Identification—
To 1 g of Cream add 40.0 mL of a 35% solution of acetonitrile in methanol, and shake until dissolved. To 20.0 mL of this solution add 10.0 mL of isooctane, and mix. Allow the layers to separate, discard the upper layer, and use the bottom layer as the test solution. Cream responds to the
Thin-Layer Chromatographic Identification Test 
201
, a preparation containing 250 µg of
USP Hydrocortisone Acetate RS per mL of methanol being used as the Standard solution, the prepared chromatographic plate being heated for 10 minutes at 105
and cooled, and the chromatogram being developed in a solvent system consisting of a mixture of ethyl acetate, toluene, and acetone (140:40:13) in a paper-lined chromatographic chamber equilibrated in an atmosphere of ammonia vapors.
Assay—
Mobile phase
, Chromatographic system, and Standard preparation—Proceed as directed in the Assay under Hydrocortisone Acetate.
Assay preparation—
Transfer an accurately weighed quantity of Cream, equivalent to about 25 mg of hydrocortisone acetate, to a suitable container, add 100.0 mL of tetrahydrofuran, and shake until the cream dissolves. Transfer 10.0 mL of the resulting solution to another container, add 15.0 mL of Mobile phase, and mix.
Procedure—
Separately inject equal volumes (about 10 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 hydrocortisone acetate (C
23H
32O
6) in the portion of Cream taken by the formula:
250C(rU / rS),
in which
C is the concentration, in mg per mL, of
USP Hydrocortisone Acetate RS in the
Standard preparation; and
rU and
rS are the hydrocortisone acetate peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.
