Identification—
Transfer a volume of Injectable Suspension, equivalent to about 5 mg of estrone, to a glass-stoppered centrifuge tube, and add 2.5 mL of a mixture of ether and benzene (1:1). Shake for 2 minutes, and allow insoluble matter to settle, centrifuging, if necessary, to obtain a clear supernatant. Apply 5 µL each of this supernatant and a 1 in 500 solution of
USP Estrone RS in a mixture of ether and benzene (1:1) to a suitable thin-layer chromatographic plate (see
Chromatography 
621
), coated with a 0.25-mm layer of chromatographic silica gel. Allow the spots to dry, and develop the chromatogram in a solvent system consisting of a mixture of benzene and acetone (4:1) 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. Spray the plate with a mixture of dehydrated alcohol and sulfuric acid (3:1), and heat in an oven at 105
for 10 minutes: the
RF value and appearance (pale orange to amber by direct observation in daylight, and fluorescing pale yellow-green under long-wavelength UV light) of the principal spot obtained from the test solution correspond to those obtained from the Standard solution.
Assay—
Mobile phase, Standard preparation, and Chromatographic system—
Prepare as directed in the
Assay under
Estrone.
Assay preparation—
Transfer an accurately measured volume of the well-mixed Injectable Suspension, equivalent to about 10 mg of estrone to a 50-mL volumetric flask. Add 30 mL of methanol and swirl for 5 minutes. Dilute with methanol to volume, and mix. Transfer 5.0 mL of this solution to a 25-mL volumetric flask, dilute with Mobile phase to volume, and mix.
Procedure—
Proceed as directed for
Procedure in the
Assay under
Estrone. Calculate the quantity, in mg, of C
18H
22O
2 in each mL of Injectable Suspension taken by the formula:
0.25(C / V)(rU / rS),
in which
V is the volume, in mL, of the Injectable Suspension taken, and the other terms are as defined therein.
