The columnar epithelium of the in vitro rat intestine was separated from the intestinal musculature by treatment with icecold edetic acid solution. The in vitro absorption kinetics through the denuded intestinal musculature for salicylate ion and acetanilide were followed in pH 7.4 sodium phosphate and potassium phosphate buffers, as well as in an isotonic sodium chloride solution where the effect of rinsing with Mg+2 and Ca+2 was also studied. Differences in directional transfer rates, as noted in previous studies with the untreated intestinal membrane, were not found. The transfer rates through the muscle layer were found to be at least twice as high as those found in the untreated membrane. As noted previously, nonionized acetanilide transfer rates were still approximately twice those seen for salicylate ion.