Medium: water;900mL.

Apparatus 1: 100rpm.

Time: 45minutes.

Phosphate buffer— Dissolve 38.0g of monobasic sodium phosphate and 2.0g of anhydrous dibasic sodium phosphate in water to obtain 1000mLof solution.

Bromocresol green solution— Dissolve 200mg of bromocresol green in 30mLof water and 6.5mLof 0.1Nsodium hydroxide.Dilute with Phosphate bufferto 500mL,mix,and add 0.1Nhydrochloric acid to adjust to a pHof 4.6±0.1.

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

Procedure— Into 3separate,glass-stoppered,50-mLcentrifuge tubes pipet (a)1mLof a filtered portion of the solution under test,(b)1mLof Standard preparation,and (c)1mLof water to provide a blank.Add 5.0mLof Bromocresol green solutionto each tube,mix,add 10.0mLof chloroform to each,insert the stoppers,and shake the mixtures vigorously.Allow the mixtures to separate,discard the upper aqueous layers,and filter the 3chloroform layers through separate pledgets of cotton.Determine the amount of C10H24N2O2·2HCl dissolved from absorbances,at the wavelength of maximum absorbance at about 415nm,obtained from the test solution in comparison with those obtained from the Standard solution,using the blank to set the instrument.

Tolerances— Not less than 75%(Q)of the labeled amount of C10H24N2O2·2HCl is dissolved in 45minutes.

Borate buffer,Standard solution,andFluorescamine solution— Prepare as directed in the test for Limit of aminobutanolunder Ethambutol Hydrochloride.

Test solution— Place a number of Tablets,equivalent to 400mg of ethambutol hydrochloride,in a beaker,cover with acetone,and allow to stand for 15minutes.Decant the acetone,dry the tablets,and remove the coating.Grind the tablet cores in a mortar to a fine powder,moisten with methanol,and triturate to a fine paste.Transfer the mixture with the aid of methanol to a 100-mLvolumetric flask,dilute with methanol to volume,and mix.Filter the mixture through a dry,folded filter paper.Pipet 25mLof the filtrate into a 200-mLvolumetric flask,and dilute with water to volume.Allow to stand for 15minutes,and filter through a dry,folded filter paper,discarding the first cloudy portions of the filtrate.The clear filtrate is the Test solution.

Procedure— Proceed as directed for Procedurein the test for Limit of aminobutanolunder Ethambutol Hydrochloride.

Buffer— Mix 1.0mLof triethylamine with 1liter of water,and adjust with phosphoric acid to a pHof 7.0.

Mobile phase— Prepare a filtered and degassed mixture of Bufferand acetonitrile (1:1).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve an accurately weighed quantity of USP Ethambutol Hydrochloride RSin water to obtain a solution having a known concentration of about 0.30mg of ethambutol hydrochloride per mL.

Assay preparation— Weigh and finely powder not fewer than 20Tablets.Transfer an accurately weighed portion of the powder,equivalent to about 30mg of ethambutol hydrochloride,to a 100-mLvolumetric flask,dissolve in water with the aid of sonication,and dilute with water to volume.Filter the solution,discarding the first 10-mLportion.

Chromatographic system (see Chromatography á621ñ)— The liquid chromatograph is equipped with a 200-nm detector and a 4.6-mm ×15-cm base-deactivated column that contains 5-µm packing L10.The flow rate is about 1.0mLper minute.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 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 ethambutol hydrochloride (C10H24N2O2·HCl)present in the portion of Tablets taken by the formula: in which Cis the concentration,in mg per mL,of USP Ethambutol Hydrochloride RSin the Standard preparation;and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.