Standard preparation— Dissolve an accurately weighed quantity of USP Benoxinate Hydrochloride RS in 0.1 N hydrochloric acid to obtain a solution having a known concentration of about 400 µg per mL.

Assay preparation— Transfer a volume of Ophthalmic Solution, equivalent to about 20 mg of benoxinate hydrochloride, to a separator containing 15 mL of water, add 1 mL of ammonium hydroxide, and extract with five 20-mL portions of ether. Wash the combined ether extracts with 10 mL of water, extract the water washing with 10 mL of ether, and add this ether extract to the main extract. Extract the ether solution with three 5-mL portions of 0.1 N hydrochloric acid, collect the acid extracts in a 50-mL volumetric flask, dilute with 0.1 N hydrochloric acid to volume, and mix.

Procedure— Transfer 5.0 mL each of the Standard preparation, the Assay preparation, and 0.1 N hydrochloric acid to provide a blank, to separate 200-mL volumetric flasks. Dilute the contents of each flask with water to volume, and mix. Concomitantly determine the absorbances of the solutions in 1-cm cells at the wavelength of maximum absorbance at about 308 nm, with a suitable spectrophotometer, using the blank to set the instrument. Calculate the quantity, in mg, of C17H28N2O3·HCl in each mL of the Ophthalmic Solution taken by the formula: in which C is the concentration, in µg per mL, of USP Benoxinate Hydrochloride RS in the Standard preparation; V is the volume, in mL, of Ophthalmic Solution taken; and AU and AS are the absorbances of the solutions from the Assay preparation and the Standard preparation, respectively.