*2.2. Synthesis of DEX-Loaded LPNCs, Fluorescent-Loaded LPNCs, and Non-Vesiculated Control Solutions*

LPNCs were produced by the Emulsion Solvent Evaporation method invented by Fessi et al. [17]. In this method, an organic phase composed of different ratios of EA and ET, EC (2.33%, *w*/*w*), MCT (0.2%, *w*/*w*), DEX (1%, *w*/*w*), and different concentrations of Span 60, were emulsified in an aqueous phase composed of different concentrations of Tween 80 and benzalkonium chloride (preservative). Emulsification was carried out under probe sonication (amplitude of 40%, 5 min; energy, 7000 W; frequency 23.88 kHz) using a UP400st ultrasonic device (Hielscher Ultrasonics, Germany), to reduce the size of the emulsion droplet. After formation of the emulsion, the system evaporated the organic solvents under vacuum in a rotary evaporator, which led to polymer precipitation and lipomer formation. The whole process was performed at room temperature.

A solution of DEX was prepared in a 1% hydroethanolic solution (90:10 *v*/*v*, water:ethanol) for use as a non-encapsulated DEX control (FREE-DEX). LRB- and C6-loaded LPNCs (LRB-C6-LPNCs) were manufactured with the same composition as previously described, but without DEX. LRB was added to the lipid nucleus in a mole-to-mole ratio of MCT:LRB of 1:1000, according to Lymberopoulos et al. [18]. In addition, 0.1% *w*/*w* C6 was incorporated into the organic phase (EA:ET), simulating the encapsulation of a hydrophobic active ingredient. Fluorochrome control aqueous solutions used in confocal microscopy were prepared with 3% Tween 80 of LRB and C6, at the same concentration as in the LRB-C6-LPNCs, for use as a control for non-vehicle fluorophores (FREE-LRB-C6).
