Identification—
Transfer a portion of powdered Tablets, equivalent to about 30 mg of timolol maleate, to a 50-mL volumetric flask, add about 2 mL of 0.1 N hydrochloric acid, and shake gently. Add about 30 mL of methanol, agitate for 20 minutes, add methanol to volume, mix, and centrifuge. Similarly prepare a Standard solution containing 0.6 mg of
USP Timolol Maleate RS per mL. Separately apply 10 µL of the test solution and 10 µL of the Standard solution to a thin-layer chromatographic plate (see
Chromatography 
621
) coated with a 0.25-mm layer of chromatographic silica gel mixture. Develop the chromatogram using a solvent system consisting of a mixture of chloroform, methanol, and ammonium hydroxide (80:20:1) until the solvent front has moved about three-fourths of the length of the plate. Air-dry, and examine under short-wavelength UV light: the
RF values of the principal spots obtained from the test solution correspond to those obtained from the Standard solution.
Dissolution, Procedure for a Pooled Sample 711—
Medium:
0.1 N hydrochloric acid; 500 mL.
Apparatus 1:
100 rpm.
Time:
20 minutes.
Procedure—
Determine the amount of timolol maleate in solution in filtered portions of the solution under test, in comparison with a Standard solution having a known concentration of
USP Timolol Maleate RS in the same medium, employing the procedure set forth in the
Assay, making any necessary modifications.
Tolerances—
Not less than 80% (Q) of the labeled amount of timolol maleate (C13H24N4O3S·C4H4O4) is dissolved in 20 minutes.
Assay—
pH 2.8 phosphate buffer—
Transfer 22.08 g of monobasic sodium phosphate to a 2-liter volumetric flask, dilute with water to volume, adjust with phosphoric acid to a pH of 2.8 ± 0.05, and filter.
Mobile phase—
Prepare a suitable degassed and filtered mixture of pH 2.8 phosphate buffer and methanol (3:2).
Standard preparation—
Transfer about 50 mg of
USP Timolol Maleate RS, accurately weighed, to a 500-mL volumetric flask. Add 50 mL of 0.05
M monobasic sodium phosphate. Sonicate until the standard is dissolved, add 100 mL of acetonitrile, shake, dilute with water to volume, and mix.
Assay preparation—
Weigh and finely powder not less than 20 Tablets. Transfer an accurately weighed portion of the powder, equivalent to about 10 mg of timolol maleate, to a 100-mL volumetric flask, add 10 mL of 0.05 M monobasic sodium phosphate, sonicate for 5 minutes, and add 20 mL of acetonitrile. Sonicate for 5 minutes, add 20 mL of water, shake for 10 minutes, dilute with water to volume, and mix.
Chromatographic system
(see
Chromatography 621)—The liquid chromatograph is equipped with a 295-nm detector and a 3.9-mm × 30-cm column that contains packing L1. The flow rate is about 1.8 mL per minute. Chromatograph five replicate injections of the
Standard preparation, and record the peak responses as directed for
Procedure: the relative standard deviation is not more than 2.0%, and the tailing factor for the main peak is not greater than 2.0.
Procedure—
Separately inject equal volumes (about 15 µL) of the
Standard preparation and the
Assay preparation into the chromatograph by means of a suitable microsyringe or sampling valve, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity, in mg, of C
13H
24N
4O
3S·C
4H
4O
4 in the portion of Tablets taken by the formula:
100C(rU / rS),
in which
C is the concentration, in mg per mL, of
USP Timolol Maleate RS in the
Standard preparation, and
rU and
rS are the peak responses obtained for timolol maleate from the
Assay preparation and the
Standard preparation, respectively.
