## 3 Results and Discussion
### Characterization of Nanoparticles
Zeta potential analysis suggests that an increase of temperature resulted in less negative charge of extract-loaded PLGA NPs over time, while the surface charge of extract-loaded PCL NPs was not apparently affected by temperature. Morphology is also a key factor for skin permeation [40]. The extract-loaded NPs (both PLGA and PCL) or empty NPs observed by SEM presented a general spherical shape, a smooth surface, and a size range below \(\approx\)471 nm for extract-loaded PLGA NPs and \(\approx\)243 nm for extract-loaded PCL NPs (Figure 5), confirming DLS data. Following the extract's encapsulation in PLGA NPs, the morphology did not changed. In the case of PCL NPs, a different density of particles between empty and extract-loaded PCL NPs was observed (Figure 5c,d, respectively). TEM observations confirm the particles' spherical shape, their nanometric dimensions and their uniform size distribution (Figure 6), which is in line with to the results obtained by the DLS method. Figure 7 shows the AFM images obtained for the different NPs, as well as their corresponding section profiles. The AFM analysis showed a mean particle size of 255 \(\pm\) 55 nm for empty PLGA NPs, a smaller particle size after loading it with 99 \(\pm\) 20 nm. PCL NPs had a larger size in comparison with PLGA NPs (367 \(\pm\) 44 nm for empty PCL NPs and 243 \(\pm\) 62 nm for extract-loaded PCL NPs). Figure 5: SEM micrographs showing the morphology of NPs. (**a**) Empty PLGA NPs, (**b**) Extract-loaded PLGA NPs, (**c**) Empty PCL NPs and (**d**) Extract-loaded PCL NPs. Scale bars = 2 μm (**a**, **b**), 10 μm (**c**,**d**). Figure 6: TEM micrographs showing the morphology of NPs. (**a**) Empty PLGA NPs, (**b**) Extract-loaded PLGA NPs, (**c**) Empty PCL NPs and (**d**) Extract-loaded PCL NPs. Scale bars = 0.2 μm (**a**,**b**,**c**), 0.1 μm (**d**). The spherical shape was reported as the ideal form for a fast endocytosis [41]. In the current study, all types of NPs showed a spherical shape when observed by SEM, TEM, and AFM. The encapsulation efficiency (EE) in NPs was determined in terms of ruin concentration. The value was very similar for PLGA NPs (75.90 \(\pm\) 31.14%) and PCL NPs (86.31 \(\pm\) 25.21%). In order to clarify the EE values, the complex of ruin and the polymers of NPs (PLGA and PCL) were analyzed. Table 1 presents the inherent molecular energy attributes for various biomolecular complexes, while Figure 8 shows the corresponding geometrical positions, which confirm a relation between the ruin and the polymers used in each NPs (PLGA and PCL). \begin{table}
\begin{tabular}{c c c c c c c} \hline \hline
**Energy** & **Rutin** & **PCL** & **PCL-Rutin** & \(\Delta\)E1 ** & **PLGA** & **PLGA-Rutin** & \(\Delta\)E1 ** \\ \hline Total2 & 23.09 & 6.41 & 11.47 & \(-\)18.04  & 2.64 & 5.37 & \(-\)20.36  \\ Bond3 & 1.16 & 0.34 & 1.41 & \(-\)0.09  & 0.29 & 1.64 & 0.19  \\ Angle4 & 7.57 & 1.33 & 9.14 & 0.24  & 2.53 & 13.55 & 3.44  \\ Dihed5 & 14.32 & 0.05 & 8.63 & \(-\)5.74  & 0.89 & 14.72 & \(-\)0.49  \\ vdW6 & 0.04 & 4.69 & \(-\)7.71 & \(-\)12.45  & \(-\)1.06 & \(-\)24.12 & \(-\)23.18  \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Inherent energy attributes of polymer-ruin complexes calculated using static-lattice atomistic simulations (SLAS) in vacuum. Figure 7: AFM topographic images of NPs under study. (**a**,**b**) Empty PLGA NPs, (**c**,**d**) Extract-loaded PLGA NPs, (**e**,**f**) Empty PCL NPs, (**g**. **h**) Extract-loaded PCL NPs. 3D images (**a**,**c**,**e**,**g**) and cross section analysis (**b**,**d**,**f**,**h**). The extract was successfully entrapped in both carriers. This observation was supported by analyzing the rutin-polymer complexes, which suggested that rutin-PLGA was the most stable molecular structure (\(\Delta\Sigma\approx-20\) kcal/mol). The rutin-PCL complex also showed considerable energy minimization (\(\Delta\Sigma\approx-18\) kcal/mol) compared to PLGA. The favorable energy stabilization can be attributed to the high encapsulation efficiency of the polymeric NPs.