Mobile phase,System suitability solution,Standard preparation,System sensitivity solution,and Chromatographic system— Proceed as directed in the Assay.

Test solution— Use the stock solution reserved from the Assay preparation.

Procedure— Inject a volume (about 10µL)of the Test solutioninto the chromatograph,record the chromatogram,and measure the peak responses.Calculate the percentage of each impurity in the volume of Injection taken by the formula: in which riis the peak response for each impurity,and rsis the sum of the responses of all of the peaks:not more than 1.0%of any individual impurity is found,and not more than 1.5%of total impurities is found.

Mobile phase— Dissolve 2.0g of monobasic potassium phosphate in 800mLof water.Add 5mLof 1.0Mtetrabutylammonium dihydrogen phosphate solution,dilute with water to 980mL,and mix.Add 20mLof acetonitrile,mix,filter,and degas.Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

System suitability solution— Dissolve accurately weighed quantities of USP Adenosine RSand inosine in warm water (50to 55),and dilute quantitatively,and stepwise if necessary,with water to obtain a solution having known concentrations of about 0.03mg each of adenosine and inosine per mL.

Standard preparation— Dissolve an accurately weighed quantity of USP Adenosine RSin warm water (50to 55),and dilute quantitatively,and stepwise if necessary,with water to obtain a solution having a known concentration of about 0.03mg per mL.If sodium chloride is present in the Injection,add 0.01mLof sodium chloride solution (0.9in 100)per mLof the anticipated final volume of the Standard preparationbefore the addition of the warm water.

System sensitivity solution— Pipet 3.0mLof the Standard preparationinto a 200-mLvolumetric flask,dilute with water to volume,and mix.

Assay preparation— Transfer an accurately measured volume of Injection,equivalent to about 30mg of adenosine,to a 100-mLvolumetric flask,dilute with water to volume,and mix.Reserve a portion of this stock solution for use in the test for Chromatographic purity.Pipet 5.0mLof the stock solution into a 50-mLvolumetric flask,dilute with water to volume,and mix.

Chromatographic system (see Chromatography á621ñ)— The liquid chromatograph is equipped with a 254-nm detector and a 3.9-mm ×30-cm column that contains packing L1.The flow rate is about 2.5mLper minute.Chromatograph the System suitability solution,and record the peak responses as directed for Procedure:the tailing factor for the adenosine peak is not more than 2.0;and the resolution,R,between adenosine and inosine is not less than 6.0.Similarly chromatograph the Standard preparation:the relative standard deviation for replicate injections is not more than 1.5%.Chromatograph the System sensitivity solution,and adjust the run time to 2½times the retention time of adenosine.

Procedure— Separately inject equal volumes (about 10µL)of the Assay preparationand the Standard preparationinto the chromatograph,record the chromatograms,and measure the areas of the adenosine peak responses.Calculate the quantity,in mg,of adenosine (C10H13N5O4)in each mLof the Injection taken by the formula: in which Cis the concentration,in mg per mL,of USP Adenosine RSin the Standard preparation;Dis the concentration,in mg per mL,of adenosine in the Assay preparation,based on the labeled quantity per mLand the extent of dilution;and rUand rSare the peak responses for adenosine obtained from the Assay preparationand the Standard preparation,respectively.