The widely used solvent hexamethylphosphoramide (HMPA) and its biological (metabolic) and chemical (abiotic) phosphoramide-based oxidation products may cause adverse health effects through occupational exposure and intake of contaminated groundwater. However, no current methods exist for the separation and the detection of the many polar HMPA oxidation products. Thus, we developed a new RPLC/ESI-TOF-MS method and further investigated the chromatographic performances of two columns (i.e., XTerra Phenyl and XBridge Phenyl). In addition, the impact of (forced) acid hydrolysis for optimized chromatographic performance of the XTerra Phenyl column is investigated. The XTerra Phenyl column showed the best separation of the less polar major metabolic oxidation products pentamethylphosphoramide and hydroxymethyl-pentamethylphosphoramide, however, only after treating the column with formic acid (acid-treated). The XTerra column separated most of the investigated HMPA oxidation products (11 of 16 compounds) in a single chromatographic run. In contrast, the XBridge Phenyl column requires one method for the less polar and another method for the more polar oxidation products. However, this results in an overall better separation performance of the XBridge Phenyl column, especially for the less polar major abiotic oxidation products hydroxymethyl-pentamethylphosphoramide and formyl-pentamethylphosphoramide, as well as for 11 highly polar oxidation products (R(S)>1.5). The RPLC/ESI-TOF-MS method presented and validated in this study is the first analytical method that can be used to separate and detect HMPA (LOD 0.10 μM without preconcentration) and all of its oxidation products.
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