Diphenylhydantoin (DPH) is extracted from plasma with ethylene dichloride, returned to alkali, and oxidized with permanganate in glass-stoppered test tubes containing a layer of iso-octane. The absorbance of the oxidation product (benzophenone) in the iso-octane layer is measured spectrophotometrically. This procedure eliminates interference by chloroform that is encountered in other methods; highly specialized reflux condensers or distillation equipment is not needed; and the cost of solvents is decreased. The procedure is highly specific; it detects less than 1 µg of DPH per milliliter of plasma, and it is suitable for routine clinical laboratory use.