Identification—
Weigh an amount of Ointment, equivalent to about 2.5 mg of fluocinonide, into a glass-stoppered, 50-mL centrifuge tube containing 20 mL of cyclohexane. Gently disperse to form a suspension. Add 5 mL of water and 10 mL of methanol. Shake vigorously, allow the phases to separate, and discard the upper phase. Add 20 mL of water and 5 mL of chloroform, shake vigorously, centrifuge, and transfer a portion of the chloroform layer to a small test tube containing about 200 mg of anhydrous sodium sulfate. Mix, and allow to stand until the extract is clear. Using the clear chloroform 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 filtered and degassed mixture of acetonitrile and water (1:1). Adjust the ratio as necessary to obtain suitable chromatographic performance.
Standard preparation—
Dissolve an accurately weighed quantity of
USP Fluocinonide RS in acetonitrile to obtain a solution having a known concentration of about 400 µg per mL. Transfer 10.0 mL of this solution to a 100-mL volumetric flask. Dilute with methanol to volume, and mix. The final concentration of
USP Fluocinonide RS is about 40 µg per mL.
Assay preparation—
Transfer an accurately weighed quantity of Ointment, containing about 1.35 mg of fluocinonide, to a round-bottom, 50-mL centrifuge tube. Add 35.0 mL of methanol. Emulsify, using an ultrasonic probe, and centrifuge to bring the insoluble matter to the bottom. The clear supernatant is the Assay preparation.
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 fluocinonide (C
26H
32F
2O
7) in the portion of Ointment taken by the formula:
0.035C(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.
755
:
meets the requirements.