## 3 Results and Discussion  
### Lipid Nanoparticle Characterization  
As shown in Table 2, unloaded and IDE-loaded lipid nanoparticles, prepared by the PIT method, showed small particle sizes (range: 23-42 nm), which were not significantly affected by IDE or VitE loading. PDI values were lower than 0.300, suggesting the presence of a homogeneous population of nanoparticles in the investigated samples. TEM analyses of the lipid nanocarriers under investigation showed roughly spherical nanoparticles (Figure 1). As this technique gave information only about the external surface of the nanocarrier, similar images were obtained for all nanoparticles. Therefore, we reported as an example in Figure 1 TEM pictures of only SLN IDE1 and NLC IDE1. Although all these lipid nanoparticles showed low \(\zeta\)-potential values (range: \(-1.56/-2.32\) mV), they proved stable after storage for two months at room temperature as no change of particle size and PDI values were observed (data not shown). In addition, we did not observe any sign of drug precipitation during storage. Similar low \(\zeta\)-potential vales were obtained in previous works on SLN with the same composition, and their good stability was attributed to their high PIT values [17; 20; 30]. IDE loading into SLN and NLC was determined as the maximum amount of drug that could be incorporated in the carriers leading to a clear colloidal system, as previously reported [17; 20; 30]. IDE loading into SLN and NLC was 1.5 and 2%, respectively. The higher IDE loading into NLC could be attributed to the presence of the oil component that could favor both IDE solubilization in the lipid matrix of the nanoparticles and a less ordered packing of the NLC core. DSC analyses of unloaded and IDE-loaded SLN and NLC pointed out different calorimetric behaviors of these nanocarriers (Figure 2). As already reported in previous studies, unloaded SLN showed a transition temperature (T\({}_{\rm m}\)) at 42 \({}^{\circ}\)C that was about 12 \({}^{\circ}\)C lower than that of the bulk lipid (cetyl palmitate), thus indicating the solid nature of the lipid in the nanocar matrix [31; 32; 33]. Unloaded SLN had a crystallinity index (CI) of 66%, which is in accordance with the results obtained in previous studies [12; 23; 34]. \begin{table}
\begin{tabular}{c c c c c} \hline \hline
**Code** & **Size \(\pm\) SD (nm)** & **PDI \(\pm\) SD** & \(\zeta\pm\) SD (mV)** & **PIT (\({}^{\circ}\)C)** \\ \hline SLN & 36.40 \(\pm\) 0.30 & 0.260 \(\pm\) 0.010 & \(-\)1.56 \(\pm\) 0.48 & 78 \\ SLN IDE1 & 23.55 \(\pm\) 1.08 & 0.266 \(\pm\) 0.023 & \(-\)2.02 \(\pm\) 0.54 & 69 \\ SLN IDE 1.5 & 42.33 \(\pm\) 0.29 & 0.230 \(\pm\)0.031 & \(-\)2.01 \(\pm\) 0.33 & 77 \\ NLC & 28.78 \(\pm\) 0.90 & 0.235 \(\pm\) 0.030 & \(-\)2.00 \(\pm\) 0.24 & 85 \\ NLC IDE1 & 26.76 \(\pm\) 0.33 & 0.178 \(\pm\) 0.041 & \(-\)1.99 \(\pm\) 0.33 & 78 \\ NLC IDE1.5 & 27.33 \(\pm\) 0.55 & 0.198 \(\pm\) 0.023 & \(-\)2.32 \(\pm\) 0.63 & 71 \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Mean size (size), polydispersity index (PDI), zeta potential (\(\zeta\)), and phase inversion temperature (PIT) of unloaded and IDE-loaded SLN and NLC. IDE = ideebenone, SLN = solid lipid nanoparticles, NLC = nanostructured lipid carriers. Values are expressed as mean \(\pm\) standard deviation (SD). Figure 1: Transmissions electron microscopy (TEM) images of SLN IDE1 (**a**) and NLC IDE1 (**b**). IDE = ideebenone, SLN = solid lipid nanoparticles, NLC = nanostructured lipid carriers. In the thermogram of unloaded NLC, we observed a T\({}_{\rm m}\) value lower than that of unloaded SLN that suggested a lesser crystalline structure of the nanocarrier core in the presence of the liquid lipid VitE. As expected, unloaded NLC had a CI (59%) lower than that of unloaded SLN. Loading 1% w/w IDE in these nanocarriers led to changes in the thermograms of the resulting lipid nanocarriers.