# Application of the _Aliium_ Test in Toxicity Studies of Lead and Copper: A Cytological Perspective
## 3 Results
### Analysis of the Effect of Cu+2 on the Mitotic Index  
The effect of copper observed in the mitotic index is substantially different, as it is significantly influenced by the dose and exposure time (\(p<0.001\)). Furthermore, the interaction between dose and time is also significant (\(p<0.001\)). Therefore, cellular division is inhibited and is dependent on both the dose used and the duration of exposure. Thus, the acute effect of copper is reflected, as increasing the dose leads to a more pronounced inhibitory effect on mitotic division over the exposure time (Table 3). According to the results regarding the effects of copper on the mitotic index, compared to the control the mitotic index (MI) decreases significantly with increasing copper concentration and exposure time (Figure 4). \begin{table}
\begin{tabular}{l l l l l l} \hline \hline
**Source** & **Df** & **Sum Sq** & **Mean Sq** & **F Value** & **Pr(\textgreater{}F)** \\ \hline Dose & 4 & 511.50 & 127.88 & 164.01 & \textless{}2 \(\times\) 10\({}^{-16}\)*** \\ \hline Time & 2 & 106.10 & 53.04 & 68.02 & \(3.75\times\) 10\({}^{-16}\)*** \\ \hline Dose–Time & 8 & 25.2 & 3.15 & 4.05 & \(6.78\times\) 10\({}^{-4}\)*** \\ \hline Residuals & 60 & 46.80 & 0.78 & & \\ \hline \hline \end{tabular}  
* Significance level: *** \(p<0.001\).  
\end{table} 
Table 3: ANOVA test for determining the factorial implications in the reduction of the mitotic index. Figure 3: Polynomial regression analyzing the influence of time on the mitotic index under lead exposure. For the control, the MI remains above 10%, regardless of the exposure duration. However, the MI decreases significantly at the first dose of 0.25 mM at a 48-72 h exposure time. Regarding the mitotic index, under copper treatment, the index was inhibited along with the concentration exposure. The results indicate a significant inhibition of the mitotic index (MI) at the highest concentration (1 mM) and the longest exposure duration of 72 h. The reduction trend is significant, with only 2.52% of the cells in division (Table 4). Regarding the 50% inhibitory concentration relative to the control (IC\({}_{50}\)), the results indicate that this level was achieved at a concentration of 0.50 mM after 72 h of exposure, 0.75 mM after 72 h of exposure, and 1.00 mM after 48 h of exposure. The observed inhibition in the MI highlights the genotoxic effects of copper on cellular division. Also, the decline in MI reflects the inhibition of cellular processes caused by the presence of numerous abnormal cells, which may disrupt chromosomal segregation during mitosis. Abnormal cells were first detected at a concentration of 0.25 mM after 72 h of exposure. As concentration and exposure time increase, the abnormalities index (AI) increases significantly, revealing the genotoxic effect of copper. The highest AI was observed at the concentration of 1.00 after 72 h of exposure. This trend reveals copper's capacity to induce chromosomal abnormalities, highlighting its genotoxic effect, which could have long-term implications. Figure 4: Polynomial regression showing the influence of time on the mitotic index under copper treatment. \begin{table}
\begin{tabular}{c c c c c c c c c c} \hline \hline
**Dose** & **Time** & \begin{tabular}{c} **Miotic\_Index** \\ **(\%)** \\ \end{tabular}  & \begin{tabular}{c} **AC\({}^{*}\)** \\ **(\%)** \\ \end{tabular}  & \begin{tabular}{c} **Abnormalities\_Index** \\ **(\%)** \\ \end{tabular}  & \begin{tabular}{c} **Interphase** \\ **(\%)** \\ \end{tabular}  & \ \begin{tabular}{c} **Prophase** \\ **(\%)** \\ \end{tabular}  & \begin{tabular}{c} **Metaphase** \\ **(\%)** \\ \end{tabular}   
Other mitotic indices, such as the probleme index, metaphase index, anaphase index, and telophase index, decrease with increasing concentration and prolonged exposure time (Table 4). The results of the polynomial regression analysis highlight the trend of a decreasing mitotic index (MI) resulting from copper treatment.