A: Infrared Absorption á197Kñ.

B: Ultraviolet Absorption á197Uñ—

C: Dissolve about 5mg in 2mLof sulfuric acid:a red color is produced.

Test solution: 5mg per mL,in dioxane.

Mobile phase— Prepare a filtered and degassed mixture of water,tetrahydrofuran,dimethylsulfoxide,and butanol (149:40:10:1).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Diluting solution— Prepare a filtered mixture of water,tetrahydrofuran,and glacial acetic acid (72:25:3).

Standard solution— Dissolve an accurately weighed quantity of USP Methylprednisolone RSin Diluting solution.Dilute quantitatively,and stepwise if necessary,with Diluting solutionto obtain a solution having a known concentration of about 0.01mg per mL.

Test solution— Transfer about 25mg of Methylprednisolone,accurately weighed,to a 25-mLvolumetric flask,dissolve in and dilute with Diluting solutionto volume,and mix.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm ×20-cm column that contains packing L1.The flow rate is about 1mLper minute.Chromatograph the Standard solution,and record the peak responses as directed for Procedure:the column efficiency is not less than 800theoretical plates;and the relative standard deviation for replicate injections is not more than 5.0%.

Procedure— Inject a volume (about 10µL)of the Test solutioninto the chromatograph,record the chromatogram,and measure all of the peak responses.Calculate the amount of each impurity in the portion of Methylprednisolone taken by the formula: in which Cis the concentration,in mg per mL,of USP Methylprednisolone RSin the Standard solution;riis the peak response for each impurity obtained from the Test solution;and rSis the peak response obtained in the Standard solution;not more than 1.0%of any individual impurity is found,and not more than 2.0%of total impurities is found.

Mobile phase— Prepare a solution containing a mixture of butyl chloride,water-saturated butyl chloride,tetrahydrofuran,methanol,and glacial acetic acid (475:475:70:35:30).

Internal standard solution— Dissolve prednisone in a 3in 100solution of glacial acetic acid in chloroform to obtain a solution having a concentration of about 0.2mg per mL.

Standard preparation— Dissolve an accurately weighed quantity of USP Methylprednisolone RSin Internal standard solutionto obtain a solution having a known concentration of about 0.2mg per mL.

Assay preparation— Using about 10mg of Methylprednisolone,accurately weighed,proceed as directed for Standard preparation.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 254-nm detector and a 4-mm ×25-cm column that contains packing L3.The flow rate is about 1mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the resolution,R,between the methylprednisolone and internal standard peaks is not less than 4.0;and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 10µL)of the Standard preparationand the Assay preparationinto the chromatograph,record the chromatograms,and measure the responses for the major peaks:the relative retention times are about 0.7for prednisone and 1.0for methylprednisolone.Calculate the quantity,in mg,of C22H30O5in the portion of Methylprednisolone taken by the formula: in which Cis the concentration,in mg per mL,of USP Methylprednisolone RSin the Standard preparation;and RUand RSare the ratios of the peak responses for the methylprednisolone peak and the internal standard peak obtained from the Assay preparationand the Standard preparation,respectively.