A: Pass about 25mLof the well-mixed Ophthalmic Suspension through a fine,sintered-glass filter,saving the filtrate.Wash the crystals in the funnel with a small amount of water.Dry the crystals at 105for 3hours:the IRabsorption spectrum of a potassium bromide dispersion of the crystals exhibits maxima only at the same wavelengths as that of a similar preparation of USP Prednisolone Acetate RS.

B: To the filtrate saved from Identificationtest A,add 6Nacetic acid dropwise until the pHis between 4and 5.Allow crystals of sulfacetamide to develop.Filter the crystals,wash with a small amount of water,and dry at 105for 2hours:the IRabsorption spectrum of a potassium bromide dispersion of the crystals so obtained exhibits maxima only at the same wavelengths as a preparation of USP Sulfacetamide Sodium RS,similarly treated.

Mobile phase— Prepare a filtered and degassed mixture of water,methanol,and glacial acetic acid (890:100:10).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve an accurately weighed quantity of USP Sulfacetamide Sodium RSin a mixture of water and methanol (4:1),and dilute quantitatively,and stepwise if necessary,with the same solvent mixture to obtain a solution having a known concentration of about 30µg per mL.

Assay preparation— Transfer an accurately measured volume of Ophthalmic Suspension,freshly mixed and free from air bubbles,equivalent to about 100mg of sulfacetamide sodium,to a 100-mLvolumetric flask,dilute with a mixture of water and methanol (4:1)to volume,and mix.Dilute 3.0mLof this solution with the same solvent mixture to 100.0mL,and mix.

System suitability preparation— Dissolve about 3mg of sulfanilamide in 100mLof the Standard preparation,and mix.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm ×25-cm column that contains packing L1.The flow rate is about 1.5mLper minute.Chromatograph the Standard preparationand the System suitability preparation,and record the peak responses as directed for Procedure:the column efficiency determined for the analyte peak is not less than 1500theoretical plates;the resolution,R,between the sulfacetamide and sulfanilamide peaks is not less than 3;and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 90µL)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 sulfacetamide sodium (C8H9N2NaO3S.H2O)in each mLof the Ophthalmic Suspension taken by the formula: in which 254.24and 236.23are the molecular weights of sulfacetamide sodium monohydrate and anhydrous sulfacetamide sodium,respectively;Cis the concentration,in µg per mL,calculated on the anhydrous basis,of USP Sulfacetamide Sodium RSin the Standard preparation;and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.

Mobile phase— Prepare a filtered and degassed mixture of water and acetonitrile (60:40).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve an accurately weighed quantity of USP Prednisolone Acetate RSin methanol to obtain a solution containing about 2mg per mL.Transfer 2.0mLof this solution to a 100-mLvolumetric flask,and dilute with a solvent mixture prepared by dissolving 2.72g of monobasic potassium phosphate in 300mLof water and 700mLof methanol.The Standard preparationhas a known concentration of about 0.04mg per mL.

Assay preparation— Using a “To contain”pipet,transfer an accurately measured volume of Ophthalmic Suspension,freshly mixed and free from air bubbles,equivalent to about 10mg of prednisolone acetate,to a 250-mLvolumetric flask.Rinse the pipet with the solvent mixture described under Standard preparation,collecting the rinsings in the flask,dilute with the same solvent mixture to volume,and mix.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 254-nm detector and a 4.0-mm ×30-cm column that contains packing L1.The flow rate is about 1.5mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the column efficiency determined from the analyte peak is not less than 3000theoretical plates;and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 30µL)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 prednisolone acetate (C23H30O6)in each mLof the Ophthalmic Suspension taken by the formula: in which Cis the concentration,in mg per mL,of USP Prednisolone Acetate RSin the Standard preparation;Vis the volume,in mL,of Ophthalmic Suspension taken;and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.