The effect of varying mucosal pH (5, 7.4 and 8) on aflatoxin B1 (AFB1) transmural transfer was studied in the everted rat jejunum. The viability of the preparation was evaluated by: light microscopy; rate of glucose uptake by tissue from the mucosal fluid; content and distribution of water and electrolytes in the tissue; oxygen uptake by the tissue; and the intestinal permeability to [14C]sucrose. These parameters revealed the structural and functional integrity of the preparation, which was not affected by incubation in the presence of AFB1 or at different pH values in the mucosal fluid. The results obtained show that AFB1 was transferred to the serosal fluid and that the cumulative transfer was greater at pH 5 than at pH 7.4 or 8. In addition, AFB1 clearance, calculated from the slope of the plot of the cumulative amount of AFB1 transferred per unit of mucosal concentration vs. incubation time, indicated a very rapid accumulation by tissue and transfer of AFB1 to the serosal fluid. This process was also faster at pH 5. The integrity of the AFB1 molecule during incubation was shown by NMR spectra. The lack of metabolization of AFB1 during its translocation was demonstrated by TLC of the serosal fluid.