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Journal of Applied Oral Science

Print version ISSN 1678-7757On-line version ISSN 1678-7765

J. Appl. Oral Sci. vol.16 no.6 Bauru Nov./Dec. 2008 



Efficacy of sodium hypochlorite and chlorhexidine against Enterococcus faecalis – a systematic review



Carlos EstrelaI; Julio Almeida SilvaII; Ana Helena Gonçalves de AlencarIII; Claudio Rodrigues LelesIV; Daniel Almeida DecurcioII

IDDS, MSc, PhD, Chairman and Professor of Endodontics, Federal University of Goiás, Goiânia, GO, Brazil
IIDDS, MSc, Professor of Endodontics, Federal University of Goiás, Goiânia, GO, Brazil
IIIDDS, MSc, PhD, Professor of Endodontics, Federal University of Goiás, Goiânia, GO, Brazil
IVDDS, MSc, PhD, Professor of Prevention and Oral Rehabilitation, Federal University of Goiás, Goiânia, GO, Brazil

Correspondence address




The efficacy of the sodium hypochlorite (NaOCl) and chlorhexidine (CHX) on Enterococcus faecalis was evaluated by systematic review and meta-analysis. The search strategies included search in electronic biomedical journal databases (MEDLINE, EMBASE, CENTRAL) and handsearching records, using different matches of keywords for NaOCl, CHX and Enterococcus faecalis. From 41 in vivo studies, 5 studies met the inclusion criteria. In a sample containing 159 teeth, E. faecalis was detected initially in 16 (10%) teeth by polymerase chain reaction (PCR) and 42 (26.4%) teeth by microbial culture techniques. After root canal disinfection, this species was observed in 11 (6.9%) teeth by PCR and 12 (7.5%) teeth by culture. Risk differences of included studies were combined as generic inverse variance data type (Review Manager Version 5.0 _ Cochrane Collaboration,, accessed 15 May 2008), taking into account the separate tracking of positive and negative cultures/PCR. The level of statistical significance was set at p<0.05. In conclusion, NaOCl or CHX showed low ability to eliminate E. faecalis when evaluated by either PCR or culture techniques.

Key words: E. faecalis. Sodium hypochlorite. Chlorhexidine. Irrigating solutions. Systematic review.




The pathogenicity of endodontic microorganisms responsible for stimulating apical periodontitis creates the need for finding effective antimicrobial medicaments16. Sodium hypochlorite (NaOCl) and chlorhexidine (CHX) are the most frequently widely studied and employed antimicrobial agents for treatment of root canal infection2,5. These medicaments present chemical characteristics that are particularly responsible for their distinct results when compared1,6,20,23. These variations occur probably due to differences in methodology, biological indicators, concentrations, exposure time, the potential for different anatomical and treatment differences between patients1,6,20,23,27.

The contemporary literature contains numerous reports on the antimicrobial efficacy of NaOCl and CHX in several experimental models - infected human teeth in vivo7,16-18,29,30, infected human teeth ex vivo4, infected dog's teeth in vivo5,21, infected bovine teeth ex vivo10, biofilm model in membrane filters22,25, direct contact and agar diffusion test6,9.

E. faecalis is an extensively evaluated biological indicator12,15,19,26,27. Some factors can explain the concern with this pathogen in endodontic infections. Its high prevalence in cases with post-treatment disease associated with virulence factors (aggregation substance, enterococcal surface proteins (Esp), gelatinase, cytolysin toxin, extracellular superoxide production, capsular polysaccharides, antibiotic resistance determinant) can facilitate the adherence of host cells and extracellular matrix, tissue invasions, immunomodulation effect and cause toxin-mediated damage19.

Actual thinking has recommended the implementation of evidence-based dentistry, which valorizes studies involving the systematic review or meta-analysis. Systematic reviews use a strict methodological approach to search, select, evaluate, and analyze original data from primary sources. Good scientific evidence is mandatory to elaborate clinical decisions, yet few systematic reviews or meta-analysis have been developed in Endodontics14,21,28.

Torabinejad and Babjri28 reported that the American Dental Association has adopted the concept of evidence-based healthcare, which requires the judicious integration of systematic assessments of clinically relevant scientific evidence.

Previous studies using in vitro experimental models have confirmed the antimicrobial efficacy of NaOCl and CHX against E. faecali6,23, while others using different study designs have not found same results4,5,7,16-18,29,30. Relevant clinical questions based on evidence regarding the resistance of E. faecalis to NaOCl or CHX should be further discussed. Therefore, the aim of this study was to review findings on the antibacterial efficacy of NaOCl or CHX against E. faecalis in endodontic infection using systematic review and meta-analysis.



Literature Review

This study was designed using an analysis of longitudinal studies from a quantitative systematic review. Prospective studies were selected towards the efficacy of NaOCl and CHX against E. faecalis identified in endodontic infections before and after root canal preparation. English-language articles were retrieved from electronic biomedical journal databases and handsearching records.

The following databases were searched on January 2nd, 2007: MEDLINE (without filter, from 1966 to January 2nd, 2007), EMBASE (without filter, from 1980 to January 2nd, 2007), Cochrane Oral Health Group Trials Register and Cochrane Central Register of Controlled Trials (CENTRAL). For the electronic search strategy, the following terms were used as keywords in several combinations: 1. faecalis and sodium hypochlorite OR, 2. faecalis and chlorhexidine OR, 3. faecalis and root canal infections OR, 4. faecalis and endodontics infections OR, 5. faecalis and root canal irrigants OR, 6. faecalis and irrigating solution OR, 7. faecalis and endodontics irrigants OR, 8. faecalis and intracanal irrigants.

A systematic review was conducted according to the guidelines of the Cochrane Collaboration, which recommend the search for the largest possible number of articles. Handsearching was conducted by the review of the reference lists of the eligible clinical trials and the review of author's personal databases of trial reports in an attempt to identify any other relevant studies.

Inclusion and Exclusion Criteria

The selected articles were identified from titles and abstracts by two independent reviewers, considering the tabulated inclusion and exclusion criteria.. The inclusion criteria were studies in humans, studies related to the efficacy of NaOCl or CHX against E. faecalis, nonsurgical root canal treatment performed during the study, subjects with a noncontributory medical history, microbiological samples collected before and after root canal preparation, English-language articles. The exclusion criteria were in vitro and animal studies, studies related to the efficacy of intracanal irrigants and medications other than NaOCl or CHX, studies without collection of microbiological samples before or immediately after root canal preparation, non-English-language articles, studies abstract only or no abstract, literature reviews, studies involving primary teeth, case reports, studies related only to microbial identification or studies involving microorganisms other than E.faecalis.

Full-text reprints were obtained from all relevant and potentially relevant studies, which seemed to meet the inclusion criteria and from those that had insufficient data in the title and abstract to make a clear decision.


The design of this meta-analysis was based on the guidelines of the Cochrane Collaboration and on the methodology of previous study21. Chi-square test was used to analyze the differences between the studies. The essential analysis of antibacterial efficacy was risk difference (difference in the ratio of positive bacterial identification obtained by PCR of culture techniques between pre- and post-disinfection). Risk differences of included studies were combined as generic inverse variance data type (Review Manager Version 5.0 _ Cochrane Collaboration,, accessed 15 May 2008), taking into account the separate tracking of positive and negative cultures/PCR. The level of statistical significance was set at p<0.05.



The search retrieved 229 related articles, being 6 literature reviews, 39 articles related to in vivo studies (27 in humans and 12 in animals), and 189 related to in vitro studies. From the 39 in vivo studies, 5 met the inclusion criteria. In these 5 papers, from a total of 159 teeth with primary or secondary endodontic infections, E. faecalis was initially detected in 16 (10%) teeth by PCR and 42 (26.4%) teeth by culture. After disinfection (effect of root canal enlargement associated with the action of chemical irrigants), E. faecalis was identified in 11 (6.9%) teeth by PCR and 12 (7.5%) teeth by culture techniques. No longitudinal studies in humans investigating the efficacy of NaOCl and CHX against E. faecalis from endodontic infections were found (Table 1).



The outcomes of the 5 selected studies are shown in Tables 2 and 3. The analysis was made between pre- and post-disinfection of the same root canals. The 5 5 studies were heterogeneous (Test of Homogeneity Chochran), considering samples evaluated by culture technique (÷2 =45.85, df=4, p<0.00001) and PCR (÷2 =1.65, df=1, p=0.20). Thus, NaOCl or CHX showed low ability to eliminate E. faecalis when evaluated by either culture or PCR techniques.



The success of endodontic treatment is closely associated to the control of endodontic microbiota. Several chemical irrigants have been suggested for use in the treatment of infected root canal1,2,4-7,9-11,16-18,20,22,23,25,28-30 .

Studies with similar results have been reported when NaOCl and CHX were compared9,30. However, other investigations have shown that NaOCl presents better antimicrobial activity than CHX1,20,23, or, conversely, that CHX has better antimicrobial activity than NaOCl23. Recently, it was demonstrated in a model of E. faecalis biofilm in human root canals that ozonated water, 2.5% NaOCl, 2% CHX and the application of gaseous ozone for 20 min were not sufficient to inactivate E. faecalis4. These differences may have been caused by differences in the experimental methodology, concentration, type of irrigating solution, patient and anatomical differences in root canal anatomy or the period of time used in the analysis.

It is important to emphasize the methodology used in the present investigation. Studies based on scientific evidence have stood out in dentistry14,21. Various advantages of systematic review can be observed: explicit methods limit bias in identifying and rejecting studies; conclusions are more reliable and accurate; more information can be rapidly assimilated by health care providers, researchers and policymakers; delay among research discoveries, implementation of effective diagnostic and therapeutic strategies is potentially reduced; results of different studies can be formally compared to establish generalization of findings and consistency of results (lack of heterogeneity); reasons for heterogeneity (inconsistency in results across studies) can be identified and new hypotheses generated about particular subgroups; quantitative systematic reviews (meta-analyses) increase the accuracy of the overall result.

The investigation model adopted in the present essay involved 5 studies, characterized by the heterogeneity of the clinical protocols. The analysis was made between pre- and post-disinfection of the same root canals considering samples evaluated by culture or PCR techniques. Thus, the NaOCl or CHX showed low ability to eliminate E. faecalis when evaluate by both techniques. An identical problem occurs when calcium hydroxide [Ca(OH)2] is tested against E. faecalis14,21.Depending on the methodology - direct contact, agar diffusion or contaminated dentin test - this intracanal medicament may either present efficacy or not. This fact was not confirmed in human studies7,17,18,29,30 (Table 1), but the limitations of the methodology employed in this study should be considered. The application and validation of the results of longitudinal studies, as far as evidence-based view is concerned, are essential to determine the scientific value of the selected studies. Knowledge of the strategies to be applied for study selection is also important. Therefore, planning and development of this study model should be done with great caution8.

The first aspect to be considered is related to the bacteria location. If bacteria are located only on the root canal surface, where the intracanal medicaments can reach, NaOCl and CHX may present efficacy against E. faecalis. However, when bacteria are lodged within the dentinal tubules or in deep layers, E. faecalis can be more resistant to the antibacterial action of NaOCl and CHX.

Five studies met the inclusion criteria established for the present investigation7,17,18,29,30. Peciuliene, et al.18 observed E. faecalis in 25 asymptomatic teeth with secondary infection. Avoiding contamination, microbiological samples were collected from the canals before and after preparation and irrigation with NaOCl and EDTA. E. faecalis was isolated from 14 of those 20 culture positive teeth, usually in pure culture or as a major component of the flora. Second samples taken after preparation revealed growth in 7 of the 20 teeth. Five of the 7 cases were E. faecalis in pure culture. Isolation of E. faecalis was not related to the use of any specific root filling material in the original root filling. Peciuliene, et al.29 determined the occurrence and role of yeasts, enteric Gram-negative rods and Enterococcus species in root-filled teeth with chronic apical periodontitis. After collection of the first microbiological sample, the root canals were prepared to a size 40 file using 2.5% NaOCl and 17% EDTA as irrigating solutions. Microbes were isolated from 33 of 40 teeth in the initial sampling. Yeasts were isolated from 6 teeth, 3 of them together with E. faecalis. E. faecalis was isolated from 21 of the 33 culture positive teeth, 11 in pure culture. Growth was detected in 10 teeth of the second samples. Six of the 10 cases were E. faecalis, with five being a pure culture. Ferrari et al.7 detected enterococci, enteric bacteria and yeast species from 25 root canals with primary endodontic infections before and after canal preparation and to test the antibiotic susceptibility of enterococcal strains isolated. The canals were instrumented using a simple stepback technique with Endo PTC cream associated 0.5% NaOCl and EDTA. Microorganisms were isolated from 92% of the samples following intracoronal access, 22% were enterococci, enteric bacteria or yeast species. After biomechanical preparation, these species were no longer detected. After 7 days without intracanal dressing, 100% of the canals contained microorganisms, 52% of which were target species. E. faecalis and E. faecium were resistant to removal by root canal preparation followed by intracanal dressing. Zerella, et al.15 compared the effect of a slurry of Ca(OH)2 mixed in aqueous 2% CHX versus aqueous Ca(OH)2 slurry alone on the disinfection of the pulp space of failed root-filled teeth during endodontic retreatment in 40 teeth. The root canal was then cleaned and shaped with endodontic files using conventional endodontic technique. A copious amount of 1.0% NaOCl solution was used for irrigation. The results of this analysis were previously reported. The teeth were nonsurgically retreated and medicated over 3 treatment visits with 7-10-day intervals with either Ca(OH)2 in water or Ca(OH)2 in 2% aqueous CHX. Of the total sample population, 12 of 40 (30%) were positive for bacteria before root filling. The control medication disinfected 12 of 20 (60%) teeth including 2 of 4 teeth originally diagnosed with enterococci. The experimental medication resulted in 16 disinfected teeth out of 20 (80%) at the beginning of the third appointment. None of the teeth originally containing enterococci showed persistent bacterial growth. Canal dressing with a mixture of 2% CHX and Ca(OH)2 slurry is as efficacious as aqueous Ca(OH)2 on the disinfection of failed root-filled teeth. Williams, et al.29 compared real-time quantitative PCR (qPCR) assay to cultivation for E. faecalis detection and quantification during endodontic treatment. Final shaping and mechanical root canal debridement was achieved using nickel-titanium files in a rotary crown-down technique. Teeth were irrigated with 1.05% NaOCl between files and after the final file. In primary infections, E. faecalis was present in Sample 1 in 7% (1/15) of cases by cultivation and 13% (2/15) by qPCR. No tooth was positive for the bacterium in either Sample 2 or Sample 3 by cultivation, indicating the removal of culturable E. faecalis by the instrumentation - irrigation protocol. Using qPCR, 3 teeth (the two teeth identified at Sample 1 and another tooth) harbored the bacterium at both Sample 2 and Sample 3. As observed with the primary infections, there was a nonsignificant trend for the number of E. faecalis positive cases detected by qPCR to increase to 57% (8/14) in Sample 2 and 50% (7/14) in Sample 3. qPCR detected up to three times more E. faecalis in Sample 1 than cultivation, but the difference was not statistically significant. At collection times in Samples 2 and 3, qPCR identified more E. faecalis infections in refractory lesions than cultivation (Table 1).

Based on the meta-analysis results, in the selected 5 studies9-13, from a total of 159 teeth with endodontic infections, E. faecalis was detected initially in 16 (10%) teeth by PCR and 42 (26.4%) teeth by microbial culture techniques. Immediately after root canal preparation using 0.5% to 2.5% NaOCl, it was possibly to identify E. faecalis in 11 teeth (6.9%) by PCR and in 12 teeth (7.5%) by culture. No longitudinal studies in humans investigating the efficacy of NaOCl and CHX against E. faecalis from endodontic infections were found

The difficulty in comparing the studies retrieved in the present search is due to differences on the methodological design of each investigation: standardization of the limit of preparation, choice of the preparation technique, standardization of tooth type and sample size, time of the initial endodontic treatment in cases of secondary infection, quality control of the chemical irrigants and variation in their concentration, criteria for the detection of the periapical lesion etc, in addition to other important data that were not mentioned in these studies (Table 1).

The selection of endodontic irrigants that aggregate the largest possible number of ideal properties was a major point in the present study. There was a concern in this systematic review10 regarding to the process of making clinical decisions in the control of microorganisms in endodontic infections.



In summary, the disinfection of the root canal system produced by emptying, enlargement and action of NaOCl reduces the remaining endodontic microbiota, which optimizes the efficacy of the intracanal dressing and favors the achievement of a higher level of success of the endodontic treatment.



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Correspondence address:
Prof. Carlos Estrela
Centro de Ensino e Pesquisa Odontológica do Brazil (CEPOBRAS)
Rua C-245, Quadra 546, Lote 9, Jardim América
74.290-200, Goiânia, GO, Brazil

Received: December 05, 2007
Modification: June 16, 2008
Accepted: June 17, 2008

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