Rapid, sensitive colorimetric method for determination of ethinyl estradiol

A colorimetric procedure, based on the formation of an azo dye by condensation of diazotized 5-chloro-2,4-dinitroaniline with ethinyl estradiol, was developed. An alkaline solution of ethinyl estradiol is reacted with the reagent, and the resulting color is measured at 450 nm. Absorbance versus concentration is linear up to 10 mug/ml; the lower limit of detection is 1 mug/ml under the conditions studied. Replicate analysis showed good agreement, and an average recovery of 99.6 +/- 0.3% was obtained for analyses of synthetic mixtures. Vitamins and minerals likely to be present along with ethinyl estradiol in certain geriatric formulations, as well as ordinary tablet excipients and coating materials, do not interfere with the precision of the method or development of the color. The method is applicable to progestin-estrogen preparations. Assay results on various single-component as well as contraceptive commercial samples are reported.