Acid resistance of human enamel in vitro after bicarbonate application during remineralization

OBJECTIVES: The aim of this study was to investigate the acid resistance of subsurface enamel lesions remineralized with bicarbonate solutions during remineralization. METHODS: Two experiments were carried out. In experiment 1, mineral uptake and acid resistance of remineralized enamel lesions were analyzed quantitatively by microradiography for mineral changes. Bicarbonate solutions of 0.5, 5.0 and 50.0 mM were used. In experiment 2, to clarify acid resistance mechanisms, the pH changes in demineralizing solutions on the remineralized enamel surfaces were measured continuously. Only a bicarbonate solution of 5.0 mM was used. RESULTS: In experiment 1, the bicarbonate-treated groups were more acid resistant than the non-treated groups (p<0.05). However, no statistically significant difference was observed among the different concentrations of bicarbonate. In experiment 2, the pH rise of the bicarbonate group was greater than the other groups. CONCLUSION: It was found that bicarbonate-treated enamel lesions were resistant to acid. It would suggest that bicarbonate ions applied during remineralization may have penetrated into the subsurface lesions. These ions may have worked as buffer agents against the acid challenge and inhibited the decrease in pH.     

Acid resistance of enamel subsurface lesions remineralized by a sugar-free chewing gum containing casein phosphopeptide-amorphous calcium phosphate

The aim of this clinical study was to investigate the acid resistance of enamel lesions remineralized in situ by a sugar-free chewing gum containing casein phosphopeptide-amorphous calcium phosphate nanocomplexes (CPP-ACP: Recaldent). The study utilized a double-blind, randomized, crossover design with two treatments: (i) sugar-free gum containing 18.8 mg of CPP-ACP, and (ii) sugar-free gum not containing CPP-ACP as control. Subjects wore removable palatal appliances with insets of human enamel containing demineralized subsurface lesions and chewed the gum for 20 min 4 times per day for 14 days. After each treatment the enamel slabs were removed and half of each lesion challenged with acid in vitro for 8 or 16 h. The level of remineralization was determined using microradiography. The gum containing CPP-ACP produced approximately twice the level of remineralization as the control sugar-free gum. The 8- and 16-hour acid challenge of the lesions remineralized with the control gum resulted in 65.4 and 88.0% reductions, respectively, of deposited mineral, while for the CPP-ACP-remineralized lesions the corresponding reductions were 30.5 and 41.8%. The acid challenge after in situ remineralization for both control and CPP-ACP-treated lesions resulted in demineralization underneath the remineralized zone, indicating that the remineralized mineral was more resistant to subsequent acid challenge. The results show that sugar-free gum containing CPP-ACP is superior to an equivalent gum not containing CPP-ACP in remineralization of enamel subsurface lesions in situ with mineral that is more resistant to subsequent acid challenge.     

Effect of addition of citric acid and casein phosphopeptide-amorphous calcium phosphate to a sugar-free chewing gum on enamel remineralization in situ

Casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) has been shown to remineralize enamel subsurface lesions in situ. The aim of this study was to investigate the effects of CPP-ACP in a fruit-flavoured sugar-free chewing gum containing citric acid on enamel remineralization, and acid resistance of the remineralized enamel, using an in situ remineralization model. The study utilized a double-blind, randomized, crossover design with three treatments: (i) sugar-free gum (2 pellets) containing 20 mg citric acid and 18.8 mg CPP-ACP, (ii) sugar-free gum containing 20 mg citric acid alone, (iii) sugar-free gum not containing CPP-ACP or citric acid. Ten subjects were instructed to wear removable palatal appliances, with 4 half-slab insets of human enamel containing demineralized subsurface lesions and to chew gum (2 pellets) for 20 min 4 times per day for 14 days. At the completion of each treatment the enamel half-slabs were removed and half of the remineralized lesion treated with demineralization buffer for 16 h in vitro. The enamel slabs (remineralized, acid-challenged and control) were then embedded, sectioned and subjected to microradiography to determine the level of remineralization. Chewing with gum containing citric acid and CPP-ACP resulted in significantly higher remineralization (13.0 +/- 2.2%) than chewing with either gum containing no CPP-ACP or citric acid (9.4 +/- 1.2%) or gum containing citric acid alone (2.6 +/- 1.3%). The acid challenge of the remineralized lesions showed that the level of mineral after acid challenge was significantly greater for the lesions exposed to the gum containing CPP-ACP.     

Dental caries: a dynamic disease process

Abstract Dental caries is a transmissible bacterial disease process caused by acids from bacterial metabolism diffusing into enamel and dentine and dissolving the mineral. The bacteria responsible produce organic acids as a by-product of their metabolism of fermentable carbohydrates. The caries process is a continuum resulting from many cycles of demineralization and remineralization. Demineralization begins at the atomic level at the crystal surface inside the enamel or dentine and can continue unless halted with the end-point being cavitation. There are many possibilities to intervene in this continuing process to arrest or reverse the progress of the lesion. Remineralization is the natural repair process for non-cavitated lesions, and relies on calcium and phosphate ions assisted by fluoride to rebuild a new surface on existing crystal remnants in subsurface lesions remaining after demineralization. These remineralized crystals are acid resistant, being much less soluble than the original mineral.     

Relationship between electrical resistance measurements and microradiographic variables during remineralization of softened enamel lesions

The purpose of this study was to investigate the relationship between electrical resistance (ECM) measurements and microradiographic variables during remineralization of softened enamel lesions. Sound bovine enamel blocks were demineralized and, subsequently, were remineralized for 3, 4, 5, 7 or 14 days. All the specimens were assessed by ECM and transversal microradiography. The mean ECM values in each group increased after remineralization. The ECM values after remineralization (ECM1), mineral loss (DeltaZ) and maximum mineral content in the surface layer (V(max)) were significantly correlated with the remineralization time. The ECM1 values were also significantly correlated with DeltaZ and V(max). It is suggested that ECM could be used as a device to monitor mineral accumulation as a result of preventive treatment in vitro. However, it may take a much longer time to follow up the remineralization changes compared with demineralization.     

Remineralization of deep enamel dentine caries lesions

Enamel remineralization is generally studied in superficial (up to 100 mum) lesions, but in vivo caries lesions may be tenfold deeper. This article addresses the question whether deep lesions, and extending into dentine, can be remineralized under optimal conditions and if this process is influenced by agents affecting calcium phosphate precipitation and dissolution. Lesions through enamel into dentine were first formed in thin sections and then continuously remineralized for periods up to 200 days. With longitudinal assessment by transversal microradiography it was showed that remineralization throughout the depth of the lesion and into the dentine was possible, although this process is very slow. Fluoride and bisphosphonate treatments affected mainly the deposition in the outer enamel. Although it was assumed that this would affect the diffusion of ions to deeper layers, the treatments had no impact on remineralization in the inner enamel or dentinal parts of the lesions. These findings are discussed with relevant theoretical considerations, and in their possible clinical implications.     

The carbonate and fluoride in surfaces of remineralized enamel.

Analyses were made of samples of surface enamel collected before and after acid demineralization and after subsequent remineralization in solutions containing calcium and phosphate and various levels of bicarbonate and fluoride ions. Demineralization caused a preferential loss of carbonates. Remineralization was increased by both fluoride and bicarbonate ions. With no bicarbonate in the calcifying solution, 0.25 to 4.0 mM NaF increased mineral formation and thereby caused a relative reduction of its carbonate content. With 0.5 mM NaF (10 ppm F-) and 5, 15, and 25 mM HCO-3 in remineralizing solutions more carbonate was deposited than in the absence of NaF. Five and 15 mM HCO-3 increased, but 25 mM HCO-3 decreased, deposit of fluoride in enamel.     

In vitro remineralization of in vivo and in vitro formed enamel lesions

Thin sections of natural white spot enamel lesions (WS) and of artificial in vitro lesions (VL) were remineralized simultaneously in vitro. The sections, clamped in a PMMA holder, were microradiographed at baseline and after remineralization in a calcium- and phosphate-containing solution (pH = 7.0; 1 ppm F) after 2 and 4 weeks. All data were analyzed with respect to baseline. The results show that the lesion depth values did not change significantly during 2 and 4 weeks of remineralization. The mineral accumulation (change in DeltaZ), however, was substantial and significant in WS and VL. In WS the change in mineral accumulation was roughly proportional to the amount of mineral at baseline. The WS accumulated more than two times the amount of mineral than VL in the same periods of remineralization. After 4 weeks of remineralization the maximum mineral value Vmax in the surface layer of the WS was nearly up to the sound enamel level thick approximate87 vol%. This study shows that the technique and calculation procedure described make this single section method attractive for longitudinal demineralization-remineralization studies in vitro or in situ. Both WS and VL samples obviously remineralized in vitro similarly with respect to the baseline. Furthermore, this in vitro work indicates that remineralization inhibitors present in saliva, and previously penetrated into the enamel tissue, do not influence remineralization later on.     

3种溶液对早期釉质龋再矿化作用的扫描电镜观察

目的观察比较碳酸氢钠溶液、氟化钠溶液及碳酸氢钠与氟化钠混合液对人工釉质龋的再矿化作用。方法选择因正畸拔除的前磨牙制备成牙釉质标本,用凝胶酸蚀法形成人工釉质龋模型,脱矿标本分别经碳酸氢钠溶液、氟化钠溶液及碳酸氢钠与氟化钠混合液及双蒸水浸泡1周后,扫描电镜观察比较经不同溶液处理的脱矿釉质标本表面形态差异。结果经碳酸氢钠溶液、氟化钠溶液及碳酸氢钠与氟化钠混合液浸泡的脱矿釉质表面均有不同程度矿物质沉积,其中,经碳酸氢钠与氟化钠混合液浸泡的脱矿釉质表面形成的沉积物更规则、明显。结论碳酸氢钠溶液能够促进脱矿釉质再矿化,且能增强氟化钠溶液对早期釉质龋的再矿化作用。     

An approach to normalizing micro-CT depth profiles of mineral density for monitoring enamel remineralization progress

To indicate the possibility of a new approach to creating mineral density profiles, and to examine longitudinal changes in 'the rate of remineralization (RA)' and 'the mineral density (DAs) at 4 different depths' (surface zone: SZ, lesion body: LB, middle zone: MZ, deep zone near to sound area: DZ) in enamel subsurface lesions, eight demineralized bovine enamel-dentin blocks were remineralized for 1 to 4 week and investigated using Micro-focus X-ray CT (micro-CT). After CT scanning, mineral density profiles were created.Mineral densities at each depth after demineralization were SZ?LBMZ>DZ. This study indicated a new approach to create a mineral density profile and suggested the greater the value of the mineral density before the remineralization, the smaller the mineral density increments.     

Effects of acid etchants used for bonding orthodontic brackets on the remineralization of enamel white-spot lesions

The objective of the present study was to examine the effects of phosphoric acid and polyacrylic acid used as pretreatment for orthodontic direct bonding procedures on changes in the overall mineral levels of the subsurface white-spot lesions, and to examine the effects on the surface structure using contact microradiography (CMR) and an Atomic Force Microscope (AFM).Enamel specimens with in vitro production of artificial subsurface white-spot lesions were used for these studies. The enamel acid conditions were as follows: non-etched (Group I), 10% polyacrylic acid (Group II), 2% phosphoric acid (group III), 10% phosphoric acid (Group IV). After each acid treatment, the four groups were exposed to a remineralization solution at pH 7.0 at 37 °C for 2 weeks. An AFM was used to observe the surface after the acid treatment and remineralization. After 2 weeks of remineralization, the mineral profiles were assessed using CMR.AFM observation revealed different acid etching patterns on the enamel surface of subsurface white-spot lesions after the different acid treatments. From the AFM observation after remineralization, Group I and Group II demonstrated deposits of mineral-like substances on the surface. From the CMR observation, the amount of minerals present increased in Groups I and II, while, in Group IV, the surface layer was removed and the amount of minerals present decreased.It was found that polyacrylic acid produced a less roughness than low concentration phosphoric acid. Moreover, the minerals were recovered during remineralization after treatment with polyacrylic acid. The minerals were not recovered after phosphoric acid treatment.     

Consumption of milk with added casein phosphopeptide-amorphous calcium phosphate remineralizes enamel subsurface lesions in situ

Background:  Casein phosphopeptide-amorphous calcium phosphate (CPP-ACP) is an anticariogenic agent that is suitable to be added to foods. The aim of this double-blind, three-way crossover randomized study was to investigate the capacity of CPP-ACP, when added to bovine milk, to remineralize enamel subsurface lesions in situ .
Methods:  Ten subjects drank 100 mL of bovine milk containing no added CPP-ACP (control milk), 0.2% (w/v) CPP-ACP or 0.3% (w/v) CPP-ACP, for 30 seconds once daily for 15 days, whilst wearing removable appliances with attached slabs of enamel containing subsurface enamel lesions. After each treatment and a one-week washout period, subjects crossed over to another treatment and this was repeated until they had consumed each of the three milk products. At the completion of each treatment the enamel slabs were removed and remineralization was determined using microradiography.
Results:  The results demonstrated that all three milk samples remineralized enamel subsurface lesions in situ . However, the two milk samples containing added CPP-ACP each produced significantly greater remineralization than the control milk.
Conclusions:  The remineralizing effect of CPP-ACP in milk was dose-dependent with milk containing 0.2% CPP-ACP and 0.3% CPP-ACP producing an increase in mineral content of 81% and 164%, respectively, relative to the control milk.     

Remineralization of caries lesions extending into dentin.

Remineralization is one aspect of the overall process of tooth decay. However, it is primarily studied in shallow lesions. The aim of this study was to explore whether caries lesions in enamel and extending into the dentin can be remineralized. A single-section model was developed for the longitudinal and non-destructive monitoring of changes in enamel and dentin. Lesions at least 200 microm into dentin were formed in undersaturated acetate buffers. Next, the lesions were divided into groups (three treatment and one control) and remineralized. The treatments were: weekly immersion in 1,000 ppm fluoride, single treatment with methanehydroxybisphosphonate, and a constant level of 1 ppm fluoride. De- and remineralization was assessed by transverse microradiography. Remineralization was observed in enamel, but also in dentin, indicating that, deep into dentin, the pores become supersaturated to apatite formation. Treatments affected remineralization only in the outer part of enamel. Both findings are explained by a relatively fast diffusion of mineral ions, with precipitation being rate-limiting. The results suggest that dentin remineralization, underneath enamel, can be achieved and could possibly be used in clinical treatment strategies.     

In situ effect of sodium fluoride or titanium tetrafluoride varnish and solution on carious demineralization of enamel

This study evaluated the effect of titanium tetrafluoride (TiF(4)) formulations on enamel carious demineralization in situ. Thirteen subjects took part in this cross-over, split-mouth, double-blind study performed in three phases of 14 d each. In each subject, two sound and two predemineralized specimens of bovine enamel were worn intra-orally and plaque accumulation was allowed. One sound and one predemineralized specimen in each subject was treated once with sodium fluoride (NaF) varnish or solution (Treatment A); TiF(4) varnish or solution (Treatment B); or placebo varnish or no treatment (Treatment C). The initially sound enamel specimens were exposed to severe cariogenic challenge (20% sucrose, eight times daily for 5 min each time), whereas the predemineralized specimens were not. Eleven subjects were able to finish all experimental phases. The enamel alterations were quantified by surface hardness and transversal microradiography. Demineralization of previously sound enamel was reduced by all test formulations except for the NaF solution, while both TiF(4) formulations were as effective as NaF varnish. For the predemineralized specimens, enamel surface hardness was increased only by TiF(4) formulations, while subsurface mineral remineralization could not be seen in any group. Within the experimental protocol, TiF(4) was able to decrease enamel demineralization to a similar degree as NaF varnish under severe cariogenic challenges, while only TiF(4) formulations remineralized the enamel surface.     

Influence of salivary macromolecules and fluoride on enamel lesion remineralization in vitro

The aim of this study was to investigate the influence of salivary macromolecules on enamel lesion remineralization in the presence or absence of fluoride. Paraffin-stimulated whole saliva was centrifuged, and the supernatant was dialyzed in 1,000 molecular-weight cutoff dialysis tubes, first against a phosphate buffer and then against a mineral solution containing Ca and phosphate. Artificial subsurface lesions of human enamel, created in pH 4.5 acetate buffer, were remineralized for 28 days in 4 remineralizing solutions: group C--mineral solution as a control; group S--mineral solution + dialyzed saliva; group F--mineral solution + 1 ppm F; group SF--mineral solution + dialyzed saliva + 1 ppm F. Changes in relative mineral concentration in the lesions were assessed by transverse microradiography. The results showed statistically significant mineral gains in the lesion body in groups C (DeltaZ = 3,254 +/- 1,562% x microm) and SF (DeltaZ = 2,973 +/- 1,349% x microm), but not in groups S (DeltaZ = 5,192 +/- 1,863% x microm) and F (DeltaZ = 4,310 +/- 1,138% x microm) compared with the baseline group (DeltaZ = 5,414 +/- 461% x microm). It was also found that the mineral density at the surface layer in group F (75.0 +/- 15.7%) was greater than that in the baseline group (30.1 +/- 12.3%) with statistical significance, but not in group SF (39.9 +/- 16.5%). It was concluded that the macromolecules inhibited lesion remineralization fundamentally but that these molecules, in the presence of fluoride, seemed to play an important role in the continuation of remineralization by reducing mineral gains at the surface layer.     

Acid-susceptibility of lesions in bovine enamel after remineralization at different pH values and in the presence of different fluoride concentrations.

Artificial caries lesions were created in sound bovine enamel slices demineralized at pH 5.0. The lesions were then remineralized at either pH 5.5 or pH 6.8 in solutions containing either 1.5, 15, or 50 mumol/L fluoride. The remineralized slices were then demineralized at pH 5.0 for investigation of acid-susceptibility by means of quantitative microradiography. The results indicated that fluoride, incorporated during the preceding remineralization, had a retarding effect on the demineralization after remineralization, and that for the lowest concentration of fluoride in the remineralizing solution, the inhibitory influence of fluoride depended on the pH of the remineralization solutions used.     

The carbonate and fluoride in surfaces of remineralized enamel

Analyses were made of samples of surface enamel collected before and after acid demineralization and after subsequent remineralization in solutions containing calcium and phosphate and various levels of bicarbonate and fluoride ions. Demineralization caused a preferential loss of carbonates. Remineralization was increased by both fluoride and bicarbonate ions. With no bicarbonate in the calcifying solution, 0.25 to 4.0 mM NaF increased mineral formation and thereby caused a relative reduction of its carbonate content. With 0.5 mM NaF (10 ppm F^?) and 5, J5, and 25 mM HCO^?₃, in remineralizing solutions more carbonate was deposited than in the absence of NaF. Five and 15 mM HCO^?₃ increased, but 25 mM HCO^?₃ decreased, deposit of fluoride in enamel.     

Remineralization and acid resistance of enamel lesions after chewing gum containing fluoride extracted from green tea

Background: The aim of this study was to evaluate enamel remineralization and the acquisition of acid resistance by using sugar‐free chewing gum containing fluoride extracted from green tea. Methods: Forty‐five volunteers participated in a crossover, double‐blind study and wore intraoral appliances with human demineralized enamel. Subjects chewed fluoride chewing gum (FCG: 50 μg fluoride) or placebo gum. Remineralization and acid resistance were evaluated using the mineral change value (ΔZ, in vol%·μm). Fluoride concentrations in saliva and remineralized enamel were analysed. Results: The peak salivary fluoride concentration was 3.93 ± 1.28 ppm (mean ± SD). The elevated salivary fluoride concentration resulted in a higher fluoride concentration of 656 ± 95 ppm in the remineralized region versus 159 ± 26 ppm for placebo gum (p < 0.001). After remineralization, the ΔZ of the FCG group was higher than that of the placebo gum group. After an acid challenge, ΔZ of the FCG group was lower than the placebo gum group. Both ΔZ were statistically significant. Conclusions: FCG produced a superior level of remineralization and acid resistance, as compared to the placebo gum. The in situ results suggest that regular use of FCG is useful for preventing dental caries.     

Effects of two fluoride gels on fluoride uptake and phosphorus loss during artificial caries formation

Blocks of human enamel were cycled through a demineralization--F-treatment-remineralization procedure and then analyzed for fluoride and the presence of caries-like lesions. Treatments with a sodium fluoride gel (5000 ppm F) increased the enamel fluoride concentration to 6500 ppm F, whereas a stannous fluoride gel (1000 ppm F) increased enamel fluoride to about 1200 ppm F. Although a control treatment (water) allowed caries-like lesions to form, as observed by microradiography, no lesions were found in either of the fluoride-treated groups. When the experiment was repeated with radioactive teeth, mineral loss, as determined by release of 32P, was again greatest in the water-treated control group, but some loss was observed in the fluoride treatment groups. The least loss was found in the sodium fluoride group. It was concluded that the fluoride treatments not only increased enamel resistance but also enhanced remineralization so that calcium phosphate was replaced during the subsequent remineralization phase. Because of the probability that stannous ions were deposited during the stannous fluoride treatments, some of the apparent calcium phosphate re-deposition in this group was probably stannous compounds.     

Early detection of white spot lesions with digital camera and remineralization therapy

BACKGROUND: In this paper, the characteristics of the early stage of dental caries are discussed and the methods we used to treat the early stage of dental caries to increase the number of caries-free patients are presented. Studies from in vitro to in situ experiments and a clinical study were carried out to support clinical remineralization therapy. METHODS AND RESULTS: To clarify the effect of time for remineralization, the degree of remineralization was assessed at 2 days, 6 days, and 10 days after 2-day demineralization in 0.01 M/L lactic acid buffer (pH 4.0 at 37 degrees ). The remineralization solution contained 3.0 mM/L Ca, 1.8 mM/L P, and 3 ppm fluoride adjusted to pH 7.0. A 10-day continuous remineralization with a 3 ppm fluoride resulted in a high fluoride concentration. To evaluate mineral loss from sound tooth structure and white spot lesions, thin sections (about 90 microm) including white spots (WS) were prepared and exposed to oral conditions for 2 weeks continuously. The mineral loss from sound tooth structure was found to be twice that from WS. In another experiment during the remineralization period, enamel samples were immersed in three different bicarbonate solutions; 0.5, 5.0 and 50 mM/L for 30 minutes, two times per day. Both the bicarbonate and fluoride applied groups showed higher improvement in acid resistance and the amount of remaining mineral was almost two times higher than the controls (p < 0.01). In a clinical study we demonstrated remineralization in patients who followed professional mechanical tooth cleaning and fluoride prophylaxis paste. Using this regime, in patients with deciduous caries present at baseline, over 80 per cent of permanent teeth were caries free at the age of 12 years. In these studies the digital camera with CasMaTCH and an image analysis system showed several advantages for monitoring in de- and remineralization. CONCLUSIONS: White spot lesions, rather than intact tooth surfaces, can be mineralized through the daily clinical procedures described in this paper.