Increased intestinal absorption of medium-sized aqueous probes has been found in patients with a variety of disorders. We studied the physiologic control mechanisms, intestinal regions, and effects of lumenal factors on the intestinal absorption of polyethylene glycol (PEG) 900 in the rat in vivo and in rabbit brush border membrane vesicles (BBMVs). The kinetics of PEG 900 intestinal absorption were compatible with simple passive diffusion. Because transport across BBMVs was minimal, we concluded that transport of PEG 900 is mostly through the paracellular tight junctions. Absorption was highest in the midcolon (104.3 +/- 9.5 mumol/100 mg protein per hour vs 9.1 +/- 1.2 mumol/100 mg protein per hour in the jejunum). Absorption was decreased by higher lumenal osmolarity (greater than 400 mOsm/L) after the additions of 2.5 to 5.0 mmol/L chenodeoxycholate or 2.5 mmol/L lysolecithin, or at higher lumenal flow rates (greater than 1 ml/minute), higher lumenal pressure (7.5 cm H2O),or higher lumenal pH (8.0). Lipid solubility of PEG 900 was less than 0.00079%. Under all experimental conditions, PEG net absorption followed changes in water transport. When water transport changed from absorption to secretion, PEG absorption decreased. When water absorption increased, PEG 900 absorption increased in parallel. We conclude that PEG 900 is absorbed by passive diffusion that is modulated by solvent drag and is maximal in the midcolon. Transport directly across cell membranes is mimimal, but overall PEG 900 permeability is closely linked to water absorption by solvent drag and takes place primarily through the paracellular junctions. We propose that these features and mechanisms of PEG 900 transport make PEG 900 a suitable probe molecule for studying intestinal permeability changes.