## 4 Discussion  
The stability of neutral electrolyzed water (NW) is a crucial factor determining its long-term efficacy as a disinfectant in oral applications. This study evaluated the effects of different storage conditions on key properties of NW, including pH, ORP, and ACC, all of which play important roles in its microbicidal performance. In electrolyzed water for disinfection, the fraction of free available chlorine (Cl\({}_{2}\), HOCl, and OCl\({}^{-}\)) as a sterilization component varies depending on the pH (Figure A1) [29; 30]. The free available chlorine in NW is mainly HOCl and secondarily hypochlorite ion (OCl\({}^{-}\)). After penetrating the cell wall and biological membrane and entering the inside of the bacterial cells, HOCl has the potential to permeate membranes into the interior of microorganisms and has an oxidizing impact on vital tissues (Figure A2) [30; 31]. In contrast, OCl\({}^{-}\) does not have membrane permeability and only oxidizes the cell membrane (plasma membrane) and cell periphery (Figure A2) [30; 31]. Therefore, HOCl has a higher bactericidal effect at lower concentrations than OCl\({}^{-}\). The dissociation constant (Ka) of HOCl at 25 \({}^{\circ}\)C is 2.95 \(\times\) 10\({}^{-8}\) mol/L (pKa: 7.53), and at pH 7.5, the ratio of HOCl and OCl\({}^{-}\) is approximately 1:1 [30; 31]. Consequently, HOCl concentration decreases in alkaline environments, reducing its bactericidal efficacy. To maximize antimicrobial activity for oral applications, the pH of NW should be less than 7.5. At a pH lower than 7.5, the bactericidal efficacy of HOCl increases. However, excessive acidity can lead to chlorine gas formation, causing odor and property deterioration. Acidic environment increases the risk of tooth tissue demineralization and corrosion of metal restorations [26; 27]. To achieve a high microbicidal effect without acid influence on tooth and metal restorations for intraocular treatment, referring to the critical pH of human enamel and dentin (5.5 and 6.0-6.2, respectively), NW should be prepared and used with a pH in the range of 6.2-7.5 for the standard-concentration type without dilution and in the range of 5.5-7.5 for the high-concentration type with dilution. In this study, standard-concentration NW preparations (NW\({}_{25}\), NW\({}_{35}\), and NW\({}_{50}\)) maintained a relatively stable pH range (6.9-7.3), remaining above the critical threshold for enamel demineralization (5.5) and dentin erosion (6.0-6.2) throughout the 126-day storage period. This stability is consistent with previous studies conducted by [PERSON] et al. (2021) and [PERSON] et al. (2022) [23; 32], which highlighted that neutral and slightly acidic electrolyzed waters maintain their stability when stored in refrigerated and shaded environments. However, high-concentration NW preparations (NW\({}_{500}\), and NW\({}_{1000}\)) showed significant pH falls, especially under non-shaded at room temperature storage (L\({}_{\text{RT}}\)). For example, the pH of NW\({}_{1000}\) fell below pH 4 after 14 days of storage under L\({}_{\text{RT}}\), posing a risk of enamel damage and chemical instability; this rapid acidification under L\({}_{\text{RT}}\) is consistent with the results from [PERSON] et al. (2020) and [PERSON] et al. (2011) [33; 34], who reported that \begin{table}
\begin{tabular}{c c c c} \hline \hline \multirow{2}{*}{**Test NW**} & \multicolumn{3}{c}{**Surviving Bacteria Number *} \\ \cline{2-4}  & **L\({}_{\text{RT}}\)** & **S\({}_{\text{RT}}\)** & **S\({}_{\text{LW}}\)** \\ \hline NW\({}_{25}\) & 2.9 (\(\pm\)1.6) \(\times\) 10\({}^{4}\) **\#** & n.d. & n.d. \\ Other 7 types of NW preparations & n.d. & n.d. & n.d. \\ \hline \hline \end{tabular}  
* Mean. The value in ( ) indicates the coefficient in SD index notation. **\#** Bacteria removal rate: 99.8 (\(\pm\)0.1)%. n.d.: not detected. Surviving bacteria number: \(<\)0, removal rate: 99.9999%.  
\end{table} 
Table 2: Bactericidal effect of NW preparations after the 126-day storage period.