A: Transfer a quantity of Suppositories,equivalent to about 150mg of bisacodyl,to a 500-mLconical flask,add 75mLof solvent hexane,and heat on a steam bath until they are melted.Filter the solution,with the aid of vacuum,through a medium-porosity,sintered-glass funnel,and wash the residue with about 100mLof warm solvent hexane until it is free from fat.Continue the vacuum until the residue appears dry.Dissolve the residue by rinsing the filter with about 50mLof warm acetone,collecting the filtrate in a 150-mLbeaker,and evaporate the filtrate on a steam bath to a volume of about 5mL.To the residual liquid add about 75mLof water,heat on a steam bath for 15minutes,and cool.Scratch the sides of the beaker to induce crystallization,filter the crystals,and dry at 100for about 15minutes:the bisacodyl so obtained melts between 129and 135,and responds to Identificationtest Aunder Bisacodyl.

B: The chromatogram of the Assay preparationobtained as directed in the Assayexhibits a major peak for bisacodyl,the retention time of which corresponds to that exhibited in the chromatogram of the Standard preparation.

Mobile phase— Prepare a filtered and degassed mixture of 0.074Msodium acetate in water [adjusted with 2.5%(v/v)acetic acid to a pHof 7.4]and acetonitrile (55:45).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve an accurately weighed quantity of USP Bisacodyl RSin acetonitrile to obtain a Standard preparationhaving a known concentration of about 0.5mg per mL.

Assay preparation— Transfer a number of Suppositories,equivalent to about 100mg of bisacodyl,to a 500-mLseparator,add 150mLof n-hexane,and shake until all the suppositories are dissolved.Add 50mLof acetonitrile,shake for 1minute,and allow the layers to separate.Drain the lower layer into a 200-mLvolumetric flask,and extract the n-hexane layer remaining in the separator with two 50-mLportions of acetonitrile,combining the lower layers in the volumetric flask.Dilute the combined extracts in the volumetric flask with acetonitrile to volume,mix,and filter.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 265-nm detector,a 3.9-mm ×30-cm column that contains packing L1,and a guard column that contains packing L2.The flow rate is about 2mLper 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 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 C22H19NO4in the Suppositories taken by the formula: in which Cis the concentration,in mg per mL,of USP Bisacodyl RSin the Standard preparation;and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.