The sulfonylurea class of compounds has demonstrated its effectiveness in treating non-insulin-dependent diabetes mellitus, although the exact mechanisms of action are still not fully defined. Clinical studies have suggested the action may be at least in part due to alterations of insulin's effect on liver. We have examined the effects of glyburide and glipizide on insulin metabolism in isolated hepatocytes and perfused livers. Our studies show that both drugs increase insulin binding to hepatocyte, but only glyburide-treated animals exhibit a concomitant increase in degradation. Studies with recycling perfused liver agreed with these results, with glyburide treatment causing a significantly more rapid clearance than control or glipizide treatment. Single-pass perfusion studies showed significantly less insulin retained by glipizide-treated animals as compared to control- and glyburide-treated animals. Further, hepatocytes from glipizide-treated animals required higher concentrations of insulin to achieve the same stimulation of amino acid transport (as measured by aminoisobutyric acid uptake) as in control- and glyburide-treated animals, suggesting that the altered processing affects insulin's action. Taken together, these studies demonstrate alterations in liver insulin metabolism that may explain clinical differences identified in these two drugs.