## 4 Discussion
As it was mentioned above, in our study, the effect of etidronate on the ability of NaOCl to degrade organic components was manufacturer-dependent. All concentrations of ID decreased this activity of NaOCl, although, for ID18%, the differences did not reach the level of statistical significance. At the same time, the DR of all concentrations promoted the tissue-dissolving capability of NaOCl. The increase in the proteolytic activity in the HEDP/NaOCl group was also reported by [PERSON] et al. [62]. The authors demonstrated that 2.25% NaOCl alone dissolved 27% of bovine muscle tissue by weight, while 5% NaOCl combined with 18% HEDP dissolved 41% of organic tissue. Previously, it has been shown that EDTA may decrease the ability of NaOCl to degrade organic components. However, there is a controversy in the literature regarding the proteolytic activity of the NaOCl + EDTA mixture. In some studies, mixtures of 5% NaOCl with 17% EDTA produced no tissue-dissolving effect at all [43; 62], while [PERSON] et al. reported that EDTA significantly reduced the capability of NaOCl to dissolve pulp tissue [72]. In the latter study, the percent weight losses were 57% in the 2.5% NaOCl group and 33% in the 2.5% NaOCl + EDTA group after a 15 min immersion. Similarly, in our study, we observed a two-fold decrease in the organic tissue degrading capacity of NaOCl mixed with EDTA as compared with NaOCl alone. However, these solutions are not used as a mixture in clinical practice; instead, they are used sequentially. We found that the sequential use of NaOCl and EDTA resulted in a significantly higher proteolytic activity compared with NaOCl used alone or mixed with EDTA. On the other hand, it is next to impossible to completely avoid contact between these solutions in root canal complexities where pulp dissolving activity is of particular importance. In most studies assessing the ability of irrigating solutions to dissolve pulp tissue remnants, saline was used as a negative control. The immersion in this solution produced no [64; 73] or a non-significant [62; 71] decrease in the weights of the specimens. [PERSON] et al. used distilled water as a negative control. They found no decrease in the weights of the specimens in the control group [74]. In our study, we found a slight decrease in the samples' weights after a 15 min immersion in distilled water. However, these changes were insignificant. For the assessment of SEM images, we modified the Hulsmann scale of the smear layer [69] based on the preliminary analysis of the images. In our scale, we described the smear layer removal pattern by focusing on the degree of dentinal tubule closure and not on the thickness of the smear layer. Dentinal tubules were graded as completely open, partially open, or closed. All cases where all tubules were completely closed were graded with the same score (score 5). In our study, we assessed smear layer removal with the help of different irrigating solutions at different levels of the root canals. A number of previous studies have underlined a greater amount of the residual smear layer in the apical root canal third compared to the middle and coronal root canal thirds [7; 58; 59; 60]. We also found less cleanliness of the root canal walls in the apical part. Previous studies evaluating the removal of the smear layer observed inconsistent results on whether the combination of NaOCl with HEDP was more effective than the standard irrigating sequence (NaOCl and EDTA). In a study by [PERSON] et al., the amount of the smear layer in the apical and coronal thirds did not differ significantly between the 6% NaOCl/HEDP (DR) and 6% NaOCl and 17% EDTA groups. However, NaOCl and EDTA removed significantly more of the smear layer in the middle thirds of the root canals [4]. [PERSON] et al. reported that the 9% HEDP-based (DR) continuous irrigation did not differ from alternating irrigation with 3% NaOCl, followed by EDTA, in terms of the ability to remove the smear layer [59].