In experimental diabetes, a number of intestinal brush-border hydrolases and transport systems are stimulated. In this study, we assessed possible effects of diabetes on the composition and membrane fluidity of rat intestinal brush-border membranes that might correlate with these functional changes. We found similar proportions of lipid and protein in the diabetic and control preparations, although there was a considerable increase in total membrane from the diabetic rats, presumably reflecting mucosal hyperplasia. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of membrane protein revealed an increase in the bands corresponding to sucrase-isomaltase, consistent with an increased enzyme activity of sucrase. Membrane lipid analysis revealed only a decrease in fatty acids of the neurtral lipid fraction of diabetics-a change that may well have occurred during membrane preparation. 1-6-Diphenyl-1,3,5-hexatriene fluorescence polarization data, obtained as a function of temperature, was similar for the diabetic and control rats, with a three-phase linear model superior to one- and two-phase linear or quadratic models. The overall composition of the intestinal brush-border membrane, unlike other plasma membranes, appears little affected by experimental diabetes.