Estradiol Tablets
» Estradiol Tablets contain not less than 90.0 percent and not more than 115.0 percent of the labeled amount of C18H24O2.
Packaging and storage— Preserve in tight, light-resistant containers.
USP Reference standards 11
USP Estradiol RS Click to View Structure
USP Estrone RS Click to View Structure
Identification— Place a quantity of finely powdered Tablets, equivalent to about 4 mg of estradiol, in a screw-capped, 20-mL vial. Add 10 mL of chloroform, and sonicate for 2 minutes. Filter through medium-porosity filter paper. Apply 20 µL each of this solution and a Standard solution of USP Estradiol RS in chloroform containing 0.4 mg per mL to a suitable thin-layer chromatographic plate (see Chromatography 621) coated with a 0.25-mm layer of chromatographic silica gel mixture. Allow the spots to dry, and develop the chromatogram in a lined chamber with a solvent system consisting of a mixture of toluene and acetone (4:1) until the solvent front has moved 10 cm beyond the starting line. Remove the plate from the developing chamber, mark the solvent front, and allow to air-dry. Spray the plate with a mixture of methanol and sulfuric acid (1:1), and heat at 100 for about 5 minutes: the principal spots obtained from the test solution and the Standard solution have the same color and RF value.
Dissolution 711
Medium: 0.3% sodium lauryl sulfate in water; 500 mL.
Apparatus 2: 100 rpm.
Time: 60 minutes.
Mobile phase— Prepare a suitable degassed and filtered solution of water and acetonitrile (55:45).
Standard solution— Prepare a solution of USP Estradiol RS in methanol having an accurately known concentration of about 0.02 mg per mL. Dilute aliquots of this solution with Medium to obtain a final solution having a concentration approximately equal to the expected concentration of drug in the Medium, assuming 100% dissolution.
Test solution— Use a filtered portion of the solution under test from the dissolution vessel.
Chromatographic system (see Chromatography 621)—The liquid chromatograph is equipped with a 205-nm detector and a 4.6-mm × 7.5-cm column that contains packing L1. The flow rate is about 1.5 mL per minute. Chromatograph replicate injections of the Standard preparation, and record the peak areas as directed for Procedure: the tailing factor is not more than 2.0; and the relative standard deviation is not more than 2.0%.
Procedure— Separately inject equal volumes (about 100 µL) of the Standard solution and the Test solution into the chromatograph, record the chromatograms, and measure the areas for the major peaks. Calculate the quantity of C18H24O2 dissolved by comparison of the peak areas obtained from the Test solution and the Standard solution.
Tolerances— Not less than 75% (Q) of the labeled amount of C18H24O2 is dissolved in 60 minutes.
Uniformity of dosage units 905: meet the requirements.
Chromatographic purity—
Solution A— Prepare a degassed mixture of water and acetonitrile (8:2).
Solution B— Prepare a degassed mixture of acetonitrile and water (8:2).
Diluent— Prepare a mixture of water and acetonitrile (6:4).
Mobile phase— Use variable mixtures of Solution A and Solution B as directed for Chromatographic system. Make adjustments if necessary (see System Suitability under Chromatography 621).
System suitability solution— Dissolve accurately weighed quantities of USP Estradiol RS and USP Estrone RS in acetonitrile to obtain a solution having concentrations of 0.5 mg per mL and 0.3 mg per mL, respectively. Pipet 2 mL of this solution into a 50-mL volumetric flask, and dilute with Diluent to volume.
Test solution— Transfer a number of Tablets, containing a combined amount of 4 to 8 mg of estradiol based on the label claim, to a suitable flask, add a volume of Diluent equivalent to about 5 mL per each mg of estradiol, swirl until the Tablets are completely disintegrated, then shake for 15 minutes, and allow the solids to settle. Pass a portion of this solution through a 0.45-µm PVDF filter, discarding the first 2 mL of the filtrate. This solution contains about 0.2 mg of estradiol per mL.
Chromatographic system (see Chromatography 621)—The liquid chromatograph is equipped with a 200-nm detector and a 4.0-mm × 12.5-cm column that contains 5-µm packing L7. The flow rate is about 1 mL per minute. The chromatograph is programmed as follows.
Solution A (%) Solution B (%) Elution
0–5 100®90 0®10 linear gradient
5–15 90®50 10®50 linear gradient
15–25 50®0 50®100 linear gradient
25–35 0 100 isocratic
35–35.1 0®100 100®0 linear gradient
35.1–40 100 0 re-equilibration
Chromatograph the System suitability solution, and record the peak responses as directed for Procedure. Identify the components based on their relative retention times which are 1.0 for estradiol and about 1.1 for estrone: the resolution, R, between estradiol and estrone is not less than 3.0; the tailing factors for estradiol and estrone peaks are not more than 1.5; and the relative standard deviation for replicate injections is not more than 2% for each peak.
Procedure— Inject a volume (about 10 µL) of the Test solution into the chromatograph, record the chromatogram, and measure the peak responses. Calculate the percentage of each impurity in the portion of Tablets taken by the formula:
100(ri / rs)
in which ri is the peak response for each impurity; and rs is the sum of the responses of all the peaks: not more than 5% of total impurities is found. Disregard any peaks observed in the blank.
Mobile phase, Internal standard solution, Standard preparation, and Chromatographic system— Proceed as directed in the Assay under Estradiol.
Assay preparation— Weigh and finely powder not fewer than 10 Tablets. Transfer a portion of the powder, equivalent to about 8 mg of estradiol, to a 100-mL volumetric flask. Add 4 mL of water, and swirl. Add 10.0 mL of Internal standard solution and about 60 mL of methanol. Shake by mechanical means for 15 minutes, dilute with methanol to volume, mix, and allow the solids to settle. Filter a portion, discarding the first 10 mL of the filtrate. Mix 5.0 mL of the subsequent filtrate with 5.0 mL of methanol and 10.0 mL of water.
Procedure— Proceed as directed for Procedure in the Assay under Estradiol. Calculate the quantity, in mg, of C18H24O2 in the portion of Tablets taken by the formula:
0.4C(RU / RS)
in which the terms are as defined therein.
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