## 2 Materials and Methods
### Antibacterial Activity  
The antibacterial activity of the _P. pinaster_ Ait. and _P. pinea_ L. bark polar extracts was assessed against two infectious agents of public health relevance, namely the Gram-positive _S. aureus_ (ATCC\({}^{\circledright}\) 6538) and the Gram-negative _E. coli_ (ATCC\({}^{\circledright}\) 25922), as illustrated in Figure 2.  
The _P. pinaster_ Ait. bark polar extracts exhibited bactericidal action in comparison with the solvent control, at a 6.25-25 mg mL\({}^{-1}\) range (Figure 2A), causing 8.3-, 6.3-, and 4.0-log (CFU mL\({}^{-1}\)) statistically significant decreases at 6.25, 12.5, and 25 mg mL\({}^{-1}\), respectively (\(p<0.05\)) against the Gram-positive bacterium _S. aureus_. Furthermore, the _P. pinea_ L. bark polar extracts showed a bactericidal effect at the same concentration range (Figure 2A), revealing 8.3-, 6.0-, and 4.6-log (CFU mL\({}^{-1}\)) reductions in relation to the solvent control, at \begin{table}
\begin{tabular}{c c c c} \hline \hline \multirow{2}{*}{**Concentration**} & \multicolumn{3}{c}{**Pearson Correlation Coefficients (\(r\))**} \\ \cline{2-4}  & **Reducing Power** & **DPPH Scavenging Effect** & **ABTS Scavenging Effect** \\ \hline Total phenolic content & \(-0.403\) & \(-0.146\) & \(0.228\) \\ Proanthocyanidin content & \(0.289\) & \(-0.146\) & \(0.625\) \\ Total identified compounds & \(-0.757\) & \(-0.257\) & \(-0.325\) \\ Total identified phenolic compounds & \(-0.642\) & \(-0.203\) & \(-0.211\) \\ Flavan-3-ols & \(-0.627\) & \(-0.226\) & \(-0.196\) \\ Flavononols & \(-0.828\) 1 & \(-0.071\) & \(-0.562\) \\ Quintic acid & \(-0.856\) 1 & \(-0.305\) & \(-0.441\) \\ Protocatechic acid & \(-0.363\) & \(0.221\) & \(0.226\) \\ B-type (epi)catechin dimer isomer 1 & \(-0.732\) & \(-0.122\) & \(-0.308\) \\ B-type (epi)catechin dimer isomer 2 & \(-0.502\) & \(-0.098\) & \(-0.108\) \\ Catechin + B-type (epi)catechin dimer isomer 3 & \(-0.484\) & \(-0.119\) & \(-0.093\) \\ B-type (epi)catechin trimer isomer 2 & \(0.013\) & \(-0.114\) & \(-0.392\) \\ A-type (epi)calcechin-(epi)catechin dimer & \(-0.770\) & \(-0.179\) & \(-0.212\) \\ B-type (epi)catechin trimer isomer 3 & \(-0.721\) & \(-0.416\) & \(-0.361\) \\ B-type (epi)catechin trimer isomer 4 & \(-0.579\) & \(-0.334\) & \(-0.071\) \\ B-type (epi)catechin dimer isomer 4 & \(0.721\) & \(0.680\) & \(0.217\) \\ Taxifolin & \(-0.823\) 1 & \(-0.029\) & \(-0.605\) \\ \hline \hline \end{tabular} \({}^{1}\) The correlation is statistically significant at \(p<0.05\).  
\end{table} 
Table 6: Pearson correlation coefficients between the concentration of compounds and the IC\({}_{50}\) values of reducing power, DPPH, and ABTS scavenging effects exhibited by _Pinus pinaster_ Ait. and _Pinus pinate_ L. bark polar extracts. 6.25, 12.5, and 25 mg mL\({}^{-1}\), respectively (\(p<0.05\)). On the one hand, _P. pinea_ L. bark polar extracts even significantly suppressed _S. aureus_ growth at 3.13 mg mL\({}^{-1}\) compared to the solvent control (\(p<0.05\)), via bactericidal action, resulting in a 3.15-log (CFU mL\({}^{-1}\)) loss; on the other hand, _P. pinaster_ Ait. bark polar extracts decreased _S. aureus_ growth, at that concentration, with no statistical difference in comparison to the solvent control (\(p>0.05\)), leading to a 2.10-log (CFU mL\({}^{-1}\)) decrease. Regarding the inhibitory effects of _Pinus_ spp. bark polar extracts against the Gram-negative bacterium _E. coli_ growth (Figure 2B), with the 24 h incubation with 12.5 mg mL\({}^{-1}\), no statistical difference (\(p>0.05\)) was observed between the extract and the solvent control. Surprisingly, when tested at 25 mg mL\({}^{-1}\), both extracts acted as strong bactericidal agents, with a significant 8.26-log (CFU mL\({}^{-1}\)) loss in relation to the solvent control (\(p<0.05\)). The essay was repeated several times, corroborating the results reported here. In fact, a previous study has already pointed out that Gram-positive bacteria are more easily inactivated by pine extracts than Gram-negative ones [67].