologies of the cellulose, gelatine- and agar-coated cellulose films after QCM-D investigation were characterized by atomic force microscopy (AFM) in tapping mode with a Nanoscope IIIa multimode scanning probe microscope from Digital Instruments Inc. (Santa Barbara, CA, USA). The 3x3 µm 2 images were acquired at room temperature by using silicon cantilevers in air atmosphere. Prior to measurements, the samples were allowed to dry in desiccator at room temperature overnight. Image analyses were performed using NanoScope Analysis 1.5 software; no image processing except flattening was done. The rms surface roughness was measured from 3x3 µm 2 scan sizes. Stress-strain measurements Tensile strength and strain at break of the paper samples were determined in accordance with the SCAN-P 67:93 standard. Testing conditions The testing conditions that were applied in this work are shown in the Table 1. Formability strain measurement and tray forming Formability strain of the paper was measured using the 2D-formability tester in VTT Jyväskylä. The procedure is described in our previous paper (Vishtal, Retulainen, 2014a). The trays were formed from the high-grammage (ca. 250 g/m 2 ) paper sprayed with 4% of gelatine and 4% of agar, both containing crosslinker. Forming was performed on the pilot scale plastic thermoforming machine (air forming) in Coveris Rigid Packaging in Hämeenlinna, Finland. 454

4 Fig 1 - The influence of agar (A) and gelatine (G) addition in the presence and absence of the crosslinker (X) on the tensile strength and strain at break of unrestrained dried paper. HCLC stands for high and low consistency refined pulp. The numerals indicate the addition level (% relative to fibre mass). Results & Discussion The aim of this work is to improve extensibility and formability of paper using combined application of gelatine and agar, and to compare the effectiveness of such protocol over that involving the addition of a single component. An adsorption study (QCM-D experiments) was performed in order to elucidate possible mechanisms of interaction between these two polymers and cellulose. At first, spraying of paper was performed to verify whether the sequential addition of gelatine and agar has any superior effect than separate addition, and once this hypothesis was confirmed, additional adsorption and imaging investigations were carried out. The stress-strain behaviour of the paper treated with agar and gelatine The influence of single addition of agar, gelatine, and combined addition of agar and gelatine, as well as the influence of the crosslinker addition on the tensile strength (tensile index) and strain at break of unrestrained dried paper can be elucidated in Fig 1. From Fig 1, addition of 4% of either agar or gelatine brings nearly the same improvement in strain at break of the paper. The strain at break was clearly higher in all cases when gelatine and agar were used together (A2G2, A4G4, and A4G4X). The order of addition (agar and gelatine) in spraying is extremely important. Gelatine should be sprayed first, followed by the application of agar. Reversing this order makes the agar to form a nearly impermeable gel layer on the surface of paper, which prevents gelatine from interacting with cellulose, thus offsetting any positive effect on extensibility. On the other hand, when gelatine is added first, it readily penetrates and adsorbs onto cellulose; subsequently, addition of agar closes the surface with a gel layer that positively affect gelatine retention in the web. For restrained dried paper (Fig 2), agar and/or gelatine addition brings the same changes in the mechanical properties as in case of unrestrained dried paper. The only difference is the lower absolute strain at break values and somewhat higher tensile strength. Agar and gelatine were equally effective in improving the extensibility, which suggests that agar improves extensibility not only by increasing the drying shrinkage. Fig 2 - The influence of agar (A) and gelatine (G) addition in presence and absence of the crosslinker (X) on the tensile strength and strain at break of restrained dried paper Fig 3 - Selected stress-strain curves of untreated unrestrained dried paper (HCLC), paper sprayed with agar (A4), gelatine (G4), gelatine followed by agar (G4A4). The speculated mechanism of agar action to improve the extensibility of restrained dried paper can be related to its hygroscopicity. Under the standard conditions (25 C, 50% RH) agar-containing paper has somewhat high moisture content (9-10% vs. 7-8% for paper without agar), which positively affects strain at break of paper (Vishtal, Retulainen 2014b). The highest strain at break (ca. 7%) and tensile strength (ca. 80 Nm/g) for restrained dried paper were obtained by addition of agar, gelatine and a crosslinker. This suggests certain additivity between these two polymers or at least the absence of negative interactions. The addition of the agar and gelatine also changes the stress-strain behaviour of paper. The differences in the stress-strain curves of the agar-, gelatine- and gelatine/agar- treated paper can be seen in Fig 3. The addition of agar to the paper does not increase the tensile strength of paper, though provides somewhat higher elongation, which can be explained by the significantly increased drying shrinkage. The addition of gelatine increases tensile strength but the extensibility of such paper is close to that of agar-treated paper. This might indicate strengthened bonding. The stress-strain behaviour of gelatine/agar treated paper may include both effects of improved bonding and extended elongation brought by increased drying shrinkage. 455

5 Fig 4 - The influence of the agar (A) and gelatine (G) addition in presence and absence of the crosslinker (X) on the drying shrinkage and tensile stiffness of unrestrained dried paper, acronyms as in Fig 1. Previously, Vishtal and Retulainen (2014a) have shown that agar increases shrinkage of unrestrained dried paper. However, addition of gelatine does not affect much the drying shrinkage of paper (Fig 4). It can be seen that the stiffness of treated paper depends directly on the extent of drying shrinkage. Agar-treated paper shrinks most, which is reflected by the lower tensile stiffness measured for such paper (Fig 4). It is clear that sequential addition of gelatine and agar to the paper have better effect on mechanical properties than the separated (single) application of these polymers. However, it is not completely evident why these two polymers have an additive action towards extensibility improvement. Thus, additional adsorption and imaging experiments were performed in order to elucidate possible mechanisms of action. Adsorption of gelatine and agar on cellulose by using QCM-D Gelatine and agar interactions were studied via QCM-D method. The Langmuir Schaefer (LS) cellulose films (Tammelin et al., 2006) were used as substrate. The absence of voids and the lower surface roughness of LS films simplify the interpretation of data from adsorption and interfacial interactions. The interactions of agar and gelatine with LS cellulose film and adsorbed amounts measured for different sequences of addition are presented in Fig 5 and Table 2. Gelatine has negative surface net charge at ph 7.8 (isoelectric point of gelatine is 5.8). Likewise, LS cellulose is negatively charged. However, gelatine adsorption is rather irreversible since no significant desorption is observed after rinsing with the MilliQ-water (red profile in Fig 5). The same observation was reported by (Khakalo et al., 2014) who speculated that hydrogen bonding and other non-specific interactions are dominant over electrostatic forces. Moreover, gelatine adsorption can be facilitated by its flexible configuration at the interface, so that the surface of cellulose can accommodate a large amount of protein molecules, i.e., by suitably changing their structural orientation on the polysaccharide interface (Halder et al., 2005). Gelatine adsorption on pre-adsorbed agar layer (blue profile in Fig 5) demonstrated the same pattern as in the Fig 5 - Single and sequential adsorption of agar and gelatine on the LS cellulose film monitored with QCM-D. Table 2 - Calculated gelatine and agar adsorbed Voigt mass (Δm, mg/m 2 ) and layer thicknesses (nm) onto LS cellulose surfaces. Adsorprion sequence Thickness (nm) Δm (mg/m 2 ) Agar on cellulose 1.18 ± ± 0.2 Gelatine on agar 7.26 ± ± 0.6 Gelatine on cellulose 5.93 ± ± 0.4 Agar on gelatine 64.3 ± ± 1.7 case of adsorption onto cellulose. Moreover, the amounts of gelatine and agar adsorbed on cellulose were relatively similar. Agar adsorption on cellulose is rather negligible as indicated by almost immediate removal upon rinsing with solvent (blue line in Fig 5). However, this is in stark contrast when agar is adsorbed onto cellulose with a preadsorbed layer of gelatine. A large polymer amount is deposited in this case, even after rinsing (Table 2). This could be associated with electrostatic interactions between gelatine and agar. In general, upon introduction of agar solution (ph 5.6) most of the amino groups of gelatine acquire positive charges (pka of arginine is 9.0) meaning that gelatine pre-adsorbed layer is positively charged under these conditions. Agar in turn consists of a mixture of agarose and agaropectin that are heavily modified with acidic side-groups, such as sulfate, pyruvate and glucuronate, which are negatively charged at ph 5.6. It should be noted that adsorbing gelatine layer formation proceeds slower than that of agar. Both agar and gelatine are prone to form gel-like structures via formation of a hydrogen-bonded, cross-linked network in water. However, agar gel sets faster than gelatine due to its higher molecular weight. Hence, under the used experimental conditions, agar might interact with the substrate already in a form of a gel, as indicated by a sharp drop in QCM-D oscillating frequency, thus rapidly covering the surface. In contrast, gelatine, being still in a dissolved state, slowly undergoes structural orientation at the interface. Irreversible adsorption of agar on gelatine improves retention of gelatine in paper during wet pressing as well as may improve the interactions at the fibre-polymer 456

6 Fig 6 - The cross-sectional SEM images of the unrestrained dried paper without any additives (A) unrestrained dried paper sprayed with 4% of agar (B), and unrestrained dried paper sprayed with 4% of gelatine and 4% of agar (C). contact zone. These phenomena improve compliance of fibre bonds, which is reflected in the increased strength and extensibility (Borodulina et al. 2012). SEM Web cross-sections SEM imaging was performed in order to elucidate the changes in the paper structure by single and sequential addition of agar and gelatine. Previously, it was shown that the agar formed a film on the surface of paper (Vishtal, Retulainen 2014a), while gelatine somewhat densified the paper structure (Khakalo et al. 2014). The cross-sectional SEM images of the untreated, agartreated, and agar- and gelatine- treated paper are shown in Fig 6. From Fig 6b, an agar film is evenly observed on the surface of paper without any signs of the agglomeration or thinning. There is also a clear indication of a close contact and high degree of adhesion between the agar layer and fibres, which supports the hypothesis of shrinkage transfer from drying agar film to fibre network proposed by Vishtal and Retulainen 2014a. SEM imaging of the paper treated with agar and gelatine (Fig 6c) appears to be somewhat densified and more consolidated on the surface layer onto which the polymers were sprayed. The agar layer on the surface can be still distinguished but not through the whole surface of the paper sample. It seems that the agar and gelatine were intermixed on the surface and the formation of an agar film was not evident. It can be suggested that the agar mixed with the gelatine has a higher potential to diffuse into the paper, which might significantly strengthen both the surface layer as well as the bulk of the paper. SEM web plane view The effects of the sequential addition of agar and gelatine are not easily observable in SEM cross-sections. Thus, plane view images were acquired to further clarify the effect of the polymers. The SEM images of the unrestrained dried paper (A), unrestrained dried paper sprayed with agar (B), gelatine (C), and their combination (D) are shown in Fig 7. As it might be expected, non-fibrillated softwood pulp fibres formed a relatively porous and bulky network. Also, a quite large number of deformations in fibres are observed, likely due to HC treatment and drying shrinkage. Addition of agar in turn leads to the formation of a film on the surface of the paper. The fibres under the agar layer appear to be somewhat swollen, which might be a consequence of high hygroscopicity of the agar film. Due to this fact and possibly due to high drying shrinkage, axial deformations in fibres appear to be more profound than in the untreated paper. The surface of the paper treated with the gelatine appears to be denser and less porous than the surface of untreated paper. This might be a consequence of the improved bonding and larger fibre contact area provided by gelatine. Unlike agar, gelatine penetrates into the paper and accumulates at the fibre crossings, thus providing a reinforcing effect to the paper (Khakalo et al. 2014). The film on the surface of paper treated with agar and gelatine is interrupted by voids which might originate from the intermixing of agar and gelatine, which prevents agar from forming a continuous film on the surface or by weakening it. The evenness of the film was improved by addition of crosslinker to the polymer solutions (Fig 7e). Evenness of the film is probably improved due to the depression of the molecular mobility and increasing viscosity of agar solution. Nanoscale investigation of the LS cellulose film surfaces modified with gelatine and agar using atomic force microscopy The changes in the nanoscale surface morphology of LS cellulose films modified with gelatine and agar can provide further insights into the nature of the interactions between these polymers and cellulose. The AFM images of pristine LS cellulose film as well as cellulose surfaces modified sequentially with gelatine and agar are shown in Fig 8. As can be seen from the AFM images, gelatine forms a thin layer on cellulose surface, which follows from slightly increased surface roughness without major change in the surface morphology. Subsequent addition of agar leads to a formation of a cross-linked polymer network on the cellulose surface. Agarose may coil to double helix and join together, thus forming a crosslinked gel. 457

7 Fig 7 - The Surface SEM images of unrestrained dried untreated paper (A), unrestrained dried sprayed with 4% of agar (B), unrestrained dried paper sprayed with 4% of gelatine (C), unrestrained dried paper sprayed with agar and gelatine (4% of each) (D), unrestrained dried paper sprayed with agar and gelatine (4% of each) and AmZrCarb (E). Fig 8 - AFM height images and corresponding roughness profiles of unmodified cellulose (A), cellulose modified with gelatine (B) and agar modified cellulose pre-treated with gelatine (C). The Z-range of all the images is 5 nm. 458

8 AFM images were taken in air, which could mask the actual size of the polymer structures in water. The pronounced cross-linked nature of the agar gel is the major distinguishing feature in contrast to the gelatine. These observations are in agreement with the changes observed in SEM images of the surface structures (Fig 7). Formability strain of paper sprayed with agar and gelatine As natural polymers, agar and gelatine have the ability to soften at elevated temperatures. In analogy to previous research (Vishtal, Retulainen 2014a, Khakalo et al. 2014) formability of treated paper was studied using 2Dformability tester (Fig 9). Fig 9 - The influence of the temperature on the formability strain of the untreated paper and paper sprayed with agar, gelatine and gelatine-agar. The formability response to the temperature of paper treated with agar and gelatine is similar to that of paper after single addition (either agar or gelatine) (Vishtal, Retulainen 2014a, Khakalo et al. 2014). This might indicate that softening is induced by weakening of hydrogen bonding, which is the primary bonding type in all investigated samples. The maximum formability strain of paper treated with agar and gelatine is approx. 18%, which translates into shapes with a depth of around 2-3 cm (depending on curvature and shape) produced in a fixed blank forming process (no compressive wrinkles are formed) (Fig 10). Fig 10 - An example of the shape produced from highgrammage paper (G4A4X, ca. 250 g/m 2 ). In sum, sequential application of gelatine and agar is more effective to improve the extensibility and strength of paper compared to single applications. The principal convertibility of such paper in 3D-forming process is demonstrated. Summary Based on the experimental results presented in this study it is possible to make the following conclusions: 1. Compared to single addition protocols, the sequential application of gelatine and agar is more effective to improve paper web extensibility and formability. 2. Sequential application is only effective if gelatine is added first followed by agar. Gelatine easily penetrates in the Z-direction and adsorbs on cellulose. Agar forms a film on the surface, which provides a strengthening effect. 3. Adsorption studies (QCM-D) revealed that agar retention is significantly improved upon gelatine preadsorption. 4. The SEM images of paper treated with gelatine and agar reveal inter-mixing of the polymers; however, an agar film is still formed. 5. Crosslinker addition to the agar and gelatine solutions further improves extensibility and tensile strength of paper. However, the possible mechanism of such effect is still to be elucidated. 6. Formability of paper treated with gelatine and agar allows to produce tray-like, 3D-shapes with 2-3 cm in depth by using fixed blank forming process. Acknowledgements The Future Biorefinery Programme at Finnish Bioeconomy Cluster Ltd. and VTT Graduate School are acknowledged for providing financial support to this study. Stora Enso RC is thanked for taking the SEM images. Coveris Rigid Packaging Ltd. (Hämeenlinna, Finland) is thanked for the opportunity to use pilot scale thermoforming equipment for the manufacturing of the trays. Literature Aulin, C., Ahola, S., Josefsson, P., Nishino, T., Hirose, Y., Osterberg, M., and Wågberg, L. (2009): Nanoscale cellulose films with different crystallinities and mesostructures their surface properties and interaction with water, Langmuir, 25 (13), Banerjee S., and Bhattacharya, S. (2012): Food gels: gelling process and new applications, Crit. Rev. Food. Sci. Nutr., 52 (4), Barrett, T., et al. (2014): Plot 1. Gelatin. Paper through time: Nondestructive analysis of 14th- through 19th-century papers. The University of Iowa, Electronic material, available via: /plot1.php. Borodulina, S., Kulachenko, A., Galland, S., and Nygårds, M. (2012): Stress-strain curve of paper revisited, Nord. Pulp Paper Res. J. 27(2), Halder, E., Chattoraj, D. K., and Das, K. P. (2005): Adsorption of biopolymers at hydrophilic cellulose-water interface, Biopolymers, 77(5),

9 Hauptmann, M., and Majschak, J.P. (2011): New quality level of packaging components from paperboard through technology Improvement in 3D forming, Packag. Technol. Sci., 24(7), Höök, F., Rodahl, M., Brzezinski, P., and Kasemo, B. (1998): Energy dissipation kinetics for protein and antibody antigen adsorption under shear sscillation on a quartz crystal microbalance, Langmuir, 14(4), Khakalo, A., Filpponen, I., Johansson, L.S., Vishtal, A., Lokanathan, A.R., Rojas, O.J., and Laine, J. (2014): Using gelatin protein to facilitate paper thermoformability, React. Funct. Polym., 85, Kontturi, E., Thuene, P. C., and Niemantsverdriet, J. W. (2003): Cellulose model surfaces - simplified preparation by spin coating and characterization by x-ray photoelectron spectroscopy, infrared spectroscopy, and atomic force microscopy, Langmuir, 19(14), Laine, J., Lindström, T., Nordmark G., and Risinger, G., (2000): Studies on topochemical modification of cellulosic fibres Part 1. Chemical conditions for the attachment of carboxymethyl cellulose onto fibres, Nord. Pulp Paper Res. J. 15(5), Pranoto, Y., Lee, C. M., and Park, H. J. (2007): Characterizations of fish gelatin films added with gellan and k- carrageenan, LWT-Food Science and Technology, 40 (5), Schaub, M., Wenz, G., Wegner, G., Stein, A., and Klemm, D. (1993): Ultrathin films of cellulose on silicon wafers, Adv Mater, 5, Svensson, A., Lindström, T., Ankerfors, M., and Östlund, S (2013): 3D-shapeable thermoplastic paper materials, Nord. Pulp Paper Res. J., 28(4), Tammelin, T., Saarinen, T., Österberg, M., and Laine, J. (2006): Preparation of Langmuir/Blodgett-cellulose surfaces by using horizontal dipping procedure. Application for polyelectrolyte adsorption studies performed with QCM-D. Cellulose, 13(5), Vishtal A. and Retulainen E. (2014a): The improvement of paper extensibility, wet and dry strength by spray addition of agar solutions, Nord. Pulp Paper Res. J., 29(3), Vishtal, A., and Retulainen, E. (2014b): Boosting the extensibility potential of fibre networks: A review, BioRes., 9(4), Vishtal A., Hauptmann M., Zelm R., Majschak J.P. and Retulainen E. (2014): 3D forming of paperboard: the influence of paperboard properties on formability, Packag. Technol. Sci., 27(9), Voinova, M. V., Rodahl, M., Jonson, M., and Kasemo, B. (1999): Viscoelastic acoustic response of layered polymer films at fluid-solid interfaces: continuum mechanics, Approach. Phys. Scripta, 59(5), Zeng, X., Vishtal, A., Retulainen, E, Sivonen, E. and Fu S. (2013): The Elongation of Paper How fibres should be deformed to make paper extensible?, BioRes 8(1), Manuscript received February 16, 2015 Accepted April 7, 2015 PAPER PHYSICS 460