Gentamicin and Prednisolone Acetate Ophthalmic Ointment
»Gentamicin and Prednisolone Acetate Ophthalmic Ointment contains the equivalent of not less than 90.0percent and not more than 120.0percent of the labeled amount of gentamicin,and not less than 90.0percent and not more than 110.0percent of the labeled amount of prednisolone acetate (C23H30O6).
Packaging and storage— Preserve in collapsible ophthalmic ointment tubes,and avoid exposure to excessive heat.
Identification—
A: Shake a quantity of Ophthalmic Ointment,equivalent to about 5mg of gentamicin,with a mixture of 200mLof chloroform and 5mLof water.Allow to separate,and filter the aqueous layer:the filtrate so obtained meets the requirements of the Identificationtest under Gentamicin Sulfate Injection.
B: The chromatogram of the Assay preparationobtained as directed in the Assay for prednisolone acetateexhibits a major peak for prednisolone acetate,the retention time of which corresponds to that obtained in the chromatogram of the Standard preparationobtained as directed in the Assay for prednisolone acetate.
Sterility á71ñ: meets the requirements.
Minimum fill á755ñ: meets the requirements.
Water,Method Iá921ñ: not more than 2.0%,20mLof a mixture of toluene and methanol (7:3)being used in place of methanol in the titration vessel.
Metal particles— It meets the requirements of the test for Metal Particles in Ophthalmic Ointments á751ñ.
Assay for gentamicin— Proceed with Ophthalmic Ointment as directed for gentamicin under Antibiotics—Microbial Assays á81ñ,using an accurately weighed quantity of Ophthalmic Ointment,equivalent to about 1mg of gentamicin,shaken with about 50mLof ether in a separator,and extracted with four 20-mLportions of Buffer No.3.Combine the aqueous extracts,and dilute quantitatively and stepwise with Buffer No.3to obtain a Test Dilutionhaving a concentration of gentamicin assumed to be equal to the median dose level of the Standard.
Assay for prednisolone acetate—
Mobile phase— Prepare a suitable mixture of water and acetonitrile (60:40),and pass through a suitable filter having a porosity of 1µm or less.Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).
Internal standard solution— Prepare a solution in methanol containing about 2.7mg of fluorometholone acetate per mL.
Standard preparation— Transfer about 38mg of USP Prednisolone Acetate RS,accurately weighed,to a 100-mLvolumetric flask,dissolve in methanol,dilute with methanol to volume,and mix.Transfer 8.0mLof this solution to a 50-mLvolumetric flask,add 25mLof n-hexane,and shake.Add 2.0mLof Internal standard solution,dilute with methanol to volume,and shake vigorously for 30seconds.Allow the layers to separate,remove the upper n-hexane layer by aspiration,and discard the aspirate.Dilute the solution in the volumetric flask with methanol to volume,and mix.Centrifuge a portion of this solution,and use the clear liquid as the Standard preparation.This solution contains about 0.06mg of USP Prednisolone Acetate RSper mL.
Assay preparation— Transfer an accurately weighed portion of Ophthalmic Ointment,equivalent to about 3mg of prednisolone acetate,to a 50-mLvolumetric flask,add 25mLof n-hexane,and shake.Add 2.0mLof Internal standard solution,dilute with methanol to volume,and shake vigorously for 30seconds.Allow the layers to separate,remove the upper n-hexane layer by aspiration,and discard the aspirate.Dilute the solution in the volumetric flask with methanol to volume,and mix.Centrifuge a portion of this solution,and use the clear liquid as the Assay preparation.
Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 254-nm detector and a 3.9-mm ×20-cm column that contains packing L1.The flow rate is about 2mLper minute.Chromatograph the Standard preparation,and measure the peak responses as directed for Procedure:the tailing factor is not more than 1.5;the resolution,R,between the prednisolone acetate peak and the fluorometholone acetate peak is not less than 2.0;the column efficiency is not less than 2500theoretical plates;and the relative standard deviation for replicate injections is not more than 2.0%.
Procedure— Calculate the quantity,in mg,of prednisolone acetate (C23H30O6)in the portion of Ophthalmic Ointment taken by the formula:
50C(RU/RS),
in which Cis the concentration,in mg per mL,of USP Prednisolone Acetate RSin the Standard preparation,and RUand RSare the ratios of the response of the prednisolone acetate peak area to that of the fluorometholone acetate peak 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 900
Phone Number:1-301-816-8335