Edition: BP 2025 (Ph. Eur. 11.6 update)
Action and use
Estrogen.
DEFINITION
Estradiol and Norethisterone Tablets contain Estradiol Hemihydrate and Norethisterone. They are coated.
The tablets comply with the requirements stated under Tablets and with the following requirements.
Content of estradiol, C18H24O2
95.0 to 105.0% of the stated amount.
Content of norethisterone, C20H26O2
95.0 to 105.0% of the stated amount.
IDENTIFICATION
A. Carry out the method for thin-layer chromatography, Appendix III A, using the following solutions.
(1) Add 0.2 mL of water to two tablets and shake to disperse. Add sufficient ethanol (96%) to produce a solution containing 0.035% w/v of Norethisterone, centrifuge and use the clear supernatant liquid.
(2) A suitable concentration of estradiol hemihydrate BPCRS and norethisterone BPCRS in ethanol (96%).
CHROMATOGRAPHIC CONDITIONS
(a) Use as the coating silica gel F254 (Merck silica gel 60 F254 plates are suitable).
(b) Use the mobile phase as described below.
(c) Apply 2 μL of each solution.
(d) Develop the plate to 8 cm.
(e) After removal of the plate, dry in air and spray with ethanolic sulfuric acid (5%). Heat the plate at 105° for 15 minutes and examine under ultraviolet light (365 nm).
MOBILE PHASE
10 volumes of acetone and 90 volumes of dichloromethane.
CONFIRMATION
The chromatogram obtained with solution (1) shows two clearly separated spots with Rf values corresponding to those observed in the chromatogram obtained with solution (2).
B. In the test for Uniformity of content, the chromatogram obtained with solution (1) shows two peaks having the same retention times as those due to estradiol and norethisterone in the chromatogram obtained with solution (2).
TESTS
Dissolution
Comply with the requirements for Monographs of the British Pharmacopoeia in the dissolution test for tablets and capsules, Appendix XII B1.
TEST CONDITIONS
(a) Use Apparatus 2, rotating the paddle at 50 revolutions per minute.
(b) Use 500 mL of a 0.3% w/v solution of sodium lauryl sulfate in water, at a temperature of 37°, as the medium.
PROCEDURE
Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) After 45 minutes withdraw a sample of the medium and filter, discarding the first 5 mL of the filtrate. Use the filtered medium.
(2) Dissolve a sufficient quantity of estradiol hemihydrate BPCRS and norethisterone BPCRS in methanol (80%) and dilute an aliquot with a 0.3% w/v solution of sodium lauryl sulfate in water; the concentration of the final solution should be the same as that expected for solution (1).
(3) 0.0017% w/v of estradiol hemihydrate BPCRS, 0.00084% w/v of norethisterone acetate BPCRS, 0.00066% w/v of estrone BPCRS and 0.00034% w/v of norethisterone BPCRS in the mobile phase.
CHROMATOGRAPHIC CONDITIONS
(a) Use a stainless steel column (25 cm × 4.6 mm) packed with end-capped octadecylsilyl silica gel for chromatography (5 μm) (Spherisorb ODS 2 is suitable).
(b) Use isocratic elution and the mobile phase described below.
(c) Use a flow rate of 2 mL per minute.
(d) Use an ambient column temperature.
(e) Use a detection wavelength of 235 nm.
(f) Inject 200 μL of each solution.
MOBILE PHASE
450 volumes of water and 550 volumes of acetonitrile.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (3), the resolution factor between each pair of peaks (estradiol and norethisterone, and estrone and norethisterone acetate) is at least 1.0.
DETERMINATION OF CONTENT
Calculate the total content of estradiol, C18H24O2, and norethisterone, C20H26O2, in the medium from the chromatograms obtained and using the declared content of C18H24O2 in estradiol hemihydrate BPCRS and C20H26O2 in norethisterone BPCRS.
Estrone
Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) Powder 20 tablets. Add 20 mL of the mobile phase to a quantity of the powdered tablets containing the equivalent of 5 mg of estradiol, mix with the aid of ultrasound and add sufficient mobile phase to produce 25 mL. Centrifuge and use the clear supernatant liquid.
(2) 0.0001% w/v of estrone BPCRS in the mobile phase.
(3) 0.0017% w/v of estradiol hemihydrate BPCRS, 0.00084% w/v of norethisterone acetate BPCRS, 0.00066% w/v of estrone BPCRS and 0.00034% w/v of norethisterone BPCRS in the mobile phase.
CHROMATOGRAPHIC CONDITIONS
The chromatographic conditions described under Dissolution may be used, but using an injection volume of 20 μL.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (3), the resolution factor between each pair of peaks (estradiol and norethisterone, and estrone and norethisterone acetate) is at least 1.0.
LIMITS
In the chromatogram obtained with solution (1):
the area of any peak corresponding to estrone is not greater than the area of the principal peak in the chromatogram obtained with solution (2) (0.5%).
Uniformity of content
Tablets containing less than the equivalent of 2 mg and/or less than 2% w/w of estradiol comply with the requirements stated under Tablets using the following method of analysis. Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) Add 20 mL of the mobile phase to one tablet, mix with the aid of ultrasound, cool, add sufficient of the mobile phase to produce 25 mL and centrifuge. Dilute the supernatant liquid, if necessary, with the mobile phase to produce a solution containing the equivalent of 0.002% w/v of estradiol.
(2) Dissolve sufficient quantities of estradiol hemihydrate BPCRS and norethisterone BPCRS in the mobile phase and dilute an aliquot with the mobile phase; the concentrations in the final solution are the same as those expected for solution (1).
(3) 0.0017% w/v of estradiol hemihydrate BPCRS, 0.00084% w/v of norethisterone acetate BPCRS, 0.00066% w/v of estrone BPCRS and 0.00034% w/v of norethisterone BPCRS in the mobile phase.
CHROMATOGRAPHIC CONDITIONS
The chromatographic conditions described under Dissolution may be used, but using an injection volume of 20 μL.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (3), the resolution factor between each pair of peaks (estradiol and norethisterone, and estrone and norethisterone acetate) is at least 1.0.
DETERMINATION OF CONTENT
Calculate the content of C18H24O2 and C20H26O2 in each tablet using the declared content of C18H24O2in estradiol hemihydrate BPCRS and the declared content of C20H26O2 in norethisterone BPCRS.
ASSAY
For estradiol
For tablets containing the equivalent of less than 2 mg and/or less than 2% w/w of estradiol
Use the average of the individual results determined in the test for Uniformity of content.
For tablets containing the equivalent of 2 mg or more and 2% w/w or more of estradiol
Weigh and powder 20 tablets. Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) Add 50 mL of the mobile phase to a quantity of powdered tablets containing the equivalent of 10 mg of estradiol, mix with the aid of ultrasound and centrifuge. Dilute 1 volume of the clear supernatant liquid to 10 volumes with mobile phase.
(2) 0.002% w/v of estradiol hemihydrate BPCRS in the mobile phase.
(3) 0.0017% w/v of estradiol hemihydrate BPCRS, 0.00084% w/v of norethisterone acetate BPCRS, 0.00066% w/v of estrone BPCRS and 0.00034% w/v of norethisterone BPCRS in the mobile phase.
CHROMATOGRAPHIC CONDITIONS
The chromatographic conditions described under Dissolution may be used, but using an injection volume of 20 μL.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (3), the resolution factor between each pair of peaks (estradiol and norethisterone, and estrone and norethisterone acetate) is at least 1.0.
DETERMINATION OF CONTENT
Calculate the total content of estradiol, C18H24O2, in the tablets using the declared content of C18H24O2 in estradiol hemihydrate BPCRS.
For norethisterone
Use the average of the individual results determined in the test for Uniformity of content.
STORAGE
Estradiol and Norethisterone Tablets should be protected from light.
LABELLING
The quantity of Estradiol Hemihydrate is stated in terms of the equivalent amount of estradiol.
IMPURITIES
The impurities limited by the requirements of this monograph include:
A. Estrone.
