A: Ultraviolet Absorption á197Uñ—

B: The retention time of the major peak in the chromatogram of the Assay preparationcorresponds to that in the chromatogram of the Standard preparation,as obtained in the Assay.

Dilute phosphoric acid solution— Prepare a solution of phosphoric acid in water (1in 1000).

Mobile phase— Prepare a filtered and degassed mixture of Dilute phosphoric acid solutionand acetonitrile (850:150).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Prepare a solution of USP Norfloxacin RSin Dilute phosphoric acid solutionhaving a known concentration of about 0.06mg per mL.

Resolution solution— Prepare a solution of USP Norfloxacin RSand pipemidic acid in Dilute phosphoric acid solutionhaving known concentrations of about 0.06mg of each per mL.

Assay preparation— Dilute an accurately measured volume of Ophthalmic Solution quantitatively and stepwise with Dilute phosphoric acid solutionto obtain a solution having a concentration of about 0.06mg of norfloxacin per mL.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 278-nm detector and a 3.9-mm ×30-cm column that contains packing L1.The column temperature is maintained at 50.The flow rate is about 0.5mLper minute.Precondition the column for about 8hours with 0.01Mmonobasic sodium phosphate buffer adjusted with phosphoric acid to a pHof 4.0.Chromatograph the Resolution solution,and record the peak responses as directed for Procedure:the relative retention times are about 0.8for pipemidic acid and 1.0for norfloxacin;and the resolution,R,between the pipemidic acid peak and the norfloxacin peak is not less than 1.2.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the tailing factor for the norfloxacin peak is not more than 2.0;and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— [NOTE—Use peak areas where peak responses are indicated.]Separately inject equal volumes (about 10µL)of the Standard preparationand the Assay preparationinto the chromatograph,record the chromatograms,and measure the responses for the major peaks.Calculate the quantity,in mg,of norfloxacin (C16H18FN3O3)in each mLof the Ophthalmic Solution taken by the formula: in which Lis the labeled quantity,in mg per mL,of norfloxacin in the Ophthalmic Solution;Dis the concentration,in mg per mL,of norfloxacin in the Assay preparation,based on the labeled quantity of norfloxacin in each mLof the Ophthalmic Solution and the extent of dilution;Cis the concentration,in mg per mL,of USP Norfloxacin RSin the Standard preparation;and rUand rSare the norfloxacin peak responses obtained from the Assay preparationand the Standard preparation,respectively.