# Olive Oil with Ozone-Modified Properties and Its Application
## 2 Results and Discussion
### Cytotoxicity of Ozonated Oils
\begin{table}
\begin{tabular}{c c c c c} \hline \multirow{2}{*}{**Cell Line**} & \multirow{2}{*}{**Oil Concentration (\(\mu\)g mL\({}^{-1}\))**} & \multicolumn{3}{c}{**Cytotoxicity (\%)**} \\ \cline{3-5}  & & & **Ozonated Olive Oil** \\ \cline{3-5}  & & **Refined Olive Oil** & **0.04 Mole O\({}_{\text{3}}\)/100 g** & **0.10 Mole O\({}_{\text{3}}\)/100 g** \\ \hline \multirow{6}{*}{HaCaT} & 39 & \(-7.14\pm\) 0.14 & \(15.05\pm\) 0.15 * & 8.06 \(\pm\) 0.37 \\  & 78 & 7.14 \(\pm\) 0.35 & 25.81 \(\pm\) 0.00 * & 12.10 \(\pm\) 0.83 \\  & 156 & 9.82 \(\pm\) 0.95 & 22.58 \(\pm\) 0.36 * & 13.71 \(\pm\) 0.43 \\  & 312 & 19.05 \(\pm\) 0.29 * & 24.19 \(\pm\) 0.07 * & 27.42 \(\pm\) 0.38 * \\  & 625 & 12.50 \(\pm\) 0.66 & 32.26 \(\pm\) 0.00 * & 26.61 \(\pm\) 0.59 * \\  & 1250 & 17.86 \(\pm\) 0.26 * & 21.51 \(\pm\) 0.67 & 22.58 \(\pm\) 0.36 * \\ \hline \multirow{6}{*}{LLC-PK1} & 39 & \(-0.51\pm\) 1.60 & \(-3.86\pm\) 2.36 & 11.29 \(\pm\) 2.27 \\  & 78 & 5.96 \(\pm\) 0.39 & 13.29 \(\pm\) 3.07 & 15.00 \(\pm\) 1.38 * \\ \cline{1-1}  & 156 & 3.08 \(\pm\) 0.62 & 22.29 \(\pm\) 1.27 * & 14.43 \(\pm\) 2.50 * \\ \cline{1-1}  & 312 & 4.81 \(\pm\) 1.04 & 23.29 \(\pm\) 2.44 * & 13.86 \(\pm\) 1.08 * \\ \cline{1-1}  & 625 & 3.85 \(\pm\) 0.14 & 15.24 \(\pm\) 0.21 * & 22.71 \(\pm\) 1.95 * \\ \cline{1-1}  & 1250 & 4.42 \(\pm\) 1.07 & 35.86 \(\pm\) 0.99 * & 36.86 \(\pm\) 2.22 * \\ \hline \multirow{6}{*}{Caco-2} & 39 & 9.87 \(\pm\) 1.20 & 14.33 \(\pm\) 2.76 & 10.19 \(\pm\) 2.72 \\  & 78 & 5.73 \(\pm\) 1.87 & 14.01 \(\pm\) 1.18 & 8.60 \(\pm\) 1.42 \\ \cline{1-1}  & 156 & 7.96 \(\pm\) 0.78 & 10.51 \(\pm\) 0.99 & 1.59 \(\pm\) 0.92 \\ \cline{1-1}  & 312 & 7.22 \(\pm\) 0.06 & 0.64 \(\pm\) 1.23 & 1.59 \(\pm\) 0.78 \\ \cline{1-1}  & 625 & 11.78 \(\pm\) 1.20 & 6.37 \(\pm\) 2.83 & 5.73 \(\pm\) 3.05 \\ \cline{1-1}  & 1250 & 7.22 \(\pm\) 0.15 & 2.71 \(\pm\) 2.63 & 1.59 \(\pm\) 0.35 \\ \hline \multirow{6}{*}{HeLa} & 39 & \(-19.96\pm\) 0.05 & 2.31 \(\pm\) 2.78 & \(-18.15\pm\) 5.00 \\ \cline{1-1}  & 78 & 7.09 \(\pm\) 0.20 & 0.64 \(\pm\) 7.35 & \(-0.72\pm\) 6.54 \\ \cline{1-1}  & 156 & 1.06 \(\pm\) 0.12 & 6.77 \(\pm\) 8.22 & 3.26 \(\pm\) 4.75 \\ \cline{1-1}  & 312 & 2.76 \(\pm\) 0.20 & 12.58 \(\pm\) 3.81 & 3.11 \(\pm\) 4.31 \\ \cline{1-1}  & 625 & 5.31 \(\pm\) 0.14 & 11.07 \(\pm\) 2.55 & 0.00 \(\pm\) 2.74 \\ \cline{1-1}  & 1250 & 6.93 \(\pm\) 0.11 & 18.23 \(\pm\) 2.66 & 15.68 \(\pm\) 6.13 \\ \hline \end{tabular}  
* results significantly different (conducted for component results) from negative control (i.e., the cells themselves not exposed to the cytotoxic agent), in the culture medium only ANOVA (\(p<0.01\)); when comparing the cytotoxicity between refined and ozonated oils (conducted for component results) at the same concentrations (for each cell line) results are not significantly different; ANOVA (\(p<0.01\)).  
\end{table} 
Table 5: Cytotoxic action of refined and ozonated olive oils towards normal (HaCaT and LLC-PK1) and cancerous (Caco-2 and HeLa) cell lines after 24 h exposure as measured in the 3-(4,5-dimethylthiazol-2)-2,5-diphenyltetrazolzolium bromide (MTT) assay comparing to the cells non-exposed to the olive oils. Each value represents the mean of four individual replications \(\pm\) SD. The increase in cytotoxicity was not linear, but it persisted at the similar level. Non-cancerous cell lines (HaCaT and LLC-PK1) seemed to demonstrate slightly higher sensitivity to ozonated oil (\(p<0.01\)) than cancerous ones (HeLa and Caco-2) (\(p<0.01\)), but generally, the oil demonstrated slight cytotoxicity--up to approximately 32 and 36% in the highest concentration for HaCaT and LLC-PK1, respectively (Table 5). The viability of HeLa and Caco-2 cancerous cell lines was persistent for all tested oil concentrations (above 80%). In addition, there were no statistically significant differences (\(p<0.01\)) between cytotoxicity induced by both ozonated oils (0.04 mole O\({}_{3}\) and 0.10 mole O\({}_{3}\)/100 g), so it did not depend on the degree of ozonation.