The intestinal epithelium represents the major barrier to absorption of orally administered drugs and peptides into the systemic circulation. Entry of molecules through the paracellular pathway is restricted by tight junctions(tj). We have previously reported that these structures can be modulated by Zonula Occludens Toxin (Zot). Aim of the study was to establish whether the permeabilizing effect of the toxin leads to increased intestinal absorption of macromolecules normally not absorbed when administered orally. To evaluate the transepithelial transport of either insulin 10-11M or IgG 156.25 ng, alone or in the presence of 1.1×10-10M Zot, in vitro and in vivo experiments were performed on rabbit intestine. To evaluate the bioactivity of insulin after enteral co-administration with Zot, acute type 1 diabetic male BB/Wor rats were orally treated with insulin, with or without Zot, and the blood glucose levels of the rats were serially measured. Zot reversibly increases rabbit intestinal permeability to insulin by 72% (p=0.034) and immunoglobulins by 52% (p=0.04), in vitro. When tested in vivo, Zot induced a 10-fold increase of insulin absorption in both the rabbit jejunum and ileum, whereas no substantial changes were detected in the colon. Similar results were obtained with immunoglobulins, whereby Zot induced 2-fold and 6-fold increases of IgG absorption in the jejunum and ileum, respectively. In diabetic rats, bioavailability of oral insulin coadministered with Zot was sufficient to lower serum glucose concentrations to levels comparable to those obtained after parenteral injection of the hormone. The survival time of diabetic animals chronically treated with oral insulin+Zot was comparable to that observed in parenterally treated rats. These studies offer a useful strategy for the oral delivery of drugs and proteins normally not absorbed through the gut.