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Effect of stannous and fluoride concentration in a mouth rinse on erosive tissue loss in enamel in vitro
archives of oral biology 54 (2009) 432–436
available at www.sciencedirect.com
journal homepage: www.intl.elsevierhealth.com/journals/arob
Effect of stannous and fluoride concentration in a mouth rinse on erosive tissue loss in enamel in vitro N. Schlueter *, J. Klimek, C. Ganss Department of Conservative and Preventive Dentistry, Dental Clinic of the Justus Liebig University, Schlangenzahl 14, D-35392 Giessen, Germany
article info
abstract
Article history:
The aim of this in vitro study was to investigate the influence of stannous and fluoride ion
Accepted 31 January 2009
concentrations in various experimental solutions on erosion progression in enamel. Human enamel specimens were subjected to a cyclic de- and remineralisation procedure for 10
Keywords:
days, with six demineralisation periods per day, of 5 min each. Erosive demineralisation was
Erosion
performed with 0.05 M citric acid (pH 2.3). Except in the control group, specimens were
Enamel
treated for 2 min with test solutions after the first and the sixth demineralisation. Test
Tin
solutions were: 1500 mg/L F groups: group 1: 2800 mg/L Sn2+; group 2: 2100 mg/L Sn2+; group
Fluoride
3: 1400 mg/L Sn2+; group 4: 700 mg/L Sn2+; 1000 mg/L F groups: group 5: 2100 mg/L Sn2+; group 6: 1400 mg/L Sn2+. All preparations were adjusted to pH 4.5. Tissue loss was determined profilometrically after the last experimental day. As expected, the greatest tissue loss (mm, mean S.D.) was found in the control group (72.6 11.5). All test solutions were able to reduce tissue loss significantly ( p 0.001). The reduction of tissue loss by test solutions depended on the ratio of the tin concentration to the fluoride concentration. Lowest values were obtained by the application of the solutions of group 1 (7.8 2.5) and group 5 (7.6 5.2). Solutions with high concentrations of tin and fluoride are very effective in reducing erosive tissue loss, and their efficacy increased with increasing ratios of tin to fluoride concentrations. # 2009 Elsevier Ltd. All rights reserved.
1.
Introduction
Fluorides are a key component in oral health promotion, especially in the prevention of damage of the oral hard tissues.1 Due to such properties, several fluoride compounds, such as NaF, SnF2 or TiF4, have been investigated for use in products for oral health. In particular, stannous fluoride containing preparations have proved to be very effective, where their therapeutic benefits were established as early as five decades ago.2 In addition to its antimicrobial properties, the efficacy of tin in protection against demineralisation is attributed to its ability to react with3,4 and to modify the tooth surface,5 and
this effect is likely to result in a greater resistance to decay. Indeed, several studies have investigated tin’s properties of conferring protection against caries, and the results have been very promising. In a caries model, the solubility of enamel, with demineralisation by lactic acid, was reduced by approximately 80% with the application of a 0.2% SnF2 solution.6 In another study, SnF2 treatment resulted in a greater reduction of enamel solubility compared with NaF treatment.7 In addition, in experiments with rats given SnF2 treatment, the caries occurrence was reduced by approximately 60% at coronal surfaces and by approximately 90% at root surfaces.8 Even in an experiment with severe etching using 2N HCl, SnF2 showed promising results. The application of SnF2 alone, but
* Corresponding author. Tel.: +49 641 99 46173; fax: +49 641 99 46169. E-mail address: [email protected] (N. Schlueter). 0003–9969/$ – see front matter # 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.archoralbio.2009.01.019
archives of oral biology 54 (2009) 432–436
also the combined application of AmF and SnF2 decreased enamel solubility to a significantly greater extent than the application of NaF or AmF.9–12 Although stannous fluoride-containing preparations have been promoted as the agent of choice for reducing enamel solubility, it has been less frequently used (at least in Europe2), which may be due to the limited stability of SnF2 in dentifrice or the poor formulation flexibility, particularly as a result of its flavour. Therefore, alternative fluorides, such as NaF, amine fluoride (AmF) or sodium monofluorophosphate have been preferred.13 In the last few years, stannous fluoride has again become the focus of research. Recently, numerous studies have investigated its properties on enamel solubility under more severe acidic conditions and its potential as a preventive agent against erosive substance loss. Without exception, studies of treatments with stannous ions combined with fluoride have shown very promising results, with 50–90% reductions in mineral loss.14–19 But even in the absence of fluoride, treatments with stannous ions showed a reduction of mineral loss by 64%.14 The studies mentioned above were greatly variable in their results, probably due to the differences in the study design, differences in the demineralisation solutions used or differences in concentrations of the stannous (400–30,000 mg/L Sn2+) and fluoride ions. Thus, these differences do not allow a direct comparison of the results or allow a statement about the efficacy of the stannous or fluoride ion. To clarify whether there is a dose–response relationship between tin or fluoride concentration and tissue loss, a systematic testing using different solutions containing various concentrations of stannous and fluoride ions for their erosion inhibiting properties under standardised erosive conditions seems to be necessary. Therefore, the aim of this in vitro study was to compare the efficacy of different pH adjusted AmF/NaF/SnCl2 containing solutions with varying Sn2+ and F concentrations in reducing erosive tissue loss in cyclic de- and remineralised enamel. The null hypothesis tested was that there is no difference between the various solutions.
2.
Material and methods
2.1.
Specimen preparation
Eighty-four longitudinal enamel specimens were prepared from 30 freshly extracted, completely impacted human third molars without cracks. All donors lived in an area with 0.03 mg/L fluoride in the drinking water. The natural surfaces of specimens were ground flat and polished under sufficient water flow (Exakt Abrasive Cutting System and Exakt Mikrogrinder, Exakt-Apparatebau, Norderstedt, Germany; P800 and P1200 silicon carbide abrasive paper, Leco, St. Joseph, USA). The preparation resulted in an experimental area of at least 3 3 mm2. Specimens were randomly divided into seven groups (n = 12 each) and mounted on microscope slides (R. Langenbrinck, Teningen, Germany) with a light curing acrylate (Technovit 7230 VLC, Kulzer-Exakt, Wehrheim, Germany) with six specimens on one slide. One half of the
433
experimental area of each specimen was covered with acrylate. This area served as the reference area for the profilometric measurement. After acrylate covering, the specimens were scrutinised under a microscope (magnification 10, SMZ-1, Zoom Stereomicroscope, Nikon GmbH, Du¨sseldorf, Germany) to ensure that there was no contamination by the acrylate on the experimental area. Specimens were stored in 100% humidity until use.
2.2.
Treatment
Specimens were subjected to a cyclic de- and remineralisation procedure with six demineralisation periods per day, 5 min each. For erosive demineralisation, a 0.05 M citric acid solution (pH 2.3, anhydrous citric acid, Merck, Darmstadt, Germany) was used. The de- and remineralisation cycles were performed over a total of 10 days. In all groups except the control group, specimens were treated with test solutions after the first and after the last demineralisation periods, for 2 min each. Six different fluoride- and tin-containing solutions (for details see Table 1) were used. All experimental solutions were adjusted to pH 4.5. Slides with specimens were put into special racks that were then transferred to a container filled with 250 mL of the respective solution to achieve a constant immersion time. Prior to the transfer to the next solution, specimens were rinsed for 1 min with tap water. During the remaining time, specimens were stored in a remineralisation solution20 (4.08 mM H3PO4, 20.10 mM KCl, 11.90 mM Na2CO3, and 1.98 mM CaCl2 (chemicals from Merck), with a pH of 6.7). Solutions were renewed at the beginning of each experimental day, and the pH of all solutions was controlled at the beginning and at the end of each experimental day. All groups were treated simultaneously. All procedures were performed under gentle agitation at room temperature.
2.3.
Tissue loss measurement
Tissue loss was measured profilometrically after the last experimental day. The acrylic cover was carefully removed and the surfaces were checked for acrylate remnants or damage. The measurements were performed with a Perthometer S8P (Mahr, Go¨ttingen, Germany), equipped with an optical stylus (Focodyn, Rodenstock, Munich, Germany). On each sample, three traces, each of 1.75 mm in length, were made at intervals of 0.25 mm. The traces were interpreted with special software (Perthometer Concept 4.0, Perthen Mahr, Go¨ttingen, Germany). Two regression lines were constructed on each trace. The first one on the reference area was 0.4 mm in length, and the second one on the experimental area was 0.6 mm in length; both lines were at a distance of 0.3 mm from the edge between the reference and experimental areas. The mid-points of both regression lines were calculated by software. The vertical distance between the mid-points was defined as tissue loss. In cases of distinct substance loss, the reproducibility (10 repeated tracings of one sample) was 0.8 mm and in cases of slight substance loss, the reproducibility was 0.9 mm. The repeated analysis of one trace showed a standard deviation of 0.1 mm. The variance of profiles of untreated specimens was 0.3 0.5 mm.
434
archives of oral biology 54 (2009) 432–436
Table 1 – Compositions of the experimental solutions, all adjusted to pH 4.5. Group
Total fluoride content (mg/L)
Compound
Fluoride concentration (mg/L)
Group 1
1500
0.98% (w/v) amine fluoridea 0.17% (w/v) NaF 0.53% (w/v) SnCl2
750 750
1500
a
750 750
Group 2
Group 3
Group 4
Group 5
Group 6
a
0.98% (w/v) amine fluoride 0.17% (w/v) NaF 0.40% (w/v) SnCl2
2100 Sn2+ 750 750
0.32% (w/v) amine fluoridea 0.28% (w/v) NaF 0.13% (w/v) SnCl2
250 1250
1000
0.64% (w/v) amine fluoridea 0.11% (w/v) NaF 0.4% (w/v) SnCl2
500 500
1000
a
250 750
1500
0.32% (w/v) amine fluoride 0.17% (w/v) NaF 0.27% (w/v) SnCl2
1400 Sn2+
700 Sn2+
2100 Sn2+
1400 Sn2+
Olaflur, all solutions provided by GABA International AG, Muenchenstein, Switzerland.
2.4.
Statistics
The statistical analysis of data was performed with SPSS 11.0 for Windows (SPSS, Chicago, IL, USA). The Kolmogorov– Smirnov test was used for checking the normal distribution of data. An analysis of variance (ANOVA) with Tukey’s post hoc test was performed to compare the groups. The level of significance was set at 0.05.
3.
2800 Sn2+
0.98% (w/v) amine fluoridea 0.17% (w/v) NaF 0.27% (w/v) SnCl2
1500
Tin concentration (mg/L)
Results
procedures were similar to our own previously performed studies, which helps to achieve comparability with our other results. The solutions were applied after the erosive impact, which reflects the clinical situation. Mostly, patients already have dental erosion, meaning that the dental hard tissue has already been affected by acids and the solutions are intended also as a therapeutic measure. The application frequency of the experimental solutions could possibly be used in clinical practice and in at home applications. The concentrations chosen for the active agents were pursuant to the directives for dental hygiene products and cosmetics. Because the current recommendation for symptomatic therapy of dental
The results are displayed in Fig. 1, and the values are given as mean S.D. The tissue loss was highest in the control group (72.6 11.5 mm). All test solutions reduced tissue loss significantly compared with the control group ( p 0.001). Within the 1500 mg/L F groups, tissue loss was highest in the 700 mg/ L Sn2+ group (group 4: 30.3 10.4 mm), and it decreased with an increasing stannous concentration (group 3 – 1400 mg/L Sn2+: 23.3 6.7 mm; group 2 – 2100 mg/L Sn2+: 19.6 4.6 mm; group 1 – 2800 mg/L Sn2+: 7.8 2.5 mm). The same trend was observed for the 1000 mg/L F solutions. The tissue loss was highest in the 1400 mg/L Sn2+ group (group 6: 18.6 2.8 mm), whereas in the 2100 mg/L Sn2+ group, lower values were found (group 5 – 2100 mg/L Sn2+: 7.6 5.2 mm).
4.
Discussion
This study investigated the impact of the tin and the fluoride concentrations of different solutions on erosion progression. Relatively severe erosive conditions were chosen to simulate the clinical conditions that occur in patients at high risk of erosion, such as vegetarians21 and patients with eating disorders, who can have frequent acid attacks. The erosive
Fig. 1 – Tissue loss (mm) in all groups after 10 days cyclic deand remineralisation (6 T 5 min erosive demineralisation with 0.05 M citric acid and 2 T 2 min treatment with test solutions per day). No statistical significance of differences between groups is indicated by corresponding indices (ANOVA with Tukey’s post hoc).
archives of oral biology 54 (2009) 432–436
erosion is frequent application of high amounts of fluoride,22– 24 so experimental solutions with high concentrations of fluoride were used. The pH was 4.5, which make the solutions potentially applicable for daily mouth rinses. However, at this pH, solutions containing stannous ions in relatively high concentrations have limited stability. Amine fluoride was used in all tested solutions, since some studies have found that it has stabilising properties.25,26 Stannous chloride, which has also shown marked efficacy in vitro in reducing erosion in enamel,14 was used as an independent source of tin. As expected, the highest tissue loss was found in the control group. All tin–fluoride preparations significantly reduced erosive tissue loss by approximately 60–90% when compared with the control group. Both results were comparable with those from a previous study.14 Concerning the fluoride concentration of the solutions, there was no indication that the solutions with higher fluoride concentrations were more effective than those with lower concentrations, as would be expected from the literature.27 Rather, in solutions with the same concentration of tin, the solution with a lower fluoride concentration was more effective than the one with a higher fluoride concentration, as evidenced by the comparison of the efficacy of groups 2 (1500 mg/L F , 2100 mg/L Sn2+) and 5 (1000 mg/L F , 2100 mg/L Sn2+). Hence, in the presence of tin ions, the fluoride concentration seems to play a secondary role, at least in the range of the investigated concentrations. The tin concentration appears more important. The efficacy of the solutions increased nearly linearly with an increasing tin concentration among the 1500 mg/L F solutions. Similar results had been found by Wachtel.6 This study investigated the effect of a single application of various stannous ion-containing solutions on enamel dissolution in a caries model. The results have clearly shown that with an increasing stannous ion concentration, the protection against enamel dissolution increases.6 However, the efficacy of the tested solutions did not depend solely on the absolute concentrations of the stannous or fluoride ions. Comparing the groups 1 (1500 mg/L F , 2800 mg/L Sn2+) and 5 (1000 mg/L F , 2100 mg/L Sn2+), it is notable that they gave nearly identical results, even though the latter solution contained one third less fluoride and one fourth less tin than the first. A similar pattern was observed when comparing groups 2 (1500 mg/L F , 2100 mg/L Sn2+) and 6 (1000 mg/L F , 1400 mg/L Sn2+). This indicated that not only were the absolute concentrations important, but the ratio between the stannous and the fluoride ions was also relevant. The calculated tin to fluoride ratios were similar or even identical in the compared groups: 1.87 and 2.1 for groups 1 and 5, respectively, and 1.4 for both groups 2 and 6. The results indicated that the higher the ratio, the better the solution. Furthermore, the results indicated that solutions with ratios above 1.8 were very effective and that those with ratios of 1.4 or below had lower efficacy. These results are in agreement with those of Barbakow et al.9 This study showed that SnF2/ AmF solutions with a tin to fluoride ratio of 1.56 or higher were of marked efficacy in reducing mineral loss in enamel after an HCl treatment (2N), but solutions with a lower ratio (0.78) were notably less effective.9 Currently, it is unclear as to which ratio can be identified as the ‘‘cut-off-point’’ for
435
maximal efficacy of fluoride solutions containing tin and to what extent the fluoride or tin concentrations can be reduced for a specific level of efficacy. These considerations, however, disregard the AmF/NaF ratio. Further studies considering these ratios of tin and the various fluoride compounds could provide more information. However, based on the present results, it is quite conceivable, that one can reduce the fluoride and tin concentrations to a certain amount in solutions to be used as a mouth rinse for the symptomatic therapy of dental erosion, in order to maximize stability and flavour.
Acknowledgement The study was supported by GABA International AG, Muenchenstein, Switzerland. Funding: Supported by GABA International AG, Muenchenstein, Switzerland. Conflict of interest: None. Ethical Approval: None.
references
1. Fejerskov O. Changing paradigms in concepts on dental caries: consequences for oral health care. Caries Res 2004;38:182–91. 2. Brambilla E. Fluoride – is it capable of fighting old and new dental diseases? An overview of existing fluoride compounds and their clinical applications. Caries Res 2001;35:6–9. 3. Babcock FD, King JC, Jordan TH. The reaction of stannous fluoride and hydroxyapatite. J Dent Res 1978;57:933–8. 4. Cooley WE. Reactions of tin(II) and fluoride ions with etched enamel. J Dent Res 1961;40:1199–210. 5. Krutchkoff DJ, Jordan TH, Wei SHY, Nordquist WD. Surface characterization of the stannous fluoride–enamel interaction. Arch Oral Biol 1972;17:923–30. 6. Wachtel LW. In vitro comparison of effects of topical stannic fluoride and stannous fluoride solutions on enamel solubility. Arch Oral Biol 1964;9:439–45. 7. Gray JA. Acid dissolution rate of sound and white-spot enamel treated with tin(II) and fluoride compounds. J Dent Res 1965;44:493–501. 8. Beiraghi S, Rosen S, Beck F. The effect of stannous and sodium fluoride on coronal caries, root caries and bone loss in rice rats. Arch Oral Biol 1990;35:79–80. 9. Barbakow F, Lutz F, Scherle W. Solubility changes of human enamel treated with precipitate-free, aged SnF2/amine fluoride 297 solutions. J Dent Child 1986;53:415–9. 10. Barbakow F, Sener B, Lutz F. Dissolution of phosphorus from human enamel pretreated in vitro using SnF2 stabilized with amine fluoride 297. Clin Prev Dent 1987;9:3–6. 11. Barbakow F, Lutz F, Sener B, Ransberger M. Dissolution rates of enamel immersed in precipitate-free, aged solutions of SnF2 and amine fluoride 297. Schweiz Mschr Zahnmed 1986;96:1494–503. 12. Barbakow F, Lutz F, Sener B. In vitro dissolution of human enamel after application of a mixture of stannous fluoride and amine fluoride 297: a pilot study. ASDC J Dent Child 1985;52:444–8. 13. White DJ. A ‘‘return’’ to stannous fluoride dentifrices. J Clin Dent 1995;6(Spec No):29–36.
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14. Ganss C, Schlueter N, Hardt M, Schattenberg P, Klimek J. Effect of fluoride compounds on enamel erosion in vitro – a comparison of amine, sodium and stannous fluoride. Caries Res 2008;42:2–7. 15. Hove L, Holme B, Ogaard B, Willumsen T, Tveit AB. The protective effect of TiF4, SnF2 and NaF on erosion of enamel by hydrochloric acid in vitro measured by white light interferometry. Caries Res 2006;40:440–3. 16. Willumsen T, Ogaard B, Hansen BF, Rolla G. Effects from pretreatment of stannous fluoride versus sodium fluoride on enamel exposed to 0.1 M or 0. 01 M hydrochloric acid. Acta Odontol Scand 2004;62:278–81. 17. Hjortsjo¨ C, Jonski G, Ro¨lla G, Thrane PS, Saxegaard E, Young A. Short term effect of NaF, SnF2 and dilute HF treatments on enamel erosion-like lesions in vivo. Caries Res 2008;42:189. 18. Young A, Thrane PS, Saxegaard E, Jonski G, Ro¨lla G. Effect of stannous fluoride toothpaste on erosion-like lesions: an in vivo study. Eur J Oral Sci 2006;114:180–3. 19. Hooper SM, Newcombe RG, Faller R, Eversole S, Addy M, West NX. The protective effects of toothpaste against erosion by orange juice: studies in situ and in vitro. J Dent 2007;35:476–81.
20. Gerrard WA, Winter PJ. Evaluation of toothpastes by their ability to assist rehardening of enamel in vitro. Caries Res 1986;20:209–16. 21. Ganss C, Schlechtriemen M, Klimek J. Dental erosions in subjects living on a raw food diet. Caries Res 1999;33:74–80. 22. Ganss C, Klimek J, Scha¨ffer U, Spall T. Effectiveness of two fluoridation measures on erosion progression in human enamel and dentine in vitro. Caries Res 2001;35:325–30. 23. Wiegand A, Attin T. Influence of fluoride on the prevention of erosive lesions—a review. Oral Health Prev Dent 2003;1:245–53. 24. Ganss C, Klimek J, Brune V, Schu¨rmann A. Effects of two fluoridation measures on erosion progression in human enamel and dentine in situ. Caries Res 2004;38:561–6. 25. Mu¨hlemann HR, Saxer UP. Reduction of plaque and gingivitis by stannous fluoride stabilised with amine fluoride. Caries Res 1981;15:186. [Abstr.]. 26. Mu¨hlemann HR, Schmid H. GABA Int. AG, editor. Orale Kompositionen mit stabilisierten Zinnsalzen. Basel/ Switzerland patent Europ. Pat. NR. 0 026 539. 1985. 27. Abdullah AZ, Strafford SM, Brookes SJ, Toumba KJ, Barlow AP, Smith SR, et al. The effect of fluoride on tooth surface loss of enamel under erosive cycling challenge. Caries Res 2008;42:190. [Abstr.].
available at www.sciencedirect.com
journal homepage: www.intl.elsevierhealth.com/journals/arob
Effect of stannous and fluoride concentration in a mouth rinse on erosive tissue loss in enamel in vitro N. Schlueter *, J. Klimek, C. Ganss Department of Conservative and Preventive Dentistry, Dental Clinic of the Justus Liebig University, Schlangenzahl 14, D-35392 Giessen, Germany
article info
abstract
Article history:
The aim of this in vitro study was to investigate the influence of stannous and fluoride ion
Accepted 31 January 2009
concentrations in various experimental solutions on erosion progression in enamel. Human enamel specimens were subjected to a cyclic de- and remineralisation procedure for 10
Keywords:
days, with six demineralisation periods per day, of 5 min each. Erosive demineralisation was
Erosion
performed with 0.05 M citric acid (pH 2.3). Except in the control group, specimens were
Enamel
treated for 2 min with test solutions after the first and the sixth demineralisation. Test
Tin
solutions were: 1500 mg/L F groups: group 1: 2800 mg/L Sn2+; group 2: 2100 mg/L Sn2+; group
Fluoride
3: 1400 mg/L Sn2+; group 4: 700 mg/L Sn2+; 1000 mg/L F groups: group 5: 2100 mg/L Sn2+; group 6: 1400 mg/L Sn2+. All preparations were adjusted to pH 4.5. Tissue loss was determined profilometrically after the last experimental day. As expected, the greatest tissue loss (mm, mean S.D.) was found in the control group (72.6 11.5). All test solutions were able to reduce tissue loss significantly ( p 0.001). The reduction of tissue loss by test solutions depended on the ratio of the tin concentration to the fluoride concentration. Lowest values were obtained by the application of the solutions of group 1 (7.8 2.5) and group 5 (7.6 5.2). Solutions with high concentrations of tin and fluoride are very effective in reducing erosive tissue loss, and their efficacy increased with increasing ratios of tin to fluoride concentrations. # 2009 Elsevier Ltd. All rights reserved.
1.
Introduction
Fluorides are a key component in oral health promotion, especially in the prevention of damage of the oral hard tissues.1 Due to such properties, several fluoride compounds, such as NaF, SnF2 or TiF4, have been investigated for use in products for oral health. In particular, stannous fluoride containing preparations have proved to be very effective, where their therapeutic benefits were established as early as five decades ago.2 In addition to its antimicrobial properties, the efficacy of tin in protection against demineralisation is attributed to its ability to react with3,4 and to modify the tooth surface,5 and
this effect is likely to result in a greater resistance to decay. Indeed, several studies have investigated tin’s properties of conferring protection against caries, and the results have been very promising. In a caries model, the solubility of enamel, with demineralisation by lactic acid, was reduced by approximately 80% with the application of a 0.2% SnF2 solution.6 In another study, SnF2 treatment resulted in a greater reduction of enamel solubility compared with NaF treatment.7 In addition, in experiments with rats given SnF2 treatment, the caries occurrence was reduced by approximately 60% at coronal surfaces and by approximately 90% at root surfaces.8 Even in an experiment with severe etching using 2N HCl, SnF2 showed promising results. The application of SnF2 alone, but
* Corresponding author. Tel.: +49 641 99 46173; fax: +49 641 99 46169. E-mail address: [email protected] (N. Schlueter). 0003–9969/$ – see front matter # 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.archoralbio.2009.01.019
archives of oral biology 54 (2009) 432–436
also the combined application of AmF and SnF2 decreased enamel solubility to a significantly greater extent than the application of NaF or AmF.9–12 Although stannous fluoride-containing preparations have been promoted as the agent of choice for reducing enamel solubility, it has been less frequently used (at least in Europe2), which may be due to the limited stability of SnF2 in dentifrice or the poor formulation flexibility, particularly as a result of its flavour. Therefore, alternative fluorides, such as NaF, amine fluoride (AmF) or sodium monofluorophosphate have been preferred.13 In the last few years, stannous fluoride has again become the focus of research. Recently, numerous studies have investigated its properties on enamel solubility under more severe acidic conditions and its potential as a preventive agent against erosive substance loss. Without exception, studies of treatments with stannous ions combined with fluoride have shown very promising results, with 50–90% reductions in mineral loss.14–19 But even in the absence of fluoride, treatments with stannous ions showed a reduction of mineral loss by 64%.14 The studies mentioned above were greatly variable in their results, probably due to the differences in the study design, differences in the demineralisation solutions used or differences in concentrations of the stannous (400–30,000 mg/L Sn2+) and fluoride ions. Thus, these differences do not allow a direct comparison of the results or allow a statement about the efficacy of the stannous or fluoride ion. To clarify whether there is a dose–response relationship between tin or fluoride concentration and tissue loss, a systematic testing using different solutions containing various concentrations of stannous and fluoride ions for their erosion inhibiting properties under standardised erosive conditions seems to be necessary. Therefore, the aim of this in vitro study was to compare the efficacy of different pH adjusted AmF/NaF/SnCl2 containing solutions with varying Sn2+ and F concentrations in reducing erosive tissue loss in cyclic de- and remineralised enamel. The null hypothesis tested was that there is no difference between the various solutions.
2.
Material and methods
2.1.
Specimen preparation
Eighty-four longitudinal enamel specimens were prepared from 30 freshly extracted, completely impacted human third molars without cracks. All donors lived in an area with 0.03 mg/L fluoride in the drinking water. The natural surfaces of specimens were ground flat and polished under sufficient water flow (Exakt Abrasive Cutting System and Exakt Mikrogrinder, Exakt-Apparatebau, Norderstedt, Germany; P800 and P1200 silicon carbide abrasive paper, Leco, St. Joseph, USA). The preparation resulted in an experimental area of at least 3 3 mm2. Specimens were randomly divided into seven groups (n = 12 each) and mounted on microscope slides (R. Langenbrinck, Teningen, Germany) with a light curing acrylate (Technovit 7230 VLC, Kulzer-Exakt, Wehrheim, Germany) with six specimens on one slide. One half of the
433
experimental area of each specimen was covered with acrylate. This area served as the reference area for the profilometric measurement. After acrylate covering, the specimens were scrutinised under a microscope (magnification 10, SMZ-1, Zoom Stereomicroscope, Nikon GmbH, Du¨sseldorf, Germany) to ensure that there was no contamination by the acrylate on the experimental area. Specimens were stored in 100% humidity until use.
2.2.
Treatment
Specimens were subjected to a cyclic de- and remineralisation procedure with six demineralisation periods per day, 5 min each. For erosive demineralisation, a 0.05 M citric acid solution (pH 2.3, anhydrous citric acid, Merck, Darmstadt, Germany) was used. The de- and remineralisation cycles were performed over a total of 10 days. In all groups except the control group, specimens were treated with test solutions after the first and after the last demineralisation periods, for 2 min each. Six different fluoride- and tin-containing solutions (for details see Table 1) were used. All experimental solutions were adjusted to pH 4.5. Slides with specimens were put into special racks that were then transferred to a container filled with 250 mL of the respective solution to achieve a constant immersion time. Prior to the transfer to the next solution, specimens were rinsed for 1 min with tap water. During the remaining time, specimens were stored in a remineralisation solution20 (4.08 mM H3PO4, 20.10 mM KCl, 11.90 mM Na2CO3, and 1.98 mM CaCl2 (chemicals from Merck), with a pH of 6.7). Solutions were renewed at the beginning of each experimental day, and the pH of all solutions was controlled at the beginning and at the end of each experimental day. All groups were treated simultaneously. All procedures were performed under gentle agitation at room temperature.
2.3.
Tissue loss measurement
Tissue loss was measured profilometrically after the last experimental day. The acrylic cover was carefully removed and the surfaces were checked for acrylate remnants or damage. The measurements were performed with a Perthometer S8P (Mahr, Go¨ttingen, Germany), equipped with an optical stylus (Focodyn, Rodenstock, Munich, Germany). On each sample, three traces, each of 1.75 mm in length, were made at intervals of 0.25 mm. The traces were interpreted with special software (Perthometer Concept 4.0, Perthen Mahr, Go¨ttingen, Germany). Two regression lines were constructed on each trace. The first one on the reference area was 0.4 mm in length, and the second one on the experimental area was 0.6 mm in length; both lines were at a distance of 0.3 mm from the edge between the reference and experimental areas. The mid-points of both regression lines were calculated by software. The vertical distance between the mid-points was defined as tissue loss. In cases of distinct substance loss, the reproducibility (10 repeated tracings of one sample) was 0.8 mm and in cases of slight substance loss, the reproducibility was 0.9 mm. The repeated analysis of one trace showed a standard deviation of 0.1 mm. The variance of profiles of untreated specimens was 0.3 0.5 mm.
434
archives of oral biology 54 (2009) 432–436
Table 1 – Compositions of the experimental solutions, all adjusted to pH 4.5. Group
Total fluoride content (mg/L)
Compound
Fluoride concentration (mg/L)
Group 1
1500
0.98% (w/v) amine fluoridea 0.17% (w/v) NaF 0.53% (w/v) SnCl2
750 750
1500
a
750 750
Group 2
Group 3
Group 4
Group 5
Group 6
a
0.98% (w/v) amine fluoride 0.17% (w/v) NaF 0.40% (w/v) SnCl2
2100 Sn2+ 750 750
0.32% (w/v) amine fluoridea 0.28% (w/v) NaF 0.13% (w/v) SnCl2
250 1250
1000
0.64% (w/v) amine fluoridea 0.11% (w/v) NaF 0.4% (w/v) SnCl2
500 500
1000
a
250 750
1500
0.32% (w/v) amine fluoride 0.17% (w/v) NaF 0.27% (w/v) SnCl2
1400 Sn2+
700 Sn2+
2100 Sn2+
1400 Sn2+
Olaflur, all solutions provided by GABA International AG, Muenchenstein, Switzerland.
2.4.
Statistics
The statistical analysis of data was performed with SPSS 11.0 for Windows (SPSS, Chicago, IL, USA). The Kolmogorov– Smirnov test was used for checking the normal distribution of data. An analysis of variance (ANOVA) with Tukey’s post hoc test was performed to compare the groups. The level of significance was set at 0.05.
3.
2800 Sn2+
0.98% (w/v) amine fluoridea 0.17% (w/v) NaF 0.27% (w/v) SnCl2
1500
Tin concentration (mg/L)
Results
procedures were similar to our own previously performed studies, which helps to achieve comparability with our other results. The solutions were applied after the erosive impact, which reflects the clinical situation. Mostly, patients already have dental erosion, meaning that the dental hard tissue has already been affected by acids and the solutions are intended also as a therapeutic measure. The application frequency of the experimental solutions could possibly be used in clinical practice and in at home applications. The concentrations chosen for the active agents were pursuant to the directives for dental hygiene products and cosmetics. Because the current recommendation for symptomatic therapy of dental
The results are displayed in Fig. 1, and the values are given as mean S.D. The tissue loss was highest in the control group (72.6 11.5 mm). All test solutions reduced tissue loss significantly compared with the control group ( p 0.001). Within the 1500 mg/L F groups, tissue loss was highest in the 700 mg/ L Sn2+ group (group 4: 30.3 10.4 mm), and it decreased with an increasing stannous concentration (group 3 – 1400 mg/L Sn2+: 23.3 6.7 mm; group 2 – 2100 mg/L Sn2+: 19.6 4.6 mm; group 1 – 2800 mg/L Sn2+: 7.8 2.5 mm). The same trend was observed for the 1000 mg/L F solutions. The tissue loss was highest in the 1400 mg/L Sn2+ group (group 6: 18.6 2.8 mm), whereas in the 2100 mg/L Sn2+ group, lower values were found (group 5 – 2100 mg/L Sn2+: 7.6 5.2 mm).
4.
Discussion
This study investigated the impact of the tin and the fluoride concentrations of different solutions on erosion progression. Relatively severe erosive conditions were chosen to simulate the clinical conditions that occur in patients at high risk of erosion, such as vegetarians21 and patients with eating disorders, who can have frequent acid attacks. The erosive
Fig. 1 – Tissue loss (mm) in all groups after 10 days cyclic deand remineralisation (6 T 5 min erosive demineralisation with 0.05 M citric acid and 2 T 2 min treatment with test solutions per day). No statistical significance of differences between groups is indicated by corresponding indices (ANOVA with Tukey’s post hoc).
archives of oral biology 54 (2009) 432–436
erosion is frequent application of high amounts of fluoride,22– 24 so experimental solutions with high concentrations of fluoride were used. The pH was 4.5, which make the solutions potentially applicable for daily mouth rinses. However, at this pH, solutions containing stannous ions in relatively high concentrations have limited stability. Amine fluoride was used in all tested solutions, since some studies have found that it has stabilising properties.25,26 Stannous chloride, which has also shown marked efficacy in vitro in reducing erosion in enamel,14 was used as an independent source of tin. As expected, the highest tissue loss was found in the control group. All tin–fluoride preparations significantly reduced erosive tissue loss by approximately 60–90% when compared with the control group. Both results were comparable with those from a previous study.14 Concerning the fluoride concentration of the solutions, there was no indication that the solutions with higher fluoride concentrations were more effective than those with lower concentrations, as would be expected from the literature.27 Rather, in solutions with the same concentration of tin, the solution with a lower fluoride concentration was more effective than the one with a higher fluoride concentration, as evidenced by the comparison of the efficacy of groups 2 (1500 mg/L F , 2100 mg/L Sn2+) and 5 (1000 mg/L F , 2100 mg/L Sn2+). Hence, in the presence of tin ions, the fluoride concentration seems to play a secondary role, at least in the range of the investigated concentrations. The tin concentration appears more important. The efficacy of the solutions increased nearly linearly with an increasing tin concentration among the 1500 mg/L F solutions. Similar results had been found by Wachtel.6 This study investigated the effect of a single application of various stannous ion-containing solutions on enamel dissolution in a caries model. The results have clearly shown that with an increasing stannous ion concentration, the protection against enamel dissolution increases.6 However, the efficacy of the tested solutions did not depend solely on the absolute concentrations of the stannous or fluoride ions. Comparing the groups 1 (1500 mg/L F , 2800 mg/L Sn2+) and 5 (1000 mg/L F , 2100 mg/L Sn2+), it is notable that they gave nearly identical results, even though the latter solution contained one third less fluoride and one fourth less tin than the first. A similar pattern was observed when comparing groups 2 (1500 mg/L F , 2100 mg/L Sn2+) and 6 (1000 mg/L F , 1400 mg/L Sn2+). This indicated that not only were the absolute concentrations important, but the ratio between the stannous and the fluoride ions was also relevant. The calculated tin to fluoride ratios were similar or even identical in the compared groups: 1.87 and 2.1 for groups 1 and 5, respectively, and 1.4 for both groups 2 and 6. The results indicated that the higher the ratio, the better the solution. Furthermore, the results indicated that solutions with ratios above 1.8 were very effective and that those with ratios of 1.4 or below had lower efficacy. These results are in agreement with those of Barbakow et al.9 This study showed that SnF2/ AmF solutions with a tin to fluoride ratio of 1.56 or higher were of marked efficacy in reducing mineral loss in enamel after an HCl treatment (2N), but solutions with a lower ratio (0.78) were notably less effective.9 Currently, it is unclear as to which ratio can be identified as the ‘‘cut-off-point’’ for
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maximal efficacy of fluoride solutions containing tin and to what extent the fluoride or tin concentrations can be reduced for a specific level of efficacy. These considerations, however, disregard the AmF/NaF ratio. Further studies considering these ratios of tin and the various fluoride compounds could provide more information. However, based on the present results, it is quite conceivable, that one can reduce the fluoride and tin concentrations to a certain amount in solutions to be used as a mouth rinse for the symptomatic therapy of dental erosion, in order to maximize stability and flavour.
Acknowledgement The study was supported by GABA International AG, Muenchenstein, Switzerland. Funding: Supported by GABA International AG, Muenchenstein, Switzerland. Conflict of interest: None. Ethical Approval: None.
references
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