The properties and applications of chlorhexidine in endodontics

The properties and applications of chlorhexidine inendodontics Z. Mohammadi1,2 & P. V. Abbott31Department of Endodontics, School of Dentistry, Hamedan University of Medical Sciences, Hamedan, Iran; 2Iranian Centre forEndodontic Research (ICER); and 3School of Dentistry, University of Western Australia, Perth, WA, Australia action of CHX, its antibacterial and antifungal activity,its effect on biofilm, its substantivity (residual antibac- Mohammadi Z, Abbott PV. The properties and applications terial activity), its tissue solvent ability, its interaction of chlorhexidine in endodontics. International Endodontic Jour- with calcium hydroxide and sodium hypochlorite, its anticollagenolytic activity, its effect on coronal and Microorganisms and their by-products are considered apical leakage of bacteria, its toxicity and allergenicity to be the major cause of pulp and periradicular and the modulating effect of dentine and root canal pathosis. Hence, a major objective in root canal components on its antimicrobial activity. A Medline treatment is to disinfect the entire root canal system, search was performed from 1981 to the end of March which requires that all contents of the root canal 2008 and was limited to English-language papers. The system be eliminated as possible sources of infection.
keywords searched on Medline were ‘chlorhexidine This goal may be accomplished using mechanical AND endodontics’, ‘chlorhexidine AND root canal instrumentation and chemical irrigation, in conjunc- tion with medication of the root canal system between ‘chlorhexidine AND toxicity’. The reference lists of treatment sessions. To reduce or eliminate bacteria, each article were manually checked for additional various irrigation solutions have been advocated.
Chlorhexidine is a cationic molecule, which can be used during treatment. It has a wide range of antimi- crobial activity. Its cationic structure provides a uniqueproperty named substantivity. The purpose of this Received: 1 May 2008; accepted: 18 November 2008 paper is to review the structure and mechanism of complicated task involving the use of various instru- mentation techniques, irrigation regimens and intra- The major causative role of microorganisms in the canal medicaments. Mechanical instrumentation alone pathogenesis of pulp and periapical diseases has clearly does not result in a bacteria-free root canal system and been demonstrated (Kakehashi et al. 1965, Mo¨ller et al.
when the complex anatomy of the root canal system 1981, Sundqvist 1992). The elimination of microor- (Hess 1925) is considered, this is not surprising. On the ganisms from infected root canal systems is a other hand, ex vivo and clinical evidence has shownthat mechanical instrumentation leaves significantportions of the root canal walls untouched (Peters Correspondence: Dr Zahed Mohammadi, Department of et al. 2001) and complete elimination of bacteria by Endodontics, Hamedan Dental School, Shahid Fahmideh instrumentation alone is unlikely to occur (Bystro¨m & Street, Hamedan, Iran (Tel.: +98 918 8729690; fax: +98351 6250344; e-mail: mohammadi_zahed@yahoo.com).
Sundqvist 1981). It is assumed, but not demonstrated, Chlorhexidine in endodontics Mohammadi & Abbott that any pulp tissue left in the root canals can serve as a nutrient source for any remaining microorganisms.
Furthermore, tissue remnants also impede the antimi- Delany et al. (1982) evaluated 0.2% CHX-gluconate in crobial effects of root canal irrigants and medica- obtained before, during, immediately after and 24 h disinfection is necessary to remove residual tissue after instrumentation, irrigation and medication either and to kill microorganisms. Chemical treatment of the with CHX-gluconate or with sterile saline. There was a root canal can be arbitrarily divided into irrigants, highly significant reduction in the number of microor- canal rinses and inter-appointment medicaments.
ganisms in the CHX-treated specimens after instrumen- Chlorhexidine (CHX) is used widely as an endodontic tation and irrigation. Basson & Tait (2001) compared irrigant and medicament, but there has not been an adequate review of the literature regarding CHX.
[Ca(OH)2], iodine potassium iodide (IKI) and a CHX Hence, the purpose of this paper is to review different solution in disinfecting root canal systems that were aspects of CHX of relevance to endodontics. The infected with Actinomyces israelii. The root canals were literature review was performed using a Medline exposed to either IKI, calcium hydroxide or 2% CHX for electronic search. The search was performed from periods of 3, 7 and 60 days. CHX was the only 1981 to the end of March 2008 and was limited to disinfectant that was able to eliminate A. israelii from English-language papers. The keywords searched on all samples at all time periods whilst 25% of the specimens treated with IKI and 50% of the specimens ‘chlorhexidine AND root canal therapy’, ‘chlorhexi- treated with Ca(OH)2 still had viable A. israelii after dine AND substantivity’ and ‘chlorhexidine AND treatment. Onc¸ag˘ et al. (2003) evaluated the antibac- toxicity’. The reference lists of each article were terial properties of 5.25% sodium hypochlorite (NaOCl), manually checked for additional articles of relevance.
2% CHX and 0.2% CHX plus 0.2% cetrimide [Cetrexidin(GABA Vebas, San Giuliano Milanese, Italy)] after5 min and 48 h in extracted human teeth after the canals had been infected by Enterococcus faecalis. The Chlorhexidine is a synthetic cationic bis-guanide that 2% CHX and Cetrexidin were significantly more effec- consists of two symmetric 4-cholorophenyl rings and tive against E. faecalis than the 5.25% NaOCl at both two biguanide groups, connected by a central hexam- time periods. Two studies (Gomes et al. 2001, Vianna ethylene chain (Greenstein et al. 1986). CHX is a et al. 2004) have investigated the ex vivo antimicrobial positively charged hydrophobic and lipophilic molecule activity against endodontic pathogens of three concen- that interacts with phospholipids and lipopolysaccha- trations (0.2%, 1% and 2%) of two forms of CHX (gel rides on the cell membrane of bacteria and then enters and liquid) and compared them with five concentra- the cell through some type of active or passive tions of NaOCl (0.5%, 1%, 2.5%, 4% and 5.25%). Both transport mechanism (Athanassiadis et al. 2007). Its the 2% gel and 2% liquid formulations of CHX efficacy is because of the interaction of the positive eliminated Staphylococcus aureus and Candida albicans charge of the molecule and the negatively charged within 15 s, whereas the gel formulation killed phosphate groups on microbial cell walls (Gomes et al.
E. faecalis within 1 min. All of the tested irrigants 2003a,b), thereby altering the cells’ osmotic equilib- eliminated Porphyromonas endodontalis, Porphyromonas rium. This increases the permeability of the cell wall, gingivalis and Prevotella intermedia within 15 s. The which allows the CHX molecule to penetrate into the time required for 1.0% and 2.0% CHX liquid to bacteria. CHX is a base and is stable as a salt. The most eliminate all microorganisms was the same as the time common oral preparation, CHX gluconate, is water- required for 5.25% NaOCl. These studies confirm that soluble and at physiologic pH, it readily dissociates and the antimicrobial action is related to the type, concen- releases the positively charged CHX component (Green- tration and presentation form of the irrigants as well as stein et al. 1986). At low concentration (0.2%), low the microbial susceptibility to the formulation used.
molecular weight substances, specifically potassium Zamany et al. (2003) examined the effects of adding and phosphorous, will leak out of the cell. On the other a 2% CHX rinse to the conventional treatment protocol.
hand, at higher concentration (2%), CHX is bactericidal Their results showed that cultivable bacteria were as precipitation of the cytoplasmic contents occurs, retrieved at the conclusion of the first visit in one of the which results in cell death (Gomes et al. 2003a).
CHX cases, whereas seven of the 12 control cases Mohammadi & Abbott Chlorhexidine in endodontics without CHX showed growth; this difference was Another interesting topic is the additive effect of CHX statistically significant. Siqueira et al. (2007) compared and hydrogen peroxide. Heling & Chandler (1998) the effectiveness of 2.5% NaOCl and 0.12% CHX as studied the antimicrobial effect of irrigant combinations irrigants in reducing the cultivable bacteria in infected within dentinal tubules ex vivo against E. faecalis and root canal systems of teeth with apical periodontitis.
found that a specific combination of 3% hydrogen They found that the two solutions had comparable peroxide (H2O2) and CHX was superior in its antibac- effects in eliminating bacteria and they suggested that terial activity in dentine compared with other regi- mens, such as CHX alone and NaOCl. Steinberg et al.
In a randomized clinical trial, Manzur et al. (2007) (1999) challenged E. faecalis suspensions in trypticase assessed the antibacterial efficacy of intracanal medi- soy broth (a culture medium rich in peptides) with cation with Ca(OH)2, 2% CHX gel and a combination of various combinations of CHX and H2O2. The experi- both [Ca(OH)2/CHX] in teeth with chronic apical ments demonstrated that the combination of the two periodontitis. Bacteriological samples were obtained substances totally killed E. faecalis at concentrations from the operative field and the root canals before and much lower than that required for each component after instrumentation in the first treatment session.
alone. According to that study, the bactericidal effect of Further samples were taken from the canals at the CHX is derived from its ability to denature the bacterial commencement of the second appointment 1 week cell wall whilst forming pores in the membrane, later. They concluded that the antibacterial efficacies of whereas H2O2 is effective against intracellular organ- Ca(OH)2, CHX and a mixture of Ca(OH)2/CHX were elles, such as DNA. Although the exact synergistic mechanism of CHX and H2O2 is not known, it can be Zerella et al. (2005) investigated the effect of a slurry postulated that the exposure of bacteria to CHX leads to of Ca(OH)2 mixed in aqueous 2% CHX versus aqueous a more permeable cell wall that the H2O2 can easily Ca(OH)2 alone on the disinfection of the root canal penetrate and hence damage the intracellular organelles system of root filled teeth that required root canal (Steinberg et al. 1999). Shabahang et al. (2008) evalu- re-treatment because the canals had become infected ated the antibacterial efficacy of the substitution of CHX again. Twelve (30%) of the 40 samples were positive for for doxycycline in MTAD against a strain of E. faecalis bacteria before root filling. The control medication ex vivo. Findings showed that the presence of doxycycline disinfected 12 (60%) of 20 teeth including two of four in the concentration included in the MTAD formulation teeth that had been originally diagnosed with entero- was effective in eliminating E. faecalis. Furthermore, the cocci. The experimental medication resulted in disinfec- addition of 0.2% CHX did not adversely affect the tion of 16 of 20 (80%) teeth at the beginning of the antibacterial action of doxycycline. On the other hand, third appointment. None of the teeth originally con- the substitution of 0.2% CHX did not allow the same taining enterococci showed remaining growth. They disinfection efficacy on E. faecalis as MTAD.
concluded that a mixture of 2% CHX and a Ca(OH)2 On the whole, although studies comparing the slurry is as efficacious as aqueous Ca(OH)2 on the antibacterial effect of CHX and NaOCl have produced disinfection of infected root filled teeth.
somewhat conflicting results, it seems that when used Ercan et al. (2004) evaluated the antibacterial activ- in identical concentrations, their antibacterial effects ex ity of 2% CHX and 5.25% NaOCl in infected root canals vivo (in infected dentine) and in vivo (in the root canal of incisors and premolars. They concluded that both CHX and NaOCl were effective irrigants for reducingthe number of microorganisms in teeth with a necrotic Tanomaru et al. (2003) evaluated the effect of Fungi (or yeasts) constitute a small proportion of the biomechanical preparation with 5% NaOCl, 2% CHX usual oral microbiota with Candida species being the and physiological saline irrigating solutions and most common of the fungi present in both healthy (30– Ca(OH)2 dressing in the root canals of dogs’ teeth that 45%) and medically compromised (95%) individuals contained bacterial endotoxin. They found that biome- (Siqueira & Sen 2004). Fungi have occasionally been chanical preparation with the irrigating solutions did found in infected root canals that have not had any not inactivate the endotoxin, but the calcium hydrox- previous endodontic treatment, but they are more ide intracanal dressing did inactivate the effects common in filled root canals in teeth that have become infected some time after treatment or in those that have Chlorhexidine in endodontics Mohammadi & Abbott not responded to endodontic treatment (23). Overall, Infected dentine cylinders were exposed to four different the occurrence of fungi reported in infected root canals medications: Ca(OH)2/glycerin; Ca(OH)2/0.12% CHX; varies between 1% and 17% (Waltimo et al. 2004).
Fungi may be involved in cases of persistent and erin and 0.12% CHX/zinc oxide. The specimens treated secondary infections associated with recalcitrant per- with the Ca(OH)2/camphorated monoparachlorophe- iradicular lesions and therefore the spectrum of anti- nol/glycerin paste or with the CHX/zinc oxide paste microbial activity of endodontic medicaments and were completely disinfected after 1 h of exposure whilst irrigants should include these organisms. Thus, med- the Ca(OH)2/glycerin paste consistently eliminated the icaments that have antifungal effectiveness may assist C. albicans after 7 days of exposure. Calcium hydroxide in the successful management of persistent or second- mixed with CHX was ineffective in disinfecting dentine ary endodontic infections caused by fungi (Siqueira & even after 1 week. In another study, Siqueira et al.
Sen 2004, Waltimo et al. 2004). To try and improve (2001) investigated the antifungal activity of several antisepsis in single-appointment endodontic treatment medicaments against C. albicans, Candida glabrata, regimes, it has been suggested to irrigate and/or ‘soak’ Candida guilliermondii, Candida parapsilosis and Saccha- the root canals with either CHX or iodine-IKI solutions romyces cerevisiae. Calcium hydroxide mixed with following irrigation with NaOCl. Aqueous CHX solution CPMC/glycerin as a paste showed the most pronounced has a wide spectrum of antimicrobial activity at low antifungal effects. Calcium hydroxide in glycerin, concentrations and is especially effective against Ca(OH)2 with CHX and CHX in detergent had less C. albicans. Furthermore, it binds to surrounding tissues antifungal activity. Ferguson et al. (2002) sought to and can then be released again slowly over extended determine the in vitro susceptibility of C. albicans to periods of time, a phenomenon known as substantivity.
various irrigants and medicaments. The minimum Interestingly, it appears that CHX can efficiently inhibit inhibitory concentrations of NaOCl, hydrogen peroxide, the initial adherence and perhaps further accumulation CHX-digluconate and aqueous Ca(OH)2 were deter- and biofilm formation of fungi and other micro- mined. Their results revealed that NaOCl, hydrogen organisms. A recent clinical study has shown that peroxide and CHX-digluconate were effective against canals that received a final rinse with a 2% CHX C. albicans even when significantly diluted. However, solution were significantly more often free of cultivable aqueous Ca(OH)2 had no anti-fungal activity. Taken microorganisms than controls irrigated with NaOCl together, it can be concluded that CHX is an effective alone (Siqueira & Sen 2004, Waltimo et al. 2004).
antifungal agent, but its efficacy is significantly less Sen et al. (1999) evaluated the antifungal properties of 0.12% CHX, 1% NaOCl and 5% NaOCl againstCandida albicans using a cylindrical dentine tube model.
They reported that C. albicans was more resistant tothese irrigant solutions when the smear layer was The term biofilm was introduced to designate the thin- present than when it was absent. When smear layer layered condensations of microbes that may occur on was absent, the NaOCl started to display antifungal various surface structures in nature. Free-floating activity after 30 min. Waltimo et al. (1999) evaluated bacteria existing in an aqueous environment, the so- the susceptibility of seven strains of C. albicans to four called planktonic form of microorganisms, are a pre- disinfectants, namely IKI, CHX-acetate (0.5%), NaOCl requisite for biofilm formation (Bowden & Hamilton (5% and 0.5%) and Ca(OH)2. Each solution was tested 1998). Biofilms may thus become established on any individually as well as in pairs using all possible pairs of organic or inorganic surface substrate where plank- these four disinfectants. All C. albicans strains tested tonic microorganisms prevail in a water-based solution.
showed similar susceptibility to these medicaments.
In dental contexts, a well-known and extensively They were highly resistant to Ca(OH)2, but the NaOCl studied biofilm structure is established during the and IKI killed all cells within 30 s and the CHX-acetate attachment of bacteria to teeth to form dental plaque.
showed complete killing after 5 min. Combinations of Here, bacteria free in saliva (planktonic organisms) disinfectants were either equally or less effective than serve as the primary source of organisms for the the more effective component of the pair tested.
organization of this specific biofilm (Bowden & Hamil- Siqueira et al. (2003) evaluated the effectiveness of ton 1998). In endodontics, the biofilm concept was four intracanal medications in disinfecting root dentine initially discussed mainly within the framework of in bovine teeth experimentally infected with C. albicans.
bacteria on the root tips of teeth with necrotic and Mohammadi & Abbott Chlorhexidine in endodontics infected pulps or pulpless and infected root canal kill was found. The percentage kills of the bacteria were: 6% NaOCl (>99.99%), 1% NaOCl (99.78%), thought to be the cause of therapy-resistant apical Smear ClearÔ (78.06%), 2% CHX (60.49%), REDTA periodontitis (Tronstad & Sunde 2003). Although not (26.99%) and BioPureÔ MTADÔ (16.08%). There was described in as much detail, bacterial condensations a significant difference between NaOCl (both concen- (that is, biofilms) on the walls of infected root canals trations of 1% and 6%) and all other agents. Therefore, both 1% NaOCl and 6% NaOCl were more efficient in Antimicrobial agents have often been developed and eliminating E. faecalis biofilm than the other solutions optimized for their activity against fast growing, tested. In another study, Lima et al. (2001) assessed the dispersed populations containing a single microorgan- effectiveness of CHX-based or antibiotic-based (clinda- ism. However, microbial communities grown in bio- mycin and metronidazole) medications in eliminating 1- and 3-day E. faecalis biofilms. Each biofilm-contain- antimicrobial agents and microorganisms in mature ing membrane was thoroughly covered with 1 mL of biofilms can be notoriously resistant for reasons that the test medications and incubated for 1 day at 37 °C.
have yet to be adequately explained (Bowden & The treated biofilms were then aseptically transferred to Hamilton 1998). There are reports showing that vials containing a neutralizing agent in saline solution microorganisms grown in biofilms could be twofold to and vortexed. Suspensions were diluted 10-fold, seeded 1000-fold more resistant than the corresponding onto Mitis salivarius agar plates and the colony-forming planktonic form of the same organisms (Svensater & units counted after 48 h of incubation. There were Bergenholtz 2004). Spratt et al. (2001) have evaluated significant differences between the formulations tested.
the effectiveness of 2.25% NaOCl, 0.2% CHX, 10% The association of clindamycin with metronidazole povidone iodine, 5 ppm colloidal silver and phosphate significantly reduced the number of cells in the 1-day buffered solution (PBS) as a control against monocul- biofilms. However, of all medications tested, only 2% ture biofilms of five root canal isolates including P.
CHX-containing medications were able to thoroughly intermedia, Peptostreptococcus miros, Streptococcus inter- eliminate most of both the 1-day and 3-day E. faecalis medius, Fusobacterium nucleatum and E. faecalis. They biofilms. On the whole, it seems that although CHX is reported that NaOCl was the most effective anti- somewhat effective against bacterial biofilms, NaOCl is microbial agent followed by the iodine solution. Clegg the only irrigation solution with the capability of et al. (2006) evaluated the ex vivo effectiveness against apical dentine biofilms of three concentrations of NaOCl(6%, 3% and 1%), 2% CHX and a commercially available mixture of a tetracycline, an acid and adetergent known as BioPure MTAD (Dentsply, Tulsa Chlorhexidine as well as tetracyclines have a unique Dental, Tulsa, OK, USA). They reported that the 6% feature in that dentine medicated with it acquires NaOCl and 3% NaOCl were capable of disrupting and antimicrobial substantivity (Khademi et al. 2006). The removing the biofilm, but the 1% NaOCl and the MTAD positively charged ions released by CHX can adsorb into were capable of disrupting the biofilm, but did not dentine and prevent microbial colonization on the eliminate the bacteria. The 2% CHX was not capable of dentine surface for some time beyond the actual the disrupting the biofilm. Viable bacteria could not be period of time of application of the medicament cultured from specimens exposed to 6% NaOCl, 2% CHX or 1% NaOCl followed by BioPure MTAD. Duna- The antimicrobial substantivity of CHX has been vant et al. (2006) evaluated the efficacy of 6% NaOCl, assessed in several periodontal and endodontic studies.
1% NaOCl, Smear ClearÔ (SybronEndo, Orange, CA, In an in vivo periodontal study, Stabholz et al. (1993) USA), 2% CHX, REDTA (Roths International Ltd, evaluated the substantivity of the human root surface Chicago, IL, USA) and BioPureÔ MTADÔ against E.
after in situ subgingival irrigation with tetracycline HCL faecalis biofilms using a novel laboratory testing system.
and CHX. They found that the substantivity of tetracy- Biofilms grown in a flow cell system were submerged in cline at 50 mg mL)1 was significantly greater than that test irrigants for either 1 or 5 min. There was a of CHX for 12 days and greater than saline for 16 days.
significant relationship between the test agent and the In a laboratory study, White et al. (1997) evaluated percentage kill of the bacteria in the biofilm. No the antimicrobial substantivity of a 2% CHX solution as significant relationship between time and percentage an endodontic irrigant. Findings showed that the Chlorhexidine in endodontics Mohammadi & Abbott substantivity lasted 72 h. In an in vivo study, Leonardo years. Haapasalo et al. (2000) introduced a new et al. (1999) evaluated the antimicrobial substantivity dentine powder model for studying the inhibitory effect of 2% CHX used as a root canal irrigating solution in of dentine on various root canal irrigants and medica- teeth with pulp necrosis and radiographically visible ments. They investigated the modulating effect of chronic periapical lesions. They reported that the CHX dentine on the antibacterial activity of saturated prevented microbial activity with residual effects in the CA(OH)2 solution, 1% NaOCl, 0.5% and 0.05% CHX root canal system for up to 48 h after application.
acetate and 2/4% and 0.2/0.4% IKI. Dentine powder However, some other studies have reported that the had an inhibitory effect on all medicaments tested. The substantivity of CHX can last for longer periods of time.
effect was dependent on the concentration of the Khademi et al. (2006) found that 5 min application of medicament as well as on the length of time the 2% CHX solution induced substantivity for up to medicament was pre-incubated with the dentine 4 weeks. Rosenthal et al. (2004) evaluated the sub- powder before adding the bacteria. Similarly, 0.2/ stantivity of 2% CHX solution within the root canal 0.4% IKI lost its effect after pre-incubation for 1 h with system after 10 min of application and they reported dentine before adding the bacteria. The effect of 0.05% that the CHX was retained in the root canal dentine in CHX and 1% NaOCl on E. faecalis was reduced, but not antimicrobially effective amounts for up to 12 weeks.
totally eliminated by the presence of dentine. No Antimicrobial substantivity depends on the number inhibition could be measured when full strength of CHX molecules available to interact with the dentine.
solutions of CHX and IKI were used in killing E. faecalis.
Therefore, medicating the canal with a more concen- Portenier et al. (2001) evaluated the inhibition of the trated CHX preparation should result in increased antibacterial effect of saturated Ca(OH)2 solution, CHX- resistance to microbial colonization. The antibacterial acetate and IKI by dentine, hydroxylapatite (HA) and substantivity of three concentrations of CHX solution bovine serum albumin (BSA). Calcium hydroxide was (4%, 2% and 0.2%) after 5 min of application has been totally inactivated by the presence of 28 mg of dentine powder or BSA. CHX (0.05%) was strongly inhibited by between the concentration of CHX and its substantivity BSA and slowed down by dentine. However, HA had (Mohammadi et al. 2008). On the contrary, Lin et al.
little or no inhibitory effect on CHX. The antibacterial (2003a) attributed the substantivity of CHX to its ability effect of 0.2/0.4% IKI on E. faecalis was totally inhibited to adsorb on to the dentine during the first hour. They by dentine (28 mg), but was practically unaffected by stated that it is only after the saturation point is reached HA or BSA. A stepwise reduction of dentine from 28 to after the first hour that the antimicrobial capability of 2.8 mg 150 lL)1 was followed by a similar reduction CHX increases with time. Furthermore, Komorowski of the inhibition of the antibacterial activity of CHX. IKI et al. (2000) revealed that 5 min application of CHX did was not inhibited at all with dentine amounts <28 mg.
not induce substantivity and that the dentine should be However, the effect of saturated calcium hydroxide treated with CHX for 7 days. Taken together, it seems solution was totally eliminated by dentine at all four that residual antimicrobial activity of CHX in the root concentrations tested. It could be assumed that inhibi- canal system remains for up to 12 weeks.
tion by dentine of the antibacterial activity of Ca(OH)2,CHX and IKI occurs by different mechanisms (Portenieret al. 2001). Surprisingly, Ca(OH) inhibitory effect of all three materials tested. The The root canal milieu is a complex mixture of a variety inhibition of Ca(OH)2 by dentine and by the other of organic and inorganic compounds. Hydroxyapatite, compounds is, of course, dependent on their quantita- the main component of dentine, is the major represen- tive relationships (Portenier et al. 2001). One major tative of inorganic components present. In addition, mechanism for resistance of survival of E. faecalis in the inflammatory exudate, entering the apical root canal in root canal filled with Ca(OH)2 may be the buffering purulent infections, is rich in proteins, such as albumin.
effect of dentine against the pH rise. Inorganic HA had The relative importance of the various organic and little or no inhibitory activity against CHX as compared inorganic compounds in the inactivation of root canal with dentine, whereas BSA was the strongest inhibitor disinfectants have been studied restrictively (Haapasalo of CHX, with more than 10% of E. faecalis cells still et al. 2000). Difficulties in designing experiments that viable after 24 h of incubation with the medicament.
will give reliable and comparable data have been some This indicates that periapical inflammatory exudate of the greatest challenges for researchers for many entering the root canal is a greater threat to the activity Mohammadi & Abbott Chlorhexidine in endodontics of CHX than the dentine walls. Dentine powder totally the frequency of shaking, the amount of organic matter eliminated the antibacterial effect of IKI; whereas the in relation to the amount of irrigant in the system and major component of dentine, HA did not affect the the surface area of tissue that was available for contact antibacterial effect of IKI, nor did BSA. In addition, it is with the irrigant. Okino et al. (2004) evaluated the generally known that blood rapidly inactivates the tissue dissolving ability of 0.5%, 1.0% and 2.5% NaOCl, antibacterial activity of iodine compounds (Portenier 2% aqueous solution of CHX-digluconate, 2% CHX et al. 2001). In another study, Portenier et al. (2002) digluconate gel and distilled water as the control.
assessed the antibacterial activity of CHX and IKI on E.
Bovine pulp fragments were weighed and placed in faecalis in the presence of dentine, dentine matrix, contact with 20 mL of each tested substance in a dentine pre-treated by ethylenediamine tetraacetic acid centrifuge at 150 rpm until total dissolution. Dissolu- (EDTA) and citric acid, collagen and heat-killed cells of tion speed was calculated by dividing the pulp weight E. faecalis and Candida albicans. Dentine matrix and by the dissolution time. Distilled water and both heat-killed microbial cells were the most effective solutions of CHX did not dissolve the pulp tissue within inhibitors of CHX, whereas dentine pre-treated by citric 6 h. The mean dissolution speeds for 0.5%, 1.0% and acid or EDTA showed only slight inhibition. Dentine and skin collagen showed some inhibition at 1 h, but 0.55 mg min)1, respectively. In another study, Naenni not after 24 h. IKI was effectively inhibited by dentine, et al. (2004) assessed the necrotic tissue dissolution dentine matrix and heat-killed microbial cells. Skin capacity of 1% (w/v) NaOCl, 10% CHX, 3% and 30% collagen and dentine pre-treated by EDTA or by citric hydrogen peroxide, 10% peracetic acid, 5% dichloro- acid showed little or no inhibitory effect on IKI. The isocyanurate (NaDCC) and 10% citric acid. Standard- inhibitory effect of dentine and BSA on the antibacterial ized necrotic tissue samples obtained from pig palates activity of CHX and MTAD was assessed in another were incubated in these solutions and their weight loss study (Portenier et al. 2006). The presence of dentine was measured over time. None of the test solutions or BSA caused a marked delay in the killing of E. faecalis except NaOCl had any substantial tissue dissolution by both medicaments. The inhibitory effect of BSA on capacity. It was concluded that this might be important the antibacterial activity of CHX and NaOCl has been when considering the use of irrigants other than confirmed by Sassone et al. (2008). Taken together, it NaOCl. On the whole, one of the major disadvantages seems that dentine, dentine components (HA and of CHX is that it has no tissue solvent activity.
collagen), killed microorganisms and inflammatoryexudate in the root canal system all reduce or inhibit the antibacterial activity of medicaments and irrigants.
On the whole, it seems that dentine, dentine compo- Chlorhexidine is a cationic biguanide whose optimal nents (HA and collagen), killed microorganisms and antimicrobial activity is achieved within a pH range of inflammatory exudates in the root canal system reduce 5.5–7.0 (Athanassiadis et al. 2007). Therefore, it is likely that alkalinizing the pH by adding Ca(OH)2 toCHX will lead to precipitation of the CHX molecules andthereby decreases its effectiveness. It has been demon- strated that the alkalinity of Ca(OH)2 when mixed with Several studies have been conducted in the search for CHX remained unchanged. Therefore, the usefulness of an irrigant that meets the four major desirable prop- mixing Ca(OH)2 with CHX still remains unclear and erties for root canal irrigants – namely: antimicrobial controversial (Athanassiadis et al. 2007).
activity, nontoxicity to the periapical tissues, water When used as an intracanal medicament, CHX was solubility and the capacity to dissolve organic matter.
more effective than Ca(OH)2 in eliminating E. faecalis Therefore, an ideal irrigant should dissolve the organic from inside dentinal tubules (Athanassiadis et al.
matter inside the root canal system. Grossman & 2007). In a study by Almyroudi et al. (2002), all of Meiman (1941) demonstrated the importance of the the CHX formulations used, including a CHX/ Ca(OH)2 solvent ability of an endodontic irrigant and empha- 50:50 mix, were efficient in eliminating E. faecalis from sized that the elimination of pulp tissue from the root the dentinal tubules with a 1% CHX gel working canal was important for the ultimate success of root slightly better than the other preparations. These canal treatment. Moorer & Wesselink (2003) showed findings were corroborated by Gomes et al. (2006) in that tissue dissolution was dependent on three factors: bovine dentine and Schafer & Bossmann (2005) in Chlorhexidine in endodontics Mohammadi & Abbott human dentine where 2% CHX gel had greater activity canal system of teeth with coronal restorations medi- against E. faecalis, followed by CHX/ Ca(OH)2 and then cated with either calcium hydroxide, 2% CHX gel or with a combination of both. The canals without a In a study using agar diffusion, Haenni et al. (2003) coronal restoration, but medicated with CHX, showed could not demonstrate any additive antibacterial effect recontamination after an average time of 3.7 days; the by mixing Ca(OH)2 powder with 0.5% CHX and they group with Ca(OH)2 after 1.8 days and the group with showed that the CHX had a reduced antibacterial action.
CHX+Ca(OH)2 after 2.6 days. The canals medicated However, Ca(OH)2 did not lose its antibacterial proper- with CHX and restored with IRM showed recontami- ties in such a mixture. This may be because of the nation within 13.5 days; the group with Ca(OH)2+IRM deprotonation of CHX at a pH >10, which reduces its after 17.2 days and the group with CHX+ Ca(O- solubility and alters its interaction with bacterial sur- H)2+IRM after 11.9 days. The group with no medica- faces as a result of the altered charge of the molecule. In tion, but restored with IRM, showed recontamination an in vitro study using human teeth, Ercan et al. (2006) after an average time of 8.7 days. There were statisti- showed 2% CHX gel was the most effective agent against E. faecalis inside dentinal tubules, followed by a Ca(OH)2/ (P < 0.05). All groups without a coronal restoration 2% CHX mix, whilst Ca(OH)2 alone was totally ineffec- were recontaminated significantly more quickly than tive, even after 30 days. The 2% CHX gel was also those restored with IRM, except those teeth that had a significantly more effective than the Ca(OH)2/2% CHX restoration, but no medicament. The groups with mix against C. albicans at 7 days, although there was no intracanal medication and a coronal restoration were significant difference at 15 and 30 days. Ca(OH)2 alone not significantly different from each other.
was completely ineffective against C. albicans. In another Vivacqua-Gomes et al. (2002) assessed ex vivo coro- in vivo study using primary teeth, a 1% CHX-gluconate nal dye penetration of extracted human teeth after root gel, both with and without Ca(OH)2, was more effective canal treatment using 1% NaOCl, 1% NaOCl + 17% against E. faecalis than CH alone over a 48-h period EDTA, 2% CHX gel, 2% CHX gel + 1% NaOCl and distilled water. After root canal filling, the teeth were Schafer & Bossmann (2005) reported that 2% CHX- incubated at 37 °C for 10 days followed by 10 days gluconate was significantly more effective against immersion in human saliva and an additional 10 days E. faecalis than Ca(OH)2 used alone or a mixture of in India ink. The teeth were cleared and the maximum the two. This was also confirmed by Lin et al. (2003b) depth of dye penetration was determined digitally in although in a study by Evans et al. (2003) using bovine millimetres. Results revealed that the least dye pene- dentine, 2% CHX with Ca(OH)2 was shown to be more tration occurred with 1% NaOCl + 17% EDTA and 2% effective than Ca(OH)2 in water. In an animal study, CHX gel. NaOCl, distilled water and 2% CHX gel + 1% Lindskog et al. (1998) reported that teeth dressed with NaOCl had more dye penetration with a significant CHX for 4 weeks had reduced inflammatory reactions difference compared with NaOCl + 17% EDTA and 2% in the periodontium (both apically and marginally) and CHX gel and compared with one another.
less root resorption. Waltimo et al. (1999) reported that Other studies have shown that viscous irrigants, 0.5% CHX-acetate was more effective at killing C.
including those containing CHX gluconate, were less albicans than saturated Ca(OH)2, whilst Ca(OH)2 com- soluble substances and they can leave residues on the bined with CHX was more effective than Ca(OH)2 used root canal surfaces, which may affect the root canal alone. The high pH of Ca(OH)2 was unaffected when filling. Lambrianidis et al. (2006) investigated the combined with CHX in this study. Taken together, it efficiency of removing Ca(OH)2/CHX gel, Ca(OH)2/ seems that the usefulness of mixing Ca(OH)2 with CHX CHX solution and Ca(OH)2/saline pastes with the use of instrumentation and irrigation with NaOCl andEDTA solutions. None of the techniques used in thisstudy removed the inter-appointment root canal med- icaments effectively (Lambrianidis et al. 2006). Overall, Because of its antimicrobial substantivity, it seems that Ca(OH)2/CHX (gel) paste was associated with signifi- CHX preparations delay entry of bacteria through the cantly larger amount of residue, whereas the Ca(OH)2/ coronal portion of the tooth into the root canal system.
CHX mixture was associated with less residue than the In a laboratory study, Gomes et al. (2003b) investi- other two medicaments. When all these studies are gated the time required for recontamination of the root considered it appears as although CHX used as an Mohammadi & Abbott Chlorhexidine in endodontics intracanal medicament and/or irrigant may delay Gomes et al. 2002). The formation of a precipitate could recontamination of the root canal system via the be explained by the acid–base reaction that occurs coronal route because of its substantivity. Overall, when NaOCl and CHX are mixed together. CHX, a because of its substantivity, CHX as an intracanal dicationic acid has the ability to donate protons whilst medicament/irrigant delays recontamination of the NaOCl is alkaline and can accept protons from the root canal system via the coronal route.
dicationic acid. This proton exchange results in theformation of a neutral and insoluble substance, referredto as the ‘precipitate’ (Basrani et al. 2007). Basrani et al.
(2007) evaluated the chemical nature of this precipitate Marley et al. (2001) assessed the effect of 0.12% and reported that there was an immediate reaction CHX-gluconate as an endodontic irrigating solution when 2% CHX was combined with NaOCl, even at the on the apical seal of root filled canals using three low concentration (0.023%). Increasing of the concen- different cements (Roth’s 801, AH26 and Sealapex).
tration of NaOCl to 0.19% (the sixth dilution in their At 90 and 180 days after root filling, apical fluid series) resulted in the formation of a precipitate, which penetration was measured using the fluid filtration consisted mainly of para-chloroaniline (PCA). This method. The results showed no significant differences occurred through a substitution of the guanidine group related to the irrigants at both the 90- and 180-day in the CHX molecule. They found that the amount of observation periods. Furthermore, the same group PCA directly increased with the increasing concentra- reported that at long-term periods (270 and 360 tion of NaOCl. PCA has been shown to be toxic with days), CHX-gluconate irrigant did not adversely affect short-term exposure of humans to PCA resulting in the apical penetration of fluid with the different root cyanosis, which is a manifestation of methemoglobin canal cements (Ferguson et al. 2003). Wuerch et al.
formation. In another study, Bui et al. (2008) evaluated the effect of irrigating root canals with a combination of Ca(OH)2 on the apical seal of root canal fillings.
NaOCl and CHX on root dentine and dentinal tubules by They reported that 2% CHX gel and Ca(OH)2 paste using the environmental scanning electron microscope did not adversely affect the apical seal. These findings (FEI Quanta 200, Hillsboro, OR, USA) and a computer were also confirmed by Engel et al. (2005). Overall, it program (photoshop cs2, Adobe Systems, San Jose, CA, seems that medication and/or irrigation with CHX USA). Their findings indicated that there were no does not adversely affect the ability of root fillings to significant differences in the amount of debris remain- prevent fluid penetration into the root canal system ing between the negative control group and the experimental groups although there were significantlyfewer patent tubules in the experimental groups whencompared with the negative control group. They concluded that the NaOCl/CHX precipitate tends to A suggested clinical protocol by Zehnder (2006) for occlude the dentinal tubules and suggested that until treating the dentine before root canal filling consists of this precipitate is studied further, caution should be irrigation with NaOCl to dissolve the organic compo- exercised when irrigating with both NaOCl and CHX.
nents, irrigation with EDTA to eliminate the smear layer Taken together, the combination of NaOCl and CHX and irrigation with CHX to increase the anti-microbial causes colour changes and formation of a neutral and spectrum of activity and to impart substantivity.
insoluble precipitate, which may interfere with the seal Although such a combination of irrigants may enhance of the root filling. Copious amounts of CHX irrigant the overall antimicrobial effectiveness (Kuruvilla & should be administered to prevent discolouring of the Kamath 1998), the possible chemical interactions tooth by this precipitate. Alternatively, the canal can be amongst the irrigants have to be considered. Some dried using paper points before the final CHX rinse studies have reported the occurrence of colour change and precipitation when NaOCl and CHX are combined(Vivacqua-Gomes et al. 2002, Zehnder 2006, Basrani et al. 2007). Furthermore, concern has been raised that the colour change may have some clinical relevancebecause of staining and that the precipitate might During the last two decades, chemical and technical interfere with the seal of the root filling (Vivacqua- advances have contributed to increases in resin–dentine Chlorhexidine in endodontics Mohammadi & Abbott bond strength. However, the premature loss of bond concluded that auto-degradation of collagen matrices strength is one of the problems that still affects adhesive can occur in resin-infiltrated dentine, but may be restorations (Mjo¨r et al. 2000) and markedly reduces prevented by the application of a synthetic protease their durability (Carrilho et al. 2005b, De Munck et al.
inhibitor, such as CHX (Carrilho et al. 2007b). On the 2005, Frankenberger et al. 2005). The loss of bond whole, because of its broad-spectrum MMP-inhibitory strength has been attributed mainly to the degradation effect, CHX can significantly improve the resin–dentine of the hybrid layer at the dentine-adhesive interface.
Numerous publications have demonstrated this lack ofbond stability (Wang & Spencer 2003, 2005, Yiu et al.
2004, Carrilho et al. 2005a). The notion that deterio-ration of dentine collagen fibrils contributes to the Results from a study of the cytotoxic effect of chloreh- mechanism responsible for bond degradation has been exidine on canine embryonic fibroblasts and Staphylo- reported (Hashimoto et al. 2003, Pashley et al. 2004).
coccus aureus showed that bactericidal concentrations In this context, it has been speculated that a decreasing of CHX were lethal to canine embryonic fibroblasts concentration gradient of resin monomer diffusion whilst noncytotoxic concentrations allowed significant within the acid-etched dentine and a subsequent resin bacterial survival (Sanchez et al. 1988). In a study by elution from hydrolytically unstable polymeric hydro- Tatnall et al. (1990), the cytotoxic effects of CHX, gels within the hybrid layers (Wang & Spencer 2003) hydrogen peroxide and NaOCl were examined on leaves the collagen fibrils unprotected and vulnerable cultured human fibroblasts, basal keratinocytes and a to degradation by endogenous metalloproteinases transformed keratinocyte line (SVK 14 cells). At con- (MMPs). The MMPs are a group of 23 mammalian centrations recommended for wound cleansing, all enzymes capable of degrading all extracellular matrix agents produced 100% killing of all cell types. Com- components. Human dentin contains at least collage- parison of the ED50 concentration for each agent on all nase (MMP-8), gelatinases MMP-2 and -9 and enam- cell types produced a ranking order of toxicity showing elysin MMP-20 (Martin-De Las Heras et al. 2000, CHX to be the least toxic antiseptic agent.
Sulkala et al. 2002, 2007, Mazzoni et al. 2006).
Results from a laboratory study on the toxicity of CHX to human gingival cells showed that the toxic (Pashley et al. 2004) can be suppressed by protease potency of CHX is dependent on the length of inhibitors (Pashley et al. 2004), indicating that MMP inhibition could be beneficial in the preservation of medium (Babich et al. 1995). The addition of foetal hybrid layers. This was demonstrated in an in vivo study, in which the application of CHX, known to have Escherichia coli reduced the cytotoxicity of CHX, a broad-spectrum MMP-inhibitory effect (Gendron et al.
presumably because of the binding of the cationic 1999), significantly improved the integrity of the CHX to the negatively charged chemical moieties/ hybrid layer in a 6-month clinical trial (Hebling et al.
2005). Carrilho et al. (2007a) evaluated the effect of 1995). These findings suggest that similar reactions CHX on the resin–dentine bond stability ex vivo. Results within a root canal may reduce the potential of a showed that with CHX, significantly better preservation cytotoxic reaction in the periapical tissues (Boyce of bond strength was observed after 6 months and et al. 1995). Boyce et al. (1995) found CHX (0.05%) protease inhibitors in the storage medium had no effect.
uniformly toxic to both cultured human cells and Failure analysis showed significantly less failure in the microorganisms. Agarwal et al. (1997) found that hybrid layer with CHX, compared with controls after CHX rapidly disrupts the cell membrane of both 6 months. Furthermore, they evaluated the effect of crevicular and peripheral blood neutrophils at con- CHX on the preservation of the hybrid layer in vivo.
centrations above 0.005% within 5 min, indicating Findings showed that bond strength remained stable in that its inhibitory effect on neutrophil function is the CHX-treated specimens, whilst bond strength mostly because of its lytic properties. Yesilsoy et al.
decreased significantly in control teeth. Resin-infil- (1995) assessed the short-term toxic effects of CHX in trated dentine in CHX-treated specimens exhibited the subcutaneous tissue of guinea pigs and found normal structural integrity of the collagen network.
moderate inflammation present after 2 days, followed Conversely, progressive disintegration of the fibrillar by a foreign-body granuloma formation at 2 weeks.
network was identified in control specimens. They Mohammadi & Abbott Chlorhexidine in endodontics (potential damage to DNA) of formocresol, para- 3. The effect of CHX on microbial biofilms is signifi- against Chinese hamster ovary cells. Results showed 4. CHX has antibacterial substantivity in dentine for that none of the mentioned agents contributed to DNA damage. Taken together, in the clinically used 5. Dentine, dentine components (HA and collagen), concentrations, the biocompatibility of CHX is accept- killed microorganisms and inflammatory exudate in the able. The potentially toxic interactions between CHX root canal system may reduce or inhibit the antibac- and NaOCl were considered previously.
terial activity of CHX.
6. CHX has little to no ability to dissolve organictissues.
7. Mixing CHX with Ca(OH)2 may enhance its antimi- Although sensitivity to CHX is rare, contact dermatitis is a common adverse reaction to CHX (Krautheim et al.
8. Medication and/or irrigation with CHX may delay 2004). Apart from this, CHX may have a number of rare the contamination of root filled teeth by bacteria side effects, such as desquamative gingivitis, discolour- entering through the coronal restoration/tooth inter- ation of the teeth and tongue or dysgeusia (distorted taste). Contact with conjunctiva can cause permanent 9. Medication and/or irrigation with CHX will not damage and accidental contact with the tympanum can adversely affect the penetration of fluid through the cause ototoxicity (Dukes 1992). Various allergic reac- tions to CHX have been described. Contact sensitivity to 10. Combination of NaOCl and CHX causes colour CHX was first reported by Calnan (1962). Today, CHX is changes and formation a precipitate, which may known to elicit allergic contact dermatitis, including interfere with the seal of the root filling.
connubial contact dermatitis, generally after prolonged 11. CHX can significantly improve the integrity of the and repeated application (Krautheim et al. 2004). It can hybrid layer andresin–dentine bond stability.
also cause contact urticaria, photosensitivity, fixed drug 12. The biocompatibility of CHX is acceptable.
eruption and occupational asthma. People at particular 13. In rare cases, CHX may cause allergic reactions.
risk of contact allergy (apart from medical and dentalstaff) are patients with leg ulcers and leg eczema (Krautheim et al. 2004). Overall, contact sensitivity toCHX seems to be rare as some large studies have shown a Agarwal S, Piesco NP, Peterson De et al. (1997) Effects of sensitization rate of about 2% (Osmundsen 1982, sanguinarium, chlorhexidine and tetracycline on neutrophil Bechgaard et al. 1985, Nomura et al. 1989). Even rarer viability and functions in vitro. Journal of Periodontal Research are reports of immediate anaphylactic reactions because of CHX. Ohtoshi et al. (1986) demonstrated IgE antibod- Almyroudi A, Mackenzie D, McHugh S, Saunders WP (2002) The effectiveness of various disinfectants used as endodontic ies in the sera of patients with anaphylaxis to CHX.
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