A: Using a similar portion of Topical Suspension,proceed as directed in Identificationtest Bunder Neomycin and Polymyxin B Sulfates Cream:the indicated result is obtained.

B: It responds to the Identificationtest under Penicillin G Procaine Intramammary Infusion.

C: The chromatogram of the Assay preparationobtained as directed in the Assay for hydrocortisone acetate and hydrocortisone sodium succinateexhibits major peaks for hydrocortisone acetate and hydrocortisone sodium succinate,the retention times of which,relative to that of the internal standard,correspond to those for hydrocortisone acetate and hydrocortisone hemisuccinate exhibited in the chromatogram of the Standard preparationobtained as directed in the Assay for hydrocortisone acetate and hydrocortisone sodium succinate.

Mobile phase— Prepare a filtered and degassed mixture of butyl chloride,water-saturated butyl chloride,tetrahydrofuran,methanol,and glacial acetic acid (544:544:58:29:25).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Extraction solution— Prepare a mixture of chloroform and glacial acetic acid (1000:30).

Internal standard solution— Prepare a solution in tetrahydrofuran containing about 1.4mg of methylprednisolone per mL.

Standard preparation— Transfer about 7.5mg of USP Hydrocortisone Acetate RS,accurately weighed,and 7.5Jmg of USP Hydrocortisone Hemisuccinate RS,accurately weighed,to a conical flask,Jbeing the ratio of the labeled amount,in mg,of hydrocortisone sodium succinate to the labeled amount,in mg,of hydrocortisone acetate in the Topical Suspension.Add 5.0mLof Internal standard solutionand about 95mLof Extraction solution,and mix.

Resolution solution— Dissolve about 3.7mg of penicillin Gprocaine in 10mLof Standard preparation.

Assay preparation— Transfer an accurately measured portion of well-mixed Topical Suspension,equivalent to about 7.5mg of hydrocortisone acetate,to a conical flask.Add 5.0mLof Internal standard solutionand about 95mLof Extraction solution,and shake by mechanical means for about 15minutes.Centrifuge a portion of this mixture,and use the clear supernatant as the Assay preparation.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 254-nm detector,a guard column containing packing L3,and a 3.9-mm ×30-cm analytical column that contains packing L3.The flow rate is about 1mLper minute.Chromatograph the Resolution solution,and measure the peak responses as directed for Procedure:the relative retention times for penicillin Gprocaine,hydrocortisone acetate,hydrocortisone hemisuccinate,and methylprednisone are about 0.3,0.4,0.7,and 1.0,respectively,and the resolution,R,between penicillin Gprocaine and hydrocortisone acetate is not less than 1.5.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the relative standard deviation of the ratios of the hydrocortisone acetate peak to the internal standard peak and the hydrocortisone sodium succinate peak to the internal standard peak is not more than 2%.

Procedure— Separately inject equal volumes (about 20µL)of the Standard preparationand the Assay preparationinto the chromatograph,record the chromatograms,and measure the responses for the major peaks.Calculate the quantity,in mg,of hydrocortisone acetate in the portion of Topical Suspension taken by the formula: in which Wis the quantity,in mg,of USP Hydrocortisone Acetate RStaken to prepare the Standard preparation,and RUand RSare the ratios of the hydrocortisone acetate peak response to the internal standard peak response obtained from the Assay preparationand the Standard preparation,respectively.Calculate the quantity,in mg,of hydrocortisone sodium succinate in the portion of Topical Suspension taken by the formula: in which 484.52and 462.54are the molecular weights of hydrocortisone sodium succinate and anhydrous hydrocortisone hemisuccinate,respectively,Wis the quantity,in mg,of USP Hydrocortisone Hemisuccinate RStaken to prepare the Standard preparation,and RUand RSare the ratios of the hydrocortisone hemisuccinate peak response to the internal standard peak response obtained from the Assay preparationand the Standard preparation,respectively.