Annatto Oil Loaded Nanostructured Lipid Carriers: A Potential New Treatment for Cutaneous Leishmaniasis

Abstract

Annatto (Bixa orellana L.) is extensively used as food pigment worldwide. Recently, several studies have found it to have healing and antioxidant properties, as well as effective action against leishmaniasis. Therefore, the purpose of this study was to incorporate the oil obtained from annatto seeds into a nanostructured lipid carrier (NLC) and evaluate its physicochemical properties and biological activity against Leishmania major. Nanoparticles were prepared by the fusion-emulsification and ultrasonication method, with the components Synperonic™ PE (PL) as the surfactant, cetyl palmitate (CP) or myristyl myristate (MM) as solid lipids, annatto oil (AO) (2% and 4%, w/w) as liquid lipid and active ingredient, and ultra-pure water. Physicochemical and biological characterizations were carried out to describe the NLCs, including particle size, polydispersity index (PDI), and zeta potential (ZP) by dynamic light scattering (DLS), encapsulation efficiency (EE%), thermal behavior, X-ray diffraction (XRD), transmission electron microscopy (TEM), Electron Paramagnetic Resonance (EPR), cytotoxicity on BALB/c 3T3 fibroblasts and immortalized human keratinocyte cells, and anti-leishmaniasis activity in vitro. Nanoparticles presented an average diameter of ~200 nm (confirmed by TEM results), a PDI of less than 0.30, ZP between −12.6 and −31.2 mV, and more than 50% of AO encapsulated in NLCs. Thermal analyses demonstrated that the systems were stable at high temperatures with a decrease in crystalline structure due to the presence of AOs (confirmed by XRD). In vitro, the anti-leishmania test displayed good activity in encapsulating AO against L. major. The results indicate that the oily fraction of Bixa orellana L. in NLC systems should be evaluated as a potential therapeutic agent against leishmaniasis.

Hybrid Lipid/Clay Carrier Systems Containing Annatto Oil for Topical Formulations

“…In addition, the results showed that the long and short spacing of the crystalline lattice were modified in the hybrid systems when compared to the lipid raw materials (CP and MM), where a reduction in peak intensity was observed, and this can be related to the reduction in the degree of crystallinity of the lipid in its nanoparticulate state, resulting in a more amorphous system ( Table 2 ). Similar results were observed by Ferreira et al, [ 42 ] without the presence of laponite.…”

“…Figure 2 shows the profile for the loss of mass during heating of the components of the systems ( Figure 2 a) and hybrid systems ( Figure 2 b,c). Figure 2 a shows similar results for loss of mass of the crystalline solids (PL bulk , MM bulk , and CP bulk ) in a single event, and these results agree with Ferreira et al, 2021 [ 42 ]. Laponite, in turn, had its mass reduced by 7.6% between 33 °C and 79 °C due to the loss of adsorption water and 4.3% slowly over the maximum heating period, due to the loss of interlamellar clay water.…”

“…No change in pH values was observed for samples with different liquid lipid concentrations ( Figure 1 b), with media values between 8.52 and 8.61 for 2 and 4% of AO, respectively. A recent study by Ferreira et al, 2021 [ 42 ] showed lower pH values for NLCs prepared with the same compounds; however, without LAP, between 5.0–6.0 for all samples. In fact, the presence of LAP provides an increase in pH due to the high pH of the raw clay.…”

“…As observed in Figure 1 b, the high concentration of AO can be responsible for forming hybrid systems with the most negative charges. The results of size, polydispersity, and zeta potential point to the presence of LAP as being an essential factor in maintaining the stability of the systems over ten weeks, mainly for formulations with MM since the samples without clay showed phase separation 48 h after their preparation, according Ferreira et al, 2021 [ 42 ]. The surface charge of the particles is one of the factors that determine physical stability.…”

“…Previous studies from our group carried out using different techniques such as DSC, XRD, and EPR helped to elucidate the arrangements of the crystalline structures, and the degree of order of the lipids present in the lipid-based nanoparticles, with and without AO. It was demonstrated that the presence of AO is one of the main factors responsible for structural modification in this lipid nanoparticle [ 42 ].…”