A: Infrared Absorption 197K.

B: Ultraviolet Absorption 197U—

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

Test solution: 5 mg 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 Suitability under Chromatography 621).

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 RS in Diluting solution. Dilute quantitatively, and stepwise if necessary, with Diluting solution to obtain a solution having a known concentration of about 0.01 mg per mL.

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

Chromatographic system (see Chromatography 621)—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 1 mL per minute. Chromatograph the Standard solution, and record the peak responses as directed for Procedure: the column efficiency is not less than 800 theoretical plates; and the relative standard deviation for replicate injections is not more than 5.0%.

Procedure— Inject a volume (about 10 µL) of the Test solution into 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 C is the concentration, in mg per mL, of USP Methylprednisolone RS in the Standard solution; ri is the peak response for each impurity obtained from the Test solution; and rS is 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.

(Official January 1, 2007)

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

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

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

Chromatographic system (see Chromatography 621)—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 1 mL per 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 10 µL) of the Standard preparation and the Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the major peaks: the relative retention times are about 0.7 for prednisone and 1.0 for methylprednisolone. Calculate the quantity, in mg, of C22H30O5 in the portion of Methylprednisolone taken by the formula: in which C is the concentration, in mg per mL, of USP Methylprednisolone RS in the Standard preparation; and RU and RS are the ratios of the peak responses for the methylprednisolone peak and the internal standard peak obtained from the Assay preparation and the Standard preparation, respectively.