Spray reagent— Dissolve 2g of bismuth subnitrate in a solution consisting of 100mLof water and 25mLof glacial acetic acid (Solution A).Dissolve 40g of potassium iodide in 100mLof water (Solution B).Add 10mLof Solution Aand 10mLof Solution Bto a solution consisting of 100mLof water and 20mLof glacial acetic acid,and mix.

Procedure— Pipet an amount of Injection equivalent to about 1mg of glycopyrrolate into a 10-mLvolumetric flask,dilute with water to volume,and mix to obtain the test solution.Prepare a Standard solution of USP Glycopyrrolate RSin water containing about 0.1mg of glycopyrrolate per mL.Apply 30µLof the test solution and 30µLof the Standard solution to a suitable 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 chromatogram in a solvent system consisting of a mixture of butyl alcohol,glacial acetic acid,and water (3:1:1)until the solvent front has moved about three-fourths of the length of the plate.Remove the plate from the developing chamber,and allow to air-dry.Spray the plate with Spray reagent,and allow to air-dry:the RFvalue and color of the principal spot obtained from the test solution correspond to those obtained from the Standard solution.

Mobile phase— Dissolve 1.0g of anhydrous sodium sulfate and 200mg of sodium 1-pentanesulfonate in 615mLof water in a 1000-mLvolumetric flask.Add 3.0mLof 1Nsulfuric acid,235mLof acetonitrile,and 150mLof methanol,and mix.Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Dissolve an accurately weighed quantity of USP Glycopyrrolate RSin Mobile phase,and dilute quantitatively with Mobile phaseto obtain a solution having a known concentration of about 0.2mg per mL.

Resolution solution— Prepare a solution of benzaldehyde in Mobile phasecontaining about 0.5mg per mL.Transfer 2.0mLof this solution to a 25-mLvolumetric flask,dilute with Standard preparationto volume,and mix.

Assay preparation— Dilute a volume of Injection,quantitatively if necessary,with Mobile phaseto obtain a solution having a concentration of about 0.2mg of glycopyrrolate per mL.

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 222-nm detector and a 3.9-mm ×30-cm column containing packing L1.The flow rate is about 2mLper minute.Chromatograph the Resolution solution,and record the peak responses as directed under Procedure:the resolution,R,between the benzaldehyde and glycopyrrolate peaks is not less than 3.0.Chromatograph the Standard preparation,and record the peak responses as directed under Procedure:the relative standard deviation for replicate injections is not more than 1.0%.

Procedure— Separately inject equal volumes (about 35µL)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 glycopyrrolate (C19H28BrNO3)in each mLof the Injection taken by the formula: in which Cis the concentration,in mg per mL,of USP Glycopyrrolate RSin the Standard preparation;Lis the labeled quantity,in mg per mL,of glycopyrrolate in the Injection;Dis the concentration,in mg per mL,of glycopyrrolate in the Assay preparation,on the basis of the labeled quantity and the extent of dilution;and rUand rSare the glycopyrrolate peak responses obtained from the Assay preparationand the Standard preparation,respectively.