Gas chromatography–electron ionization mass spectrometry and liquid chromatography–electrospray tandem mass spectrometry for determination of impurities in the anti-cancer drug isophosphoramide mustard

Abstract

Isophosphoramide mustard (IPM) is known to have substantial anti-cancer activities in various animal models. Liquid chromatography–electrospray mass spectrometry (LC–ES–MS) and LC–ES–MS/MS methodologies have been developed and applied to the analysis of synthesized preparations of IPM. Our studies reveal that the principal impurity in IPM is N-(2-chloroethyl)- N′-ethylphosphorodiamidic acid (MC-IPM) formed by dehydrochlorination of IPM with subsequent hydrogenation during synthesis. This impurity is present at levels in the range of 2–5% depending upon synthesis conditions. In addition, a second IPM derivative has been characterized by LC–ES–MS/MS and has been shown to be the product of a reaction of IPM with the dilute perchloric acid mobile phase used for liquid chromatography separations. The LC–ES–MS/MS method has been successfully employed to detect IPM spiked into a blood plasma sample. This work establishes that LC–ES–MS/MS is a viable tool for the detailed characterization of IPM and related products.