## 3 Results and Discussion
\begin{table}
\begin{tabular}{c c c c c c} \hline \hline
**Step** & **Total Activity** & **Total Protein** & **Specific Activity** & **Yield** & **Purification** \\  & **[U]** & **[mg]** & **[mU/mg]** & **[\%]** & **Fold** \\ \hline Crude extract & 0.35 & 65.79 & 5.29 & 100 & 1 \\ Salting out & 0.17 & 25.8 & 6.43 & 47.65 & 1.21 \\ Sephadex G-75 & 0.05 & 0.65 & 75.65 & 14.21 & 14.3 \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Partial purification procedure of PPO from stored (10 days) iceberg lettuce. Figure 1: Effects of pH (**A**) and temperature (**B**) on the activity of iceberg lettuce PPO. Means with different letters are significantly different (\(n=9\); \(\alpha=0.05\)). The inhibitory effect of the studied pure chemical compounds and plant infusions is presented in Table 2. The lowest values of the IC50 (pyrocatechol as a substrate) were noted for chemical compounds. Even 0.03% solutions of ascorbic acid and L-cysteine inhibit the activity of an enzyme by 50% (with pyrocatechol as a substrate). Interestingly, the application of natural extracts in low concentrations also effectively limited the activity of the tested enzyme from the oxidoreductase class. Among the water plant extracts, the best results were obtained for lavage (IC50 = 0.09%), marjoram (IC50 = 0.13%), and orange peel (IC50 = 0.14%) with pyrocatechol as a substrate. Usually, synthetic substrates (catechol and its derivatives) are used to study PPO activity. In our research, to confirm the inhibitory properties of the selected extracts, additionally, an endogenous substrate isolated from iceberg lettuce was used. Similar results were obtained; the lowest IC50 values were in the case of water extract obtained from oregano (0.04%), lavage (0.09%), and orange peel (0.11%). Interestingly, in the case of L-cysteine, kojic acid, and ascorbic acid, the obtained IC50 values in reaction with the endogenous substrate were significantly higher than in the assays conducted using pyrocatechol. To clarify the inhibitory effects of the studied extracts on PPO activities, kinetic studies were performed with the Lineweaver-Burk method (Table 3, Figure 2). The majority of the studied extracts suppressed PPO activities in a dose-dependent manner; only in the case of chamomile extract and fresh parsley was it impossible to determine the IC50 values because the inhibition was dose-independent. \begin{table}
\begin{tabular}{c c c} \hline \hline
**Inhibitor** & \multicolumn{2}{c}{**IC50**} \\ \hline  & **Pyrocatechol** & **Endogenous Substrate** \\ \hline Chemical compounds & & [\%] \\ \hline L-cysteine & 0.00085 \(\pm\) 0.00002 k & 0.34 \(\pm\) 0.01 c \\ \hline kojic acid & 0.00043 \(\pm\) 0.00001 k & 0.0067 \(\pm\) 0.0002 k \\ \hline ascorbic acid & 0.00053 \(\pm\) 0.00001 k & 0.026 \(\pm\) 0.007 jk \\ \hline Plant infusions & & [\%] \\ \hline basil (_Ocimum basilicum_ L.) & 0.22 \(\pm\) 0.01 de & 0.21 \(\pm\) 0.01 e \\ \hline lovage (_Levisticum officinal_) & 0.09 \(\pm\) 0.003 hi & 0.09 \(\pm\) 0.004 i \\ \hline oregano (_Origanum vulgare_) & 0.15 \(\pm\) 0.007 fg & 0.04 \(\pm\) 0.002 j \\ \hline marjoram (_Origanum majorana_ L.) & 0.13 \(\pm\) 0.005 fg & 0.16 \(\pm\) 0.07 f \\ \hline orange peel (_Citrus sinensis_) & 0.14 \(\pm\) 0.002 fg & 0.11 \(\pm\) 0.03 gh \\ \hline lemon peel (_Citrus limon_) & 0.24 \(\pm\) 0.01 d & na. \\ \hline dried parsley leaves & 0.58 \(\pm\) 0.02 b & na. \\ (_Petroselinum crispum_) & dose-independent & na. \\ \hline chamomile (_Matricaria chamomilla_) & dose-independent & na. \\ \hline wheat bran from common wheat & 1.06 \(\pm\) 0.045 a & na. \\ (_Triticum aestivum_ L.) & & \\ \hline \hline \end{tabular}  
Means (\(\pm\) standard deviation) with different letters are statistically significant (\(n=9\); \(\alpha=0.05\), 2-way ANOVA), IC50— the concentration of an inhibitor that inhibits the enzyme activity by 50%; na.—not analyzed (the reactions with endogenous substrate were carried out only for the most active inhibitors).  
\end{table}