A: Infrared Absorption 197K.

B: Ultraviolet Absorption 197U—

Test solution: 10 mg per mL, in dioxane.

Adsorbent: a suitable thin-layer chromatographic plate (see Chromatography 621) coated with a 0.25-mm layer of chromatographic silica gel mixture.

Test solution— Dissolve an accurately weighed amount of Medroxyprogesterone Acetate in methylene chloride to obtain a solution having a concentration of about 20 mg per mL.

Standard solution— Prepare a solution of USP Medroxyprogesterone Acetate RS and USP Medroxyprogesterone Acetate Related Compound A RS in methylene chloride containing 20 mg per mL and 0.1 mg per mL, respectively.

Application volume: 10 µL.

Developing solvent system: a mixture of hexanes, tert-butyl methyl ether, and tetrahydrofuran (45:45:10).

Spray reagent— Prepare a solution of 20 g of p-toluenesulfonic acid in 100 mL of alcohol.

Procedure— Proceed as directed for Thin-Layer Chromatography under Chromatography 621. Develop the chromatogram until the solvent front has moved about 10 cm. Allow the plate to air-dry, and develop the chromatogram again until the solvent front has moved about 10 cm. Allow the plate to dry at 120 for 10 minutes. Spray the plate with Spray reagent. Heat the plate for 10 minutes at 120, and examine the plate under UV light at 365 nm. Any blue fluorescent spot with an RF value higher than that of the principal spot due to medroxyprogesterone acetate in the chromatogram obtained from the Test solution is not more intense than the corresponding blue fluorescent spot in the chromatogram obtained from the Standard solution: not more than 0.5% is found.

Mobile phase— Prepare a filtered and degassed mixture of acetonitrile and water (3:2). Make adjustments if necessary (see System Suitability under Chromatography 621).

Standard solution— Dissolve an accurately weighed quantity of USP Medroxyprogesterone Acetate RS in Mobile phase, and dilute quantitatively, and stepwise if necessary, with Mobile phase to obtain a solution having a known concentration of about 50 µg per mL.

System suitability solution— Dissolve suitable quantities of megestrol acetate and USP Medroxyprogesterone Acetate RS in Mobile phase to obtain a solution containing about 40 µg of each per mL.

Test solution— Transfer about 62.5 mg of Medroxyprogesterone Acetate, accurately weighed, to a 25-mL volumetric flask, dissolve in and dilute with Mobile phase to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 25-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the System suitability solution, and record the peak responses as directed for Procedure: the resolution, R, between megestrol acetate and medroxyprogesterone acetate is not less than 1.5. Chromatograph the Standard solution, and record the peak responses as directed for Procedure: the relative standard deviation for replicate injections is not more than 3.0%.

Procedure— Inject a volume (about 20 µL) of the Test solution into the chromatograph, record the chromatogram, and measure the peak responses. Calculate the percentage of each impurity in the portion of Medroxyprogesterone Acetate taken by the formula: in which C is the concentration, in mg per mL, of USP Medroxyprogesterone Acetate RS in the Standard solution; W is the weight, in mg, of Medroxyprogesterone Acetate taken to prepare the Test solution; ri is the peak response for each impurity obtained from the Test solution; and rS is the response from the major peak obtained from the Standard solution: not more than 1.0% of any individual impurity is found; and not more than 1.5% of total impurities is found.

(Official January 1, 2007)

Mobile phase— Prepare a filtered and degassed mixture of water and acetonitrile (60:40). Make adjustments if necessary (see System Suitability under Chromatography 621).

Standard preparation— Dissolve an accurately weighed quantity of USP Medroxyprogesterone Acetate RS in acetonitrile to obtain a solution having a known concentration of about 1 mg per mL.

Assay preparation— Dissolve about 25 mg of Medroxyprogesterone Acetate, accurately weighed, in 25.0 mL of acetonitrile, 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 packing L1. The flow rate is about 2 mL per minute. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the tailing factor is not more than 2; and the relative standard deviation of the peak responses for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 10 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of C24H34O4 in the portion of Medroxyprogesterone Acetate taken by the formula: in which C is the concentration, in mg per mL, of USP Medroxyprogesterone Acetate RS in the Standard preparation; and rU and rS are the peak responses obtained from the Assay preparation and the Standard preparation, respectively.