| With the development of society, aesthetics of the teeth is of great importance to individuals. An increasing number of people who are dissatisfied with the appearance of their teeth are seeking for esthetic treatment. Under the principle of saving tooth structure, the noninvasive vital tooth bleaching is widely used for normal discolored teeth. Although there is little doubt about the efficacy of tooth bleaching, the safety of this technique has been questioned. Based on our previous investigations, the first and second parts of this thesis evaluated the effects of commercial in-office bleaching agents with different pH values and at-home bleaching agents with different concentrations on the structure, components and color of tooth enamel in vitro and in situ. Attenuated total reflectance-infraed (ATR-IR) spectroscopy, Raman spectroscopy, atomic force microscopy (AFM), microhardness (MHS) test and color measurements were used for investigations before and after the bleaching procedure. The obtained results gave new evidences for the safety of bleaching agents. Moreover, it was found that the laser-induced fluorescence and tooth color always together changed during bleaching, which provide the inspiration for the origin of fluorescence within enamel.Besides the bleaching treatment for normal tooth, the invasive prosthetic treatment can also be chosen for abnormal tooth. However, choosing veneer or crown is determined by enamel structure. Inspired by the previous two parts, the third part of this thesis compared enamel structure and related mechanical property between normal enamel and hypomaturation enamel by using FTIR, Raman, micro-CT, microzone X-ray diffraction, scanning electron microscope and thermal gravimetric analysis techniques. The thorough comparisons between normal enamel and hypomaturation enamel explained clinical manifestations of hypomaturation enamel, provided fundamental evidences for dentists to choose optimal therapeutic strategy and deepened our understanding about fluorescence origin and biomineralization process. Moreover, these results provided inspirations for enamel regeneration and novel biomaterials design.Part I Effects of in-office bleaching agents with different pH on the color, structure and mechanical properties of enamel in situ and in vitro.Objectives:To investigate in situ and in vitro effects of in-office bleaching agents with different pH values on human enamel in terms of tooth color, chemical structure and mechanical property. There are two experiments in this part. Experiment I:both acidic and neutral in-office bleaching agents were used to bleach the enamel from the extracted tooth. After in-office bleaching procedure, the bleached enamel samples were placed into different storage environments (distilled water, artificial saliva or human oral cavity) to evaluate the effects of two in-office bleaching agents on the structure and mechanical properties of human enamel in vitro and in situ. Experiment Ⅱ:with the same bleaching mode from experiment I, effects of two in-office bleaching agents with different pH on the color and chemical component of human enamel were evaluated using in situ/in vitro methodologies.Experiment I Effects of two in-office bleaching agents with different pH on the structure and mechanical properties of enamel in situ and in vitro.Methods:Fifty-four freshly extracted orthodontic premolars were selected and stored in0.2%thymol at4℃until required. Two dental slabs (4mm x3mm×2mm) were obtained from the middle third of the buccal surface of each tooth using a low speed water-cooled diamond saw under water cooling. The obtained enamel slabs were randomly divided into9groups (n=12):group acidic bleaching agents-distilled water (DW), group acidic bleaching agents-artificial saliva (AS), group acidic bleaching agents-human saliva (HS), group neutral bleaching agents-DW, group neutral bleaching agents-AS, group neutral bleaching agents-HS, group control-DW, group control-AS and group control-HS. Samples with DW or AS storage were placed in vitro and samples with HS storage were fixed in the palatal part of bleaching tray and then placed in the volunteers’ oral cavities. The bleaching treatments were performed on the1st and8th day. Each in-office bleaching treatment contained "three bleaching episode" and each episode was lasted for15minutes. Baseline and final AFM surface detection, MHS and fracture toughness (FT) were measured before the treatment and on the15th day, respectively.Results:Compared with control groups, surface alterations on enamel were only found in Beyond+AS and Beyond+DW under AFM evaluation, which induced the significantly increased surface roughness in these two groups (p<0.001, p<0.001). No obvious changes of surface roughness were found in other groups (p>0.05). MHS and FT revealed no significant alteration between the baseline and final measurements in all groups (p>0.05).Conclusion:In-office bleaching agents with low pH values could induce enamel morphology alterations under in vitro conditions. However, with the protective effects of acquired membrane and remineralization ability of HS, the adverse effect of acidic bleaching agent was minimized in situ. The neutral bleaching agents did not induce the demineralization of enamel. Within the limitation of this study, in-office bleaching did not influence the MHS and FT of enamel.Experiment II Effects of in-office bleaching agents with different pH on the color, structure and mechanical properties of enamel in situ and in vitro.Methods:Forty-five freshly extracted orthodontic premolars were selected and stored in0.2%thymol at4℃until required. Two dental slabs (4mm x3mm x2mm) were obtained from the middle third of the buccal surface of each tooth using a low speed water-cooled diamond saw under water cooling. The obtained enamel slabs were randomly divided into6groups (n=15):group acidic bleaching agents-DW, group acidic bleaching agents-HS, group neutral bleaching agents-DW, group neutral bleaching agents-HS, group control-DW and group control-HS. Samples with DW storage were placed in vitro and samples with HS storage were fixed in the palatal part of bleaching tray and then placed in the volunteers’oral cavities. The bleaching treatments were performed on the1st and8th day. Each in-office bleaching treatment contained "three bleaching episode" and each episode was lasted for15minutes. Baseline and final ATR-IR spectroscopy, Raman spectroscopy and color measurement were performed before the bleaching treatment and after one week post-treatment, respectively.Results:V2CO32-: V1V3PO43-in ATR-IR spectrum and percentage Relative Raman Intensity (RRI) in Raman spectrum showed significantly decrease in group Beyond+DW (p<0.001, p<0.001) while little variation was observed in the other groups (p>0.05, p>0.05). Percentage fluorescence intensity (FI) and AE revealed statistical difference in all bleached groups (p<0.001, p<0.001) while no statistical difference in control groups (p>0.05, p>0.05).Conclusion:Acidic and neutral in-office bleaching agents had the same whitening efficiency in situ and in vitro. Acidic agents could induce demineralisation of human enamel in vitro and the presence of natural human saliva could minimise this adverse effect.Part Ⅱ Effects of at-home bleaching agents with different concentrations on the structure, component and mechanical properties of enamel in situ and in vitro.Objectives:To investigate in situ and in vitro effects of at-home bleaching agents with different concentrations on human enamel in terms of tooth structure, chemical structure and mechanical property. There are two experiments in this part. Experiment I:at-home bleaching agents with different concentrations were used to bleach the tooth enamel from the extracted tooth. After bleaching procedure, the bleached enamel samples were placed into DW to evaluate the effects of at-home bleaching agents with different concentrations on the structure, chemical component and mechanical properties of human enamel in vitro and in situ. Experiment Ⅱ:with the same bleaching mode from experiment Ⅰ, effects of at-home bleaching agents with different concentrations on the structure, chemical component and mechanical properties of enamel were evaluated using in situ/in vitro methodologies.Experiment III Effects of three at-home bleaching agents with different concentrations on the structure, component and mechanical properties of enamel in vitro.Methods:Thirty freshly extracted orthodontic premolars were selected and stored in0.2%thymol at4℃until required. Two dental slabs (4mm×3mm×2mm) were obtained from the middle third of the buccal surface of each tooth using a low speed water-cooled diamond saw under water cooling. Sixty obtained enamel specimens were randomly divided into four groups and treated with10%carbamide peroxide (CP),15%CP,20%CP and DW, respectively. The bleaching treatment was8h/day. Subsequently, enamel samples were stored in DW for the other14h/day. The whole experiment lasted for14consecutive days. Baseline and final AFM surface detection, Raman spectroscopy, ATR-IR spectroscopy, MHS and FT measurements were carried out before and after bleaching experiments.Results:CP didn’t change the morphology of enamel. Meanwhile, the three bleached groups and the control group had no significant difference in RMS detection (p=0.774), v2CO32":v1v3PO43-(p=0.263), RRI percentage (p=0.062) and MHS (p=0.829). However, the percentage of laser-induced fluorescence in the three bleached groups decreased significantly when compared with the control group (p<0.001, p<0.001, p<0.001). Moreover, FT declined significantly in the three groups when compared with the control group (p=0.024, p=0.005, p=0.013).Conclusion:Under in vitro condition, three differently concentrated at-home bleaching agents wouldn’t induce the demineralization and the decline of microhardness on enamel. However, the decrease of FT on enamel seemed to be inevitable after bleaching.Experiment IV Effects of three at-home bleaching agents with different concentrations on the structure, component and mechanical properties of enamel in situ.Methods:Thirty freshly extracted orthodontic premolars were selected and stored in0.2%thymol at4℃until required. Two dental slabs (4mm×3mm×2mm) were obtained from the middle third of the buccal surface of each tooth using a low speed water-cooled diamond saw under water cooling. Sixty obtained enamel specimens were randomly divided into four groups and fixed at palatal part of bleaching tray. Bleaching groups were treated with10%CP,15%CP and20%CP and the control group was applied with DW to make the enamel surface keep moist. The bleaching treatment was8h/day and then the bleaching agents were removed by DW. Subsequently, the bleaching tray were placed at the volunteers’oral cavity and stored for the other16hours. The whole experiment lasted for14consecutive days. Baseline and final AFM surface detection, Raman spectroscopy, ATR-IR spectroscopy, MHS and FT measurements were carried out before and after bleaching experiments.Result:CP didn’t change the morphology of enamel. Meanwhile, the three bleached groups and the control group had no significant difference in RMS detection (p0.784), v2CO32-:v1v3PO43-(p=0.917), RRI percentage(p=0.187) and MHS (p=0.935). However, the percentage of laser-induced fluorescence in the three bleached groups decreased significantly when compared with the control group (p<0.001, p<0.001, p<0.001). Moreover, FT declined significantly in the three groups when compared with the control group (p=0.028. p=0.015. p=0.019).Conclusion:Under in situ condition, three differently concentrated at-home bleaching agents wouldn’t induce the demineralization and the decline of microhardness on enamel. However, the decrease of FT on enamel seemed to be inevitable after bleaching even the acquired membrane could protect the enamel surface.Part Ⅲ Structural, compositional and mechanical comparison of normal enamel and hypomaturation enamelExperiment V Structural, compositional and mechanical comparison of normal enamel and hypomaturation enamelMethods:Under a protocol approved by the local Ethics Committee of the School and Hospital of Stomatology, Wuhan University, PR China, three human permanent molars (two from maxillary and one from mandibular) diagnosed with hypomaturation amelogenesis imperfecta were obtained with informed consent from patients. And for minimizing the variation of teeth location, three human permanent normal molars from same location were chosen as control ones. All teeth were cleaned thoroughly and then stored in0.2%thymol solution at4℃until use. After removing the teeth roots with a low speed water-cooled diamond saw under water cooling, micro-CT was used to scan and reconstruct the3D model of tooth crown. Subsequently, each crown was longitudinally sectioned into two halves. Fourier transform infrared (FTIR), Raman, microzone X-ray diffraction (microzone XRD), thermal gravimetric analysis (TGA), scanning electron microscope (SEM)/energy diffraction spectrum (EDS), MHS measurements were used to analyze the structure, component, mechanical property of normal enamel and hypomaturation enamel.Result:From FTIR spectra, hypomaturation enamel showed significantly higher matrix:mineral ratio and relative carbonate content but significantly lower crystallinity than normal enamel (p<0.001, p<0.001and p<0.001, respectively). From Raman spectra, hypomaturation enamel showed significantly lower RRI but significantly higher full width at half maximum (FWHM) and FI than normal enamel (p<0.001, p<0.001and p<0.001, respectively). In addition, normal enamel displayed clear tendencies of FWHM and RRI from surface towards the dentin-enamel junction, whereas hypomaturation enamel not. A closer look found that FWHM was inversely correlated with RRI in normal enamel. When RRI increases, FWHM decreases progressively (r=-0.717, p<0.001). However, no significant correlation was found in hypomaturation enamel between FWHM and RRI (r=0.173, p=0.256). Microzone XRD showed hypomaturation enamel had samaller crystallite size than normal enamel (p=0.007). TGA result revealed that the hypomaturation enamel showed higher weight loss than normal enamel from200℃to600℃. Micro-CT images showed hypomaturation enamel had thin thickness and denuded surface at the macroscale; SEM images showed that hypomaturation enamel had porous structure, widened interprismatic gaps and even the lack of enamel prisms at the macroscale and randomly and loosely arrayed crystals at the nanoscale. MHS test revealed that hypomaturation enamel had significantly lower MHS than normal enamel.Conclusion:Side-by-side comparisons between hypomaturation enamel and normal enamel revealed that the retention of organic matrix influenced the quantity, quality and distribution of mineral crystals in enamel, which further affected the hierarchical architecture and the corresponding mechanical property of enamel. |
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