» Clotrimazole Lozenges contain not less than 90.0 percent and not more than 110.0 percent of the labeled amount of clotrimazole (C22H17ClN2) in a suitable molded base.
Packaging and storage Preserve in well-closed containers.
USP Reference standards 11
A: The retention time of the major peak in the chromatogram of the Assay preparation corresponds to that in the chromatogram of the Standard preparation, both relative to the internal standard, as obtained in the Assay.
B: Place a quantity of finely powdered Lozenges, equivalent to about 50 mg of clotrimazole, into a screw-capped 50-mL test tube, add 20.0 mL of dichloromethane, and mix. Shake by mechanical means for 10 minutes, and allow the suspension to settle. Use the supernatant as the test solution. Separately apply 20 µL of this solution and 20 µL of a Standard solution of USP Clotrimazole RS in dichloromethane containing 2.5 mg per mL to a thin-layer chromatographic plate (see Chromatography 621) coated with a 0.25-mm layer of chromatographic silica gel mixture. Position the plate in a chromatographic chamber, and develop the chromatograms in a solvent system consisting of a mixture of diethyl ether and ammonium hydroxide (8:1) until the solvent front has moved about three-fourths of the length of the plate. Remove the plate from the developing chamber, mark the solvent front, and allow the solvent to evaporate. Examine the plate under short-wavelength UV light: the RF value of the principal spot obtained from the test solution corresponds to that obtained from the Standard solution. Dissolve 3 g of bismuth subnitrate and 30 g of potassium iodide in 10 mL of dilute hydrochloric acid (1 in 4), dilute with water to 100 mL, mix, and prepare a spray reagent by diluting 10 mL of this solution and 10 mL of dilute hydrochloric acid (1 in 4) with water to 100 mL. Spray the plate with the spray reagent, and visually locate the clotrimazole spots on the plate: the spots are orange.
Medium: 0.1 N hydrochloric acid; 500 mL, deaerated.
Apparatus 2: 50 rpm.
Time: 45 minutes.
Determine the amount of C22H17ClN2 dissolved by employing the following method.
25 mM Phosphate buffer Dissolve 4.4 g of dibasic potassium phosphate in 1000 mL of water.
100 mM Phosphate buffer Dissolve 17.4 g of dibasic potassium phosphate in 1000 mL of water.
Diluent Prepare a filtered and degassed mixture of methanol and 100 mM Phosphate buffer (60:40).
Mobile phase Prepare a filtered and degassed mixture of methanol and 25 mM Phosphate buffer (4:1). Make adjustments if necessary (see System Suitability under Chromatography 621).
Standard solution Dissolve an accurately weighed quantity of USP Clotrimazole RS, and dilute quantitatively, and stepwise if necessary, with Medium to obtain a solution having a concentration of about 0.02 mg per mL.
Working standard solution Transfer 5.0 mL of the Standard solution to a 25-mL volumetric flask, dilute with Diluent to volume, and mix.
Test solution Withdraw 25 mL of the solution under test from the vessel. Pass through a 0.45-µm polyvinylidene difluoride filter, discarding the first 10 mL of the filtrate. Transfer 5.0 mL of the filtrate to a 25-mL volumetric flask. Dilute with Diluent to volume, and mix.
Chromatographic system (see Chromatography 621) The liquid chromatograph is equipped with a 215-nm detector and a 3.9-mm × 7.5-cm column that contains packing L1. The flow rate is about 1.0 mL per minute. Chromatograph the Working standard solution, and record the peak responses as directed for Procedure: the tailing factor is not more than 2.0, and the relative standard deviation for replicate injections is not more than 2.0%.
Procedure Separately inject equal volumes (about 20 µL) of the Working standard solution and the Test solution into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the amount, in percentage, of C22H17ClN2 released by the formula: in which rU and rS are the peak responses for the Test solution and the Working standard solution, respectively; CS is the concentration, in mg per mL, of the Working standard solution; 500 is the volume, in mL, of Medium; D is the dilution factor of the Test solution; 100 is the conversion factor to percentage; and LC is the label claim, in mg.
Tolerances Not less than 80% (Q) of the labeled amount of C22H17ClN2 is dissolved in 45 minutes.
Uniformity of dosage units 905: meet the requirements.
Buffer solution Dissolve 1 g of ammonium carbonate in 1000 mL of water. Adjust with 10% sulfuric acid solution to a pH of 6.0.
Mobile phase Prepare a filtered and degassed mixture of methanol and Buffer solution (75:25). Make adjustments if necessary (see System Suitability under Chromatography 621).
Internal standard stock solution Dissolve an accurately weighed quantity of triphenylmethane in methanol, and dilute quantitatively and stepwise with methanol to obtain a solution having a concentration of 5 mg per mL.
Internal standard solution Pipet 10.0 mL of Internal standard stock solution into a 250-mL volumetric flask, and dilute with Mobile phase to volume.
Standard preparation Transfer about 20 mg of USP Clotrimazole RS, accurately weighed, to a 100-mL volumetric flask. Add 4.0 mL of Internal standard stock solution, dissolve in and dilute with Mobile phase to volume, and mix.
Assay preparation Weigh and pulverize not fewer than 10 Lozenges. Transfer an accurately weighed portion of the powder, equivalent to about 5 mg of clotrimazole, to a 50-mL screw-capped centrifuge tube. Pipet 25 mL of Internal standard solution into the tube. Sonicate for 10 minutes, then shake for 10 minutes. Centrifuge at 2500 rpm for 30 minutes. Use the clear supernatant layer as the Assay preparation.
Chromatographic system (see Chromatography 621) The liquid chromatograph is equipped with a 215-nm detector and a 3.9-mm × 30-cm column that contains packing L1. The flow rate is about 2 mL per minute. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the relative retention times are about 1.0 for clotrimazole and 1.4 for triphenylmethane; the resolution, R, between clotrimazole and triphenylmethane is not less than 1.5; the column efficiency is not less than 1500 theoretical plates; the tailing factor is not more than 2; and 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 responses for the major peaks. Calculate the quantity, in mg, of clotrimazole (C22H17ClN2) in the portion of Lozenges taken by the formula:
25C(RU / RS)in which C is the concentration, in mg per mL, of USP Clotrimazole RS in the Standard preparation; and RU and RS are the ratios of the peak responses for clotrimazole to triphenylmethane obtained from the Assay preparation and the Standard preparation, respectively.
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USP35NF30 Page 2743Pharmacopeial Forum: Volume No. 32(1) Page 78