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Table of Contents
ORIGINAL ARTICLE
Year : 2020  |  Volume : 4  |  Issue : 3  |  Page : 105-109

Eugenia polyantha (Wight) infusion against oral microorganisms on toothbrushes


1 Department of Dental Health, Bandung Polytechnique of Health, Bandung, Indonesia
2 Kulliyyah of Dentistry, International Islamic University Malaysia, Kuantan, Malaysia

Date of Submission03-Aug-2020
Date of Decision24-Aug-2020
Date of Acceptance17-Sep-2020
Date of Web Publication17-Oct-2020

Correspondence Address:
Susi Sukmasari
Kulliyyah of Dentistry, International Islamic University Malaysia, Kuantan
Malaysia
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/SDJ.SDJ_38_20

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  Abstract 


Background: Toothbrushes could get contaminated with pathogenic bacteria from inside or outside the oral cavity. Eugenia polyantha (Wight) has been reported to exhibit antibacterial activity. Objectives: The objective was to assess the potential of E. polyantha infusion to inhibit the growth of oral microorganisms on used toothbrushes. Materials and Methods: Fresh E. polyantha leaves were washed, drained, sliced, boiled in distilled water, and filtered. The filtrate was diluted to 60%, 70%, 80%, 90%, and 100% concentrations with distilled water. Each participant (n = 4) brushed their teeth properly under supervision. The bacterial solution from the used toothbrushes was inoculated into tryptic soy broth (TSB) and incubated. Following this, the bacterial suspension in TSB was evenly inoculated on Muller–Hinton agar plates and incubated. Both the incubations were stored at 37°C for 24 h under aerobic conditions. The diameter of the inhibition zone around the disc containing E. polyantha was measured. Results: The mean diameters of the inhibition zones at 0%, 60%, 70%, 80%, 90%, and 100% concentrations of E. polyantha infusion were 6.10, 13.85, 14.47, 14.61, 15.18, and 15.57 mm, respectively. The inhibition zone at an E. polyantha concentration of 0% statistically significantly differed from those concentrations of 60%, 70%, 80%, 90%, and 100% (P < 0.05 for all). There was no significant difference of measurement growth zone among the different concentrations; there was no inhibition in the control plate. Conclusion: Our results revealed that E. polyantha infusion has the potential to inhibit the growth of oral microorganisms on used toothbrushes; however, this effect is not concentration dependent.

Keywords: Antibacterial activity, Eugenia polyantha (Wight), inhibition zone, toothbrushes


How to cite this article:
Nurjanah N, Herijulianti E, Putri MH, Sukmasari S. Eugenia polyantha (Wight) infusion against oral microorganisms on toothbrushes. Sci Dent J 2020;4:105-9

How to cite this URL:
Nurjanah N, Herijulianti E, Putri MH, Sukmasari S. Eugenia polyantha (Wight) infusion against oral microorganisms on toothbrushes. Sci Dent J [serial online] 2020 [cited 2020 Oct 25];4:105-9. Available from: https://www.scidentj.com/text.asp?2020/4/3/105/298445




  Background Top


Millions of microorganisms present in the oral cavity can cause oral and dental diseases.[1] These diseases can be prevented by different means, such as chemical or mechanical removal of the microorganisms by tooth brushing or gargling with an antiseptic mouthwash solution. Tooth brushing removes accumulated food remains from the teeth and plaque (which contains bacteria) from the tooth surface.[2] Used toothbrushes can get contaminated with potentially pathogenic bacteria present in dental plaque.[2]

Streptococcus mutans plays a role in plaque formation; after tooth brushing, S. mutans remains on used toothbrushes for more than 24 h.[3],[4],[5] The use of unclean water to rinse toothbrushes, inappropriate storage of toothbrushes, or handling of toothbrushes with dirty hands facilitate the attachment of bacteria present outside the oral environment to toothbrushes. Several methods can be used to avoid and reduce the bacterial contamination of toothbrushes. One of the methods involves flushing toothbrushes with clean flowing water or soaking them in an antiseptic or decontamination solution.[3],[6] Another method for the decontamination of toothbrushes is via ultraviolet sanitization and the use of solutions, such as chlorhexidine, garlic extract, green tea, and neem fruit extract.[3],[7],[8],[9],[10],[11],[12],[13],[14] However, some of these methods are relatively expensive and are not commonly used in rural areas.

In addition to the increasing awareness of the adverse effects associated with the use of chemical products in the health-care industry, there is an increasing awareness of the use of natural products as alternatives. Natural products are thought to be safer,[15],[16] inexpensive, relatively easy to find, and readily available/prepared at home, making them an interesting topic of research in the health-care industry. Southeast Asia is blessed with a treasure of natural resources. It is second only to Brazil in having the highest abundance and diversity of medicinal plants in the world. Traditional medicines have been used and developed in Southeast Asia. In Indonesia, the use of the traditional medicine jamu is backed by scientific research.[17]Eugenia polyantha (Wight) (E. polyantha is also a medicinal plant).

E. polyantha or E. lucidula Miq. (also known as Syzygium polyanthum [Wight] Walp.) is a tropical tree belonging to the kingdom Plantae, division Spermatophyta, subdivision Pinophyta, class Coniferopsida, order Myricales, family Myrtaceae, genus Eugenia, and species Polyanthum (Wight). It is known as Indonesian bayleaf or Indonesian laurel in Indonesia.[18] Moreover, it is known as ubar serai and meselangan in Sumatera; manting and salam in Jawa; and salam in Madura.[18]

E. polyantha extract has been reported to exhibit antibacterial activity against oral microorganisms, such as Staphylococcus aureus and S. mutans, and antifungal activity against spoilage fungi.[19],[20],[21],[22],[23],[24]E. polyantha leaf has long been known as a spice that can be used for traditional healing and is easy to find in the market. This leaf consists of tannins, flavonoids, saponins, and essential oils. Infusion is an easy method for preparing E. polyantha extract; it is practiced at home by locals.

Therefore, the present study aimed to assess the potential of E. polyantha infusion to inhibit the growth of microorganisms on used toothbrushes. The results of the study can be used as reference for the development of antimicrobial solutions for toothbrush decontamination.


  Materials and Methods Top


Ethical approval was obtained from the Ethical Committee of Bandung Polytechnic of Health prior to the study with approval number 07/KEPK/PE/V/2017.

Preparation of infusion

E. polyantha leaves were purchased from the traditional market of Bandung, Indonesia. The samples were identified and deposited in the Natural Products Laboratory, Institute of Technology, Bandung. Fresh E. polyantha leaves were washed, drained, and sliced. In total, 200 g of the sliced leaves were weighed and placed into a beaker. Following this, 200 mL of distilled water was added to the leaves, and the mixture was boiled at 90°C for 20 min. The infusion was cooled and then filtered. The filtrate was placed into a 200-mL beaker and diluted to 60%, 70%, 80%, and 90% concentrations. Sterile distilled water was used as the negative control.

Media and reagents

Tryptic soy broth (TSB) was used to grow the bacteria collected from used toothbrushes, while Mueller–Hinton agar (MHA) was used to test the efficacy of the herbal infusion against oral microorganisms. The microorganisms from toothbrushes were cultured for 24 h at 37°C under aerobic conditions.

Tooth brushing

The study participants (n = 4) were primary schoolchildren aged 9–12 years who had more than three teeth caries. We gave each participant a toothbrush and taught them how to brush their teeth properly. Then, after applying a toothpaste to each toothbrush, the participants brushed their teeth under supervision. The participants systematically brushed their teeth for 2 min, which consisted of 30 s of brushing of each dental region, namely the upper right, upper left, lower left, and posterior lower right regions (in this order). Following this, each toothbrush was washed under running tap water. The dry toothbrushes were stored in a clean and closed storage space until the next day. After brushing the teeth twice, the bacteria on the used toothbrushes were collected on the 2nd day.

Measurement of inhibition zones

Bacteria from the used toothbrushes were collected by soaking the toothbrushes in physiological NaCl solution. Then, 50 μL (1.5 × 10[8] colony-forming unit/mL) of the bacterial suspension was inoculated into TSB and incubated at 37°C for 24 h. The bacterial suspension in TSB was evenly inoculated on the surface of MHA plates using sterile cotton bud. Holes were made using a sterile tube with a diameter of 6 mm. Different concentration of E. polyantha infusion; 60%, 70%, 80%, 90% and 100% was dropped into each hole. The hole containing sterile distilled water was used as the negative control, whereas the hole containing sterile distilled water with 5-μg ciprofloxacin was used as the positive control. The wrapped plates were incubated at 37°C for 24 h, following which the diameter of the inhibition zones was measured. The antibacterial activity of different concentrations of E. polyantha infusion was compared by measuring the diameter of the bacteria-free area (clear area) around each hole in millimeters (mm). The diameter of the clear inhibition zones was measured from two different directions.

Data analysis

Normality and homogeneity were tested using Shapiro–Wilk and Levene tests, respectively. The data were analyzed using t-test and analysis of variance (ANOVA).


  Results Top


As shown in [Table 1], ANOVA revealed that F = 31.288 and P = 0.000. The diameters of the inhibition zones (mm) varied significantly between the five concentrations of E. polyantha infusion and the negative control (0%).
Table 1: Inhibition zone with Eugenia polyantha (Wight) infusion

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The mean diameters of the inhibition zones at 0%, 60%, 70%, 80%, 90%, and 100% concentrations of E. polyantha infusion as shown in the [Figure 1] were 6.10, 13.85, 14.47, 14.61, 15.18, and 15.57 mm, respectively. Therefore, as referred in [Figure 2], the mean diameters of the inhibition zones increased on increasing the infusion. The mean and standard deviation after four times' repetition of each concentration were 0% = 6.1000 ± 0.14142, 60% = 13.8500 ± 1.52630, 70% = 14.4688 ± 1.50491, 80% = 14.6063 ± 1.65433, 90% = 15.1813 ± 0.92610, and 100% = 15.5688 ± 1.26267. The inhibition zone at an E. polyantha concentration of 0% statistically significantly differed from those at E. polyantha concentrations of 60%, 70%, 80%, 90%, and 100% (P < 0.05 for all).
Figure 1: Measurement of inhibition zones of oral microorganisms with Eugenia polyantha (Wight) infusion

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Figure 2: Mean diameters of inhibition zones with different concentrations of Eugenia polyantha (Wight) infusion

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  Discussion Top


E. polyantha leaves are primarily used as a spice or for fragrance in several countries in Southeast Asia. These leaves have also been used in traditional medicine. Several studies have mentioned that these leaves have many benefits for the treatment of various diseases, such as diabetes, hypertension, hypercholesterolemia, gastritis, diarrhea, hyperuricemia, eczema, scabies, and urticaria.[18] However, the present study is the first to test the potential of E. polyantha infusion to inhibit the growth of oral microorganisms on used toothbrushes.

Tooth brushing is an oral hygiene practice aimed at cleaning the oral cavity by removing plaque in order to prevent caries and periodontal disease. A previous study reported that regular toothbrushes can be contaminated with microorganisms from the oral cavity.[25] In addition, toothbrushes can be contaminated with microorganisms from the surrounding environment outside the oral cavity. The humidity in bathrooms can facilitate the growth of bacteria, particularly through aerosols from the toilet, rinsing water, contaminated fingers, and commensals on the skin. In a previous study, S. mutans, Candida albicans, Lactobacillus, Klebsiella, Streptococcus pyogenes, Streptococcus aureus, and Pseudomonas were detected on toothbrushes placed in the bathroom, while Escherichia coli was detected on toothbrushes placed adjacent to the toilet.[26] Our findings are in line with these findings; we detected microorganisms that exhibit growth on used toothbrushes.

E. polyantha contains active substances that have effective antibacterial/antimicrobial potential. Based on the existing data, we aimed to assess the potential of E. polyantha to inhibit the growth of bacterial colonies on used toothbrushes. The active substances found in E. polyantha include tannins, flavonoids, and 0.05% essential oils consisting of eugenol.[23] Phenols in essential oils exert toxicity by denaturing proteins in the bacterial wall and forming a tertiary protein structure with nonspecific bonds or disulfide bonds. Eugenol as an essential oil of E. polyantha has anesthetic and antiseptic properties.[27]

Infusion is a general term used to describe the method of soaking plant material in liquids in order to impart its flavor to the liquid medium. This method can easily be performed at home. Infusions can be prepared from a wide range of ingredients. The present study aimed to assess the potential of E. polyantha infusion to inhibit the growth of bacterial colonies on used toothbrushes. E. polyantha infusion was found to significantly inhibit the growth of bacterial colonies derived from used toothbrushes. This finding is in accordance with the finding of Adrianto, who reported the antibacterial activity of E. polyantha extract in a toothpaste formulation against S. mutans. However, specific oral microorganisms were not tested in the present study.[28]

Our findings clearly prove that E. polyantha infusion has potential antimicrobial effects against oral microorganisms. The mean inhibition zone in the E. polyantha-treated groups was larger than that in the positive control group. The diameter of the inhibition zones of bacterial colonies was found to increase with an increase in the infusion concentration. However, there was no significant difference among the different concentrations of E. polyantha infusion. This can be attributed to the diversity of bacteria present in the plaque, periodontal pocket, or saliva, such as Porphyromonas gingivalis, Aggregatibacter actinomycetemcomitans, and Treponema denticola.[29] Different microorganisms have different properties and resistance. Gram-positive (+) and Gram-negative (−) bacteria have different susceptibilities to antimicrobial agents at different concentrations because of the difference in the permeability of their cell walls.[30] Therefore, the minimal inhibitory concentrations of the isolated oral microorganisms need to be tested for further research.


  Conclusion Top


The present study revealed that E. polyantha (Wight) infusion has the potential to inhibit the growth of oral microorganisms on used toothbrushes. However, this effect is not concentration dependent.

Acknowledgments

We offer our sincere thanks to the Director of Health Polytechnic, Ministry of Health Bandung, for their excellent technical assistance and support throughout this research.

Financial support and sponsorship

Riset Pembinaan Tenaga Kesehatan Politeknik Bandung.

Conflicts of interest

There are no conflicts of interest.



 
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