Biochemical characterization of human gingival crevicular fluid during orthodontic tooth movement using Raman spectroscopy

Abstract

This study used Raman spectroscopy to report the first human gingival crevicular fluid (GCF) biochemical characterization during the early phase of orthodontic tooth movement. This technique allows for label-free and noninvasive biochemical change monitoring in GCF during orthodontic tooth movement. Ten orthodontic patients (20.8 ± 2.5 years) participated in the study. GCF samples were obtained before (baseline, 0 days) and during orthodontic treatment at 1, 7 and 28 days. For Raman spectroscopic measurement, GCF samples (5 µl) were deposited onto a gold-coated substrate, then dried at room temperature. Raman spectra GCF analysis during orthodontic treatment indicated that the hydroxyapatite to primarily collagen-dominated matrix band (phosphate 984 cm(-1)/amide I 1667 cm(-1)) intensity ratio decreased at day 7 (P < 0.05). The carbonate apatite to hydroxyapatite ratio (carbonate 1088 cm(-1)/phosphate 984 cm(-1)) was significantly higher on day 7 compared to day 0 (P < 0.05). These results indicate that demineralization occurs during the alveolar bone remodeling process. We also found notable peak shifts in the amide I range during orthodontic tooth movement. The 1658 cm(-1) in baseline red shifted to 1667 cm(-1) at orthodontic treatment day 7. Curve fitting in the amide I (1615-1725 cm(-1)) range demonstrated that increased random coil conformation was accompanied by a decrease in β-sheet structure during orthodontic tooth movement. Thus, we suggest Raman spectroscopy could be used for label-free, non-invasive GCF quality assessment during orthodontic tooth movement. Furthermore, this method may prove to be a powerful diagnostic and prognostic tool for monitoring orthodontic tooth movement in a clinical setting.

Keywords: (170.0170) Medical optics and biotechnology; (170.1610) Clinical applications; (170.1850) Dentistry; (170.5660) Raman spectroscopy.

Fig. 1

Collection procedures for gingival crevicular fluid (GCF). (a) Prophylaxis, (b) drying the target area, (c) GCF collection before orthodontic force loading, (d) GCF collection after orthodontic force loading, (e) paper point with absorbed GCF, and (f) paper point in Eppendorf tubes.

Fig. 2

Normalized average GCF Raman spectrum without orthodontic treatment

Fig. 3

Normalized average Raman spectra for control GCF (0 d) and early-phase GCF tooth movement at day 1, 7 and 28 (1 d, 7 d and 28 d).

Fig. 4

Hydroxyapatite to collagen (phosphate 984 cm⁻¹/ amide I 1667 cm⁻¹) ratio in GCF during orthodontic treatment. *Significantly different from day 0 (P < 0.05, one-way ANOVA with Turkey’s HSD post-hoc procedure)

Fig. 5

Carbonate apatite to collagen (carbonate 1088 cm⁻¹/ amide I 1667 cm⁻¹) ratio in GCF during orthodontic treatment. *Significantly different from day 0 (P < 0.05, one-way ANOVA with Turkey’s HSD post-hoc procedure)

Fig. 6

Carbonate apatite to hydroxyapatite (carbonate 1088 cm⁻¹/ phosphate 984 cm⁻¹) ratio in GCF during orthodontic treatment. *Significantly different from day 0 (P < 0.05, one-way ANOVA with Turkey’s HSD post-hoc procedure)

Fig. 7

(A) Relative intensities in 1667 cm⁻¹ Raman peaks during orthodontic treatment. *Significantly different from day 0 (P < 0.05, one-way ANOVA with Turkey’s HSD post-hoc procedure). (B) Normalized average GCF Raman spectra in the 1625-1715 cm⁻¹ range during orthodontic treatment.

Fig. 8

Normalized average GCF Raman spectra curve-fitting during orthodontic tooth movement (0, 1, 7 and 28 days) in the 1625-1715 cm⁻¹ range. The black lines represent experimental data. The blue and red lines illustrate fitted peaks and summation of fitting curves, respectively.