A high-performance liquid chromatographic (HPLC) analysis is described for dienestrol as a drug substance and in cream, foam, and tablet dosage forms. After incorporation of the drug or dosage form into a solvent mixture containing an internal standard, biphenyl, an aliquot was chromatographed using a reversed-phase medium, followed by UV spectrophotometric detection at 254 nm. The response of the chromatographic system was linear over a concentration range corresponding to 50-200% of the labeled amount of dienestrol. Satisfactory accuracy and precision were confirmed by analyzing cream by the standard addition method. The advantages of the HPLC method are its simplicity, speed, and sensitivity, which permit direct analysis of single-dose quantities of dienestrol.