TRH immunoreactivity levels were measured in human blood, urine, saliva, spinal fluid, amniotic fluid and gastric juice. Urinary TRH excretion during a 48-h period was measured in 11 healthy persons. Blood and urinary TRH immunoreactivity were measured at 2 and 5 h, respectively, after administration of 40 mg of TRH. All the samples were prepurified by SP-Sephadex-C-25 cation-exchange chromatography and subjected to reverse-phase high-pressure liquid chromatography (HPLC). TRH immunoreactivity levels were then measured by our TRH radioimmunoassay. The TRH immunoreactivity (TRH-ir) levels found in urine were 14.6 +/- 2.5 pmol/l; in blood 7.5 +/- 2.0 pmol/l; in spinal fluid 2.8 +/- 1.4 pmol/l, and in gastric juice 23.2 +/- 7.1 pmol/l. In all of the amniotic fluid and saliva samples, in almost one half of the blood and spinal fluid samples, and in almost one third of the gastric juice samples, TRH-ir was below the detectable limit. In blood and urine samples taken after oral administration of TRH, TRH-ir was eluted at the same time as synthetic TRH. The recovery of synthetic TRH added to the samples ranged from 36 to 99%. In all of the biological fluid samples, endogenous TRH-ir was eluted at the same time in HPLC, at 15-18 min, as was synthetic TRH which had been added to the samples. Urine was found to contain two TRH immunoreactive peaks, the second of which was eluted at the same time as synthetic TRH. No diurnal variation in urinary TRH excretion or TRH-ir levels was found.(ABSTRACT TRUNCATED AT 250 WORDS)