Chromatographic tubes ,Citrate buffer,and Standard preparation—Prepare as directed in the Assayunder Paregoric.

Assay preparation— Transfer about 2g of Opium,accurately weighed,to a 250-mLbeaker,add 20mLof methyl sulfoxide,and heat for 20minutes on a steam bath,intermittently dispersing the substance with a flat-end stirring rod.Allow to stand for 15minutes to permit undissolved material to settle,and carefully decant the supernatant into a 100-mLvolumetric flask.Add another 20mLof methyl sulfoxide to the residue,rinsing the sides of the beaker with the methyl sulfoxide.Disperse and heat the substance as before,allow to settle,and decant into the volumetric flask.Repeat the dissolution one or two times,until the opium is dissolved (other than for small leaf fragments,sand-like particles,gelatinous materials,etc.).Rinse the beaker,and transfer the residue to the flask with the aid of water.Dilute with water to about 90mL,and mix.If necessary,add 1drop of alcohol to dispel any foam.Cool to room temperature,adjust with water to volume,and mix.Filter the resulting solution through a medium-porosity filter paper,discarding the first 20mLof the filtrate.

Chromatographic columns— Pack a pledget of glass wool at the base of each of the three tubes,and fill with adsorbent prepared as follows,using chromatographic siliceous earth as the base of the adsorbent,and tamp it firmly in place.Pack Column Iin two layers,the lower layer consisting of 3g of chromatographic siliceous earth mixed with 2mLof Citrate bufferand the upper layer of 3g of chromatographic siliceous earth mixed with 2.0mLof the Assay preparationand 0.5mLof Citrate buffer.Dry-rinse the beaker in which the components of the two layers have been mixed with 1g of chromatographic siliceous earth,and add it also to the top of Column I.Pack Column IIwith 3g of chromatographic siliceous earth mixed with 2mLof dibasic potassium phosphate solution (1in 5.75).Pack Column IIIwith 3g of chromatographic siliceous earth mixed with 2mLof sodium hydroxide solution (1in 50).Place a small pad of glass wool above each column packing.

Procedure— Proceed as directed in the Assayunder Paregoric.Calculate the percentage of anhydrous morphine in the Opium taken by the formula: in which Cis the concentration,in µg per mL,of anhydrous morphine in the Standard preparation;Wis the weight,in g,of Opium taken,and AUand ASare the corrected absorbances of the solutions from the Assay preparationand the Standard preparation,respectively.