Identification of new aqueous chemical degradation products of isophosphoramide mustard

NMR (31P, 1H and 13C) spectroscopy was used to study the products of the degradation of isophosphoramide mustard (IPM) in buffered solutions at pH ranging from 1 to 13. At pH < or = 1, the only degradation compounds detected were phosphate ion (Pi) and chloroethylammonium chloride (CEA-HCl), resulting from the breakdown of the two P-N bonds (pathway Ia). At pH 9.3 and 13, only the products of 1,3-cyclization of the N-chloroethyl group (monoaziridinylIPM (monoAzIPM) and a very low level of bisaziridinylIPM (bisAzIPM)) were found after approximately 15 h of reaction (pathway II). At intermediate pH, the two pathways coexist. At pH 3.5 and 5.0, the P-N bond hydrolysis is the major pathway, but two final phosphorylated products were detected, Pi which represented 67% (pH 3.5) and 17% (pH 5.0) of all the IPM phosphorylated degradation products after approximately 15 h of reaction, and phosphorylethanolamine (PEA) which represented 16% (pH 3.5) and 46% (pH 5.0) of the same sum. PEA formation can be explained by the 1,5-cyclization of a transient compound giving a 1,3,2-oxazaphospholidine intermediate whose P-N bond is exclusively cleaved in acidic medium. The presence of monohydroxyIPM (monoOHIPM) (whose percentage increases with pH from 5% (pH 3.5) to approximately 28% (pH 5.0) of all the IPM phosphorylated degradation compounds), probably coming from the alkylation by water of an aziridine/aziridinium intermediate, demonstrates the occurrence of pathway II. At pH 7.0 and 7.4, the pathway II is initiated first, leading to 1,3-cyclization(s), followed by water alkylation of the aziridines formed. The sequences are IPM 1-->monoAzIPM 5-->bisAzIPM 9; IPM 1-->monoAzIPM 5-->monoOHIPM 6-->monoAzIPM with a N-hydroxyethylchain (presumed structure) 7-->dihydroxyIPM 8. Nevertheless, PEA and Pi are the final products observed, which implies the P-N bond hydrolysis of products 5-9 as demonstrated by the presence in the medium of CEA, aziridine and ethanolamine.