Chromatographic purity
Buffer solution, Mobile phase, and Chromatographic system
Prepare as directed under
Assay.
Test preparation
Use the Assay preparation.
Procedure
Inject a volume (about 20 µL) of the
Test preparation into the chromatograph, record the chromatogram obtained for a period of not less than twice the retention time of cyclopentolate, and measure the peak responses. Calculate the percentage of each peak, other than the solvent peak and the cyclopentolate peak, in the specimen of Cyclopentolate Hydrochloride taken by the same formula:
100ri / rt,
in which
ri is the response of each peak and
rt is the sum of the responses of all of the peaks, excluding that of the solvent peak: not more than 1.0% individual impurity and not more than 2.0% total impurities are found.
Assay
Buffer solution
Dissolve 660 mg of dibasic ammonium phosphate in 1000 mL of water. Adjust with phosphoric acid to a pH of 3.0 ± 0.1, and mix.
Mobile phase
Prepare a suitable filtered and degassed mixture of acetonitrile and
Buffer solution (7:3). Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Standard preparation
Dissolve an accurately weighed quantity of
USP Cyclopentolate Hydrochloride RS in water, dilute quantitatively, and stepwise if necessary, with water, and mix to obtain a solution having a known concentration of about 0.1 mg per mL.
Assay preparation
Transfer about 100 mg of Cyclopentolate Hydrochloride, accurately weighed, to a 100-mL volumetric flask, dilute with water to volume, and mix. Transfer 5.0 mL of this solution to a 50-mL volumetric flask, dilute with water to volume, and mix.
Chromatographic system
(see
Chromatography 621)The liquid chromatograph is equipped with a 220-nm detector and a 4.6-mm × 15-cm column that contains packing L15. The flow rate is about 2 mL per minute. Chromatograph the
Standard preparation, and record the peak responses as directed for
Procedure: the column efficiency determined from the analyte peak is not less than 3000 theoretical plates, the tailing factor for the analyte peak 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
17H
25NO
3·HCl in the portion of Cyclopentolate Hydrochloride taken by the formula:
1000C(rU / rS),
in which
C is the concentration, in mg per mL, of
USP Cyclopentolate Hydrochloride RS in the
Standard preparation; and
rU and
rS are the cyclopentolate peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.