A: Infrared Absorption 197K.

B: Ultraviolet Absorption 197U—

C: The principal spot from the Test preparation found in the test for Related alkaloids corresponds in RF value to that obtained from the Standard preparation.

Test solution: 25 mg per mL, in a mixture of chloroform, alcohol, and ammonium hydroxide (10:10:1).

Solvent mixture— Mix 10 volumes of chloroform, 10 volumes of methanol, and 1 volume of ammonium hydroxide.

Test solution— Prepare a solution of Dihydroergotamine Mesylate in Solvent mixture to contain 20 mg per mL.

Standard solution and Standard dilutions—Prepare a solution of USP Dihydroergotamine Mesylate RS in Solvent mixture to contain 20 mg per mL (Standard solution). Prepare a series of dilutions of the Standard solution in Solvent mixture to contain 0.40 mg, 0.20 mg, and 0.10 mg per mL (Standard dilutions).

Procedure— In a suitable chromatographic chamber arranged for thin-layer chromatography place a volume of a solvent system consisting of a mixture of chloroform and alcohol (9:1) sufficient to develop the chromatogram, cover, and allow to equilibrate for 30 minutes. Apply 5-µL portions of the Test solution, the Standard solution, and each of the three Standard dilutions to a suitable thin-layer chromatographic plate coated with a 0.25-mm layer of chromatographic silica gel. Allow the spots to dry, and develop the chromatogram until the solvent front has moved about three-fourths of the length of the plate. Remove the plate from the developing chamber, mark the solvent front, and allow the solvent to evaporate. Locate the spots on the plate by lightly spraying with a solution prepared by dissolving 800 mg of p-dimethylaminobenzaldehyde in a cooled mixture of 80 g of alcohol and 20 g of sulfuric acid. The RF value of the principal spot obtained from the Test solution corresponds to that obtained from the Standard solution. Estimate the concentration of any other spots observed in the lane for the Test solution by comparison with the Standard dilutions. The spots from the 0.40-, 0.20-, and 0.10-mg-per-mL dilutions are equivalent to 2.0%, 1.0%, and 0.50% of impurities, respectively. The sum of the impurities is not greater than 2.0%.

(Official January 1, 2007)

Change to read:

Diluent 1— Prepare a solution of 0.1 mL of phosphoric acid in 1000 mL of water.

Diluent 2— Prepare a mixture of Diluent 1 and acetonitrile (60:40).

Solution A— Prepare a filtered and degassed mixture of water, 25 percent ammonia water, and 98% formic acid (1000:10:5). Adjust the pH to 8.50.

Solution B— Prepare a filtered and degassed mixture of acetonitrile and Solution A (80:20).

Mobile phase— Use variable mixtures of Solution A and Solution B as directed for Chromatographic system. Make adjustments to either solution as necessary (see System Suitability under Chromatography 621).

Standard preparation— Dissolve an accurately weighed quantity of USP Dihydroergotamine Mesylate RS in acetonitrile, and dilute quantitatively, and stepwise if necessary, with Diluent 1 to obtain a solution having a known concentration of about 0.6 mg per mL. [NOTE—The final ratio of acetonitrile and Diluent 1 should be similar to the final ratio obtained in the Assay preparation.]

Assay preparation— Transfer about 30 mg of Dihydroergotamine Mesylate, accurately weighed, to a 50-mL volumetric flask, dissolve in 20 mL of acetonitrile, dilute with Diluent 1 to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 280-nm detector and 4.0-mm × 25-cm column that contains packing L1. The flow rate is about 1.5 mL per minute. The chromatograph is programmed as follows. Chromatograph the Standard preparation, and record the peak areas as directed for Procedure: the tailing factor is between 0.8 and 1.5; 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 areas for the analyte peaks. Calculate the quantity, in mg, of C33H37N5O5·CH4O3S in the portion of Dihydroergotamine Mesylate taken by the formula: in which C is the concentration, in mg per mL, of USP Dihydroergotamine Mesylate RS in the Standard preparation; and rU and rS are the peak areas obtained from the Assay preparation and the Standard preparation, respectively.USP29