STEM and HRTEM studies of accumulated deposits on human tooth surface

The aim of this study was to clarify the fine structure of accumulated deposits on the surface of teeth that are considered to affect the gloss of teeth. The study was carried out using, as specimens, human incisor teeth having gloss, which were extracted from teenage donors and those incapable of showing gloss even by brushing which were extracted from donors in their 50s. Thin longitudinal sections of tooth enamel with accumulated deposits on the surface were prepared by focused ion beam (FIB) milling, and the fine structure was analyzed using a scanning transmission electron microscope (STEM) and a high resolution transmission electron microscope (HRTEM). By FIB, thin longitudinal sections could be prepared from tooth enamel together with organic and inorganic substances accumulated on the surface without artifacts. The accumulated deposits on the surface of teeth having gloss were composed of organic substances. However, it was first revealed by STEM observation that the accumulated solid deposits on the surface of teeth having no gloss had a complicated structure wherein inorganic and organic substances coexisted. It is suggested that the organic substances contain proteins derived from saliva. The inorganic substances were spherical and needle‐like hydroxyapatites (HAs). It is considered that amino acids constituting the proteins affected the nucleus formation and the crystal formation of HA. It is considered that the unevenness of the accumulated deposits existing on the surface of tooth enamel having no gloss causes the decrease in gloss of teeth due to diffuse reflection of light.     

Microstructure and mineral composition of dental enamel of permanent and deciduous teeth

Purpose: This study evaluated and compared in vitro the microstructure and mineral composition of permanent and deciduous teeth's dental enamel. Methods: Sound third molars (n = 12) and second primary molars (n = 12) were selected and randomly assigned to the following groups, according to the analysis method performed (n = 4): Scanning electron microscopy (SEM), X‐Ray diffraction (XRD) and Energy dispersive X‐ray spectrometer (EDS). Qualitative and quantitative comparisons of the dental enamel were done. The microscopic findings were analyzed statistically by a nonparametric test (Kruskal‐Wallis). The measurements of the prisms number and thickness were done in SEM photomicrographs. The relative amounts of calcium (Ca) and phosphorus (P) were determined by EDS investigation. Chemical phases present in both types of teeth were observed by the XRD analysis. Results: The mean thickness measurements observed in the deciduous teeth enamel was 1.14 mm and in the permanent teeth enamel was 2.58 mm. The mean rod head diameter in deciduous teeth was statistically similar to that of permanent teeth enamel, and a slightly decrease from the outer enamel surface to the region next to the enamel‐dentine junction was assessed. The numerical density of enamel rods was higher in the deciduous teeth, mainly near EDJ, that showed statistically significant difference. The percentage of Ca and P was higher in the permanent teeth enamel. Conclusions: The primary enamel structure showed a lower level of Ca and P, thinner thickness and higher numerical density of rods. Microsc. Res. Tech., 2010. © 2009 Wiley‐Liss, Inc.     

Effect of laser preparation on adhesion of a self‐adhesive flowable composite resin to primary teeth

The aim of the study was to evaluate the adhesion of a self‐adhering flowable composite resin to primary tooth enamel and dentin after silicon carbide paper (SiC) and laser pretreatment. Adhesive properties were evaluated as shear bond strength (SBS) and scanning electron microscopic (SEM) characteristics. A total 120 primary canine teeth were randomly divided into two groups to study enamel and dentin. Each group was divided into 6 subgroups (n = 10) according to type of surface preparation (SiC or Er:YAG laser) of enamel or dentin. Three methods were used to build cylinders of restoration on tooth surface: OptiBond All‐In‐One + Premise Flowable composite, OptiBond All‐In‐One + Vertise Flow and Vertise flow. After restoration, samples were tested for SBS and failure mode. Twenty eight samples were examined by SEM. The results of the study showed SBS of Vertise Flow was lower than others in enamel and dentin samples pretreated with SiC and in dentin samples pretreated with laser (P < 0.001). Compared to SiC pretreatment, laser pretreatment led to a significantly higher SBS with Vertise Flow on enamel (P < 0.001). Vertise Flow associated with the adhesive led to a higher SBS in enamel and dentin compared to Vertise Flow alone. Adhesive and mixed failure modes were observed more frequently in Vertise Flow groups. SEM images showed that Vertise Flow led to more irregularities on enamel and more open dentinal tubules after laser ablation compared SiC pretreatment. Microsc. Res. Tech. 79:334–341, 2016. © 2016 Wiley Periodicals, Inc.     

Enamel alteration following tooth bleaching and remineralization

The purpose of this study was to compare the effects of professional tooth whitening agents containing highly concentrated hydrogen peroxide (with and without laser activation), on the enamel surface; and the potential of four different toothpastes to remineralize any alterations. The study was performed on 50 human molars, divided in two groups: treated with Opalescence^® Boost and Mirawhite^® Laser Bleaching. Furthermore, each group was divided into five subgroups, a control one and 4 subgroups remineralized with: Mirasensitive^® hap+, Mirawhite^® Gelle?, GC Tooth Mousse? and Mirafluor^® C. The samples were analysed by SEM/3D‐SEM‐micrographs, SEM/EDX‐qualitative analysis and SEM/EDX‐semiquantitative analysis. The microphotographs show that both types of bleaching cause alterations: emphasized perikymata, erosions, loss of interprizmatic substance; the laser treatment is more aggressive and loss of integrity of the enamel is determined by shearing off the enamel rods. In all samples undergoing remineralization deposits were observed, those of toothpastes based on calcium phosphate technologies seem to merge with each other and cover almost the entire surface of the enamel. Loss of integrity and minerals were detected only in the line‐scans of the sample remineralized with GC Tooth Mousse?. The semiquantitative EDX analysis of individual elements in the surface layer of the enamel indicates that during tooth‐bleaching with HP statistically significant loss of Na and Mg occurs, whereas the bleaching in combination with a laser leads to statistically significant loss of Ca and P. The results undoubtedly confirm that teeth whitening procedures lead to enamel alterations. In this context, it must be noted that laser bleaching is more aggressive for dental substances. However, these changes are reversible and can be repaired by application of remineralization toothpastes.     

Evaluation of the effect of food and beverages on enamel and restorative materials by SEM and Fourier transform infrared spectroscopy

Objectives To examine different types of restorative materials used in children as well as primary and permanent teeth enamel when affected by erosive foods. Materials and Method Buttermilk, fruit yoghurt, Coca‐cola, fruit juice, Filtek Z‐250, Dyract Extra, Fuji II LC, and Fuji IX and tooth enamel were used. Measurements were performed on 1‐day, 1‐week, 1‐month, 3‐month, 6‐month time periods by using ATR‐FTIR technique and surface of the specimens were examined with SEM. Results Permanent tooth showed the least change among human tooth samples when compared to restorative materials. Among filler materials, the most change was observed in Fuji IX. In terms of beverages the most changes on absorption peaks obtained from spectra were seen on the samples held in Coca‐Cola and orange‐juice. Conclusion The exposure of human enamel and restorative materials to acidic drinks may accelerate the degradation process and so reduce the life time of filler materials at equivalent integral exposure times longer than three months. Clinical Relevance Erosive foods and drinks having acidic potential destroy not only tooth enamel but also restorative materials. Microsc. Res. Tech. 77:79–90, 2014. © 2013 Wiley Periodicals, Inc.     

Microhardness and SEM after CO2 laser irradiation or fluoride treatment in human and bovine enamel

Background: It remains uncertain as to whether or not CO₂ laser is able to hinder demineralization of enamel. The possibility to use bovine instead of human teeth on anticariogenic studies with laser has not yet been determined. Purpose: To compare the ability of CO₂ laser and fluoride to inhibit caries‐like lesions in human enamel and to test whether a similar pattern of response would hold for bovine enamel. Study Design: Ninety‐six enamel slabs (2 × 2 × 4 mm) (48 from bovine and 48 from human teeth) were randomly distributed according to surface treatment (n = 12): CO₂ laser, 5% sodium fluoride varnish (FV), 1.23% acidulated phosphate fluoride (APF) gel, or no treatment (control). Specimens were subjected to a 14‐day in vitro cariogenic challenge. Microhardness (SMH) was measured at 30 μm from the surface. For ultrastructural analysis, additional 20 slabs of each substrate (n = 5) received the same treatment described earlier and were analyzed by SEM. Results: ANOVA and Tukey test ascertained that CO₂ laser promoted the least mineral loss (SMH = 252^a). Treatment with FV resulted in the second highest values (207^b), which was followed by APF (172^c). Untreated specimens performed the worst (154^d). SEM showed no qualitative difference between human and bovine teeth. APF and control groups exhibited surfaces covered by the smear layer. A granulate precipitate were verified on FV group and fusion of enamel crystals were observed on lased‐specimens. Conclusions: CO₂ laser may control caries progression more efficiently than fluoride sources and bovine teeth may be a suitable substitute for human teeth in studies of this nature. Microsc. Res. Tech. 73:1030–1035, 2010. © 2010 Wiley‐Liss, Inc.     

Thermal,morphological, and spectral changes after Er,Cr:YSGG laser irradiation at low fluences on primary teeth for caries prevention

The aim of this study was to determine the temperature increase in the pulp chamber and possible thermal effects on molecular structure of primary teeth during the irradiation with Er,Cr:YSGG laser. Primary central incisors were divided into three groups (n = 20). Labial surfaces in each group were irradiated by Er,Cr:YSGG laser within different power and frequencies as following groups: I: 0.25 W, 20 Hz, II: 0.50 W, 20 Hz, III: 0.75 W, 20 Hz. A thermocouple was placed inside the pulp chamber so that the temperature increments were recorded during the enamel irradiation. Morphological changes of enamel surfaces were experimentally evaluated by SEM. Fourier‐transform infrared spectroscopy and RAMAN analyses were carried out to determine the differentiations in the molecular structure. The experimental results obtained were analyzed statistically by means of one‐way analysis of variance. Statistically significant differences were detected between groups (p < .05). Group III exhibited the highest values for the temperature parameters. Besides, the conical craters, cracks, and formation of ablation areas were observed for all the groups. Also, it was obtained that the hydroxyapatite lost the hydroxyl ions due to the thermal effect of the laser. Temperature rise throughout the Er,Cr:YSGG laser irradiation for prevention of primary enamel demineralization presented a positive correlation with the laser output power level. The formations of adverse morphological and spectral changes were detected on the surface of teeth after the laser application. On this basis, the Er,Cr:YSGG laser applications should be treated with much more caution considering enamel surface and pulpal tissues in primary teeth.     

Wear of dental enamel

Teeth wear for a number of reasons. One of the causes for wear is the use of abrasive tooth pastes. A study is reported in which enamel was damaged by single pass sliding of a diamond indenter. In single crystals of fluorapatite, wear occurs by flaking out of chips of material and severely fractured substrate is left behind. On the other hand teeth are made of small rods of polycrystalline hydroxyapatite of the order of 5 μm in diameter oriented perpendicular to the tooth surface. This rod structure of human teeth prevents large scale flaking out of material. Apparently the subsurface cracks do not extend from one rod to the other so that subsequent passes do not result in severe wear. Small abrasive particles may do considerable damage to rods, but the overall effect is small. Large abrasive particles with high loads cause separation of rods, and cracks appear to propagate but without severe permanent damage.     

Resin-tooth interfacial morphology and sealing ability of one-step self-etch adhesives: microleakage and SEM study

OBJECTIVES: To compare microleakage of three self‐etch adhesives and to analyze enamel surface morphology and interfacial morphology of resin–enamel and resin–dentin interface under scanning electron microscope (SEM). EXPERIMENTAL DESIGN: Study was conducted in 65 extracted human premolars. Class V cavities were prepared in 45 teeth and assigned to three groups (n = 15) according to three self‐etch adhesives (OptiBond All‐in‐One, iBond, and Adper Prompt L‐Pop). After restoration, 10 samples from each group were used to assess microleakage at enamel and dentin margin. Five samples from each group were used for analysis of interfacial morphology at resin–enamel and resin–dentin interface under SEM. Remaining 20 teeth were used to prepare flat enamel buccal surfaces to analyze the difference in surface morphology after treatment with three adhesives (n = 5 each) and 36% phosphoric acid treatment (n = 5). PRINCIPAL OBSERVATIONS: At enamel margin, Prompt L‐Pop depicted least leakage of all the three adhesives and also showed best interfacial adaptation under SEM. At dentin margin, OptiBond All‐in‐One showed significant less leakage than iBond and Prompt L‐Pop. On flat enamel surface, phosphoric acid produced the most retentive etching pattern when compared with the three adhesives. CONCLUSION: Prompt L‐Pop showed the best bonding effectiveness in enamel, whereas OptiBond All‐in‐One performed significantly better in dentin. Microsc. Res. Tech. 2012. © 2012 Wiley Periodicals, Inc.     

High resolution SE-I SEM study of enamel crystal morphology

Until recently high resolution TEM was the only imaging mode capable of probing the atomic lattice structure of crystals composing tooth enamel. Studies designed to determine the polyhedral shape of normal enamel crystals and initiation of carious lesions in enamel crystals were hampered and limited by interpretation of two-dimensional TEM images from thin section and freeze fracture replica specimens lacking depth of field. The newly developed SE-I signal mode for SEM (SE-I/SE-II ratio) can produce images of enamel crystals approaching beam diameter dimensions (0.7–2.0 nm), rivaling the resolution of the TEM technique and generating topographic contrasts for three dimensional imaging at very high magnification (≈?1,000,000 X). Ultrathin chromium (Cr) films generate enriched high resolution SE-I contrasts of enamel crystal surfaces and when imaged using an immersion lens field emission SEM operated at high voltage (20–30 KeV) produce unsurpassed topographic contrasts. Since the grain size of Cr is below the resolution of any SEM and is ultrathin (≈?1 nm), then SE-I images can provide a more accurate representation of enamel crystal structure than TEM methodologies. Our SE-I SEM observations of normal human enamel crystals reveal fractured spicules which contain angled flat surfaces delineated by a prominent 2 nm wide SE-I edge brightness contrast. Although microscopic observations often show crystals which are hexagonal in cross-section, in both SEM and TEM many other growth habits, including rectangular or irregular crystals (30–40 nm in width) which contain “notches,” are also observed. More detailed morphological studies are therefore required to determine the most likely habit planes and their relevance to the function of the enamel crystals. The granular appearing fine structural contrast imposed onto <100> lattice planes of sectioned enamel in TEM micrographs is also resolved with topographic contrasts in SE-I micrographs. These granules probably represent one or both of the enamel protein classes.     

Active application of primer acid on acid‐treated enamel: Influence on the bond effectiveness of self‐etch adhesives systems

Aim: Evaluate the composite‐to‐enamel bond after passive or active application of self‐etching primer systems on polished or pre‐etched enamel with phosphoric acid. Materials and Methods: Two self‐etch adhesives systems (SEAS) were used: Clearfil SE Bond and Easy Bond. Third human molars were divided into 8 groups (N = 10). The crown of each tooth was sectioned into halves and the mesial/distal surfaces were used. The adhesives were actively or passively applied on enamel with or without prior phosphoric‐acid etching. Resin composite cylinders were built after adhesive application. After stored in relative humidity for 24 hr/37°C the specimens were subjected to microshear test in universal testing a machine at a crosshead speed of 0.5 mm/minute. The results were analyzed with three‐way ANOVA and the Tukey test. The enamel‐etching pattern was evaluated under SEM. Results: The 2‐step SEAS system presented significantly higher adhesive bond strength means (47.37 MPa) than the 1‐step (36.87 MPa). A poor enamel‐ etching pattern was observed in active mode showing irregular and short resin tags, however there was not compromised the bond strength. Conclusion: Active or passive application produced similar values of bond strength to enamel regardless of enamel pretreatment and type of SEAS.     

Comparative analysis of dental enamel polyvinylsiloxane impression and polyurethane casting methods for SEM research

Dental casting is a very common procedure for making high-quality replicas of paleo-anthropological remains. Replicas are frequently used, instead of original remains, to study both fossil and extant Primate teeth in morphological and metrical analyses. Several commercial products can be used in molds. This study analyzed SEM image resolution and enamel surface feature definition of tooth molds at various magnification levels and obtained, with both Coltène and 3M low-viscosity body polyvinylsiloxane impression, materials and polyurethane casts. Results, through comparison with the original teeth, show that both the negative molds and the positive casts are highly reliable in replicating enamel surfaces. However, positive cast quality is optimal for SEM observation only till the fourth consecutive replica from the original mold, especially at high SEM magnification levels.     

Friction and wear behavior of human teeth under various wear conditions

Previous reports have described the differences in the friction and wear behavior between different zones of human teeth. The objective of this research was to study the friction and wear behavior of human teeth under different wear conditions to extend the understanding of the tooth wear process, as well as to provide a more rational explanation for wear mechanism of teeth. Two typical wear tests, namely two- and three-body wear, were conducted on human tooth enamel using a reciprocating apparatus. The effect of food particles was of particular interest. Three loads, 10, 20 and 40 N, were used. Wear was assessed by sample wear volume. The results show that human tooth enamel exhibits lower friction and smaller wear volume under three-body wear conditions than under two-body wear conditions. Under three-body wear conditions, although increasing normal load results in a progressive increase in the wear volume of enamel, the increasing rate is lower at high load than that under two-body wear conditions. Further analysis of wear surfaces indicates that human tooth enamel experiences different wear mechanisms under different wear conditions.     

Marginal adaptation of composite resins under two adhesive techniques

In the present research, different adhesive techniques were used to set up fillings with composite resins. After the application of etch and rinse or self etch adhesive technique, marginal adaptation of composite fillings was estimated by the length of margins without gaps, and by the microretention of resin in enamel and dentin. The study material consisted of 40 extracted teeth. Twenty Class V cavities were treated with 35% phosphorous acid and restored after rinsing by Adper Single Bond 2 and Filtek Ultimate‐ASB/FU 3M ESPE composite system. The remaining 20 cavities were restored by Adper Easy One‐AEO/FU 3M ESPE composite system. Marginal adaptation of composite fillings was examined using a scanning electron microscope (SEM). The etch and rinse adhesive technique showed a significantly higher percentage of margin length without gaps (in enamel: 92.5%, in dentin: 57.3%), compared with the self‐etch technique with lower percentage of margin length without gaps, in enamel 70.4% (p < .001), and in dentin‐22.6% (p < .05). In the first technique, microretention was composed of adhesive and hybrid layers as well as resin tugs in interprismatic spaces of enamel, while the dentin microretention was composed of adhesive and hybrid layers with resin tugs in dentin canals. In the second technique, resin tugs were rarely seen and a microgap was dominant along the border of restoration margins. The SEM analysis showed a better marginal adaptation of composite resin to enamel and dentin with better microretention when the etch and rinse adhesive procedure was applied.     

X‐ray microdiffraction,TEM characterization and texture analysis of human dentin and enamel

Human tooth is a complex bioceramic composite, which consists of enamel, dentin and the interface, the dentin–enamel junction (DEJ). The crystal properties and ultrastructure of the inorganic phase through the thickness of healthy human molar teeth were investigated using X‐ray microdiffraction (μXRD), electron diffraction and transmission electron microscopy (TEM) techniques. The XRD data were analysed using the Le Bail profile fitting approach. The size and the texture of the crystallites forming enamel and dentin in the crown part of teeth were measured using both techniques and then compared. Results showed that the thickness of dentin crystallites was found to decrease towards the DEJ, whereas the thickness of the enamel crystallites increased from the DEJ towards the outer layers. It was demonstrated that enamel exhibited an increase of texture in 002 lattice planes from the DEJ towards the outer layers. Texture was also detected in 102 lattice planes. The texture effect in 002 planes at the scale of less than 1 μm was also demonstrated in dentin. The variation of lattice parameters as a function of the position within the thickness of dentin and enamel was also observed. The values of the nonuniform microstrain in the dentin and enamel crystallites were from 1.40 × 10^?6% to 4.44 × 10^?5%. The good correlation between XRD and TEM indicated that μXRD is a useful technique to study crystallography and microstructure of heterogeneous enamel and dentin. The observed gradient characteristics of texture and crystallite size in enamel and dentin maybe an evolutionary outcome to resist wear and fracture, thereby contributing to the excellent mechanical properties of teeth.     

天然牙釉质/纯钛摩擦磨损行为的研究

对天然牙釉质与TA2纯钛球组成的摩擦副进行了实验研究，结合显微分析，研究了牙釉质的摩擦磨损行为。结果表明牙冠外层釉质的摩擦学特性与釉质的基本结构釉柱的排列方向密切相关，釉质在He面上的耐磨性明显优于垂直方向。     

On the method of revealing enamel structure by acid etching. Aspects of optimization and interpretation

The study aimed at finding an optimal combination of acid concentration and etching time when nitric acid is used as etchant for the study of the finer details of human dental enamel structure. Four hundred 2–3‐mm‐thick segments of facio‐lingually sectioned human third molar crowns were assigned to 20 groups with 20 specimens in each group, each group differing with respect to acid concentration (0.1, 1, 2.5, and 5%) and etching time (15, 30, 45, 90, and 180 s). After etching and preparation, specimens were observed in the scanning electron microscope (SEM). Surface roughness/topography increased with increasing acid concentration and increasing etching time, but not in a linear fashion; generally, prisms tended to go from flat‐surfaced to cone‐shaped and prism sheaths from fissure‐like to wedge‐shaped. Intragroup variations and intergroup similarities were considerable. The two major enamel factors determining the etch effect are crystal orientation and prism sheath properties. Other factors, such as distribution of porosities and crystal quality, also contribute probably. Slight to moderate topography is best for observing the finer enamel structure, for example, etching with concentrations in the range 0.1–1% and with etching times in the range 15–90 s, the stronger the acid, the shorter the time. The depth effect of nitric acid is judged to be relatively small. Considerable variations in expression of prism cross‐striations were observed. SEM observations of acid‐etched enamel in carefully selected planes are a powerful method for the study of enamel structure, bearing in mind the artifactual aspects of the observed surface.     

The relation between structural,rugometric and fractal characteristics of hard dental tissues at micro and nano levels

Human tooth exhibits a structure of a mixture of inorganic hydroxyapatite nanocrystals and organic phases. The aim of this study is to investigate different tissues of human canine teeth surface along with the micro structure parameters of each tissue. X‐ray diffraction (XRD) is used to study the amorphous or crystalline nature of each tissue with different mineral compositions and crystalline structures where the highest crystalline quality is related to enamel. The surfaces are also examined by energy‐dispersive X‐ray spectrometry. Moreover, crystalline quality factor is carried out to estimate the crystallinity of the tissues. Also, based on the basic Scherrer equation, the Williamson–Hall equation is applied to extend the formula for the XRD. Enamel and cementum tissues of a typical human tooth, which look similar, are composed of a large variety of wide lines with different widths through Raman spectra analysis. In addition, the applied scanning electron microscopy extracts similar morphology for all tissues with round granular structures which are denser in the cementum. Atomic force microscopy is finally used for investigation of micro‐morphologies of the different tissues and the results are compared with the fractal analysis which ends to the bifractal and anisotropic nature of enamel and cementum along with monofractal and isotropic nature of dentin.     

Stress and strain distribution in demineralized enamel: A micro‐CT based finite element study

Physiological oral mechanical forces may play a role on the progression of enamel carious lesions to cavitation. Thus, the aim of this study was to describe, by 3D finite element analysis, stress, and strain patterns in sound and carious enamel after a simulated occlusal load. Micro‐CT based models were created and meshed with tetrahedral elements (based on an extracted third molar), namely: a sound (ST) and a carious tooth (CT). For the CT, enamel material properties were assigned according to the micro‐CT gray values. Below the threshold corresponding to the enamel lesion (2.5 g/cm³) lower and isotropic elastic modulus was assigned (E = 18 GPa against E₁ = 80 GPa, E₂ = E₃ = 20 GPa for sound enamel). Both models were imported into a FE solver where boundary conditions were assigned and a pressure load (500 MPa) was applied at the occlusal surface. A linear static analysis was performed, considering anisotropy in sound enamel. ST showed a more efficient transfer of maximum principal stress from enamel to the dentin layer, while for the CT, enamel layer was subjected to higher and concentrated loads. Maximum principal strain distributions were seen at the carious enamel surface, especially at the central fossa, correlating to the enamel cavity seen at the original micro‐CT model. It is possible to conclude that demineralized enamel compromises appropriate stress transfer from enamel to dentin, contributing to the odds of fracture and cavitation. Enamel fracture over a dentin lesion may happen as one of the normal pathways to caries progression and may act as a confounding factor during clinical diagnostic decisions. Microsc. Res. Tech. 78:865–872, 2015. © 2015 Wiley Periodicals, Inc.