*3.1. Preparation and Characterization of the Control and DOX-Loaded SLPs*

The SLPs were synthesized using the melt emulsification method described in the Materials and Methods section (Figure S1). The hydrophobic organic matrix of the SLPs consisted of carnauba wax containing iron oxide nanoparticles. The wax matrix chosen is a natural complex wax obtained from a Brazilian palm tree widely used in food [36], pharmaceutical applications [8], and other technologies [37]. The SLPs were labeled with a fluorescent dye (3,30 -dioctadecyloxacarbocyanine perchlorate (DiO)), which was used for detection and imaging purposes. DiO has been widely used due to its biocompatibility and great stability in living and fixed tissues [38]. This dye allows particle localization using fluorescence technologies in processed cells and tissues [39]. The encapsulation of iron nanoparticles inside the wax matrix allowed us to know the biodistribution of SLPs in in vivo studies. Concentrations of iron were measured in different tissues of mice by the ICP technique. In this study, particles were loaded with 20% (*w*/*w*) of DOX to study if a drug delivery system can decrease the toxicity and improve the antineoplastic and antimetastatic effects of traditional chemotherapy (Figure 1a).

**Figure 1.** Solid lipid particle (SLP) characterization. (**a**) Composition of the SLPs. (**b**) Hydrodynamic particle size (right) and ζ potential (left) of the control SLPs (yellow) and SLPs loaded with doxorubicin (DOX) (red). (**c**) Representative TEM micrographs of SLPs. The Fe3O<sup>4</sup> nanoparticles are visible as dark structures inside the composite (inset).

Upon SLPs-DOX synthesis, particle characterization revealed a DOX and Fe3O<sup>4</sup> nanoparticle encapsulation efficiency of 86.5% ± 1.4 and 99%, respectively. The SLPs presented a hydrodynamic diameter of ca. 200 nm and an estimated ζ potential value of −10 and +23 mV for the control SLPs and SLPs-DOX, respectively. This charge change can be attributed to the positive charge of DOX at physiological pH (Figure 1b). The values of the polydispersity index were 0.36 ± 0.04 for the control SLPs and 0.16 ± 0.1 for the SLPs-DOX (Figure S2). Electron microscopy images show the rounded shape of particles. This technique also allowed the identification of the iron oxide nanoparticles inside the wax matrix (Figure 1c, dark hypointense spots).
