A Rapid Methodology for the Characterization of Dialkyl Tertiary Amine-N-Oxide Metabolites Using Structurally Dependent Dissociation Pathways and Reconstructed Ion Current Chromatograms

摘要

A high-performance liquid chromatography-electrospraynionization-tandem mass spectrometry (HPLC-ESI-MS/MS)napproach to the characterization of dialkyl tertiary amine-nN-oxides is presented. The methodology is based uponnforming reconstructed ion current chromatograms (RICCs)nof m/z values of product ions known to form throughndiagnostic losses from dialkyl tertiary amine-N-oxides. Thendiagnostic losses of N,N-dimethylhydroxylamine and N,Ndiethylhydroxylaminenwere identified through the analysisnof a structurally diverse library of compounds by ESI-lowenergyncollision-induced dissociation (CID)-MS/MS usingnquadrupole ion trap-mass spectrometry (QIT-MS) and quadrupolentime-of-flight-mass spectrometry (QqTOF-MS). Thenlibrary consisted of dialkyl tertiary amine-containing commerciallynavailable pharmaceuticals, along with a numbernof model, synthetic N-oxides. The loss of the nitrogencontainingngroup was observed in 89% of the low-energy CIDnproduct ion spectra acquired using various collision energies.nFurther, the resultant product ions, formed throughnthe loss of the nitrogen-containing group, were shown to benunstable because of the observation of second-generationndissociation. These observations regarding gas-phase ionnchemistry could be useful to developers of in silico programsnfor fragmentation prediction by allowing the creation ofnimproved algorithms and models for predicting dissociation.nUsing the information derived from the library analysis, thencharacterization methodology was developed and demonstratednusing tetracaine. The approach is rapid, MS/MSnplatform independent, utilizes existing technology, and couldnbe automated. Further, it is definitive and overcomes thenlimitations of other tools for N-oxide identification by localizingnthe site of oxidation. Thus, it provides a useful additionnto the existing approaches for metabolite identification.