Unusual MS n fragmentation patterns of 2,4‐dinitrophenylhydrazine and its propanone derivative

During the structural elucidation of carbonyl‐2,4‐dinitrophenylhydrazone derivatives in the low‐ng range by multiple mass spectrometry combined with high performance liquid chromatography (HPLC/MS n ) some fragments were detected which were difficult to explain by ordinary fragmentation rules. Atmospheric pressure chemical ionization in the negative ion mode was applied. The ion formation was studied by labelling the parent compounds with D, 13 C and 15 N and recording the MS/MS, MS 3 and MS 4 spectra. This allowed an investigation of the detailed fragmentation of 2,4‐dinitrophenylhydrazine and its propanone derivative as well as the structures of the MS 2 and MS 3 product ions. A highly reactive product ion was found which undergoes unexpected reactions inside the ion trap of an HPLC/MS n system. In addition, reactions of MS n fragments with solvents or species generated in situ in the ion source and/or the ion trap were observed. It is demonstrated that unusual fragments in the MS/MS or MS n spectra of a compound may be formed by such reactions and that the eluent used for HPLC might have an influence on the observed fragmentation even if the mass spectrum itself is completely free from adducts.