Quantification of minerals associated to enamel caries process by raman spectroscopy

Abstract

Background: Stimulated Raman spectroscopy (SRS) is a nondestructive tool for biochemical characterization of tissues. The aims were: 1) To evaluate the ability of SRS and Spontaneous Raman spectroscopy (SpRS) to differentiate among sound, demineralized and remineralized bovine enamel by phosphate and carbonate ratio (P/C-Ratio); and 2) To determine the correlation between the outcomes of transverse microradiography (TMR: Integrated mineral loss (ΔZ) and lesion depth) and P/C-Ratio. Material and Methods: Thirty, 5×5×2-mm ground and polished bovine enamel blocks were prepared. The surface was divided into 3 equal areas. Each area was chemically demineralized (demin) by Carbopol demineralized solution for 0 (Sound-Demin), 24 (24h-Demin), and 48h (48h-Demin), respectively. Then, specimens were sectioned for TMR analysis, and the remaining one part of each specimen was remineralized (remin) for 15days using a pH-cycling model (Sound-Remin, 24hD-Remin=24h-Demin and remineralization, 48hD-Remin = 48h-Demin and remineralization). Demin and remin groups were scanned to obtain P/C-Ratio by SpRS and SRS. SRS was further scanned from 0 (surface) up to 100 µm into the dentine at 10-µm intervals. Remineralized specimens were sectioned for TMR analysis. Wilcoxon signed-rank tests were used to compare between TMR and SpRS/SRS. Spearman correlation coefficients were used to correlate among TMR, SpRS, and SRS. A 5-percent significance level was used for each test. Results: As demin time increased, both ΔZ and lesion depth were increased. After remineralization, both values were decreased. There were significant differences between demine and remin groups and between demin times. For SpRS, Sound-Demin had significantly larger P/C-Ratio than 24h-Demin and 48h-Demin (p ≤ 0.001). The 24h-Demin had significantly larger values than 48h-Demin (p = 0.048). Sound-Remin had larger P/C-Ratio than 24hD-Remin (p = 0.316) and 48hD-Remin (p = 0.015). 24hD-Remin was larger than 48hD-Remin (p = 0.269). 24hD-Remin had significantly larger P/C-Ratio than 24h-Demin (p ≤ 0.001). 48hD-Remin had significantly larger P/C-Ratio than 48h-Demin (p ≤ 0.001). For SRS, at surface (0 µm), for demin group, Sound-Demin had significantly larger P/C-Ratio than 24h-Demin (p = 0.020) and 48h-Demin (p = 0.032). 24h-Demin had larger value than 48h-Demin; but no significant difference (p = 0.117). Among remin groups, Sound-Remin was not statistical significance different for 24hD-Remin (p = 0.172) and 48hD-Remin (p = 0.134). However, 24hD-Remin was smaller; but not statistical significance different from 48hD-Remin (p = 0.688). At deeper levels (10 µm to 100µm), it was found that 1) After demineralization, Sound-Demin had significantly larger P/C-Ratio than 24h-Demin and 48h-Demin at 0 µm to 20 µm, and 80 µm to 100µm; Sound-Demin had significantly larger P/C-Ratio than 48h-Demin; and no statistical significance differences were found among Sound-Demin and 24h-Demin, 24h-Demin and 48h-Demin. 2) After remineralization, no statistical significance differences were found among Sound-Remin, 24hD-Remin, and 48hD-Remin. 3) Sound-Demin had significantly larger P/C-Ratio than Sound-Remin at 0 µm ,10 µm, 20 µm; and no statistical significance differences were found at levels deeper than 30 µm. 4) No statistical significance differences were found between 24h-Demin and 24hD-Remin from 0 µm to 70µm; and 24hD-Remin had significantly larger P/C-Ratio than 24h-Demin from 80 µm to 100 µm. 5) No statistical significance differences were found between 48h-Demin and 48hD-Remin. For correlation, moderate correlation was found between SpRS demineralized/remineralized groups and ΔZ, and between SpRS demineralized groups and lesion depth. Conclusion: SpRS and SRS have the potential to quantify demineralization through calculation of the phosphate and carbonate ratio. In addition, SpRS can detect the change of remineralization. A nondestructive caries detection approach using SpRS and SRS would be beneficial in clinical practice.