Identification—
Transfer an amount of finely ground Tablets, equivalent to 4 mg of trimethoprim, to a 10-mL volumetric flask, add 8 mL of methanol, warm for several minutes on a steam bath with frequent shaking, cool, dilute with methanol to volume, mix, and centrifuge briefly. Apply 5 µL each of this solution, a Standard solution of
USP Trimethoprim RS in methanol containing 0.4 mg per mL, and a Standard solution of
USP Sulfamethoxazole RS in methanol containing 2 mg per mL to separate points about 3 cm from one end of a thin-layer chromatographic plate coated with chromatographic silica gel mixture. Dry the spots in a current of warm air, and develop the plate with a mixture of chloroform, isopropyl alcohol, and diethylamine (6:5:1) in a chamber lined with filter paper. Remove the plate, dry, and examine under short-wavelength UV light: the trimethoprim and sulfamethoxazole spots from the solution under test have the same
RF values as the spots from the corresponding Standard solutions.
Dissolution 
711
—
Medium:
0.1 N hydrochloric acid; 900 mL.
Apparatus 2:
75 rpm.
Time:
60 minutes.
Procedure—
Determine the amounts of sulfamethoxazole (C
10H
11N
3O
3S) and trimethoprim (C
14H
18N
4O
3) dissolved, employing the procedure set forth in the
Assay, making any necessary volumetric adjustments (see
Chromatography 621). Calculate the percentage of each active component dissolved by comparison of the peak responses obtained from a filtered aliquot of the solution under test with the peak responses from the corresponding component obtained from the
Standard preparation.
Tolerances—
Not less than 70% (Q) of the labeled amounts of C10H11N3O3S and C14H18N4O3 is dissolved in 60 minutes.
Assay—
Assay preparation—
Weigh and finely powder not fewer than 20 Tablets. Transfer an accurately weighed portion of the powder, equivalent to about 160 mg of sulfamethoxazole, to a 100-mL volumetric flask. Add about 50 mL of methanol and sonicate, with intermittent shaking, for 5 minutes. Allow to equilibrate to room temperature, dilute with methanol to volume, mix, and filter. Transfer 5.0 mL of the clear filtrate to a 50-mL volumetric flask, dilute with Mobile phase to volume, and mix.
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 quantities, in mg, of trimethoprim (C
14H
18N
4O
3) and sulfamethoxazole (C
10H
11N
3O
3S) in the portion of Tablets taken by the formula:
1000C(rU / rS),
in which
C is the concentration, in mg per mL, of the appropriate USP Reference Standard in the
Standard preparation; and
rU and
rS are the responses of the corresponding analyte obtained from the
Assay preparation and the
Standard preparation, respectively.
