Separation of pharmaceutically important estrogens by micellar electrokinetic chromatography

Baseline separation of ten estrogens of the type found in conjugated estrogens tablets from pregnant mares' urine were separated by micellar electrokinetic chromatography with a 20 mM sodium borate-sodium phosphate buffer (pH 8) containing either 75 mM sodium cholate and 15 mM beta-cyclodextrin or 50 mM sodium dodecyl sulfate and 20 mM gamma-cyclodextrin. The system containing sodium cholate was used to identify estrogens in a pharmaceutical tablet formulation of conjugated estrogens and to determine the amounts and ratio of sodium equilin sulfate to sodium estrone sulfate to confirm tablet conformity with regulatory requirements. Baseline separation of the seven ethynyl steroids registered as oral contraceptives in the USA was obtained with the system 20 mM sodium borate-sodium phosphate buffer (pH 8) containing 50 mM sodium cholate and 10% (v/v) methanol. Factors that affect selectivity and reproducibility of the above separation systems are discussed.