Identification—
Weigh an amount of Gel, equivalent to about 2.5 mg of fluocinonide, into a glass-stoppered, 50-mL centrifuge tube containing 20 mL of sodium chloride solution (1 in 10). Add 5 mL of chloroform and 15 mL of methanol, and shake vigorously. Centrifuge to clarify the chloroform layer, and remove the solid material present at the interphase. Discard the upper phase. Dry a portion of the chloroform layer over anhydrous sodium sulfate. Using the dried extract as the
Test preparation, proceed as directed in the
Identification test under
Fluocinonide Cream, beginning with “Apply 10 µL of the
Test solution.”
Assay—
Mobile phase—
Prepare a mixture of acetonitrile and water (1:1). Make adjustments if necessary (see
System Suitability under
Chromatography 
621
).
Standard preparation—
Dissolve an accurately weighed quantity of
USP Fluocinonide RS in acetonitrile to obtain a solution having a known concentration of about 200 µg per mL. Transfer 10.0 mL of this solution to a 100-mL volumetric flask, dilute with acetonitrile to volume, and mix. The final concentration is 20 µg per mL.
Assay preparation—
Transfer an accurately weighed quantity of Gel, containing about 2 mg of fluocinonide, to a 100-mL volumetric flask. Add about 60 mL of acetonitrile, and dissolve the gel by heating on a steam bath. Cool to room temperature, dilute with acetonitrile to volume, and mix. Centrifuge a portion at about 2500 rpm for about 5 minutes. Filter a portion of the centrifugate through an acetonitrile-insoluble membrane filter. The filtrate is the Assay preparation.
Chromatographic system
(see
Chromatography 621)—The liquid chromatograph is equipped with a 254-nm detector and a 3.9-mm × 30-cm column that contains packing L1. The flow rate is about 2 mL per minute. Chromatograph the
Standard preparation, and record the peak responses as directed for
Procedure: the relative standard deviation for replicate injections is not more than 1.5%.
Procedure—
Separately inject equal volumes (about 20 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
26H
32F
2O
7 in the portion of Gel taken by the formula:
0.1C(rU / rS),
in which
C is the concentration, in µg per mL, of
USP Fluocinonide RS in the
Standard preparation; and
rU and
rS are the peak responses due to fluocinonide obtained from the
Assay preparation and the
Standard preparation, respectively.
