Ordinary impurities 466
Test solution:
methanol.
Standard solution:
methanol.
Application volume:
10 µL.
Eluant:
a mixture of toluene and isopropyl alcohol (90:10), in a nonequilibrated chamber.
Visualization:
5.
Assay
Mobile phase
Prepare a filtered and degassed mixture of water, acetonitrile, and glacial acetic acid (800:700:1.5). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Internal standard solution
Transfer about 35 mg of progesterone to a 50-mL volumetric flask, add Mobile phase to volume, and mix.
Standard preparation
Dissolve an accurately weighed quantity of
USP Betamethasone Acetate RS in
Mobile phase, and quantitatively dilute with
Mobile phase to obtain a solution containing about 0.5 mg per mL. Transfer 10.0 mL of the resulting solution to a 50-mL volumetric flask, add 10.0 mL of
Internal standard solution, dilute with
Mobile phase to volume, and mix to obtain a solution having a known concentration of about 0.1 mg of
USP Betamethasone Acetate RS per mL.
Assay preparation
Transfer about 50 mg of Betamethasone Acetate, accurately weighed, to a 100-mL volumetric flask, add Mobile phase to volume, and mix. Transfer 10.0 mL of the resulting solution to a 50-mL volumetric flask, add 10.0 mL of Internal standard solution, dilute with Mobile phase to volume, and mix.
Chromatographic system (see Chromatography 621)
The liquid chromatograph is equipped with a 254-nm detector and a 4-mm × 30-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the
Standard preparation, and record the peak responses as directed for
Procedure: the relative retention times are about 3 for progesterone and 1.0 for betamethasone acetate; the resolution,
R, between the analyte and internal standard peaks is not less than 2; and the relative standard deviation for replicate injections is not more than 2.0%.
Procedure
Separately inject equal volumes (about 25 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 C
24H
31FO
6 in the portion of Betamethasone Acetate taken by the formula:
500C(RU / RS)
in which
C is the concentration, in mg per mL, of
USP Betamethasone Acetate RS in the
Standard preparation; and
RU and
RS are the peak response ratios obtained from the
Assay preparation and the
Standard preparation, respectively.