3.3.2 XRD analysis and FTIR after immersion in SBF
The XRD of the prepared scaffolds after soaking in SBF for different time intervals is demonstrated in Fig. 6. XRD spectra of Ch/MB composite scaffolds showed sharp peaks, 25.88°, 31.8°, 39.89° and 46.7° (2θ) attributed to 022, 211, 221 and 222 reticular planes of HA [30, 31]. The increase in the intensity of peaks from 2 days to 30 days was indicative of the increase in the deposition of HA. It also confirmed that the presence of MB increased the deposition of HA on the scaffolds  (Fig. 6). The IR spectrum of synthetic hydroxy apatite was used as references to evaluate the structural evolution and the bioactivities of the prepared scaffolds . After soaking in SBF solution, the initial characteristic bands of the Ch/MB biocomposite are modified strongly because of the interfacial reactions between scaffolds and the SBF. Consequently, the spectra of these biomaterials reveal new bands as demonstrated in Fig. 7.
In detail, the spectrum of Ch/MB biocomposite shows three new, well-defined phosphate bands at 565, 603 and 1039cm–1 after 2 days of soaking in physiological solution for Ch/MB scaffolds. They are assigned to stretching vibrations of the PO4 3– group in phosphate crystalline phases. This result confirms the formation of a calcium phosphate layer; this spectrum is quite similar to that of hydroxyl apatite except for the two bands located at 1620 and 3423 cm–1. These bands are characteristic of the presence of water related to the hygroscopic feature of the apatite formed. In addition, the carbonate band at 1420 cm–1 is also observed. This band attributes to a stretching vibration of the C−O liaisons in carbonate groups. The presence of carbonate bands indicates the formation of a layer of carbonated hydroxyl apatite on the surface of Ch/MB biocomposite. The results obtained highlight the rapid formation of the apatite layer on the surface of the Ch/MB biocomposite. In addition, Ch/MB scaffolds reveal three Si–O–Si bands at 470 cm–1 tion), 799 cm–1 (bending vibra-(bending vibration) and 1075 cm–1 (stretching vibration). These confirm the presence of a silica gel . The appearance of apatite mineral and a silica gel indicate the interactions between the scaffolds and SBF as described by Hench et al. . The results obtained confirm the bioactivity of the Ch/MB biocomposite.
3.3.3 SEM with EDS after immersion in SBF
The bioactive character of the composite scaffolds was tested in vitro by analyzing the ability to form apatite at their surface after being immersed in SBF. Two compositions of the prepared scaffolds were investigated by SEM coupled with EDS (Fig. 8) to evaluate their surface changes after soaking in SBF for 21 days. The 1Ch:2MB sample was chosen with reference to Ch alone due to the high MB content, which induces a much better formation of the Ca–P layer on their surfaces, as confirmed by XRD and FTIR. A marked apatite formation can be seen on the surface of Ch/MB...