Trace analysis of ethinyl estradiol in casein diet using gas chromatography with electron capture detection

Ethinyl estradiol (EE2) is an extremely potent synthetic estrogen and a common component in oral contraceptives. The drug has a well-characterized pharmacological profile and is used as a positive control in toxicological investigations of compounds having estrogenic activity. An analytical method developed for the determination of low microg/kg levels of EE2 in a casein-based rodent diet is presented. A methanol extract of casein diet is purified for instrumental analysis by a 3-fold solid-phase extraction process. The sample extract is derivatized with pentafluoropropionic anhydride to the pentafluoropropionyl product and analyzed by capillary gas chromatography with electron-capture detection. Recoveries of EE2 from casein diet fortified at 5, 10, and 50 microg/kg average 88.8% and have a relative standard deviation (%) of 7.2. The method limit of detection in a casein-based diet is 1 microg/kg.