Effect of Disinfectant Solutions on Gutta-Percha andResilon ConesMAIRA PRADO,1 HELOISA GUSMAN,2 BRENDA P.F.A. GOMES,3* AND RENATA A. SIMAO4

1Department of Metallurgic and Materials Engineering, Federal University of Rio de Janeiro, Rio de Janeiro, RJ, Brazil2Department of Endodontics, Federal University of Rio de Janeiro, Rio de Janeiro, RJ, Brazil3Department of Restorative Dentistry, Endodontics Division, State University of Campinas- UNICAMP, Piracicaba, SP, Brazil4Department of Metallurgic and Materials Engineering, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil

KEY WORDS atomic force microscopy; chlorhexidine; gutta-percha; sodium hypochlorite;MTAD; Resilon

ABSTRACT The aim of this study was to evaluate the effects of the 5.25% sodium hypochlorite(NaOCl), 2% chlorhexidine (CHX), and MTAD solutions on the surface of gutta-percha and Resiloncones by using atomic force microscopy (AFM). Accessory cones were washed and dried. The coneswere randomly divided into six groups: gutta-percha immersed in NaOCl, CHX, and MTAD, andResilon immersed in NaOCl, CHX, and MTAD. AFM images of the same area were made in differ-ent periods of time. JPK2 Image Processing Software was used to evaluate the images. The param-eters used to evaluate the changes were RMS and line profiles. No statistically significant changewas observed in the RMS values. The line profiles detected changes only for gutta-percha surfacesafter immersion in NaOCl and MTAD solutions. In conclusion, 5.25% NaOCl and MTAD are associ-ated with local changes in surface roughness of gutta-percha cones. No change was observed when2% CHX was used. The use of all tested solutions did not produce any changes on Resilon surface.Microsc. Res. Tech. 00:000–000, 2012. VVC 2012 Wiley Periodicals, Inc.

INTRODUCTION

A successful endodontic therapy is related to elimi-nating microorganisms that are found in the root canalsystem and blocking their re-entry (Isci et al., 2006).The prevention of contamination in the root canal sys-tem is an important therapeutic aspect, for it is essen-tial to maintain the aseptic chain (Short et al., 2003).

Although most instruments used in endodontic treat-ment can be sterilized by autoclaving or dry heat beforetreatment, gutta-percha and Resilon cones, because oftheir thermoplastic characteristics, cannot be sterilizedby these conventional processes (Ozalp and Ozcelika,2006). Therefore, a rapid chair-side chemical disinfec-tion is needed (Gomes et al. 2005; Senia et al., 1975).Some chemical agents commonly used are sodium hy-pochlorite, chlorhexidine, and MTAD—a mixture ofdoxycycline, 10% citric acid, and a detergent, Tween 80(Cardoso et al., 1999; Da Motta et al., 2001).

A 5.25% sodium hypochlorite (NaOCl) solution is aneffective decontaminant; however, it has been demon-strated that this solution is a strong oxidizing agentthat can cause deterioration of various materials(Yamauti et al., 2003), such as gutta-percha andResilon cones (Isci et al., 2006; Valois et al., 2005). Ithas been reported in the literature that 2% chlorhexi-dine is equally effective and a valuable alternative dis-infectant (Pang et al., 2007). MTAD has been alsointroduced as a solution for rapid disinfection of cones,but there are no data evaluating its effect on the sur-face of gutta-percha and Resilon cones (Royal et al.,2007).

Atomic force microscopy (AFM) is a well-establishedand documented tool for structural characterization ofmaterials (Arvidsson et al., 2002; Jagger et al., 2000).

A number of different AFM operation modes, such ascontact mode imaging, intermittent contact, and forcemodulation microscopy, can be used to optimize thestudy of surface topography and physical properties.AFM offers the opportunity to image the three-dimen-sional surface topography of biological specimens withhigh spatial resolution under a wide variety of condi-tions. These include exposure to air, water, and otherstorage solutions at elevated or reduced temperatures(Eliades et al., 1999; Marshall et al., 1993). Moreover,sensitive nonconducting samples, which are difficult toexamine using traditional SEM due to the high vac-uum requirement, can be studied through AFM (Mar-shall et al., 1993).

The aim of this study was to evaluate the effects of5.25% sodium hypochlorite, 2% chlorhexidine, andMTAD on the surface of gutta-percha and Resiloncones by using AFM.

MATERIALS ANDMETHODS

Nine gutta-percha (Odous De Deus1, Belo Hori-zonte, MG, Brazil) and Nine Resilon (Pentron ClinicalTechnologies, Wallingford, CT, USA) cones of mediumsize were randomly selected from the same batch,washed with ultrapure water, and dried with nitrogen

*Correspondence to: Brenda P.F.A. Gomes, Department of Restorative Den-tistry, Piracicaba Dental School State University of Campinas, Avenida Limeira,901, Piracicaba, SP, Brazil. E-mail: bpgomes@fop.unicamp.br

Received 5 September 2011; accepted in revised form 17 October 2011

Contract grant sponsor: Brazilian agencies FAPERJ and CNPq

DOI 10.1002/jemt.21126

Published online inWiley Online Library (wileyonlinelibrary.com).

VVC 2012 WILEY PERIODICALS, INC.

MICROSCOPY RESEARCH AND TECHNIQUE 00:000–000 (2012)

gas. The cones were attached to a glass base withrapid-setting cyanoacrylate glue for analysis.

The samples were randomly divided into six groupsaccording to disinfecting solutions and material used,as described below:

1. Gutta-percha immersed in 5.25% NaOCl2. Gutta-percha immersed in 2% chlorhexidine3. Gutta-percha immersed in MTAD4. Resilon immersed in 5.25% NaOCl5. Resilon immersed in 2% chlorhexidine6. Resilon immersed in MTAD

Three analyses were made in a same cone. AFMimages of the samples were recorded by using a JPKNano Wizard I atomic force microscope (JPK Instru-ments - Nanotechnology for Life Science, Berlin, Ger-many) operating at contact-mode under room condi-tions and relative humidity between 45 and 55%. A rec-tangular cantilever CSC17/AlBS (MikroMasch2, SanJose, CA, USA) was used for obtaining all images.

First, images of 30 lm 3 30 lm were taken and a10 lm3 10 lm area was selected and imaged. After that,the sample was covered for 1 min with 2 mL of the solu-tion according to the groups described above, washedwith ultrapure water, and dried with nitrogen gas. A newimage of 30 lm 3 30 lm was taken to establish the samearea of 10 lm 3 10 lm, as determined previously. Next,an image of this area was taken. The protocol describedbefore was used in the 3-min and 6-min periods.

In this way, the AFM analyses were performedexactly in the same area of 10 lm 3 10 lm in differentperiods (0-control, 1, 3, and 6 min). It was possiblebecause the equipment described above (JPK) allowsremoving the head of the microscope without movingthe sample from its position and replace it for subse-quent analysis of the same site. This procedure is possi-ble due to closed-loop scanners that return the AFM tipexactly to the same point of the first analysis, thuscompensating the thermal drift. During treatment, thesample is kept in the same position and the systemallows us to remove the head of the microscope andreturn it to the same position, enabling analysis of thesame site.

JPK2 Image Processing Software (JPK InstrumentsAG, Berlin, Germany) was used to evaluate the images.The parameter used to measure the roughness of thesamples was RMS (roughness mean square). RMS isthe root-mean-square calculation of all values of theroughness profile. These data were analyzed statisti-cally using Kruskal-Wallis and Mann-Whitney tests(P < 0.05).

Finally, 10 selected line profiles of the each image,ranging between from 1.5 to 3.0 lm, were generatedand also compared at the same exact location after dif-ferent treatment times. Using line profiles, it is possi-ble evaluate exactly the same feature in differentperiod of time. After the line profiles were created inthe images, Origin software was used to evaluate thepossible overlap of the lines and evaluate real localchanges on the cone surface

RESULTS

Figure 1 presents images of the gutta-percha andResilon cones treated with 5.25% sodium hypochlorite,

2% chlorhexidine, and MTAD in different periods. Byvisual inspection, it is not possible to observe changesin the surface of the cones.

Table 1 shows the roughness parameter RMSobtained for the same cones observed in Figure 1. Inthis case, no significant change was observed in theRMS values. It occurred because the difference wassmaller than the standard deviation.

Figure 2 indicates one of the line profiles obtainedfor exactly the same site of the gutta-percha conetreated with sodium hypochlorite, chlorhexidine, andMTAD, respectively, in different periods. Significantchanges were observed in the line profile of the gutta-percha cone after contact with sodium hypochloriteand MTAD. No changes were observed when chlorhexi-dine was used. Although no significant variations wereobserved in the roughness parameters, local variationswere found when line profiles were used. These datawere found in the three cones evaluated. The sameresult was found in the other two cones of the group.

Figures 3 shows one of the line profiles obtained forexactly the same site of the Resilon cones treated withsodium hypochlorite, chlorhexidine, and MTAD,respectively. No changes were observed in the cones af-ter immersion in all solutions. The three parametersused to evaluate the Resilon surface, i.e., visual inspec-tion, roughness parameter, and line profiles did notdetect changes before immersion in the three solutions.The same result was found in the other two cones ofthe group.

DISCUSSION

A decontamination program begins with a cleaningmethod that assures the reduction of natural bio-bur-den, which is the initial population of viable microor-ganisms present on the material. This is also the guar-antee of a proper and successful application of a disin-fectant, a sanitizer, or a sterilant for use in health care.Therefore, the correct term for the use of chemicalcleaning agents is disinfection (Gomes et al., 2005).Gutta-percha and Resilon cones cannot be sterilized byheat because of their thermoplastic characteristics. Asa result, a rapid chair-side disinfection using chemicalagents such as sodium hypochlorite, chlorhexidine,and MTAD should be adopted in the endodontic prac-tice (Royal et al., 2007).

Previous studies have related that NaOCl and chlo-rhexidine cause changes on the surface of the cones(Isci et al., 2006; Valois et al., 2005). The importance ofcarefully following up such changes is related to thetopographic effects induced by cleaning procedures andto the chemical attack on the surface that can be re-sponsible for changes in surface properties (Turker andBiskin, 2003). According to Wenzel law, the increase inthe roughness can favor the spread of a liquid in thesurface. This way, the modifications on the cones surfa-ces can favor or disfavor the sealer spread (Hu et al.,2010).

In this study, three cones were used. However, thedata were evaluated separately, one by one. This proce-dure was made because different cones presented avery large roughness variation. This is clearly demon-strated in the Table 1. Different cones, i.e. differenttreatment, presented variations on the initial values ofroughness, for example in case of gutta-percha one

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sample has RMS values of (108.70 6 5.6) nm for onecone and for the other cone of the same brand andbatch the RMS values measured were (47.25 6 1.1)nm. Although the data had been evaluated separately,the results found were the same for all the analyzedsamples. When gutta-percha cones were evaluated, theuse of NaOCl and MTAD led to local changes in thesurface roughness. However, the increases and

decreases in the roughness of some areas resulted inno changes in the global roughness. Chlorhexidine didnot cause any changes on the cones surfaces. Thisresult was found for all the three cones evaluated.

Valois et al. (2005) evaluated the action of sodium hy-pochlorite and chlorhexidine on the surface of gutta-percha and observed that these solutions resulted inthe roughness modifications of the cones. However,

Fig. 1. Images of the gutta-percha and Resilon cones treated with 5.25% sodium hypochlorite, 2%chlorhexidine, and MTAD in different periods.

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these modifications can be due to the area selection,not being related to the surface attack itself, as theseauthors evaluated different small areas (2.5 lm 32.5 lm) and always different areas that can present dif-ferent roughness. In this study, evaluating the samearea, no global changes (RMS values) were observed,but local changes were presented in the cases of NaOCland MTAD. It can be associated with several localchanges in different areas of the sample, increasing ordecreasing the roughness, resulting in absence ofglobal changes.

The three solutions evaluated in this study did notcause changes on the Resilon surfaces in all cones.These data are in disagreement with Isci et al. (2006),

who observed changes in the roughness after contactwith NaOCl and chlorhexidine. The use of Resiloncones is associated with a resin sealer, creating a mono-block. According to the literature, 2% chlorhexidine so-lution interacts positively with the adhesion processbetween dentin and sealers (Lindblad et al., 2010; Ricciet al., 2010), and cone and sealer (Prado et al., 2011).

Gomes et al. (2005) and Prado et al. (2011) have eval-uated surface alterations in cones by using SEM, andthey observed that sodium hypochlorite and chlorhexi-dine did not cause changes on the cone surface. Thesefindings are, partially, consistent with this study as nochanges in the Resilon surface were detected. The dif-ferent results are due to the methodological approach

TABLE 1. Mean and standard deviation values of RMS (roughness mean square)

Gutta-percha RMS values (mean 6 SD) Resilon RMS values (mean 6 SD)

NaOCl Chlorhexidine MTAD NaOCl Chlorhexidine MTAD

Control 108.70 6 5.6 47.256 1.1 79.71 6 0.9 62.6 6 0.3 53.10 6 1.0 43.1 6 0.51 min 109.60 6 5.4 47.126 1.1 79.72 6 0.9 62.87 6 0.6 53.13 6 1.0 43.03 6 0.33 min 109.33 6 5.8 47.286 0.8 79.70 6 0.9 62.63 6 0.6 53.13 6 0.6 43,00 6 0.66 min 109.33 6 6.2 47.496 0.7 79.72 6 0.8 62.97 6 0.4 53.00 6 1.0 43.07 6 0.5

Fig. 2. Line profiles obtained for exactly the same site of the gutta-percha cones treated with so-dium hypochlorite, chlorhexidine, and MTAD in different periods.

Fig. 3. Images of the Resilon cones immersed in 5.25% sodium hypochlorite, 2% chlorhexidine, andMTAD for 0, 1, 3, and 6 min.

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because SEM evaluated different bigger areas,whereas AFM the same smaller ones.

In conclusion, 5.25% sodium hypochlorite and MTADare associated with local changes in surface roughnessof gutta-percha cones. No change was observed when2% chlorhexidine was used. The use of all tested solu-tions did not produce any changes on Resilon surface.Therefore, the use of chlorhexidine solution to disinfectgutta-percha cones is preferred. For Resilon disinfec-tion, all solutions can be used.

ACKNOWLEDGMENTS

The authors thank Heleno de Souza da Silva andJackson Belmiro for technical support.

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