Preliminary Assessment of Polysaccharide-Based Emulgels Containing Delta-Aminolevulinic Acid for Oral Lichen planus Treatment

Abstract

Photodynamic therapy using delta-aminolevulinic acid is considered a promising option in the treatment of oral lichen planus. In the present work, three emulgel compositions prepared from natural polysaccharide gums, tragacanth, xanthan and gellan, were preliminarily tested for oromucosal delivery of delta-aminolevulinic acid. Apart from cytotoxicity studies in two gingival cell lines, the precise goal was to investigate whether the presence of the drug altered the rheological and mucoadhesive behavior of applied gelling agents and to examine how dilution with saliva fluid influenced the retention of the designed emulgels by oromucosal tissue. Ex vivo mucoadhesive studies revealed that a combination of xanthan and gellan gum enhanced carrier retention by buccal tissue even upon dilution with the saliva. In turn, the incorporation of delta-aminolevulinic acid favored interactions with mucosal tissue, particularly formulations comprised of tragacanth. The designed preparations had no significant impact on the cell viability after a 24 h incubation in the tested concentration range. Cytotoxicity studies demonstrated that tragacanth-based and gellan/xanthan-based emulgels might exert a protective effect on the metabolic activity of human gingival fibroblasts and keratinocytes. Overall, the presented data show the potential of designed emulgels as oromucosal platforms for delta-aminolevulinic acid delivery.

Keywords: delta-aminolevulinic acid; emulgel; gellan gum; oral lichen planus; oromucosal delivery; tragacanth; xanthan.

Figure 1

Rheograms (A,B) and plots of viscosity vs. shear rate (C,D) of emulgels containing ALA (A,C) and corresponding placebo formulations (B,D) diluted with simulated saliva fluid at the weight ratio of 1:0.5 assessed at 37 ± 1 °C (mean ± SD, n = 3).

Figure 2

In vitro mucoretention profile of ALA-loaded emulgels (F1–F3) (A,C,E) and placebo (P1–P3) (B,D,F) expressed as % of weight of undiluted emulgel or formulation diluted with SSF in the ratio of 1:0.25 or 1:0.5 that remained on the porcine buccal mucosa; water was used as control (mean ± SD, n = 3).

Figure 3

MTT viability (expressed as % of control) of gingival fibroblasts (A,C) or gingival keratinocytes (B,D) after 4 h (A,B) or 24 h (C,D) incubation with emulgels containing delta-aminolevulinic acid (F1–F3) and corresponding placebo formulations (P1–P3) in concentration range 0.01–1.0 mg/mL in comparison to untreated cells (100%) and pure ALA (blue line) (mean ± S.D.; n = 4).

Figure 4

Intensity of luminescence (RLU)) displaying the mitochondrial membrane potential (ΔΨM) of gingival fibroblasts (A) or keratinocytes (B) after 24 h incubation with 0.1 or 0.3 mg/mL of drug-free or ALA-loaded emulgels measured by JC-1 assay in comparison to untreated cells (blue line); RLUs of cells treated with pure drug (ALA) at concentration corresponding to those encapsulated in 0.3 mg/mL drug-loaded formulations were 8832 ± 954 and 8644 ± 1375 for fibroblasts and keratinocytes, respectively (mean ± S.D.; n = 4).

Figure 5

Evaluation of apoptosis measured by intensity of luminescence (A,B) and necrosis level (C,D) of gingival fibroblasts (A,C) or gingival keratinocytes (B,D) after 24 h incubation with emulgels containing delta-aminolevulinic acid (F1–F3) and corresponding placebo formulations (P1–P3) in concentrations 0.1 and 0.3 mg/mL in comparison to untreated cells (blue line); for the positive control (50 mM H₂O₂), RLU was 964,542 ± 32,319 and RFU was 35,750 ± 2511; for pure drug (ALA) RLU was 6585 ± 293 and RFU was 2513 ± 350 (mean ± S.D.; n = 4).

Figure 6

The scheme of the mucoretention test using thermostated inclined steel plate (modified according to [13]).