## 3 Results
### Effects of Metal Ions and Chemical Agents on Keratinases Stability  
The effects of various metal irons with different concentrations (1 mmol/L and 5 mmol/L) on keratinocyte activity were studied and the results were shown in Table 4 Mg\({}^{2+}\) and Li\({}^{+}\) had stimulatory effect on residual activity of DgeKer; 5 mmol/L K\({}^{+}\) caused increased activity of DgeKer (104.11 \(\pm\) 0.14%). By contrast, other tested metal ions prompted lower stability of DgeKer. 1 mmol/L and 5 mmol/L Ca\({}^{2+}\) affected the DgeKer stability with residual activity of 94.07 \(\pm\) 0.72% and 83.06 \(\pm\) 0.36%, respectively. DgeKer-C was stable (101.07 \(\pm\) 1.37%) at 5 mmol/L Co\({}^{2+}\). The other tested metal ions had a different decrease on the stability of DgeKer-C with residual activity of 88.25 \(\pm\) 0.58% after pretreatment with 5 mmol/L K\({}^{+}\). Unfortunately, DgeKer and DgeKer-C showed complete inhibition with 5 mmol/L Cr\({}^{3+}\). There was no significant difference (\(p\) > 0.05) between DgeKer and DgeKer-C after pretreatment with 1 mmol/L K\({}^{+}\), Ca\({}^{2+}\), Mn\({}^{2+}\), Ni\({}^{2+}\) and 5 mmol/L Cd\({}^{2+}\). DgeKer showed better tolerance than DgeKer-C to Li\({}^{+}\) and Mg\({}^{2+}\). However, DgeKer-C was more stable than DgeKer with the pretreatment Co\({}^{2+}\) and Cu\({}^{2+}\). In general, DgeKer and DgeKer-C had good tolerance to some metal ions. \begin{table}
\begin{tabular}{c c c c} \hline \hline \multirow{2}{*}{**Metal Irons**} & **Concentration** & \multicolumn{2}{c}{**Residual Activity (\%)**} \\ \cline{3-4}  & **(mmol/L)** & **DgeKer** & **DgeKer-C** \\ \hline Control & & 100.00 \(\pm\) 0.27 \({}^{\text{m}}\) & 100.00 \(\pm\) 3.11 \({}^{\text{k}}\) \\ K\({}^{+}\) & 1 & 91.77 \(\pm\) 0.49 \({}^{\text{j}}\)k & 87.45 \(\pm\)1.41 \({}^{\text{s}}\) \\  & 5 & 104.11 \(\pm\) 0.14 \({}^{\text{o}}\) & 88.25 \(\pm\) 0.58 \(\pm\) 0.58 \(\pm\) 0.58 \(\pm\) 0.56 \({}^{\text{sh},*}\) \\ Li\({}^{+}\) & 5 & 113.97 \(\pm\) 0.62 \({}^{\text{p}}\) & 88.81 \(\pm\) 1.72 \({}^{\text{sh},*}\) \\ Mg\({}^{2+}\) & 1 & 103.35 \(\pm\) 1.07 \({}^{\text{n,o}}\) & 93.80 \(\pm\) 1.12 \({}^{\text{j}}\)* \\ Ca\({}^{2+}\) & 5 & 121.53 \(\pm\) 0.75 \({}^{\text{q}}\) & 87.09 \(\pm\) 0.71 \({}^{\text{s}}\)* \\ Ca\({}^{2+}\) & 1 & 94.07 \(\pm\) 0.72 \({}^{\text{k}}\) & 92.13 \(\pm\) 1.18 \({}^{\text{j}}\)j \\ Ca\({}^{2+}\) & 5 & 83.06 \(\pm\) 0.36 \({}^{\text{g}}\) & 90.65 \(\pm\) 0.88 \({}^{\text{h},*}\) \\ Co\({}^{2+}\) & 1 & 91.20 \(\pm\) 1.66 \({}^{\text{j}}\) & 99.20 \(\pm\) 3.37 \({}^{\text{k},*}\) \\ Co\({}^{2+}\) & 5 & 92.82 \(\pm\) 0.49 \({}^{\text{k,l}}\) & 101.07 \(\pm\) 1.37 \({}^{\text{k},*}\) \\ Mn\({}^{2+}\) & 1 & 85.07 \(\pm\) 0.89 \({}^{\text{h}}\) & 80.94 \(\pm\) 0.45 \({}^{\text{f}}\) \\ Mn\({}^{2+}\) & 5 & 64.69 \(\pm\) 0.27 \({}^{\text{d}}\) & 60.39 \(\pm\) 0.96 \({}^{\text{b},*}\) \\ Ni\({}^{2+}\) & 1 & 88.33 \(\pm\) 0.82 \({}^{\text{i}}\) & 86.67 \(\pm\) 0.87 \({}^{\text{g}}\) \\  & 5 & 91.29 \(\pm\) 0.23 \({}^{\text{i}}\) & 77.51 \(\pm\) 0.66 \({}^{\text{e}}\)* \\ Cd\({}^{2+}\) & 1 & 74.93 \(\pm\) 0.62 \({}^{\text{e}}\) & 86.34 \(\pm\) 0.83 \({}^{\text{g}}\)* \\ Cd\({}^{2+}\) & 5 & 63.16 \(\pm\) 1.02 \({}^{\text{c}}\) & 65.32 \(\pm\) 1.23 \({}^{\text{c}}\) \\ \hline \hline \end{tabular}
\end{table} 
Table 4: Effects of various metal ions on keratinocyte activity of DgeKer and DgeKer-C. \begin{table}
\begin{tabular}{c c c} \hline \hline \multirow{2}{*}{**Substrate**} & \multicolumn{2}{c}{**Activity of Different Keratinases (U/mg)**} \\ \cline{2-3}  & **DgeKer** & **DgeKer-C** \\ \hline Soluble keratin & 40,192.30 \(\pm\) 230.48 & 40,408.89 \(\pm\) 342.73 \\ Feather powder & 1494.78 \(\pm\) 25.78 & 1462.54 \(\pm\) 71.19 \\ \hline \hline \end{tabular}
\end{table} 
Table 3: Activities of different keratinases on several proteinaceous substrates. As shown in Table 5, all the surfactants and organic solvent employed negatively impacted on the stability of DgeKer and DgeKer-C, with more drastic effect obtained in the presence of \(\beta\)-mercaptoethanolhanol.