Assay
[NOTEConduct this procedure with a minimum exposure to light.
]
Mobile phase
Prepare a filtered and degassed mixture of 0.015
M monobasic potassium phosphate and acetonitrile (4:1). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Solvent mixture
Transfer 2.5 g of tartaric acid to a 1000-mL volumetric flask, add 500 mL of water, and mix with shaking. Dilute with methanol to volume, and allow the mixture to cool before use.
Standard preparation
Transfer about 20 mg of
USP Methylergonovine Maleate RS, accurately weighed, to a 200-mL volumetric flask. Add 150 mL of
Solvent mixture, and shake by mechanical means for 15 minutes. Dilute with
Solvent mixture to volume, and mix. Quantitatively dilute a portion of this solution with
Solvent mixture to obtain a solution having a known concentration of about 4 µg of
USP Methylergonovine Maleate RS per mL.
Assay preparation
Transfer about 100 mg of Methylergonovine Maleate, accurately weighed, to a 500-mL volumetric flask. Add 300 mL of Solvent mixture, and shake by mechanical means for 15 minutes or until completely dissolved. Dilute with Solvent mixture to volume, and mix. Quantitatively dilute a portion of this solution with Solvent mixture to obtain the Assay preparation having a known concentration of about 4 µg of Methylergonovine Maleate per mL.
Chromatographic system
(see
Chromatography 621)The liquid chromatograph is equipped with a fluorometric detector set at an excitation wavelength of 315 nm, and an emission wavelength that is set to zero, using a cutoff filter that passes light from about 418 to 700 nm, and a 4.6-mm × 25-cm column that contains packing L7 maintained at 30
. The flow rate is about 2 mL per minute. Chromatograph the
Standard preparation, and record the peak responses as directed under
Procedure: the column efficiency is not less than 1000 theoretical plates, the tailing factor is not more than 2.0, and the relative standard deviation for replicate injections is not more than 2.0%.
Procedure
Separately inject equal volumes (about 20 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 C
20H
25N
3O
2·C
4H
4O
4 in the Methylergonovine Maleate taken by the formula:
25C(rU / rS),
in which
C is the concentration, in µg per mL, of
USP Methylergonovine Maleate RS in the
Standard preparation, and
rU and
rS are the fluorescence intensity responses obtained from the
Assay preparation and the
Standard preparation, respectively.