In the chromatogram obtained from the Assay preparation
in the Assay
, the sum of the responses of any peaks detected, other than the major peak due to trioxsalen, is not more than 2.0% of the total of all the peak responses and the response of the peak occurring at retention time relative to trioxsalen of about 0.75 is not more than 1.5% of the total of all responses.
Prepare a filtered and degassed mixture of methanol and water (70:30). Make adjustments if necessary (see System Suitability
under Chromatography 621
Dissolve an accurately weighed quantity of USP Trioxsalen RS
in tetrahydrofuran to obtain a solution having a known concentration of about 1 mg per mL. Transfer 5.0 mL of this solution to a 100-mL volumetric flask, dilute with Mobile phase
to volume, and mix.
Transfer about 100 mg of Trioxsalen, accurately weighed, to a 100-mL volumetric flask, dissolve in tetrahydrofuran, dilute with tetrahydrofuran to volume, mix, and filter. Transfer 5.0 mL of this solution to a 100-mL volumetric flask, dilute with Mobile phase to volume, and mix.
(see Chromatography 621
)The liquid chromatograph is equipped with a 254-nm detector and a 4.6-nm × 25-cm column that contains packing L1. The flow rate is about 1 mL per minute. Chromatograph the Standard preparation
, and record the peak responses as directed for Procedure:
the tailing factor for the trioxsalen peak is not more than 2.0, and the relative standard deviation for replicate injections is not more than 2.0%.
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 C14
in the portion of Trioxsalen taken by the formula:
2000C(rU / rS),
in which C
is the concentration, in mg per mL, of USP Trioxsalen RS
in the Standard preparation
, and rU
are the peak responses obtained from the Assay preparation
and the Standard preparation