Proximal and distal tubule suspensions were prepared from kidneys of Sprague-Dawley rats by an isolation procedure on a Percoll gradient. The marker enzymes alkaline phosphatase (brush border) and hexokinase (cytoplasmic) as well as p-aminohippurate transport capacity, gluconeogenic activity and electron microscopy were used to characterize the two kidney tubule suspensions. The results of this study indicate that cytochrome P-450 is localized to the proximal tubular cells and that the O-deethylation of 7-ethoxycoumarin was higher in the proximal than distal fraction. Both proximal and distal tubules showed glucuronidation and deacetylation capacities and a relatively equal distribution of non-protein sulfhydryls. These studies demonstrate metabolic heterogeneity of the nephron, the proximal tubule being the main site of renal xenobiotic metabolism. Understanding of metabolic heterogeneity of proximal and distal kidney tubules should provide important information regarding cell specific mechanisms of nephrotoxicity.