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.

Mobile phase— Prepare as directed in the Assay.

Blank solution— Transfer 12.5mLof water to a 25-mLvolumetric flask,dilute with methanol to volume,and mix.

Standard thiol solution— Transfer about 7mg of USP Amifostine Thiol RS,accurately weighed,to a 100-mLvolumetric flask.Dissolve in and dilute with Mobile phaseto volume,and mix.[NOTE—Inject immediately after preparation,or refrigerate until use.]

Test solution— Transfer about 150mg of Amifostine,accurately weighed,to a 10-mLvolumetric flask.Dissolve in 5.0mLof water,dilute with methanol to volume,and mix.[NOTE—Inject immediately after preparation,or refrigerate until use.]

Chromatographic system (see Chromatography á621ñ)—Prepare as directed in the Assay.Chromatograph the Standard thiol solution,and record the peak responses as directed for Procedure:the relative standard deviation for replicate injections is not more than 4.0%.

Procedure— Separately inject equal volumes (about 10µL)of the Standard thiol solution,the Test solution,and the Blank solutioninto the chromatograph,record the chromatograms,and measure the responses of all the peaks,excluding the peaks corresponding to those obtained from the Blank solution.Calculate the percentage of amifostine thiol in the portion of Amifostine taken by the formula: in which 134.24and 207.17are the molecular weights of amifostine thiol and amifostine thiol dihydrochloride,respectively;Cis the concentration,in mg per mL,of amifostine thiol dihydrochloride in the Standard thiol solution;Wis the weight,in mg,of Amifostine taken to prepare the Test solution;rUand rSare the amifostine thiol peak responses obtained from the Test solutionand the Standard thiol solution,respectively.Calculate the percentage of each of the other impurities in the portion of Amifostine taken by the formula: in which riand rAare the peak responses for each impurity and amifostine,respectively,obtained from the Test solution:not more than 0.1%of any individual impurity,excluding amifostine thiol,is found;and not more than 0.3%of total impurities,including amifostine thiol,is found.

Mobile phase— Dissolve 0.54g of sodium 1-octanesulfonate in 500mLof water.Adjust with phosphoric acid to a pHof 2.5.Prepare a filtered and degassed mixture of this solution and methanol (1:1).Make adjustments if necessary (see System Suitabilityunder Chromatography á621ñ).

Standard preparation— Transfer about 30mg of USP Amifostine RS,accurately weighed,to a 10-mLvolumetric flask.Dissolve in 5mLof water,dilute with methanol to volume,and mix.[NOTE—Inject immediately after preparation,or refrigerate until use.]

Assay preparation— Transfer about 30mg of Amifostine,accurately weighed,to a 10-mLvolumetric flask.Dissolve in 5mLof water,dilute with methanol to volume,and mix.[NOTE—Inject immediately after preparation,or refrigerate until use.]

Chromatographic system (see Chromatography á621ñ)—The liquid chromatograph is equipped with a 220-nm detector and a 4.6-mm ×25-cm column that contains 5-µm packing L1.The flow rate is about 1mLper minute.Chromatograph the Standard preparationand the Assay preparation,and record the peak responses as directed for Procedure:the column efficiency is not less than 1000theoretical plates;the tailing factor is not more than 2;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 C5H15N2O3PSin the portion of Amifostine taken by the formula: in which Cis the concentration,in mg per mL,of USP Amifostine RSin the Standard preparation;and rUand rSare the peak responses obtained from the Assay preparationand the Standard preparation,respectively.