Preparation and characterization of a transparent amorphous cellulose film

Abstract

Amorphous cellulose film (ACF) was prepared from cellulose solution in lithium chloride (8 wt%)/N,N-dimethylacetamide by regeneration with acetone. The obtained ACF possessed a dense, smooth surface, and excellent transparency. The X-ray diffraction results indicated that ACF was highly amorphous, which was further confirmed by solid-state ¹³C-NMR and Fourier transform infrared (FT-IR) spectra. The tensile analysis implied that the elongation at break (23.9%) and the maximum stress (157 MPa) of ACF derived from the Whatman CF11 fibrous cellulose were higher than those of cellophane (19.9% and 135 MPa, respectively). In addition, the enzymatic hydrolysis of ACF and cellophane showed that the former had a higher hydrolysis rate (about 7 times higher than the latter), indicating its outstanding environmental friendliness. This work provides a simple, less-destructive, and universal method to prepare transparent ACF, which could serve as a promising packaging material to replace cellophane.

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Introduction

With the depletion of fossil fuel and the ever-increasing environment concerns, cellulose has again attracted researchers' interest as a raw material over the last few decades, due to it being an abundant resource, and having extraordinary renewability, biodegradability, and a unique molecular structure.^1,2 Cellulose possesses great potential for application in fibers, film, coatings, and the matrix of control-release systems, especially in the food packaging area.³ Nowadays, commercial cellulose film (cellophane) is mainly produced by the viscose method. Another two methods (the carbamate and Lyocell technologies) developed in recent years are also used to produce cellulose film.³ However, most cellulose films prepared by the existing methods possess a cellulose II structure.

It is well known that cellulose is composed of a group of crystalline allomorphs (I, II, III_I, III_II, IV_I, and IV_II) and has a disordered (amorphous) structure with two polymorphs (I_α and I_β) in the cellulose I.⁴ The molecular chains in amorphous cellulose are loosely arranged, unlike their tight compaction in cellulose's crystalline counterpart, which should cause a significant difference in some aspects, such as in its mechanical properties,⁵ reaction kinetics,⁶ and enzymatic hydrolysis rate.^7–9 Some special applications, such as enzyme screening and displaying material, could be developed using amorphous cellulose film (ACF). Meanwhile, it is of great importance to investigate the behaviors of ACF for better utilization of this cellulose resource. However, most cellulose films reported to date possess the crystalline structure with cellulose II, since it is thermodynamically more stable than the other allomorphs.^3,10–12 In contrast, ACF with a good performance has rarely been reported, even though many methods have been developed to prepare amorphous cellulose samples, such as ball milling,¹³ hydrolysis of cellulose triacetate,¹⁴ regeneration from cadmium ethylenediamine,¹⁵ sodium cellulose xanthates,¹⁶ cuprammonium hydroxide,¹⁶ dimethylsulfoxide/paraformaldehyde,¹⁷ phosphoric acid,¹⁷ and from SO₂/diethylamine/dimethylsulfoxide solution.¹⁸ Moreover, most of these methods either used toxic reagents or inevitably caused degradation of the cellulose, which were the major disadvantages for scientific studies and for the practical application of ACF.

The cellulose solvent of LiCl–N,N-dimethylacetamide (DMAc) was first reported by McCormick and Lichatowich in 1979.¹⁹ Initially, water swelled and opened the structure, and the intermolecular and intramolecular hydrogen bonds were replaced by hydrogen links with H₂O. Then, methanol and DMAc were introduced subsequently to remove water and to impede the re-formation of the intermolecular and intramolecular hydrogen bonds. In the final step, the swollen sample was added into LiCl–DMAc solvent, with stirring until dissolved.^20,21 Although the mechanism of dissolution remained controversial, one generally accepted principle was that [DMAc_n + Li]⁺ macrocation evolved, leaving the chloride anion (Cl⁻) free. Thereby, Cl⁻, being highly active as a nucleophilic base, was able to play a major role by breaking up the intermolecular and intramolecular hydrogen bonds.^19–24 The whole process was operated under mild conditions, and no appreciable degradation occurred. In addition, the cellulose solution in LiCl–DMAc was reported to be extremely stable,^20,21 which made it attractive for practical application. However, only a few reports were related to the preparation of cellulose film from LiCl–DMAc solution.^25–32 Moreover, none of them mentioned the fabrication of ACF.

In the present study, ACF with excellent transparency was prepared by regeneration from LiCl–DMAc solution. The relationships between the concentration of cellulose solution and the mechanical properties were systematically investigated. We also compared the enzymatic hydrolysis rate of ACF and commercially available cellophane. This study aims to provide a simple, less-destructive, and universal method to prepare amorphous cellulose film, and, in addition, to enhance our understanding about the behaviors of amorphous cellulose to open it up to new practical applications.

Experimental section

Materials

Whatman CF11 fibrous medium cellulose powder (CF11, cotton origin, 50–350 μm, GE Healthcare Life Science Corp., Piscataway, NJ, USA), microcrystalline cellulose powder (Merck, cotton origin, 20–160 μm, ≥80%, Merck KGaA, Darmstadt, Germany), Avicel SF microcrystalline cellulose powder for thin layer chromatography (Avicel, pulp origin, mean particle size around 10 μm, Funakoshi, Co. Ltd., Tokyo, Japan), and bacterial cellulose prepared as described previously, except for under static condition (BC, Gluconacetobacter xylinus (Brown) Yamada et al. ATCC 53524),³³ were used as the cellulose resource. For reference, an amorphous cellulose sample derived from CF11 was prepared by a vibrating ball-mill in a N₂ atmosphere for 48 h by using ceramic balls (Ball-mill, Type MB-1 Vibrating mill, Chuo Kakohki, Co. Ltd., Nagoya, Japan).¹³ Cellophane (thickness ≈ 22 μm) without any additives and coating was supplied by Futamura Chemical Co. Ltd., Nagoya, Japan. N,N-Dimethylacetamide (DMAc, purity > 99%) was obtained from Tokyo Chemical Industry Co. Ltd., Japan. Anhydrous lithium chloride (LiCl), D-glucose, anhydrous citric acid, 3,5-dinitrosalicylic acid (DNS), potassium sodium L-(+)-tartrate tetrahydrate (Rochelle salt), methanol, and acetone were obtained from Wako Pure Chemical Industries Ltd., Japan. Cellulase from Aspergillus niger (activity ≥ 60 000 units per mg) was obtained from MP Biomedicals, LLC., Santa Ana, CA, USA. All other reagents not specially mentioned were used as received.

Preparation of cellulose solution

The first step was the fabrication of cellulose solution from different cellulose resources. To facilitate mass production, the reported method^20,21 was simplified (Fig. S1†). In a typical run, 3 g CF11 was immersed in deionized water for 4 h at room temperature (RT, 25 °C) and filtered to remove water, followed by successive solvent exchange with methanol and DMAc, each for 2 h. Then, the activated cellulose was soaked in 47 g LiCl (8 wt%)–DMAc solution under the protection of a N₂ atmosphere. After mechanical stirring for 12 h, a clear cellulose solution was obtained. To complete the dissolution of cellulose, the solution was kept overnight at 4 °C.²⁰ Finally, a transparent cellulose solution with 6 wt% concentration was obtained. The solution was stored at 4 °C until use. For concentrations below 6 wt%, the solution became clear only after stirring for several hours. With respect to 8 wt%, 24 h were needed for complete dissolution. According to the same procedure, 6 wt% of Merck and 6 wt% of Avicel cellulose solutions were obtained. The dissolution time was less than 2 h for both samples. On the contrary, even for 1 wt% BC solution, the dissolution took at least 24 h, and the viscosity of the solution was higher than the other samples.

Preparation of cellulose film

The cellulose solution was degassed by centrifugation at 10 000 rpm for 10 min at RT, then cast on a glass plate. The thickness was controlled at 0.5 mm using an applicator. After the glass plate was gently immersed into 100 ml of acetone bath, a transparent cellulose gel was immediately formed. The cellulose gel was kept in acetone for 1 h, and washed with 100 ml deionized water five times to remove the salt completely, each time for 1 h. For the preparation of cellulose films, the usability of various kinds of organic solvents other than acetone was checked as regeneration solvents, with water, methanol, and ethanol. The washed sample was fixed on the poly(methyl methacrylate) (PMMA) plate with adhesive tape to prevent shrinkage¹⁰ and was then dried in the oven at 40 °C for 2 h. The glass and Teflon plates were also employed as the substrate for this drying process (Fig. S1†). The sample was further dried in a desiccator containing phosphorus(V) oxide at RT for at least 48 h. Finally, for 6 wt% of CF11 solution, a transparent cellulose film was obtained with a thickness of about 22 μm. In the following content, the samples prepared from different kinds and concentration of cellulose solutions are referred to as CF11 4%, CF11 5%, CF11 6%, CF11 7%, CF11 8%, Merck 6%, Avicel 6%, and BC 1%, respectively.

Enzymatic hydrolysis of CF11 6% and cellophane

CF11 6% and cellophane with similar thicknesses of 22–23 μm were treated with cellulolytic enzymes. Hydrolysis experiments were run concurrently. To minimize the difference in specific area, CF11 6% and the cellophane were cut into square shapes with the same size of about 2 cm × 2 cm. For each film, 150 mg of sample, 10 ml of sodium citrate buffer solution (0.05 M, pH 4.8), and 20 mg of cellulase were added in this order to a 50 ml vial. The vials were capped and put into a bioshaker at 40 °C with a shaking speed 200 rpm. To monitor the content of released reducing sugar, 100 μl of the supernatant was transferred from the vial to a test tube periodically and diluted with 2.9 ml of Milli-Q water, followed by blending with 3 ml of DNS reagent, which was prepared according to the method reported by Miller.³⁴ The test tubes were heated in a boiling water bath for 15 min. After the development of color, 1 ml of 40 wt% Rochelle salt solution was added immediately. The test tubes were rapidly cooled down to RT using running water. The absorbance of the solution was measured at 575 nm using a Hitachi U2810 UV-visible spectrophotometer. Finally, the released reducing sugar content was calculated as D-glucose.

Characterization

Fourier transform infrared (FT-IR) spectra in the attenuated total reflection (ATR) mode were recorded on a Nicolet iS5 FT-IR Spectrometer with iD5 ATR accessory (Thermo Fisher Scientific Inc., Waltham, MA, USA). The optical transmittance of the films were measured from 200 to 900 nm using a Hitachi U2810 UV-visible spectrophotometer. Scanning electron microscopy (SEM) analysis was carried out by a HITACHI SU-3500 instrument (Hitachi High-Technologies Corp., Tokyo, Japan). Wide-angle X-ray diffraction (XRD) was performed on an X-ray diffractometer (Shimadzu XRD-6100) at a rate of 2° (2θ) min⁻¹ over the 2θ range from 5° to 40°. The X-ray radiation used was Ni-filtered CuKα with a wavelength of 0.15406 nm. The voltage and current were set at 40 kV and 30 mA, respectively. Solid-state ¹³C-NMR spectra with cross polarization/magic angle spinning (CP/MAS) were recorded on a 600 MHz NMR spectrometer (150.95 MHz for ¹³C, Advance III, Brucker BioSpin GmbH, Rheinstetten, Germany) at RT. The chemical shift was calibrated by the carbonyl carbon of glycine at 176.46 ppm. The cellulose distribution in cellulose films was observed by an X-ray computed tomography (XCT) instrument at 80 kV and 100 μA with an isotropic voxel of 600 nm (SKY Scan 1172, High resolution micro-CT, Brucker AXS GmbH, Karlsruhe, Germany). The tensile properties were measured by a Shimadzu EZ Graph instrument equipped with a 500 N load cell (Shimadzu Corp., Kyoto, Japan). A cross-head speed of 1 mm min⁻¹ was used. The sample was cut into rectangular strips of 40 mm × 5 mm and tested with a span length of 10 mm.

Results and discussion

Characterization of cellulose film

To prepare cellulose film with a good appearance, three substrates were employed during the drying process (Fig. S1†). The film was well attached to the glass plate, but the bonding force between the surfaces was so strong that the film could not be peeled off from the plate. In contrast, the bonding force between the film and Teflon was too weak to maintain the shape of the film, which was easily deformed after drying. The best result was obtained by using the PMMA plate. The bonding force between the surfaces was strong enough to fix the cellulose film. Meanwhile, the film can be easily detached from the plate. Considering the cost and environmental friendliness, four common solvents: water, methanol, ethanol, and acetone, were chosen as the regeneration solvent. The first three kinds of solvents caused a drastic shrinkage of the cellulose film. Only in the case of acetone, however, was a transparent, flat and smooth cellulose film obtained. The usability of acetone as a regeneration solvent has been previously reported,^10,35 but no description about the preparation of transparent films has been noted using the LiCl–DMAc solvent system. In addition, it was reported that acetone will lead to a better amorphous cellulose structure.¹⁸ Based on the above reasons, acetone was chosen as the regeneration solvent. All of the cellulose films regenerated individually from CF11, Merck, Avicel, and BC cellulose solutions in LiCl–DMAc by acetone possessed good optical appearance. Among them, CF11 6% was taken as a typical example, and its image is shown in Fig. 1. A smooth and dense surface was observed by SEM in the micron level (Fig. S2†). The thickness of the cellulose films increased with the increasing concentration of cellulose solution from 4 wt% to 7 wt% (16, 18, 22, and 29 μm, respectively), and a slight decrease appeared at 8 wt% (27 μm), because of the incomplete dissolution of cellulose into the solvent.

Fig. 1 Image of a transparent film of CF11 6%.

The crystalline structure of the native CF11, Merck, Avicel, and BC samples was studied by XRD (Fig. 2a). The typical diffractions due to I_β-rich natural cellulose for the former three were observed at 2θ = 14.8°, 16.3°, and 22.6°, which corresponded to the (11̄0), (110), and (200) planes,³⁶ respectively. In the case of I_α-rich BC, three distinct diffractions (100), (010), and (110) were observed at 2θ = 14.6°, 16.9°, and 22.7°, respectively.³³ After regeneration, these diffractions disappeared, showing a broad peak at 2θ ≈ 20° (Fig. 2b), which indicated that the cellulose I structure was transformed to amorphous cellulose during the dissolution, regeneration, and drying process. Compared to Ball-mill cellulose, the regenerated samples showed similar diffractions, except that, for Avicel 6%, there were weak peaks appearing at around 2θ = 12.1° and 22.0°. These diffractions were attributed to the cellulose II structure, indicating that a small amount of the cellulose II structure was also formed apart from just amorphous cellulose.

Fig. 2 X-ray diffractions of (a) native samples, (b) regenerated samples and a ball-milled sample.

The amorphous structure of the cellulose films was further confirmed by CP/MAS ¹³C-NMR (Fig. 3). The native cellulose showed characteristic signals assignable to cellulose I (Fig. 3a): the signals around 105 ppm were assigned to the most deshielded anomeric carbon atom C1; the sharp signal at 89 ppm and the broad signal between 86 ppm and 80 ppm were assigned to C4 in the crystalline and amorphous regions, respectively; the signals from 79 ppm to 70 ppm belonged to C2, C3, and C5; similar to C4, C6 displayed a sharp signal at 65 ppm and a broad signal around 63 ppm, corresponding to the crystalline and amorphous regions, respectively.³⁷ After regeneration (Fig. 3b), all the signals showed a decrease in sharpness, especially for C4. The sharp peaks at 89 ppm totally disappeared for CF11 6%. With respect to the other regenerated samples, only two small signals appeared in this area because of the regeneration of a small amount of the cellulose II structure. Moreover, the strength of signals from 86 ppm to 80 ppm increased for all samples. These changes stemmed from the differences between the crystalline and amorphous structure, including conformational differences, differences in bond geometries, and non-uniformities of the neighboring chain environments.³⁸ The results for the regenerated samples were similar to the ball-milled sample, indicating that highly amorphous cellulose films were obtained. Moreover, for CF11, the transformation from cellulose I to amorphous cellulose was more completely achieved by regeneration from the LiCl–DMAc solution, compared to the ball-milling method, since there were still two small signals around 89 ppm displayed for the ball-milled sample, due to the remaining cellulose I structure.

Fig. 3 CP/MAS ¹³C-NMR spectra of (a) native samples, (b) regenerated samples and a ball-milled sample.

The FT-IR results (Fig. 4) also provided evidence of the transformation from the crystalline to the amorphous structure. The absorption at 1429 cm⁻¹ was assigned to the CH₂ symmetrical bending vibration, and the absorption at 897 cm⁻¹ responded to the change in molecular conformation due to rotation about the β-(1 → 4)-D-glucosidic linkage.³⁹ Normally, these two bands were used to measure the crystallinity of cellulose. In the native cellulose (Fig. 4a), a sharp absorption at 1429 cm⁻¹ and a weak band at 897 cm⁻¹ appeared. In the regenerated cellulose film (Fig. 4b), on the other hand, only a broad absorption at 1429 cm⁻¹ could be seen and the intensity of the absorption at 897 cm⁻¹ increased, proving the low crystallinity of the regenerated film. In addition, the intensity of the other peaks at 1335, 1315, 1111, 1057, and 1033 cm⁻¹ decreased after the regeneration. The broad absorption in the 3600–3000 cm⁻¹ region, due to the OH– stretching vibration, could reflect changes of the hydrogen bonds. A narrow peak appeared at 3340 cm⁻¹ for native cellulose, which was caused by the regular arrangement of intramolecular and intermolecular hydrogen bonds. After regeneration, the regularity of hydrogen bonds was disturbed, and the peak shifts to a higher wavenumber at 3350 cm⁻¹ and broadening were also detected. Since it was reported that unbounded or “free” OH groups absorb infra-red light at 3584 to 3650 cm⁻¹,⁴⁰ which was higher than that observed in the prepared films, we could conclude that the hydroxyl groups in the amorphous structure existed in an irregular arrangement of hydrogen bonds rather than in the free mode.

Fig. 4 FT-IR spectra of (a) native samples, (b) regenerated samples and a ball-milled sample.

In conclusion, all of the cellulose samples, namely CF11, Merck, Avicel, and BC, could be transformed from cellulose I to a highly amorphous structure. Among these, the best result was obtained with CF11, whereas there was only a small amount of cellulose II structure regenerated in the case of Merck, Avicel, and BC. Therefore, in the following content, the properties of the ACF derived from CF11 were investigated and compared with those of cellophane.

Mechanism of the formation of ACF

We have attempted to give an explanation about the formation of ACF. Cellulose is mainly composed of two parts, namely, crystalline and disordered regions. In most cases, the latter is referred to as “amorphous”. Compared to the amorphous parts, the crystalline structure is more difficult to access and is the main obstacle to dissolution. First, water is used to swell the crystalline lattice, making the LiCl–DMAc solvent easy to penetrate. During the dissolution process, the [DMAc_n + Li]⁺ macrocation is evolved, leaving the chloride anion (Cl⁻) free, which disturbs the intermolecular and intramolecular hydrogen bonds by forming new hydrogen bonds with the hydroxyl groups of the cellulose chain.²⁴ Afterwards, the cellulose chains become much easier to tear off from the crystalline lattice and drag into solution. This process is repeated until the “true” solution is formed, in which the cellulose chains are freely extended, unlike in the other kinds of solvents such as aqueous NaOH/urea.⁴¹ When this cellulose solution is immersed into a poor solvent, cellulose is immediately reprecipitated from the solution through the entanglement of the molecular chains, leading to the formation of cellulose gel. Followed by the drying process, water quickly evaporates accompanying the collapse of the pores in the hydrogel, due to the high surface energy of water. In addition, regeneration of the hydrogen bonds between the cellulose chains provides another driving force. Finally, ACF with a dense structure was obtained. Although cellulose II is thermodynamically more stable, the drying process is so fast that the kinetic control takes advantage, and not enough time is left to rearrange the cellulose chains, which are more likely aligned in a bent and twisted conformation. A large amount of intramolecular hydrogen bonds replace the intermolecular hydrogen bonds existing in native cellulose to stabilize this conformation, making the ACF stable in common conditions unless exposed to high temperature, moisture, or pressure.

The influence of the cellulose resources on its solubilization and formation of the amorphous structure is worth mentioning. Three plant celluloses with different particle sizes (CF11 > Merck > Avicel) were chosen. According to the XRD (Fig. 2) and ¹³C-NMR (Fig. 3) results, the sequence of the perfection of the amorphous structure was CF11 > Merck > Avicel, which is consistent with their particle size. To some extent, particle size is related with the molecular chain length or degree of polymerization (DP). In the case of Avicel, the short chain length causes a large specific surface area contactable with the solvent, which promotes their high mobility leading them to form the thermodynamically favored cellulose II structure during the regeneration and drying process. With respect to BC, because of its distinct complex entangled structure, the solubilization is difficult. Moreover, the viscosity of solution is obviously higher than those of the other three plant celluloses, reflecting the longest chain length of BC among the chosen cellulose resources. The molecular chains of BC probably still remain orientated, to some extent, in the solubilized state, which easily leads to the formation of the crystalline structure. Therefore, only for the sample with a median particle size, such as CF11, more perfect amorphous structure could be obtained.

Transparency of CF11 and cellophane

The transparency of cellulose films was investigated by UV-visible spectroscopy. As shown in Fig. S3,† all of the cellulose films from CF11 4% to CF11 8% possessed high transparency, not only in the visible region (transmittance is about 90%), but also in the near ultraviolet region (transmittance is above 70%), which was better than the commercial cellophane (Fig. 5) and the other cellulose films reported in the ref. 10, 42 and 43. The reason may be due to the difference of the crystalline structure between CF11 films and cellophane, the latter being characterized as cellulose II by XRD (Fig. S6†) and ¹³C-NMR spectra (Fig. S7†). To further investigate the reason, XCT was measured, as it was recently used in the cellulose materials area.^44–46 With the help of XCT, a volumetric map of the specimen in three dimensions could be obtained. Meanwhile, the distribution of different components and pores could be differentiated. As the XCT images (Fig. 6) showed, CF11 6% was more homogeneous compared with cellophane, in the order of ≥600 nm. In the latter case, the presence of cloudy aggregates that may be composed of the small crystal grains could be clearly detected. Such aggregates would cause the scattering of light, resulting in the inferior transparency of cellophane.

Fig. 5 Transmittance of CF11 6% and cellophane in the UV-visible wavelength region.

Fig. 6 X-ray CT image of (a) CF11 6% and (b) cellophane.

Mechanical properties of CF11 and cellophane

The tensile properties of cellulose films were investigated. For reference, cellophane was tested. Fig. 7 shows the typical stress–strain curves of cellulose samples. Table 1 summarizes the tensile properties of the measured samples. The elongation at break and maximum stress for CF11 4% were 15.9% and 133 MPa, respectively. With the increasing concentration of the cellulose solution, the elongation at break increased. After the maximum value of 23.9% was obtained for CF11 6%, an obvious decrease was shown for CF11 8%, because of the incomplete dissolution of cellulose, which was confirmed by the XRD results (Fig. S4†). The undissolved grain will function as a defect and is detrimental to the tensile performance. The largest maximum stress value was about 160 MPa, belonging to CF11 5% and CF11 6%. Because CF11 6% and cellophane possess similar thicknesses, the tensile properties of them were compared. The elongation at break (23.9%) and the maximum stress (157 MPa) of CF11 6% were higher than those of cellophane (19.9% and 135 MPa, respectively). Although the cellulose resource would affect the mechanical properties, such a rarely reported performance is probably attributed to the distinctive amorphous structure of ACF. In the amorphous structure, cellulose chains are assumed to be bent and twisted, intermolecular hydrogen bonds are ripped off and regenerated under stretching, leading to the extension and rearrangement of cellulose chains in a regular way, and ultimately a higher elongation at break and maximum stress are desirably obtained.

Fig. 7 Stress–strain curves of CF11 6% and cellophane.

Table 1 Tensile properties of ACFs and cellophane

	ACF 4%	ACF 5%	ACF 6%	ACF 7%	ACF 8%	Cellophane
a Standard deviation (SD). For each group experiment, ten samples were tested and at least three samples were chosen.
Elongation (%)	15.9 ± 1.1^a	20.7 ± 1.2	23.9 ± 3.2	22.5 ± 2.2	17.6 ± 3.3	19.9 ± 3.7
Max stress (MPa)	132 ± 7	161 ± 8	157 ± 8	145 ± 9	145 ± 9	135 ± 6

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Enzymatic hydrolysis of CF11 6% and cellophane

The results of the enzymatic hydrolysis of CF11 6% and cellophane are shown in Fig. 8. In the initial 8 h, the concentration of the reducing sugar released by CF11 6% rapidly rose to 3.7 mg ml⁻¹, showing a little lower rise in the following time. After 48 h, the concentration increased up to 11.9 mg ml⁻¹. Assuming that the released reducing sugar was only comprised of glucose, it can be calculated that about 107 mg of CF11 6% (71.5% of the total amount) was hydrolyzed. Moreover, it was observed that CF11 6% was partially hydrolyzed into small pieces after 48 h. In contrast, the concentration of reducing sugar released by cellophane rapidly increased to 1.0 mg ml⁻¹ in the initial 4 h, showing only a small increase to 1.7 mg ml⁻¹ after 48 h. About 15.0 mg of cellophane (10.0% of the total amount) was hydrolyzed. In addition, the films remained intact. The enzymatic hydrolysis rate of CF11 6% was above 7 times higher than that of cellophane. To explain this phenomenon, the mechanism of enzymatic hydrolysis would be focused. Generally, the activity of cellulolytic enzymes largely depends on their types (endo- and exo-glucanases) and accessibility on the surface of cellulose as a subtrate.^47,48 Usually cellulase derived from Trichoderma and Aspergillus spp. are used for the degradation of natural cellulose I and more soft cellulosic materials, respectively. By considering the amorphous nature of the present films, we selected a cellulase originated from Aspergillus niger for testing their biodegradability. For the cellophane (cellulose II), only cellulose chains on the surface are available for the attachment of the cellulase, since cellulose chains stack closely, and the film will be decomposed layer by layer. This process will greatly inhibit the hydrolysis of cellophane. The rapid increase in the beginning is attributed to the amorphous region in the surface of cellophane. With respect to CF11 6%, cellulase does not only function on the surface but also acts on internal chains because of their more open and accessible structure. Under similar conditions, CF11 6% will provide more active sites and chain ends for attack by cellulase. Eventually, CF11 6% shows a higher efficiency of enzymatic hydrolysis. Therefore, it is reasonable to conclude that CF11 6% will be decomposed much faster in the natural world and be friendlier to the environment than cellophane or other crystalline types of cellulose products. Moreover, cellulosic waste derived from ACF can be recycled and converted to liquid fuels,⁴⁹ due to its higher efficiency of enzymatic hydrolysis compared to the other cellulose resource, which will completely release the burden to the environment.

Fig. 8 Time profile of the enzymatic degradation of CF11 6% and cellophane.

Conclusions

Cellulose films with excellent transparency were regenerated from LiCl–DMAc solutions by using acetone as the regeneration solvent. The cellulose films were highly amorphous, which was confirmed by XRD, ¹³C-NMR, and FT-IR measurements. According to the best of our knowledge, this was the first time that one had prepared such amorphous cellulose films with good performance through a simple, less-destructive, and universal method. Compared with commercial cellophane, ACF possessed a comparable mechanical performance, but much faster enzymatic hydrolysis rate, due to its distinctive amorphous structure, which is more open and accessible, indicating its prevailing environmental friendliness. Based on the present results, we can conclude that the ACF possesses a great potential for replacing cellophane used in packaging materials. Moreover, it has importance to serve as a new standard sample for the study of cellulose structure and enzyme activity.

Acknowledgements

B.Z. would like to thank China Scholarship Council (CSC) for a scholarship support.

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Footnote

† Electronic supplementary information (ESI) available. See DOI: 10.1039/c4ra14090g

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