Standard preparation— Transfer an appropriate amount of USP Colestipol Hydrochloride RSinto the probe.If necessary to keep the colestipol hydrochloride in the probe,mix 4parts with 1part of n-eicosane.Grind in a mortar with chloroform until the colestipol hydrochloride is uniformly coated with the n-eicosane.This preparation is stable indefinitely but may require wetting with a small amount of chloroform before each use.

Test preparation— Proceed as directed under Standard preparation,using an appropriate amount of the test specimen.

Chromatographic system (see Chromatography á621ñ)—The gas chromatograph is equipped with a flame-ionization detector and a 3-mm ×180-cm column packed with 80-to 100-mesh support S1Awhich has been coated with 0.25%potassium hydroxide and 5%phase G16.The carrier gas is helium at a flow rate of about 60mLper minute.Maintain the detector and column temperatures at about 270and about 85,respectively.The pyrolysis unit is capable of reaching 1100when equipped with a platinum probe,and pyrolysis time is not less than 10seconds.

Procedure— Install the pyrolysis unit on the chromatograph,and position the probe so that it is just above but not touching the column packing.Set the pyrolysis temperature to about 1100.

Test preparation— Using about 20mg of Colestipol Hydrochloride,accurately weighed,proceed as directed under Oxygen Flask Combustion á471ñ,10mLof 0.05Nsodium hydroxide being used as the absorbing liquid.Do not allow the paper specimen wrapper to come in contact with the liquid,and ignite the paper with an IRigniter.After combustion is complete,shake the flask vigorously,and allow to stand,with frequent shaking,for about 40minutes or until no cloudiness is present.Transfer the solution to a 50-mLbeaker.Wash the flask with two 5-mLportions of water and two 10-mLportions of alcohol,adding each washing to the beaker,and add 0.2mLof nitric acid.

Reagent blank preparation— Using a paper specimen wrapper,complete the combustion,and allow the mixture to stand for about 40minutes or until no cloudiness is present,as directed under Test preparation.Transfer the solution so obtained to a 50-mLbeaker.Wash the combustion flask with two 5-mLportions of water and two 10-mLportions of alcohol,adding the washings to the beaker,and add 0.2mLof nitric acid.

Procedure— Titrate the Test preparationand the Reagent blank preparationwith 0.05Nsilver nitrate VS,determining the endpoint potentiometrically,using a silver-silver chloride electrode and a glass reference electrode (see Titrimetry á541ñ).Determine the volume of 0.05Nsilver nitrate VSconsumed by the test specimen taken by the formula: in which Vand VBare the volumes,in mL,of titrant used for the Test preparationand the Reagent blank preparation,respectively.Each mLof 0.05Nsilver nitrate is equivalent to 1.773mg of Cl:the chloride content is between 6.5%and 9.0%,calculated on the dried basis.

Cholate solution— Dissolve accurately weighed quantities of sodium cholate and sodium chloride in water,and quantitatively dilute with water to obtain a solution having known concentrations of 10.0mg of sodium cholate per mLand 9.0mg of sodium chloride per mL.Adjust the solution by the dropwise addition of hydrochloric acid to a pHof 6.4±0.1.

Test preparation— Transfer 1.0±0.01g of Colestipol Hydrochloride to a glass-stoppered,125-mLconical flask.Add 100.0mLof Cholate solution,insert the stopper securely in the flask,shake vigorously for 90minutes with the flask positioned horizontally on a platform shaker,remove the flask from the shaker,and allow the solids to settle for 5minutes.

Procedure— Transfer 20.0mLof supernatant from the Test preparationto a 40-mLbeaker,transfer 20.0mLof Cholate solutionto a second 40-mLbeaker,and adjust both solutions by the dropwise addition of 1Nsodium hydroxide to a pHof 10.5±0.5.Titrate both solutions with 0.1Nhydrochloric acid VS,determining the endpoints potentiometrically,and measure the titrant volume corresponding to the difference between the midpoints of the two inflections in the titration curves obtained for each solution (see Titrimetry á541ñ).Determine the volume of titrant equivalent to the bound cholate by subtracting the volume of 0.1Nhydrochloric acid VSused in titrating the Test preparationfrom that used in titrating the Cholate solution.Calculate the Cholate binding capacity,in mEq per g,taken by the formula: in which Vis the volume,in mL,of titrant equivalent to the bound cholate;Nis the normality of the 0.1Nhydrochloric acid VS;and Wis the weight,in g,of Colestipol Hydrochloride taken for the Test preparation.The Cholate binding capacityis between 1.1mEq per g and 1.6mEq per g.

Resin base preparation— Combine not less than 2g of Colestipol Hydrochloride and 100mLof 1Nsodium hydroxide in a 125-mLconical flask,insert a stopper in the flask,secure the flask on a platform shaker,and shake the mixture for 3to 4hours.Filter the suspension through a coarse-porosity,fritted-glass funnel,and wash the resin with 500mLof water.Transfer the resin to a 1000-mLbeaker,add 200mLof water,allow to stand for 10minutes,filter the suspension,and check the pHof the filtrate.Repeat the washing procedure with 200-mLportions of water until the pHof the filtrate is below 8(as much as 5000mLof water may be required).Dry the colestipol base resin so obtained and the funnel at a pressure of about 5mm of mercury at 60for 16hours.Break up any aggregates,and store the Resin base preparationin a desiccator.

Procedure— Transfer about 1.0g of the Resin base preparationto a 125-mLconical flask,add 100.0mLof 0.20Nhydrochloric acid,insert a stopper in the flask,and shake the mixture by mechanical means for 2.5hours.Filter a portion of the suspension through a pledget of glass wool,and transfer 8.0mLof the filtrate (test preparation)to a 25-mLbeaker.Transfer 5.0mLof the same 0.20Nhydrochloric acid that was used to equilibrate the resin to a second 25-mLbeaker,and add 5.0mLof water.Titrate both solutions with 0.2Nsodium hydroxide VS,determining the endpoints potentiometrically (see Titrimetry á541ñ),and calculate the Colestipol exchange capacity,in mEq per g,taken by the formula: in which Nis the normality of the sodium hydroxide VS;Wis the weight,in g,of the Resin base preparationtaken;Vbis the volume,in mL,of titrant used to neutralize the 5.0-mLaliquot of 0.20Nhydrochloric acid;and Vais the volume,in mL,of titrant used to neutralize the residual acid in the test preparation.Each g exchanges not less than 9.0mEq and not more than 11.0mEq of sodium hydroxide,calculated as colestipol exchange capacity.