Thalidomide Click to View Image C13H10N2O4 258.23
1H-Isoindole-1,3(2H)-dione,2-(2,6-dioxo-3-piperidinyl)-,(±)-.
(±)-N-(2,6-Dioxo-3-piperidyl)phthalimide.
a-(N-Phthalimido)glutarimide [50-35-1]. »Thalidomide contains not less than 98.0percent and not more than 101.5percent of C13H10N2O4,calculated on the anhydrous basis. Packaging and storage— Preserve in tight containers,protected from light,at controlled room temperature. USP Reference standards á11ñ USP Thalidomide RS. Identification,Infrared Absorption á197Kñ. Microbial limits á61ñ: meets the requirements. Water,Method Ic á921ñ: not more than 0.5%. Solvent: anhydrous dimethyl sulfoxide. Heavy metals,Method IIá231ñ: 0.002%. Chromatographic purity— Solution A— Prepare a filtered and degassed mixture of water,acetonitrile,and phosphoric acid (95:5:0.1). Solution B— Prepare a filtered and degassed mixture of water,acetonitrile,and phosphoric acid (85:15:0.1). Diluent— Prepare a mixture of water,acetonitrile,and phosphoric acid (50:50:0.1). Mobile phase— Use variable mixtures of Solution Aand Solution Bas directed for Chromatographic system.Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ). Phthalic acid stock solution— Transfer about 100mg of phthalic acid to a 100-mLvolumetric flask,dissolve in a mixture of acetonitrile and water (80:5),and dilute with acetonitrile to volume.Mix,and dilute quantitatively,and stepwise if necessary,with acetonitrile to obtain a solution having a concentration of about 0.1mg per mL. Standard stock solution— Dissolve,with the aid of sonication,an accurately weighed quantity of USP Thalidomide RSin acetonitrile to obtain a solution having a known concentration of about 1mg per mL. Standard solution— Pipet 2.0mLof the Standard stock solutionand 2.0mLof the Phthalic acid stock solutioninto a 100-mLvolumetric flask,dilute with Diluentto volume,and mix.Pipet 10.0mLof this solution into a 100-mLvolumetric flask,add 10.0mLof phosphoric acid solution (1in 100),dilute with water to volume,and mix to obtain a solution having a known concentration of about 0.0002mg of phthalic acid per mL. Test solution— Transfer about 100mg of Thalidomide,accurately weighed,to a 50-mLvolumetric flask,and dissolve,with the aid of sonication,in 40mLof a mixture of water,acetonitrile,and phosphoric acid (50:50:0.1).Dilute with Diluentto volume,and mix.Pipet 10.0mLof this solution into a 100-mLvolumetric flask,add 10.0mLof phosphoric acid solution (1in 100),dilute with water to volume,and mix. Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 218-nm detector and a 3.9-mm ×15-cm column that contains 4-µm packing L1.The flow rate is about 2mLper minute.The chromatograph is programmed as follows. Time
(minutes) Solution A
(%) Solution B
(%) Elution 0 100 0 equilibration 0-15 100®50 0®50 linear gradient 15-20 50®100 50®0 linear gradient 20-30 100 0 isocratic Chromatograph the Standard solution,and record the peak responses as directed for Procedure:the relative retention times are about 0.35for phthalic acid and about 1.0for thalidomide;the tailing factor for the phthalic acid and thalidomide peaks is not more than 2.0;and the relative standard deviation determined from the phthalic acid peak for replicate injections is not more than 2.0%. Procedure— Separately inject equal volumes (about 200µL)of the Standard solutionand the Test solutioninto the chromatograph,record the chromatograms,and measure the areas for all the peaks.Calculate the percentage of each impurity in the portion of Thalidomide taken by the formula: 500(CP/W)(ri/rP), in which CPis the concentration,in mg per mL,of phthalic acid in the Standard solution;Wis the amount,in mg,of Thalidomide taken to prepare the Test solution;riis the peak response for each impurity obtained from the Test solution;and rPis the phthalic acid peak response obtained from the Standard solution:not more than 0.1%of any individual impurity is found;and not more than 0.3%of total impurities is found. Ordinary impurities á466ñ Test solution— Dissolve an accurately weighed quantity of Thalidomide in acetonitrile to obtain a solution having a concentration of about 2mg per mL. Standard solution— Dissolve an accurately weighed quantity of glutamine in a mixture of acetonitrile and water (1:1)to obtain a solution having a known concentration of about 0.1mg per mL. Eluant: a mixture of methylene chloride,methanol,and acetic acid (75:25:0.05). Application volume: 2µL(Standard solution)and 100µL(Test solution). Visualization: 4. Limit: 0.1%. Organic volatile impurities,Method Iá467ñ: meets the requirements. Assay— Mobile phase— Prepare a filtered and degassed mixture of water,acetonitrile,and phosphoric acid (85:15:0.1).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ). Internal standard preparation— Transfer about 150mg of phenacetin,accurately weighed,to a 100-mLvolumetric flask,dissolve in about 80mLof acetonitrile,dilute with acetonitrile to volume,and mix. Standard preparation— Dissolve,with the aid of sonication,an accurately weighed quantity of USP Thalidomide RSin acetonitrile to obtain a solution having a known concentration of about 1mg per mL.Transfer 10.0mLof this solution and 5.0mLof Internal standard preparationto a 100-mLvolumetric flask,add 10.0mLof phosphoric acid solution (1in 100),dilute with water to volume,and mix. Assay preparation— Transfer about 100mg of Thalidomide,accurately weighed,to a 100-mLvolumetric flask,and dissolve,with the aid of sonication,in 80mLof acetonitrile.Dilute with acetonitrile to volume,and mix.Pipet 10.0mLof this solution and 5.0mLof Internal standard preparationinto a 100-mLvolumetric flask,add 10.0mLof phosphoric acid solution (1in 100),dilute with water to volume,and mix. Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 237-nm detector and a 3.9-mm ×15-cm column that contains 4-µm packing L1.The flow rate is about 1.0mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the resolution,R,between thalidomide and phenacetin is not less than 3.0;the column efficiency determined from the thalidomide and phenacetin peaks is not less than 7000and 9000theoretical plates,respectively;the tailing factor is not more than 2.0;and the relative standard deviation for replicate injections is not more than 1.0%. Procedure— Separately inject equal volumes (about 20µL)of the Standard preparationand the Assay preparationinto the chromatograph,record the chromatograms,and measure the areas for the major peaks.Calculate the quantity,in mg,of C13H10N2O4in the portion of Thalidomide taken by the formula: 1000C(RU/RS), in which Cis the concentration,in mg per mL,of USP Thalidomide RSin the Standard preparation;and RUand RSare the peak area ratios obtained from the Assay preparationand the Standard preparation,respectively. Auxiliary Information— Staff Liaison:Lawrence Evans,III,Ph.D.,Scientist Expert Committee:(PA6)Pharmaceutical Analysis 6 USP28–NF23Page 1893 Pharmacopeial Forum:Volume No.27(1)Page 1818 Phone Number:1-301-816-8389