100(CS/CA)(AU/AS),

100(CS/CA)(AU/AS),

NOTE—Use freshly prepared solutions.Measure the peak responses at the following wavelengths:prostaglandin A1at 224nm;prostaglandin B1at 280nm;and all other foreign prostaglandin impurities at 200nm.

Mobile phase— Proceed as directed in the Assay.

Standard solution— Dissolve accurately weighed quantities of USP Alprostadil RS,USP Prostaglandin A1RS,and USP Prostaglandin B1RSin a mixture of methanol and water (9:1),and dilute quantitatively,and stepwise if necessary,with a mixture of methanol and water (9:1)to obtain a solution having known concentrations of about 6µg per mL,15µg per mL,and 6µg per mL,respectively.

Test solution— Dissolve about 15mg of Alprostadil,accurately weighed,in 5mLof a mixture of methanol and water (9:1),and mix.

Chromatographic system— Proceed as directed in the Assay.Chromatograph the Standard solution,and record the peak responses as directed for Procedure:the resolution between prostaglandin A1and alprostadil is not less than 7.5,and the relative standard deviation from the peaks at their respective wavelength for replicate injections is not more than 4.0%.

Procedure— Separately inject equal volumes (about 20µL)of the Standard solutionand the Test solutioninto the chromatograph,record the chromatograms,and measure the peak responses at 200nm,224nm,and 280nm.Calculate the percentage of prostaglandin A1and prostaglandin B1in the portion of Alprostadil taken by the formula: in which CSis the concentration,in mg per mL,of USP Prostaglandin A1RSor USP Prostaglandin B1RSin the Standard solution;Wis the weight,in mg,of Alprostadil taken for the Test solution;riis the peak response for prostaglandin A1or prostaglandin B1obtained from the Test solution;and rSis the peak response of prostaglandin A1or prostaglandin B1obtained from the Standard solution:not more than 1.5%of prostaglandin A1is found;and not more than 0.1%of prostaglandin B1is found.Calculate the percentage of each impurity occurring at 200nm and eluting before prostaglandin A1in the portion of Alprostadil taken by the formula: in which CSis the concentration,in mg per mL,of USP Alprostadil RSin the Standard solution;riis the peak response for each impurity obtained from the Test solution;rSis the peak response for alprostadil obtained from the Standard solution;and the other terms are as defined herein:not more than 0.9%of any foreign prostaglandin impurity is found.Calculate the percentage of any impurity having a relative retention time of 0.6,relative to the prostaglandin A1peak detected at 224nm,in the portion of Alprostadil taken by the formula: in which CSis the concentration,in mg per mL,of USP Prostaglandin A1RSin the Standard solution;rIis the peak response for any impurity having a relative retention time of 0.6,relative to the prostaglandin A1peak,obtained from the Test solution;rSis the peak response of prostaglandin A1obtained from the Standard solution;and the other terms are as defined herein:not more than 0.9%of any impurity having a relative retention time of 0.6,relative to the prostaglandin A1peak,is found.

Mobile phase— Prepare a filtered and degassed mixture of methanol,acetonitrile,and 0.02Mmonobasic potassium phosphate (2:1:1),and adjust with phosphoric acid to a pHof 3.Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard solution— Dissolve an accurately weighed quantity of USP Alprostadil RSin a mixture of acetonitrile and water (1:1)to obtain a solution having a known concentration of about 10µg per mL.

Test solution— Dissolve about 25mg of Alprostadil,accurately weighed,in 5mLof a mixture of acetonitrile and water (1:1),using ultrasound if necessary.

Identification solution— Use the Standard solutionunder Test 1.

Chromatographic system (see Chromatography á621ñ)— The liquid chromatograph is equipped with a photodiode array detector or equivalent capable of detecting UVwavelengths between 200nm and 300nm and a 4.6-mm x 25-cm column that contains packing L1.The flow rate is about 1.2mLper minute.Chromatograph the Identification solutionas directed for Procedure:the relative retention times for prostaglandin A1and alprostadil are about 1.2and 1.0,respectively;the resolution between prostaglandin A1and alprostadil is not less than 4.0.Chromatograph the Standard solution,and record the peak responses as directed for Procedure:the relative standard deviation determined from the main peak for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 20µL)of the Standard solutionand the Test solutioninto the chromatograph,record the chromatograms,and measure the peak responses at 200nm,224nm,and 280nm.Calculate the percentage of each impurity occurring at 200nm and eluting after prostaglandin A1,excluding prostaglandin B1,in the portion of Alprostadil taken by the formula: in which CSis the concentration,in mg per mL,of USP Alprostadil RSin the Standard solution;Wis the weight,in mg,of Alprostadil taken for the Test solution;rIis the peak response for each impurity obtained from the Test solution;and rSis the peak response for alprostadil obtained from the Standard solution:the sum of the peaks having relative retention times of 2.0and 2.3is not more than 0.6%;not more than 0.9%of any other foreign prostaglandin impurity is found;and not more than 2.0%of total impurities is found,the results for Test 1and Test 2being added.

25C(RU/RS),