Internal standard— Butabarbital.

Internal standard solution— Dissolve an accurately weighed quantity of Butabarbital in chloroform,and quantitatively dilute with chloroform to obtain a solution having a known concentration of about 0.7mg per mL.

Standard preparation— Dissolve accurately weighed quantities of USP Secobarbital RSand Butabarbital in chloroform,and quantitatively dilute with chloroform to obtain a solution that contains,in each mL,known amounts of about 1.2mg of USP Secobarbital RSand about 0.9mg of Butabarbital.

Assay preparation— Transfer an accurately measured volume of Oral Solution,equivalent to about 22mg of secobarbital,to a separator,add 1mLof dilute hydrochloric acid (1in 5),and extract with four 10-mLportions of chloroform.Filter the extracts through about 15g of anhydrous sodium sulfate that is supported on a funnel by a small pledget of glass wool.Collect the combined filtrate in a 50-mLvolumetric flask,wash the sodium sulfate with 5mLof chloroform,dilute with chloroform to volume,and mix.Combine 4.0mLof this solution with 2.0mLof Internal standard solutionin a suitable container,and reduce the volume to about 1.5mLby evaporation,with the aid of a stream of dry nitrogen,at room temperature.

Chromatographic system andSystem suitability— Proceed as directed for Chromatographic Systemand System Suitabilityunder Barbiturate Assay á361ñ,the resolution,R,between secobarbital and butabarbital being not less than 3.0.[NOTE—Relative retention times are,approximately,0.6for butabarbital and 1.0for secobarbital.]

Procedure— Proceed as directed for Procedureunder Barbiturate Assay á361ñ.Calculate the quantity,in mg,of secobarbital (C12H18N2O3)in each mLof the Oral Solution taken by the the formula: in which Vis the volume,in mL,of Oral Solution taken;and the other terms are as defined therein.