## 4 Discussion  
The present study aimed to investigate the physicochemical characterization, stability, and release kinetics of nanostructured lipid carriers (NLCs) and conventional emulsions (CEs) loaded with essential oils (EOs), as well as the efficacy and duration of the insect-repellent action of the NLC EO. The characterization of droplet size is crucial in understanding the physical stability and performance of emulsions. The droplet sizes of the CEs and NLCs, both with and without EO incorporation, were characterized to assess the impact of EO on the dispersed phase. Interestingly, the droplet size distribution of the CE did not differ significantly from that of the CE EO, showing that the incorporation of EO did not impact the size of emulsion droplets. This suggests that the formulation process effectively stabilized the emulsion, preventing coalescence or Ostwald ripening, even in the presence of EO. The substantial difference in droplet size between the CE (18.46 \(\pm\) 1.78 \(\upmu\)m) and NLC (136 \(\pm\) 10.7 nm) has important implications for stability and application effectiveness. Smaller droplet sizes in the NLC contribute to enhanced physical stability by minimizing the risk of phase separation and sedimentation, which is often observed in conventional emulsions [25]. Furthermore, the nanoscale size of the NLC allows for a more homogeneous and prolonged distribution of essential oils on the skin, enhancing the sustained release and overall effectiveness of the repellent action [26]. This size advantage ensures that NLC formulations remain stable and effective over time, even under varying environmental conditions. The addition of EO did not alter the droplet size distribution or \(\zeta\)-potential of the NLC, indicating the stability of these formulations following incorporation. The consistent size distribution and surface charge suggest that the lipid-based matrix effectively incorporated the EO, maintaining the integrity of the NLC structure. Furthermore, the \(\zeta\)-potential, an indicator of the stability of colloidal dispersions, remained consistent in both the NLC and NLC EO, even after centrifugation and accelerated aging. These findings suggest that the stability of the NLC formulation was maintained despite the incorporation of EO, indicating its potential for long-term stability. These findings are in accordance with a previous study from our group investigating the incorporation of lavender essential oil in conventional emulsions and lipid nanocarriers. In that study, the mean droplet size of the conventional emulsions was 16.46 \(\pm\) 5.73 \(\upmu\)m, while the nanoemulsions exhibited a mean droplet size of 103.04 \(\pm\) 12.25 nm and a \(\zeta\)-potential of \(-\)60.5 \(\pm\) 2.8 mV. Notably, the incorporation of lavender essential oil did not significantly impact the physicochemical properties of the formulations [23]. Gas chromatography mass spectrometry (GC-MS) analysis was conducted to identify the components of the EO incorporated in the CE EO and NLC EO. Eucalyptustold emerged as the predominant constituent, with other significant ingredients being eugenol, linalool, and geraniol. Interestingly, the composition of the EO differed between the CE EO and NLC EO, highlighting the influence of formulation type on the encapsulation and retention of EO compounds [23]. This underscores the importance of formulation optimization to achieve the desired EO loading and mosquito-repellent efficacy [27]. Stability studies were conducted to assess the long-term stability of the CE EO and NLC EO formulations. Centrifugation tests demonstrated that both formulations remained stable, with no signs of phase separation or changes in droplet size distribution. Additionally, accelerated aging tests and storage at 4 \({}^{\circ}\)C over 90 days revealed no significant alterations in droplet size or \(\zeta\)-potential (for NLC EO), indicating the robustness of these formulations under various conditions.