Electrospray ionization tandem mass spectrometry of protonated and alkali-cationized Boc-N-protected hybrid peptides containing repeats of D-Ala-APyC and APyC-D-Ala: Formation of [bn–1 + OCH3 + Na]+ and [bn–1 + OH + Na]+ ions


R. Srinivas, National Centre for Mass Spectrometry, CSIR – Indian Institute of Chemical Technology, Hyderabad 500 007, India.




Differentiation and structural characterization of positional isomers of non-natural amino acid hybrid peptides by using electrospray ionization tandem mass spectrometry (ESI-MSn) is desirable because of their fundamental importance from the view point of peptide mass spectrometry and also of their increasing importance in the area of research towards biomedical and material applications; hence, the present study is undertaken.


Electrospray ionization ion-trap tandem mass spectrometry (ESI-MSn) was used to characterize and differentiate three pairs of positional isomers of Boc-N-protected hybrid peptides containing repeats of D-Ala-APyC and APyC-D-Ala (D-Ala = D-alanine and APyC = trans-3-aminopyran-2-carboxylic acid).


ESI-MSn spectra of protonated and alkali-cationized positional isomeric peptides display characteristic fragmentation involving the peptide backbone, the Boc group, and the side chain. It is observed that abundant rearrangement ions [bn–1 + OCH3 + Na]+ or [bn–1 + OH + Na]+ are formed when D-Ala is present at C-terminus and the presence of APyC at the C-terminus inhibits the formation of rearrangement ions. In addition, abundant bn–1+ ions are formed, presumably with stable oxazolone structures, when the C-terminus of bn–1+ ions possessed D-Ala.


The present study demonstrates that ESI tandem mass spectrometry is very useful for differentiating positional isomers of hybrid peptides containing D-Ala and APyC amino acids. While the protonated peptides give rise to characteristic sequencing ions, the cationized peptides produce additional rearrangement ions ([bn–1 + OCH3 + Na]+ and [bn–1 + OH + Na]+) which helps distinguish between the presence of D-Ala and APyC amino acids at the C-terminus. Copyright © 2012 John Wiley & Sons, Ltd.