Pathophysiologic changes in mucosal protein expression inflammatory bowel disease (IBD) may affect drug concentration in mucosal tissue making it highly relevant to drug dose at the site of action and subsequently for success of the therapy. Tissue samples from an experimental colitis rat model were mounted in Ussing chambers and intratissue concentrations of diverse compounds were quantified. Studies with healthy versus colitis tissue samples and respective microsomal fractions made it possible to assess the involvement of P-glycoprotein (P-gp) and cytochrome P450 3A (CYP3A) on tissue penetration behavior. P-gp-related efflux was slightly increased for colitis tissue. Metabolism studies exhibited higher tacrolimus and testosterone mucosal metabolism in inflamed tissue. However, similar metabolic activity was observed for healthy and colitis groups with equivalent CYP3A expression levels in respective microsome fractions. Severity of colitis as determined by myeloperoxidase activity was found to have linear correlation to changes in tacrolimus degradation (R2 = 0.8299). It is hypothesized that increased drug metabolism is dependent on the number of cells infiltrating inflamed tissue. A dominant contribution of immune-related cells to observed variations in mucosal drug metabolism has been determined. This observed pathophysiologic mechanism may have a significant influence on available drug concentrations at the inflammation site, thus modifying anti-inflammatory efficiency of the therapy.