## 4 Discussion  
We compared the effect of PUI on the efficacy of the elimination of organic material remaining into infected, narrow and moderated curved mandibular root canals after their instrumentation using the RaCe rotary files 0.04 taper with apical enlargements of #25 and #35, following an instrumentation protocol similar to the one we used in clinical practice. Our results show that when PUI was combined with the RaCe rotary system instrumentation, there was no difference in the elimination of organic material between groups 1A and 1B (with apical enlargements of #25 and #35, respectively). Our results agree with the study of [PERSON] et al. who compared different apical diameters with and without PUI [18]. They found that PUI reduces remnant pulp tissues in canals to small preparations. However, in the present study we performed an instrumentation and irrigation protocol which is closer to the real clinical practice than the one used by [PERSON] et al., who used a single file protocol. In addition, we used infected and moderate curved root canals instead of vital pulp teeth. Regarding the latter, there is a consensus that PUI is more effective than traditional syringe irrigation, removing more efficiently the organic tissue, planktonic bacteria and dentine debris from the root canal. These phenomena could be explained because the ultrasound creates a higher speed and flow volume of the irrigant in the canal during irrigation, thereby eliminating more debris, producing less apical packing, allowing better access of the chemical product to accessory canals and even causing a flush effect to be produced by ultrasound but not manual irrigation [3; 13]. On the other hand, in our study when the instrumentation was performed without PUI in apical enlargements of #25 and #35, the difference in the elimination of organic material was significant, with a pronounced reduction in the 2B group (with apical enlargement of #35) compared with 2A group (with apical enlargement of #25). This is in agreement with several studies regarding apical enlargement that establish a directly proportional relationship between the increase of last instrument diameter and the reduction of organic and bacterial remaining [5; 6; 7; 10; 15; 21; 25; 26; 27; 28]. In fact, during canal preparation, apical size has been crucial in defining the successful debridement of the RCS because the penetration of the irrigant into the apical one-third of the canal and the removal of debris depend on the final size of the instrument used in the canals; so, an increase in the apical preparation size significantly enhances the root canal disinfection [10; 25]. In addition, several studies have shown the relevance not only of the apical preparation but also of the taper in the instrumentation of the apical one-third of root canals. Regarding this, a previous study by [PERSON] et al. (2014) showed that the cleanliness of the apical canal walls was better when the apical preparation was performed after a basic preparation of a \begin{table}
\begin{tabular}{c c c c} \hline \hline  & **Irigation System** & \multicolumn{2}{c}{**Diameter of the Instrument**} \\  & & & **(RaCe 0.04 Taper)** \\ \cline{2-4}  & & **PUI** & **CI** & **\#25** & **\#35** \\ \cline{2-4}  & & + & – & – & + \\ \cline{2-4}  & & + & – & + & – \\ \hline \hline \end{tabular}
\end{table} 
Table 3: Summary of the experimental design to assess the effect of three relevant variables on RCS cleanness: (1) Distance from the apex (depth); (2) Irrigation system (PUI vs. Conventional syringes); and (3) Diameter of the instrument used during biomechanical instrumentation (#25 vs. #35). A “+” sign indicates the presence of a significant effect of each of the analysed variables; its absence means that there are no differences between them. RCS = Root canal system; CI = Conventional irrigation. size #25/0.06 taper, irrespective of the technique used to perform the apical preparation [29]. [PERSON] et al.