»Levodopa contains not less than 98.0percent and not more than 102.0percent of C9H11NO4,calculated on the dried basis.
Packaging and storage Preserve in tight,light-resistant containers,in a dry place,and prevent exposure to excessive heat.
A:Infrared Absorption á197Mñ.
B:Ultraviolet Absorption á197Uñ
Solution: 40µg per mL.
Medium: 0.1Nhydrochloric acid.
Absorptivities at 280nm,calculated on the dried basis,do not differ by more than 3.0%.
C: The retention time of the major peak in the chromatogram of the Assay preparationcorresponds to that in the chromatogram of the Standard preparation,as obtained in the Assay.
Specific rotation á781Sñ: between -160and -167.
Test solution Transfer about 500mg of Levodopa,accurately weighed,to a 25-mLvolumetric flask,add 10mLof 1Nhydrochloric acid to dissolve the solid,then add 5g of methenamine,swirl the contents to dissolve the methenamine,add 1Nhydrochloric acid to volume,and mix.Allow to stand in the dark at 25for 3hours,and measure the rotation.
Loss on drying á731ñ Dry it at 105for 4hours:it loses not more than 0.5%of its weight.
Residue on ignition á281ñ: not more than 0.1%.
Heavy metals,Method IIá231ñ: 0.002%.
Related compounds [NOTEProtect all solutions from light and maintain them at 10until they are injected into the chromatograph.]
Diluent,Mobile phase,System suitability solution,andChromatographic system Proceed as directed in the Assay.
Standard solution Prepare as directed for the Standard preparationin the Assay.
Test solution Use the Assay preparation.
Procedure Separately inject equal volumes (about 20µL)of the Standard solutionand the Test solutioninto the chromatograph,record the chromatograms,and measure the peak responses.Calculate the percentage of each impurity in the portion of Levodopa taken by the formula:
100F(ri/rs),in which Fis the correction factor and is equal to 2.5for peaks with a relative retention time of 0.9or 1.3,and is equal to 1for peaks with a relative retention time of 1.6or for any other peak;riis the peak response of each impurity obtained from the Test solution;and rsis the sum of the responses of all the peaks:not more than 0.1%of levodopa related compound Ais found;not more than 0.1%of L-tyrosine is found;not more than 0.1%of any unknown impurity is found;not more than 0.5%of 3-methoxytyrosine is found;and the sum of all impurities is not more than 1.1%.
Organic volatile impurities,Method IVá467ñ: meets the requirements.
Assay [NOTEProtect all solutions from light and maintain them at 10until they are injected into the chromatograph.]
Diluent Prepare a mixture of trifluoroacetic acid and water (1in 1000).
Mobile phase Prepare a filtered and degassed mixture of Diluentand tetrahydrofuran (97:3).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).
System suitability solution Dissolve suitable quantities of USP Levodopa RS,USP3-Methoxytyrosine RS,and USPL-Tyrosine RSin Diluentto obtain a solution containing about 10µg of each per mL.
Standard preparation Dissolve an accurately weighed quantity of USP Levodopa RSin Diluent,and dilute quantitatively,and stepwise if necessary,with Diluentto obtain a solution having a known concentration of about 0.4mg per mL.
Assay preparation Transfer about 40mg of Levodopa,accurately weighed,to a 100-mLvolumetric flask,dissolve in and dilute with Diluentto 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 double-endcapped packing L1.The flow rate is about 1.0mLper minute.Chromatograph the System suitability solution,and record the peak responses as directed for Procedure:the relative retention times are about 1.0for levodopa,1.3for L-tyrosine,and 1.6for 3-methoxytyrosine;the resolution,R,between levodopa and L-tyrosine is not less than 3.0;the tailing factor is not more than 2.0for levodopa;and the relative standard deviation determined from levodopa for replicate injections is not more than 2.0%.
Procedure Separately inject equal volumes (about 20µL)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 C9H11NO4in the portion of Levodopa taken by the formula:
100C(rU/rS),in which Cis the concentration,in mg per mL,of USP Levodopa RSin the Standard preparation;and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.
Auxiliary Information Staff Liaison:Salvador C.Salado,M.S.,Scientist and Latin American Liaison
Expert Committee:(PA3)Pharmaceutical Analysis 3
USP28NF23Page 1121Pharmacopeial Forum:Volume No.30(3)Page 888