Goblet cell mucin (GCM) has been purified for the first time from mucosal scrapings of human small intestine. Proteolytic enzymes and organic solvents were avoided during the isolation procedure. The mucin was purified by Sepharose 4B and 2B column chromatography of high-speed supernatant fractions. The most purified fraction was compared with rat intestinal GCM. The two were similar with respect to chemical composition, antigenic features, and polyacrylamide disc gel electrophoresis. The major chemical differences included a higher hexosamine-fucose and hexosamine-sialic acid ratio in human mucin. The two mucins showed strong concentration dependence in sedimentation velocity studies. Human mucin at a concentration of 0.2 to 1.5 mg protein per millilitre gave multiple associated peaks with variable So values (10.8-36.6). Rat mucin, in contrast, gave a constant (although polydisperse) pattern with So = 15.15. To explore these differences both mucins were stained with periodic acid - Schiff reagent and subjected to band ultracentrifugation at concentrations of 0.6-1.9 mug protein per millilitre. At this low concentration, rat mucin did not change in its sedimentation characteristics. In contrast, human GCM produced a single peak with So = 37.9. Thus dilution abolished polydispersity in the human but not the rat mucin, suggesting that intermolecular bonding forces in the human mucin are weaker.