Scanning Microradiography Research Articles

Validation of a Real-Time ISE Methodology to Quantify the Influence of Inhibitors of Demineralization Kinetics in vitro Using a Hydroxyapatite Model System

The aim was to validate a novel protocol to measure the cariostatic efficacies of demineralization inhibitors by repeating previous SMR (scanning microradiography) studies investigating the dose response of Zn²⁺ and F^– on demineralization kinetics in vitro using real-time Ca²⁺ ion selective electrodes (ISEs). In this study, Ca²⁺ release was used as a proxy for the extent of demineralization. Forty-eight hydroxyapatite (HAP) discs were allocated into 16 groups (n = 3) and adding either increasing [Zn²⁺], or [F^–], similar to those used in the previous SMR studies. Each HAP disc was immersed in 50 mL, pH 4.0, buffered acetic acid for 1 h, and real-time ISE methodology was used to monitor the rate of increase in [Ca²⁺] in the demineralization solution. Next, either zinc acetate or sodium fluoride was added into each demineralization solution accordingly. Then after each [Zn²⁺] or [F^–] addition, the HAP disc was further demineralized for 1 h, and ISE measurements were continued. The percentage reduction in the rate of calcium loss from hydroxyapatite (PRCL_HAP) at each [Zn²⁺] or [F^–] was calculated from the decrease in Ca²⁺ release rate, similar to that used in the previous SMR studies. A log-linear relationship between mean PRCL_HAP and inhibitor concentration was found for both Zn²⁺ and F^–, similar to that reported for each ion in the previous SMR studies. In conclusion, real-time Ca²⁺ ISE systems can be used to measure the cariostatic efficacies of demineralization inhibitors.

A Novel Kinetic Method to Measure Apparent Solubility Product of Bulk Human Enamel.

Introduction: Tooth enamel mineral loss is influenced by its solubility product value, which is fundamental to the understanding of de- and remineralization resulting from a carious or erosive challenge. Published pKsp values for human enamel and hydroxyapatite range from 110 to 126 suggesting a heterogeneous nature of enamel solubility. However, this range of values may also result from the variety of methods used, e.g., some authors reporting values for suspensions of enamel powder and others for bulk enamel. The aim of this study was to develop a method to measure the solubility of bulk human enamel under controlled in vitro conditions simulating demineralization behavior of enamel within the oral environment using scanning microradiography (SMR). SMR was used to monitor real-time changes in enamel demineralization rates at increasing calcium concentrations in a caries simulating demineralization solution until the concentration at which thermodynamic equilibrium between enamel and solution was achieved.Method: 2 mm thick caries free erupted human enamel slabs with the natural buccal surfaces exposed were placed in SMR cells exposed to circulating caries-simulating 2.0 L 0.1 M pH = 4.0 acetic acid, at 25°C. SMR was used to continuously measure in real-time the decrease in mineral mass during the demineralization at 5 different points from on each slab. Demineralization rates were calculated from a linear regression curve of projected mineral mass against demineralization time. Changes in the demineralization rates were monitored following a series of successive increases in calcium (and phosphate at hydroxyapatite stoichiometric ratios of Ca:P 1.67) were added to the demineralizing solution, until demineralization ceased. The pH was maintained constant throughout.Results: Demineralization halted when the calcium concentration was ~30 mM. At higher calcium concentrations, mineral deposition (remineralization) occurred. By comparison with results from speciation software calculations for the calcium phosphate ternary system, this result suggests that the bulk solubility product of enamel (pKspBEnamel) under the conditions used is 121.Discussion: The apparent pKspBEnamel under these conditions was higher than many previous reported values, and much closer to those previously reported for HAp. However, this is a bulk value, and does not reflect that enamel is a heterogeneous material, nor the influence of ionic inclusions.

Inhibitory Effects of Zinc Ions on Enamel Demineralisation Kinetics in vitro

Metal ions including zinc have the ability to influence enamel demineralisation. However, there is a paucity of data regarding reductions in demineralisation effected by zinc ions (Zn²⁺) in the literature. Therefore the aim was to measure the effects of zinc ion concentration ([Zn²⁺]) on the real-time in vitro demineralisation of enamel, during exposure to caries-simulating conditions, using scanning microradiography (SMR). Human enamel blocks were fixed in SMR environmental cells, through which acidic solutions (0.1 <smlcap>M</smlcap> acetic acid, pH 4.0) were circulated for periods of 50 h. SMR was used to quantitatively measure continuous mineral mass loss. Subsequently, the effects of sequentially increasing [Zn²⁺] (0.1-3,565 ppm) in the acidic solutions were measured on the rate of enamel demineralisation. This study demonstrated that Zn²⁺ even at low concentrations significantly reduces enamel demineralisation. There was a log-linear relationship between the mean percentage reduction in demineralisation and increasing [Zn²⁺] up to 3,565 ppm, i.e. the change in the overall percentage reduction in demineralisation was greater at lower concentrations than at higher concentrations, with 60% reduction at 36 ppm increasing to 90% at 3,565 ppm. In conclusion, SMR demonstrated the ability of Zn²⁺ to reduce the rate of enamel demineralisation under real-time in vitro acid conditions simulating dental caries. The results suggest that Zn²⁺ in the oral fluids could protect against enamel demineralisation during an acidic challenge. The log-linear relationship between [Zn²⁺] and demineralisation suggests that the reduction in enamel dissolution is limited by the saturation of surface sites on the enamel surface.

Recovery of crystallographic texture in remineralized dental enamel.

Dental caries is the most prevalent disease encountered by people of all ages around the world. Chemical changes occurring in the oral environment during the caries process alter the crystallography and microstructure of dental enamel resulting in loss of mechanical function. Little is known about the crystallographic effects of demineralization and remineralization. The motivation for this study was to develop understanding of the caries process at the crystallographic level in order to contribute towards a long term solution. In this study synchrotron X-ray diffraction combined with scanning electron microscopy and scanning microradiography have been used to correlate enamel crystallography, microstructure and mineral concentration respectively in enamel affected by natural caries and following artificial demineralization and remineralization regimes. In particular, the extent of destruction and re-formation of this complex structure has been measured. 2D diffraction patterns collected at the European Synchrotron Radiation Facility were used to quantify changes in the preferred orientation (crystallographic texture) and position of the (002) Bragg reflection within selected regions of interest in each tooth slice, and then correlated with the microstructure and local mineral mass. The results revealed that caries and artificial demineralization cause a large reduction in crystallographic texture which is coupled with the loss of mineral mass. Remineralization restores the texture to the original level seen in healthy enamel and restores mineral density. The results also showed that remineralization promotes ordered formation of new crystallites and growth of pre-existing crystallites which match the preferred orientation of healthy enamel. Combining microstructural and crystallographic characterization aids the understanding of caries and erosion processes and assists in the progress towards developing therapeutic treatments to allow affected enamel to regain structural integrity.

Effects of fluoride concentration on enamel demineralization kinetics in vitro

ObjectivesThe aim of the present study was to measure the effects of fluoride concentration on the real-time in vitro demineralization of enamel during exposure to caries-simulating conditions using Scanning Microradiography (SMR). MethodsEnamel blocks obtained from non-carious human molars were fixed in SMR environmental cells, through which acidic solutions (0.1M acetic acid, pH 4.0) were circulated for periods of 48h. SMR was used to quantitatively measure continuous mineral mass loss. Subsequently, the effects of sequentially increasing fluoride concentration (0.1–4500mg/L [F−]) in the acidic solutions were measured on the rate of enamel demineralization. ResultsThe data shows a log-linear relationship between [F−] and reduction in demineralization up to 135mg/L [F−]. Above 135mg/L, no further significant decrease in demineralization occurred. ConclusionThe optimum range of local fluoride concentration for reducing enamel demineralization was in the range 0.1–135mg/L [F−] under the conditions studied. Clinical significanceRelatively low [F−] can exhibit near-optimum protection. Increasing the fluoride concentrations above 135mg/L may not necessarily give an increased cariostatic benefit. Improving the means of delivery of relatively low fluoride concentrations to the oral fluids through slow releasing mechanisms, such as the oral fluoride reservoirs, is the more appropriate way forward for sustaining long-term clinical efficacy.

Scanning Microradiography: Measurement of X-ray Attenuation Coefficient of Fissure Sealants Containing Tin Methacryloxytri-n-butyltin (SnM) as Radio-opaquer

Objective: Radio-opacity is a fundamental requirement for fissure sealants in order to be clearly visible radiographicallyand may be well differentiated from caries. Many attempts have been made to make resin based fissure sealants radio-opaque. Researchers achieved radio-opacity of the sealants by blending heavy metals as fillers into the polymeric matrix. Keeping the demerits of using heavy metals in view, a novel fissure sealant was developed by chemically incorporating Tin (Sn) in monomer of methacrylate. This study has been done to assess under scanning micro-radiography (SMR), the radiopacity of the indigenously prepared fissure sealants containing increasing amount of SnM until the consistency of the mix remains workable and clinically useful. Design: Experimental study. Place of study: Biophysical Lab Queen Mary University of London, United Kingdom. Methodology: An organo-tin compound – Methacryloxytri-n-butyltin (SnM) and Ethylene Glycol Dimethacrylate (EGDMA) in varying quantities were mixed to prepare fissure sealant indigenously. Camphorquinone (CQ) and N, N-Dimethyl-P-Toluidine (DMPT) were added to above mixture. The prepared sealants were polymerized in cuvettes and mounted on Scanning microradiography machine. The machine was run for 100 seconds repeatedly for 60 runs. Obtained results were processed and calculated in Microsoft Excel software. Results: It was observed that increasing the weight of Tin (Sn) content in a sealant increases the radio-opacity of the sealant but SnM more than 9.5 gm renders the material unworkable. Conclusion: This study will not only help in enhancing the radio-opacity of fissure sealants but also the radio-opacity of other clinical composite materials maybe enhanced by this method.

An in vitro scanning microradiography study of the reduction in hydroxyapatite demineralization rate by statherin‐like peptides as a function of increasing N‐terminal length

Enamel demineralization is slowed by salivary proteins that inhibit calcium hydroxyapatite (HA) demineralization. Statherin (StN43), a 43-residue phosphorylated salivary protein with primary sequence similarities to osteopontin and caseins, binds calcium and HA. The aim of this study was to identify the minimum length of the functional domain of the statherin molecule required for cariostatic function by measuring the efficacy of peptides of progressively shorter length (i.e. containing only the N-terminal 21 (StN21), 15 (StN15), 10 (StN10), or 5 (StN5) residues) to reduce HA demineralization rates (RD(HA) ). Porous HA blocks were used as enamel analogues, and were exposed to 0.1 M acetic acid at pH 4 for 120 h, rinsed, and treated with StN21, StN15, StN10, or StN5 peptides (1.88 × 10(-5) M) for 24 h, then demineralized for a further 120 h. The RD(HA) was measured, before and after peptide treatment, using scanning microradiography. Hydroxyapatite blocks treated with StN21 and StN15 demonstrated a 50-60% reduction in the RD(HA) . However, no reduction in the RD(HA) was observed following treatment with either StN10, StN5, or buffer only. The mechanism by which statherin-like peptides reduce RD(HA) may be associated with their binding to HA surfaces. Comparisons with previously published binding energies of statherin to HA also suggest that statherin-like peptides containing 15 N-terminal residues or more, are required for binding, suggesting a link between binding and demineralization reduction.

The influence of proteins on demineralization kinetics of hydroxyapatite aggregates

Salivary proteins influence the biomineralization of hydroxyapatite (HAp) within enamel. Their effect on the crystal growth has been extensively studied, but, their effect on demineralization kinetics is less well investigated. In this study bovine serum albumin (BSA) was used as a model protein to measure its effect on demineralization kinetics of hydroxyapatite aggregates using scanning microradiography (SMR). HAp aggregates (8 and 20% porous) were cut into 5 x 5 x 2 mm blocks. SMR cells were prepared containing hydroxyapatite blocks. BSA was added to demineralising solutions (0.1 mol L(-1) acetic acid, buffered to pH 4.0; degree of saturation zero and 0.062 respectively) at a concentration range 0.76-75.8 micromol L(-1). Demineralising solution without added BSA was used as a control. The demineralising solutions were circulated past the samples at 0.4 mL min(-1). SMR was used to measure the rate of mineral loss (RML(HAp)) at 14 points in each sample repeatedly for 3 weeks. The results show that BSA increases or decreases the RML(HAp) depending on; BSA concentration, HAp porosity, and the degree of saturation of the demineralising solution. It is suggested that BSA influences demineralization kinetics of HAp either by modifying solution properties, or, by affecting the surface energy of hydroxyapatite.

Comparison of demineralisation rates in pre- and postnatal enamel and at the neonatal line

AimsThe neonatal line, which is an exaggerated incremental layer line, separates pre- and postnatal enamel. It has been suggested that this layer may be a barrier to the progress of a carious lesions. The objective was to measure the rate of demineralisation in pre- and postnatal enamel and within the neonatal layer using scanning microradiography (SMR). Permanent enamel and compressed permeable hydroxyapatite samples were used as controls. Methods and resultsEnamel specimens from deciduous incisors were cut into mesiodistal blocks of 2mm thickness without altering the labial surface and located within SMR cells. Permanent enamel and hydroxyapatite specimens were similarly prepared. Artificial caries-like lesions were created by exposing the specimens to 0.1moll−1 acetic acid (pH 4.0) within the SMR cells. SMR was used to measure the rate of mineral loss at 10 points either side of and at the neonatal line in the deciduous enamel, and 20 points across in the control specimens.The rate of demineralisation was almost the same in pre- and postnatal enamel ((6.0–8.0)×10−4gcm−2h−1), but much lower in the vicinity of the neonatal line (2.0×10−4gcm−2h−1). The rate of demineralisation was lower in permanent (5.0×10−4gcm−2h−1) than in deciduous enamel, and even lower in the permeable hydroxyapatite specimen (2.5×10−4gcm−2h−1). ConclusionsThis study showed no difference in the rate of demineralisation between pre- and postnatal enamel, but a reduced rate within the region that contained the plane of the neonatal line. This supports the hypothesis that the neonatal line may act as a barrier to the propagation of carious lesions.

A Synthetic Peptide Based on a Natural Salivary Protein Reduces Demineralisation in Model Systems for Dental Caries and Erosion

The salivary protein statherin is an inhibitor of spontaneous and secondary precipitation of hydroxyapatite (HAp). It is also detected in enamel pellicle. The N-terminal region of statherin is involved in its adsorption onto tooth surfaces, and, calcium binding. A peptide (StN21) was designed with a 21 amino acid sequence identical to the N-terminus of statherin. The aim was to measure the effect of StN21 on the rate of mineral loss in a model system for dental caries and erosion using HAp subjected to artificial carious and erosive conditions. StN21 was synthesised using Fmoc chemistry. A surface of each HAp block was exposed to solution containing StN21 at concentrations 9.4–376 μmol L−1 (in phosphate buffer) for 24 h. Controls were HAp exposed to buffer only, and HAp exposed to lysozyme. Demineralising solution (0.1 mol L−1 acetic acid, pH 4.5, 1.0 mmol L−1 calcium and 0.6 mmol L−1 phosphate) was circulated past the HAp blocks at 0.4 mL min-1 to mimic carious and erosive conditions. Scanning microradiography was used to measure the rate of mineral loss for demineralisation periods of 3 weeks. The rate of mineral loss of the samples exposed to StN21 was reduced by ∼40% compared to the controls, but no dependence on the concentration of StN21 was observed at the concentrations used. StN21 has been shown to be a potent and stable peptide that has potential as a preventive/therapeutic agent in the treatment of enamel erosion and dental caries.

Acidic demineralization of apatites studied by scanning X-ray microradiography and microtomography

AbstractThe mineral in bones and teeth is an impure form of hydroxylapatite (HAP), the principal impurity being 2—5 wt.% carbonate. This mineral dissolves during remodelling of bone and also in dental caries as a result of the action of acids produced by osteoclasts and by bacteria, respectively. In enamel, demineralization proceeds with preferential loss of carbonate relative to phosphate. Surprisingly, in the early stages, the demineralization is subsurface. In order to facilitate the understanding of physical chemical aspects of these processes, we have undertaken studies of demineralization in model systems. We give three examples here. The first two used scanning microradiography in which the specimen is stepped across a 10—30 μm diameter X-ray beam. Intensity measurements allow calculation of the mineral mass per unit area in the X-ray path through the specimen. In the first experiment, porous HAP sections were separated from a reservoir of acidic buffer by a column initially filled with water (the diffusion length) and scanned with the X-ray beam perpendicular to the axis of the diffusion length. The rate of total loss of mineral along each profile was calculated from the scans. The rate of demineralization fell as the diffusion length increased. We believe the explanation is that the rate-controlling step is the diffusion of dissolved HAP away from the solid to the buffer reservoir. In the second experiment, demineralizing solution and water were pumped alternately, for equal lengths of time, past blocks of porous HAP or enamel. The X-ray beam was perpendicular to the exposed surface. As the rate of switching between solutions decreased, the mean rate of demineralization also fell. We propose that this effect is due to retention of acid in the pores of the HAP during the time when water flows, allowing further demineralization to take place during this time. The third study used X-ray microtomography, a form of 3D microscopy, to study the loss of mineral in compacted carbonate apatite powders. The powders were packed in six 10 mm internal diameter acrylic cylinders to a depth of 4 mm (after pressing). One end was covered with a porous polyethylene disc and each tube placed in acidic buffer for 70 days. Periodic examination by microtomography showed the development of subsurface demineralization. Infrared spectroscopy of the dissected-out surface layers showed preferential loss of carbonate over phosphate by comparison with deeper layers. Rietveld analysis of X-ray powder diffraction data showed changes in the crystallographic structures of the apatites between the initial and dissected-out apatite.

Scanning microradiographic study on the influence of diffusion in the external liquid on the rate of demineralization in hydroxyapatite aggregates

Subsurface demineralization in enamel caries is known to entail diffusion of reagents and products both within the lesion and within the plaque biofilm external to the lesion. However, development of a predictive mathematical model for subsurface demineralization is hindered by limited quantitative understanding of the effects of these diffusion processes. The purpose of this quantitative study was to investigate and understand the effect of external diffusion length on the rate of demineralization in a simple model system. Ten, 500-microm thick sections cut from a porous hydroxyapatite (HAP) pellet were inserted in scanning microradiography (SMR) cells. The exposed thin edges of the sections were initially separated by columns of water (diffusion lengths) of 0-0.9 cm from a 1-l reservoir of demineralizing buffer (pH 4). Buffer was found to diffuse from the reservoir through the increasing diffusion lengths to the exposed HAP surface, whilst dissolved product diffused along the reverse path. Rates of HAP loss (from SMR measurements) decreased as the diffusion length increased. Experimental data were fitted to a general diffusion-reaction model. This showed that the solution near the HAP surface was almost completely saturated with HAP, and that the diffusion of dissolution products, rather than of buffer species, was rate limiting.

Demineralization in enamel and hydroxyapatite aggregates at increasing ionic strengths

Subsurface demineralization of dental enamel is a curious feature of both in vivo and in vitro lesion formation. Numerous explanations have been proposed to explain this. One general hypothesis is that subsurface demineralization in enamel and synthetic hydroxyapatite (HAP) aggregates may result from the phenomenon of coupled diffusion between the inward transport of acid and the outward transport of dissolution products. The aim of this study was to test the validity of this explanation. Inert electrolyte was added to demineralizing solutions in order to reduce electrostatic coupling between the diffusive flows that occur during lesion formation. Scanning microradiography (SMR) was used to examine surface layer formation, and to measure the rate of mineral loss at increasing ionic strengths. It was found that surface layer formation was significantly reduced as the concentration of inert electrolyte was increased. Further, the rate of mineral loss from the developing lesion increased as the concentration of inert electrolyte (and therefore the ionic strength) in the demineralizing solution increased. It is concluded that electrostatic coupling between counter diffusing acid and dissolution products during lesion formation can significantly influence the mineral concentration within the surface layer.

Effects of sodium hypochlorite solution on root dentine composition.

Sodium hypochlorite (NaOCl) solution,<or=5% w/v available chlorine (abbreviated subsequently to %), is widely used as an irrigant in root canal treatment of teeth, so its effects on dentine are of clinical importance. The effects of approximately 0.5%, 3% and 5% NaOCl solution on the composition of root dentine were studied at ambient temperature. For dentine powder treated for 30 min, depletion of the organic phase was confirmed by infrared spectroscopy. Apatite lattice parameters showed no significant change, but NaCl was also detected by X-ray powder diffraction. The low solubility of apatite mineral in the NaOCl solutions was demonstrated by the constant weight of bulk enamel specimens immersed for seven days. The stability of the mineral phase was confirmed by scanning microradiography (SMR), an X-ray attenuation method employing photon counting. Repeated SMR measurements of the local mineral content of bulk samples of root dentine and a synthetic hydroxyapatite aggregate during exposure to pumped NaOCl solutions for 100 h showed no mineral loss. As predicted from apatite chemistry, reaction of NaOCl with the mineral phase can be excluded as a primary factor in changes in mechanical properties of treated dentine. Effects of retention of NaCl on endodontic sealants requires further investigation.

Real-time measurement of in vitro enamel demineralization in the vicinity of the restoration-tooth interface.

An X-ray attenuation method using photon counting (scanning microradiography) is presented for the real-time study of in vitro demineralization of dental tissues in the vicinity of the restoration-tooth interface. By repeated measurement of mineral content profiles during the course of demineralization, the pattern of lesion development and the rate of mineral loss can be studied. The method is illustrated by comparison of enamel demineralization near a polyacid-modified composite resin restoration, near a bis-GMA/TEGMA composite resin restoration, and in an unrestored control. The method has potential for study of the influence of restorative materials on susceptibility of tooth tissue to demineralization.

Scanning microradiographic studies of rates of in vitro demineralization in human and bovine dental enamel

The aim was to measure frequently and with precision the local integrated mineral loss through small areas of the natural surface of human and bovine enamel during in vitro demineralization using an X-ray photon-counting system (scanning microradiography). The method used was an adaptation of photographic longitudinal microradiography in which the attenuation of X-rays through the enamel is measured in the direction of acid attack, i.e., normal to the enamel surface. The mass of mineral (assumed to be hydroxyapatite) per unit exposed area was measured over 15 μm dia. circles at a series of positions as a function of time in blocks of human and bovine enamel immersed in 0.1 mol/l acetic acid buffered to pH 4.0 with NaOH. There was an initial period (approx. 45 h for human, approx. 75 h for bovine enamel) during which the mineral loss with time was sigmoidal, followed by a nearly linear loss for the remainder of the experiment, in some cases up to 500 h. The initial sigmoidal period may be due to properties of surface enamel or be associated with the development of a surface layer overlying subsurface demineralization. The essentially constant rate of mineral loss after the surface layer has formed confirms earlier observations and is consistent with a rate-limiting process occurring at the dissolving enamel surfaces of the advancing front, and not by transport of ions within the lesion. Small perturbations from a linear loss were seen, which were approximately periodic for human enamel. The slope of the linear period was rather constant within one human or bovine block, but variable between blocks without a clear distinction between human and bovine enamel.

Determination of Mineral Concentration in Dental Enamel from X-ray Attenuation Measurements

The mineral content of dental enamel is commonly measured by X-ray attenuation experiments. Most studies have used contact microradiography in which intensities are measured with photographic film which is convenient and gives high spatial resolution. However photon counting intensity measurements are to be preferred in many experiments (longitudinal and scanning microradiography, and microtomography), as illustrated here, because they have a larger dynamic range and greater sensitivity to small intensity changes. Additionally, the detector and specimen are well separated which allows the pseudo-continuous study of de- and remineralization. The mineral content is often quoted as 95 wt% or 87 vol% hydroxyapatite for permanent human enamel. This determination from attenuation experiments requires accurate values of elemental mass attenuation coefficients and a number of assumptions. The effects of possible choices of these are considered and it is shown that the most important is the density of enamel mineral used in conversion of wt% to vol%. If the density is taken as 2.99 g cm−3 as recently suggested (J.C. Elliott, Dental Enamel, Ciba Foundation Symposium 205, Wiley, Chichester, pp. 54–72, 1997), instead of 3.15 g cm−3 as for hydroxyapatite, the calculated vol% is ∼93 instead of ∼87.

Application of scanning microradiography and X-ray microtomography to studies of bones and teeth

In scanning microradiography (SMR), a thin section is stepped across a 15- μ m diameter X-ray beam and the transmitted intensity measured at each point. This technique has permitted more accurate measurements of the spatial variation of the mineral concentration in sections of dentin and enamel than conventional photographic microradiography. Moreover, because the section is not in close contact with an emulsion, SMR allows continuous study while the specimen is bathed in a reaction solution. The present studies have been particularly directed to gaining an understanding of the formation and repair of carious lesions in teeth: one particular puzzle is subsurface demineralization, in which the initial loss of mineral appears to take place some 20 to 50 μ m below the tooth surface. SMR studies are reported here on the demineralization in dilute acids and the subsequent partial remineralization in supersaturated calcium phosphate solutions in model systems for dental caries. In order to develop a theoretical model for de- and remineralization of carious lesions, it is necessary to quantify transport processes within the tooth. To this end, we are developing a method of measuring effective diffusion coefficients of strongly X-ray-absorbing ions in water within permeable solids in which the diffusion coefficient varies with position. The method uses sequential concentration/distance profiles determined by SMR. As a test, diffusion coefficients of potassium iodide in water within a permeable glass frit have been measured. X-ray microtomography (XMT) can be carried out by adding an axis of rotation to the SMR apparatus. Using this method, linear absorption coefficients, and hence mineral concentrations, can be measured in 15 × 15 × 15- μ m 3 voxels. This has advantages over SMR in that superposition within the depth of the section and errors in determining its thickness are avoided. XMT studies of de- and remineralization similar to those described above for SMR, and also XMT studies of the variation in mineral concentration in the cortical bone of a rat femur along its length, are reported.

Application of Scanning Microradiography and X-Ray Microtomography to Studies of Bones and Teeth

In scanning microradiography (SMR), a thin section is stepped across a 15-μm diameter X-ray beam and the transmitted intensity measured at each point. This technique has permitted more accurate measurements of the spatial variation of the mineral concentration in sections of dentin and enamel than conventional photographic microradiography. Moreover, because the section is not in close contact with an emulsion, SMR allows continuous study while the specimen is bathed in a reaction solution. The present studies have been particularly directed to gaining an understanding of the formation and repair of carious lesions in teeth: one particular puzzle is subsurface demineralization, in which the initial loss of mineral appears to take place some 20 to 50 μm below the tooth surface. SMR studies are reported here on the demineralization in dilute acids and the subsequent partial remineralization in supersaturated calcium phosphate solutions in model systems for dental caries. In order to develop a theoretical model for de- and remineralization of carious lesions, it is necessary to quantify transport processes within the tooth. To this end, we are developing a method of measuring effective diffusion coefficients of strongly X-ray-absorbing ions in water within permeable solids in which the diffusion coefficient varies with position. The method uses sequential concentration/distance profiles determined by SMR. As a test, diffusion coefficients of potassium iodide in water within a permeable glass frit have been measured. X-ray microtomography (XMT) can be carried out by adding an axis of rotation to the SMR apparatus. Using this method, linear absorption coefficients, and hence mineral concentrations, can be measured in 15 X 15 X 15-μm3 voxels. This has advantages over SMR in that superposition within the depth of the section and errors in determining its thickness are avoided. XMT studies of de- and remineralization similar to those described above for SMR, and also XMT studies of the variation in mineral concentration in the cortical bone of a rat femur along its length, are reported.

Scanning Microradlographic Study of the Kinetics of Subsurface Demineralization In Tooth Sections under Constant-composition and Small Constant-volume Conditions

The kinetics of subsurface demineralization of tooth sections has been studied in real-time by scanning microradiography (SMR). Demineralization was carried out: (1) with a large volume of solution buffered to pH = 4 to maintain a constant composition; and (2) in a small constant volume (approximately 3 mL), buffered initially at pH = 4, so that the degree of saturation at the tooth surface increased as the tooth dissolved. At constant composition, the change in lesion depth, Y, with time, T, followed a linear relation, Y = a + bT, for T > 44 +/- 5 h. Before this time, the relation could be approximated by a linear one with different a and b constants. At constant volume, Y = q(1-e-r(T + s)) for all T, where q, r and s are constants. Similar relations, with different constants, were found for the mineral loss per unit area of lesion exposed to acid. These results showed that the process of demineralization under the rather severe conditions used was essentially a surface-controlled process. The change of slope at approximately 44 h and the presence of the constant s in the exponential function were attributed to a change in kinetics after formation of the surface layer.