Figure 4. Fourier-transformed infrared spectroscopy of NLP and NB reveals the presence of both carbonate and phosphate groups.

(A–D) NLP were prepared as in Fig. 3 and submitted to FTIR spectroscopy (A) immediately after formation, or after the following periods of incubation at 37°C in DMEM: (B) overnight; (C) one week; and (D) one month. The FTIR spectrum of NLP revealed peaks characteristic of carbonate ions at 875 cm⁻¹ (ν2) and 1,430 cm⁻¹ (ν3). Phosphate peaks were also present at 566 cm⁻¹, 604 cm⁻¹ (ν4), and between 1,033–1,100 cm⁻¹ (ν3). The peak seen at 1,651 cm⁻¹ was due to the presence of water in the sample. (E and F) NB showing FTIR spectra similar to those of NLP. (E) NB cultured from DMEM containing 10% healthy HS, after one month. (F) “Nanons” after one week in DMEM without serum. Commercial powders of (G) CaCO₃, (H) Ca₃(PO₄)₂, and (I) HAP were included as controls. CaCO₃ produced an additional peak near 650 cm⁻¹ attributed to calcite that was not found in our experimental samples.