Skip to main content

Advertisement

SpringerLink
Log in

In situ antimicrobial activity on oral biofilm: essential oils vs. 0.2 % chlorhexidine

  • Original Article
  • Published:
Clinical Oral Investigations Aims and scope Submit manuscript

Abstract

Objectives

This study aims to evaluate the in situ antibacterial activity of a mouthwash containing essential oils (M-EO) on undisturbed de novo plaque-like biofilm (PL-biofilm) up to 7 h after its application.

Patients and methods

An appliance was designed to hold six glass disks on the buccal sides of the lower teeth, allowing PL-biofilm growth. Fifteen healthy volunteers wore the appliance for 48 h and then performed a M-EO. Disks were removed after 30 s and at 1, 3, 5, and 7 h later. After a washout period, the same procedure was repeated with a M-WATER and a M-0.2 % chlorhexidine. After PL-biofilm vital staining, samples were analyzed using a confocal laser scanning microscope.

Results

At 30 s after M-EO, the levels of bacterial vitality were 1.18 %, significantly lower than that of the basal sample (p < 0.001). After 7 h, the antibacterial effect of essential oils was still patent with a 47.86 % difference in bacterial vitality compared to the basal sample (p < 0.001).

Conclusion

A single M-EO presents high antibacterial immediate activity and penetration capacity in situ and a substantivity which lasts for at least 7 h after its application over de novo biofilm. These results were better than those observed with 0.2 % chlorhexidine under the same conditions.

Clinical relevance

A single M-EO is an effective measure against the de novo biofilm, presenting a good alternative to clorhexidine such as a preoperative rinse, in periodontal procedures or post-treatment applications.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Vlachojannis C, Winsauer H, Chrubasik S (2012) Effectiveness and safety of a mouthwash containing essential oil ingredients. Phytother Res 27(5):685–691. doi:10.1002/ptr.4762

    Article  PubMed  Google Scholar 

  2. FDA (2003) Oral health care drug products for over-the-counter human use; antigingivitis/ antiplaque drug products; establishment of a monograph; proposed rules part III. Vol 21 CFR

  3. Sliepen I, Van Essche M, Quirynen M, Teughels W (2010) Effect of mouthrinses on Aggregatibacter actinomycetemcomitans biofilms in a hydrodynamic model. Clin Oral Investig 14(3):241–250. doi:10.1007/s00784-009-0286-0

    Article  PubMed  Google Scholar 

  4. Fine DH, Furgang D, Barnett ML (2001) Comparative antimicrobial activities of antiseptic mouthrinses against isogenic planktonic and biofilm forms of Actinobacillus actinomycetemcomitans. J Clin Periodontol 28(7):697–700. doi:10.1034/j.1600-051x.2001.028007697.x

    Article  PubMed  Google Scholar 

  5. Filoche SK, Coleman MJ, Angker L, Sissons CH (2007) A fluorescence assay to determine the viable biomass of microcosm dental plaque biofilms. J Microbiol Methods 69(3):489–496

    Article  PubMed  Google Scholar 

  6. Pan PC, Harper S, Ricci-Nittel D, Lux R, Shi W (2010) In-vitro evidence for efficacy of antimicrobial mouthrinses. J Dent 38(Suppl 1):S16–S20. doi:10.1016/S0300-5712(10)70006-3

    Article  PubMed Central  PubMed  Google Scholar 

  7. Auschill TM, Hellwig E, Sculean A, Hein N, Arweiler NB (2004) Impact of the intraoral location on the rate of biofilm growth. Clin Oral Investig 8(2):97–101. doi:10.1007/s00784-004-0255-6

    Article  PubMed  Google Scholar 

  8. Auschill TM, Hein N, Hellwig E, Follo M, Sculean A, Arweiler NB (2005) Effect of two antimicrobial agents on early in situ biofilm formation. J Clin Periodontol 32(2):147–152. doi:10.1111/j.1600-051X.2005.00650.x

    Article  PubMed  Google Scholar 

  9. Watson PS, Pontefract HA, Devine DA, Shore RC, Nattress BR, Kirkham J, Robinson C (2005) Penetration of fluoride into natural plaque biofilms. J Dent Res 84(5):451–455

    Article  PubMed  Google Scholar 

  10. Al-Ahmad A, Wunder A, Auschill TM, Follo M, Braun G, Hellwig E, Arweiler NB (2007) The in vivo dynamics of Streptococcus spp., Actinomyces naeslundii, Fusobacterium nucleatum and Veillonella spp. in dental plaque biofilm as analysed by five-colour multiplex fluorescence in situ hybridization. J Med Microbiol 56(Pt 5):681–687. doi:10.1099/jmm.0.47094-0

    Article  PubMed  Google Scholar 

  11. Hannig C, Hannig M (2009) The oral cavity—a key system to understand substratum-dependent bioadhesion on solid surfaces in man. Clin Oral Investig 13(2):123–139. doi:10.1007/s00784-008-0243-3

    Article  PubMed  Google Scholar 

  12. Pan P, Barnett ML, Coelho J, Brogdon C, Finnegan MB (2000) Determination of the in situ bactericidal activity of an essential oil mouthrinse using a vital stain method. J Clin Periodontol 27(4):256–261. doi:10.1034/j.1600-051x.2000.027004256.x

    Article  PubMed  Google Scholar 

  13. Charles CH, Pan PC, Sturdivant L, Vincent JW (2000) In vivo antimicrobial activity of an essential oil-containing mouthrinse on interproximal plaque bacteria. J Clin Dent 11(4):94–97

    PubMed  Google Scholar 

  14. Fine DH, Furgang D, Sinatra K, Charles C, McGuire A, Kumar LD (2005) In vivo antimicrobial effectiveness of an essential oil-containing mouth rinse 12 h after a single use and 14 days’ use. J Clin Periodontol 32(4):335–340. doi:10.1111/j.1600-051x.2005.00674.x

    Google Scholar 

  15. Dong WL, Zhou YH, Li CZ, Liu H, Shang SH, Pan BQ (2010) Establishment and application of an intact natural model of human dental plaque biofilm. Shanghai Kou Qiang Yi Xue 19(2):196–201

    PubMed  Google Scholar 

  16. Gosau M, Hahnel S, Schwarz F, Gerlach T, Reichert TE, Burgers R (2010) Effect of six different peri-implantitis disinfection methods on in vivo human oral biofilm. Clin Oral Implants Res 21(8):866–872. doi:10.1111/j.1600-0501.2009.01908.x

    PubMed  Google Scholar 

  17. Hannig C, Hannig M, Rehmer O, Braun G, Hellwig E, Al-Ahmad A (2007) Fluorescence microscopic visualization and quantification of initial bacterial colonization on enamel in situ. Arch Oral Biol 52(11):1048–1056

    Article  PubMed  Google Scholar 

  18. Wood SR, Kirkham J, Marsh PD, Shore RC, Nattress B, Robinson C (2000) Architecture of intact natural human plaque biofilms studied by confocal laser scanning microscopy. J Dent Res 79(1):21–27. doi:10.1177/00220345000790010201

    Article  PubMed  Google Scholar 

  19. Arweiler NB, Hellwig E, Sculean A, Hein N, Auschill TM (2004) Individual vitality pattern of in situ dental biofilms at different locations in the oral cavity. Caries Res 38(5):442–447. doi:10.1159/000079625

    Article  PubMed  Google Scholar 

  20. Wright SJ, Wright DJ (2002) Introduction to confocal microscopy. Methods Cell Biol 70:1–85

    Article  PubMed  Google Scholar 

  21. Hahnel S, Rosentritt M, Burgers R, Handel G (2008) Surface properties and in vitro Streptococcus mutans adhesion to dental resin polymers. J Mater Sci Mater Med 19(7):2619–2627. doi:10.1007/s10856-007-3352-7

    Article  PubMed  Google Scholar 

  22. Berney M, Hammes F, Bosshard F, Weilenmann HU, Egli T (2007) Assessment and interpretation of bacterial viability by using the LIVE/DEAD BacLight Kit in combination with flow cytometry. Appl Environ Microbiol 73(10):3283–3290. doi:10.1128/AEM.02750-06

    Article  PubMed Central  PubMed  Google Scholar 

  23. Arweiler NB, Lenz R, Sculean A, Al-Ahmad A, Hellwig E, Auschill TM (2008) Effect of food preservatives on in situ biofilm formation. Clin Oral Investig 12(3):203–208. doi:10.1007/s00784-008-0188-6

    Article  PubMed  Google Scholar 

  24. Netuschil L, Reich E, Unteregger G, Sculean A, Brecx M (1998) A pilot study of confocal laser scanning microscopy for the assessment of undisturbed dental plaque vitality and topography. Arch Oral Biol 43(4):277–285. doi:10.1016/S0003-9969(97)00121-0

    Article  PubMed  Google Scholar 

  25. Auschill TM, Arweiler NB, Netuschil L, Brecx M, Reich E, Sculean A (2001) Spatial distribution of vital and dead microorganisms in dental biofilms. Arch Oral Biol 46(5):471–476. doi:10.1016/S0003-9969(00)00136-9

    Article  PubMed  Google Scholar 

  26. Zaura-Arite E, van Marle J, ten Cate JM (2001) Confocal microscopy study of undisturbed and chlorhexidine-treated dental biofilm. J Dent Res 80:1436–1440. doi:10.1177/00220345010800051001

    Article  PubMed  Google Scholar 

  27. Tawakoli PN, Al-Ahmad A, Hoth-Hannig W, Hannig M, Hannig C (2013) Comparison of different live/dead stainings for detection and quantification of adherent microorganisms in the initial oral biofilm. Clin Oral Investig 17(3):841–850. doi:10.1007/s00784-012-0792-3

    Article  PubMed  Google Scholar 

  28. WHO (1997) Oral health surveys, basic methods, 4th edn. WHO, Geneva

    Google Scholar 

  29. Tomás I HB, Diz P, Donos N (2010) In vivo oral biofilm analysis by confocal laser scanning microscopy: methodological approaches. In: A M-V (ed) Microscopy. Science, technology, applications and education. Formatex, Badajoz (Spain), pp 597-606

  30. Tomás I, García-Caballero L, Cousido MC, Limeres J, Álvarez M, Diz Dios P (2009) Evaluation of chlorhexidine substantivity on salivary flora by epifluorescence microscopy. Oral Dis 15(6):428–433. doi:10.1111/j.1601-0825.2009.01570.x

    Article  PubMed  Google Scholar 

  31. Roberts SKBC, Brading M, Lappin-Scott H, Stoodley P (1999) Biofilm formation and structure; what’s new? In: Newman HNWM (ed) Dental plaque revisited—oral biofilms in health and disease. BioLine, Cardiff, pp 1–36

    Google Scholar 

  32. Diaz PI, Chalmers NI, Rickard AH, Kong C, Milburn CL, Palmer RJ Jr, Kolenbrander PE (2006) Molecular characterization of subject-specific oral microflora during initial colonization of enamel. Appl Environ Microbiol 72(4):2837–2848. doi:10.1128/AEM.72.4.2837-2848.2006

    Article  PubMed Central  PubMed  Google Scholar 

  33. Dige I, Nyengaard JR, Kilian M, Nyvad B (2009) Application of stereological principles for quantification of bacteria in intact dental biofilms. Oral Microbiol Immunol 24(1):69–75. doi:10.1111/j.1399-302X.2008.00482.x

    Article  PubMed  Google Scholar 

  34. Jung DJ, Al-Ahmad A, Follo M, Spitzmuller B, Hoth-Hannig W, Hannig M, Hannig C (2010) Visualization of initial bacterial colonization on dentine and enamel in situ. J Microbiol Methods 81(2):166–174. doi:10.1016/j.mimet.2010.03.002

    Article  PubMed  Google Scholar 

  35. Wood S, Nattress B, Kirkham J, Shore R, Brookes S, Griffiths J, Robinson C (1999) An in vitro study of the use of photodynamic therapy for the treatment of natural oral plaque biofilms formed in vivo. J Photochem Photobiol B 50(1):1–7

    Article  PubMed  Google Scholar 

  36. Robinson C, Strafford S, Rees G, Brookes SJ, Kirkham J, Shore RC, Watson PS, Wood S (2006) Plaque biofilms: the effect of chemical environment on natural human plaque biofilm architecture. Arch Oral Biol 51(11):1006–1014. doi:10.1016/j.archoralbio.2006.04.010

    Article  PubMed  Google Scholar 

  37. Palmer RJ Jr, Gordon SM, Cisar JO, Kolenbrander PE (2003) Coaggregation-mediated interactions of streptococci and actinomyces detected in initial human dental plaque. J Bacteriol 185(11):3400–3409. doi:10.1128/JB.185.11.3400-3409.2003

    Article  PubMed  Google Scholar 

  38. Chalmers NI, Palmer RJ Jr, Du-Thumm L, Sullivan R, Shi W, Kolenbrander PE (2007) Use of quantum dot luminescent probes to achieve single-cell resolution of human oral bacteria in biofilms. Appl Environ Microbiol 73(2):630–636. doi:10.1128/AEM.02164-06

    Article  PubMed Central  PubMed  Google Scholar 

  39. Hannig C, Follo M, Hellwig E, Al-Ahmad A (2010) Visualization of adherent micro-organisms using different techniques. J Med Microbiol 59(Pt 1):1–7. doi:10.1099/jmm.0.015420-0

    Google Scholar 

  40. Decker EM (2001) The ability of direct fluorescence-based, two-colour assays to detect different physiological states of oral streptococci. Lett Appl Microbiol 33(3):188–192. doi:10.1046/j.1472-765x.2001.00971.x

    Article  PubMed  Google Scholar 

  41. von Ohle C, Gieseke A, Nistico L, Decker EM, DeBeer D, Stoodley P (2010) Real-time microsensor measurement of local metabolic activities in ex vivo dental biofilms exposed to sucrose and treated with chlorhexidine. Appl Env Microbiol 76(7):2326–2334. doi:10.1128/AEM.02090-09

    Article  Google Scholar 

  42. Pan PH, Finnegan MB, Sturdivant L, Barnett ML (1999) Comparative antimicrobial activity of an essential oil and an amine fluoride/stannous fluoride mouthrinse in vitro. J Clin Periodontol 26(7):474–476

    Article  PubMed  Google Scholar 

  43. Fine DH, Furgang D, Barnett ML, Drew C, Steinberg L, Charles CH, Vincent JW (2000) Effect of an essential oil-containing antiseptic mouthrinse on plaque and salivary Streptococcus mutans levels. J Clin Periodontol 27(3):157–161. doi:10.1034/j.1600-051X.1999.260710.x

    Article  PubMed  Google Scholar 

  44. Jenkins S, Addy M, Wade W, Newcombe RG (1994) The magnitude and duration of the effects of some mouthrinse products on salivary bacterial counts. J Clin Periodontol 21(6):397–401. doi:10.1111/j.1600-051X.1994.tb00736.x

    Article  PubMed  Google Scholar 

  45. Stoeken JE, Paraskevas S, van der Weijden GA (2007) The long-term effect of a mouthrinse containing essential oils on dental plaque and gingivitis: a systematic review. J Periodontol 78(7):1218–1228. doi:10.1902/jop.2007.060269

    Article  PubMed  Google Scholar 

  46. Cortelli SC, Cortelli JR, Wu MM, Simmons K, Charles CA (2012) Comparative antiplaque and antigingivitis efficacy of a multipurpose essential oil-containing mouthrinse and a cetylpyridinium chloride-containing mouthrinse: a 6-month randomized clinical trial. Quintessence Int 43(7):e82–e94

    PubMed  Google Scholar 

  47. Van Leeuwen MP, Slot DE, Van der Weijden GA (2011) Essential oils compared to chlorhexidine with respect to plaque and parameters of gingival inflammation: a systematic review. J Periodontol 82(2):174–194. doi:10.1902/jop.2010.100266

    Article  PubMed  Google Scholar 

  48. Neely AL (2012) Essential oil mouthwash (EOMW) may be equivalent to chlorhexidine (CHX) for long-term control of gingival inflammation but CHX appears to perform better than EOMW in plaque control. J Evid Based Dent Pract 12(3 Suppl):69–72. doi:10.1016/S1532-3382(12)70017-9

    PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by project PI11/01383 from Carlos III Institute of Health (General Division of Evaluation and Research Promotion, Madrid, Spain), which is integrated in National Plan of Research, Development and Innovation (PN I + D + I 2008-2011). This project was co-financed by European Regional Development Fund (ERDF 2007–2013). The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Conflict of interest

The authors declare that they have no conflict of interest.

Author information

Authors and Affiliations

  1. Oral Sciences Research Group, School of Medicine and Dentistry, Santiago de Compostela University, C./ Entrerrios s/n, 15872, Santiago de Compostela, Spain

    Victor Quintas, Isabel Prada-López & Inmaculada Tomás

  2. Department of Stomatology, Faculty of Health Sciences, Rey Juan Carlos University, Madrid, Spain

    Juan Carlos Prados-Frutos

Authors
  1. Victor Quintas
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Isabel Prada-López
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Juan Carlos Prados-Frutos
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Inmaculada Tomás
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Inmaculada Tomás.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Quintas, V., Prada-López, I., Prados-Frutos, J.C. et al. In situ antimicrobial activity on oral biofilm: essential oils vs. 0.2 % chlorhexidine. Clin Oral Invest 19, 97–107 (2015). https://doi.org/10.1007/s00784-014-1224-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00784-014-1224-3

Keywords

Search

Navigation