Kanamycin Sulfate
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C18H36N4O11·H2SO4 582.58

D-Streptamine,O-3-amino-3-deoxy-a-D-glucopyranosyl-(1®6)-O-[6-amino-6-deoxy-a-D-glucopyranosyl(1®4)]-2-deoxy-,sulfate (1:1)(salt).
Kanamycin sulfate (1:1)(salt)
[133-92-6;25389-94-0].
»Kanamycin Sulfate has a potency equivalent to not less than 750µg of kanamycin (C18H36N4O11)per mg,calculated on the dried basis.
Packaging and storage— Preserve in tight containers.
Labeling— Where it is intended for use in preparing injectable dosage forms,the label states that it is sterile or must be subjected to further processing during the preparation of injectable dosage forms.
Identification—
A: Dissolve about 10mg in 1mLof water,add 1mLof a solution of ninhydrin in butyl alcohol (1in 500),and add 0.5mLof pyridine.Heat in a steam bath for 5minutes,and add 10mLof water:a deep purple color is produced.
B: It meets the requirements of the tests for Sulfate á191ñ.
C: The retention time of the peak for kanamycin in the chromatogram of the Assay preparationcorresponds to that in the chromatogram of the Standard preparation,as obtained in the Assay.
Crystallinity á695ñ: meets the requirements.
pHá791ñ: between 6.5and 8.5,in a solution (1in 100).
Loss on drying á731ñ— Dry about 100mg,accurately weighed,in a capillary-stoppered bottle in vacuum at a pressure not exceeding 5mm of mercury at 60for 3hours:it loses not more than 4.0%of its weight.
Residue on ignition á281ñ: not more than 1.0%,the charred residue being moistened with 2mLof nitric acid and 5drops of sulfuric acid.
Chromatographic purity— Dissolve a quantity of Kanamycin Sulfate in water to obtain a test solution having a concentration of 30mg per mL.Dissolve a suitable quantity of USP Kanamycin Sulfate RSin water to obtain a Standard solution having a known concentration of 30mg per mL.Dilute a portion of this solution quantitatively with water to obtain a Diluted standard solutionhaving a concentration of 0.90mg per mL.Apply separate 1-µLportions of the three solutions to the starting line of a suitable thin-layer chromatographic plate (see Chromatography á621ñ)coated with a 0.25-mm layer of chromatographic silica gel and heated at 110for 1hour and cooled immediately before use.Allow the spots to dry,and develop the chromatogram in a suitable chamber,previously equilibrated for 90minutes with a developing solvent of monobasic potassium phosphate solution (7.5in 100),until the solvent front has moved about three-fourths of the length of the plate.Remove the plate from the chamber,and air-dry.Spray the plate with a solution of ninhydrin in butyl alcohol (1in 100).Dry the plate at 110for 10minutes,and examine the chromatograms:the chromatograms show principal spots at about the same RFvalue,and no secondary spot,if present in the chromatogram from the test solution,is more intense than the principal spot obtained from the Diluted standard solution.
Other requirements— Where the label states that Kanamycin Sulfate is sterile,it meets the requirements for Sterilityand Bacterial endotoxinsunder Kanamycin Injection.Where the label states that Kanamycin Sulfatemust be subjected to further processing during the preparation of injectable dosage forms,it meets the requirements for Bacterial endotoxinsunder Kanamycin Injection.
Assay—
Mobile phase— Use a 0.115Nsodium hydroxide solution.Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).
Resolution solution— Prepare a solution in water containing about 0.02mg of USP Amikacin RSper mLand 0.008mg of USP Kanamycin Sulfate RSper mL.
Standard preparation— Quantitatively dissolve an accurately weighed quantity of USP Kanamycin Sulfate RSin water to obtain a solution having a known concentration of about 0.008mg per mL.
Assay preparation— Transfer about 40mg of Kanamycin Sulfate,accurately weighed,to a 250-mLvolumetric flask,dilute with water to volume,and mix.Transfer 5.0mLof this solution to a 100-mLvolumetric flask,dilute with water to volume,and mix.
Chromatographic system (seeChromatography á621ñ)— The liquid chromatograph is equipped with an electrochemical detector,a gold working electrode,a pHsilver–silver chloride reference electrode,a guard column that contains packing L47,and a 4-mm ×25-cm analytical column that contains packing L47.The electrochemical detector is used in the integrated amperometric mode with a range of 300nCand an output of 1Vfull-scale.The potential is programmed as follows.
Time (seconds) Potential (V) Integration
0.00 +0.04
0.30 +0.04 begins
0.50 +0.04 ends
0.51 +0.80
0.70 +0.80
0.71 –0.80
0.90 –0.80
The flow rate is about 0.5mLper minute.Chromatograph the Resolution solution,and record the peak responses as directed for Procedure:the relative retention times are about 1.0for kanamycin and 1.3for amikacin;and the resolution,R,between kanamycin and amikacin is not less than 3.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the tailing factor is not more than 2;and the relative standard deviation 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 areas for the major peaks.Calculate the quantity,in µg,of kanamycin (C18H36N4O11)in each mg of Kanamycin Sulfate taken by the formula:
5000(CP/W)(rU/rS),
in which Cis the concentration,in mg per mL,of USP Kanamycin Sulfate RSin the Standard preparation;Pis the designated content,in µg per mg,of kanamycin in USP Kanamycin Sulfate RS;Wis the weight,in mg,of Kanamycin Sulfate taken to prepare the Assay preparation;andrUandrSare the kanamycin peak areas obtained from the Assay preparationand the Standard preparation,respectively.
Auxiliary Information— Staff Liaison:William W.Wright,Ph.D.,Scientific Fellow
Expert Committee:(PA7)Pharmaceutical Analysis 7
USP28–NF23Page 1094
Pharmacopeial Forum:Volume No.27(6)Page 3312
Phone Number:1-301-816-8335