Short‐term antimicrobial properties of mineral trioxide aggregate with incorporated silver‐zeolite

Abstract – The purpose of this in vitro study was to determine whether adding silver‐zeolite (SZ) to mineral trioxide aggregate (MTA) would enhance the antimicrobial activity of MTA against Staphylococcus aureus (ATCC #25923), Enterococcus faecalis (ATCC #29212), Escherichia coli (ATCC#25922), Pseudomonas aeruginosa (ATCC #27853), Candida albicans (ATCC #90028), Porphyromonas gingivalis (ATCC #33277), Actinomyces israelii (ATCC #12102), and Prevotella intermedia (ATCC# 15032). SZ was added at 0.2% and 2% mass fraction concentration to MTA powder. The control group was MTA powder with no SZ. The antimicrobial effect test was accomplished by placing freshly mixed MTA specimens on agar plates inoculated with microorganisms and comparing the zones of inhibition at 24, 48, and 72 h. The amounts of silver ion release from MTA specimens were measured with atomic absorption spectrophotometry at 10‐min, 24‐, 48‐, and 72‐h periods. The pH of MTA specimens was measured with a pH meter at 10‐min, 24‐, 48‐, and 72‐h periods. MTA with 2% and 0.2% SZ specimens showed inhibitory effects on some microorganisms at all time periods, whereas no antimicrobial activity showed for P. intermedia and A. israelii. MTA without SZ inhibited C. albicans, E. Coli, and P. intermedia. The highest silver release was detected in 2% SZ MTA at 24 h. The incorporation of SZ may enhance the antimicrobial activity of MTA.     

COMPARATIVE EVALUATION OF ANTIMICROBIAL ACTION OF MTA,CALCIUM HYDROXIDE AND PORTLAND CEMENT

The present study aimed to evaluate and compare the antimicrobial effect of MTA Dentsply, MTA Angelus, Calcium Hydroxide and Portland cement. Four reference bacterial strains were used: Pseudomonas aeruginosa, Escherichia coli, Bacteroides fragilis, and Enterococcus faecalis. Plates containing Mueller-Hinton agar supplemented with 5% sheep blood, hemin, and menadione were inoculated with the bacterial suspensions. Subsequently, wells were prepared and immediately filled with materials and incubated at 37°C for 48 hours under anaerobic conditions, except P. aeruginosa. The diameters of inhibition zones were measured, and data analyzed using ANOVA and the Tukey test with 1% level of significance. MTA Dentsply, MTA Angelus and Portland cement inhibited the growth of P. aeruginosa. Calcium Hydroxide was effective against P. aeruginosa and B. fragillis. Under anaerobic conditions, which may hamper the formation of reactive oxygen species, the materials failed to inhibit E. faecalis, and E. coli.     

Antimicrobial action of four herbal plants over mixed-species biofilms of Candida albicans with four different microorganisms

This study aimed to evaluate the antimicrobial effect of four herbal plants glycolic extracts over mixed-species biofilm composed of Candida albicans (C. albicans) and another pathogenic bacterium as alternative therapy to be investigated. Four plants extract of Pfaffia paniculata roots; Hamamelis virginiana leaf, Stryphnodendron barbatiman tree bark and Gymnema sylvestre stem and leaves were tested over multi-species biofilm of C. albicans (ATCC 18804) and Streptococcus mutans (ATCC 35688), Staphylococcus aureus (ATCC 6538), Enterococcus faecalis (ATCC 4083) or Pseudomonas aeruginosa (ATCC 15442) for 5 min and 24 h and colony forming units per millilitre was calculated. The data were analysed using Kruskal–Wallis with Dunn's test (p ≤ 0.05). All tested extracts showed antimicrobial action over the mixed-species biofilms after 24 h. Some extracts eliminated totally the biofilms. The glycolic extract of P. paniculata, H. virginiana, S. barbatiman and G. sylvestre are effective over mixed-species biofilms and may be indicated as endodontic irrigant or intracanal medication.     

Antigen I/II mediates interactions between Streptococcus mutans and Candida albicans

Streptococcus mutans and Candida albicans are frequently co‐isolated from dental plaque of children with early childhood caries (ECC) and are only rarely found in children without ECC, suggesting that these species interact in a manner that contributes to the pathogenesis of ECC. Previous studies have demonstrated that glucans produced by S. mutans are crucial for promoting the formation of biofilm and cariogenicity with C. albicans; however, it is unclear how non‐glucan S. mutans biofilm factors contribute to increased biofilm formation in the presence of C. albicans. In this study we examined the role of S. mutans antigen I/II in two‐species biofilms with C. albicans, and determined that antigen I/II is important for the incorporation of C. albicans into the two‐species biofilm and is also required for increased acid production. The interaction is independent of the proteins Als1 and Als3, which are known streptococcal receptors of C. albicans. Moreover, antigen I/II is required for the colonization of both S. mutans and C. albicans during co‐infection of Drosophila melanogaster in vivo. Taken together, these results demonstrate that antigen I/II mediates the increase of C. albicans numbers and acid production in the two‐species biofilm, representing new activities associated with this known S. mutans adhesin.     

Mineral trioxide aggregate enriched with iron disulfide nanostructures: an evaluation of their physical and biological properties

The purpose of this study was to characterize mineral trioxide aggregates (MTA) enriched with iron disulfide (FeS₂) nanostructures at different concentrations, and to investigate their storage modulus, radiopacity, setting time, pH, cytotoxicity, and antimicrobial activity. Iron disulfide nanostructures [with particle size of 0.357 ± 0.156 μm (mean ± SD)] at weight ratios of 0.2, 0.4, 0.6, 0.8, and 1.0 wt% were added to white MTA (wMTA). The radiopacity, rheological properties, setting time, and pH, as well as the cytotoxicity (assessed using the MTT assay) and antibacterial activity (assessed using the broth microdilution test) were determined for MTA/FeS₂ nanostructures. The nanostructures did not modify the radiopacity values of wMTA (~6 mm of aluminium); however, they reduced the setting time from 18.2 ± 3.20 min to 13.7 ± 1.8 min, and the storage modulus was indicative of a good stiffness. Whereas the wMTA/FeS₂ nanostructures did not induce cytotoxicity when in contact with human pulp cells (HPCs) and human gingival fibroblasts (HGFs), they showed bacteriostatic activity against Staphylococcus aureus, Escherichia coli, and Enterococcus faecalis. Adding FeS₂ nanostructures to MTA might be an option for improving the root canal sealing and antibacterial effects of wMTA in endodontic treatments.     

In Vitro Evaluation of Antimicrobial Efficacy of Iontophoresis against Enterococcus faecalis,Candida albicans,Pseudomonas aeruginosa and Bacillus subtilis

This in vitro study examined the antimicrobial efficacy of iontophoresis against Enterococcus faecalis, Candida albicans, Pseudomonas aeruginosa and Bacillus subtilis, all of which are often found in the root canals of teeth with refractory apical periodontitis. An experimental periapical lesion model was made from a transparent plastic root canal model. Iontophoresis was performed with Kantop Junior®; with (1) diammine silver fluoride solution (2) ammoniacal silver hydroxide solution and (3) iodine zinc iodide solution as the iontophoretic solutions. Iontophoresis using diammine silver fluoride solution eradicated all four microorganisms whereas iontophoresis using ammonium silver solution and iodine zinc iodide solution could not. The results of this study suggested that iontophoresis with diammine silver fluoride is an effective treatment for E. faecalis, C. albicans, P. aeruginosa and B. subtilis, all of which often found in the root canals of teeth with refractory apical periodontitis.     

The development and validation of a rapid genetic method for species identification and genotyping of medically important fungal pathogens using high‐resolution melting curve analysis

Accurate, rapid and economical fungal species identification has been a major aim in mycology. In this study, our goal was to examine the feasibility of a high‐resolution melting curve analysis (HRMA) of internal transcribed regions ITS1 and ITS2 in ribosomal DNA (rDNA) for a rapid, simple and inexpensive differentiation of eight clinically relevant Candida species (Candida albicans, Candida glabrata, Candida parapsilosis, Candida krusei, Candida tropicalis, Candida guilliermondii, Candida dubliniensis and Candida lusitaniae). In addition, for the first time, we tested the applicability of HRMA to classify C. albicans strains into four previously described genotypes (A, B, C and D) using a primer set that spans the transposable intron region of 25S of rDNA. Type and unknown clinical oral isolates were used in this study and the melting curve analysis was compared with both amplicons' sequencing and agarose gel electrophoresis analysis. Real‐time PCR and subsequent HRMA of the two described rDNA regions generated distinct melting curve profiles that were in accord with sequencing and gel electrophoresis analysis, highly reproducible, and characteristic of each of the eight Candida species and C. albicans genotypes. Moreover, results were obtained in 4 h and without the need for any post‐amplification handling, so reducing time and cost. Owing to its simplicity and speed, this technique is a good fit for genotypic analysis of hundreds of clinical strains in large epidemiological settings.     

Adherence of Candida albicans to silicone is promoted by the human salivary protein SPLUNC2/PSP/BPIFA2

Interactions between Candida albicans, saliva and saliva‐coated oral surfaces are initial events in the colonization of the oral cavity by this commensal yeast, which can cause oral diseases such as candidiasis and denture stomatitis. Candida albicans also colonizes silicone voice prostheses, and the microbial biofilm formed can impair valve function, necessitating frequent prosthesis replacement. We have previously shown that saliva promoted binding of C. albicans cells to silicone in vitro, and that the selective binding of specific salivary proteins to voice prosthesis silicone mediated attachment of C. albicans cells. The C. albicans cells adhered to a polypeptide (or polypeptides) of ~36 kDa eluted from saliva‐treated silicone. We show here that a protein of similar size was identified in replicate blots of the eluate from saliva‐treated silicone when the blots were probed with antibodies to human SPLUNC2, a salivary protein with reported microbial agglutination properties. In addition, SPLUNC2 was depleted from saliva that had been incubated with silicone coupons. To determine whether SPLUNC2 is a yeast‐binding protein, SPLUNC2 cDNA was expressed in Escherichia coli. Purified recombinant His‐tagged protein (SPLUNC2r) bound to silicone as demonstrated by immunoblot analysis of an eluate from SPLUNC2r‐treated silicone coupons and ³⁵S‐radiolabelled C. albicans cells adhered in a dose‐dependent manner to SPLUNC2r‐coated silicone. We conclude that SPLUNC2 binds to silicone and acts as a receptor for C. albicans adherence to, and subsequent colonization of, voice prosthesis silicone.     

Antibacterial effects of blackberry extract target periodontopathogens

Background and Objective: Antimicrobial agents provide valuable adjunctive therapy for the prevention and the control of oral diseases. Limitations in their prolonged use have stimulated the search for new, naturally occurring agents with more specific activity and fewer adverse effects. Here we sought to determine the antibacterial properties of blackberry extract (BBE) in vitro against oral bacterial commensals and periodontopathogens. Material and Methods: The effects of whole and fractionated BBE on the metabolism of 10 different oral bacteria were evaluated using the colorimetric water‐soluble tetrazolium‐1 assay. The bactericidal effects of whole BBE against Fusobacterium nucleatum were determined by quantitating the numbers of colony‐forming units (CFUs). Cytotoxicity was determined in oral epithelial (OKF6) cells. Results: BBE at 350–1400 μg/mL reduced the metabolic activity of Porphyromonas gingivalis, F. nucleatum and Streptococcus mutans. The reduced metabolic activity observed for F. nucleatum corresponded to a reduction in the numbers of CFUs following exposure to BBE for as little as 1 h, indicative of its bactericidal properties. An anthocyanin‐enriched fraction of BBE reduced the metabolic activity of F. nucleatum, but not of P. gingivalis or S. mutans, suggesting the contribution of species‐specific agents in the whole BBE. Oral epithelial cell viability was not reduced following exposure to whole BBE (2.24–1400 μg/mL) for ≤ 6 h. Conclusion: BBE alters the metabolic activity of oral periodontopathogens while demonstrating a minimal effect on commensals. The specific antibacterial properties of BBE shown in this study, along with its previously demonstrated anti‐inflammatory and antiviral properties, make this natural extract a promising target as an adjunct for prevention and/or complementary therapy of periodontal infections.     

Antimicrobial effectiveness of grape seed extract against Enterococcus faecalis biofilm: A Confocal Laser Scanning Microscopy analysis

This study evaluated the antimicrobial effectiveness of 6.5% Vitis vinifera grape seed extract (GSE) against Enterococcus faecalis biofilm using confocal laser scanning microscopy (CLSM). Saline solution (SS), 5.25% sodium hypochlorite (NaOCl) and 2% chlorhexidine (CHX) were used for comparison. Dentin discs were inoculated with E. faecalis strain establishing a 3‐week‐old biofilm. Discs (n = 10) were exposed to 5.25% NaOCl, 2% CHX, 6.5% GSE and SS (negative control) for 10 min. Discs were stained with the fluorescent LIVE/DEAD‐BacLight? dye and analysed using CLSM. The proportion of dead cells in biofilm was analysed using one‐way anova and Tukey tests (P < 0.05). A higher proportion of dead cells was found in GSE group compared with CHX and SS (P < 0.05). NaOCl group was associated with the highest proportion of dead cells (P < 0.05). GSE presented antimicrobial activity against E. faecalis; however, NaOCl was the most effective irrigant solution. GSE was more effective than CHX and SS.     

Tricalcium silicate cements: osteogenic and angiogenic responses of human bone marrow stem cells

Tricalcium silicate cements (TSCs) are used in endodontic procedures to promote wound healing and hard tissue formation. The aim of this study was to evaluate and compare the effect of commonly used TSCs [mineral trioxide aggregate (MTA), Biodentine, and TotalFill] on cellular metabolism and osteogenic/angiogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) in vitro. We tested the null hypothesis of no difference between MTA, Biodentine, and TotalFill in stem cell responses. Cells were subjected to eluates of the tested materials for up to 14 d. Cell viability was evaluated using the 3‐(4,5‐dimethyl‐thiazoyl)‐2,5‐diphenyl‐tetrazolium bromide (MTT) assay. Real‐time PCR was used to determine the levels of expression of the osteogenic factors alkaline phosphatase (ALP), osteoprotegerin (OPG), osteocalcin (OC), and collagen 1A (COL1A1), and the angiogenic factors vascular endothelial growth factor A (VEGFA) and fibroblast growth factor 1 (FGF1). ELISAs were used to measure the levels of VEGFA and ALP in culture supernatants. Mineralization in vitro of hBMSCs was assessed using Alizarin Red staining. The hBMSCs tolerated exposure to TSCs well, with Biodentine showing the most favorable effect on cell viability. Expression of ALP, COL1A1, OPG, and VEGFA were differentially affected by the materials, with Biodentine and TotalFill inducing earlier changes at gene level. Increased mineralization was observed with time, after exposure to all TSCs tested, with MTA showing the greatest effect. The results revealed different responses of hBMSCs to TSCs in vitro.     

Bioactivity evaluation of calcium silicate‐based endodontic materials used for apexification

This study aims to compare the bioactivity of Biodentine, ProRoot MTA and NeoMTA Plus with regard to their element uptake (Ca, Si and Ca/P) by root canal dentine in a simulated apex (n = 30 each) and evaluate the correlation between the dentine fracture resistance (n = 30 each) and interfacial layer thickness. Specimens immersed in a corrected simulated body solution (c‐SBF) for 1, 30 and 90 days were used. In all test materials, the Ca and Si concentrations in the root dentine were found to be significantly higher, whereas the Ca/P and Si concentrations increased over time (P < 0.05). The dentine fracture resistance showed a difference at only day 30. The dentine fracture resistance of Biodentine and ProRoot MTA was positively correlated with the Si and Ca/P values, and the mean interfacial layer thickness of all specimens. A high biomineralisation capacity of ProRoot MTA and Biodentine, and their positive effects on the dentine fracture resistance during the first 30 days suggest that they may present more advantages than NeoMTA Plus in apexification treatment.     

Induction of apoptosis in oral epithelial cells by Candida albicans

During infection, interactions between Candida albicans and oral epithelial cells result in oral epithelial cell death. This is clinically manifested by the development of oral mucosal ulcerations generally associated with discomfort. In vitro studies have shown that C. albicans induces early apoptotic alterations in oral epithelial cells; however, these studies have also shown that treatment of infected cells with caspase inhibitors does not prevent their death. The reasons for these contradictory results are unknown and it is still not clear if C. albicans stimulates oral epithelial signaling pathways that promote apoptotic cell death. Activation of specific death pathways in response to microbial organisms plays an essential role in modulating the pathogenesis of a variety of infectious diseases. The aim of this study was to (i) characterize C. albicans‐induced apoptotic morphological alterations in oral epithelial cells, and (ii) investigate the activation of apoptotic signaling pathways and expression of apoptotic genes during infection. Candida albicans induced early apoptotic changes in over 50% of oral epithelial cells. However, only 15% of those showed mid‐late apoptotic alterations. At the molecular level, C. albicans caused a loss of the mitochondrial transmembrane potential and translocation of mitochondrial cytochrome c. Caspase‐3/9 activities increased only during the first hours of infection. Moreover, poly[ADP ribose] polymerase 1 was cleaved into apoptotic and necrotic‐like fragments. Finally, five anti‐apoptotic genes were significantly upregulated and two pro‐apoptotic genes were downregulated during infection. Altogether, these findings indicate that epithelial apoptotic pathways are activated in response to C. albicans, but fail to progress and promote apoptotic cell death.     

Antifungal Activity of Endosequence Root Repair Material and Mineral Trioxide Aggregate

Introduction

The purpose of this study was to investigate the antifungal activity of Endosequence Root Repair Material (ERRM; Brasseler USA, Savannah, GA) as compared with mineral trioxide aggregate (MTA) using Candida albicans.

Methods

All materials were packed into sterilized intravenous tubing to obtain standardized samples and allowed to set for 3 or 24 hours and then exposed to a suspension of C. albicans for incubations of 24 or 48 hours. To analyze the mechanisms of the material's antifungal activity, additional samples of each test material were prepared in the same manner and allowed to set for 24 hours; these were then incubated in a culture medium for 24 hours. The pH of each conditioned media was measured before transferring to wells containing C. albicans. The development of biofilm was analyzed after 24 and 48 hours with 2,3-bis (2-methoxy-4-nitro-5-sulfophenyl)-5-([phenyl amino] carbonyl)-2H-tetrazolium hydroxide reduction assay.

Results

Materials in both experimental groups significantly limited biofilm formation at each interval (ie, 24 and 48 hours). After incubating for a 24-hour period in the presence of C. albicans, ERRM in both experimental groups showed a reduction in biofilm formation that was statistically significant in comparison with MTA. However, when set for 24 hours and incubated for 48 hours, gray MTA and white MTA showed a more substantial reduction in biofilm formation than comparable samples of ERRM. Cultured media conditioned with test materials showed statistically significant antifungal biofilm activity after 48 hours.

Conclusions

All materials tested have comparable antifungal biofilm activity. It appeared that changing the environment, such as the pH, contributed to this activity.     

Fermentative 2‐carbon metabolism produces carcinogenic levels of acetaldehyde in Candida albicans

Acetaldehyde is a carcinogenic product of alcohol fermentation and metabolism in microbes associated with cancers of the upper digestive tract. In yeast acetaldehyde is a by‐product of the pyruvate bypass that converts pyruvate into acetyl‐Coenzyme A (CoA) during fermentation. The aims of our study were: (i) to determine the levels of acetaldehyde produced by Candida albicans in the presence of glucose in low oxygen tension in vitro; (ii) to analyse the expression levels of genes involved in the pyruvate‐bypass and acetaldehyde production; and (iii) to analyse whether any correlations exist between acetaldehyde levels, alcohol dehydrogenase enzyme activity or expression of the genes involved in the pyruvate‐bypass. Candida albicans strains were isolated from patients with oral squamous cell carcinoma (n = 5), autoimmune polyendocrinopathy–candidiasis–ectodermal dystrophy (APECED) patients with chronic oral candidosis (n = 5), and control patients (n = 5). The acetaldehyde and ethanol production by these isolates grown under low oxygen tension in the presence of glucose was determined, and the expression of alcohol dehydrogenase (ADH1 and ADH2), pyruvate decarboxylase (PDC11), aldehyde dehydrogenase (ALD6) and acetyl‐CoA synthetase (ACS1 and ACS2) and Adh enzyme activity were analysed. The C. albicans isolates produced high levels of acetaldehyde from glucose under low oxygen tension. The acetaldehyde levels did not correlate with the expression of ADH1, ADH2 or PDC11 but correlated with the expression of down‐stream genes ALD6 and ACS1. Significant differences in the gene expressions were measured between strains isolated from different patient groups. Under low oxygen tension ALD6 and ACS1, instead of ADH1 or ADH2, appear the most reliable indicators of candidal acetaldehyde production from glucose.     

Effectiveness of ozone against endodontopathogenic microorganisms in a root canal biofilm model

Aim To assess the antimicrobial efficacy of aqueous (1.25–20 μg mL^?1) and gaseous ozone (1–53 g m^?3) as an alternative antiseptic against endodontic pathogens in suspension and a biofilm model. Methodology Enterococcus faecalis, Candida albicans, Peptostreptococcus micros and Pseudomonas aeruginosa were grown in planctonic culture or in mono‐species biofilms in root canals for 3 weeks. Cultures were exposed to ozone, sodium hypochlorite (NaOCl; 5.25%, 2.25%), chlorhexidine digluconate (CHX; 2%), hydrogen peroxide (H₂O₂; 3%) and phosphate buffered saline (control) for 1 min and the remaining colony forming units counted. Ozone gas was applied to the biofilms in two experimental settings, resembling canal areas either difficult (setting 1) or easy (setting 2) to reach. Time‐course experiments up to 10 min were included. To compare the tested samples, data were analysed by one‐way anova . Results Concentrations of gaseous ozone down to 1 g m^?3 almost and aqueous ozone down to 5 μg mL^?1 completely eliminated the suspended microorganisms as did NaOCl and CHX. Hydrogen peroxide and lower aqueous ozone concentrations were less effective. Aqueous and gaseous ozone were dose‐ and strain‐dependently effective against the biofilm microorganisms. Total elimination was achieved by high‐concentrated ozone gas (setting 2) and by NaOCl after 1 min or a lower gas concentration (4 g m^?3) after at least 2.5 min. High‐concentrated aqueous ozone (20 μg mL^?1) and CHX almost completely eliminated the biofilm cells, whilst H₂O₂ was less effective. Conclusion High‐concentrated gaseous and aqueous ozone was dose‐, strain‐ and time‐dependently effective against the tested microorganisms in suspension and the biofilm test model.     

The potential of reuterin derived from Indonesian strain of Lactobacillus reuteri against endodontic pathogen biofilms in vitro and ex vivo

ObjectivesDespite the use of common irrigating solution with antimicrobial properties, failed root canal treatment remains a significant problem in endodontics. In the present study, we examined the efficacy of reuterin derived from probiotic bacteria, Lactobacillus reuteri on the biofilms of major endodontic pathogens using ex vivo model of root canal infections.MethodsBiofilms of major endodontic pathogens namely Enteoroccus faecalis, Fusobacterim nucleatum, Porphyromonas gingivalis, and Candida albicans were formed on root canals of 60 human premolar tooth samples accordingly a standard protocol. Thereafter, teeth were treated with either 2.5 % NaOCl (positive control), various concentrations of reuterin (test-group) or sterilized-distilled water (negative control) in a time-dependent assay. The efficacy of irrigation was evaluated by a time-dependent assay at 5 min and 30 min after irrigation by colony-forming units assay. The findings were further confirmed by species-specific real-time PCR. Data were statistically analysed using one way ANOVA with a significance level of P < 0.05.ResultsReuterin isolated from L. reuteri was effective against E. faecalis, C. albicans, F. nucleatum, and P. gingivalis biofilms, with a concentration of 100 µg/mL being the most effective compared to the negative control (P < 0.05) and also showed similar efficacy when compared with NaOCl.ConclusionReuterin isolated from L. reuteri has ability to inhibit in vitro and ex-vivo biofilms of endodontic pathogens, namely E. faecalis, F. nucleatum, P. gingivalis, and C. albicans. Reuterin has potential as a root canal irrigating solution due to its antibiofilm activity. Further research is warranted to determine the potential of probiotic bacteriotherapy in root canal systems.     

Evaluation of the antimicrobial activities of chlorhexidine gluconate,sodium hypochlorite and octenidine hydrochloride in vitro

The objective of this study was to compare the antimicrobial activity of sodium hypochlorite (NaOCl), chlorhexidine gluconate (CHX) and octenidine hydrochloride (OCT) in different concentrations against endodontic pathogens in vitro. Agar diffusion procedure was used to determine the antimicrobial activity of the tested materials. Enterococcus faecalis, Candida albicans and the mixture of these were used for this study. In the agar diffusion test, 5.25% NaOCl exhibited better antimicrobial effect than the other concentrations of NaOCl for all strains. All concentrations of OCT were effective against C. albicans and E. faecalis. Some 0.2% CHX was ineffective on all microorganisms. Antibacterial effectiveness of all experimental solutions decreased on the mixture of all strains. Decreasing concentrations of NaOCl resulted in significantly reduced antimicrobial effect.