The stability of rat (human) CRF in serum, urine and tissue incubation medium was examined using Sephadex gel filtration and CRF radioimmunoassay with anti-rat (human) CRF serum. Human serum after incubation with rat (human) CRF for 1 h at 37 degrees C showed two peaks of CRF immunoreactivity on a Sephadex G-50 fine column. Most of the immunoreactivity coeluted with the rat (human) CRF marker. When rat (human) CRF was incubated with rat liver, kidney or hypothalamus, only 3.1-14.9% of the CRF was recovered at the rat (human) CRF position on gel filtration, and two to four CRF-immunoreactive peaks appeared after the rat (human) CRF marker. When rat (human) CRF was incubated with human urine (pH 6.0) for 24 h at room temperature, one peak of CRF immunoreactivity coeluted with the rat (human) CRF marker on Sephadex gel filtration. The urine extracts of normal rats showed some small peaks of CRF-like immunoreactivity on the Sephadex column, with the main peak appearing after authentic CRF. These results suggest that rat (human) CRF is relatively stable in serum and urine, but is easily degraded by tissue enzymes, with the degraded CRF fragments being excreted in the urine.