A: Infrared Absorption 197S—

B: Ultraviolet Absorption 197U—

C: Add 100 mg to a mixture of 10 mL of water and 2 mL of 1 N sodium hydroxide, boil the mixture for 3 minutes, cool, and add 1 mL of glacial acetic acid and 1 mL of lead acetate TS: a brown to black precipitate of lead sulfide is formed.

Test solution: 10 mg per mL, in chloroform.

Test solution: chloroform.

Standard solution: chloroform.

Eluant: butyl acetate.

Visualization: 5.

Solvent— Use dimethyl sulfoxide.

(Official January 1, 2007)

Mobile phase— Prepare a filtered and degassed mixture of methanol and water (60:40).

Standard preparation— Dissolve an accurately weighed quantity of USP Spironolactone RS in a mixture of acetonitrile and water (50:50), and quantitatively dilute with the same mixture to obtain a solution having a known concentration of about 0.5 mg of USP Spironolactone RS per mL.

Assay preparation— Transfer about 50 mg of Spironolactone, accurately weighed, to a 100-mL volumetric flask, add a mixture of acetonitrile and water (50:50) to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 230-nm detector and a 4.6-mm × 15-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the tailing factor is not more than 2.0; and the relative standard deviation for replicate injections is not more than 1.5%.

Procedure— Separately inject equal volumes (about 20 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 C24H32O4S in the portion of Spironolactone taken by the formula: in which C is the concentration, in mg per mL, of USP Spironolactone RS in the Standard preparation; and rU and rS are the spironolactone peak responses obtained from the Assay preparation and the Standard preparation, respectively.