## 4 Discussion  
Previous studies have reported that the bone-like apatite formed on biomaterials exhibits a spherical shape [18]; calcium-, phosphorus-, carbon-, and oxygen-rich elemental composition [19]; and a v1 PO\({}_{4}\)[3]- Raman band at 960 cm\({}^{-1}\)[20]. In the present study, aged NBG samples, except for F-NBG aged in 10-g/L BSA-SBF, displayed surfaces covered by apatite-like spherical precipitates composed mainly of calcium, phosphorus, carbon, and oxygen elements, showing clear v1 PO\({}_{4}\)[3]- Raman bands at 960 cm\({}^{-1}\) (Figure 1). In contrast, the surface of F-NBG aged in 10-g/L BSA-SBF exhibited few spherical precipitates, bismuth elements originating from the NBG content, and a weak Raman band at 960 cm\({}^{-1}\) (Figure 1). These findings indicated that both F-NBG and P-NBG formed apatite in all tested media, although only a scant amount of apatite was formed on F-NBG in 10-g/L BSA-SBF. The positive AFA values of NBGs in SBF [3; 10; 11] and the negative effects of BSA on apatite formation [14; 15] are consistent with previous reports. Next, we further evaluated the phosphate ion consumption rate after immersing NBG samples in SBF and BSA-SBF, as this reflects apatite growth on NBGs [21; 22]. The rates of phosphate ion consumption of P-NBG were greater than that of F-NBG in all tested media (Figure 2), which indicated faster apatite growth on P-NBG than on F-NBG. Based on these findings, it can be concluded that P-NBG has greater AFA than F-NBG. The null hypothesis that the AFA of P-NBG does not differ from that of F-NBG was, thus, rejected. The greater AFA of P-NBG may be associated with its greater capacity to expose apatite nucleation sites on the surface. In the present study, BG-like crystals appeared on the as-prepared P-NBG at a higher density than on the as-prepared F-NBG. In addition, the surface elemental composition of the as-prepared P-NBG was silicon- and oxygen-rich but carbon- and magnesium-deficient compared with that of as-prepared F-NBG (Figure 3). This result indicated that more BG (SiO\({}_{2}\)-Na\({}_{2}\)O-CaO-P\({}_{2}\)O\({}_{5}\) glass) was exposed on the as-prepared P-NBG than on the as-prepared F-NBG. The BG partially degrades after contact with body fluids and negatively charged silanol groups are formed on the surface. The silanol groups then act as nucleation sites of apatite, binding calcium ions in the surrounding environment [23]. It is, therefore, reasonable to draw the conclusion that  
Figure 4: Calcium ion concentrations (**a**) and pH levels (**b**) of distilled water after immersion of flowable and putty-form Nishika Canal Sealer BG Multi (F-NBG and P-NBG). Asterisk indicates \(p<0.05\) (unpaired \(t\)-test, \(n=6\)). silon groups, acting as apatite nucleation sites, were more abundant on P-NBG than on F-NBG and enabled faster apatite formation on P-NBG. In the present study, distilled water was used to evaluate the calcium ion release and alkalizing ability of NBGs. In physiological solutions, NBGs consume calcium and hydroxyl ions for apatite formation, which obscures the calcium ion release and alkalizing effect of NBGs; therefore, distilled water is optimal for this purpose. Apatite growth on biomaterials is accelerated by increases in the calcium ion concentration and pH of the surrounding environment [24]. We found that the calcium ion release from P-NBG was significantly greater than that from F-NBG, whereas no significant difference in alkalizing ability was observed between NBGs (Figure 4). The greater calcium ion release may also be responsible for the greater AFA of P-NBG. Albumin inhibits apatite formation by obstructing apatite nucleation sites [25]. The sparse formation of apatite on F-NBG aged in 10-g/L BSA-SBF may have resulted from the almost complete blockage of apatite nucleation sites by albumin. In addition, it has been reported that albumin binds calcium and phosphate ions in body fluids and reduces the number of ions available for apatite nucleation [26].