Figure 2. a) Schematic
diagram of the optical path of Raman spectrometer. b) The relevant study
conditions of Raman spectroscopy for biomineralization.
Physicochemical study of Raman spectroscopy on
biomineralization
It is known that each peak illustrated by Raman spectroscopy corresponds
to a specific molecular bond vibration, which includes both the single
chemical bond and the vibration of a group composed of several chemical
bonds involved in biomineralization. The Raman spectral peaks in
biomineralized tissues include those attributed to inorganic and organic
substances.[123–125]The main mineral component in teeth
and bones is hydroxyapatite, which exhibits
ν1PO43-, ν2PO43-, ν3PO43-, and ν4PO43- fundamental frequency modes in
the Raman spectrum (Table 1 ). The sharpest and most intense
band, ν1 PO43-, is
associated with the symmetric stretching of the oxygen atom tetrahedra
around the phosphorus atom, with the main peak located at 960
cm-1, representing the characteristic peak of
hydroxyapatite.[59,126] Collagen plays a role in
the induction of hydroxyapatite nucleation and growth in bone and tooth
mineralization.[127–129] The three main amide
bands of collagen are amide I (1675 cm-1), amide II
(1565 cm-1), and amide Ⅲ (1272
cm-1). Among them, ν1PO43- and amide Ⅲ are susceptible to
the direction of incident light and the direction of polarization. This
phenomenon can be used to analyze the orientation of the crystal and
collagen fibers.[3,33,130,131] In addition to the
characteristic peaks of collagen and hydroxyapatite, there are other
peaks, including ν1CO32-, ν2CO32-, ν4CO32-, ν(C-C), δ(CH2)
and ν(O-H), etc. (Table 1 ).
Pathological mineralization, such as tumor micro mineralization, urinary
stones, and atherosclerosis, is distinct from the mineral and organic
composition of physiological
mineralization.[7,8,132–134] In atherosclerosis,
the characteristic marker bands of the protein are located at 1181,
1209, 1272, 1410, 1565, and 1675 cm-1, corresponding
to ν(C-O-C), ω(CH2), amide III, β(CH2),
amide II and amide I, respectively (Table 1 ). One of the
minerals belongs to carbonate apatite, which also has the characteristic
peak of ν1 PO43- at
960 cm-1.[135]Microcalcification of tumor tissue is usually due to oversized tumor
tissue, resulting in deep tumors not receiving nutrients These tumor
cells eventually calcify into minerals after
necrosis.[66,136] The calcified composition is
mainly that of apatite. However, due to the heterogeneity of the tumor,
differences in organic matter composition can exist. The cause of
urinary stones is the nucleation of insoluble microcrystals in the
urethra and kidney tubules.[64,137] These
insoluble microcrystals are various types of calcium oxalate
crystals.[138,139] In the urinary environment, the
nucleation and dissolution processes of calcium oxalate crystals are in
a dynamic equilibrium, while the true stone formation process is still
unknown.
Morphology and other characterization of biomineralization
As described above, the specific mineralization information could be
readily obtained by Raman spectroscopy. Meanwhile, other
characterization strategies have been also utilized to analyze the
morphology and crystal morphology of the biomineralization. Scanning
electron microscopy (SEM) provides nanoscale resolution of sample
morphological features and, together with an energy dispersive
spectrometer (EDS), it can provide quantitative elemental analysis of
mineralized tissues.[140,141] X-ray diffraction
(XRD) is the main method for studying the physical phase and crystal
structure of minerals.[9,142,143] The
crystallographic characteristics such as composition, crystalline shape,
intra-molecular bonding patterns, molecular conformation, and
conformation are obtained by the diffraction phenomenon of minerals
irradiated by X-rays to different degrees. X-ray computed tomography
uses precisely collimated x-ray beams to obtain macroscopic structural
images of biomineralized tissues.[144,145]Multiple analysis techniques are needed to obtain comprehensive
nano-structural information on the general nature of relevant
biomineralization.