N-TRIMETHYL CHITOSAN CHLORIDE AS A POTENTIAL ABSORPTION ENHANCER ACROSS MUCOSAL SURFACES - IN-VITRO EVALUATION IN INTESTINAL EPITHELIAL-CELLS (CACO-2)

Purpose. Previous studies have established that chitosan hydrochloride and glutamate are potent absorption enhancers for large hydrophilic c ompounds across mucosal surfaces. However, these compounds lack solubi lity at neutral pH values. A partially quaternized and well-soluble de rivative of chitosan, N-trimethyl chitosan chloride, was synthesized a nd the effects of this polymer on the transepithelial electrical resis tance and permeability of intestinal epithelial cells were investigate d in vitro. Methods, N-trimethyl chitosan chloride was synthesized by reductive methylation and characterized with NMR. The effect of this p olymer (1.0-2.5% w/v) on the transepithelial electrical resistance of intestinal epithelial cells, using Caco-2 cell monolayers, was investi gated. Permeation of the hydrophilic model compounds [C-14]-mannitol ( MW 182.2), FITC-Dextran (MW 4400) and the peptide drug buserelin (MW 1 299.5), in the presence of N-trimethyl chitosan chloride (1.5-2.5% w/v ), was followed for 3 hours. The transport process of the fluorescent marker, FITC-Dextran 4400, across the cell monolayers was visualised w ith confocal laser scanning microscopy. Viability of the cells was che cked with the trypan blue exclusion technique. Results. N-trimethyl ch itosan chloride was found to be a perfectly water-soluble, partially q uaternized (about 12%) derivative of chitosan. This polymer (1.5-2.5% w/v) caused a pronounced and immediate reduction (25-85%) in the trans epithelial electrical resistance of Caco-2 cells. Large increases in t he transport rate of [C-14]-mannitol (32-60 fold), FITC-Dextran 4400 ( 167-373 fold) and buserelin (28-73 fold) were demonstrated. Confocal l aser scanning microscopy confirmed that N-trimethyl chitosan chloride opens the tight junctions of intestinal epithelial cells to allow incr eased transport of hydrophilic compounds through the paracellular tran sport pathway. No deleterious effects to the cells could be demonstrat ed with trypan blue. Conclusions. The potential use of N-trimethyl chi tosan chloride as an absorption enhancer across mucosal surfaces could be an important contribution towards the development of effective del ivery systems for hydrophilic drugs.