A: Infrared Absorption 197K.

B: Ultraviolet Absorption 197U—

Test solution: 20 mg per mL, in dioxane.

(Official January 1, 2007)

Mobile phase— Dissolve 0.8 g of ammonium nitrate in 300 mL of water, add 700 mL of acetonitrile, and mix.

Internal standard solution— Prepare a solution of testosterone benzoate in tetrahydrofuran containing 2.0 mg per mL.

Standard preparation— Accurately weigh about 10 mg of USP Estradiol Cypionate RS, and transfer to a 10-mL volumetric flask. Add Internal standard solution to volume, and shake vigorously to dissolve.

Assay preparation— Using 10 mg of Estradiol Cypionate, accurately weighed, proceed as directed under Standard preparation.

Procedure— Separately inject 10-µL aliquots of the Assay preparation and the Standard preparation into a suitable high-pressure liquid chromatograph fitted with a 280-nm detector, a 4-mm × 30-cm column containing packing L1 and operated at room temperature. The Mobile phase is maintained at a pressure and flow rate capable of giving the required resolution and a suitable elution time. In a suitable system, the resolution factor R (see Chromatography 621) is not less than 3.0 between the peaks for estradiol cypionate and the internal standard. Five replicate injections of the Standard preparation show a relative standard deviation that is not more than 1.5%. Calculate the quantity, in mg, of C26H36O3 in the portion of Estradiol Cypionate taken by the formula: in which C is the concentration, in mg per mL, of USP Estradiol Cypionate RS in the Standard preparation; and RU and RS are the ratios of the peak responses of the estradiol cypionate and internal standard peaks obtained from the Assay preparation and the Standard preparation, respectively.