Mobile phase, Standard solution, System suitability solution, and Chromatographic system— Proceed as directed in the test for Limit of ondansetron related compound D under Ondansetron Hydrochloride.

Test solution— Transfer an accurately measured volume of Injection, equivalent to about 10 mg of ondansetron, to a 25-mL volumetric flask, dilute with Mobile phase to volume, and mix.

Procedure— Separately inject equal volumes (about 20 µL) of the Standard solution and the Test solution into the chromatograph, record the chromatograms, and measure the responses for the major peaks. Calculate the quantity of ondansetron related compound D in the volume of Injection taken by the formula: in which V is the volume, in mL, of Injection taken; CS is the concentration, in µg per mL, of ondansetron related compound D in the Standard preparation; CA is the concentration, in mg per mL, of ondansetron in the Injection, as determined in the Assay; and rU and rS are the peak responses obtained from the Test preparation and the Standard preparation, respectively: not more than 0.12% is found.

Mobile phase and Chromatographic system— Proceed as directed in the Assay.

Standard solution— Proceed as directed for Standard preparation in the Assay.

Test solution— Use the Assay preparation.

Procedure— Inject a volume (about 10 µL) of the Test solution into the chromatograph, record the chromatogram, and measure the peak responses. Calculate the percentage of each impurity in the volume of Injection 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 0.2% of any individual impurity is found, and the total of all impurities is not more than 0.5%.

(Official January 1, 2007)

Mobile phase— Prepare a filtered and degassed mixture of 0.02 M monobasic potassium phosphate (previously adjusted with 1 M sodium hydroxide to a pH of 5.4), and acetonitrile (50:50). Make adjustments if necessary (see System Suitability under Chromatography 621).

Standard preparation— Dissolve an accurately weighed quantity of USP Ondansetron Hydrochloride RS in Mobile phase, and dilute quantitatively, and stepwise if necessary, with Mobile phase to obtain a solution having a known concentration of about 0.1 mg per mL.

System suitability solution— Dissolve suitable quantities of USP Ondansetron Hydrochloride RS and USP Ondansetron Related Compound A RS in Mobile phase, and dilute quantitatively, and stepwise if necessary, with Mobile phase to obtain a solution containing about 0.1 mg per mL and 50 µg per mL, respectively.

Assay preparation— Transfer an accurately measured volume of Injection, equivalent to about 2 mg of ondansetron, to a 25-mL volumetric flask, dilute with Mobile phase to volume, and mix.

Chromatographic system (see Chromatography 621)— The liquid chromatograph is equipped with a 216-nm detector and a 4.6-mm × 20-cm column that contains packing L10. The flow rate is about 1.5 mL per minute. Chromatograph the System suitability solution, and record the peak responses as directed for Procedure: the relative retention times are about 1.0 for ondansetron and 1.1 for ondansetron related compound A; and the resolution, R, between ondansetron related compound A and ondansetron is not less than 1.5. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the tailing factor is not more than 2.0; and the relative standard deviation for replicate injections is not more than 1.5%.

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 responses for the major peaks. Calculate the quantity, in mg, of ondansetron (C18H19N3O) in each mL of the Injection taken by the formula: in which 293.36 and 329.82 are the molecular weights of ondansetron and anhydrous ondansetron hydrochloride, respectively; C is the concentration, in mg per mL, on the anhydrous basis, of USP Ondansetron Hydrochloride RS in the Standard preparation; V is the volume, in mL, of Injection taken; and rU and rS are the peak responses obtained from the Assay preparation and the Standard preparation, respectively.