11 Research Report Dental Journal (Majalah Kedokteran Gigi) 2017 March; 50(1): 1–5 Rinsing effect of 60% bay leaf (Syzygium polyanthum wight) aqueous decoction on the accumulation of dental plaque during fixed orthodontic treatment Fitria Avriliyanti, Sri Suparwitri, and Ananto Ali Alhasyimi Department of Orthodontics Faculty of Dentistry, Universitas Gadjah Mada Yogyakarta - Indonesia abstract Background: Fixed orthodontic appliance patients have a high risk to increasing plaque accumulation in tooth surface due to the presence of fixed appliance components. Using mouthwash with antibacterial agent from herbal material can control dental plaque accumulation. Bay leaf (BL) contains active compounds such as essential oil, tannin and flavonoid that have been known as an antibacterial agent. Purpose: The purpose of this study was to determine the effect of rinsing with 60% of BL aqueous decoction to the accumulation of dental plaque in fixed orthodontic appliance patients. Method: This research was an experimental clinical research with pretest and post-test control group design conducted on 20 subjects with age ranged between 18-25 years old. All subjects were undergoing the last stage (finishing) of fixed active orthodontic treatment. The subjects were instructed to rinse with 60% of BL aqueous decoction and 0.2% chlorhexidine as a control. Wash out period that needed between rinsing with 60% of BL aqueous decoction and rinsing with 0.2% chlorhexidine was 7 days. Each mouthwash used routinely for 7 days with same duration and intensity. Plaque scoring was measured before and after each treatment using Orthodontic Plaque Index (OPI). Result: One-way Anova test showed that there was a significant difference in the plaque index before and after treatment between the group of rinsing with 60% of BL aqueous decoction and group of rinsing with chlorhexidine (p<0.05). Conclusion: Rinsing with 60% BL aqueous decoction can decrease the accumulation of dental plaque in fixed orthodontic appliance patients. Keywords: fixed ortodontic appliance; plaque accumulation; 60% bay leaf aqueous decoction Correspondence: Ananto Ali Alhasyimi, Department of Orthodontics, Faculty of Dentistry, Universitas Gadjah Mada. Jl. Denta Sekip Utara, Bulaksumur, Yogyakarta 55281, Indonesia. E-mail: anantoali@ugm.ac.id introduction Orthodontic treatment used to be considered cosmetic, but nowadays, dentists and patients alike realize that orthodontic treatment may be a necessity. There are two kinds of orthodontic appliances; removable and fixed appliances. Fixed orthodontic appliance has more complicated design compared to removable appliance which makes cleaning procedure more difficult and increasing the risk of developing plaque retention.1,2 Accumulation of plaque can be found on fixed orthodontic appliance such as the bracket, wire, interface between bonding material and bracket. An orthodontic bracket may create difficulties to maintain good oral hygiene, leading to plaque accumulation and increase of enamel demineralization. Wire that is used for a long time during orthodontic treatment tends to plaque accumulation which can increase the level of microorganism in oral cavity. The roughness of composite surface as bonding material for orthodontic bracket also can be a predisposition factor for the attachment and growth of intraoral microorganisms.3,4 Plaque accumulation that is not properly cleaned during fixed orthodontic treatment may result a bad oral hygiene which leads to various oral diseases and failure of orthodontic treatment. Negligence in maintaining good oral hygiene will result in several negative effects, such as the destruction of periodontal Dental Journal (Majalah Kedokteran Gigi) p-ISSN: 1978-3728; e-ISSN: 2442-9740. Accredited No. 56/DIKTI/Kep./2012. Open access under CC-BY-SA license. Available at http://e-journal.unair.ac.id/index.php/MKG DOI: 10.20473/j.djmkg.v50.i1.p1-5 http://dx.doi.org/10.20473/j.djmkg.v50.i1.p1-5 2 Avriliyanti, et al./Dent. J. (Majalah Kedokteran Gigi) 2017 March; 50(1): 1–5 tissue (gingivitis, periodontitis), caries, halitosis and affecting the length of orthodontic treatment time.5 Around 5-10% of patients failure in fixed orthodontic treatments are caused by poor oral hygiene.6 One of the indicators for dental and mouth hygiene is dental plaque. Plaque control is an attempt to prevent the formation of plaque on the tooth surface. Plaque control can be performed mechanically or chemically. Mechanical plaque control includes tooth brushing and flossing, while the use of mouthwash is an example of chemical plaque control.7 Mouthwash is a type of plaque control that is easily accessible dan practical to use. This solution is a necessity for most people because it is believed to prevent the formation of dental plaque and occurrence of gingivitis mechanically and chemically. Rinsing using mouthwash can help the solution to reach more areas of teeth and intraoral surfaces, especially the interproximal areas which are not accessible mechanically.8 The example of mouthwash that is easily accessible in the market is chlorhexidine (CHX). CHX has been proven to decrease the accumulation of plaque due to its broad spectrum antimicrobial, bactericidal, and bacteriostatic properties to any kind of microorganisms, including bacteria, fungi, and virus.9 CHX is still the gold standard for its antimicrobial action and high substantiveness, but side effects in long- term used such as tooth and restoration staining, soft tissue staining, increased calculus deposition, unpleasant taste, taste alteration, burning sensation, desquamation and mucosal irritation limit its continued use. CHX can also lead oral discomfort in some of patients with chemotherapy- induced mucositis, xerostomia or ulcerative oral mucosal conditions.8,10 Nowadays, natural materials are being used and developed as more safe and cheaper alternatives with lesser side effects compared to chemical based products.11 A type of traditional herbal plant that is commonly seen in Indonesia is bay leaf (Syzygium polyanthum weight). Bay leaf (BL) that is also used as cooking spice, has many pharmacological activities that are useful in the field of dentistry. The chemical properties of BL consist of tannins, flavonoid and essential oils (0.05%), including citric acid and eugenol.12 Tannin is an active compound that has antibacterial activity. Tannin can inhibit the growth of bacteria by reacting with protein due to solidification on cell protein of bacterium (occurrence of protein denaturation). Flavonoid has biological and pharmacologic activities, including antibacterial activity.13 Essential oil also can inhibit the growth of some bacteria. BL is very effective against several bacteria such as Salmonella enterica and E. coli.14,15 The antibacterial activity of BL can be also due compounds non-flavonoid origin. The high contents of eugenol, methyl eugenol and fatty acid methyl esters together with other active components could contribute to its overall antibacterial activity.16,17 This research was carried out to investigate the effect of rinsing with 60% of BL aqueous decoction to the decreasing accumulation of dental plaque in fixed orthodontic appliance patients. materials and methods This research was a clinical experimental with pretest and post-test control group design conducted on 20 subjects with age ranged between 18-25 years old. The study was approved by The Ethics Committee of Faculty of Dentistry, Universitas Gadjah Mada with number 00789/KKEP/FKG- UGM/EC/2016. A total of 100 grams of fresh bay leaves were minced and boiled in 200 ml of boiling water until 100 ml of decoction was left. The concentration of the decoction was 100%. It was separated from the minced leaves and left to chill in room temperature (25O C). The decoction was filtered afterward with a 0.45 µ pore (millipore) diameter filter. Sixty percent of BL decoction was obtained by adding 40 ml of aquades into 60 ml of 100% BL decoction. All eligible subjects were given oral and written informations about the products and the purpose of the study and were asked to sign an informed consent prior to the studied procedures. All subjects were in the last stage (finishing) of fixed active orthodontic treatment. Exclusion criteria were considered: pregnancy, systemic disorders, patients under preventive treatment, and absence of studied evaluated teeth. Subjects were instructed to rinse their mouths with 60% BL aqueous decoction and 0.2% CHX mouth rinse as a control. The time interval between mouth rinsing with 60% BL aqueous decoction and 0.2% CHX mouth rinse was 7 days. Each mouthwash was routinely used for 7 days with the intensity of 2 times a day (after tooth brushing in the morning and at night) for 30 seconds strongly, using 10 ml of the solution in each rinsing. After a week of washout period, each group was instructed to use the opposite mouthwash for 7 days. The subjects in the study were still instructed to brushing (with bass technique) using pumice toothpaste for seven days of treatment. Plaque measurement was performed before and after application each mouth rinse using the Orthodontic Plaque Index (OPI) method. The OPI developed by Declerk in 1989, is a special index for patients with fixed orthodontic appliances. OPI has a higher diagnosis performance and accuracy compared to Quigley and Hein Index, and Modified Navy Plaque Index.18,19 In this study, we used disclosing agents that were applied on vestibular surfaces of teeth with orthodontic brackets and then the presence of dyed plaque was evaluated with the Yes-No system in 3 areas of vestibular surfaces of a tooth (Figure 1). All measurements were carried out under the same conditions and were performed by two calibrated examiners who were blinded to the applied regimen. The examiners performed their analysis with a satisfactory intraexaminer and interexaminer reliability Kappa index value (0.75). Each area has its own level of difficulty depending on the accessibility for cleaning (occlusal area = 1 = easily accessible; cervical area = 2 = accessible with certain difficulty; central area = 3 = poorly accessible). The values found are entered into a work table (Table 1).20 Dental Journal (Majalah Kedokteran Gigi) p-ISSN: 1978-3728; e-ISSN: 2442-9740. Accredited No. 56/DIKTI/Kep./2012. Open access under CC-BY-SA license. Available at http://e-journal.unair.ac.id/index.php/MKG DOI: 10.20473/j.djmkg.v50.i1.p1-5 http://dx.doi.org/10.20473/j.djmkg.v50.i1.p1-5 33Avriliyanti, et al./Dent. J. (Majalah Kedokteran Gigi) 2017 March; 50(1): 1–5 Resulting index is obtained by means of the formula:20 OPI % = sum of values from dyed areas x 100% total number of teeth x 6 The condition of oral hygiene then evaluated according to the following scheme: 0-30%= excellent hygiene; 30-50%= average hygiene; over 50% = insufficient hygiene.20 4 decoction and 0.2% of CHX mouth rinse as a control. The time interval between mouth rinsing with 60% BL aqueous decoction and 0.2% CHX mouth rinse was 7 days. Each mouthwash was routinely used for 7 days with the intensity of 2 times a day (after tooth brushing in the morning and at night) for 30 seconds strongly, using 10 ml of the solution in each rinsing. After a week of washout period, each group was instructed to use the opposite mouthwash for 7 days. The subjects in the study were still instructed to brushing (with bass technique) using pumice toothpaste for seven days of treatment. Plaque measurement was performed before and after application each mouth rinse using the Orthodontic Plaque Index (OPI) method. The Orthodontic Plaque Index developed by Declerk in 1989, is a special index for patients with fixed orthodontic appliances. Orthodontic Plaque Index has a higher diagnosis performance and accuracy compared to Quigley and Hein Index, and Modified Navy Plaque Index.18,19 In this study, we used disclosing agents that were applied on vestibular surfaces of teeth with orthodontic brackets and then the presence of dyed plaque was evaluated with the Yes-No system in 3 areas of vestibular surfaces of a tooth (Figure 1). All measurements were carried out under the same conditions and were performed by two calibrated examiners who were blinded to the applied regimen. The examiners performed their analysis with a satisfactory intraexaminer and interexaminer reliability Kappa index value (0.75). Each area has its own level of difficulty depending on the accessibility for cleaning (occlusal area = 1 = easily accessible; cervical area = 2 = accessible with certain difficulty; central area = 3 = poorly accessible). The values found are entered into a work table (Table 1).20 Figure 1. Scheme distribution of vestibular surface for evaluation of OPI: I = occlusal area=easily accessible; cervical area = 2 = accessible with certain difficulty; central area = 3 = poorly accessible21 Figure 1. Scheme distribution of vestibular surface for evaluation of OPI: I = occlusal area=easily accessible; cervical area = 2 = accessible with certain difficulty; central area = 3 = poorly accessible21 Table 1. Orthodontic Plaque Index: recorded dental plaque in individual areas multiplied by the relevant factor of cleaning difficulty20 Cervical 2× Total Central 3× Total Occlusal 1× Total Teeth 5 4 3 2 1 1 2 3 4 5 Occlusal 1× Total Central 2× Total Cervical 3× Total 5 Table 1. Orthodontic Plaque Index: recorded dental plaque in individual areas multiplied by the relevant factor of cleaning difficulty20 Resulting index is obtained by means of the formula:20 The condition of oral hygiene then evaluated according to the following scheme: 0-30%= excellent hygiene; 30-50%= average hygiene; over 50% = insufficient hygiene.20 RESULTS The result showed decreasing of plaque index before and after rinsing with 60% of BL aqueous decoction as well as rinsing with chlorhexidine. Decreasing of plaque index on group using BL aqueous decoction 60% amounts 43.1% ± 4.02%, while other group rinsing with CL amounts 42.1% ± 4.3%. Figure 2 shows the coloured dental plaque after application of disclosing agent solution in patients rising with 60% of BL aqueous decoction and rinsing with 0.2% CL. Figure 2. Coloured dental plaque after application of disclosing agent solution in patients rinsing with 60% of bay leaf aqueous decoction (A) and rinsing with 0.2% CHX (B). 5 Table 1. Orthodontic Plaque Index: recorded dental plaque in individual areas multiplied by the relevant factor of cleaning difficulty20 Resulting index is obtained by means of the formula:20 The condition of oral hygiene then evaluated according to the following scheme: 0-30%= excellent hygiene; 30-50%= average hygiene; over 50% = insufficient hygiene.20 RESULTS The result showed decreasing of plaque index before and after rinsing with 60% of BL aqueous decoction as well as rinsing with chlorhexidine. Decreasing of plaque index on group using BL aqueous decoction 60% amounts 43.1% ± 4.02%, while other group rinsing with CL amounts 42.1% ± 4.3%. Figure 2 shows the coloured dental plaque after application of disclosing agent solution in patients rising with 60% of BL aqueous decoction and rinsing with 0.2% CL. Figure 2. Coloured dental plaque after application of disclosing agent solution in patients rinsing with 60% of bay leaf aqueous decoction (A) and rinsing with 0.2% CHX (B). Figure 2. Coloured dental plaque after application of disclosing agent solution in patients rinsing with 60% of bay leaf aqueous decoction (A) and rinsing with 0.2% CHX (B). results The result showed decreasing of plaque index before and after rinsing with 60% BL aqueous decoction as well as rinsing with CHX. Decreasing of plaque index on group using 60% BL aqueous decoction 60% amounts 43.1% ± 4.02%, while other group rinsing with CHX was 42.1% ± 4.3%. Figure 2 shows the coloured dental plaque after application of disclosing agent solution in patients rising with 60% of BL aqueous decoction and rinsing with 0.2% CHX. Normality test result by Shapiro-Wilk showed normal distribution of data (p>0.05), while homogeneity test by Levene’s Test showed all the data have a homogeneous variance (p>0.05). The result of normality test and homogeneity test showed analysis could be done by parametric test using One-way Anova. Table 2 showed the result of One-way Anova analysis obtained p-value p<0.05. The result showed that there were significant differences in plaque indexes before and after rinsing between treatment groups. Differences between treatment groups could be determined by post-hoc test using Least Significant Difference (LSD). The results of LSD test in Table 2 showed that there were significant differences in group before and after rinsing with BL, group after rinsing with BL and before rinsing with CHX, group before and A B Dental Journal (Majalah Kedokteran Gigi) p-ISSN: 1978-3728; e-ISSN: 2442-9740. Accredited No. 56/DIKTI/Kep./2012. Open access under CC-BY-SA license. Available at http://e-journal.unair.ac.id/index.php/MKG DOI: 10.20473/j.djmkg.v50.i1.p1-5 http://dx.doi.org/10.20473/j.djmkg.v50.i1.p1-5 4 Avriliyanti, et al./Dent. J. (Majalah Kedokteran Gigi) 2017 March; 50(1): 1–5 after group rinsing with CHX (p<0.05). Whereas in the group before rinsing with BL and before rinsing with CHX, group after rinsing with BL and after rinsing with CHX had no significant difference (p>0.05). discussion Based on the study there were decreasing of plaque indexes in groups rinsing with 60% BL aqueous decoction as well as groups rinsing with CHX in fixed orthodontic appliance patients. Percentage of plaque index reduction in groups gargling by 60% BL aqueous decoction was 43.1% ± 4.02%, while other group rinsing with CHX was 42.1% ± 4,3%. Both of groups belongs to average hygiene category (31-50%).20 The study used pumice as a toothpaste thus anti-bacterial agent only came from mouthwash (BL and CHX) so reduction of plaque index was not optimal and only achieved moderate level. The used of pumice toothpaste without detergent and abrasive materials aimed to minimalize a bias observation. Results of the study in each group showed rinsing with 60% BL aqueous decoction as well as rinsing with 0.2% CHX could decrease accumulation of dental plaque in fixed orthodontic appliance patients (p<0.05). Another study showed that chlorhexidine was a gold standard of mouthwash and proven to be broad-spectrum of anti-bacterial agent and also having bactericidal and bacteriostatic towards all of the type microbes, fungi and virus.22 In this study, BL potentially proved on reduction plaque index in fixed orthodontic appliance patients. It is supported by a previous study conducted by Sumono and Wulan12 that rinsing with BL aqueous decoction could decrease the colony number of Streptococcus sp. BL are rich in essential oils, flavonoids and tannins which have been known as antibacterial agent.23 The activity of essential oils can affect both the external envelope of the cell and the cytoplasm of bacteria. The specific mechanisms action of essential oils in decreasing bacterial accumulation is via hydrophobicity. The typical of essential oils allows them to partition in the lipids of the bacterial cell membrane and mitochondria and leading to leakage of its cell contents and responsible for the disruption of bacterial structures.24,25 Tannin also has antibacterial activity and it is related to the ability to inactivated adhesin bacterial cell, enzyme and interfered protein transport on cell layer so that bacterial metabolism becomes impaired.26 Tannin have the ability to reduce the attachment of bacteria by binding to proline-rich protein of the salivary pellicle or to the cell-surface lipoteichoic acid.27 Tannin can also inhibit growth and kill bacteria by reacting with the cell membrane, resulting in leakage of essential metabolites that inactivate the bacterial enzyme system.28 BL also contained flavonoid besides essential oil and tannin. Flavonoids are well-known plant compounds that have antibacterial property.29 Action mechanism of flavonoid as an antibacterial is to interfere motility of bacteria, synthesis of nucleic acid, damage fluidity of membrane therefore membrane fluidity of outer and inner layer will decrease and interfere energy metabolism of bacteria.30,31 Moreover, their mode of antimicrobial action may be related to their ability to inactivated microbial adhesion, enzymes and cell envelope transport proteins.32 The antibacterial activity of BL can be also due to compounds non-flavonoid origin. The high contents of eugenol, methyl eugenol and fatty acid methyl esters together with other active components could contribute to its overall antibacterial activity.16,17 Plaque index score in group before rinsing using BL with group before rinsing with CHX showed no significant differences (p>0.05), while plaque index score in group after rinsing using BL and group before rinsing with CHX showed significant differences (p<0.05). The result showed washout process period in this study was successful. Washout period aimed to ensure the effect of prior exposure of first mouthwash had been stopped before the second mouthwash applied.33 Plaque index score in group after rinsing with BL and after rinsing with CHX showed no significant difference (p>0.05), the means rinsing used 60% BL aqueous decoction have the same effect with chlorhexidine as gold standard mouthwash to decrease the accumulation of dental plaque. One of excellences rinsing with BL was no alteration in taste sensation, while rinsing with CHX raised bitter tastes, caused change of temporary sensation and burning sensation and also long-term use of CHX can be associated with local side effects such as impaired sense of taste, tooth staining and occasional irritation and desquamation of mucous membranes.8,10,34,35 Table 2. Descriptive statistics and results of the Anova and LSD tests comparing the plaque index in the 4 groups tested Group n Plaque index (%) Significance* p-value After BL Before CHX After CHX Before BL 10 56.4±3.05 p=0.001 0.001* 0.621 0.001* After BL 10 43.1±4.02 0.,001* 0.741 Before CHX 10 57.0±3.78 0.001* After CHX 10 42.7±4.30 Values are presented as mean ± standard deviation or p-value only. *by ANOVA, *Significant differences between groups (p < 0.05). ANOVA: Analysis of variance; BL: 60% Bay leaf aqueous decoction, CHX: 0.2% Chlorhexidine Dental Journal (Majalah Kedokteran Gigi) p-ISSN: 1978-3728; e-ISSN: 2442-9740. Accredited No. 56/DIKTI/Kep./2012. Open access under CC-BY-SA license. Available at http://e-journal.unair.ac.id/index.php/MKG DOI: 10.20473/j.djmkg.v50.i1.p1-5 http://dx.doi.org/10.20473/j.djmkg.v50.i1.p1-5 55Avriliyanti, et al./Dent. J. (Majalah Kedokteran Gigi) 2017 March; 50(1): 1–5 Finally, it can be concluded that rinsing with 60% bay leaf (Syzygium polyanthum wight) aqueous decoction can reduce the accumulation of dental plaque in fixed orthodontic appliance patients. references 1. Singh G. Fixed orthodontic appliances. Text book of ortodontics. 1st ed. New Delhi: Jaypee Publishers; 2007. p. 449. 2. Karadas M, Cantekin K, Celikoglu M. Effects of orthodontic treatment with a fixed appliance on the caries experience of patients with high and low risk of caries. J of Dental Sciences 2011; 6: 195- 99. 3. Lee HJ, Park HS, Kim KH, Kwon TY, Honge SH. Effect of garlic on bacterial biofilm formation on orthodontic wire. Angle Orthod 2011; 81(5): 895–900. 4. Sukontapatipark W, El-Agraudi MA, Seliseth NJ, Thunold K, Selvig KA. Bacterial colonization associated with fixed orthodontic appliances. A scanning electron microscopy study. Eur J of Orthod 2001; 23(5): 475-84. 5. Al-Jewaira T, Suri S, Tompson BD. Predictors of adolescent compliance with oral hygiene instr uctions dur ing two-a rch multibracket fixed orthodontic treatment. Angle Orthod 2011; 81(3): 525-31. 6. Mehra T, Nanda RS, Sinha PK. Orthodontists’ assessment and management of patient compliance. Angle Orthod 1998; 68: 115–22. 7. Carranza FA, Newman MG. Clinical periodontology. 9th ed. Philadelphia: WB Saunders; 2006. p. 76. 8. Camile SF, McIntosh L, McCullough MJ. Mouthwash. Aust Prescr 2009; 32: 162-4. 9. Fajriani, Andriani JN. Reduction of salivary Streptococcus mutans colonies in children after rinsing with 2.5% green tea solution. JDI 2014; 21(3): 79-84. 10. Eley BM, Manson JD. Periodontic. 5th edition. Saunders: Elsevier; 2004. p. 21-7, 209. 11. Alam MT, Parvez N, Sharma PK. FDA-Approved natural polymers for fast dissolving tablets. Journal of Pharmaceutics 2014; 952970: 1-6. 12. Sumono A, Wulan SDA. The use of bay leaf (Eugenia polyanthum Weight) in dentistry. Dent J (Majalah Kedokteran Gigi) 2008; 41(3): 147-50. 13. Lestari IKA, Nazip K, Estuningsih S. Test of effectiveness of antibacterial of ethanol extract of loranthus of tea (Scurulla atropurpurea BL danser) on the growth of Enterobacter sakazakii. International Conference on Food, Biological and Medical Sciences, Bangkok, Thailand: 2014. p. 10-15. 14. Santos FA, Rao VSN . Antiinflammatory and antinociceptive effects of 1.8-cineole a terpenoid oxide present in many plant essential oils. Phytotherapy Research 2000; 14: 240-4. 15. Friedman M, Henika PR and Mandrell RE. Bactericidal activities of plant essential oils and some of their isolated constituents against Campylobacter jejuni, Escherichia coli, Listeria monocytogenes, and Salmonella enterica. J Food Prot 2002; 65: 1545-60. 16. Marzouki H, Piras A, Marongiu B, Assunta R, Assunta M. Extraction and separation of volatile and fixed oils from berries of Laurus nobilis L. by Supercritical CO2 Dessì 3. Molecules 2008; 13: 1702- 11. 17. Ivanović J, Mišić D, Ristić M, Pešić O, Žižović I. Supercritical CO2 extract and essential oil of bay (Laurus nobilis L.) – chemical composition and antibacterial activity. J Serb Chem Soc 2010; 75(3): 395–404. 18. Paschos E, Bucher K, Huth CK, Crispin A, Wichelhaus A, Dietel T. Is there a need for orthodontic plaque indices? Diagnostic accuracy of four plaque indices. Clin Oral Invest 2014; 18(4): 1351-8. 19. Beberhold K, Sachse-Kulp A, Schwestka-Polly R, Hornecker E, Ziebolz D. The orthodontic plaque index: an oral hygiene index for patients with multibracket appliances. Orthodontics 2012; 13(1): 94-9. 20. Ticha R, Bohmova H. Influence of fixed orthodontic appliance on the level of patient’s oral hygiene. Orthodoncie 2005;14(4): 29-34. 21. Heintze SD, Jost-Brinkmann PG, Finke C, Miethke RR. Oral health for the orthodontic patient. Illinois: Quintessenz Publishing Co, Inc; 1999. p. 39. 22. Fajriani, Andriani JN. Reduction of salivary Streptococcus mutans colonies in children after rinsing with 2,5% green tea solution. JDI2014; 21(3): 79-84. 23. Said CM, Hussein K. Determination of the chemical and genetic differences of laurus collected from three different geographic and climatic areas in lebanon. European Scientific J 2014; 2: 412-9. 24. Nazzaro, Fratianni F, Martino LD, Coppola R, Feo VD. Effect of essential oils on pathogenic bacteria filomena. Pharmaceuticals 2013; 6: 1451-74. 25. Burt S. Essential oils: their antibacterial properties and potential applications in foods – a review. Int J Food Microbiol 2004; 94(3): 223–53. 26. Akiyama H, Fujii K, Yamasaki O, Oono T, Iwatsuki K. Antibacterial action of several ta nnins aga inst Staphylococcus aureus. J Antimicrob Chemother. 2011; 48(4): 487-91. 27. Islam B, Khan AN, Khan AU. Dental caries: from infection to prevention. Med Sci Monit 2007; 13 (11): 196-203. 28. Mailoa MN, Mahendradatta M, Laga A, Djide N. Antimicrobial activities of tannins extract from guava leaves (Psidium Guajava L) on pathogens microbial. Int J Sci Technol 2014; 3(1): 236-41. 29. Sabir A. In vitro antibacterial activity of flavonoids Trigona sp propolis against Streptococcus mutans. Dent J (Majalah Kedokteran Gigi) 2005; 38(3): 135–41. 30. Mirzoeva OK, Grishanin RN, Calder PC. Antimicrobial action of propolis and some of its components: the effects on growth, membrane potential, and motility of bacteria. Microbiol Res 1997; 152: 239-46. 31. Vasconcelos LCS, Sampaio FC, Sampaio MCC, Pereira MSV, Higino JS, Peixoto MHP. Minimum inhibitory concentration of adherence of Punica granatum Linn (Pomegranate) gel againts S. mutans, S. mitis, and C albicans. Braz Dent J 2006; 17(13): 223-7. 32. Alghazeer R, Elmansori A, Sidati M, Gammoudi F, Azwai S, Naas H, Garbaj A, Eldaghayes I. In vitro antibacterial activity of flavonoid extracts of two selected libyan algae against multi-drug resistant bacteria isolated from food products. JBM 2017; 5: 26-48. 33. Roberts AW, Dusetzina SB, Farley JF. Revisiting the washout period in the incident user study design: why 6–12 months may not be sufficient. J Comp Eff Res 2015; 4(1): 27–35. 34. Charles CH, Sharma NC, Galustians HJ, Qaqish J, McGuire JA, Vincent JW.Comparative efficacy of an antiseptic mouthrinse and an antiplaque or antigingivitis dentifrice. A six-month clinical trial. J Am Dent Assoc 2001;132: 670-5. 35. Graziani F, Gabriele M, D’Aiuto F, Suvan J, Tonelli M, Cei S. Dental plaque, gingival inflammation and tooth discolouration with different commercial–formulations of 0.2% chlorhexidine rinse: a double-blind randomized controlled clinical trial. Oral Health Prev Dent 2015; 13: 101-11. Dental Journal (Majalah Kedokteran Gigi) p-ISSN: 1978-3728; e-ISSN: 2442-9740. Accredited No. 56/DIKTI/Kep./2012. Open access under CC-BY-SA license. Available at http://e-journal.unair.ac.id/index.php/MKG DOI: 10.20473/j.djmkg.v50.i1.p1-5 http://dx.doi.org/10.20473/j.djmkg.v50.i1.p1-5