## 4 Discussion  
In the current experimental study, the attenuation properties of five different photosensizers were evaluated. The laser wavelengths examined were 445 nm, 635 nm, 660 nm and 808 nm. These wavelengths, and their corresponding light sources, were selected since they represent those which are most commonly employed in aPDT dental applications. Except for toluidine blue illuminated by an 808 nm laser showing the lowest intensity loss, all the others gave attenuation ranges of 63% to 99%.  
It is evident that the concentration of the photosensizer clearly affects the results. Higher concentrations of methylene blue and indocyanine green revealed no measurable light intensity after passing through the tested photosensizer. This can be explained by the two merged laws, specifically the [PERSON]'s law. Indeed, the applied formula is provided in Equation (2).  
\[\log\mathrm{I}_{0}/\mathrm{I}_{\mathrm{d}}=\mathrm{A}=\varepsilon*\mathrm{C}* \mathrm{d} \tag{2}\]  
\begin{table}
\begin{tabular}{c c c c} \hline
**Group** & **Mean** & **Lower Bound (95\%)** & **Upper Bound (95\%)** \\ \hline CUR + 445 & 0.880 & 0.879 & 0.881 \\ \hline ICG0.1 + 808 & 0.683 & 0.682 & 0.683 \\ \hline ICG1 + 808 & 1.119 & 1.118 & 1.120 \\ \hline MB0.05 + 660 & 0.705 & 0.704 & 0.706 \\ \hline MB0.1 + 660 & 1.096 & 1.095 & 1.096 \\ \hline MB10 + 660 & 1.238 & 1.237 & 1.239 \\ \hline PL + 660 & 1.437 & 1.436 & 1.438 \\ \hline PL + 808 & 1.092 & 1.091 & 1.092 \\ \hline TB + 635 & 0.940 & 0.939 & 0.941 \\ \hline TB + 808 & 0.118 & 0.117 & 0.119 \\ \hline \end{tabular}
\end{table}
Table 2: Arcsine-transformed mean values and their 95% CIs for all photosensitizer /laser wavelength treatment combinations featured in this study. Abbreviations: CUR, Curcumin; MB, Methylene blue; PL, Photolase (methylene blue derivative); TB, Toluidine blue; ICG, Indocyanine green.  
where I0 is the initial intensity of the light, Id is the intensity after travelling the distance d in the medium, A is the absorbance, \(\varepsilon\) is the extinction coefficient of the medium at a selected wavelength, c is the molar concentration and d is the travelling distance in the medium or light path length [32].  
Additionally, higher concentrations (c) of the applied photosensitizer can limit the efficacy of the treatment through \"photobleaching\". This phenomenon mainly occurs when the generated singlet oxygen chemically reacts with the photosensitizer, and hence hinders any further photodynamic processes [33; 34].  
Besides added concentration, another parameter that determines the absorbance value is, of course, the wavelength-dependent molar extinction coefficient (\(\varepsilon\)) of each photosensitizing agent. These values should be noted, since photosensitizers with higher values of this parameter can dramatically increase absorbance, a phenomenon affecting light penetration and leading to \"optical shielding\" at their corresponding wavelengths of maximum absorption [23]. This phenomenon refers to light blocking attributable to high superficial absorbance, and hence inhibition of the light from reaching deeper layers of the target tissue involved [35].  
Regarding the light path length (d), in this experimental set-up, a 1-mm path-length cuvette was used for all evaluations; this is a previously described optimal path length for spectrophotometric studies to allow accurate measurements [30]. Additionally, this is the approximate thickness of the photosensitizer typically applied in clinical dental procedures [9]. Therefore, this parameter can be considered as a constant in the experiments conducted.  
As noted in the Introduction, photodynamic therapy is based on the combination of three elements: the photosensitizer involved, light source and available oxygen level [5]. Therefore, the nature of each photosensitizer, the light pathway into the surrounding tissues of the oral cavity and the presence, concentration and availability of oxygen (for \({}^{1}\)O\({}_{2}\) generation) should be thoroughly examined.