A: Infrared Absorption 197K.

B: Ultraviolet Absorption 197U—

Solution: 10 µg per mL.

Medium: methanol.

Test solution: 20 mg per mL, in dioxane.

(Official January 1, 2007)

Mobile phase— Prepare a filtered and degassed mixture of water and isopropyl alcohol (72:28). Make adjustments if necessary (see System Suitability under Chromatography 621).

Internal standard solution— Transfer about 66 mg of methyltestosterone to a 10-mL volumetric flask, add dilute alcohol (85 in 100) to volume, and mix.

Standard preparation— Dissolve an accurately weighed quantity of USP Progesterone RS in dilute alcohol (85 in 100) to obtain a solution having a known concentration of about 2.5 mg per mL. Transfer 4.0 mL of this solution to a 10-mL volumetric flask, add 1.0 mL of Internal standard solution, then add dilute alcohol (85 in 100) to volume, and mix to obtain a solution having a known concentration of about 1 mg of USP Progesterone RS per mL.

Assay preparation— Transfer about 10 mg of Progesterone, accurately weighed, to a 10-mL volumetric flask, add 1.0 mL of Internal standard solution, then add dilute alcohol (85 in 100) to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 254-nm detector and a 4-mm × 30-cm column that contains 10-µm packing L1. The flow rate is about 1.5 mL per minute. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the resolution, R, between the analyte and internal standard peaks is not less than 3.5; and the relative standard deviation for replicate injections is not more than 1.5%.

Procedure— Separately inject equal volumes (5 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks. The relative retention times are about 2.0 for progesterone and 1.0 for methyltestosterone. Calculate the quantity, in mg, of C21H30O2 in the portion of Progesterone taken by the formula: in which C is the concentration, in mg per mL, of USP Progesterone RS in the Standard preparation; and RU and RS are the peak response ratios obtained from the Assay preparation and the Standard preparation, respectively.