A: The chromatogram of the Assay preparationobtained as directed in the Assayexhibits a major peak for dihydrocodeine,the retention time of which corresponds to that in the chromatogram of the Standard preparationsimilarly determined.

B: It responds to the tests for Tartrate á191ñ.

C: To a solution of 20mg in 5mLof sulfuric acid in a test tube add 1drop of ferric chloride TS,and heat in a boiling water bath for 2minutes:although the solution may darken,no blue color is produced (distinction from codeine and morphine).

Test solution: 10mg per mL,in water.

Test solution: water.

Standard solution: water.

Eluant: a mixture of methylene chloride,methanol,and ammonium hydroxide (90:10:1).

Visualization: 17,and view under short-wavelength UVlight.

Mobile phase— Prepare a mixture of acetonitrile,water,and diethylamine (800:4:1).Prepare a suitable mixture of this solution and methanol (55:45).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Resolution solution— Prepare a solution in methanol containing about 0.4mg of USP Dihydrocodeine Bitartrate RSand 0.6mg of hydrocodone bitartrate per mL.Prepare a mixture of this solution and water (1:1).

Standard preparation— Transfer about 20mg of USP Dihydrocodeine Bitartrate RS,accurately weighed,to a 50-mLvolumetric flask,add 25mLof methanol,swirl to dissolve,dilute with water to volume,and mix.

Assay preparation— Transfer about 20mg of Dihydrocodeine Bitartrate,accurately weighed,to a 50-mLvolumetric flask,add 25mLof methanol,swirl to dissolve,dilute with water to volume,and mix.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 280-nm detector and a 4.6-mm ×25-cm column that contains 5-µm packing L3.The flow rate is about 1.5mLper minute.Chromatograph the Resolution solution,and record the responses as directed under Procedure:the relative retention times are about 0.8for hydrocodone and 1.0for dihydrocodeine,and the resolution,R,between the hydrocodone and dihydrocodeine peaks is not less than 1.8.Chromatograph the Standard preparation,and record the responses as directed under Procedure:the column efficiency determined from the dihydrocodeine peak is not less than 900theoretical plates,the tailing factor for the dihydrocodeine peak is not more than 1.7,and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 25µL)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 C18H23NO3·C4H6O6in the portion of Dihydrocodeine Bitartrate taken by the formula: in which Cis the concentration,in mg per mL,of USP Dihydrocodeine Bitartrate RSin the Standard preparation,and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.