This study was performed to determine whether corticosterone (B), the major glucocorticoid of rat, is metabolized to its 6 beta-OH derivative (6 beta-OH-B) and whether this derivatives has any effects on Na+ or K+ transport in rat kidney. Normal and adrenalectomized (adx) rats were injected with [3H]B and urine was collected for 5 h and examined for metabolites of B. Metabolites were collected by solid phase extraction on mu Bondapak C18 cartridges and fractionated by reversed phase high performance liquid chromatography. Fractions coeluting with 6 beta-OH-B were rechromatographed by normal phase thin layer chromatography. Approximately 5% of the radioactivity recovered from the urine of both intact and adx rats cochromatographed with 6 beta-OH-B on the two systems. Mass spectra of this fraction were virtually identical to those of authentic 6 beta-OH-B, demonstrating that rats do metabolize B to its 6 beta-OH derivative. To evaluate the biological activity of this metabolite, adx rats were injected with NaCl and KCl and with varying dosage of either 6 beta-OH-B or reference steroids (aldosterone, B, 6 beta-OH-F). 6 beta-OH-B produced a significant antinatriuresis at all doses. Kaliuresis occurred only at the highest dose and creatinine excretion increased, suggesting increased glomerular filtration from a glucocorticoid effect. Although 6 beta-OH-B may simply be exerting mineralo- and glucocorticoid actions there are two unexplained findings. First, 6 beta-OH-B (10 micrograms/100 g) significantly decreased urinary K excretion with associated antinatriuresis, an effect which has not been seen with Aldo administration. Second, neither a kaliuretic nor antinatriuretic effect of 6 beta-OH-B could be demonstrated in experiments using a method to enhance mineralocorticoid induced K+ excretion (K+ deprivation and NaCl loading only). Yet, the dose used was clearly antinatriuretic in the initial bioassay. It is concluded that the rat is capable of metabolizing B to its 6 beta-OH-B derivative which appears in substantial quantity in the urine. This metabolite produces antinatriuresis in the adrenalectomized rat.