The urinary excretion of 6Β-hydroxycortisol (6Β-OHF), free cortisol, and estriol was studied at 4-hourly intervals in 9 subjects between the 36th and 38th week of pregnancy. The ratio of 6Β-OHF to creatinine showed a significant diurnal rhythm with a 40% fall from a peak at 06.00–10.00 h to a trough at 22.00–02.00 h. There was a significant positive correlation between the excretion of 6Β-OHF and free cortisol. Estrogen excretion showed no diurnal variation. No increase was observed in the 6Β-OHF:creatinine ratio in early morning urine samples from 5 subjects followed daily prior to the onset of spontaneous labour at term. Normal nonpregnant females were also found to excrete significant quantities of 6Β-OHF. These findings suggest that the clinical significance of 6Β-OHF levels measured in a single void sample of urine has to be reassessed in the face of such a marked diurnal variation.