Edition: BP 2025 (Ph. Eur. 11.6 update)
Action and use
Estrogen.
DEFINITION
Estriol Cream contains Estriol in a suitable basis.
The cream complies with the requirements stated under Topical Semi-solid Preparations and with the following requirements.
Content of estriol, C18H24O3
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 in methanol. (1) To a quantity of cream containing 2 mg of Estriol add 50 mL of solvent, heat on a water bath and mix until the cream has completely dispersed, cool the contents in ice, centrifuge and use the supernatant liquid.
(2) 0.004% w/v of estriol BPCRS.
(3) 0.004% w/v each of estriol BPCRS and prednisolone BPCRS.
CHROMATOGRAPHIC CONDITIONS
(a) Use a silica gel 60 high performance precoated plate (Merck HPTLC silica gel 60 is suitable).
(b) Use the mobile phase as described below.
(c) Apply 5 μL of each solution.
(d) Develop the plate to 15 cm.
(e) After removal of the plate, dry the plate in a horizontal position at 100 to 105°. Dilute 2 mL of sulfuric acid to 100 mL with ethanol (96%) and spray the solution evenly onto the plate. Examine the plate in daylight.
MOBILE PHASE
10 volumes of methanol and 90 volumes of dichloromethane.
SYSTEM SUITABILITY
The test is not valid unless the chromatogram obtained with solution (3) shows two clearly separated spots.
CONFIRMATION
The principal spot in the chromatogram obtained with solution (1) is similar in position, colour and size to that in the chromatogram obtained with solution (2).
B. In the Assay, the chromatogram obtained with solution (1) shows a peak with the same retention time as the principal peak in the chromatogram obtained with solution (2).
TESTS
Acidity
pH, 3.5 to 4.5, Appendix V L.
Disperse 10 g of the cream in 10 mL of water, with continuous stirring throughout the pH determination.
Related substances
Carry out the method for liquid chromatography, Appendix III D, using the following solutions in methanol.
(1) To a quantity of cream containing 0.5 mg of Estriol add 20 mL of solvent, heat on a water bath and mix until the cream has completely dissolved, allow to cool to room temperature and add sufficient solvent to produce 25 mL. Cool the contents in ice for 15 minutes, centrifuge and use the supernatant liquid.
(2) Dilute 5 volumes of solution (1) to 100 volumes, further dilute 10 volumes of this solution to 100 volumes.
(3) Dilute 10 volumes of solution (2) to 100 volumes.
(4) 0.01% w/v of estriol impurity standard BPCRS.
CHROMATOGRAPHIC CONDITIONS
(a) Use a stainless steel column (25 cm × 4.6 mm) packed with octadecylsilyl silica gel for chromatography (5 μm) (Nucleosil C18 is suitable).
(b) Use isocratic elution and the mobile phase described below.
(c) Use a flow rate of 1 mL per minute.
(d) Use a column temperature of 35°.
(e) Use a detection wavelength of 280 nm.
(f) Inject 100 μL of each solution.
MOBILE PHASE
5 volumes of methanol, 38 volumes of acetonitrile and 57 volumes of water.
For solutions (3) and (4), when the chromatograms are recorded under the prescribed conditions, the retention times for estriol and 16-epi-estriol are about 5 minutes and about 7 minutes respectively.
SYSTEM SUITABILITY
The test is not valid unless, (a) in the chromatogram obtained with solution (4), the resolution factor between the peaks due to estriol and 16-epi-estriol is at least 2.5 and (b) in the chromatogram obtained with solution (3) the signal-to-noise ratio of the principal peak is at least 10.
LIMITS
In the chromatogram obtained with solution (1):
the area of any secondary peak is not greater than the area of the principal peak in the chromatogram obtained with solution (2) (0.5%);
the sum of the areas of any secondary peaks is not greater than twice the principal peak in the chromatogram obtained with solution (2) (1%).
Disregard any peak with an area less than the area of the principal peak in the chromatogram obtained with solution (3) (0.05%).
ASSAY
Carry out the method for liquid chromatography, Appendix III D, using the following solutions in methanol.
(1) To a quantity of cream containing 0.5 mg of Estriol add 20 mL of methanol, heat on a water bath and mix until the cream has completely dispersed, allow to cool to room temperature and add sufficient methanol to produce 25 mL, transfer to a centrifuge tube and cool the contents in ice for 15 minutes, centrifuge and dilute 5 volumes of the supernatant liquid to 100 volumes with methanol.
(2) 0.0001% w/v of estriol BPCRS.
(3) 0.01% w/v of estriol impurity standard BPCRS.
CHROMATOGRAPHIC CONDITIONS
The chromatographic procedure described under Related substances may be used.
SYSTEM SUITABILITY
The test is not valid unless, in the chromatogram obtained with solution (3), the resolution factor between the peaks due to estriol and 16-epi-estriol is at least 2.5.
DETERMINATION OF CONTENT
Calculate the content of C18H24O3 in the cream using the declared content of C18H24O3 in estriol BPCRS.
