A: The UVabsorption spectrum of the Assay preparationobtained in the Assayexhibits maxima and minima at the same wavelengths as that of the Standard preparation,concomitantly measured.

B: Transfer a quantity of Injection,equivalent to about 50mg of flunixin,to a 50-mLcentrifuge tube.Add 10mLof Acetate buffer,prepared as directed in the Assay,and extract with 25mLof ethyl acetate.Use the upper phase as the test solution.Separately apply 10µLof the test solution and 10µLof a Standard solution of USP Flunixin Meglumine RSin methanol containing 3mg per mLto a thin-layer chromatographic plate (see Chromatography á621ñ)coated with a 0.25-mm layer of chromatographic silica gel mixture.Allow the spots to dry,and develop the chromatograms in a solvent system consisting of a mixture of toluene,ethyl acetate,glacial acetic acid,and water (75:25:10:1)until the solvent front has moved about three-fourths of the length of the plate.Remove the plate from the developing chamber,mark the solvent front,and allow the spots to air-dry.Examine the plate under short-wavelength UVlight:the RFvalue of the principal spot obtained from the test solution corresponds to that obtained from the Standard solution.

Acetate buffer— Dissolve 4.1g of anhydrous sodium acetate in 500mLof water.Add 2.9mLof glacial acetic acid,dilute with water to 1000mL,and mix.

Standard preparation— Dissolve an accurately weighed quantity of USP Flunixin Meglumine RSin 0.1Nsodium hydroxide,and dilute quantitatively,and stepwise if necessary,with 0.1Nsodium hydroxide to obtain a solution having a known concentration of about 1.65mg per mL.Transfer 4.0mLof this solution to a 100-mLvolumetric flask,dilute with 0.1Nhydrochloric acid to volume,and mix.

Assay preparation— Transfer an accurately measured volume of Injection,equivalent to about 100mg of flunixin,to a 50-mLcentrifuge tube.Add 20mLof Acetate buffer,and extract with three 25-mLportions of ethyl acetate.Combine the extracts,filter,and evaporate to dryness on a steam bath under a stream of nitrogen.Dissolve the residue in 0.1Nsodium hydroxide,and transfer to a 100-mLvolumetric flask with the aid of 0.1Nsodium hydroxide.Dilute with 0.1Nsodium hydroxide to volume,and mix.Transfer 4.0mLof this solution to a second 100-mLvolumetric flask,dilute with 0.1Nhydrochloric acid to volume,and mix.

Procedure— Concomitantly determine the absorbances of the Assay preparationand the Standard preparationat the wavelength of maximum absorbance at about 327nm.Calculate the quantity,in mg,of flunixin (C14H11F3N2O2)in each mLof Injection taken by the formula: in which 296.25and 491.46are the molecular weights of flunixin and flunixin meglumine,respectively;Cis the concentration,in mg per mL,of USP Flunixin Meglumine RSin the Standard preparation;Vis the volume,in mL,of Injection taken to prepare the Assay preparation;and AUand ASare the absorbances of the Assay preparationand the Standard preparation,respectively.