## 3 Results
### The Ozone Degradation of Contaminants
#### 3.2.3 The Biotoxicity Assays: The Bioluminescent Bacteria  
The biotoxicity test based on the inhibition of the _Vibrio fisher_ light emission has been widely used to determine the presence of toxic compounds in liquid or solid samples by this rapid screening test. Bacterial bioluminescence is an energy-consuming phenomenon and light is emitted only when the organisms are in their best metabolic and physiologic conditions. Any component in the environment injuring the bacterial integrity or functions will produce a reduction or the disappearance of the light emission in a way directly proportional to the intensity of the suffered damage. As expected, the mixture of the six pollutants was highly toxic to the luminescent bacteria reducing the light emission intensity to zero. The ozone treatment was effective in the cleavage of the parent molecules, leading to a partial restoration of luminescence emission, as shown in Figure 4. Prolonged-time treatment seems to increase and not reduce the toxicity of the mixture. Probably, the degradation process produces smaller \begin{table}
\begin{tabular}{c c c c c c c} \hline
**Time** & **Cyprodinil** & **Bisphenol-A** & **Carbamazepine** & **Imalcloprid** & **Benzophenone-4** & **Ofloxacin** \\
**(h)** & **mg/L \(\pm\) SD** & **mg/L \(\pm\) SD** & **mg/L \(\pm\) SD** & **mg/L \(\pm\) SD** & **mg/L \(\pm\) SD** & **mg/L \(\pm\) SD** \\ \hline
**LOD\(\rightarrow\)** & **(25 ng/L)** & **(0.1 mg/L)** & **(25 ng/L)** & **(25 ng/L)** & **(5 ug/L)** & **(25 ng/L)** \\ \hline
0 & 0.225 \(\pm\) 0.003 & 0.535 \(\pm\) 0.014 & 0.561 \(\pm\) 0.015 & 0.500 \(\pm\) 0.011 & 0.385 \(\pm\) 0.002 & 0.640 \(\pm\) 0.003 \\
1 & \textless{LOD} & \textless{LOD} & \textless{LOD} & 0.433 \(\pm\) 0.005 & \textless{LOD} & 0.012 \(\pm\) 0.002 \\
2 & – & – & – & 0.419 \(\pm\) 0.002 & – & 0.009 \(\pm\) 0.004 \\
4 & – & – & – & 0.356 \(\pm\) 0.007 & – & \textless{LOD} \\
6 & – & – & – & 0.303 \(\pm\) 0.005 & – & – \\
8 & – & – & – & 0.284 \(\pm\) 0.005 & – & – \\ \hline \end{tabular}
\end{table} 
Table 4: Concentration of the pollutants before (T = 0) the treatment and in the treated samples. \begin{table}
\begin{tabular}{c c c} \hline
**Samples** & **COD (mg/mL)** & **TOC (mg/L)** \\ \multicolumn{2}{c}{**(\(\pm\)Uncertainty)**} \\ \hline Untreated & 19.24 \(\pm\) 5.12 & 21.6 \(\pm\) 2.40 \\ Treated 6 h & & 14.6 \(\pm\) 1.80 \\ Treated 8 h & 15.57 \(\pm\) 1.52 & \\ \multicolumn{2}{c}{\(\Delta\)COD = 21.1 \(\pm\) \%} & \(\Delta\)TOC = 32.4 \(\pm\) 2.2\% \\ \hline \end{tabular}
\end{table} 
Table 5: The COD and the TOC values in secondary effluent treated and untreated samples. fragments able to interact more effectively with bacterial physiology. The increased toxicity of pollutants after degradation treatment is a well-known phenomenon. Figure 4 shown that the chronic toxicity assay can offer more precise information with respect to the acute one. After one hour, the untreated sample looked very similar to the control, but after 24 h the situation was completely different. The sensitivity of the assay is underlined by the fact that its results indicate that even the tap water, employed to prepare the solution, produced a 20% of light emission inhibition with respect to the control, probably due to the sanitizing treatment residues. By testing the effluent samples, no significant toxicity of both untreated and treated samples was revealed. On the other hand, in the chronic toxicity assays, treated samples frequently showed a light emission higher than that of the control. This was not surprising since WW was rich in organic compounds and the ozone degradation process produced smaller molecules acting as nutrients. The eutrophic effect produced by these nutrients must be carefully considered when planning the discharge of treated effluents in water bodies. Figure 5 reports the emissions of _Vibrio fisher_ in presence of the effluent samples. On some occasions, the results were quite different: some samples were more toxic than expected or were greatly stimulated the light emission.