Solid Lipid Nanoparticles as Formulative Strategy to Increase Oral Permeation of a Molecule Active in Multidrug-Resistant Tuberculosis Management

Abstract

The role of mycobacterial efflux pumps in drug-resistant tuberculosis has been widely reported. Recently, a new compound, named SS13, has been synthesized, and its activity as a potential efflux inhibitor has been demonstrated. In this work, the chemical-physical properties of the SS13 were investigated; furthermore, a formulative study aimed to develop a formulation suitable for oral administration was performed. SS13 shows nonintrinsic antitubercular activity, but it increases the antitubercular activity of all the tested drugs on several strains. SS13 is insoluble in different simulated gastrointestinal media; thus, its oral absorption could be limited. Solid lipid nanoparticles (SLNs) were, therefore, developed by using two different lipids, Witepsol and/or Gelucire. Nanoparticles, having a particle size (range of 200-450 nm with regards to the formulation composition) suitable for intestinal absorption, are able to load SS13 and to improve its permeation through the intestinal mucosa compared to the pure compound. The cytotoxicity is influenced by the concentration of nanoparticles administered. These promising results support the potential application of these nanocarriers for increasing the oral permeation of SS13 in multidrug-resistant tuberculosis management.

Keywords: Gelucire@; Witepsol@; antitubercular activity; drug-resistant tuberculosis; oral permeation; solid lipid nanoparticles.

Figure 1

Schematic representation of plates used for ex vivo permeation test. The test was carried out using three different plates, one for each time point. Blue box reports the arrangement of intestinal mucosa and samples over each well. All samples were tested in triplicate (n = 3).

Figure 2

TEM images of SLN-W (A) and SLN-Wb (B).

Figure 3

TEM images of SLN-G (A) and SLN-Gb (B).

Figure 4

Topographic images of SLN-W (A) and SLN-Wb (B). Three-dimensional views of 20 × 20 µm scan of SLN-W (C) and SLN-Wb (D).

Figure 5

Topographic images of SLN-G (A) and SLN-Gb (B). Three-dimensional views of 20 × 20 µm scan of SLN-G (C) and SLN-Gb (D).

Figure 6

FTIR spectra of SS13, SLN-Wb, SLN-W and physical mixture of SLN-Wb and SS13.

Figure 7

FTIR spectra of SS13, SLN-Gb, SLN-G and physical mixture of SLNGb and SS13.

Figure 8

XRD pattern of SLNs prepared with Witepsol E85 (W) (A,B) and with the mixture of W/G (C,D). Magnification of XRD pattern of SLNs with W (B) and with W/G (D).

Figure 9

Influence of storage (at 4 and 25 °C) on the particle size and PDI of SLN-Wb and SLN-W. Particle size: SLN-Wb at 25 °C, 0 days vs. 15 and 30 days (p < 0.05); SLN-Wb at 4 °C, 0 days vs. 7, 15 and 30 days (p < 0.05); SLN-W at 25 °C, 0 days vs. 7, 15 and 30 days (p < 0.05); SLN-W at 4 °C, 0 days vs. 1, 7, 15 and 30 days (p < 0.05). PDI: SLN-Wb at 25 °C, 0 days vs. 7, 15 and 30 days (p < 0.05); SLN-Wb at 4 °C, 0 days vs. 1, 7, 15 and 30 days (p < 0.05); SLN-W at 25 °C, 0 days vs. 30 days (p < 0.05); SLN-W at 4 °C, 0 days vs. 7, 15 and 30 days (p < 0.05).

Figure 10

Influence of storage (at 4 and 25 °C) on the particle size and PDI of SLN-Gb and SLN-G. Particle size: SLN-G at 25 °C, 0 days vs. 1, 7, 15 and 30 days (p < 0.05); SLN-G at 4 °C, 0 days vs. 1, 7, 15 and 30 days (p < 0.05). PDI: SLN-Gb at 25 °C, 0 days vs. 1 day (p < 0.05); SLN-Gb at 4 °C, 0 days vs. 1, 7, 15 and 30 days (p < 0.05); SLN-G at 25 °C, 0 days vs. 1, 7, 15 and 30 days (p < 0.05); SLN-G at 4 °C, 0 days vs. 1, 7, 15 and 30 days (p < 0.05).

Figure 11

In vitro release kinetics of SS13 from SLN-W and SLN-G compared to the dissolution of free molecule. Data are reported as mean ± SD (n = 3).

Figure 12

Ex vivo distribution of SS13 during the permeation test (30, 60 and 120 min) through the intestinal mucosa (n = 3). * p < 0.05, SLN-G on mucosa 120 min vs. SLN-G on mucosa 60 min; ⁺ p < 0.05, SLN-G inside mucosa 120 min vs. SLN-G inside mucosa 30 and 60 min; ^° p < 0.05, SS13 on mucosa 120 min vs. SLN-W on mucosa 120 and SLN-G on mucosa 120 min; ^& p < 0.05, SLN-G inside mucosa 120 min vs. SLN-W inside mucosa 120 min and SS13 inside mucosa 120 min; ^# p < 0.05, SS13 permeated 120 min vs. SLN-W permeated 120 min and SLN-G permeated 120 min; ^§ p < 0.05, SLN-W permeated 120 min vs. SLN-G permeated 120 min.

Figure 13

Effect of increasing concentrations (0.5, 1 and 3 µM) of SS13, SLN-Wb, SLN-W, SLN-Gb and SLN-G on CACO-2 cells at 6 and 24 h of exposure. Data are reported as mean ± SD (n = 3). * p < 0.05 vs. Control; ^& p < 0.05 vs. SS13 0.5 and 1 µM; ^# p < 0.05 vs. SLN-W 0.5 and µM; ^§ p < 0.05 vs. SLN-Gb 3 µM and SLN-G 0.5 µM; ^φ p < 0.05 vs. SLN-G 0.5 and 1 µM; ^θ p < 0.05 vs. SLN-W 0.5, 1 and 3 µM.