5
dental materials 23 ( 2 0 0 7 ) 900–904 available at www.sciencedirect.com journal homepage: www.intl.elsevierhealth.com/journals/dema Thirty-five percent carbamide peroxide application causes in vitro demineralization of enamel Neslihan Efeoglu a,, David J. Wood b , Candan Efeoglu c a Department of Fixed & Removable Prosthodontics, Level 6 Worsley Building, Leeds Dental Institute, University of Leeds, Clarendon Way, Leeds LS2 9LU, UK b Department of Oral Biology, Leeds Dental Institute, University of Leeds, Clarendon Way, Leeds LS2 9LU, UK c Department of Oral & Maxillofacial Surgery, Leeds Dental Institute, University of Leeds, Clarendon Way, Leeds LS2 9LU, UK article info Article history: Received 25 August 2005 Received in revised form 12 April 2006 Accepted 22 June 2006 Keywords: Carbamide peroxide Bleaching Enamel Dentin Micro-computerized tomography Demineralization Power bleaching In-office bleaching In vitro abstract Objectives. The objective of this in vitro study was to investigate whether a high concentration ‘in-office’ bleaching agent affected the mineral content of enamel and dentin. Methods. A commercially available 35% carbamide peroxide bleaching agent was applied for 2 h to sectioned teeth (n = 11). Specimens were then immersed in artificial saliva at 37 C for a further 24h to simulate the oral environment. Tomographic images of these sections were obtained (micro-CT 80, Scanco, Switzerland) prior to and post-bleach application. Eight three-dimensional regions of interest (ROI), starting from the enamel surface extending to the dentinoenamel junction, were selected for each section. The hydroxyapatite equivalent mineral concentrations (g/cm 3 ) of the ROIs were calculated. Any changes in mineral content as a consequence of the bleaching procedure were calculated in relation to each ROI. Results. There was a significant reduction in the mineral content of enamel specimens post- bleach application extending to a depth of 250 m (paired t-test, p < 0.05); this reduction in mineral content was greatest in the ROI’s closest to the tooth surface. There was, however, no significant difference in the mineral content of dentin as a consequence of bleaching. Significance. This in vitro study has shown that significant demineralization of enamel occurred following bleaching with 35% carbamide peroxide. The concept that ‘in-office’ bleaching is a non-destructive cosmetic procedure should be reconsidered. © 2006 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved. 1. Introduction In-office power bleaching has been available to dentists for nearly a century [1]. Its current popularity is due to its claimed ability to produce immediate results. Despite the widespread use of in-office bleaching [2], there is still controversy in the literature as to whether these agents could adversely affect dental hard tissues. Bleaching materials for in-office use contain high concen- trations of hydrogen peroxide or carbamide peroxide; typi- cally, 35% carbamide peroxide is used as the active bleaching Corresponding author. Tel.: +44 113 343 6316; fax: +44 113 343 6129. E-mail addresses: [email protected], [email protected] (N. Efeoglu). agent. This high concentration of carbamide peroxide can be used either as a pre-treatment or in combination with at-home bleaching [1]. Morphological alteration of enamel following 35% carbamide peroxide bleaching has been shown by several researchers. Cavalli et al. [3] showed that 35% carbamide per- oxide application increased the roughness of enamel surfaces. Oltu and Gurgan [4] reported a change of inorganic compo- sition of enamel after 35% carbamide peroxide application. Bitter [5] stated that teeth that were bleached in vivo with 35% carbamide peroxide lost their aprismatic enamel layer and that the damage was not repaired after 90 days. Ernst et 0109-5641/$ – see front matter © 2006 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.dental.2006.06.032

Thirty-five percent carbamide peroxide application causes in vitro demineralization of enamel

Embed Size (px)

Citation preview

d e n t a l m a t e r i a l s 2 3 ( 2 0 0 7 ) 900–904

avai lab le at www.sc iencedi rec t .com

journa l homepage: www. int l .e lsev ierhea l th .com/ journa ls /dema

Thirty-five percent carbamide peroxide applicationcauses in vitro demineralization of enamel

Neslihan Efeoglua,∗, David J. Woodb, Candan Efeogluc

a Department of Fixed & Removable Prosthodontics, Level 6 Worsley Building, Leeds Dental Institute,University of Leeds, Clarendon Way, Leeds LS2 9LU, UKb Department of Oral Biology, Leeds Dental Institute, University of Leeds, Clarendon Way, Leeds LS2 9LU, UKc Department of Oral & Maxillofacial Surgery, Leeds Dental Institute, University of Leeds, Clarendon Way, Leeds LS2 9LU, UK

a r t i c l e i n f o

Article history:

Received 25 August 2005

Received in revised form

12 April 2006

Accepted 22 June 2006

Keywords:

Carbamide peroxide

Bleaching

Enamel

Dentin

a b s t r a c t

Objectives. The objective of this in vitro study was to investigate whether a high concentration

‘in-office’ bleaching agent affected the mineral content of enamel and dentin.

Methods. A commercially available 35% carbamide peroxide bleaching agent was applied for

2 h to sectioned teeth (n = 11). Specimens were then immersed in artificial saliva at 37 ◦C

for a further 24 h to simulate the oral environment. Tomographic images of these sections

were obtained (micro-CT 80, Scanco, Switzerland) prior to and post-bleach application. Eight

three-dimensional regions of interest (ROI), starting from the enamel surface extending to

the dentinoenamel junction, were selected for each section. The hydroxyapatite equivalent

mineral concentrations (g/cm3) of the ROIs were calculated. Any changes in mineral content

as a consequence of the bleaching procedure were calculated in relation to each ROI.

Results. There was a significant reduction in the mineral content of enamel specimens post-

bleach application extending to a depth of 250 �m (paired t-test, p < 0.05); this reduction in

Micro-computerized tomography

Demineralization

Power bleaching

In-office bleaching

mineral content was greatest in the ROI’s closest to the tooth surface. There was, however,

no significant difference in the mineral content of dentin as a consequence of bleaching.

Significance. This in vitro study has shown that significant demineralization of enamel

occurred following bleaching with 35% carbamide peroxide. The concept that ‘in-office’

stru

emy

sition of enamel after 35% carbamide peroxide application.

In vitro bleaching is a non-de

© 2006 Acad

1. Introduction

In-office power bleaching has been available to dentists fornearly a century [1]. Its current popularity is due to its claimedability to produce immediate results. Despite the widespreaduse of in-office bleaching [2], there is still controversy in theliterature as to whether these agents could adversely affectdental hard tissues.

Bleaching materials for in-office use contain high concen-trations of hydrogen peroxide or carbamide peroxide; typi-cally, 35% carbamide peroxide is used as the active bleaching

∗ Corresponding author. Tel.: +44 113 343 6316; fax: +44 113 343 6129.E-mail addresses: [email protected], [email protected] (N. Efeo

0109-5641/$ – see front matter © 2006 Academy of Dental Materials. Pudoi:10.1016/j.dental.2006.06.032

ctive cosmetic procedure should be reconsidered.

of Dental Materials. Published by Elsevier Ltd. All rights reserved.

agent. This high concentration of carbamide peroxide can beused either as a pre-treatment or in combination with at-homebleaching [1]. Morphological alteration of enamel following35% carbamide peroxide bleaching has been shown by severalresearchers. Cavalli et al. [3] showed that 35% carbamide per-oxide application increased the roughness of enamel surfaces.Oltu and Gurgan [4] reported a change of inorganic compo-

glu).

Bitter [5] stated that teeth that were bleached in vivo with35% carbamide peroxide lost their aprismatic enamel layerand that the damage was not repaired after 90 days. Ernst et

blished by Elsevier Ltd. All rights reserved.

2 3

aidetoc

thsaaia

dthtoqtmse

asie3e

2

2

Fwsepwscwosn

2

Tau

cb

d e n t a l m a t e r i a l s

l. [6] showed that a high concentration of carbamide perox-de was detrimental to enamel surface integrity; however, theamage was less than that was seen after phosphoric acidtching. However, Gultz et al. [7] have reported that followingreatment with 35% carbamide peroxide, no difference wasbserved in enamel surface morphology between treated andontrol specimens.

Previously reported work to investigate the demineraliza-ion effects of in-office bleaching agents on tooth structuresas also been controversial. Suleiman et al. [8] reported noignificant changes in hardness values for enamel and dentinfter bleaching with 35% hydrogen peroxide whereas Attin etl. [9] and Lewinstein et al. [10] reported a significant reductionn Knoop hardness of enamel after 35% carbamide peroxidepplication.

Micro-computerized tomography (micro-CT) is a new andeveloping technology that can be used to map the distribu-ion of mineral in teeth non-destructively [11]. The authorsave previously described a new in vitro micro-CT methodo show the demineralization effect of a 10% carbamide per-xide bleaching agent on enamel [12]. This method enableduantification of the mineral content of tooth specimens inhree dimensions. Comparisons were made to investigate the

ineral content prior to and post-bleach application. Resultshowed micro-CT was indeed a reliable tool to investigate theffects of bleaching agents.

Present knowledge of the effects of in-office bleachinggents is still limited and controversial [3–10]. Therefore in thistudy the aim was, using micro-CT, to investigate the effects ofn-office bleaching on enamel surface layers, subsurface lay-rs and the dentinoenamel junction. It was hypothesized that5% carbamide peroxide application would decrease the min-ral content of enamel.

. Materials and methods

.1. Preparation of the specimens

reshly extracted sound human upper second molar teethere used for the study. The teeth were stored in physiological

olution at room temperature until required. Eleven tooth rodsach 2 mm × 3 mm in cross section and 4 mm in length wererepared under water-cooling with a reciprocating diamondire saw (precision wire diamond saw, Well, Germany). All the

ections were taken from the buccal mid-1/3 of the anatomi-al crown. Any debris was removed from the teeth by brushingith a soft toothbrush (Oral-B no. 35, soft bristles; Oral B Lab-ratories, Belmont, USA) under running deionised water. Allurfaces except the natural enamel surface were coated withail varnish.

.2. Micro-CT measurements and evaluations

he mineral content of the tooth specimens both prior tond post-35% carbamide peroxide application was quantified

sing a micro-CT scanner (�CT 80, Scanco, Switzerland).

After the first scan, specimens were transferred to a sterileell culture well. 0.01 ml bleaching gel containing 35% car-amide peroxide (Opalescence Quick, Ultradent, USA) was

( 2 0 0 7 ) 900–904 901

applied on the natural enamel surfaces with a 1ml syringe.To accelerate the activity the gel was heated under runninghot water for 2 min. Immediately after opening the gel, thepH was measured with a calibrated pH meter (Orion 920A,Thermo Electron, USA) and found to be 6.7. A clinically realis-tic application time of 2 h was chosen as this corresponded tothe maximum application time recommended by the manu-facturer. During this time, specimens were kept in a humidenvironment at 37 ◦C. Specimens were then washed underrunning deionised water in order to remove the gel. Sub-sequently, 2.5 ml of artificial saliva (Batch number 17336,Saliveze, Wyvern, UK) was placed in the wells to simulatethe oral environment. The saliva contained calcium chloride,magnesium chloride, sodium chloride, potassium chloride,dibasic sodium diphosphate, sorbitol and carboxymethyl cel-lulose (as listed by the manufacturer). Specimens were incu-bated in a humid environment at 37 ◦C for a further 24 h. ThepH of the saliva substitute was 6.9.

Tooth specimens were scanned twice: before and afterbleach application. The same scanning parameters wereapplied in both scans. The X-ray source was set at 45 kvP,and 177 �A. Integration time was 400 s. The entire thicknessof the tooth rods were scanned at high resolution. The datacollected were used to reconstruct images with a resolution of2048 × 2048 pixels and with an isotropic voxel size of 25 �m.

A custom sample holder was used to position the speci-mens in the sample holder of the micro-CT scanner. Duringscanning, a damp sponge was placed in the sample holderand the holder was sealed with cling film to maintain a humidenvironment therefore preventing any cracks that might occurin a dry environment [12].

The ‘optimum threshold procedure’ was run and thethreshold gray values for enamel and dentin were calculatedas 296 and 580, respectively. The evaluation software avail-able in the workstation of the scanner was used to defineeight regions of interest (ROI) per tooth rod. The enamel sur-face and immediate subsurface were divided into two 25 �mthick regions of interest; ROI-1 started from the enamel sur-face extending to a depth of 25 �m followed by ROI-2, whichextended a further 25 �m. Other ROIs had a thickness of 50 �m(Fig. 1). ROI-8 extended from the dentinoenamel junction (DEJ)towards enamel and ROI-7 extended towards the dentin. Thethreshold value for enamel was utilized in defining the ‘innervalue’ during the automatic contouring of the ROI-1, ROI-2,ROI-3, ROI-4, ROI-5 and ROI-6. ROI-7 and ROI-8 were definedmanually as for practical reasons it was easier to define theDEJ. The ‘outer value’ corresponded to the nail varnish, air andthe damp sponge therefore excluding these from the evalua-tions. These were previously described in detail [12].

Evaluations were carried out on each ROI both prior to andpost-bleach application. One hundred and seventy-six regionswere evaluated using the image processing language availableon the workstation. Gray level median values for each ROI wereconverted to g/cm3 assuming that the component absorbingthe X-rays was calcium hydroxyapatite. An in house producedcylindrical hydroxyapatite block (Plasma Biotal Ltd., Buxton,

UK) with 2.9 g/cm3 density was scanned with the specimensand the data was used to calculate the hydroxyapatite equiv-alent density (g/cm3). In addition, the percentage of mineralloss in relation to each ROI was calculated.

902 d e n t a l m a t e r i a l s 2 3 ( 2 0 0 7 ) 900–904

Fig. 1 – 3D micro-CT images of a tooth specimenillustrating eight regions of interest (ROI). The mineral

Fig. 2 – Demineralization gradient seen in this graphrepresents the percentage of the mineral loss after bleach

concentration of each ROI before and after bleachapplication was compared (p = 0.05).

The data were then subjected to statistical analysis. Pairedt-tests were used to determine if any of the observed changeswere significant at a 95% confidence level.

3. Results

Mean values of the hydroxyapatite equivalent density of theROIs are given in Table 1. Demineralization gradients were cal-culated as the percentage of mineral loss in any given ROI(Fig. 2). The mineral concentration of each ROI before and after

Table 1 – Hydroxyapatite equivalent densities for eachROI before and after bleaching

Region ofinterest (ROI)

Mean (g/cm3) S.D. S.E.M.

ROI 1 1.83a 0.30814 0.092911.36b 0.33000 0.09950

ROI 2 2.61a 0.53669 0.161821.94b 0.47219 0.14237

ROI 3 2.21a 0.37712 0.113711.68b 0.35920 0.10830

ROI 4 3.31a 0.63424 0.191232.94b 0.61292 0.18480

ROI 5 3.91a 0.25498 0.076883.72b 0.33974 0.10244

ROI 6 4.09a 0.09557 0.028824.00b 0.20223 0.06098

ROI 7 1.89a 0.60063 0.181101.73b 0.34482 0.10397

ROI 8 2.58a 0.64973 0.195902.50b 0.39362 0.11868

S.D.: standard deviation; S.E.M.: standard error mean.a Values obtained before bleach application (n = 11).b Values obtained after bleach application (n = 11).

application in each ROI. Dots/lines show means, error barsshow 95% confidence interval of mean.

bleach application was compared (p = 0.05). There was a sig-nificant difference in the hydroxyapatite equivalent mineralconcentration in the ROI-1, ROI-2, ROI-3, ROI-4, ROI-5 and ROI-6 corresponding to the outer 250 �m of enamel (paired t-test,p < 0.05). No significant difference was found for ROI-7 and ROI-8 which corresponded to the dentinoenamel junction (pairedt-test, p > 0.05) (Table 2).

4. Discussion

Thirty-five percent carbamide peroxide gel is the most com-monly used in-office bleaching agent. However, there is stillsome controversy in the dental literature as to whether 35%carbamide peroxide bleaching causes demineralization ofteeth [8–10]. The main advantage of micro-CT for the currentin vitro study was that because it is a non-destructive tech-nique, paired comparisons of the same tooth sections beforeand after bleaching could be made. The main drawback of thetechnique is the large amount of time required to reconstructand analyze the data.

For the present study, the reconstructed microtomographic3D image had a resolution of 2048 × 2048 pixels with anisotropic voxel size of 25 �m. Other X-ray attenuation meth-ods like transverse microradiography (TMR) can only providequantitative measurements of demineralized lesions of a thinplanoparallel tooth section (typically 100 �m) so that a 1D(line) or 2D (area) array of mineral dissolution is obtained [13].Atomic force microscopy (AFM) has also been reported as asuitable tool for measuring early stages of enamel deminer-alization in high-resolution (10−10 m) by providing a detailedsurface topography. The micro-CT used in this study cannotprovide a surface topography however; it has the ability toevaluate the mineral content of both the surface and subsur-face enamel layers quantitatively in 3D [14].

The authors have previously used micro-CT to investi-gate demineralization of enamel caused by application ofa 10% carbamide peroxide agent [12]. The present studyused the same methodology, which enabled non-destructive

d e n t a l m a t e r i a l s 2 3 ( 2 0 0 7 ) 900–904 903

Table 2 – Paired t-test statistics (p = 0.05)

Paired differences

Mean S.D. S.E.M. Significance(two-tailed)

Pair 1 ROI1a–ROI1b 0.4730 0.30210 0.09109 0.000×

Pair 2 ROI2a–ROI2b 0.6701 0.45792 0.13807 0.001×

Pair 3 ROI3a–ROI3b 0.5309 0.35396 0.10672 0.001×

Pair 4 ROI4a–ROI4b 0.3711 0.33240 0.10022 0.004×

Pair 5 ROI5a–ROI5b 0.1868 0.16680 0.05029 0.004×

Pair 6 ROI6a–ROI6b 0.0919 0.12481 0.03763 0.035×

Pair 7 ROI7a–ROI7b 0.1643 0.54354 0.16388 0.340Pair 8 ROI8a–ROI8b 0.0779 0.70713 0.21321 0.723

S.D.: standard deviation; S.E.M.: standard error mean.

ptoc

sm(ghDiatsaciesaaoOaKs

2chctp

ecobpbb

a Values obtained before bleach application (n = 11).b Values obtained after bleach application (n = 11).×Significant differences (p < 0.05).

aired evaluations, gave quantifiable results and excludedhe possible influence of nail varnish, damp sponge and airn the measurements by using the “outer value” thresholdorresponding to these structures.

In this study, the hypothesis was confirmed as there was aignificant reduction in the mineral content of enamel speci-ens post-bleach application extending to a depth of 250 �m

paired t-test, p < 0.05); this reduction in mineral content wasreatest in the ROI’s closest to the tooth surface. There was,owever, no significant difference in the mineral content ofEJ as a consequence of bleaching. Previously reported work to

nvestigate the demineralization effects of in-office bleachinggents on tooth structures has primarily used microhardnessechniques. Suleiman et al. [8] reported that there were noignificant changes in hardness values for enamel and dentinfter bleaching with 35% hydrogen peroxide after 30 min appli-ation. Microhardness of enamel is linearly correlated withts mineral content; however, the method is destructive onnamel samples [15], therefore it is not possible to test theame volume for test and control. Differences between thisnd the authors’ current study may be due to the differentpplication times. In the present study, 35% carbamide per-xide gel was applied for 2 h. In a study by Attin et al. [9]palesence Quick was applied to bovine enamel in vitro fortotal of 2 h and the results showed a significant reduction innoop hardness of enamel up to 700 �m deep. These resultsupport the current findings.

Following bleaching, enamel samples were immersed for4 h in artificial saliva with a neutral pH that contained cal-ium and phosphate; therefore, it might be expected to havead a remineralization effect [4]. In addition, the pH of thearbamide peroxide gel was 6.7 therefore the demineraliza-ion effect seen in this study cannot be attributed to the lowH of the gel.

Demineralization was clearly greater in the outer 150 �m ofnamel (Fig. 2). In the authors’ previous study [12] the appli-ation of 10% carbamide peroxide caused demineralizationf the enamel surface up to 50 �m deep. The mechanism of

leaching is surprisingly ambiguous; however, 10% carbamideeroxide yields 3.3% hydrogen peroxide whereas 35% car-amide peroxide yields 10% hydrogen peroxide [1] and it haseen reported that a higher peroxide concentration caused

r

higher diffusion of the peroxide towards the pulp [16]. Thedeeper demineralization presented in this study supports thishypothesis. Therefore, it can be argued that the demineral-ization effect presented could be the result of the possiblealteration in the structure of the enamel caused by uncon-trolled reaction of the peroxide radical [17]. On the other hand,it has been shown that one of the breakdown products of car-bamide peroxide “urea” can have detrimental effects on theintraprismatic enamel which may also explain this deminer-alization effect [18].

Demineralization seen in this study may increase suscep-tibility to tooth wear caused by abrasive factors like toothbrushing [19]. Whether this demineralization can be reminer-alized in the longer term by saliva and/fluorides should befurther investigated. In addition, future in vivo studies withtreatment controls are needed to verify these results.

5. Conclusion

Within the limitations of this study in vitro application of 35%carbamide peroxide on enamel for 2 h, followed by storage inartificial saliva for 24 h, resulted in demineralization of theenamel extending to a depth of 250 �m below the enamel sur-face. It is recommended that application of high concentrationbleaching agents should be carefully considered in patientssusceptible to tooth wear.

Acknowledgements

This research study has been supported by the Departmentof Restorative Dentistry of University of Leeds. We would liketo thank Dr. Nigel Bubb for his intellectual and technical sup-port in preparation of the hydroxyapatite discs, Mr. Ian Smithfor his technical support in preparation of the sample hold-ers and Mr. Nigel Willmott for his useful comments regardingpresentation of the data.

e f e r e n c e s

[1] Greenwall L. Bleaching techniques in restorative dentistry.London: Martin Dunitz; 2001. p. 29.

l s 2

904 d e n t a l m a t e r i a

[2] Attin T, Hannig C, Wiegand A, Attin R. Effect of bleaching onrestorative materials and restorations a systematic review.Dental Mater 2004;20(9):852–61.

[3] Cavalli V, Arrais CAG, Giannini M, Ambrosano GMB. Highconcentrated carbamide peroxide bleaching agents effect onenamel surface. J Oral Rehabil 2004;31:155–9.

[4] Oltu U, Gurgan S. Effects of three concentrations ofcarbamide peroxide on the structure of enamel. Journal ofOral Rehabilitation 2000;27:332–40.

[5] Bitter NC. A scanning electron microscope study of thelong-term effect of bleaching agents on the enamel surfacein vivo. Gen Dent 1998:84–8.

[6] Ernst CP, Marroquin BB, Willershausen-Zonnchen B. Effectsof hydrogen peroxide-containing bleaching agents on themorphology of human enamel. Quintessence Int1996;27(1):53–6.

[7] Gultz J, Kaim J, Scherer W, Gupta H. Two in-office bleachingsystems: a scanning electron microscope study. CompendContin Educ Dent 1999;20(10):965–8, 970 [quiz 972].

[8] Suleiman M, Addy M, Macdonald E, Rees JS. A safety study invitro for the effects of an in-office bleaching system on theintegrity of enamel and dentine. J Dent 2004;32:581–90.

[9] Attin T, Vollmer D, Wiegand A, Attin R, Betke H. Subsurfacemicrohardness of enamel and dentine after different

external bleaching procedures. Am J Dent 2005;18:8–12.

[10] Lewinstein I, Fuhrer N, Churaru N, Cardash H. Effect ofdifferent peroxide bleaching regimens and subsequentfluoridation on the hardness of human enamel and dentin. JProsthet Dent 2004;92:337–42.

3 ( 2 0 0 7 ) 900–904

[11] Elliot JC, Davis GR, Anderson P, Wong FSL, Dowker SEP,Mercer C. Application of laboratory microtomography to thestudy of mineralised tissues. Anales de Quimica Int Ed1997;93:S77–82.

[12] Efeoglu N, Wood D, Efeoglu C. Microcomputerisedtomography evaluation of 10% carbamide peroxide appliedto enamel. J Dent 2005;33(7):561–7.

[13] Dowker SEP, Elliott JC, Davis GR, Wassif HS. Longitudinalstudy of the three-dimensional development of subsurfaceenamel lesions during in vitro demineralization. Caries Res2003;37:237–45.

[14] Finke M, Jandt KD, Parker D. The early stages of nativeenamel dissolution studied with atomic force microscopy. JColloid Interf Sci 2000;232:156–64.

[15] White DJ, Faller RV, Bowman WD. Demineralization andremineralization evaluation techniques-addedconsiderations. J Dent Res 1992;71(Spec Iss):929–33.

[16] Gokay O, Yilmaz F, Akin S, Tuncbilek M, Ertan R. Penetrationof the pulp chamber by bleaching agents in teeth restoredwith various restorative materials. J Endo 2000;26(2):92–4.

[17] Seghi RR, Denry I. Effects of external bleaching onindentation and abrasion characteristics of human enamelin vitro. J Dent Res 1992;71(6):1340–4.

[18] Arends J, Jongebloed WI, Goldberg M, Schuthof J. Interactionof urea and human enamel. Caries Res 1984;(18):17.

[19] Attin T, Kielbassa M, Schwanenberg M, Hellwig E. Effect offluoride treatment on remineralization of bleached enamel.J Oral Rehab 1997;24:282–6.