A: Infrared Absorption á197Kñ.

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.

Buffer solution— Dissolve 790mg of ammonium carbonate in 1000mLof water.Adjust,if necessary,with a solution of formic acid (1in 10)or ammonium hydroxide (1in 10)to a pHof 8.0±0.05.

Diluent— Prepare a mixture of methanol and water (65:35).

Mobile phase— Prepare a filtered and degassed mixture of methanol and Buffer solution(65:35).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Transfer about 25mg of USP Altretamine RS,accurately weighed,to a 50-mLvolumetric flask.Dissolve in 35mLof methanol,dilute with water to volume,and mix.Transfer 5.0mLof this solution to a 50-mLvolumetric flask,dilute with Diluentto volume,and mix.This solution contains about 0.05mg per mL.

Assay preparation— Transfer about 25mg of Altretamine,accurately weighed,to a 50-mLvolumetric flask.Dissolve in 35mLof methanol,dilute with water to volume,and mix.Transfer 5.0mLof this solution to a 50-mLvolumetric flask,dilute with Diluentto volume,and mix.

Chromatographic system— The liquid chromatograph is equipped with a 227-nm detector and a 4.6-mm ×30-cm column that contains packing L1.The flow rate is about 2mLper minute.Chromatograph the Standard preparation,and record the peak responses as directed for Procedure:the tailing factor is not greater than 1.5,and the relative standard deviation for replicate injections is not more than 2.0%.

Procedure— Separately inject equal volumes (about 10µL)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 C9H18N6in the portion of Altretamine taken by the formula: in which Cis the concentration,in mg per mL,of USP Altretamine RSin the Standard preparation;and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.