A: Transfer an amount of powdered Tablets,equivalent to about 3mg of clonidine hydrochloride,to a beaker,add 30mLof water,stir for 5minutes,and filter,using a medium-porosity,sintered-glass funnel.Transfer the filtrate to a separator,add 5mLof 0.1Nsodium hydroxide,and extract with 20mLof chloroform,collecting the chloroform extract in a separator.Extract the chloroform phase with 15mLof 0.01Nhydrochloric acid,collecting the acid extract in a beaker.Remove any residual chloroform from the acid extract by heating on a steam bath:the UVabsorption spectrum of the solution so obtained exhibits maxima and minima at the same wavelengths as that of a similar solution of USP Clonidine Hydrochloride RS,concomitantly measured.

B: Transfer 10powdered Tablets to a 50-mLbeaker,add 10mLof methanol,boil on a steam bath for 5minutes,and filter.Add 20mLof water to the filtrate,and boil on a steam bath for 5minutes under a current of air.Cool,with stirring,in ice until crystals form,filter the crystals,and dry at 105for 1hour:the IRabsorption spectrum of a mineral oil dispersion of the dried crystals exhibits maxima only at the same wavelengths as that of a similar preparation of USP Chlorthalidone RS.

C: The retention times of the chlorthalidone and clonidine hydrochloride peaks in the chromatogram of the Assay preparationcorrespond to those of the Standard preparationas obtained in the Assay.

Medium: water;900mL.

Apparatus 2: 100rpm.

Time: 60minutes.

Procedure— Pipet 20mLof a centrifuged portion of the solution under test into a 25-mLvolumetric flask,and dilute with 0.5%monobasic ammonium phosphate solution to volume.Use the resulting solution as the Assay preparation.Determine the amounts of chlorthalidone (C14H11ClN2O4S)and clonidine hydrochloride (C9H9Cl2N3·HCl)dissolved,employing the procedure set forth in the Assay,making any necessary volumetric adjustments.

Tolerances— Not less than 50%(Q)of the labeled amount of C14H11ClN2O4Sand not less than 80%(Q)of the labeled amount of C9H9Cl2N3·HCl are dissolved in 60minutes.

Mobile phase— Dissolve 800mg of monobasic ammonium phosphate in 800mLof water,add 100mLof methanol and 100mLof acetonitrile,mix,filter,and degas.Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve an accurately weighed quantity of USP Clonidine Hydrochloride RSin 0.1%monobasic ammonium phosphate solution,and dilute quantitatively with the same solvent to obtain a solution having a known concentration of about 1500Jµg per mL,Jbeing the ratio of the labeled amount,in mg,of clonidine hydrochloride to the labeled amount,in mg,of chlorthalidone per Tablet (Solution P).Transfer about 15mg of USP Chlorthalidone RS,accurately weighed,to a 100-mLvolumetric flask,dissolve in 10mLof methanol,add 25mLof 0.1%monobasic ammonium phosphate solution and 10.0mLof Solution P,dilute with 0.1%monobasic ammonium phosphate solution to volume,and mix to obtain a solution having known concentrations of about 150Jµg of USP Clonidine Hydrochloride RSper mLand about 150µg of USP Chlorthalidone RSper mL.

Assay preparation— Weigh and finely powder not fewer than 20Tablets.Transfer an accurately weighed portion of the powder,equivalent to about 15mg of chlorthalidone,to a 100-mLvolumetric flask,add 10mLof methanol,and sonicate for 5minutes.Add 40mLof 0.1%monobasic ammonium phosphate solution,sonicate until the solution is free from agglomerates,allow to cool to ambient temperature,dilute with 0.1%monobasic ammonium phosphate solution to volume,mix,and centrifuge.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 220-nm detector and a 4.6-mm ×10-cm column that contains packing L7.The flow rate is about 2mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the relative retention times are about 0.2for clonidine hydrochloride and 1.0for chlorthalidone;the resolution,R,between the clonidine hydrochloride and chlorthalidone peaks is not less than 3;and the relative standard deviation for replicate injections is not more than 2%.

Procedure— Separately inject equal volumes (about 20µL)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 clonidine hydrochloride (C9H9Cl2N3·HCl)in the portion of Tablets taken by the formula: in which Cis the concentration,in µg per mL,of USP Clonidine Hydrochloride RSin the Standard preparation;and rUand rSare the peak responses of clonidine hydrochloride obtained from the Assay preparationand the Standard preparation,respectively.Calculate the quantity,in mg,of chlorthalidone (C14H11ClN2O4S)in the portion of Tablets taken by the same formula,changing the terms to refer to chlorthalidone.