Hydroxyapatite-Based Magnetic Bionanocomposite as Pharmaceuticals Carriers in Chitosan Scaffolds

: Hydroxyapatite (HA) is a bioceramic very similar to the mineral component of bones and teeth. It is well established that osteoblasts grow better onto HA-coated metals than on metals alone. Herein, the preparation of a new system consisting of magnetite (Fe 3 O 4 ) and HA functionalized with oleic acid and simvastatin (SIMV), and incorporated in chitosan (CHI) scaffolds, was undertaken. HA was synthesized by the hydrothermal method, while Fe 3 O 4 was synthesized by co-precipitation. The polymer matrix was obtained using a 2% CHI solution, and allowed to stir for 2 h. The ﬁnal material was freeze-dried to produce scaffolds. The magnetic properties remained unchanged after the formation of the composite, as well as after the preparation of the scaffolds, maintaining the superparamagnetism. CHI scaffolds were analyzed by scanning electronic spectroscopy (SEM) and showed a high porosity, with very evident cavities, which provides the functionality of bone growth support during the remineralization process in possible regions affected by bone tissue losses. The synthesized composite showed an average particle size between 15 and 23 nm for particles (HA and Fe 3 O 4 ). The scaffolds showed considerable porosity, which is important for the performance of various functions of the tissue structure. Moreover, the addition of simvastatin in the system can promote bone formation. N.M.P.S.R. and P.B.A.F.; data curation, A.V.C. and P.B.A.F.; writing—original draft preparation, A.V.C.; writing—review and editing, A.V.C. and P.B.A.F.; visualization, R.M.F., D.M.A.N. and P.B.A.F.; supervision, V.P.F. and P.B.A.F.; project administration, A.V.C. and P.B.A.F.; funding


Introduction
Natural and synthetic bioceramics calcium phosphates are currently presented as one of the main materials used in medicine and dentistry. They are applied to the repair of bone loss due to their physical, chemical, and biological characteristics, which stimulates an adequate response of living tissue [1].

Synthesis of Fe 3 O 4 Nanoparticles
The synthesis of MNPs was carried out following the methodology used by Barreto et al. [24]. Briefly, 30 mL of Milli-Q ®® water was acidified in a beaker with 5% HCl solution until pH adjustment between 3 and 4 under constant stirring at 70 • C. Thereafter, metal salts were weighed and dissolved in Milli-Q ®® water as precursors of Fe +2 and Fe +3 in a molar ratio of 1:2, respectively. For this, 1.08 M FeCl 3 ·6H 2 O and 0.54 M FeSO 4 ·7H 2 O aqueous solutions were produced and heated in a previously acidified solution. A 5% NH 4 OH solution was added to adjust the pH to 10. After the adjustment, a black precipitate was observed, which denotes the formation of magnetite. The reaction that occurred during this process can be summarized as follows: After 30 min of stirring, the mixture containing the black precipitate was naturally cooled to room temperature. Magnetic decantation was used to separate the product, and then the solution was discarded. The process was repeated several times to wash the material using Milli-Q ®® water and methanol. Finally, the powder was dried and stored under a vacuum for further application and characterization.

HA Synthesis via Hydrothermal Treatment
The required concentration of H 3 PO 4 and CaCl 2 ·2H 2 O solution was mixed under stirring to obtain a Ca/P molar ratio equals 1.67. After this, a white precipitate was formed by adding NH 4 OH solution to adjust the pH to 9. The white suspension was stirred for 2 h. The precipitate was washed with Milli-Q ®® water and vacuum filtered. The powder was then re-suspended in a solution of NH 4 Cl 0.1 M, with the pH maintained at 9 to undergo the hydrothermal bath. The weight ratio between the precipitate and the solution in this step was 1:10. The suspension was transferred for a Teflon autoclave with a stainless steel support to receive the hydrothermal treatment at 150 • C for 5 h. After this, the autoclave was cooled to room temperature and the product was filtered, washed, and dried at 80 • C [25]. Equation (2) describes the synthesis of HA.

Composite Synthesis Fe 3 O 4 -HA
The first step for the synthesis of magnetic composite consists primarily of the production of Fe 3 O 4 through co-precipitation. Thus, the iron oxide was dispersed in the 0.5 M CaCl 2 ·2H 2 O solution under constant agitation using a mechanical stirring. Afterward, 0.3 M H 3 PO 4 were slowly dripped, maintaining a Ca/P molar ratio equals 1.67, and then 5% (v/v) NH 4 OH was used to adjust the pH to 9.0. In this step, the color of the solution changed from black to gray. The precipitate was separated by filtration, washed, and dried. The dry powder was dispersed in 0.1 M NH 4 Cl (90% w/w) and the solution was transferred to a Teflon-lined autoclave to perform the hydrothermal treatment (150 • C for 5h). Finally, the product was filtered, washed, and dried at 80 • C. The weight ratio of the composite was 90:10 (HA:Fe 3 O 4 ) [26].

Surface Modification of Fe 3 O 4 -HA with Oleic acid (Fe 3 O 4 -HA@OA)
500 mg of the synthesized magnetic nanocomposite were dispersed in 50 mL of chloroform using an ultrasonic treatment for 30 min, is then heated until it reached 75 • C. Afterward, 4 mL of OA was slowly added, and the mixture was mechanically stirred for 30 more minutes. The mixture was centrifuged at 3000× g rpm for 20 min and the precipitate was washed repeatedly with distilled water and ethanol. Finally, the product was dried at room temperature and stored under vacuum.

Simvastatin Adsorption on the Surface of Fe 3 O 4 -HA@OA
First, some commercially available pharmaceutical capsules (Sinvasmax ® , Globo Laboratory, São José da Lapa, Brazil) were macerated, and the powder was dispersed in 20 mL of acetonitrile (ACN). Unlike the excipient, SIMV is soluble in acetonitrile. Thus, the suspension was filtered to collect the excipient (insoluble solid), separating the SIMV, which was recrystallized at room temperature, macerated, and stored under vacuum and darkness. After this, in a falcon tube, 100 mg of previously synthesized Fe 3 O 4 -HA@OA was mixed with 10 mL solution of 2000 µg.mL −1 SIMV/ACN. The tube was subjected to strong horizontal stirring with shaker equipment for 24 h. Figure 1 describes the process of synthesis and adsorption of the drug in the composite. darkness. After this, in a falcon tube, 100 mg of previously synthesized Fe3O4-HA@OA was mixed with 10 mL solution of 2000 μg.mL −1 SIMV/ACN. The tube was subjected to strong horizontal stirring with shaker equipment for 24 h. Figure 1 describes the process of synthesis and adsorption of the drug in the composite.

Preparation of Chitosan Scaffolds with Fe3O4-HA@OA-SIMV
4 g of chitosan (Sigma-Aldrich, deacetylation degree 86%), low molecular weight powder, was dissolved in 200 mL of acetic acid solution 1% (v/v) under vigorous stirring for 3 h. Then, the solution was centrifuged at 4000 rpm for 30 min for the separation of possible impurities [27]. Using this solution as a solvent, 0.8 g of the composite was dispersed in 8 g of CHI solution. The mixture was sonicated for 30 min and then transferred to the template (5 cm × 5 cm). Finally, the filled template was frozen and lyophilized to obtain the scaffold.

Characterizations
The structural analysis and verification of the single-phase nature of the samples were obtained using X-ray diffraction (XRD). These measurements were carried out using XPert Pro MPD-Panalytical X-ray diffractometer equipped with CuKα radiation tube (k = 1.5406 Å) operated at a voltage of 30 kV and current of 15 mA. The identification analysis phase was conducted by comparing powder diffractograms with standard patterns from the International Centre for Diffraction Data (ICDD). The Rietveld refinement procedure [28] was applied to all diffraction patterns using the DBWS 2.4 [29]. The average crystallite sizes of NPs were calculated from the XRD data using the Scherrer equation.
The samples were also surveyed through Fourier-transform infrared (FTIR) spectra. The samples were macerated and pressed with KBr to form tablets with a 1:10 (sample:KBr) ratio. The spectra were obtained between 4000 and 400 cm −1 in a FTIR spectrophotometer (FTIR 8300, Shimadzu, Tokyo, Japan) by using 2 cm −1 resolution and 32 scans per analyzes.   [27]. Using this solution as a solvent, 0.8 g of the composite was dispersed in 8 g of CHI solution. The mixture was sonicated for 30 min and then transferred to the template (5 cm × 5 cm). Finally, the filled template was frozen and lyophilized to obtain the scaffold.

Characterizations
The structural analysis and verification of the single-phase nature of the samples were obtained using X-ray diffraction (XRD). These measurements were carried out using XPert Pro MPD-Panalytical X-ray diffractometer equipped with CuKα radiation tube (k = 1.5406 Å) operated at a voltage of 30 kV and current of 15 mA. The identification analysis phase was conducted by comparing powder diffractograms with standard patterns from the International Centre for Diffraction Data (ICDD). The Rietveld refinement procedure [28] was applied to all diffraction patterns using the DBWS 2.4 [29]. The average crystallite sizes of NPs were calculated from the XRD data using the Scherrer equation.
The samples were also surveyed through Fourier-transform infrared (FTIR) spectra. The samples were macerated and pressed with KBr to form tablets with a 1:10 (sample:KBr) ratio. The spectra were obtained between 4000 and 400 cm −1 in a FTIR spectrophotometer (FTIR 8300, Shimadzu, Tokyo, Japan) by using 2 cm −1 resolution and 32 scans per analyzes.
The magnetic measurements were obtained using a vibrating sample magnetometer (Cryogenic VSM 5 Tesla system, Machida-shi, Tokyo, Japan) with a magnetic field of 5 Tesla and the temperature range 2-300 K (Department of Physics, University of Santiago de Chile-USACH). The VSM had been previously calibrated using a pure Ni wire, and, after measuring the mass of each sample, the magnetization was given in emu·g −1 .
Transmission electron microscope (TEM) images were obtained using a JEOL JEM 2100 LaB6 (JEOL, Tokyo, Japan) operating at accelerating voltage equals 200 kV and equipped with a TV (Gatan ES500W); CCD (TVips-16 MP). TEM samples were prepared by dropcasting a hexane dispersion of the nanocrystals onto carbon and carbon-coated copper grids. After deposition, the samples were dried to a temperature of 60 • C overnight before the obtention of the images. These analyzes were carried out in partnership with the Università del Salento, Italy.
The characterization of the SIMV was done using the spectra in the ultraviolet region in the wavelength range 190-300 nm, using a solution of 16 mg/mL SIMV in ACN. The maximum absorption was compared with those reported in the literature.
The morphology and porosity of the functionalized nanocomposite were evaluated using microscopy scanning Quanta 450 FEG (FEI, Amsterdam, The Netherlands). The samples (scaffolds) were fixed on carbon tapes, and then air-dried and coated with a thin layer of gold to gain better conductivity, as is required for high-quality SEM imaging. The powder was fixed on silver glue and examined using the same instrument.
The thermogravimetric analysis (TGA) curves are usually obtained on TGA Q50 V20.13 Build 39 of thermal analysis (TA) Instruments at a heating rate of 10 • C min −1 , starting at room temperature to 950 • C in a dynamic atmosphere of nitrogen and flow 25 mL.min −1 alumina crucible and sample mass approximately 5.0 mg. The differential scanning calorimetry (DSC) curves were obtained on DSC Q20 V24.9 Build 121 equipment under a dynamic nitrogen atmosphere with a flow rate of 50 mL·min −1 , heating rate of 10 • C·min −1 , with heating from room temperature to 400 • C, partially closed aluminum pan and sample mass about 0.5 mg. The equipment must be calibrated with indium (MP 156.6 • C; ∆H melting = 28.54 J·g −1 ) and lead (MP 327.0 • C), or other standards recommended by the manufacturer. The purity by the DSC evaluation is performed by applying the Van't Hoff equation to the melting peak of the drug. Additionally, one might note that in the Rietveld method, some of the most important data to be evaluated are the R WP (by obtaining the error percentage) and S (ratio of error provided by the expected error), since they refer to the agreement and the progress of refinement (Table 1) [30,31]. The observed peaks are approximately coincident with the calculated peaks, which can be seen in a line representing the difference between the intensity observed and intensity calculated (Figure 2a). This justifies the values shown in Table 1 and demonstrates the satisfactory character of refinement.  According to the calculated data presented in Table 1, it was shown that the average crystallite sizes of the samples were 18.37 nm for HA, 15.5 nm for Fe 3 O 4 and 22.53 nm for the composite. It was observed that the particle dimensions presented small variation even after the formation of the composite, maintaining its size on a nanometric scale. A similar size in the refinement by the Rietveld method has been reported in the literature [32]. Figure 2b shows the FTIR spectra of precursors (Fe 3 O 4 and HA) to confirm the binding band assignments of the synthesized compounds. Regarding the Fe 3 O 4 , bands related to the bonds for iron spinel were observed, with a wavenumber of 582 cm −1 , referring to the stretching of the Fe bonds in the tetrahedral site, 1400 cm −1 referring to the angular vibration related to the Fe-O bond, 1635 cm −1 and 3410 cm −1 agreeing with the angular vibration and stretching of OH bonds respectively [33,34]. All vibrational modes of the HA functional groups were identified. The very strong bands observed in 1097 and 1033 cm −1 , respectively, were associated with vibrational stretches outside the P-O plane, whereas the weak band centered in 960 cm −1 was attributed to the symmetrical stretch vibration of O-P-O. Strong O-P-O vibrational waves can be seen in the wavenumbers between 563 and 603 cm −1 [35,36]. In this way, the characteristic bands of the two materials are also identified in the Fe 3 O 4 -HA spectra.

Magnetization Measurements
The magnetization curves were used to investigate the magnetic properties of the nanocomposite. In this sense, Figure 3a,b displayed the measurements recorded at room temperature for Fe 3 O 4 and Fe 3 O 4 -HA, respectively. The absence of hysteresis was observed for both nanomaterials, which indicates a superparamagnetic behavior [37,38]. However, the Fe 3 O 4 -HA nanocomposite was found to have inferior magnetic properties compared to Fe 3 O 4 NPs. This can be explained by the presence of non-magnetic material [39]. Therefore related to the bonds for iron spinel were observed, with a wavenumber of 582 cm , referring to the stretching of the Fe bonds in the tetrahedral site, 1400 cm −1 referring to the angular vibration related to the Fe-O bond, 1635 cm −1 and 3410 cm −1 agreeing with the angular vibration and stretching of OH bonds respectively [33,34]. All vibrational modes of the HA functional groups were identified. The very strong bands observed in 1097 and 1033 cm −1 , respectively, were associated with vibrational stretches outside the P-O plane, whereas the weak band centered in 960 cm −1 was attributed to the symmetrical stretch vibration of O-P-O. Strong O-P-O vibrational waves can be seen in the wavenumbers between 563 and 603 cm −1 [35,36]. In this way, the characteristic bands of the two materials are also identified in the Fe3O4-HA spectra.

Magnetization Measurements
The magnetization curves were used to investigate the magnetic properties of the nanocomposite. In this sense, Figure 3a,b displayed the measurements recorded at room temperature for Fe3O4 and Fe3O4-HA, respectively. The absence of hysteresis was observed for both nanomaterials, which indicates a superparamagnetic behavior [37,38]. However, the Fe3O4-HA nanocomposite was found to have inferior magnetic properties compared to Fe3O4 NPs. This can be explained by the presence of non-magnetic material [39]. Therefore, the HA component does not affect the magnetic nature of the composite. However, its influence on the saturation magnetization (Ms) is clear, since the nonmagnetic mass was taking into consideration to normalize the M parameter. Due to this,  To deepen further our study, the experimental data obtained through magnetic measurements of the samples were described by the Langevin function [40], using the following equation: where N, KB, T e H indicates, respectively, the number of the ions of Fe 2+ , Boltzmann constant, temperature, and the magnetic field applied. By using this function, an average diameter related to the magnetic-ordered structure (dL) can also be inferred by setting a parameter / . Once this parameter was calculated, the dL was obtained using To deepen further our study, the experimental data obtained through magnetic measurements of the samples were described by the Langevin function [40], using the following equation: where N, K B , T e H indicates, respectively, the number of the ions of Fe 2+ , Boltzmann constant, temperature, and the magnetic field applied. By using this function, an average diameter related to the magnetic-ordered structure (d L ) can also be inferred by setting a parameter a = m/K B . Once this parameter was calculated, the d L was obtained using a = 4p(d L /2) 3 M 0 /3K B [41]. It was observed that the particle sizes found by the magnetiza-  Figure 4A) [42,43]; while HA NPs exhibited rod-like morphology ( Figure 4B), which corroborates with a c-axis preferential orientation demonstrated by the XRD; and for the sample Fe 3 O 4 -HA we could evidence NPs with morphologies such as Fe 3 O 4 and HA, as pointed with green and red arrows ( Figure 4C). TEM analysis confirms that the mixture of the two components of the magnetic composite is mixed without the core-shell structure. However, even without the core-shell structure, we demonstrated that the magnetic composite behaved, as a whole, as magnetic material, as shown in Figure 1 C in Section 2.6. 4 ( /2) /3 [41]. It was observed that the particle sizes found by the magnetization curve crystallites are close to the values found by the PXRD, showing an approximate average size of 13.3 nm to Fe3O4 and 27.5 nm to Fe3O4-HA. Figure (4A-C) shows TEM images for Fe3O4, HA, and Fe3O4-HA. The sample Fe3O4 is composed by NPs with pseudo spherical morphology, which is expected for Fe3O4 NPs synthesized by co-precipitation using NH4OH ( Figure 4A) [42,43]; while HA NPs exhibited rod-like morphology ( Figure 4B), which corroborates with a c-axis preferential orientation demonstrated by the XRD; and for the sample Fe3O4-HA we could evidence NPs with morphologies such as Fe3O4 and HA, as pointed with green and red arrows ( Figure 4C). TEM analysis confirms that the mixture of the two components of the magnetic composite is mixed without the core-shell structure. However, even without the core-shell structure, we demonstrated that the magnetic composite behaved, as a whole, as magnetic material, as shown in Figure 1 C in Section 2.6. XRD refinement indicated that the biomaterial is composed of 93.5% of hydroxyapatite and 6.5% of Fe3O4 (Table 1). However, in Figure 4, it is possible to see that the number of NPs with rod-like morphology does not correspond to the molar fraction found in the XRD. This indicates that the presence of Fe3O4 in the reaction medium to produce HA affected the growth of HA NPs, decreasing the c-axis preferential orientation. This fact was also demonstrated in XRD refinement. Therefore, our results suggest that physical interaction occurred between Fe3O4 and HA, probably through electrostatic interactions.

Thermal Analysis
Simvastatin was characterized using the DSC technique for measuring the heat flow difference between a substance and a reference material as a function of a heating or cooling program. This thermal characterization and determination of the purity of drugs are widely used in the pharmaceutical industry [44]. The DSC curve of the SIMV ( Figure  5a) shows the first endothermic event at temperatures ranging from 52.46 °C with power consumption, which probably refers to the boiling point of the solvent used in the recrystallization, in this case, ACN. The second event, characteristic of the melting process of the substance, SIMV, starts at 122.85 °C, with power consumption. Thus, according to the curve, SIMV showed a melting temperature of 134.5 °C. The spectrum in the ultraviolet (UV) region is shown in Figure 5b and agrees with the spectrum described in the literature for simvastatin absorption maxima at 231, 238, and 247 nm [45]. XRD refinement indicated that the biomaterial is composed of 93.5% of hydroxyapatite and 6.5% of Fe 3 O 4 (Table 1). However, in Figure 4, it is possible to see that the number of NPs with rod-like morphology does not correspond to the molar fraction found in the XRD. This indicates that the presence of Fe 3 O 4 in the reaction medium to produce HA affected the growth of HA NPs, decreasing the c-axis preferential orientation. This fact was also demonstrated in XRD refinement. Therefore, our results suggest that physical interaction occurred between Fe 3 O 4 and HA, probably through electrostatic interactions.

Thermal Analysis
Simvastatin was characterized using the DSC technique for measuring the heat flow difference between a substance and a reference material as a function of a heating or cooling program. This thermal characterization and determination of the purity of drugs are widely used in the pharmaceutical industry [44]. The DSC curve of the SIMV (Figure 5a) shows the first endothermic event at temperatures ranging from 52.46 • C with power consumption, which probably refers to the boiling point of the solvent used in the recrystallization, in this case, ACN. The second event, characteristic of the melting process of the substance, SIMV, starts at 122.85 • C, with power consumption. Thus, according to the curve, SIMV showed a melting temperature of 134.5 • C. The spectrum in the ultraviolet (UV) region is shown in Figure 5b and agrees with the spectrum described in the literature for simvastatin absorption maxima at 231, 238, and 247 nm [45].
The melting point of SIMV, according to the literature, ranging from 135-138 • C [46]. Therefore, the purity of this drug recrystallized showed up around 94.37 to 97.53%.
The TGA of the modified sample was performed to verify the presence of oleate and SIMV molecules through mass-loss events. Therefore, Figure 6 shows the thermograms of samples with and without surface modification, respectively. The first thermogram indicates that up to 850 • C, the weight loss of Fe 3 O 4 -HA is 5.8% of the original. Furthermore, the thermogram finds the stability at elevated temperatures of bioceramic [47]. Initially, the loss corresponds to the thermal desorption of water from the surface of NP. Near 200 • C, the weight loss is attributed to the elimination of structural H 2 O lattice. This long-range of temperature, which occurs in weight loss, indicates that the water does not interact evenly with the structure, which is compatible with the existence of different structural sites. The total mass loss for the sample modified with OA was 10.75%. The slight weight loss observed next to 250 • C indicates some OA molecules were adsorbed on the sample surface Fe 3 O 4 -HA@OA (Figure 6b) [48]. The melting point of SIMV, according to the literature, ranging from 135-138 °C [46]. Therefore, the purity of this drug recrystallized showed up around 94.37 to 97.53%.
The TGA of the modified sample was performed to verify the presence of oleate and SIMV molecules through mass-loss events. Therefore, Figure 6 shows the thermograms of samples with and without surface modification, respectively. The first thermogram indicates that up to 850 °C, the weight loss of Fe3O4-HA is 5.8% of the original. Furthermore, the thermogram finds the stability at elevated temperatures of bioceramic [47]. Initially, the loss corresponds to the thermal desorption of water from the surface of NP. Near 200 °C, the weight loss is attributed to the elimination of structural H2O lattice. This long-range of temperature, which occurs in weight loss, indicates that the water does not interact evenly with the structure, which is compatible with the existence of different structural sites. The total mass loss for the sample modified with OA was 10.75%. The slight weight loss observed next to 250 °C indicates some OA molecules were adsorbed on the sample surface Fe3O4-HA@OA (Figure 6b) [48]. However, any weight loss above 400 °C may indicate distinct oleate interactions between molecules and atoms' surface, leading to differences in the intensities of these interactions. Therefore, it is possible to determine that the weight loss compared to OA was 5.85%, i.e., the amount of the OA adsorbed on Fe3O4-HA@OA sample is approximately this value. The thermogram shows four events for functionalized composite samples with SIMV, indicating the presence of physiosorbed species, total weight loss of 15.4%. In the temperature range of approximately 175-600 °C, in which 14.5% of the mass was eliminated, three events can be attributed to SIMV decomposition However, any weight loss above 400 • C may indicate distinct oleate interactions between molecules and atoms' surface, leading to differences in the intensities of these interactions. Therefore, it is possible to determine that the weight loss compared to OA was 5.85%, i.e., the amount of the OA adsorbed on Fe 3 O 4 -HA@OA sample is approximately this value. The thermogram shows four events for functionalized composite samples with SIMV, indicating the presence of physiosorbed species, total weight loss of 15.4%. In the temperature range of approximately 175-600 • C, in which 14.5% of the mass was eliminated, three events can be attributed to SIMV decomposition and loss of OA chemisorbed and physiosorbed. Figure 6 also shows a comparison between the thermograms of the three samples, presenting the different weight losses. The improved thermal stability given by the Fe 3 O 4 -HA indicates that the incorporation of OA and SIMV on their surface was successfully carried out and it possibly indicates that the weight loss of SIMV was 4.65% compared to composites with and without this drug.

Chitosan Bionanocomposite
SEM micrographs of the pure CHI matrix and CHI/Fe 3 O 4 -HA@OA-SIMV bionanocomposite are presented in Figure 7a,c, respectively. The micron-size elements are distributed in the scaffold. The visualization of nano-sized particles is not possible in Figure 7, but can be observed on grains with the presence of all chemical items for the magnetic composite structures and the CHI matrix in the micrographs. It is possible to verify the high porosity of the scaffold distributed all over the area of the polymer, which benefits a better efficiency of the composite face to magnetic material adhesion, and the degradability and assimilation function in the bone area to be regenerated. The chemical analysis by energy-dispersive X-ray (EDX) (Figure 7b,d) showed the chemical elements of the CHI matrix and the presence of iron, calcium, and phosphate ions in the bionanocomposite respectively, including through CHI fibers. This allows us to infer that the NPs were distributed all over the scaffold matrix.

Conclusions
The hydroxyapatite-based magnetic composite functionalized with OA and simvastatin was synthesized according to the proposed methodology, in addition to the It is possible to verify the high porosity of the scaffold distributed all over the area of the polymer, which benefits a better efficiency of the composite face to magnetic material adhesion, and the degradability and assimilation function in the bone area to be regenerated. The chemical analysis by energy-dispersive X-ray (EDX) (Figure 7b,d) showed the chemical elements of the CHI matrix and the presence of iron, calcium, and phosphate ions in the bionanocomposite respectively, including through CHI fibers. This allows us to infer that the NPs were distributed all over the scaffold matrix.

Conclusions
The hydroxyapatite-based magnetic composite functionalized with OA and simvastatin was synthesized according to the proposed methodology, in addition to the production of CHI scaffolds and their incorporation of the composite. Based on VSM, as well as particle size analysis by diffraction, the size of magnetite with hydroxyapatite NPs was in the range of 15-30 nm. According to the TEM images, the size of the particles was 40-60 nm; and the SEM images showed the presence of all compounds synthesized, in addition to showing high porosity of the scaffolds. The magnetization of the composite after the addition of HA suggests that the structure of magnetite remained unchanged at around 4.85 emu·g −1 . The adsorption of SIMV on the composite can be observed in the TGA, which showed a total weight loss of 4.65% of this drug. The literature review supports the hypothesis that the compounds used in this work induce osteogenesis and the production of a magnetic composite, aiming at the creation of better material, capable of being applied in bone regeneration in the future. Therefore, the association of SIMV with calcium phosphates and magnetite encourages research in the area of tissue engineering.