Mobile phase— Prepare as directed in the Assay.

Standard solution— Dissolve an accurately weighed quantity of USP Isradipine RSin Mobile phase,with the aid of sonication if necessary,and dilute quantitatively and stepwise with Mobile phaseto obtain a solution having a known concentration of about 6µg per mL.[NOTE—If necessary,use 1mLof methanol per 20mLof Mobile phase to dissolve the Reference Standard prior to diluting with Mobile phase.]

Resolution solution— Use the Standard preparationprepared as directed in the Assay.

Test solution— Transfer about 50mg of Isradipine,accurately weighed,to a 25-mLvolumetric flask,and add 5.0mLof methanol to dissolve,using sonication if necessary.Dilute with Mobile phaseto volume,and mix.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 230-nm detector and a 4.6-mm ×10-cm column that contains packing L1.The flow rate is about 1.7mLper minute.Chromatograph replicate injections of the Resolution solution,and record the peak responses as directed under Procedure:the resolution,R,between isradipine and isradipine related compound Ais not less than 1.5,and the relative standard deviation for replicate injections is not more than 1.5%.

Procedure— Separately inject equal volumes (about 25µL)of the Standard solutionand the Test solutioninto the chromatograph,and allow the Test solutionto elute for not less than three times the retention time of isradipine.Record the chromatograms,and measure the responses for all of the peaks:the sum of the peak responses,other than that of isradipine,in the chromatogram of the Test solutionis not more than four times the isradipine response obtained from the Standard solution(1.2%),the response of the largest peak,other than that of isradipine,in the chromatogram of the Test solutionis not more than 1.6times greater than the isradipine response obtained from the Standard solution(0.5%),and no other peak response,other than that of isradipine,is greater than the isradipine response obtained from the Standard solution(0.3%).

Mobile phase— Prepare a filtered and degassed mixture of water,methanol,and tetrahydrofuran (50:40:10).Make adjustments,if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve,with the aid of sonication,if necessary,accurately weighed quantities of USP Isradipine RSand USP Isradipine Related Compound A RSin Mobile phase,and dilute quantitatively,and stepwise,if necessary,with Mobile phaseto obtain a solution having known concentrations of 0.2mg and 10µg of USP Isradipine RSand USP Isradipine Related Compound A RS,respectively,per mL.[NOTE—If necessary,1mLof methanol per 20mLof Mobile phase may be added to dissolve the Reference Standards.]

Assay preparation— Transfer about 20mg of Isradipine,accurately weighed,to a 100-mLvolumetric flask.Add sufficient methanol to dissolve,using sonication,if necessary.Dilute with Mobile phaseto volume,and mix.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 326-nm detector and a 4.6-mm ×10-cm column that contains packing L1.The flow rate is about 1.7mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed under Procedure:the resolution,R,between isradipine and isradipine related compound Ais not less than 1.5,and the relative standard deviation for replicate injections is not more than 1.5%.

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 C19H21N3O5in the portion of Isradipine taken by the formula: in which Cis the concentration,in mg per mL,of USP Isradipine RSin the Standard preparation,and rUand rSare the isradipine peak responses obtained from the Assay preparationand the Standard preparation,respectively.