A highly sensitive isotope-coded derivatization method and its application for the mass spectrometric analysis of analytes containing the carboxyl group

A novel isotope labeling reagent d0-/d6-2, 4-dimethoxy-6-piperazin-1-yl pyrimidine (DMPP) has been developed for derivatization toward the carboxyl group based on carbodiimide chemistry for mass spectrometry (MS) analysis. The strengths of this derivatization strategy involve fast labeling (15 s), low chemical background and general access to most carboxylic analytes. This has been demonstrated using a series of compounds containing carboxylic acids, including peptides and proteins. To enhance the MS response of the derivatized analytes, the design of DMPP has been based on integration of the theoretical consideration of high gas-phase hydrogenation capacity and hydrophobicity. In addition, the high abundance product ions at m/z 225 and m/z 231 from d0-/d6-DMPP labeled carboxylic acids indicate high efficiency of the gas-phase cleavage induced by the labeling reagent. Quantitative determination of these ions can also be used in single reaction monitoring to achieve extremely high sensitivity toward the target analytes. This has subsequently been used to determine trace free fatty acids in human urine. Furthermore, the DMPP labeled peptides also provide additional sequence information in MALDI–MS/MS because of the formation of sequence-related isotope fragment ions. This DMPP-oriented labeling technique is expected to be a promising tool for the MS detection of many varieties of compounds containing carboxyl groups.

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► A novel MS-based isotope reagent DMPP was developed for the derivatization toward carboxyl group.
► The simple and rapid labeling reaction was carried out under mild conditions with high specificity.
► The LODs of analytes containing the carboxyl and excess labeling reagent were greatly reduced.
► Enhanced accuracy in peptide sequencing was achieved by the introduction of isotope-coded DMPP.