Visualization and Quantification of the Oral Hygiene Effects of Brushing, Dentifrice Use, and Brush Wear Using a Tooth Brushing Simulator

Abstract

Approaches that reproduce dental hygiene regimens under controlled conditions have applications in preclinical research. We have applied standardized, reproducible brushing regimes to typodonts coated in simulated or biological plaques to assess the effects on tooth cleaning of toothbrush/dentifrice regimens. Replicated typodonts were coated with Occlude^TM or Glogerm^TM indicators to simulate plaque, and brushed reproducibly using a mechanical brushing simulator to compare the cleaning of occlusal surfaces before and after brushing with water or a dentifrice. An in vitro model using salivary inocula to cultivate oral biofilms on typodont surfaces was then developed to evaluate removal of disclosed plaque by new toothbrushes in comparison to toothbrushes with wear equivalent to 3 months of use. Analyses of typodonts brushed under controlled conditions significantly (p < 0.01) distinguished between brushed and unbrushed surfaces and between the use of water vs. dentifrice for the removal of simulated interproximal plaque (p < 0.05). New toothbrushes removed significantly (p < 0.05) more biological plaque from typodont surfaces than brushes that had been worn by repeated brushing. Through controlled and defined brushing of typodonts with simulated and biological plaques, the effectiveness of dental hygiene regimens was compared under reproducible conditions. Data indicate that the cleaning effectiveness of brushing was augmented by the addition of dentifrice and that new brushes were significantly more effective than brushes with substantial wear from previous use. Whilst we have focussed on the occlusal surfaces of molars and worn brushes, the method could be applied to a range of other tooth surfaces and oral hygiene regimens.

Keywords: biofilm model simulated plaque; brush wear; brushing simulator; dental plaque; typodont.

Figure 1

A typical experimental set-up. Up to eight typodonts were coated with simulated plaque and mounted with brushes on the brushing simulator. Typodonts were brushed with a constant brushing pattern under constant pressure for a specified period of time. Typodonts were then photographed and the images analyzed to quantify removal of plaque.

Figure 2

Development of a custom-built drip-flow model for the cultivation of dental plaque on typodont surfaces. (A) Medium vessel containing artificial saliva; (B) peristaltic pump running at approximately 4 mL h-1; (C) capillary delivering medium, drop-wise, onto a plastic typodont surface (D) in a sterile housing comprising a growth vessel and a waste vessel (E). (F) Shows an image of the assembled model. The model was run for 72 h at 37°C and surfaces were inoculated daily with the saliva of a healthy adult volunteer (2 mL). Visible plaque was evident following this growth phase.

Figure 3

Brushing caused significant changes in simulated plaque. The molars of plastic typodonts were coated with simulated plaque and brushed with an unused toothbrush under constant pressure and brushing pattern. Simulated plaque was visualized under controlled light conditions. Plaque removal is indicated by absence/reduction of green coloration (A) and quantification (mean and standard deviations) (B) for occlude and by fluorescence (C,D). Significantly less plaque was detected on brushed surfaces (p < 0.01, n = 9 typodonts).

Figure 4

A fluoride toothpaste (FTP) removed more simulated interproximal plaque than water. Example images are shown. Representative images (A) and data (mean and standard deviations) (B) show removal of simulated plaque from the interproximal regions. FTP removed significantly more simulated plaque than water (12 separate experiments) (p = 0.017; n = 12).

Figure 5

New brushes remove more plaque than worn brushes. Oral biofilm was cultivated, stained and brushed with a wetted toothbrush (new or worn) under constant pressure and brushing pattern. Images show stained plaque before and after brushing (A). The red channel was isolated, converted to gray-scale and inverted to differentiate between areas with or without plaque. The occlusal surfaces of two molars were isolated and light intensity was measured using ImageJ. Data show background-corrected mean signal intensity. Higher numbers indicate higher levels of plaque. Black bars represent plaque levels before brushing, while light bars represent plaque levels after brushing. Error bars show standard deviations (B). Data represent two separate experiments; mean values and standard deviations. Both brush types caused significant reductions (p < 0.05). The new brush removed significantly more biofilm than the worn brush (p < 0.05; n = 6 typodonts).