Structural Characterization of Nanocellulose/Fe3O4 Hybrid Nanomaterials

The rise of innovation in the electrical industry is driven by the controlled design of new materials. The hybrid materials based on magnetite/nanocellulose are highly interesting due to their various applications in medicine, ecology, catalysis and electronics. In this study, the structure and morphology of nanocellulose/magnetite hybrid nanomaterials were investigated. The effect of nanocellulose loading on the crystal structure of synthesized composites was investigated by XRD and FTIR methods. The presented study reveals that the interaction between the cellulose and magnetic nanoparticles depends on the nanocellulose content. Further, a transition from cellulose II to cellulose I allomorph is observed. SEM and EDS are employed to determine the variation in morphology with changes in component concentrations. By the calculation of magnetic interactions between adjacent Fe3+ and Fe2+ ions within composites, it is determined that ferromagnetic coupling predominates.


Introduction
Development of hybrid organic/inorganic composites attracts great attention in the scientific community. Hybrid multifunctional materials have advantages, due to the combination of different properties of organic-inorganic ingredients, where an organic component, such as nanocellulose, contributes with high flexibility, dielectric and piezoelectric properties, while an inorganic component may contribute with magnetic and electric properties [1]. Several studies emphasized the importance of achieving improved material properties through the prevention of the mutual inhibition of different origin fillers and the promotion of compatibility at the interface matrix/added components. The resulting hybrid material has demonstrated diverse properties, which can be tailored by changing components, loadings, morphology, and the arrangement of an individual component, etc. [2,3]. As a result, the materials with a broad range of applications, such as drug delivery, magnetic resonance imaging (MRI), sensors, magnetic storage media, photocatalysts and electromagnetic absorption materials, etc., can be developed [4][5][6][7][8][9].

Materials
Commercially available cellulose was used for nanocellulose preparation. Components used for co-precipitation, FeSO 4 × 7H 2 O and FeCl 3 × 6H 2 O, were purchased from Merck (Darmstadt, Germany). Deionized water (DW, Sigma Aldrich, Saint Louis, MO, USA), with a resistivity of 18 MΩ cm, was used for NC washing, aqueous solution preparation and for dish washing.

Preparation of NC/Fe 3 O 4
Nanocellulose (NC) was prepared by the acid hydrolysis (H 2 SO 4 ) of commercially available cellulose, according to the established procedure [30]. Obtained NC was separated from the liquid phase by centrifugation and washed with deionized water until pH reached the value of 5.5. The functionalized nanocellulose was prepared by the co-precipitation of Fe(II) and Fe(III) ions in an aqueous solution containing NC with ammonia. NC, 1.5 g, was added to 200 mL of distilled water and stirred for 10 min. FeCl 3 × 6H 2 O and SO 4 × 7H 2 O were added to a mixture as sources of ferro and ferri ions, maintaining the ratio 2:1 and heated to 60 • C. Chemical precipitation was achieved by adding 8.0 M of the ammonia solution dropwise with vigorous stirring and at a constant pH of 10. Precipitation was followed with changes of orange color suspension to a black precipitate. Different concentrations of NC were achieved by reducing or increasing the amount of NC in aqueous solutions, while maintaining the same concentrations of Fe(II) and Fe(III) ions. After incubation for 4 h at 60 • C, the mixture was cooled to room temperature with stirring.
Fourier-transform infrared (FTIR) spectra of NC/Fe 3 O 4 samples were recorded in the transmission mode, within the region 400 to 4000 cm −1 , using a BOMEM (Hartmann and Braun, Quebec City, QC, Canada) spectrometer with a resolution of 4 cm −1 .
The dispersion state of Fe 3 O 4 on the nanocellulose surface and morphology of final composite films has been investigated by scanning electron microscopy (SEM, Tokyo, Japan, model: JEOL JSM 6610LV).
For the theoretical calculation of magnetic interactions and magnetic couplings between iron atoms in the magnetite structure within investigated composite materials, we performed the Density Functional Theory-Broken Symmetry [31][32][33][34] using B3LYP [35,36] and OPBE [37] functionals and a TZP basis set.

Results and Discussion
Changes in the crystal structure and phase purity, due to the variation of NC loadings, are examined by the diffraction of X-rays. The XRD diffractogram of the pure NC and the investigated Fe 3 O 4 /NC composite samples are shown in Figure 1. The diffractogram of the pure NC shows a typical cellulose II structure, with a low intensity peak at 12 • and strong doublet of 20 and 22 • [38,39]. A similar behavior was obtained by Lani et al. in [40], after the acid hydrolysis of cellulose was isolated from a fruit bunch. With the precipitation of Fe 3 O 4 in alkali, the conversion of cellulose II into cellulose I occurs. The characteristic signals at 30  Fourier-transform infrared (FT-IR) spectroscopy in the transmission mode was employed to investigate the variation in the infrared optical properties of NC with the precipitation of Fe3O4 on the nanocellulose surface. The FT-IR transmittance spectra of the NC and composite samples are shown in the Figure 2. The FTIR spectrum of pure nanocellulose was explained in detail in our previous publication [42]. In the spectra of magnetite/NC composites (Figure 2), the bands originating from nanocellulose are the most dominant bands. Characteristic bands, assigned to the O-H stretching vibration at 3430 cm −1 , C-H stretching vibrations of the -CH2 group at 2900 cm −1 and -OH bending vibration at 1640 cm −1 , are observed in the spectra of all samples. A high intensity broad band, at around 563 cm −1 , is a characteristic band of Fe-O in the tetrahedral sites [43]. Normally, this band appears at around 600 cm −1 or a little below this value, as suggested by the literature [44]. A shift of this band to a lower wave number is due to the occurrence of Fe-O bonds on the surface of the nanocellulose. The decrease in the intensity of this signal is caused by the lowering of the Fe3O4 concentration, from the sample Fe3O4/NC1 to Fe3O4/NC8, respectively. The shift of the O-H peak after the deposition of Fe3O4 on the NC surface suggests the existence of an interaction between the cellulose and magnetic nanoparticles [45]. Fourier-transform infrared (FT-IR) spectroscopy in the transmission mode was employed to investigate the variation in the infrared optical properties of NC with the precipitation of Fe 3 O 4 on the nanocellulose surface. The FT-IR transmittance spectra of the NC and composite samples are shown in the Figure 2. The FTIR spectrum of pure nanocellulose was explained in detail in our previous publication [42]. In the spectra of magnetite/NC composites ( Figure 2), the bands originating from nanocellulose are the most dominant bands. Characteristic bands, assigned to the O-H stretching vibration at 3430 cm −1 , C-H stretching vibrations of the -CH 2 group at 2900 cm −1 and -OH bending vibration at 1640 cm −1 , are observed in the spectra of all samples. A high intensity broad band, at around 563 cm −1 , is a characteristic band of Fe-O in the tetrahedral sites [43]. Normally, this band appears at around 600 cm −1 or a little below this value, as suggested by the literature [44]. A shift of this band to a lower wave number is due to the occurrence of Fe-O bonds on the surface of the nanocellulose. The decrease in the intensity of this signal is caused by the lowering of the Fe 3 O 4 concentration, from the sample Fe 3 O 4 /NC1 to Fe 3 O 4 /NC8, respectively. The shift of the O-H peak after the deposition of Fe 3 O 4 on the NC surface suggests the existence of an interaction between the cellulose and magnetic nanoparticles [45]. The region between 3600 and 2800 cm −1 was additionally investigated and depicted in Figure 3a. The maximum absorbance of the OH stretching vibration is positioned at 3440 cm −1 , indicating intramolecular hydrogen bonding of O2-H---O6 for CN sample. In the cases of the modified samples, the O3-H---O5 and O6-H---O3 intramolecular hydrogen bonding was detected at 3338 cm −1 and 3270 cm −1 , respectively [46]. It was observed that the sample marked as NC exhibits a typical cellulose II structure with a low intensity band at 3480 cm −1 and a strong band at 3440 cm −1 . These bands are characteristic of the cellulose II allomorph, and as it can be noticed from Figure 3a, these bands were not detected in FTIR spectra of the samples modified with Fe3O4 nanoparticles. In the spectra of the hybrid materials (Fe3O4/NC1, Fe3O4/NC2, Fe3O4/NC4 and Fe3O4/NC8), bands at 3338 and 3270 cm −1 were observed, indicating a transition from cellulose II to cellulose I allomorph, which is in agreement with the results obtained by XRD analysis. Further, the band at 1922 cm −1 was significantly shifted to a higher wavenumber after the addition of magnetic particles into the structure, suggesting an exchange in the arrangements caused by the changes of angles around β-1,4-D-glycosidic linkages rearrangement [47].  The region between 3600 and 2800 cm −1 was additionally investigated and depicted in Figure 3a. The maximum absorbance of the OH stretching vibration is positioned at 3440 cm −1 , indicating intramolecular hydrogen bonding of O2-H-O6 for CN sample. In the cases of the modified samples, the O3-H-O5 and O6-H-O3 intramolecular hydrogen bonding was detected at 3338 cm −1 and 3270 cm −1 , respectively [46]. It was observed that the sample marked as NC exhibits a typical cellulose II structure with a low intensity band at 3480 cm −1 and a strong band at 3440 cm −1 . These bands are characteristic of the cellulose II allomorph, and as it can be noticed from Figure 3a, these bands were not detected in FTIR spectra of the samples modified with Fe 3 O 4 nanoparticles. In the spectra of the hybrid materials (Fe 3 O 4 /NC1, Fe 3 O 4 /NC2, Fe 3 O 4 /NC4 and Fe 3 O 4 /NC8), bands at 3338 and 3270 cm −1 were observed, indicating a transition from cellulose II to cellulose I allomorph, which is in agreement with the results obtained by XRD analysis. Further, the band at 1922 cm −1 was significantly shifted to a higher wavenumber after the addition of magnetic particles into the structure, suggesting an exchange in the arrangements caused by the changes of angles around β-1,4-D-glycosidic linkages rearrangement [47]. The region between 3600 and 2800 cm −1 was additionally investigated and depicted in Figure 3a. The maximum absorbance of the OH stretching vibration is positioned at 3440 cm −1 , indicating intramolecular hydrogen bonding of O2-H---O6 for CN sample. In the cases of the modified samples, the O3-H---O5 and O6-H---O3 intramolecular hydrogen bonding was detected at 3338 cm −1 and 3270 cm −1 , respectively [46]. It was observed that the sample marked as NC exhibits a typical cellulose II structure with a low intensity band at 3480 cm −1 and a strong band at 3440 cm −1 . These bands are characteristic of the cellulose II allomorph, and as it can be noticed from Figure 3a, these bands were not detected in FTIR spectra of the samples modified with Fe3O4 nanoparticles. In the spectra of the hybrid materials (Fe3O4/NC1, Fe3O4/NC2, Fe3O4/NC4 and Fe3O4/NC8), bands at 3338 and 3270 cm −1 were observed, indicating a transition from cellulose II to cellulose I allomorph, which is in agreement with the results obtained by XRD analysis. Further, the band at 1922 cm −1 was significantly shifted to a higher wavenumber after the addition of magnetic particles into the structure, suggesting an exchange in the arrangements caused by the changes of angles around β-1,4-D-glycosidic linkages rearrangement [47].  In the spectra shown in Figure 3b, there are few obvious differences between the NC and modified NC. First of all, the bands at 1720 cm −1 and 1510 cm −1 , which are present in the spectrum of pure nanocellulose, disappear in the spectra of modified hybrid material. According to the literature, these bands originate from the C=C stretching vibration of the aromatic ring and the C=O stretching vibration of carboxylic groups of the lignin and hemicellulose [48]. It was demonstrated earlier that these bands can despair after the treatment of cellulose with NaOH [49]. We have also used an alkali environment during the modification of nanocellulose with magnetic particles, and we consider that as a reason for the disappearance of the earlier mentioned bands.
The shift to lower wavenumbers and broadening of the FTIR bands usually appears due to a presence of hydrogen bonds between cellulose and filler particles [50]. It was noticed that after the introduction of Fe 3 O 4 particles, bands of hydroxyl and carboxyl groups were downshifted, indicating a formation of the hydrogen bonds among NC and the Fe 3 O 4 . A shift of the bands at 1640 cm −1 and 1420 cm −1 was detected in the spectra presented in Figure 3b. With an increasing amount of NC, the shift becomes smaller, and for the sample Fe 3 O 4 /NC8, this shift is almost indistinguishable. This indicates that a lower number of hydrogen bonds between NC and the Fe 3 O 4 was formed with a decreasing percentage of magnetic nanoparticles in composites.
In addition, the SEM analysis was used for a morphological investigation of Fe 3 O 4 /NC ( Figure 4). The SEM micrographs indicate that the lower amount of NC resulted in an aggregation and formation of agglomerates of magnetite particles, which are different in size and shape (Figure 4a An improvement in the homogeneity of the prepared sample continues with an increase in the NC content (Fe 3 O 4 /NC4), i.e., the availability of its larger surface area. Within the Fe 3 O 4 /NC8 sample, the higher substrate area leads to an equal distribution of magnetite nanoparticles on the NC surface.
Cellulose fibers modified by paramagnetic elements (deposition of iron) may enable an improvement in cellulose orientation in a polymer matrix when exposed to lower strength electric and magnetic fields [28]. Having in mind the significant influence of the NC functionalization with Fe 3 O 4 and its influence on the magnetic properties of such materials, magnetic interactions in the investigated magnetite structures were analyzed by the determination of exchange coupling constants between neighboring iron atoms. Exchange couplings are described with the model of spin Hamiltonian, namely, the Heisenberg-Dirac-van Vleck (HDVV) Hamiltonian [51,52], which can be expressed by Equation (1): whereŜ A andŜ B stand for the effective spin operators, and J represents the exchange coupling constant. For the ferromagnetic interactions, with the parallel spin orientation, J has a positive sign, while the negative sign indicates the antiferromagnetic interactions with the antiparallel spin alignment.
In addition, the SEM analysis was used for a morphological investigation of Fe3O4/NC (Figure 4). The SEM micrographs indicate that the lower amount of NC resulted in an aggregation and formation of agglomerates of magnetite particles, which are different in size and shape (Figure 4a-c). The irregular Fe3O4 crystallites grew larger with lower loadings of NC in the composite sample. Apparently, a higher amount of NC favors a more uniform deposition of Fe3O4 nanoparticles. The homogeneous deposition of Fe3O4 on the surface of nanocellulose was found for the sample with the highest content of NC (Figure 4d). Generally, it seems that a nanocellulose high-surface area plays a crucial role in their merging into homogeneous layer-like structures.    The effective spin operators are related to the total spin operator (Equation (2)): Using the previous two equations, one can rewrite the Heisenberg-Dirac-Van Vleck Hamiltonian in the final form (Equation (3)): Hence, the expression for the eigenvalues of H EX for the different spin states is presented in Equation (4).
where S, S A and S B are the quantum numbers associated with the spin operators, while E 2S+1 represents the energy of a spin multiplet. In order to determine the magnetic couplings between iron atoms in an investigated magnetite structure, we performed the Density Functional Theory-Broken Symmetry (DFT-BS) method developed by Noodleman [31][32][33][34]. The exchange coupling constants were estimated according to the Yamaguchi approach (Equation (5)) [53,54].
where E HS is the energy of the high spin, E BS is the energy of the broken symmetry, while S 2 HS and S 2 BS are the expectation values of the high-spin and broken symmetry spin operators.
All calculations were conducted using the B3LYP [32,33] hybrid functional and TZP basis set. For comparison purposes, coupling constants were also calculated with the OPBE [37] functional. All calculations were performed with the ORCA program package, version 2.8-20 [55].
The exchange coupling constants between Fe 2+ and Fe 3+ ions were calculated on Fe 3 O 4 model systems, in which the first coordination sphere around the iron atoms was taken directly from the crystal structures ( Figure 5). The other atoms were replaced with zinc ions and hydrogen atoms. This represents a very useful method for the calculation of exchange coupling constants in polynuclear metal complexes. The exchange coupling constants between Fe 2+ and Fe 3+ ions were calculated on Fe3O4 model systems, in which the first coordination sphere around the iron atoms was taken directly from the crystal structures ( Figure 5). The other atoms were replaced with zinc ions and hydrogen atoms. This represents a very useful method for the calculation of exchange coupling constants in polynuclear metal complexes. The calculated exchange coupling constants, as well as relevant bond distances, are presented in Table 1. The calculated exchange coupling constants, as well as relevant bond distances, are presented in Table 1. In all cases, the predominant ferromagnetic coupling was obtained. Although the B3LYP functional represents the method of choice for the calculation of exchange coupling constants, the results obtained with the OPBE functional are quite similar. The calculated values are slightly lower at the B3LYP level of theory, in comparison to the OPBE level of theory.
Magnetic couplings between paramagnetic centers are the consequence of the electronic structure and coordination environment around metal ions. In the inverse spinel structure of Fe 3 O 4 , half of the Fe 3+ ions occupy tetrahedral voids with a high-spin e(2)t 2 (3) electronic configuration, while the Fe 2+ ions have octahedral coordination with a high-spin t 2g (4)e g (2) electronic configuration. The net interactions between Fe 3+ ions with tetrahedral coordination and Fe 2+ ions with octahedral environment can be explained through the analysis of molecular orbitals with predominant metal character (Figures 6 and 7).
Polymers 2022, 14, x FOR PEER REVIEW Figure 6. Singly occupied molecular orbitals (SOMO) in high-spin state with S = 9/2. A more fied structure was used, in which the negative oxygen charge was neutralized by hydrogen Figure 6. Singly occupied molecular orbitals (SOMO) in high-spin state with S = 9/2. A more simplified structure was used, in which the negative oxygen charge was neutralized by hydrogen atoms. Figure 6. Singly occupied molecular orbitals (SOMO) in high-spin state with S = 9/2. A more simplified structure was used, in which the negative oxygen charge was neutralized by hydrogen atoms.  Although ferromagnetic couplings have been observed, an orbital analysis indicates the possible antiferromagnetic interactions via a superexchange mechanism, through bridging the oxygen ligand. Moreover, the electron density is delocalized over both metal centers. It is well known that magnetic interactions between paramagnetic centers strongly depend on geometrical parameters, namely on bond distances and angles. In most cases, magnetic interactions decrease with increasing bond distances and angles. The change in the percentage of NC does not drastically influence a change of Fe 2+ -O and Fe 3+ -O bond distances (Table 1). Hence, the differences in magnetic couplings between Fe 2+ and Fe 3+ ions in the investigated structures are very small and demonstrate weak dependence on the percentage of NC. In the case of the Fe 3 O 4 /NC1 sample, the Fe 2+ -O and Fe 3+ -O bond distances are slightly longer in comparison to other structures, i.e., 2.081 and 1.846 Å, respectively. Indeed, the ferromagnetic interactions in this case are slightly lower.

Conclusions
In this study, the investigation of the influence of a nanocellulose amount on features of hybrid composite materials, based on functionalized nanocellulose with magnetic particles, has been presented. We focused on the investigation of the structural and morphological properties of nanocellulose/magnetite (Fe 3 O 4 /NC) composites. The XRD analysis confirmed the transformation from cellulose II into cellulose I allomorph with the addition of a magnetic particle. Further, a small but noticeable shift of the diffraction line at the Bragg angle of 22 • indicates coupling between phases. Detailed FTIR analysis confirmed the results obtained by XRD, i.e., the transition among allomorphs, along with changes in structure due to the covering of NC with Fe 3 O 4 . In order to enable the further development of multicomponent hybrid materials based on NC/Fe 3 O 4 , a theoretical calculation of mag-netic interactions and magnetic couplings between iron atoms in the magnetite structure within the investigated composite materials were performed. A B3LYP hybrid functional and TZP basis set were used for the calculation of bond distances and for the determination of coupling constants between neighboring iron atoms in Fe 3 O 4 /NC composites. The derived model confirmed the predominant ferromagnetic coupling. It was demonstrated that the amount of NC has an influence on the bound distance and angles. This conclusion suggests that the NC content may affect magnetic couplings between neighboring Fe 3+ and Fe 2+ ions.
This research provides expellant inputs for further investigation in the area of biosensors. The Fe 3 O 4 component of the composite can be easily manipulated with the permanent magnet, and the NC component is biodegradable and possesses piezoelectric properties. The MNP/NC is comprised of a multifferoic material, which electric and magnetic properties can be tailored to by different amounts of NC, and in that way, this composite is promoted as a good candidate for various sensor applications. We believe this material should be further examined in the domain of its magnetic and electrical properties, as well as for its use as a biosensor for appropriate antigen and antibody systems.
Author Contributions: A.J., writing-original draft and investigation; V.P.P., formal analysis, writing-review and editing and visualization; D.K., resources and investigation; M.P., software; B.V., writing-review and editing, and supervision and methodology; V.B.P., writing-review and editing, and supervision, methodology and project administration; S.F., writing-review and editing, and supervision and visualization. All authors have read and agreed to the published version of the manuscript.

Data Availability Statement:
The raw/processed data required to reproduce these findings cannot be shared at this time as the data also form part of an ongoing study.

Conflicts of Interest:
The authors declare no conflict of interest.