A: Infrared Absorption 197K.

B: Ultraviolet Absorption 197U—

Buffer solution— Dissolve 1.80 g of anhydrous sodium acetate in 1 L of water. Adjust, if necessary, with glacial acetic acid to a pH of 4.5 ± 0.2.

Mobile phase— Prepare a filtered and degassed mixture of Buffer solution and acetonitrile (86:14). Make adjustments if necessary (see System Suitability under Chromatography 621).

Standard preparation— Dissolve about 20 mg of USP Methazolamide RS, accurately weighed, in 20 mL of acetonitrile contained in a 200-mL volumetric flask. Dilute with Buffer solution to volume, and mix. Quantitatively dilute an accurately measured volume of this solution with Buffer solution to obtain a solution having a known concentration of about 50 µg of USP Methazolamide RS per mL.

Resolution solution— Prepare a solution of acetaminophen and USP Methazolamide RS in acetonitrile containing 0.3 mg of acetaminophen and 0.5 mg of methazolamide per mL. Quantitatively dilute an accurately measured volume of this solution with Buffer solution to obtain a solution containing 30 µg of acetaminophen and 50 µg of methazolamide per mL.

Assay preparation— Transfer about 100 mg of Methazolamide, accurately weighed, to a 200-mL volumetric flask, dissolve in 20 mL of acetonitrile, dilute with Buffer solution to volume, and mix. Quantitatively dilute an accurately measured volume of this solution with Buffer solution to obtain a solution having a known concentration of about 50 µg per mL.

Chromatographic system (see Chromatography 621)—The liquid chromatograph is equipped with a 265-nm detector and a 3.9-mm × 15.0-cm column containing packing L1. The flow rate is about 1.5 mL per minute. Chromatograph the Resolution solution, and record the peak responses as directed for Procedure: the relative retention times are about 0.6 for acetaminophen and 1.0 for methazolamide, the resolution, R, between the acetaminophen peak and the methazolamide peak is not less than 4.0, and the tailing factor is not more than 2.0. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the relative standard deviation for replicate injections is not more than 2%.

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 areas of the responses for the major peaks. Calculate the quantity, in mg, of C5H8N4O3S2 in the portion of Methazolamide taken by the formula: in which C is the concentration, in µg per mL, of USP Methazolamide RS in the Standard preparation; and rU and rS are the peak responses obtained from the Assay preparation and the Standard preparation, respectively.