Related compounds—
TEST 1—
Mobile phase, System suitability solution, and Chromatographic system—
Proceed as directed in
Test 1 for
Related compounds under
Brinzolamide.
Test solution—
Transfer an accurately weighed volume of Ophthalmic Suspension, equivalent to about 10 mg of brinzolamide, to a 25-mL volumetric flask, dilute with alcohol to volume, and mix.
Procedure—
Proceed as directed in
Test 1 for
Related compounds under
Brinzolamide: not more than 1.5% of brinzolamide related compound A is found.
TEST 2—
Buffer solution and Mobile phase—
Proceed as directed in the Assay.
Standard solution—
Dissolve an accurately weighed quantity of
USP Brinzolamide Related Compound B RS in
Mobile phase, and dilute quantitatively, and stepwise if necessary, to obtain a solution having a known concentration of about 2.5 µg per mL.
Test solution—
Use the Assay preparation.
Chromatographic system—
Proceed as directed in the Assay, and chromatograph the System suitability preparation, prepared as directed in the Assay, and the Standard solution, instead of the Standard preparation.
Procedure—
Separately inject equal volumes (about 20 µL) of the
Standard solution and the
Test solution into the chromatograph, record the chromatograms, and measure the areas for all the peaks. Calculate the quantity, in mg, of each impurity in the portion of Ophthalmic Suspension taken by the formula:
(356.46/445.49)50C(ri / rS),
in which 356.46 and 445.49 are the molecular weights of des-ethyl brinzolamide and des-ethyl brinzolamide oxalate, respectively;
C is the concentration, in mg per mL, of
USP Brinzolamide Related Compound B RS in the
Standard solution; ri is peak response for each impurity obtained from the
Test solution; and
rS is the peak response for
USP Brinzolamide Related Compound B RS obtained from the
Standard solution: not more than 0.5% of any individual impurity is found; and not more than 2.0% of total impurities is found.
Assay—
Buffer solution—
Dissolve 11.75 g of ammonium acetate in about 1000 mL of water. Adjust with acetic acid to a pH of 5.2.
Mobile phase—
Prepare a filtered and degassed mixture of
Buffer solution and methanol (65:35). Make adjustments if necessary (see
System Suitability under
Chromatography 
621
).
Standard preparation—
Dissolve an accurately weighed quantity of
USP Brinzolamide RS in
Mobile phase to obtain a solution having a known concentration of about 0.2 mg per mL.
System suitability preparation—
Dissolve an accurately weighed quantity of
USP Brinzolamide Related Compound B RS in
Standard preparation to obtain a solution having a known concentration of about 0.06 mg per mL.
Assay preparation—
Transfer an accurately measured volume of Ophthalmic Suspension, equivalent to about 10 mg of brinzolamide, to a 50-mL volumetric flask, 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 × 15-cm column that contains 5-µm packing L1. The flow rate is about 1.0 mL per minute. Chromatograph the
System suitability preparation, and record the peak responses as directed for
Procedure: the relative retention times are between 0.48 and 0.61 for brinzolamide related compound B and 1.0 for brinzolamide; the resolution,
R, between brinzolamide and brinzolamide related compound B is not less than 4.5; the column efficiency is not less than 2500 theoretical plates; 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 2.0%.
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 areas for the major peaks. Calculate the quantity, in mg, of brinzolamide (C
12H
21N
3O
5S
3) in the portion of Ophthalmic Suspension taken by the formula:
50C(rU / rS),
in which
C is the concentration, in mg per mL, of
USP Brinzolamide RS in the
Standard preparation; and
rU and
rS are the peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.
and 30