Characterization of the teratogenic form of phenytoin in serum

Laboratory studies have implicated phenytoin (5,5 diphenylhydantoin) as a teratogen but its mechanism of action remains unknown. Serum collected from monkeys 10 hrs following phenytoin exposure (275 mg/kg p.o.) induced abnormalities and growth retardation in 9.5 day rat embryos cultured for 48 hrs when used as media. Embryonic abnormalities included exencephaly, anophthalmia and partial curvature. Serum was fractionated using ammonium sulfate. The restriction of teratogenic activity to ammonium sulfate precipitates of treated serum suggested that a modified serum protein of phenytoin-protein complex may be responsible for the teratogenicity. Additional serum fractionation using DEAE-Sphacel chromatography has localized this activity to a specific protein eluate which contains predominantly albumin as shown by SDS polyacrylamide gel electrophoresis. However, preliminary results using a hydantoin-ring specific antibody indicated that it was not serum albumin which was binding phenytoin in vivo. The teratogenic activity of serum samples from human patients taking phenytoin is also being characterized.