*3.1. Morphological Characterization of Raw OPEFB Fiber, Treated cellulose, Microcrystalline Cellulose and Nano-Bentonite*

SEM analysis was performed to have a better understanding of the microstructural changes of the OPEFB caused by the pre-treatment and acid hydrolysis process. Figure 1a shows the surface morphology of the raw OPEFB fiber, while Figure 1b presents the structure of the OPEFB cellulose (after it underwent alkaline pre-treatment and the bleaching process). Apparently, there are changes in the OPEFB morphology detected through the comparison of both micrographs indicating the success of the chemical pre-treatment procedure in removing non-cellulosic materials such as hemicellulose, lignin and waxy substances from the raw OPEFB fiber.

The raw OPEFB fiber in Figure 1a possesses an irregular shape and the surface of the microfibrils cellulose (pointed by the blue arrow) was covered up by non-cellulosic materials which act as the protective surface (as marked by red circles). In contrast, Figure 1b shows the smooth, clear and individualized rod-like microfibrils surface of cellulose. The different morphology of the surface structure in the raw OPEFB (Figure 1a) and the treated OPEFB cellulose (Figure 1b) indicates that chemical pretreatment has successfully removed the non-cellulosic materials such as hemicellulose, lignin and waxy substances from the raw OPEFB fiber, which is in line with the FTIR results. Due to these removals, the microfibrils surface of the cellulose has been exposed and the crystallinity region of the cellulose was increased, as confirmed through the XRD analysis. Figure 1c represents the morphology of the microcrystalline cellulose after being dried up and observed under SEM with ×30k magnification. It can be observed that the structure of the OPEFB cellulose's microfibril was completely destroyed and the size of the cellulose was significantly being reduced. However, the SEM diagram suggests that the microcrystalline cellulose tends to aggregate together into large particles of cellulose. Agglomeration of cellulose usually occurs after being dried from the suspension form [15].

**Figure 1.** Scanning electron microscope (SEM) micrographs of: raw oil palm empty fruit bunch (OPEFB) fiber taken at ×5000 (**a**); treated OPEFB cellulose taken at ×5000 (**b**); microcrystalline cellulose taken at ×30k (**c**).

Figure 2a,b display the SEM images of the pristine nano-bentonite and ultra-sonicated bentonite, respectively. By comparing both figures, we can prove that the particle size of the nano-bentonite had been reduced significantly upon the ultra-sonication process. Before ultra-sonication, the nano-bentonite appears in large particles with an irregular shape and size. This was due to the high stacking of platelets forming large tactoids of the phyllosilicate structure. After being ultra-sonicated, the particle size of the nano-bentonite was seen to greatly reduce, with rougher surface morphology. This observation was due to the tactoid size reduction of the nano-clay as a result of the delamination of the nano-platelets. However, in agglomerated form, it is difficult to get a clear image of the individual platelet of the nano-bentonite and determine its particle size. However, the results can confirm that the ultra-sonication process has successfully reduced the tactoid size of the nano-bentonite. *Polymers* **2021**, *13*, x FOR PEER REVIEW 2 of 22

> FTIR spectroscopy is a non-destructive method to study the chemical composition of material by analyzing its functional groups. The FTIR spectra of the raw OPEFB fiber, treated OPEFB cellulose and OPEFB microcrystalline cellulose are illustrated in Figure 3. Generally, the FTIR spectra of all samples were almost the same, indicating that they pos-

> The FTIR spectra of the raw OPEFB and treated OPEFB cellulose represent a broad peak at the region of 3353 and 2891 cm−1 due to the –OH stretching of the aliphatic structure and aliphatic saturated C-H stretching of CH2 and CH3. The 3353 cm−1 can also be attributed to moisture content where the hydroxyl group is found in cellulose, hemicellulose and lignin. This shows that the cellulose is hydrophilic in nature [16]. The peak at 1035 cm−1 appears in both samples and represents the characteristics of anhydroglucose chains with the C-O-C stretch [17]. The treated cellulose has a sharper peak than the raw OPEFB due to its higher surface area caused by the removal of non-cellulosic components in the cellulose. Meanwhile, the peaks at 1634 and 2891 cm−1 represent the –OH bending vibration of absorbed water from the strong interaction between cellulose and water [18]. A peak at 896 cm−1 is related to β-glycosidic linkage between the glucose rings of cellulose. [19] This indicates that the cellulose is made up of a linkage bond of saccharides that formed polysaccharides. Meanwhile, the peak that appears at 1330–1369 cm−1 represents the bending vibration of the C-H and C-O groups of aromatic rings in polysaccharides [20].

> The disappearance of a peak at 1723 cm−1 in the spectrum of the treated cellulose and microcrystalline cellulose samples verifies the removal of hemicellulose and lignin. This is because the presence of lignin and hemicellulose can be detected through the appearance of those peaks due to the functional group of carbonyl ester and the C=O acetyl group of the uronic ester, respectively [4,8,21]. In addition, the peak at 1228 cm−1, which represents the syringyl ring unit and C-O stretching at lignin and xylan, disappeared in the spectra of the treated cellulose, suggesting the removal of lignin and hemicellulose after various treatments [15,22]. These clearly show that most non-cellulosic materials have been removed upon the alkaline treatment, bleaching and acid hydrolysis processes.

> Figure 4 shows the FTIR spectra of the nano-bentonite (before and after ultra-sonication). By comparing the results of both samples, no significant change in the pattern of the FTIR spectra can be observed. This indicates that the ultra-sonication process does not alter the chemical structure of the nano-bentonite. Both samples exhibit peaks at 3614 and

*Nano-Bentonite* 

sess the same chemical composition and structure.

**Figure 2.** SEM picture of: nano-bentonite (before ultrasonication) (**a**); nano-bentonite (after ultrasonication) (**b**), taken at ×10 k. **Figure 2.** SEM picture of: nano-bentonite (before ultrasonication) (**a**); nano-bentonite (after ultrasonication) (**b**), taken at ×10k.

#### *3.2. Chemical Structure of Raw OPEFB Fiber, Treated Cellulose, Microcrystalline Cellulose and Nano-Bentonite* water in the bentonite di-octahedral surface [23]. The sharp and intense peak near 1000 cm−<sup>1</sup> represents the Si-O-Si stretching in the nano-bentonite [24]. The peaks at 790 and 997 cm−<sup>1</sup> are the characteristic signals of Al-O-Si and Si-O stretching vibrations, respectively [24].

3435 cm-1, which indicate the possibility of the hydroxyl linkage from alumino-silicate layers [23]. The peak at 3614 cm−1 represents the interlayer hydrogen bonding, corresponding to the stretching vibration of the OH coordinated octahedral layer of Al+Mg [18]. The bands at 1636 and 3435 cm−1 indicate the –OH stretching vibration due to the absorption of

FTIR spectroscopy is a non-destructive method to study the chemical composition of material by analyzing its functional groups. The FTIR spectra of the raw OPEFB fiber, treated OPEFB cellulose and OPEFB microcrystalline cellulose are illustrated in Figure 3. Generally, the FTIR spectra of all samples were almost the same, indicating that they possess the same chemical composition and structure. Meanwhile, the peaks at 3614 cm−1, 3435 cm−1, 1636 cm−1, 997 cm−1 and 790 cm−1 imply the presence of montmorillonite (MMT) in the bentonite [23]. The ultra-sonicated nano-bentonite has a more intense peak related to Si-O stretching around 790 cm−1, indicating that the ultra-sonication treatment has increased the surface area of the nano-filler by delamination or the stacking platelets, which is also proved through the SEM and XRD analyses.

*Polymers* **2021**, *13*, x FOR PEER REVIEW 2 of 22

**Figure 3.** Fourier transform infrared spectroscopy (FTIR) spectra of raw OPEFB fiber, treated OPEFB cellulose fiber and OPEFB microcrystalline cellulose. **Figure 3.** Fourier transform infrared spectroscopy (FTIR) spectra of raw OPEFB fiber, treated OPEFB cellulose fiber and OPEFB microcrystalline cellulose.

The FTIR spectra of the raw OPEFB and treated OPEFB cellulose represent a broad peak at the region of 3353 and 2891 cm−<sup>1</sup> due to the –OH stretching of the aliphatic structure and aliphatic saturated C-H stretching of CH<sup>2</sup> and CH3. The 3353 cm−<sup>1</sup> can also be attributed to moisture content where the hydroxyl group is found in cellulose, hemicellulose and lignin. This shows that the cellulose is hydrophilic in nature [16]. The peak at 1035 cm−<sup>1</sup> appears in both samples and represents the characteristics of anhydroglucose chains with the C-O-C stretch [17]. The treated cellulose has a sharper peak than the raw OPEFB due to its higher surface area caused by the removal of non-cellulosic components in the cellulose. Meanwhile, the peaks at 1634 and 2891 cm−<sup>1</sup> represent the –OH bending vibration of absorbed water from the strong interaction between cellulose and water [18]. A peak at 896 cm−<sup>1</sup> is related to β-glycosidic linkage between the glucose rings of cellulose. [19] This indicates that the cellulose is made up of a linkage bond of saccharides that formed polysaccharides. Meanwhile, the peak that appears at 1330– 1369 cm−<sup>1</sup> represents the bending vibration of the C-H and C-O groups of aromatic rings in polysaccharides [20].

**Figure 4.** FTIR spectra of nano-bentonite (before and after ultra-sonication). The disappearance of a peak at 1723 cm−<sup>1</sup> in the spectrum of the treated cellulose and microcrystalline cellulose samples verifies the removal of hemicellulose and lignin. This is because the presence of lignin and hemicellulose can be detected through the appearance of those peaks due to the functional group of carbonyl ester and the C=O acetyl group of the uronic ester, respectively [4,8,21]. In addition, the peak at 1228 cm−<sup>1</sup> , which represents the syringyl ring unit and C-O stretching at lignin and xylan, disappeared in the spectra of the treated cellulose, suggesting the removal of lignin and hemicellulose after various treatments [15,22]. These clearly show that most non-cellulosic materials have been removed upon the alkaline treatment, bleaching and acid hydrolysis processes.

Figure 4 shows the FTIR spectra of the nano-bentonite (before and after ultra-sonication). By comparing the results of both samples, no significant change in the pattern of the FTIR

spectra can be observed. This indicates that the ultra-sonication process does not alter the chemical structure of the nano-bentonite. Both samples exhibit peaks at 3614 and 3435 cm−<sup>1</sup> , which indicate the possibility of the hydroxyl linkage from alumino-silicate layers [23]. The peak at 3614 cm−<sup>1</sup> represents the interlayer hydrogen bonding, corresponding to the stretching vibration of the OH coordinated octahedral layer of Al+Mg [18]. The bands at 1636 and 3435 cm−<sup>1</sup> indicate the –OH stretching vibration due to the absorption of water in the bentonite di-octahedral surface [23]. The sharp and intense peak near 1000 cm−<sup>1</sup> represents the Si-O-Si stretching in the nano-bentonite [24]. The peaks at 790 and 997 cm−<sup>1</sup> are the characteristic signals of Al-O-Si and Si-O stretching vibrations, respectively [24]. Meanwhile, the peaks at 3614 cm−<sup>1</sup> , 3435 cm−<sup>1</sup> , 1636 cm−<sup>1</sup> , 997 cm−<sup>1</sup> and 790 cm−<sup>1</sup> imply the presence of montmorillonite (MMT) in the bentonite [23]. The ultra-sonicated nano-bentonite has a more intense peak related to Si-O stretching around 790 cm−<sup>1</sup> , indicating that the ultra-sonication treatment has increased the surface area of the nano-filler by delamination or the stacking platelets, which is also proved through the SEM and XRD analyses. **Figure 3.** Fourier transform infrared spectroscopy (FTIR) spectra of raw OPEFB fiber, treated OPEFB cellulose fiber and OPEFB microcrystalline cellulose.

3435 cm-1, which indicate the possibility of the hydroxyl linkage from alumino-silicate layers [23]. The peak at 3614 cm−1 represents the interlayer hydrogen bonding, corresponding to the stretching vibration of the OH coordinated octahedral layer of Al+Mg [18]. The bands at 1636 and 3435 cm−1 indicate the –OH stretching vibration due to the absorption of water in the bentonite di-octahedral surface [23]. The sharp and intense peak near 1000 cm−<sup>1</sup> represents the Si-O-Si stretching in the nano-bentonite [24]. The peaks at 790 and 997 cm−<sup>1</sup> are the characteristic signals of Al-O-Si and Si-O stretching vibrations, respectively [24]. Meanwhile, the peaks at 3614 cm−1, 3435 cm−1, 1636 cm−1, 997 cm−1 and 790 cm−1 imply the presence of montmorillonite (MMT) in the bentonite [23]. The ultra-sonicated nano-bentonite has a more intense peak related to Si-O stretching around 790 cm−1, indicating that the ultra-sonication treatment has increased the surface area of the nano-filler by delamination or the stacking platelets, which is also proved through the SEM and XRD analyses.

*Polymers* **2021**, *13*, x FOR PEER REVIEW 2 of 22

**Figure 4. Figure 4.** FTIR spectra of nano-bentonite (b FTIR spectra of nano-bentonite (before and after ultra-sonication). efore and after ultra-sonication).
