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Objective: To evaluate cytotoxicity and effect on protease activity of epigallocatechin-gallate extracted from experimental restorative dental copolymers in comparison to the control compound chlorhexidine., Methods: Copolymer disks were prepared from bis-GMA/TEGDMA (70/30 mol%) containing no compound (control) or 1% w/w of either epigallocatechin-gallate or chlorhexidine. MDPC-23 odontoblast-like cells were seeded with the copolymer extracts leached out into deionized water. Cell metabolic activity was quantified by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay at 24, 48, 72 h. Inhibition of protease activity by resin extracts was measured by a collagenolytic/genatinolytic enzyme activity assay and gelatin zymography. Data for MTT and protease inhibition were analyzed using two-way ANOVA followed by Tukey or Bonferroni post hoc tests (α=0.05)., Results: The MTT revealed that at 72 h, extracts from control (16.7%) and chlorhexidine (22.3%) copolymers induced significant reduction in cell metabolism (p<0.05). All copolymer extracts caused enzymatic inhibition in a dose dependent manner (p<0.01). Even when highly diluted, epigallocatechin-gallate extract had a significant antiproteolytic activity (p<0.05). Zymograms showed that all extracts reduced activity of MMP-2 and MMP-9 (pro- and active forms), with MMP-9 exhibiting the highest percentage inhibition revealed by densitometry., Conclusions: Epigallocatechin-gallate and chlorhexidine extracts did not exert cytotoxicity on evaluated cells when compared to control extracts. Both compounds retained antiproteolytic activity after extraction from a dental copolymer., Clinical Significance: Once extracted from a dental copolymer, epigallocatechin-gallate is not cytotoxic and retains antiproteolytic activity. These results may allow incorporation of epigallocatechin-gallate as a natural-safe alternative to chlorhexidine in functionalized restorative materials., (Copyright © 2016. Published by Elsevier Ltd.)

Objective: This study attempted to clarify the mechanisms of action of fluoridated acidic liquid dentifrices against dental caries., Design: In the in vitro leg, enamel specimens were submitted to a pH-cycling model, treated with distinct dentifrices (0, 550 μgF/g pH 4.5 and pH 7.0, 1100 or 5000 μgF/g pH 7.0) and analyzed using hardness. Alkali-soluble fluoride (F) deposition was quantified on pre-demineralized specimens treated with the dentifrices. In the clinical leg, 2-to-4-year-old children who had been using liquid dentifrices for 6 months (550 μgF/g pH 4.5 or pH 7.0 or 1100 μgF/g pH 7.0) had their plaque samples collected 5 and 60 min after the last brushing. Fluoride uptake in whole plaque was evaluated., Results: The reduction of the pH had a partial preventive effect on subsurface hardness loss only. [F] had a significant influence on the deposition of fluoride, surface and subsurface hardness loss. In vivo, the reduction of the pH was able to significantly increase plaque F uptake, leading to similar levels as those found for the neutral dentifrice containing twice [F]., Conclusion: The results obtained from in vitro studies whose design does not include the presence of dental plaque should be interpreted with caution., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Objective: To evaluate the effect of ferrous sulphate on enamel demineralization and remineralization, using pH-cycling models., Design: Fifty blocks were selected by their initial surface hardness and subjected to a pH-cycling demineralization process. Artificially demineralized lesions were produced in 60 blocks; out of these blocks, the surface hardness of 50 blocks and the cross-sectional hardness of 10 blocks were determined. The 50 blocks were then subjected to a remineralization pH-cycling process. Treatments were carried out using ferrous sulphate solutions of different concentrations (0.333, 0.840, 18.0, and 70.0 μg Fe/mL) and a control group (deionized water). The final surface hardness (SH(2)) was determined, and the integrated subsurface hardness (ΔKHN) was calculated. The enamel blocks were analysed for fluoride, calcium, phosphorus, and iron. The obtained data were distributed heterogeneously and were analysed using the Kruskal-Wallis test (p<0.05)., Results: In demineralization pH cycling, the group treated with the 18.0 μg Fe/mL solution had higher secondary surface hardness and lower integrated subsurface hardness (ΔKHN) than the other groups. In remineralization pH cycling, the control group showed the lowest value of ΔKHN. A decline in Ca and P concentration was observed when the Fe concentration increased (p<0.05). There was no significant difference in the F concentration (p>0.05) and an increase in Fe concentration (p<0.05) in the enamel was observed when the Fe concentration increased in both the demineralization and remineralization experiments., Conclusion: The results suggest that iron reduces demineralization but does not allow remineralization to occur., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Objective: This in situ/ex vivo study assessed the effect of titanium tetrafluoride (TiF(4)) on permanent human enamel subjected to erosion., Design: Ten volunteers took part in this study performed in two phases. In the first phase (ERO), they wore acrylic palatal appliances containing two enamel blocks, divided into two rows: TiF(4) (F) and no-TiF(4) (no-F). During the 1st day, the formation of a salivary pellicle was allowed. In the 2nd day, the TiF(4) solution was applied on one row (ERO+F), whereas on the other row no treatment was performed (ERO+no-F). From 3rd until 7th day, the blocks were subjected to erosion, 4x per day. In the 2nd phase (no-ERO), the volunteers wore acrylic palatal appliances containing one enamel block, during 2 days, to assess the effect of TiF(4) only (no-ERO+F). Enamel alterations were determined using profilometry (wear), microhardness (%SMHC) tests, scanning electron microscope and microprobe analysis. The %SMHC and wear were tested using ANOVA and Tukey's post hoc tests (p<0.05)., Results: The mean of %SMHC and wear (microm) values (+/-S.D.) were, respectively: ERO+F -73.32+/-5.16(A)/2.40+/-0.60(a); ERO+no-F -83.49+/-4.59(B)/1.17+/-0.48(b) and no-ERO+F -67.92+/-6.16(A)/0.21+/-0.09(c). In microscope analysis, the no-F group showed enamel with honeycomb appearance. For F groups, it was observed a surface coating with microcracks. The microprobe analysis revealed the presence of the following elements (%) in groups ERO+F, ERO+no-F and no-ERO+F, respectively: Ca (69.9, 72.5, 66.25); P (25.9, 26.5, 26.06); Ti (3.0, 0, 5.93)., Conclusions: The TiF(4) was unable to reduce dental erosion.

Objective: This in situ blind crossover study investigated the effect of calcium (Ca) rinse prior to the use fluoride (F) dentifrice on remineralisation of artificially demineralised enamel and on the composition of biofilm., Design: During four phases of 14 days, 10 volunteers wore appliances containing two artificially demineralised bovine enamel blocks. Three times a day, they rinsed with 10 mL of Ca (150 mM) or placebo rinse (1 min). A slurry (1:3, w/v) of F (1030 ppm) or placebo dentifrice was dripped onto the blocks. During 1 min, the volunteers brushed their teeth with the respective dentifrice. The appliance was replaced into the mouth and the volunteers rinsed with water. The biofilm formed on the blocks was analysed for F and Ca. Enamel alterations were evaluated by the percentage of surface microhardness change (%SMHC), cross-sectional microhardness (% mineral volume) and alkali-soluble F analysis. Data were analysed by ANOVA (p<0.05)., Results: The use of the Ca pre-rinse before the F dentifrice produced a six- and four-fold increase in biofilm F and Ca concentrations, respectively. For enamel, the remineralisation was significantly improved by the Ca pre-rinse when compared to the other treatments. There was a significantly higher concentration of alkali-soluble F in enamel when the F dentifrice was used, but the Ca pre-rinse did not have any significant additive effect., Conclusions: According to our protocol, the Ca pre-rinse significantly increased biofilm F concentration and, regardless the use of F dentifrice, significantly enhanced the remineralisation of artificially demineralised enamel.

Objectives: The aim of this study was to evaluate, in vitro, the effect of iron on the acid demineralisation of bovine enamel blocks by a soft drink., Design: Twenty-four blocks of bovine enamel were randomly assigned to two groups (experimental and control), which were exposed to 4 cycles of demineralisation in Coke containing or not 10 mmol/L of iron, respectively, and another 4 cycles of remineralisation in artificial saliva. The softening of enamel was evaluated by % superficial microhardness change (SMHC) and wear analysis. Data were analysed using t-test (p<0.05)., Results: With regard to %SMHC, the experimental group presented average values (+/-S.E.) of -85.3+/-0.9, that were significantly higher (t=4.7, p<0.01) than the ones observed for control group (-76.7+/-1.6). The experimental group presented a mean wear (+/-S.E.) of 2.3+/-0.2 microm, that was significantly lesser (t=3.7, p<0.01) than the one observed for the control group (3.5+/-0.2 microm)., Conclusions: According to this protocol, iron at 10 mmol/L significantly reduced the wear, but significantly enhanced the %SMHC of enamel blocks submitted to erosion by Coke.

Objectives: The aim of this study was to evaluate, in vitro, the effect of iron on the dissolution of bovine enamel powder, when added to two carbonated beverages., Design: Powdered enamel was produced by griding enamel fragments of bovine incisor in a steel pestle and mortar. Particles between 75 and 106 microm were selected using appropriated meshes. At time zero, the carbonated beverage (Coke or Sprite Zero) was added to powdered enamel (1 mg enamel powder/10 microL of beverage) and vortexed for 30 s. The sample was immediately centrifuged (11,000 rpm) for 30 s and the supernatant was removed at 1 min 40 s. This procedure was repeated five times with the beverage containing increasing ferrous sulphate concentrations (1.25, 2.5, 5, 10, 15, 30 and 60 mmol/L). The phosphate released in the medium was analysed spectrophotometrically. Data were analysed using ANOVA and Tukey's test (p<0.05)., Results: When iron at 30 and 60 mmol/L was added to Coke, a significant reduction in the dissolution of powdered enamel was observed when compared to control (11 and 17%, respectively), while lower iron concentrations did not have any effect on enamel powder dissolution. Regarding Sprite Zero, iron concentrations up to 10 mmol/L had no significant effect, while higher concentrations significantly increased enamel powder dissolution., Conclusions: The results suggest that iron can interfere with the dissolution of dental enamel powder in the presence of acidic beverages and the type of acid in these beverages seems to modulate this effect.

Objectives: Iron ions (Fe(2+)) have been shown to be cariostatic in many studies particularly by their ability to reduce bacterial metabolism. Nevertheless, the role of iron ions on dissolution of enamel is unexplored. The aim of the present study was therefore to investigate the protective effect of increasing concentrations (0-120mmol/L) of Fe(2+) on the dissolution of enamel., Design: Enamel powder was subjected to acetic acid made with increasing concentrations with respect to FeSO(4)x7H(2)O. In order to determine the amount of enamel dissolved, the phosphate released in the medium was analysed spectrophotometrically using the Fiske-Subarrow method. Data were tested using Kruskall-Wall and Dunn's tests (p<0.05). The degree of protection was found to approach maximum at about 15mmol/L Fe(2+). Higher concentrations of Fe(2+) did not have an extra effect on inhibition of dissolution of enamel powder. In the next step, the protective effect of 15mmol/L Fe(2+) against mineral dissolution of the bovine enamel was evaluated using a simple abiotic model system. Enamel blocks were exposed to a sequence of seven plastic vials, each containing 1mL of 10mmol/L acetic acid. The acid in vial 4 was made 15mmol/L with respect to FeSO(4)x7H(2)O. The mineral dissolved during each challenge was thus determined by phosphate released as described above. Data were tested using two-way ANOVA (p<0.05)., Results: Lower demineralisation (around 45%) was found in vial 4 (with Fe) that continued stable until vial 7., Conclusions: Thus, our data suggest that Fe(2+) can be effective on inhibition of dissolution of enamel and that this effect may be durable.

Objectives: This study investigated in situ the effect of iron (Fe) on the reduction of demineralization of bovine enamel, as well as on the composition of dental biofilm., Design and Methods: Twelve volunteers were included in this blind crossover study, which was conducted in two stages of 14 days each. For each stage, the volunteers received palatal appliances containing four blocks of bovine enamel (4 mm x4 mm x 2.5 mm). Six volunteers dripped a solution of 15 mmol L(-1) ferrous sulphate onto the fragments and the remaining six dripped deionized water (eight times per day). After five minutes, a fresh 20% (w/v) sucrose solution was dripped onto all enamel blocks. During the experimental period the volunteers brushed their teeth with non-fluoridated dentifrice. After each stage, the percentage of surface microhardness change (%SMHC) and area of mineral loss (DeltaZ) were determined on enamel and the dental biofilm formed on the blocks was collected and analysed for F, P, Ca, Fe and alkali-soluble carbohydrates. The concentrations of F, Ca and Fe in enamel were also analysed after acid biopsies., Results: There was a statistically significant increase in the P and Fe concentrations in the biofilms treated with ferrous sulphate (p<0.05), which was not observed for F, Ca and alkali-soluble carbohydrates. The group treated with ferrous sulphate had significantly lower %SMHC and DeltaZ when compared to control (p<0.05)., Conclusions: These results showed that ferrous sulphate reduced the demineralization of enamel blocks and altered the ionic composition of the dental biofilm formed in situ.