The effect of 24-h metyrapone administration on adrenal mineralocorticoid and glucocorticoid pathways has been studied in normal children. Urine and serum steroid concentrations were evaluated. Simultaneous determinations of deoxycorticosterone (DOC), corticosterone (B), cortisol (F), and aldosterone (aldo), in both urine and serum, were carried out by celite column chromatography and radioimmunoassay. 11-deoxycortisol (Compound S) in serum and urine, urinary free 18-hydroxydeoxycorticosterone (18-OH-DOC), urinary free 18-hydroxycorticosterone (18-OH-B), and urinary aldosterone-18-glucuronide (pH 1 aldo) were measured separately by specific methods. Conclusions drawn were that (1) Useful indices of metyrapone effect were the marked rise of 11-deoxysteroids (DOC and S), and the fall of aldo and 18-OH-B; no changes occurred in 18-OH-DOC and the 11-hydroxylated steroids (B, F); (2) Serum and urine values were significantly correlated, indicating that both were equally reliable in the evaluation of the metyrapone test; (3) Hormonal ratios indicate that metyrapone inhibition affected the 11-hydroxylation of DOC and Compound S, and the 18-hydroxylation of DOC and B; and (4) The correlation between DOC/B and S/F ratios was highly significant (r = 0.86; P less than 0.0001), showing the same degree of inhibition of 11-hydroxylase functions for both 17-hydroxy (17-OHS) and 17-deoxysteroids.