Mobile phase and Diluent— Prepare as directed in the Assay.

System suitability solution— Prepare a solution in Diluent containing about 0.04 mg of USP Glimepiride RS per mL and about 0.02 mg each of USP Glimepiride Related Compound B RS and USP Glimepiride Related Compound C RS per mL. Pipet a 5.0-mL aliquot of this solution into a 50-mL volumetric flask, dilute with Diluent to volume, and mix.

Sensitivity solution— Pipet a 5.0-mL aliquot of the System suitability solution into a 100-mL volumetric flask, dilute with Diluent to volume, and mix.

Test solution— Weigh and finely powder not fewer than 10 Tablets, and transfer an accurately weighed portion of the powder to a 50-mL centrifuge tube. Add Diluent to prepare a solution containing about 0.1 mg of glimepiride per mL, based on the label claim. Sonicate in a water bath at a temperature not to exceed 20 for not less than 5 minutes and not more than 10 minutes, with occasional mixing. Centrifuge the samples, and use the clear supernatant.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 228-nm detector and a 4-mm × 25-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the System suitability solution, and identify the glimepiride peak and the peaks due to the related compounds on the basis of the relative retention times, which are about 0.2 for glimepiride related compound B, about 0.3 for glimepiride related compound C, and 1.0 for glimepiride. Record the peak responses as directed for Procedure: the resolution, R, between glimepiride related compounds B and C is not less than 4, and the relative standard deviation of the glimepiride peak for replicate injections is not more than 2.0%. Chromatograph the Sensitivity solution, and calculate the signal-to-noise ratio, S/N, for the peaks of glimepiride related compounds B and C by the formula: in which H is the measured height of the respective related compound peak, and h is the amplitude of the average measured baseline noise. The S/N for each peak is not less than 10.

Procedure— Inject a volume (about 10 µL) of the Test solution into the chromatograph, record the chromatograms, and measure the peak responses. Continue the elution for at least two times the retention time of the glimepiride peak. Calculate the percentage of each impurity in the portion of Tablets taken by the formula: in which F is the relative response factor, which is equal to 1.3 for glimepiride related compound B and 1.0 for any other impurity; rU is the peak response for each impurity obtained from the Test solution; and rs is the sum of the responses of all the peaks in the Test solution. Disregard any peak less than 0.1%. Not more than 2.5% of glimepiride related compound B is found, not more than 0.5% of any other individual impurity is found, not more than 1.0% of total impurities excluding glimepiride related compound B is found, and not more than 3.5% of overall total impurities (including glimepiride related compound B) is found.

Mobile phase— Dissolve 0.5 g of monobasic sodium phosphate in 500 mL of water. Adjust with 10% phosphoric acid to a pH of 2.1 to 2.7, add 500 mL of acetonitrile, and mix.

Diluent— Prepare a mixture of acetonitrile and water (9:1).

System suitability preparation— Prepare a solution in Diluent containing 0.1 mg of USP Glimepiride RS per mL and 0.02 mg each of USP Glimepiride Related Compound B RS and USP Glimepiride Related Compound C RS per mL.

Standard preparation— Dissolve an accurately weighed quantity of USP Glimepiride RS in Diluent to obtain a solution having a known concentration of about 0.1 mg per mL.

Assay preparation— Transfer five whole Tablets into a suitable volumetric flask to prepare a solution of approximately 0.1 mg of glimepiride per mL, based on the label claim. Add water to 10% of the volume of the flask. Shake the flask to completely dissolve the tablets. Add acetonitrile to about 70% of the volume of the flask, and swirl. Sonicate the samples in a water bath not to exceed 20 for not less than 5 minutes and not more than 10 minutes with occasional shaking. Allow the solutions to come to room temperature, dilute with acetonitrile to volume, mix, and filter.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 228-nm detector and a 4-mm × 12.5-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the System suitability preparation, and identify the glimepiride peak and the peaks due to the related compounds based on their relative retention times, which are about 0.25 for glimepiride related compound B, about 0.35 for glimepiride related compound C, and 1.0 for glimepiride. Record the peak responses as directed for Procedure: the resolution, R, between glimepiride related compounds B and C is not less than 1.5, and the tailing factor for the glimepiride peak 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 10 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 percentage of the labeled amount of glimepiride (C24H34N4O5S) in the portion of Tablets taken by the formula: in which CS is the concentration, in mg per mL, of glimepiride in the Standard preparation; CU is the concentration of glimepiride in the Assay preparation, based on the labeled quantity per Tablet and the extent of dilution; and rU and rS are the peak responses for glimepiride obtained from the Assay preparation and the Standard preparation, respectively.