Characterization of the aqueous decomposition products of (+)1,2-bis(3,5-dioxopiperazinyl-1-yl)-propane (ICRF-187) by liquid chromatographic and mass spectral analysis.

High-performance liquid chromatography (HPLC) and fast atom bombardment mass spectrometry (FAB-MS) were employed to separate and identify the aqueous decomposition products of (+)1,2-bis(3,5-dioxopiperazinyl-1-yl)-propane (ICRF-187; 1), a drug active against several forms of human cancer and which also has recently been shown to display potent cardioprotective activity in patients treated with the antitumor antibiotic doxorubicin. Two reversed-phase HPLC columns were used to separate the hydrolysis products of 1, a Waters muBondapak phenyl column and an LKB Spherisorb ODS2 column. Incubation of 20 microM 1 in phosphate-buffered saline (PBS) at 37 degrees C for 21 h resulted in 47% decomposition, with three hydrolysis products detected (compound 2, Waters column retention time (RT) = 3.7 min, observed monoisotopic protonated molecular ion (MH+) m/z value of 305.1; compound 3, RT = 4.1 min, MH+ m/z value of 287.1; compound 4, RT = 4.8 min, MH+ m/z value of 287.1). The RT and MH+ m/z values for 1 were 17.1 min and 269.1, respectively. Based on the FAB-MS data, 2 corresponds to ICRF-198, the polar diacid diamide derivative of 1, while peaks 3 and 4 represent the monoacid monoamide derivatives of 1. Using B/E linked scan daughter FAB-MS analysis, 3 displayed a prominent fragment with a m/z value of 160, indicating that it corresponds to the monoacid monoamide derivative of 1, with the methyl group adjacent to the hydrolyzed ring. Compound 4, displaying a fragment with a m/z value of 142 in its B/E linked scan daughter ion spectrum, corresponds to the monoacid monoamide derivative of 1, with the methyl group adjacent to the closed ring.