pH7.5buffer— Dissolve 6.8g of monobasic potassium phosphate in 1000mLof water,add 5mLof triethylamine,and adjust with 5Npotassium hydroxide to a pHof 7.5.Filter through a filter having a porosity of 0.5µm or finer,and degas.

Mobile phase— Prepare a mixture of pH7.5bufferand acetonitrile (60:40).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Transfer about 25mg of USP Proparacaine Hydrochloride RS,accurately weighed,to a 25-mLvolumetric flask,dissolve in and dilute with water to volume,and mix.Transfer 5.0mLof this stock solution to a 50-mLvolumetric flask,dilute with Mobile phaseto volume,and mix.[NOTE—Use this solution within 6hours.]

Assay preparation— Transfer an accurately measured volume of Ophthalmic Solution,equivalent to about 10mg of proparacaine hydrochloride,to a 100-mLvolumetric flask,dilute with Mobile phaseto volume,and mix.[NOTE—Use this solution within 6hours.]

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 270-nm detector and a 4.6-mm ×15-cm column that contains 5-µm spherical packing L10.The flow rate is about 1.5mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the tailing factor is not more than 1.5,the column efficiency is not less than 3000theoretical plates,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 C16H26N2O3·HCl in each mLof the Ophthalmic Solution taken by the formula: in which Cis the concentration,in mg per mL,of USP Proparacaine Hydrochloride RSin the Standard preparation,Vis the volume,in mL,of Ophthalmic Solution taken,and rUand rSare the proparacaine peak responses obtained from the Assay preparationand the Standard preparation,respectively.