Towards refining Raman spectroscopy-based assessment of bone composition

Various compositional parameters are derived using intensity ratios and integral area ratios of different spectral peaks and bands in the Raman spectrum of bone. The [Formula: see text]₁-, [Formula: see text]₂-,[Formula: see text]₃-, [Formula: see text]₄ PO₄^3-, and [Formula: see text] CO₃^2- bands represent the inorganic phase while amide I, amide III, Proline, Hydroxyproline, Phenylalanine, δ(CH₃), δ(CH₂), and [Formula: see text](C-H) represent the organic phase. Here, using high-resolution Raman spectroscopy, it is demonstrated that all PO₄^3- bands of bone either partially overlap with or are positioned close to spectral contributions from the organic component. Assigned to the organic component, a shoulder at 393 cm^-1 compromises accurate estimation of [Formula: see text]₂ PO₄^3- integral area, i.e., phosphate/apatite content, with implications for apatite-to-collagen and carbonate-to-phosphate ratios. Another feature at 621 cm^-1 may be inaccurately interpreted as [Formula: see text]₄ PO₄^3- band broadening. In the 1020-1080 cm^-1 range, the ~ 1047 cm^-1 [Formula: see text]₃ PO₄^3- sub-component is obscured by the 1033 cm^-1 Phenylalanine peak, while the ~ 1076 cm^-1 [Formula: see text]₃ PO₄^3- sub-component is masked by the [Formula: see text]₁ CO₃^2- band. With [Formula: see text]₁ PO₄^3- peak broadening, [Formula: see text]₂ PO₄^3- integral area increases exponentially and individual peaks comprising the [Formula: see text]₄ PO₄^3- band merge together. Therefore, [Formula: see text]₂ PO₄^3- and [Formula: see text]₄ PO₄^3- band profiles are sensitive to changes in mineral crystallinity.