Optimizing cellulose microfibrillation with NaOH pretreatments for unbleached Eucalyptus pulp

Microfibrillated cellulose (MFC) is a form of biopolymer that stands out because of its versatile use. However, the process of obtaining MFC requires adjustments to provide the increase of microfibrillation in industrial scale. Thus, this study aimed to apply pretreatments with NaOH in unbleached Eucalyptus sp. fibers to optimize the microfibrilation process, in order to evaluate the effect of drying and not drying the fibers after pretreatments for reduction of energy consumption. Treatments of MFC with NaOH at 5% with 2 h of reaction and by 10 wt% were evaluated for the resulting morphology, chemical composition, and energy consumption. The length and width of the pulp fibers pretreated with NaOH decreased significantly, mainly with hasher pretreatments. Pretreating fibers with 5 wt% NaOH for 2 h increased the water retention value (WRV), in addition to presenting the lowest energy consumption for fibrillation, promoting energy savings of up to 48%. Pulps that were non-dried after the NaOH pretreatments resulted in easier microfibrillation and lower energy consumption when compared to dried pulp, which shows the negative impact of drying on the fibers to obtain MFC.


Introduction
Cellulose is a biopolymer that stands out for several reasons, such as its abundance, renewability, and nontoxicity to the environment and biodegradability after disposal. Through the processing of cellulose, it is possible to produce different materials in nanometric scale (Liu 2018). Nanostructured cellulose can be obtained from several sources, such as bacterial cellulose (Andriani et al. 2020), or from lignocellulosic materials spread into cellulose microfibrils or nanofibrils (CNFs) (Osong et al. 2016), cellulose nanocrystals (CNCs) (Pereira and Arantes 2020), and microfibrillated cellulose (MFC) (Guimarães Junior et al. 2018).
Microfibrillated cellulose (MFC) is a fibrillated material with fibrils with average diameter of less than 100 nm and lengths that can reach more than 10 lm (Zambrano et al. 2020). This material has high specific surface area due to the large amount of individual micro/nanofibrils, which increases bonding with each other or with other matrices, forming high-strength films (Guimarães Junior et al. 2018). Several industrial sectors and researchers are developing applications for MFC, such as those reported elsewhere for pharmaceutical (Salimi et al. 2019), food, paint and cosmetics (Fujisawa et al. 2017), films and coating (Matos et al. 2019), and fiber-cement composite (Fonseca et al. 2019(Fonseca et al. , 2021 fields, as well as bacterial nanocellulose for medical uses (Amorim et al. 2020).
MFC can be obtained by mechanical, chemical or enzymatic methods (Rol et al. 2019). The most common method for producing MFC involves a mechanical approach. However, the high energy consumption of the operation is the main limiting factor for scaling up this procedure. Therefore, pretreatments of cellulosic pulp are necessary to optimize the process. When submitting cellulose pulp to pretreatment, hydrogen bonds are weakened, which makes the process more efficient (Durães et al. 2020).
Previous studies have tested alkaline pretreatments of cellulosic pulp using sodium hydroxide (NaOH) at different concentrations to obtain MFC. Wang et al. (2014) stated that pretreatments with high alkaline concentrations impair the MFC acquisition process due to the conversion of cellulose I into cellulose II (mercerization). This process generates greater aggregation through the interdigitation of MFC in cell walls, which significantly hinders its individualization. Dias et al. (2019) studied alkaline pretreatments of bleached Eucalyptus sp. and showed that the use of 5% NaOH for 2 h was the most effective to obtain MFC and reduce energy consumption during the fibrillation process. Thus, alkaline pretreatments associated with mechanical methods are considered a viable alternative to reduce operational energy consumption.
However, if pulp fibers are dried, some cellulose chains can irreversibly bond, modifying their structure and properties, such as reducing the capacity to retain water. This process is usually called hornification. Regarding this process, Ballesteros et al. (2017) studied the water retention values of bleached and unbleached Eucalyptus and Pinus fibers after successive hornification cycles. When the fiber cell wall is submitted to the drying process, the cellulose microfibrils increase the degree of physical crosslinking between them, which decreases the pore size and the water retention capacity of the fibers. When the fibers retain less water in their structure, the cellulose microfibrils are less exposed due to the lower number of hydrogen bonds formed between them, making the fibrillation process more difficult (García-Iruela et al. 2019).
Studies related to the drying effect on fiber and the drying process have already been reported (Zimmermann et al. 2016;Silva et al. 2021). However, there is still a lack of information about the joint effect of pretreatment and microfibrillation process of unbleached Eucalyptus kraft pulp and how it could more easily produce MFC, with less energy consumption. This information could help optimize the fibrillation process and facilitate the scaling up of MFC production.
Given the above considerations, the present study aimed to investigate the drying effect on unbleached Eucalyptus cellulosic fibers after applying NaOH pretreatments to optimize the microfibrillation process, as well as assessing the efficiency of pretreatments to reduce the energy consumption for production of microfibrillated cellulose (MFC).

Alkaline pretreatment
The cellulosic pulp was oven-dried at 50°C for 24 h prior to the pretreatments. Then, the fibers were pretreated with 5 wt% NaOH solution for 2 h and 10 wt% NaOH for 1 h at 80°C while under continuous agitation at 800 rpm. Afterwards, the fibers were subjected to repeated sifting and washing with deionized water until neutral pH was reached according to the information reported by Aguado et al. (2019). The pH was monitored by pH indicator strips.
After the pulp was pretreated and thoroughly washed, it was divided into two samples: pulp ovendried at 50°C for 24 h and wet pulp (refrigerated at 5 ± 3°C until fibrillation). As a control, pulp without pretreatment was immersed in water until fibrillation. The pulps were subjected to continuous stirring (* 800 rpm) at 80°C for 1 h and 2 h. All treatments were coded to be easily assessed throughout the manuscript (Table 1).

Fiber characterization
The pulps were chemically characterized before and after the alkaline pretreatments. Lignin was evaluated according to Tappi UM 250 standard (1976) and soluble monosaccharides and disaccharides were quantified according to Wallis et al. (1996). An ion chromatography system (Dionex ICS 5000, Sunnyvale, CA, USA) was used. The average length, weighted average length, width and curvature were measured using a fiber image analyzer (Valmet FS5, Finland). The curvature index measures the degree of curvature of the fiber and is defined as follows: (contour fiber length/projected fiber length -1) 9 100 (Alimadadi et al. 2018).
The water retention value (WRV) of the pulp was calculated according to the Scandinavian test method SCAN-C 62:00 (2000), dispersing them in water with a fiber content of 0.5 wt% after boiling for 5 min. The water suspension was filtered by a Heraeus Megafuge 16R centrifuge (Thermo Fisher Scientific, Waltham, MA, USA), at 3000 9 g for 15 min to dehydrate, and the wet pulps were weighed immediately after. After oven-drying at 110°C for 5 h, the weight of the pulp was determined again. WRV was calculated based on Eq. (1): where W 0 and W 1 are the weights of wet and ovendried pulps, respectively.

Pulp mechanical fibrillation
The cellulose pulps with and without pretreatment were soaked in deionized water at a concentration of 2 wt% for 6 days to maximize the swelling of the fibers. After this procedure, the fibers were subjected to mechanical fibrillation using a Super MassColloider fibrillator (Masuko Sangyo MKCA6-2, Kawaguchi, Japan) with 5 passes at 1500 rpm (Tonoli et al. 2016;Lago et al. 2020).

Light microscopy (LM) of pulp fibers
A Nikon Eclipse E20 light microscope was used to analyze the fibrillation levels of the pretreated fibers and without pretreatments after 5 passes through the equipment. The samples were diluted in deionized water (0.75 wt%) and stained with ethanol-safranin (1% v/v) to enhance the contrast of the images.

Transmission electron microscopy (TEM) of MFC
The morphology of the MFC was analyzed by a Zeiss EM 109 microscope with an accelerated voltage of 80 kV. The sample preparation and equipment settings followed the information reported by Tonoli et al. (2016). Uranyl acetate was added during sample preparation to improve the contrast of the samples. A few drops of the suspension with the dye were added to copper grids (400 mesh) with formvar film (thermoplastic resin) and dried before visualization by TEM.

Energy consumption (EC) during fibrillation
The EC to fibrillate the pulps was calculated as suggested by Xu et al. (2020), considering the average amperage used for each cycle (dispersion passage through the fibrillator), the equipment tension and the fibrillation time per ton of microfibrillated material, with a consistency of 2.0 wt% (Eq. 2). The EC was calculated until the material exhibited a gel-like structure in aqueous suspension, corresponding to the successful fibrillation of the pulps.
where EC is the energy consumption (kW h ton -1 ); P is the power (in kW, voltage 9 electric current); h is the time (in hours) spent during fibrillation; and m is the mass (in tons) of the cellulose material processed in the fibrillator.

Results and discussion
Chemical characterization of the pulps  note that the chemical characterization of dried and non-dried pretreated pulps was the same because alkaline pretreatments of the cellulosic pulps were the same in both cases. It is clear that all chemical pretreatments performed resulted in a relative increase of the glucose content as a probable consequence of the decreased content of hemicellulose, mainly xylose (Castro et al. 2020). The higher were the NaOH concentration and pretreatment time of the fibers, the greater the relative amount of cellulose and lower the xylose content (Bufalino et al. 2015). 8% increase in the relative glucose content was found for the E10 samples. With harsher pretreatment, the relative xylose content decreased. After E10 pretreatment, the initial xylose content of 12% decreased to around 5%. This effect happens because xylose is more present on outer layers than in the corresponding internal layers (Dahlman et al. 2003), and the pretreatment reaches the fibers more superficially.
As reported by Dias et al. (2019), the mechanical fibrillation process is more efficient in pre-treatments that preserve part of the hemicellulose, since this component prevents direct contact between the microfibrils, which allows the internal structures of the fibers to be easily exposed (Albornoz-Palma et al. 2020). The results found suggest that the relative levels of glucose in the E5 samples are more suitable, since in these pretreatments the fibers are exposed to lower concentrations of NaOH, and therefore, greater amounts of hemicellulose are preserved in the fiber cell wall.

Morphology of fibers and MFC
Water retention values (WRVs) of fibers Table 3 shows the morphological properties of cellulosic fibers before and after chemical pretreatments. It is possible to observe that there was a small decrease in the weighted average width of the fibers.
Regarding the fiber length, a small reduction was observed in all oven-dried pretreated fibers, especially those that were subjected to more intense pretreatments with NaOH. Decreased fiber length may be related to the increase in the curvature of the fibers, which may have influenced the measurement values. Stronger alkaline hydrolysis removes more hemicellulose from the fiber cell wall, a fact that directly interferes with the internal strength of the fibers, which makes them more flexible and susceptible to increase its curvature and, consequently, reduces their longitudinal size (Martin-Sampedro et al. 2014). A greater number of curved fibers can directly interfere with the microfibrilation process due to reduced contact area between the fibers and the grinder stone discs as well as increased accumulation of fibers trapped inside the grinder (Dias et al. 2019).
The WRVs of dried and non-dried fibers are presented in Table 3. The WRV is the most commonly used index to assess the swelling of cellulose fibers (Olejnik et al. 2017). The WRVs found for E5-D and E5-ND indicate that there was no reduction of water retention of the fibers. According to Klemm et al. (2006), the alkaline environment contributes significantly to fiber swelling due to osmotic effects and expansion pressure that disrupts hydrogen bonds formed between fibrils in the fiber cell wall, which facilitates the water absorption.
Regarding the E10 fibers, the WRVs found were below those of the E5 fibers, for both dried and nondried. These results corroborate the findings of Dias et al. (2019), where the higher concentration of NaOH promoted an increase in the removal of fiber cell wall components, mainly hemicellulose, which have a large number of hygroscopic sites capable of forming hydrogen bonds (Pacaphol and Aht-Ong 2017). The presence of functional groups loaded in the fiber cell wall as uronic acids, for example, increase WRV due to osmotic pressure. The reduction of the content of changed groups of pulps extracted with 5% alkali is offset by the swelling effect generated by the alkali, resulting in the WRV increase. In the other hand, for pulps treated with 10% alkali, the uronic acid removal is high and the WRV decreases as consequence (Lund et al. 2012). Compared to the non-dried fibers, the dried samples had decreased WRVs. According to Ballesteros et al. (2017), the drying and subsequent wetting of the fibers tends to reduce the WRV due to the collapse and stiffening of the fiber surface generated by the hornification process. In addition, the number of pores on the fiber surface decreases due to the greater crosslinking of cellulose fibrils, which further reduces the WRVs (Mo et al. 2019). Studies also point out that the reduction of hemicellulose and/or charged groups in the interfibrillar spaces reduces the tendency of irreversible bonds between adjacent cellulose fibrils (Santmarti et al. 2020). When fibers are rehydrated, the hemicellulose contribute to some extent to increase the amount of water absorbed by the fiber cell wall. However, the remaining amount of charged groups is the dominant factor in the fiber swelling behavior (Lund et al. 2012). Therefore, pretreatments that removed large amount of hemicelluloses and charged groups (E10) led to, higher fiber hornification and the WRV tends to be lower. Figure 1 presents the curvature of dried and non-dried fibers. Figures 1a, 1c, and 1e show the oven-dried fibers after the chemical pretreatments, while those in Figs. 1b,1d,and 1f show the fibers that were nondried after the chemical pretreatments.

Light microscopy of pulp fibers
Fibers that were subjected to chemical pretreatments presented a considerable increase in curvature when compared to untreated fibers. Pulps that were subjected to pretreatments in higher alkaline concentrations (10% NaOH) had even greater curvature, with curvature values of 30.4 ± 0.2% for E10-D and 29.7 ± 0.3% for E10-ND (Table 3).
The increase of curvature mainly resulted from the greater removal of hemicellulose and lignin from the fiber cell wall, and with higher concentrations of NaOH, the curvature value of the fibers increased (Tarrés et al. 2017). Fibers that were oven-dried after the pretreatments had higher curvature than the nondried fibers. The hornification of the fibers is responsible for the closer proximity of the cellulosic chains (Luo et al. 2018), which contributes to the shrinkage and bending of the fibers. Figure 2 shows the MFC morphology observed by TEM. The MFC produced from the E0 fibers (Fig. 2a,  b) was less individualized than that from the fibers subjected to pretreatments with NaOH. The E5 fibers were easily converted to MFC, presenting average diameter of 26 ± 14 nm for E5-D (Fig. 2c) and 29 ± 15 nm for E5-ND (Fig. 2d). Due to the intense removal of hemicellulose from the cell wall of the fibers, the E10 fibers presented less individualized MFC, regions with aggregation and fibers that were not fibrillated, which is not desired in this process. This fact indicates the importance of hemicellulose during the fibrillation process (Durães et al. 2020). According to Chaker et al. (2013), hemicellulose hinders the formation of irreversible hydrogen bonds between microfibrils, acting as a physical barrier that prevents direct contact and further aggregation (Fig. 3). Figure 4 shows the MFC distribution according to the diameter classes for pulps with and without pretreatment, as well as dried and non-dried pulps. Most of the MFC obtained showed diameters that ranged from 15 to 30 nm. The MFC content for nondried fibers with average diameters below 45 nm was approximately 74% for E0-ND, 86% for E5-ND and 82% for E10-ND. However, fibers that were dried presented larger particles, with average diameters below 45 nm representing 72% for E0-D, 84% for E5-D and 78% for E10-D. Non-dried fibers presented smaller MFC diameters than dried fibers. The hornification of the fibers during oven-drying implies the close proximity of the cellulose chains and the reduced number of pores in the fiber microstructure, which decreases the amount of individualized microfibrils in the MFC suspension (Duan et al. 2015). Energy consumption for obtaining MFC Figure 5 shows the evolution of energy consumption during fibrillation with the increase of the number of passes through the mechanical fibrillator.

Transmission electron microscopy (TEM) of MFC
All pretreated samples reached a gelatinous appearance on the 3rd pass through the fibrillator, exception for E10-ND pulp, that was able to achieve it on the 2nd pass. This gel-like appearance of the MFC suspension is due to its increased viscosity, which is caused by micro/nanofibrils ability to retain water in its internal and external structures (Ioelovich 2008). With more exposed and individualized micro/nanofibrils, the surface area increases, allowing the suspension to retain significantly more water than the starting pulp, leading to a shear resistance behavior in MFC suspension, modifying its viscosity and hence its appearance became gelatinous.
The E10 pretreatments, both dried and non-dried, were not as effective in the fibrillation process, therefore, (despite the E10-D pretreatment showed a great reduction in energy consumption) the efficiency in the microfibrils individualization process was not satisfactory. As previously noted in Dias et al. (2019), preserved structure and still with large dimensions after chemical pretreatments and mechanical fibrillation; arrow 2: Well individualized microfibrils after chemical pretreatments and mechanical fibrillation this harsher pretreatment removes most of the hemicellulose present in the pulp fibers. The presence of these polysaccharides in the pulp contributes to increased fibrillation since the carboxylic groups of the hemicellulose contribute to electrostatic repulsion between MFC chains in water (Iwamoto et al. 2008).
E5-D pretreatment was proved to be efficient in reducing energy consumption, promoting savings of up to 22% compared to E0-D fibers (Table 4). However, E5-ND fibers demonstrated greater efficiency in the process, reaching savings of up to 48% compared to E0-ND fibers. The procedures conducted prior to fibrillation that preserve most of the Fig. 3 Description of the importance of hemicellulose during fibrillation and the hindrance caused by the hornification process. a Depicts the unbleached fibers and their deconstruction into MFC; b is a representation of the microstructure and composition of the MFC into the pulper fiber; c shows the individualization process happening more easily when hemicellulose is present. The proximity of cellulose bundles without hemicellulose led to a stiffer structure, incurring in cutting of fibrils instead of individualization; d depicts the hornification process and subsequent structural change happening after MFC drying hemicellulose in the cell walls of the fibers are more efficient in the fibrillation process (Berglund et al. 2016).
A superior performance was achieved in reducing the energy consumption for obtaining MFC from the fibers that were non-dried after pretreatment, apart from fibers E10-D and E10-ND. The fiber swelling of non-dried samples allowed a greater reduction in energy consumption than that of oven-dried fibers (Ireana Yusra et al. 2018). Thus, non-dried fibers with mild alkaline pretreatment E5-ND have been proven to be the ideal path to produce MFC with a relatively low energy consumption.

Conclusions
This study sought to evaluate the effect of chemical pretreatments to optimize the process of obtaining MFC. In addition, it was evaluated the effect of drying and not drying the pretreated unbleached Eucalyptus fibers before grinding to obtain MFC.
E5-ND fibers were more easily fibrillated with reduced energy consumption, being influenced mainly by the hornification and swelling processes of the fibers. Morphological analyses have also shown that the E5-ND samples were more effective in producing MFC. E10-D and E10-ND pretreatment produced intense removal of hemicellulose and charged groups from the fiber cell wall, which are responsible for part of the water retention in the structure, reducing fiber swelling, which compromised fibrillation.
The E5-ND fibers presented a very significant reduction in energy consumption, reaching approximately 48% savings. Even though the E10-D fibers showed a 53% reduction in energy consumption, the pretreatment was not effective in obtaining MFC. Thus, removing part of the hemicellulose and charged groups seems to facilitate the fibrillation process, while intense removal of these components impairs the obtention of MFC.
This study contributes to improve the methodology for obtaining MFC. It was shown that oven-drying fibers after pretreatment is detrimental to lowering the energy consumption and improving the MFC obtaining process, which is desired when scaling up the production.