Neomycin and Polymyxin B Sulfates and Pramoxine Hydrochloride Cream
»Neomycin and Polymyxin B Sulfates and Pramoxine Hydrochloride Cream contains the equivalent of not less than 90.0percent and not more than 130.0percent of the labeled amounts of neomycin and polymyxin B,and not less than 90.0percent and not more than 110.0percent of the labeled amount of pramoxine hydrochloride (C17H27NO3·HCl).
Packaging and storage— Preserve in well-closed containers,preferably at controlled room temperature.
Identification—
A:Thin-Layer Chromatographic Identification Test á201ñ—
Test solution— Disperse a quantity of Cream,equivalent to about 25mg of neomycin,with 20mLof chloroform in a 60-mLseparator.Add 0.2mLof 2.5Nhydrochloric acid,and shake.Allow the layers to separate for about 30minutes.Discard the lower chloroform layer,and centrifuge the upper aqueous layer.Use a portion of the centrifuged aqueous layer.
Standard solution— Dissolve suitable quantities of USP Neomycin Sulfate RSand USP Polymyxin B Sulfate RSin 0.1Nhydrochloric acid to obtain a solution containing the equivalent of about 3.5mg of neomycin and 10,000USP Polymyxin B Units per mL.
Developing solvent system— Dissolve 0.1g of benzalkonium chloride in a mixture of isopropyl alcohol,water,and ammonium hydroxide (60:40:10).
Procedure— Proceed as directed in the chapter.Place the plate in a chromatographic chamber saturated with Developing solvent system,and develop the chromatogram.Dry the plate at 105for about 10minutes,spray with a solution of ninhydrin in butyl alcohol (1in 200),and heat the plate at 105for about 15minutes.The RFvalues of the two principal spots in the chromatogram obtained from the Test solutioncorrespond to those of the two principal spots in the chromatogram obtained from the Standard solution.
B: 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 for pramoxine hydrochloride.
pHá791ñ— Transfer 1g of Cream to a small beaker,add 10mLof carbon dioxide-free water,and mix:the pHis between 3.3and 6.0.
Assay for neomycin— Proceed as directed for neomycin under Antibiotics—Microbial Assays á81ñ,using an accurately weighed portion of Cream,equivalent to about 3.5mg of neomycin,blended for 3to 5minutes in a high-speed blender with 249mLof Buffer No.3and 1mLof polysorbate 80.Quantitatively dilute an accurately measured volume of this solution with Buffer No.3to obtain a Test Dilutionhaving a concentration of neomycin assumed to be equal to the median level of the Standard (1µg of neomycin per mL).
Assay for polymyxin— Proceed as directed for polymyxin Bunder Antibiotics—Microbial Assays á81ñ,using an accurately weighed portion of Cream,equivalent to about 10,000USP Polymyxin B Units,blended for 3to 5minutes in a high-speed blender with 199mLof Buffer No.6and 1mLof polysorbate 80.Quantitatively dilute an accurately measured volume of this solution withBuffer No.6to obtain a Test Dilutionhaving a concentration of polymyxin Bassumed to be equal to the median dose level of the Standard (10USP Polymyxin B Units per mL).
Assay for pramoxine hydrochloride—
Mobile phase— Dissolve 3.5g of dibasic potassium phosphate in 1000mLof water.Prepare a mixture of this solution,acetonitrile,and triethylamine (700:300:2),and adjust with phosphoric acid to a pHof 4.0±0.1.Filter and degas.Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).
Standard preparation— Prepare a solution of USP Pramoxine Hydrochloride RSin methanol to obtain a solution having a known concentration of about 0.2mg per mL.
Assay preparation— Transfer an accurately weighed portion of Cream,equivalent to about 10mg of pramoxine hydrochloride,to a 50-mLvolumetric flask,add about 5mLof chloroform,and sonicate at about 40to disperse the Cream.Allow to cool to room temperature,dilute with methanol to volume,and mix.Pass a portion of this solution through a glass fiber filter and a PTFEfilter having a 0.45-µm porosity,discarding the first few mLof the filtrate.
Chromatographic system(see Chromatography á621ñ)— The liquid chromatograph is equipped with a 280-nm detector,a guard column that contains packing L7,and a 4.6-mm ×25-cm analytical column that contains packing L7.The column is maintained at a constant temperature of about 40.The flow rate is about 2mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure: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 mg,of pramoxine hydrochloride (C17H27NO3·HCl)in each g of Cream taken by the formula:
50(C/W)(rU/rS),
in which Cis the concentration,in mg per mL,of USP Pramoxine Hydrochloride RSin the Standard preparation;Wis the weight,in g,of Cream taken to prepare the Assay preparation;and rUand rSare the peak areas for pramoxine 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 1360
Pharmacopeial Forum:Volume No.28(3)Page 774
Phone Number:1-301-816-8335