Simple fluorometric determination of aldosterone in urine without use of isotopes or chromatography

Aldosterone 18-glucuronide in urine is hydrolyzed by adjusting the pH to 1.0 and allowing the mixture to stand overnight at room temperature. The free aldosterone is then extracted into dichloromethane, which is washed with carbonate to remove acidic compounds and evaporated. The residue is partitioned between a nonpolar organic phase and an aqueous phase, and the aldosterone oxidized to the 13-carboxylic acid derivative with Benedict's qualitative glucose reagent. Neutral compounds are extracted from this oxidation mixture with dichloromethane at pH 7.5, the mixture is acidified, and the oxidized aldosterone extracted into dichloromethane. After washing with pH 3.5 buffer, this extract is evaporated and the oxidized aldosterone determined fluorometrically via a two-stage reaction with sulfuric acid/water (85/15 by vol) and methanol containing ferric chloride.