## 3 Results and Discussion
### Assessment of CTAB and DDAB-Decorated SLNs Cytotoxicity  
Different cell lines were exposed to different concentrations of CTAB-SLNs and DDAB-SLNs, as shown in Table 2. The wt%, as concentration unit, is commonly used as a parameter in nanoparticles production. As these two surfactants have different molecular weights, Table 2 also shows the concentration (in \(\upmu\)M) of surfactants in SLNs. Figure 1 shows the effect of exposure to CTAB-SLNs and DDAB-SLNs in the different cell lines. CTAB-SLNs produced a drastic decrease of cell viability to values close to zero in all cell lines (Figure 1), which is translated by the low IC\({}_{50}\) (Table 3) calculated for 24 and 48 h of exposure. For all cell lines, IC\({}_{50}\) was \(<\)10 \(\upmu\)g/mL in both exposure times. In a study using Y-79 cells, [PERSON] et al. [4] demonstrated that the concentration of CTAB compromises the cell viability at concentrations above 0.5% (wt%) [4]. However, CTAB concentration is critical for the mean particle size. Indeed, that the mean particle size decreases with the increase of CTAB concentration (\(239.5\pm 0.61\) nm to \(144.7\pm 1.61\) nm, at 0.25 and 1.0% (wt%), respectively [4]). The increase of CTAB concentration also increased the ZP of SLNs, from +24 mV (at 0.25%) to +48 mV (at 1.0%) favoring the particles' stability and their interactions with the cells [4]. \begin{table}
\begin{tabular}{c c c c c} \hline \hline \multirow{2}{*}{**Formulation**} & \multicolumn{2}{c}{**DLS**} & \multicolumn{2}{c}{**LD**} \\ \cline{2-5}  & **Z-Ave (nm)** & **PI** & **ZP (mV)** & **d50 (nm)** \\ \hline
**CTAB-SLNs** & \(135.1\pm 0.22\) & \(0.196\pm 0.02\) & \(+28.20\pm 1.33\) & \(118.3\pm 0.04\) \\
**DDAB-SLNs** & \(134.2\pm 1.12\) & \(0.179\pm 0.07\) & \(+28.20\pm 2.29\) & \(119.8\pm 0.01\) \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Mean particle size (Z-Ave), polydispersity index (PI), and zeta potential (ZP) of solid lipid nanoparticles (SLNs), recorded by dynamic light scattering (DLS), and d50% measured by laser diffraction (LD). Results are indicated as mean \(\pm\) S.D. (n = 3). \begin{table}
\begin{tabular}{c c c c c c} \hline \hline
**CTAB (*)** & \(\upmu\)g/mL & **66.0** & **165.0** & **330.0** & **660.0** \\  & \(\upmu\)M & 181.1 & 452.7 & 905.5 & 1810.9 \\
**DDAB (*)** & \(\upmu\)g/mL & **66.0** & **165.0** & **330.0** & **660.0** \\  & \(\upmu\)M & 104.6 & 261.5 & 523.0 & 1046.0 \\ \hline \hline \end{tabular}
\end{table} 
Table 2: CTAB (cetyltrimethylammonium bromide) and DDAB (dimethyldioctadecylammonium bromide) concentrations of each test solution (in \(\upmu\)g/mL and in \(\upmu\)M) used in this study. See methods for details (wt% of CTAB and DDABB (g/100 mL) in formulation was 0.5). The results from cytotoxicity and IC\({}_{50}\) suggest that the choice of surfactant is relevant when designing formulations aiming the direct contact with the cells (i.e., intravenous injections or mucosal application), and thus caution has to be taken concerning the final concentration of CTAB. [PERSON] et al. produced gold nanoparticles (Au-NRs or nanorods) stabilized with CTAB [29]. The cytotoxic effect of both Au-NRs and CTAB was assessed against keratinocytes. The authors reported that the inherent toxicity of Au-NRs was mainly due to the presence of the cationic surfactant. CTAB decreased the cell viability at concentrations as low as 10 \(\upmu\)M (or 3.64 \(\upmu\)g/mL) [29], a value within the same range of concentrations here reported for CTAB-SLNs (Figure 1, Table 2). For some time, the toxicity of Au-NRs was attributed to the particle shape, but [PERSON] et al. [30] described that  
Figure 1: Cell viability profiles of CTAB-SLNs and DDAB-SLNs in Caco-2 **(A)**, HepG2 **(B)**, MCF-7 **(C)**, SV-80 **(D)**, and Y-79 **(E)** cells. Cells were exposed to CTAB-SLNs and DDAB-SLNs test solution, indicated in \(\upmu\)g/mL of surfactant in test solution (66.0, 165.0, 330.0, and 660.0), for 24 h. Results, expressed as % of control (non-exposed cells), indicate the mean \(\pm\) SD of n = 3 independent experiments (see methods).