Mobile phase— Prepare a filtered and degassed mixture of pH2.5Bufferand acetonitrile (850:150).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve an accurately weighed quantity of USP Epinephrine Bitartrate RSin water,and dilute quantitatively,and stepwise if necessary,with water to obtain a solution having a known concentration of about 0.1mg per mL.

Assay preparation— Transfer an accurately measured volume of Ophthalmic Solution,freshly mixed and free from air bubbles,equivalent to about 50mg of epinephrine bitartrate,to a 500-mLvolumetric flask,dilute with water to volume,and mix.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 280-nm detector and a 3.2-mm ×25-cm column that contains packing L1.The flow rate is about 1mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed under Procedure:the column efficiency determined from the analyte peak is not less than 2000theoretical plates,the tailing factor for the analyte peak is not more than 2.5,and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 50µL)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 epinephrine (C9H13NO3)in each mLof the Ophthalmic Solution taken by the formula: in which Cis the concentration,in mg per mL,of USP Epinephrine Bitartrate RSin the Standard preparation,Vis the volume,in mL,of Ophthalmic Solution taken,and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.