We have developed a new tool to measure the acid production by plaque oral bacteria. Many species of oral bacteria metabolize sugars in food and produce organic acids that demineralize the dental enamel leading to the formation of cavities. Measuring the acidity level before and after a sugar rinse can indicate the susceptibility of an individual to tooth decay and location of active caries. In a case study on two subjects, a non-contact optics-based pH device was able to track pH before and after a sugar rinse. The fiber optic probe measures acidity level in difficult to access dental locations such as occlusal pits and fissures based on changes in the spectral fluorescence profile of fluorescein (FL) dye.
Fiber coupled 420 nm LED excites 200uM aqueous FL solution in the mouth. The fluorescence spectrum in 450-650 nm range is obtained using an adjacent fiber optic cable coupled to a spectrometer. Chemometric analysis of endmember dianion and anion species using least-square fitting is performed to determine the pH of the FL absorbed into the extracellular region of the oral biofilm. Other unwanted noise, like background light and auto-fluorescence in the range of 450-650 nm is removed before calculating biofilm pH.
Using this device in a darkened room on two subjects, we were able to measure resting pH (before a sugar rinse) and track time dependent change in pH (after a sugar rinse) in the range of pH 4-7 paving the way for first clinical optical pH measurement in the mouth.
Sugar-rich diets and poor dental hygiene promote the formation of a biofilm (plaque) that strongly adheres to the dental enamel surface and fosters the evolution of aciduric bacteria. The acid contributes to demineralization of the exterior tooth enamel, which accelerates after the pH drops below a critical value (∼5.5) for extended time periods resulting in the need for restorative procedures. Preventative techniques to alert the dentist and caries-susceptible patients regarding vulnerability to dental decay require a clinical measure of plaque activity. Therefore, there is a need to evaluate the acid production capability of plaque deposits in the pits and fissures of occlusal and interproximal regions. A ratiometric fluorescence pH-sensing device has been developed using an FDA-approved dye and LED excitation. Fluorescein spectral profiles were collected using a spectrometer and analyzed with a spectral unmixing algorithm for calibration over the pH range of 4.5 to 7. An in vivo pilot study on human subjects was performed using a sucrose rinse to accelerate bacterial metabolism and to measure the time-dependent drop in pH. The optical system is relatively immune to confounding factors such as photobleaching, dye concentration, and variation in excitation intensity associated with earlier dye-based pH measurement techniques.
A pH measurement of oral biofilms is helpful for monitoring the impact of acidogenic bacteria in the caries process. Demineralization of dental enamel is closely related to the time dependent pH of human plaque. Therefore, providing a means to easily measure the local pH of biofilms is a useful clinical diagnostic in the arsenal of caries prevention tools. Optical measurement methods of plaque metabolism can use intrinsic fluorescence or extrinsic fluorescence from added dyes. Autofluorescence spectral features of human oral biofilms at green (500 nm) and red (634 nm) fluorescence wavelengths using 405 nm excitation did not demonstrate a spectral or intensity shift between neutral and acidic conditions. Chlorin e6, an ingredient in chlorophyllin food supplement, exhibited a spectral and intensity shift of fluorescence emission in buffered solutions, but this quantitative pH-dependence was not transferable to a human plaque environment. Finally, a ratiometric quantitative pH measure was achieved by exciting (405 nm laser) a mixture of two dyes, fluorescein and rhodamine B. This two-dye mixture produced two strong fluorescent bands centered at 515 nm (fluorescein) and 580 nm (rhodamine B), where the 515 nm band was pH sensitive and the 580 nm band served as a pH insensitive reference. This dual-dye fluorescence ratio exhibited a linear response over pH 7 to 5 in human oral biofilms during a sugar challenge. We have explored methods to use non-contact, optical measures of local acidity levels in difficult to access dental locations such as occlusal fissures using various pH sensitive fluorescent dye systems.
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