The inhibitory effects of different mouthwash brands on the zone of inhibition of. Micrococcus luteus and Escherichia coli

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1 The inhibitory effects of different mouthwash brands on the zone of inhibition of Micrococcus luteus and Escherichia coli George Kendros Bio 2290 Section 001 TM Gray

2 Abstract The use of mouthwash has been shown to be an effective tool for maintaining oral health when used in combination with daily toothbrushing and flossing. Mouthwash functions as an antibacterial, with cetylpyridinium chloride (CPC), essential oils (EO) and sodium fluoride being the active ingredients found in common household mouthwash brands. Because mouthwashes vary in their active ingredients, this experiment will investigate the antibacterial effects of various mouthwash brands (Crest, Scope, Listerine and Exact) on Micrococcus luteus and Escherichia coli. Bacteria were first isolated and plated on growth medium. Wells were cut into plates, and each treatment was pipetted into the well, and were incubated for 48 hours. Our results show that most mouthwash brands (Listerine, Crest, and Scope) significantly increased the diameter of the zone of inhibition for both species. The study concluded that the use of mouthwashes containing either CPC or EO reduces oral bacteria loads. Key words: Antibacterial, mouthwash, CPC, Essential Oils, E. coli, M. luteus Introduction Oral hygiene is an essential component to one s overall health, and can help prevent gingivitis, cavities, plaque and other oral maladies (Axelsson and Lindhe, 1978). Today, the most common forms of oral hygiene are brushing with toothpaste and the occasional use of dental floss. Most agree that daily toothbrushing is adequate to prevent oral maladies, however most individuals cannot effectively remove the biofilm from their teeth (Cumming and Loe, 1973). It is this biofilm that covers teeth that is teeming with oral bacteria, and plays an important role in the advancement of oral maladies

3 (Sreenivasan et al. 2012). The prevalence of oral bacteria in combination with ineffective brushing techniques has led many to use another, widely-available oral hygiene product, mouthwash. Currently, there are a cohort of oral mouthwashes available on the market, with various prices and ingredients. Daily use of mouthwash has been shown to be effective in reducing the number of bacteria in the oral cavity (Albert-Kiszely et al. 2007). However, not all mouthwashes are equal, and their effectiveness is dependent on the active ingredients they contain, and how susceptible oral fauna are to these ingredients. Cetylpyridinium chloride (CPC) is a common active ingredient in mouthwash, and has been shown to use its positive charge to enter negatively charged bacterial membranes and disrupt cell growth, metabolism and cause the secretion of intracellular constituents (Albert-Kiszely et al. 2007; Sreenivasan et al. 2012). Other mouthwashes contain essential oils (EO), which can inhibit the aggregation of Gram-positive species and remove endotoxins from Gram-negative species (Albert-Kiszely et al. 2007). Other active ingredients such as sodium fluoride also possess antibacterial properties (Aminbadi et al. 2007). Overall, different ingredients have different effects on the various bacteria found in the mouth. Manufacturers of mouthwash claim that their products are the most effective antibacterials. With the saturation of mouthwash on the market, the objective of this present study is to determine which common, commercial mouthwashes (Crest Pro- Health, Scope, Exact, and Listerine) are the most effective antibacterials. Each mouthwash contains different ingredients, and their effectiveness can be measured by determining the effect they have on the zone of inhibition on bacterial species. The

4 antibacterial ability of mouthwash will be tested against Micrococcus luteus, a Grampositive bacteria that is commonly found in the mouth and the upper respiratory tract and Escherichia coli, a Gram-negative bacteria, which when ingested orally can cause disease. Due to the effectiveness of the active ingredients, all mouthwash brands should inhibit the growth of bacteria, and mouthwash should have a greater effect on Grampositive bacteria. Furthermore, more expensive mouthwash brands should exhibit higher antibacterial capability due to their increased price. Results The mean diameter of the zone of inhibition was significantly greater for M. luteus when exposed to the Crest and Scope treatments compared to the other treatments and the control (F =26.3, P <0.001). E. coli growth was significantly inhibited by Crest, Scope and Listerine compared to Exact and the control (F =28.4, P <0.001). Overall, both species of bacteria had wider zones of inhibition when exposed to all brands of mouthwash compared to the control, and M. luteus was more susceptible to mouthwash and E. coli (Figure 1).

5 Fig. 1 The effects of different mouthwash brands on the mean diameter (mm ± SE) of the zone of inhibition for Escherichia coli and Micrococcus luteus. Means followed by different letters within each variable are significantly different (p <0.05) according to Tukey s HSD test. Discussion The results from this study indicate that only Scope and Crest brands of mouthwash were effective antibacterials against both E. coli and M. Luteus. Listerine was also shown to be an effective antibacterial agent against E. coli, and not M. luteus. Exact brand was the only mouthwash shown not to be an effective antibacterial agent against either species. Studies conducted by Albert-Kiszely et al. in 2007 found no significant difference in antibacterial efficacy between essential oil containing mouthwash, such as Exact and Listerine, and mouthwashes containing the more potent CPC. Interestingly, only Listerine significantly decreased the zone of inhibition for E. Coli and not Exact, despite containing the same active essential ingredients. This may indicate that

6 antibacterial efficacy may not just be attributed to the active ingredients in mouthwash, but the other ingredients that may affect the mouthwash s ability to kill or inhibit bacterial growth. Crest and Scope exhibit the largest zone of inhibition diameter for both species, indicating that against the bacterial species tested, CPC is an effective antibacterial agent. Furthermore, both mouthwashes had a greater effect on M. luteus than E. coli, which can be attributed to its biology. Being a Gram-positive bacterium, it lacks an outer membrane which makes it more susceptible to the mode of action of CPC (Albert-Kiszely et al. 2007). This finding further supports the hypothesis that Gram-negative bacteria will be more affected by antibacterial agents compared to the Gram-positive E. coli. Two of the four mouthwash brands tested are in accord with the initial price prediction. Exact, the least expensive brand exhibited the least antibacterial activity, and was just as effective as the deionized water control. Listerine, the second least expensive brand tested and was only more effective than the control against E. Coli and just as effective as control treatment against M. luteus. In contrast to the price prediction, Scope the most expensive mouthwash brand did not possess significantly greater antibacterial capabilities than Crest, the second most expensive brand tested. Furthermore, Crest exhibited the widest diameter of the zone of inhibition for both species, further rules out the concept that cost is an indicator of antibacterial effectiveness. In conclusion, Crest and Listerine are the only brands that showed exceptional antibacterial activity against both strains of bacteria tested. Exact was the only mouthwash brands to be ineffective against E. coli and M. luteus. Listerine was only shown to be effective against E. coli. Furthermore, this study showed that Gram-positive

7 species are more susceptible to the antibacterial agents found in mouthwash than Grampositive species. CPC, the active ingredient in both Crest and Scope was the most potent mouthwash against bacterial species, suggesting that mouthwashes containing CPC are the more effective than those containing essential oils or other ingredients. Word Count: 1184 Literature Cited Albert-Kiszely A, Pjetursson BE, Salvi GE, Witt J, Hamilton A, Persson GR, Lang NP (2007) Comparison of the effects of cetylpyridinium chloride with an essential oil mouth rinse on dental plaque and gingivitis a six-month randomized controlled clinical trial. J Clin Periodontol 34: Aminabadi NA, Balaei E, Pouralibaba F (2007) The effect of 0.2% sodium fluoride mouthwash in prevention of dental caries according to the DMFT index. J Dent Res Dent Clin Dent Prospects 1: Axelsson P, Lindhe J (1978) Effect of controlled oral hygiene procedures on caries and periodontal disease in adults. J Clin Periodontol 5: Cumming BR, Loe H (1973) Consistency of plaque distribution in individuals without special home care instruction. J Periodontal Res 8: Sreenivasan PK, Haraszthy VI, Zambon JJ (2012) Antimicrobial efficacy of 0.05% cetylpyridinium chloride mouthrinses. Lett Appl Microbiol 56: 14 20

Citation for published version (APA): Otten, M. P. T. (2011). Oral Biofilm as a Reservoir for Antimicrobials Groningen: University of Groningen

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