Sephadex LH 20 columns have been investigated for the separation of initial prostaglandin metabolites. Solvent systems are described for the separation of the free acids of 15-keto-dihydro-PGE, 15-keto-PGE, PGE, and PGF(1a). Further, one of the solvent systems is described for the separation of pulmonary metabolites of PGE and PGF(1a), and another one for separation of dihydro-PGE and PGE.