A: It meets the requirements under Identification—Organic Nitrogenous Bases 181.

B:Ultraviolet Absorption 197U—

C: It responds to the tests for Citrate 191.

Mobile phase, Standard preparation, System suitability solution, and Chromatographic system— Proceed as directed in the Assay.

Test solution— Use the Assay preparation prepared as directed in the Assay.

Procedure— Inject a volume (about 50 µL) of the Test solution into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the percentage of (Z)-isomer in the portion of Clomiphene Citrate taken by the formula: in which rZ is the peak response of the (Z)-isomer in the Test solution, and ri is the sum of all of the peak responses obtained from the Test solution.

Mobile phase and System suitability solution— Proceed as directed in the Assay.

Standard solution— Dissolve an accurately weighed quantity of USP Clomiphene Citrate RS in Mobile phase, and dilute quantitatively, and stepwise if necessary, with Mobile phase to obtain a solution having a known concentration of about 1.0 µg per mL.

Test solution— Use the Assay preparation.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 290-nm detector and a 4.6-mm × 25-cm column that contains packing L26. The flow rate is about 1 mL per minute. Chromatograph the System suitability solution, and record the peak responses as directed for Procedure: the relative retention times are about 0.9 for clomiphene related compound A, 1.0 for the (Z)-isomer, and 1.2 for the (E)-isomer; the resolution, R, between clomiphene related compound A and the (Z)-isomer is not less than 1.0; and the resolution, R, between the (Z)-isomer and the (E)-isomer is not less than 1.5. Chromatograph the Standard solution, and record the peak responses as directed for Procedure: the column efficiency is not less than 2000 theoretical plates for the (E)-isomer; the tailing factor is not more than 3.0 for the (E)-isomer; and the relative standard deviation for replicate injections is not more than 2.0% determined from both the (E)- and (Z)-isomers.

Procedure— Inject a volume (about 50 µL) of the Test solution into the chromatograph, record the chromatogram, and measure the peak responses. Calculate the percentage of each impurity in the portion of Clomiphene Citrate taken by the formula: in which ri is the peak response for each impurity; and rs is the sum of the responses of all of the peaks: not more than 2.0% of clomiphene related compound A is found; not more than 0.5% of any other individual impurity is found; and not more than 2.0% of total impurities is found.

Solvent: dimethyl sulfoxide.

(Official January 1, 2007)

Mobile phase— Prepare a filtered and degassed mixture of methanol, water, and triethylamine (55:45:0.3). Adjust with phosphoric acid to a pH of 2.5. Make adjustments if necessary (see System Suitability under Chromatography 621).

System suitability solution— Dissolve suitable quantities of USP Clomiphene Related Compound A RS and USP Clomiphene Citrate RS in Mobile phase to obtain a solution containing about 0.002 and 0.05 mg per mL, respectively.

Standard preparation— [NOTE—Use actinic glassware for all solutions.] Dissolve an accurately weighed quantity of USP Clomiphene Citrate RS in Mobile phase, and dilute quantitatively, and stepwise if necessary, with Mobile phase to obtain a solution having a known concentration of about 0.05 mg per mL.

Assay preparation— Transfer about 50 mg of the Clomiphene Citrate, accurately weighed, to a 100-mL volumetric flask, dilute with Mobile phase to volume, mix, and filter. Transfer 10.0 mL of this solution to a 100-mL volumetric flask, dilute with Mobile phase to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 233-nm detector and a 4.6-mm × 25-cm column that contains packing L26. The flow rate is about 1 mL per minute. Chromatograph the System suitability solution, and record the peak responses as directed for Procedure: the relative retention times are about 0.9 for clomiphene related compound A, 1.0 for (Z)-isomer, and 1.2 for (E)-isomer; and the resolution, R, between clomiphene related compound A and (Z)-isomer is not less than 1.0 and between (Z)-isomer and (E)-isomer is not less than 1.5. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the column efficiency is not less than 2000 theoretical plates for the (E)-isomer; the tailing factor is not more than 3.0 for the (E)-isomer; and the relative standard deviation for replicate injections is not more than 2.0% for both (E)- and (Z)-isomers.

Procedure— Separately inject equal volumes (about 50 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 C26H28ClNO·C6H8O7 in the portion of Clomiphene Citrate taken by the formula: in which C is the concentration, in mg per mL, of USP Clomiphene Citrate RS in the Standard preparation; rUE and rUZ are the peak responses obtained from the Assay preparation for the (E)- and (Z)-isomers, respectively; and rSE and rSZ are the peak responses obtained from the Standard preparation for the (E)- and (Z)-isomers, respectively.