To characterize the transformation process of ACP to HAP (ACP-HAP) regulated by poly(acrylic acid) (PAA), an in situ fluorescence dualprobe method was established using Eu3+ and tetracarboxylic acid tetraphenylethylene (TCPE). Based on the principles of the Eu3+ hypersensitive transition matching the asymmetry of the crystal field environment and the aggregation-induced emission (AIE) of TCPE, the characteristics of the ACP-HAP process were successfully visualized through the Eu3+ fluorescence ratio (I614/I591) for the crystallization feature and the TCPE fluorescence intensity (I468) for the state of PAA. During the crystallization process in the cycling system, the I614/I591 ratio of Eu3+ first decreased and then increased, and I468 of TCPE continued to decrease. This indicated that the Ca2+ ions bonded to the carboxyl groups of PAA were transferred to the phosphate group, and PAA was gradually separated during the ACP-HAP process. Based on the Eu3+ fluorescence probe, the relationship between the normalized I614/I591 (y) and the mass ratio (x) of Eu-HAP/Eu-PAA-ACP was established (y = 0.347 + 0.673 × 0.874x, R2 = 0.993). The HAP content was quantitatively analyzed during the early stage of the ACP-HAP process in a cycling system simulating the blood flow. On the second, third, and fourth days, the content of HAP was approximately 0.900, 1.732, and 3.288%, respectively, which could not be detected by XRD. It can be speculated that this in situ fluorescence probe method is helpful to investigate the early stages of biomineralization.
Tan et al. (Thu,) studied this question.