Although the liver is recognized as a major site of glutathione (GSH) synthesis, it is thought to play only a minor role in GSH catabolism. This is primarily because in the rat, the most commonly used experimental animal, hepatic gamma-glutamyltransferase (gamma-GT) activity is very low, whereas kidney activity is quite high. gamma-GT is the only enzyme known to catalyze the initial step in GSH degradation. The present work compares gamma-GT and dipeptidase activities in liver, kidney, and gallbladder of six mammalian species to assess the importance of hepatobiliary catabolism of GSH, relative to renal degradation. Marked species differences were observed in gamma-GT activities, and in kidney to liver (K/L) ratios for both gamma-GT concentration (milliunits/mg protein) and whole organ activities (total activity per liver or two kidneys). The K/L concentration ratios for gamma-GT activities ranged from 875 in the rat to 15 in the guinea pig. Whole organ gamma-GT ratios were approximately 150 in mouse and rat, and only 2-5 in guinea pig. pig, and macaque. Human K/L ratios calculated from gamma-GT activities reported previously were similar to those of the guinea pig. Species differences were also observed in K/L ratios for dipeptidase activities, though these differences were not as large as those for gamma-GT, gamma-GT and dipeptidase activities were also measured in gallbladders of all species examined (except rat which does not have this organ), and were found to be comparable to those of liver. These results suggest that in species such as the guinea pig and perhaps humans, the liver and biliary tree play a prominent role in GSH turnover. Because of the low hepatic and high renal gamma-GT activities of the rat, and because it does not have a gallbladder, this species may not be the best model for studying the catabolism of GSH and GSH conjugates. Use of the rat model may underestimate the contribution of liver, and overestimate that of kidney, in these degradative processes.