Bilosomes and Biloparticles for the Delivery of Lipophilic Drugs: A Preliminary Study

Abstract

In this study, bile acid-based vesicles and nanoparticles (i.e., bilosomes and biloparticles) are studied to improve the water solubility of lipophilic drugs. Ursodeoxycholic acid, sodium cholate, sodium taurocholate and budesonide were used as bile acids and model drugs, respectively. Bilosomes and biloparticles were prepared following standard protocols with minor changes, after a preformulation study. The obtained systems showed good encapsulation efficiency and dimensional stability. Particularly, for biloparticles, the increase in encapsulation efficiency followed the order ursodeoxycholic acid < sodium cholate < sodium taurocholate. The in vitro release of budesonide from both bilosytems was performed by means of dialysis using either a nylon membrane or a portion of Wistar rat small intestine and two receiving solutions (i.e., simulated gastric and intestinal fluids). Both in gastric and intestinal fluid, budesonide was released from bilosystems more slowly than the reference solution, while biloparticles showed a significant improvement in the passage of budesonide into aqueous solution. Immunofluorescence experiments indicated that ursodeoxycholic acid bilosomes containing budesonide are effective in reducing the inflammatory response induced by glucose oxidase stimuli and counteract ox-inflammatory damage within intestinal cells.

Keywords: NLC; SLN; bile acids; drug solubility; immunofluorescence; inflammasome; nanoparticles; nanotechnology; nanovesicles; poorly water-soluble drugs.

Figure 1

Cryo-TEM images of Bilos_PCU without (a) and with (b) budesonide prepared via a warm film hydration method followed by sonication (FhWS).

Figure 2

Dimensional parameters of bilosomes with (squares) and without (plain) budesonide, expressed as average diameter (histograms) and dispersity index (circles). Values are the mean of 3 batches ± s.d.

Figure 3

Cryo-TEM images of BilopS_UB (a) and BilopN_UB (b).

Figure 4

Mean diameter (Z-Average, histograms) and polydispersity (PdI, circles) of BilopS (dark color) and BilopN (light color) containing different bile acids, as determined via PCS. Each value is the average of 5 batches ± s.d.

Figure 5

Budesonide release kinetics in FaSSGF (a,c,e) and FaSSIF (b,d,f) from Bilos (a,b), BilopS (c,d) and BilopN (e,f). Bilos_PCUB, BilopS_UB and BilopN_UB: closed circle; Bilos_PCCB, BilopS_CB and BilopN_CB: closed square; Bilos_PCTB, BilopS_TB and BilopN_TB: closed diamond; For comparison, the releases from Susp B in FaSSGF/FaSSIF (open circle) and Sol B (open square) are also reported. Each datum is the average of 2 experiments ± s.d.

Figure 6

Budesonide releases kinetics through rat small intestine from Bilos (a), BilopS (b) and BilopN (c) suspended in PBS/FaSSIF (50:50 v:v). Bilos_PCUB, BilopS_UB and BilopN_UB: closed circle; Bilos_PCCB, BilopS_CB and BilopN_CB: closed square; Bilos_PCTB, BilopS_TB and BilopN_TB: closed diamond; The release from FaSSIF Susp B (open circle) is also reported for comparison. Each value is the mean of 3 experiments ± s.d.

Figure 7

MTT assay on CaCo2 cells treated for 24 h with budesonide formulations (Sol B, Bilos_PCUB) at different doses ranging from 1 to 20 μg/mL. Cell viability is presented as percentage with respect to untreated control cells. Data are given as mean ± SD, representative of three independent experiments with at least three technical replicates each time. A value of 80% (red dotted line) was considered the reference accepted cell viability.

Figure 8

Immunofluorescence staining for NLRP3 (green) and ASC (red) on CaCo2 cells treated with formulations containing budesonide and exposed to 0.5 U/mL of GO for 1 h. Blue staining (DAPI) represents nuclei. Images were taken at 40 × magnification (scale bar = 40 μm), and the fluorescent signal was quantified using ImageJ software. Data are the results of the averages of at least three different experiments, obtained via 2-way ANOVA followed by Tukey’s post hoc comparison test. ^$ p < 0.05; ** p < 0.01; *** p < 0.005; ****^,#### p < 0.0001 (* with respect to Ctrl and # with respect to Ctrl + GO).

Figure 9

Released levels of IL- 1β in media of CaCo2 cells pre-treated with formulations containing budesonide, exposed to 0.5 U/mL of GO for 1 h and collected after 3 h (T3). Data are the results of the averages of at least three different experiments, obtained via 2-way ANOVA followed by Tukey’s post hoc comparison test. ^$ p < 0.05; ^## p < 0.01; *** p < 0.005; ^#### p < 0.0001 (* with respect to Ctrl and # with respect to Ctrl + GO).