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 Table of Contents  
Year : 2021  |  Volume : 12  |  Issue : 3  |  Page : 139-143

Effect of Different Disinfecting Agents on Dental Impressions Contaminated with Candida albicans

1 Department of Microbiology, Medical Microbiology Research Center, Qazvin University of Medical Sciences, Qazvin, Iran
2 Department of Prosthodontics, Faculty of Dentistry, Qazvin University of Medical Sciences, Qazvin, Iran
3 Student Research Committee, Qazvin University of Medical Sciences, Qazvin, Iran
4 Department of Prosthodontics, Dental Caries Prevention Research Center, Qazvin University of Medical Sciences, Qazvin, Iran

Date of Submission16-Sep-2021
Date of Decision23-Sep-2021
Date of Acceptance30-Sep-2021
Date of Web Publication2-Nov-2021

Correspondence Address:
Shima Aalaei
Dental Caries Prevention Research Center, Qazvin University of Medical Sciences, Bahonar Blvd, Qazvin
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/denthyp.denthyp_10_21

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Introduction: This research was carried out to compare the effects of 0.525% and 5.25% NaOCl, 2% H2O2, and 16-mg/mL propolis on additional silicone impression materials contaminated with Candida albicans at 5- and 10-minute exposure times, to evaluate the hypotheses of this study that propolis can be used for disinfecting of dental impressions. Materials and methods: A total of 122 silicone impressions were prepared. Two samples were considered the negative controls, and the rest were contaminated with C. albicans fungal species and disinfected with 0.525% NaOCl, 5.25% NaOCl, 2% H2O2, 16-mg/mL propolis, and 96% ethyl alcohol for 5 and 10 minutes (10 samples for each interval in each group). The data were analyzed with Kruskal–Wallis and Mann–Whitney U tests in all the groups at 5- and 10-minute intervals. Results: All the disinfecting agents significantly decreased C. albicans colony counts at both intervals compared to the control groups (P = 0.00). The differences in the disinfecting effects were significant between the four study groups (P = 0.00). The most significant effects were related to NaOCl at both intervals and concentrations and 2% H2O2 at the 10-minute interval (P = 0.001), followed by 2% H2O2 at the 5-minute interval and propolis (P = 0.001). Concerning propolis (P = 0.001) and 2% H2O2 (P = 0.004), the effect of exposure time was significant. Conclusion: All the disinfecting agents in the present study can be used to disinfect impressions contaminated with C. albicans; however, concerning propolis, although it resulted in significant decreases in C. albicans colony counts, the elimination of the remaining colonies was ineffective.

Keywords: Candida albicans, disinfection, hydrogen peroxide, propolis, sodium hypochlorite

How to cite this article:
Aslanimehr M, Gholami F, Torbati S, Aalaei S. Effect of Different Disinfecting Agents on Dental Impressions Contaminated with Candida albicans. Dent Hypotheses 2021;12:139-43

How to cite this URL:
Aslanimehr M, Gholami F, Torbati S, Aalaei S. Effect of Different Disinfecting Agents on Dental Impressions Contaminated with Candida albicans. Dent Hypotheses [serial online] 2021 [cited 2021 Nov 29];12:139-43. Available from:

  Introduction Top

There is a risk of transmission of pathogenic microorganisms to dental laboratories through impressions. Several disinfecting agents have been introduced for disinfection of impressions, some of which are chemicals, which might affect some essential properties of impressions, including their dimensional stability. Recently, attempts have been made to use new methods and materials, such as hydrogen peroxide, ozone,[1] microwave radiation,[2],[3] and propolis, in dental procedures to eliminate microorganisms.

Propolis or bee glue is a resinous mixture that honey bees produce by mixing saliva and beeswax with exudate gathered from tree buds snap flows or botanical sources. It is used as a sealant for unwanted open space in beehive, reduces microbes, and waterproofs the hive.[4]

At present, the antimicrobial effects of propolis have been established. In this context, Arslan et al. evaluated the antimicrobial effects of propolis, BioPure MTAD (Dentsply Sirona, USA), 5.25% NaOCl, and 2% chlorhexidine on Enterococcus faecalis and Candida albicans.[5] The results showed that the antimicrobial effect of propolis on fungal species. Therefore, propolis has been used in various dental procedures, such as endodontics, preventive dentistry, periodontics, orthodontics, etc. However, to date, it has not been used for the disinfection of dental impressions. As it is easy to prepare this material, it is discarded as a waste material in many cases. If it is shown that it is useful for disinfecting dental impressions, its use will be cost-effective.

No studies to date have evaluated the disinfecting effect of propolis exclusively on impressions contaminated with fungal species. NaOCl, too, is used for comparison as a useful material[5] for disinfection. On the other hand, H2O2 is routinely used in dental clinics and dental schools to disinfect impressions. Therefore, the present study was undertaken to compare the antifungal effects of 0.525% NaOCl, 5.25% NaOCl, 2% H2O2, and propolis. As time is an essential factor for these agents’ efficacy, their effect was evaluated at 5- and 10-minute exposure times using the spray technique.

  Materials and Methods Top

This study was approved by the Ethics Committee of Qazvin University of Medical Sciences under the code IR.QUMS.REC.1394.477. There was no conflict with ethical considerations.

The sample size of the present in vitro study was estimated at 20 samples of additional silicone impression material in each group based on a previous study[6] and statistical calculations.

Each experimental and positive control group was subdivided into two groups (n = 10) to evaluate disinfecting agents’ properties at two exposure times: 5 and 10 minutes. Therefore, 122 samples were prepared in four groups for disinfecting agents and the control groups.

Group 1: 0.525% NaOCl

Group 2: 5.25% NaOCl

Group 3: 2% H2O2

Group 4: 16 mg/mL propolis

The positive control group for propolis: 96% ethyl alcohol

The positive control group: 20 samples

The negative control group: two samples

As propolis is an alcohol-based material and is a disinfecting agent, to avoid any errors, 20 samples were disinfected by 96% ethyl alcohol, and the effect of propolis on decreasing colony counts was compared with the ethyl alcohol group as a positive control at 5- and 10-minute exposure times. Concerning the positive control group, the samples were not disinfected after contamination, and the colonies were only counted at 5- and 10-minute intervals. In the negative control group, to confirm the impressions’ sterilization, no procedures were carried out, and sampling was carried out at 5- and 10-minute intervals.

Procedural steps

Preparation of additional silicone impression materials

In the present study, heavy- and light-body additional impression materials (Zhermack, DC, Germany) and quadrant metallic trays were used to prepare the samples. The trays were sterilized in an autoclave (121°C for 20 minutes) before the procedures. The impression materials were mixed according to the manufacturer’s instructions under sterile conditions and placed in the trays. Impressions were taken from dentures that had already been sterilized in an autoclave, using the two-step putty/wash technique.

Sterilization of the silicone impression materials

The prepared impressions were sterilized in an autoclave at 121°C for 20 minutes. Two samples were sterilized as negative controls and were not contaminated.

Contamination of silicone impressions with C. albicans

The sterlized silicone impressions were separately immersed and stored in sterile glass containers containing 150 mL of the microbial suspension at 0.5 McFarland concentration at 37°C for 60 minutes.[7]

After contamination, all the samples were rinsed with sterile distilled water for 30 seconds to simulate the irrigation procedure carried out by dentists. Then the samples were shaken gently to eliminate excess water.

Disinfection of impressions

To disinfect all the samples, the disinfecting agent was sprayed for 15 seconds (10 puffs in 15 seconds), and the impressions were kept in a closed sterile environment containing a sterile wet piece of cotton cloth to create a moist environment for 5 or 10 minutes.

Because of the accuracy of impressions, a ≤10-minute exposure has been recommended[8]; on the other hand, hypochlorite sodium can be effective in 5 minutes,[9],[10],[11] and the silicon impression material loaded lower microflora than irreversible hydrocolloid material; therefore, we used 5 and 10 minutes exposure time.[12]

After the disinfection periods, the samples were retrieved from the sterile closed environment and rinsed with sterile distilled water for 30 seconds. The samples were then shaken gently to eliminate the excess water and disinfecting agent to simulate the irrigation procedure carried out by dentists.

The impressions were transferred into sterile glass containers containing 150 mL of physiologic serum and vortexed for 1 minute, followed by a rest period of 1 minute. This procedure was repeated three to five times. A sampler was used to remove 25 μL of physiologic serum to culture on Sabouraud dextrose agar, followed by incubation at 37°C for 72 hours. C. albicans colony counts were then determined.

Statistical analysis

The data were analyzed with SPSS 24.0 (SPSS Inc., Chicago, IL, USA). The data were analyzed with the Kolmogorov–Smirnov test at a 95% confidence interval, and since they were not distributed normally (P < 0.05), the Kruskal–Wallis nonparametric test was used to analyze differences in mean colony counts among all the groups. In case of significant differences, the Mann–Whitney U test was used for two-by-two comparisons of the groups and two different exposure times. Statistical significance was set at P ≤ 0.05.

  Results Top

No fungal colonies were detected in the negative controls. [Table 1] presents the colony counts after disinfection at 5- and 10-minute exposure times. As summarized in the [Table 1], NaOCl at both concentrations and exposure times and H2O2 at 10-minute exposure time were significantly the most effective, and the colonies had almost completely been eliminated where the effects of them were significantly more than that of 2% H2O2 at 5-minute exposure time (P = 0.00). Propolis exhibited the minimum effect (P = 0.00) and its effect at 10-minute exposure time was higher than that at 5-minute exposure time (P = 0.001).
Table 1 The means and standard deviations of Candida albicans colony counts in all the study groups, and comparison of colony count decreases with the controls

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All the groups exhibited significant decreases in colony counts compared to the positive controls (P = 0.00). Propolis, too, resulted in a significant decrease in colony counts at both exposure times compared to the ethyl alcohol control group (P = 0.00).

Concerning propolis and 2% H2O2, an increase in exposure time resulted in a significant increase in the disinfecting effect (P = 0.001 and P = 0.004, respectively).

  Discussion Top

As dental impressions cannot be sterilized with heat, the use of chemical agents is the only technique of choice for the elimination of microorganisms in these impressions.[13] However, new materials and techniques, too, have been used which have seldom been more effective than NaOCl.[14] Propolis is one of the materials used for disinfection in recent years.[15] It affects the adherence of C. albicans to the surfaces like resin denture base surface[16]; therefore, it is believed that this disinfecting effect is influenced by surface characteristics. In this study, the disinfecting effect of propolis on dental impressions contaminated with C. albicans was compared with other disinfecting agents, as a novel idea. Since the Iranian propolis in alcohol has useful antiseptic activity,[15] this solution was used in the present study.

NaOCl is the most effective disinfecting agent against C. albicans, and it was used in the current study as a gold standard. Its 5.25% concentration in 5 minutes and its 0.525% concentration in 10 minutes eliminated all the C. albicans colonies. This disinfecting agent has some advantages, including low cost, significant efficacy, and the ability to disinfect tools and instruments,[17] with a rapid activity against a broad spectrum of microorganisms[18]; however, one of its problems is its high contact angle, resulting in low wettability.[1]

The significantly higher disinfecting efficacy of NaOCl has been confirmed in various studies.[9],[10],[19] In a study by Badrian et al., NaOCl was reported to be an effective antifungal agent and eliminated 90.62% of C. albicans colonies on alginate impressions after 5 minutes and 96.7% after 10 minutes with the use of the spray technique.[9] In studies by Azevedo et al.[10] and Jeyapalan et al.,[19] NaOCl was 100% effective in eliminating microorganisms.

Therefore, in most studies, NaOCl has been used for comparison, and in the majority of studies, its fungicidal effect on C. albicans has been close to 100%, consistent with the present study.

Based on the present study results, 2% H2O2 exhibited significantly less disinfecting efficacy against C. albicans compared to NaOCl, but it was more effective than propolis. H2O2 can produce nascent oxygen after its application, resulting in a strong antimicrobial activity by damaging the microbial cell and interfering with cell division.[20]

Azevedo et al. evaluated the antimicrobial effect of 1% and 5.25% NaOCl and 3% H2O2 on additional silicon impressions for 10 minutes and concluded that all the three solutions decreased the microbial load of the impressions >99.9%. They used addition silicone impressions using the immersion technique.[10] Concerning 5.25% NaOCl, the results were similar to the present study; however, concerning H2O2, the results were different; it should be noted that the immersion technique was used in that study, a higher concentration of H2O2 was used, and C. albicans was not specifically evaluated. Badrian et al. evaluated the effect of 0.525% NaOCl and Epimax (50% H2O2 + alcohol + organic acid) on alginate impressions contaminated with different microorganisms, including C. albicans, using the spray technique for 5 and 10 minutes.[9] The effect of the disinfecting agents was similar to the present study over time. However, the effect of Epimax at the two time intervals was similar to that of 0.525% NaOCl, which is different from the present study because a lower concentration of H2O2 was used in the present study, and Epamax contained alcohol and acid, too. The decrease in C. albicans colonies in 0.525% NaOCl at 5- and 10-minute intervals was 90.62% and 96.09%, respectively, close to the present study. Badrian et al. evaluated these materials’ effects on alginate impressions.[9]

Based on the results, propolis exhibited the least disinfecting activity significantly on additional silicone impression materials. However, it significantly reduced colony counts compared to the positive and ethyl alcohol control groups, and an increase in the exposure time increased its antifungal activity.

No study is available on the disinfecting effect of propolis on dental impressions. However, various studies are available on the treatment of candidiasis[21],[22] and periodontitis, in almost all of which propolis has exhibited proper fungicidal effects. For example, in a study by Siqueira et al.,[21] all the C. albicans samples in patients with chronic periodontitis were sensitive to red propolis. Martins et al.,[22] too, reported that propolis was effective in the treatment of resistant candidiasis in HIV patients. Yildirim-Bicer et al.[23] compared the antifungal effect of NaOCl- and propolis-containing mouthwash. The minimum antifungal effect was related to propolis, and 1% NaOCl exhibited a significantly higher antifungal effect. However, the difference between these two groups and the positive control group was significant.

Some of the propolis ingredients include aliphatic aldehydes, acid esters, carboxylic acids, cinnamic acid, and esters such as ketone, terpene, alcohol, hydrocarbons, phenolics, and isoflavone formononetin, each with antibacterial and antifungal activities.[24] The isoflavone formononetin acts as a fungicidal agent.[25]

Differences in the chemical structure of propolis samples and their flavonoid content might cause differences in their antifungal properties.[26],[27] However, a study by Shokri et al. showed that some Candida species are not affected by Iranian propolis.[28]

Carvalho et al., in a review literature study, analyzed the evidence of the use of two types of propolis in dentistry. In their study, propolis exhibited antifungal properties, but there were differences between the different kinds of propolis because of differences in their chemical compositions.[29]

As this was the first time propolis was used to disinfect dental impressions, it was designed in vitro so that the results would serve as basic data for subsequent clinical studies.

Propolis affects the adhesion and cell wall characteristics of C. albicans.[16] Therefore, the properties of the contaminated surfaces affect the antifungal activity of propolis. This material has never been used to disinfect dental impressions. Therefore, it was tested in the present study.

  Conclusion Top

Under the limitations of the present study, propolis can be used as a disinfectant agent for dental impressions, especially at 10-minute exposure. However, its antifungal effect was lower than 5.25% NaOCl, 0.525% NaOCl, and 2% H2O2. An increase in exposure time from 5 to 10 minutes increased the disinfecting efficacy of 2% H2O2 and propolis.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

  References Top

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